Location: Meat Safety and QualityTitle: Identification of phosphoglucomutase as an enteropathogen growth stimulating factor
|Bosilevac, Joseph - Mick|
|RICHARDSON, ANDREW - Paradigm Diagnostics, Inc|
|OLSTEIN, ALAN - Paradigm Diagnostics, Inc|
Submitted to: Nutrition Research and Food Science Journal
Publication Type: Research Notes
Publication Acceptance Date: 5/4/2021
Publication Date: 5/17/2021
Citation: Bosilevac, J.M., Richardson, A.J., Olstein, A.D. 2021. Identification of phosphoglucomutase as an enteropathogen growth stimulating factor. Nutrition Research and Food Science Journal. 4(1):1-12.
Interpretive Summary: In order to isolate pathogens like E. coli and Salmonella contaminating foods, a food sample must be enriched in media to get the pathogen to grow to a level that can be detected. After noticing pathogens reached higher levels in meat samples compared to spinach samples, which had the same amount of pathogen to start, we investigated what was responsible. After experiments to isolate what was increasing pathogen growth, the protein phosphoglucomutase was identified. Results show that adding this protein to enrichment media can speed detection of pathogen contamination.
Technical Abstract: A highly selective Salmonella and Shiga toxin-producing E. coli (STEC) enrichment medium broth (SSS; commercially known as PDX-STEC), in comparative studies of low level E. coli O157:H7 inoculated ground beef and spinach, showed a 50- to 100- fold increase in STEC recoveries of the pathogen from ground beef compared to spinach enrichments. These observations suggested that either a soluble component of spinach inhibited the growth of the E. coli O157:H7 or a soluble component of ground beef stimulated the growth of the pathogen. The growth stimulating effect was linked to a soluble component of ground beef by comparing the growth of STEC and Salmonella in SSS conditioned with ground beef by passive extraction to their growth in control SSS containing traditional powdered beef extract media supplement. Then attempts were made to isolate and identify the responsible compound(s). A 20 to 60% ammonium sulfate fraction of ground beef extract maintained the growth stimulation of STEC and Salmonella. Further purification using affinity chromatography and preparative polyacrylamide gel electrophoresis identified three specific protein bands (52kD, 35kD and 20kD) associated with the growth stimulating activity. Mass spectral analysis of the trypsin-digested peptides of these proteins provided a putative identification of the proteins as the glycolytic protein, phosphoglucomutase (E.C. 22.214.171.124). Finally, commercially prepared rabbit muscle phosphoglucomutase (PGM) was shown to have the same growth stimulating activity thereby confirming the identity of the active protein. The possible mechanisms of growth stimulation by PGM may be through increasing bacterial fitness and environmental adaptability. Inclusion of PGM in food safety test protocols can enhance detection and isolation of contaminating STEC and Salmonella.