Study of toxigenic Aspergillus in feed and impact of bovine breed on aflatoxins carryover in milk and urine in dairy cows: a case of Bulawayo, Zimbabwe

Abstract

This thesis explored the biodiversity and phylogenetic relationships of aflatoxigenic Aspergillus isolated from different feeds used for feeding dairy cows under intensive, semi-intensive and extensive feeding systems in Zimbabwe during the dry and rainy seasons. It also looked at the relationship between AFB₁ dietary intake and AFM₁ output in milk on cows fed with naturally contaminated feed on a daily basis. The work also involved the determination of effect of breed on aflatoxin carryover from feed into milk and urine. In Chapter 3, both morphological and molecular methods were used for the identification of aflatoxigenic speciess isolated from the different feed types. The bulk of of the feeds were contaminated by moulds from the Aspergillus genus with the potential of producing aflatoxins. The most contaminated feed was the mixed rations and most of the toxigenic strains were isolated during the rainy season. Phylogenetic trees constructed based on single gene were not able to distinguish the isolates to species level or group them into their respective sections. However gene concatenation was able to cluster the isolates into the individual clades as well as grouping them based on the time of isolation giving a true reflection of the evolution aspect of the isolates with time. In Chapter 4, quantification of aflatoxins in the feeds was done by high performance liquid chromatography which indicated the presence of all the four naturally occurring aflatoxins namely aflatoxin B₁, B₂, G₁ and G₂ in the feeds. The mixed rations had the highest average total aflatoxin concentration of 29.0μg/kg. Aflatoxin B₁, the most potent aflatoxin was present in all feeds with an average concentration of 9.0μg/kg which was above the European Union (EU) standard of 5.0μg/kg for lactating cows which the country adopted and grass had the lowest aflatoxin concentrations of 2.5μg/kg. Chapter 5 showed that AFM₁ was present in 70.6% of the milk samples with the bulk of of the samples coming from the dry season. Milk samples from the rainy season had a higher percentage (88%) of compliance to the EU limit of 0.05μg/L compared to the dry season which had 58% of the milk samples below the EU limit. Aflatoxin M₁ concentrations in urine were between 0 and 2.36μg/L with samples from the dry season having higher concentrations than those from the rainy season. Although most of the milk samples complied with the EU and FDA limits, the consumers are still at the risk of having chronic aflatoxicosis. In Chapter 6, regression analysis showed that the concentration of AFM₁ in milk on any given day is governed by the AFB₁ concentration in the cow's diet on that particular day and that AFM₁ carryover from the previous day usually remains in the system from the previous day(s). This is because 3-4 days are required for the AFM₁ to get cleared from the system; however, this does not happen in a real life situation as the cows feed on naturally contaminated feed daily. Therefore there is a cumulative effect of the milk aflatoxin on a daily basis depending on the amount of AFB₁ in the feed. Multivariate analysis of variance and correlation analysis showed that the effect of diet on aflatoxin carryover into milk diet is significant but breed does not have any significant effect of AFM₁ in milk. Repeated measures one way ANOVA also showed that breed had no effect on the amount of AFM₁ secreted into milk. Findings from this study showed that feeds used by the farmers are contaminated by aflatoxigenic Aspergillus. Daily consumption of the contaminated feeds by the dairy cows results in the release of the aflatoxins in milk on a daily basis posing a threat to the population. The association of breed on both AFM₁ secretion into milk and excretion into urine was insignificant whereas diet effect was significant at 0.05% significance level.