Submitted to: National Center for Biotechnology Information (NCBI)
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 12/11/2006
Publication Date: 12/31/2006
Citation: Guard, J.Y., Morales, C. 2006. Evolutionary trends in two strains of Salmonella enterica subsp. I serovar Enteritidis PT13a that vary in virulence potential. National Center for Biotechnology Information (NCBI). Available at:http://www.ncbi.nlm.nih.gov/genomes/static/Salmonella_SNPS.html. Interpretive Summary: Researchers at the Egg Safety and Quality Research Unit (ESQRU) of the U. S. Department of Agriculture, Agricultural Research Service (USDA-ARS) have used microarray technology to identify a set of potential single nucleotide polymorphisms (SNPs) that distinguish two PT13a strains of S. Enteritidis that are genetically related but phenotypically divergent. Using the S. Enteritidis PT4 genomic sequence from the Sanger Institute as a starting point, a set of overlapping primers were generated and used in conjunction with Nimblegen Systems technology to detect SNPs that distinguish S. Enteritidis PT13a strain 21027 (bf), which forms biofilm-forming but does not contaminate eggs, from S. Enteritidis PT13a strain 21046 (wt), which contaminates eggs but does not form biofilm. BLAST comparisons of the SNPs with other Salmonella enterica annotated databases revealed potentially relevant phenotypic traits matched to genetic loci. Since this data is considered preliminary it should be used with caution. Version 3 includes results from BLAST searches and alignments to evaluate if a SNP occurred within a protein and if it altered amino acid sequence. Nimblegen states that comparative resequencing detects within 5% of true SNP content (+/- 10 SNPs). Mutational mapping previously detected 3 of 3 SNPs selected for confirmatory sequencing (rrlC, rrlA, cyaA); thus, mutational mapping was reliable for detection of SNPs. Initial resequencing parameters that were used to confirm version 3 SNPs missed selection of all three of the control sequences. However, these sites gave a substantial signal that moved them to the category referred to as “non-called ROI.” Therefore, SNPs detected by mutational mapping but that fell within the ROI category during resequencing require further analysis and will be added to the database as completed. SNP locations are subject to change if the genomic database for S. Enteritidis PT4 is revised by the Sanger Institute after May 22, 2006. Another 261 SNPs that were identified by mutational mapping remain to be sequenced.
Technical Abstract: Salmonella enterica subsp. I serovar Enteritidis (S. Enteritidis) is the world's leading cause of salmonellosis. Eggs that are contaminated by apparently healthy hens and that have been improperly cooked can result in illness in humans who consume them. Although the incidence of this pathogen within the United States has not been as high as it has been in Europe, current estimates from the Centers for Disease Control and from the USDA-Food Safety Inspection Service respectively suggest that this pathogen is causing more human illness and that it has begun to contaminate broilers in the States (Altekruse et al, 2006). Egg contamination studies in hens, analysis of cell surface properties and analysis of the global physiology of strains using phenotype microarrays have shown that some strains complement each other in the infection pathway that results in egg contamination (Morales et al, 2005). However, S. Enteritidis has also evolved dimorphism, which is the ability to exhibit multiple phenotypes from one genome in response to environmental conditions. To investigate the genetic basis for the emergence of dimorphism and phenotypic variation that alters virulence potential in the Salmonellae, the genomes of two variant strains of S. Enteritidis PT13a that were descended from a dimorphic parent over ten years were subjected to virtual subtraction hybridization. Direct comparison of the closely related genomes was possible by using the genomic database for S. Enteritidis PT4 NCTC 13349 (http://www.sanger.ac.uk/Projects/Salmonella/) to generate overlapping sets of primers (Nimblegen, Inc.) that were capable of detecting single nucleotide polymorphisms (SNPs). The table lists the location for 195 SNPs and the probable change in amino acid sequence if any. Of these 195 confirmed SNPs, 3 occurred in the virulence plasmid, 102 altered predicted amino acid sequences, and 33 occurred in either non-coding or ribosomal DNA regions. Two deletions were detected in wt strain 21046, which does not form biofilm but retains the ability to contaminate eggs. Three SNPs in bf strain 21027, which forms biofilm but does not contaminate eggs, removed stop codons, whereas two single base pair SNPs in the wt strain introduced stop codons. ZipA of wt strain 21046 accumulated numerous SNPs that altered amino acid sequence.