A panel of diverse Klebsiella pneumoniae clinical isolates for research and development.

Klebsiella pneumoniae are a leading cause of healthcare-associated infections worldwide. In particular, strains expressing extended-spectrum ß-lactamases (ESBLs) and carbapenemases pose serious treatment challenges, leading the World Health Organization (WHO) to designate ESBL and carbapenem-resistant Enterobacteriaceae as 'critical' threats to human health. Research efforts to combat these pathogens can be supported by accessibility to diverse and clinically relevant isolates for testing novel therapeutics. Here, we describe a panel of 100 diverse K. pneumoniae isolates that are publicly available to assist the research community in this endeavour. Whole-genome sequencing (WGS) was performed on 3878 K. pneumoniae clinical isolates housed at the Multidrug-Resistant Organism Repository and Surveillance Network. The isolates were cultured from 63 facilities in 19 countries between 2001 and 2020. Core-genome multilocus sequence typing and high-resolution single-nucleotide polymorphism-based phylogenetic analyses captured the genetic diversity of the collection and were used to select the final panel of 100 isolates. In addition to known multidrug-resistant (MDR) pandemic lineages, the final panel includes hypervirulent lineages and isolates with specific and diverse resistance genes and virulence biomarkers. A broad range of antibiotic susceptibilities, ranging from pan-sensitive to extensively drug-resistant isolates, are described. The panel collection, and all associated metadata and genome sequences, are available at no additional cost and will be an important resource for the research community and for the design and development of novel antimicrobial agents and diagnostics against this important pathogen.

Molecular characterization of multidrug-resistant ESKAPEE pathogens from clinical samples in Chonburi, Thailand (2017-2018).

BACKGROUND: ESKAPEE pathogens Enterococcus faecium, Staphylococcus aureus, Klebsiella pneumoniae, Acinetobacter baumannii, Pseudomonas aeruginosa, Enterobacter spp. and Escherichia coli are multi-drug resistant (MDR) bacteria that present increasing treatment challenges for healthcare institutions and public health worldwide. METHODS: 431 MDR ESKAPEE pathogens were collected from Queen Sirikit Naval Hospital, Chonburi, Thailand between 2017 and 2018. Species identification and antimicrobial resistance (AMR) phenotype were determined following CLSI and EUCAST guidelines on the BD Phoenix System. Molecular identification of antibiotic resistant genes was performed by polymerase chain reaction (PCR), real-time PCR assays, and whole genome sequencing (WGS). RESULTS: Of the 431 MDR isolates collected, 1.2% were E. faecium, 5.8% were S. aureus, 23.7% were K. pneumoniae, 22.5% were A. baumannii, 4.6% were P. aeruginosa, 0.9% were Enterobacter spp., and 41.3% were E. coli. Of the 401 Gram-negative MDR isolates, 51% were carbapenem resistant, 45% were ESBL producers only, 2% were colistin resistance and ESBLs producers (2%), and 2% were non-ESBLs producers. The most prevalent carbapenemase genes were blaOXA-23 (23%), which was only identified in A. baumannii, followed by blaNDM (17%), and blaOXA-48-like (13%). Beta-lactamase genes detected included blaTEM, blaSHV, blaOXA, blaCTX-M, blaDHA, blaCMY, blaPER and blaVEB. Seven E. coli and K. pneumoniae isolates showed resistance to colistin and carried mcr-1 or mcr-3, with 2 E. coli strains carrying both genes. Among 30 Gram-positive MDR ESKAPEE, all VRE isolates carried the vanA gene (100%) and 84% S. aureus isolates carried the mecA gene. CONCLUSIONS: This report highlights the prevalence of AMR among clinical ESKAPEE pathogens in eastern Thailand. E. coli was the most common MDR pathogen collected, followed by K. pneumoniae, and A. baumannii. Carbapenem-resistant Enterobacteriaceae (CRE) and extended spectrum beta-lactamases (ESBLs) producers were the most common resistance profiles. The co-occurrence of mcr-1 and mcr-3 in 2 E. coli strains, which did not affect the level of colistin resistance, is also reported. The participation of global stakeholders and surveillance of MDR remain essential for the control and management of MDR ESKAPEE pathogens.

Comparison of Pseudomonas aeruginosa strains reveals that Exolysin A toxin plays an additive role in virulence.

BACKGROUND: Pseudomonas aeruginosa possesses an array of virulence genes ensuring successful infection development. A two-partner secretion system Exolysin BA (ExlBA) is expressed in the PA7-like genetic outliers consisting of ExlA, a pore-forming toxin and ExlB transporter protein. Presence of exlBA in multidrug-resistant (MDR) strains has not been investigated, particularly in the strains isolated from wounded soldiers. METHODS: We screened whole genome sequences of 2439 MDR- P. aeruginosa strains for the presence of exlBA. We compiled all exlBA positive strains and compared them with a diversity set for demographics, antimicrobial profiles and phenotypic characteristics: surface motility, biofilm formation, pyocyanin production and hemolysis. We compared the virulence of strains with comparable phenotypic characteristics in Galleria mellonella. RESULTS: We identified 33 exlBA-positive strains (1.5%). These strains have increased antibiotic resistance, they are more motile, produce more robust biofilms and have comparable pyocianin production with the diversity set despite the phenotypic differences within the group. In in vivo infection models, these strains were less virulent than Type III Secretion System (T3SS) positive counterparts. CONCLUSIONS: exlBA-positive strains are wide spread among the PA7-like outliers. While not as virulent as strains possessing T3SS, these strains exhibit phenotypic features associated with virulence and are still lethal in vivo.