Transcriptional expression of PmrB and arnA within polymyxin-resistant nosocomial isolates of Pseudomonas aeruginosa from India

Year 2018, , 61 - 68, 20.06.2018

Sayani Roy Deepjyoti Paul Rajkumari Elizabeth Debadatta Dhar Atanu Chakarvarty Amitabha Bhattacharjee

https://doi.org/10.5799/jmid.434595

Abstract

ABSTRACT

Objective: The polymyxin group of antibiotics is considered to be
one of the most effective antimicrobial agents against many serious pathogenic
bacteria, but the excessive use of these antibiotics has led to the development
of drug resistance among bacteria. This study was designed to characterize polymyxin-resistant
P. aeruginosa and to explore the role
of PmrB and arnA in resistant phenotype.

Methods:
mRNA and cDNA of five selected polymyxin-resistant strains representing
different MIC range; isolated under normal condition of  strain growth, after treating sample/media
with FeCl3 and MgCl2 alone, or after treating with FeCl3 and polymyxin
antibiotic. The transcriptional expression was observed for PmrB and arnA by quantitative real time RT-PCR in reference to P. aeruginosa PAO1. The presence of
plasmid mediated colistin resistance determinants mcr-1 was screened by PCR. Susceptibility of the strains was
determined by disc-diffusion method and DNA fingerprinting was carried out by
performing REP-PCR.

Results: A
down regulated expression of PmrB and
arnA was observed even after the
unique induction with FeCl3 and MgCl2. All the isolates were found to be
resistant against cefepime and different clonal patterns of resistance were
found among the isolates.

Conclusion: This study has drawn a new insight into polymyxin
resistance which will help in the detection and control of infections caused by
multidrug resistant P. aeruginosa.
The low susceptibility rate to aminoglycoside, piperacllin-tazobactam and
ciprofloxacin was found and in addition, detection of PmrB and arnA as
molecular markers in the follow up of infections caused by multidrug resistant P. aeruginosa. J Microbiol Infect Dis 2018; 8(2):61-68.

Keywords

arnA, mcr-1, PmrB, Polymyxin, Pseudomonas aeruginosa, Multidrug resistance

References

• REFERENCES 1) Gupta S, Govil D, Kakar PN, et al. Colistin and polymyxin B: A re-emergence. Indian J Crit Care Med 2009; 13: 49-53. 2) Vaara M. Polymyxins and their novel derivatives. Curr Opin Microbiol 2010; 13: 574-581. 3) Biswas S, Brunel JM, Dubus JC, Reynaud-Gaubert M, Rolain JM. Colistin: An update on the antibiotic of the 21st century. Expert Rev Anti Infect Ther 2012;10:917-934. 4) Michalopoulos A, Falagas ME. Colistin and Polymyxin B in critical Care. Crit Care Clin 2008; 24: 377-391. 5) Balaji V, Jeremiah SS, Baliga PR. Polymyxins: Antimicrobial susceptibility concerns and therapeutic options. Indian J Med Microbiol 2011; 29:230-242. 6) Falagas ME, Michalopoulos A. Polymyxins: old antibiotics are back. Lancet 2006; 367:633-634. 7) Falagas ME, Rfailidis PI, Matthaiou DK. Resistance to Polymyxins: Mechanisms, frequency and treatment options. Drug Resist Updat 2010; 13:132-138. 8) Gupta V, Bansal N, Palta S, Gombar S, Chaner J. Comparative in vitro activity of polymyxins against carbapenem susceptible and resistant non-fermenters from critically ill-patients. Indian J Med Microbiol 2014;32:203-204. 9) Fernandez L, Gooderham WJ, Bains M, McPhee JB, Wiegand I, Hancock REW. Adaptive Resistance to the “Last Hope” antibiotics polymyxin B and Colistin in Pseudomonas aeruginosa is mediated by the novel two-component regulatory system ParR-ParS. Antimicrob Agents Chemother 2010; 54: 3372-82. 10) Paul D, Dhar D, Maurya AP, et al. Occurrence of co‑existing blaVIM‑2 and blaNDM‑1 in clinical isolates of Pseudomonas aeruginosa from India. Ann Clin Microbiol Antimicrob (2016) 15:31.doi 10.1186/s12941-016-0146-0. 11) Moskowitz SM, Brannon MK, Dasgupta N, et al. PmrB Mutations Promote Polymyxin Resistance of Pseudomonas aeruginosa Isolated from Colistin-Treated Cystic Fibrosis Patients. Antimicrob Agents Chemother 2012; 56:1019-30. 12) Deng D, Phan M, Sharma A, Sharma M. RpoS and PmrA induced resistance to polymyxin B is independent of arnT expression in Pseudomonas aeruginosa. J Exp Microbiol Immun 2013; 17: 55-59. 13) Liu YY, Wang Y, Walsh TR, 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. 14) Colee JG, Diguid JP, Fraser AG. Mackie and McCartney Practical Medical Microbiology. 1996; 14th ed, Edinburgh: Churchill, Livingstone. 15) Clinical Laboratory Standard Institute. Performance Standards for Antimicrobial Susceptibility Testing; Twenty-Third Informational Supplement. 2013; M100-S23. CLSI, Wayne, USA. 16) Elnahriry SS, Khalifa HO, Soliman AM, et al. Emergence of Plasmid-Mediated Colistin Resistance Gene mcr-1 in a Clinical Escherichia coli Isolate from Egypt. Antimicrob Agents Chemother 2016; 60:3249-3250. 17) Paul D, Bhattacharjee A, Bhattacharjee D, Dhar D, Maurya AP, Chakravarty A. Transcriptional analysis of blaNDM-1 and copy number alteration under carbapenem stress. Antimicrob Resist Infect Control 2017; 6, 26. 18) Mesaros N,Glupczynski Y, Avrain L, Caceres NE, Tulkens PM, Bambeke FV. A combined phenotypic and genotypic method for the detection of Mex efflux pumps in Pseudomonas aeruginosa. J Antimicrob Chemother 2007; 59:378-386. 19) Versalovic J, Koeuth T, Lupski JR. Distribution of repetitve DNA sequences in eubacteria and application to fingerprinting of bacterial genomes. Nucleic Acids Res1991; 19:6823-6831. 20) Paul D, Maurya AP, Chanda DD, Sharma GD, Chakravarty A, Bhattacharjee A. Carriage of blaNDM-1 in Pseudomonas aeruginosa through multiple Inc type plasmids in a tertiary referral hospital of northeast India. Indian J Med Res 2016; 143: 826-829. 21) Suresh KP, Chandrashekara S. Sample size estimation and power analysis for clinical research studies. J Hum Reprod Sci 2012; 5:7-13. 22) Grewal US, Bakshi R, Walia G, Shah PR. Antibiotic susceptibility profiles of non-fermenting Gram-negative bacilli at a tertiary care hospital in Patiala, India. Niger Postgrad Med J 2017; 24: 121-125. 23) Adams MD, Nickel GC, Bajaksouzian S, et al. Resistance to colistin in Acinetobacterbaumanii associated with mutations in the PmrAB two component systems. Antimicrob Agents Chemother 2009;53: 3628-3634. 24) Zavascki AP, Goldani LZ, Natiion RL. Polymyxin-B for the treatment of multidrug-resistant pathogens: a critical review. J Antimicrob Chemother 2007; 60:1206-15.