Figure 3 In vivo gene expression at 12
h (A), 24 h (B), and 36 h (C) relative to the highest level of expression in vitro by real-time PCR analysis. Total bacterial RNA extracted from strain ZY05719 grown in LB broth media was used as the template to assay the in vitro expression levels of the 10 newly identified genes. Temsirolimus cell line SPF minipigs were employed as model to study the in vivo expression levels. Pigs were inoculated intravenously with strain ZY05719, and bacterial cells recovered from blood at 12 h, 24 h, and 36 h post-inoculation were considered as in vivo growth bacteria. Total bacterial RNAs extracted from in vivo growth bacterial cells were further analyzed by real-time PCR. To determine whether RNA expression level
is induced or upregulated under in vivo conditions, we compared in vivo gene expression with the highest level of expression in vitro. The standard deviations are presented from three pigs each, blood collected at 12, 24 and 48 h. 1, ss-1616; 2, trag; 3, nlpa; 4, srt; 5, cwh; 6, hprk; 7, ysirk; 8, ss-1955; 9, sdh; 10, ss-1298; gapdh was used as reference gene. Location of the IVI genes on the SS2 chromosome To learn about location of the 48 IVI genes on the SS2 chromosome, we used BLAST to identify them in the S. suis strain P1/7 genomic sequence (genomic sequence data were generated by the S. suis strain P1/7 Sequencing Group at the Sanger Institute, and can be obtained from ftp://ftp.sanger.ac.uk/pub/pathogens/ss/.
Thirty-eight IVI genes were located (data not shown). Four genes (trag, exc-b, lac, and ppc) did not have high homology with CHIR-99021 solubility dmso P1/7, but demonstrated homology with strains S. suis 89/1591, 98HAH33, and 05ZYH33. The remaining six genes could not be located because their sequences were short and 3-mercaptopyruvate sulfurtransferase did not show high homology with any other sequence in the database. click here Pathogenicity islands (PAIs) are clusters of genes that may contribute to virulence in pathogens, sometimes by responding to environmental signals [25, 26]. Wei et al. (2006) predicted eight possible SS2 pathogenicity islands based on a systematic analysis of the SS2 strain P1/7 genomic sequence [27]. In this study, five IVI genes (sdh, srt, ss-1955, ss-1829, and ss-802) were found to be distributed in four pathogenicity islands (Figure 4) when located on the SS2 chromosome. Figure 4 Graphical representation of the locations of five IVI genes on the pathogenicity islands of S. suis serotype 2 strain P1/7. Based on a complete analysis of the SS2 reference strain P1/7 genomic sequence, W. Wei et al. predicted eight putative pathogenicity islands (PAIs). When we determined the locations of the 48 IVI genes identified by IVIAT, we found five IVI genes (sdh, ss-1955, srt, ss-1829, and ss-802) located in four pathogenicity islands in SS2 reference strain P1/7. The genomic map was published by W. Wei et al., 2006 (gray bars the third ring represent eight possible pathogenicity islands).