ABSTRACT The aim of this work was to determine the bacteriological status of selected RTE foods, associated preparation surfaces and cleaning tools sampled from four retail delicatessens in Johannesburg. An initial pilot study of four RTE foods such as filled baguettes, assorted salads, sliced processed meats and hot meals was conducted in order to set the parameters for the remainder of the study. Results showed that filled baguettes and assorted salads contained the highest bacterial counts and incidences of potential foodborne pathogens. Bacterial counts were obtained from the associated preparation surfaces, whilst cleaning tools were associated with coliform and Escherichia coli, suggesting they may harbour potential foodborne pathogens. For the main study, 60% filled baguettes and assorted salads complied with the microbiological guidelines recommended by the retailer, however fruit salads had the lowest bacterial counts overall. Furthermore, of the food contact surfaces plastic chopping boards were identified as the greatest reservoir for RTE food contamination. Bacteriological analysis in conjunction with scanning electron microscopy showed potential foodborne pathogens associated with cleaning tools. Low numbers of aerobic bacteria and Staphylococcus aureus were associated with disposable plastic gloves, suggesting that good glove practices are used by the food handlers.

Master of Science / Food Science Institute -- Animal Science & Industry / Daniel Y.C. Fung / In February 2008, the FDA released a draft Compliance Policy Guide (CPG) on Listeria monocytogenes and proposed that ready-to-eat (RTE) foods that do not support the growth of L. monocytogenes may contain up to 100 CFU/g of this pathogen. Frozen foods such as ice cream fall in that category since they are consumed in the frozen state. However, other frozen foods, such as vegetables and seafood that are thawed and served at salad and food bars, may support the growth of Listeria and would not be allowed to contain 100 CFU/g according to the draft CPG. In the current study, growth curves were generated for L. monocytogenes inoculated onto four thawed frozen foods - corn, green peas, crabmeat, and shrimp - stored at 4, 8, 12, and 20ºC. Growth parameters, lag phase duration (LPD), and exponential growth rate (EGR) were determined using a two-phase linear growth model and the Square Root Model. The results demonstrated that L. monocytogenes has a very short LPD on these thawed frozen foods during refrigerated storage and that there would be several orders of magnitude of growth (i.e., more than 1.7 log increase at 4 ºC) of the organism before the product is found to be organoleptically unacceptable. Although it would not be possible to take advantage of any extended lag phase duration caused by freeze injury to the organism, frozen foods containing less than 100 CFU/g of L. monocytogenes that are thawed, or thawed and cooked, and then consumed immediately, should not represent a public health hazard.

Listeriosis caused by Listeria monocytogenes (L. monocytogenes) has been of public concern since the 1980s. Among all the RTE food, deli meats are the major carrier for this pathogen. Eliminating or lowering the initial level of L. monocytogenes in RTE meat and poultry product in the plants is an important practice in reducing the risk of L. monocytogenes to the public due to the growth potential of L. monocytogenes in the RTE food product during storage. Research identifying the contamination at plants provided information for the Food Safety and Inspection Service (FSIS) to establish the Interim Final Rule, requiring the food processing plants that produce post-lethality exposed RTE meat and poultry product choose one of the three alternative plans to ensure good sanitation conditions during food processing or suppress the growth of L. monocytogenes during storage: post-processing treatment and use of growth inhibitor (Alternative1), post-processing alone (Alternative 2a) or use of growth inhibitor and sanitation program (Alternative 2b), and sanitation program alone (Alternative 3). This research developed a comprehensive model that simulated the entire processes of RTE food production, taking into account potential transfer and growth of L. monocytogenes in RTE meat and poultry products. This plant-to-consumption model analyzed the effectiveness of the three alternative processes on reducing the L. monocytogenes in the RTE food products and also investigated the optimal sampling and sanitizing program. Results showed that formulation of food products with growth inhibitor has the greatest impact on reducing the risk of L. monocytogenes, followed by the post-processing treatment and sanitation intervention. Risk can also be reduced depending on alternatives. For example, 70% reduction if all are switched to alternative 2b and 91% reduction if all are switched to Alternative 1, compared with the current alternative selection by food establishments. This study investigated several important factors in the sanitation program, analyzed the sensitivities of these factors, and proposed the reasonable improvement of the hold-and-test strategies by the plant-to-consumer mathematic model. Holding all the lots during the food contact surface (FCS) testing period instead of holding lots after finding the positive FCS would increase the detection rate of positive lots by three "times. These results may help the food establishments under Alternative 3 choosing the proper sampling and sanitation program or switching to Alternative 1 or 2. / Ph. D.

