Methods of Testing of Pathogens

  1. The FDA CVM cultures raw pet food samples in an enrichment broth which selects and enhances the growth of pathogens like Salmonella, while selectively killing microorganisms that would naturally compete with the pathogen and prevent its propagation.  The broth is then incubated for anywhere from 16 hours to 5 days, at up to 115.2⁰F (this is a very wide range for a scientific analysis).  Scientists are unable to differentiate live cells from dead cells.  Therefore, it’s likely that samples with large amounts of dead cells would still result in a “false positive” for pathogens. (44) FDA guidance advises, “you are cautioned that significant deviations between actual microbiological data in specific products and the predictions do occur.” (45) “Time and Temperature abuse at one step alone might not result in an unsafe product. However…abuse that occurs at successive processing steps (including storage steps) might be sufficient to result in unsafe levels of pathogenic bacteria or toxins.” The document also states that pathogens are only likely to grow in raw food products where “(pathogenic) growth is not prevented by a condition of the food” (such as fermentation, ozone, essential oils, naturally occurring competitive bacteria, freezing/refrigeration, etc). FDA acknowledges that “the preventative measures that can be applied for pathogenic bacteria growth and toxin formation due to time and temperature abuse include 1) refrigeration of the product and controlling refrigeration temperatures 2) proper icing of the product 3) controlling the amount of time that the product is exposed to temperatures that would permit pathogenic bacteria growth or toxin production 4) rapid cooling of the product.” Ultimately, they acknowledge that under normal handling conditions, raw foods are not likely to accumulate pathogens. They also admit that “pathogenic bacteria also could be introduced during processing, even after cooking. However, in most cases it is not reasonable to assume that they will fully prevent the introduction of pathogenic bacteria.” (45)(46) This indicates that they are fully aware that even cooked kibble and sterilized products can harbor high levels of pathogens.

Dulcitol Fermentation Test – http://www.vumicro.com/vumie/help/VUMICRO/Dulcitol_Fermentation_Test.htm

ONPG Test (for ?-galactosidase): Principle, procedure and results: https://microbeonline.com/onpg-test-%CE%B2-galactosidase-principle-procedure-results/

Malonate Broth for the identification of enteric bacteria – https://catalog.hardydiagnostics.com/cp_prod/content/hugo/malonatebroth.htm

Gelatin Hydrolysis Test: Principle, Procedure and expected results – https://microbeonline.com/gelatin-hydrolysis-test-principle-procedure-expected-results/

Sorbitol Fermentation Test – http://www.vumicro.com/vumie/help/VUMICRO/Sorbitol_Fermentation_Test.htm

Growth in KCN – http://www.vumicro.com/vumie/help/VUMICRO/KCN_Growth_Test.htm

Toward pectin fermentation by Saccharomyces cerevisiae: expression of the first two steps of a bacterial pathway for D-galacturonate metabolism – PubMed – NCBI – https://www.ncbi.nlm.nih.gov/pubmed/23079077

Gamma-Glutamyl Transferase (GGT) – Understand the Test & Your Results – https://labtestsonline.org/tests/gamma-glutamyl-transferase-ggt

Abnormalities in liver enzyme levels during Salmonella enteritidis enterocolitis. PubMed – NCBI – https://www.ncbi.nlm.nih.gov/pubmed/15449987

MUG (beta-Glucoronidase) test for rapid identification of e.coli – https://microbeonline.com/mug-test-b-glucuronidase-test-for-rapid-identification-of-e-coli/

Enzyme Characteristics of ?-d-Galactosidase- and ?-Glucoronidase – Positive Bacteria and Their Interference in Rapid Methods for Detection of Waterborne Coliforms and Escherichia coli – https://www.ncbi.nlm.nih.gov/pmc/articles/PMC106360/

Salicin Fermentation Test – http://www.vumicro.com/vumie/help/VUMICRO/Salicin_Fermentation_Test.htm

Mucate Broth – http://www.himedialabs.com/TD/M1226.pdf

The galacturonate TRAP Transporter GaaPQM of Agrobacterium tumefaciens and its involvement in virulence gene expression – https://aem.asm.org/content/early/2015/12/03/AEM.02891-15 

Malonate Inhibition of Oxidation in the Krebs Tricarboxylic Acid Cycle – http://www.jbc.org/content/178/1/241.full.pdf

Utilization of d-tartaric acid by Salmonella paratyphi B and Salmonella java: comparison of anaerobic plate test, lead acetate test and turbitidy test – https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2129498/

Diagnostic and Public Health Dilemma of Lactose-Fermenting Salmonella enterica Serotype Typhimurium in Cattle in the Northeastern United States – https://www.ncbi.nlm.nih.gov/pubmed/10699026

Lysis Timing and Bacteriophage Fitness – https://www.ncbi.nlm.nih.gov/pubmed/16219778

Phage lysis: three steps, three choices, one outcome – PubMed – NCBI – https://www.ncbi.nlm.nih.gov/pubmed/24585055

Bacteriophages lytic for Salmonella rapidly reduce Salmonella contamination on glass and stainless steel surfaces – https://www.ncbi.nlm.nih.gov/pubmed/24228226

Improved lysis efficiency and immunogenicity of Salmonella ghosts mediated by co-expression of phage – https://www.nature.com/articles/srep45139

Beneficial effect of Salmonella typhimurium infection and of immmunoglobulins from S. typhimurium-infected mice on the autoimmune disease of (NZB x NZW) F1 Mice – https://www.ncbi.nlm.nih.gov/pubmed/8625513

Beneficial effects of live and dead Salmonella-based vector strain on the course of colitis in mice – https://www.ncbi.nlm.nih.gov/pubmed/8625513

Salmonella bacteria used to stop viruses – https://newatlas.com/salmonella-bacteria-used-to-stop-viruses/17827/

HACCP Validation Resources – https://www.fsis.usda.gov/wps/portal/fsis/topics/regulatory-compliance/haccp/resources-and-information/haccp-validation/haccp-validation-resources

Methods to Reduce/ Eliminate Pathogens from Fresh and Fresh-Cut Produce – https://onlinelibrary.wiley.com/doi/abs/10.1111/j.1541-4337.2003.tb00033.x

Department of Agriculture Pathogen Reduction: HACCP Systems Final Rule, Part II – https://www.fsis.usda.gov/wps/wcm/connect/e113b15a-837c-46af-8303-73f7c11fb666/93-016F.pdf?MOD=AJPERES

Compendium of analytical laboratory methods for food and feed safety

Detection of salmonella spp by a loop-mediated isothermal amplification (LAMP)

PmrAB a Two-component regulatory system of pseudomonas aeruginosa that modulates…

Biofilm Formation and Detachment in Gram-Negative Pathogens is Modulated by Select Bile Acids – Laura M Sanchez, Andrw T. Cheng, and Roger G. Lining

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