• TABLE OF CONTENTS
HIDE
 Front Cover
 Front Matter
 Frontispiece
 Front Matter
 Preface
 List of contributors
 Table of Contents
 Taxonomy and nomenclature...
 Fruit characters in young trees...
 Classifying certian diseases as...
 The color test for exocortis indexing...
 Evidence for strain differences...
 Relationship between exocortis...
 A quick field test for exocort...
 Testing for exocortis
 Reaction of rootstocks to...
 Can phytophthora spp. transmit...
 Psorosis in Venezuela - an...
 Chemical studies on stubborn-affected...
 Stunting and chlorosis induced...
 Virus content of citrus trees with...
 Morhological modifications induced...
 Response of stubborn-infected trees...
 Differences in temple orange color...
 Some possible anatomical and serological...
 Tristeza in the Philippines
 Tristeza in Florida
 The danger of introducing tristeza...
 Tristeza and stem pitting...
 Studies on likubin
 Variations in aphid transmission...
 Is there tristeza in Andhra Pradesh,...
 Anatomical aspects of tristeza-diseased...
 Seedling yellows in California
 Comparative reactions or Orlando...
 Evaluation of indicators for xyloporosis...
 A disorder of rangpur lime and...
 Scion-rootstock incompatibilities...
 Impietratura in mediterranean...
 Observations and research...
 Responses of citrus to concurrent...
 Mechanical transmission of infections...
 Lemon crinkly leaf virus
 Bud certification in Arizona
 The budwood registration program...
 The California citrus variety improvement...
 Indexing citrus for viruses in...
 Citrus virus diseases in Argen...
 Occurrence of citrus virus diseases...
 Citrus virus diseases in Corsi...
 Citrus virus diseases in the region...
 Citrus virus diseases in Japan
 Citrus virus diseases in Tunis...
 Suggested procedures and differential...
 Index
 Back Cover






Title: Proceedings of the second conference of the International Organization of Citrus Virologists
CITATION THUMBNAILS PAGE IMAGE ZOOMABLE
Full Citation
STANDARD VIEW MARC VIEW
Permanent Link: http://ufdc.ufl.edu/UF00089519/00001
 Material Information
Title: Proceedings of the second conference of the International Organization of Citrus Virologists
Physical Description: xiv, 265 p. : ill., port. ; 24 cm.
Language: English
Creator: International Organization of Citrus Virologists -- Conference, 1960
Price, W. Conway ( William Conway ), 1906-
Publisher: University of Florida Press,
University of Florida Press
Place of Publication: Gainesville
Publication Date: 1961
Copyright Date: 1961
 Subjects
Subject: Citrus -- Diseases and pests -- Congresses   ( lcsh )
Virus diseases of plants -- Congresses   ( lcsh )
Genre: conference publication   ( marcgt )
non-fiction   ( marcgt )
 Notes
Statement of Responsibility: edited by W.C. Price.
General Note: Includes index and bibliographical references.
 Record Information
Bibliographic ID: UF00089519
Volume ID: VID00001
Source Institution: University of Florida
Holding Location: University of Florida
Rights Management: All rights reserved by the source institution and holding location.
Resource Identifier: oclc - 221248208

Table of Contents
    Front Cover
        Front cover
    Front Matter
        Page i
    Frontispiece
        Page ii
    Front Matter
        Page iii
        Page iv
    Preface
        Page v
        Page vi
    List of contributors
        Page vii
        Page viii
        Page ix
        Page x
    Table of Contents
        Page xi
        Page xii
        Page xiii
        Page xiv
    Taxonomy and nomenclature in citrus
        Page 1
        Page 2
        Page 3
        Page 4
        Page 5
        Page 6
        Page 7
    Fruit characters in young trees of long-established nucellar lines
        Page 8
        Page 9
        Page 10
        Page 11
        Page 12
        Page 13
        Page 14
    Classifying certian diseases as inhereited
        Page 15
        Page 16
        Page 17
        Page 18
        Page 19
        Page 20
        Page 21
    The color test for exocortis indexing in Florida
        Page 22
        Page 23
        Page 24
        Page 25
    Evidence for strain differences and stunting with exocortis virus
        Page 26
        Page 27
        Page 28
        Page 29
        Page 30
        Page 31
        Page 32
        Page 33
    Relationship between exocortis and stunting of citrus varieties on poncirus trifoliata rootstock
        Page 34
        Page 35
        Page 36
        Page 37
        Page 38
        Page 39
    A quick field test for exocortis
        Page 40
        Page 41
        Page 42
    Testing for exocortis
        Page 43
        Page 44
        Page 45
        Page 46
        Page 47
        Page 48
        Page 49
    Reaction of rootstocks to exocortis
        Page 50
        Page 51
        Page 52
        Page 53
        Page 54
        Page 55
    Can phytophthora spp. transmit psorosis
        Page 56
    Psorosis in Venezuela - an emendation
        Page 57
        Page 58
        Page 59
    Chemical studies on stubborn-affected marsh grapefruit and washington navel oranges
        Page 60
        Page 61
        Page 62
        Page 63
        Page 64
        Page 65
        Page 66
        Page 67
        Page 68
    Stunting and chlorosis induced in young-line citrus plants by inoculations from navel orange trees having symptoms of stubborn disease
        Page 69
        Page 70
        Page 71
        Page 72
        Page 73
        Page 74
        Page 75
        Page 76
    Virus content of citrus trees with symptoms of stubborn disease
        Page 77
        Page 78
    Morhological modifications induced by stubborn disease on citrus fruits
        Page 79
        Page 80
        Page 81
        Page 82
        Page 83
    Response of stubborn-infected trees to iron chelates
        Page 84
        Page 85
        Page 86
        Page 87
        Page 88
        Page 89
        Page 90
        Page 91
        Page 92
    Differences in temple orange color and quality associated with stylar-end greening
        Page 93
        Page 94
        Page 95
        Page 96
    Some possible anatomical and serological techniques in diagnosing stubborn disease in citrus
        Page 97
        Page 98
        Page 99
        Page 100
    Tristeza in the Philippines
        Page 101
        Page 102
        Page 103
        Page 104
        Page 105
        Page 106
    Tristeza in Florida
        Page 107
        Page 108
        Page 109
        Page 110
        Page 111
        Page 112
    The danger of introducing tristeza and its most efficient vector into mediteranean area
        Page 113
        Page 114
        Page 115
    Tristeza and stem pitting in Brazil
        Page 116
        Page 117
        Page 118
        Page 119
        Page 120
    Studies on likubin
        Page 121
        Page 122
        Page 123
        Page 124
        Page 125
    Variations in aphid transmission of tristeza virus
        Page 126
        Page 127
        Page 128
        Page 129
        Page 130
        Page 131
    Is there tristeza in Andhra Pradesh, India
        Page 132
        Page 133
        Page 134
        Page 135
    Anatomical aspects of tristeza-diseased citrus, aeglopsis, and afraegle
        Page 136
        Page 137
        Page 138
        Page 139
        Page 140
    Seedling yellows in California
        Page 141
        Page 142
        Page 143
        Page 144
        Page 145
        Page 146
        Page 147
        Page 148
        Page 149
    Comparative reactions or Orlando tangelo and palestine sweet lime to cachexia and xyloporsis
        Page 150
        Page 151
        Page 152
        Page 153
        Page 154
        Page 155
        Page 156
        Page 157
        Page 158
    Evaluation of indicators for xyloporosis and exocortis in Texas
        Page 159
        Page 160
        Page 161
        Page 162
        Page 163
        Page 164
        Page 165
    A disorder of rangpur lime and citron on sweet orange
        Page 166
        Page 167
        Page 168
        Page 169
        Page 170
        Page 171
    Scion-rootstock incompatibilities in Brazil
        Page 172
        Page 173
        Page 174
        Page 175
        Page 176
    Impietratura in mediterranean countries
        Page 177
        Page 178
        Page 179
        Page 180
        Page 181
    Observations and research on impietratura
        Page 182
        Page 183
        Page 184
        Page 185
        Page 186
    Responses of citrus to concurrent infection with two or more unrelated viruses
        Page 187
        Page 188
        Page 189
        Page 190
        Page 191
        Page 192
        Page 193
        Page 194
        Page 195
        Page 196
    Mechanical transmission of infections variegation virus in citrus and noncitrus hosts
        Page 197
        Page 198
        Page 199
        Page 200
        Page 201
        Page 202
        Page 203
        Page 204
    Lemon crinkly leaf virus
        Page 205
        Page 206
        Page 207
        Page 208
        Page 209
        Page 210
    Bud certification in Arizona
        Page 211
        Page 212
        Page 213
        Page 214
        Page 215
    The budwood registration program for the rio citrus area
        Page 216
        Page 217
        Page 218
        Page 219
    The California citrus variety improvement program
        Page 220
        Page 221
        Page 222
        Page 223
        Page 224
        Page 225
    Indexing citrus for viruses in Texas
        Page 226
        Page 227
        Page 228
        Page 229
        Page 230
    Citrus virus diseases in Argentina
        Page 231
        Page 232
        Page 233
        Page 234
        Page 235
        Page 236
        Page 237
    Occurrence of citrus virus diseases in the state of São Paulo
        Page 238
        Page 239
        Page 240
        Page 241
    Citrus virus diseases in Corsica
        Page 242
        Page 243
        Page 244
    Citrus virus diseases in the region of Fondi, Italy
        Page 245
        Page 246
    Citrus virus diseases in Japan
        Page 247
        Page 248
        Page 249
        Page 250
        Page 251
        Page 252
    Citrus virus diseases in Tunisia
        Page 253
        Page 254
        Page 255
    Suggested procedures and differential hosts for identifying viruses
        Page 256
        Page 257
        Page 258
        Page 259
    Index
        Page 261
        Page 262
        Page 263
        Page 264
        Page 265
    Back Cover
        Page 266
Full Text



Americas, and the Mediterranean.
Most of them are experienced field
plant pathologists, bringing into their
writings a wealth of citrus crop
knowledge to complement the more
technical aspects of their research re-
ports. Few groups of crop plant dis-
eases have been so exhaustively treat-
ed in comparable compact volumes.
The two volumes of papers pre-
sented at the International Confer-
ences are evidence that the study of
citrus virus diseases shares in the cur-
rent high degree of international vol-
untary cooperative effort to solve ag-
ricultural Iroblems throughout the
world. Their publication results not
only in prompt di,-semination of all
presently available information, but
also will stimulate additional research
and encourage the solution of local
problems throughout the world.
This book will be an absolutely nec-
essary reference for all research
workers studying citrus diseases. All
plant pathologists, but especially
those dealing with diseases of tree
crops, will find in the book a wealth
of observational data. This book
should also be required reading for
all students and other technical per-
sonnel associated in any way with
citrus crop production. Finally, all
well-informed citrus growers, nurs-
erymen, and citriculturists will find
this book both interesting and in-
formative.


UNIVERSITY OF FLORIDA PRESS


I
.j J,









i',











jI
4fi



k .


Gainesville


a -
























Proceedings
of the
SECOND CONFERENCE
of the
International Organization of
CITRUS VIROLOGISTS


























CITRUS EYPERIlJFNT FIIII


DELEGATES TO THE SECOND CONFERENCE OF THE INTERNATIONAL ORGANIZATION OF CITRUS VIROLOGISTS


:ai

"i























Proceedings
of the
SECOND CONFERENCE
of the
International Organization of
CITRUS VIROLOGISTS

Edited by W. C. Price

UNIVERSITY OF FLORIDA PRESS / GAINESVILLE / 1961













PUBLICATION OF THIS VOLUME, The Proceedings of the Second Con-
ference of the International Organization of Citrus Virologists, was
made possible through the generosity of the
FLORIDA CITRUS NURSERYMEN'S ASSOCIATION
who sponsored its publication, and the following members of the Asso-
ciation, who underwrote the costs of publication:


ADAMS CITRUS NURSERY
Winter Haven
Florida

BROOKSVILLE CITRUS GROWERS
ASSOCIATION
Brooksville, Florida

ELOISE GROVES ASSOCIATION
P. O. Box 1312
Winter Haven, Florida

FLORIDA PONKAN CORPORATION
Citrus Nurseries
Apopka, Florida

GRAND ISLAND NURSERIES
P. O. Box 906
Eustis, Florida


H & D NURSERY
Tavares
Florida

LAKE GARFIELD NURSERIES
COMPANY
Bartow, Florida

MCMULLEN GROVE
2711 Jetton Avenue
Tampa, Florida

MINUTE MAID CORPORATION
Soils and Horticulture Division
Plymouth, Florida

SOUTHERN GROVES NURSERY
P. O. Box 1400
Winter Haven, Florida


WARD'S NURSERY, INC.
Avon Park
Florida


A University of Florida Press Book
TITLE PAGE DRAWING BY ANNE HAYDEN VALK
COPYRIGHT, 1961, BY THE BOARD OF COMMISSIONERS
OF STATE INSTITUTIONS OF FLORIDA
LIBRARY OF CONGRESS CATALOG CARD NO. 61-64183
PRINTED BY THE H. & W. B. DREW COMPANY
JACKSONVILLE, FLORIDA













Preface


CITRUS VIRUS DISEASES are such important problems that laboratories
for their study have been established in all the important citrus-growing
regions of the world. The largest of these laboratories is at Riverside,
California, where nine or more specialists devote much of their time to
study of these virus diseases.
When the fiftieth anniversary of the Citrus Experiment Station of the
University of California was celebrated, citrus virologists from around
the world were invited to meet at Riverside in November, 1957, and
discuss their mutual problems. One result of this Conference was pub-
lication of 35 papers in Citrus Virus Diseases, edited by James M.
Wallace and published by the University of California Division of Agri-
cultural Sciences at Berkeley. Another result was the banding together
of the citrus virologists into an informal organization called The Inter-
national Organization of Citrus Virologists. Dr. Wallace was elected
chairman of the new organization.
The Second Conference of the International Organization of Citrus
Virologists was held in Florida November 7-11, 1960. Formal papers
were presented at meetings at the Citrus Experiment Station of the
University of Florida, Lake Alfred, on Monday and Tuesday and in
Orlando on Thursday and Friday; a field trip arranged by the Florida
State Plant Board through an area having the largest concentration of
citrus trees in the world was made on Wednesday; a business meeting
was held at the USDA Horticultural Station in Orlando on Saturday
morning; and another field trip was made in the Orlando area on
Saturday afternoon. Some of the delegates elected to take an optional
field trip to the Indian River area on Saturday and Sunday. Dr. T. J.
Grant was elected to succeed Dr. Wallace as chairman.
Delegates from Argentina, Brazil, France, Israel, Italy, Japan,
Morocco, Peru, Philippines, Spain, Taiwan, Trinidad, Tunisia, Turkey,
United States, and Venezuela attended the Second Conference. Forty-







PROCEEDINGS of the IOCV


one papers were presented and 12 additional papers were read by title
because the authors were unable to attend in person.
The 35 papers in Citrus Virus Diseases summarize the observations,
research findings, and development of control methods from the begin-
ning to 1957. The 47 papers in the present volume represent progress
made since then. Consequently, reviews of literature and literature
citations have been kept to a minimum; the reader is referred to Citrus
Virus Diseases for more extensive citations.
This volume, Proceedings of the Second Conference of the Interna-
tional Organization of Citrus Virologists, and other volumes to follow,
will bring together in one place the most recently accumulated knowl-
edge about a very important group of diseases-the citrus virus diseases.
The volumes will be of interest to many individuals who are not special-
ists in virus diseases-to pathologists, horticulturists, citrus growers, pro-
duction managers, nurserymen, service and regulatory service personnel,
and teachers in all countries where citrus is grown.
Many of the manuscripts submitted for publication were too long to
be published here in their entirety; had they been so published, the Pro-
ceedings would have been almost twice as long as it now is. Authors
were very generous in permitting their manuscripts to be cut to pieces
and shortened and in allowing me to delete many excellent illustrations
that could not be included because of the cost that would have been in-
volved in reproducing them. The changes made, including those of
literature citations, are in keeping with the policies adopted by the Com-
mittee on Form and Style of the Conference of Biological Editors and
published in the Style Manual for Biological Journals.
In editing the Proceedings, I attempted to make the language as clear
and concise as possible, but avoided changing an author's conclusions or
point of view. Many of the papers are written by those to whom the
English language is not native and some of them contain expressions
that may sound peculiar to the American ear; when such expressions do
not present an ambiguity, they add a certain flavor to which the author
is entitled.
W. C. PRICE
Gainesville, Florida



When citing this book, use the following abbreviation:
Proc. 2nd Conf. Intern. Organization Citrus Virol.













Contributors


Ross M. ALLEN, University of Arizona, Yuma Branch Experiment Sta-
tion, Yuma, Arizona
AVELINO E. BIGORNIA, Guinobatan Experiment Station, Guinobatan,
Albay, Philippines
COLETTE BOVE, Institut Francais de Recherches Frutieres Outre-Mer,
Paris 16, France
JOSEPH M. BovI, Institut Francais de Recherches Frutieres Outre-Mer,
Paris 16, France
RALPH T. BROWN, Plaquemines Parish Experiment Station, Sulphur,
Louisiana
GORDON BUFFINGTON, Texas Agricultural Experiment Station, Carrizo
Springs, Texas
HARRY C. BURNETT, Division of Plant Industry, Florida Department of
Agriculture, Winter Haven, Florida
E. C. CALAVAN, University of California, Riverside, California
CARLOS A. CALICA, Guinobatan Experiment Station, Guinobatan, Al-
bay, Philippines
J. W. CAMERON, University of California, Riverside, California
J. B. CARPENTER, U.S. Date Field Station, Indio, California
H. CHAPOT, Institut Francais de Recherches Fruiti&res Outre-Mer 5,
Avenue de la Gare, Rabat, Morocco
J. F. L. CHILDS, U.S. Horticultural Field Station, Orlando, Florida
D. W. CHRISTIANSEN, University of California, Riverside, California
MORTIMER COHEN, University of Florida Citrus Experiment Station,
Fort Pierce, Florida
M. K. CORBETT, University of Florida, Gainesville, Florida
J. Cox, New South Wales Department of Agriculture, Sydney, N.S.W.,
Australia
T. A. DEWOLFE, University of California, Riverside, California
R. J. DRAKE, University of California, Riverside, California
vii







PROCEEDINGS of the IOCV


M. V. FERNANDEZ VALIELA, Estaci6n Experimentale Agropecuaria dcl
Delta del ParanA, Campana, Argentina
LILIAN R. FRASER, New South Wales Department of Agriculture, Rydal-
mere, N.S.W., Australia
E. F. FROLICH, University of California, Los Angeles, California
D. C. GIACOMETTI, Brazilian Ministry of Agriculture, Rio de Janeiro,
Brazil
INGVAR GRANHALL, European and Mediterranean Plant Protection Or-
ganisation, 142, Avenue des Champs-Slys6es, Paris, France
THEODORE J. GRANT, U.S. Horticultural Field Station, Orlando, Florida
R. H. HILGEMAN, University of Arizona, Citrus Branch Station, Tempe,
Arizona
ROBERT W. HODGSON, University of California, Los Angeles, California
BECHIR JAMOUSSI, Ecole Sup6rieure d'Agriculture de Tunis, Route de
l'Ariana, Tunis
L. J. KLOTZ, University of California, Riverside, California
L. C. KNORR, University of Florida, Citrus Experiment Station, Lake
Alfred, Florida
NORBERTO LEITE, Brazilian National Research Council, Rio de Janeiro,
Brazil
E. C. LEVITT, New South Wales Department of Agriculture, Sydney,
N.S.W., Australia
WILLIAM G. LONG, U.S. Horticultural Field Station, Orlando, Florida
GINO MALAGUTI, Centro de Investigaciones Agronomicas, Maracay,
Venezuela
T. MATSUMOTO, National Taiwan University, Taipei, Formosa
FRANCOISE MONIER, Institut Franqais de Recherches Frutieres Outre-
Mer, Paris 16, France
SYLVIO MOREIRA, Instituto Agronomico, Campinas, Brazil
GEORGES MOREL, Centre Nationale de Recherches Agronomiques, Ver-
sailles (S & O), France
PAUL A. NORMAN, U.S. Horticultural Field Station, Orlando, Florida
EDWARD O. OLSON, Agricultural Research Service, U.S. Department of
Agriculture, Weslaco, Texas
P. GOVINDA RAO, Department of Agriculture, Government of Andhra
Pradesh, India
G. S. REDDY, Department of Agriculture, Government of Andhra
Pradesh, India
WALTER REUTHER, University of California, Riverside, California








CONTRIBUTORS


PEDRO RODRIGO, Bureau of Plant Industry, Manila, Philippines
VICTORIA ROSSETTI, Instituto Biol6gico, Sao Paulo, Brazil
GAETANO RUGGIERI, Stazione Sperimentale Agrumicultura, Acireale,
Italy
ARY A. SALIBE, Instituto Agron6mico, Campinas, Brazil
HENRY SCHNEIDER, University of California, Riverside, California
ART SHULL, Citrus Experimental Department, Rio Farms, Inc., Ed-
couch, Texas
CESARE SIBILIA, Plant Pathology Station, Via Casal de' Pazzi, 250,
Rome, Italy
JAMES B. SINCLAIR, Louisiana State University, Baton Rouge, Louisiana
BAILEY SLEETH, Texas Agricultural Experiment Station, Weslaco, Texas
R. K. SoosT, University of California, Riverside, California
L. W. STORM, University of Arizona, Tucson, Arizona
R. B. STREETS, University of Arizona, Tucson, Arizona
H. J. Su, National Taiwan University, Taipei, Formosa
SHOICHI TANAKA, Tokai-Kinki Agricultural Experiment Station,
Okitsu-Machi, Schizuoka-Ken, Japan
H. H. THORNBERRY, University of Illinois, Urbana, Illinois
R. VOGEL, Station Exp6rimentale d'Agrumiculture, San Giuliano,
Corsica
J. M. WALLACE, University of California, Riverside, California
M. C. WANG, National Taiwan University, Taipei, Formosa
L. G. WEATHERS, University of California, Riverside, California
SHUNICHI YAMADA, T6kai-Kinki Agricultural Experiment Station,
Okitsu-Machi, Schizuoka-Ken, Japan












International Organization of Citrus Virologists


M EMBERSHIP IS OPEN to anyone who is interested in the virus dis-
eases of citrus. A membership fee of $5.00, payable to E. O. Olson, is
required for the period between the second and third international
meetings.

