UVM Theses and Dissertations
Format:
Print
Author:
Petrova, Vera K.
Dept./Program:
Animal, Nutrition and Food Sciences Program
Year:
2008
Degree:
Ph. D.
Abstract:
Preventing illnesses resulting from consumption of contaminated food products is a major food safety objective. It has been estimated that each year one in four Americans has a significant foodborne illness. Listeria monocytogenes, a foodborne pathogen with the highest mortality rate, is found widely in foods, mostly in dairy products, meat, poultry, and seafood products. To prevent cases of product contamination with L. monocytogenes, it is essential to find sensitive and suitable methods for identification and isolation of the organism. This dissertation focuses on investigations of detection and differentiation approaches for L. monocytogenes and other foodborne microorganisms. More specifically, the focus of this work was.
1) to investigate the use of Flow Cytometry (FCM) for assessment of dead, viable and injured L. monocytogenes cells after their exposure to heat and acid stresses; 2) to perform a comparative analysis of six procedures (a PCR based detection method (BAX); a visual-immuno-assay (VIA); a visual-immunoprecipitate method (VIP); 3M[superscript TM] Petrifilm[superscript TM] Environmental Listeria Plates (Petrifilm); the USDNFSIS standard cultural procedure (USDA); and a modified USDAIFSIS procedure (mUSDA)) for detection of Listeria spp. in environmental samples; and 3) to assess the effect of chemical interferents, such as food matrices, different sanitizing compounds and growth media, on the ability of Fourier Transform Infrared (FTIR) spectroscopy to accurately identify and classify foodborne microorganisms.
We demonstrated that the assessment of the environmental stress and physiological heterogeneity of Listeria populations can be successfully done with use of differential dyes and molecular probes. The results of the method comparison study revealed that the highest levels of identification were achieved by the procedure incorporating a nonselective pre-enrichment period (mUSDA), which indicates the importance of a repair step for resuscitation of stressed cells in environmental samples. Investigations of FTIR spectroscopy demonstrated that 100% correct identification and differentiation of studied bacterial strains can be achieved in the presence of chemical interferents or food matrices, and a>99% identification rate in the presence of media matrices. This work thus provides crucial new insights in the monitoring systems for L. monocytogenes, knowledge that can assist the prevention and control of human infection by this notorious pathogen.
1) to investigate the use of Flow Cytometry (FCM) for assessment of dead, viable and injured L. monocytogenes cells after their exposure to heat and acid stresses; 2) to perform a comparative analysis of six procedures (a PCR based detection method (BAX); a visual-immuno-assay (VIA); a visual-immunoprecipitate method (VIP); 3M[superscript TM] Petrifilm[superscript TM] Environmental Listeria Plates (Petrifilm); the USDNFSIS standard cultural procedure (USDA); and a modified USDAIFSIS procedure (mUSDA)) for detection of Listeria spp. in environmental samples; and 3) to assess the effect of chemical interferents, such as food matrices, different sanitizing compounds and growth media, on the ability of Fourier Transform Infrared (FTIR) spectroscopy to accurately identify and classify foodborne microorganisms.
We demonstrated that the assessment of the environmental stress and physiological heterogeneity of Listeria populations can be successfully done with use of differential dyes and molecular probes. The results of the method comparison study revealed that the highest levels of identification were achieved by the procedure incorporating a nonselective pre-enrichment period (mUSDA), which indicates the importance of a repair step for resuscitation of stressed cells in environmental samples. Investigations of FTIR spectroscopy demonstrated that 100% correct identification and differentiation of studied bacterial strains can be achieved in the presence of chemical interferents or food matrices, and a>99% identification rate in the presence of media matrices. This work thus provides crucial new insights in the monitoring systems for L. monocytogenes, knowledge that can assist the prevention and control of human infection by this notorious pathogen.