CHAPTER 1

INTRODUCTION
Aflatoxins are a toxic metabolites of Aspergillus flavus and Aspergillus parasiticus
fungi which, are certainly occurring contaminants of food. Even though aflatoxins
were a hassle throughout records, they have got most, effectively diagnosed as
large contaminants in agriculture since the Nineteen Sixties. Establishing
regulatory limits on traded foods, enforcing those limits through food tracking, and
implementing the most appropriate drying and garage practices have generally
removed dangerous exposures in evolved international locations (Brown et al.
1999). The software of those strategies in developing international locations is
difficult due to differences in food production, such as the prominence of
subsistence farming in developing international locations. Furthermore, those
international locations frequently lack the sources, generation, and infrastructure
necessary for habitual food tracking as well as the most appropriate drying and
garage practices. Therefore, 5 billion people in developing international locations
worldwide are at risk of continual publicity to aflatoxins through contaminated
foods (Williams et al. 2004). Aflatoxin associated health outcomes pervade the
developing global. Those outcomes might be mitigated or averted through the
powerful and integrated use of modern agricultural expertise and public health
exercise. The dialogue of this hassle and its remedies need to consist of the
underlying query of food insufficiency and extra well-known economic demanding
situations in developing international locations. Outbreaks of acute aflatoxin
poisoning are a recurrent public health hassle. In 2004, one of the most important,
most intense aflatoxicosis outbreaks occurred in Kenya, accompanied by another
outbreak in 2005 [centers for sickness manage and Prevention (CDC) 2004,
unpublished records]. Each outbreak was because of infection of inadequately
stored, homegrown maize. Given that diseases inside the developing global
frequently pass unreported, the Kenya outbreaks in all likelihood represent the
most effective portion of the hassle. The whole burden of sickness because of
continual aflatoxin publicity [e.g., hepatocellular carcinoma (HCC), impaired
increase, immune suppression] stays undefined. Those outbreaks emphasize the
want to quantify and manage aflatoxin publicity in developing international
locations and highlight the ability function of public health services. In July 2005,
the CDC and the world health organization (WHO) convened a workgroup of
experts to discover culturally appropriate, long- time period public health strategies
to reduce aflatoxin publicity in developing international locations. The forty
individuals blanketed internationally diagnosed scientists from various
backgrounds (public health, agriculture, animal health, change, and social
technology). Additionally, they blanketed key public health officers and
stakeholders from international locations closely suffering from aflatoxins.
The workgroup individuals identified gaps in modern expertise approximately the
acute and continual human health outcomes of aflatoxins. Additionally, they
reviewed surveillance and food tracking schemes, analytic techniques, and the
efficacy of intervention strategies. Individuals discussed public health strategies
that would complement agricultural efforts to reduce or prevent public exposure to
aflatoxins in the developing global community. Ultimately, the workgroup
discussed areas wherein efforts should be concentrated to reduce aflatoxin
publicity and fill gaps in modern expertise. Aflatoxins are toxic secondary
metabolites produced through Aspergillus fungi. Aflatoxin B1 (AFB1), a
recognized human carcinogen, is the most powerful and doubtlessly lethal
metabolite. Over forty years Agricultural scientists were reading about aflatoxins
because of the big prevalence of their contaminants and their large impact on crops
(Williams et al. 2004). Aflatoxins can affect a huge range of commodities, which
include cereals, oilseeds, spices, tree nuts, milk, meat, and dried fruit. Maize and
groundnuts are important sources of human publicity due to their extra
susceptibility to infection and frequent consumption throughout the sector.
Aflatoxins are most common every day in latitudes between 40° N and 40° S of the
equator, but the best health threat lies within developing international locations in
tropical areas, which depend upon those commodities as their staple food source.
Food insufficiency and the absence of diversity drastically make a contribution to
the susceptibility of people and groups to aflatoxins. Infection is prompted by
many factors and might occur at any degree of food production, from pre-harvest
to garage (Wilson and Payne 1994). Factors that affect aflatoxin infection consist
of the climate of the place, the genotype of the crop planted, soil kind, minimal and
most everyday temperatures, and everyday internet evaporation (Bankole and
Mabekoje 2004; Fandohan et al. 2005).
Aflatoxin infection is also promoted through strain or harm to the crop because of
drought before harvest, insect hobby, negative timing of harvest, heavy rains at and
after harvest, and insufficient drying of the crop before garage (Hawkins et al.
2005; Hell et al. 2000; Ono et al. 2002; Turner et al. 2005). Tiers of humidity,
temperature, and aeration during drying and garage also are critical factors. Acute
publicity to aflatoxins can bring about aflatoxicosis, which manifests as intense,
acute hepatotoxicity with a case fatality rate of approximately 25% (Cullen and
Newberne 1994). Early hepatotoxicity symptoms from aflatoxicosis can include
anorexia, malaise, and low-grade fever. Acute high-level public exposure can
progress to doubtlessly lethal hepatitis with vomiting, belly pain, jaundice,
fulminant hepatic failure, and loss of life. Outbreaks of acute aflatoxicosis are a
routine public health hassle in many developing international locations, including
Kenya and India. (CDC 2004; Krishnamachari et al. 1975a, 1975b; Lye et al. 1995;
Ngindu et al. 1982). HCC due to continual aflatoxin publicity has been properly
documented, usually in affiliation with hepatitis B virus (HBV) or other threat
factors (Chen et al. 2001; Henry et al. 2002; Omer et al. 2004; Qian et al. 1994;
Wang et al. 1996). The worldwide employer for research on most cancers (IARC)
first diagnosed aflatoxins as carcinogenic in 1976. It ultimately reaffirmed
certainly occurring combinations of aflatoxins and AFB1 as organization 1 cancer-
causing agents (carcinogenic to humans) (IARC 2002). Extra outcomes of
continual publicity have no longer been widely studied, however, are thought to
consist of immunologic suppression, impaired increase, and nutritional interference
(Cullen and Newberne 1994; Fung and Clark 2004; Patten 1981; Williams et al.
2004).
CHAPTER 2
2.0 AFLATOXINS

Aflatoxins are one of the highly toxic secondary metabolites derived from
polyketides produced via fungal species which includes Aspergillus flavus, a.
parasiticus, and a. nomius (Payne and Brown, 1998). these fungi generally infect
cereal crops such as wheat, walnut, corn, cotton, peanuts, and tree nuts (Severns et
al., 2003), and may result in serious threats to human and animal health through
causing numerous complications that include toxicity, teratogenicity, and
immunotoxicity) (Roze et al., 2013).
The predominant aflatoxins are B1, B2, G1, and G2, which can poison the body
through respiratory, mucous, or cutaneous routes, ensuing in the over-activation of
the inflammatory reaction (Romani, 2004).
The safety of food is one of the primary issues presently facing the agriculture and
health sector; as a result, a variety of research has been conducted to discuss
methods of addressing client issues with numerous elements of meal protection
(Nielsen et al., 2009). Seeing that 1985, the United States Food and Drug
Administration (USFDA) confined the number of mycotoxins approved in meal
products. The USDA Grain and Plant Inspection carrier (GPIS) has carried out a
carrier laboratory for the inspection of mycotoxins in grains. Additionally, the
Food and Agricultural Organization (FAO) and the world health organization
(WHO) have diagnosed many pollutions present in agricultural products. While
mycotoxins are infected into ingredients, they can't be destroyed by using regular
cooking techniques. but, there had been many latest advances in food processing
advanced to hold very last meals products secure and healthy, such as risk analysis
of critical management factors (HACCP) and correct manufacturing practices
(Maldonado-Siman et al., 2014). Furthermore, several bodily, chemical, and
organic strategies may be applied to in part or absolutely dispose of the pollution
from food and guarantee food safety and health issues.

