TATE PL/-NT BOARD
TECHNICAL SERIES NO. 19, PART V.
U. S. DEPARTMENT OF AGRICULITU RE,
BUI1EAU OF ENTO MOLOGY.
L. O. HOWARD, Entomologist and Chief of Bureau.
TECHNICAL RESULTS FROM THE G(IPSY MOTH
V. EXPERIMENTAL PARASITISM:
A STUDY OF THE BIOLOGY OF LIMNERIUM VALIDUMI (C1tESSON,
By P. H. TIMBER.LAKE, A. M.,
Agent and Expert. Gipsy Mloth Parasite Laboratory.
ISSUED MAY 29, 1912.
GOVERNMENT PRINTING OFFICE. 3L3 1912.
BUREAU 01' ENTOMOLOGY.
L. 0. HOWARD, 1,7140111olOffixt (OW ChiCf Of B111-C(I'll.
C. L. MARLATT, I"'WOM0109ist (1101 ACNII!j ChiCf ill Absclicc of Cllicf.
R. S. CLIFTON, k,'XCClltiI'C ASSiNI(Int.
11'. 1. TASTET, Chicf Clerk. F. 11. ill, ('1101-tIC of h w-l". (111d sim-cd product i116 cct ill 1-csfigatIMIS,
A. 1). 1101-KINS. ill (-Wlt'gO of fol-CSt hl,, C('t ill I'('slig(Itiolix, AN'. 1). ill charge (if solithcr)l fivId cr)p insccl ill 1-cstigatiolls.
F. 11. in charge of ccl-c(II (Ind forage illscct hlrcxliyatioll, .
A. 1,. QVAINTANCE, in charge of (1ccidloms fruit ia8c(-t inrcstigations. 11]. F. I'mrrirs, in ch(tryc of bcc cultarc.
Ro(;) ,I S, in Char (Ic ()f prercittitig sprca.ll of laoths, ficId work.
11. Cujmii in charge of c(liforilil 11-ork. MAI'LL COLCORD, in chm-gc of library.
PIUNENTIN(. SPREAD OF 'MOTHS.
W. F. Fjsi i,. in charyc; A. F. UUW;ISS, IIAIIRY S. SNIITII,' 1'. 11. TINIMAILAKE!
C. W. COLLINS, 11. 1':. SMITH. V. 11. Mwsm it, J. N. Sum'mi-,Ils. CII..'s. W.
.111NOTT, 4'. W (ISSINI(III/S.
in char.uc: 11. B. D--turoN, 11. W. VINP)rq. D. C. 'MURI-11Y. 1. L.
BAILIN, 11. L. NtVINTYRE, (INSiShMIS.
TnITISI".1-red to :111d fm-;I,- e illsoct ill ve"'t t ions.
.111-4ol-l-cd to citi'll-, fruit illsect illv' Sti-aliou,6.
C 0 NT EN T S.
Introduction ....................................................... 71
Experim ents ............................................................... 72
General methods of experimentation .................................... 72
Behavior of Limnerium validum (Cresson) in confineiiet ---------------- 73
Limnerium validum as a parasite of Euproctis chrysorrhtaa (Lii uaeus) ..... 73
First experiment............................................. 74
Second experiment ................................................. 74
Amcebocytosis as a protective reaction of the host ..................... 75
Adaptation of parasite to host .........--------------------------. 76
Further experiments with Euproctis chrysorrhwa ---------------------- 77
Limnerium validum as a parasite of Malacosoma americana (Fabricius) ..... 78
Results of experiments as determined by dissection ................. -78
Emergence of adults of Limnerium from reproduction ................. 79
Length of the stages in the life history of Limnerium .....------------ 80
A possibly normal parasite of Malacosoma --------------------------- 80
Limneriurn validum as a parasite of Notolophus antiques (Linnaeus) ----- 81 Limnerium validum as a parasite of Porthetria dispar (Linnaeus ---------- 82 Experiments with other hosts ------------------------------------------ 83
Notes on development and anatomy of the larva --------------------------- 83
The egg ---------------------------------------------------------------- 83
The first-stage larva -------------------------------------------------- -84
Description of the newly hatched larva ------------------------------ 84
Function of the tail appendage -------------------------------------- 85
Appearance of older larvae ---------------------------------------- 85
Notes on the anatomy of the larva ------------------------------------ 86
The second-stage larva ------------------------------------------------ 87
The third-stage larva -------------------------------------------------- 87
Description of the full-grown larva---------------------------. 87
Feeding habits of the imprisoned larva- --------------------------88
Respiration of the larva in the third stage -------------------------- 89
The cocoon ----------------------------------------------------------- 90
Results of the dissections of Hyphantria during the fall of 1910 ................ 90
General summary -------------------------------------------------------- 91
Bibliography ............................................................... 92
I L LUS TRAT IONS.
FIG. 3'.. Limneriumin validum: Adult female, abdomen of male, metascutum,
3:3. Limlieriuin 'alidumi: Egg......................................---- 83
34. Limneriuln rabalin: Eclosion of larva............................. 83
35. Lirnneriuin ralidum: First-stage larva, newly hatched ................84
36. Limiterium ralidurn: Firsit-stage larva of large size, showing- ,ilk glands
and ner-vous systemli .......................................... 5
37. Limneriurn ialidumi: Mouthiparts o)f first-stage larva ..................8S6
38. Lim neriint ra lidu iu: S'-econd-stage lav..............7
39. Linmrium caiduhint: Mouthparts of second-stage larva ................871
40. Lineiriurn culiduin: Third-stage larva ............................. 8
41, Lirnncriuin r~diditi: Anterior view of head of third-stage larva, showi n 1outhpart............................................... 88
U. S. D. A., B. E. Tech. Ser. 19. Pt. V. Issued May 29, 1912.
TECHNICAL RESULTS FROM THE GIPSY MOTH PARASITE LABORIATOR1Y.
V. Experimental Parasitism:
A Study of the Biology of Limnerium Validum (Cresson).
By P. H. TIMBERLAKE, A. M.,
Agent and Expert. Gip.sy Moth Pa rasite Labo'atory.
The biology of none of our American parasitic Ilymenoptera is too well known to demand an apology for the appearance of this article, and it is the hope of the author that the results of the studies here set forth will call to the attention of our professional and amateur entomologists this exceedingly rich but almost untouched field for investigation. A little of the ecology, especially of the host relationship, of many of our parasites has been recorded, but the various and interesting adaptations in the larval structure and habits of our different species are almost unknown.
In connection with the practical work at the Gipsy Moth Parasite Laboratory of importing and liberating parasites of the gipsy a-nd brown-tail moths, considerable attention is paid to the habits and biology not only of the imported parasites, but also of our native parasites of caterpillars that resemble the gipsy moth or brown-tail moth in habits or destructiveness. In this way considerable knowledge has accumulated which, besides its intrinsic and theoretical value, has been useful in perfecting the methods of handling the imported species and in showing which of our American species, if any, may be expected to become contributing factors in the control of these pests.
It was especially with this latter idea in mind that in the spring of 1910 a series of experiments was undertaken to work out the biology of our native Limnerium validum (Cresson) (fig. 32) and its behavior toward unusual hosts. The experiments grew out of a successful attempt to foster oviposition on small brown-tail moth caterpillars (Euproctis chrysorrhoea Linnaeus), which yielded such unexpected complications that the work was continued on other hosts as they became available in the course of the spring.
