The Presence, Effect, and Diagnosis of Zearalenone
in Dairy Cattle
Background
Mycotoxins are toxic substances that are produced
by fungal species. These fungi can be found in a wide variety
of plants and soil types. Toxigenic fungi are thought to
be ubiquitous in the environment. Several characteristics
are thought to be important for fungal growth, but the exact
mechanisms of their proliferation are still unknown. The
mycotoxins of major importance in Indiana are zearalenone,
aflatoxin, DAS (vomitoxin) and fumonisin. Because the production
of mycotoxins is strongly influenced by weather patterns and
climates, the prevalence of each toxin varies with geographical
location.
Zearalenone is naturally produced by the fungus Fusarium
roseum and by some isolates of Fusarium moniliforme.
Zearalenone is most commonly reported in the north central
cornbelt of the United States and southeastern Canada. Zearalenone’s
estrogenic effects can be attributed to its complex chemical
structure which makes it a phytoestrogenic molecule. Phytoestrogens
are estrogenic compounds produced by plants/fungi. The fungus
responsible for zearalenone production, Fusarium spp.,
has also been shown to produce the nonestrogenic toxins deoxynivalenol
and T-2 under appropriate conditions. Therefore, veterinarians
and producers should be aware that mycotoxin contamination
can be a multi-factorial problem.
Clinical Effects
A wide variety of clinical effects attributed to zearalenone
have been described in the literature. Decreased fertility,
abnormal estrus cycles, swollen vulvas, vaginitis, reduced
milk production and mammary gland enlargement are the most
common findings reported in cattle and swine. From the aforementioned
changes, single or multiple effects have been observed. A
change in the estrus cycle can manifest itself in various
forms. Prolonged, skipped, or irregular heats are commonly
associated with zearalenone effects. While these abnormal
estrus changes are not exclusively specific to zearalenone
toxicity, one should investigate feed related causes when
increases in abnormal estrus cycles are observed on farm.
In one report from a 150 cow dairy herd, an increased artificial
insemination index (decreased fertility) was reported after
the herd had been fed moldy hay. Zearalenone concentrations
in the hay extracts were reported to be 14mg/kg of hay. Upon
removal of the infected hay, the AI index returned to the
previous level. A 20 cow Brown Swiss herd was observed showing
anestrus, false estrus, or nymphomania with a gray vaginal
discharge. Infectious causes were ruled out, and feed samples
were taken. Zearalenone concentrations of
50 ppb and 100 ppb were detected in the corn silage and haylage,
respectively. Based on feed intakes, it was determined that
these cows were receiving 1.6 mg of zearalenone per animal.
Finally, an abnormal estrus cycle, nonresponsive to lutenizing
hormone, was reported to have occurred in a dairy herd. Feed
analysis yielded a zearalenone concentration of 25 mg/kg.
The clinical effects observed on heifers vary somewhat
from cows. Mammary gland enlargement, swollen vulvas, and
vaginitis are frequently observed more often in heifers as
compared to cows. After being fed a ration containing moldy
corn, 17 of 20 prepubertal dairy heifers developed enlarged
mammary glands in at least one quarter. The secretion had
a consistency of skim milk and appeared an off-white color.
Zearalenone was determined to be present in the corn. Seven
weeks after removal of the affected corn, all heifers were
clinically normal. Weaver reported that when dietary zearalenone
was greater than 12.5 ppm, a reduced conception rate was observed
in virgin heifers.
Clinical effects of zearalenone vary by animal breed, age,
and environment. While the majority of clinical symptoms
observed mimic that of estrogenic stimulation, at the present
time there is no way to determine what form of the syndrome
will manifest in an affected group of animals. Multiple subclinical
changes are probably occurring in affected animals. A combination
of genetic and environmental factors most likely determines
what outward clinical signs will be observed. Because the
Fusarium spp. can produce estrogenic as well as nonestrogenic
toxins, a variety of clinical signs are possible.
Feed Sampling and Diagnosis
Diagnosis can be quite difficult and frustrating to the
veterinarian and owner. Animals may have consumed the offending
agent prior to the veterinary investigation, thus making collection
of diagnostic samples difficult or impossible. This problem
is further exacerbated by the sporadic and variable nature
of the toxin. Zearalenone, as with other mycotoxins, can
only be detected in feed or feed products. No individual
animal serologic or tissue test, antemortem or postmortem,
exists at this time. Histopathological changes indicative
of mycotoxicosis can be observed in individual animal tissues;
however, this is a subjective assessment and does not support
a definitive diagnosis. Clinical signs, such as vaginal discharge
or vulva enlargement, can serve to further support zearalenone
toxicity, but this too is a nonspecific indication. Sampling
feed or feed products is the only diagnostic tool available.