OFFICERS
Chairman-T. J. GRANT, U.S. Horticultural Field Station, 2120 Camden Road,
Orlando, Florida
Secretary-Treasurer-E. O. OLSON, U.S.D.A. Laboratory, P. O. Box 267, Wes-
laco, Texas

CHAIRMEN OF COMMITTEES
COMMITTEE ON TIME AND LOCATION OF THE THIRD CONFERENCE
J. M. WALLACE, Citrus Experiment Station, Riverside, California

COMMITTEE ON BIBLIOGRAPHY
JOSEPH M. BovE, I.F.A.C., 6, rue du General-Clergerie, Paris,
France

COMMITTEE ON FINANCES AND MEMBERSHIP
P. W. ROHRBAUGH, Texas College of Arts and Industries, Citrus and
Vegetable Training Center, Weslaco, Texas

COMMITTEE ON INDEXING PROCEDURES, DIAGNOSIS, AND NOMENCLATURE
J. F. L. CHILDS, U.S. Horticultural Field Station, 2120 Camden
Road, Orlando, Florida

COMMITTEE ON LIAISON WITH OTHER ORGANIZATIONS
I. REICHERT, Division of Plant Pathology, Agricultural Research
Station, Rehovot, Israel

COMMITTEE ON ROOTSTOCKS AND NUCELLAR CLONES
W. P. BITTERS, Citrus Experiment Station, Riverside, California

COMMITTEE ON PUBLICATIONS
E. P. DUCHARME, Citrus Experiment Station, Lake Alfred, Florida

PAST OFFICERS
Chairman, 1957-1960-J. M. WALLACE













Contents


PREFACE ..--..-....................-- ----.....-- v
LIST OF CONTRIBUTORS-...........-...... ..--- .......- ...-............... vii
TAXONOMY AND NOMENCLATURE IN CITRUS
Robert W. Hodgson---.....--------- --....... .. ---....--..... 1
FRUIT CHARACTERS IN YOUNG TREES OF LONG-
ESTABLISHED NUCELLAR LINES
R. K. Soost and J. W Cameron......................----- ....--........... 8
CLASSIFYING CERTAIN DISEASES AS INHERITED
Henry Schneider, J. W. Cameron,
R. K. Soost, and E. C. Calavan...................... ................ 15
THE COLOR TEST FOR EXOCORTIS INDEXING IN
FLORIDA
Harry C. Burnett..- -........-------..... .. ...............---....... 22
EVIDENCE FOR STRAIN DIFFERENCES AND STUNTING
WTH EXOCORTIS VIRUS
E. C. Calavan and L. G. Weathers....----................................ 26
RELATIONSHIP BETWEEN EXOCORTIS AND STUNTING
OF CITRUS VARIETIES ON PONCIRUS TRIFOLIATA
ROOTSTOCK
L. R. Fraser, E. C. Levitt, and J. Cox-----------..................................... 34
A QUICK FIELD TEST FOR EXOCORTIS
Sylvio Moreira .................. ---------------------- 40
TESTING FOR EXOCORTIS
Victoria Rossetti ---........ .........................------- ---.. 43
REACTION OF ROOTSTOCKS TO EXOCORTIS
James B. Sinclair and Ralph T. Brown-----------................................... 50
CAN PHYTOPHTHORA SPP. TRANSMIT PSOROSIS
L. J. Klotz and T. A. DeWolfe--------------................ ----................ 56
PSOROSIS IN VENEZUELA-AN EMENDATION
Gino Malaguti and L. C. Knorr......................--------.--..- 57

xi








PROCEEDINGS of the IOCV


CHEMICAL STUDIES ON STUBBORN-AFFECTED MARSH
GRAPEFRUIT AND WASHINGTON NAVEL ORANGES
Colette Bov6, Georges Morel, Francoise
Monier, and Joseph M. Bov-...........................-.......- 60
STUNTING AND CHLOROSIS INDUCED IN YOUNG-LINE
CITRUS PLANTS BY INOCULATIONS FROM NAVEL
ORANGE TREES HAVING SYMPTOMS OF STUB-
BORN DISEASE
E. C. Calavan and D. W. Christiansen..............-.-----. 69
VIRUS CONTENT OF CITRUS TREES WITH SYMPTOMS
OF STUBBORN DISEASE
J. B. Carpenter -..... ...-- ......-.-----........ 77
MORPHOLOGICAL MODIFICATIONS INDUCED BY
STUBBORN DISEASE ON CITRUS FRUITS
H. Chapot ........-....-...-......... ... -- --- 79
RESPONSE OF STUBBORN-INFECTED TREES TO IRON
CHELATES
R. H. Hilgeman ...-~.......- ...-..-.......-- 84
DIFFERENCES IN TEMPLE ORANGE COLOR AND QUAL-
ITY ASSOCIATED WITH STYLAR-END GREENING
William G. Long and J. F. L. Childs.... ..........---... .. 93
SOME POSSIBLE ANATOMICAL AND SEROLOGICAL
TECHNIQUES IN DIAGNOSING STUBBORN DISEASE
IN CITRUS
L. W Storm and R. B. Streets .................... ......... 97
TRISTEZA IN THE PHILIPPINES
Avelino E. Bigornia and Carlos A. Calica-_ ............ ..........--- 101
TRISTEZA IN FLORIDA
Mortimer Cohen and Harry C. Burnett............. ~.-........ 107
THE DANGER OF INTRODUCING TRISTEZA AND ITS
MOST EFFICIENT VECTOR INTO THE MEDITER-
RANEAN AREA
Ingvar Granhall .... ... .............. ............ .- 113
TRISTEZA AND STEM PITTING IN BRAZIL
T. J. Grant, S. Moreira, and Ary A. Salibe... .~......--............. 116
STUDIES ON LIKUBIN
T. Matsumoto, M. C. Wang, and H. J. Su ...--...- -------..... 121
xii








CONTENTS


VARIATIONS IN APHID TRANSMISSION OF TRISTEZA
VIRUS
Paul A. Norman and Theodore J. Grant..---....-..----.................... 126
IS THERE TRISTEZA IN ANDHRA PRADESH, INDIA
G. S. Reddy and P. Govinda Rao ...................... ....-.......... 132
ANATOMICAL ASPECTS OF TRISTEZA-DISEASED
CITRUS, AEGLOPSIS, AND AFRAEGLE
Henry Schneider--- .............. ---- --....-....... .......--........ 136
SEEDLING YELLOWS IN CALIFORNIA
J. M W allace and R. J. Drake--.............. .... -- ........- -..... 141
COMPARATIVE REACTIONS OF ORLANDO TANGELO
AND PALESTINE SWEET LIME TO CACHEXIA AND
XYLOPOROSIS
E. C. Calavan, D. W. Christiansen, and L. G. Weathers--.......... 150
EVALUATION OF INDICATORS FOR XYLOPOROSIS
AND EXOCORTIS IN TEXAS
Edward O. Olson, Art Shull, and Gordon Buffington.............. 159
A DISORDER OF RANGPUR LIME AND CITRON ON
SWEET ORANGE
E. F. Frolich and R. W Hodgson.....................--............... 166
SCION-ROOTSTOCK INCOMPATIBILITIES IN BRAZIL
Ary A. Salibe...----...-. ---- .........--..----.......... 172
IMPIETRATURA IN MEDITERRANEAN COUNTRIES
H Chapot ............ .....-- ... -- ............ 177
OBSERVATIONS AND RESEARCH ON IMPIETRATURA
Gaetano Ruggieri --......-------------...........- --- ----..... 182
RESPONSES OF CITRUS TO CONCURRENT INFECTION
WITH TWO OR MORE UNRELATED VIRUSES
L. G. Weathers-.......--....--- ----- .........---..... 187
MECHANICAL TRANSMISSION OF INFECTIOUS VARIE-
GATION VIRUS IN CITRUS AND NONCITRUS
HOSTS
T. J. Grant and M. K. Corbett------.............- ..---......... 197
LEMON CRINKLY LEAF VIRUS
L. R. Fraser......- .....------ ----- ---- ........-- ...... 205
BUD CERTIFICATION IN ARIZONA
Ross M Allen and R. B. Streets.........~........ ..-- ........... 211








PROCEEDINGS of the IOCV


THE BUDWOOD REGISTRATION PROGRAM FOR THE
RIO CITRUS AREA
D. C. Giacometti and Norberto Leite................. ... .....-----------. 216
THE CALIFORNIA CITRUS VARIETY IMPROVEMENT
PROGRAM
Walter Reuther ......--..............--------.....------220
INDEXING CITRUS FOR VIRUSES IN TEXAS
Bailey Sleeth .............................. ------ ---------- 226
CITRUS VIRUS DISEASES IN ARGENTINA
M. V. Fernandez Valiela---.-----.....-----.................. 231
OCCURRENCE OF CITRUS VIRUS DISEASES IN THE
STATE OF SAO PAULO
Victoria Rossetti and Ary A. Salibe ...-------........------....-..----- 238
CITRUS VIRUS DISEASES IN CORSICA
R. Vogel ......................---. ............- --- 242
CITRUS VIRUS DISEASES IN THE REGION OF FONDI,
ITALY
Cesare Sibilia .......- --- -...........--- ----------. 245
CITRUS VIRUS DISEASES IN JAPAN
Shoichi Tanaka and Shunichi Yamada.---..-.~.-......----......-... ... -247
CITRUS VIRUS DISEASES IN TUNISIA
B6chir Jamoussi .---............------...--... ---..--........... 253
SUGGESTED PROCEDURES AND DIFFERENTIAL HOSTS
FOR IDENTIFYING VIRUSES
H. H. Thornberry..---............-......----- ... ...... ..---- 256
INDEX ..-..-..-----.................--------------.--- 261











ROBERT W. HODGSON


Taxonomy and Nomenclature in Citrus








M ANY HORTICULTURISTS are inclined to question the utility of knowl-
edge concerning the botanical classification of the plants with which
their studies or work are concerned, and for those engaged in certain
fields of horticultural research this attitude may have some justification.
On the other hand, such knowledge is almost indispensable to the
breeder and finds important applications in certain other fields. An
illustration is afforded by the problem of rootstock-scion relations in
general and graft compatibility and virus reactions in particular. In both
of these cases, knowledge of natural relationships, as reflected in botani-
cal classification, provides a basis for both understanding and predicting
rootstock-scion relations and virus reactions.
Indeed the tristeza virus rootstock-scion reactions so extensively re-
ported by virologists and horticulturists in recent years have provided
information of great value concerning the degree of natural relationship
between the rootstocks employed. Likewise the colorimetric identifica-
tion tests developed a few decades ago and considerably extended and
refined in recent years have contributed useful information in this con-
nection. An example is afforded by the rough lemon, which commonly
has been grouped with the true lemons and assigned to Citrus limon.
Its reactions to the tristeza virus and colorimetric tests cast extreme
doubt on the validity of this classification and suggest that the rough
lemon may deserve species standing, a belief long held by some botanists
and horticulturists. Indeed these reactions actually indicate possible re-
lationship to the Rangpur lime, likewise a conclusion reached by several
botanists.







PROCEEDINGS of the IOCV


Whatever their opinion of the value of taxonomic botany, plant scien-
tists in general and horticulturists and plant pathologists in particular,
in reporting research results, are confronted with the question of the
botanical nomenclature to be used in describing the plant materials with
which they have worked. Obviously the objective should be to employ
that nomenclature which indicates most accurately and precisely the
species or botanical and horticultural varieties involved so that the
reader may recognize them as discrete entities (taxa) with which he is
acquainted or can become so by consulting authoritative reference works.
Standardized botanical nomenclature, while a highly desirable objec-
tive, has not yet been achieved however. The practical problem, there-
fore, to which this discussion is addressed, is to select and use that system
of nomenclature which most nearly attains the objective in question.


The Systems of Citrus Taxonomy and Nomenclature
Currently Available
There are only two modern systems available, of sufficient comprehen-
siveness to deserve consideration, namely those of W. T. Swingle (4) and
Tyozaburo Tanaka (5), and hence choice is restricted to one or the
other. Both represent the culmination of studies spread over approxi-
mately four decades during which each author published extensively.
Swingle's final and complete system of classification and nomenclature
appeared in 1943; that of Tanaka became available in English in 1954.
For obvious reasons, the former is that best known and currently most
widely used, at least in the English-speaking world.

The Comparative Backgrounds and Experience of
the Originators
Swingle's interest in citrus taxonomy was an outgrowth of the breed-
ing program of the U.S. Department of Agriculture, established by him
and H. J. Webber in Florida following the great freeze of 1894-95. In
pursuit of this interest, he travelled widely-principally in Japan, China,
the Philippines, and the Mediterranean-and assembled extensive study
collections of citrus materials from nearly all parts of the world. Prior to
the appearance of his monograph he had published 30 taxonomic papers
on plants of the orange subfamily. In addition to his work on citrus,
however, he made many other contributions of great horticultural im-
portance, notably those relating to the fig and date industries of the








HODGSON


Pacific Southwest. He was essentially a subtropical horticulturist with a
special interest in citrus breeding and taxonomy.
Tanaka became interested in citrus taxonomy while still a student in
the Imperial University, just prior to World War I, and has continued to
work actively in that field ever since. At various times during the period
1915-1930, he was associated with Swingle, both in this country and
Japan. In following up this interest, he has travelled more widely than
any other worker and has become better acquainted with the original
literature and herbarium materials. He has assembled what is probably
the best library in existence in this field, now sequestered in the Univer-
sity of Taiwan. He is undoubtedly acquainted at firsthand with a wider
range of citrus materials than anyone else. Prior to the appearance of his
monograph he had published at least 30 papers on citrus taxonomy. He
is primarily a systematist, who has specialized in Citrus, and secondarily
an economic botanist.
Despite the early association and long collaboration of Swingle and
Tanaka, their respective systems reflect widely divergent viewpoints and
conclusions.
THE TWO SYSTEMS BRIEFLY SUMMARIZED.-According to the system
of Swingle, the citrus fruits are assigned to 3 genera; namely Fortunella
(kumquats) with 2 subgenera and 4 species, Poncirus trifoliatee orange)
with 1 species and 1 botanical variety, and Citrus (citrus fruits) with 2
subgenera, 16 species and 8 botanical varieties. The citrus fruits of cur-
rent economic importance are assigned to 8 of the 10 species which con-
stitute the subgenus Eucitrus. Papeda is the other subgenus. Thus his
classification involves a total of 3 genera, 21 species, of which 16 are in
Citrus, and 9 botanical varieties.
Tanaka's treatment of the genera Fortunella and Poncirus corresponds
approximately to that of Swingle but he treats the genus Citrus quite
differently. He recognizes 2 subgenera, 8 sections, and 144 species. The
subgenus Archicitrus contains the following sections: Papeda-12
species, Limonellus-16, Citrophorum-21, Cephalocitrus-21, and
Aurantium-28, a total of 98 species. The subgenus Metacitrus consists
of the following sections: Osmocitrus-9 species, Acrumen-36, and
Pseudofortunella-1, a total of 46. Thus his classification of the citrus
fruits involves a total of 3 genera and 151 species, of which 144 are
in Citrus.
VALID CRITICISMS OF EACH SYSTEM.-While the writer does not claim
special competence in the field of citrus taxonomy, during the past 30

3







PROCEEDINGS of the IOCV


years he has made two trips to the Orient, involving nearly a year's
residence in India and visits to the principal collections there (1, 3),
and brought in and studied a large number of introductions from many
parts of the world (2). Additionally, he enjoyed the privilege of close
association with Tanaka for the better part of the year, the latter spent
recently as a Fulbright research scholar in the United States. From the
background of these experiences, it is his conclusion that each of these
systems is subject to valid criticisms. Most certainly, insofar as their
treatment of the genus Citrus is concerned, they represent extremes.
The justifiable criticisms of Swingle's treatment include the following:
1. It is not sufficiently comprehensive to cover the materials with
which horticulturists and plant pathologists are concerned, not to men-
tion the known gamut of the genus. Thus numerous ancient forms of the
Orient are ignored, some of which are of economic importance. Illustra-
tive of the Indian forms not treated are gajanimma, kichili (vadlapudi),
amilbed, attani, kimb, and sadaphal.
2. It denies species standing to many ancient, well-known, and highly
distinctive forms, some of which are of great economic importance.
Among these are the rough lemon, Indian (Palestine) sweet lime, Sat-
suma and kunembo mandarins, Rangpur lime, natsudaidai, and yuzu.
3. Its treatment of certain distinctive and important forms is compli-
cated, speculative, and cumbersome. This arises from denial of species
standing and the consequent necessity for their inclusion under the most
appropriate species. This in turn involves speculation as to probable or
hypothetical parentage. The result is that the user, if he wishes to be
exact and accurate, must employ a terminology which is speculative-in
that the parentage indicated cannot be proved-or cumbersome, or both.
Thus the rough lemon is indicated as C. limon hybrid, the Indian sweet
lime C. aurantifolia hybrid, the King mandarin C. reticulata-sinensis
hybrid, the Rangpur lime C. reticulata var. austera hybrid, and the
calamondin C. reticulata var. austera-Fortunella sp. hybrid.
4. It is lacking in consistency to a remarkable degree. Separate species
standing is not questioned for certain forms which are obviously closely
related but others equally obviously much less closely related are placed
in a single species. Examples of the former are provided by the pummelo
and grapefruit, the sour and sweet oranges, and the oval and round kum-
quats. Illustrations of the latter are the placing of all the kinds of man-
darin-Satsuma, kunembo, Nagpur, Mediterranean, tangerine, etc.-
in a single species, and similar treatment for the limes and lemons.

Wolfe (6) has recently called attention to the validity of some of these
criticisms.








HODGSON


In the writer's opinion, the principal valid criticism of Tanaka's sys-
tem is concerned with what appears to be an excessive number of species
in the genus Citrus-certainly more than the range of known citrus ma-
terials seems to require. Thus he recognizes 35 species in the mandarin
group alone. Obviously these must be based on very minor differences-
so small in fact as to be detectable with great difficulty if at all. It is
doubtful that some of these constitute valid species. Additionally, he has
granted species standing to numerous cultigens, some of which are un-
doubtedly justifiable but others of questionable validity. Likewise species
standing has been given to some natural hybrids and forms for which
such treatment seems unnecessary or inappropriate.

Discussion
It seems clear that the basic difference which led these two investi-
gators to such divergent conclusions relates to their respective concepts
as to what constitutes a Citrus species. Essentially one is a "lumper"
whereas the other is a "splitter." In the writer's opinion, the major
reason for this difference arises from their respective experiences and
interests.
Swingle's interest in citrus taxonomy was an outgrowth of his partici-
pation in the first citrus breeding program ever undertaken, which in-
volved materials belonging to what are now agreed upon as 3 genera and
early gave rise to a series of hybrids of striking variety and interest. In-
deed he soon faced the problem of nomenclature for his hybrids, which
he solved by the coinage of a series of new names indicating their general
parentage-citrange, tangelo, tangor, etc.
Because of his awareness of the rich variation which could arise from
citrus crosses, Swingle early reached the tentative conclusion that any
citrus form which exhibited characters similar to those observed in
another must be considered to be a probable hybrid relative, and hence
should be assigned to the species it most closely resembled. As he became
acquainted with the literature and when he visited the Orient and later
studied his numerous introductions, he encountered many forms with
characters which to him suggested hybrid origin. Indeed, he found some
which somewhat resembled hybrids created in his breeding program. As
a consequence, he was led to reject a large number of species established
or accepted by other workers. Still other species he appears to have
ignored.
Swingle's assumption that citrus forms which exhibit characters found
5








PROCEEDINGS of the IOCV


in other forms cannot be considered for species standing unless it can be
established that they are not of hybrid origin, while understandable in
the light of his experience as a citrus breeder, is not in accord with the
views of most botanical taxonomists. Hybridization is generally regarded
as one of the important modes of origin of new species. Many existing
species which the taxonomists do not hesitate to recognize and accept are
considered to have arisen from natural intercrossing at some time in the
past. Indeed, in recent decades a number of plant species have been sub-
jected to cytogenetic analysis, their component parentage determined,
and then resynthesized.
Tanaka, on the other hand, approached the problem of citrus classifi-
cation and nomenclature from the strictly botanical viewpoint. In the
writer's opinion, however, a dominating influence, almost from the out-
set, was his interest in and desire to determine the facts concerning the
center of origin, dissemination, and evolutionary development of Citrus.
It would appear that the latter objective has caused him to search for the
connecting links and to refine his species treatment to an unnecessary
degree, which, together with acceptance of cultigens and certain hybrids,
has led to the establishment or recognition of an excessive number of
species.
Since these two systems reflect such extreme divergence in species con-
cept, the possibility clearly exists that an intermediate system might be
developed which would not be subject to their valid criticisms and hence
of maximum utility to all concerned. For some years past, as the details
of the two systems have become available and the opportunity to study
the materials in the field has permitted, this has been an objective of the
writer. His conclusions to date are set forth below.


Citrus Species not Accepted by Swingle but Considered
to be Valid
ACID MEMBERS GROUP
Citrus bergamia Risso-bergamot
Citrus jambhiri Lushington-rough or Mazoe lemon, citronelle, jam-
buri
Citrus latifolia Tanaka-Tahiti or Persian lime
Citrus limetta Risso-Mediterranean sweet lemon
Citrus limettioides Tanaka-Palestine sweet lime, mitha nimbu
ORANGE GROUP
Citrus myrtifolia Rafinesque-chinnoto, chinois, hazara








HODGSON


MANDARIN GROUP
Citrus deliciosa Tenore-Mediterranean or Willowleaf mandarin
Citrus nobilis Loureiro-King mandarin, kunembo
Citrus reshni hort. Tanaka-Cleopatra mandarin
Citrus sunki hort. Tanaka-sunki
Citrus tangerina hort. Tanaka-tangerine
Citrus unshiu Marcovitch-Satsuma mandarin
OTHERS
Citrus junos Siebold-yuzu
Citrus karna Rafinesque-karna, kharna khatta, khatta nimbu, id
nimbu
Citrus limonia Osbeck-Rangpur lime, Canton or cravo lemon, Ota-
heite orange, mandarin-lime
Citrus macrophylla Wester-colo, alemow
Citrus maderaspatana hort. Tanaka-kichili, vadlapudi, Guntur sour
orange
Citrus madurensis Loureiro-calamondin
Citrus natsudaidai Hayata-natsudaidai, dai dai mikan, Japanese
summer grapefruit
Citrus pennivesciculata Tanaka-gajanimma. Identical with C. Moi.


Literature Cited

1. HODGSON, R. W. 1937. The citrus fruits of India. Calif. Citrograph 22: 504,
513-514, 517.
2. HODGSON, R. W. 1955. Citrus introductions at UCLA. Calif. Citrograph 40:
164, 172-176.
3. HODGSON, R. W. 1961. Citriculture in India. Calif. Citrograph 46: 66, 76,
78-79, 81.
4. SWINGLE, W. T. 1943. Botany of citrus and its wild relatives of the orange
subfamily, p. 129-474. In H. J. Webber and L. D. Batchelor [ed.], The
Citrus Industry, Vol. I. Univ. Calif. Press, Berkeley and Los Angeles.
5. TANAKA, T. 1954. Species problem in citrus. Published by the Japanese So-
ciety for the Promotion of Science, Uemo, Tokyo. 152 p. 3 pl.
6. WOLFE, H. S. 1959. Some problems of citrus nomenclature. Proc. Amer. Soc.
Hort. Sci. (Caribbean Region) 7: 18-21.










R. K. SOOST and J. W. CAMERON


Fruit Characters in Young Trees of
Long-Established Nucellar Lines











THE VALUE of nucellar embryony for obtaining virus-free lines of
citrus was considered by Weathers and Calavan at the First International
Citrus Virus Conference (4). At the same time, Cameron et al. (2) dis-
cussed horticultural aspects of nucellar seedling lines which are being
used in California. The need to maintain high fruit quality and trueness
to type within citrus varieties makes it important to study critically the
fruit characters of lines which may be used commercially. If these
characters differ from those of established lines, it should be determined
whether the differences are genetic, and therefore permanent, or are
continuing manifestations of juvenility. Some citrus virus diseases also
affect fruit characters directly or indirectly, and it is useful to know the
range of expression of all these effects.
Earlier studies (1) on mature nucellar-line trees of several varieties
indicated that in most characters they were equal or superior to their
old-line parents. However, certain fruit character differences were still
detectable although the lines were then about 35 years old from seed.
By 1957 (2), data on young orchard trees repropagated from some of
these nucellar lines indicated that some differences were still being ex-
pressed. In the present paper we present data taken between 1957 and
1960 on young plantings of the Frost (nucellar) Marsh grapefruit and
Washington Navel orange, compared with old lines of these varieties.