Limnerium validum proved to be an excellent subject for experimentation for several reasons, but chiefly on account of its docility in 71
72 RF. T-ITS FRo.\r GIPSY MOTH PARASITE LABORATORY.
coTifimiiient and the readiness with which it attacks any suitable ,lwcie- of caterpillar and because the heavily chitinized eborion of Ilic egg is reimirl ably durable. This last fact wag an advantage in deterinininor the number of eggs deposited in a caterpillar, even when coiisiderable time had elapsed after the hatching of the larva. Foritinately for the success of the experiments an abundance of adults of this species was at hand, reared from cocoons obtained the preeedijl _(r fall frolli it!:z 11slial Ilost, Hypantr;a citnea (Drury).
This L;i1oi(r;iii1i is one of the hymenopterous parasites alfectinor the f all welm-orin (Hyphanfila citnea) and although not so important a.,, Jloteoris and Apaideles, it is frequently ound. In the northern st'ltc- it, en-lel-fres from the fall w bworm and spins its cocoon mostly duriii Ole iiioiitli. of September. Unlike L;mnej4wI1, J);7o, 1171111? (Pro1-:111clier). ajiotlier species con-iiiion on Hyphantria, it hibernates 111 its cocoon until late
the following spring
or summer Mid consequentlv -it has been
considered to have
only a single generation a year. 11"artlier
South, however, as
pointed out bi Dr.
L. 0. Howard (1897)
in his bulletin oil the
white marked tiissock-moth (11cmerocampa 1eu(-o,,4;fpm.
Fit,. :.2. Limm-rittin ralithim: (l, Adult feluale, side view Sinith and Abbot) I
1A, abdoineii of inale; c, inehiscutuin froin above; d, of wlii'ch it is an iiiicocooll. itt, 1). (1, Nedrawn froul 11oward; inipoi-tant, parasite, it
e, orit"ifitil.) probably biberiia tes
"is t1lil "Idlilti as its iiil)t y c x -oons were found associated witli -'spim111) 11-v'r of the Ili, Ill C I W(WIll p(i in the middle of 1 eceint)(w. Dr. 11,MV11111 al- o i-econis rezii-ina tlieadtilt as eai-1 t -, tIl ill I Ile ) J t_ y a e I ( ( ( f IlIv,
SO t1lat in flie viciiiity of Wa.sliiilgrton, D. C., two (rViieratiolls, if not, 1110IT11 0CCM* 41111111,111Y. Besides its faxoied host. HypIwIlb.;(I (www, wi(] tlle 1j(1Ilcr0( milp(t, it probably atfilcks other cateqJllars, ))lit so fial. :is tli(. is aNvare no (Aller rearijigs liave been recorded.
BIOLOGY OF LIMNERIUM VALIl'M. 73
number of caterpillars, a small, flat tray was used(l, wire scrieellned on the bottom and covered with glass. A small hole in the side of the tray was found useful in introducing fresh foliage s)rayed with sweetened water, as food for the caterpillars and p)arasites. In the first experiments several females of the Limn,riutm. were confined with a limited number of caterpillars, but later, wheni it was discovered how readily the parasites attacked the hosts, the number of the latter was proportionately increased. )issetin of
the caterpillars was chiefly relied upon in working out the results of each experiment.
BEHAVIOR OF LIMNERIUM VALIDUM (CRESSON) IN CONFINEMENT.
This species did not become overrestless in confinement, and hardly betrayed the excitement that some parasites are said to do when in proximity to their hosts. This tranquility may possibly be explained by the fact that the Limneriim was always confined with species of caterpillars that do not naturally serve it as hosts, and it might have been stimulated, perhaps, to greater activity by fall webworms. The females, however, readily oviposited in 4 of the 6 species of caterpillars with which they were confined. The posterodorsal part of the host was usually chosen as the most suitable point for attack, and there seems to be a more or less well-developed instinct in this as well as other parasites to keep away from the head of the caterpillar in ovipositing. This instinct is not so necessary for large. strong parasites like Limnerium raliduim, but may be of great advantage to small, weak species like Aleteoruts or .tpanteles. These frequently attack caterpillars that are many times their size, ind might easily fall a victim to one vicious sweep of the host's head. or become so daubed up by juices exuding from the mouth of the excited caterpillar that they could not escape. As circumstantial evidence of the existence of this instinct, comparatively few eggs were found near the head in dissected caterpillars, and by far the greater number was found in the posterior half of the body. A favored site for them was in the extreme tip of the postero-dorsal part of the body cavity just beneath the integument.
The males of Limnerium were slightly more active than the females in running about the cage, but they paid no attention to the caterpillars, and but little attention to the females. None was ever seen attempting to mate.
LIMNERIUM VALIDUM AS A PARASITE OF EUPROCTIS CHRYSORRH(EA (LINNAEUS).
Three experiments were undertaken during February and March, 1910, to determine whether Limnerium ?alidtum would successfully attack brown-tail caterpillars, and to collect data, if possible, on the early stages.
74 RESULTS FROM GIPSY MOTH PARASITE LABORATORY.
In the first experiment, on February 2, 5 females of Lim,nwrium were confined in a small glass cylinder with 10 active brown-tail caterpillars about S nun. long. The experiment was closed on February 15, at which time the last of the caterpillars died. Nine of the caterpillars were carefully dissected and a count made of the easily discovered eggs and larv' of Linnwrim. The largest number found in one caterpillar was 23 eggs and 11 larve with the empty eggshells dissected from one that died on February 11. The smallest number was 4 eggs and 5 larva taken from a dead caterpillar on IFebruiary. 15. One hundred and forty-six eggs and 72 larva, were found in all, making a total of 218 eggs deposited by the 5 females in the course of the 4 to 6 days that they remained alive, or an average of nearly 44 eggs for each female, and about 24 to each caterpillar.
With this unusual number of eggs and larve present, it is no wonder that the caterpillars failed to survive. As the caterpillars in every case died before the larvae had grown to any appreciable extent and even before all the eggs had hatched, it is not probable that they were killed l)y the feeding of the parasites. It is much more probable that they succumbed to the mechanical irritation produced by so many eggs and larva, which not only may have Caused a violent disturbance or pathological condition of the body tissues, but also tended to keep themni from feeding.
A second experimnient wis started oni February 5, with 5 females and 13 active )rown-tail caterpillars. By February 9, all the parasites were dead, and 3 caterpillars were found dead on the 11th and 5 more on the 19th. In the S dead caterpillars 61 unhatched and 5 hatched eggs were found, hut only 1 larva, which had grown to be aboit five times the size of newly hatched larva,, although still in the first stage. About S eggs, on an average, were thus found in each (ate rpillar, and they ,were undoubtedly at least a strong contrilj)utingr factor in causing the death of the latter.
