Article history: Received: 07-12-2017, Accepted: 12-03-2018, Published online: 13-04-2018
Corresponding author: B. Joseph
E-mail: firstname.lastname@example.orgCitation: Rathore K, Joseph B, Sharma DK, Gaurav A, Sharma SK, Milind M, Patel P, Prakash C, Singh L (2018) Evaluation of multiplex polymerase chain reaction as an alternative to conventional antibiotic sensitivity test, Veterinary World, 11(4): 474-479.
Aim: This study was designed to evaluate the potential of the use of multiplex polymerase chain reaction (PCR) as an alternative to conventional antibiotic sensitivity test.
Materials and Methods: Isolates of Staphylococcus aureus (total = 36) from clinical cases presented to Teaching Veterinary Clinical Complex of College of Veterinary and Animal Sciences (CVAS), Navania, Udaipur, were characterized by morphological, cultural, and biochemical methods. Then, the isolates were further subjected to molecular characterization by PCR targeting S. aureus-specific sequence (107 bp). Phenotypic antibiotic sensitivity pattern was analyzed by Kirby Bauer disc diffusion method against 11 commonly used antibiotics in veterinary medicine in and around Udaipur region. The genotypic antibiotic sensitivity pattern was studied against methicillin, aminoglycosides, and tetracycline targeting the gene mecA, aacA-aphD, and tetK by multiplex PCR.
Results: There was 100% correlation between the phenotype and genotype of aminoglycoside resistance, more than 90% correlation for methicillin resistance, and 58.3% in the case tetracycline resistance.
Conclusion: As there is a good correlation between phenotype and genotype of antibiotic resistance, multiplex PCR can be used as an alternative to the conventional antibiotic susceptibility testing, as it can give a rapid and true prediction of antibiotic sensitivity pattern.
Keywords: antimicrobial resistance, genotype, phenotype, Staphylococcus aureus.
1. Carlet, J., Jarlier, V., Harbarth, S., Vos, A., Goossens, H. and Pittet, D. (2012) Ready for a world without antibiotics? The pensieres antibiotic resistance call to action. Antimicrob. Resist. Infect. Control, 1: 11. [Crossref] [PubMed] [PMC]
2. Helmuth, R. (2000) In: Wary, C. and Wary, A., editors. Antibiotic Resistance in Salmonella in Domestic Animals. CABI Publishing, Oxon, United Kingdom. p89-106. [Crossref]
3. Ray, B. (2004) Foodborne Infections: Fundamental Food Microbiology. 3rd ed. CRC Press, Washington DC.
4. Salem-Bhekhit, M.M. (2014) Phenotypic and Genotypic characterization of nosocomial isolates of Staphylococcus aureus with reference to methicillin resistance. Trop. J. Pharm. Res., 13(8): 1239-1246. [Crossref]
5. Mohanasoundaram, K. and Lalitha, M. (2008) Comparison of phenotypic versus genotypic methods in the detection of methicillin resistance in Staphylococcus aureus. Ind. J. Med. Res., 127: 78-84. [PubMed]
6. Evans, S.R., Hujer, A.M., Jiang, H., Hill, C.B., Hujer, K.M., Mediavilla, J.R., Manca, C., Tran, T.T.T., Domitrovic, T.N., Higgins, P.G., Seifert, H., Kreiswirth, B.N., Patel, R., Jacobs, M.R., Chen, L., Sampath, R., Hall, T., Marzan, C., Fowler, V.G. Jr., Chambers, H.F. and Bonomo, R.A. (2017) Informing antibiotic treatment decisions: Evaluating rapid molecular diagnostics to identify susceptibility and resistance to carbapenems against Acinetobacter spp. in PRIMERS III. J. Clin. Microbiol., 55: 134-144. [Crossref]