SOOST and CAMERON


Since much California citrus goes to the fresh market, particular atten-
tion has been paid to physical characters of the fruit.

Materials and Methods
The data on the Washington Navel oranges are from plantings by
commercial growers in Tulare County, where this fruit is marketed from
mid-November through February or later. These plantings consisted of
one to a few acres of nucellar-line trees, usually with greater acreages of
old-line trees nearby. At the McMaster ranch, all trees were planted in
1951 on sweet orange rootstock. At the Chase ranch, nucellar-line trees
were planted in 1953 on Troyer citrange rootstock; unrelated old-line
trees were planted in 1948 on sour orange. The Johnston trees (all
nucellar-line) were planted in 1952 on trifoliate orange, Cleopatra
mandarin, and Troyer rootstocks. Except at McMaster's, old-line trees
were thus older from planting than trees of the nucellar line.
A sample (usually 20 fruits) was taken from each of 10 trees of each
line in each planting early in the season, and sometimes again late in the
season. Only healthy-appearing trees carrying relatively good crops were
sampled. The fruits were visually graded for physical characters while
spread out in groups so that several old-line and nucellar samples could
be viewed at once.
Most of the Marsh grapefruit data are from the Coachella Valley, a
hot desert area. The plantings varied from a few trees to several acres.
At the Price Ranch, nucellar-line trees were planted in 1954 on Cleo-
patra mandarin and Savage citrange rootstocks. The compared old-line
trees had been planted in 1932, apparently on sour orange stock. At the
Seaview ranch nucellar trees were planted in 1953 on Troyer rootstock,
while the old-line trees were perhaps 30 years of age, on undefined root-
stock. In the Scanlon and Brown plantings, all trees were planted in
1956 on rough lemon rootstock.
Ten fruits were usually taken from each tree of each line on 3 sampling
dates, for 3 seasons. With the young Scanlon and Brown trees, however,
only one sampling was obtained (in April, 1960). Fruit was graded in
the same manner as with the oranges.
Chi squares were calculated for all enumeration data. F values were
calculated for testing the significance of the differences of peel measure-
ments and juice percentage and composition data from the McMaster
ranch and the Scanlon ranch. Because of fewer trees sampled, analyses
were not made for these measurement data in the other plantings.








PROCEEDINGS of the IOCV


Navel Orange Comparisons

PHYSICAL CHARACTERS.-In the earliest crops of some nucellar-line
plantings, considerable numbers of fruits were tapered or rough at the
stem end. Data on these and other characters for three seasons at the
McMaster ranch and for the 1960 season at the Chase ranch are shown
in Table 1. Fruit size has been rather similar between old-line and

TABLE 1. PHYSICAL CHARACTERS OF NUCELLAR- AND OLD-LINE WASHINGTON
NAVEL ORANGES"
Percentage of fruits with Peel
Year thick-
Line" planted stem end Loose ness
tapered furrowed core (mm)
McMaster Ranch, 1958
Old 1951 11 1 5.4
Frost nucellar 1951 11 14** 5.8**
1959
Old 1951 1 56 0 5.5
Frost nucellar 1951 7** 27** 4 5.7
1960
Old 1951 7 25 2 5.5
Frost nucellar 1951 16** 6** 10** 5.4
Chase Ranch, 1960
Old 1948 9 17 7 6.3
Frost nucellar 1953 21** 20 3 6.1
"Data from 20 fruits of each of 10 trees of each line in each planting; November
picks at McMaster's, December picks at Chase's. Asterisks indicate the 1 per cent
level of significance of a difference of a value from the old-line value immediately
above it.
bMcMaster trees on sweet orange rootstock; old-line trees at Chase Ranch on
sour stock, nucellar-line on Troyer.


nucellar-line in most samplings, and can scarcely account for differences
in other characters. Fruit shape index (the ratio of transverse to longi-
tudinal diameter) has been nearly identical at any one location in any
one year, indicating that the nucellar-line fruits are not more elongate
than old-line fruits. Despite this there has been a higher percentage of
tapering stem ends in nucellar fruit at nearly every sampling. In some
comparisons (Table 1) the differences have been significant. The fruit
have had slightly rougher stem ends in most samplings. In contrast, fur-
rowing or grooving of the peel at the stem end has seemed to be nega-
tively correlated with tapering and roughness; that is, there has usually
been a greater percentage of old-line fruits with furrowing. At the







SOOST and CAMERON


McMaster planting the differences in furrowing have been significant.
Peel thickness has not been a problem in these nucellar-line plantings.
The McMaster data suggest a slight decrease in peel thickness over the
3-year period, and in the other plantings very little difference between
lines is evident. A greater tendency to loose cores has been evident in
nucellar fruit at McMaster's, with the differences being significant in
1958 and 1960. In two other plantings not shown in Table 1, nucellar-
line fruits in 1960 showed percentages of tapered or rough stem ends and
core looseness which were in the range of those at the Chase planting.
Differences in other characters which would affect the appearance of the
fruit have not been consistently found. The effects of location, season,
and age of tree on the expression of physical fruit characters have been
apparent.
Observations indicated that unfavorable characters in nucellar-line
fruits might be accentuated late in the season. Where fruit was available,
samples were therefore taken late in the season as well as early. In 1959,
stem-end taper, roughness, and core looseness were considerably greater
late in the season. There was also a considerable increase in rind thick-
ness at McMaster's in 1959. However, in 1960 there was little change in
the incidence of any of the characters except loose core, over the season.
Such changes are no doubt mainly due to continued growth of the peel
and consequent tendency of the peel and fruit segments to separate. The
incidence of unfavorable characters also increased in old-line fruit, but
remained lower than with the nucellar-line.
JUICE PERCENTAGE AND COMPOSITION.-Three years of data from the
McMaster planting show the nucellar line to be very similar to the
McMaster old line in percentage of juice and in total soluble solids. The
percentage of acid has been slightly higher in the nucellar line. In 1959,
the difference was just significant at the 5 per cent point; in 1958 and
1960 it was not significant. These higher acid values result in slightly
lower soluble solids:acid ratios, but the ratios have been well above the
required 8: 1 at the beginning of each marketing season. At the Johnston
planting nucellar-line fruits have always been higher in soluble solids and
acid than at McMaster's, and the ratios have always been 9:1 or higher
at the beginning of the season.

Marsh Grapefruit Comparisons
PHYSICAL CHARACTERS.-First crops of young trees of the Frost Marsh
grapefruit in Coachella Valley, like those of the Washington Navel







PROCEEDINGS of the IOCV


orange in Tulare County, have shown certain undesirable fruit charac-
ters. Beginning in 1958, data were taken from the Price and Seaview
plantings. Young trees of old-line grapefruit were not available in these
plantings, so that compared old-line trees are much older from planting
than the Frost nucellar trees. The nucellar-line trees have shown con-
sistently higher percentages of tapered and rough stem end fruit than the
older old-line trees. They have also shown more fruit with somewhat
thick peel at Price's, and the differences in taper and peel thickness have
usually been significant. The degree of expression of the characters has
definitely declined during the 3 years of sampling. Unlike the Washing-
ton Navel oranges, these nucellar-line grapefruits have shown lower
average fruit-shape indices, that is, less flattening. With light crops, they
have also often shown larger fruit size.
In the Scanlon and Brown plantings, where all trees were young, both
lines had generally greater percentages of fruit with poor physical
characters than did the older plantings. However, the nucellar-line trees
had higher percentages of such fruits than the old line. The differences
in incidence of core hollowness and thick peel were significant. The
nucellar-line trees also had lighter relative crops and larger fruit size.
One older Frost Marsh tree, planted in 1933 at the Citrus Experiment
Station at Riverside, has been studied for several years in comparison
with one old-line companion tree and some other old-line trees from
another bud source of Marsh. The Frost Marsh tree and its companion
are planted side by side, on sour rootstock, in a variety orchard. The trees
of the other old line, on sweet stock, are part of a long-time fertilizer
experiment, in treatments where favorable conditions have been main-
tained. The physical characters of the fruit of the Frost Marsh and its
companion tree have been very similar, but the characters of the fer-
tilizer-experiment trees have been somewhat better. At present, it is not
known whether the difference is due to environment or to a genetic
difference in the Frost Marsh.

JUICE PERCENTAGE AND COMPOSITION.-Fruit composition tests have
been made for several years from the grapefruit trees discussed above.
The older trees, at Riverside, have not shown consistent differences in
percentage of juice or total soluble solids. Acidity, however, has been
slightly lower in the Frost Marsh tree in most assays. In the Coachella
Valley, from 1958 to 1960, the percentages of juice and soluble solids in
the nucellar line have improved as the trees gained age. Percentages of
acid have regularly been lower than in the older old-line trees. At the







SOOST and CAMERON


Price and Seaview Ranches, old-line and nucellar fruit were rather
similar in percentages of juice and soluble solids; acids were slightly
lower in the nucellar line. At the Scanlon and Brown ranches, where all
trees were only 4 years from planting, fruit composition characters were
poor in both lines. Soluble solids in the nucellar line were equal to those
in the old line, and acid was slightly lower in the nucellar line. Per-
centage of juice was also lower, but this is probably due to the lighter
crops of larger-size fruit. Analyses of the data from the Scanlon Ranch
indicate that the differences in percentage of juice, percentage of acid,
and soluble solids: acid ratio are significant at the 1 per cent level.

Discussion
The reasons for the observed differences in fruit characters are not
entirely clear. Because of greater vigor, it can be expected that newly
repropagated trees of either old or young lines will produce fruit in-
ferior to that of older trees. This undoubtedly accounts for some of the
differences found. However, the higher numbers of these fruits from
nucellar-line trees when both lines are of the same age from recent
planting suggests that effects of nucellar juvenility are still operating.
The facts that two different citrus varieties exhibit these tendencies, and
that physical characters seem to be improving with tree age and heavier
crops, fit this supposition. Older trees of these lines at Riverside have
differed little from compared old lines in these characters. Although the
two nucellar lines had not been often repropagated until about 1950,
it is surprising that such a degree of juvenility could still be discernible.
Unfavorable fruit characters have been more prominent in the grape-
fruit than in the navel orange. This has been correlated with extremely
vigorous growth of the young grapefruit trees, and with slowness to come
into heavy bearing.
Genetic differences may yet be found in either nucellar line. The
slightly lower acid in the nucellar Marsh is suggestive of such a differ-
ence.
The contribution of the several rootstocks to differences between lines
cannot be adequately evaluated. However, the vigorous growth of the
nucellar lines on Troyer appears to have accentuated adverse fruit
characters. Since differences were present even when the rootstock was
the same for both lines, rootstock does not appear to be the primary
factor involved.
Symptoms suggestive of stubborn disease have recently been observed








PROCEEDINGS of the IOCV


in some Frost Marsh trees in California. This disease has not been proved
absent in the trees reported here, but they have been among the most
vigorous in the plantings and the fruit characters discussed are not ones
presently associated with stubborn disease.
The adverse characters noted in the 2 varieties have not been promi-
nent in recent plantings of nucellar Valencia oranges and lemons.
Similarly, nucellar Red Blush grapefruit lines in California do not seem
to show these characters. Cooper et al. (3) state that in Texas the fruit
size, shape, and quality of 8-year-old seedlings of Webb Red Blush grape-
fruit are essentially as good as in budded old-line trees.
More data are needed on the performance of nucellar lines of all the
major varieties. Data are being taken at Riverside on a 1954 planting of
additional Washington Navel lines originated by Dr. H. B. Frost, and on
seedlings from the same lines. Dr. W. P. Bitters is also establishing
nucellar lines of many of the better-known strains of Washington Navel.
ACKNOWLEDGMENTS.-The authors gratefully acknowledge the as-
sistance of Donald Cole, Jr., and R. H. Burnett, Laboratory Technicians
in the Citrus Experiment Station, in collecting the data; and the co-
operation of D. D. Halsey and K. W. Opitz, University of California
Agricultural Extension Service.


Literature Cited

1. CAMERON, J. W., and R. K. SoOST. 1952. Size, yield and fruit characters of
orchard trees of citrus propagated from young nucellar-seedling lines and
parental old lines. Proc. Am. Soc. Hort. Sci. 60:255-264.
2. CAMERON, J. W., R. K. SOOST, and H. B. FROST. 1959. The horticultural
significance of nucellar embryony in citrus, p. 191-196. In J. M. Wallace
[ed.], Citrus Virus Diseases. Univ. Calif. Div. Agr. Sci., Berkeley.
3. COOPER, W. C., E. O. OLSON, N. MAXWELL, and A. V. SHULL. 1958. Nursery
and orchard performance of nucellar seedling clones of citrus in the Rio
Grande Valley of Texas. J. Rio Grande Valley Hort. Soc. 12:44-52.
4. WEATHERS, L. G., and E. C. CALAVAN. 1959. Nucellar embryony-a means of
freeing citrus clones of viruses, p. 197-202. In J. M. Wallace [ed.], Citrus
Virus Diseases. Univ. Calif. Div. Agr. Sci., Berkeley.











HENRY SCHNEIDER, J. W. CAMERON, R. K. SOOST,
and E. C. CALAVAN



Classifying Certain Diseases as Inherited








RESEARCH WORKERS tend to group apparently-inherited disorders
with virus diseases, under such categories as virus-like, nontransmissible,
and noninfectious. Apparently-inherited disorders often have many of
the characteristics of virus diseases, but their inclusion with virus diseases
does little to stimulate further study of them. Plant pathology textbooks
classify toxicities and nutritional disorders, some of which are virus-like,
as distinct diseases, but apparently-inherited disorders usually are not
mentioned.
In several tree crops, apparently-inherited disorders are of major im-
portance. For example, much of the decline of maturing lemon trees that
has plagued the lemon industry in California from its beginning is due to
lemon sieve-tube necrosis and to sour orange rootstock necrosis. Dr.
George Nyland (personal communication) states that in California
sweet cherry crinkle leaf (5) and sweet cherry deep suture (9) equal or
surpass the cherry virus diseases in causing crop reduction in older Bing
and Tartarian cherry trees.
The purpose of this paper is to present criteria for judging whether
a disorder is inherited; to describe the characteristics and behavior of
some of the inherited and probably-inherited disorders in citrus and
other plants; and to discuss control measures for them.


Criteria Indicating Inheritance
DIRECT EVIDENCE.-Mendelian segregation of a character is strong
evidence that the character is inherited. Characters controlled by single
15







PROCEEDINGS of the IOCV


genes show simple inheritance patterns. Examples are bush-type growth
versus vine-type in beans, and sugary versus starchy endosperm in corn.
Simple inheritance patterns do not however distinguish between the di-
rect inheritance of a disorder and inheritance of susceptibility to some
unknown inciting agent. The inheritance of susceptibility is well illus-
trated in wheat, where many varieties differ in resistance to rust or bunt
fungi, and where the fungus inciting the symptoms can be easily detected.
On the other hand, if an undemonstrable virus is responsible, it may not
be clear whether susceptibility to the virus or direct inheritance of a
disorder is involved.
When several or many genes control a heritable character, proof of
inheritance is difficult. For example, the mode of inheritance of trifoliate
versus unifoliate leaves in hybrids of Citrus x Poncirus is not certain, even
though the 3-leaf condition is regularly dominant in F, hybrids. Popu-
lations of F, plants from open pollination show far more 3-leaved
zygotic plants than would be expected from a single, dominant gene.

INDIRECT EVIDENCE.-It is difficult to use Mendelian segregation to
prove inheritance of citrus disorders. Many zygotic seedlings do not
survive, and populations for study are incomplete. The occurrence of
nucellar seedlings further complicates the problem. The time and land
required to raise sufficient zygotic individuals of several successive gen-
erations to an age where most disorders become apparent is excessive.
Therefore, classifying disorders as inherited usually becomes a matter of
determining whether affected plants express characteristics and behavior
of inherited disorders, and a matter of eliminating other possible causes.
An inherited disorder should normally be expected to exhibit the fol-
lowing characteristics: it should be limited to certain varieties or clones
of a crop, in some cases being traceable to a single source plant; it should
not be transmissible from affected to normal plants either by propagative
parts or by external agents; and it should not spread in the field. In citrus,
all nucellar lines from affected clones should show the disorder. Histo-
chemical tests for viruses should be negative. The disorder should appear
on many soil types and under many cultural and climatic conditions,
although it might not develop under certain climatic conditions.
In the aggregate, these characteristics point strongly toward inherit-
ance, but they do not completely eliminate the possibility that a virus is
involved. If a virus were present in all members of a variety and if all
other varieties were tolerant or immune, evidence of transmission would
be difficult to obtain. A virus disease can fail to spread in the field due







SCHNEIDER, CAMERON, SOOST, and CALAVAN


to the lack of a vector. Viruses in plants are not known to be 100 per cent
seed transmitted; if, however, 100 per cent transmission did occur, the
disease, like an inherited disorder, would be present in all nucellar clones.
A disorder due to a virus might also falsely appear to be inherited if all
nucellar seedlings should immediately become infected.
Malnutrition or injuries due either to deficiencies or excesses of various
elements and compounds can cause symptoms resembling those of in-
herited disorders. If the disease occurs in orchards on different soil types,
and in orchards irrigated with water of various salt contents, and if other
varieties and species of the crop growing nearby do not show symptoms,
the possibility of faulty nutrition is greatly reduced.
Parasitic organisms, such as bacteria, fungi, and nematodes, can
usually be eliminated as causal factors if none are found associated with
early symptoms of the disease. There remains the possibility that or-
ganisms on one part of the plant could indirectly induce what appear to
be primary symptoms in another part.


Examples of Inherited Disorders of Plants
There are numerous examples of simple inheritance of disorders that
are virus-like. In tomato, the single recessive gene, sun dwarf, produces
a stunting and corking of the internodes when the plants are grown in
light of high intensity (8). Another recessive gene in tomato, curly
mottled, causes a curling and mottling of the leaves that is very similar to
symptoms caused by tobacco mosaic virus (8). Symptoms are produced
only under certain environmental conditions. A disorder of maize being
investigated by J. W. Cameron is genetically controlled, yet has some
virus-like characters. In one line of sweet corn, the leaves develop numer-
ous chlorotic spots early in the life of the plant. The spots enlarge longi-
tudinally, and necrosis follows until a third or more of the leaf surface
dies. Other varieties of sweet corn, grown adjacent to this line, do not
develop these spots. When the affected line is crossed with other varieties,
the F, hybrids never develop the spotting, but F2 populations from self-
ing segregate in a ratio of 3 normals to 1 spotted, indicating that the
character is recessive.
In woody plants, genetic information is often incomplete, but there are
examples of disorders which are probably inherited. Noninfectious bud
failure in almonds occurs in the varieties Peerless and Nonpareil. This
disorder also occurs in Jordanola, a hybrid between Nonpareil and a
nonaffected variety. Seedlings from Jordanola x Ne Plus Ultra, another







PROCEEDINGS of the IOCV


unaffected variety, also carry the disorder. Attempts to transmit the
disease have failed (15). It seems likely that the genes controlling bud
failure are passed from variety to variety.
Sweet cherry crinkle leaf occurs in some trees of the Bing and Black
Tartarian varieties, and transmission has not been obtained (5). Symp-
toms occur in scattered portions of trees as if frequent somatic mutations
were responsible. The incidence of crinkle leaf shoots is frequent in old
wood of trees low in vigor. Some hybrid seedlings have developed crinkle
leaf when one parent was affected and one was normal.

Some Apparently Inherited Disorders of Citrus
Apparently-inherited disorders of citrus may be divided into two main
types: those which are independent of rootstock-scion interaction, and
those which are not. The former includes lemon sieve-tube necrosis and
wood pocket, and the latter includes various incompatibilities.
LEMON SIEVE-TUBE NECROSIS.-This disorder of the scion portion of
the lemon tree trunk is one of several diseases having decline as an ulti-
mate symptom. It affects all Eureka lemon trees as well as some strains
of Lisbon and Villafranca (12). A characteristic symptom is necrosis of
older sieve tubes, which may begin when trees are 3 or 4 years old; after
several more years, necrosis may increase and involve even the youngest
sieve tubes. As a result of the blockage of translocation, reserve starch is
used from the xylem; unless new phloem forms, the feeder roots die.
Entire blocks of trees in apparent good health may suddenly decline or
wilt from this disorder.
Lemon sieve-tube necrosis has many of the distinguishing characteris-
tics of inherited disorders. Only certain clones are affected. It occurs in
all Eureka trees regardless of rootstock, and is found throughout the
diverse environmental conditions of southern California's coastal and
semicoastal areas. Most of the trees in a planting are affected simul-
taneously. One Lisbon lemon strain and 10 Eureka lemon clones derived
from nucellar seedlings have been examined; all of them, like the clone
from which derived, are affected by lemon sieve-tube necrosis. Symptoms
have not been produced in closely related disease-free clones by budding
and grafting, nor has it been possible to demonstrate a consistent rela-
tion of any known virus to this disorder (2, 14). Strains of lemons that
are consistently free of sieve-tube necrosis do not develop it when used
as rootstocks for affected clones; this indicates nontransmissibility. There
is no evidence that soil fungi, nematodes, or soil fertility are involved.







SCHNEIDER, CAMERON, SOOST, and CALAVAN

WooD POCKET.-This is a disorder which affects a few strains of
Lisbon lemons and the Tahiti lime (1, 7). Chlorotic blotching occurs
on one or both sides of the midrib of the leaves, and pockets of dead bark
and wood appear on the limbs and trunks. Warm interior climates
favor symptom production. Although this disorder was thought to be a
virus disease when first described (4), it is now believed to be an in-
herited disorder for the following reasons. It occurs in both zygotic and
nucellar seedlings; although present for 41 years in diseased lemon tops,
it has not moved down into healthy lemon interstocks; efforts to transmit
the disease by grafting to lemons during a 9-year period have failed (1);
indexing for known virus diseases has shown no one virus to be con-
sistently present in affected trees; and in lemons the disease is clonally
limited. The variable pattern of symptoms suggests that wood pocket
may be due to an unstable gene or a chimeral condition (1).
OTHER DISORDERS.-Disorders which may be inherited are: crinkle-
scurf of Valencia orange (6); chimeric breakdown of Tahiti lime fruits
(7); and a seed-perpetuated disorder of trifoliate orange with symptoms
resembling those of exocortis (3).
INCOMPATIBILITIES.-When Eureka lemons and certain Lisbon lemon
clones are grown on sour orange rootstocks, the trees go through cycles
of decline and recovery, beginning at about 12 to 15 years of age. The
decline is due to necrosis of sieve tubes immediately below the bud
union; hence the name sour orange rootstock necrosis (10, 11). After 8
years, symptoms of this disease were not transmitted by grafts from
affected lemon strains to strains compatible with sour orange rootstocks.
Frost nucellar Eureka trees are affected but certain Lisbon lemon strains
are consistently free of the disease. The disease occurs in many areas,
and affected strains decline when growing next to nonaffected strains.
Experimental and commercial attempts to grow Eureka lemons on
the trifoliate orange and on Troyer citrange (sweet orange x trifoliate)
have failed because of a degeneration of tissues at the bud union. Eureka
trees on Troyer rootstock decline when 3 or more years old (13). Other
lemon varieties incompatible with Troyer are Genoa and 2 varieties
developed from crossing a hybrid of Genoa with an unnamed clone of
Lisbon lemon. Sweet orange is a compatible rootstock for lemons, so it
is apparently the trifoliate orange that carries an incompatibility factor.
Orlando and Minneola tangelos (tangerine x grapefruit) are ap-
parently compatible with Troyer citrange. When these varieties were
crossed with a grapefruit hybrid called Sukega by R. K. Soost, progeny








PROCEEDINGS of the IOCV


were obtained which were not compatible with Troyer. Some hybrids
obtained from crosses of the Frua tangerine with Sukega and of the
Sunrise tangelo with Sukega were also incompatible with Troyer. Al-
though it is not known whether Frua and Sunrise are compatible with
Troyer it appears that the Sukega carries germ plasm for incompatibility.

