Evaluation of Expression Levels of Virulence Associated luxS, and ctxM Genes in Escherichia coli Isolated from Dairy Products Co-cultured with Bacillus coagulans

Elahianfiroz, Zahra; Baserisalehi, Majid; Ghane, Masood

doi:10.22101/JRIFST.2021.258117.1202

Evaluation of Expression Levels of Virulence Associated luxS, and ctxM Genes in Escherichia coli Isolated from Dairy Products Co-cultured with Bacillus coagulans

Document Type : Original Paper

Authors

¹ PhD. Student, Department of Microbiology, College of Science, Shiraz Branch, Islamic Azad University, Shiraz, Iran

² Associate Professor, Department of Microbiology, Kazeron Branch, Islamic Azad University, Kazeron, Iran

³ Associate Professor, Department of Microbiology, Tonekabon Branch, Islamic Azad University, Tonekabon, Iran

10.22101/JRIFST.2021.258117.1202

Abstract

The purpose of this study was the evaluation of expression levels of virulence associated luxS, and ctxM genes in Escherichia coli co-cultured with domestic probiotics. E. coli was isolated from 20 samples of dairy products (raw milk and cheeses). Then probiotic characteristics of Bacillus spp. isolated from intestinal tracts of poultry were assessed for ability to survive at acidic pH, bile salts, simulated gastric juice, production of bioactive compound and proteolytic activity. The isolate exhibited favorable characters, was identified by molecular method using 16SrDNA gene sequencing. Then expression levels of luxS, and ctxM genes in E. coli isolates in co-cultured with the probiotic isolates was evaluated by Real time-PCR method. The results obtained indicated that out of 16 E. coli isolates, two strains carried the genes. In addition, out of 12 Bacillus spp. isolates, one strain showed probiotic characters. The results on molecular identification verified the isolation of two strains of E. coli (carried luxS, and ctxM genes) and one strain of Bacillus coagulans. The results on expression levels of luxS, and ctxM genes in E. coli in co-cultured with Bacillus coagulans isolates indicated that expression levels of the genes diminished significantly (P-value<0.05). Hence, it can be interpreted that native spore-forming probiotic can greatly decrease virulence association gene expression in E. coli. Therefore, based on our finding it can be interpreted that the consumption of native probiotics as food supplement can greatly eliminate gastrointestinal diseases caused by E. coli. Hence, consumption of native probiotics potentially improves gut health. However, it needs more evaluation.

Keywords

References

Barbosa, T. M., Serra, C. R., La Ragione, R. M., Woodward, M. J., & Henriques, A. O. (2005). Screening for bacillus isolates in the broiler gastrointestinal tract. Appl Environ Microbiol, 71(2), 968-978. doi:https://doi.org/10.1128/aem.71.2.968-978.2005

Baserisalehi, M., & Bahador, N. (2013). Evaluation of soil origin Pseudomonas sp. for production of bioactive compounds. Journal of Biological Sciences, 13(3), 152. doi:https://doi.org/10.3923/jbs.2013.152.157

Chelius, M. K., & Triplett, E. W. (2001). The Diversity of Archaea and Bacteria in Association with the Roots of Zea mays L. Microb Ecol, 41(3), 252-263. doi:https://doi.org/10.1007/s002480000087

Croxen, M. A., Law, R. J., Scholz, R., Keeney, K. M., Wlodarska, M., & Finlay, B. B. (2013). Recent advances in understanding enteric pathogenic Escherichia coli. Clin Microbiol Rev, 26(4), 822-880. doi:https://doi.org/10.1128/cmr.00022-13

Djuikoue, I. C., Woerther, P. L., Toukam, M., Burdet, C., Ruppé, E., Gonsu, K. H., . . . Ngogang, J. (2016). Intestinal carriage of Extended Spectrum Beta-Lactamase producing E. coli in women with urinary tract infections, Cameroon. J Infect Dev Ctries, 10(10), 1135-1139. doi:https://doi.org/10.3855/jidc.7616

Elshaghabee, F. M. F., Rokana, N., Gulhane, R. D., Sharma, C., & Panwar, H. (2017). Bacillus As Potential Probiotics: Status, Concerns, and Future Perspectives. Frontiers in Microbiology, 8. doi:https://doi.org/10.3389/fmicb.2017.01490

Fijan, S. (2014). Microorganisms with claimed probiotic properties: an overview of recent literature. Int J Environ Res Public Health, 11(5), 4745-4767. doi:https://doi.org/10.3390/ijerph110504745