2.1 CHEMISTRY AND BIOSYNTHESIS OF AFLATOXINS

The chemical nature of aflatoxins is such that a bifuran group is attached at one
side of the coumarin nucleus, while a pentanone ring is attached to the other side in
the case of the AFTs and AFTs-B series, or a six-membered lactone ring is
attached in the AFTs-G series (Nakai et al., 2008). The physical, biological, and
chemical conditions of Aspergillus influence the production of toxins. Among the
20 identified AFTs, AFT-B1, and AFT-B2 are produced by A. flavus, while AFT-
G1 and AFT-G2 along with AFT-B1 and AFT-B2 are produced by A.
parasiticus (Bennett and Klich, 2003). AFT-B1, AFT-B2, AFT-G1, and AFT-G2 are
the four major naturally produced aflatoxins (Pitt, 2000). AFTs-M1 and AFTs-
M2 are derived from aflatoxin B types through different metabolic processes and
are expressed in animals and animal products (Wolf-Hall, 2010). AFT-B1 is highly
carcinogenic (Squire, 1981), as well as heat resistant over a wide range of
temperatures, including those reached during commercial processing conditions
(Sirot et al., 2013).
The biosynthetic pathway of aflatoxins consists of 18 enzymatic steps for
conversion from acetyl-CoA, and at least 25 genes encoding the enzymes and
regulatory pathways have been cloned and characterized (Yu et al., 2002; Yabe
and Nakajima, 2004). The gene comprises 70 kb of the fungal genome and is
regulated by the regulatory gene, (Price et al., 2006). The metabolic grid is
involved in aflatoxin biosynthesis (Yabe et al., 1991, 2003). Hydroxyversicolorone
(HVN) is converted to versiconal hemiacetal acetate (VHA) by a cytosol
monooxygenase, in which NADPH is a cofactor (Yabe et al., 2003).
Monooxygenase is encoded by the moxY gene, which catalyzes the conversion of
HVN to VHA and the accumulation of HVN and versicolorone (VONE) occurs in
the absence of the moxY gene (Wen et al., 2005).

2.2.0 SOURCES OF AFLATOXINS

The fundamental sources of aflatoxins are fungi including a. flavus, a. parasiticus,
and a. nomius (Kurtzman et al., 1987), however they're also produced through
other species of Aspergillus as well as Emericella spp. (Reiter et al., 2009). There
are more than 20 known aflatoxins, however, the four principal ones are aflatoxin
B1 (AFB1), aflatoxin B2 (AFB2), aflatoxin G1 (AFG1), and aflatoxin G2 (AFG2;
Inan et al., 2007), at the same time as aflatoxin M1 (AFM1) and M2 (AFM2) are
the hydroxylated metabolites of AFB1 and AFB2 (Giray et al., 2007; Hussain and
Anwar, 2008).

2.2.1 OCCURRENCE OF AFLATOXINS IN FOOD

There are six common mycotoxins that affect animals: aflatoxins, fumonisins,
ochratoxins (which like aflatoxins affect liver function), trichothecenes, and
zearalenone. Diagnosis of aflatoxin exposure in animals is difficult, especially in
large farms that use mixed feed, which may also comprise quite various
combinations of feedstuffs.
Cereals and cereal-based products are the primary foods for human consumption
worldwide (Temba et al., 2017). Amongst cereals, rice and corn are generally
infected by AFs in natural conditions due to modifications in agricultural practices.
The AFs are produced both in pre and submit harvest conditions (Hesseltine,
1974). Filazi and Sireli (2013) mentioned rice is more susceptible to AFs
contamination in comparison to other cereals. The fungal growth happens due to
improper drying of rice grains maintaining better moisture content greater than
14%. As a result, these fungi cause discoloration of grain and/or husk together with
deteriorating the high-quality of the grains. Groundnut and beans, however, are
often used in lots of African diets to supplement cereal diets (Soro-Yao et al.,
2014). But, these are highly susceptible to AFs infection both in field and storage
conditions (Lombard, 2014). The quantity of fungal growth and AFs production in
cereals depends on temperature, moisture, soil type, and storage conditions
(Achaglinkame et al., 2017). Similarly, spices are susceptible to AFs
contamination and are significantly affected by storage and processing conditions.
Elshafie et al. (2002) mentioned the AFs contamination in an extensive type of
spices which includes black pepper, cardamom, cinnamon, clove, cumin,
coriander, and ginger within the Sultanate of Oman. Moreover, Tchana et al.
(2010) mentioned the presence of AFs in eggs accumulated from the chicken farm
and in raw cow milk in Cameroon. Consequently, the affected plants permit AFs to
enter the meals chain, which may be very much influenced by climatic situations.
As in humans, animals exposed to high levels of aflatoxin-contaminated feed have
been known to exhibit a severe form of “intoxication,” which can lead to death.
Usually, however, exposure in animals is of a “sub-clinical” level, which leads to
liver damage, reduced weight gain, and lost productivity (declines in egg and milk
production) resulting in economic losses to the industry. Aflatoxins affect livestock
growth, reproduction, immune functioning, and ability to metabolize vaccines
Anthony Makun Hussaini, et al. (2012). Poultry and fish are most affected, but
there is also a lot of concern about B1 contamination in milk, given that it is often
fed to infants and young children (Fink Gremmels et al. 2012).
The effects of animal feed on meat and other byproducts: Consumption of
aflatoxin-contaminated animal feeds varies with preparation and the type of
product. Anthony et al. reviewed studies in which the prevalence of aflatoxins was
compared across beef and edible organs that were either fresh or sun dried.
Consistently, organ meats (especially kidney) were more contaminated than beef,
while fresh products (as opposed to sundried products) maintained higher aflatoxin
contamination levels.
Time of harvest has been shown to be important in influencing the occurrence and
levels of aflatoxin because Aspergillus does not compete well with other molds
when corn presents more than 20% moisture. Harvesting corn when moisture
content is above 20% followed by rapid drying to at least 14% moisture content
within 24 to 48 hours of harvest can inhibit Aspergillus growth and toxin
production. Contaminated grains and their byproducts are the most common
sources of aflatoxin. Corn silage may also be a source of aflatoxins because the
ensiling process does not destroy toxins already present in silage (Cassel EK et al.
2012).
On the farm, more than one mold or toxin may be present in the contaminated feed,
which often makes the definitive diagnosis of aflatoxicosis is difficult. The
prognosis of aflatoxicosis depends on the severity of liver damage. Once overt
symptoms are noticed the prognosis is poor. Treatment should be directed at the
severely affected animals in the herd and further poisoning prevented. Aflatoxicos
is typically a herd rather than an individual cow problem. If aflatoxicos is
suspected, feed should be analyzed immediately. If aflatoxins are occurred, the
source should be eliminated immediately. Levels of protein in feed and vitamins A,
D, E, K and B should be increased as the toxin binds vitamins and affects protein
synthesis. Good management practices to alleviate stress are essential to reduce the
risk of secondary infections which must receive immediate attention and treatmen
(Cassel EK et al. 2012).
Importantly, it has been demonstrated that simple measures can significantly
reduce the risk of mycotoxin exposure on farms. Storage of grain at the appropriate
moisture content (below 130 kg-1), and inspection of grain regularly for
temperature, insects, and wet spots will limit the possibility of fungal development
in feeds and feedstuffs as discussed before. The risk of feed contamination will be
reduced in animal units with rapid turnover of feed because there will be less time
for fungal growth and toxin production (Bryden WL, 2012).  Aflatoxin is just one
of many mycotoxins that can adversely affect animal health and productivity. Care
regarding animal feed must be extended not only to the nutritional and economic
value, but also to fod quality (Bryden WL, 2012).  
The presence of molds in foodstuffs causes the appearance of flavors and odors
that reduce palatability and affect feed consumption by animals as well as reduce
the nutritional value of foods. Mycotoxins, in turn, affect the digestion and
metabolism of nutrients in animal production, resulting in nutritional and
physiological disorders, besides a negative effect on the immune system. Huff WE.
(2018)

