Analysis of evolutionary patterns of genes in Campylobacter jejuni and C. coli

BACKGROUND: The thermophilic Campylobacter jejuni and Campylobacter coli are considered weakly clonal populations where incongruences between genetic markers are assumed to be due to random horizontal transfer of genomic DNA. In order to investigate the population genetics structure we extracted a set of 1180 core gene families (CGF) from 27 sequenced genomes of C. jejuni and C. coli. We adopted a principal component analysis (PCA) on the normalized evolutionary distances in order to reveal any patterns in the evolutionary signals contained within the various CGFs. RESULTS: The analysis indicates that the conserved genes in Campylobacter show at least two, possibly five, distinct patterns of evolutionary signals, seen as clusters in the score-space of our PCA. The dominant underlying factor separating the core genes is the ability to distinguish C. jejuni from C. coli. The genes in the clusters outside the main gene group have a strong tendency of being chromosomal neighbors, which is natural if they share a common evolutionary history. Also, the most distinct cluster outside the main group is enriched with genes under positive selection and displays larger than average recombination rates. CONCLUSIONS: The Campylobacter genomes investigated here show that subsets of conserved genes differ from each other in a more systematic way than expected by random horizontal transfer, and is consistent with differences in selection pressure acting on different genes. These findings are indications of a population of bacteria characterized by genomes with a mixture of evolutionary patterns

Whole genome genetic variation and linkage disequilibrium in a diverse collection of Listeria monocytogenes isolates.

We performed whole-genome multi-locus sequence typing for 2554 genes in a large and heterogenous panel of 180 Listeria monocytogenes strains having diverse geographical and temporal origins. The subtyping data was used for characterizing genetic variation and evaluating patterns of linkage disequilibrium in the pan-genome of L. monocytogenes. Our analysis revealed the presence of strong linkage disequilibrium in L. monocytogenes, with ~99% of genes showing significant non-random associations with a large majority of other genes in the genome. Twenty-seven loci having lower levels of association with other genes were considered to be potential "hot spots" for horizontal gene transfer (i.e., recombination via conjugation, transduction, and/or transformation). The patterns of linkage disequilibrium in L. monocytogenes suggest limited exchange of foreign genetic material in the genome and can be used as a tool for identifying new recombinant strains. This can help understand processes contributing to the diversification and evolution of this pathogenic bacteria, thereby facilitating development of effective control measures

Evolutionary Implications of Microbial Genome Tetranucleotide Frequency Biases

We compared nucleotide usage pattern conservation for related prokaryotes by examining the representation of DNA tetranucleotide combinations in 27 representative microbial genomes. For each of the organisms studied, tetranucleotide usage departures from expectations (TUD) were shared between related organisms using both Markov chain analysis and a zero-order Markov method. Individual strains, multiple chromosomes, plasmids, and bacteriophages share TUDs within a species. TUDs varied between coding and noncoding DNA. Grouping prokaryotes based on TUD profiles resulted in relationships with important differences from those based on 16S rRNA phylogenies, which may reflect unequal rates of evolution of nucleotide usage patterns following divergence of particular organisms from a common ancestor. By both symmetrical tree distance and likelihood analysis, phylogenetic trees based on TUD profiles demonstrate a level of congruence with 16S rRNA trees similar to that of both RpoA and RecA trees. Congruence of these trees indicates that there exists phylogenetic signal in TUD patterns, most prominent in coding region DNA. Because relationships demonstrated in TUD-based analyses utilize whole genomes, they should be considered complementary to phylogenies based on single genetic elements, such as 16S rRNA

Multilocus Sequence Typing of Listeria monocytogenes by Use of Hypervariable Genes Reveals Clonal and Recombination Histories of Three Lineages

In an attempt to develop a method to discriminate among isolates of Listeria monocytogenes, the sequences of all of the annotated genes from the fully sequenced strain L. monocytogenes EGD-e (serotype 1/2a) were compared by BLASTn to a file of the unfinished genomic sequence of L. monocytogenes ATCC 19115 (serotype 4b). Approximately 7% of the matching genes demonstrated 90% or lower identity between the two strains, and the lowest observed identity was 80%. Nine genes (hisJ, cbiE, truB, ribC, comEA, purM, aroE, hisC, and addB) in the 80 to 90% identity group and two genes (gyrB and rnhB) with approximately 97% identity were selected for multilocus sequence analysis in two sets of L. monocytogenes isolates (a 15-strain diversity set and a set of 19 isolates from a single food-processing plant). Based on concatenated sequences, a total of 33 allotypes were differentiated among the 34 isolates tested. Population genetics analyses revealed three lineages of L. monocytogenes that differed in their history of apparent recombination. Lineage I appeared to be completely clonal, whereas representatives of the other lineages demonstrated evidence of horizontal gene transfer and recombination. Although most of the gene sequences for lineage II strains were distinct from those of lineage I, a few strains with the majority of genes characteristic of lineage II had some that were characteristic of lineage I. Genes from lineage III organisms were mostly similar to lineage I genes, with instances of genes appearing to be mosaics with lineage II genes. Even though lineage I and lineage II generally demonstrated very distinct sequences, the sequences for the 11 selected genes demonstrated little discriminatory power within each lineage. In the L. monocytogenes isolate set obtained from one food-processing plant, lineage I and lineage II were found to be almost equally prevalent. While it appears that different lineages of L. monocytogenes can share habitats, they appear to differ in their histories of horizontal gene transfer