Discussion
Classifying apparently-inherited disorders with virus diseases has
been a convenient way of handling them, but such classification has not
always motivated satisfactory investigations. Some citrus diseases have
not been transmitted nor has it been possible to prove them to be in-
herited, but this should not prevent them from being placed in the
category where they most likely belong. Classification of diseases has
not always awaited procedures for proving their cause. The existence of
citrus virus diseases is accepted, but Koch's postulates have not been
carried out. Although graft transmission proves that an infectious agent
is present, it gives no evidence of the nature of the agent. Classifying
disorders as inherited should be based upon the disorders having char-
acteristics like those of known inherited disorders. Likewise, evidence
for virus is largely a matter of the disease having characteristics of
known virus diseases. These are similarity of symptoms of the disease to
other virus diseases, the manner in which the disease spreads, transmis-
sion by an insect vector, absence of parasitic organisms, and histo-
chemical evidence for virus in affected tissue. Recognition that a disease
is probably caused by virus stimulates appropriate investigation and
control measures, and recognition that a disorder is probably inherited
would direct more attention to its investigation and control.
Certain methods used to control virus diseases can be applied to in-
herited disorders. For example, affected scion varieties, rootstocks, or
scion-rootstock combinations can be avoided. In a cherry budwood cer-
tification program, stone fruit virologists are eliminating some ap-
parently-inherited disorders that arise spontaneously. If all members of
a variety are affected, and if there is no satisfactory substitute variety,
this approach is not possible. Cultural practices, such as pruning, may
be used at times to alleviate the effects of a disorder, or varieties may
be selectively grown in environments where the disorder does not occur.
Anatomical studies can sometimes detect the presence of slowly de-
veloping disorders many years before they become limiting to the health
of the tree, thus aiding in the selection of new varieties and scion-root-
stock combinations by early elimination of undesirable ones. Breeding








SCHNEIDER, CAMERON, SOOST, and CALAVAN


may sometimes eliminate inherited disorders, although with tree crops
this is a slow and tedious process.
Additional techniques should be sought to alleviate inherited dis-
orders, but this might require more knowledge about the physiological
and biochemical effects of the disorders. Perhaps the application of
some compound, which the plant cannot synthesize, would enable it to
grow normally; or some chemical might be used to neutralize a toxic
substance.
Literature Cited
1. CALAVAN, E. C. 1957. Wood pocket disease of lemons and seedless limes.
Calif. Citrograph 42: 265-268, 300-304.
2. CALAVAN, E. C., and L. G. WEATHERS. 1959. Transmission of a growth-
retarding factor in Eureka Lemon trees, p. 167-177. In J. M. Wallace
[ed.], Citrus Virus Diseases. Univ. Calif. Div. Agr. Sci., Berkeley.
3. CALAVAN, E. C., R. K. SOOST, and J. W. CAMERON. 1959. Exocortis-like
symptoms on unbudded seedlings and rootstocks of Poncirus trifoliata with
seedling-line tops, and probable spread of exocortis in a nursery. Plant
Disease Reptr. 43: 374-379.
4. FAWCETT, H. S., and E. C. CALAVAN. 1947. Wood pocket, a newly reported
disease of lemons. (Abstr.) Phytopathology 37: 843.
5. KINMAN, C. F., W. B. HEWITT, T. B. LOTT, G. L. PHILIP, E. L. REEVES,
R. S. WILLISON, and S. M. ZELLER. 1951. Sweet Cherry Crinkle Leaf,
pp. 195-200. In Virus diseases and other disorders with virus-like symp-
toms of stone fruits in North America. U.S. Dept. Agr. Handbook 10.
Illus.
6. KNORR, L. C. 1953. Transmission trials with crinkle-scurf of citrus. Plant
Disease Reptr. 37: 503-507.
7. KNORR, L. C., and J. F. L. CHILDS. 1957. Occurrence of wood pocket
(blotch), chimeric breakdown, and endoxerosis in Florida, with particular
reference to the Tahiti lime. Proc. Florida State Hort. Soc. 70: 75-81.
8. RICK, C. M., and L. BUTLER. 1956. Cytogenetics of the tomato. Advances in
Genetics 8: 267-381.
9. REEVES, E. L., E. C. BLODGETT, C. F. KINMAN, T. B. LOTT, J. A. MILBRATH,
and S. M. ZELLER. 1951. Sweet cherry deep suture, p. 201-204. In Virus
diseases and other disorders with virus-like symptoms of stone fruits in
North America. U.S. Dept. Agr. Handbook 10.
10. SCHNEIDER, H. 1952. Bud union problems of lemon trees on sour orange
rootstock. Calif. Citrograph 37(5): 208-212.
11. SCHNEIDER, H. 1956. Decline of lemon trees on sour orange rootstock. Calif.
Citrograph 41(3): 117-120.
12. SCHNEIDER, H. 1960. Sieve-tube necrosis in nucellar lemon trees. Calif. Citro-
graph 45(7): 208, 219-222.
13. WEATHERS, L. G., E. C. CALAVAN, J. M. WALLACE, and D. W. CHRISTIAN-
SEN. 1955. A bud-union and rootstock disorder of Troyer citrange with
Eureka lemon tops. Plant Disease Reptr. 39(9): 665-669.
14. WEATHERS, L. G., and E. C. CALAVAN. 1959. Nucellar embryony-a means
of freeing citrus clones of viruses, p. 197-201. In J. M. Wallace [ed.], Citrus
Virus Diseases. Univ. Calif. Div. Agr. Sci., Berkeley.
15. WILSON, E. E., and R. D. SCHEIN. 1956. The nature and development of
noninfectious bud failure of almonds. Hilgardia 24: 519-542.











HARRY C. BURNETT


The Color Test for Exocortis Indexing in Florida








THE FLORIDA Budwood Registration Program was established in 1952
to enable growers to obtain a source of citrus budwood free from several
serious virus diseases known to be present in Florida. Citrus trees become
registered when they have been screened for tristeza, psorosis, xyloporo-
sis, and exocortis. Indexing for tristeza can be completed within 6
months; for psorosis, 2 years are required; for xyloporosis, 4 years; but
for exocortis, 8 years.
Since the period required for accurate detection of exocortis by
nursery indexing methods only was so prolonged, a color test was de-
veloped by Childs, Norman, and Eichhorn (1) and is now a part of
the indexing program. It is used in conjunction with other tests for
exocortis in Poncirus trifoliata in the Division of Plant Industry.


Method of Using the Color Test in the Florida Program
The steps involved in screening trees for exocortis in the Florida
Budwood Registration Program are as follows:
1. Buds from each candidate tree in the Budwood Registration Pro-
gram are inserted in two Poncirus trifoliata seedlings. Both seedlings are
given the same accession number, but one is designated as W (west)
tree and the other as E (east) tree. The color test for exocortis is made
on seedlings budded at least two years previously in the Florida Program.
2. A piece of bark approximately 3/4-inch long by 1/4-inch wide is
cut from the tree. If one of the seedlings is not sufficiently large in
diameter that the bark sample can be taken without endangering the







BURNETT


life of the tree, the bark specimen is taken only from the larger of the
two trees. Childs et-al. (1) suggest taking these samples at the bud union.
However, Sinclair and Brown (3) have reported quite recently that
scaling due to exocortis first appears on the side of the large roots near
the soil line and spreads during the next 4 to 6 years over the entire
rootstock, up to the bud union. Since publication of their paper, our
bark samples have been taken below the bud union, near the soil line.
As yet there is little evidence that any one point between the soil line
and the bud union is to be preferred more than another. Positive read-
ings have been made from all areas within the above limits. Perhaps the
chief advantage in taking the bark sample near the ground level is that
there is more likelihood of getting only Poncirus trifoliata bark and
none of the scion top, since it is often difficult to determine the location
of the bud union on young budded trees.
Immediately after excision, the bark samples are placed in a formalde-
hyde-acetic acid-alcohol solution, FAA No. 2 (2), for killing and
fixation.
3. Transverse sections of bark samples about 20 to 30 microns thick
are made on the sliding microtome. Ten sections are placed on a
microslide.
4. A few drops of phloroglucinol-HCI reagent are added to the bark
sections and a cover glass is eased into place. The staining process is
complete within 3 or 4 minutes and results remain substantially un-
changed for 4 hours. The phloem ray cells of the bark are the chief
objects of study. The phloroglucinol-HCl reagent produces a red color
reaction in some to many of the ray cells in Poncirus trifoliata bark if
exocortis infection is present.

An arbitrary system of evaluating the sections was arrived at after
consultation with Dr. J. F. L. Childs. When 3 or more cells are stained
red in 10 sections, a positive diagnosis is given. When 2 are found, the
diagnosis is called "probably positive" or "positive?". A "probably nega-
tive" designation is given when only one cell is stained in 10 sections,
and, of course, a negative determination is given when no cells are
found stained red.


Results Obtained with the Color Test
As of September 15, 1960, a total of 570 samples involving 322 can-
didate trees has been examined for the presence of exocortis. Of these,







PROCEEDINGS of the IOCV


150 were diagnosed as positive, 45 as probably positive, 333 as negative,
and 42 as probably negative.
All except 6 of the trees were budded between April, 1955, and June,
1956. The 6 exceptions were budded in 1957. Final field determinations
cannot be made until April, 1963, and June, 1964, for most of the trees
under study. However, a correlation of the results obtained with the
color test with the corresponding plants in the test plots is as follows:
Of the plants that were diagnosed as negative or probably negative
325 are still symptomless in the test plots; 98 plants that were diagnosed
as positive or probably positive are now showing bark scaling in the test
plots; and 97 read as positive are still symptomless in the nursery test
plots. In 22 of the latter cases, the corresponding west or east tree is
showing exocortis scaling symptoms. Color test results showed most of
these to be positive 3 or 4 years after they were budded, yet after 4 or 5
years they still are without visible bark symptoms in the test plots.
There are 50 cases where a negative diagnosis by the color test was
given, and the plants later showed positive bark symptoms in the nursery.
The plants in question had been budded 3 or 4 years, and in most cases
the field symptoms did not show until a year after the use of the color
test. Tests were rerun on 17 of these 50 trees soon after field symptoms
were noted. All 17 gave a positive reaction when retested. For 10 of
these 50 trees, the color test gave a positive reaction for the adjoining
east or west tree approximately one year before the field symptoms were
noted.
Childs et al. (1) report that (a) the reaction to phloroglucinol is not
exhibited uniformly in all the rays, (b) in a given specimen, particularly
from a young tree, there may be only a few rays with reactive cells, and
(c) there is some chance that reactive ray cells will not be present in a
given bark specimen. It seems probable that, in the case of these 50
trees, if another sample had been taken, either at the same time or a
few months later, we might have secured a positive test.
As pointed out earlier in this paper, the limiting factor for resampling
a given tree is the danger of girdling too large a part of the trunk. How-
ever, since it was found that a color reaction may be obtained at any
point from the soil line to just below the bud union, we have a much
larger area from which we may secure our bark samples for testing.
Therefore, if bark samples could be taken in the summer of the third
year after budding, and again in the spring and summer of the fourth
year, the chances for correct diagnosis, both negative and positive, would
be greatly increased.







BURNETT


This, of course, can be confirmed only by waiting until 1963 and 1964,
the end of the 8-year test period. If the results obtained from the appli-
cation of the color test still correspond to the symptoms in their coun-
terparts in the test plots, then we will know that indexing candidate
citrus trees for exocortis can be completed by use of the color test within
4 years from the date of the original budding in Poncirus trifoliata.


Literature Cited

1. CHILDS, J. F. L., G. G. NORMAN, and J. L. EICHHORN. 1958. A color test for
exocortis infection in Poncirus trifoliata. Phytopathology 48:426-432.
2. RAWLINGS, T. E. 1933. Phytopathological and botanical research methods.
John Wiley & Sons, Inc., New York. 156 p.
3. SINCLAIR, J. B., and R. T. BROWN. 1960. Effect of exocortis disease on four
citrus rootstocks. Plant Disease Reptr. 44:180-183.










E. C. CALAVAN and L. G. WEATHERS


Evidence for Strain Differences and Stunting
with Exocortis Virus








RANGPUR LIME DISEASE, described by Olson (8) in 1952, is generally
believed to be caused by the same virus, or virus complex, as that which
causes exocortis (7, 10, 11). Therefore the name exocortis is used here
irrespective of host variety.
Reports from various countries have indicated that exocortis virus
causes stunting (1, 6, 7, 9, 12). Either exocortis virus from some sources
evidently causes more stunting than that from other sources or some
trifoliate orange (Poncirus trifoliata (L.) Raf.) and Rangpur lime
(Citrus limonia Osbeck) rootstocks are less stunted by exocortis than
others (1, 5).
This paper reports effects of exocortis virus upon the growth of 16
stionic combinations of citrus and presents evidence for the existence of
strains of exocortis virus.


Materials and Methods
Experimental trees were propagated by grafting buds of parent trees
free of exocortis virus onto seedling rootstocks; some were inoculated
and others maintained as controls. Inoculations were made by grafting
2 or more buds from exocortis-infected trees (exocortis sources) into an
experimental tree, care being taken that each tree received buds from
only one source; exceptions were a few trees in Plot 1, which received
buds from more than one source. Growth from buds used for inoculation
was suppressed.
26







CALAVAN and WEATHERS


Plot 1, described previously (2), was set out in 1948 near Oxnard.
Plot 2, planted near Ventura in 1953, contains 3 6-tree blocks. Three
trees of each block were inoculated in 1955, the others were left as
controls.
Plot 3 contains 66 trees planted in two rows near Ventura in 1954.
Alternate trees in almost 3/4 of the plot were inoculated in 1955. Alter-
nate trees in the other portion were grafted with buds from trees
presumed to be free of viruses. The other half of the trees in the plot
were left as controls.
Plot 4 consists of greenhouse-grown plants set out in 1956 in River-
side, with 24' x 10' spacing. Half of the plants had been inoculated in
1955; some of the controls, following propagation, had been grafted
with buds from trees presumably free of viruses. Growth from buds
inserted after propagation was suppressed.
Plot 5 contains trees propagated in 1957 and planted in 1958 in
Riverside, with 10' x 12' spacing. Each of the 2 varieties used as tops in
this plot had been grown from a single parent tree. Half of the trees
were inoculated in 1957; the remaining noninoculated trees were kept
as controls.

Parent trees used as sources of buds for propagating the stionic com-
binations, those used as sources of buds for inoculations, and those used
as sources for postpropagative budding of controls were indexed for
viruses during the past 3 to 7 years. Experimental trees found to have
been inoculated with buds from trees with mixed infections are omitted
from Table 1; exceptions are trees inoculated with buds from one
exocortis source used in Plot 1 (2) and some trees inoculated with buds
from sources with both exocortis and vein enation viruses.
The parent trees used as budwood sources for propagating the stionic
combinations have proved to be free of detectable viruses, excepting the
young-line navel orange (C. sinensis (L.) Osbeck) which carried vein
enation virus in many buds used for propagation.
The exocortis sources used-1 Clementine mandarin (C. reticulata
Blanco), 9 Lisbon lemon (C. limon (L.) Burm.), 30 Eureka lemon-
were indexed for viruses of exocortis, psorosis, tristeza, vein enation, and
yellow vein. All 40 sources were found to contain exocortis virus; one
source contained psorosis and cachexia viruses; 10 other sources had
vein enation virus; 29 sources apparently carried only exocortis virus.
The sources were also indexed for xyloporosis virus, using sweet lime (C.
limettioides Tanaka) as an indicator, and for cachexia virus, using











p-. *'`'or- cor'-ommo es-es oi c c

F a * "*


0 z

I ** *4 -
o 0
ol)

o r cc c n- U. o o -

0 0 a .
"og e > aS

















H a- a-- - 0


o ^ *
i*** ***





















ao `0
0
E c0 ) cc kn -n tn Ln M cc c o r, c. cc I
c e u -? ? ? ......I u 0


0 "1 .- cu





0 M 0 50 -
un L c g i n c t7> o cc cc r j ^ D co Ct^ o n i '-* C m





>
o | 'IF I1 P 1- S 5 0












0-------No-lnooo~------------------------,NC 0J 0C
0 - .











-0 o :r-, -0



z o 2 oj c > n c ,



uZ a) n 0 ^ [ Lno i1 n cmn O kn u- i L C n c Ln n n n b| .S -a 3
0aaa a crr







CALAVAN and WEATHERS


Orlando tangelo (C. reticulata x C. paradisi Macf.) as an indicator.
None of the sources has been found to carry xyloporosis virus, although
insufficient time has elapsed to rule out its presence in more than 6 of
the sources. Thirty-nine of the 40 sources are thus far negative for
cachexia virus.
Vein enation virus has spread by vectors to some extent in all plots;
tristeza virus has spread by vectors to some trees in Plots 4 and 5 only.
Most of the spread occurred in 1959-60 and involved less than half of
the experimental trees, as judged by random indexing.


Results
STUNTING.-Stunting was sometimes observed in inoculated indicator
trees before external symptoms developed but sometimes could not be
detected until bark symptoms were well developed. Most of the inocu-
lated trees became stunted 1 /2 to 5 years after inoculation. The degree
of stunting of trunks may be judged by consideration of the data in
Table 1. Stunting of tops having exocortis-indicator rootstocks (Fig. 1)
usually was more severe than indicated by cross-section areas of trunks.


FIGURE 1. A. Noninoculated control tree of Valencia orange/Trifoliate
orange. B. Inoculated tree of same combination stunted by exocortis virus. Photo-
graphed 30 months after planting.







PROCEEDINGS of the IOCV


Budling controls having postpropagative grafts from trees free of
exocortis virus were not stunted.
Stunting of the same order of magnitude as that illustrated in Figure
1 occurred in the tops of inoculated navel orange and grapefruit (C.
paradisi Macf.) on trifoliate orange stocks; the degree of stunting was
slightly less for grapefruit on Rangpur lime stocks, much less for navel
orange on Rangpur lime stocks.

STRAINS OF EXOCORTIS VIRUs.-Indicator trees inoculated with buds
from some exocortis sources gummed and developed bark cracking and
shelling more quickly and severely than trees inoculated with buds from
other sources. Since the time required for symptoms to develop, as well
as the severity of these symptoms, was consistent for each exocortis
source, it is concluded that different sources carry different strains of
exocortis virus. A strain carried by the old-line CES Eureka lemon is


FIGURE 2. Comparative reactions of Morton citrange stocks (under calamon-
din) to different strains of exocortis virus. A. Severe exocortis on tree inoculated
with severe strain. B. Mild exocortis (arrow)just below original soil line on tree
inoculated from mild strain. Photographed 5 years after inoculation, in Plot 4.







CALAVAN and WEATHERS


especially virulent and has a shorter incubation period in most exocortis
indicators, 2 years or less (15), than the strains carried by many other
sources (Fig. 2); some strains of exocortis virus have an incubation
period of 4 years or more. A strain of virus carried by the CES Eureka
lemon causes more stunting and more seasonal chlorosis of trees having
sensitive rootstocks than some other strains studied. Similar results were
obtained in several field plots with trees having Cuban shaddock (a
lemon hybrid), Morton citrange (P. trifoliata x C. sinensis), Palestine
sweet lime, Rangpur lime, or trifoliate orange rootstocks.


Discussion
The early reports of stunting by exocortis were based mostly on ob-
servations in rootstock plots and commercial orchards. Some recent
reports compare infected trees with trees containing no exocortis virus;
others are based on observations of trees having mixtures of viruses, or
on trees not indexed for viruses. An association between exocortis and
stunting of some trees having indicator rootstocks already has been
clearly established, but it has not been previously determined whether
exocortis virus alone caused the stunting. Even those experiments based
on inoculated trees and comparable controls apparently were done
without indexing to determine whether other detectable viruses were
present in the experimental trees.
Measurable stunting in most of the inoculated trees in our experiments
is attributed to exocortis and to no other virus. The influence of viruses
other than exocortis on the stunting indicated by data in Table 1 is
assumed to have been small for the following reasons: less than 1/3 of
the exocortis sources used contained any other detectable virus, as
judged by the results of indexing (14); few calamondin (C. mitis
Blanco), lemon, and grapefruit (C. paradisi Macf.) trees became in-
fected with vein enation virus, none with tristeza virus; fewer than half
the Valencia orange trees acquired vein enation virus and less than 1/5
acquired tristeza virus; no effect of tristeza or vein enation viruses on
the growth of the experimental trees could be detected.
Exocortis lesions and associated gum may directly affect the functions
of the vascular system and reduce growth. Stunting, however, is some-
times evident before exfoliation occurs (4), and exocortis often can be
detected before shelling appears (3). We have observed milky or water-
soaked plaques of tissue developing near the cambium in the bark of
exocortis indicator stocks before the bark cracks and exfoliates. No







PROCEEDINGS of the IOCV


specific symptoms of exocortis developed in 3 nonindicator stocks
(grapefruit, sour orange, and sweet orange) having Eureka lemon tops
stunted by exocortis virus, nor did external symptoms develop in most
of the stunted trees of Lisbon lemon on Troyer citrange stock. It is as-
sumed, therefore, that in many nonindicator plants exocortis virus causes
inconspicuous internal changes which lead to stunting without specific
symptoms or any severe effect.
Fraser and Levitt (4) suggested that exocortis virus exists in a num-
ber of strains. They noted variations in degree of stunting, location and
amount of bark scaling, and in rate of external symptom development
which might be related to differences in strains or mixtures of virus.
Although the same kinds of variations occur in California, only those
involving conspicuous differences in incubation period (15) and severity
of symptom development in genetically uniform indicator plants grown
in similar environments have proved to be due to differences in virus
strains. Other factors also affect exocortis symptom development. The
influence of nutritional factors upon exocortis symptoms has been dem-
onstrated by Weathers (13); high levels of nitrogen and phosphorus
reduced the incubation period of exocortis to less than 1 year in some
cases, whereas low levels of nitrogen and phosphorus favored stunting
without scaling. Variable incubation periods and stunting without
scaling in trees infected with exocortis virus may be due to environ-
mental factors and are not necessarily dependent upon variations in
host or in virus strains.



