Genotypic and phenotypic characterization of Escherichia coli isolated from indigenous individuals in Malaysia

Document Type : Original Article

Authors

1 Centre of Toxicology and Health Risk Studies (CORE), Faculty of Health Sciences, Universiti Kebangsaan Malaysia, 50300 Kuala Lumpur, Malaysia

2 Department of Medical Microbiology, Faculty of Medicine, Universiti Malaya, 50603 Kuala Lumpur, Malaysia

3 Department of Parasitology, Faculty of Medicine, Universiti Malaya, 50603 Kuala Lumpur, Malaysia

4 Centre for Malaysian Indigenous Studies (CMIS), Universiti Malaya, Kuala Lumpur, Malaysia

5 Department of Biomedical Science, Faculty of Medicine, Universiti Malaya, 50603 Kuala Lumpur, Malaysia

Abstract

Objective(s): The occurrence of asymptomatic verocytotoxin (VT)-producing Escherichia coli (VTEC) infections among humans in recent years is posing a high risk to public health. Thus, the role of asymptomatic human carriers as a source of dissemination should not be underestimated. This study aimed to elucidate the phenotypic and genotypic characteristics of E. coli in the stool samples collected from indigenous individuals in Malaysia. 
Materials and Methods: E. coli strains (n=108) were isolated from stool samples obtained from 41 indigenous individuals. All strains were subjected to Repetitive Extragenic Palindromic-Polymerase Chain Reaction (REP-PCR) typing and confirmation of VTEC variants. Non-duplicate strains were selected based on REP-PCR profiles and further subjected to antimicrobial susceptibility test (AST). The genotypic and phenotypic characteristics of the strains were then correlated with the demographic data of the subjects.  
Results: A total of 66 REP-PCR profiles grouped in 53 clusters (F=85%) were obtained. Four genetically distinct strains were confirmed as VTEC (eaeA-positive). The predominant resistance was against ampicillin (34.2%), followed by trimethoprim-sulfamethoxazole (32.9%), ampicillin-sulbactam (5.5%), and ciprofloxacin (1.4%). All isolates were sensitive to amoxicillin-clavulanate, cefuroxime, ceftriaxone, imipenem, and meropenem. 
Conclusion: Genetically diverse E. coli and VTEC strains were found to colonize the intestines of the indigenous populations. This study is important for the prospective surveillance of E. coli among the indigenous individuals in Malaysia, especially in asymptomatic VTEC infection and antimicrobial resistance phenomenon.  

