Title: Legume inoculation in Florida
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Title: Legume inoculation in Florida
Series Title: Legume inoculation in Florida
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Creator: Hubbell, D. H.
Publisher: Florida Cooperative Extension Service, Institute of Food and Agricultural Sciences, University of Florida
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Full Text
Circular 376


Legume Inoculation


In Florida


HmH^^^-
o4 UIBRARY
D e Thornton
VLj L U


Florida Cooperative Extension Service
Institute of Food and Agricultural Sciences
University of Florida, Gainesville






















D. H. Hubbell, Assistant Professor (Soil Microbiologist), Department of
Soil Science
D. Jones, Professor, (Agronomist), Department of Agronomy
G. D. Thornton, Professor, (Soil Microbiologist, Retired), Department of
Soil Science







Cover Photo Soybean roots with good nodulation by efficient legume
bacteria.







What Does Legume Inoculation Mean?
The term "legume inoculation"
refers to the practice of introducing
specific bacteria into the soil with
legume seed, commonly at the time
of planting. The bacteria used
belong to the genus Rhizobium,
but they are commonly referred to
as "rhizobia," "legume bacteria" or
"root nodule bacteria". This last
term is particularly apt as it de-
scribes a visible and characteristic
effect of the bacteria on the plant.
These bacteria begin to infect the
roots of leguminous plants within a
few days after seed germination.
Within one or two weeks, small
"nodules" appear on the roots as a
result of these infections. The
mature nodules may be round, cy-
lindrical, or lobed, depending on the
plant species.
What Is Successful Inoculation?
The presence of these nodules on
the roots of legumes is proof that
the bacteria have successfully in-
fected the roots. This is of great
importance since the rhizobia living
in the nodule are able to enter into
a "symbiotic" or mutually bene-
ficial association with the plant.
The plant and the bacteria, acting
together, are able to reduce or "fix"
nitrogen gas (N2) from the atmos-
phere into ammonium (NH4+) in
the plant root. In this form it can
be readily used by both the bacteria
and the plant as a nutrient for
growth. The presence of a pink
color within the nodule indicates
that nitrogen is being fixed. This
can be observed by crushing or
slicing nodules which are approxi-
mately one week old. Very young
nodules will have a white interior
and very old nodules will have a
brown interior color. Little or no
nitrogen is fixed in these nodules.


Why Is Inoculation Important?
The ability of the plant-bacteria
combination to "fix," and utilize
an otherwise unavailable form of
nitrogen is of considerable econom-
ic importance to farmers. It sup-
plements soil nitrogen, which is low
in the majority of soils. This is
especially true for the mineral soils
of the coastal areas of the South-
east, where leaching and erosion
have severely reduced the supply of
soil nitrogen. Nitrogen is a key
element for every living cell because
it is an essential constituent of
protein, which is the building block
for plant and animal tissue.
Nitrogen is the plant nutrient
which is most often limiting to crop
growth. There are two main rea-
sons for this observation. First,
plants have a higher demand for
nitrogen, as compared with the
other required plant nutrients, in
order to achieve good growth. Sec-
ond, nitrogen present in or added to
soil is constantly being lost through
run-off, leaching, removal of nitro-
gen-containing crop residues from
the soil, and denitrification. "Deni-
trification" is the process, carried
out by some soil bacteria, of con-
verting plant-available nitrate ni-
trogen to atmospheric nitrogen,
which then diffuses as a gas.to the
atmosphere. Thus, "dentrification"
results in a loss of soil nitrogen and
is therefore opposite in effect to the
process of "nitrogen fixation" car-
ried on by the Rhizobium-legume
association.
Soil nitrogen is replenished pri-
marily from two sources, fertilizers
and nitrogen fixation. Chemical ni-
trogen fertilizers may be in the
form of non-protein organic sub-
stances such as urea, or inorganic
materials such as ammonium ni-






