Toxic and Nontoxic
MOLDS
Found in Food Materials
Botany and Plant Pathology
Departmental Series No. I
May 1966
AGRICULTURAL EXPERIMENT STATION
AUBURN UNIVERSITY
2. V. SMITH, Director AUBURN, ALABAMA
TOXIC and NONTOXIC MOLDS
FOULIND in FOOD MATERIALS'
URBAN L. DIENER, Professor of Plant Pathology
IN THIS REPORT the terms toxic and toxin refer to sub-
stances produced by fungi that have a detrimental effect
on the metabolism of animals and man. The term food
materials is quite broad, but here it is restricted to food
of plant origin, such as seeds, seed products like flour,
meals, mash, bran, and in a few cases forage, hay, straw,
and fodder. Mold is used in the collective sense and
synonymous with fungus, generally referring to saprophytic
or weakly pathogenic fungi rather than aggressive plant
pathogens.
This report is a summation of many papers and several
excellent reviews of literature that deserve special men-
tion: 1) George Semeniuk, chapter on "Microflora" in
Storage of Cereal Grains and Their Products, 1954 (15);
2) C. M. Christensen, "Deterioration of Stored Grains by
Fungi" in Botanical Review, 1957 (5); and 3) Forgacs and
Carll, "Mycotoxicoses" in Advances in Veterinary Science,
1962 (7). For anyone interested primarily in the subject
of aflatoxin, the bibliography of Skau, Robertson, and
Mayne (16) encompasses the literature from 1960 through
1963.
Fungi (Molds) Associated with Food
Materials of Plant Origin
Many species of fungi and bacteria make up the micro-
flora of seed and seed products. Semeniuk (15) reported
nearly 70 species of bacteria, 7 species of Actinomycetes,
more than 40 species of yeasts and yeastlike fungi, and
more than 50 species of fungi have been found externally
on cereal grains and their products. About 25 species of
parasitic fungi and bacteria were reported to be carried
internally by cereal grain seeds alone. Colony counts
ranged from a few hundred on some seed to as high as
41,600,000 colonies per gram of moldy grain (6, 15). The
fungi associated with cereals were divided by Christensen
(5) into: 1) the field fungi - major genera consist of Fu-
sarium, Rhizoctonia, Alternaria, Helminthosporium, Di-
plodia, Cladosporium, and Rhizopus; and 2) the storage
fungi - principally Aspergillus and Penicillum.
Thus, a great number of genera and species of fungi and
bacteria are associated with seed and food materials. Jack-
son (9) recently listed 132 species of fungi that have been
reported to be associated with peanut pods and kernels.
Because of their great reproductive capacity in favorable
environments, these molds occur in huge numbers.
Origin, Nature, and Determination of Fungi
Fungi found in and on food materials are identical to
those found in the soil and air, and on living and dead
plants and animals. Some are pathogens of living organ-
1 Originally presented as an invitational paper at the Sym-
posium on Mycotoxins, 56th annual meeting of the American
Oil Chemists' Society, Houston, Texas, April 28, 1965. Pre-
search Grant No. EF-00590-02 from Division of Environmental
Engineering and Food Protection.
isms, whereas others are primarily saprophytes (living on
soil or organic matter). Field fungi invade developing
plants and seeds in the field prior to harvest, especially
under moderately warm and moist weather conditions.
In order to grow in seeds, fungi require moist environ-
ments with a moisture content in equilibrium with rela-
tive humidities from 90 to 100 per cent. Most seed crops
are harvested at moisture contents and relative humidities
below the level favorable for further invasion of seed
by these fungi. Therefore, field fungi gradually diminish
in quantity as seed moisture declines in curing and de-
pending upon temperature and moisture of the environ-
ment around the seed.
