Livestock Research for Rural Development 31 (1) 2019 Guide for preparation of papers LRRD Newsletter

Citation of this paper

Prevalence of aflatoxins contamination in commercial broiler feeds in Kenya

F N Thuita, J K Tuitoek, A M King’ori and M A Obonyo1

Department of Animal Sciences, Egerton University, P O Box 536-20115, Egerton, Kenya
jtuitoek@egerton.ac.ke
1 Department of Biochemistry and Molecular Biology, Egerton University, P O Box 536 -20115 Egerton, Kenya.

Abstract

Aflatoxin contamination of broiler feed is a major barrier to sustained agricultural productivity and trade. Aflatoxins are a type of mycotoxins (secondary fungal metabolites) produced by fungi of the genus Aspergillus (A), mainly A. flavus and A. parasiticus, in cereals and grains when conditions are favorable. The aim of this study was to determine the levels of total aflatoxins (AFs) in common commercial broiler feeds among feed manufacturers in Nakuru town, Kenya. Forty compounded broiler feed samples were randomly collected from ten feed mill companies in Nakuru town (ten broiler starter and ten broiler finisher feed samples per company) in two phases. Each collection phase was determined by the frequency of purchase of raw materials by the individual milling companies. The total aflatoxin levels in the feed were analyzed using the ELISA technique in the Mycotoxin Research Laboratory in Egerton University. The data was subjected to SAS procedures using two way analysis of variance. All the feeds collected contained aflatoxins within a range of 1.07- 41.01 g/kg. The samples (92.5%) contained total aflatoxin levels which exceeded the WHO limits of 5 g/kg in animal feeds. Of the samples collected, 52.5% exceeded the FDA limits of 20 g/kg in poultry feeds. To avoid high levels of AFs in broiler feeds, feed manufacturers should test for aflatoxins in the raw materials and avoid the fungal contamination in the broiler feeds at all stages of handling.

Key Words: ELISA, mycotoxins


Introduction

Livestock production contributes approximately 10 and 30% of the national and agricultural GDP respectively (FAO 2013). Poultry population in Kenya is estimated at 32.5 million (FAO 2013), with 8 million being commercial poultry population (where, 60% of these is comprised of broiler chicken) (Atherstone et al 2016). About 500,000 tons of animal feed is produced in Kenya annually and roughly 70% is poultry feed (Atherstone et al 2016). Animal feed manufacturing industries compete for feed materials with humans, thus, low quality raw materials are often used for poultry feed production (Abidin et al 2011). In Kenya, food ingredients rejected for human consumption (off-color and feed grade cereals and grains) are normally utilized for feed production. These off-color cereals and grains are often contaminated with mycotoxins and there are risks that contamination may exceed acceptable standards. Poultry have been reported to be very sensitive to the effects of mycotoxins (Denli and Okan 2006). Mycotoxins are secondary fungal metabolites with very different chemical configurations (Bryden 2012). They exist in many forms but Aflatoxins (AFs) and Ochratoxins (Abidin et al 2011) which are rated as Class 1 human carcinogens by the International Agency for Research on Cancer (IARC) (Binder 2007) are considered the most important. Aflatoxins are produced by members of the genus Aspergillus (A), mainly A. flavus and A. parasiticus, in cereals and grains when moisture and ambient temperature are favorable (Bryden 2012; Marchioro et al 2013; Spragg and Watts 2013). The poultry sector therefore is at risk of economic losses due to the negative impact of AFs on their performance (Marchioro et al 2013; Atherstone et al 2016). Aflatoxins may cause oxidative stress, reductions in growth parameters, increased susceptibility to disease and adversely affect sexual maturity and hatchability in chicken (Denli and Okan 2006; Karaman et al 2010). In addition to the negative effects of AFs in poultry feed on the performance of broilers, contamination of feeds with aflatoxins may result in the presence of AFs residues in foods of animal origin such as broiler meat posing a serious threat to public health (Herzallah 2009). AFs can cause mutagenic effects, are carcinogenic, teratogenic and hepatotoxic and they also depress immunity in both human beings and animals (Leslie et al 2008 ; Herzallah 2009 ; Abrar et al 2013). Standards have been set by different countries and regulatory bodies to govern AF levels in human and animal foods/feeds. The limits of aflatoxin B1 and total AFs in foods are 2 and 4 g/kg respectively in the European Union whilst they are 5 and 10 g/kg, respectively, in more than 75 countries around the world (Herzallah 2009). WHO has set AFs limits for animal feeds at 5 g/kg (Kajuna et al 2013) and FDA at 20 g/kg (Reddy and Raghavender 2007). The aim of this study was to determine the prevalence and levels of AFs in commercial broiler feeds manufactured in Nakuru town, Kenya.