trate or ammonium sulphate. Due
to the nature of the processes in-
volved in their manufacture, chemi-
cal nitrogen fertilizers are rather
expensive. Nitrogen applied in
these forms is available immediately
for plant uptake, but the portion
not used by the plant soon after
application is more subject to loss
by run-off, leaching, and denitrifi-
cation.
Properly inoculated leguminous
plants convert atmospheric nitrogen
into organic nitrogen of plant pro-
tein. Large quantities of nitrogen
may be added to the soil by the in-
corporation of the high-nitrogen
residues of these plants. This or-
ganic nitrogen is at first unavailable
for plants. The organic residues
must first be decomposed by numer-
ous soil microorganisms. This re-
sults in a slow but continuous
liberation of inorganic nitrogen
which may be used by growing
plants as it is released, thereby
reducing the amount of nitrogen
lost through leaching.
Do All Leguminous Plants
Provide The Same Benefit?

A leguminous plant is one that
bears its seed in pods. Beans, peas,
soybeans, lupine, peanuts, and cow-
peas are common legumes with
prominent seed pods. Other plants
belonging to this family with seed
pods not so conspicuous are clover,
lespedeza, beggarweed, crotalaria,
hairy indigo, trefoil, alfalfa, and
vetch.
Almost all of the leguminous
plants have the ability to form
nodules when in association with
the proper kind of root-nodule
bacteria. Some of the few excep-
tions are honey locust, redbud, and
Kentucky coffee tree.


Leguminous plants differ with
respect to the amount of nitrogen
they return to the soil. Obviously,
where such crops as peanuts are
harvested for nuts and hay little
nitrogen may be added to the soil.
Frequently such crops actually de-
plete the soil of some of its nitrogen,
because the nodules may fail to fix
all of the nitrogen that is needed by
the plant. Other crops, such as
soybeans harvested for seed, or
lespedeza harvested for hay with a
sizable portion of the plant left on
the land as stubble, may fix as
much nitrogen as that removed in
the harvested crop, leaving the soil
nitrogen at its original level. Still
other crops, such as clover pastures
and hairy indigo for grazing, may
add a considerable amount of nitro-
gen to the soil. Mature animals
on a legume pasture leave most
of the nitrogen on the land. The
largest increases in soil nitrogen are
obtained when the entire plant is
returned to the soil as full grown,
succulent green manure.
Some excellent green manure
crops, with the approximate a-
mounts of fixed nitrogen that can
be expected per acre, are as follows:
lupine, 75 pounds; crimson clover,
80 pounds; hairy indigo, 70 pounds;
crotalaria, 72 pounds; cowpeas, 86
pounds.


How Is Successful Inoculation
Accomplished?
Many years ago legume bacteria
were introduced into new areas by
using a quantity of soil from a field
where the desired leguminous plant
had grown successfully the previous
season. This laborious process is
now unneccessary. Commercial in-






oculants of legume bacteria are now
commonly available from seed deal-
ers. These inoculants are a mixture
of strains especially selected by the
manufacturer for their ability to
"fix" large amounts of nitrogen in
association with a particular legume
or legumes. This practice helps to
insure that different legume species
or varieties of the same Cross In-
oculation Group (or the same le-
gume grown in different localities,
where growth conditions such as
temperature, moisture, pH, nutrient
status, etc. may vary) can be suc-
cessfully inoculated with the same
inoculant.
Each container of commercial
inoculant bears a written set of
instructions for its use. Check the
following items carefully: The age
of the culture as indicated by the
date on the container; the list of
plants the culture will satisfactorily
inoculate; the quantity of seed for
which the contents of the container
are intended; and directions for
applying the inoculant to the seed.
Purchase the inoculant from a
reliable dealer who can assure you
that the material is fresh and has
been stored in a cool place in the
warehouse. Inoculants are PERISH-
ABLE. The legume bacteria are
killed by excessive drying and high
temperatures. For this reason the
expiration date does not always in-
sure a strong and viable culture.
Therefore, not only the AGE of the
culture, but also the STORAGE
CONDITIONS prior to inoculation
(in the warehouse and on the re-
tailer's shelf) are vital factors to be
considered.
When the seeding date is close at
hand, it is a good idea to purchase
the seed and a sufficient quantity of
inoculant at the same time. This


