22 Egypt. J. Bot., Vol. 52, No. 2. pp.

353-369 (2012)

Toxigenic Variation in Fusarium Species Isolated

from Imported Raw Materials Used in Locally
Processed Feed
R. M. Othman1, D. A. M. Abdou1, N. E. El-Bordeny2, N. A.
Ibrahim1 and M. A. Abouzeid1*
1
Department of Microbiology, Faculty of Science, Ain Shams
University, and 2 Department of Animal Production, Faculty of
Agriculture, Ain Shams University, Cairo, Egypt.

E ighty six samples including imported raw materials and locally

processed feed were mycologically examined. Total fungal counts
in the examined samples ranged from <102 to 7.4 x 104 CFUg-1 with
counts in 34% exceeding the feed hygienic quality limits. Fifty two
Fusarium isolates belonging to 10 different species were recovered, F.
proliferatum, F. verticillioides, , F. nygamai, F. oxysporum and F.
sporotrichioides were the most important recorded species, among
which, the most prevalent ones were F. proliferatum (22 isolates)
followed by F. verticillioides (10 isolates). The mycotoxigenic ability
of these isolates was tested under laboratory conditions using HPLC
analyses of organic extracts from solid cultures. Five isolates proved to
be toxigenic, 3 belonging to F. proliferatum, in addition to one from
each of F. oxysporum and F. nygamai. Six Fusarium mycotoxins were
detected by the applied chromatographic technique. Three chemically
different mycotoxins: moniliformin (0.0847mmol/L), beauvericin (0.206
mmol/L) and fusaproliferin (0.0355mmol/L) in addition to its derivative
deacetylfusaproliferin (0.0425mmol/L) were produced by three different
isolates of F. proliferatum. Fusaric acid was detected in the extract of
the F. oxysporum culture filtrate (0.268 mmol/L) whereas the isolate of
F. nygamai was found to produce both fusaric acid (0.191mmol/L) and
9,10-dehydrofusaric acid (0.0966mmol/L). The obtained results could
be considered as an important step in evaluating materials used in
animal feeding especially the imported ones. Also it may indicate that
both raw and processed materials used in animal feeding could be a
potential source for contamination with different mycotoxins, thats why
they should be subjected to strict regulations and regular monitoring in
order to ensure feed safety and avoid the negative economic and health
consequences.

Keywords: Fusarium Species, Feed Materials, Mycotoxins, HPLC

analyses, Beauvericin, Fusaproliferins.
*
Corresponding author, Tel: +0123570802; Fax: +20226857769, E-mail: m_abouzaid @ sci.asu.edu.eg
354 RASHA. M. OTHMAN et al.

Mold and mycotoxin contamination of feed and feed ingredients occurs

worldwide and because of the ubiquitous nature of these micro-organisms they
cannot be totally eliminated from feeds and ingredients (Trenholm et al. 2000). Most
fungal contaminants in stored feed materials usually arise from infestations
that began in the field, although some can directly infest storage grains as well
when conditions are right (Vieira 2003 & Mabbett 2003).
More than 100.000 fungal species are considered as natural contaminants of
agricultural and food products. However, due to genetical and ecological factors,
relatively few can actually generate mycotoxins (Jemmali 1979). According to
Leibetseder (1989), 30 to 40 % of existing moulds can elaborate toxic substances
under favorable conditions. Presence of the fungus is not an evidence that it is
the producer of the toxin; a given toxin may persist in the substrate when the
producer fungus is no longer present (Fink Gremmels 1999).
Moldy feed may lead to mycotoxin buildup reaching to injurious levels for
farm animals and human health. Effects on animals may be detrimental as noted
by Oyedeji (1986) including reduction in energy, degradation of essential
nutrients (amino acids and vitamins) and mycotoxicosis (hepatotoxicity,
nephrotoxicity, immunotoxicity, oncogenesis and genotoxicity (Dierheimer
1998 & Gabal and Azzam, 1998).
The most frequently toxinogenic fungal species found in food and feed
commodities belong to the genera Aspergillus, Fusarium and Penicillium
(Bankole and Kpodo, 2005). Fusarium species frequently contaminate many
crops such as corn, wheat, rye, tricale, oats and rice, on which they are capable of
elaborating a wide range of toxins. Fusarium toxins have been shown to cause a
variety of toxic effects in animals and, in some occasions, they have also been
suspected of toxicity for humans (Nair 1998).
Moniliformin is only known to be produced by species of Fusarium. This
metabolite works by suppression of the pyruvate dehydrogenase enzymatic
system, thus, inhibiting the Krebs cycle with consequent inhibition of cellular
respiration. Moniliformin is known to be toxic to poultry based on laboratory
toxicity studies (Leslie et al. 1996 and 2005). Among chickens, ducklings, and
turkeys, ducklings were found to be the most sensitive and necropsy revealed
ascites, hydropericardium, and myocardial pallor (onkova et al. 2003).
Beauvericin is a non-ribosomal, cyclohexadepsipeptide cation chelating agent
and ionophore with insecticidal properties (Logrieco et al. 1998). It was first
isolated from the entomopathogenic fungus Beauveria bassiana (Desjardins 2006).
Other beauvericin-producing species include F. avenaceum, F. fujikuroi,
F. globosum, F. nygamai, F. poae, F. proliferatum, and F. subglutinans.
Beauvericin has not been associated with any animal disease outbreaks or
disease in experimental animals (Desjardins 2006) but has been shown to induce
apoptosis in mammalian cell lines (Logrieco et al. 2002).
Egypt. J. Bot., Vol. 52, No. 2. (2012)
 TOXIGENIC VARIATION IN FUSARIUM SPECIES ISOLATED FROM 355