2.2.2 AFLATOXINS OCCURRENCE IN ANIMAL PRODUCTS

When focusing on how mycotoxins play a role in food safety, attention should be
limited to mycotoxins that are known to be transferred from feed to food of animal
origin, as this food represents a significant route of exposure for humans (FAO and
IFIF 2010)Apart from their toxicological effects in affected animals, the carry-over
through animal derived products, such as meat, milk, and eggs into the human food
chains is an important aspect of mycotoxin contamination. FAO has estimated that
up to 25% of the world’s food crops and a higher percentage of the world’s animal
feedstuffs are significantly contaminated by mycotoxins.
Aflatoxin or ochratoxin residues in meat are uncommon and rarely found.
However, it’s more common in organs, especially the liver. This organ may have
its lipid content increased over threefold when 20 ppm aflatoxin is incorporated in
broiler feed (ASR, et al. 2010)
The problem the egg production is that the long-term or short-term hen’s exposure,
via dietary sources, too low concentrations of certain mycotoxins causes
contamination of eggs. This is the wit of aflatoxins, which have a high impact in
both, human and animal health, causing significant losses in the egg industry,
considering the deleterious effect on egg production and quality. In laboratory
studies, it was proved that aflatoxin can decrease egg production and increase liver
fat (fatty liver syndrome). This classical study established the typical symptoms
associated with acute or chronic aflatoxicosis is, observed until today in field
conditions. Huff WE.(2018)
A distinctive sequence of events during acute aflatoxicosis is in laying hens (30
weeks old) in a four week experiment with increasing aflatoxin doses in the diet of
0; 1.25; 2.5; 5.0 and 10.0μg g-1 Huff WE.(2018) Results indicated that egg
production was decreased by about 70% from the control value at 10μg g-1
concentration in the diet and the liver size was increased significantly by 5 and 10
μg g-1 dietary concentrations of aflatoxin and the liver lipid increasing
dramatically by a smaller dose of 2.5 μg g-1. Shows the dramatic effect of
aflatoxin on liver function (Hamilton PB and Gralich JD, 1971). The obtained data
suggest that plasma and yolk lipids respond to the inhibition of lipid synthesis and
transport from the liver during aflatoxicosis induced by the dietary treatments. The
liver malfunction results in an increase in its fat content and a decrease in the levels
of plasma lipids.
A review carried out in Brazil showed high variability among the results. For
instance, corn contamination with aflatoxins reached 906 ppb, above those levels
allowed by legislation (20 ppb). This fact indicates the need for quality control in
the reception of this ingredient in the feed mill with the use of rapid tests for
mycotoxins. Regarding products of animal origin, major problems were not
observed in the eggs and tissues of swine and poultry. However, among chicken
liver samples 50% tested positive but with relatively low levels. Anyway, attention
should be paid to liver consumption when there are pieces of evidence of corn
contamination. (Rodriguez-Amaya DB, Sabino M, 2002)