will insure having the culture on
hand when weather conditions are
favorable for planting. It is the
purchaser's responsibility to check
the container carefully at the time
of purchase to be sure he is getting
the proper inoculant. Otherwise,
an extra trip to the seed store will
be necessary; or the seed will be
planted with the all-too-common
"I'll take a chance this time" atti-
tude. The newly purchased inocu-
lant should be stored in a refrigera-
tor or an air conditioned room if it
is not used immediately.
NOTE: TWICE the manufac-
turer's recommended amount for
large seeded legumes and FIVE
TIMES the recommended amount
for small seed have given more
satisfying results under Florida's
soil and climatic conditions. Hot,
dry weather kills many of the
legume bacteria. Using extra in-
oculant, under cool, moist, condi-
tions if possible, is cheap insurance
against inoculation failure.
Care of Inoculated Seed. Inoculated
legume seed should be planted as
soon as possible after the inoculant
has been added. They should never
be exposed to direct sunlight or
allowed to dry before planting.
Never inoculate more seed than can
be conveniently planted within a
few hours. If there is any delay in
the seeding operation for more than
two or three hours, keep the seed
moist and protected from unfavora-
ble temperatures. If the seed be-
come excessively dry for any reason,
REINOCULATE. It will pay.
Seedbed Preparation. Inoculated
seed should be planted in a well
prepared and moist soil. To insure
quick germination of the seed and
to prevent the loss of inoculant by






slow IWO-- -


Fig. 2. Portion of experimental area where individual strains of legume bacteria are being
tested for their nitrogen-fixing ability under Florida conditions.


drying, roll the planted soil with a
cultipacker equipped with a seeding
attachment.
Frequently it is necessary to plant
clover seed in a grass sod. This is a
very hazardous condition for secur-
ing good germination and satisfac-
tory inoculation. The chances of
obtaining a good "catch" of in-
oculated clover on such areas will
be increased if the grass is closely
clipped and the surface of the soil
scarified by light disking just prior
to or at the time of seeding. It will
be beneficial if the seeding can be
timed so as to coincide with a per-
iod of light rains and cloudy skies.
The rain will keep the ground moist
until the seedlings are established
and nodulation has taken place.


Seed Treatments and Fertilizers.
Many seed treatment chemicals (pes-
ticides) have been found to be
toxic to legume bacteria. Although
some of these chemicals may be
compatible with legume bacteria
the evidence to date indicates that
these chemicals and inoculant
should not be used together at
planting. If it becomes necessary to
plant treated legume seed in large
scale operations, the soil may be
"preinoculated." This is accom-
plished by mixing the inoculant,
moist soil, moist sawdust, or other
inert material and sowing or drilling
the mixture in advance of seeding.
This may be carried out on plowed
fields at the last harrowing for weed
control and final, preparation just






prior to seeding.
Inoculated seed should not come
in contact with caustic lime, acid
fertilizers, or fertilizers containing
micronutrients as chemical salts. It
is permissible to allow them to mix
in the drill spout with such mate-
rials as superphosphate and basic
slag. Ordinarily, concentrations of
fertilizer in the soil that do not
affect seed germination or injure
the roots of seedlings will not harm
the root-nodule bacteria, but mix-
ing of fertilizer with inoculated
seed should be avoided.

HOW IS THE PROPER
INOCULANT CHOSEN?