Fusaproliferin is a sesterterpene metabolite produced by only a few Fusarium

species within the G. fujikuroi species complex. In relation to feed grain, F.
proliferatum is a common pathogen of maize, and isolates of this species
consistently produce high levels of fusaproliferin (Desjardins 2006 & Shephard
et al. 1999). This metabolite has not been linked to any animal or human disease
outbreaks. Experimental analysis found fusaproliferin cytotoxic to mammalian
and insect cell lines (Fornelli et al. 2004 & Logrieco et al. 1996). Some strains of
F. globosum and F. pseudonygamai also are able to produce fusaproliferin (Fotso
et al. 2002 & Shephard et al. 1999).
Fusaric acid is a picolinic acid derivative. It is typically isolated from various
Fusarium species, and has low to moderate toxicity, which is of concern since it
might be synergistic with other cooccurring mycotoxins. Fusaric acid is
widespread especially on corn and corn-based food and feeds and is frequently
found in grain, where Fusarium species are also dominant (Bacon et al. 1996).
Fusaric acid mechanism of action depends on inhibition of Dopamine beta-
hydroxylase which converts dopamine to noradrenaline, lowering the blood
pressure. It may also have other actions, such as the inhibition of cell
proliferation and DNA synthesis (Nagasaka et al. 1985).
Regular monitoring of toxigenic mycoflora of the agricultural based feeds
and foods is an essential pre-requisite for development of control strategies.
Study of prevalence of toxigenic fungi of animal and poultry feed is regularly
reported from many countries including Spain (Accensi et al. 2004), Brazil
(Oliveira et al. 2006 & Rosa et al. 2006) and Nigeria (Osho et al. 2007).
However, in Egypt, limited studies have been reported upon presence of
mycotoxins in feeds and agricultural products (Abdelhamid 1990, El-Shanawany
et al. 2005, Mahmoud 2007 & Abo El Yazeed et al. 2011).
For this reason, the aim of this study was to isolate, enumerate and identify
toxigenic Fusarium species contaminating processed poultry and animal feed or
some imported raw ingredients used in feed manufacturing. Likewise, the
mycotoxin production by some of the identified species was also investigated.
Material and methods
Sample collection
Eighty six samples including processed feed samples (56) and feed
ingredients (30) were collected from different feed factories in Elfayume and
Qalubea governorates, Egypt. Raw ingredients used in poultry feed
manufacturing included: yellow corn, soybean meal, sunflower meal and corn
gluten. Processed feed samples included: Ruminant fattening and dairy
concentrate feed mixture (CFM), Rabbit does and growing CFM , broiler starter
and finisher feed and Poultry layer feed. Samples were cultured immediately
upon arrival.

Egypt. J. Bot., Vol. 52, No. 2 (2012)

