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Objectives Emergence of carbapenem-resistant Pseudomonas aeruginosa is limiting current treatment options. Carbapenemases and their association with integrons can cause rapid dissemination of resistance traits. We report here the co-existence and chromosomal inheritance of all four classes of β-lactamase and the presence of a unique class 1 integron (intI1) harbouring blaVIM-5 within a single isolate of P. aeruginosa, DMC-27b. Methods DMC-27b, isolated from urine, was characterized for carbapenem resistance both phenotypically and genotypically. The orientation of gene cassette structures of class 1 integrons was determined using referenced and designed overlapping primers and complete genome sequence (CGS) data. The antimicrobial resistance profile, porin protein mutations and the presence of active efflux activity were studied from the CGS. Results P. aeruginosa DMC-27b was resistant to a total of 20 antibiotics, with imipenem and meropenem MIC90s of >512 mg/L. The isolate harboured all four classes of β-lactamase: VEB-1 (class A), VIM-5 (class B), PDC-35 (class C) and OXA-2 and OXA-50 (both class D). Chromosomal harbouring of blaVIM-5 was associated with the intI1 gene cassette as the sole gene, a unique cassette so far reported. A total of 11 mutations, among them some mutations causing extra folds and changes in binding sites, in porin protein OprD might also affect its functionality regarding the transportation of antibiotics. Conclusions This is one of the earliest reports of its kind on the co-existence of all four β-lactamase classes in P. aeruginosa DMC-27b. Acquisition of multiple resistance determinants is paving the way for the development of MDR. This superbug is a model for rapid dissemination of resistance traits both horizontally and vertically.

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ObjectivesPseudomonas aeruginosa is a key opportunistic pathogen causing a wide range of community- and hospital-acquired infections in immunocompromised or catheterized patients. Here, we report the complete genome sequence of a multidrug resistant (MDR) P. aeruginosa DMC30b in order to elucidate the genetic diversity, molecular epidemiology, and underlying mechanisms for antimicrobial resistance and virulence.MethodsP. aeruginosa DMC30b was isolated from septic wound swab of a severe burn patient. Whole-genome sequencing (WGS) was performed under Ion Torrent platform. The genome was annotated using the SPAdes v. 3.12.01 in an integrated Genome Analysis Platform (IonGAP) for Ion Torrent sequence data. The genome was annotated using the NCBI Prokaryotic Genome Annotation Pipeline (PGAP). In-silico predictions of antimicrobial resistance genes (ARGs), virulence factor genes (VFGs) and metabolic functional potentials were performed using different curated bioinformatics tools.ResultsP. aeruginosa DMC30b was classified as MDR and belongs to sequence type 244 (ST244). The complete genome size is 6,994,756 bp with a coverage of 76.76x, G+C content of 65.7% and a BUSCO (Benchmarking Universal Single-Copy Orthologs) score of 100. The genome of P. aeruginosa DMC30b harboured two plasmids (e,g., IncP-6 plasmid p10265-KPC; 78,007 bp and ColRNAI_pkOIISD1; 9,359 bp), 35 resistomes (ARGs) conferring resistance to 18 different antibiotics (including four beta-lactam classes), and 214 VFGs. It was identified as the 167th ST244 strain among ∼ 5,800 whole-genome sequences of P. aeruginosa available in the NCBI database.ConclusionP. aeruginosa DMC30b belongs to ST244 and was identified as the 167th such isolate to be submitted to NCBI, and the first complete ST244 genome from Bangladesh. The complete genome data with high genetic diversity and underlying mechanisms for antimicrobial resistance and virulence of P. aeruginosa DMC30b (ST244) will aid in understanding the evolution and phylogeny of such high-risk clones and provide a solid basis for further research on MDR or extensively drug resistant strains.

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