Several species of leguminous
plants may be effectively nodulated
by a single species of root-nodule
bacteria. The groups of plants
nodulated by different species of
bacteria are called CROSS INOCU-
LATION GROUPS. Containers of
commercial inoculant are clearly
labeled with the proper Cross In-
oculation Group designation. It is
the responsibility of the customer
to request inoculant for the Cross
Inoculation Group which includes
the legume he wishes to plant. The
more commonly recognized groups
are as follows:


Alfalfa Group

alfalfa
sweetclover
bur clover
black medic

Cowpea Group

cowpea
crotalaria


lespedeza
peanut
kudzu
alyce clover
beggarweed
velvet bean
lima bean
pigeon pea
partridge pea
mung bean
hairy indigo
Aeschynomene
Stylosanthes humilis


Lupine Group

blue lupine
yellow lupine
white lupine
serradella

Soybean Group

soybean

Clover Group

Berseem clover
white clover
red clover
crimson clover
subterranean clover
other true clovers

Pea and Vetch Group

field pea
garden pea
Austrian winter pea
common vetch
monantha vetch
sweet pea
rough pea
(Singletary, Caley)
tangier pea







Bean Group

garden bean
scarlet runner bean
Trefoil Group

birdsfoot trefoil

Frequently strains of bacteria
effective on one member of a group
are ineffective on other members of
the same group. An ineffective
strain of bacteria is often termed
"parasitic", meaning that it will
nodulate a particular host plant but
will fix little or no nitrogen. Good
examples of this may be pointed
out in both the clover and trefoil
groups. In the clover group many
effective strains of bacteria isolated
from crimson clover are ineffective
on white clover. The converse is
also true. Bacteria isolated from
either big trefoil or birdsfoot trefoil
may be totally ineffective on the
opposite host. However, within
each group there are strains of bac-
teria which are known to be effec-
tive on one or more of the members.
Therefore, satisfactory group cul-
tures can be prepared by combining
the strains which have been proven
effective on all members of the
group in question.

WHAT ARE SOME POSSIBLE
CAUSES OF INOCULATION
FAILURE?

1. Use of wrong inoculant. It is the
buyer's responsibility to be certain
of the type of inoculant he needs,
to request this inoculant specifically,
and to ascertain that he has received
what he requested. If in doubt
concerning the type of inoculant
required he should consult his Coun-


ty Extension Director.
2. Unfavorable soil conditions.
In general, legumes will nodulate
and fix nitrogen best when grown in
soil which will permit normal plant
growth -- with the exception that
soil nitrogen may be quite low. In
other words, conditions (other than
high soil N levels) which favor
plant growth also favor the proc-
cesses of nodulation and nitrogen
fixation.
3. Use of "dead" inoculant. In-
oculant as purchased by the farmer
may be "dead" due to faulty man-
ufacture, although this is unlikely.
It is more likely to be the result of
poor shipping or storage conditions
prior to purchase. In particular,
exposure of the inoculant to high
temperatures even for very brief
periods of time, or exposure to
direct sunlight may kill so many of
the bacteria as to severely reduce the
success of inoculation.
4. Faulty handling of inoculant
after purchase. Inoculant must be
transferred and stored under cool
conditions after purchase. Careful
attention should be given to follow-
ing proper inoculation procedure
as previously outlined.
NOTE: Excessive heating or drying
of the inoculant at ANY time prior
to infection of the seedling will
result in inoculation failure.

WHAT BENEFITS ARE DERIVED
FROM LEGUME INOCULATION?

Improved Quality of Forage. The
protein content of pasture and hay
is increased by the presence of
inoculated legumes.
Increased Animal Gains. Higher
protein content and often improved
palatability with legumes results in
increased consumption of total di-