356 RASHA. M. OTHMAN et al.

Isolation and identification of Fusarium species

Quantitative enumeration of fungal propagules was done on a solid medium
using the surface spread method. Ten grams of each sample were homogenized
in 90 mL 0.1% peptone water solution for 30 min. Serial dilutions (10-2 - 10-4)
were prepared and 0.1 mL aliquots were inoculated in triplicates onto the
following media: dichloran rose bengal chloramphenicol agar (DRBC) (Pitt and
Hocking 1997) to estimate total culturable mycobiota and dichloran
chloramphenicol peptone agar (DCPA) (Andrews and Pitt 1986) to enumerate
Fusarium species. DRBC plates were incubated at 25 oC for 710 days. DCPA
plates were incubated at 25 oC for 7 days. Only plates containing 10100 CFU
were used for counting and the results were expressed as CFU per gram of
sample. On the last day of incubation, individual CFU g-1 counts for each colony
type, considered to be different, were recorded. Colonies representative of each
type were transferred for subculturing on plates with fresh agar media. Fusarium
species were identified on the basis of the macroscopic and microscopic
characteristics according to the key of Leslie and Summerell (2006).
Mycotoxin production and analysis
Corn and rice grain solid cultures were prepared according to the method
described by Idris et al. (2003) as follows: 100 g corn/rice was placed in 500 mL
Erlenmeyer flasks with 45 mL of distilled water. The flasks were stored
overnight at room temperature, steamed for 20 min., and then sterilized for 20
min at 121 oC. The flasks were inoculated with 5 pieces of agar containing
actively growing mycelia and incubated at 25 C for 4 weeks. Duplicates were
made for each isolate.
Toxin extraction from Fusarium solid cultures
Corn or rice culture of each isolate were dried in a forced-air draft oven at
55 oC for 48 h, crushed with a mortar and pestle, packed in plastic bags, labeled
and stored at 4 oC until use (Idris et al. 2003). Extraction was carried out as
follows: 25 g was dried and ground, extracted with 100 mL methanol/water
(55:45, v/v, 1% NaCl), filtered through a filter paper. The filtrate (50 mL) was
defatted with n-hexane (50 mL x 2) and extracted with methylene chloride (50
mL x 3). Extracts were combined, dried (Na2 SO4), evaporated under reduced
pressure and dissolved in 1 mL methanol for further analysis (Idris et al. 2003).
Mycotoxin analysis using high Performance Liquid Chromatography (HPLC)
Authentic samples of the mycotoxins: Moniliformin, Beauvericin,
Fusaproliferin, Deacetylfusaproliferin and Fusaric acid were purchased from
Sigma Aldrich, St Louise, USA, whereas Dehyrdofusaric acid was kindly
provided by Dr Anna Andolfi, DISPA, Italy. Instruments and running conditions
used in HPLC analysis of the organic extracts of different Fusarium solid
cultures are described in Table 1.

Egypt. J. Bot., Vol. 52, No. 2. (2012)

 TOXIGENIC VARIATION IN FUSARIUM SPECIES ISOLATED FROM 357

TABLE 1: Instrumentation and running conditions of HPLC analysis

HPLC MONILIFORMIN BEAUVERICIN FUSAPRO- DEACETYL- FUSARIC
-LIFERIN FUSAPRO- ACIDS
LIFERIN
INSTRUMENTA The HPLC system employed was a Shimadzu (Kyoto, Japan) consisting of:
TION - An LC-10 AD VP pump
- A SPD-10 AV VP UV spectrophotometric detector
COLUMN Macherey-Nagel Shiseido Capcel C18 PREP. Macherey-
(Dren, Pak Reverse- Nagel (Dren,
Germany) Phase C18 Germany)
Reverse- Phase (150x4.6 mm) high-density
C18 (150x4.6 Reverse- Phase
mm, 5 m) C18 100-5
Nucleosil
column
(150x4.6 mm)
SOLVENT Ion pair modifier ACETONITRIL ACETONITRILE (80): (A) Methanol
SYSTEM 1) 1L H2O E: H2O (20) (B) H2O
(Milli-Q System) H2O (Milli-Q (C) 1%
pH 6.5 50 mL System) dipotassium
40% Tetrabutyl (65:35 5min) hydrogen
ammonium (70:30 7 min) phosphate in
hydrogen (65:35 4 min) H2O (pH 7.4
sulphate + 100 with conc.
mL 1.1 M phosphoric
Potassium acid
dihydrogen
sulphate Elution:
2) Initially with
ACETONITRIL A:B:C
E 8% (50:10:40)
transformed
according to a
linear gradient
over 20 min
to A:B:C
(75:10:15);
initial
conditions
were restored
according to a
linear gradient
for 5 min, and
the column
was re-
equilibrated
under these
conditions for
10 min before
the next run
FLOW RATE 1.5. mL/min 1.5 mL/min 1 mL/min 1 mL/min
WAVE LENGTH 229 nm 202 nm 192 nm 268 nm
RETENTION 6.18 min 13.63 min 4.78 min 8.68 min DFA: 7.04 min
TIME FA: 8.98 min
TOTAL TIME 40 min 35 min 35 min
RUN
3 injections of 20 L for each sample

Egypt. J. Bot., Vol. 52, No. 2 (2012)

