Yersinia pestis pFra shows biovar-specific differences and recent common ancestry with a Salmonella enterica serovar Typhi plasmid

Abstract

Population genetic studies suggest that Yersinia pestis, the cause of plague, is a clonal pathogen that has recently emerged from Yersinia pseudotuberculosis. Plasmid acquisition is likely to have been a key element in this evolutionary leap from an enteric to a flea-transmitted systemic pathogen. However, the origin of Y. pestis-specific plasmids remains obscure. We demonstrate specific plasmid rearrangements in different Y. pestis strains which distinguish Y. pestis bv. Orientalis strains from other biovars. We also present evidence for plasmid-associated DNA exchange between Y. pestis and the exclusively human pathogen Salmonella enterica serovar Typhi.

FIG. 1

IS100-associated deletion in pFra of Y. pestis CO-92 compared with Y. pestis KIM5 pMT1 (GenBank accession no. AF053947) and S. enterica serovar Typhi CT18 plasmid pHCM2. A region of 6,668 bp present in KIM5 is shown above the corresponding region in CO-92, which lacks this insertion. Thick black lines above and below the two Y. pestis plasmids and linked to them by dotted lines indicate the regions of pHCM2 showing more than 95% identity to the adjacent Y. pestis sequence. Locations of PCR primers for the reactions shown in Fig. 2 are indicated by arrowheads. Numbers shown at an angle to the horizontal correspond to base-pair locations in the labeled plasmid. In one KIM5 sequence (GenBank accession no. AF074611), the regions shown on either side of this IS100 copy are separated by 24 kb due to IS100 recombination.

FIG. 2

PCR for IS100-flanking regions from Y. pestis KIM5 pMT1 (bv. Medievalis) showing absence in Y. pestis bv. Orientalis strains and presence in other biovar Medievalis and Antiqua strains. DNA fragment sizes are given in base pairs. (A) PCR with primers P121 and P221. (B) PCR with primers P226 and P126. Lanes: 1, strain A1122; 2, PEXU2; 3, Harbin 35; 4, PB6; 5, F361/66; 6, Nepal 516; 7, 15–91; 8, 16–34; 9, AZ94-0666; 10, ZE94-2122; 11, TWS; 12, CO-92; 13, GB; 14, JAVA 9; 15, 195P; C, KIM (control); M, molecular weight marker. Additional text labels indicate lanes containing DNA templates from Y. pestis bv. Antiqua (An) and bv. Medievalis (Me) strains. All other numbered lanes contain DNA from Y. pestis bv. Orientalis strains. A 0.9% agarose gel was used. The digital image was obtained with GelDoc (Bio-Rad) and was cropped and resized in Adobe Photoshop.

FIG. 3

Comparison of the Y. pestis CO-92 pFra plasmid (A) with Y. pestis KIM5 pMT1 (GenBank accession no. AF053947) (B) and S. enterica serovar Typhi CT18 pHCM2 (C). Numbers on the horizontal scales are base pairs. Numbers on the vertical scale correspond to the plasmid GC percentage determined on a 200-bp window shown graphically along the top of each panel. The three numbers shown in each panel correspond to the mean percent GC for the whole plasmid and regional maxima and minima. Each panel contains the same features in the following order below the GC percentage graph. (i) Selected ORFs are shown as labeled blocks. Those transcribed to the right are shown above those transcribed to the left. ORF block shades are coded as follows: black, insertion sequence or phage related; dark gray, plasmid or DNA replication; light gray, non-DNA metabolism; white, virulence related. ORF abbreviations: DNA pol, DNA polymerase III alpha subunit; int, phage integrase; caf1R, F1 capsular operon and regulatory genes; ymt, murine toxin. (ii) Arrowheads corresponding to the location of E. coli Chi sequences are shown. Those pointing to the right are 5′-GCTGGTGG-3′, and those to the left are 5′-CCACCAGC-3′. (iii) Clear blocks in panels A and B correspond to regions of the Y. pestis plasmid sequence with sequence identity to S. enterica serovar Typhi pHCM2. The grey shading between the clear blocks corresponds to regions of the Y. pestis plasmids with no sequence identity to pHCM2. In panel C, the clear blocks represent regions of S. enterica serovar Typhi pHCM2 with significant sequence identity to Y. pestis (pFra and pMT1). The dark gray blocks represent regions with no sequence identity with Y. pestis pFra/pMT1. This figure is based on Artemis (63) views of individual annotated plasmid sequences arranged in a composite image using Adobe Photoshop.