Detection of extended spectrum β-lactamase (ESBLA), screening of AmpC β-lactamase and detection of CTX-M and aacA-aphD genes among the multidrug resistant bacteria found in two tertiary hospitals of Dhaka city

Abstract

Klebsiella spp. and Pseudomonas spp. are opportunistic pathogens that are responsible for severe infections. These infections can be treated by antibiotics of different classes individually or in combination. However, lately, these pathogens have become resistant to antibiotics through its increased exposure or conjugal transfer of antibiotic resistant genes, making treatment difficult. Production of β-lactamases like extended spectrum β-lactamases (ESBL) or AmpC β-lactamases by pathogens could be one of the reasons for the occurrence of multidrug resistant (MDR) bacteria. The objective of this study was to molecularly detect the presence of CTX-M and aacA-aphD genes in the samples responsible for multidrug resistance. Isolates of sputum and pus origin were collected from two tertiary hospitals and antimicrobial susceptibility test was performed to detect the multidrug resistant strains. The ESBLA producers were detected by the double disk synergy test. The AmpC β-lactamase producers were also screened simultaneously by checking its susceptibility to cefoxitin. Polymerase chain reaction was then performed to detect CTX-M gene for the samples resistant to both third-generation cephalosporins and monobactams and to detect aacA-aphD for the samples resistant to aminoglycosides. All the samples were observed to be completely resistant to cloxacillin (100%) followed by kanamycin (61.5%) whereas, the samples showed least resistance to imipenem (7.7%). Of the total samples, 76.9% were screened to be AmpC β-lactamase positive and were the most prevalent (96%) among Pseudomonas spp. However, 65.4% of the samples were phenotypically detected to be ESBLA positive and were the most prevalent among Klebsiella spp. (74.1%). When PCR was carried out, 2 out of 28 samples, that were found to be resistant to both third-generation cephalosporins and monobactams, were confirmed to have the CTX-M gene of size 800bp, whereas none of the samples, that were resistant to aminoglycosides, were found positive for aacA-aphD gene. From the results, it can be concluded that the percentage of ESBLA and AmpC β-lactamase producers were high among pathogens found in the tertiary hospitals. Hence, ESBL and AmpC β-lactamase detection methods of high sensitivity and specificity, as well as its molecular detection, should be made compulsory so that appropriate antibiotics can be used for the treatment of infection.