Prevalence and Serotype comparison of ESBL and Non-ESBL Klebsiella pneumoniae in Malaysian public hospitals

Introduction: Klebsiella pneumoniae is a common nosocomial pathogen that can be found in the hospital setting and is often implicated in hospital outbreaks. A more virulent strain of K. pneumoniae, known as hypervirulent K. pneumoniae (hvKp), is frequently found in K1 and K2 capsular serotypes. The emergence and spread of these strains are considered a major public health issue as it compromises the management and treatment of infections caused by this pathogen. This condition is further worsened by the presence of co-resistance in extended-spectrum β-lactamase (ESBL)-producing K. pneumoniae strains. Infections caused by hypervirulent ESBL-producing K. pneumoniae strains have been extensively reported to be associated with higher mortality rates, clinical complications, prolonged and costly hospitalizations due to limited treatment options. Objective: This study aims to determine the distribution and molecular characteristics of ESBL-producing K. pneumoniae clinical isolates in Hospital Sultanah Aminah (HSA), Johor Bahru and Hospital Pengajar Universiti Putra Malaysia (HPUPM), Serdang, Selangor. Methodology: A total of 192 K. pneumoniae isolates were collected from HSAJB and HPUPM. Antibiotic disc diffusion test was then carried out on all isolates to detect ESBL-producing phenotype among them. Subsequently, hypermucoviscosity test (string test) was performed to observe the formation of viscous string of hypervirulent K. pneumoniae strain with more than 5 mm in length. Multiplex PCR was then conducted to detect isolates possessing K1 and K2 capsule-associated genes, magA and K2A genes, respectively. Isolates found to be positive for ESBL phenotype and K1 or K2 serotypes were selected for DNA sequencing and multilocus sequence typing (MLST). Finally, phylogenetic analysis was performed to characterise the genetic linkage between the K1 and K2 serotypes among ESBL-producing isolates. Result: A total of 87 out of 192 (46%) ESBL-producing K. pneumoniae were detected in the clinical isolates collected from HSAJB and HPUPM. They showed a higher resistance rate towards aztreonam, ceftriaxone, ceftazidime and cefotaxime compared to non-ESBL producing isolates. Among the ESBL isolates, only 12 were positive for K1 (n= 4) and K2 (n= 8) capsular serotypes, respectively. Statistical analysis showed that K1 and K2 capsular serotypes were not significantly associated with ESBL phenotype (P= 0.196). However, they were significantly associated with hypermucoviscosity, as demonstrated by the positive string test (P<.001). MLST analysis identified seven different sequence types (ST) in this study, with ST23 predominantly associated with the K1 serotype. Meanwhile, the K2 serotype is widespread among different sequence types (ST14, ST65, ST86, ST628, ST657 and ST792). Phylogenetic analysis of K1 and K2 serotypes revealed two distinct clusters. Cluster A consists of all K1 serotypes, and cluster B consists of all K2 serotypes. Conclusion: A high prevalence of 44.2% and 50% of the K. pneumoniae isolates collected from HSAJB and HPUPM respectively, were ESBL producers. However, only 8.3% and 10.9% of the total isolates were detected to carry K1- and K2-serotype associated genes respectively. In the current study, seven different sequence types were identified by MLST analysis, with ST23 being predominantly associated with the K1 serotype. Meanwhile, the K2 serotype is widespread among ST14, ST65, ST86, ST628, ST657 and ST792.

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