The Role of Cell Wall Mutation in the Pathogenicity of E. coli H7:O157: A Molecular Evolution Study
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Abstract
- coli H7:O157, the cauWallsative pathogen of many disease outbreaks and food poisoning cases, is the subject of many studies. The bacterium’s pathogenicity is highly associated with cell wall modi-fications and changes. A total of 20 fecal samples from patients who showed typical symptoms of the infection and tested positive for E. coli H7:O157 were collected. Another 20 samples from ani-mals that showed signs of infection with this bacterium were also collected and processed. The bac-terium was isolated and identified using cultural and molecular methods. The waa K, waa L, and waa Y sites were subjected to site-directed mutagenesis, and the effect of these mutations was studied and analyzed through their influence on the pathogenicity compared to the wild type. We found that the invasiveness and morbidity of mutant E. coli H7:O157 increased significantly when ingest-ed by laboratory animals. This may be attributed to waa K and waa L since they led to a significant change in the transmembrane helix ratio compared to the wild type, enabling the uncontrolled re-lease of the Shiga toxin into the infected animals and causing their death in 6 h. Specific sites in the waa operon, namely waa K and waa L, play the leading role in controlling the progress of patho-genicity. Mutations in these sites may increase the virulence of this bacterium.
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References
[1] Fontaine, R.; Arnon, S.; Martin, W. Raw hamburger: An interstate common source of human salmonellosis. Am. J. Epidemiol. 1978, 107, 36–45. https://doi.org/10.1093/oxfordjournals.aje.a112505.
[2] Ahmed, S.T.; Jasim, A.N.; Ali, A.M. Isolation and characterization E. coli O157:H7 from meat and cheese and detection of Stx1, Stx2, hlyA, eaeA using multiplex PCR. JOBRC 2011, 5, 15–24.
[3] Thompson, J.S.; Hodge, D.S.; Borczyk, A.A. Rapid biochemical test to identify verocytotoxin-positive strains of Escherichia coli serotype O157. J. Clin. Microbiol. 1990, 28, 2165–2168.
[4] Blanco, M.; Blanco, J.; Mora, A.; Dahbi, G.; Alonso, M.; González, E. Serotypes, virulence genes, and intimin types of Shiga toxin (verotoxin)-producing Escherichia coli isolates from cattle in Spain and identification of a new intimin variant gene (eae-xi). J. Clin. Microbiol. 2004, 42, 645–651.
[5] Al-Khyat, F. Enterohaemorrhagic E. coli O157 in locally produced soft cheese. Iraqi J. Vet. Med. 2008, 32, 88–99.
[6] Guth, B.; Prado, V.; Rivas, M. Shiga toxin-producing Escherichia coli. In Pathogenic Escherichia coli in Latin America, Torres, A.G., Ed.; Department of Microbiology and Immunology, Sealy Institute for Vaccine Sciences, University of Texas Medical Branch: Galveston, TX, USA, 2010.
[7] Paton, A.W.; Paton, J.C. Direct detection of Shiga toxigenic Escherichia coli strains belonging to serogroups O111, O157, and O113 by multiplex PCR. J. Clin. Microbiol. 1999, 37, 3362–3365.
[8] Heiman, K.E.; Mody, R.K.; Johnson, S.D.; Griffin, P.M.; Gould, L.H. Escherichia coli O157 outbreaks in the United States, 2003–2012. Emerg. Infect. Dis. 2015, 21, 1293–1301. https://doi.org/10.3201/eid2108.141364.
[9] Van, T.T.H.; Yidana, Z.; Smooker, P.M.; Coloe, P.J. Antibiotic use in food animals worldwide, with a focus on Africa: Pluses and minuses. J. Glob. Antimicrob. Resist. 2020, 20, 170–177. https://doi.org/10.1016/j.jgar.2019.07.031.
[10] Mohammed, Z.A. Isolation and Identification of Escherichia coli O157:H7 from locally minced meat and imported minced and chicken meat. Iraqi J. Vet. Med. 2008, 32, 100–113.
[11] Hasan, J.M.; Najim, S.S. A review of the Prevalence of Enterohemorrhagic E. coli in Iraq. JOBRC 2024, 18, 33–39.
[12] Khalil, Z.K. Isolation and identification of Staphylococcus aureus, Listeria monocytogenes, E. coli O157:H7 and Salmonella specious from raw beef and lamb meat in Baghdad by PCR. Iraqi J. Sci. 2016, 57, 1891–1897.
