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Introduction bla OXA-48, bla NDM-1 and bla CTX-M-3 are clinically relevant resistance genes, frequently associated with the broad-host range plasmids of the IncL/M group. The L and M plasmids belong to two compatible groups, which were incorrectly classified together by molecular methods. In order to understand their evolution, we fully sequenced four IncL/M plasmids, including the reference plasmids R471 and R69, the recently described bla OXA-48-carrying plasmid pKPN-El.Nr7 from a Klebsiella pneumoniae isolated in Bern (Switzerland), and the bla SHV-5 carrying plasmid p202c from a Salmonella enterica from Tirana (Albania).MethodsSequencing was performed using 454 Junior Genome Sequencer (Roche). Annotation was performed using Sequin and Artemis software. Plasmid sequences were compared with 13 fully sequenced plasmids belonging to the IncL/M group available in GenBank.ResultsComparative analysis of plasmid genomes revealed two distinct genetic lineages, each containing one of the R471 (IncL) and R69 (IncM) reference plasmids. Conjugation experiments demonstrated that plasmids representative of the IncL and IncM groups were compatible with each other. The IncL group is constituted by the bla OXA-48-carrying plasmids and R471. The IncM group contains two sub-types of plasmids named IncM1 and IncM2 that are each incompatible.ConclusionThis work re-defines the structure of the IncL and IncM families and ascribes a definitive designation to the fully sequenced IncL/M plasmids available in GenBank.

The Mycobacterium tuberculosis genome includes the large family of pe_pgrs genes, whose functions are unknown. Because of precedents in other pathogens in which gene families showing high sequence variation are involved in antigenic variation, a similar role has been proposed for the pe_pgrs genes. However, the impact of immune selection on pe_pgrs genes has not been examined. Here, we sequenced 27 pe_pgrs genes in 94 clinical strains from five phylogenetic lineages of the M. tuberculosis complex (MTBC). We found that pe_pgrs genes were overall more diverse than the remainder of the MTBC genome, but individual members of the family varied widely in their nucleotide diversity and insertion/deletion (indel) content: some were more, and others were much less, diverse than the genome average. Individual pe_pgrs genes also differed in the ratio of nonsynonymous to synonymous mutations, suggesting that different selection pressures act on individual pe_pgrs genes. Using bioinformatic methods, we tested whether sequence diversity in pe_pgrs genes might be selected by human T cell recognition, the major mechanism of adaptive immunity to MTBC. We found that the large majority of predicted human T cell epitopes were confined to the conserved PE domain and experimentally confirmed the antigenicity of this domain in tuberculosis patients. In contrast, despite being genetically diverse, the PGRS domains harbored few predicted T cell epitopes. These results indicate that human T cell recognition is not a significant force driving sequence diversity in pe_pgrs genes, which is consistent with the previously reported conservation of human T cell epitopes in the MTBC.

Background Despite the adoption of strict infection prevention and control measures, many hospitals have reported outbreaks of multidrug-resistant organisms (MDRO) during the Coronavirus 2019 (COVID-19) pandemic. Following an outbreak of carbapenem-resistant Acinetobacter baumannii (CRAB) in our institution, we sought to systematically analyse characteristics of MDRO outbreaks in times of COVID-19, focussing on contributing factors and specific challenges in controlling these outbreaks. Methods We describe results of our own CRAB outbreak investigation and performed a systematic literature review for MDRO (including Candida auris) outbreaks which occurred during the COVID-19 pandemic (between December 2019 and March 2021). Search terms were related to pathogens/resistance mechanisms AND COVID-19. We summarized outbreak characteristics in a narrative synthesis and contrasted contributing factors with implemented control measures. Results The CRAB outbreak occurred in our intensive care units between September and December 2020 and comprised 10 patients (thereof seven with COVID-19) within two distinct genetic clusters (both ST2 carrying OXA-23). Both clusters presumably originated from COVID-19 patients transferred from the Balkans. Including our outbreak, we identified 17 reports, mostly caused by Candida auris (n = 6) or CRAB (n = 5), with an overall patient mortality of 35% (68/193). All outbreaks involved intensive care settings. Non-adherence to personal protective equipment (PPE) or hand hygiene (n = 11), PPE shortage (n = 8) and high antibiotic use (n = 8) were most commonly reported as contributing factors, followed by environmental contamination (n = 7), prolonged critical illness (n = 7) and lack of trained HCW (n = 7). Implemented measures mainly focussed on PPE/hand hygiene audits (n = 9), environmental cleaning/disinfection (n = 9) and enhanced patient screening (n = 8). Comparing potentially modifiable risk factors and control measures, we found the largest discrepancies in the areas of PPE shortage (risk factor in 8 studies, addressed in 2 studies) and patient overcrowding (risk factor in 5 studies, addressed in 0 studies). Conclusions Reported MDRO outbreaks during the COVID-19 pandemic were most often caused by CRAB (including our outbreak) and C. auris. Inadequate PPE/hand hygiene adherence, PPE shortage, and high antibiotic use were the most commonly reported potentially modifiable factors contributing to the outbreaks. These findings should be considered for the prevention of MDRO outbreaks during future COVID-19 waves.