2.3 HUMAN AFLATOXICOSIS

Assisting the priority for the feasible carcinogenicity of aflatoxin turned into the
truth that the rat and rainbow trout have been observed to be quite liable to
aflatoxin tumorigenesis. But, the excretion sample of the rhesus monkey turned
into observed to be extraordinary from that of the rat, and outcomes in primates
have been scarce and handiest proved they have been lots much less liable to
aflatoxin tumorigenesis than rats. Adamson et al. (1973) initiated a mixed
intercourse and species take a look at related forty rhesus and cynomolgus
monkeys. through 44 monkeys of age, one lady who had received as much as 800
f-lg/kg three times every 2 weeks has become progressively jaundiced and
inactive, turned into killed, and primary liver carcinoma turned located. Campbell
and Stoloff (1974) collated the primate studies, and their research emphasized the
variability in outcomes of those studies.
In 1966- 67, potential human epidemiology studies have been initiated in Africa
when Alpert and coworkers (1969) surveyed the prevalence of primary hepatoma
in Uganda and determined aflatoxin tiers in meal samples stored for consumption
between harvests (Alpert et al. 1969). The frequency of aflatoxin infection of
market meal samples turned into definitely associated with liver most cancers
prevalence in localized populace agencies. Coincident with the considerations of
the feasible role of aflatoxins in hepatocellular carcinoma turned into an attempt to
tie it to different distressing human ailments of unknown etiology, for example,
kwashiorkor and Reye's syndrome.
There is not any commonly agreed definition of kwashiorkor, but edema associated
with hypoalbuminemia is universally commonplace because of the vital minimum
criterion for diagnoses (Hendrickse 1984). Kwashiorkor turned into the first notion
to be due to protein deficiency in the presence of relative carbohydrate excess.
when this speculation have become untenable, it turned referred to those animals
receiving aflatoxin experienced several outcomes continuously observed in
kwashiorkor, i.e., hypoalbuminemia, fatty liver, and immunosuppression, and that
" the sector distribution and seasonal fluctuations of mycotoxins infection of meals
are remarkably just like the geographical distribution and seasonal presence of
kwashiorkor which has a tendency to height in the moist season in many regions"
(Hendrickse et al. 1983). For example, aflatoxin AFB1 turned into detected from
post-mortem liver specimens in all 36 children who had kwashiorkor (Hendrickse
1985). But, Hendrickse (1985) concluded "that our findings in those studies set up
an affiliation between aflatoxins and kwashiorkor. The nature of this affiliation is
difficult to understand and is being investigated." Frustration from the failure of
medicine to prevent and reliably treat kwashiorkor made the affiliation with dietary
aflatoxin easy to take to the next step of motive and effect courting. Reye's
syndrome, an encephalopathy with fatty degeneration of the liver, turned into
defined by Reye et al. (1963). Following scattered reviews of aflatoxin in tissues of
sufferers of Reye's syndrome, Ryan et al. (1979) the use of formalin-fixed tissue
observed a really strong affiliation between children’s death with Reye's syndrome
and aflatoxin isolation from the liver. but, Rogan et al. (1985), following research
through the centers for sickness Management, concluded that "we do now not
agree with that most or many instances of Reye's syndrome in the U.S. are due to
aflatoxin exposure; right now we think it prudent to view reviews of aflatoxin in
the tissues as signs of exposure of uncertain significance for etiology." In any other
look for institutions, the workplace of Rural fitness in Georgia (Caster et al. 1986)
stated that the consumption of huge amounts of com, rice, peanuts, and milk
( meals considered to be probably high in aflatoxins) turned into notably related to
the mental retardation of children in one county having high tiers of aflatoxins in
the meals supply. No such courting turned into observed in any other county
having trivial tiers of aflatoxins «20 ppb) in its food plan. Acute poisonous liver
damage in people has been ascribed to aflatoxin. The most vast and widely
publicized case turned into a plague in Western India in October 1974, where
unseasonal rains led to vast mildew damage to some crops with extremely high
concentrations of aflatoxin observed to be 6.3-15.6 ppm from affected households.
The ailments have been characterized by jaundice, rapidly developing ascites,
portal high blood pressure, and a high mortality rate.
Dietary calculations and the evaluation of infected samples showed that affected
human beings ought to have been fed between 2 and 6 mg of aflatoxin daily over a
period of a month. Vast bile duct proliferation, periportal fibrosis, and occasional
multinucleated giant cellular are usual for aflatoxin-caused sickness in animals, it
has been observed in the histopathologic examination of liver specimens
(Krishnamachari et al. 1975b). There may be inhalation exposure to aflatoxin
among business and agricultural employees. The presence of aflatoxin in dust from
infected crops has been set up (Burg and Shotwell 1984; Burg et al. 1981; Silas et
al. 1987). Because of the sporadic distribution of aflatoxin in com and different
infected commodities, it turned difficult to as it should be quantitated the infection.
In business settings, two Czechoslovakian chemical engineers died running on a
technique for sterilizing peanut meal (Dvorackova et al. 1981) and two British
biochemists advanced adenocarcinomas of the colon following exposure to
purified aflatoxins (Deger 1976). Van Nieuwenhuizen et al. (1973) diagnosed
several most cancers instances among a small group of employees in a peanut
processing plant in the Netherlands and associated them with the locating of
aflatoxins at three 300- 400 mg/kg in the peanut meal.
Hayes et al. (1984) carried out a follow-up take a look at employees on this plant.
The test subjects extracted oil from peanuts and linseed oil, while the controls
labored in a nearby com processing plant and, because of the coping with
equipment in use, possibly had lots much less dust exposure. The located mortality
due to most cancers turned into stated to be better than expected for the aflatoxin
exposed employees in the course of the entire period. But, the tumors have been at
a variety of infected sites without a primary tumor of the liver, but there was a
much more than expected number of tumors of the respiration device in the
aflatoxin exposed group, with seven such deaths. The amount to which the
development of tumors at individual turned into associated with aflatoxin exposure
couldn't be detennined. There are several widely stated epidemiological studies
trying to hyperlink hepatocellular carcinoma with high intakes of aflatoxin B 1
from the food plan. In the early group of studies, aflatoxin intakes have been
sampled in populations of extraordinary geographical regions and the outcomes
correlated with p.c prevalence in those regions. those studies have been through
Shank et al. (1972) in Thailand, friends and Linsell (1973) in Kenya, Van
Rensburg et al. (1974) in Mozambique, friends et al. (1976) in Swaziland, and Van
Rensburg et al. (1985) in Mozambique/Transkei. In each of those places except
Mozambique, a couple of regions have been studied and aflatoxin exposure
assessed through the measurement of dietary intake. intake turned into now not
proven through measuring metabolites, DNA, or protein adducts in the urine of
subjects. problems preventing strict adherence to right protocol design in those
studies are mentioned through Wagstaff (1985, 1990). those encompass brief-tenn
migration of running-age adult males in Africa, nonavailability of subjects, and
problems in sporting out dependable analyses. In Kenya ( friends and Linsell
1973), the male populace over 15 year of age turned into 29% much less than the
lady populace of that group and the handiest liver most cancers instances recorded
have been those individuals who voluntarily provided themselves to a health
facility or health center. The three take a look at agencies have been now not
similar, e.g., there have been differences in altitude of the respective regions,
proximity to the health facility, economic and agricultural situations and crops
grown, and diets. In Thailand, snacks and meals in brief supply in houses,
consisting of meats, have been now not sampled. problems in case locating have
been so terrific in the Thailand take a look at that prevalence information in rural
take a look at regions have been taken from nearby urban regions. Hepatitis B virus
(HBV) popularity turned into now not detennined in any of those studies, although
the feasible role of HBV infection turned into mentioned through Van Rensburg et
al. (1985). As a consequence, except this latter take a look at from Mozambique,
the thesis turned into that aflatoxin is probably the primary causative issue in