[13] Lim, J.M.; Singh, S.R.; Duong, M.C.; Legido-Quigley, H.; Hsu, L.Y.; Tam, C.C. Impact of national interventions to promote responsible antibiotic use: A systematic review. J. Antimicrob. Chemother. 2020, 75, 14–29. https://doi.org/10.1093/jac/dkz348.
[14] Nataro, J.P.; Kaper, J.B. Diarrheagenic Escherichia coli. Clin. Microbiol. Rev. 1998, 11,142–201. https:// 10.1128/cmr.11.1.142. Caprioli A, Morabito S, Brugere H, Oswald E. Enterohaemorrhagic Escherichia coli: Emerging issues on virulence and modes of transmission. Vet. Res. 2005, 36, 289–311. https://doi.org/10.1051/vetres:2005002.
[15] Beata, S.; Michał, T.; Mateusz, O.; et al. Norepinephrine affects the interaction of adherent-invasive Escherichia coli with intestinal epithelial cells. Virulence 2021, 12, 630–637. https://doi.org/10.1080/21505594.2021.1882780.
[16] Perna, N.T.; Plunkett, G.; Burland, V.; Mau, B.; Glasner, J.D.; Rose, D.J.; Mayhew, G.F.; Evans, P.S.; Gregor, J.; Kirkpatrick, H.A.; et al. Genome sequence of enterohaemorrhagic Escherichia coli O157:H7. Nature 2001, 409, 529–533.
[17] Dobrindt, U.; Agerer, F.; Michaelis, K.; Janka, A.; Buchrieser, C.; Samuelson, M.; Svanborg, C.; Gottschalk, G.; Karch, H.; Hacker, J. Analysis of genome plasticity in pathogenic and commensal Escherichia coli isolates by use of DNA arrays. J. Bacteriol. 2003, 185, 1831–1840.
[18] Wick, L.M.; Qi, W.; Lacher, D.W.; Whittam, T.S. Evolution of genomic content in the stepwise emergence of Escherichia coli O157:H7. J. Bacteriol. 2005, 187, 1783–1791.
[19] Amézquita-López, B.A.; Quiñones, B.; Soto-Beltrán, M.; et al. Antimicrobial resistance profiles of Shiga toxin-producing Escherichia coli O157 and non-O157 recovered from domestic farm animals in rural communities in Northwestern Mexico. Antimicrob. Resist. Infect. Control. 2016, 5, 1. https://doi.org/10.1186/s13756-015-0100-5.
[20] Aklilu, F.; Daniel, K.; Ashenaf, K. Prevalence and antibiogram of Escherichia coli O157 isolated from bovine in Jimma, Ethiopia: Abattoir-based survey. Ethiop. Vet. J. 2017, 21, 109–120.
[21] Tarekegn, A.A.; Mitiku, B.A.; Alemu, Y.F. Escherichia coli O157:H7 beef carcass contamination and its antibiotic resistance in Awi Zone, Northwest Ethiopia. Food Sci. Nutr. 2023, 11, 6140–6150.
[22] Gambushe, S.M.; Zishiri, O.T.; E El Zowalaty, M. Review of Escherichia coli O157:H7 Prevalence, Pathogenicity, Heavy Metal and Antimicrobial Resistance, African Perspective. Infect. Drug Resist. 2022, 15, 4645–4673.
[23] Banerjee, S.; Lo, K.; Ojkic, N.; Stephens, R.; Scherer, N.F.; Dinner, A.R. Mechanical feedback promotes bacterial adaptation to antibiotics. Nat. Phys. 2021, 17, 403–409.
[24] Harris, L.K.; Theriot, J.A. Relative rates of surface and volume synthesis set bacterial cell size. Cell 2016, 165, 1479–1492.
[25] Assefa, A.; Garcias, B.; Mourkas, E.; Molina-López, R.; Darwich, L. Global distribution of antimicrobial resistance genes in Escherichia coli isolated from wild animals using genomes available in public databases. Sci. Total Environ. 2025, 985, 179742.
[26] Ojkic, N.; Banerjee, S. Bacterial cell shape control by nutrient-dependent synthesis of cell division inhibitors. Biophys. J. 2021, 120, 2079–2084.
[27] Ojkica, N.; Serbanescua, D.; Banerjee, S. Antibiotic Resistance via Bacterial Cell Shape-Shifting. mBio 2022, 13, e0065922. https://doi.org/10.1128/mbio.00659-22.