Storage fungi consist mainly of 10 to 18 species of
Aspergillus and Penicillium that are saprophytic on a wide
variety of plant and animal residues as well as seed and
seed products. Seed invasion occurs primarily following
harvest when moisture contents are in equilibrium with
relative 'humidities from 75 to 90 per cent and frequently
accompanied by temperatures of 25 to 385 C. The determ-
ination of fungi present is made by isolation, cultivation on
laboratory media, and subsequent identification using a
microscope. Field fungi can be cultured on most common
laboratory media, such as potato-dextrose agar and
Czapek's Dox medium. However, the storage fungi, es-
pecially the important Aspergillus glaucus group of five to
eight species, cannot be cultured satisfactorily on these
media. For good growth, sporulation, and development of
perithecia, they require culture media containing high
concentrations of sucrose or salt (NaCI), such as Thom
and Raper's Czapek solution agar with 20 per cent sucrose
(17), or Christensen's malt agar with 8 to 20 per cent salt
(4). A. flavws, the A. glaucus group, other Aspergilli, and
many Penicillium spp. grow well on these media of high
osmotic tension.
Factors That Determine Mold Invasion
Moisture is the most critical factor, whether it is the
water content of the material or the relative humidity
around the food stuff, since in a restricted, unaerated, or a
stable environment they come to equilibrium (i.e.-hygro-
scopic equilibrium). Most fungus species remain dormant
and/or slowly die out under conditions of low moisture.
Under these conditions they perish faster at high tempera-
tures than at low. The degree of invasion by the fungus
prior to storage of food materials will affect the moisture
relationship, since fungi directly produce water as a prod-
uct of respiration as do most other living things. Heavily-
infested seeds, dried to safe moisture levels, have been
reported to continue being deteriorated by fungi.
Storage fungi and field fungi generally grow well at
temperatures of 25 to 80' C. However, many species of
storage fungi grow well up to 85 to 38 C. A. flaws grows
well on certain substrates up to 45' C. (118 F). Growth
rates of most mesophilic fungi are greatly reduced at tem-
peratures below 20' C., although certain Penicillium
species and a few Aspergilli will grow slowly at 2 to 50 C.
under conditions of high moisture.
Time is an important factor as it relates to storage of
food materials with suboptimal environments that permit
only limited mold development. During extended periods,
fluctuations of moisture and temperature result in chang-
ing environmental conditions that may favor mold growth
for short periods of time, during which deterioration is
progressive and accumulative in its effect.
Contamination of food materials with foreign matter of
high moisture content, or of a hygroscopic riature can be
a factor in promoting mold development. Insect and/or
animal infestation may introduce moisture and produce
substrates favorable for mold development.
Molds are typically aerobic in nature and growth rates
decrease with lowering of oxygen tensions. High levels
of CO
2 
concentration (above 14 per cent) inhibit growth
of many molds.
Deteriorative Effects of Fungi
Seeds lose germinability, germs become discolored, and
musty odors develop in stored grain as a result of the
presence of molds. Christensen (5), Milner and Geddes
(11), and others have documented these facts.
Food nutritive value is destroyed by molds. Zeleny (19)
has reviewed the literature on chemical, physical, and
nutritive changes during storage. Nagel and Semeniuk
(13), Milner and Geddes (10), and Ward and Diener
(18) have demonstrated the biochemical changes in corn,
soybeans, and peanuts caused by fungi. These and other
investigations have shown that losses of organic matter,
carbohydrates, and oil are accompanied by increases in
free fatty acids and development of off-odor, changes in
flavor, and changes in color of oil. Thus, quality goes
down as rancidity goes up. Edibility or palatability of
food or feed may be reduced.
Changes in respiration, moisture content, and heating
of damp grain as caused by fungi have been reviewed by
Milner and Geddes (11).
Economic losses in terms of value of food for animal
and human consumption, caused by deterioration as result
of mold, have been estimated at 1-2 per cent of the an-
nual crop. Seed losses, poor stands, and land replanting
are all economic losses caused by fungus deterioration of
seed germinability.
Production of Toxins by Fungi
Mycotoxicoses are poisonings after entrance into the
host's body of toxic substances of fungal origin, Forgacs
and Carll (7). Most micro- and macro-organisms secrete
or excrete a toxin or toxins in normal metabolic processes
that are toxic or have detrimental effects on one or more
other organisms. However, this discussion is limited to
fungus-toxins affecting animals and man.