Keywords


1.    Kaper JB, Nataro JP, Mobley HLT. Pathogenic Escherichia coli. Nat Rev Microbiol 2004; 2:123-140. 
2.    Sperandio V, Nguyen Y. Enterohemorrhagic Escherichia coli (EHEC) pathogenesis. Front Cell Infect Microbiol 2012; 2:1-7. 
3.    Karmali MA. Infection by verocytotoxin-producing Escherichia coli. Clin Microbiol Rev 1989; 2:15-38. 
4.    Paton AW, Paton JC. Detection and characterization of Shiga Toxigenic Escherichia coli by using multiplex PCR assays for stx1; stx2, eaeA, Enterohemorrhagic Escherichia coli hlyA, rfbO111, and rfbO157. J Clin Microbiol 1998; 36:598-602. 
5.    Boerlin P, McEwen SA, Boerlin-Petzold F, Wilson JB, Johnson RP, Gyles CL. Associations between virulence factors of Shiga Toxin-Producing Escherichia coli and disease in humans. J Clin Microbiol 1999; 37:497-503. 
6.    Fagan PK, Hornitzky MA, Bettelheim KA, Djordjevic SP. Detection of Shiga-like toxin (stx1 and stx2), intimin (eaeA), and Enterohemorrhagic Escherichia coli (EHEC) hemolysin (EHEC hlyA) genes in animal feces by multiplex PCR. Appl Environ Microbiol 1999; 65:868-872. 
7.    Beutin L, Geier D, Zimmermann S, Karch H. Virulence markers of Shiga-like toxin-producing Escherichia coli strains originating from healthy domestic animals of different species. J Clin Microbiol 1995; 33:631-635.
8.    Chong CW, Ahmad AF, Lim YAL, Teh CSJ, Yap IKS, Lee SC, et al. Effect of ethnicity and socioeconomic variation to the gut microbiota composition among pre-adolescent in Malaysia. Sci Rep 2015; 5:13338. 
9.    Rajoo Y, Ambu S, Lim YAL, Rajoo K, Tey SC, Lu CW, et al. Neglected intestinal parasites, malnutrition and associated key factors: A population based cross-sectional study among indigenous communities in Sarawak, Malaysia. PLoS One 2017; 12:e0170174. 
10.    Chin YT, Lim YAL, Chong CW, Teh CSJ, Yap IKS, Lee SC, et al. Prevalence and risk factors of intestinal parasitism among two indigenous sub-ethnic groups in Peninsular Malaysia. Infect Dis Poverty 2016; 5:77-91. 
11.    Teh CSJ, Chua KH, Lim YAL, Lee SC, Thong KL. Loop-mediated isothermal amplification assay for detection of generic and verocytotoxin-producing Escherichia coli among indigenous individuals in Malaysia. Sci World J 2014; 2014:457839. 
12.    Institute of Medical Research (IMR). National Surveillance of Antimicrobial Resistance, Malaysia. Kuala Lumpur, Malaysia: Ministry of Health, MOH; 2019. Available from: https://www.imr.gov.my/MyOHAR/index.php/site/archive_rpt. 
13.    Fazlul MKK, Rashid SS, Nazmul MHM, Zaidul ISM, Baharudin R, Ramli ANM. A clinical update on antibiotic resistance Gram-negative bacteria in Malaysia - a review. J Int Pharm Res 2019; 45:270-283.
14.    Liu Y-Y, Wang Y, Walsh TR, Yi L-X, Zhang R, Spencer J, et al. Emergence of plasmid-mediated colistin resistance mechanism MCR-1 in animals and human beings in China: A microbiological and molecular biological study. Lancet Infect Dis 2016; 16:161-168. 
15.    Bradford PA, Kazmierczak KM, Biedenbach DJ, Wise MG, Hackel M, Sahm DF. Colistin-resistant Enterobacteriaceae: Correlation of β-lactamase production and colistin resistance among isolates from a global surveillance program. Antimicrob Agents Chemother 2015; 60:1385-1392. 
16.    Mobasseri G, Teh CSJ, Ooi PT, Thong KL. The emergence of colistin-resistant Klebsiella pneumoniae strains from swine in Malaysia. J Glob Antimicrob Resist 2019; 17:227-232. 
17.    Malchione MD, Torres LM, Hartley DM, Koch M, Goodman JL. Carbapenem and colistin resistance in Enterobacteriaceae in Southeast Asia: Review and mapping of emerging and overlapping challenges. Int J Antimicrob Agents 2019; 54:381-399. 
18.    Ong L, Muniandy K, How S, Tang S, Lim B. A report on antibiotic resistance of Escherichia coli isolated from veterinary samples in Malaysia from 2010 to 2013. Malays J Vet Res (Putrajaya) 2014; 5:70-71.
19.    Yap PSX, Ahmad Kamar A, Chong CW, Yap IKS, Thong KL, Choo YM, Md Yusof MY, Teh CSJ. Intestinal carriage of multidrug-resistant gram-negative bacteria in preterm infants during hospitalization in neonatal intensive care unit (NICU). Pathog Glob Health 2016; 110:238-246. 
20.    Kong RYC, So CL, Law WF, Wu RSS. A sensitive and versatile multiplex PCR system for the rapid detection of enterotoxigenic (ETEC), enterohaemorrhagic (EHEC) and enteropathogenic (EPEC) strains of Escherichia coli. Mar Pollut Bull 1999; 38:1207-1215. 
21.    Dashti AA, Jadaon MM, Abdulsamad AM, Dashti HM. Heat treatment of bacteria: A simple method of DNA extraction for molecular techniques. Kuwait Med J 2009; 41:117-122.
22.    Navia MM, Capitano L, Ruiz J, Vargas M, Urassa H, Schellemberg D, et al. Typing and characterization of mechanisms of resistance of Shigella spp. isolated from feces of children under 5 years of age from Ifakara, Tanzania. J Clin Microbiol 1999; 37:3113-3117. 
23.    Schmidt H, Scheef J, Morabito S, Caprioli A, Wieler LH, Karch H. A new Shiga toxin 2 variant (Stx2f) from Escherichia coli isolated from pigeons. Appl Environ Microbiol 2000; 66:1205-1208. 
24.    Clinical and Laboratory Standards Institute (CLSI). Performance Standards for Antimicrobial Susceptibility Testing; Twenty-Fifth Informational Supplement. CLSI document M100-S25. Wayne, Pennsylvania, United States of America: Clinical and Laboratory Standards Institute (CLSI); 2015.
25.    Tenover FC, Arbeit RD, Goering RV, Mickelsen PA, Murray BE, Persing DH, et al. Interpreting chromosomal DNA restriction patterns produced by pulsed-field gel electrophoresis: Criteria for bacterial strain typing. J Clin Microbiol 1995; 33:2233-2239.
26.    Tenaillon O, Skurnik D, Picard B, Denamur E. The population genetics of commensal Escherichia coli. Nat Rev Microbiol 2010; 8:207-217. 
27.    Adiba S, Nizak C, van Baalen M, Denamur E, Depaulis F. From grazing resistance to pathogenesis: The coincidental evolution of virulence factors. PLoS One 2010; 5:e11882. 
28.    Tadesse DA, Zhao S, Tong E, Ayers S, Singh A, Bartholomew MJ, et al. Antimicrobial drug resistance in Escherichia coli from humans and food animals, United States, 1950-2002. Emerg Infect Dis 2012; 18:741-749. 
29.    Ab Rahman N, Teng CL, Sivasampu S. Antibiotic prescribing in public and private practice: A cross-sectional study in primary care clinics in Malaysia. BMC Infect Dis 2016; 16:208-215.
30.    Teng CL, Tong SF, Khoo EM, Lee V, Zailinawati AH, Mimi O, et al. Antibiotics for URTI and UTI prescribing in Malaysian primary care settings. Aust Fam Physician 2011;40:325-329.