Detection target genes in combating biofilm forms in $\textit{Salmonella}$ Typhimurium 14028

Abstract

In this study, the relationship of $\textit{hilA, invA, adrA, spiC, otsB}$ and $\textit{csgD}$ genes, which are known to play critical roles in the pathogenicity and virulence of $\textit{Salmonella}$ strains, with biofilm formation was investigated by examining the changes in the expression levels of these genes during the transition from planktonic form to biofilm form. When the virulence gene expressions between the S. Typhimurium 14028 mutant, which lost its ability to form biofilms due to $\textit{csgD}$ gene deletion, and the wild type strain were compared, it was determined that the expression levels of $\textit{hilA, invA}$ and $\textit{adrA}$ genes increased, whereas the expression levels of $\textit{spiC, otsB}$ and $\textit{csgD}$ genes decreased. These data indicate that all examined genes play critical activation or inhibition roles in biofilm regulation as well as pathogenicity and virulence. On the other hand, in the mutant strain; The increase in the expression levels of $\textit{hilA, invA}$ and $\textit{adrA}$ genes shows that inhibitors of the proteins encoded by these genes have the potential to be of practical use in the prevention and control of infections caused by both biofilm-forming and non-biofilm-forming $\textit{Salmonella}$ strains.

Keywords

• Salmonella
• biofilm
• gene expression
• qRT-PCR

References

1. Brasil, A. Agência Nacional de Vigilância Sanitária., Programa Nacional de Monitoramento de Prevalência e da Resistência Bacteriana em Frangos - PREBAF. Monitoramento de prevalência e do perfil de susceptibilidade aos antimicrobianos em enterococos e salmonelas isolados de carcaças de frango congeladas comercializadas no Brasil. Brasília, DF, ANVISA, (2013), p.1-171.
2. Cheng, R.A., Eade, C.R., Wiedmann, M., Embracing diversity: differences in virulence mechanisms, disease severity, and host adaptations contribute to the success of nontyphoidal Salmonella as a foodborne pathogen. Frontiers in Microbiology, 10 (2019), 1368-1378. https://doi.org/10.3389/fmicb.2019.01368.
3. Scallan, E., Hoekstra, R.M., Angulo. F.J., Tauxe, R.V., Widdowson, M.A., Roy, S.L., Jones, J.L., Griffin, P.M., Foodborne illness acquired in the United States--major pathogens. Emerging Infectious Diseases. 17 (2011), 7-15. doi: 10.3201/eid1701.P11101.
4. Xu, H., Lee, H.Y., Ahn, J., Growth and virulence properties of biofilm-forming Salmonella enterica serovar. Typhimurium under different acidic conditions. Applied Environmental Microbiology, 76 (2010), 910-917. doi: 10.1128/AEM.01508-10.
5. Hasegawa, A., Hara-Kudo, Y., Kumagai, S., Survival of Salmonella strains differing in their biofilm-formation capability upon exposure to hydrochloric and acetic acid and to high salt. Journal of Veterinary Medical Science, 73 (2011), 1163-1168. doi: 10.1292/jvms.11-0090.
6. Yang, Y. C., Lii, C. K., Wei, Y. L., Li, C. C., Lu, C. Y., Liu, K. L., Docosahexaenoic acid inhibition of inflammation is partially via cross-talk between Nrf2/heme oxygenase 1 and IKK/NFkappaB pathways. The Journal of Nutritional Biochemistry. 24 (2013), 204–212. doi: 10.1016/j.jnutbio.2012.05.003.
7. Akçelik, M., Akçelik, N., What makes another life possible in bacteria? Global regulators as architecs of bacterial biofilms. World Journal of Microbiology and Biotechneology, 28 (2023), 26-36. doi: 10.1007/s11274-022-03376-4.
8. Allen, N. E., From vancomycin to oritavancin: the discovery and development of a novel lipoglycopeptide antibiotic. Anti-Infective Agents in Medicinal Chemistry. 9 (2010), 23–47. doi: 10.2174/187152110790886745.