358 RASHA. M. OTHMAN et al.

Results

Isolation and identification of Fusarium species

Total fungal counts ranged from < 102 to 7.4 x 104 CFU g-1of feed samples
with an average of 9.8 x 103 CFU g-1. Majority of samples analyzed (52.3%)
contained 9.4 x 103 to 1 x 103 CFU g-1. A total of 52 isolates of Fusarium were
recovered from the investigated samples. The predominant Fusarium species was
F. proliferatum with a total number of 22 isolates followed by F. verticillioides (10
isolates), F. napiforme (7 isolates), and F. globosum (4 isolates). The incidence of
other species in samples was relatively and was only represented by 1-2 isolates for
each. The total number and frequency of Fusarium species recovered from fed
samples during this study is recorded in Table 2..
Mycotoxin production and analysis
Only 5 isolates were able to produce mycotoxins. These mycotoxigenic
isolates belonged to 3 species namely F. proliferatum, F. oxysporum and F.
nygamai. Of which one isolate (F. proliferatum) was found to produce
fusaproliferin and its derivative deacetylfusaproliferin and another isolate (F.
nygamai) produced fusaric acid and dehydrofusaric acid (Table, 3). The highest
concentration of mycotoxins 0.268 mmol/L was recorded for fusaric acid
detected in the culture of an isolate of F. oxysporum recovered from a sample of
locally processed ruminant dairy feed, while the lowest concentration detected
(0.0355 mmol/L) was recorded for fusaproliferin detected in the culture of an
isolate of F. proliferatum recovered from a sample of corn imported from
Ukrainia (Table 3). In comparison to authentic samples of the detected
mycotoxins, the precision of different measurements for aliquots of organic
extracts from solid cultures of Fusarium isolates was determined by triplicate
injections under the HPLC conditions demined previously in the experimental
section (Table 1). Chromatograms obtained showed that the six mycotoxins
detected have different retention time. Figures (1-5) shows six well defined
peaks of the detected mycotoxins. The used method being rapid and allowed to
detect the mycotoxins by the coincidence of their retention times with those of
the standards. Linearity was confirmed using the calibration curve per each
mycotoxin concentration (Figure 6). A linearity was obtained for the range
between 0.5 to 2.0 g/mL with a correlation coefficient r = 0.99.

Egypt. J. Bot., Vol. 52, No. 2. (2012)

 TOXIGENIC VARIATION IN FUSARIUM SPECIES ISOLATED FROM 359

TABLE 2: Frequency of isolated Fusarium species from different feed

materials

Number of
Fusarium Number of samples
samples of Frequency* Frequency*
species of feed ingredientsb
processed feed a
F. proliferatum 15 26.78 7 23.33
F. 3.57 26.6
2 8
verticillioides
F. napiforme 3 5.35 4 13.33
F. nygamai 1 1.78 1 3.33
F. fujikouroi 1 1.78 0 0
F. oxysporum 1 1.78 1 3.33
F. Solani 0 0 2 6.66
F. andiyazi 1 1.78 0 0
F. globosum 4 7.14 0 0
F. 0 3.33
0 1
sporotrichioides
a: total collected samples = 56 b: total collected samples = 30
ns: the number of samples where a species occurred
N: the total number of collected samples

TABLE 3: Toxigenic potential of different Fusarium species associated with

different feeds
Type of mycotoxins
Parameters Moniliformin Beauvericin Fusa- Deasetyl- Fusaric acid Dehydro-
prolliferin
fusaproliferin fusaric acid

No. of
positive
1 1 1 1 2 1
samples

Producing
species F. F. F. F. F. F.
F. nygamai
proliferatum proliferatum proliferatum proliferatum oxysporum nygamai

Detected level
of mycotoxins
(mmol/L) 0.0847 0.206 0.0355 0.0425 0.268 0.191 0.0966

Source feed
product Starter poultry Ruminant
corn Corn Corn Corn Corn
feed dairy feed

Origin Processed in Ukrainian Ukrainian Processed in

Ukrainian American American
Egypt Egypt
No. of tested samples = 49

Egypt. J. Bot., Vol. 52, No. 2 (2012)

360 RASHA. M. OTHMAN et al.

Fig. 1: HPLC chromatographic pattern of the organic extract of F. proliferatum

showing a characteristic peak of Beauvericin at rt of 13.63 min.

Fig. 2: HPLC chromatographic pattern of the organic extract of F.proliferatum

showing a characteristic peak of Moniliformin at rt of 6.18 min.

Egypt. J. Bot., Vol. 52, No. 2. (2012)

 TOXIGENIC VARIATION IN FUSARIUM SPECIES ISOLATED FROM 361

Fig.3: HPLC chromatographic pattern of the organic extract of F. proliferatum

showing 2 characteristic peaks of fusaproliferin and deacetylfusaproliferin at
rt of 4.78 min and 8.68 min

Fig .4: HPLC chromatographic pattern of the organic extract of F. nygamai showing
2 characteristic peaks of dehydrofusaric and acid Fusaric acid at rt of 7.04 min
and 8.98 min.