In order to maximize the probability of isolating zearalenone
or any mycotoxin, the investigating veterinarian should utilize
several steps to aid in his/her efforts. Feed samples
taken should reflect the feed/forage utilized during the time
period of the problem. This is not always possible, especially
if low-level contamination is present. Clinical signs indicating
a problem may not appear until weeks after the infected feed
was consumed. Samples should be representative of the entire
product being fed. Zearalenone levels can vary dramatically
from areas in the same storage unit, or even among kernels
on the same ear of corn! Even if moldy appearing areas are
present, sampling of normal appearing feed (representative
sample) should occur because normal appearing areas can be
more severely affected. A good time to sample is after blending
(such as auguring grain) has occurred. Periodic sampling
of flowing grain/feed is recommended. Once individual stream
samples have been collected, these can be combined and a subsample
of at least 10 pounds should be submitted. Samples should
be submitted dry or frozen and protected from light.
Heat, chemicals, and sunlight all have the potential to alter
mold metabolites from their original structure and activity.
Once samples have been frozen or refrigerated they should
remain that way. Submit individual feeds rather than mixed
feeds if at all possible. Isolation of toxins in mixed
feed is often difficult because of the complex nature of mixed
feeds. If individual components are not available, a list
of ingredients should be supplied to the diagnostic lab.
As with any other diagnostic test, quality sampling and sample
submission is of the utmost importance.
Several testing options are available for the diagnosis
of zearalenone. Thin Layer Chromatography (TLC) is one of
the earliest and simplest analytical methods that has been
developed. Because TLC does not require expensive analytical
machines and is relatively uncomplicated, most laboratories
can utilize TLC as a diagnostic tool. The Purdue ADDL utilizes
this method for determination of mycotoxins. The process
of TLC consists of extracting solvents, sample clean-up/purification,
solvents for separation, and detection methods. Because of
the reliability, cost effectiveness, and quick results, TLC
is utilized in many diagnostic laboratories A disadvantage
of TLC is that the results are semi-quantitative.
High Performance Liquid Chromatography (HPLC) is another
option for the diagnosis of mycotoxins. The sensitivity,
accuracy, and quantification ability of this test has made
HPLC popular in recent years. Cost of sampling is increased
utilizing this method due to the expense of owning/operating
the HPLC machine. HPLC samples also require extensive clean
up, similar to TLC; however, an advantage is that quantitative,
rather than semi-quantitative results, are obtained with HPLC.
Conclusion:
Dairy cow and heifer productivity can be greatly altered
by the presence of zearalenone in feedstuffs. Diagnosing
and isolating zearalenone is often a difficult and frustrating
problem. A basic knowledge of fungal dynamics, proper sampling
techniques, sample handling, and persistence are important
aids in the diagnosis of mycotoxicosis. Several testing modalities
are available at various diagnostic laboratories. Diagnosis
of zearalenone-induced estrogenic effects is based on history,
clinical signs, and detection of zearalenone in feed. Treatment
is based on removal of the contaminated feed and replacement
with high-quality feedstuffs. While the incidence of zearalenone
toxicity varies greatly, producers and veterinarians should
be aware of this estrogenic substance and its effects on dairy
reproductive health.
-by Christopher Witte, Class of 2003
-edited by Dr. Steve Hooser, Chief of Toxicology, ADDL
References
1. Abramson et al: 1997. Mycotoxins in Fungal Contaminated
Samples from Animal Feed from Western Canada, 1982-1994.
Canada J of Vet Research 61:49-52.
2) Diekman M and Green M: 1992. Mycotoxins and Reproduction
in Domestic Livestock. J of Anim Sci 70 : 1615-1627.
3) Osweiler et al: 1985. Clinical and Diagnostic Veterinary
Toxicology, 3rd ed. 410-436
4) Schuh M: 1997. Clinical and Subclinical Events Related
to the Presence of Mycotoxins in Cattle Feed. Israel J of
Vet Med 52: 93-97.
5) Sharma R and Dattajiao S: 1991. Mycotoxins and Phytoalexins.
36-40.
6) Weaver et al: 1986. Effect of Zearalenone on Dairy Cows.
Am J of Vet Research 47: 1826-1828.
7) Whitlow et al: 1998. Mycotoxin Occurrence in Farmer
Submitted Samples of North Carolina Feedstuffs: 1989-1997.
J of Dairy Science 81,4: 1189.
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