Literature Cited

1. BENTON, R. J., F. T. BOWMAN, LILIAN FRASER, and R. G. KEBBY. 1950.
Stunting and scaly butt of citrus associated with Poncirus trifoliata root-
stock. N. S. Wales, Dept. Agr. Sci. Bull. 70: 1-20.
2. CALAVAN, E. C., and L. G. WEATHERS. 1959. Transmission of a growth-
retarding factor in Eureka lemon trees, p. 167-177. In J. M. Wallace [ed.],
Citrus Virus Diseases, Univ. Calif. Div. Agr. Sci., Berkeley.
3. CHILDS, J. F. L., G. G. NORMAN, and J. L. EICHHORN. 1958. A color test for
exocortis infection in Poncirus trifoliata. Phytopathology 48: 426-432.
4. FRASER, LILIAN R., and E. C. LEVITT. 1959. Recent advances in the study of
exocortis (scaly butt) in Australia, p. 129-133. In J. M. Wallace [ed.],
Citrus Virus Diseases, Univ. Calif. Div. Agr. Sci., Berkeley.
5. KNORR, L. C., and H. J. REITZ. 1959. Exocortis in Florida, p. 141-150. In
J. M. Wallace [ed.], Citrus Virus Diseases, Univ. Calif. Div. Agr. Sci.,
Berkeley.
6. McCLEAN, A. P. D., R. H. MARLOTH, and A. H. P. ENGELBRECHT. 1958.
Exocortis in South African citrus trees. S. African J. Agr. Sci. 1: 293-299.









CALAVAN and WEATHERS


7. MOREIRA, S. 1955. A mol6stia "exocortis" e o cavalo de limoeiro cravo. Rev.
Agr. (Piracicaba) 30: 99-112.
8. OLSON, E. O. 1952. Investigations of citrus rootstock diseases in Texas.
Proc. Rio Grande Valley Hort. Inst. 6: 28-34.
9. OLSON, E. O., W. C. COOPER, and A. V. SHULL. 1957. Effect of bud-
transmitted diseases on size of young Valencia orange trees on various root-
stocks. J. Rio Grande Valley Hort. Soc. 11: 28-33.
10. OLSON, E. O., B. SLEETH, and A. V. SHULL. 1958. Prevalence of viruses
causing xyloporosis (cachexia) and exocortis (Rangpur lime disease) in
apparently healthy citrus trees in Texas. J. Rio Grande Valley Hort.
Soc. 12: 35-43.
11. REITZ, H. J., and L. C. KNORR. 1957. Occurrence of Rangpur lime disease
in Florida and its concurrence with exocortis. Plant Disease Reptr. 41:
235-240.
12. SINCLAIR, J. B., and R. T. BROWN. 1960. Effect of exocortis disease on four
citrus rootstocks. Plant Disease Reptr. 44: 180-183.
13. WEATHERS, L. G. 1960. The effect of host nutrition on the development of
exocortis in Poncirus trifoliata. (Abstr.) Phytopathology 50: 87.
14. WEATHERS, L. G., and E. C. CALAVAN. 1959. The occurrence of cachexia
and xyloporosis in California lemon varieties, with particular reference to
the old-line Eureka lemon. Plant Disease Reptr. 43: 528-533.
15. WEATHERS, L. G., and E. C. CALAVAN. 1961. Additional indicator plants for
exocortis and evidence for strain differences in the virus. Phytopathology
51: 262-264.











L. R. FRASER, E. C. LEVITT, and J. COX


Relationship Between Exocortis and Stunting of
Citrus Varieties on Poncirus Trifoliata Rootstock






FRASER AND LEVITT (2) reported that stunting of trees on P. trifoliata
stock can occur without scaling, with results almost as unsatisfactory as
when exocortis is present. They suggested that stunting and exocortis
may possibly be related. This paper reports the details of experiments by
means of which the hypothesis is being tested, and discusses the results so
far obtained.


Symptoms of Stunting
The degree of stunting varies from severe, where affected trees are
comparable in size with those affected by exocortis, to moderate or slight.
In extreme cases, cropping is poor and the foliage sparse.
Stunting is common in Washington Navel orange and Marsh grape-
fruit, relatively uncommon in Valencia orange. In Emperor mandarins,
some variation in tree size and in the nature of the bud union is found in
trees on P. trifoliata but it is not certain that true stunting of the type
seen in Washington Navel oranges and Marsh grapefruit occurs.
Field observations suggest that, in trees affected by stunting, the slight
depression in growth rate that occurs at the age of 3 or 4 years is hard to
distinguish from normal variability. By the time trees are 8 to 10 years
old, there are detectable differences in size between those which will be
permanently stunted and nonstunted trees, and these size differences
become increasingly pronounced with age. In stunted trees, the stock is
broader than the scion trunk and often prominently shouldered or
benched at the bud union, in contrast with the butt of nonstunted trees,







FRASER, LEVITT, and COX

in which the stock is fluted and fairly evenly expanding from the bud
union to the crown roots. All nucellar scions on P. trifoliata rootstocks
which have been examined show the smoothly expanding type of bud
union.
The presence of an indented ring in the surface of the wood at the
bud union, with a corresponding projecting ridge and brown deposits in
the bark, is relatively common in stunted trees, but is not constantly as-
sociated. It has also been seen fairly frequently in nonstunted trees.
The colour test found by Childs et al. (1) to be a reliable means of
forecasting the appearance of exocortis has been used in theexamination
of very many stunted trees, but no positive reaction has been seen.
The development of yellow blotches, which is a constant feature of
exocortis-affected wood of P. trifoliata older than 2 years (Frolich, per-
sonal communication), has not been seen in material propagated from
stock suckers from a number of stunted trees.


Experimental Work
During the period 1950-1960, experimental propagations were made
to obtain information on the hypothesis that stunting and exocortis may
be related. The problem has been attacked from a number of aspects.
PERPETUATION.-To determine whether the stunting factor is carried
in budwood, trees have been propagated from stunted, exocortis-free
trees of Washington Navel orange, Valencia orange, and Marsh grape-
fruit; and these are under observation in Experiment Station orchards
at Yanco and Somersby. The Valencia orange parent was an extremely
stunted mature tree. Twenty progeny trees 9 years old are not noticeably
smaller than trees of the same age of vigorous budlines. The Washington
Navel orange parent was an extremely stunted tree 38 years old. Twenty
progeny trees 9 years old average 2/3 the size of trees of vigorous bud-
lines. The Marsh grapefruit parents were (a) a nonstunted vigorous tree,
(b) a moderately stunted tree, and (c) a severely stunted tree, all 16
years old. Five progeny trees of each are commencing to show slight
differences in size and habit.
TRANSMISSION OF A STUNTING FACTOR.-A nucellar navel-type
orange, the Bellamy navel, which produces a vigorous, large tree on P.
trifoliata, was used to obtain information on the effect of inoculation
with buds from stunted exocortis-free trees. Nursery stocks were budded
in 1953 and inoculating buds from stunted sources inserted in February,







PROCEEDINGS of the IOCV


1955. The trees were planted in the experimental orchard of the Gosford
Citrus Experiment Station in September, 1955.
Nine bud sources were used, comprising 3 severely stunted and 3
moderately stunted Washington Navel orange trees, 1 severely stunted
and 1 moderately stunted Marsh grapefruit, and 1 severely stunted
Valencia orange. No effect of inoculation on size or vigour was observed
in 1956, 1957, or 1958. In 1959, depression of vigour was apparent in
trees inoculated from 2 sources, and in 1960 all inoculated trees with the
exception of 1 series inoculated from a stunted Washington Navel orange
and 1 series inoculated from the stunted Valencia orange were showing
marked effects. The degree of stunting induced varied according to the
source of inoculum. The most severe reaction was shown by trees inoc-
ulated with buds from a Marsh grapefruit source. These were little more
than half the size of noninoculated trees, and their foliage was paler and
less dense. Other selections have so far caused somewhat less stunting.
The effect of inoculation on butt circumference of 4 of the selections is
shown in Table 1.


TABLE 1. EFFECT OF INSERTION OF INOCULATING BUD FROM STUNTED
SOURCE ON GROWTH OF BELLAMY NUCELLAR NAVEL ORANGE
Mean butt
circumference
in cm, taken
at 6 inches
Source of Number above bud
inoculum of trees union, 19.1.60
Noninoculated 11 15.8
Severely stunted Marsh grapefruit 5 12.5
Moderately stunted Marsh grapefruit 5 11.7
Moderately stunted Washington
Navel orange 5 13.4
Severely stunted Valencia orange 5 14.4


INTERACTION OF STUNTING AND EXOCORTIS.-Budwood from severely
stunted, moderately stunted, and very vigorous nonstunted trees of
Marsh grapefruit on P. trifoliata stock was used for the production of
trees on P. trifoliata stock in 1953. Ten trees of each were inoculated with
a severe type of exocortis by bud insertion in February, 1955. These and
5 noninoculated trees of each budline were planted in the experimental
orchard at Somersby in September, 1955.
No effect of source of budwood was discernible in 1956, 1957, 1958,







FRASER, LEVITT, and COX

or 1959, but in 1960 slight differences in size had begun to show between
noninoculated trees of stunted and nonstunted origin. During 1959-60,
symptoms of scaling were observed on exocortis-inoculated trees of the
severely stunted and nonstunted budlines, but not on trees of the mod-
erately stunted budline. The strain of exocortis virus used has in other
transmission work produced scaling symptoms in 4 to 5 years from inocu-
lation. There are indications of a slight retardation in time of the onset
of the scaling symptom in trees of stunted parentage compared with trees
of nonstunted parentage. There are also slight differences in the effect
of exocortis on tree size, between trees of stunted and nonstunted bud-
lines. The effect of inoculation with exocortis on growth rate was first
apparent in 1957, 2 years after inoculation, and has become progressively
more marked. The trees of nonstunted origin inoculated with exocortis
are smaller than similarly infected trees of severely stunted and mod-
erately stunted origin (see Table 2) and have somewhat sparser foliage.

TABLE 2. THE EFFECT OF INOCULATION WITH EXOCORTIS ON SIZE OF
GRAPEFRUIT TREES OF VERY STUNTED, MODERATELY STUNTED, AND NONSTUNTED
ORIGIN
Girth of trunk in cm, at
height of 6 inches above Number of trees
bud union, 19.1.60 showing scaling
Budwood source inoculated symptoms,
noninoculated with 22.9.60
exocortis
Nonstunted 18.2 12.3 7
Moderately stunted 17 12.7 0
Severely stunted 16.7 12.9 5


THE EFFECT OF STOCK SELECTION.-Strains of P. trifoliata.-Varia-
tion within the species of P. trifoliata is not great in the material avail-
able in New South Wales, suggesting that original imports of this stock
were few in number and of uniform type. However, the possibility that
strains of P. trifoliata may perform differently as stocks and that some
could produce stunted trees is being investigated. A total of 33 stock
strains are under trial for Washington Navel and Valencia orange,
Marsh grapefruit, and Ellendale mandarin in experiment orchards at
Griffith, Yanco, and Somersby. These trees are from 6 to 14 years old
and differences in size of trees of the same age on these various stocks are
very slight as yet. None of the trees is stunted. In all these trials, the
stocks were selected in the nursery for uniformity before budding.







PROCEEDINGS of the IOCV


Selection within a strain of P. trifoliata.-It is possible to distinguish
categories of seedlings in the seedbed, based on size and on degree of
deciduousness. A percentage of seedlings, which varies somewhat with
the seed source, will become completely leafless during their first winter,
others will retain their leaves until the spring. In 1952, seedlings of a
single tree source were graded into 5 classes based on size and presence or
absence of the deciduous habit as follows: (a) deciduous, over 18 inches;
(b) deciduous, 12-18 inches; (c) leafy, 12-18 inches; (d) leafy, 6-12
inches; (e) leafy, under 6 inches.
These were budded with a vigorous Valencia orange selection and the
trees planted in the experimental orchard at the Gosford Citrus Experi-
ment Station. Slight differences in tree size are becoming apparent be-
tween the different classes. The smallest trees are those on the smallest
deciduous stocks, the largest those on the leafy stocks. There is no dif-
ference between the trees on the different grades of leafy stock. None of
the trees, however, can be classed as stunted.


Discussion
Experiments on the stunting of citrus trees on P. trifoliata rootstock
have not yet reached the stage where the cause of the condition can be
assigned. It is thought, however, that stunting may be due to several
different causes. Variations in the vigour of both stock and scion almost
certainly are responsible for some of the size differences seen in commer-
cial orchards. Severe stunting in occasional cases is probably due to
accidental use of nonnucellar stocks, as has been demonstrated by Web-
ber (3) to occur with sour orange stocks. It is possible that the few cases
of stunting known in Valencia orange are due to this cause, since the
progeny of a severely stunted Valencia have grown well on P. trifoliata
stock, and inoculation from a stunted Valencia to nucellar navel orange
has produced no marked stunting effect in the inoculated tree.
Transmission to vigorous nucellar navel orange of a factor causing
stunting has been obtained with budwood of Marsh grapefruit and
Washington Navel orange and it is probable, therefore, that a virus is
involved in these cases. The prompt development of exocortis in a pro-
portion of stunted Marsh grapefruit trees inoculated with exocortis virus
may be taken to indicate a lack of relationship between the two condi-
tions, though this is not supported by the relatively greater stunting pro-
duced by exocortis when inoculated into vigorous Marsh grapefruit than
when inoculated into trees of a stunted budline. This seemed to show







FRASER, LEVITT, and COX


that the presence of the stunting factor reduced somewhat the stunting
effect of exocortis, but at this stage it cannot be taken as proof of rela-
tionship.
The psorosis virus is not involved in the stunting problem, for none of
the experimental material used carried psorosis. The possible association
of xyloporosis has not yet been fully investigated.
From the practical aspect of providing clean budwood for use with
P. trifoliata, quick tests for exocortis alone are insufficient because it is
not possible by this means to eliminate all unsatisfactory scions.


Literature Cited

1. CHILDS, J. F. L., G. G. NORMAN, and J. L. EICHHORN. 1959. Early diagnosis of
exocortis infection in Poncirus trifoliata by a laboratory test, p. 155-161.
In J. M. Wallace [ed.], Citrus Virus Diseases. Univ. Calif. Div. Agr. Sci.,
Berkeley.
2. FRASER, L. R., and E. C. LEVITT. 1959. Recent advances in the study of
exocortis (scalybutt) in Australia, p. 129-133. In J. M. Wallace [ed.], Citrus
Virus Diseases. Univ. Calif. Div. Agr. Sci., Berkeley.
3. WEBBER, H. J. 1948. Rootstocks, their character and reactions, p. 69-161. In
L. D. Batchelor and H. J. Webber [ed.], The Citrus Industry, Vol. 2. Univ.
Calif. Press., Berkeley and Los Angeles.










SYLVIO MOREIRA


A Quick Field Test for Exocortis








AT THE Experiment Station in Limeira, tests were undertaken to find
a method that would reduce the time required to produce easily visible
symptoms of exocortis; such a test is of great importance in a budwood
certification program. The new tests and the results obtained are describ-
ed in this paper.


Top-Working of Plants in the Nursery
Nursery plants of Caipira sweet orange were budded February 17,
1956, with buds from infected and healthy clones representing 4 varieties
as follows: Marsh seedless grapefruit, 5 with exocortis and 6 without;
Baianinha (navel) orange, 7 with exocortis and 6 without; Hamlin
orange, 3 with exocortis and 3 without; and Pera orange, 3 with exocortis
and 5 without.
Almost two years later, December 6, 1958, these plants were top-
worked with buds from seedlings of Rangpur lime. Within 5 months,
some of the Rangpur lime branches growing on infected plants showed
yellow elongated blotches and subsequently developed splits with raised
borders typical of exocortis. The branches of Rangpur lime that de-
veloped on plants without exocortis grew with normal green colour in
the bark.


Top-Working Old Plants
In this test, buds of seedlings of Rangpur lime were top-worked on old







MOREIRA


trees of the Hamlin, Pera, and Baianinha orange varieties. Two plants of
each variety were of nucellar seedling origin and free from the virus. The
top-working was done August 25, 1959, and the budded branches were
cut back a month later. The Rangpur lime branches that developed from
buds in trees with exocortis showed the elongated yellow blotches in the
bark in February of 1960. The Rangpur lime buds in the nucellar clonal
trees developed normal green branches.


Top-Working with Trifoliate Orange
Replication of these two tests using P. trifoliata as indicator showed
the same results as with the Rangpur lime, but the yellow blotches ap-
peared 1-2 months later. The trifoliate sprouts were frequently weaker
than those from Rangpur lime.


Inoculation of Seedlings in the Nursery
In the third test, buds from infected trees of Hamlin and Baianinha
orange varieties were budded directly into 18 seedlings of Rangpur lime
that were 2-year-old vigorous nursery plants. These inoculations were
made October 12, 1959, and a month later the main stems were cut back
to 10 cm above the point of budding. The buds used for inoculation were
allowed to develop but were later cut back to within 5 cm of their base,


FIGURE 1. Branches from Rangpur lime sprouts. A. Bark splitting with curled
edges. B. Two exocortis-inoculated seedlings (left) and 2 noninoculated ones.







PROCEEDINGS of the IOCV


and 4 young Rangpur lime seedling branches were allowed to grow. The
rest of the seedling sprouts were removed.
By February of 1960, 17 of the 18 seedlings showed the elongated
yellow blotches on the Rangpur lime branches. About 2-3 months later,
some of the infected Rangpur lime branches showed bark splitting with
curled edges similar to the symptoms recognized as due to exocortis
(Fig. 1, A). Leaves of the infected branches show chlorotic symptoms
similar to those recognized as due to mineral deficiencies. However, the
Rangpur lime seedlings in the same nursery and the same age, but bud-
ded with buds from nucellar clones of the Hamlin and Baianinha orange
varieties and cut back and subjected to the same treatment as the in-
fected plants, had vigorous normal branches. In fact, growth of the
plants budded with the nucellar clones was much greater than growth of
plants inoculated with the infected buds (Fig. 1, B).


Results
The search for a field test for exocortis has been based on the bark
splitting symptoms in the trunk of trifoliate and Rangpur lime rootstocks.
One of the tests made by Moreira (1) demonstrated that top-working
with Rangpur lime or with trifoliate orange was also a good way to ob-
tain an indication whether the tree is or is not an exocortis carrier. It was
mentioned that the first exocortis symptoms were "several yellowing spots
on the bark" of the Rangpur lime or of the trifoliate orange branches.
Later the bark splits and shelling appears.
Replications of this test in nursery trees and in old plants showed that
it is possible to see the yellow elongated blotches within 5-6 months after
top-working. The more vigorous the branches the less time is required for
the yellow blotches to appear.
Earlier results were obtained in the nursery by inoculating (budding)
vigorous Rangpur lime seedlings and cutting back. The 3 to 4 sprouts
growing from the seedlings showed the yellow blotches within 4 months.
This later procedure seems to be the best and quickest field test for
exocortis in a budwood certification program.


Literature Cited
1. MOREIRA, S. 1959. Rangpur lime disease and its relationship to exocortis,
p. 135-140. In J. M. Wallace [ed.], Citrus Virus Diseases. Univ. Calif. Div.
Agr. Sci., Berkeley.










VICTORIA ROSSETTI


Testing for Exocortis







SYMPTOMS OF EXOCORTIS are usually visible 4 to 8 years after graft
inoculation (1, 2, 3). This fact has led several authors to undertake
investigations for a quicker detection of the virus infection.
Childs, Norman, and Eichhorn in 1958 (4) showed color reactions of
the ray cells in cross sections of the bark of exocortis-infected Poncirus
trifoliata treated with certain reagents such as phloroglucinol-HC1.
In 1957 Moreira (5) observed scaling of the bark on branches of
Rangpur lime which had developed from healthy budwood grafted to
exocortis-affected citrus trees. More recently he has observed that symp-
toms, starting with yellow patches and followed by typical cracking and
scaling, will develop within a few months (6). This method should pro-
vide for a valuable field test for the quick detection of exocortis virus on
citrus trees to be selected as a source of healthy budwood.
To verify the efficiency of this method as a field test, an experiment
was carried out at the Limeira Citrus Experiment Station of the Instituto
Agronomico, the results of which are reported here.


Methods and Material
The experiment was carried out in a number of 3-year-old trees
originating from the following budwood sources:
1. A 14-year-old healthy Bahianinha orange tree derived from a
nucellar seedling, grafted on Rangpur lime-Limeira Citrus Experiment
Station.
2. A 20-year-old diseased Bahianinha orange tree grafted on Rangpur
Lime-Limeira Citrus Experiment Station.







PROCEEDINGS of the IOCV


3. A 20-year-old diseased Bahianinha orange tree grafted on trifoliate
orange-Limeira Citrus Experiment Station.
4. A 7-year-old diseased Bahianinha orange tree grafted on Rangpur
lime-Limeira Citrus Experiment Station.
5. A 20-year-old apparently healthy Pera orange tree grafted on
Rangpur lime-Limeira Citrus Experiment Station.
6. A 20-year-old apparently healthy Pera orange tree grafted on tri-
foliate orange-Limeira Citrus Experiment Station.
7. A 4-year-old diseased Pera orange tree grafted on Rangpur lime-
Limeira, Fazenda Citra.
8. A 16-year-old healthy Hamlin orange nucellar seedling-Limeira
Citrus Experiment Station.
9. A 6-year-old diseased Hamlin orange tree grafted on Rangpur lime
-Araras, Fazenda Campo Alto.
10. A 16-year-old healthy Marsh-seedless grapefruit seedling-
Limeira Citrus Experiment Station.
11. A 6-year-old healthy nucellar Marsh-seedless grapefruit tree graft-
ed on trifoliate orange-Limeira Citrus Experiment Station.
12. A 20-year-old diseased Marsh-seedless grapefruit tree grafted on
Rangpur lime-Limeira Citrus Experiment Station.
13. A 20-year-old diseased Marsh-seedless grapefruit tree grafted on
trifoliate orange-Limeira Citrus Experiment Station.


In all cases of diseased trees, exocortis was identified from the bark
symptoms.
In February and March, 1956, buds from each source were grafted on
4 rootstocks-Cleopatra tangerine, Caipira sweet orange, Rangpur lime,
and trifoliate orange-which were then kept in the nursery until Novem-
ber, 1957, when 7 plants of each stock-scion combination were planted
in an orchard at a distance of 4 x 4 meters. In May, 1959, the trees of
different stock-scion combinations were top-worked with healthy buds
from seedlings of Rangpur lime and trifoliate orange. A total of 12 bud-
dings, 6 each of those 2 varieties, were made on the branches of a number
of trees of each stock-scion combination, except on the trees on Trifoliata
rootstock because their poor development permitted only a small number
of grafts.


Results and Discussion
In September, 1959, some of the grafts of Rangpur lime were already
showing yellow patches on the bark; and in October, 5 months after







ROSSETTI


FIGURE 1. Young branches of Rangpur lime showing different intensity of
symptoms of exocortis. The first branch at left is healthy.


budding, most of those budded on diseased trees were showing cracks in
the bark (Figs. 1 and 2).
Table 1 gives the results read 9 months after the trees were top-
worked. Only the number of trees on which the top-worked shoots had a
satisfactory growth are recorded; most of the Trifoliata buddings de-
veloped poorly or not at all. Because of its fast development, Rangpur
lime gives a better test than trifoliate orange. Tests in which symptoms
were observed, such as yellow patches, cracks, and scaling of the bark
along the shoots of the test variety, are recorded as positive, and those
in which no such abnormalities were found as negative. In most cases,
the shoots of Rangpur lime showed symptoms when the top-worked tree
had originated from diseased budwood, and developed normally healthy
when the tree was from healthy budwood, thus proving that the test is
reliable for early detection of symptoms of exocortis virus in citrus trees.







