High incidence of carbapenemase-producing Pseudomonas aeruginosa clinical isolates from Lagos, Nigeria.

Background: Carbapenem-resistant Pseudomonas aeruginosa strains are on the rise worldwide. This study characterized clinical isolates of P. aeruginosa from three Nigerian hospitals for carbapenem resistance. Methods: Strains isolated from wounds (n = 88), urine/catheter tips (n = 25), sputum/tracheotomy aspirates (n = 5), ear swabs (n = 4) and vaginal swabs (n = 1) were identified by MALDI-TOF and antibiotic susceptibility testing was performed using the VITEK 2 system. The genomic DNA of each isolate was subject to sequencing using Illumina and Oxford nanopore technology. Bioinformatics analyses were performed to detect antimicrobial resistance genes, clonal affiliations and phylogenetic relations of 123 non-duplicate P. aeruginosa isolates, whereas assembly of the nanopore reads using the plasmIDent pipeline enabled the identification of plasmids. Results: Forty-three percent of the isolates were resistant to all antibiotic categories tested. More than 40% of the isolates were resistant to the carbapenems imipenem and/or meropenem (39% and 44%, respectively). Among the meropenem-resistant isolates, 48 (89%) carried at least one carbapenemase gene. The predominant one was bla NDM-1 (n = 34), which conferred resistance to all five antibiotic categories and highly increased the MICs of both meropenem and imipenem. The other recurrent carbapenemase genes were bla VIM-2 (n = 4), and bla VIM-5-like (n = 11), which co-existed with bla NDM-1 in two isolates. Conclusions: The study revealed a high rate of carbapenem resistance and conjugative, broad host range plasmids carrying carbapenemase-encoding genes, especially the NDM-1 type, among isolates of P. aeruginosa. This may forebode the emergency of ubiquitous carbapenem resistance urging the implementation of infection control and antimicrobial stewardship strategies in Nigerian hospitals.

Surveillance of Enterobacter cloacae complex colonization and comparative analysis of different typing methods on a neonatal intensive care unit in Germany.

BACKGROUND: Enterobacter cloacae complex is a group of common opportunistic pathogens on neonatal intensive care units. Active microbiological screening to guide empirical antimicrobial treatment or to detect transmission events is recommended in high-risk preterm neonates. A rise in colonization with E. cloacae complex was observed in a German perinatal centre. The aim of this study was to evaluate the performance of different typing techniques using whole genome sequencing (WGS) as a reference. METHODS: Enterobacter cloacae complex isolates from clinical and screening specimens with an epidemiological link to the neonatal intensive care units were further assessed. Identification and antibiotic susceptibility testing was performed by a combination of VITEK2 (bioMérieux) and MALDI-TOF (Bruker Daltonics), followed by RAPD/rep-PCR and PFGE (XbaI). Retrospectively, all isolates were analyzed by Fourier-transform infrared (FTIR) spectroscopy (IR Biotyper, Bruker Daltonics). Whole genome sequencing with SNP-based clustering was used as the reference method. Furthermore, resistome analysis, sequence type and species identification were derived from the WGS data. Transmission analysis was based on epidemiological and typing data. RESULTS: Between September 2017 and March 2018 32 mostly preterm neonates were found to be colonized with E. cloacae complex and 32 isolates from 24 patients were available for further typing. RAPD/rep-PCR and PFGE showed good concordance with WGS whereas FTIR displayed mediocre results [adjusted rand index (ARI) = 0.436]. A polyclonal increase and two dominant and overlapping clonal clusters of two different E. hormaechei subspecies were detected. Overall, four different species were identified. Genotyping confirmed third-generation cephalosporin resistance development in isolates of the same patient. During the six-month period several infection prevention interventions were performed and no E. cloacae complex isolates were observed during the following months. CONCLUSIONS: Interpretation of the microbiological results alone to detect transmission events is often challenging and bacterial typing is of utmost importance to implement targeted infection control measures in an epidemic occurrence of E. cloacae complex. WGS is the most discriminatory method. However, traditional methods such as PFGE or RAPD/rep-PCR can provide reliable and quicker results in many settings. Furthermore, research is needed to quickly identify E. cloacae complex to the species level in the microbiological laboratory.

Acinetobacter geminorum sp. nov., isolated from human throat swabs.

Two isolates of a non-fermenting, Gram-negative bacterial strain were cultured from two throat swabs that were taken from a pair of twins during routine microbiological surveillance screening. As these isolates could not be unambiguously identified using routine diagnostic methods, whole genome sequencing was performed followed by phylogenetic analysis based on the rpoB gene sequence and by whole genome datasets. The two strains compose a separate branch within the clade formed by the Acinetobacter calcoaceticus-baumannii (ACB) complex with Acinetobacter pittii CIP 70.29T as the most closely related species. The average nucleotide identity compared to all other species of the ACB complex was below 94.2% and digital DNA-DNA hybridization values were less than 60%. Biochemical characteristics confirm affiliation to the ACB complex with some specific phenotypic differences. As a result of the described data, a new Acinetobacter species is introduced, for which the name Acinetobacter geminorum sp. nov. is proposed. The type strain is J00019T with a G+C DNA content of 38.8 mol% and it is deposited in the DSMZ Germany (DSM 111094T) and CCUG Sweden (CCUG 74625T).

Fourier-Transform Infrared (FTIR) Spectroscopy for Typing of Clinical Enterobacter cloacae Complex Isolates.

Members of the Enterobacter (E.) cloacae complex have emerged as important pathogens frequently encountered in nosocomial infections. Several outbreaks with E. cloacae complex have been reported in recent years, especially in neonatal units. Fast and reliable strain typing methods are crucial for real-time surveillance and outbreak analysis to detect pathogen reservoirs and transmission routes. The aim of this study was to evaluate the performance of Fourier-transform infrared (FTIR) spectroscopy as a fast method for typing of clinical E. cloacae complex isolates, when whole genome sequencing (WGS) analysis was used as reference. First, the technique was used retrospectively on 24 first isolates of E. cloacae complex strains from neonatal patients and showed good concordance with SNP-based clustering [adjusted rand index (ARI) = 0.818] and with the sequence type (ST) (ARI = 0.801). 29 consecutive isolates from the same patients were shown by WGS analysis to almost always belong to the same SNP cluster as the first isolates, which was only inconsistently recognized by FTIR spectroscopy. Training of an artificial neural network (ANN) with all FTIR spectra from sequenced strains markedly improved the recognition of related and unrelated isolate spectra. In a second step, FTIR spectroscopy was applied on 14 strains during an outbreak with E. cloacae complex and provided fast typing results that were confirmed by WGS analysis. In conclusion, FTIR spectroscopy is a promising tool for strain typing of clinical E. cloacae complex strains. Discriminatory power can be improved by implementing an ANN for spectrum analysis. Due to its low costs and fast turnaround times, the method presents a valuable tool for real-time surveillance as well as outbreak analysis.