Evaluation of Antibiotic Resistance of Salmonella Serotypes and Whole-Genome Sequencing of Multiresistant Strains Isolated from Food Products in Russia

Food products may be a source of Salmonella, one of the main causal agents of food poisoning, especially after the emergence of strains resistant to antimicrobial preparations. The present work dealt with investigation of the occurrence of resistance to antimicrobial preparations among S. enterica strains isolated from food. The isolates belonged to 11 serovars, among which Infantis (28%), Enteritidis (19%), and Typhimurium (13.4%) predominated. The isolates were most commonly resistant to trimethoprim/sulfamethoxazole (n = 19, 59.38%), cefazolin (n = 15, 46.86%), tetracycline (n = 13, 40.63%), and amikacin (n = 9, 28.13%). Most of the strains (68.75%) exhibited multiple resistance to commonly used antibiotics. High-throughput sequencing was used to analyse three multidrug-resistant strains (resistant to six or more antibiotics). Two of them (SZL 30 and SZL 31) belonged to S. Infantis, while one strain belonged to S. Typhimurium (SZL 38). Analysis of the genomes of the sequenced strains revealed the genes responsible for antibiotic resistance. In the genomes of strains SZL 30 and SZL 31 the genes of antibiotic resistance were shown to be localized mostly in integrons within plasmids, while most of the antibiotic resistance genes of strain SZL 38 were localized in a chromosomal island (17,949 nt). Genomes of the Salmonella strains SZL 30, SZL 31, and SZL 38 were shown to contain full-size pathogenicity islands: SPI-1, SPI-2, SPI-4, SPI-5, SPI-9, SPI-11, SPI-13, SPI-14, and CS54. Moreover, the genome of strain SZL 38 was also found to contain the full-size pathogenicity islands SPI-3, SPI-6, SPI-12, and SPI-16. The emergence of multidrug-resistant strains of various Salmonella serovars indicates that further research on the transmission pathways for these genetic determinants and monitoring of the distribution of these microorganisms are necessary.

Genomic Analysis of Global Staphylococcus argenteus Strains Reveals Distinct Lineages With Differing Virulence and Antibiotic Resistance Gene Content

Infections due to Staphylococcus argenteus have been increasingly reported worldwide and the microbe cannot be distinguished from Staphylococcus aureus by standard methods. Its complement of virulence determinants and antibiotic resistance genes remain unclear, and how far these are distinct from those produced by S. aureus remains undetermined. In order to address these uncertainties, we have collected 132 publicly available sequences from fourteen different countries, including the United Kingdom, between 2005 and 2018 to study the global genetic structure of the population. We have compared the genomes for antibiotic resistance genes, virulence determinants and mobile genetic elements such as phages, pathogenicity islands and presence of plasmid groups between different clades. 20% (n = 26) isolates were methicillin resistant harboring a mecA gene and 88% were penicillin resistant, harboring the blaZ gene. ST2250 was identified as the most frequent strain, but ST1223, which was the second largest group, contained a marginally larger number of virulence genes compared to the other STs. Novel S. argenteus pathogenicity islands were identified in our isolates harboring tsst-1, seb, sec3, ear, selk, selq toxin genes, as well as chromosomal clusters of enterotoxin and superantigen-like genes. Strain-specific type I modification systems were widespread which would limit interstrain transfer of genetic material. In addition, ST2250 possessed a CRISPR/Cas system, lacking in most other STs. S. argenteus possesses important genetic differences from S. aureus, as well as between different STs, with the potential to produce distinct clinical manifestations.