PROCEEDINGS of the IOCV


FIGURE 2. A. A lesion showing cracking and scaling of the bark in a branch
of Rangpur lime. B. A lesion when the bark is removed; gum is formed in the
wood, inside the crack.




Nevertheless, some exceptions were found. With 4 of 12 trees originating
from healthy nucellar Bahianinha, the test was unexpectedly positive
on the Rangpur lime shoots. To be sure that those trees were diseased,
buds from 3 of them and buds from healthy Rangpur lime were grafted
on Caipira sweet orange seedlings in 5 replications each. In a few
months, the Rangpur lime shoots showed marked symptoms of the
disease in all replications in 2 of the cases, thus proving that those 2 trees
had been infected with exocortis virus, whereas for the third tree, only 1
of the 5 replications is now-10 months after budding-showing some
discoloration of the bark. More recently, still another test was made:
buds from the same 4 nucellar Bahianinha trees, as well as from the
mother tree from which they had originated, were budded on Rangpur
lime seedlings, but none of them are showing symptoms yet, 3 months
after budding.
Before being transplanted in the orchard, those trees had stayed in the










o,





0
ci







0
0'-
C "
o o
cc2Z
ai<*

















0
cic
3 a

a o


0 a



















rz
H
14
0 0



HZ
a <
ci-








Ho.



CC



ZS

o a



.am
3e,
Hci
ci-












ci~
-ci
t S
Sa


< 0

> Q
Bgo
-ci
aci
0>'

0<
Cci






ci
"2Z



0


Cd -V
6hi

o+




b E+




0$

C











Co d i












P +


oc
c^, 0













0'-
C-
ci
-, w
s "f











6 0.-'I






0< ^
00O
o




s 5+






s :+





u -

C 5





cS
S





4 "

b1
0^


0


010 00 M 00

0- C00 0 --





0 0404 en -0

S-0 0 0


mCOOO)Ln '-LnO DCe --CD

004q0 en 00 O D0 I- 00- en





00 C00 -

-- C 00-




S moI- cen 00- 0 o 00
COOOO MO ON OO



en c'd co' -' o co 14-




0 0- 0

0 0o -



















B es o ( a co

e u u n u uc u en
4-C 3







PROCEEDINGS of the IOCV


nursery for over a year, close to diseased grapefruit trees, and root graft-
ing may have taken place, thus transmitting the virus to healthy nucellar
Bahianinha plants. However, it seems improbable that natural root
grafts should have occurred so frequently, and other means of transmis-
sion of the exocortis virus should be investigated. The occurrence of root
grafts could not be verified because almost all the trees had been taken
out of the nursery.
On 4 trees originating from diseased Pera orange buds budwoodd
source n.7), the test was negative when the 9 months' readings were
made; but 12 months after budding, the symptoms started to show very
definitely on 3 of them; on the fourth one they have not yet appeared.
Buds taken from those trees were grafted on Rangpur lime seedlings; and
3 months after budding, the Rangpur lime shoots were already showing
some discoloration and cracks of the bark in some of the replications.
The same was observed on 1 tree of infected grapefruit.
The fact that symptoms of variable intensity and variable periods of
incubation were observed in the test applied to trees originating from
different budwood sources seems to point to the possible occurrence of
different strains of the exocortis virus.
Objections may be presented that other bud-transmitted virus diseases
might cause the same bark symptoms on the Rangpur lime and trifoliate
orange bud-developed branches. It is well known, and was confirmed by
a recent survey carried out in the State of Sao Paulo by Rossetti and
Salibe (7), that the Bahianinha variety in S. Paulo orchards is free from
xyloporosis and Hamlin is free from psorosis. Since the test has shown
positive symptoms on exocortis-infected trees of those two varieties and
none on the noninfected ones, it seems that at least xyloporosis and
psorosis can be ruled out.
In the State of Sio Paulo, a Budwood Certification Program is now
being developed to provide for virus-free budwood for citrus nurseries.
The results of the experiments reported in this paper seem to promise
the availability of a reliable field test to be employed in that program for
the selection of citrus trees free from exocortis.


Acknowledgment
The author is indebted to Dr. Carlos Roessing, head of the Citrus Ex-
periment Station at Limeira, and to his employees for their valuable help
with the grafting work; to Dr. A. A. Bitancourt, S. Moreira, and T. J.
Grant for reading the manuscript.








ROSSETTI


Literature Cited

1. BENTON, R. J., F. T. BOWMAN, L. FRASER, and R. J. KEBBY. 1949. Selection
of citrus budwood to control scaly butt on Trifoliata rootstock. Agr. Gaz.
N.S. Wales 60: 31-34.
2. BENTON, R. J., F. T. BOWMAN, L. FRASER, and R. J. KEBBY. 1949. Stunting
and scaly butt of citrus associated with Poncirus trifoliata rootstock. Agr.
Gaz. N.S. Wales 60: 521-526, 577-582, 641-645, 654.
3. BENTON, R. J., F. T. BOWMAN, L. FRASER, and R. J. KEBBY. 1950. Stunting
and scaly butt of citrus associated with Poncirus trifoliata rootstock. Agr.
Gaz. N.S. Wales 61:20-22,40.
4. CHILDS, J. F. L., G. G. NORMAN, and J. L. EICHHORN. 1958. A color test for
exocortis infection in Poncirus trifoliata. Phytopathology 48: 426-432.
5. MOREIRA, S. 1957. Rangpur lime disease and its relationship to exocortis,
p. 135-40. In J. M. Wallace [ed.], Citrus Virus Diseases. Univ. Calif. Div.
Agr. Sci., Berkeley.
6. MOREIRA, S. 1961. A quick field test for exocortis, p. 40-42. In W. C. Price
[ed.], Proc. 2nd Conf. Intern. Organization Citrus Virol. Univ. Fla. Press,
Gainesville.
7. ROSSETTI, V., and A. A. SALIBE. 1961. Occurrence of citrus virus diseases
in the State of Sio Paulo, pp. 238-241. In W. C. Price [ed.], Proc. 2nd
Conf. Intern. Organization Citrus Virol. Univ. Fla. Press, Gainesville.










JAMES B. SINCLAIR and RALPH T. BROWN


Reaction of Rootstocks to Exocortis







IT WAS RECENTLY REPORTED (2) that trifoliate orange (Poncirus
trifoliata) and Rangpur lime (Citrus sp.) in Louisiana show typical
symptoms of exocortis when infected, whereas Cleopatra mandarin
(Citrus reticulata) and sweet orange (C. sinensis) are symptomless
carriers of the virus. This report presents results from investigations
of effects of the virus on growth (trunk circumference) and yield of
Valencia orange (C. sinensis) scions budded on these and other root-
stocks and describes the symptoms produced on the stock.


Methods and Materials
Three series of rootstock tests were planted at the Plaquemines
Parish Experiment Station, Port Sulphur, Louisiana. The first series
was budded in the spring of 1951 with Valencia orange, Washington
Navel orange (Citrus sinensis), Owari satsuma (C. nobilis), and Pine-
apple sweet orange (C. sinensis) onto the rootstocks listed in Table
1. The second series was budded in the spring of 1952. Valencia
orange, Washington Navel orange, Owari satsuma, and Ruby Red
grapefruit (C. paradisi) were budded onto the rootstocks listed in
Table 2. In the third series, Valencia orange, Owari Satsuma, Ponkan
mandarin (C. nobilis), Orlando tangelo (Citrus sp.), and Washington
Navel orange, budded in the spring of 1956 onto the rootstocks listed
in Table 3, were planted in the field in January, 1957. All 3 series were
planted in 2 replications with 4 trees per replicate. Trunk circum-
ferences in cm taken 15 cm above the bud union, total yield in pounds
per tree, and symptoms produced were recorded.














o g


0 Pc1










4a 0
-g-








XSi
o 0I


z














"a
0 xix









--F-
z


g m






-0
a







Z0




z <








Woz





s


>'


CO

H

















>'


uc

EH


cu~doi-^omoo
en c -.14 e in en t


oocmr^ M ISC


00mm4 t4 In
s en en f




COCliOcOO-^'*-'





cc o- cc cc 0 co




en (M C0) 00- on M
Le) IriT CDco cM
4 Ce Cm 004




cot cc c-i c\
t -Oen 04 en en







r04cC) en In enc cc
t- In i-iC cc) 0) (D
r- en n I enccd


I O- c t.1 -- r-0 4 I1\


en I>


I cM4 n In 2
C04td i-on


en O4 c O Oe n 0 w
-.. .C ....aoJiocM
[H -04 ^CCCCIM


0
04
en
_i


(


- o -o .-



u 04






PROCEEDINGS of the IOCV


There were 2 plantings made for the purpose of recording the effects
of exocortis on Rusk citrange and to determine whether or not exocortis
virus could be transmitted through Owari satsuma budwood. In the
first, Rusk citrange (Citrus sp.) rootstocks were budded with exocortis-
infected Valencia orange scions and used as guard trees in the 1953
planting. In the second, budwood from Owari satsuma trees growing
on P. trifoliata rootstocks, some of which showed typical exocortis
symptoms and some that did not show symptoms, were budded on
exocortis-free P. trifoliata and planted as guard trees in 1953. The
source of infected budwood for this second test came from Owari sat-
suma grafted onto P. trifoliata rootstock previously infected by grafting
exocortis-infected Valencia orange budwood onto the stock and then
cutting the latter out before grafting Owari satsuma budwood onto the
same stock.


Experimental Results and Conclusions
No trees in these 3 series were free from exocortis that could be used
for comparison with infected trees. In order to evaluate these data it
was necessary to use previously reported results from another test plot
on the station (2). It was shown in this test that Rangpur lime and
Cleopatra mandarin were affected by exocortis, the former showing
typical exocortis symptoms, the latter no symptoms, but both showed
significant reductions in trunk circumferences when compared to their
virus-free counterpart. Exocortis-infected Rangpur lime had trunk
circumferences significantly below those of exocortis-infected Cleopatra
mandarin. There were no significant differences in trunk circumferences
between the 2 rootstocks, when both were free of exocortis, until the
eighth year after budding. Since the reaction of these 2 rootstocks to
the virus was known, Cleopatra mandarin was included in series 1 and
2 and Rangpur lime was included in the first series, and all other root-
stocks in the first series were compared to them and to Cleopatra
mandarin in the second series. In the third series, the rootstocks were
compared to Troyer citrange, which was shown to be very susceptible in
the second series.
Symptoms of exocortis, consisting of bark scaling along roots near the
soil line, appeared 2 years after budding on Rubidoux trifoliata and
Morton citrange; symptoms developed more rapidly on the latter.
Rangpur lime showed gumming and bark sloughing 2 years after
budding. All rootstocks of Rubidoux trifoliata, Carrizo, Morton, and







SINCLAIR and BROWN


Troyer citranges, Rangpur lime, and Williams tangelo were showing
bark scaling within 5 years after budding. Of these, Rubidoux trifoliata
and Carrizo citrange produced no gumming. Bark scaling did not pro-
gress so rapidly on Carrizo, Morton, and Troyer citranges as on Rangpur
lime. The amount of scaling increased each year on all susceptible root-
stocks until scaling reached the bud union. Rootstocks of Williams
tangelo, whether budded with exocortis-free or exocortis-infected scions,
produced gumming and bark sloughing, but these symptoms were not
like those produced on other rootstocks infected with exocortis. Bark
scaling and gumming did not appear on Cleopatra mandarin, sour
orange, Citremon C46216, rooted cuttings of Valencia orange, or Nor-
ton and Uvalde citranges. Data on trunk circumference and yield by
years after budding are summarized in Tables 1, 2, and 3.

TABLE 2. TRUNK CIRCUMFERENCE MEANSa (T) IN CM AND YIELD MEANS
(Y) IN POUNDS OF FRUIT PER TREE OF EXOCORTIS-INFECTED VALENCIA ORANGE
BUDDED IN 1952 ON CLEOPATRA MANDARIN, CARRIZO CITRANGE, MORTON CIT-
RANGE, TROYER CITRANGE, NORTON CITRANGE, AND UVALDE CITRANGE FOR THE
YEAR AFTER BUDDING INDICATED. SERIES 2 OF EXPERIMENTS AT PORT SULPHUR,
LOUISIANA
Years after budding
Rootstocks 1 4 5 6 7
T Y T Y T Y T Y
Cleopatra 1.6 61 33 134 38 236 44 247
Carrizo 1.6 58 34 153 40 274 46 302
Morton 1.9 32 26 73 29 169 34 205
Troyer 2.3 76 33 145 39 285 43 307
Norton 2.0 78 39 123 44 302 49 324
Uvalde 1.7 77 38 142 44 267 48 337
LSD .05 0.1 9 1 ns 1 29 2 19
.01 0.2 14 2 ns 2 ns 3 29
"Mean of 2 replications.

Observations on guard trees budded with exocortis-infected Valencia
orange on Rusk citrange rootstock failed to reveal any exocortis
symptoms on the rootstock 8 years after budding, indicating that it
may be a symptomless carrier or resistant to the virus. In the case of
the Owari satsuma test, typical exocortis symptoms appeared on those
rootstocks budded with scions from inoculated trees in the third year
after budding. Where exocortis-free scions were used, no symptoms ap-
peared 7 years after budding.
It was reported that Cleopatra mandarin was a symptomless carrier







PROCEEDINGS of the IOCV


TABLE 3. TRUNK CIRCUMFERENCE MEANS' (T) IN CM AND YIELD MEANS
(Y) IN POUNDS OF FRUIT PER TREE OF EXOCORTIS-INFECTED VALENCIA ORANGE
BUDDED IN 1956 ON TROYER CITRANGE, RUBIDOUX TRIFOLIATA, SOUR ORANGE,
ROOT CUTTINGS OF VALENCIA ORANGE, AND CITREMON C46216 FOR THE YEAR
AFTER BUDDING INDICATED. SERIES 3 OF EXPERIMENTS AT PORT SULPHUR,
LOUISIANA
Years after budding
Rootstocks 2 3
T Y T Y
Troyer 26 82 32 116
Rubidoux 15 19 19 38
Sour 27 91 33 126
Valencia 27 126 35 147
Citremon C46216 30 140 33 170
LSD .05 ns 33 ns ns
.01 ns ns ns ns
aMean of 2 replications.


of exocortis virus and was affected by the disease as shown by trunk cir-
cumferences and yield (2). Results from the present work substantiated
these findings about Cleopatra mandarin. They also suggest that 4 addi-
tional rootstocks and rooted cuttings of Valencia orange may be
symptomless carriers of the virus or resistant to it. These are Norton and
Uvalde citranges, sour orange, Citremon C46216, and rooted cuttings
of Valencia orange. Norton and Uvalde citrange budded with Valencia
orange gave yields significantly above those of Cleopatra mandarin in
the sixth and seventh year after budding.
It may be too early to give a positive evaluation based on these tests
for sour orange, Citremon C46216, and rooted cuttings of Valencia
orange. These 2 rootstocks and the rooted cuttings of Valencia orange
have given yields greater than those of the susceptible Troyer citrange
and Rubidoux trifoliata, with the differences showing up in the second
year. Norton and Uvalde citranges had significantly larger trunks than
either Cleopatra mandarin or the 3 other citranges in the fifth, sixth,
and seventh year.
Trunk circumferences of sour orange, rooted cuttings of Valencia
orange, and Citremon C46216 were not significantly larger than those
of Troyer citrange and Rubidoux trifoliata in the third series.
Although Carrizo citrange showed symptoms, it has consistently
yielded significantly higher than Cleopatra mandarin and its trunk cir-
cumferences have been close to those of Cleopatra mandarin or exceeded







SINCLAIR and BROWN


them. It may be that this rootstock will become more affected as it
grows older.
Observations on guard trees budded with exocortis-infected Valencia
orange on Rusk citrange rootstock failed to reveal any exocortis
symptoms on the rootstock 8 years after budding, indicating that it
may be a symptomless carrier or resistant to the virus. In the case of
exocortis-infected scions of Owari satsuma budded on P. trifoliata, ob-
servations showed that Owari satsuma can carry the virus and may have
escaped becoming infected in Florida as suggested by Knorr and
Reitz (1).


Literature Cited

1. KNORR, L. C., and H. J. REITZ. 1959. Exocortis in Florida. p. 141-150. In
J. M. Wallace [ed.], Citrus virus diseases. Univ. Cal. Div. Agr. Sci., Berke-
ley.
2. SINCLAIR, J. B., and R. T. BROWN. 1960. Effect of exocortis disease on four
citrus rootstocks. Plant Disease Reptr. 44:180-183.











L. J. KLOTZ and T. A. DEWOLFE


Can Phytophthora spp. Transmit Psorosis












SWEET ORANGE TREES with psorosis A lesions were inoculated with
Phytophthora spp. at the margins of the lesions. On February 9, 1960,
a 5 mm disk of trunk bark was removed with a cork borer, a 5 mm disk
of P. citrophthora on potato-dextrose agar was placed on the cambium,
and the inoculation sites were covered with paraffined paper and ad-
hesive tape to prevent drying. Seventy-one days later, when symptoms
of gummosis were evident, the fungus was isolated from the gummosis
margins advancing into psorosis lesions, and these isolates were used for
inoculating the stems of sweet orange seedlings which, when infected
with psorosis, readily show symptoms of psorosis on new leaf growth.
Gummosis lesions on the seedlings were allowed to involve about half
the circumference of the stems, and the disease was then checked by
excising the diseased bark. On June 9, 1960, margins of psorosis lesions
of the same orchard trees were also inoculated with P. parasitica. Sev-
enteen days later the fungus was isolated from the gummosis margin
advancing into the psorosis lesion. The remainder of the procedure was
carried out as with P. citrophthora..
Leaves of the inoculated seedlings are similar to those of the check
trees which received only agar on the cambium. Both sets show diffuse
clearings somewhat resembling psorosis A symptoms but which are prob-
ably caused by copper deficiency. To date there is no evidence that the
fungi transmitted psorosis.











GINO MALAGUTI and L. C. KNORR


Psorosis in Venezuela-An Emendation









IN 1954, following a survey of Venezuelan citrus groves, Malaguti and
Stoner (5) announced the presence of psorosis in the country. The
original report that most types of psorosis are present in Venezuela rested
on a comparison of symptoms in citrus groves with those described in
the literature (1, 2, 3). Conclusions based on these comparisons are now
considered specious in view of (a) subsequent observations by the senior
author, (b) recent publications of various investigators (4, 6, 7), and
(c) findings of the junior author during a visit to Venezuela in January
and February of 1960.
Bark lesions of various types are common in Venezuelan citrus trees.
Some resemble those described for psorosis "A." When, however, sub-
sequent to 1954, trees bearing such bark lesions were examined peri-
odically at different seasons of the year, none-regardless of type of
bark shelling-developed young leaf symptoms of psorosis. Nor did such
symptoms develop when buds from suspected trees were grafted into
sweet-orange indicator plants.
Circular leaf spots in hardened leaves and in fruits were interpreted
in the 1954 report as being the chlorotic spots described by Fawcett
et al. (2) under mature leaf and fruit symptoms of psorosis "B." Circu-
lar spot came to attention originally in the citrus groves of the Escuela
Practice at Maracay; since then the trouble spread in the same groves
and became known in other citrus-growing areas of the country.
In 1956, to test the transmissibility of circular spot and to attempt
confirmation of the diagnosis of psorosis "B," 16 seedlings each of sweet
orange, sour orange, grapefruit, rough lemon, and sweet lime were







PROCEEDINGS of the IOCV


grafted with buds from sweet-orange trees affected by circular spot. A
year later, circular spot was observed wherever the foliage of grafted
plants was sweet orange (whether in the stock or in the scion), but
failed to appear in any of the other varieties mentioned. This reappear-
ance of circular spot in the foliage of stock portions of trees on sweet
orange suggested at first the transmissibility of psorosis "B." Further
inspection, however, revealed the presence of circular spot also in nearby
sweet-orange seedlings that had never been budded, thus casting doubt
on the relationship between circular spot and psorosis. Further inquiry
into the nature of circular spot showed it actually to be lepra explosive,
the South American form of leprosis (4); confirmation lay in finding
the causal Brevipalpus mites in association with circular spot and in
controlling the further spread of the disease with applications of sulfur-
containing acaricides.
The above considerations settle certain questions regarding psorosis
in Venezuela; but they also raise additional ones. Why, for instance,
have the immature leaf symptoms of psorosis not yet been observed in
Venezuela-neither in the course of examinations by the first author
extending over the past 6 years, nor during the survey made by the
second author the early part of 1960 when the "spring" flush was in
excellent condition for production of symptoms? It can hardly be
imagined that psorosis virus has not yet been introduced into the coun-
try for, after all, much of Venezuela's citrus originated from importa-
tions of budwood from California, Florida, and other areas where
psorosis is troublesome. Might it be that psorosis virus is inactivated
under the high-temperature conditions prevalent in the tropics or that
leaf symptoms are obscured or masked? In this connection, it would be
interesting to learn the experience of others with psorosis in the tropics.


Conclusion and Summary
In view of the above considerations, much, if not all, of what had
previously been described as psorosis in Venezuela is actually foot rot,
crotch rot, Rio Grande gummosis, concentric canker, mechanical in-
jury, and physiological gumming rather than psorosis.
The circular spots in fruits and hardened leaves, which were thought
to be symptoms of psorosis "B," are now known to be symptoms of
lepra explosive.
There is as yet no evidence-whether of psorosis scaling, psorosis
decline, flecking, or oakleaf patterns in grove trees, or from transmission








MALAGUTI and KNORR


studies with sweet-orange indicator plants in the greenhouse-to indi-
cate that psorosis virus is present in Venezuela.


Literature Cited

1. FAWCETT, H. S. 1936. Citrus diseases and their control. McGraw-Hill, New
York. 656 p.
2. FAWCETT, H. S., and A. A. BITANCOURT. 1943. Comparative symptomatology
of psorosis varieties on citrus in California. Phytopathology 33: 837-864.
3. FAWCETT, H. S., and L. J. KLOTZ. 1938. Types and symptoms of psorosis and
psorosis-like diseases of citrus. (Abstr.) Phytopathology 28: 670.
4. KNORR, L. C., and E. P. DuCHARME. 1951. Relationship between Argentina's
lepra explosive and Florida's scaly bark, with implications for the Florida
citrus grower. Plant Disease Reptr. 35: 70-75.
5. MALAGUTI, G., and W. N. STONER. 1954. Psorosis de las citricas en Venezuela.
Agron. Tropical (Venezuela) 4: 127-149.
6. WALLACE, J. M. 1957. Virus-strain interference in relation to symptoms of
psorosis disease of citrus. Hilgardia 27: 223-246.
7. WALLACE, J. M. 1959. A half century of research on psorosis, p. 5-21. In J. M.
Wallace [ed.], Citrus Virus Diseases, Univ. Cal. Div. Agr. Sci., Berkeley.

This paper is the Florida Agricultural Experiment Stations' Journal Series
No. 1249.











