Aflatoxin Contamination of Milk Produced in Peri-urban Farms of Pakistan: Prevalence and Contributory Factors.

Aflatoxin M1 contamination of milk in Pakistan, like many developing countries, is poorly understood. The present study was therefore conducted to determine AFM1 contamination of milk and its contributory factors in Pakistan. We sampled milk and feedstuffs from 450 peri-urban dairy farms in seven major cities following a cross-sectional study design. Analysis of milk using ELISA revealed high contamination with an overall average of 3164.5 ng of AFM1/L, and significant differences (p < 0.001) between cities. The milk sampled from Gilgit, in northern hilly areas, had an average AFM1 level of 92.5 ng/L. Milk from other cities had 3529.7 ng/L average contamination, with only 5.7% samples qualifying the maximum tolerable limit of 500 ng of AFM1/L. Heavy mean aflatoxin contamination was found in bakery waste (724.6 µg/kg), and cottonseed cake (600.8 µg/kg). Rest of the other feedstuffs had moderate to low mean aflatoxin contamination, ranging from 66.0 µg/kg in maize stover to 3.4 µg/kg in wheat bran. The mean aflatoxin level in commercial dairy concentrates was 32.7 µg/kg. About 80% of the total aflatoxin intake of dairy animals was contributed by cottonseed cake alone due to its high aflatoxin contamination and proportion in dairy rations. On-farm storage time of oilseed cakes varied (p < 0.01) in different cities but was not associated with aflatoxin contamination. The exceptionally high AFM1 contamination suggests that milk from peri-urban dairy farms is a serious public health threat in Pakistan. This situation can be mitigated by reducing aflatoxin contamination in cottonseed cake and promoting the use of commercial concentrates and other feedstuffs with low contamination.

Aflatoxin Contamination of Milk Marketed in Pakistan: A Longitudinal Study.

A longitudinal one-year study was conducted to determine aflatoxin M1 levels in different types of milk marketed in Pakistan. Processed and raw liquid milk from 21 sources, two milk powder and six tea whitener brands were sampled on monthly basis from Islamabad. The aflatoxin M1 levels in liquid milk were lower (p < 0.05) in summer (April to July) compared with the levels in winter (January, November and December). The mean aflatoxin M1 levels were 254.9, 939.5, and 1535.0 ng/L in UHT, pasteurized, and raw milk, respectively (differing at p < 0.001). The mean toxin level in powdered milk after reconstitution was 522.1 ng/L. Overall, 12.9, 41.0, 91.9 and 50.0% of the UHT, pasteurized, raw and powdered milk samples, respectively, exceeded the Codex maximum tolerable limit of 500 ng of aflatoxin M1/L. It was estimated that consumers of raw and processed milk were exposed to 11.9 and 4.5 ng aflatoxin M1, respectively, per kg of body weight daily. The study indicates potential aflatoxin M1 exposure risks for the consumers of raw milk in the country. The levels of the toxin though comparatively lower in milk powder, requires attention as this type of milk is consumed by infants.

Comparison of Some ELISA Kits for Aflatoxin M1 Quantification.

Background: ELISA is a widely used method for aflatoxin M1 (AFM1) quantification in dairy products. This study was conducted to compare ELISA kits from different manufacturers for AFM1 quantification in milk. Methods: High sensitivity ELISA kits (for up to 250 ng AFM1/L), including AgraQuant (Romer Labs), Bioshield M1ES (Prognosis Biotech), Helica 96 (Helica Biosystems), Veratox (Neogen, Inc.), and the medium sensitivity kit Immunolab AM1E01 (Immunolab GmbH; for 10-1000 ng AFM1/L) were tested against a certified reference AFM1 whole milk having 44 ng AFM1/L and its 10-fold dilution, and a 50 ng AFM1/L standard. In another experiment, Prognosis Bioshield M1UF (70-1000 ng AFM1/L), Romer's AgraquantPlus (10-2000 ng AFM1/L), and Immunulab's AM1E01 kits were compared to test a 500 ng AFM1/L solution. Results: In both of the experiments, the quantification of the tested AFM1 levels did not differ (P ≥ 0.310) between the kits from various manufacturers. In the case of 4.4 ng AFM1/L, recovery of the toxin was close to the assigned value under the kit by Neogen, Inc. In case the of 44 ng level, better recoveries were seen under the kits by Immunolab and Prognosis Biotech. In both the cases, low relative standard deviation (RSD) values were obtained only for the kit by Prognosis Biotech. In the case of the 50 and 500 ng AFM1/L solutions, the kits by Prognosis Biotech and Romer Labs provided recoveries close to the assigned values as well as low RSD. Conclusions: These data indicate that all the studied ELISA kit brands had comparable efficacy for AFM1 quantification. Recovery of the toxin and RSD may, however, differ between kits from various manufacturers.