TlIhree of the 5 living caterpillars were dissected on February 19 and ithe Iremaining two o()n March 4. In these 5 caterpillars only 1 unmatched egg and 12 eggshells with only 4 nlarvn were found. This, on an average, is only 2 or 3 to each caterpillar, for although 1 had been victimized( S times, 1 had escaped altogether. Only 4 nlarv, were found, and of these 3 had died shortly after hatching ,before grolwilig to aIy extent, whereas 1 had managed to live 1ild i4creal in size aomit five times. In lthe 13 caterpillars dise ,teld. 72 ii uihallchd eggs. 17 hatcl(led eggs, aind 5 larvae were found, (or 14early 7 to a cater)illar and is for each female Linwru.
BIOLOGY OF LIMNERIUM VALIDUM. 75
AMGBOCYTOSIS AS A PROTECTI'IVE REACTION OFT II 1osT.
The remarkable fact in the array of figures jtist given is ihe disparity between the number of eggshells or cast clhoria found a11d the number of larva. The latter were large enough so that they could not have been overlooked, and the only explanation is that they had been killed by the host and absorbed by the blood tissues, or destroyed, in other words, by an(nbocytosis. The chitinolis choria, on the other hand, escaped unharmed. In the preceding experiment, in one of the dead caterpillars dissected on February 15 the dead larvae and some of the eggs and eggshells were found inclosed in a firm, thick, homogeneous-appearing, transparent capsule of tissue. The significance of this was not realized at that time, but when on February 19 the same phenomenon was noticed in the dissection of living caterpillars, the mystery of the missing larvw was explained, for when the capsule was stained in methyl green and mounted in glycerin, it was resolved under the high power of the microscope into innumerable, densely packed, minute, roundish and spindleshaped cells. These were undoubtedly blood-tissue cells or amnebocytes, and whereas they were not true phagocytes, yet their function must have been to break down and absorb the tissues of the parasites, for within the capsules larvw were found in several stages of dissolution.
Later experiments with the same and other hosts, such as Jlalacosoma and Notolophus, brought to light the fact that this amcebocytic reaction takes place regularly when the Lirnnerdam occurs in these hosts to which it seems to be unaccustomed and unadapted. In most cases it was found that the parasite was able to hatch from its egg, but that it perished generally soon afterwards before growing to any extent. Many times the larva was found to have disappeared entirely through the action of the amcbocytes, yet the story of its death and dissolution was told by the encapsulated remains of the much more resistant eggshell. Whether the amwbocytes gathered around and killed the living larvoe and eggs or attacked only parasites that had been killed by some other factor was not certainly determined; but the former supposition is probably correct, as unhatched eggs containing embryos in different stages of dissolution were found sometimes inclosed in sheaths of these cells. In one instance, also, a larva that had grown to be about six times the size of newly hatched larvw, was found entirely inclosed except for its head in a characteristic sheath. This larva was probably living when its host was killed for dissection, or at least, judging from its fresh and uninjured condition, it could not have been dead for more than a few hours.
An extended series of dissections of Hypliantria canea, on the other hand, made by J. D. Tothill during the fall of 1910, showed that the
341760-No. 19, Pt. V-12- 2
76 RESULTS FROM GIPSY MOTH PARASITE LABORATORY.
larvw of Llll lwre*llia are free froin the slightest trace of a inoebocy t i e attack NA-hen ocetiri-ing in their natural host.
Other pha c of this so-called pliagrocytosis have been studied hy -,evei-al Ettropean investigators, especially by Cuenot (189(0, Janet (1906, 1907), aiid Pantel (1910). The latter author has gii-eii
-icv were known to him
stininnary of those ca,, es of all bocytosis that
in Ills excellent "Rech(rolics 8ttr 7(,., Diptcres a, L(irves Entaill'obies." He colieludes that this defensive reactioii of the host is aroused i)ot
()iil-%- bv bacterial infection but. also by sick or dead parasitic lai.va', an(i bN*' niolt skins. and states that free and healthy larvw are retntlarly exempt.
The only ca 4es of sick larvae that Pantel seems to have observed tire hii-N-te of Tachinidw that have fallen accidentally from their 1weathing holes. geiienally during the ii-iolting of the host. Such
do not, affix theniseIA-es anew, but wander about in the body c-1166- of the host until overcome bv stiffocation. Panel further ob! el'Ve.s that the anal sheaths of such larvtt begin to turn brown and becoine inclosed by a great number of amwbocytes, which finally ahnost coiiipletely cover the larN-a.
In the ctlse Of dead larvae. which have perished in the striiagle for the I)o se.,- sioii of the host, Pantel ol)-erves that their bodies onlY exceptioiudly become incapstilated by amwbocytes. The molt skins, oil the othe I- litand, lie furthermore states, are attacked, not always, btit most fre(pient1v bv aino(,bocytes.
'I"he shnilar act ioii of"' aiii(,Pbocvtes in bi-eaking down the will(Y ilillscles of (Itieen 'ants has been described by Janet (1906, 1907). The imiscles -tire not. devoiired in sniall fragments by pliaorocytes, but are dishite,ri-tited cyradtially and absorbed by ainwbocytes, which creep in laiumig the fibi-ilh-v in large ntunbers.
Ave liaN-e seemingly in this amcebocytic reaction a manifestation of the protective factiltv of the host in guarding against the attael of elltopal.,.I 'It(-'. s siiown by Pantel (1910) and confirmed by oitr ONN H 01)-cl \-atiolls of VaillOlls species of parasitic Dil)tvi-a atiol HN1111(11optei',i. iiormal or adaptc(l parasitic when liealthy. ral:ely if
ev(,r tar(')tise this defensive reactioti, or, in other possible 11101T (10111'('Ct Nvords. they ( ble to i-opel the ainwbocytes by oiiie ctirioos adaptatioll. 1-11a(1:1pted parasite ; oil the (.olltl..,;r y. 1)(:M(Y unable to r(,I)vI the aII Iwh(wvt iiiiv i-eptLirly sticeiiiiib to their The histilict of
1111(ler or(lillarv (q)ji(litiolls, well fits tjj(qll fql- (-hoo ijig t1lell,
-toilled hosts, I)Ilt in the million,., of illAallces whel-C this illstinct
illto plaN1, all (,(Y()* May be occasioli:llly ill all unsuitable Ill sitch (-"VC th(I FC Itlltillg I'll-va eellls to he (1(miiied to a Iiiigel-il)g death. :111d it,, tissile" 411.0 ahs(>I-bed by the aillwbocYtes With I)() 111tilliate (101-1111elit to Ow 11()-t.
ADAPTATION 01-' PARAS4111' TO 11014T.
()II the hnII(L brillfr- 111) the
(11111-10111. W 111%. (10) 1104 '111 -Ilffcl. the "aille fate, and NvIiat
BIOLOGY OF LIMNERIUM VALIIDUM. 77
constitutes adaptation? We begin here to sink deep) into tle
FURTHER EXPERIMENTS WITH EUPROCTIS CHRYSORRH(EA.
A third experiment was started on February 15 with about 100 brown-tail moth caterpillars that had been feeding for 20 days and were consequently about 10 mm. long. These caterpillars were placed in a small tray, and 15 females of Limnneributm were confined with them between the 15th and 27th of February, fresh females being added as the first ones died.