One of the earliest known fungus toxins is that found
in "ergot" sclerotia on ryegrass produced by Claviceps
purpurea. Rye was the main food cereal in the Middle
Ages. Ergot contains alkaloids that affect the central
nervous system causing convulsions and gangrene. Rye
bread eaten by peasants in France in severe ergot years
resulted in gangrenous infections and death to many peo-
ple. Ergot, caused by other species of Claviceps, occurs on
barley, wheat, dallis, brome, and other grasses. Ergot also
causes death and abortions in livestock.
Barley and corn infected with head smut or scab con-
tains toxic substances formed by the pathogenic fungus,
Gibberella zeae (Fusarium spp.) during invasion of the
living plant (12). Scabby grain is prevalent in warmer
parts of humid Eastern and Central States. Swine losses
are greatest following severe scab years because of feed-
ing infested grain, especially barley.
Stachybotryotoxicosis is the result of ingestion of feeds
(hay, straw, fodder) upon which strains of Stachyboirys
atra have grown and formed a toxin. Horses were severely
affected in Russia and Siberia in 1981. Cattle, domesti-
cated and experimental animals (mice, guinea pigs, rab-
bits and dogs) as well as humans are susceptible to this
toxin. Man may develop a rash.
Various Russian workers reported that extracts of A.
flavus, A. fumigatus, A. nidulans, A. niger, A. calyptratus,
Mucor albo-alter, and M. hiemalis were toxic to rabbits
and other animals. Other workers have reported that
various species of Aspergillus, Penicillium, and Rhizopus
were toxic to swine, ducks, rabbits, and guinea pigs. Carll
and coworkers implicated A. chevalieri, A. clavatus, and
A. fumigatus as causes of bovine hyperkeratosis in cattle,
horses, rabbits and mice.
Moldy corn toxicosis in swine was reported by Burn-
side, Sippel et al. (3) in Georgia and Florida. They found
corn cultured with A. flavus, P. purpurogenum, and P.
rubrum was toxic to swine and P. rubrum also affected the
horse and goat. This toxicosis was probably identical to
hepatitis X in dogs (1).
Moldy feeds are toxic to poultry - hemorrhagic syn-
drome. Alternaria sp., Aspergillus clavatus, A. flavus, A.
fumigatus, A. glaucus, P. purpurogenum, P. rubrum, P.
citrinum, and several other fungi have been shown to in-
duce mycotoxicoses in chickens by Forgacs and Carll (7).
Facial eczema in ruminants (sheep and cattle) was first.
reported in 1942 to be caused by Sporodesmium bakeri
growing on dead perennial ryegrass in New Zealand and
Australia. Moldy bermudagrass toxicosis in fields of South-
eastern United States also produces facial eczema in cattle.
The fungus, Periconia minutissima, (Alabama, Florida)
develops on bermudagrass after a killing frost or drouth is
followed by warm rain and the dead grass becomes in-
fested with various fungi. Any similar combination of con-
ditions caused by drouth or other environmental factors
will produce an outbreak in ruminants foraging such pas-
tures.
Alimentary toxic aleukia (ATA) 
develops in human
beings, who have ingested overwintered moldy grain or
its by-products. Russia reported high mortality rates
among those afflicted in 1913 in far eastern Siberia; in
1932 in western Siberia; and in 1941-45 in the Ukraine,
the area south and west of the Ural Mountains, in central
Asia, and in far eastern Siberia. The causal fungus was
Fusarium sporotrichioides growing on cereal grains pri-
marily proso millet. The fungus develops after a mild
winter with abundant snows followed by frequent alter-
nate freezing and thawing in the spring on millet allowed
to overwinter on the ground. Farm animals that eat toxic
grain are also affected. Mostly rural people are affected.
It has been experimentally produced in cats, guinea pigs,
dogs, and monkeys. Uniquely, this fungus grows slowly
at -10' C., optimally at 24' C., with an optimal tempera-
ture for toxin production between 1.5 to 4' C. The toxin
is stable for 30 minutes at 125' C. and 18 hours or longer
at 1100 C. Moldy grain has remained toxic for 6. years.
