Determination of Antibiotic Susceptibility Profile and Int1, blaSHV and blaTEM Genes of Raw Milk Origin Enterobacteriaceae Isolates

In this study, a total of 68 raw milk samples were used to investigate the prevalence of Enterobacteriaceae in milk samples obtained from different dairy and supermarket in the province of Amasya (Turkey), as well as to determine the antibiotic resistance profile, the presence of Int1, blaTEM and blaSHV gene. In this study, isolates were obtained using classical culture technique. Then, detection of antibiotic resistance profile was carried out using disc diffusion methods. 12 different antibiotics were used as antibiotics including meropenem, cefotaxime, nalidixic acid, ceftriaxone, chloramphenicol, ceftazidime, streptomycin, ampicillin, gentamicin, tetracycline, levofloxacin and trimethoprim-sulfamethoxazole. Final, single strain PCR was created for the detection of ESBLs. For the aims, blaTEM and blaSHV genes (for determination of extended-spectrum beta-lactamase) were demonstrated by PCR assay. Then, for determination of Int1 was determined by using PCR assay. As a result, 50 isolates belonging to the Enterobacteiaceae family were obtained. Isolates against 41 (82%) ampicillin, 38 (76%) trimethoprim-sulfamethoxol, 7 (14%) ceftazidime, 6 (12%) cefotaxime, 2 (4%) meropenem and nalidixic acid and 1 (2%) ceftriaxone and streptomycin was determined as resistant. In addition, isolates were found to be 49 (98%), 50 (100%) 49 (98%) and 50 (100%) sensitive to chloramphenicol, gentamicin tetracycline and levofloxacin antibiotics, respectively. Among Enterobacteriaceae isolates, 22 (44%), 6 (12%) and 2 (4%) rates of strains were carrying Int1, blaSHV and blaTEM gene, respectively. In conclusion, the resistance of Enterobacteriaceae isolates isolated from milk samples to many antibiotics poses a potential danger in terms of public health

Keywords:

