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Background: The whole-genome sequencing (WGS) is becoming an increasingly effective tool for rapid and accurate detection of drug resistance in Mycobacterium tuberculosis complex (MTBC). This approach, however, has still been poorly evaluated on strains from Central and Eastern European countries. The purpose of this study was to assess the performance of WGS against conventional drug susceptibility testing (DST) for the detection of multi-drug resistant (MDR) phenotypes among MTBC clinical strains from Poland and Lithuania. Methods: The study included 208 MTBC strains (130 MDR; 78 drug susceptible), recovered from as many tuberculosis patients in Lithuania and Poland between 2018 and 2021. Resistance to rifampicin (RIF) and isoniazid (INH) was assessed by Critical Concentration (CC) and Minimum Inhibitory Concentration (MIC) DST as well as molecular-based techniques, including line-probe assay (LPA) and WGS. The analysis of WGS results was performed using bioinformatic pipeline- and software-based tools. Results: The results obtained with the CC DST were more congruent with those by LPA compared to pipeline-based WGS. Software-based tools showed excellent concordance with pipeline-based analysis in prediction of RIF/INH resistance. The RIF-resistant strains demonstrated a relatively homogenous MIC distribution with the mode at the highest tested MIC value. The most frequent RIF-resistance conferring mutation was rpoB S450L. The mode MIC for INH was two-fold higher among double katG and inhA mutants than among single katG mutants. The overall rate of discordant results between all methods was calculated at 5.3%. Three strains had discordant results by both genotypic methods (LPA and pipeline-based WGS), one strain by LPA only, three strains by MIC DST, two strains by both MIC DST and pipeline-based WGS, and the remaining two strains showed discordant results with all three methods, compared to CC DST. Conclusions: Considering MIC DST results, current CCs of the first-line anti-TB drugs might be inappropriately high and may need to be revised. Both molecular methods demonstrated 100% specificity, while pipeline-based WGS had slightly lower sensitivity for RIF and INH than LPA, compared to CC DST. [ABSTRACT FROM AUTHOR]

Bacteria of the genus Paracoccus are common components of the microbiomes of many naturally- and anthropogenically shaped environments. One species, Paracoccus yeei , is unique within the genus because it is associated with opportunistic human infections. Therefore, strains of P. yeei may serve as an interesting model to study the transition from a saprophytic to a pathogenic lifestyle in environmental bacteria. Unfortunately, knowledge concerning the biology, genetics and genomic content of P. yeei is fragmentary; also the mechanisms of pathogenicity of this bacterium remain unclear. In this study we provide the first insight into the genome composition and metabolic potential of a clinical isolate, P. yeei CCUG 32053. This strain has a multipartite genome (4,632,079 bp) composed of a circular chromosome plus eight extrachromosomal replicons pYEE1–8: 3 chromids and 5 plasmids, with a total size of 1,247,173 bp. The genome has been significantly shaped by the acquisition of genomic islands, prophages (Myoviridae and Siphoviridae phage families) and numerous insertion sequences (ISs) representing seven IS families. Detailed comparative analysis with other complete genomic sequences of Paracoccus spp. (including P. yeei FDAARGOS_252 and TT13, as well as non-pathogenic strains of other species in this genus) enabled us to identify P. yeei species-specific genes and to predict putative determinants of virulence. This is the first attempt to identify pathoadaptive genetic information of P. yeei and to estimate the role of the mobilome in the evolution of pathogenicity in this species. [ABSTRACT FROM AUTHOR]

Bacteria of the genus Listeria (phylum Firmicutes) include both human and animal pathogens, as well as saprophytic strains. A common component of Listeria spp. genomes are plasmids, i.e., extrachromosomal replicons that contribute to gene flux in bacteria. This study provides an in-depth insight into the structure, diversity and evolution of plasmids occurring in Listeria strains inhabiting various environments under different anthropogenic pressures. Apart from the components of the conserved plasmid backbone (providing replication, stable maintenance and conjugational transfer functions), these replicons contain numerous adaptive genes possibly involved in: (i) resistance to antibiotics, heavy metals, metalloids and sanitizers, and (ii) responses to heat, oxidative, acid and high salinity stressors. Their genomes are also enriched by numerous transposable elements, which have influenced the plasmid architecture. The plasmidome of Listeria is dominated by a group of related replicons encoding the RepA replication initiation protein. Detailed comparative analyses provide valuable data on the level of conservation of these replicons and their role in shaping the structure of the Listeria pangenome, as well as their relationship to plasmids of other genera of Firmicutes, which demonstrates the range and direction of flow of genetic information in this important group of bacteria. [ABSTRACT FROM AUTHOR]

Bordetella bronchiseptica, an emerging zoonotic pathogen, infects a broad range of mammalian hosts. B. bronchiseptica-associated atrophic rhinitis incurs substantial losses to the pig breeding industry. The true burden of human disease caused by B. bronchiseptica is unknown, but it has been postulated that some hypervirulent B. bronchiseptica isolates may be responsible for undiagnosed respiratory infections in humans. B. bronchiseptica was shown to acquire antibiotic resistance genes from other bacterial genera, especially Escherichia coli. Here, we present a new B. bronchiseptica lytic bacteriophage—vB_BbrP_BB8—of the Podoviridae family, which offers a safe alternative to antibiotic treatment of B. bronchiseptica infections. We explored the phage at the level of genome, physiology, morphology, and infection kinetics. Its therapeutic potential was investigated in biofilms and in an in vivoGalleria mellonella model, both of which mimic the natural environment of infection. The BB8 is a unique phage with a genome structure resembling that of T7-like phages. Its latent period is 75 ± 5 min and its burst size is 88 ± 10 phages. The BB8 infection causes complete lysis of B. bronchiseptica cultures irrespective of the MOI used. The phage efficiently removes bacterial biofilm and prevents the lethality induced by B. bronchiseptica in G. mellonella honeycomb moth larvae. [ABSTRACT FROM AUTHOR]