Finally the fungus more or less responsible for this
symposium - Aspergillus flavus - and its mycotoxin, afla-
toxin, which is also produced by a closely related species,
A. parasiticus, and possibly by Penicillium puberulum.
In 1960 (2) an apparently new disease occurred in tur-
key poults in England causing a loss of more than 100,000
birds at 500 locations. Outbreaks in ducklings, young
pheasants, swine, and calves were subsequently reported.
Brazilian groundnut (peanut) meal was found to be in-
volved in all cases. Similar disease outbreaks occurred in
East Africa and India from local peanuts. Sargeant et al.
(14) isolated a toxin-producing strain of Aspergillus flavus
from toxic Uganda peanuts. Considerable data have es-
tablished that toxic groundnut meals and feeds had been
present in England since 1951. In the USA, moldy corn
toxicosis in 1957 (3) and hepatitis in dogs in 1959 (1) had
already been shown to be caused by A. flavus and P.
rubrum.
Aspergillus flavus has been found to be prominent 
in
the microflora of cereal grains, oilseeds, other seed crops
and seed products (5, 15). It grows profusely and produces
aflatoxin on most seed crops and seed products in the lab-
oratory. A fairly adequate method for determining afla-
toxin has been developed and aflatoxin has subsequently
been found in low quality stocks of most cereals, oilseed
crops, and their products. Toxin-producing strains are
probably country-wide and world-wide in distribution.
Elimination and Control
The first step in control is the recognition and awareness
that the problem and threat exists. Prevention and control
of mycotoxins are dependent on control of factors influenc-
ing mold growth, such as the following:
Harvest seed crops at maturity and follow curing and
drying techniques to rapidly reduce seed moistures to
safe levels for storage. Utilize modem drying methods.
Store cleaned and sound seed in dry, aerated, insect-
free storage facilities. Foreign matter, whether 
soil or
plant debris, damaged seed, and immature seed 
may af-
fect aeration and moisture availability unfavorably.
Feed and food processors should determine toxin pres-
ence by chemical procedures before products 
are manu-
factured and processed.
Research may eventually yield methods for detoxifying
mycotoxin-contaminated feeds and food materials. Im-
proved techniques for growing, harvesting, drying, and
storing food materials to prevent mold development 
are
being further investigated. Basic research on the 
influence
of environment on mold growth in food products and 
ef-
fect of chemicals on mold development 
are being studied.
Summary
Despite thousands of plant diseases and species of fungi
associated with food materials, relatively few have proved
to be toxic to man and animals. Many species of fungi
contribute to the welfare of mankind in destroying organic
waste, formation of mycorhizae, alcohol and organic acid
production by fermentation, direct utilization as food
(mushrooms), food production such as cheese and bread
making, livestock feed as fermentation byproducts, anti-
biotics, medicinal compounds such as 
vitamins, and chem-
ical synthesis of enzymes, glycerol, and fats. 
These bene-
fits are in opposition to harmful activities of 
fungi such as
plant diseases, decay of timber, tropical deterioration of
textiles, food spoilage, allergens, and toxin-production.
Storage fungi are important in stored food materials
because they grow at low moistures, fairly high tempera-
tires, and are universally present in air and soil. Fungi
adversely affect seed viability, storage quality, nutritive
value, edibility, and occasionally produce toxins in seed
and food products.
Although toxin production is characteristic of most
micro-organisms, only a few toxins affect animals. The
present concern over mycotoxins has been emphasized
because aflatoxin is a carcinogen at very low levels in the
diet of experimental animals.
Control probably will be accomplished by utilizing
knowledge of moisture, temperature, and aeration to pre-
vent development of fungi during harvest, curing, storage,
and processing of food materials. Fungicidal chemicals
may afford some promise for mold and toxin control.
Literature Cited
(1) BAILEY, W. S., AND GRoTH, A. H., JR. The Relationship
of Hepatitis X of Dogs and Moldy Corn Poisoning of
Swine. J. Amer. Vet. Med. Assn. 184:514-516. 1959.
(2) BLOUNT, W. P. Turkey "X" Disease. J. Brit. Turkey
Federation. 9:52. 1961.
