ASPERGILLUS SPECIES AND ASSOCIATED AFLATOXINS IN DAIRY FEED AND RAW MILK FROM SPECIALIZED DAIRY FARMS AND JOCAL MARKETS IN SELECTED URBAN CENTERS OF EASTERN ETHIOPIA

Abstract

The contamination of dairy cattle feed with Aspergillus fungi and their aflatoxins poses significant global food safety challenges, impacting the dairy industry and public health. Despite this, data on Aspergillus species and aflatoxins in dairy feeds and milk in eastern Ethiopia remain limited. This study aimed to determine the prevalence and levels of Aspergillus species and aflatoxins in feed and milk samples collected from specialized dairy farms and local markets in Chiro town, Dire Dawa, and Harar cities. Additionally, the study evaluated the awareness of aflatoxin contamination among dairy farmers and feed retailers, as well as explored the associations between the demographic variables of the respondents and aflatoxin levels in feeds. A total of 360 samples (180 each of feed and milk) were randomly collected from the specialized dairy farms and local markets. The specialized dairy farms were selected based on lactating cows and feed types (i.e., wheat bran (WB), maize feed (MF) and concentrate mix (CM)), which are susceptible to fungal and their toxin contamination. Similarly, feed retailers/shops selling feed (WB, MF, CM) and milk vendors selling raw milk of dairy cows in local markets were identified for sample collection. Interviews with 180 respondents from dairy farmers (n=90) and feed retailers/shops (n=90) were conducted to assess awareness of aflatoxin contamination. The Aspergillus species in feed samples were cultured using Yeast Extract Sucrose Agar (YES) and Aspergillus Flavus and Parasiticus Agar (AFPA). The identification of Aspergillus species was based on macroscopic and microscopic morphological characteristics. Their aflatoxigenic potential was evaluated using Coconut Agar Media (CAM) under UV fluorescence. Aflatoxins (AFB1, AFB2, AFG1, and AFG2) in feed and aflatoxin M1 (AFM1) in milk samples were analyzed using high-performance liquid chromatography with fluorescence detector (HPLC/FLD), following thorough extraction and immunoaffinity column sample clean-up. The study results revealed that the prevalence of Aspergillus species in feed samples was 80.6% (mean count: 3.04 log10CFU/g), with A. flavus being the most common species (80.0%, xviii mean count: 2.73 log10CFU/g). Approximately 37.2% of feed samples exceeded Good Manufacturing Practices (GMP) limits for fungal counts (1×104 CFU/g). Mean fungal counts were significantly higher in feeds from Dire Dawa city compared to other locations (P < 0.05). Concentrate mix (CM) showed the highest fungal contamination among feed types, with mean counts for total Aspergillus species, A. flavus, and A. parasiticus of 3.65 log10CFU/g, 3.26 log10CFU/g, and 2.98 log10CFU/g, respectively. A significant proportion of aflatoxigenic isolates (81.4%), corresponding to A. flavus and A. parasiticus were detected. Total aflatoxin (TAF) was detected in 82.8% of feed samples, with mean levels of AFB1, AFB2, AFG1, and AFG2 being 28.15 μg/kg, 3.30 μg/kg, 19.87 μg/kg, and 2.70 μg/kg, respectively. However, the levels and contamination of aflatoxin varied across sampled sites and feed types (P<0.05). Thus, in Chiro town, the significantly (P<0.001) lowest mean of AFB1 was observed in WB compared to MF and CM. In Dire Dawa and Harar, the highest significant (P<0.001) mean of AFB1 was detected in CM compared to other feed types. Moreover, in Dire Dawa and Harar, CM had a significantly (P<0.001) higher mean of AFG1 compared to other feed types. Whereas the mean levels of TAF, AFB2, and AFG2 were found to be significantly highest in feeds from Dire Dawa city as well as CM. Overall, 33.3% and 38.9% of feed samples exceeded regulatory limits for AFB1 (20 μg/kg) and TAF (40 μg/kg), respectively. In milk samples, AFM1 contamination was found to be 63.9%, with a mean level of 0.18 μg/L. Of these, 39.1% and 26.1% exceeded EU and FDA regulatory limits, respectively. Dire Dawa recorded the highest mean of AFM1 (0.34 μg/L), and milk from specialized dairy farms was more frequently contaminated compared to milk vendors. A strong positive correlation (r = 0.67) and significant linear regression association (R2 = 0.935, P < 0.001) were found between AFB1 levels in feed and AFM1 levels in milk. Awareness of aflatoxin risks among dairy farmers and feed retailers was low. Key factors contributing to feed contamination in dairy feed and milk, with significant public health and food safety implications were observed. The findings emphasize the need for enhanced education for dairy farmers and feed retailers, rigorous monitoring, and mitigation strategies to address aflatoxin contamination in animal feeds and milk. Further research on detoxification technologies, such as the use of bentonite clay and others, is recommended.