Genetic Stability and Evolution of thesigBAllele, Used for Listeria Sensu Stricto Subtyping and Phylogenetic Inference

Abstract

Sequencing of single genes remains an important tool that allows the rapid classification of bacteria. Sequencing of a portion of , which encodes a stress-responsive alternative sigma factor, has emerged as a commonly used molecular tool for the initial characterization of diverse isolates. In this study, evolutionary approaches were used to assess the validity of allelic typing for For a data set of 4,280 isolates, allelic typing showed a Simpson's index of diversity of 0.96. Analyses of 164 allelic types (ATs) found among the 6 species, representing these 4,280 isolates, indicate that neither frequent homologous recombination nor positive selection significantly contributed to the evolution of, confirming its genetic stability. The molecular clock test provided evidence for unequal evolution rates across clades; displayed the lowest diversity and was the only species in which evolved in a clocklike manner, implying a unique natural history. Among the four lineages, evolution followed a molecular clock only in lineage IV. Moreover, displayed a significant negative Tajima value in lineage II, suggesting a recent population bottleneck followed by lineage expansion. The absence of positive selection along with the violation of the molecular clock suggested a nearly neutral mechanism of evolution. While comparison with a whole-genome sequence-based phylogeny revealed that the phylogeny did not correctly reflect the ancestry of lineage IV, the availability of a large AT database allowed accurate species classification. allelic typing has been widely used for species delineation and subtyping of However, an informative evaluation of this method from an evolutionary perspective was missing. Our data indicate that the genetic stability of is affected by neither frequent homologous recombination nor positive selection, which supports that allelic typing provides reliable subtyping and classification of strains. However, multigene data are required for accurate phylogeny reconstruction of This study thus contributes to a better understanding of the evolution of and confirms the robustness of the subtyping system for.

Detection and Prevalence of Listeria in U.S. Produce Packinghouses and Fresh-Cut Facilities

“…In addition, sampling sites representing drains showed repeat positives in three operations, with three drain-related sites showing repeat positives in packinghouse A. Whereas all isolates were characterized by sigB allelic typing, the data are consistent with findings from previous studies (1,5,16) that sigB sequencing alone has limited discriminatory power and is typically insufficient for establishment of persistent contamination, as supported by the fact that LM with sigB AT57 was found in all six operations that had LM positives. However, for two sites ("square drain 1" in packinghouse A and "rot bin under rollers" in packinghouse C; see Table 3), isolates with identical sigB ATs were obtained on three separate sampling dates; these sites had high priority for follow-up, including follow-up higher resolution subtyping.…”

“…The sequences were then trimmed for analysis of the 660-bp internal fragment to determine the sigB allelic type (AT) for each isolate. The sigB gene was used for initial subtyping and characterization due to its genetic stability and accuracy in species and lineage classifications, as previously detailed (16). Sequences were edited and assembled using Sequencher sequence analysis software (version 5.4.6, Gene Codes Corporation, Ann Arbor, MI) and were analyzed using MEGA7 (13).…”

“…Sequences were edited and assembled using Sequencher sequence analysis software (version 5.4.6, Gene Codes Corporation, Ann Arbor, MI) and were analyzed using MEGA7 (13). The assignment of ATs to each sequence was performed as previously described, using a sigB database available on Food Microbe Tracker (http://www.foodmicrobetracker.com) (16,26). All Listeria isolates were preserved at À808C in 15% glycerol.…”

Sequence-Based Characterization of <i>Listeria monocytogenes</i> Strains Isolated from Domestic Retail Meat in the Tokyo Metropolitan Area of Japan

“…In a previous study, we reported that two strains, 76P1 (Table 3) and 1E1 (sigB AT 37; derived from the skin of beef cattle), possessed nonsense mutations in the sigB gene (6). According to genetic analysis of the sigB allele, this sequence was found to be relatively conserved in L. monocytogenes (14). We confirmed that only two strains, FSL J1-0086 (AT 103, isolate source: human sporadic listeriosis) and Cornell University (Ithaca, NY, USA) (14,15).…”

“…According to genetic analysis of the sigB allele, this sequence was found to be relatively conserved in L. monocytogenes (14). We confirmed that only two strains, FSL J1-0086 (AT 103, isolate source: human sporadic listeriosis) and Cornell University (Ithaca, NY, USA) (14,15). In addition to these nonsense mutation-containing types of sigB (ATs 36, 37, 39-41, and 43), a new type of sigB (ATs 38 and 42) was identified that is not currently registered in either of the databases at the Pasteur Institute or Cornell University.…”