A presente dissertação, apresentada em dois estudos, buscou verificar as condições de boas práticas de higiene e manipulação (BPHM) e de infraestrutura (IE) de oito restaurantes/lanchonetes e de sete pontos de comércio ambulante localizados na Cidade Universitária Armando de Salles Oliveira (CUASO-USP) e a qualidade higiênico-sanitária de 45 amostras de alimentos prontos para o consumo colhidas nos mesmos. O primeiro estudo traz uma abordagem exploratória e qualitativa através da aplicação de listas de verificação; o segundo analisa laboratorialmente aspectos higiênico-sanitários dos alimentos comercializados pelos estabelecimentos alvo do estudo anterior. Concluiu-se que 1) os estabelecimentos de comércio alimentício da CUASO-USP apresentaram índices regulares de cumprimento de BPHM e adequação de IE, sendo que o eixo de higiene e manipulação de alimentos mostrou-se em melhor situação quando comparado ao de infraestrutura; 2) os ambulantes analisados apresentaram melhores resultados no cumprimento das normas de BPHM e IE e na avaliação das condições higiênico-sanitárias quando comparados aos estabelecimentos fixos de comércio alimentício. Constatou-se que é possível a prática do comércio de alimentos de rua com qualidade higiênico-sanitária, sem caracterizar uma ameaça à saúde publica, desde que o empreendedor conheça e aplique os procedimentos necessários e críticos à obtenção da garantia dos produtos comercializados, assumindo responsabilidade social ao realizar o seu modo de produção mercantil simples, porém comprometido moralmente com a sociedade. / This investigate, presented in two studies, examined the good hygiene practices and handling (GHPH) and the infrastructure (IS) conditions from eight restaurants and seven street vendors located in University City Armando de Salles Oliveira (UCASO-USP) and the sanitary conditions of 45 samples of ready to eat food harvested in them. The first study provides an exploratory and qualitative approach through the application of checklists; the second laboratory analyzes the hygienic and sanitary aspects of the food sold by the previously discussed establishments. We concluded that 1) the UCASO-USP food trade establishments showed regular compliance rates of GHPH and appropriateness of IS, and the axis of hygiene and food handling proved to be better off compared to the infrastructure; 2) the street vendors analyzed showed better results in meeting the GHPH and IS standards and in the evaluation of sanitary conditions when compared to fixed establishments of food trade. It was found that it is possible to practice the street foods trade in sanitary conditions, without characterizing a public health threat, provided that the enterprising know and apply the necessary and critical procedures to obtaining the products marketed assurance, with social responsibility while make their commodity production simple mode.

A presente dissertação, apresentada em dois estudos, buscou verificar as condições de boas práticas de higiene e manipulação (BPHM) e de infraestrutura (IE) de oito restaurantes/lanchonetes e de sete pontos de comércio ambulante localizados na Cidade Universitária Armando de Salles Oliveira (CUASO-USP) e a qualidade higiênico-sanitária de 45 amostras de alimentos prontos para o consumo colhidas nos mesmos. O primeiro estudo traz uma abordagem exploratória e qualitativa através da aplicação de listas de verificação; o segundo analisa laboratorialmente aspectos higiênico-sanitários dos alimentos comercializados pelos estabelecimentos alvo do estudo anterior. Concluiu-se que 1) os estabelecimentos de comércio alimentício da CUASO-USP apresentaram índices regulares de cumprimento de BPHM e adequação de IE, sendo que o eixo de higiene e manipulação de alimentos mostrou-se em melhor situação quando comparado ao de infraestrutura; 2) os ambulantes analisados apresentaram melhores resultados no cumprimento das normas de BPHM e IE e na avaliação das condições higiênico-sanitárias quando comparados aos estabelecimentos fixos de comércio alimentício. Constatou-se que é possível a prática do comércio de alimentos de rua com qualidade higiênico-sanitária, sem caracterizar uma ameaça à saúde publica, desde que o empreendedor conheça e aplique os procedimentos necessários e críticos à obtenção da garantia dos produtos comercializados, assumindo responsabilidade social ao realizar o seu modo de produção mercantil simples, porém comprometido moralmente com a sociedade. / This investigate, presented in two studies, examined the good hygiene practices and handling (GHPH) and the infrastructure (IS) conditions from eight restaurants and seven street vendors located in University City Armando de Salles Oliveira (UCASO-USP) and the sanitary conditions of 45 samples of ready to eat food harvested in them. The first study provides an exploratory and qualitative approach through the application of checklists; the second laboratory analyzes the hygienic and sanitary aspects of the food sold by the previously discussed establishments. We concluded that 1) the UCASO-USP food trade establishments showed regular compliance rates of GHPH and appropriateness of IS, and the axis of hygiene and food handling proved to be better off compared to the infrastructure; 2) the street vendors analyzed showed better results in meeting the GHPH and IS standards and in the evaluation of sanitary conditions when compared to fixed establishments of food trade. It was found that it is possible to practice the street foods trade in sanitary conditions, without characterizing a public health threat, provided that the enterprising know and apply the necessary and critical procedures to obtaining the products marketed assurance, with social responsibility while make their commodity production simple mode.