gestible nutrients by the animal.
Compared with nonleguminous for-
ages the leguminous plants as a class
are higher in calcium, phosphorus,
and other elements essential for
growth.
Added Soil Nitrogen. From 50
to more than 100 pounds of nitro-
gen per acre may be added to the
soil as a result of growing an ef-
fectively inoculated legume crop.
The exact amount added will de-
pend on the effectiveness of the
fixation process, soil reaction, phos-
phorus and potash levels in the soil,
amount of nitrogen already avail-
able in the soil, size of the crop, and
portion of the crop left on the land.
Increased Subsequent Crop
Yields. Yields of corn and other
crops have been doubled and tre-
bled by growing them in rotation
with leguminous green manure
crops.
Improved Soil Conditions. Pro-
perly inoculated legumes are rich in
nitrogen, often containing several
times as much of this element as
was withdrawn from the soil. Due
to the presence of certain high-
nitrogen chemical compounds in
leguminous plants, they decompose
rapidly, leaving finely divided or-
ganic matter in the soil which im-
proves its physical, chemical, and
biological condition.

Reduced Erosion. Legumes in-
oculated with effective strains of
root-nodule bacteria thrive in soils
poor in nitrogen. If adequate
amounts of lime, phosphate, and
potash are present, water and soil
losses may be reduced to a mini-
mum on poorer and more easily
eroded areas by growing legumes.
Properly inoculated kudzu, beggar-
weed, perennial lespedeza, hairy


indigo, and crotalaria are excep-
tionally well adapted for this pur-
pose.

OTHER FACTORS IN
GROWING LEGUMES

Although proper inoculation is
the key to economic production of
legumes, it alone cannot assure
success. Other essential factors are:

1. Good seedbed preparation.
2. Use of lime where needed.
(Usually not required for legumes
of the Cowpea Group).
3. Adequate supply of phosphorus
and potash.
4. Use of micronutrients where
needed. Many Florida soils fail to
support satisfactory growth of le-
guminous plants without supple-
menting the regular fertilizers with
either copper, manganese, zinc, bor-
on, or a combination of several of
these essential micronutrients, such
as F.T.E. Frit 503 (applied at a rate
of 20 lb/acre, pre-mixed with one
ton of lime before broadcasting)*.
5. Selection of the kind and variety
of legume that is best suited for the
existing soil conditions and locality
within the state.

CURRENT INFORMATION ON
LEGUMES AND INOCULATION
Additional information on this sub-
ject is contained in USDA Farmers'
Bulletin No. 2003 entitled "Legume
Inoculation: What It Is -- What It
Does" by L. W. Erdman. This
publication may be obtained at a
cost of $0.10 per copy, prepaid,
from the U.S. Government Printing
Office, Public Documents Depart-
ment, Washington, D.C. 20402.







There are several current publica-
tions on the growing of various
legume crops in Florida. The pub-
lications listed below may be ob-
tained by writing to: Institute of
Food and Agricultural Sciences, Bul-
letin Room, G044, University of
Florida, Gainesville, Florida 32601.


Soybeans in Florida
Bulletin 716
Floranna Sweet Clover
Circular S-46
Big Trefoil A New Pasture Legume
Circular S-49
Inoculated Legumes in the Farm
Program Circular S-61
Hairy Indigo
Circular S-98
Ritchey An improved Seed Pro-
ducing Variety of Bitter Blue
Lupines Circular S-143
Blue Lupines for Grazing and for
Soil Improvement in Florida
Circular S-146
Florida 66 Alfalfa, An Improved
Variety for Well Drained Soils in
Florida Circular S-191
Peanut Production Guide
Ext. Circular 145C
Hairy Indigo as a Cover Crop in
Florida Citrus Ext. Circular 227
Soybean Production Guide
Ext. Circular 277A


*The use of trade names in this publi-
cation is solely for the purpose of pro-
viding information. It does not constitute
an endorsement of the products named
and does not signify that they are ap-
proved to the exclusion of others of
suitable composition.




































COOPERATIVE EXTENSION WORK IN AGRICULTURE AND HOME ECONOMICS
(Acts of May 8 and June 30, 1914)
Cooperative Extension Service, IFAS, University of Florida
and United States Department of Agriculture, Cooperating
Joe N. Busby, Dean


June 1972




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