regions of high primary liver most cancers ( p.c) prevalence and all the studies did
reveal an effective correlation between aflatoxin intake and p.c prevalence. But, in
the u.s.a., Stoloff (1983) observed a small ageadjusted excess p.c mortality ratio for
the Southeast and he puzzled whether this small excess may be attributed to
aflatoxin. The early retrospective epidemiology studies depended on the accuracy
of dietary surveys and analyses to set up aflatoxin consumption in the various
populations surveyed. Similarly, they assumed no infectious complicating elements
in the etiology of p.c. in the years for the reason that spherical of epidemiology
studies have been initiated in 1970. Epidemiological studies can now be done on a
greater clinical foundation, as a consequence assuring greater self-belief in their
outcomes. The initial locating main to the development of alternative etiologies
turned into that of Sherlock et al. (1970), who referred to the high prevalence of
hepatitis B floor antigen (HBsAg) in a clinical collection of % sufferers. The
evidence for the affiliation of p.c and HBV, aflatoxin, and different feasible chance
elements has been summarized by Wagstaff (1985) and Bassendine (1987). Of
significance in evaluating the role of HBV are such elements because the ecologic
affiliation of HBV in numerous regions in the course of the sector and case
management studies evaluating the prevalence of HBV serologic markers in p.c
instances and controls.
In the early studies, aflatoxin ingestion needed to be estimated through taking
appropriate samples of the food plan and then assaying them through a dependable
approach before the aflatoxin content modified. Understandably, there was lots
room for blunders on this system. affirmation of aflatoxin ingestion has now been
made feasible through the elucidation of its metabolism and detoxing pathways,
identification of adducts, and development of rapid, sensitive, and accurate
strategies for adduct identification summarized through Groopman et al. (1988).
An early locating turned into that of Campbell et al. (1970), who stated that
aflatoxin M, turned into detectable in the urine of individuals who fed on greater
than 30 mg AFB,/d. Later, after it turned into recognized that reactive electrophilic
epoxides of many xenobiotics can covalently react with various nucleophilic
centers in macromolecules, like DNA, RNA, and protein, this type of pathway
turned into additionally recognized for aflatoxin. the primary foremost AFB,-DNA
adduct turned into diagnosed through Essigmann et al. (1977) in an in vitro device;
but, its presence turned into soon showed in vivo. The 1/2- lifestyles of this adduct
is approximately 12 hr, which critically limits its usefulness in confirming
aflatoxin ingestion. Autrup et al. (1983) stated the presence of a putative AFB,-
Guanine product in urine accumulated from people residing in regions with high
liver most cancers chance and suspected exposure to aflatoxin. Aflatoxin B, binds
to proteins as well as to DNA. The response with one or greater lysine residues in
serum albumin money owed for greater than 1/2 of the general binding to this
protein (Sabbioni et al. 1987).
This adduct is a completely changed aflatoxin structure maintaining handiest the
coumarin and cyclopentene jewelry of the figure compound. Protein adducts are
substantial in organic monitoring because they constitute the integrated dose of
aflatoxin received over many preceding weeks. The average 1/2- lifestyles of
albumin in people is 20 d; as a consequence, an accumulated dose of aflatoxin will
be present after dietary exposure has ceased (Groopman et al. 1988). Monoclonal
antibodies are correctly used as the idea for assessments to decide aflatoxin in
meals and organic samples. Those fast, noticeably cheaper, and without problems
performed assessments allow the practical monitoring of human exposure to
aflatoxin in field epidemiological studies (Groopman and Donahue 1988 and
Gamer et al. 1985). Monoclonal antibody columns, coupled with HPLC, can
quantify aflatoxin-DNA adducts in human urine samples received from
environmentally exposed people (Groopman et al. 1988). Those assessments
require strict satisfactory manage requirements, e.g., using an internal general is
vital for meaningful outcomes; otherwise, variability in the assay may be high. The
usefulness of this sensitive test nonetheless depends on ok sampling of the food
plan of all test agencies and the constancy of this food plan through the years,
prospectively and retrospectively, as required through the specific protocol. a
monoclonal antibody- based totally approach to measure M, in urine turned into
utilized by solar et al. (1986) and Zhu et al. (1987) to document and quantify
human exposure. Zhu observed right correlation with food plan handiest in male
subjects. The international corporation for research in most cancers (IARC) has
made several reviews on the carcinogenicity of aflatoxin beginning in 1976. this
primary report said that " studies of liver most cancers prevalence concerning
aflatoxin intake offer circumstantial evidence of a casual courting." but, through
1987, any other IARC running group concluded that there was " sufficient
evidence that aflatoxin is a likely human carcinogen." a later group of
epidemiological studies- solar and Chu (1984), Yeh et al. (1989), and friends et al.
(1987)-which stated an aflatoxin effect impartial of HBsAg prevalence, have been
to be had to the latter running group and possibly influenced their conclusion
(Campbell et al. 1990). But, the solar and Chu report contained no aflatoxin intake
information and the Yeh report as compared handiest one “high" and one "low"
village in the Guangxi region of China. In the 1/3 take a look at, friends and
coworkers reinvestigated 4 regions in Swaziland studied in advance and have been
the primary to utilize each meals measurements of aflatoxin intake and HBV
exposure of the equal populace. But, they needed to estimate the consumption of
peanuts, which have been through a long way the most infected food, and the
prevalence of primary liver most cancers turned into notably low, being handed
through cervical and esophageal most cancers. In any other current take a look at,
Autrup et al. (1987) stated no aflatoxin effect on %prevalence quotes when all
ethnic, social, and cultural agencies have been combined but a effective affiliation
turned into referred to many of the Bantu human beings. Autrup et al. (1987)
additionally measured HBsAg carrier quotes. Yeh et al. (1989) stated on their take
a look at of HBV popularity, aflatoxin exposure, and %prevalence carried out in
southern Guangxi China between July 1982 and June 1987. In a cohort of
seven,917 men aged 25- 64 year, a complete of 149 deaths occurred, seventy six of
which ( fifty one %) have been due to p.c. prevalence of HBsAg positivity in %
deaths turned into ninety one %, in contrast to 23% of all individuals of the cohort.
but, the authors saw no HBsAg effect in the move-sectional case evaluation. there
was an almost linear courting between estimated aflatoxin B J exposure in the
subpopulations and corresponding mortality quotes of p.c. but, handiest the
average nearby aflatoxin exposure values have been to be had and now not the
individual exposure values. The brand new epidemiology take a look at to be stated
that tested the affiliation with dietary aflatoxin, among different elements, is
happily a long way and away the most comprehensive, with 6,500 participants.
This take a look at through Campbell et al. (1990) turned into a comprehensive
move-sectional survey in the human beings's Republic of China on the feasible
chance elements for primary liver most cancers. It blanketed forty eight survey
web sites, an aflatoxin exposure range of about six hundred-fold, a 39-fold range of
% mortality quotes, and a 28-fold range of HBsAg+ carrier prevalence. Although
the accuracy of the urinary assay used for measuring aflatxoin metabolites has
been puzzled, this take look at did estimate exposures for a huge number of
dietary, dietary, and lifestyle variables. It represents a much larger number of
survey web sites, a much wider range of p.c mortality quotes, and a much wider
range of aflatoxin exposure than in the preceding surveys. Similarly, aflatoxin
consumption turned into estimated through measuring aflatoxin metabolites in
urine. In Campbell's take a look at, in contrast to the stated outcomes of some of
the preceding studies, there was no affiliation between aflatoxin exposure and p.c
mortality. There was additionally a lack of affiliation of % mortality with com and
moldy peanuts, the two meals in the food plan typically infected with aflatoxin.
But, % mortality turned into definitely correlated to the prevalence of HBsAg and
different elements. Notably, in the USA Stoloff (1983) observed a small age-
adjusted excess %mortality ratio for the Southeast. But, there has been a serious
question of whether the excess % mortality may be attributed to aflatoxin.