Materials and methods

Experimental sites and sampling procedure

This study was carried out in Nakuru town and Egerton University, Kenya. Broiler feed samples were collected from ten randomly selected millers out of the twenty feed milling companies in the town that manufacture broiler feeds. A total of forty compounded broiler feed samples were collected randomly. Ten broiler starter and ten broiler finisher feed samples were collected per feed miller in two phases, the first phase was April and May, 2017 and the second phase between June and July, 2017. Each collection phase was determined by the frequency of purchase of raw material by the individual milling companies. The samples were collected in different khaki bags, labelled, transported to the Mycotoxin Research Laboratory in Egerton University and stored at 4oC as per (Nemati et al 2014) awaiting analysis. All samples were analysed using the ELISA technique.

Analysis of samples

For each collected sample, one kilogram composite sample was collected by picking random portions of samples from the same feed batch and mixing thoroughly to form a homogeneous sample (Rodrigues et al 2011). The samples were then subjected to ELISA technique to determine the concentration of total aflatoxins.

Extraction of aflatoxins from the samples

Representative feed samples were individually finely ground such that 95% passed through a 20 mesh screen. Extraction and assay for AFs was carried out according to the manufacturer’s procedure (Helica Biosystems Inc). The extraction solution (70% methanol: 30% distilled water) was prepared for each sample to be tested. A 20 g ground portion of feed samples were weighed and added to 100 ml of the extraction solvent. The mixture was mixed by shaking in a sealed container for a minimum of 2 minutes. The particulate matter was allowed to settle, then filtered (Whatman #1) and the filtrate collected for testing. The Absorbance optical density (OD) of each microwell was read with a Thermo Scientific™ microtiter plate reader at 450 nm.

Assay Procedure

The levels of total AFs in each sample was determined using ELISA technique following manufacturer’s instructions (Helica Biosystems Inc).

Statistical data analysis

Graph pad prism 7 software was used to convert the optical density (OD) data to g/kg. The data were then subjected to a two way analysis of variance using the GLM procedures of SAS (version 9.13) and the means were separated using the paired T-Test. The model was;

Yijk = άi + βj + βij + Ɛk

Where; Yijk represents AFs levels the samples,

άi represents the feed type i.e. either broiler starter or finisher

βj represents phase of sample collection i.e. either phase 1 or phase 2

άβij represents the interaction between feed type and phase and,

Ɛi is the error term.

Furthermore, frequencies and percentages were also calculated.


Results

The results of these analyses are summarized in Tables 1, 2 and 3. All the compounded broiler feed samples collected in Nakuru town contained total AFs levels ranging between 1.07 - 41.01 g/kg. The range for broiler starter feed was 1.07- 41.01 g/kg whereas those for broiler finisher were 4.69-35.76 g/kg. Of the samples, 92.5% (90% broiler starter and 95% broiler finisher) exceeded the WHO recommended level of 5g/kg total AFs limits in animal feeds. Whilst 52.5% of the samples (50% broiler starter and 55% broiler finisher) exceeded the FDA 20g/kg total AFs limits in poultry feeds. The mean total AFs levels for the broiler starter and broiler finisher feed samples were 19.37 2.45 and 19.86 2.21 g/kg respectively. On the other hand, the mean total AFs levels for feed samples collected in phase 1 and phase 2 were 18.00 2.03 and 21.22 2.54 g/kg respectively. The aflatoxin levels in the starter and finisher samples were not different (p=0.88). Aflatoxin levels in samples collected in phase 1 were not different from those collected in phase 2 of sample collection (p=0.34). There was no interaction between the feed type and sample collection phase (p=0.57).