9. Barraud, N., Kelso, M. J., Rice, S. A., Kjelleberg, S., Nitric oxide: a key mediator of biofilm dispersal with applications in infectious diseases. Current Pharmaceutical Design. 21 (2015), 31–42. doi: 10.2174/1381612820666140905112822.
10. Özdemir, C., Akçelik, N., Özdemir, F.N., Evcili, İ., Kahraman, T., Gürsel, İ., Akçelik, M., The role of bcsE gene in the pathogenicity of Salmonella. Pathogens and Disease. 79 (2021),156-165. doi: 10.1093/femspd/ftab037
11. Özdemir, F.N., Buzrul, S., Özdemir, C. Akçelik, N., Akçelik, M., Determination of an effective agent combination using nisin against Salmonella biofilm. Archives of Microbiology. 204 (2022), 167-172. doi: 10.1007/s00203-022-02766-4
12. Kurtz, J.R., Nieves, W., Bauer, D.L., Israel, K.E., Adcox, H.E., Gunn, J.S., Morici, L.A., McLachlan, J.B., Salmonella persistence and host immunity are dictated by the anatomical microenvironment. Infection and Immunity. 21(2020) 3507-3512. doi: 10.1128/IAI.00026-20
13. Yoon, B. K., Jackman, J. A., Valle-González, E. R., Cho, N.J., Antibacterial free fatty acids and monoglycerides: biological activities, experimental testing, and therapeutic applications. International Journal of Molecular Sciences. 19 (2018), 1114-1121. doi: 10.3390/ijms19041114.
14. Gualdi, L., Tagliabue, L., Bertagnoli, S., Ierano, T., De Castro, C., Landini, P., Cellulose modulates biofilm formation by counteracting curli-mediated colonization of solid surfaces in Escherichia coli. Microbiology, 154 (2008). 2017–2024. doi: 10.1099/mic.0.2008/018093-0.
15. Monteiro, C., Papenfort, K., Hentrich, K., Ahmad, I., Le Guyon, S., Reimann, R., Grantcharova, N., Römling, U., Hfq and Hfq-dependent small RNAs are major contributors to multicellular development in Salmonella enterica serovar Typhimurium. RNA Biology. 94 (2012), 489-502. doi: 10.4161/rna.19682
16. Mizusaki, H., Takaya, A., Yamamoto, T., Aizawa, S., Signal pathway in salt-activated expression of the Salmonella pathogenicity island 1 type III secretion system in Salmonella enterica serovar Typhimurium. Journal of Bacteriology, 190 (2008), 4624–4631. doi: 10.1128/JB.01957-07.
17. D'Souza, D., Critzer, F., Golden, D., Real-time reverse-transcriptase polymerase chain reaction for the rapid detection of Salmonella using invA primers. Foodborne Pathogens and Disease, 6 (2009), 1097–1106. doi: 10.1089/fpd.2009.0322.
18. Balaji, B., O'Connor, K., Lucas, J., Anderson, J., Csonka, L., Timing of induction of osmotically controlled genes in Salmonella enterica Serovar Typhimurium, determined with quantitative real-time reverse transcription-PCR. Applied and Environmental Microbiology, 71 (2005), 8273–8283. doi: 10.1128/AEM.71.12.8273-8283.2005.
19. Appiah, T., Boakye,Y.D., Agyare, C., Antimicrobial activities and time-kill kinetics of extracts of selected Ghanaian mushrooms. Evidence-based Complementary and Alternative Medicine, 17 (2017), 1-15. https://doi.org/10.1155/2017/4534350.
20. Hall-Stoodley, L., Costerton, J.W., Stoodley, P., Bacterial biofilms: From the natural environment to infectious diseases. Nature Reviews Microbiology. 2 (2004), 95–108.doi: 10.1038/nrmicro821.
21. Lee, C.K., De Anda, J., Baker, A.E., Bennett, R.R., Luo, Y., Lee, E.Y., Multigenerational memory and adaptive adhesion in early bacterial biofilm communities. Proceedings of the National Academy of Sciences, 115 (2018), 4471–4476. doi: 10.1073/pnas.1720071115.
22. Krukiewicz, K., Kazek-Kesik, A., Brzychczy-Włoch, M., Los, M.J., Ateba, C.N., Mehrbod, P., Ghavami, S., Shyntum, D.Y., Recent advances in the control of clinically important biofilms. International Journal of Molecular Sciences, 23 (2022), 4-26. doi: 10.3390/ijms23179526.
23. Bajaj, V., Hwang, C., Lee, C.A., hilA is a novel ompR/toxR family member that activates the expression of Salmonella typhimurium invasion genes. Molecular Microbiology, 18 (1995), 715-727. doi: 10.1111/j.1365-2958.1995.mmi_18040715.x
24. Kim, S., Jung, U.T., Kim, S.K., Lee, J.H., Choi, H.S., Kim, C.S., Jeong, M.Y., Nanostructured multifunctional surface with antireflective and antimicrobial characteristics. ACS Applied Materials & Interfaces, 7 (2015), 326–331. doi: 10.1021/am506254r.
25. Tsai,M. H., Liang, Y.H., Chen, C.Y., Chiu, C. H., Characterization of Salmonella resistance to bile during biofilm formation. Journal of Microbiology, Immunology and Infection, 53 (2020), 518-524. doi: 10.1016/j.jmii.2019.06.003.
26. Wang,Y., Cai, Y., Zhang, J., Liu, D., Gong, X., Pan, Z., Geng, S., Jiao, X., Controvesy surrounding the function of SpiC protein in Salmonella. Frontiers in Microbiology, 10 (2019), 331-340. https://doi.org/10.3389/fmicb.2019.01784.
27. Aviles, B., Klotz, C., Eifert, J., Williams, R., Ponder, M., Biofilms promote survival and virulence of Salmonella enterica sv. Tennessee during prolonged dry storage and after passage through an in vivo digestion system. International Journal of Food Microbiology, 163 (2013), 252-259. doi: 10.1016/j.ijfoodmicro.2013.01.026.
28. Sokaribo, A. S., Hansen, E. G., McCarthy, M., Desin, T. S., Waldner, L. L., MacKenzie, K. D., Mutwiri, G., Jr, Herman, N. J., Herman, D. J., Wang, Y., White, A. P., Metabolic activation of CsgD in the regulation of Salmonella Biofilms. Microorganisms, 8 (2020), 964-871. doi: 10.3390/microorganisms8070964