(3) BURNSIDE, J. E., FORGACS, J., SIPPEL, 
W. L., JR., CARLL,
W. T., ATwooD, M. B., AND DOLL, E. R. A Disease of
Swine and Cattle Caused by Eating Moldy Corn. Amer.
J. Vet. Res. 18:817-824. 1957.
(4) CHRISTENSEN, C. M. The Quantitative Determination 
of
Molds in Flour. Cereal Chem. 23:322-829. 1946.
(5) ------------------ Deterioration of 
Stored Grains by
Fungi. Bot. Rev. 23:108-184. 1959.
(6) DIENER, U. L. The Mycoflora 
of Peanuts in Storage.
Phytopath. 50:220-228. 1960.
(7) FORGACS, J., AND CALL, W. T. Mycotoxins. 
Adv. in Vet.
Sci. 7:278-382 (180 references). 1962.
(8) HODGES, F. A., ZuST, J. R., SMITH, H. R., 
NELSON, A. A.,
ARMBRECHT, B. H., AND CAMPBELL, A. D. Mycotoxins:
Aflatoxin Isolated from Penicillium puberulum. Sci. 145:
1439. 1964.
(9) JACKSON, C. R. A List of Fungi Reported on Peanut Pods
and Kernels. Ga. Coastal Plain Expt. Sta., Tifton. Mimeo
Series N. S. 234. 1965.
(10) MILNER, M., AND GEDDES, W. F. Grain Storage Studies:
III The Relation Between Moisture Content, Mold
Growth, and Respiration of Soybeans. Cereal Chem.
28:225-247. 1946.
(11) ------------------- ....----------- Respiration and Heat-
ing: In Storage of Cereal Grains and Their Products.
Amer. Assn. Cereal Chem., St. Paul, Minn. pp. 152-220.
( 178 references). 1954.
(12) MITCHELL, H. H., AND BEADLES, J. R. The Impairment
in Nutritive Value of Corn Grain Damaged by Specific
Fungi. Jour. Agr. Res. 61:135-141. 1940.
(13) NAGEL, C. M., AND SEMINIUK, G. Some Mold-Induced
Changes in Shelled Corn. 
Plant Physiol. 22:20-38. 
1947.
(14) SARGEANT, K., SHERIDAN, A., O'KELLEY, J., AND CARNA-
GHAN, R. B. A. Toxicity Associated with Certain Samples
of Groundnuts. Nature 192:1096-1097. 1961.
(15) SEMENIUK, G. Microflora: In Storage of Cereal Grains
and Their Products. Amer. Assn. Cereal Chem. St. Paul,
(16) SKAu, DOROTHY B., ROBERTSON, J. A., J., AND MAYNE,
Rurn Y. Bibliography of Aflatoxin from 1960. U.S. Dept.
Agr., So. Util. Res. & Dev. Div. Mimeo. (188 references).
March 1964.
(17) THOM, C., AND RAPER, K. B. A Manual of the Aspergilli.
(18) WARD, H. S., AND DIENER, U. L. Biochemical Changes
in Shelled Peanuts Caused by Storage Fungi. Phytopath.
51:244-250. 1961.
(19) ZELENY, LAWRENCE. Chemical, Physical, and Nutritive
Changes During Storage: In Storage of Cereal Grains
and Their Products. Amer. Assn. Cereal Chem. St. Paul,
Minn. pp. 46-76. (91 references). 1954.
Histological Studies
of Root and Shoot from
Trifluralin-Treated Corn Seedlings
Botany and Plant Pathology
Departmental Series No. 2
August 1967
Agricultural Experiment Station
Auburn University
E. V. Smith, Director Auburn, Alabama
Ilistological Studies of Root and Shoot fron
Trifluralin-Treated Corn Seedlings
N. S. NEGI, H. H. FUNDERBURK, and D. P. SCHULTZ
Department of Botany and Plant Pathology
TRIFLURALIN (a,a,a-trifluoro-2,6-dinitro-N, N-dipropyl-
p-toluidine) is known to cause marked effects on morpho-
logy of roots (1,3,4,5).
