Genetic Identification of Shiga Toxin Encoding Gene from Cases of Multidrug Resistance (MDR) Escherichia coli Isolated from Raw Milk
Abstract
Escherichia coli is one of bacteria which have resistant to three or more classes of antimicrobial agents. E. coli having resistant to three or more classes of antimicrobial drugs can be defined as multidrug-resistant (MDR) bacteria. The aim of the study was to evaluate the expression of Shiga toxin gen in MDR E. coli. A total of 250 raw milks samples were taken from dairy farms in Kediri, Probolinggo, Pasuruan, Blitar, and Batu Region, East Java Province, Indonesia. Each sample was cultured into enrichment media Brilliant Green Bile Lactose Broth and Eosin Methyllen blue agar, then identified with TSIA agar and IMVIC biochemistry test. Antibiotic sensitivity testing was done using Kirby-Bauer disc diffusion assay on medium Mueller-Hinton agar (Oxoid, CM0337). Antibiotics disks used were 30 µg of Tetracycline (Oxoid, CT0054), 10 µg of Streptomycin (Oxoid, CT0047), 30 µg of Chloramphenicol (Oxoid, CT0013), 5 µg of Trimethoprim (Oxoid, CT0057), and 30 µg of Aztreonam (Oxoid, CT0264). Isolate showing resistance to at least 3 antibiotics disk were then continued with PCR assay to identify Shiga toxin E. coli (STEC) encoding stx2 gene. The study was designed to evaluate the nucleotide analysis of STEC gene. The result showed that 6.25% (1/16) of STEC encoding gene was found in MDR E. coli. This report of molecular identification on the presence of STEC gene in MDR E. coli confirmed a wider spread of MDR E. coli that can threaten animal health and human health.
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Putra, A. R. S., M. H. Effendi, S. Koesdarto, & W. Tyasningsih. 2019. Molecular identification of Extended Spectrum Beta-Lactamase (ESBL) producing Escherichia coli isolated from dairy cows in East Java Province, Indonesia. Indian Vet. J. 96: 26-30.
Riyanto, J., Sunarto, B. S. Hertanto, M. Cahyadi, R. Hidayah, & W. Sejati. 2016. Produksi dan kualitas susu sapi perah penderita mastitis yang mendapat pengobatan antibiotik. Sains Peternak. 14: 30-41. https://doi.org/10.20961/sainspet.14.2.30-41
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Tahamtan, Y., M. Hayati, & M. Namavari. 2010. Prevalence and distribution of the stx1, stx2 genes in Shiga toxin producing E. coli (STEC) isolates from cattle. Iran. J. Microbiol. 2: 8-13.
Tekiner, I. H. & H. Özpinar. 2016. Occurrence and characteristics of extended spectrum Beta-Lactamases-producing Enterobacteriaceae from foods of animal origin. Braz. J. Microbiol. 47: 444-451. https://doi.org/10.1016/j.bjm.2015.11.034
Ventola, C. L. 2015. The antibiotic resistance crisis: part 1: causes and threats. Pharm. Ther. 40: 277-283.
Ahmed, A. M. & T. Shimamoto. 2015. Molecular analysis of multidrug resistance in Shiga toxin-producing Escherichia coli O157:H7 isolated from meat and dairy products. Int. J. Food Microbiol. 193: 68-73. https://doi.org/10.1016/j.ijfoodmicro.2014.10.014
Arya, R., B. Antonisamy, & S. Kumar. 2012. Sample size estimation in prevalence studies. Indian J. Pediatr. 79: 1482-1488. https://doi.org/10.1007/s12098-012-0763-3
Barriere, S. L. 2014. Clinical, economic and societal impact of antibiotic resistance. Expert Opin. Pharmacother. 16: 151-153. https://doi.org/10.1517/14656566.2015.983077
Brenjchi, M., A. Jamshidi, N. Farzaneh, & MR.Bassami. 2011. Identification of Shiga toxin-producing Escherichia coli O157:H7 in raw milk samples from dairy farms in Mashhad using multiplex PCR assay. Iranian Journal of Veterinary Research. 12: 145-149.
Chui, L., M. C. Lee, K. Malejczyk, L. Lim, D. Fok, & P. Kwong. 2011. Prevalence of Shiga toxin-producing Escherichia coli as Detected by Enzyme-Linked immunoassays and real-time PCR during the summer months in Northern Alberta, Canada. J. Clin. Microbiol. 49: 4307-4310. https://doi.org/10.1128/JCM.05211-11
CLSI. 2018. M100 Performance Standards for Antimicrobial Susceptibility Testing. 28th ed. Clinical and Laboratory Standards Institute, USA.