Genomic Changes within a Subset of IncI2 Plasmids Associated with Dissemination of <em>mcr-1</em> Genes and Other Important Antimicrobial Resistance Determinants

IncI2 plasmids appear to have only recently become associated with resistance genes; however, their tendency to carry resistance to the antibiotics of last resort and their widespread distribution increase their relative importance. In this study, we describe lineages within this plasmid family that have an increased likelihood of acquisition of antimicrobial resistance genes. Globally distributed mcr-1-carrying IncI2 plasmids were found to cluster with other IncI2 plasmids carrying extended-spectrum beta-lactamase genes, and separately from the non-resistant IncI2 plasmids. In addition, insertion sequence (IS) elements with no direct association with the acquired resistance genes also clustered with the resistance plasmids in the phylogenetic tree. In recognition of the biased sequencing of resistant plasmids globally, the analysis was also performed on resistant and non-resistant IncI2 plasmids sequenced in the USA through government surveillance efforts that do not rely on antibiotic selection. This analysis confirmed a distinct clustering associated with both resistance and mobile elements and identified possible genomic changes in core genes that correlate with increased acquisition of foreign DNA. This work highlights a potential genetic mechanism for increased uptake of foreign DNA within this prevalent family of plasmids

Prevalence and Subtype Characterization of Campylobacter in Ceca of Commercial Broiler Chickens at Processing – A 452 Flock, Seven-year Survey

Human Campylobacter infections have been associated with chicken and other poultry meat products. Environmental conditions such as temperature and season can affect Campylobacter recoverability from chicken meat products. In the presented study, we sought to investigate the relationship between ambient weather conditions and the isolation of Campylobacter from chicken flocks, as well as the subtype of these isolates. Campylobacter was isolated from the ceca of broilers collected in a commercial processing facility over 7 years, representing 452 flocks. Isolates were subjected to whole-genome sequencing and subtyping by multilocus sequence typing (MLST). Approximately 60% (269/452) of flocks sampled were positive for Campylobacter. There was no significant effect on the presence of detectable Campylobacter by month, season, temperature, or rainfall during grow-out or transportation. Sixty-eight different STs were detected; 45 C. jejuni and 23 C. coli. Diversity as measured by Shannon’s diversity index was higher in the spring and fall than in mid-winter and summer. We concluded that in the warm temperate climate of the Southeastern U.S., seasonality does not affect the rate of Campylobacter isolation from broilers, but the diversity of isolates was higher in the milder spring and fall seasons

Exudate From Retail Chicken Liver Packaging Allows for Survival of Naturally Occurring Campylobacter, Coliforms, and Aerobic Microorganisms Under Drying Conditions

Campylobacter spp. are a leading cause of human foodborne illness associated with chicken meat products in the United States. Chicken livers, including exudate from packaging, commonly carry Campylobacter and could be a source of illness if mishandled. Survivability of naturally occurring Campylobacter, total aerobic bacteria, and coliforms was determined under drying conditions in two consumer simulated environments: moist sponge and solid surface. Fresh chicken liver exudate was dispensed onto sponges and glass slides and allowed to dry under ambient conditions for 7 days. Bacterial concentration was measured at 0, 6, 24, 48, 72, and 168 h. Total aerobic population did not decrease by more than one log over 7 days and did not correlate to water activity or time in either simulation. Coliform concentrations increased in sponge simulations but decreased in solid surface simulations. Further, coliform concentrations were significantly higher in sponge simulations than in solid surface. Campylobacter was naturally present in exudate and survived at least to 6 h in every trial. Campylobacter was recoverable at 24 h in some sponge trials. However, Campylobacter concentration was strongly correlated to water activity. Fresh chicken liver exudate could present a risk of campylobacteriosis to consumers if mishandled even after drying

Inc A/C Plasmids Are Prevalent in Multidrug-Resistant Salmonella enterica Isolates▿ †

Bacterial plasmids are fragments of extrachromosomal double-stranded DNA that can contain a variety of genes that are beneficial to the host organism, like those responsible for antimicrobial resistance. The objective of this study was to characterize a collection of 437 Salmonella enterica isolates from different animal sources for their antimicrobial resistance phenotypes and plasmid replicon types and, in some cases, by pulsed-field gel electrophoresis (PFGE) in an effort to learn more about the distribution of multidrug resistance in relation to replicon types. A PCR-based replicon typing assay consisting of three multiplex PCRs was used to detect 18 of the 26 known plasmid types in the Enterobacteriaceae based on their incompatibility (Inc) replicon types. Linkage analysis was completed with antibiograms, replicon types, serovars, and Inc A/C. Inc A/C plasmids were prevalent in multidrug-resistant isolates with the notable exception of Salmonella enterica serovar Typhimurium. Cluster analysis based on PFGE of a subset of 216 isolates showed 155 unique types, suggesting a variable population, but distinct clusters of isolates with Inc A/C plasmids were apparent. Significant linkage of serovar was also seen with Inc replicon types B/O, I1, Frep, and HI1. The present study showed that the combination of Salmonella, the Inc A/C plasmids, and multiple-drug-resistant genes is very old. Our results suggest that some strains, notably serovar Typhimurium and closely related types, may have once carried the plasmid but that the resistance genes were transferred to the chromosome with the subsequent loss of the plasmid