Egypt. J. Bot., Vol. 52, No. 2 (2012)

362 RASHA. M. OTHMAN et al.

Fig. 5: HPLC chromatographic pattern of the organic extract of F. oxysporum

showing a characteristic peaks of Fusaric acid at rt of 7.04min

Fig. 6: Calibration curves of the different mycotoxins detected by HPLC injection.

Egypt. J. Bot., Vol. 52, No. 2. (2012)

 TOXIGENIC VARIATION IN FUSARIUM SPECIES ISOLATED FROM 363

Discussion

Total fungal count is one of the criteria in evaluating hygienic quality and it
is very important for determining the probability that the feed contains
mycotoxins. According to the regulation on maximal quantities of harmful
substances and components in livestock, feed mixtures and feed ingredients are
not in compliance with standards of the hygiene quality if they contain above
300 X 103CFU g-1 of forage mixture for older animal categories or 50 X 103
CFU g-1 for younger animals (Krnjaja et al. 2008). By applying these defined
criteria on feed samples collected during this study, it was found that 34% of the
feed samples didnt meet standards of microbiological adequacy as they
exceeded the feed hygienic quality limits (1 x 104 CFU g-1) (GMP 2005). The
total fungal loads in the analyzed finished feed samples in the present study were
around 9.8x103 CFU g-1 which is higher than those reported in many studies
from different countries like Slovakia, Turkey, Spain and Argentina (Labuda and
Taninov 2006, Bragulat et al. 1995 & Dalcero et al. 1998).
Members of the genera Aspergillus, Penicillum, Fusarium, Alternaria and
Cladosporium are of great importance because of potential mycotoxin
production (Kurata & Ueno 1984). Species of Fusarium are among the typical
fungal contaminants inhabiting poultry feed mixtures (Labuda & Taninov,
2006). The present work was concerned with Fusarium contamination in animal
and poultry feeds. Fusarium was detected in 52 out of 86 feed samples with a
percentage reaching 60.46%. If compared with other studies concerning the
mycobiota of feed, this result is in accordance with the findings of Cvetnic et al.
(2004) who found that Fusarium spp. were the most common fungi found in
maize with the frequency of 78.6% in 1999 and 85% in 2003 at Croatia. On the
other hand, this value is higher than those obtained by Abo El Yazeed et al.
(2011) who detected Fusarium in 23% of feed samples and Khosravi et al.
(2008) who isolated Fusarium in 6% of animal feed and is much higher than that
recorded by Buckley and Fogarty (2007) who detect Fusarium only in 2.6% of
equine concentrated feed. Furthermore, results of the present study confirm the
results of Cvetnic et al. (2004) and Khosravi et al. (2008) that corn, the main
ingredient of animal and poultry feeds, was the most commonly contaminated
feed source.
In this study and in many previous studies and surveys (Labuda & Taninov
2006 & Abo El Yazeed et al. 2011), it was found that the most frequently
encountered species of Fusarium in feed samples was F. proliferatum followed
by F. verticillioides.
It has been estimated that 25% of the worlds crop production is
contaminated with mycotoxins which results in significant economic losses in
crops and animals (Abo El Yazeed et al. 2011). Toxins produced by different
species of Fusarium in particular are of most concern for animal health (Oliveira
et al. 2006). However, there are only a limited number of surveys concerning
Fusarium mycotoxins but they all clearly show that feed should be of bigger
Egypt. J. Bot., Vol. 52, No. 2 (2012)
364 RASHA. M. OTHMAN et al.
concern from the mycotoxicological point of view. Furthermore, statutory
regulations do not exist for any of the Fusarium mycotoxins. Only a selection of
advisory and tolerance limits are scarcely available in literature (D Mello et al.
1999).
It is important to mention that in general the percentage of positive samples is
depending on the detection limit. The lower the limit of detection is the higher
percentage of positive samples and vice versa. Methods having more sensitive
limits will give more positive results and consequently a higher percentage of
positive samples (Martins and Martins 2001). In the present work, 3 Fusarium
isolates were excluded from mycotoxin analyses as they belong to species not
know for mycotoxin production. 5 Fusarium isolates out of the 52 (13.46%)
recovered from 5 out of the 86 (5.8 %) investigated samples was found to be
mycotoxigenic. Most of the detected mycotoxins were produced mainly by
isolates of Fusarium proliferatum, the most predominant Fusarium species
enumerated in this study. The highest concentration of mycotoxins 0.206
mmol/L was recorded for beauvericin detected in the culture of an isolate of F.
proliferatum recovered from a sample of Starter poultry feed processed in Egypt
whereas the lowest concentration was recorded for fusaproliferin detected in
culture of another isolate of F. proliferatum recovered from a sample of corn
imported from Ukrainia. Moniliformin production was observed in a third isolate
of F. proliferatum recovered from a sample of corn imported from Ukrainia. An
isolate of F. oxysporum, recovered from a sample of Ruminant dairy feed
processed in Egypt, produced fusaric acid, while F. nygamai recovered from a
sample of corn imported from America, produced fusaric acid and its derivative,
dehydrofusaric acid.
A matter for concern is undoubtedly the findings of F. proliferatum isolates
in 22 out of the 86 feed samples tested (nearly 26%), followed by F.
verticillioides in 10 out of the 86 analyzed samples (nearly 11%). The presence
of such seriously toxicogenic species in feeds may be indicative of their
potentiality to produce carcinogenic mycotoxins such as fumonisin
when the storage conditions are not appropriate (Oliveira et al. 2006). In Egypt,
the quality of manufactured mixed-feed varies from feed mill to another. The
lack of quality control measures lead sometimes to using poor quality ingredients
(El-Sayed 1998) In addition, the processing and handling methods of some of
the feed sources are poor and primitive. This adversely affects the quality of the
used sources. Moreover, the storage systems of imported feed ingredients and
produced mixed-feeds lack the basic requirements for proper storing where
thermal control and ventilation regimes are totally lacking. Instead, feed
ingredients and produced feeds are stored mainly outdoor (mostly uncovered).
Extreme weather conditions, rodents, insects and wild birds cause a great loss of
these sources and seriously affect their quality and durability (El-Sayed 1998).
In conclusion, the results obtained in the present study clearly showed that
poultry and animal feed mixtures should be regarded as a potential source for a
possible contamination with Fusarium mycotoxins and also highlight the need
Egypt. J. Bot., Vol. 52, No. 2. (2012)
 TOXIGENIC VARIATION IN FUSARIUM SPECIES ISOLATED FROM 365

