A M . ZOOLOCIST, 12:577-587 (1972).
The Fungus-culturing Behavior of Ants
NEAL A. WEBER
Biology Department, Swarthmore College, Swarthmore, Pennsylvania 19081
SYNOPSIS. A colony of attine ants begins with a recently fecundated female carrying
hyphae from the parental garden in a pellet in an infrabuccal pocket. All future food
of the colony will be derived from this nucleus. She digs a cavity in the ground, ejects
this pellet and manures it with her liquid excrement. As the hyphae proliferate, eggs
are laid on them and the colony is launched. She continually licks both the hyphae
and the brood. Thus, both salivary and anal excretions play a vital role in the
beginning of a colony and this pattern is repeated by the resulting workers. About
60-65% of them in Atta are the minima and these are intimately involved in brood
and fungus care. Their excretions are disproportionately large. About 1/3 of the
workers in Atta are 4-6 mm media and these cut and prepare the substrate. The 7-9
mm maxima sizes and the soldiers (over 9 mm) are less directly involved in culturing
the fungus.
The effectiveness of fungus culturing is shown by the rapid build-up of gardens.
The ants maintain their garden despite surrounding contamination after a fragment
with ants is introduced to a plate of sterile nutrient agar.
A new colony of attine ants begins with
a recently fecundated female (Figs. 1, 2)
who has left her parental nest with a bit of
the fungus garden. This is usually in the
form of fragments of t'he mycelium, short,
irregular threads that are stored in the
infrabuccal pocket, a blind pouch off the
fore part of the pharynx. In all cases it is
the vegetative form of the fungus that the
female takes, in contrast to the adventitious
spores of the fungus in some beetles and
termites; no spores are present in the
healthy garden. It is not a case of pieces of
the mycelium adhering to the body of the
female or taken between the mandibles as
she leaves t'he nest, for such ants have
been carefully examined to determine
these points. Were the fungus to be held
between the mandibles it would prevent
the ant from digging into the soil. The
fungus is destined to become t'he sole food.
After she has dug into the soil she
ejects the contents of the infrabuccal pocket as a pellet. This initiation of the attine
garden was reported by Huber (1905) in
the widespread Atta sexdens of Brazil.
Goeldi (1905) and earlier von Ihering
(1898) studied the same species. Investigators in general concentrated their studies
on Atta because of its conspicuousness and
economic importance. The size of the
Atta queen also makes for ease of study.
However, the fact that the attines represent a compact tribe of a dozen genera
and some 200 species showing considerable
evolutionary divergence, suggested to me
that a comparative study of the genera
would be worthwhile. This was first performed in 1934-35 in Trinidad (Weber,
1945) and followed by similar studies in
Guyana (then British Guiana) in 1935 and
1936 (Weber, 1946) and in 1938 in Panama (Weber, 1941). Numerous studies followed in later years and verified the Atta
method of colony formation in the other
and much smaller genera.
The ejection of the infrabuccal pellet is
followed by the manuring of it with clear
amber fecal droplets. In different genera,
the female, when she lacks a viable pellet',
will gather together soil grains and manure
them in a vain attempt to start the garden. Females of Acromyrmex lundi in Argentina, immediately following the nuptial
flight, were confined in the same container and ejected their pellets close together, thus forming a temporary communal garden.
When the pellets contain viable bits of
The author has been aided by N'SF Grant GB
the hyphae of trie fungus they start
31131.
577
578
NEAL A. WEBER
FIG. 1. Female of Alia sexdens L. on her young
garden and tended by her first broods of workers.
They are in continual attendance on her. By lick-
ing her frequently they may acquire phcromoncs
that are vital to the integration of llie colony.
growing and in a day or two the fungus
shows as a halo around the dark ejecta.
The female then lays eggs on it.
There are, thus, two critical behavioral
acts in the continuum between one attine
generation and the next: the female must
leave the parental nest with a viable bit of
fungus and she must start the fungus
growing by manuring it. The best place for
the new garden usually is a cavity in soil,
and the ant' will dig down for a distance
characteristic of the genus and then enlarge a chamber off the side of the end,
rather than at the end, and start her garden and colony here. There are variations
among the genera and species and in one
widespread species it is a yeast' form of the
fungus, but the important fact is that all
have the behavioral pattern outlined
above and this is the unique feature of
at tines.
The workers have extensive salivary
glands and gastric enlargements (Paparo,
1971; Martin and Martin, 1971) that enable them to carry on the next most critical behavioral pattern, their own manuring and care of the ant fungi. The female
has these glandular characters also but her
worker offspring, no matter how disproportionately small they are, possess these.