2.4 MECHANISM OF TOXICITY AND HEALTH EFFECTS OF

AFLATOXIN
Aflatoxin are specifically target the liver organ (Abdel-Wahhab et al., 2007). Early
symptoms of hepatotoxicity of liver caused by aflatoxins comprise fever, malaise
and anorexia followed with abdominal pain, vomiting, and hepatitis; however,
cases of acute poisoning are exceptional and rare (Etzel, 2002). Chronic toxicity by
aflatoxins comprises immunosuppressive and carcinogenic effects. Evaluation of
the effects of AFT-B1 on splenic lymphocyte phenotypes and inflammatory
cytokine expression in male F344 rats have been studied (Qian et al., 2014). AFT-
B1 reduced anti-inflammatory cytokine IL-4 expression, but increased the pro-
inflammatory cytokine IFN-γ and TNF-α expression by NK cells. These findings
indicate that frequent AFT-B1 exposure accelerates inflammatory responses via
regulation of cytokine gene expression. Furthermore, Mehrzad et al.
(2014) observed that AFT-B1 interrupts the process of antigen-presenting capacity
of porcine dendritic cells, suggested this perhaps one of mechanism of
immunotoxicity by AFT-B1.
Aflatoxins cause reduced efficiency of immunization in children that lead to
enhanced risk of infections (Hendrickse, 1997). The hepatocarcinogenicity of
aflatoxins is mainly due to the lipid peroxidation and oxidative damage to DNA
(Verma, 2004). AFTs-B1 in the liver is activated by cytochrome p450 enzymes,
which are converted to AFTs-B1-8, 9-epoxide, which is responsible for
carcinogenic effects in the kidney (Massey et al., 1995). Among all major
mycotoxins, aflatoxins create a high risk in dairy because of the presence of their
derivative, AFTs-M1, in milk, posing a potential health hazard for human
consumption (Van Egmond, 1991; Wood, 1991). AFTs-B1 is rapidly absorbed in
the digestive tract and metabolized by the liver, which converts it to AFT-M 1 for
subsequent secretion in milk and urine (Veldman et al., 1992). Although AFTs-
M1 is less mutagenic and carcinogenic than AFTs-B 1, it exhibits high genotoxic
activity. The other effects of AFTs-M1 include liver damage, decreased milk
production, immunity suppression and reduced oxygen supply to tissues due to
anemia (Aydin et al., 2008), which reduces appetite and growth in dairy cattle
(Akande et al., 2006). Several studies have shown the detrimental effects of
aflatoxins exposure on the liver (Sharmila Banu et al., 2009), epididymis (Agnes
and Akbarsha, 2001), testis (Faisal et al., 2008), kidney and heart (Mohammed and
Metwally, 2009; Gupta and Sharma, 2011). It has been found that aflatoxin
presences in post-mortem brain tissue (Oyelami et al., 1995), suggested that its
ability to cross the blood brain barrier (Qureshi et al., 2015). AFTs also cause
abnormalities in the structure and functioning of mitochondrial DNA and brain
cells (Verma, 2004). The effects of aflatoxin on brain chemistry have been
reviewed in details by Bbosa et al. (2013). Furthermore, few reports have described
the effects of AFTs-B1 administration on the structure of the rodent central nervous
system (Laag and Abdel Aziz, 2013).
The liver toxicology of aflatoxin is also a critical issue (IARC, 2002; Iqbal et al.,
2014). Limited doses are not harmful to humans or animals; however, the doses
that do cause-effects diverse among Aflatoxin groups. The expression of aflatoxin
toxicity is regulated by factors such as age, sex, species, and status of nutrition of
infected animals (Williams et al., 2004). The symptoms of acute aflatoxicosis
include oedema, haemorrhagic necrosis of the liver and profound lethargy, while
the chronic effects are immune suppression, growth retardation, and cancer (Cotty
and Jaime-Garcia, 2007).

2.5 METHODS OF DETECTING AFLATOXIN

The detection and quantification of aflatoxin in food and feed is a very important
aspect for the safety concerns.
The detection of AFs has been performed by the Association of Official Analytical
Chemists (AOAC) official method in food and feed samples (Kumar et al., 2017).
Among the most commonly employed methods are chromatographic methods like
thin layer chromatography (TLC) (Fallah et al., 2011), high performance liquid
chromatography (HPLC) and liquid chromatography mass spectroscopy (LCMS)
(McDanell et al., 1988; Samarajeewa et al., 1991; Herzallah, 2009), besides the
enzyme-linked immunosorbent assay (ELISA) (Tabari et al., 2011; Andrade et al.,
2013; Sulyok et al., 2015). However, the drawbacks of these standard methods are
that they are unsuitable for rapid and real-time applications in food and feed
samples as they are tedious, time-consuming and require skilled personnel to
operate. Therefore, rapid and robust methods like polymerase chain reaction (PCR)
and non-destructive methods based on fluorescence/near-infrared spectroscopy
(FS/NIRS) and hyperspectral imaging (HSI) have emerged for the quick and easy
detection of AFs (Tao et al., 2018).
Hussain et al. (2015) utilized the PCR technique for the molecular detection of AF
producing A. flavus from peanuts. Similarly, the avfA, omtA, and ver-1 genes
encoding the major enzymes in AF-biosynthesis were used as target genes for
detecting AFs using multiplex PCR (Yang et al., 2004). Further, PCR was
employed to detect AF-producing genes in Aspergillus species in Iranian pistachio
nuts for their aflatoxigenic effect (Rahimi et al., 2008). In addition, Kim et al.
(2014) utilized PCR, ELISA and HPLC for the detection of AFs from A.
oryzae isolated from different Korean foods. HSI uses the integration of both
imaging and spectroscopy to record spatial and spectral characteristics of a given
sample (Siche et al., 2016). The visible/near-infrared (VNIR) HSI has been utilized
for the identification of maize kernels of different varieties (Zhang et al.,
2012; Wang et al., 2016). VNIR or short-wave (SWNIR) HSI techniques are
feasible for the detection of AFs as well as identification of different fungal species
in maize (Wang et al., 2015a, b). Later, Kimuli et al. (2018b) used the VNIR-HSI
system to detect AFB1 on surfaces of maize kernels from Georgia, Illinois, Indiana
and Nebraska of United States. Chu et al. (2017) used short-wave infrared (SWIR)
HIS to detect AFB1 in single maize kernels. But as the image quality could not
effectively classify AFB1 level qualitatively in individual maize kernels, therefore,
to improve this Kimuli et al. (2018a) further combined the SWIR-HSI system with
chemometric data analysis for the better detection of AFB1 on the surfaces of
maize kernels. Furthermore, the color-encoded lateral flow immunoassay (LFIA)
technique has been used for the simultaneous detection of AFB1 as well as
fumonisins in a single test line (Di Nardo et al., 2019).
To further enhance the sensitivity and detection of AFs in food and feed,
nanoparticles (NPs) based on Au/Ag, carbon (CBNs), magnetic (MNPs), Quantum
dots (QDs), up-conversion (UCNPs), metal-organic frameworks (MOFs) as well as
hybrid nanostructures have been utilized (Xue et al., 2019). Rui et al.
(2019) prepared molecular imprinted polymers (FDU-12@MIPs) using structural
analog of AFs. This highly selective surface was used as an extraction sorbent in
conjunction with HPLC for the detection of AFs in different food and feed
samples. In addition to this, the use of biosensors compared to other
spectrophotometric or chromatographic methods allow for higher selectivity, direct
detection with minimal sample pretreatment, minimal cost, portability and on-field
analysis of mycotoxins (Rotariu et al., 2016). Selvolini et al. (2019) utilized an
electrochemical enzyme-linked oligonucleotide array for easy and quick multi-
detection of AFB1 in maize. Furthermore, assays based on aptamer have been
developed for the rapid detection of AFB1. Wang et al. (2019) successfully
detected the AFB1 spiked in wine, methanol and corn flour samples using the
simple aptamer molecular beacon assay, which has the potential for the rapid
detection of AFs in the food and feed.