[28] Nikolic, P. and Mudgil P. The Cell Wall, Cell Membrane and Virulence Factors of Staphylococcus aureus and Their Role in Antibiotic Resistance. Microorganisms 2023, 11, 259. https://doi.org/10.3390/microorganisms11020259.
[29] WHO. WHO Estimates of the Global Burden of Foodborne Diseases: Foodborne Disease Burden Epidemiology Reference Group 2007–2015; WHO: Geneva, Switzerland, 2016.
[30] March, S.B.; Ratnam, S. Sorbitol-MacConkey Medium for Detection of Escherichia coli 0157:H7 Associated with Hemorrhagic Colitis. J. Clin. Microbiol. 1986, 23, 869–872.
[31] Leininger, D.J.; Roberson, J.R.; Elvinger, F. Use of eosin methylene blue agar to differentiate Escherichia coli from other gram-negative mastitis pathogens. J. Vet. Diagn. Invest. 2001, 13, 273–275.
[32] Ally, C. Antony, Mini K. Paul, Reshma Silvester, Aneesa P.A., Suresh K., Divya P.S., Simmy Paul, Fathima P.A. and Mohamed Hatha Abdulla. Comparative Evaluation of EMB Agar and Hicrome E. coli Agar for Differentiation of Green Metallic Sheen Producing Non-E. coli and Typical E. coli Colonies from Food and Environmental Samples. J. Pure Appl. Microbiol. 2016, 10, 2863–2870.
[33] Rani, A.; Ravindran, V.B.; Surapaneni, A.; Mantri, N.; Ball, A.S. Review: Trends in point-of-care diagnosis for Escherichia coli O157:H7 in food and water. Int. J. Food Microbiol. 2021, 349, 109233.
[34] Pagnout, C.; Sohm, B.; Razafitianamaharavo, A.; Caillet, C.; Offroy, M.; Leduc, M.; Gendre, H.; Jomini, S.; Beaussart, A.; Bauda, P.; et al. Pleiotropic effects of rfa-gene mutations on Escherichia coli envelope properties. Sci. Rep. 2019, 9, 9696. https://doi.org/10.1038/s41598-019-46100-3.
[35] Raetz, C.R.; Whitfield, C. Lipopolysaccharide endotoxins. Annu. Rev. Biochem. 2002, 71, 635–700.
[36] Han, W.; Wu, B.; Li, L.; Zhao, G.; Woodward, R.; Pettit, N.; Cai, L.; Thon, V.; Wang, P. Defining Function of Lipopolysaccharide O-antigen Ligase WaaL Using Chemoenzymatically Synthesized Substrates. Microbiology 2012, 287, 5357–5365.
[37] Abeyrathne, P.D.; Daniels, C.; Poon, K.K.H.; Matewish, M.J.; Lam, J.S. Functional Characterization of WaaL, a Ligase Associated with Linking O-Antigen Polysaccharide to the Core of Pseudomonas aeruginosa Lipopolysaccharide. J. Bacteriol. 2005, 187, 3002–3012. https://doi.org/10.1128/JB.187.9.3002-3012.2005.
[38] Klena, J.D.; Ashford, R.S., II; Schnaitman, C.A. Role of Escherichia coli K-12 rfa genes and the rfp gene of Shigella dysenteriae 1 in generation of lipopolysaccharide core heterogeneity and attachment of O antigen. J. Bacteriol. 1992, 174, 7297–7307. doi.org/10.1128/jb.174.22.7297-7307.1992
[39] Kaniuk, N.A.; Monteiro, M.A.; Parker, C.T.; Whitfield, C. Molecular diversity of the genetic loci responsible for lipopolysaccharide core oligosaccharide assembly within the genus Salmonella. Mol. Microbiol. 2002, 46, 1305–1318.
[40] Frirdich, E.; Lindner, B.; Holst, O.; Whitfield, C. Overexpression of the waaZ Gene Leads to Modification of the Structure of the Inner Core Region of Escherichia coli Lipopolysaccharide, Truncation of the Outer Core, and Reduction of the Amount of O Polysaccharide on the Cell Surface. J. Bacteriol. 2003, 185, 1659–1671. https://doi.org/10.1128/JB.185.5.1659-1671.2003.
