Molecular Investigaton of Carbapenem Resistant Psevdomonas Aeruginosa Isolated from Tertiary care Hospital of Nepal

Abstract

Background: P. aeruginosa are categorized as second most critical group of pathogen by WHO. These opportunistic pathogens mainly affect patients with compromised host defense mechanisms. Infections caused by P. aeruginosa can be life threatening. Several resistance mechanisms make them able to resist multiple classes of antibiotics including β-lactams. Carbapenem are potent beta lactam antibiotics and last resort of drug. However, resistance to this drug also has been reported lately. Loss of OprD porin have significant role against carbapenem. Similarly, β – lactamase (MBL and ESBL) have potential to hydrolyze carbapenem of which gene bla NDM has great concern. In addition, AmpC beta lactamase with combination to other mechanism also showed resistance to carbapenem. Due to such mechanism attributed by P. aeruginosa against carbapenem it has become very difficult to combat them, leaving fewer options for antibiotic therapy. Aims: This study aims to provide knowledge on carbapenem resistance mechanism including loss of porin, overexpression of AmpC beta -lactamase and carbapenemase at molecular level. Methods: In this study, 95 isolates of P.aeruginosa were collected. Antimicrobial susceptibility profile based on the disk-diffusion tests was performed to detect multidrug resistance isolates. Carbapenem resistance isolates were selected and classified into IMPR, MRP R,, IMPR MRP R types. Phenotypic detection for MBL-producer, ESBL-producer and AmpC producer were performed. Molecular identification of carbapenem resistant gene of P. aeruginosa was carried out by PCR and followed by sequencing. Results and Conclusion: Among 95 isolates, total 73 (76.84%) were found to be Multidrug resistance P. aeruginosa and out of 73 MDR, 61 (83.56%) were carbapenem resistant isolates. Phenotypic test revealed that 55 (90.1%) MBL-producer and 45(61.64% )were ESBL producer according to the standard microbiological method CLSI. The PCR analysis result showed that most of the carbapenem resistant isolates were found to have (42.62%) ampC gene while (31.14%) were found to lack OprD gene. Among 61 carbapenem resistant isolates, 7 (11.47%) and 6 (9.8%) were found to have detection to bla NDM gene. The sequence of these genes showed mutation that could potentially lead to stronger carbapenemase activity. Our results confirmed that multiple resistance mechanism such as OprD loss, carbapenemase and AmpC beta lactamase production conferred resistance to the carbapenem in P. aeruginosa, isolated from hospital settings. Keywords: Antibiotics resistance, Carbapenem, P. aeruginosa, Outer membrane porins.