Dissection of living caterpillars was undertaken on the 4th of March and continued on the 14th. Thirty-four caterpillars in all were dissected and 15 were found to be parasitized by Liminerintm. 'Not more than 2 larve or eggs were found, on an average, in a single caterpillar, and the larvae were in most cases dead. Only one larva, in fact, had obtained any size, and it was only about six times as large as the newly hatched larvae. Although inerusted with a capsule of amoebocytes it may have been still alive, but could not have survived
78 RESULTS FROM GIPSY MOTH PARASITE LABORATORY.
much longer. All the other larve that were found were newly hatched. or had been killed soon after hatching, and were commonly inclosed in a sheath of amwebocytes.
The rest of the caterpillars were kept alive as long as possible in the hope that one or two of the Limnerium might be able to pass through their transformations, but none was reared. The experimient therefore furnished data similar to those obtained in the preceding experiments, and conclusively confirmed the belief that Lbnim,Werium alidum is unable to live as a parasite within brown-tail moth caterpillars.
LI.MNERIUM VALIDUM AS A PARASITE OF MALACOSOMA AMERICANA (FABRICIUS).
On April 20 an experiment was started with Limhnwrium alidum
as aI parasite of the conmmnon tent caterpillar (Malacosoma americana Fabricius). Three females were confined in a large, flat, tanglefooted tray, with three nestfuls of caterpillars, mostly in the second stage, collected in the open at Melrose Highlands, Mass. A fourth nest was added on the 21st to insure an abundance of caterpillars for the Limnerbum. to attack. By the 26th the Limwnerium had all died,
but 2 more females were placed in the tray on the 28th. Many of the caterpillars at that time were passing into the third stage.
RESULTS OF EXPERIMENTS AS DETERMINED BY DISSECTION.
On May 3 dissections were made of 6 second-stage and 11 thirdstage caterpillars, of which all of the former and 9 of the latter were found to be parasitized. The results of the dissections are given in Table I.
TABiE I.-cults of di.sret~i(om of caterpillars of .1alacooima americatla par sitf(l-1ed by Limnnerium ralidum.
Number of Number of Number of Condition of
st1ge of host. eggs laid. eggshells larva, larva'. Other data.
Saeof host. eggs laid. foundll found, larv-.
Third ................. 1 1 1 Living.....
I)o ................ 1 1 1 .....do.....
l)o............... 3 3 2 1 living..... Dead larva only partially
I dead...... I larva already destroyed.
D)o ................ 1 1 1 Living.....
Second ................ 1 ...................................... I dead egg covered with amohocytes.
Do................ 1 1 1 Living.....
1o ................ 1 ............ 1 .....do...... Eggshell not found.
Do ................ I I I .....do.....
Third ................. 2 2 1 .....do...... I larva already destroyed.
Second............... 2 1 1 ..... do ...... I dead egg covered with anehwotes.
lo ............... 1 1 1 .... do.....
ThirdI ................ I I I .....do.....
)o ............. ....................................... .............. H ost not parasitized.
Io .. ...... ..... ..... 1 Living...... Eggshell not found.
I,... ..... I I. i 1 .....do.....
o................................... ....... ......... Host not parasitized.
I'.... I I I lIW ing.
ToI... (.7 hos.s) 19 .. .
BIOLOGY OF LIMNERIUM VALIDUM. 79
According to the dissections 19 eggs were deposited in the 17 caterpillars, distributed as follows: One caterpillar was parasitized lihree times, 2 twice, 12 once, and 2 escaped parasitism altogether. Of the 19 eggs deposited, 2 failed to hatch and became covered with amcebocytes, as did also the eggshells except in one doubtful instance, and in case of a third the embryo or larva was killed during the process of eclosion, becoming also densely encapsulated with amoebocytes. Two other larvu were killed after hatching, and were entirely destroyed by the action of the am(ebocytes, but their former presence was revealed by the empty eggshells. The remaining 14 larve were in a living and apparently healthy condition. Some were recently hatched, but others had grown to an appreciable extent. They lay free in the body cavity of the host, generally in the posterior half of the body, either above or below the digestive tube, and with the head directed caudad in respect to the host.
On the 7th of May, 5 caterpillars in the fourth stage were dissected, and in 2 of them was found a living first-stage larva; on the 11th, 10 more caterpillars were dissected, but none was parasitized; again on the 19th 2 caterpillars out of 10 in the fourth and fifth stages were found to be parasitized. One contained a living, first-stage larva of Limnei-bum, the other a living larva in the second stage, each host being about 20 mm. long.
EMERGENCE OF ADULTS OF LIMNERIUM FROM REPRODUCTION.
On the 21st of May, 2 unspun, full-grown larva- of Lim.neribm were discovered in the tray, and between that time and the 1st of June, 29 in all were found, 6 of which died before spinning cocoons. The rest spun perfect cocoons, and 6 passed through their transformations to the imagos, in every case males, from June 7 to June 14. The remaining 17 cocoons remained unissued up to December 13, when they were cut open and examined. All were found to contain dead larvae, some perfectly dry and shriveled, although 4 seemed only recently dead, as they were still plump although much discolored.
In all cases in this experiment where remains of the host were found associated with the larva or cocoon, the Linnerbtm issued from half-grown caterpillars, which were mostly in the fourth stage, although a few appeared to be small and stunted specimens of the fifth stage. On May 28 the remains of a host were examined. This caterpillar was killed by the parasite when about 25 mm. long, while .still in the fourth stage. It had a large, round ole in the integument on the ventral side, just back of the head, through which the larva escaped, after destroying all of the internal organs. The integument inside was left perfectly dry and bare, except for a small amount of broken-down tissue at the extreme posterior tip of the body.
ZSO REsU'LTS FROM-\ GIPSY MAOTH PARASITE LABORATORY.
LE-NGTH1 OF THE STAGES IN THE LIFE HISTORY OF LIMNERIUM.
From thbs land the preceding experiments we may estimate that the (tltat ion of the eijibryonic, development of Lim-neii ealidilm., or the ti iie fromwi oviposition to the hatching of the egg, is about 6 to 8 daiys: that the larval developmental period lasts for about 24 to 33 dav"N_: Iand that. the pupal period, or, more exactly, the time from -the SpTiningll of the cocoon to the eclosion of the adult, is about 13 to 22 dav- or much longer. The length of the larval period within the
cooowbfore. pupation was: not determined, but, judg0ing from the uwon~VXanilinedl onl December 13, it may be extended several months. ri'll( minimumii- time from egg to imago is thus about 7 weeks, or 50 days, whereas the maximum time to the eclosion of the adult may Ihe many months, due to quiiescence within the cocoon. This species hi1therto has 1)een considered to have a single generation a year, but thiis last experiment seems to indicatfe that. under certain conditions ait least it is able to pass through two generations annually. Under ioriii1al (iditionis, however, there is much doubt whether more than one 4rclierat ion aettially occuirs in the Northern States.
