Conjugative transposons in Gram-negative bacteria have a significant role in the dissemination of antibiotic-resistance-conferring genes between bacteria. This study aims to genomically characterize plasmids and conjugative transposons carrying integrons in clinical isolates of Klebsiella pneumoniae. Genetic composition of conjugative transposons and phenotypic assessment of 50 multidrug-resistant K. pneumoniae isolates from Tertiary-care Hospital (SQUH), Muscat, Oman was investigated. Horizontal transferability was investigated by filter- mating conjugation experiments. Whole genome sequencing (WGS) was performed to determine the sequence type (ST), acquired resistome and plasmidome of integron-carrying strains. Class 1 integron was detected in 96% of isolates and among integron-positive isolates, 18 stains contained variable regions. Horizontal transferability by conjugation confirmed the successful transfer of integrons between cells and WGS confirmed their presence in conjugative plasmids. Dihydrofolate reductase (dfrA14) was the most prevalent (34.8%) gene cassettes in class 1 integrons. MLST analysis detected predominantly ST-231 and ST-395. BlaOXA-232 and blaCTX-M-15 were the most frequently detected carbapenems and beta-lactamases in sequenced isolates. This study highlighted the high transmissibility of MDR-conferring conjugative plasmids in clinical isolates of K. pneumoniae. Therefore, the wise use of antibiotics and the adherence to effective infection control measures are necessary to limit further dissemination of multidrug-resistant organisms.

Hypervirulent Klebsiella pneumoniae (hvKp) is a variant that has been increasingly linked to severe, life threatening infections including pyogenic liver abscess and bloodstream infections. HvKp belonging to the capsular serotypes K1 and K2 have been reported worldwide, however very scarce studies are available on the their genomics and virulence. In the current study we report four hypermucoviscous hvKp ESBL clinical strains of capsular serotype K1 and K2 isolated from pus and urine of critically ill patients in tertiary care hospitals in Oman. These strains belong to diverse sequence types (STs) namely ST-23(K1), ST-231(K2), ST-881(K2), and ST-14(K2). To study their virulence, Galleria Mellonella model and resistance to human serum killing were used. The G. mellonella model revealed that the K1/ST-23 isolate is the most virulent as 50% of the larvae died in the first day, followed by isolate K2/ST-231 for which 75% of the larvae died in the second day and isolate K2/ST-14 for which 50% of the larvae died in the second day. Resistance to human serum killing showed there was complete inhibition of bacterial growth of all four isolates by the end of the first hour and up to the third hour. For ST-23(K1), ST-231(K2), ST-881(K2), and ST-14(K2), there was a gradual decrease in the number of colonies up to the second to third hour, which indicates neutralization of bacterial cells by the serum components. However, this was followed by sudden increase of bacterial growth, indicating possible resistance of bacteria against human serum bactericidal activity. Whole genome sequencing (WGS) revealed that hvKp strains display unique genetic arrangement of k-loci. Whole genome SNP-based phylogenetic analysis revealed that these hvKp isolates are phylogenetically distinct, belonging to diverse clades and belong to different STs in comparison to global isolates. This is the first report from Oman detailing the WGS of hvKp clinical isolates and assessing their resistance and virulence genomics, which reinforce our understanding of their epidemiology and dissemination in clinical settings.