Study of intestinal microbial flora of local chickens to investigate the effect of probiotics Bacillus subtilis and Bacillus coagulans on the expression of ctxM and luxS pathogenic genes in isolates of Escherichia coli

Abstract

Background and objective: Antibiotics are widely used worldwide. However, due to the emergence of antibiotic resistance in a wide range of microorganisms, their use worldwide has failed. Probiotics are suggested as complementary and alternative medicine. The present study aimed to investigate the effects of probiotics isolated from local chickens on the expression of luxS and ctxM genes in resistant Escherichia coli. Materials and methods: 300 fecal samples were taken from patients referring to Imam Khomeini Hospital in Tehran during May–September 2016 and Escherichia coli samples were isolated using specific culture media and biochemical tests and then the presence of luxS and ctxM genes were identified using PCR with specific primers. In order to extract the probiotics forming spores, the intestinal contents of 10 poultry that had not used any antibiotics and probiotics were cultured, isolated, and identified using biochemical and PCR methods. Commercial strains of Bacillus subtilis and Bacillus coagulans were purchased to compare their effects with native bacteria. These strains were then co-cultured with resistant Escherichia coli strains containing ctxM and luxS genes. Real-time PCR was used to evaluate the effect of these probiotics on gene expression Results: The results indicated that 40 isolates (7.5%) of Escherichia coli were obtained from the 300 fecal samples. Thirteen samples (32.5%) were outpatients and 27 (67.5%) were inpatients. All isolates were isolated from men and women aged 21–62. Four Escherichia coli strains were isolated from patients carrying ctxM and luxS genes. Isolation of Bacillus coagulans and Bacillus subtilis from samples was confirmed by biochemical and molecular experiments. The commercial and native strains of Bacillus coagulans reduced the expression of the luxS and ctxM genes by 3.60, 3.30, 1.58, and 2.70 times respectively. Also, the commercial and native strains of Bacillus subtilis decreased the expression of the luxS and ctxM genes by 1.37, 1.10, 2.20, and 2.80 times respectively. The results of statistical analysis showed a significant relationship between the presence of native and commercial probiotics in culture and reduced expression of ctxM and luxS genes. Conclusion: According to the results, supplements of Bacillus coagulans and Bacillus subtilis increase the effect of antibiotics resistance in Escherichia coli by reducing the expression of resistance genes.