Testing assembly strategies of Francisella tularensis genomes to infer an evolutionary conservation analysis of genomic structures

Background We benchmarked sequencing technology and assembly strategies for short-read, long-read, and hybrid assemblers in respect to correctness, contiguity, and completeness of assemblies in genomes of Francisella tularensis. Benchmarking allowed in-depth analyses of genomic structures of the Francisella pathogenicity islands and insertion sequences. Five major high-throughput sequencing technologies were applied, including next-generation “short-read” and third-generation “long-read” sequencing methods. Results We focused on short-read assemblers, hybrid assemblers, and analysis of the genomic structure with particular emphasis on insertion sequences and the Francisella pathogenicity island. The A5-miseq pipeline performed best for MiSeq data, Mira for Ion Torrent data, and ABySS for HiSeq data from eight short-read assembly methods. Two approaches were applied to benchmark long-read and hybrid assembly strategies: long-read-first assembly followed by correction with short reads (Canu/Pilon, Flye/Pilon) and short-read-first assembly along with scaffolding based on long reads (Unicyler, SPAdes). Hybrid assembly can resolve large repetitive regions best with a “long-read first” approach. Conclusions Genomic structures of the Francisella pathogenicity islands frequently showed misassembly. Insertion sequences (IS) could be used to perform an evolutionary conservation analysis. A phylogenetic structure of insertion sequences and the evolution within the clades elucidated the clade structure of the highly conservative F. tularensis.

Shape shifter: redirection of prolate phage capsid assembly by staphylococcal pathogenicity islands

Staphylococcus aureus pathogenicity islands (SaPIs) are molecular parasites that hijack helper phages for their transfer. SaPIbov5, the prototypical member of a family of cos type SaPIs, redirects the assembly of ϕ12 helper capsids from prolate to isometric. This size and shape shift is dependent on the SaPIbov5-encoded protein Ccm, a homolog of the ϕ12 capsid protein (CP). Using cryo-electron microscopy, we have determined structures of prolate ϕ12 procapsids and isometric SaPIbov5 procapsids. ϕ12 procapsids have icosahedral end caps with Tend = 4 architecture and a Tmid = 14 cylindrical midsection, whereas SaPIbov5 procapsids have T = 4 icosahedral architecture. We built atomic models for CP and Ccm, and show that Ccm occupies the pentameric capsomers in the isometric SaPIbov5 procapsids, suggesting that preferential incorporation of Ccm pentamers prevents the cylindrical midsection from forming. Our results highlight that pirate elements have evolved diverse mechanisms to suppress phage multiplication, including the acquisition of phage capsid protein homologs.

Staphylococcal phages and pathogenicity islands drive plasmid evolution

Conjugation has classically been considered the main mechanism driving plasmid transfer in nature. Yet bacteria frequently carry so-called non-transmissible plasmids, raising questions about how these plasmids spread. Interestingly, the size of many mobilisable and non-transmissible plasmids coincides with the average size of phages (~40 kb) or that of a family of pathogenicity islands, the phage-inducible chromosomal islands (PICIs, ~11 kb). Here, we show that phages and PICIs from Staphylococcus aureus can mediate intra- and inter-species plasmid transfer via generalised transduction, potentially contributing to non-transmissible plasmid spread in nature. Further, staphylococcal PICIs enhance plasmid packaging efficiency, and phages and PICIs exert selective pressures on plasmids via the physical capacity of their capsids, explaining the bimodal size distribution observed for non-conjugative plasmids. Our results highlight that transducing agents (phages, PICIs) have important roles in bacterial plasmid evolution and, potentially, in antimicrobial resistance transmission.

Russian strains of group B streptococci are different in the content and organization of the PAI-A and PAI-A1 pathogenicity islands

Group B streptococci, or Streptococcus agalactiae, are the major cause of severe diseases in newborns and adults. The PAI-A and PAI-A1 pathogenicity islands containing the sspB1 and sspB1a genes, respectively, were found among group B streptococci mobile genetic elements. The presence of sspB genes correlates with urogenital tract infections. The aim of this study was to determine the frequency of group B streptococci strains with the PAI-A and PAI-A1 pathogenicity islands, circulating in Moscow, in comparison with strains from St. Petersburg. The sspB1 gene, and hence the PAI-A pathogenicity island, was not found in the genomes of strains from Moscow. The frequency of the sspB1a gene and the PAI-A1 pathogenicity island in the genomes of clinical strains was three times higher than in the genomes of colonizing strains. Thus, it can be assumed that the genes of the sspB family are more specific of group B streptococci colonizing pregnant women and newborns.