7. Jamal, W., Roomi, E.A., Abdul-Aziz, L.R. and Rotimi, V.O. (2014) Evaluation of curetis unyvero, a multiplex PCR based testing system for rapid detection of bacteria and antibiotic resistance and impact of the assay on management of severe nosocomial pneumonia. J. Clin. Microbiol., 52: 2487-2492. [Crossref]
8. McClure J., DeLongchamp, J.Z., Conly, J.M. and Zhang, K. (2017) Novel multiplex PCR assay for detection of chlorhexidine-quarternary ammonium, mupirocin and methicillin resistance genes with simultaneous discrimination of Staphylococcus aureus from coagulase-negative staphylococci. J. Clin. Microbiol., 55: 1857-1864. [Crossref] [PubMed] [PMC]
9. Stormmenger, B., Kettlitz, C., Werner, G. and Witte, W. (2003) Multiplex PCR assay for simultaneous detection of nine clinically relevant antibiotic resistance genes in Staphylococcus aureus. J. Clin. Microbiol., 41: 4089-4094. [Crossref] [PMC]
10. Vannuffel, P., Gigi, J., Ezzedine, H., Vandercam, B., Delmee, M., Wauters, G. and Gala, J. (1995) Specific detection of methicillin-resistant Staphylococcus species by multiplex PCR. J. Clin. Microbiol., 33: 2864-2867. [PubMed] [PMC]
11. Quinn, P.J., Carter, M.E., Markey, B.K. and Carter, G.R. (1994) Clinical Veterinary Microbiology. Wolfe Publishing, Mosby-Year Book Europe Lynton House, Tavistock Square, London WCH 9LB, England. p7-12.
12. Wilson, K. (1987) In: Ausubal, F.M., Brent, R., Kirston, R.L., Moore, D.D., Seidman, J.G., Smith, J.A. and Struhe, K., editors. Preparation of genomic DNA from bacteria. In: Current Protocols in Molecular Biology. Vol. 1. John Wiley and Sons, New York. p241-242.
14. Maple, P.A., Hamilton-Miller, J.M. and Brumfitt, W. (1989) Worldwide antibiotic resistance in methicillin-resistant Staphylococcus aureus. Lancet, 1: 537-540. [Crossref]
15. Verma, S., Joshi, S., Chitnis, V., Hemwani, N. and Chitnis, D. (2000) Growing problem of methicillin-resistant staphylococci: Indian scenario. Indian J. Med. Sci., 54: 535-540. [PubMed]
16. Vidhani, S., Mehndiratta, P.L. and Mathur, M.D. (2001) Study of MRSA isolates from high-risk patients. Indian J. Med. Microbiol., 19: 87-90.
17. Anupurba, S., Sen, M.R., Nath, G., Sharma, B.M., Gulati, A.K. and Mohapatra, T.M. (2003) Prevalence of methicillin-resistant Staphylococcus aureus in a tertiary care referral hospital in eastern Uttar Pradesh. Indian J. Med. Microbiol., 21: 49-51. [PubMed]
18. Saikia, L., Nath, R., Choudhury, B. and Sarkar, M. (2009) Prevalence and antimicrobial susceptibility pattern of methicillin-resistant Staphylococcus aureus in Assam. Indian J. Crit. Care Med., 13: 156-158. [Crossref] [PubMed] [PMC]
19. Torres, G.M., Tejedor, J.M.T., Gonzalez, M.M. and Gonzalez, L.Z. (1996) Selection of subpopulations resistant to amikacin and netilmicin of gentamicin-resistant clinical strains of Staphylococcus aureus and Staphylococcus epidermidis. Zentbl. Bakteriol., 284: 58-66. [Crossref]
20. Roschanski, N., Fischer, J., Guerra, B. and Roesier, U. (2014) Development of a multiplex real-time PCR for the rapid detection of the predominant beta-lactamase genes CTX-M, SHV, TEM and CIT-type AmpCs in Enterobacteriaceae. PLoS One, 9: e100956-e100956. [Crossref]21. Chavda, K.D., Satlin, M.J., Chen, L., Manca, C., Jemkins, S.G., Walsh, T.J., Kreiswirth, B.N. (2016) Evaluation of multiplex PCR assay to rapidly detect Enterobacteriaceae with a broad range of beta-lactamase directly from perianal swabs. Antimicrob. Agents Chemother., 60: 6957-6961. [Crossref] [PubMed] [PMC]