A POSSIBLY NORMAL PARASITE OF MALACOqOM.\A.
liis exp~erimelit shows that Lirnner;im~ ?alidumi is a possily normial l):lnasit&' of the tent caterpillar, although thsnvrbe on attacking fltat species in the field. The reason for this is obvious: 1lw to'iit caterpillars hatch early in the spring, long before the Urniitr~w leaves its cocoon, and therefore are not in season for this parasite. Fromt the fact, however, that nearly one-half of the caterpillars (Iis -ected InI this experiment we~re found to be pairasitized, and( that con1ll'ativelV fewv were killed by Lbnner~wm, we may conclude that L;,uucrb;ii tlyl;duwv is only partially adapted to this host. In the dissectiiltg wNork only dead larv-a' or eggs were discovered, but many iti'ire withltt iuiiicli (douibt succunbedl to the defensive reactions of thie liost. In the 42 caterp~illars dissected, ,23 eggs or larv'w were folot 11. and1( 29 ot her larva' were s1uCce-ssful ill passing throulghir Iiial Ilxlq)Iet Illki a total of only .52 pa.-rasites that came
ill ) icr oh ser-a t ion. I 4 the .5 femlales used inl this exp~erinwuet oNvi) )i I ed 1i a -c reelowver,'I n5 t hose inl the prece(ling expe(rimlenits, and there lS- Itorasl to believe othierwise soic 100 to 200 eggs or even ItIO)e Werjtol )allyj Iaido. laking 11( 100o as a conIserva~tive estimate, :111d1 wvit Ii (I1c ti UI owa ii1ce for thle (lest iuct ionl wroulght by thle (lissecti oii-. we( 111,1Y t111its figure, onI a m1or-tality of at least about 37 perI
-cIll. th'le c, st eggr~sl telkI were a 1 most i arabyfound1( thlickly enlOwIrthiel with a '114( 0oce. 011( t lie fit net ion of the latter wlas wilthlout 41)ht t s;i tu ~III he ust of thle browni-tail mloth cat erpil lars,
it) ivititegrratIc if iiot to kill the( youg lr\-axp In the present Iinstatiie,
BIOLOGY OF LIMNERlIUM VALIDUM. 81
however, a few of the larva' were able to withstand the action of the amoebocytes and reach maturity. This immunity of the few is of theoretical interest, as it would furnish a basis under proper conditions for the evolutionary development of complete host relationship. It is not difficult to conceive that this parasite in the course of not many generations might adapt itself so as to insure a minimiun mortality, and thus become an effective enemy of the tent caterpillar.
LIMNERIUM VALITI-M AS A PARASITE OF NOTOI )I1II S ANTIQI 1 TUS
One of the most interesting of the experiments was started May 9, 1910, with the rusty vaporer moth (Xotoloplot. antiqitus Linnaeus) as the host. One unfertilized female Limnrium was confined with 25 smqll, third-stage caterpillars. The female remained alive for 12 days and oviposited freely in the caterpillars.
Dissection work was commenced on June 17 and was continued at intervals until June 30. In all, 10 caterpillars. 1 pupa, and 7 moths were dissected. In the 10 caterpillars, a total of 40 eggs and eggshells was found, and 8 larvoe which were all dead; in the 1 pupa, 1 eggshell was found; and in the 7 moths, a total of 16 dead eggs and eggshells was discovered, making a total of 57) eggs laid by the single female in only 18 of the hosts. The results of the dissections are given mor, in detail in Table II.
This experiment proved conclusively that this parasite is not at all adapted to live at the expense of Xotoloph ts. The larvve, even if they were able to hatch, were killed by the host soon afterward, and at least 6 of the eggs were unable to hatch. Two other eggs were found with the dead larva only partially out of the shell. The larvae in most cases were entirely destroyed by the host, so that all trace of them was lost, except for the cast eggshell, yet a few resisted total disintegration for a long period. The body tissues of the latter. however, were broken down, so that nothing was left but the integument. The eggshells, or dead eggs with the embryo inside, resisted destruction, on the other hand, to a remarkable degree, and were found practically unharmed for a month or even longer after they were laid, not only in the body cavity of the caterpillar and pupa but even in that of the adult moth, and invariably well incased in a sheath of amcebocytes. The eggshell in the single pupa dissected was found adhering to the ovarian tubules of the host; in the case of the moths, the eggs or shells were found many times just beneath the integument of the abdomen, or among the ovarian tubules. One female moth, which was in perfect condition and in no way inconvenienced, carried 6 of these eggshells.
82 RESULTS FROM GIPSY MOTH PARASITE LABORATORY.
TABLE II.-Results of dssectfons of cterpilldlars and later stages of Notolophus
an tiqutus parasitized by Linneriaim validumn.
l o ~Num- Number NumDate of ber of of un- ber of Nurdissec- Stage and sex of host. erof of un- berof ber of Remarks.
eggs hatched egg- larva.
laid. eggs. shells.
May 17... Fourth-stage caterpillar. 2 2 ................ Embryos not noticed within
Do.......... do.................. 5 5 ........ ........ Do.
May A0... Fifth-stage caterpillar... 3 ........ 3 ........ No trace of larvae.
Io..... ....o................... 2 .......... 2 2 Larve much broken down.
June 14... Sixth-stagecaterpillar... 2 .......... 2 1 Larva killed before complete
Do..... Male moth .............. 1 .. .......... 1 No trace of larva.
June 13. .....do................... 1 1 .......Egg contained dead embryo.
D)o..... Female pupa ..... ...1 .......... 1.. ......Shell among egg tubes of host.
June 17... Sixth-stage caterpillar... 72 5 1 Larva broken down. Host dead.,
Junels... Male moth.............. 1 .......... 1 No trace of larva.
June 20)... Fifth-stage caterpillar... 8 .......... 8 1 Larva killed before complete
eclosion. Host dead.
Do..... .. (......... H......... .. ost dead. not parasitized.
Do..... .....do................... 6 ..........6. 2 Larve broken down; killed soon
after ecl Hosion. Host dead.
)Do..... Female moth............ 6 .......... f .t No trace of larv, s.
June30 ... ..... do ................... 1 1 ........ ....... Egg contained dead embryo.
Do........... do .................. 5 2 3 ....... Two eggs contained dead embrvos.
Do..... Male moth.............. 1 ........ No face of larva.
Do... Sixth-stage caterpillar.. 5 .......... 5 1 Larva all destroyed except skin.
Total.............. 57 13 44 8
LIMNERIUM VALIDUM AS A PARASITE OF PORTHETRIA DISPAR (LINNAEUS).
Two experiments were undertaken to determine whether this species of Limnerium can parasitize successfully the caterpillars of
the gipsy mnath (Porthetria dispar Linnaeus). Both were, unfortunatelyv, unsatisfactory in that the caterpillars were subject to disease and soon died in confinement. The second experiment only gave
any results at all, and was started June 10, when 1 female Limnerium was confined with 20 caterpillars in the third and fourth stages, collected at Melrose Highlands, Mass.
On June 17 the female Lintnerinum. was dead, and 19 of the caterpillars had succumbed to disease. These dead caterpillars were carefully dissected, and in 4 of them were found 18 eggs of Limnerbim,
oc u(rring 2, 3, 5, and 8 in number, respectively. The eggs were all unillatched, thie embryos undeveloped, and in some of the eggs the contents appeared to be broken down, such eggs being undoubtedly dead. Wletller they had died through contneact with dead and dise(aed"I( tissues of the caterpillars or had been killed previously by s(M ra(,10tion Of the ll(t is Iot clear. Inll experiments with browntail 11(tlI caterpillars earlier in the sea son, living eggs land even larva' were found within dead hosts; but in that case the caterpillars had not died from disease, but were killed by superparasitism.
