MOLECULAR CHARACTERIZATION OF PATHOGENS AND THEIR RESPONSES TO ENVIRONMENTAL FACTORS
Location: Molecular Characterization of Foodborne Pathogens
Title: Differential gene expression of Escherichia coli O157:H7 in ground beef extract compared to tryptic soy broth
Submitted to: Journal of Food Science
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: September 30, 2010
Publication Date: February 1, 2011
Citation: Fratamico, P.M., Yan, X., Li, Y. 2011. Differential gene expression of Escherichia coli O157:H7 in ground beef extract compared to tryptic soy broth. Journal of Food Science. 76:79-87.
Interpretive Summary: Escherichia coli O157:H7 is a bacterial food-borne pathogen that causes serious human illness. Cattle are an important reservoir for this pathogen, and ground beef is a common vehicle of human infection. To control the growth of E. coli O157:H7 in meat, it is important to understand which genes play a role in its growth and survival. Studies examining gene expression of bacteria under different conditions are generally performed in non-complex laboratory media, since these types of analyses are difficult to perform in complex matrices such as food. The behavior of E. coli O157:H7 can vary considerably in laboratory medium compared to food. Therefore, gene expression of E. coli O157:H7 in a laboratory medium was compared to that in a ground beef model system (ground beef extract, GBE) using DNA microarrays (glass slides to which regions of genes complementary to all of the genes found in E. coli O157:H7 are bound). There were a number of differences in gene expression of E. coli O157:H7 in GBE compared to the laboratory medium. Genes that have a role in various cellular functions of E. coli O157:H7 were affected. This study demonstrated that microarray analyses can be performed with bacteria in complex food matrices, and gene expression of E. coli O157:H7 differs in laboratory medium compared to GBE. The information will be useful for identification of genes that can be employed as potential targets for interventions to control E. coli O157:H7 in meat.
Escherichia coli O157:H7 causes sporadic cases and outbreaks of hemorrhagic colitis and hemolytic uremic syndrome, and ground beef is a common vehicle of infection. DNA microarrays have been used for transcriptomic studies of E. coli O157:H7 using laboratory media; however, analysis of gene expression in complex matrices such as food are lacking. This study compared gene expression profiles of E. coli O157:H7 Sakai strain in raw ground beef extract (GBE) and tryptic soy broth (TSB). Total RNA was isolated from GBE and TSB after 2-h incubation with E. coli O157:H7. Following reverse transcription (RT) of the RNA, labeled cDNA was hybridized to microarrays spotted with 70-mer oligonucleotide probes representing 9608 open reading frames (Operon; Genome Array-Ready Oligo Set, Version 2.0) corresponding to four genomes of E. coli strains and three plasmids. There were 74 up-regulated and 54 down-regulated genes in E. coli O157:H7 grown in GBE compared to TSB (greater than two-fold increase or decrease), respectively. Up-regulated genes included genes involved in protein and polysaccharide biosynthesis, transcription factors, inner and periplasmic membrane transport proteins, and acid shock proteins. Down-regulated genes included genes encoding proteins for energy metabolism, biosynthesis of cofactors, transporters for degraded small molecules, amino acids, and peptides, transcription factors responsible for protein and macromolecule/small molecule degradation, and enzymes for protein degradation. The expression levels of 10 genes were confirmed by real-time RT-PCR. This study demonstrated that microarray analyses can be performed from bacteria in complex food matrices, and gene expression of E. coli O157:H7 differs in TSB compared to GBE. The information will be useful for identification of genes that can be employed as potential targets for interventions to control E. coli O157:H7.