Enterobacteriaceae, antibiotic resistance, Int1, blaTEM, blaSHV raw milk,

PDF

___

1. Mitchell J, Yehuda C. 2008. Carbapenem-resistant Enterobacteriaceae. A potential threat. Journal of the American Medical Association. 300(24): 2911–2913.
2. AlTamimi M, AlSalamah A, AlKhulaifi M, AlAjlan H. 2017. Comparison of phenotypic and PCR methods for detection of carbapenemases production by Enterobacteriaceae, Saudi Journal of Biological Sciences, 24(1): 155-161.
3. Jarzab A, Gorska-Fraczek S, Rybka J, Witkowaka D. 2011. Enterobacteriaceae infection-diagnosis, antibitioc resistance and prevention. Postępy Higienyi Medycyny Doświadczalnej. 16(65): 55–72.
4. Ye Q, Wu Q, Zhang S, Zhang J, Yang G, Wang J, Xue L, Chen M. 2018. Characterization of Extended-Spectrum β-Lactamase-Producing Enterobacteriaceae From Retail Food in China. Frontiers in microbiology. 9: 1-12.
5. Salman A, Hamad I. 2011. Enumeration and identification of Coliform bacteria from raw milk in Khartoum State, Sudan. Journal of Cell and Animal Biology, 5(7): 121-128.
6. El-Mokadem EA, El-Leboudy AA, Amer AA. 2020. Occurrence of Enterobacteriaceae in Dairy Farm Milk. Alexandria Journal of Veterinary Sciences. 64(2): 66-71.
7. Tasci F. 2011. Microbiological and chemical properties of raw milk consumed in Burdur. Journal of Animal and Veterinary Advances. 10(5): 635-641.
8. Kornacki JL, Johnson JL. 2001. Enterobacteriaceae, coliforms, and Escherichia coli as quality and safety indicators. In: Downes P, Ito K (eds) Compendium of methods for the microbiological examination of foods. 4th edn. APHA, Washington. 69–82.
9. Mullane NR, Murray J, Drudy D, Prentice N, Whyte P, Wall PG, Parton A, Fanning S. 2006. Detection of Cronobacter sakazakii in dried infant milk formula by cationic-magnetic-bead capture. Applied Environmental Microbiology. 72(9): 6325–6330.
10. Kačániová M, Terentjeva M, Godočíková L, Puchalski C, Kluz M, Kordiaka R, Kunová S, Haščík P. Rapid Identification of Enterobacteriaceae in Milk and Dairy Products with the Matrix-Assisted Laser Desorption Ionization–Time of Flight Mass Spectrometry (MALDI-TOF MS). Animal Science and Biotechnologies. 2017; 50: 41-46.
11. Al S, Hızlısoy H, Onmaz NE, Karadal F, Barel M, Yıldırım Y, Gönülalan Z. 2020. A Molecular Investigation of Carbapenem Resistant Enterobacteriaceae and blaKPC, blaNDM and blaOXA-48 Genes in Raw Milk. Kafkas Üniversitesi veteriner fakültesi dergisi. 26(3): 391-396.
12. Laxminarayan R, Duse A, Wattal C, Zaidi AKM, Wertheim HFL, Sumpradit N. 2013. Antibiotic resistance-the need for global solutions. Lancet Infectious Dissease. 13(12): 1057-1098. 13. Bassetti M, Pecori D, Sibani M, Corcione S, Rosa FG. 2015. Epidemiology and Treatment of MDR Enterobacteriaceae. Current Treatment Options in Infectious Disease. 7, 291-316.
14. Bali EB, Açık L, Sultan N. 2010. Phenotypic and molecular characterization of SHV, TEM, CTX-M and extended-spectrum β-lactamase produced by Escherichia coli, Acinobacter baumannii and Klebsiella isolates in a Turkish hospital. African Journal of Microbiology Research. 4(8): 650-654.
15. Schultsz C, Geerlings S. 2012. Plasmid-mediated resistance in Enterobacteriaceae: changing landscape and implications for therapy. Drugs. 72(1): 1-16.
16. Vitas AI, Naik D, Pérez-Etayo L, González D. 2018. Increased exposure to extended-spectrum β-lactamase-producing multidrugresistant Enterobacteriaceae through the consumption of chicken and sushi Products. International Journal of Food Microbiology. 269: 80–86.
17. Martínez LR, Jiméneza LL, Fustéa E, Vinuesaa T, Martínezb JP, Vinaas M. 2011. Class 1 integrons in environmental and clinical isolates of Pseudomonas aeruginosa. International Journal of Antimicrobial Agents. 38(5): 398–402.
18. Hanau BB, Podglajen I, Casin I, Collatz E. 2002. An intrinsic control element for translational initiation in class 1 integrons. Molecular Microbiology. 44(1): 119-130.
19. Tekiner IH, Özpinar H. 2016. Occurrence and characteristics of extended spectrum beta-lactamases-producing Enterobacteriaceae from foods of animal origin. Brazilian Journal of Microbiology. 47(2): 444–451.
20. Clinical and Laboratory Standards Institute (CLSI). Performance Standards for Antimicrobial Susceptibility Testing. Twentieth Informational Supplement. 2016; M100-S20, 30, 1 USA.
21. Hijazi SM, Fawzi MA, Ali FM, El Galil KHA. 2016. Prevalence and characterization of extended‑spectrum beta‑lactamases producing Enterobacteriaceae in healthy children and associated risk factors. Annals of Clinical Microbiology and Antimicrobials. 15(1): 1-9.
22. Bass L, Liebert CA, Lee MD, Summers AO, White DG, Thayer SG, Maurer JJ. 1999. Incidence and characterization of integrons, genetic elements mediating multiple-drug resistance, in Avian Escherichia coli. Antimicrob Agents Chemother. 43(12): 2925-2929.