()n tfhe 21st of Jumne, ( more of the (aterpillars had died. They
were dlis$e'(ed', and in one 5 eggs of L;mnwr;,eium, were found still unIhacild. ly .lllne 31 the remaining 5 caterpillars had succumbed
BIOLOGY OF LIMNERIUM VALIIDUM. 83
to disease, and the experiment was closed without any coinltsivye evidence being gained, although the inference may he drawn that Limneri4um ialidumin is not adapted to the gipsy inotih.
EXPERIMENTS WITi OTILERI HOSTS.
Several experiments were also started with caterpillars of the common tussock moth (IHemerocampa leuctostigmtta Smith anid Abbot) and of the mourning-cloak butterfly (Euranessa antiopa Linnaeus), but for various reasons they furnish no satisfactory evidence to show whether these hosts can be parasitized successfully by Limneri4tm, validum. It is rather unfortunate that the experiment with the Hemerocampa did not give any FIG. 33.-Limnerium validum: Egg. Enresults on account of disease prev- larged about 120 times. (Original.) alent among the caterpillars, in view of the fact that this species of Limner-iumn has been reared by Dr. L. O. Howard from the tussock moth at Washington, D. C. In Massachusetts, however, it has never been surely reared from this host, although large numbers of the tussock moth have been collected in various parts of the State and carefully studied at the Gipsy Moth Parasite Laboratory. It seems likely, therefore, that the Limnerium, varies in habits in different parts of the country, or this apparent difference in host relationship may be ascribable merely to
changes in its seasonal history brought about by a
NOTES ON DEVELOPMENT
AND ANATOMY OF THE
FIG. 34.-Limnerium validum: Eclosion of larva. The egg (figs. 33. 34) of
Enlarged about 120 times. (Original.) Limnerim calilm is elonLimnerwhm calidumni iseln
gate kidney-shaped or subcylindrical and rather convex on the dorsal side, slightly concave ventrally, with both poles bluntly rounded. The chorion is heavily chitinized, comparatively thick and resistant, and with a perfectly smooth surface. Its color is pale brown, or sometimes nearly white in the case of freshly deposited eggs, but not always, as fully colored eggs may be found in the oviducts of dissected females. In size the egg is about 0.35 to 0.41 mm. long and
0.13 to 0.14 mm. in transverse diameter.
84 RESULTS FROM GIPSY MOTH PARASITE LABORATORY.
Adult females freshly issued from cocoons were dissected on several occasions, and fully developed eggs to the number of 15 to 20 were found in the oviducts ready for oviposition. The female of this species therefore may perpetuate its' kind almost immediately after its eclosion from the cocoon (fig. 32). It may also be parthenogenetic at times, as eggs that were laid by unfertilized females in the preceding experiments hatched freely. Not enough evidence was gained to state positively whether unfertilized eggs always produce males, but this is probably true.
TIHE FIRST-STAGE LARVA.
DESCRIPTION OF TIHE NEWLY IIATCHED LARVA.
The first stage of the larva of Limncrium is characterized by 13 segments. including the comparatively heavily chitinized head, and a long tapering ventral appendage of the last body segment. In the case of newly hatched larvae
(fig. 35) the head is about
one-half as long as the body,
excluding the tail appendage,
and is bent to the axis of the
body at an angle of about
450. Its dorsal margin is
strongly curved, especially
anteriorly; the ventral margin is much shorter and only
slightly curved. The integuFlo. 35.-Limnerium validumn: First-stage larva, mIent of the head is rather newly hatched. Enlarged about 120 times. heavily chitinized and is Original) slightly pigmented with
brown. It has a slight ridge on each side running backward from the insertion of the mandibles, and separating the gula from the cheeks. The gula seems to be but slightly hollowed out and is as heavily armored as the rest of the head.
The mouthparts (fig. 37) consist of prominent, curved and sharply pointed mandibles. crossing each other at the tips, and projecting soinewhat downward into the large mouth cavity. The aperture of ti mouth is surrounded by a raised, chitinized, circular rim, about 0.037 nm1n. in d(ialmeter, and with a broad inner margin. The posterior inner margin is a heavily chitinized plate, with two prominent te eth separated by an angular median indentation.
lThe )ody in the thoracic region is nearly as broad as the head bult rapidly tapers posteriorly. The thin, delicate, transparent integu(inet i t lr()\wn illto f)lds (on the d)rsutin and venter, but appar-
BIOLOGY OF LIMNIERIt1' M VAL[DU l. 85
ently not on the sides. Each fold oil tle venter represents a body segment, though some may be more or less double; the folds on the dorsum, however, are plainly double, as two occur on each segtieiit. The body is thus found to be 12-segmente(1, excluding the tail appendage which is plainly a ventral outgrowth of the last segment.
The appendage itself sometimes appears to )e distinctly ritlged. due undoubtedly to the folding of the integument. It is slender, tapering, and about four-fifths as long as the body. I)urin enibryonic development it is bent sharply forward and appressed to the venter; after eclosion it is bent to the axis of the body for a short period at an angle of about 90', though later in life it extends straight backward. Its color and that of the body is transparent whitish.
The total length of the newly hatched larva is about 0.64 im.; without tail, only 0.41 nmi.; the width of the head is 0.11 inn. and that of the thorax 0.10 ram.
FUNCTION OF THE TAIL APPENDAGE.
In case of larve observed immediately after hatching, the tracheal system could be made out easily, and was filled with air without doubt, though
the closed or
one fine trache a 1 b r a n e 11 FIG. 36.--LimnCrium ralidut" First-stage larva of large size, showcould be d ing silk glands and nervous system. Enlarged 50 times. (Original.)
tinguished in the tail, and it was clearly not important enough to indicate that the tail is a tracheal gill. The function of the tail, however, is probably respiratory, and the organ might properlyy be termed a blood gill. There is nothing in its structure to contradict this view, as it is a simple, hollow tube lined with hypodernal cells, and undoubtedly filled with blood a greater part of the time. Since the larva lies free in the body cavity of the host it is constantly bathed in blood and lymph fluids, from which the oxygen of its own blood must be derived through the delicate integument of the tail, or other parts of the body, especially while still small.
APPEARANCE OF OLDER LARV--E.
The larva makes its escape from the egg by bursting open the chorion irregularly at the anterior pole (fig. 34)1 possibly with the aid of the mandibles. After hatching it develops rapidly if located in a suitable host, and molts for the first time in probably about 7 to 10
86 RESULTS FROM GIPSY MOTH PARASITE LABORATORY.
days. Before molting it attains a length of nearly if not quite 2 num., for some larve were found, evidently nearly full-sized, about 1.7 mm. long. It now differs considerably in appearance from newly hatched larvm, as a comparison of figures 35 and 36 will indicate. The head, being heavily chitinized, has remained the same size, but the body has grown until it is about five times as long as the head instead of only twice as long, as at first. The tail has also increased in size, but not proportionately to the growth of the body. Because of this rapid growth the folds of the integument so noticeable in newly hatched larve have largely disappeared. but are still discerni)le as slight creases. and afford the only means whereby the segments of the body may be distinguished.
NOTES ON THE ANATOMY OF THE LARVA.
