Bacillus spp. Contamination: A Novel Risk Originated From Animal Feed to Human Food Chains in South-Eastern Bangladesh

Abstract

The current study provides information on <i>Bacillus</i> spp. contamination along with present status in commercially available poultry and animal feeds as well as animal-derived products in Bangladesh. The research has been conducted to determine if animal feed and its components are a source of <i>Bacillus</i> spp. contamination in feed and food chain. Out of 180 different feeds, milk, egg, and human stool samples, 218 <i>Bacillus</i> spp. were isolated and identified by cultural morphology, microscopic, biochemical, and molecular characteristics where <i>B. cereus</i>, <i>B. subtilis</i>, <i>B. amyloliquefaciens</i>, <i>B. licheniformis</i>, <i>B. thuringiensis</i>, <i>B. megaterium</i>, and <i>B. coagulans</i> accounted for 51, 22, 9.1, 5.9, 5, 3.6, and 2.2%, respectively. Regarding the enumeration of total viable count and total <i>Bacillus</i> count, correspondingly 67 and 39% samples were found to be contaminated with above 10,000 CFU/g, while highest contamination was 85 and 75% in broiler feed, respectively. The total number of bacteria above the regulatory limits in commercially available feeds indicates a poor compliance with regulation and abuse administration in the Bangladeshi market. Moreover, a hospital-based survey showed that food-borne <i>Bacillus</i> spp. contributed to 4.5% human diarrhea cases and 25% food contamination associated with vegetables, rice, RTE food, milk, and egg, accounting for 46, 34, 14, 4, and 2%, respectively. <i>B. cereus</i> was the dominant isolate correspondingly accounting for 56 and 51% egg and milk contamination followed by <i>B. amyloliquefaciens</i> (32%) and <i>B. thuringiensis</i> (12%) in egg and <i>B. subtilis</i> (25%), <i>B. amyloliquefaciens</i> (12%), <i>B. thuringiensis</i> (6.4%), and <i>B. coagulans</i> (3.2%) in milk, respectively. Toxin gene profiling of <i>Bacillus</i> spp. revealed that <i>B. cereus</i> constituted a principal part of virulence, while <i>B. thuringiensis</i>, <i>B. licheniformis</i>, <i>B. megaterium</i>, <i>B. coagulans</i>, and <i>B. subtili</i>s showed genetic diversity and <i>B. amyloliquefaciens</i> had not carried any toxin gene. Detection rate of enterotoxin genes (<i>nheA</i>, <i>nheB</i>, <i>nheC</i>, <i>cytK</i>, <i>hblA</i>, <i>hblC</i>, <i>hblD</i>, and <i>entFM</i>) showed that 55% isolates carried <i>nheABC</i> genes, 80% <i>entFM</i>, and 71% <i>cytK</i>, whereas only 33% of the isolates contained <i>hblACD</i> gene clusters. These virulence genes were posing a threat to human health due to spread across the food and feed chain. Finally, our findings support the hypothesis that <i>B. cereus</i> might contribute to clinical diarrhea, gizzard erosion, and lung infection in duck and poultry, and that it contaminates animal-derived foods resulting in toxicity and antibacterial resistance to humans. Therefore, maximal tolerance limits of <i>Bacillus</i> spp. and their potential risks to the animal industry are urgently needed to clarify. Moreover, <i>Bacillus</i> spp.-induced toxin residual must be altered for human health via food chain transmission.