2.5.1 MASKED MYCOTOXINS AS A MAJOR CONCERN IN

DETECTION
Masked mycotoxins pose a major concern in food and feed as they are not
identified and detected by the usually employed detection techniques (Kamle et al.,
2019). These are the mycotoxins produced by fungi but are modified by plant
enzymes during the infection stages. They are present in vacuoles in the soluble
form or bound to macromolecules, therefore, are unable to be identified by routine
analysis processes and referred to as masked mycotoxins (Berthiller et al., 2013).
However, the modified AFs can hydrolyze back into the toxic forms during food
processing and/or digestion process (Gareis et al., 1990; Nagl et al.,
2014; Broekaert et al., 2015). Some of these modified toxins are present in
different forms as complexes with matrix compounds, hence also referred to as
matrix-associated mycotoxins (Rychlik et al., 2014). The masked mycotoxins have
been reported to occur in Asia, Africa, America and Europe. Therefore, a high
amount of masked mycotoxins prevailing in various food and feed can pose serious
health issues to both humans and animals (Zhang et al., 2019). Therefore, the
detection of masked mycotoxins is an essential part to ensure food and feed safety.
Masked fumonisins were determined through hydrolysis where modified forms
were converted back to their free forms and subsequently analyzed and detected
through LC/MS/MS (Dall’Asta et al., 2008; Dall’Asta et al., 2009). The hydrolytic
process may involve either alkaline, acidic or enzymatic treatments (Dall’Asta et
al., 2009; Beloglazova et al., 2013; Vidal et al., 2018). However, there is less
information available on the masked AFs as most of the preference is given for the
detection of free AFs in agricultural food and feed. Therefore, methods like in
vitro digestion and hydrolysis, as applied in case of masked fumonisins, can be
carried out for masked AFs in food and feed followed by detection with
LC/MS/MS and confirmation by other methods like ELISA to ensure the food and
feed safety.

3.0 CHAPTER 3
3.1 MEASURES TO CONTROL AFLATOXIN
CONTAMINATION
Aflatoxin contamination in crops caused a serious threat to production, the food
market, health, and economics. Several approaches have been manifested to reduce
the aflatoxin contamination in crops which include various physical, chemical, and
biological methods.

3.1.1 PHYSICAL METHODS

Physical methods such as steam under pressure, dry roasting, and other cooking
methods are found to be effective in the control or to reduce the aflatoxin
contamination in many crops (Peng et al. 2018). About 40–73% reduction in
aflatoxin level was also observed by heating the seed samples on 180 °C
(Opoku 2013). When groundnut and corn seed was roasted with 30% moisture at
100 °C temperature for 2 h, there is a reduction of aflatoxin by 85% (Leong et
al. 2010). Roasting also reduced the concentration of AFB1 and AFG1 by 70 and
79% when the seeds are roasted at 150 °C for 15 min (Jalili 2016). An effective
reduction in aflatoxin concentration was also observed when the seeds were treated
with various radiations such as UV and Infrared radiation (Surekha et al. 2015).
Sunlight also plays an important role in the detoxification of AFB1 in many crops.
A reduction in AFB1 content was found in artificially infected maize (80%) and
groundnut (17%) when the seeds are exposed to sunlight for a 10–12 h period
(Rushing & Selim 2019). Consequently, gamma radiation exposure was also found
to reduce the aflatoxin level (Aquino 2011). Moisture is a significant factor in
fungal growth which leads to the production of aflatoxin in crops. Aflatoxigenic
fungi growth and aflatoxin production inhibited by drying groundnut to 6.6% and
maize to 15% or much lower moisture level within 24–48 h (Awuah & Ellis 2002;
Eziah, & &Afreh-Nuamah, K. 2015). UV irradiation (220–400 nm) also degrades
the aflatoxins particularly AFB1, AFB2, and AFM1 in various crops with a
degradation potential ranging from 77 to 99.12% (Diao et al. 2015).

3.1.2 CHEMICAL METHODS

Several chemicals such as acids, alkalis, oxidizing agents, aldehydes, and several
gasses are also proved to mitigate the aflatoxigenic fungal growth and aflatoxin
production when used in appropriate quantity (Udomkun et al. 2017). Among
gasses, ozone was found to be most effective in maximizing the aflatoxin
degradation on legumes and cereals by an electrophilic attack on carbon bonds of
furan ring (Jalili 2016). However, due to the high-cost factor, ozone treatment is of
less use in the post-harvest treatment of crops. Aflatoxin production can also be
inhibited by the ammoniation process in corn and other food commodities
(Karlovsky et al. 2016). The positive aspect of the ammoniation process is that
high-pressure ammoniation (0.25, 0.5, 1.5, and 2%) minimizes the time required to
reduce the aflatoxin production in several crops and food commodities (Temba et
al. 2016). Certain chemicals such as sodium bisulfite, calcium hydroxide,
formaldehyde, sodium hypochlorite, sodium borate, and sorbents also reduce the
aflatoxin in many food commodities at a significant level (Carvajal &
Castillo 2009). The groundnut cake and poultry feed were detoxified from
aflatoxin by treating them with sodium bisulfite (0.5%) and sodium hydroxide
(1%) (Bedi & Agarwal 2014). Some food additives are also used for the inhibition
of fungal growth and aflatoxin production in combination with some physical
factors like temperature and moisture. Treatment of citric acid in conjunction with
high temperature and pressure leads to the inhibition of fungal growth as well as
aflatoxin production in sorghum (Méndez-Albores et al. 2009). Some scientists
also observed that some food preservatives such as propionic acid, crystal violet, p-
amino benzoic acid, benzoic acid, boric acid, and sodium acetate were also
inhibited the A. flavus growth and aflatoxin production (Aiko & Mehta 2015). In
peanut and maize, AFB1 production was also inhibited by the treatment of sodium
chloride (10%), acetic acid (5%), and propionic acid (5%) (Brožková et
al. 2015). Even under the favorable conditions for the growth of A. flavus, several
salts of propionic acid-like calcium, sodium propionates were able to reduce the
aflatoxin formation in maize (Hassan et al. 2015). In groundnut cake, aflatoxin
production by A. flavus was also foreclosed by certain week acids such as citric
acid, propionic acid, acetic acid, and sorbic acid at several concentrations like 0.25,
0.5, 0.75, and 1% (Verma et al. 2000). Azole fungicides are also used as a tool to
control fungus growth and aflatoxin production with prochloraz being more
effective than tebuconazole (Mateo et al. 2017).

3.1.3 BIOLOGICAL METHODS

The aflatoxin contamination in agricultural products and many other food
commodities are also minimized by the use of various microorganisms. Biological
agents such as bacteria, yeasts, molds, and algae exhibit the different potential to
degrade the aflatoxin in the emulative environment. Detoxification of aflatoxins by
using biological agents is divided into two procedures named absorption and
enzymatic decadence (Jard et al. 2011). Aflatoxins can be absorbed by
microorganisms directly either by concatenating to their cell wall contents through
effectual internalization or congregation (Motawe et al. 2014). Aflatoxins can also
be absorbed by dead microorganisms; this ability can be helpful for the fabrication
of bio-filters in the form of probiotics and found application in fluid
decontamination (Mwakinyali et al. 2019). Degradation of aflatoxins can also be
done by intra or extracellular enzymes; the end products of such enzymatic
degradation are mostly water and CO2 (Aliabadi et al. 2013). In a study on testing
the efficacy of different microbes strains, only Flavobacterium aurantiacum B-184
was found to be effective enough for aflatoxin degradation (Darsanaki &
Miri 2013). However, different bacteria strains were also found to be effective
against degrading the AFB1. The cell-free supernatant of strain Bacillus
velezensis DY3108 exhibits a strong AFB1 degradation activity of 91.5% (Shu et
al. 2018). The thermophilic bacterial strains (Geobacillus and Tepidimicrobium)
also play a major role in AFB1 degradation when used as a microbial consortium
(Wang, Zhao, et al. 2017). Moreover, a significant reduction in A. flavus growth
was observed in pre-harvest crops by inoculation of antagonistic strains
of Pseudomonas, Trichoderma, Ralstonia, Lactobacilli, Burkholderia, and Bacillus
spp. (Akocak et al. 2015; Yang et al. 2017). Surprisingly aflatoxin contamination
was also found to be inhibited by using the non-toxigenic strains of A. flavus and
other molds as a chief controlling agent (Udomkun et al. 2017).