for regular monitoring of toxigenic moulds and their toxic compounds through
analysis of processed feed especially imported raw materials. This is achievable by
following hygienic practices during the processing and storing of feeds.
Further studies and analysis are now in progress for aliquots of organic
extracts of imported raw materials to assess the presence of any of the
mycotoxins detected under laboratory conditions. This could help in accurate
estimate of the quality of feed raw materials imported from different sources
which significantly affect the quality of feed.

References

Abdelhamid, A. M. (1990) Occurrence of some mycotoxins (aflatoxin, ochratoxin

A, citrinin, zearalenone and vomitoxin) in various Egyptian feeds. Arch
Tierernahr. 40,647-64.
Abo El Yazeed, H., Hassan, A., Moghaieb, R.E, Hamed, M and Refai, M.
(2011) Molecular Detection of Fumonisin-producing Fusarium Species in
Animal Feeds Using Polymerase Chain Reaction (PCR). Journal of Applied
Sciences Research,7(4): 420-427
Accensi, E., Abarca, M.L. and Cabanes, F.J. (2004). Occurrence of Aspergillus
species in mixed feeds and component raw materials and their ability to produce
ochratoxin A. Food Microb. 21, 623-627.
Andrews, S and Pitt, J.I. (1986) Selective medium for isolation of Fusarium species
and dematiaceous hyphomycetes from cereals. Appl Environ Microbiol 51(6):
12351238.
Bacon, C. W., Porter, J. K., Norred, W. P. and Leslie, J. F. (1996) .Production of
fusaric acid by Fusarium species. Appl. Environ. Microbiol. (62)11, 4039-4043
Bankole, S. A. and Kpodo, K. A. (2005) Mycotoxin contamination in food systems
in West and Central Africa. In Reducing the impact of Mycotoxin in Tropical
Agriculture with emphasis on health and trade in Africa, Accra, Ghana, pp: 13-
16.
Bragulat, M.R,. Abarca, M.L., Castella, and G., Cabanes, F.J. (1995) A
mycological survey on mixed poultry feeds and mixed rabbit feeds. J Sci Food
Agric, 67, 215-220.
Buckley, T., A. and Fogarty, F. (2007) Analysis of Canadian and Irish forage, oats
and commercially available equine concentrate feed for pathogenic fungi and
mycotoxins. Irish Vet. J., 60(4), 231-236.
onkova, E., Laciakova, A., Kova, G., and Seidel, H. (2003) Fusarial Toxins and
their Role in Animal Diseases. Vet J. , 165, 214 220.
Cvetnic, Z., Pepeljnjak, S. and egvic, M. (2004) Toxigenic potential of Fusarium
species isolated from nonharvested maize. Arh Hig Rada Toksikol., 56, 275-280.
D Mello, J. P. F., Placinta, C. M. and Macdonald, A. M. C. (1999) Fusarium
mycotoxins: a review of global implications for animal health, welfare and
productivity. Animal Feed Science and Technology 80, 183-205.
Egypt. J. Bot., Vol. 52, No. 2 (2012)
366 RASHA. M. OTHMAN et al.