An egg-laying female of Alia ccphalotes
may weigh 346 milligrams and her workers
with lengths of 1.6 to 12 mm will weigh
O.i2 to 90 mg. Females of other species of
At la may be smaller, that of fhe egg-la) ing
colombica tonsipes averaging 172.2 nig.
While the largest of the workers are the
soldiers (Fig. 3), these will not be U.MJUI
FL'NGUS-CULTURING BEHAVIOR OF ANTS
5f9
FIG. 2. Underside of head of female of Atta
cephaloles L. showing structures involved in fungus
culture. The apices of the antennae appear to
carry the receptors chiefly used in identification
and care of the fungi and in evaluation of the
garden condition. The antennae are kept immaculate by drawing them between the strigil or comb
on each fore leg and the adjacent tarsal hairs. The
mouthparts are used for actual feeding and lacerating the fungal hyphae and for applying salivary
or pheromonal excretions. Glands discharge at the
base of them; their main mass may be in the head
or thorax.
FIG. 3. A soldier of Atta cephaloles surrounded by
various worker sizes from the author's laboratory table (photo. C. W. Rettenmeyer). A live
soldier may weight 90 mg and the smallest worker
0.42 mg.
580
NEAL A. WEBER
FIG. 4. Large worker of Atta sexdens using its
antennal apices to determine whether a fungus
culture is suitable to eat. It was. Had it been an
alien fungus it would have retracted the antennae
and walked away. Many ant species have been
tested with one another's fungus.
in care of the fungus and brood. The
smaller workers (Fig. 3) down to 1.6 mm
(the 0.42 mg size) have effective roles according to their size. Those of 1.6-3 mm
(the minima), and making up 60-65% of
the population of a mature garden in
Atta, tend the fungus and the brood, those
of 4-6 mm (the media) cut sections out of
leaves and reduce them to smaller sizes.
They excavate soil and are the general
workers of the nest, comprising about 1/3
of the population. The maxima workers
(7-9) (Fig. 4), about two per cent, and
the soldiers (over 9 mm), less than one per
cent, make up the remainder of the adults
of the colony except when a sexual brood
is being raised.
Culturing and maintaining fungi are exacting tasks for human beings and usually
require sterile conditions, to keep out contaminating organisms, and proper humidity and temperature. It is a hazard to add
new substrate and all too often results in a
few spores or a single spore of a more
vigorous "weed" being introduced and
ruining the culture. The task may be made
easier sometimes by adding an antibiotic to
the nutrient medium but' still the maintenance indefinitely of a culture is not usually attempted without repeated transfers.
Another risk is mutation, so that after a
time the culture has evolved into something else than what was first created.
Culturing and maintaining the ant garden appears to be a much easier task for
the ants to perform, but their behavior
contributes significantly to their success.
This behavior includes the following: (1)
Constant grooming of themselves and each
other so that the ants are immaculate.
Licking with the mouthparts adds salivary
excretions that may at the least be bacteriostatic and fungistatic. (2) As the cut
leaf sections are brought on the trails back
to the nest, the smallest workers often attach themselves to the section carried by a
much larger ant, and these small ants start
to swab every part of the leaf section. This
swabbing continues after the section is
brought to the garden. Of 99 consecutive
sections of leaves taken back to a laboratory nest, 95 had the small ants riding on
them for a total of 131 minima (Weber,
1972a). This early licking must promote
the cleanliness of the substrate. (3) Adding their excretions to the new substrate
and to the garden as described below. (4)
Planting hyphae as tufts on the new particles. A leaf section 0.8 X 1-5 mm may have
10 mycelial tufts planted by an ant within
five minutes (Weber, 1956&). Each tuft
becomes an island of growth and expands
rapidly.
The division of labor and the large populations of a mature colony enable Atta to
have large nests that are well known to all
people living in their area, which includes
Texas, Louisiana, and Arizona, south to
Latitude 33°, just north of Buenos Aires
Province, Argentina. Many chambers are
excavated in the soil (Fig. 5) for the gardens and sometimes for refuse. The brood
is kept in the gardens. The ants forage
widely for fresh leaves, creating conspicu-
FUNGl'S-CULTURING BEHAVIOR OF ANTS
581
FIG. 5. Large mound nest of Atta cephalotes
adapted to the periodic flooding of the Orinoco
Delta, Venezuela, by having the fungus gardens
above high tide level. Seven or more gardens have
been exposed by the axe. The diameter of the
mound was 3.7 meters and the height 1.4 meters.
ous trails (Figs. 6, 7). The ants rapidly cut
sections of leaves employing a scissor-like
action of the mandibles (Fig. 8).