23. Estaji M, Tabasi M, Sadeghpour HF, Khezerloo JK, Radmanesh A, Raheb J, Ghadirzadeh MR, Tabatabaei A. 2019. Genotypic Identification of Pseudomonas aeruginosa Strains Isolated from Patients with Urinary Tract Infection. Comparative ımmunology microbiology & Infectious Disease. 65: 1-20.
24. El Zubeir IEM, Ahmed MIA. 2007. The Hygienic Quality of Raw Milk Produced by Some Dairy Farms in Khartoum State, Sudan. Research Journal of Microbiology. 2(12): 988-991.
25. El-Diasty EM, El-Kaseh RM. 2009. Microbiological monitoring of raw milk and yoghurt samples collected from El-Beida city. Arab Journal of Biotechnology. 12(1): 57-64.
26. Kar D, Bandyopadhyay S, Bhattacharyya D, Samanta I, Mahanti A, Nanda PK, Mondal B, Dandapat P, Das AK, Dutta TK, Bandyopadhyay S, Singh RK. 2015. Molecular and phylogenetic characterization of multidrug resistant extended spectrum beta-lactamase producing Escherichia coli isolated from poultry and cattle in Odisha, India. Infection, Genetics and Evolution. 29: 82-90.
27. Casella T, Rodríguez MM, Takahashi JT, Ghiglione B, Dropa M, Assunção E, Nogueira ML, Lincopan N, Gutkind G, Noguerira MCL. 2015. Detection of blaCTX-M-type genes in complex class 1 integrons carried by Enterobacteriaceae isolated from retail chicken meat in Brazil. International Journal of Food Microbiology. 16(197): 88–91.
28. Otter JA, Natale A, Batra R, Auguet OT, Dyakova E, Goldenberg SD, Edgeworth JD. 2019. Individual- and community-level risk factors for ESBL Enterobacteriaceae colonization identified by universal admission screening in London. Clinical Microbiology and Infection. 25(10): 1259-1265.
29. Steward AG, Harris PNA, Henderson A, Schembri MA, Paterson DL. 2020. Oral cephalosporin and β-lactamase inhibitor combinations for ESBL-producing Enterobacteriaceae urinary tract infections. Journal of Antimicrobial Chemotherapy. 75(9): 2384–2393
30. Mishra M, Panda S, Barik S, Sarkar S, Singh DV, Mohapatra H. 2020. Antibiotic Resistance Profile, Outer Membrane Proteins, Virulence Factors and Genome Sequence Analysis Reveal Clinical Isolates of Enterobacter Are Potential Pathogens Compared to Environmental Isolates. Frontiers in Cellular and Infection Microbiology. 10(54): 54-67.
31. Dib AL, Agabou A, Chahed A, Kurekci C, Moreno E, Espigares M, Espigares E. 2018. Isolation, molecular characterization and antimicrobial resistance of enterobacteriaceae isolated from fish and seafood. Food Control. 88: 54-60.
32. Faife SL, Zimba T, Sekyere JO, Govinden U, Chenia HY, Simonsen GS, Sundsfjord A, Essack SY. 2020. β-lactam and fluoroquinolone resistance in Enterobacteriaceae from imported and locally produced chicken in Mozambique. The Journal of Infection in Developing Countries. 14(5): 471-478.
33. Seedy FRE, Samy AA, Khairy EA, Koraney AA, Salam HHS. 2020. Molecular Determinants of Multiple Antibiotic Resistant E. coli Isolated from Food of Animal Origin. Advances in Animal and Veterinary Sciences. 8(4): 347-353.
34. Dixit S, Gupta S. 2019. Antıbıogram Of Mıcrobes Assocıated Wıth Enterıc Infectıon Targetıng Incıdences of Multıdrug Resıstance. Internatonal Knowledge Press. 12, 41-45.
35. Amador P, Fernandes R, Prudêncio C, Duarte, I. 2019. Prevalence of Antibiotic Resistance Genes in Multidrug-Resistant Enterobacteriaceae on Portuguese Livestock Manur., Antibiotics. 8(1): 1-18.
36. Davies J, Amabile CCF. 2003. The rise of antibiotic resistance. In: Amabile-Cuevas CF, ed. Multiple Drug Resistant Bacteria. Wiltshire: Horizon Scientific Press, 1: 1–7.
37. Çolakoğlun F, Özgümüş OB, Sandal C, Sevim EÇ, Karaoğlu SA. 2010. Class 1 and Class 2 Integron Gene Cassettes and Characterization of Antibiotic Resistance in Coliforms of Sea Water Origin. Türk Mikrobiyoloji Cemiyeti Dergisi. 40(2): 97-108.
38. Leverstein van Hall MA, Box ATA, Blok HEM, Paauw A Fluit AC, Verhoef J. 2001. Evidence of extensive interspecies transfer of integron-mediated antimicrobial resistance genes among multidrug resistant Enterobacteriaceae in a clinical setting. Journal of Infectious Diseas. 186(1): 49-56.
39. Thongkao K, Sudjaroen Y. Beta-lactamase and integron-associated antibiotic resistance genes of Klebsiella pneumoniae isolated from Tilapia fishes (Oreochromis niloticus). Journal of Applied Pharmaceutical Science. 9(1): 125-130.
40. Özgümüş OB, Tosun AF, Kılıç AO, Ertürk M. 2006. Carriage of mobilizable plasmid-mediated β-lactamase gene in ampicillin-resistant Escherichia coli strains with origin of normal fecal flora. Turkish Journal of Medical Sciences. 36(5): 307–14.
41. Pehlivanoğlu F, Turutoğlu H, Öztürk D, Yardımcı H. 2016. Molecular Characterızatıon of Esbl-Producıng Escherıchıa colı Isolated From Healthy Cattle And Sheep. Acta Veterinaria-Beograd. 66(4), 520-533.
42. Shahid M, Singh A, Sobia F, Rashid M, Malik A, Shukla I, Khan HM. 2011. blaCTX-M, blaTEM, and blaSHV in Enterobacteriaceae from North-Indian tertiary hospital: high occurrence of combination genes. Asian Pacific Journal of Tropical Medicine. 4(2): 101-105.