Cointe, A., A. Birgy, P. Mariani-Kurkdjian, S. Liguori, C. Courroux, J. Blanco, S. Delannoy, P. Fach, E. Loukiadis, P. Bidet, & S. Bonacorsi. 2018. Emerging multidrug-resistant hybrid pathotype Shiga toxin-producing Escherichia coli O80 and related strains of clonal complex 165, Europe. Emerg. Infect. Dis. 24: 2262-2269. https://doi.org/10.3201/eid2412.180272
Colavecchio, A., B. Cadieux, A. Lo, & L. D. Goodridge. 2017. Bacteriophages contribute to the spread of antibiotic resistance genes among foodborne pathogens of the Enterobacteriaceae family - A Review. Front. Microbiol. 8: 1108. https://doi.org/10.3389/fmicb.2017.01108
Dairy, N. Z. 2019. Dairy Cow Housing-A Good Practice Guide for Dairy Housing in New Zealand, Version 2. DairyNZ and Ministry for Primary Industries, Hamilton, New Zealand.
Economou, V. & P. Gousia. 2015. Agriculture and Food Animals as A Source of Antimicrobial-Resistant Bacteria. Infect. Drug Resist. 8: 49-61. https://doi.org/10.2147/IDR.S55778
Effendi, M. H., N. Harijani, S. M. Yanestria, & P. Hastutiek. 2018. Identification of Shiga toxin-producing Escherichia coli in raw milk samples from dairy cows in Surabaya, Indonesia. Philipp. J. Vet. Med. 55: 109-114.
Effendi, M. H., N. Harijani, Budiarto, N. P. Triningtya, W. Tyasningsih, & H. Plumeriastuti. 2019. Prevalence of pathogenic Escherichia coli isolated from subclinical mastitis in East Java Province, Indonesia. Indian Vet. J. 96: 22-25.
Etcheverría A. I. & N. L. Padola. 2013. Shiga toxin-producing Escherichia coli, factors involved in virulence and cattle colonization. Virulence 4: 366-372. https://doi.org/10.4161/viru.24642
Falagas, M.E., P.K. Koletsi, & I. A. Bliziotis. 2006. The diversity of definitions of multidrug-resistant (MDR) and pandrug-resistant (PDR) Acinetobacter baumannii and Pseudomonas aeruginosa. J. Med. Microbiol. 55: 1619-1629. https://doi.org/10.1099/jmm.0.46747-0
Goma, MKE, A. Indraswari, A. Haryanto, & D.A. Widiasih. 2019. Detection of Escherichia coli O157:H7 and Shiga toxin 2a gene in pork, pig feces, and clean water at Jagalan slaughterhouse in Surakarta, Central Java Province, Indonesia, Vet. World, 12(10): 1584-1590. https://doi.org/10.14202/vetworld.2019.1584-1590
Gould, I. M. 2008. The epidemiology of antibiotic resistance. Int. J. Antimicrob. Agents. 32: 2-9. https://doi.org/10.1016/j.ijantimicag.2008.06.016
Hunt, J. M. 2010. Shiga toxin-producing Escherichia coli (STEC). Clin. Lab. Med. 30: 21-45. https://doi.org/10.1016/j.cll.2009.11.001
Hunter, P. A., S. Dawson, G. L. French, H. Goossens, P. M. Hawkey, E. J. Kuijper, D. Nathwani, D. J. Taylor, C. J. Teale, R. E., Warren, M. H. Wilcox, N. Woodford, M. W. Wulf, & L. J. Piddock. 2010. Antimicrobial-resistant pathogens in animals and man: Prescribing, Practices and Policies. J. Antimicrob. Chemother. 65: i3-i17. https://doi.org/10.1093/jac/dkp433
Hussein, H. S. & T. Sakuma. 2005. Invited Review: Prevalence of Shiga Toxin-producing Escherichia coli in dairy cattle and their products. J. Dairy Sci. 88: 450-465. https://doi.org/10.3168/jds.S0022-0302(05)72706-5
Kallen, A. J., A. I. Hidron, J. Patel, & N. A. Srinivasa. 2010. Multidrug resistance among gram-negative pathogens that caused healthcare-associated infections reported to The National Healthcare Safety Network, 2006-2008. Infect. Control Hosp. Epidemiol. 31: 528-531. https://doi.org/10.1086/652152
Kanayama, A., Y. Yahata, Y. Arima, T. Takahashi, T. Saitoh, K. Kanou, K. Kawabata, T. Sunagawa, T. Matsui, & K. Oishi. 2015. Enterohemorrhagic Escherichia coli outbreaks related to childcare facilities in Japan, 2010-2013. BMC Infectious Diseases. 15: 539. https://doi.org/10.1186/s12879-015-1259-3
Kristianingtyas, L., M. H. Effendi, W. Tyasningsih, & F. Kurniawan. 2020. Genetic identification of blactx-M gene and blatem gene on extended spectrum beta lactamase (ESBL) producing Escherichia Coli from Dogs. Indian Vet. J. 97:01:17-21.
