After India and the USA, Pakistan is the third country leading in global dairy production, a sector of very high socioeconomic relevance in Asia. Mycotoxins can affect animal health, reproduction and productivity. This study analysed a broad range of co-occurring mycotoxins and fungal secondary metabolites derived from Alternaria, Aspergillus, Fusarium, Penicillium and other fungal species. To complete this, a validated multi-metabolite liquid chromatography/electrospray ionization-tandem mass spectrometric (LC/ESI–MS/MS) method was employed, detecting 96 of > 500 tested secondary fungal metabolites. This first preliminary study demonstrated that total mixed rations (TMRs) (n = 30) from big commercial dairy cattle farms (> 200 lactating cows) in Punjab, Pakistan, presented ubiquitous contamination with mixtures of mycotoxins. The mean of mycotoxins per sample was 14, ranging from 11 to 20 mycotoxins among all TMR samples. Metabolites derived from other fungi and Fusarium spp. showed the highest levels, frequency and diversity among the detected fungal compounds. Among the most prevalent mycotoxins were Fusarium toxins like fumonisins B1 (FB1) (93%), B2 (FB2) (100%) and B3 (FB3) (77%) and others. Aflatoxin B1 (AFB1) was evidenced in 40% of the samples, and 7% exceeded the EU maximum limit for feeding dairy cattle (5 µg/kg at 88% dry matter). No other mycotoxin exceeds the EU guidance values (GVs). Additionally, we found that dietary ingredients like corn grain, soybean meal and canola meal were related to increased contamination of some mycotoxins (like FB1, FB2 and FB3) in TMR from the province of Punjab, Pakistan. Among typical forage sources, the content of maize silage was ubiquitous. Individually, the detected mycotoxins represented relatively low levels. However, under a realistic scenario, long-term exposure to multiple mycotoxins and other fungal secondary metabolites can exert unpredictable effects on animal health, reproduction and productivity. Except for ergot alkaloids (73%), all the groups of metabolites (i.e. derived from Alternaria spp., Aspergillus spp., Fusarium spp., Penicillium spp. and other fungi) occurred in 100% of the TMR samples. At individual levels, no other mycotoxins than AFB1 represented a considerable risk; however, the high levels of co-occurrence with several mycotoxins/metabolites suggest that long-term exposure should be considered because of their potential toxicological interactions (additive or synergistic effects).

继印度和美国之后，巴基斯坦是全球乳制品生产领先的第三个国家，乳制品生产在亚洲具有非常高的社会经济相关性。霉菌毒素会影响动物健康、繁殖和生产力。这项研究分析了来自链格孢属、曲霉属、镰刀菌属、青霉属和其他真菌物种的多种共存霉菌毒素和真菌次生代谢物。为了完成这一任务，采用了经过验证的多代谢物液相色谱/电喷雾电离串联质谱 (LC/ESI-MS/MS) 方法，检测了超过 500 种测试的次生真菌代谢物中的 96 种。第一项初步研究表明， 巴基斯坦旁遮普省大型商业奶牛场（> 200 头泌乳牛）的总混合日粮 (TMR)（ n = 30）普遍存在霉菌毒素混合物污染。每个样品的霉菌毒素平均值为 14，所有 TMR 样品中霉菌毒素的含量为 11 至 20 种。来自其他真菌和镰刀菌属的代谢物。在检测到的真菌化合物中显示出最高的水平、频率和多样性。最常见的霉菌毒素包括镰刀菌毒素，如伏马菌素 B1 (FB1) (93%)、B2 (FB2) (100%) 和 B3 (FB3) (77%) 等。40% 的样品中发现了黄曲霉毒素 B1 (AFB1)，其中 7% 超过了欧盟饲养奶牛的最高限量（88% 干物质时为 5 µg/kg）。没有其他霉菌毒素超过欧盟指导值 (GV)。此外，我们发现玉米粒、豆粕和菜籽粕等膳食成分与巴基斯坦旁遮普省 TMR 中某些霉菌毒素（如 FB1、FB2 和 FB3）污染增加有关。在典型的饲料来源中，玉米青贮饲料的含量无处不在。单独而言，检测到的霉菌毒素水平相对较低。然而，在现实情况下，长期接触多种霉菌毒素和其他真菌次生代谢物可能会对动物健康、繁殖和生产力产生不可预测的影响。除麦角生物碱（73%）外，所有代谢物组（即源自链格孢属、曲霉属、镰刀菌属、青霉属和其他真菌）在 100% 的 TMR 样品中出现。在个体水平上，除了 AFB1 之外，没有其他霉菌毒素具有相当大的风险；然而，多种霉菌毒素/代谢物的高水平共存表明应考虑长期接触，因为它们存在潜在的毒理学相互作用（相加或协同效应）。