The development of the viscera as made out in stained and mounted specimens is not without interest. Toward the end of the first instar
the sericteries become the most conspicuous organs in the body, thus forecasting the prime importance of the cocoon-slpinning habit as a protection during
.the pupal period. There is a pair of these silk
glands which seem to start blindly in the first body segment and extend backward after branching once to the eleventh or twelfth segment. In reality,
however, they are connected anteriorly with a minute tube which runs forward to the mouth. FIG. 37a.-Limnerium The glands themselves are also tubular and com"tialiu: Mouth- )posed of comparatively enormous cells, with large parts of first-stafe
lara. Enlarged oval nuclei.
abNt 250 times. The proctodaiium is also conspiciotis and extends (Original.)i
(Origina forward only to the anterior end of the twelfth
body >eginiit. It is a thick-walled, ovoid organ, and although cont igioiis with the posterior end of the mniesenteron. it does not coinmiiinicate therewith. At the anterior end it gives rise to four large Malp)ighian tibiles which extend forward into the ninth or even the eightli seinelnt. The anus appears as a distinct opening at the end of the twelfth seIgment on the dorsal side. thus proving that the tail is a ventral olutgroowth of the last seg(ilent. As the anus is distinctly open. there iv no reason why' the secretions of the Malpighian tubules may not he plased off into the blood of the host.
lht brain toward the end of the first instar is crowded partly out o(f the head and becomes a conspicuous organ of the first body se(gnwIlleit. T'Ihe samIlle l)henolllelion takes place in regard to the infraesophiageal ganglion. Twelve ventral ganglia may be distinguished, of w hichl the fir-t 3 are closely united to each other and to the infrnerphagnt ganglion, thw following 5 being epIaated by comnmissures, but the lat 2 are also conjoined.
BIOLOGY OF LIMNERIUM VALIDUM. 87
TIE SECOND-STAGE LARVA.
The larva after the first molt differs conspicuously from the firststage larva in respect to the soft, unarmored head, and the much shorter tail appendage (fig. 38). The mouth and its parts are also considerably changed.
The head is still large and prominent and has much the same general shape as in the first stage, although considerably shorter. Its
FIG. 38.-Limnerium v'talidum: Second-stage larva. Enlarged about 30 times. (Original.)
integotment is soft and pliable like that of the body. The mouthparts (fig. 39) consist of a slightly bilobed, simple labium, and two strong, curved mandibles, which project downward into the large funnel-shaped mouth cavity. The body is distinctly cylindrical and tapers but little behind. Twelve segments in the body may be easily distinguished, the last one of which is prolonged ventrally into -a short, conical tail appendage.
Only a few larvae in this stage were found, and they were all about 2.2 mm. long. The duration of this stage is probably short, perhaps not more than from 5 to 7 days.
THE THIRD-STAGE LARVA. Fi(;. 39. Liinneriurn
DESCRIPTION OF THE FULL-GROWN LARVA. validum: Mouthparts
of second-stage larva.
When the larva has obtained its full size and Enlarged about 100
leaves the host it is from 9 to 10 mm. long, and times. (Original.)
pinkish white in color. It is like the usual hymenopterous type of larva (fig. 40), and the body is rather deeply twelve-segmented. The head is comparatively small and inconspicuous, and the tail appendage, so characteristic of the first stage and still persistent in the second stage, is now entirely lacking.
The mouth parts (fig. 41) are decidedly different from those in the preceding stages. They consist of strong mandibles, supported by two short, longitudinal, heavily chitinized ridges, and two long, transverse ridges that extend nearly to the lateral margins of the
88 RESULTS FROM GIPSY MOTH PARlASITE LABORATORY.
lead. Below the mandibles is a more or less circular labium with heavily chitinized margins, supported by two short transverse ridges. The mouth opening is small and hardly distinguishable, but probably lies directly beneath the tips of the mandibles, toward which
Fia. 40.-Limneriun raldum: Third-stage larva. Enlarged about 8 times. (Original.)
point there is a gentle declivity. Just in front of the deeply concave, anterior end of the labium, in fact at the bottom of this concavity, is located the external opening of the sericteries. Above the mandibles are two large, circular, slightly pigmented spots, which undoubtedly mark the place
where the compound eyes
of the adult are beginning
FEEDING HABITS OF THE IMPIRISONED LARVA.
With a clear understand% ing of the mouth structures
in the three stages, we may
briefly consider the feeding
habits of the imprisoned
la rva. The old accepted
theory up to the time of
Ratzeburg was that inFra. 41. -fimnerium varlidum: Anterior view of ternal parasitic larvir feed head of third-stage larva, showing mouthparts. Ilpon the fat-body of the Enlarged about GO times. (Original.)
host. Ratzeburg (1844),
however, was forced to replace this theory with one that is probalbly more nearly correct. Hie concluded that such larve feed upon the lymph and )l]bxd of the host rather than upon any of
the solid tissues. This view seems especially applicable to the feeding llab)its of 1/tlpantles and related genera, which often leave the host in a living though comatose condition, but as the mlandibles are
BIOLOGY OF LIMNERIUM VALIDUM. 89
well developed in all three larval stages in the case of Limrnwiq hlm, we hesitate to apply it in entirety to this species. We do not believe, however, that the larvee of Limnerium while small actively attack and devour any of the solid tissues of the host including the fatbodies, but rather institute pathological changes whereby these tissues become .Available for food. The first tissues to be broken down are the fat-bodies, whereas the vital organs, including the digestive tube, resist dissolution until the last. The very apparent, early disappearance of the fat-body in parasitized caterpillars prolally accounts for the old theory that the parasite consumes it directly. The parasite certainly does consume the fat-body, but only after it has been broken down. We have often noticed this condition of the fatbody in apparently healthy caterpillars that harbored the early stage of almost any hymenopterous parasite. In such cases the fat-bodies were broken down more or less completely into their component parts, and the blood of the host was filled with the perfectly globular fat cells of varying sizes, and sometimes of a truly enormous size for individual cells. These small, solid particles, we believe, are as readily eaten by the larvw of Limnerbum, at least, as the fluid medium in which they float about within the body of the host. The mouthparts in the first and second stages of LimneriiTm are admirably adapted for this purpose, being in fact a sucking apparatus, with strong, pointed mandibles, so placed as to aid in swallowing small, solid particles. In the third stage the mouthparts are still essentially sucking, but as the mouth opening is surrounded by chitinized, supporting or possibly rasping ridges, there is some ground for believing that even solid tissues may now be taken in, even before they have been wholly disintegrated. When the parasite reaches this stage, the host has become somewhat weakened, its fat-body having been almost entirely, and its body fluids greatly, depleted. Tnlike the condition produced by some larva of tachinids at this stage in their development, the remaining tissues never dissolve into a putrid or semiputrid mass, but at all events remain clear and wholesome. To just what extent the tissues are broken down before being consumed has not been definitely determined, but we believe usually to a semiliquid condition in the case of Limnerium. A caterpillar forsaken by a larva of the Limnerbum was closely examined, and a small amount of tissue was found at the posterior end of its body. This tissue was probably the remains of the muscles of that region, and appeared to be of a mucilaginous consistency.
RESPIRATION OF THE LARVA IN THE THIRD STAGE.