Pathogenicity Island in Salmonella

Considering a complex set of interplay with its host, Salmonella needs numerous genes for its full virulence. These genes responsible for invasion, survival, and extra intestinal spread are located on pathogenicity islands known as Salmonella pathogenicity islands (SPIs) that are thought to be acquired by horizontal gene transfer. A total of 17 SPIs (1–17) are recognized so far. The type III secretion system (T3SS) encoded by SPI-1 is considered as the most important virulence factor for Salmonella that delivers effector proteins necessary for invasion and production of enteritis. Among various SPIs, the role in virulence is well proven for SPI1 and SPI2 and further insight into the complex regulatory network of SPIs can contribute to drug investigation and prevention of infection.

Continuous Genomic Surveillance Monitored the In Vivo Evolutionary Trajectories of Vibrio parahaemolyticus and Identified a New Virulent Genotype

ABSTRACT Our ability to predict evolutionary trajectories of pathogens is one of the promising leverages to fight against the pandemic disease, yet few studies have addressed this question in situ, due to the difficulty in monitoring the milestone evolutionary events for a given pathogen and in understanding the evolutionary strategies. In this study, we monitored the real-time evolution of Vibrio parahaemolyticus in response to successive antibiotic treatment in three shrimp farms in North China from 2011 to 2018 by whole-genome sequencing. Results showed that the stepwise emergence of resistance was associated with the antibiotic usage. Genomic analysis of resistant isolates showed that the acquisition of the resistant mobile genetic elements flanked by an insertion sequence (ISVal1) closely mirrored the antibiotics used in shrimp farms since 2014. Next, we also identified 50 insertion sites of ISVal1 in the chromosome, which facilitated the formation of pathogenicity islands (PAIs) and fitness islands in the following years. Further, horizontal transfers of a virulent trh-nik-ure genomic island (GI) and two GIs improving the fitness have been observed in two farms since 2016. In this case study, we proposed that the insertion sequence triggered four major evolutionary events during the outbreaks of shrimp disease in three farms, including horizontal transfer of transposon (HTT) (stage 1), the formation of resistance islands (stage 2) and the PAIs (stage 3), and horizontal transfer of the PAIs (stage 4). This study presented the first in vivo evolutionary trajectories for a given bacterial pathogen, which helps us to understand the emergence mechanisms of new genotypes. IMPORTANCE Most human infectious diseases originate from animals. Thus, how to reduce or prevent pandemic zoonoses before they emerge in people is becoming a critical issue. Continuous genomic surveillance of the evolutionary trajectories of potential human pathogens on farms is a promising strategy to realize early warning. Here, we conducted an 8-year surveillance of Vibrio parahaemolyticus in three shrimp farms. The results showed that the use of antibiotics and horizontal transfer of transposons (HTT) drove the evolution of V. parahaemolyticus, which could be divided into four stages: HTT, formation of resistance islands, formation of pathogenicity islands (PAIs), and horizontal transfer of PAIs. This study presented the first in vivo monitoring of evolutionary trajectories for a given bacterial pathogen, providing valuable information for the prevention of pandemic zoonoses.

Horizontally acquired papGII-containing pathogenicity islands underlie the emergence of invasive uropathogenic Escherichia coli lineages

Escherichia coli is the leading cause of urinary tract infection, one of the most common bacterial infections in humans. Despite this, a genomic perspective is lacking regarding the phylogenetic distribution of isolates associated with different clinical syndromes. Here, we present a large-scale phylogenomic analysis of a spatiotemporally and clinically diverse set of 907 E. coli isolates, including 722 uropathogenic E. coli (UPEC) isolates. A genome-wide association approach identifies the (P-fimbriae-encoding) papGII locus as the key feature distinguishing invasive UPEC, defined as isolates associated with severe UTI, i.e., kidney infection (pyelonephritis) or urinary-source bacteremia, from non-invasive UPEC, defined as isolates associated with asymptomatic bacteriuria or bladder infection (cystitis). Within the E. coli population, distinct invasive UPEC lineages emerged through repeated horizontal acquisition of diverse papGII-containing pathogenicity islands. Our findings elucidate the molecular determinants of severe UTI and have implications for the early detection of this pathogen.