3.2.0 REDUCTION OF AFLATOXIN IN LIVESTOCK FEED

Milk contaminated with aflatoxins is produced mostly from use of infected feed.
Therefore, reducing aflatoxin contamination indirectly via control of livestock feed
hygiene is possible. To achieve the aim, principles and health considerations
during farming and crop production in farms and livestock feed factories, storage
of livestock feed in traditional and industrial warehouses is necessary (Desjardins
AE, et al. 2003)Livestock feed is mostly include corn, cotton seed and canola,
dietary supplements, wheat bran, dried bread, fat powder, and alfalfa. Given that a
significant percentage of the feed composition is derived from crops, health
considerations during planting insects and, harvesting and storage of crops are
factors affecting grain quality. Drought, rainfall, infection by insects, and high
moisture during flowering can be referred to as major causes of aflatoxin
occurrence in farms.
Adherence to appropriate irrigation programs, planting varieties resistant to
moisture and molds weed control, insecticides application, harvesting at the
appropriate time, crop rotation in order to reduce the risk of pathogens, transfer
from the current farming year to the next year, and fertilizing soil are useful ways
of preventing pre-harvest aflatoxin infection. In addition, appropriate storage of
crops which includes placing crops on clean and dry surfaces, protecting crops
from moisture, heat, and insects, and use of fungicides are effective ways of
reducing the infection (Wu F, and Bhatnagar D. 2008) another important way of
controlling and reducing aflatoxin infection is the adherence of hygienic condition
in factories and livestock feed warehouses both in traditional and industrial levels
are.
Controlling mold growth and aflatoxin formation in traditional farms and
warehouses is highly important. In this regard, several studies have been carried
out on the quality of livestock feed and the amount of aflatoxin in feed produced
milk. For example, it has been shown that the amount of aflatoxin in milk
produced in (Creppy EE, 2002) autumn and winter is higher compared to spring
and summer. This is because in cold seasons feed, (Panariti E.2001). Feeding
livestock on fresh forages is not possible due to unfavorable weather conditions
and farmers have to use stored forages. Regarding that warehouse improper
temperature and moisture conditions favor mold growth; therefore, it is necessary
to improve storage conditions of livestock feed (Panariti E.2001) Findings in some
countries have shown that meeting safety conditions of livestock feed has led to
decrease aflatoxin infection (TA Adegbola, et al. 2006).

3.3.0 DIRECT METHODS OF AFLATOXIN REDUCTION IN

MILK
According to Bovo et al. (2012) toxin absorbents, chemical and biological methods
are used directly for reducing aflatoxin in milk). The use of toxin absorbents is one
of the main methods to reduce aflatoxin amounts in milk. Absorbent soils such as
bentonite, vermiculite, hydrated sodium calcium aluminosilicate (HSCAS), and
active carbon are known as absorbent compounds for absorbing various toxins in
aqueous environments. For instance, bentonite has been known as an effective
reducer of aflatoxin M1 in infected milk. The binding capacity and stability of the
compound formed between absorbent and toxin are highly variable and influenced
by temperature and pH. Information about the effect of absorbents on milk
constituents is scarce; however, it has been shown that these substances have a
slight effect on the nutritional quality of milk.37 in a study, the effect of bentonite
on milk protein content was not a considerable and maximum reduction in protein
content was only 5%. It should be mentioned that with respect to the acceptability
of absorbents as healthy additives by international authorities and the ability to
separate them from milk after absorption of aflatoxin, further investigations are in
progress.

3.3.1 PREVENTION OF AFLATOXIN IN DAIRY CATTLE

Aflatoxicosis can only be prevented by feeding rations free of aflatoxin. Preventing
aflatoxin contamination requires an ongoing and thorough sampling and testing
program.
1) Purchase feed from reputable persons and companies experienced in
aflatoxin prevention and who have a proven record of properly
monitoring their feed products. A reliable feed company will carry
insurance to cover misfortunes with aflatoxins or other problems.
2) Don't buy poor quality feed or feed ingredients. A good deal on feed
prices can be the most expensive buy a dairy farmer ever makes if it
proves to contain aflatoxin.
3) Store feed at proper moisture levels.
4) Develop a systematic inspection and clean-up program to keep bins,
delivery trucks and other equipment free of adhering or caked feed
ingredients.
5) Minimize dust accumulation in milling and mixing areas. Keep all feed
equipment free of caked feed.
6) Check feed storage bins for leaks.
7) Implement effective rodent and insect control programs in grain storage
areas.
8) Grains contaminated with aflatoxins have been successfully treated with
ammonia but it is expensive and dangerous to do.
CONCLUSION
Aflatoxin is a type of mycotoxin produced by Aspergillus mold. Aflatoxin is the
most well-known and researched mycotoxin. Ethiopia is most favorable for
aflatoxigenic fungi and aflatoxin contamination, especially AFB1. Reports show
that maize and groundnut are the most contaminated commodities in the country.
These two commodities are most important in day to day dietary sources of the
people in most regions of Ethiopia. The level of contamination for most
commodities in the country is also very much greater than the international
standard. Similarly milk and milk products are also contaminated with AFM1
above the standard level.
There are basically six groups of aflatoxins, AFB1, AFB2, AFG1, AFG2, AFM1
and AFM2; from which AFB1 are the most potent aflatoxins to cause health
damage to human and animals. AFB1 is the most common contaminant of most
Ethiopian commodities. Aflatoxins are toxic to human and animal and cause
different diseases. There are two main ways people are usually exposed to
aflatoxin. The first is when someone takes in a high amount of aflatoxins in a very
short time. This can cause liver damage, liver cancer, mental impairment,
abdominal pain, vomiting, death and others. The other way people suffer aflatoxin
poisoning is by taking in small amounts of aflatoxins at a time but over a long
period. This may happen if a person’s diet contains a small amount of aflatoxin, as
a result, it may cause growth and development impairment, liver cancer, DNA and
RNA mutation, and others
The effect of aflatoxin on humans depends on age, gender, level of exposure,
duration of exposure, health condition, and strength of their immune system, diet,
and environmental factors. In Ethiopia and many other developing countries,
aflatoxin causes huge health effects and economic loss. Commodities can be
contaminated with aflatoxicogenic fungi and aflatoxin at any time, before harvest
and after harvest. The prevention of aflatoxin once occurs and treatment of
aflatoxicosis is difficult. However, there are some mitigation mechanisms pre- and
post-harvest, especially proper storage is essential with proper moisture and
temperature. Moreover, awareness creation on aflatoxin contamination, its effect,
and management is essential.

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