Table 1. The level of total AFs (g/kg) in broiler feeds according to feed type in Nakuru town, Kenya

Broiler feed type

n

Minimum

Maximum

Mean

SEM

 p

Starter

20

1.07

41.0

19.4

2.44

0.88

Finisher

20

4.69

35.8

19.9

2.20

p-value >0.05 means that the aflatoxin levels in samples are not different.



Table 2. The level of total AFs (g/kg) in broiler feeds according to collection phase in Nakuru town, Kenya

Collection phase

n

Minimum

Maximum

Mean

SEM

p

Phase 1 (April-May)

20

1.07

29.2

18.0

2.03

0.34

Phase 2 (June-July)

20

3.76

41.0

21.2

2.54

p-value >0.05 means that the aflatoxin levels in samples are not different.



Table 3. The proportion (%) of commercial broiler feeds containing various concentrations of total aflatoxins (n=40)

Level of Aflatoxin
(g/kg)

Type of Feed

% proportion-
all samples

Cumulative
% proportion

Broiler Starter

Broiler Finisher

Undetected

0

0

0

0

< 5

10

5

7.5

7.5

5-10

15

15

15

22.5

10.001-20

25

25

25

47.5

20.001-30

35

45

40

87.5

30.001-40

10

10

10

97.5

40.001-50

5

0

2.5

100


Discussion

The results corroborate previous studies which reported that compounded animal feeds, specifically poultry feeds, had both high prevalence and concentration levels of aflatoxins. In a study conducted by Kajuna et al (2013) in Tanzania, 78.1% of all the compounded feed samples collected were contaminated with aflatoxins with broiler feeds having the highest contamination percentage (91.7%). A study conducted for a three-year period in Kenya reported that all animal feed samples were contaminated with AFs, ninety five percent (95%) of samples exceeding 10 g/kg and while 35% exceeded 100 g/kg and AFs levels ranging from 5.13 -1123 g/kg (Okoth and Kola 2012). In another study, 324 samples of grains, finished animal feeds and other feed commodities were collected from thirteen countries in the Middle East and Africa and tested for various mycotoxins including aflatoxins (Rodrigues et al 2011). Fumonisins were the main contaminant per country in all the samples collected except for samples from Nigeria and Kenya which had AFs as the main contaminant. The prevalence and level were 94 and 78%, 115 and 52 g/kg respectively for Nigeria and Kenya (Rodrigues et al 2011). AFs prevalence in agricultural products such as cereals and grains, and hence animal feeds compounded from these agricultural products, is relatively higher in tropical and subtropical regions due to warm and humid weather conditions which provide optimal conditions for the growth of the moulds (Bryden 2012; Abrar et al 2013). Poultry feed production and costs are a major issue faced by international as well as local industries due to competition for feed materials by animals and humans (Abidin et al 2011). Typically lower quality raw materials are used in poultry feed production (Abidin et al 2011) such as off-color and feed grade cereals and grains. Overall, 92.5% of broiler feeds manufactured in Nakuru town contain AFs levels higher than accepted by WHO standard. These levels may negatively impact on the performance of broilers and may end up in broiler meat and the human food with disastrous consequences to human health.


Conclusion and Recommendations


Acknowledgements

The authors would first like to thank the National Research Fund-Kenya for the Post Graduate Students Funding 2016/2017 grant for funding this work. Second, the Mycotoxin Research Laboratory, Egerton University, funded by the Canadian Government for Aflatoxin Analysis for providing the required equipment and Collins Omondi and Micah Lagat for the technical expertise rendered in this study.


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Received 17 July 2018; Accepted 12 December 2018; Published 1 January 2019

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