Aims: The high prevalence of multidrug resistance (MDR) and extended-spectrum β-lactamase (ESBL)-producing Klebsiella pneumoniae associated with nosocomial infections has caused serious therapeutic challenges. The objectives of this study were to determine the genotypic and phenotypic characteristics of K. pneumoniae strains isolated from Malaysian swine farms and the transferability of ESBL genes by plasmids. Results: A total of 50 K. pneumoniae strains were isolated from 389 samples, which were collected from healthy and unhealthy pigs (swine rectum and oral cavities), healthy farmers (human rectum, urine, and nasal cavities), farm's environment, and animal feeds from seven Malaysian swine farms. Antimicrobial susceptibility analysis of these 50 K. pneumoniae strains showed that the majority (86%) were resistant to tetracycline, while 44% and 36% of these strains were MDR and ESBL producers, respectively. PCR and DNA sequencing of the amplicons showed the occurrence of blaTEM (15/18), blaSHV (15/18), blaCTX-M-1 group (7/18), and blaCTX-M-2 group (2/18), while only class 1 integron-encoded integrase was detected. Conjugation experiments and plasmid analysis indicated that the majority of the ESBL genes were plasmid encoded and the plasmids in 11 strains were conjugative. Genotyping by pulsed-field gel electrophoresis and repetitive extragenic palindrome-polymerase chain reaction (REP-PCR) showed that these 50 strains were genetically diverse with 44 pulsotypes and 43 REP-PCR subtypes. Conclusions: ESBLproducing K. pneumoniae strains showed high resistance to tetracycline as this antibiotic is used for prophylaxis and therapeutic purposes at the swine farms. The findings in this study have drawn attention to the issue of increasing MDR in animal husbandry and it should be taken seriously to prevent the spread and treatment failure due to antimicrobial resistance.
Multidrug-resistant (MDR) and extended-spectrum β-lactamase (ESBL)-producing Klebsiella pneumoniae associated with nosocomial infections have caused serious problems in antibiotic management with limited therapeutic choices. This study aimed to determine the genotypic and phenotypic characteristics of K. pneumoniae strains isolated from a tertiary hospital in Malaysia. Ninety-seven clinical K. pneumoniae strains were analyzed for antimicrobial susceptibility, all of which were sensitive to amikacin and colistin (except one strain), while 31.9 % and 27.8 % were MDR and ESBL producers, respectively. PCR and DNA sequencing of the amplicons indicated that the majority of MDR strains (26/27) were positive for bla TEM, followed by bla SHV (24/27), bla CTX-M-1 group (23/27), bla CTX-M-9 group (2/27), and mcr-1 (1/27). Thirty-seven strains were hypervirulent and PCR detection of virulence genes showed 38.1 %, 22.7 %, and 16.5 % of the strains were positive for K1, wabG, and uge genes, respectively. Genotyping by pulsed-field gel electrophoresis (PFGE) and multilocus sequence typing (MLST) showed that these strains were genetically diverse and heterogeneous. Sequence types, ST23, ST22, and ST412 were the predominant genotypes. This is the first report of colistin-resistant K. pneumoniae among clinical strains associated with mcr-1 plasmid in Malaysia. The findings in this study have contributed to the effort in combating the increase in antimicrobial resistance by providing better understanding of genotypic characteristics and resistance mechanisms of the organisms.
Multidrug-resistant (MDR) Klebsiella pneumoniae (K. pneumoniae) poses a serious public health threat. K. pneumoniae strains that produce extended-spectrum beta-lactamases (ESBL) are becoming increasingly reported in nosocomial and community-acquired infections. Besides resistance genes, integrons, and plasmids, altered membrane permeability caused by porin loss and energy-dependent efflux have also contributed to antibiotic resistance in K. pneumoniae. The objective of this study was to determine the correlation between the reduction of antibiotic susceptibility and overexpression of efflux pump as well as the lack of outer membrane proteins (OMPs) among clinical ESBLs resistant K. pneumoniae. The expression levels of ramA, acrA, ompK35 and ompK36 in 12 MDR K. pneumoniae strains with varying MICs levels were analyzed using quantitative real time-Polymerase Chain Reaction (qRT-PCR). The role of efflux pump on antibiotic resistance was also studied by using minimum inhibitory concentration (MICs) method with//without efflux pump inhibitor. The result indicated that the strains with highest resistance to cefotaxime showed the lowest level of ompK35 and ompK36 genes expression while the strains with lowest MIC level of resistance to cefotaxime showed the highest level of expression of acrA and ramA. Our finding also revealed the effect of efflux pump inhibitor phenyl-arginine-b-naphthylamide (PAβN) on the MIC levels of ceftazidime, amoxicillin-clavulanate and cefotaxime which were significantly reduced around 1–7 folds MIC levels. These results suggest that Efflux pump system and deficiently of OMPs contributing role in antibiotic susceptibility which should be taken seriously to prevent the treatment failure due to antimicrobial resistance.
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