[41] Yethon, J.A.; Heinrichs, D.; Monteiro, M.A.; Perry, M.B.; Whitfield, C. Involvement of waaY, waaQ, and waaP in the Modification of Escherichia coli Lipopolysaccharide and Their Role in the Formation of a Stable Outer Membrane. J. Biol. Chem. 1998, 273, 26310–26316.
[42] Taylor, C.M.; Goldrick, M.; Lord, L.; Roberts, I.S. Mutations in the waaR Gene of Escherichia coli Which Disrupt Lipopolysaccharide Outer Core Biosynthesis Affect Cell Surface Retention of Group 2 Capsular Polysaccharides. J. Bacteriol. 2006, 188, 1165–1168. https://doi.org/10.1128/JB.188.3.1165-1168.2006.
[43] Al wendawi, S.H.A.; Al Rekaby, S.M. The Effeciency of Enteric Lactobacillus in Preventing Hemorrhagic Colitis and Blocking Shiga Toxins Productions in Rats Models Infected with Enterohemorrhagic Escherichia coli (EHEC). Iraqi J. Agric. Sci. 2021, 52, 1346–1355.
[44] Al-Taii, D.H.F. Effects of E. coli O157:H7 Experimental Infections on Rabbits. Iraqi J. Vet. Med. 2019, 43, 34–42.
[45] Wang, J.; Ma, W.; Wang, X. Insights into the structure of Escherichia coli outer membrane as the target for engineering microbial cell factories. Microb. Cell Fact. 2021, 20, 73. https://doi.org/10.1186/s12934–021–01565–8.
[46] Kaper, J.B.; Nataro, J.P.; Mobley, H.L. Pathogenic Escherichia coli. Nat. Rev. Microbiol. 2004, 2, 123–140.
[47] Croxen, M.A.; Finlay, B.B. Molecular mechanisms of Escherichia coli pathogenicity. Nat. Rev. Microbiol. 2010, 8, 26–38.
[48] Sora, V.M.; Meroni, G.; Martino, P.A.; Soggiu, A.; Bonizzi, L.; Zecconi, A. Extraintestinal Pathogenic Escherichia coli: Virulence Factors and Antibiotic Resistance. Pathogens 2021, 10, 1355, https://doi.org/10.3390/pathogens10111355.
[49] Khaleq, M.A.A.; Abd, A.H.; Dhahi, M.A. Efficacy of Combination of Meropenem with Gentamicin, and Amikacin against Resistant E. coli Isolated from Patients with UTIs: In vitro Study. Iraqi J. Pharm. Sci. 2011, 20, 66–74.
[50] Khan Md, F.; Rashid, S.S. Molecular Characterization of Plasmid-Mediated Non-O157 Verotoxigenic Escherichia coli Isolated from Infants and Children with Diarrhea. Baghdad Sci. J. 2020, 17, 710–719.
[51] Wang, X.; Yu, D.; Chui, L.; Zhou, T.; Feng, Y.; Cao, Y.; Zhi, S. A Comprehensive Review on Shiga Toxin Subtypes and Their Niche-Related Distribution Characteristics in Shiga-Toxin-Producing E. coli and Other Bacterial Hosts. Microorganisms 2024, 12, 687. https://doi.org/10.3390/microorganisms12040687.
[52] O’Neil, J. Review on Antimicrobial Resistance Antimicrobial Resistance: Tackling a Crisis for the Healthand Wealth of Nations. Available online: https://amr-review.org/sites/default/files/AMR%20Review%20Paper%20-%20Tackling%20a%20crisis%20for%20the%20health%20and%20wealth%20of%20nations_1.pdf (accessed on 7 2023).
[53] Gray, D.A.; Wenzel, M. Multitarget approaches against multiresistant superbugs. ACS Infect. Dis. 2020, 6, 1346–1365. https://doi.org/10.1021/acsinfecdis.0c00001.
[54] Zhao, M.; Li, Y.; Wang, Z. Mercury and mercury-containing preparations: History of use, clinical applications, pharmacology, toxicology, and pharmacokinetics in traditional Chinese medicine. Front. Pharmacol. 2022, 13, 807807. https://doi.org/10.3389/fphar.2022.807807.
[55] Lekhan, A.; Turner, R.J. Exploring antimicrobial interactions between metal ions and quaternary ammonium compounds toward synergistic metalloantimicrobial formulations. Antimicrob. Chemother. 2024, 12, e0104724. https://doi.org/10.1128/spectrum.01047-24