The shortening of the tail appendage in the second stage and its entire disappearance in the third stage must necessitate a gradual change in the respiratory habit of the larva, if, indeed, the tail is a
90 RESULTS FROM GIPSY MOTH PARASITE LABORATORY.
truly respiratory organ, as we think it must be. This change is perhaps correlated with the more ravenous appetite of the parasite in the last two stages of its larval life,. and also with the gradual disappearance of the blood and lymph of the host. With the disappearance of the fluids of the host, the tail as a blood gill must necessarily become useless, as it is fitted for life in a fluid medium only. Nor does it seem possible, for much the same reason, that the larva's whole supply of oxygen is gained by osmosis through the integument of the body itself, for as the larva grows older the integument becomes thicker and tougher, especially in the last stage. The only alternative left is to consider that the oxygen is derived from the comparatively enormous amount of food taken in during this period, and that it is absorbed by the blood of the larva through the walls of its digestive tube. In other words, if the larva stopped feeding it would not only starve but also suffocate. Toward the end of the third stage, however, when the host is nearly or possibly not entirely consumed the stigmata become open, and the larva is able to breathe air directly, as it certainly does after leaving the host to spin its COCOOn.
The cocoon has been aptly described by Dr. L. O. Howard (1897) in his bulletin on the parasites of the white-marked tussock moth, and we take the liberty of transcribing his words here: The cocoon is rather long ellipsoidal, averaging 7.5 mm. in length by 2.8 mm. in greatest diameter. It is composed of two distinct coverings, the outer one of weak, close-spun, crinkly, gray, or grayish-brown silk, readily peeling off in a sheet, and the inner one close, tough, parchmentlike, dark brown in color, with golden reflections, of the type common among the Ophioninw."
RESULTS OF THE DISSECTIONS OF HYPHANTRIA DURING THE FALL OF 1910.
During the late summer and the fall of 1910 a large series of collections of the fall webworm (Hyp/atria cunea Drury) was brought together at the Gipsy Moth Parasite Laboratory, and an extensive study made of its parasites by means of dissection and rearing work. Most of the work was conducted bv Mr. J. D). Tothill, who kindly turned over to the writer, together with his notes, the series of larve of Limnberium that were found during the course of the dissections. Inasmuch as another species of Limer ium. is found attacking the fall webworm inII Massachusetts rather abundantly, the collection of larva' may have included L. pilosulum. as well as ralidum, but the most careful scrutiny of the larva, all of which were mounted in balsaon and inl the first stage, failed to reveal any characters to; sepa-
BIOLOGY OF LIMNERIUM VALIDUM. 91
rate the series into the two species, so that somIe dotibt remains whether L. pilo8ulunt was actually represented.
The two most important lots of the larva were obtained respl)ectively on August 18, from third-stage hosts collected the day previously at Reading Highlands, Mass., and on September 6 to 9, from mostly sixth-stage hosts collected near the laboratory at MeI"ose, Highlands. The former lot may have been L. pilos'lurm, as this species spins its cocoon inside of the skin of small hosts and issues therefrom throughout the month of September. The latter lot of larvw seems to have been L. validtum without any doubt, as they were in the first stage when L. pilosulum was already beginning to issue as imagos, and inasmuch as they were found in rather large sixthstage hosts. It seems reasonable to conclude, therefore, that the firststage larvae of these two species are practically identical in structure and appearance.
The disposition of the larvae in their natural host is not without interest. According to Mr. Tothill's notes, they were found free in the body cavity, between the walls of the body and the alimentary canal, either inferior or superior to the latter. No indication of amcebocytosis was encounteredduring the course of the dissections.
Limnerium, validum (Cresson), a common parasite of the fall webworm (Hyplantria cunea Drury), readily attacks the caterpillars of the brown-tail moth, gipsy moth, and rusty vaporer moths, and also the tent caterpillar, when placed in confinement with these hosts, but is able to complete its transformations in the last species only and even then in but a small percentage of cases. Its larve seem to be totally unadapted for life in the caterpillars of the three former species, and fail to survive the protective reactions of the host, which are visibly manifested by an accumulation of active blood cells or amcebocytes around the larve, the cast eggshells, and even the eggs themselves. The amcobocytes presumably attack the living eggs and larve, or at least ultimately efface the latter entirely. The same reaction takes place in the case of the tent caterpillar, but a few of the larva are able to complete their transformations. Adaptation here is partially in evidence, and may be due to larval secretions which ward off the protective reactions of the host.
The egg of Lifmnerium has a thick, chitinized chorion, which resists decomposition in the body fluids of the host to a remarkable degree. The first-stage larva is characterized by a long, tapering tail appendage, evidently adapted for a respiratory function and acting as a blood gill. The second-stage larva has a much shorter appendage, and the third-stage larva has none, so that respiration very likely occurs through the walls of the digestive tube, oxygen
92 RESULTS FROM GIPSY MOTH PARASITE LABORATORY.
being absorbed from the constantly ingested supply of fresh tissues of the host.
The larva while still small assumes a more or less definite position in the body cavity of the host, and'generally lies toward the posterior half of the body, either above or below the digestive tract and with its head directed caudad in respect to the host. During its whole life the larva feeds on blood and lymph and on small solid particles which result from the disintegration of the host's tissues, probably pathologically induced by some larval secretion. There is no evidence to show that such definite organs of the host as the digestive tube and muscular tissue can be consumed by the larva unless they are broken down, inasmuch as the mouthparts of the larva throughout life are essentially sucking.
Under artificial conditions the minimum time needed for the developmnient of the insect from the egg to the adult was found to be about 50 days, but the maximum time may be extended many months. Under natural conditions. in the Northern States as a parasite of [lyphantia, the females are probably active in parasitizing the caterpillars throughout the month of August and the first part of Septemiber. The larvw issue from the caterpillars and spin their cocoons during September and the first part of October, but the cocoons always overwinter, and the adults emerge the following summer.
1864. ('nussoN, EI. T.-Proc. Ent. Soc. Phila., vol. 3, p. 258.
1879. PIOVANCHER, L.-Nat. Canad., vol. 11, p. 174, tig. 9, a. 18 3. PROVANCIHER, L.-Fatuna Ent. Can. Ilymn., p. 367, fig. 44, a. 18I90. ASuMEAD, W. II.-Proc. F. S. Nat. Mus., vol. 12, p. 429.
1844. ItATZEBU, F. T. C.-Die Forstinsecten. 189Is. CUENOT, L.-l tudes physiologiques sur les Orthoptcbres. (Phagocytose.)
Arch. Biol., vol. 14, pp. J315-425.
1897. Howain, L. O.-A study in insect lmparasitism. Tech. Ser. 5, Bur. Ent.,
U. S. D)ept. Agr.. p. 23, fig. 10.
1!$. JANET, (C.-Itemplaeemnent des muscles vibrateurs duii vol par des colonnes
d'Adipocytes, ciez les fourmis nuprbs le vol nuptial. C. Rt. Acad. Sel.,
vol. 142, pp. 1095-1098.
1107. JANET, C.-I IIstolyse suns phagocytose, des muscles vibrateurs dui vol,
c(ez des relies des fouris. ('. It. Acad. Sci., vol. 144, pp. 393-390.
1910. P'ANTEL, J.-IPcherches stir les I )iptres a lrves entomIobles. (Manlfestationus Oventuelles de la r6action defensive.) La Cellule, vol.
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