Yersinia pestis , the causative agent of plague, is a highly uniform clone that diverged recently from the enteric pathogen Yersinia pseudotuberculosis . Despite their close genetic relationship, they differ radically in their pathogenicity and transmission. Here, we report the complete genomic sequence of Y. pseudotuberculosis IP32953 and its use for detailed genome comparisons with available Y. pestis sequences. Analyses of identified differences across a panel of Yersinia isolates from around the world reveal 32 Y. pestis chromosomal genes that, together with the two Y. pestis -specific plasmids, to our knowledge, represent the only new genetic material in Y. pestis acquired since the the divergence from Y. pseudotuberculosis . In contrast, 149 other pseudogenes (doubling the previous estimate) and 317 genes absent from Y. pestis were detected, indicating that as many as 13% of Y. pseudotuberculosis genes no longer function in Y. pestis . Extensive insertion sequence-mediated genome rearrangements and reductive evolution through massive gene loss, resulting in elimination and modification of preexisting gene expression pathways, appear to be more important than acquisition of genes in the evolution of Y. pestis . These results provide a sobering example of how a highly virulent epidemic clone can suddenly emerge from a less virulent, closely related progenitor.

DNA microarrays encompassing the entire genome of Yersinia pestis were used to characterize global regulatory changes during steady-state vegetative growth occurring after shift from 26 to 37°C in the presence and absence of Ca ²⁺ . Transcriptional profiles revealed that 51, 4, and 13 respective genes and open reading frames (ORFs) on pCD, pPCP, and pMT were thermoinduced and that the majority of these genes carried by pCD were downregulated by Ca ²⁺ . In contrast, Ca ²⁺ had little effect on chromosomal genes and ORFs, of which 235 were thermally upregulated and 274 were thermally downregulated. The primary consequence of these regulatory events is profligate catabolism of numerous metabolites available in the mammalian host.

The established phylogeny of the etiological agent of plague, Yersinia pestis, is not perfect, as it does not take into account the strains from numerous natural foci of Commonwealth of Independent States (CIS). We have carried out PCR and SNP typing of 359 strains and whole genome sequencing of 51 strains from these plague foci and determined the phylogenetic diversity of the strains circulating here. They belong to 0.ANT3, 0.ANT5, 2.ANT3, 4.ANT branches of antique biovar, 2.MED0, 2.MED1 branches of medieval biovar and to 0.PE2, 0.PE4a. 0.PE4h, 0.PE4t branches. Based on the studies of 178 strains from 23 plague foci of CIS countries, it was determined that the population structure of 2.MED strains is subdivided into Caucasian–Caspian and Central Asian–Chinese branches. In Central-Caucasian high-mountain plague foci in the Russian Federation (RF) the most deeply diverged branch of medieval biovar, 2.MED0, has been found. With the data obtained, the current population structure of Y. pestis species has been refined. New subspecies classification is developed, comprising seven subspecies: pestis, caucasica (0.PE2), angolica (0.PE3), central asiatica (0.PE4), tibetica (0.PE7), ulegeica (0.PE5), and qinghaica (0.PE10).

We used a whole-genome scanning technique to identify the NADH dehydrogenase gamma subunit ( nuoG ) primer set that is sensitive and specific enough to detect a diverse number of Bartonella species in a wide range of environmental samples yet maintains minimal cross-reactivity to mammalian host and arthropod vector organisms.

A PCR-based genotyping system that detects divergence of IS 100 locations within the Yersinia pestis genome was used to characterize a large collection of isolates of different biovars and geographical origins. Using sequences derived from the glycerol-negative biovar orientalis strain CO92, a set of 27 locus-specific primers was designed to amplify fragments between the end of IS 100 and its neighboring gene. Geographically diverse members of the orientalis biovar formed a homogeneous group with identical genotype with the exception of strains isolated in Indochina. In contrast, strains belonging to the glycerol-positive biovar antiqua showed a variety of fingerprinting profiles. Moreover, strains of the biovar medievalis (also glycerol positive) clustered together with the antiqua isolates originated from Southeast Asia, suggesting their close phylogenetic relationships. Interestingly, a Manchurian biovar antiqua strain Nicholisk 51 displayed a genotyping pattern typical of biovar orientalis isolates. Analysis of the glycerol pathway in Y. pestis suggested that a 93-bp deletion within the glpD gene encoding aerobic glycerol-3-phosphate dehydrogenase might account for the glycerol-negative phenotype of the orientalis biovar. The glpD gene of strain Nicholisk 51 did not possess this deletion, although it contained two nucleotide substitutions characteristic of the glpD version found exclusively in biovar orientalis strains. To account for this close relationship between biovar orientalis strains and the antiqua Nicholisk 51 isolate, we postulate that the latter represents a variant of this biovar with restored ability to ferment glycerol. The fact that such a genetic lesion might be repaired as part of the natural evolutionary process suggests the existence of genetic exchange between different Yersinia strains in nature. The relevance of this observation on the emergence of epidemic Y. pestis strains is discussed.

Yersinia pestisevolved fromY. pseudotuberculosisto become the causative agent of bubonic and pneumonic plague. We identified a homolog of theSalmonella entericaserovar Typhimurium lipoprotein (lpp) gene inYersiniaspecies and preparedlppgene deletion mutants ofY. pseudotuberculosisYPIII,Y. pestisKIM/D27 (pigmentation locus minus), andY. pestisCO92 with reduced virulence. Mice injected via the intraperitoneal route with 5 × 10⁷CFU of the ΔlppKIM/D27 mutant survived a month, even though this would have constituted a lethal dose for the parental KIM/D27 strain. Subsequently, these ΔlppKIM/D27-injected mice were solidly protected against an intranasally administered, highly virulentY. pestisCO92 strain when it was given as five 50% lethal doses (LD₅₀). In a parallel study with the pneumonic plague mouse model, after 72 h postinfection, the lungs of animals infected with wild-type (WT)Y. pestisCO92 and given a subinhibitory dose of levofloxacin had acute inflammation, edema, and masses of bacteria, while the lung tissue appeared essentially normal in mice inoculated with the Δlppmutant of CO92 and given the same dose of levofloxacin. Importantly, while WTY. pestisCO92 could be detected in the bloodstreams and spleens of infected mice at 72 h postinfection, the Δlppmutant of CO92 could not be detected in those organs. Furthermore, the levels of cytokines/chemokines detected in the sera were significantly lower in animals infected with the Δlppmutant than in those infected with WT CO92. Additionally, the Δlppmutant was more rapidly killed by macrophages than was the WT CO92 strain. These data provided evidence that the Δlppmutants of yersiniae were significantly attenuated and could be useful tools in the development of new vaccines.