Sample Preparation: The Forgotten Beginning

Advances in molecular technologies and automated instrumentation have provided many opportunities for improved detection and identification of microorganisms; however, the upstream sample preparation steps needed to apply these advances to foods have not been adequately researched or developed. Thus, the extent to which these advances have improved food microbiology has been limited. The purpose of this review is to present the current state of sample preparation, to identify knowledge gaps and opportunities for improvement, and to recognize the need to support greater research and development efforts on preparative methods in food microbiology. The discussion focuses on the need to push technological developments toward methods that do not rely on enrichment culture. Among the four functional components of microbiological analysis (i.e., sampling, separation, concentration, detection), the separation and concentration components need to be researched more extensively to achieve rapid, direct, and quantitative methods. The usefulness of borrowing concepts of separation and concentration from other disciplines and the need to regard the microorganism as a physicochemical analyte that may be directly extracted from the food matrix are discussed. The development of next-generation systems that holistically integrate sample preparation with rapid, automated detection will require interdisciplinary collaboration and substantially increased funding

Food microbiological analysis: new technologies/ Edit.: Mary Lou Tortorello

xii, 360 hal.: ill.; 21 cm

Food microbiological analysis: new technologies/ Edit.: Mary Lou Tortorello

xii, 360 hal.: ill.; 21 cm

Sample Preparation: The Forgotten Beginning

Advances in molecular technologies and automated instrumentation have provided many opportunities for improved detection and identification of microorganisms; however, the upstream sample preparation steps needed to apply these advances to foods have not been adequately researched or developed. Thus, the extent to which these advances have improved food microbiology has been limited. The purpose of this review is to present the current state of sample preparation, to identify knowledge gaps and opportunities for improvement, and to recognize the need to support greater research and development efforts on preparative methods in food microbiology. The discussion focuses on the need to push technological developments toward methods that do not rely on enrichment culture. Among the four functional components of microbiological analysis (i.e., sampling, separation, concentration, detection), the separation and concentration components need to be researched more extensively to achieve rapid, direct, and quantitative methods. The usefulness of borrowing concepts of separation and concentration from other disciplines and the need to regard the microorganism as a physicochemical analyte that may be directly extracted from the food matrix are discussed. The development of next-generation systems that holistically integrate sample preparation with rapid, automated detection will require interdisciplinary collaboration and substantially increased funding.This article is from Journal of Food Protection 72, no. 8 (2009): 1774–1789.</p

Roles of a novel Crp/Fnr family transcription factor Lmo0753 in soil survival, biofilm production and surface attachment to fresh produce of Listeria monocytogenes.

Listeria monocytogenes is a foodborne bacterial pathogen and the causative agent of an infectious disease, listeriosis. L. monocytogenes is ubiquitous in nature and has the ability to persist in food processing environments for extended periods of time by forming biofilms and resisting industrial sanitization. Human listeriosis outbreaks are commonly linked to contaminated dairy products, ready-to-eat meats, and in recent years, fresh produce such as lettuce and cantaloupes. We identified a putative Crp/Fnr family transcription factor Lmo0753 that is highly specific to human-associated genetic lineages of L. monocytogenes. Lmo0753 possesses two conserved functional domains similar to the major virulence regulator PrfA in L. monocytogenes. To determine if Lmo0753 is involved in environmental persistence-related mechanisms, we compared lmo0753 deletion mutants with respective wild type and complementation mutants of two fully sequenced L. monocytogenes genetic lineage II strains 10403S and EGDe for the relative ability of growth under different nutrient availability and temperatures, soil survival, biofilm productivity and attachment to select fresh produce surfaces including romaine lettuce leaves and cantaloupe rinds. Our results collectively suggested that Lmo0753 plays an important role in L. monocytogenes biofilm production and attachment to fresh produce, which may contribute to the environmental persistence and recent emergence of this pathogen in human listeriosis outbreaks linked to fresh produce

Listeria monocytogenes Growth Kinetics in Milkshakes Made from Naturally and Artificially Contaminated Ice Cream

