Letter to the Editor

Emergence and spread of KPC-2-producing Pseudomonas aeruginosa isolates in a Brazilian teaching hospital☆

The high rates of carbapenem resistance among Brazilian Pseudomonas aeruginosa isolates are mainly associated with the clone ST277 producing the carbapenemase SPM-1. Here, the complete genetic composition of a IncP plasmid harboring bla_KPC-2 in isolates of this endemic clone carrying chromosomal bla_SPM-1 was described using whole genome sequencing. These results confirm the association of these two carbapenemases in ST277 and also describe the genetic composition of a novel bla_KPC-2-plasmid. Considering the fact that this association occurs in a high-risk clone, monitoring the dissemination of this plasmid should be a public health concern.

The Klebsiella pneumoniae carbapenemase (KPC) is disseminated worldwide mostly by plasmids. However, in Pseudomonas aeruginosa chromosomal mutations are more frequently responsible for resistance to carbapenems than the acquisition of mobile elements harbouring carbapenemases genes. Indeed, although uncommon, KPC-2-producing P. aeruginosa has appeared more frequently, including in Brazil. Here we report the first genomic analysis of a plasmid-mediated KPC-2 in an extensively drug-resistant (XDR) P. aeruginosa isolated in Santa Catarina, Brazil.

Antimicrobial susceptibility testing was performed according to CLSI 2020 guidelines. The genome was sequenced using an Illumina MiSeq platform and the data were analysed using SPAdes and Prokka. In silico predictions were fulfilled using curated bioinformatics tools.

Pseudomonas aeruginosa strain MIMA_PA2.1 (JACGTM000000000) was classified as XDR, belongs to sequence type 312 (ST312) and harbours the bla_KPC-2 gene located on a small (7975 bp) IncU plasmid. This plasmid showed 86.3% identity with a non-conjugative plasmid (KC609322) carrying the bla_KPC-2 gene from a multidrug-resistant P. aeruginosa (ST1006) from Colombia isolated in 2006. Besides the bla_KPC-2 gene, other resistance genes to β-lactams, aminoglycosides, phenicol, fosfomycin and quinolones were detected, the last two also associated with mobile genetic elements other than the IncU plasmid described here.

This is the first genomic report of the presence of the bla_KPC-2 gene carried by Pseudomonas in Southern Brazil and highlights the adaptability of bla_KPC-2 to different mobile elements. This draft genome might be useful for comparative genomic analyses to monitor the spread of plasmid-mediated bla_KPC in P. aeruginosa in Latin America.

Pseudomonas aeruginosa is a major health concern globally and treating infections caused by MDR-isolates unarguably a humongous challenge that remains an unmet need in modern medicine. To determine patterns and mechanisms of antimicrobial resistance and its spread over the years in Rio de Janeiro, Brazil, 88 P. aeruginosa isolates were selected from 1995 to 2015. Phenotypic and genotypic characterization of antimicrobial resistance was evaluated and isolates were submitted to clonality by PFGE and MLST. PFGE analysis showed a great variability of clonal groups mainly over the past 10 years of this study. STs predominant in the early years (ST804, ST1860, ST487 and ST1602) associated to multidrug resistance (MDR) phenotype were replaced by ST277, ST244, ST1945, ST1791 with extensive drug resistance (XDR) in last years, with significant increase in resistance to carbapenems, fluoroquinolones and aminoglycosides. Colistin resistance was detected in 3.5%. The main mechanisms of antimicrobial resistance were mutational mechanisms (mutations in oprD, mexT and gyrA genes). We found the ESBL genes bla_TEM (n = 2), bla_SHV (n = 3) and bla_CTX (n = 1).The carbapenemases genes was present in ST277 (bla_SPM, n = 3), ST1560 (bla_KPC, n = 3) and ST1944 (bla_KPC, n = 2). The 16S RNA methylase gene (rmtD) was found in five isolates belonged to ST277. In conclusion, molecular epidemiological investigation reveals an increase of antimicrobial resistance in P. aeruginosa over 21 years in Rio de Janeiro with higher population structure and occurrence of high risk clone in the last years. The mutational mechanisms of resistance were present in all XDR isolates.

Klebsiella pneumoniae carbapenemase (KPC) is the most widespread carbapenemase in Enterobacteriaceae in Brazil. Although its presence is not common in Pseudomonas aeruginosa, it has been increasingly reported. Here we report a draft genome sequence of a KPC-producing P. aeruginosa strain recovered from a bloodstream infection sample in Brazil.

The antimicrobial susceptibility of KPC-producing P. aeruginosa CCBH17348 was evaluated by the disk diffusion method, Etest and broth microdilution. Carbapenemase production was confirmed by colorimetric assay (Carba NP) and PCR. Genomic DNA was sequenced by Illumina MiSeq sequencing and was assembled using the A5-Miseq pipeline, and gene annotation was performed using RAST 2.0. The database ResFinder 2.1, CRISPRFinder and MLST website were used to identify resistance genes, clustered regularly interspaced short palindromic repeats (CRISPRs) and sequence types (STs), respectively.

Isolate CCBH17348 was considered multidrug-resistant, was susceptible to fluoroquinolones, gentamicin and polymyxin, and belonged to a newly described ST (ST2584), carrying an IncQ1 plasmid with bla_KPC-2 and aph(3′)-VI genes. Other genes associated with resistance and virulence found in the genome were bla_OXA-50, bla_PAO, fosA, catB, mutation in oprD and mexT (MexEF–OprN efflux regulator), and exotoxin-encoding genes (exoS, exoY and exoT).

This study highlights the potential risk of new STs of P. aeruginosa carrying bla_KPC-2 and the potential spread of bla_KPC-2 in an IncQ1 plasmid.

We described for the first time an extensively drug-resistant Pseudomonas aeruginosa isolate belonging to ST1284 carrying a plasmid-mediated bla_VIM-7 in Brazil. The bla_VIM-7 was harbored by an integron that also carried aacA4 and bla_OXA-46. Multiple virulence factors were also detected.