Dalcero, A., Magnoli, C., Luna, M., Ancasi G, Reynoso M.M., Chiachiera, S.,
Miazzo, R. and Palacio, G. (1998) Mycoflora and naturally occurring
mycotoxins in poultry feeds in Argentina. Mycopathologia. 141, 37-43.
Desjardins, A.E. (2006) Fusarium Mycotoxins: Chemistry, Genetics, and Biology.
APS Press, St. Paul, Minnesota, USA.
Dierheimer, G. (1998) Recent advances in the genotoxicity of mycotoxins. Rev
Md Vt. 149, 585-590.
El-Sayed, A.F.M. (1998) Mixed-feed manufacturing in Egypt . In proceeding of
conference of Feed Manufacturers of the Mediterranean. Brufau J., Tacon A.
(eds.).
El-Shanawany, A.A, Eman Mostafa M, and Barakat, A. (2005) Fungal
populations and mycotoxins on silage in Assiut and Sohag governorates in
Egypt, with a special eference to characteristic Aspergilli toxin.
Mycopathologia, 159, 281-289.
Fink-Gremmels, J. (1999) Mycotoxins: their implications for human and animal
health. Vet Q, 21 (4): 115-20.
Fornelli, F., Minervini, F., and Logrieco A. (2004) Cytotoxicity of fungal
metabolites to lepidopteran (Spodoptera frugiperda) cell line (SF-9). J.
Invertebr. Pathol. 85, 7479.
Fotso, J., Leslie, J.F. and Smith, J.S. (2002) Production of beauvericin,
moniliformin, fusaproliferin, and fumonisins B1, B2, and B3 by fifteen ex-type
strains of Fusarium species. Appl. Environ. Microbiol. 68, 5195 5197.
Gabal, M. A. and Azzam A. H. (1998) Interaction of aflatoxin in the feed and
immunization against selected infectious diseases in poultry. II. Effect on one
day old layer chicks simultaneously vaccinated against Newcastle diseases
infectious bronchitis and infectious bursal disease. Av Pathol. 27, 290-295.
GMP. (2005) Regulations on Product Standards in the Animal Feed Sector, GMP14.
17-08-05.
Idris, A.E., Abouzeid, M.A., Boari, A., Vurro, M. and Evidente, A. (2003)
Identification of phytotoxic metabolites of a new Fusarium sp. inhibiting
germination of Striga hermonthica seeds. Phytopathologia Mediterranea. 42, 65
70.
Jemmali, M. (1979). Les moisissures et leurs toxines. La Recherche, 10: 124-131.
Khosravi, A.R., Dakhili M. and Shokri, H. (2008) A mycological survey on feed
ingredients and mixed animal feeds in Ghom Province, Iran. Pakistan. J. Nutrit.,
7(1), 31-34.
Krnjaja, V., Stojanovi, Lj, Cmiljani, R., Trenkovski ,S. and Tomaevi, D.
(2008) The presence of potentially toxigenic fungi in poultry feed. Biotechnol.
Anim. Husbandry, 24, 87-93.

Egypt. J. Bot., Vol. 52, No. 2. (2012)