The liquid necessary for the extensive
salivary and anal excretions are maintained normally through the ingestion of
the protoplasm of the ant fungus. During
dry periods, or in the laboratory when the
air is drier than it would be in nature, the
ants may make up the deficiency in liquids
by imbibing water (Fig. 9). This may
also be true when the anfe are using dry
substrate, such as dead leaves or flowers,
since the fungus also requires much moisture. The workers quickly approach a pool
of water and stand at the edge, insert
their mouthparts into the water and pump
it into the digestive tract with their
pharyngeal muscles. The muscles may also
be seen at work when the ants imbibe
water on the ceiling of a glass plate under
the microscope.
Normally, the ants defecate only on the
garden or on the new substrate. A particle
of the latter may be so treated during the
late stages of the manipulation of it or the
ants may defecate on it after it has been
imbedded.
Young colonies, taken to the Swarthmore laboratory under U.S. Department of
Agriculture permit, have expanded markedly (Fig. 10). In recent years they
have been confined to plastic boxes of 2]^
liter capacity, connected by tubing of 2.5
cm diameter. Boxes are added as necessary.
The ants tend to fill an empty chamber
with exhausted substrate, the leaf sections
after the fungus has abstracted much of
the cellulose content, and has been cut out
from the base of a garden (the oldest
part) by the ants. This behavioral feature,
determination by the ants of the point
when old substrate should be removed, is a
worthy study in itself. A box full of garden will contain some 8,000-9,000 ants,
fully 60% of which are the minimas.
The gardens of the other chief leafcutting genus, Acromyrmex, are similar to
582
NEAL A. WEBER
FIG. 6. Trail made by Atta cephalotes in Trinidad. The nest was in the background along a
railroad right of way that was relatively undisturbed. The ants foraged for suitable leaves on the
campus of the University of the West Indies.
I K . . /. Irail made b\ Alln tcfihtilalt* in C.mana
and crossing a human foot trail al right anglrs.
l u o ant trails at. the left t o m e i g i d to a single-
those of Atta (Fig. 11), and the brood is
also kept in the upper cells of the garden.
The larvae are fed strands of the mycelium
or staphylae, aggregates of modified hyphae.
The external appearance of the aitinc
nest varies markedly with the species and
tends to be species- to genus-specific
(Weber, \912a,b). The young Atta nest
usually is surmounted by a chimney or
turret. This is reduced to a crater when
the colony is slightly older (Fig. 12). Such
turrets or craters are often closed by the
ants filling the entrance with dried leaf
sections or nearby vegetal debris. Very
young colonies close the entrance for the
night, older colonies may close it during
the clay during a dry period. By this behavior the ants conserve moisture for the
young garden or inhibit the entrance of
other animals.
The substrate used in nature, usually
green leaves in Atta and Acromyrmex,
comprises a great; variety of material. This
includes caterpillar and beetle feces, which
are dry and compact and contain recognizable particles of leaves (caterpillar) and
one leading
right,
the IILM silt
in
toicst
at
Ft'XGLS-CL'LTLRIXC BEHAVIOR OF ANTS
583
Ants of .-Ilia ceplialoles cutting fresh leaves in the laboratory.
wood fibers (beetle) in addition to the
metabolic products of the insect. The ants
may dissect tlie feces before planting them
on the garden. Insect carcasses are regularly found in gardens of small genera. Grass
is preferred by several species and Alia
may use woody twigs. This heterogeneous
plant and animal material contains a variety of bacteria, fungi, and other organisms.
In all cases the ants treat them by licking
FIG. 9. Alia cephalotes workers imbibing water on
the laboratory table during a period when the
relative humidity of the room was low and the
leafy substrate was relatively dry. At such times the
ants need much moisture to produce copious
.salivary and anal excretions.
l-'IG. 10. Colony of Alia cephalotes occupying connected chambers of 2i/J liter capacity in the laboratory. The holes in the top of the chambers permit
access to the ants or gardens. The older chamber
in the upper right was an inverted mouse nest of
1600 ml capacity on a plaster of Paris base.
584
NEAL A. WEBER
FIG. 11. Upper cells of a garden of Acromyrmex
octospinosus (Reich) with brood.
and defecating. The ant fungus is cellulose-digesting and in several cases known
to be basidiomycete.