Llor, C. & L. Bjerrum. 2014. Antimicrobial resistance: risk associated with antibiotic overuse and initiatives to reduce the problem. Ther. Adv. Drug Saf. 5: 229-241. https://doi.org/10.1177/2042098614554919
Magiorakos, A. P., A. Srinivasan, R. B. Carey, Y. Carmeli, M. E. Falagas, C. G. Giske, S. Harbarth, J. F. Hindler, G. Kahlmeter, B. Olsson-Liljequist, D. L. Paterson, L. B. Rice, J. Stelling, M. J. Struelens, A. Vatopoulos, J. T. Weber, & D. L. Monnet. 2012. Multidrug-resistant, extensively drug-resistant and pandrug-resistant bacteria: An international expert proposal for interim standard definitions for acquired resistance. Clin. Microbiol. Infect. 18: 268-281. https://doi.org/10.1111/j.1469-0691.2011.03570.x
Martin, A. & L. Beutin. 2011. Characteristics of Shiga toxin-producing Escherichia coli from meat and milk products of different origins and association with food producing animals as main contamination sources. Int. J. Food Microbiol. 146: 99-104. https://doi.org/10.1016/j.ijfoodmicro.2011.01.041
Mc Collum, J. T., N. J. Williams, S. W. Beam, S. Cosgrove, P. J. Ettestad, T. S. Ghosh, A. C. Kimura, L. Nguyen, S. G. Stroika, R. L. Vogt, A. K. Watkins, J. R. Weiss, I. T. Williams, & A. B. Cronquist. 2012. Multistate outbreak of Escherichia coli O157:H7 infections associated with in-store sampling of an aged raw-milk Gouda cheese. J. Food Prot. 75: 1759-1765. https://doi.org/10.4315/0362-028X.JFP-12-136
Melton-Celsa, A. R. 2014. Shiga Toxin (Stx) classification, structure and function. Microbiol. Spectr. 2: EHEC-0024-2013. https://doi.org/10.1128/microbiolspec.EHEC-0024-2013
Mohammadi, P., R. Abiri, M. Rezaei, & S. Salmanzadeh-Ahrabi. 2013. Isolation of Shiga toxin-producing Escherichia coli from raw milk in Kermanshah, Iran. Iran. J. Microbiol. 5: 233-238.
Newell, D. G. & R. M. La Ragione. 2018. Enterohaemorrhagic and other Shiga toxin-producing Escherichia coli (STEC): where are we now regarding diagnostics and control strategies?. Transbound. Emerg. Dis. 65: 49-71. https://doi.org/10.1111/tbed.12789
Ningrum, S. G., R. D. Soejoedono, H. Latif, W. Arnafia, & I. W. T. Wibawan. 2016. Prevalence and characterization of Shiga toxin-producing Escherichia coli isolated from slaughtered qurban animal in Jakarta Province. Med. Pet., 39: 90-94. https://doi.org/10.5398/medpet.2016.39.2.90
Noris, M., F. Mescia, & G. Remuzzi. 2012. STEC-HUS, Atypical HUS and TTP are all diseases of complement activation. Nat. Rev. Nephrol. 8: 622-633. https://doi.org/10.1038/nrneph.2012.195
Paton J. C. & A. W. Paton. 1998. Pathogenesis and diagnosis of Shiga toxin-producing Escherichia coli infections. Clin. Microbiol. Rev. 11: 450-479. https://doi.org/10.1128/CMR.11.3.450
Putra, A. R. S., M. H. Effendi, S. Koesdarto, & W. Tyasningsih. 2019. Molecular identification of Extended Spectrum Beta-Lactamase (ESBL) producing Escherichia coli isolated from dairy cows in East Java Province, Indonesia. Indian Vet. J. 96: 26-30.
Riyanto, J., Sunarto, B. S. Hertanto, M. Cahyadi, R. Hidayah, & W. Sejati. 2016. Produksi dan kualitas susu sapi perah penderita mastitis yang mendapat pengobatan antibiotik. Sains Peternak. 14: 30-41. https://doi.org/10.20961/sainspet.14.2.30-41
Tadesse, D. A., S. Zhao, E. Tong, S. Ayers, A. Singh, M.J. Bartholomew, and P.F. McDermott. 2012. Antimicrobial drug resistance in Escherichia coli from humans and food animals, United States, 1950-2002. Emerging Infectious Diseases. 18: 741-749. https://doi.org/10.3201/eid1805.111153
Tahamtan, Y., M. Hayati, & M. Namavari. 2010. Prevalence and distribution of the stx1, stx2 genes in Shiga toxin producing E. coli (STEC) isolates from cattle. Iran. J. Microbiol. 2: 8-13.
Tekiner, I. H. & H. Özpinar. 2016. Occurrence and characteristics of extended spectrum Beta-Lactamases-producing Enterobacteriaceae from foods of animal origin. Braz. J. Microbiol. 47: 444-451. https://doi.org/10.1016/j.bjm.2015.11.034
Ventola, C. L. 2015. The antibiotic resistance crisis: part 1: causes and threats. Pharm. Ther. 40: 277-283.
Authors
AnsharietaR., EffendiM. H., & PlumeriastutiH. (2021). Genetic Identification of Shiga Toxin Encoding Gene from Cases of Multidrug Resistance (MDR) Escherichia coli Isolated from Raw Milk. Tropical Animal Science Journal, 44(1), 10-15. https://doi.org/10.5398/tasj.2021.44.1.10
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