 TOXIGENIC VARIATION IN FUSARIUM SPECIES ISOLATED FROM 367
Kurata, A. and Ueno, Y. (1984) Toxigenic fungi: Their toxins and health hazard.
Odansha-Elseviers Tokyo-Amsterdam-Oxford-NewYork; 247 p.
Labuda R, and Tancinov, D. (2006) Fungi recovered from Slovakian poultry feed
mixtures and their toxinogenity. Ann Agric Environ Med. 13, 193200
Leibetseder, J., (1989) Die bedeutung der Mykotoxine fr Mensch und Tier. Ernhr
Nut.13, 739.
Leslie, J. F., Marasas, W. F. O., Shephard, G.S., Sydenham, E.W., Stockenstrom,
S., and Thiel, P.G. (1996) Duckling toxicity and the production of fumonisin
and moniliformin by isolates in the A and E mating populations of Gibberella
fujikuroi (Fusarium moniliforme). Appl. Environ. Microbiol., 62, 11821187.
Leslie, J.F. and Summerell, B.A. (2006) The Fusarium Laboratory Manual. Ames:
Blackwell publishing Chichester, UK, 388pp.
Leslie, J.F., Zeller, K.A., Lamprecht, S.C., Rheeder, J.P., and Marasas, W.F.O.
(2005) Toxicity, pathogenicity, and genetic differentiation of five species of
Fusarium from sorghum and millet. Phytopathology, 95, 275283.
Logrieco, A., Moretti, A., Castella, G., Kostecki, M., Golinski, P., Ritieni, and
Chelkowski, J. (1998) Beauvericin production by Fusarium species. Appl.
Environ. Microbiol., 64, 30843088.
Logrieco, A., Moretti, A., Fornelli, F., Fogliano,V., Ritieni, A., Caiaffa,M.F.,
Randazzo, G., Bottalico, A. and Macchia, L. (1996) Fusaproliferin
production by Fusarium subglutinans and its toxicity to Artemia salina, SF-9
insect cells, and IARC/LCL 171 human B lymphocytes. Appl. Environ.
Microbiol. 62, 33783384.
Logrieco, A., Mul`e, G., Moretti, A., and Bottalico, A. (2002). Toxigenic Fusarium
species and mycotoxins associated with maize ear rot in Europe. Eur. J. Plant
Pathol. 108, 597609.
Mabbett, T. (2003) Contamination control in feeds. African Farming, 28-29.
Mahmoud , L. E. (2007) Toxigenic fungi and mycotoxin content in poultry feedstuff
ingredients. Journal of Basic Microbiology 33(2): 101-104.
Martins, M.L. and Martins, H.M. (2001) Occurrence of the mycotoxins content in
raw materials in Portugal. In: Book of Abstracts of the International
Symposium - Bioactive fungal metabolites Impact and exploitation. University
of Wales Swansea, Swansea, UK, 76.
Nagasaka, A., Hara, I., Imai, Y., Uchikawa, T., Yamauchi, K., Suzuki, S.,
Takatsuki, K., Tomita, A, Niinomi, M. and Nakagawa, H. (1985) Effect of
fusaric acid (a dopamine beta-hydroxylase inhibitor) on phaeochromocytoma.
Clin Endocrinol (Oxf). 22(4):437-44.
Nair, M. G. (1998) Fumonisins and human health. Annals Trop Paed Supp.,5: 47-52
Oliveira, G.R., Ribeiro, J.M., Fraga, M.E., Cavaglieri, L.R., Direito, G.M.,
Keller, K.M., Dalcero A.M. and Rosa., C.A.R. (2006) Mycobiota in poultry
feeds and natural occurrence of aflatoxins, fumonisins and zearalenone in the
Rio de Janeiro State, Brazil. Mycopathologia, 162, 355-362.
Egypt. J. Bot., Vol. 52, No. 2 (2012)
 368 RASHA. M. OTHMAN et al.

Osho, I.B., Awoniyi T.A.M. and Adebayo, A.I. (2007) Mycological investigation
of compound poultry feeds used in poultry farms in south west Nigeria. African
J. Biotech., 6, 1833-1836.
Oyedeji, A. N. (1986) Prevalence of aflotoxin B1 in commercial poultry rations in
Nigeria. J. Inst. Fur Tropishe Land.
Pitt, J.I. and Hocking, A.D. (Eds.) (1997) Fungi and Food Spoilage. second ed.
Blackie Academic Press, London.
Rosa, C.A.R., Riberio, J.M.M., Fraga, M.J. Gatti, M. Cavaglieri, L.R.
Magnoli, C.E Dalcero, . A.M. and Lopes., C.W.G. (2006) Mycoflora of
poultry feed and ochratoxin- producing ability of isolated Aspergillus and
Penicillium species. Vet. Microb., 113: 89-96.
Shephard, G.S., Sewram, V., Nieuwoudt, T.W., Marasas, W.F.O. and Ritieni, A.
(1999) Production of the mycotoxins fusaproliferin and beauvericin by South
African isolates in the Fusarium section Liseola. J. Agric. Food Chem. 47,
51115115.
Trenholm, H. L., Charmley and PreLusky, L. L (2000) Mycotoxin binding agents:
An Update Farming today. 1, 11.
Vieira, S. L. (2003) Nutritional implications of mould development on feedstuffs and
alternatives to reduce the mycotoxins problem in poultry feeds. World Poult.
Sci. J., 59(1): 111- 122.

(Received 5/3/2012;
accepted 12/4/2012)

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