The presence of multinucleate cells and the interference
in cell division in roots following trifluralin treatment have
been reported in several species (1,5). Laboratory studies
at Auburn (3) showed that in some species trifluralin also
caused swelling of shoots. The study reported here was
designed to examine the nature of swelling caused by
trifluralin both in roots and shoots of corn (Zea mays
var. Dixie 18).
The corn used in this study was germinated and grown
in the dark for 3.5 days in 5 p.p.m. trifluralin solution and
in water at 82 F. Treated corn showed an abnormal swell-
ing of root tips and first internodes (Fig. 1) Three 5
mm. segments of root beginning at the tip and three 5 mm.
segments of shoot from the base of first internode were
sampled and killed in FAA. The tissue was dehydrated,
infiltrated, and finally embedded in wax. Transverse and
longitudinal microtome sections 10 jt in thickness were
cut. Sections were stained by the tannic acid-ferric chlor-
ide method of Foster (2). Radii of the sections were
measured from a camera lucida sketch. The number of
cells were counted in these radii and the average cell
diameter was calculated.
Swelling in roots was maximum at about 3 mm. from
the tip. In this region the radius of roots in treated seed-
lings was about 3 times larger than those of the check (Fig.
2). Increase in size of the cortex was larger than in the vascu-
lar tissue. The number of cells was slightly higher in tri-
fluralin-treated seedlings, but the diameter of the cortex
cells was more than double and they were also longer.
Similar effects of trifluralin treatment on radius, number,
and diameter of cells were noticed, in next two segments
(5 to 10 mm. and 10 to 15 mm.) of the roots. Length of
the cortex cells, however, did not increase because of
treatment in these segments.
In shoots the characteristic swelling from the trifluralin
treatment was more obvious in 5 to 10 mm. segments of
1Assistant Professor, Associate Professor, and NDEA Fellow,
Department of Botany and Plant Pathology, Agricultural Ex-
periment Station, Auburn University.
first internode (Fig. 3). As in roots the larger part of the
swelling was the result of increase in cortex cell size.
The epidermis in both roots and shoots of treated seed-
lings was either disorganized or not present (Fig. 2b and
3b). This could be attributed to the fact that the increase
in epidermal cell size could not keep pace with the rate
of increase in size of the cortex. Cortex cells near the edge
were also very irregular in shape.
Multinucleate cells were noticed both in roots and the
first internode sections of treated seedlings (Fig. 4).
Nuclei, in some cases four to five, were generally aggre-
gated and some of them were different in size. The
frequency of multinucleate cells was greater in the apical
5 mm. section of roots to the 5 to 10 mm. segment of
the first internode.
ACKNOWLEDGMENTS
The authors thank Dr. Edward M. Clark and Dr.
Amelie C. Blyth for use of equipment and Mrs. Martha P.
Bennett for technical assistance. Financial support of this
investigation by WP-00636-03 from the Federal Water
Pollution Control Administration and by Eli Lilly Co. is
also acknowledged.
REFERENCES
1. AMATO, V. A., R. R. HOVERSON, and J. HACSKAYLO.
1965. Micro-anatomical and Morphological Responses
of Corn and Cotton to Trifluralin. Proc. Assoc. South-
ern Agr. Workers. 62:234.
2. FOSTER, A. S. 1984. The Use of Tannic Acid and Iron
Chloride for Staining Cell Walls in Meristematic Tissue.
Stain Tech. 9:91-92.
3. NEGI, N. S. and H. H. FUNDERBURK, Jr. 1967. Response
of Various Plant Species to Different Rates of Triflu-
ralin and Benefin. Proc. Southern Weed Conf. 20:369.
4. STANDIFER, L. C., L. W. SLOANE, and M. E. WRIGHT.
1965. The Effect of Repeated Trifluralin Applications
on Growth of Cotton Plants. Proc. Southern Weed
Conf. 18:92-93.
5. TALBERT, R. E. 1965. Effects of Trifluralin on Soybean
Root Development. Proc. Southern Weed Conf. 18:652.
18:652.
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