This study assessed the growth of Listeria monocytogenes in milkshakes made using the process-contaminated ice cream associated with a listeriosis outbreak in comparison to milkshakes made with artificially contaminated ice cream. For all temperatures, growth kinetics including growth rates, lag phases, maximum populations, and population increases were determined for the naturally and artificially derived contaminants at 5, 10, 15, and 25°C storage for 144 h. The artificially inoculated L. monocytogenes presented lower growth rates and shorter lag phases than the naturally contaminated populations at all temperatures except for 5°C, where the reverse was observed. At 25°C, lag phases of the naturally and artificially contaminated L. monocytogenes were 11.6 and 7.8 h, respectively. The highest increase in population was observed for the artificially inoculated pathogen at 15°C after 96 h (6.16 log CFU/mL) of storage. Growth models for both contamination states in milkshakes were determined. In addition, this study evaluated the antimicrobial effectiveness of flavoring agents, including strawberry, chocolate and mint, on the growth of the pathogen in milkshakes during 10°C storage. All flavor additions resulted in decreased growth rates of L. monocytogenes for both contamination states. The addition of chocolate and mint flavoring also resulted in significantly longer lag phases for both contamination states. This study provides insight into the differences in growth between naturally and artificially contaminated L. monocytogenes in a food product

Listeria monocytogenes growth kinetics in refrigerated ready-to-eat dips and dip components.

Refrigerated ready-to-eat (RTE) dips often have pH and water activity combinations conducive to the proliferation of foodborne pathogens, including Listeria monocytogenes. This study conducted product assessments of five refrigerated RTE dips: baba ghanoush, guacamole, hummus, pesto, and tahini, along with individual dip components including avocado, basil, chickpeas, cilantro, eggplant, garlic, and jalapeno pepper. Dips and dip components were inoculated with 2 log CFU/g of L. monocytogenes and stored at 10°C for 28 days. The pathogen was enumerated throughout storage and growth rates were determined using the DMFit program to compute the time required for L. monocytogenes to achieve a 1 log CFU/g increase in population. Survival and growth rates varied significantly between the refrigerated RTE dips and dip components assessed in this study. For dips, L. monocytogenes progressively decreased in baba ghanoush, pesto, and tahini. In contrast, the pathogen proliferated in both hummus and guacamole and the highest growth rate was observed in guacamole (0.34±0.05 log CFU/g per day) resulting in a 1 log CFU/g increase in population in 7.8 days. L. monocytogenes proliferated in all dip components with the exception of eggplant and garlic. The pathogen achieved the highest growth rate in chickpeas (2.22±1.75 log CFU/g per day) resulting in a computed 1 log CFU/g increase in only 0.5 days. Results from this study can aid in understanding how L. monocytogenes behaves in refrigerated RTE dips and dip components and data can be utilized in understanding product formulations and in risk assessments

Transcriptomic Response of Escherichia coli O157:H7 to Oxidative Stress▿ †

Chlorinated water is commonly used in industrial operations to wash and sanitize fresh-cut, minimally processed produce. Here we compared 42 human outbreak strains that represented nine distinct Escherichia coli O157:H7 genetic lineages (or clades) for their relative resistance to chlorine treatment. A quantitative measurement of resistance was made by comparing the extension of the lag phase during growth of each strain under exposure to sublethal concentrations of sodium hypochlorite in Luria-Bertani or brain heart infusion broth. Strains in clade 8 showed significantly (P < 0.05) higher resistance to chlorine than strains from other clades of E. coli O157:H7. To further explore how E. coli O157:H7 responds to oxidative stress at transcriptional levels, we analyzed the global gene expression profiles of two strains, TW14359 (clade 8; associated with the 2006 spinach outbreak) and Sakai (clade 1; associated with the 1996 radish sprout outbreak), under sodium hypochlorite or hydrogen peroxide treatment. We found over 380 genes were differentially expressed (more than twofold; P < 0.05) after exposure to low levels of chlorine or hydrogen peroxide. Significantly upregulated genes included several regulatory genes responsive to oxidative stress, genes encoding putative oxidoreductases, and genes associated with cysteine biosynthesis, iron-sulfur cluster assembly, and antibiotic resistance. Identification of E. coli O157:H7 strains with enhanced resistance to chlorine decontamination and analysis of their transcriptomic response to oxidative stress may improve our basic understanding of the survival strategy of this human enteric pathogen on fresh produce during minimal processing