COLETTE BOVE, GEORGES MOREL, FRANQOISE MONIER,
and JOSEPH M. BOVE



Chemical Studies on Stubborn-Affected Marsh
Grapefruit and Washington Navel Oranges








Ir IS GENERALLY ASSUMED that stubborn is a virus disease, although
its cause is still not clearly established (3). Presence of acorn-shaped
fruit is the most specific expression of stubborn on orange trees. On
grapefruit, blue albedo has been considered a valuable indicator of
stubborn, although nonspecific. Stubborn symptoms on citrus trees can
be very severe, but an indicator plant for stubborn, such as Mexican
lime for tristeza, is not known yet.
Tests for virus diseases are based on specific anatomical or chemical
changes that are induced by the virus in the host plant. One way of
detecting these changes has been successfully opened by Childs, Nor-
man, and Eichhorn (6, 7) for exocortis when they developed a staining
technique giving a "specific color reaction in the phloem ray cells of
the bark of exocortis-infected trifoliate orange." Advantages of this
colorimetric test over the indicator plant method reside in the fact
that it provides a specific test before the morphological symptoms are
visible on the indicator plant.
One other approach to the problem would be to detect an early virus-
induced, biochemical modification, as for instance the presence in the
virus-infected plant of a compound that would be absent in the healthy
one, such as the phenols noticed by Martin and Morel (10) in potatoes
and tobacco affected with virus.
This paper reports studies on organic acids and amino acids in the
juice from normal and stubborn-affected Marsh grapefruit and Wash-
ington Navel oranges, as a first step towards this approach.
60







C. BOVE, MOREL, MONIER, and J. BOVE


Materials
The grapefruit for these studies were kindly supplied by Dr. J. B.
Carpenter. The blue albedo fruit, as well as the normal fruit, came from
the same old-line Marsh grapefruit tree, numbered J.B.C. 436; it should
be considered to have stubborn.
The fruit, picked on March 16, 1960, were packed in clean new
vermiculite and shipped by air parcel post to Versailles (France) where
they arrived on March 22, 1960. Four normal fruit weighed an average
of 342 g whereas 13 blue albedo fruit weighed an average of 186 g.
The Washington Navel oranges used in the studies to be reported
came from two 11-year-old trees, designated A and C, in a 10-acre
block in the Sidi Slimane district in Morocco. Tree A had been given
the same cultural treatment as those in the remainder of the block, but
tree C was given a yearly "rich additional application" of fertilizer,
starting when the tree was 4 years old. Fruit from each tree was picked
in 1959 and sorted into 2 categories, normal fruit and stubborn fruit
(very small size or misshapen). Tree A yielded 33 normal fruit (average
weight, 197 g) and 80 acorn fruit (average weight, 188 g); tree C
yielded 341 normal fruit (average weight, 158 g) and 440 acorn fruit
(average weight, 104 g) (6). We chose 6 normal fruit (average weight,
199 g) and 8 acorn fruit (average weight, 170 g) from tree A, and 9
normal fruit (average weight, 167 g) and 17 acorn fruit (average
weight, 94 g) from tree C.


Methods
PREPARATION OF FRUIT JUICE.-Marsh grapefruit.-The peeled fruit
were homogenized in a Waring Blendor and the homogenate centri-
fuged. The supernatant fluid was kept frozen for one month at -100C
before being worked up. Before use, the unfrozen juice was filtered in
a folded filter paper. This filtrate is referred to as juice and all results
are expressed per 100 ml of this juice.
Washington Navel oranges.-Each fruit was cut into 2 parts-a
peduncular half and a stylar half. The normal fruit were cut into 2
equal parts. The stubborn-affected fruits were cut at the level of the
acorn constriction, so that the peduncular half corresponded to the part
of the fruit where the peel had a normal thickness, whereas the stylar
half corresponded to the narrow-skinned part of the fruit. The juice was







PROCEEDINGS of the IOCV


extracted with a lemon squeezer. Juices from similar half-fruits were
combined and filtered, first over a cheesecloth, then over folded filter
paper. Results are expressed per 100 ml of this filtrate, referred to as
juice.
Free acidity.-Aliquots of juice were titrated with 0.1 NaOH in the
presence of phenolphthalein in the case of the grapefruit, and in the
presence of phenol red in the case of the oranges. From titration curves,
it was found that phenol red was a better pH indicator than phe-
nolphthalein in the sense that the pH zone of the color change corre-
sponds more closely with the equivalent point.
TOTAL ACIDITY AND SEPARATION OF SOLUBLE CONSTITUENTS INTO
THREE GROUPS.-The constituents of the juice were separated into 3
groups: (a) acidic components, including the organic acids; (b) basic
components, including the free amino acids; and (c) neutral com-
ponents, including the soluble sugars, by passing the juice successively
through a column of Permutite 50, H* form, (a strongly acid, sul-
fonated polystyrene cation exchange resin) and a column of Dowex 2,
CO,-- form (a quaternary ammonium, strongly basic, anion exchange
resin).
The basic components are retained on Permutite 50, and the acidic
ones on Dowex 2. The neutral substances, including the sugars, are not
retained, and end up in the effluent from the Dowex 2 column. Total
acidity was determined on an aliquot of the effluent from the Permutite
50 column, before passing it through the Dowex 2 column. The titra-
tions were done with the same pH indicators used for free acidity
determinations.
The amino acids were eluted from the Permutite 50 with 1 N NH4OH
and the ammonium salts of the acid components from the Dowex 2 by
0.5 N CO, (NH,),.
The sugar solution, the amino-acid solution, and the organic acid
ammonium-salt solution were respectively taken to dryness under
vacuum at 250C. Excess NH4OH or CO,(NH,)2 was thereby elim-
inated. The sugars were redissolved in a small volume of water. So
were the amino acids and the organic-acid-ammonium salts. The latter
were passed furthermore through a Permutite 50 column to free the
organic acids from their ammonium salts.
Thus one obtains 3 solutions: an amino-acid solution, a free organic-
acid solution, and a sugar solution.
SEPARATION AND DETERMINATION OF ORGANIC ACIDS.-The organic







C. BOVE, MOREL, MONIER, and J. BOVE

acids were separated by partition chromatography on a silicic acid
column (1). They were eluted from the column by mixtures of tertiary
butanol in chloroform; the eluate was collected with a fraction collector,
and the fractions were titrated with 0.005 N NaOH.
The great amount of citric acid present in certain citrus-fruit juices
and its percentage, very high in comparison with that of other organic
acids present, render it generally difficult to detect and determine the
other organic acids. We overcame the problem by separating the organic
acids into 2 groups: one containing only malic, citric, and isocitric acids;
the other containing fumaric, glutaric, itaconic, succinic, lactic, alpha-
ketoglutaric, aconitic, malonic, oxalic, and glycolic acids (2).
The nature of the organic acids separated on silicic acid column was
furthermore checked and confirmed by two-dimensional paper chroma-
tography (4).
SEPARATION AND DETERMINATION OF THE FREE AMINO ACIDS.-The
amino acids were separated by two-dimensional paper chromatography
(9). The purple spots obtained after spraying with ninhydrin were
eluted and the optical density measured at 570 mu (11).
SEPARATION OF SOLUBLE SUGARS.-The sugars were separated by one-
dimensional paper chromatography (8).
TOTAL SOLUBLE NITROGEN.-Total soluble nitrogen was determined
on the amino-acid solution by the Kjeldahl method.


Results
FREE ACIDITY, TOTAL ACIDITY, AND ORGANIC AcID.--Data for grape-
fruit are summarized in Table 1; those for oranges in Table 2. In the
grapefruit, there seemed to be little difference in acid content between

TABLE 1. ACIDITY DATA ON GRAPEFRUIT JUICE FROM NORMAL AND BLUE-
ALBEDO FRUIT (FIGURES ARE MILLI-EQUIVALENTS (MEQ) PER 100 ML OF JUICE)
Normal fruit Blue albedo fruit
Free acidity 21.25 21.80
Total acidity 27.10 28.30
Citric acid 21.72 25.60
Malic acid 0.65 0.49
Oxalic + glycolic acids 0.14 0.20
Aconitic + malonic + alpha-
keto-glutaric acid 0.13 0.19
Succinic acid 0.05 0.06

















ac a

Q- 0r-e -
g cora rco 0
- I1
cl Ca

Z u a







u H



C -





Sco c- j c aO
C 00x-r-- 0
:j C D C) C0


2 c! c




oce ci
m c6
<








o I --








0 C)




Ui j C)





0 O U
C
CC




-< 3 -3 D u
CZC -c-+0
C c
0W









" C ^ a
EiG o*








C. BOVE, MOREL, MONIER, and J. BOVE

normal and albedo fruit. With respect to the oranges, total acidity was
higher in acorn fruit of tree C than in normal fruit and this was mainly
due to the higher citric acid content of the stylar half of the acorn fruit.
The same trends seemed to show up in the case of tree A, but to a
much smaller extent. As a matter of fact, there was a greater difference
in acidity between tree A and tree C than between normal and acorn
fruit from tree A.
Besides citric, malic, oxalic, and glycolic acids, the following acids
have been chromatographically identified and tentatively estimated in
the oranges, but so far no significant differences concerning these acids
have been found between the normal and the stubborn-affected fruit:
succinic acid (0.05 meq/100 ml juice) alpha-ketoglutaric acid (0.01
meq/100 ml juice), aconitic + malonic acids (0.07 meq/100 ml juice),
and isocitric acid (traces). Quinic, shikimic, and glyceric acids have not
been looked for. A few acids, present in microtrace amounts, have not
been identified yet.
TOTAL SOLUBLE NITROGEN AND AMINO ACIDs.-Tables 3 and 4 sum-
marize the results concerning total soluble nitrogen and amino acids,
respectively, for the grapefruit and the oranges. The blue albedo grape-
fruit contained much less arginine, proline, and gamma-amino-butyric
acid than the normal fruits. In the case of the oranges, the total soluble
nitrogen was higher in the acorn fruit than in the normal ones. There
was an increase in arginine, aspartic acid, alanine, and gamma-amino-
butyric acid in the stylar half of the acorn fruit and a decrease of most
of these acids in the peduncular half of the same fruit.
Besides the amino acids listed in Tables 3 and 4, the following free
amino acids have been identified in grapefruit and oranges, stubborn-
affected or not: lysine, asparagine, glutamine, glycine, threonine, tyro-

TABLE 3. TOTAL SOLUBLE NITROGEN, AND NITROGEN OF VARIOUS AMINO
ACIDS IN GRAPEFRUIT JUICE FROM NORMAL AND BLUE ALBEDO FRUIT (FIGURES
ARE MILLIGRAMS OF NITROGEN PER 100 ML JUICE)
Normal fruit Blue albedo fruit
Total soluble nitrogen 105.2 111.7
Arginine 53.2 30.8
Proline 17.5 13.0
Aspartic acid 11.9 9.5
Alanine 14.0 11.9
Gamma-amino butyric acid 13.3 6.7
Serine 6.3 5.6
Glutamic acid 2.6 2.1











SO

) omn r-
e c 0 -~ c 0


0s


0





0

0


0



0

0:

0











0,
0


0


0
0
F-


00
0
0








z


0 0
mtis o om n T
CLS c C0 ) m cX. co -


0

co -


0- 0

c', 4-


0 l


0


0
0 C' C - c
r- m C C
cnL


00 0


S- o
-cc


* O
ce C 13 IIR C
cd i nrl c


000
Lo r-, c6 cn cM
a -


- C -!


0 6






^ c 0
o o-


0
0 r- 00
Cflri .fr Cr c<







C. BOVE, MOREL, MONIER, and J. BOVE


sine, valine, leucine, phenylalanine, and cysteine. Asparagine was present
in great amounts, but has not been estimated yet. Tyrosine, valine,
leucine, phenylalanine, and cysteine were present in very low amounts.
Three undetermined compounds-I, II, and III-have been detected
on the ninhydrin sprayed chromatograms from the orange juices. They
could not be detected in grapefruit juices. Compound I moves close to
asparagine. Compound II lies between threonine, glutamine, and
alanine. Compound III has an Rf value of 0.98 in the phenol solvent.
Two undetermined compounds, IV and V, have been noted in trace
amounts on the chromatograms from grapefruit juices, but not on
those from orange juices.
SOLUBLE SUGARs.-Fructose, glucose, and sucrose were separated by
paper chromatography. From the size of the spots on the developed
chromatograms, it was very apparent that the blue-albedo-grapefruit
juice contained less sucrose than the normal one, and that the stylar
juice from the acorn oranges also contained much less sucrose than the
normal stylar juice. Work is underway to check, with accurate sugar
determination, the preceding rough estimations.


Discussion and Conclusion
From the data presented, it appears that stubborn induces rather
clear-cut chemical modifications in the juice from affected grapefruit
and oranges. That the chemical differences between the normal and
affected fruit are due to stubborn seems highly probable, since the af-
fected fruit showed severe stubborn symptoms. However, the results
obtained so far indicate that the chemical modifications induced by
stubborn respectively on oranges and on grapefruit are not similar.
The observed chemical differences are only quantitative, not qualita-
tive. But one should remember that the so-called "normal" fruits come
from the same tree as the affected fruits. The probability of finding a
qualitative difference, the only one of real value in trying to establish
a virus test, would be greater if one were to look for differences between
diseased material and 100 per cent healthy material. At the present time,
it is difficult, if not impossible, to find such healthy material, at least
in Morocco.
Tree C has received during 7 years a "rich additional application" of
fertilizer. Tree A did not receive it. The data on organic acids and
amino acids show that the chemical modifications induced by stubborn







PROCEEDINGS of the IOCV


are greater on tree C than on tree A. Thus it seems as if applying more
fertilizer has resulted in enhancing the effects of the disease.
We think that the ultimate chemical work intended to find a virus
test for stubborn, or for any other citrus virus disease, should not be done
on fruit, because it may take a long time before a tree bears a fruit. But,
in a preliminary phase, work on severely affected fruit can open the way
to a test that would have to be worked out on vegetative material. In
the case of stubborn, the use of fruit can be helpful because the disease
affects the fruit so very much.


Literature Cited

1. Bovi, J., et R. RAVEUX. 1957. La separation et la determination des acides
carboxyliques de C1 A Ce par chromatographie de partage sur colonne de
silice. II-Nouvelle technique propose. Bull. soc. chim. France 1957:
376-381.
2. Bovi, J., et F. MONIER. Manuscript in preparation.
3. CARPENTER, J. B. 1959. Present status of some investigations on stubborn
disease of citrus in the United States, p. 101-107. In J. M. Wallace [ed.],
Citrus Virus Diseases. Univ. Calif. Div. Agr. Sci., Berkeley.
4. CHEFTEL, R. J., R. MUNIER, et M. MACHEBOEUF. 1952. Microchromato-
graphie sur paper des acides aliphatiques hydrosolubles et non volatils.
II-Utilisation d'une phase solvante alcaline puis d'une phase solvant e
acide pour la microchromatographie A deux dimensions. Bull. soc. chim.
biol. 34. p. 380-387.
5. CHILDS, J. F. L., and J. B. CARPENTER. 1960. Observations on stubborn
disease of citrus in Morocco. Report obtained from the authors.
6. CHILDS, J. F. L., G. G. NORMAN, and J. L. EICHHORN. 1959. Early diagnosis
of exocortis infection in Poncirus trifoliata by a laboratory test, p. 155-161.
In J. M. Wallace [ed.], Citrus Virus Diseases Univ. Calif. Div. Agr. Sci.,
Berkeley.
7. CHILDS, J. F. L., G. G. NORMAN, and J. L. EICHHORN. 1958. A color test for
exocortis infection in Poncirus trifoliata. Phytopathology 48: 426-432.
8. COLOMBO, P., A. CORBETTA, A. PIROTTA, G. RUFFINI, and A. SARTORI. 1960.
A solvent for qualitative and quantitative determination of sugars using
paper chromatography. J. Chromatog. 3: 343-350.
9. LEVY, A. L., and G. D. CHUNG. 1953. Two-dimensional chromatography of
amino acids on buffered papers. Anal. Chem. 25: 396-399.
10. MARTIN, C., et G. MOREL. 1958. Accumulation de composes ph6noliques chez
les plants atteintes de maladies A virus. Compt. rend. 246: 2283-2286.
11. MENORET, Y. 1960. Action du 2.4.D. sur le m6tabolisme azot6 des tissues de
Carotte cultiv6s in-vitro. These de Doctorat, Paris.











E. C. CALAVAN and D. W. CHRISTIANSEN


Stunting and Chlorosis Induced in Young-Line
Citrus Plants by Inoculations from Navel
Orange Trees Having Symptoms of
Stubborn Disease




THE TERM "stubborn" was applied, about 1921, to nonproductive
navel orange [Citrus sinensis (L.) Osbeck] trees which had responded
poorly to top-working (6). Much later, Fawcett, Perry, and Johnston,
after 2-years' observation of experimentally top-worked young navel
orange trees, described stubborn disease in 1944, and indicated its "prob-
able virus nature." The acorn symptom of navel orange fruit was
ascribed to stubborn disease at that time and possible relationships of
blue albedo, crazy top, and acorn symptoms of grapefruit (C. paradisi
Macf.) to stubborn disease were suggested (3, 6, 8). Haas (7) later
found no relationship of acorn-shaped grapefruits to crazy top, blue
albedo, and low yield, and Carpenter (1) recently considered blue
albedo to be a valuable but nonspecific indication of stubborn disease.
Chlorosis and stunting-symptoms frequently associated with stubborn
disease (6)-were observed by us in 2 lots of citrus seedlings in 1958
during indexing operations. In the first lot, these symptoms were noted
on 6 seedlings within 2 months after inoculation with buds from tree
29-36, a twiggy, stunted, nonproductive 15-year-old navel orange on a
sweet orange rootstock inoculated by Fawcett et al. (6) in the first ex-
periment with stubborn disease. The plants affected were Eureka lemon
[C. limon (L.) Burm.], West Indian lime [C. aurantifolia (Christm.)
Swingle], and calamondin (C. mitis Blanco). They developed no leaf or
stem symptoms specific for psorosis, tristeza, vein enation, or yellow vein,
and all had sound roots. About half the seedlings used for indexing tree
29-36 remained normal, and affected seedlings made partial recovery.
69







PROCEEDINGS of the IOCV


In the second lot, stunting and chlorosis were observed in a seedling
of Tien Chieh mandarin (C. reticulata Blanco) inoculated with buds
from shoots of Frost Washington Navel orange top-worked in 1956 on
an old, medium-sized, nonproductive navel orange, tree C-189, having
symptoms of stubborn disease.
This paper reports some preliminary results of studies made from
1957 to 1960 with the agent or agents present in the 2 source trees, and
discusses their possible significance in relation to stubborn disease.


Experiments and Results
Forty seedlings of Koethen sweet orange in 1-gal. cans were inoculated
as follows: (a) 10 received 1 bud and 1 side graft, on February 16,
1960, from tree 16A-1, a severely stunted, 2/2-year-old Frost Washing-
ton Navel orange tree with symptoms of stubborn; this tree had been
inoculated in the nursery with two buds from tree 29-36; (b) 10 received
1 bud, on April 14, from a West Indian lime plant with symptoms of
tristeza; (c) 10 received 1 bud from tree 16A-1, on February 16, and
1 bud from tristeza-infected lime, on April 14; (d) 10 received 1 bud,
on April 14, from Meyer lemon 1-54, which carries seedling yellows
virus (9). Ten plants were retained as noninoculated controls. Each of
the 50 plants was then budded, on April 14, with a single bud from a
2-year-old field-grown Eureka lemon seedling, the lemon buds being
forced to grow after the sweet orange seedling tops were cut back on
May 19. All groups were kept in a screened greenhouse.
Some of the trees inoculated from 16A-1 developed, within 10 days
after they began to grow in May, 1960, weak lemon scions with small
chlorotic or mottled leaves which tended to remain stiffly erect or more
nearly at right angles to the upright stem than the larger and heavier
leaves on control plants. Some leaves became mottled, with pale chlorotic
areas extending across secondary veins. Other leaves on the same plants
developed a pale creamy yellow color, except for a wedge-shaped area
of green along the midvein and occasional specks of green in the chlorotic
portion. Many chlorotic leaves gradually became less chlorotic after
they ceased expanding. Some of the stunted plants inoculated with
buds from 16A-1 made partial recovery, and others which had started
to grow normally became stunted later. Reactions of the plants inocu-
lated from 16A-1 and those inoculated from both 16A-1 and tristeza-
affected lime appeared to be almost identical.
Some plants inoculated only with tristeza virus soon developed mild









CALAVAN and CHRISTIANSEN


chlorosis in the young lemon leaves, but 5 months after inoculation these
plants appeared normal.
Plants inoculated with seedling yellows virus from Meyer lemon 1-54
forced short chlorotic lemon shoots which soon developed severe shock
reaction, abscised their leaves, and within a few weeks dropped part or
all their stems. Six of these plants later improved and some produced
lemon shoots equal to those on some of the plants inoculated with buds
from 16A-1, or superior to them. All control plants made normal growth.
Plants representing each treatment are shown in Figure 1; specific
measurements are summarized in Table 1.


TABLE 1. MEASUREMENTS OF DISEASED AND APPARENTLY NORMAL PLANTS
OF SEEDLING-LINE EUREKA LEMON/KOETHEN SWEET ORANGE


Inoculation group Number


and apparent


condition of plants
Seedling yellows
1-54:
Diseased
Normal
Navel 16A-1:
Diseased
Normal
16A-1 + tristeza:
Diseased
Normal
Tristeza:
Diseased
Normal
None: (controls)
Diseased
Normal


Leaf blade
dimensions


of in mm"
plants Length Width


75d 34"


Shootb
length
in mm


33d 52


4 79 32 71 88
6 140 61 138 85

5 97 43 79 76
5 122 60 131 81


0
10 134


62 133 50


58 143 60


"Average of 2 leaves per plant on September 28. Measured leaves were the first
2 below a point 6 inches from the tip.
bAverage length of lemon shoots on September 28.
'Averages calculated from diameters at pot level.
'Based on the 6 seedling yellows plants which retained lemon shoots.


Field experiments were started in 1957, in which buds from trees 29-36
and C-189 were placed in seedlings of several varieties and in the seedling
rootstocks of several stionic combinations. The buds were grown into
tops on some of the seedlings; on other seedlings and on the budlings
they were used only for inoculation. All the inoculated seedlings sub-

71


Size' Growth' of stock
of stock in mm2
in mm2 May 19 to
May 19 September 28









PROCEEDINGS of the IOCV


/ f


FIGURE 1. Sister plants of seedling-line Eureka lemon/Koethen sweet orange.
Inoculations from left to right as follows: none (control); tristeza alone; tristeza
-navel orange 16A-1; 16A-1 alone; seedling yellows from Meyer lemon 1-54.








CALAVAN and CHRISTIANSEN


sequently were converted into budlings by postinoculative propagation
from young-line trees presumably free of viruses. Comparable noninocu-
lated trees were grown as controls.
Preliminary results of these field experiments agree in general with
results obtained from lemons in the greenhouse. Some plants inoculated
with buds from stubborn-affected trees 29-36 and C-189 were dwarfed
in the nursery. Other plants, similarly inoculated, were apparently
normal in the nursery, but failed to grow well after being transplanted
to the field (Fig. 2) or began to grow poorly within 2 years after trans-
planting. The fibrous roots of stunted trees appeared to be in sound
condition when examined. Eleven of the 18 young-line trees inoculated
in 1957 and 10 of 16 trees propagated in the nursery in 1957-58 with
buds from trees 29-36 and C-189 were visibly stunted or dwarfed by
May, 1960. Inoculated trees that became stunted were: grapefruit and
Satsuma on Rangpur lime (C. limonia Osbeck); Navel orange on Sun-
shine tangelo (C. reticulata x C. paradisi), on trifoliate orange (Pon-
cirus trifoliata (L.) Raf.), and on Troyer citrange (C. sinensis x P. tri-
foliata); Shamouti orange on Palestine sweet lime (C. limettioides

.' I 0. 3


FIGURE 2. A. Noninoculated control tree of Frost Washington Navel orange/
Pomeroy trifoliate propagated in 1957 and planted in 1958. B. Sister tree in
juxtaposition in same row. The rootstock of this tree was inoculated from stubborn
navel orange tree 29-36 in the nursery in 1957. Pole height, 6 ft., 3 in. Photo-
graphed September 22, 1960.