Fooks, L. J., & Gibson, G. R. (2002). Probiotics as modulators of the gut flora. Br J Nutr, 88 Suppl 1, S39-49. doi:https://doi.org/10.1079/BJN2002628

Guo, X., Li, D., Lu, W., Piao, X., & Chen, X. (2006). Screening of Bacillus strains as potential probiotics and subsequent confirmation of the in vivo effectiveness of Bacillus subtilis MA139 in pigs. Antonie Van Leeuwenhoek, 90(2), 139-146. doi:https://doi.org/10.1007/s10482-006-9067-9

Kaur, A., Capalash, N., & Sharma, P. (2020). Expression of Meiothermus ruber luxS in E. coli alters the antibiotic susceptibility and biofilm formation. Appl Microbiol Biotechnol, 104(10), 4457-4469. doi:https://doi.org/10.1007/s00253-020-10480-8

Kim, K., He, Y., Xiong, X., Ehrlich, A., Li, X., Raybould, H., . . . Liu, Y. (2019). Dietary supplementation of Bacillus subtilis influenced intestinal health of weaned pigs experimentally infected with a pathogenic E. coli. Journal of Animal Science and Biotechnology, 10(1), 52. doi:https://doi.org/10.1186/s40104-019-0364-3

Köhler, C. D., & Dobrindt, U. (2011). What defines extraintestinal pathogenic Escherichia coli? International journal of medical microbiology, 301(8), 642-647. doi:https://doi.org/10.1016/j.ijmm.2011.09.006

Lin, S., Hung, A., & Lu, J. (2011). Effects of supplement with different level of Bacillus coagulans as probiotics on growth performance and intestinal microflora populations of broiler chickens. Journal of Animal and Veterinary Advances, 10(1), 111-114. doi:https://doi.org/10.3923/javaa.2011.111.114

Livak, K. J., & Schmittgen, T. D. (2001). Analysis of Relative Gene Expression Data Using Real-Time Quantitative PCR and the 2−ΔΔCT Method. Methods, 25(4), 402-408. doi:https://doi.org/10.1006/meth.2001.1262

Medellin-Peña, M. J., Wang, H., Johnson, R., Anand, S., & Griffiths, M. W. (2007). Probiotics Affect Virulence-Related Gene Expression in Escherichia coli O157:H7. Appl Environ Microbiol, 73(13), 4259-4267. doi:https://doi.org/10.1128/AEM.00159-07

Nakayama, T., Kawahara, R., Kumeda, Y., & Yamamoto, Y. (2018). Extended-spectrum β-lactamase-producing Escherichia coli contributes to the survival of cefotaxime-susceptible E. coli under high concentrations of cefotaxime by acquisition of increased AmpC expression. FEMS Microbiol Lett, 365(5). doi:https://doi.org/10.1093/femsle/fny009

Piri, F., Tajabadi Ebrahimi, M., & Amini, K. (2019). Molecular investigation of CTX-M gene in Extended Spectrum β Lactamases (ESBLs) producing Pseudomonas aeruginosa isolated from Iranian patients with burn wound infection. Archives of Medical Laboratory Sciences, 4(1). doi:https://doi.org/10.22037/amls.v4i1.23078

Saitou, N., & Nei, M. (1987). The neighbor-joining method: a new method for reconstructing phylogenetic trees. Molecular Biology and Evolution, 4(4), 406-425. doi:https://doi.org/10.1093/oxfordjournals.molbev.a040454

Shen, Z., Ding, B., Bi, Y., Wu, S., Xu, S., Xu, X., . . . Wang, M. (2017). CTX-M-190, a Novel β-Lactamase Resistant to Tazobactam and Sulbactam, Identified in an Escherichia coli Clinical Isolate. Antimicrob Agents Chemother, 61(1). doi:https://doi.org/10.1128/aac.01848-16

Ur Rahman, S., Ali, T., Ali, I., Khan, N. A., Han, B., & Gao, J. (2018). The Growing Genetic and Functional Diversity of Extended Spectrum Beta-Lactamases. Biomed Res Int, 2018, 9519718. doi:https://doi.org/10.1155/2018/9519718

Wang, X., Li, S., Lu, X., Hu, P., Chen, H., Li, Z., . . . Wang, X. (2016). Rapid method of luxS and pfs gene inactivation in enterotoxigenic Escherichia coli and the effect on biofilm formation. Molecular Medicine Reports, 13(1), 257-264. doi:https://doi.org/10.3892/mmr.2015.4532