When substrate is incorporated into the
ant garden and transferred to a sterile nutrient agar plate, many organisms start to
grow. Under the care of the ants, however,
the ants maintain a flourishing culture of
their own fungus despite surrounding contamination on the general agar surface
(Weber, 1956a). During the past year this
type of experiment was repeated with a
number of ant species (Weber, 1972i>) and
since continued. An experiment with
Trachymyrmex urichi (Weber, 19726) was
noteworthy in the extent to which the
ants defecated their amber droplets on the
general opalescent agar surface so that it
became a uniform amber color itself. Contamination was particularly slow to start in
this experiment. Ordinarily the ants defecate only on the garden or on substrate.
In this and other examples with other ant
species the ants cut' agar blocks, planted
them on their garden, defecated on them,
and grew their fungus successfully for periods of several weeks. Control plates became overwhelmed with alien sporulating fungi.
The most recent experiments (Figs. 13,
11, 15) also demonstrated behavior vit'al in
the maintenance of the garden. The ants
cut and discarded agar blocks from the
vicinity of the garden and continued until
a clear space surrounded it. This procedure is followed in nature: soil is
removed from all sides of the garden ex-
cept die base and even here they excavate
around stones where present so that the
garden touches only clean stones or roots.
Not only does this facilitate the isolation
of the garden from contaminants but it
inhibits easy access of pests such as Collembola and mites which bring contamination
with them, especially through their fecal
pellets. The ants regularly placed a few
agar blocks on the garden and these were
treated as substrate; the ant fungus grew
normally on them.
Such experiments succeed better with
monomorphic (workers all of similar size)
attine genera such as Sericomyrmex and
Trachymyrmex, than with the polymorphic Acromyrmex and Atta. In small samples
of the latter genera, the right proportion
of the worker sizes may not be obtained,
FIG. 12. Entrance to a young nest of Atta
cephalotes in the form of a crater 30 cm in diameter. The ants closed the entrance during the day
with leaf sections. Ants of still younger colonies
may close the entrance between 5:00 and 6:00 pm
tor the night. Mature colonif. mu\ be mostly
diurnal but at times will work into the night it a
rain during the day has prevented leaf ruttiny.
FUNGUS-CULTURING BEHAVIOR OF ANTS
585
FIG. 13. A fragment of a garden of Alia cephaloles
with workers (not visible) placed on a sterile
potato dextrose agar plate (pH 5.6) three days
after they were introduced. The first response of
the ants was to cut agar blocks in the immediate
vicinity of the garden. The blocks remained clean
in the interval at 25-26°C. Eventually the ants cut
all of the agar in the vicinity of the garden and
cast it to one side.
with a general inefficiency resulting. The
ants are also more quick in their movements in responding to external stimuli,
such as lifting the cover or jarring the
container.
The net effect of the behavioral pattern
described above is to produce a flourishing
culture of the ant fungus, a culture that
thrives despite constant' possibilities for
contamination. The salivary and anal ex-
cretions were long believed to be growthpromotional and to have at least bacteriostatic and fungist'atic qualities if they
were not actually antibiotic for alien organisms (Weber, 1947, 1956a).
Investigators faited to find any appreciable antibiotic activity of the excretions or
in ground ants, but others ascribed clear
antibiotic production by the metapleural
glands (reviewed in Weber, 1972a). At the
586
NEAL A. WEBER
. .-Alt "
FIG. 14. A fragment of a garden of Trachymyrmex
urichi Forel with workers placed on a sterile
potato dextrose agar plate (pH 5.6) three days
after they were introduced. These ants were much
faster in cutting the agar around the vicinity of the
garden down to the glass floor than was Alia
cephalotes above. The blocks of agar remained
clean in the interval at 25-26°C although there
were several islets of contamination on the uncut
agar.
FIG. 15. The garden of Trachymyrmex urichi on
the plate of Figure 14 but 17 days (at 25-26°C)
after it was introduced. In the meanwhile, the
form of the garden has been changed by the ants
and the fungus became a dense mycelium. The
dark spots are fecal droplets. The surrounding agar
has become generally contaminated by alien
fungi or bacteria but this has not affected the
garden. The cover of the dish had been lifted onh
on the 13th day, thus facilitating contaminalion.
present time, the main principle unquestionably clear is that the ants produce a
flourishing culture of one fungus, their
sole food, that enables them In fin he
FuNGUS-CULTURING BEHAVIOR OF AN'TS
throughout much of the Western Hemisphere despite persistent attempts to eradicate them by man.
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587
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