PROCEEDINGS of the IOCV


Tanaka); and Valencia orange on Troyer citrange. Propagations of
29-36 grew poorly on Orlando tangelo and rough lemon (C. jambhiri
Lush.); those of C-189 grew poorly on Cunningham citrange, trifoliate
orange, Rangpur lime, Orlando tangelo, and Palestine sweet lime stocks.
Most leaves of the dwarfed trees are abnormally small and stiff, and
some are distorted. Considerable shoot growth has occurred and some
nearly normal shoots have been produced, but most shoots have many
multiple buds. Foliage is crowded or clustered and leaves usually drop
prematurely. Although tree 29-36 and the young dwarfed trees are non-
productive, they have produced many off-season blooms, which soon
dropped. All noninoculated controls have made normal growth.
The trees used as sources of buds for inoculations in these experiments
were indexed for known citrus viruses, with the following results: (a)
lemon seedling, apparently virus free; (b) trees 29-36, 16A-1, and C-189,
stubborn; trees 29-36 and C-189 are negative for exocortis and cachexia
3V2 years after indexing; (c) West Indian lime, tristeza; (d) tree 1-54,
seedling yellows and vein enation.


Discussion
The original stubborn experiments (3, 4, 5, 6), by themselves, neither
proved transmission nor demonstrated the nature of any causal agent of
the disease. Instead, they confirmed observations that healthy buds on
stubborn interstocks grow poorly (6) and indicated that stubborn disease
is bud-perpetuated and may be transmissible. In those experiments, tree
29-36 represented a method which, with adequate controls, might have
proved transmissibility of a causal agent of stubborn disease; tree 29-36,
like most inoculated trees in our experiments, consisted of an inoculated
seedling rootstock on which a stunted top was grown from a bud of a
normal tree (6). The close resemblance of symptoms developing in our
young trees, inoculated by buds from trees 29-36 or C-189, to those
described and illustrated for young trees in early reports of stubborn
disease (4, 5, 6) and the relationship of tree 29-36 to the original ex-
periments with stubborn disease have led us to conclude that the causal
factor or factors of one kind of stubborn disease studied by Fawcett et al.
(6) have been transmitted to several stionic combinations of citrus.
The results from 3/2 years of indexing indicate that the graft-trans-
missible agent or agents responsible for chlorotic, small-leaved growth
and general stunting of inoculated trees are different from the viruses of
psorosis, vein enation, yellow vein, and tristeza; probably different from








CALAVAN and CHRISTIANSEN


those causing exocortis, cachexia, and xyloporosis. The possibility that
combinations or variant forms of these viruses may have been present
has not been eliminated.
Inoculations from 29-36 and from C-189 have induced dwarfing in
some stionic combinations that are considered tolerant of tristeza and
seedling-yellows viruses, in some tolerant of exocortis virus, and in some
tolerant of cachexia and xyloporosis viruses. However, the effect of
16A-1 inoculations in some plants of Eureka lemon on sweet orange re-
sembled that of seedling-yellows, except that the initial severe shock
following seedling-yellows inoculations was absent in these plants. Re-
duction in leaf size, similar to that caused by 16A-1 inoculations, has
been reported for orange trees affected by stubborn disease (2). The
transmissible agent or agents from dwarf Navel tree 16A-1 differ from
tristeza virus, and there has been little or no synergistic effect between
tristeza virus and the dwarfing agent from 16A-1 in budlings of Eureka
lemon on Koethen sweet orange.
The occurrence of apparently normal individuals among bud progeny
of diseased sources, as well as mild or seemingly negative reactions in
nearly half the inoculated young-line trees, might be due to (a) slow
movement or irregular distribution of the causal agent in the host, (b)
dominance of a mild form of the agent in some buds, (c) host tolerance
due to physiological or genetic factors.
Considerable variation in symptoms occurs among trees assumed to
be affected by stubborn disease (1, 6), and it has been questioned
whether a single disease is responsible for all the symptoms ascribed to
stubborn disease (1, 2). Environmental factors can cause dwarfing and
poor tree condition (1), and large Navel orange trees having stubborn
disease may be confused with nonproductive strains of this variety (6).
The most constant symptoms attributed to stubborn disease (6) are not
sufficiently specific to be diagnostic. In our experience, many trees
tentatively diagnosed as having stubborn disease on the basis of gross
symptomatology have proved to be adversely affected principally by
factors such as root rot, cachexia, exocortis, inferior rootstocks, stionic
incompatibility, and heredity.


Literature Cited
1. CARPENTER, J. B. 1959. Present status of some investigations on stubborn
disease of citrus in the United States, p. 101-107. In J. M. Wallace [ed.],
Citrus Virus Diseases. Univ. Calif. Div. Agr. Sci., Berkeley.
75








PROCEEDINGS of the IOCV


2. CHAPOT, H. 1959. First studies on the stubborn disease of citrus in some Medi-
terranean countries, p. 109-117. In J. M. Wallace [ed.], Citrus Virus Dis-
eases. Univ. Calif. Div. Agr. Sci., Berkeley.
3. FAWCETT, H. S. 1946. Stubborn disease of citrus, a virosis. Phytopathology
36: 675-677.
4. FAWCETT, H. S., and L. J. KLOTZ. 1948. Stubborn disease. Calif. Citrograph
33: 229.
5. FAWCETT, H. S., and L. J. KLOTZ. 1948. Stubborn disease one cause of non-
bearing in navels. Citrus Leaves 28(8) : 8-9.
6. FAWCETT, H. S., J. C. PERRY, and J. C. JOHNSTON. 1944. The stubborn
disease of citrus. Calif. Citrograph 29(6) : 146-147.
7. HAAS, A. R. C. 1950. Acorn disease in grapefruit. Citrus Leaves 30(9) : 10-11.
8. HAAS, A. R. C., L. J. KLOTZ, and J. C. JOHNSTON. 1944. Acorn disease in
oranges. Calif. Citrograph 29(6): 148, 168-169.
9. WALLACE, J. M. 1957. Tristeza and seedling yellows of citrus. Plant Disease
Reptr. 41: 394-397.











J. B. CARPENTER


Virus Content of Citrus Trees with
Symptoms of Stubborn Disease






SINCE 1955 some citrus trees in California and Arizona that showed
symptoms of stubborn disease (1) have been indexed for the following:
tristeza, psorosis, vein enation, exocortis, cachexia, and xyloporosis
viruses. Trees representing severe, moderate, and mild types of stubborn
disease (1) and growing on various rootstocks were selected from the
following varieties: sweet orange [Citrus sinensis (L.) Osbeck]-'Wash-
ington Navel', 'Robertson Navel', a local navel orange selection, 'Va-
lencia', 'Mediterranean Sweet', 'Hamlin', 'Shamouti', 'Rico No. 1',
'Koethen', and an unnamed sweet orange; grapefruit (C. paradisi
Macfad.) -'Marsh', 'Redblush', and 'Webber's Java Pink', an unlisted
introduction made by W. T. Swingle; and Minneola tangelo (C.
paradisi 'Bowen' x C. reticulata Blanco 'Dancy').
Although the indexing will not be completed for several years, pre-
liminary results from 36 trees provide the first information on the
virus content of trees affected by stubborn disease.
No evidence of tristeza, psorosis, or vein enation was found in any of
the trees indexed for adequate periods on Mexican lime [C. aurantifolia
(Christm.) Swingle], on sweet orange, or on sour orange (C. aurantium
L.), indicating that the causal viruses of these diseases are probably not
involved in the development of stubborn disease. Exocortis, as indicated
by symptoms on seedlings of Rangpur lime (C. limonia Osbeck) and
trifoliate orange [Poncirus trifoliata (L.) Raf.], was present in 19 trees.
Cachexia, as expressed on Orlando tangelo (C. paradise 'Bowen' x C.
reticulata 'Dancy'), was found in 20 trees. Exocortis and cachexia oc-
curred together in 15 trees. Indexing for xyloporosis on Palestine sweet








PROCEEDINGS of the IOCV


lime (C. limettiodes Tanaka) was abandoned as unreliable because at
2V2 years of age all the 6 nonbudded control seedlings had developed
conspicuous wood pitting that could be confused with symptoms of
xyloporosis. No suitable substitute has been found for Palestine sweet
lime unless one concurs with the view that xyloporosis and cachexia are
caused by the same virus, or closely related viruses, and that, therefore,
Orlando tangelo is an adequate indicator plant for both.
By August, 1960, 12 of the 36 trees had not yielded any evidence of
an identifiable virus disease other than stubborn; this group included 5
trees with severe symptoms of stubborn disease, 5 with moderate
symptoms, and 2 with mild symptoms. Indexing of these trees is being
continued and in some cases will be repeated.
In searching for a specific indicator plant for a possible stubborn
disease virus, budwood of 2 Marsh grapefruit trees with stubborn disease
was used to inoculate seedlings of 59 species, varieties, hybrids, and
relatives of citrus. The uninoculated control plants of 5 kinds failed to
grow well. Plants of the other 54 kinds have grown satisfactorily for 3
to 5 years and have shown no definite symptoms of virus infection or
symptoms that might be attributed to some unknown virus.
These preliminary results suggest that stubborn disease does not result
from infection by the causal viruses of tristeza, psorosis, vein enation,
exocortis, or cachexia, either singly or in combination. Likewise, these
results provide no evidence that stubborn disease is caused by an un-
known virus. Nevertheless, virus studies will be continued to complete
current and planned investigations to determine the cause of stubborn
disease and to clarify Fawcett's (2) report that stubborn disease is graft-
transmissible and caused by the virus Citrivir pertinaciae Fawcett.


Literature Cited
1. CARPENTER, J. B. 1959. Present status of some investigations on stubborn
disease of citrus in the United States, p. 101-107. In J. M. Wallace [ed.],
Citrus Virus Diseases. Univ. Calif. Div. Agr. Sci., Berkeley.
2. FAWCETT, H. S. 1946. Stubborn disease of citrus, a virosis. Phytopathology
36: 675-677.










H. CHAPOT


Morphological Modifications Induced by
Stubborn Disease on Citrus Fruits





STUBBORN DISEASE causes characteristic modifications that seem to be
specific for the disease. Nevertheless, so long as no unquestionable in-
dicator plant is available, only the presence of acorn-shaped fruits,
according to certain authors, is diagnostic. We think, however, that the
acorn deformation can show up in a slightly modified way and still be
specific for stubborn, and that, inversely, stubborn induces on citrus
fruits other modifications just as typical.


Acorn Deformation
For a long time, in the Mediterranean area, acorn deformation re-
mained, if not rare, at least not frequent, even on trees showing all the
leaf and blooming symptoms. Recently, it has shown up more and more
frequently, even on varieties, like Valencia orange, considered as pro-
ducing typical acorn fruits scarcely ever. For instance, in the citrus
collection of the Iskenderun Experiment Station (Turkey), of 17 varie-
ties, 16 showed acorn fruits in February, 1960, on practically each of
the 15 trees of the following 16 varieties: Marocchini sanguigno, Khalili
red, Valencia Late (from California), Valencia Late (from Palestine),
Valencia Late (from Egypt), Ovale calabrese, Hamlin, Tarocchino,
Tarocco dal muso, Tarocco liscio, Sanguigno, Washington (from Flor-
ida), Shamouti, Ananas Signorelli, Tounsi, Akgay sekerlisi. In Morocco,
typical acorn fruits were found on the following orange varieties:
Washington, Surprise, Golden Buckeye, Hamlin, Petite Jaffa, Shamouti,
Grosse Sanguine, Tarocco, Vernia, and an unnamed variety from
Florida.







PROCEEDINGS of the IOCV


On the other hand, we have not found the acorn deformation on
mandarin (especially Willow-Leaf and Clementine), or on grapefruit,
or on sour orange, although these species, especially the first two, show
all the other stubborn symptoms. Neither has acorn deformation been
seen on lemon, but this species has never presented leaf or blooming
symptoms. While looking at the citrus collection of the College of Agri-
culture in Athens (Greece), we observed acorn deformation on all the
fruits from two small trees of an undetermined variety, probably an
orange-lemon hybrid.
Acorn deformation is accompanied by a complete necrosis of the tissues
corresponding to the stylar half. The cells die and the cuticle becomes
wrinkled and whitish, probably due to the presence of air between the
disorganized cells. The intensity of this whitish stain is increased by
rubbing the fruit. The necrosis of the fruit's distal hemisphere can be
observed rather early, and there are cases of very young fruits (size of
a hazelnut) that show rotting from June on. All these fruits drop prema-
turely. During October-November from the aspect of the tissues of cer-
tain fruits when they are still green, it is possible to tell which ones will
show the typical acorn modification when mature. This necrosis is di-
rectly related to the phenomena of color inversion (see below) and to
pronounced modifications of the pulp.


Columella Curvature ("Lopsided Fruits")
All the cases of columella curvature, yielding lopsided fruits, have
been observed on trees showing other stubborn symptoms, on branches,
leaves, and blooms. These trees often produce acorn fruit also. Thus,
one can assume that the curvature is a symptom of stubborn, particularly
since we have observed in Morocco, all the transition stages between a
typical acorn fruit and a lopsided fruit, especially one stage where the
fruits show laterally the acorn deformation plus necrosis. Furthermore,
on that part of the lopsided fruit where the meridian is shortest, a
modification of the oil gland is noticeable, identical with the one on the
acorn fruits. However, it should be noticed that only the lopsided fruits
have a greyish stained albedo (see below).
The columella curvature is extremely frequent on mandarin (Willow-
Leaf and Clementine) and on grapefruit, in contrast to the scarcity of
the acorn modification. It is also frequent on oranges.
While in Morocco, J. B. Carpenter drew our attention to the fruit
deformation of Sampson tangelo affected by internal breakdown. In







CHAPOT


studying 5 trees in the Sidi Kacem district (Morocco), we observed
fruit deformations consisting of the "sun burn" symptom of internal
breakdown, but also acorn deformation, very typical of stubborn, with
all the transitions between sun burn fruits and acorn fruits.


Decrease of the Fruit Size
One of the effects of stubborn is a decrease in size of fruit, particu-
larly with Valencia orange. For some time, we have noted on stubborn-
affected trees, fruits of a size much smaller than normal. These fruits
do not come from an out-of-season blooming, but from the normal
March-April blooming; thus, they have had a sufficient growing
period.
Furthermore, the epidermis of these fruits is extremely rough and
makes them resemble small sour oranges. On almost all these small-
sized fruits, the albedo is stained either blue or grey. This phenomenon
is extremely frequent on oranges, mandarins, and grapefruit. In
Turkey, Dr. Adil Cengiz, of the Plant Protection Service in Adana,
has observed it on seedlings of sour orange trees, more than 100 years
old, planted in the public garden of Tarsus. Looking at those trees
in February, 1960, we made the same observations; the trees seem to
manifest very typical symptoms of stubborn. It is interesting to note
that this disease affects seedlings and that these seedlings have had a
normal behaviour up to a recent period.


Coloration of the Albedo
For a long period, the abnormal color of the albedo appeared not to
exist in the Mediterranean countries. The reason for this is that one
looked for it only on mature and normal-sized fruits. Even today, in
Morocco, blue albedo is not found on mature and normal-sized fruits,
not even on fruits harvested from trees showing other symptoms of
stubborn.
Since J. B. Carpenter's observations (2) and especially since his
demonstrations in the Moroccan groves in November, 1959, the facts
appear differently; the abnormal albedo colorations have often been
noticed, but only on immature fruits and on mature but small-sized
fruits. These fruits show a rough epidermis.
These observations were made in groves where hormonal treatments
were never carried out and in countries where they never are used







PROCEEDINGS of the IOCV


whatever the culture might be; thus we are faced here with a phenome-
non due to stubborn and not to chemical treatments, such as those men-
tioned by J. B. Carpenter (1).
On the other hand, the blue staining of the albedo is much less fre-
quent than in the stubborn cases noted in Arizona and in California,
although one observes it more and more frequently in the Mediter-
ranean area; the most widespread staining is a blue-greyish color, or
more frequently, a clear-cut grey. This greyish stain can be found on
the lopsided part of the fruit, the one corresponding to the shortest
meridian; it is almost always accompanied by an abnormal abundance
of fruit vessels, which contribute to a fibrous appearance in a cross
section through the albedo.


Color Inversion
In a presumably healthy fruit, the orange color of the rind starts at
the stylar scar and moves towards the peduncle in such a way that, at
half maturity, the peduncular hemisphere is still green whereas the
stylar end is perfectly orange. In a stubborn-affected fruit, the color
pattern is just the opposite; the stylar end does not color up or remains
orange-green, and the peduncular hemisphere is orange-colored. This
phenomenon is very clear-cut, and very marked on navel oranges, and
is accompanied by profound chemical modifications of the pulp. The
color inversion is a regular fact on any fruit, mainly oranges, showing
acorn deformation.
In connection with the color inversion, another phenomenon may be
observed on stubborn-affected blood oranges; the normally red part of
the rind and flesh remains uncolored (or the color has been destroyed).
This lack of color is found on acorn fruit as well as on lopsided fruit;
in acorn fruit the blood-orange color is lacking only from the affected
stylar part; that is, the peduncular part is normally colored on the out-
side and shows red flesh. In lopsided fruit, only the affected lateral part
is uncolored; the opposite one is colored and has red flesh. In this way,
actually bicolored fruits are obtained.
This lack of anthocyanic pigments (or their destruction) is intimately
connected with the other effects of the causal agent of the disease on the
fruit: acorn shape, thinning down of the epidermis, necrosis of tissues,
delayed maturity of the stylar part for the acorn fruit, deviation of
columella, modifications of the shape and of the number of essential
oil glands for the lopsided fruit.








CHAPOT


Another symptom, probably connected with color inversion, is dis-
played by all the out-of-season fruits harvested from stubborn-affected
trees: the stylar half of the fruit remains green, sometimes dark green.
It was observed on oranges, grapefruits, and chiefly mandarins (Clemen-
tine, Willow-Leaf, and Wilking). Similar observations were made by
J. F. L. Childs while examining in November, 1959, the Sanguine man-
darin trees in the citrus collection of the Marrakech Experiment Station
in Morocco. On all the fruits, the stylar hemisphere was completely green
whereas the peduncular hemisphere was orange-red. The limit between
the two colors was extremely marked. Such fruits were perfectly ripened,
and from a regular blooming, not from an out-of-season period as for
the fruits described above. At first sight, the trees did not show any
other stubborn symptoms, whether on leaves, shoots, or flowers; how-
ever, they grow in an area heavily contaminated by this disease.
Thus, some similarities can be observed between this "green bottom"
in the Mediterranean area, and the "greening" observed by P. C. J.
Oberholzer in South Africa.


Literature Cited
1. CARPENTER, J. B. 1958. Accentuation of the blue albedo in Marsh grapefruit
by sizing sprays with 2, 4, 5-T. Plant Disease Reptr. 42, 63-64.
2. CARPENTER, J. B. 1959. Present status of some investigations on stubborn
disease of citrus in the United States, p. 101-107. In J. M. Wallace [ed.],
Citrus Virus Diseases. Univ. Calif. Div. Agr. Sci., Berkeley.










R. H. HILGEMAN


Response of Stubborn-Infected Trees
to Iron Chelates







W HILE STUBBORN DISEASE affects several varieties of sweet oranges
and grapefruit in Arizona, it is most widely prevalent in the Washington
Navel oranges. Surveys show from 4 to 43 per cent affected trees.
Several commercial groves planted since 1948 have more than 20
per cent infected trees. Budwood for these groves came from highly
productive old-line trees without recognizable stubborn symptoms. This
high incidence indicates that stubborn is the most serious citrus disease
in Arizona and suggests that it is becoming worse.
Although stubborn disease was not designated and reported as such
until 1944 (3), many Arizona growers recognized these offtype "runt"
trees between 1920 and 1930 and attempted to improve them by various
fertilizer, pruning, and top-working treatments. Records of the develop-
ment of the disease and its effect on fruit production have not been
reported.


Diagnosis of Stubborn Disease
The chief characteristic for the identification of stubborn disease is
abnormal growth which produces atypical development of shoots,
leaves, and fruit. This is an uncertain basis for diagnosis. Acorn-shaped
fruit associated with stubborn in other areas occurs infrequently in the
desert area. Young trees may develop vegetative symptoms before the
trees begin to set fruit, so that only leaf and shoot abnormalities can be
used.
The writer distinguishes two types of stubborn disease in the Wash-







HILGEMAN


ington Navel orange. Type A includes trees with a general restriction of
growth that produces a stunted tree. Symptoms may develop on trees
3 years old or may be delayed until the tree is 25 to 30 years old. In old
trees, symptoms usually develop on one part of the tree and gradually
spread. Characteristic symptoms are as follows: small, upright spring
leaves; short, stubby summer shoot growth with round type, thick leaves
with prominent veins; growth from multiple buds and twig dieback.
Other symptoms are: leaf abcission in December; reduced sucker de-
velopment; less tolerance to cold; increased injury from citrus thrips
and off-season blossoming.
Type B is characterized by abnormal vigor of primary shoots which
develop into main scaffolds. Secondary shoot growth is restricted and
similar to that of Type A so that an open type tree develops. As the
disease advances, individual limbs may defoliate badly, weak shoot
growth then follows or the limb may die back. Iron chlorosis is prevalent.
Acorn-type fruit are more likely to occur than on Type A trees.

Development of Stubborn Disease
Observations have been made on 417 trees budded on sour orange
rootstock and planted in 1933 in Block F at the Citrus Experiment Sta-
tion in Tempe, Arizona. The original owner of the grove obtained the
trees from two commercial nurseries and they are typical of the com-
mercial trees planted at that time. Yield records began in 1943 when
the University obtained the grove and observations on stubborn disease
were started in December, 1948.
The development of Type A stubborn disease symptoms and yield
records from 5 affected trees are presented in Table 1. These records
illustrate typical variations in the development of the disease and
emphasize the difficulty in evaluating treatment responses.
Tree I is considered a normal healthy tree. Varying yields apparently
are caused by climatic conditions.
Tree 2 is a typical tree that developed stubborn disease early in its
life so that yields were always low.
Tree 3 was never fruitful, but failed to develop definite stubborn
symptoms until 1951. After many low producing years it produced a
moderate crop in 1954, and then declined rapidly.
Tree 4 is a tree with delayed deterioration from stubborn disease. In
1954 two main limbs developed symptoms rapidly. The opposite side
of the tree remained normal for 2 more years before stubborn became
general and yields decreased. This type of disease manifestation is be-




University of Florida Home Page
© 2004 - 2010 University of Florida George A. Smathers Libraries.
All rights reserved.

Acceptable Use, Copyright, and Disclaimer Statement
Last updated October 10, 2010 - - mvs