[ES] La seguridad alimentaria es una prioridad para la población y en la actualidad cobra mayor importancia por ciertas tendencias alimentarias como el consumo de alimentos crudos y la distribución generalizada de alimentos orgánicos, que pueden ser la causa de enfermedades transmitidas por alimentos. Para garantizar la seguridad alimentaria, la detección de estos microorganismos debe realizarse de manera rápida y eficiente. Par eso, el método de cultivo microbiológico se considera el oficial para la detección de estos patógenos. Sin embargo, adolece de importantes inconvenientes, ya que no solo requiere mucho tiempo, sino que también es laborioso y consume muchos recursos. Además, puede ser limitado con respecto a la detección de bacterias fisiológicamente alteradas y/o estresadas durante el almacenamiento y la conservación. En este trabajo se ha desarrollado un protocolo sencillo y rápido para la detección simultánea de E. coli, L. monocytogenes, S. aureus y S. enterica en alimentos, mediante la combinación de una etapa de co-cultivo en medio líquido y la detección por PCR múltiple. Se ha evaluado la eficiencia de varios medios de enriquecimiento y se seleccionó el agua de peptona tamponada como el medio óptimo para el co-cultivo de las cuatro bacterias diana. También se optimizaron las condiciones de PCR múltiple y se aplicaron tanto a co-cultivos como a muestras de alimentos inoculados artificialmente, lechuga orgánica y carne picada. Después de la optimización, la PCR múltiple desarrollada fue capaz de detectar las cuatro bacterias simultáneamente, hasta con una inoculación inicial de 10^0 UFC/mL. En presencia de ambas matrices alimentarias inoculadas, tras la etapa de co-cultivo, la PCR múltiple pudo detectar simultáneamente las 3 bacterias E. coli, S. enterica y L. monocytogenes, mientras que S. aureus se ha detectado por PCR simplex, a partir del mismo extracto de ADN del co-cultivo. Los resultados obtenidos permiten concluir que el uso de un paso de co-cultivo en Agua peptona tamponada, antes de la detección por PCR simple y múltiple, puede facilitar la detección simultánea de las cuatro bacterias potencialmente presentes en las matrices alimentarias. La presencia o ausencia de la bacteria diana en los alimentos se confirma en unas 30 horas, lo que reduce el tiempo requerido para la detección en comparación con el tiempo mínimo de 7 días por método cultural. Asimismo, permite reducir el número de medios de cultivo y reactivos, para el aislamiento e identificación de bacterias que no son detectadas por PCR y que no están presentes en las matrices alimentarias, lo que supone un importante ahorro económico. / [CA] La seguretat alimentària sempre és una prioritat per a la població i en l' actualitat cobra major importància per certes tendències alimentàries, com el consum d' aliments crus i la distribució generalitzada d' aliments orgànics, que poden ser la causa de malalties transmeses per aliments. Per garantir la seguretat alimentària, la detecció d' aquests microorganismes s' ha de realitzar de manera ràpida i eficient. Per a això, el mètode de cultiu microbiològic es considera l' oficial per a la detecció d' aquests patògens. Però, hi ha importants inconvenients, ja que no només requereix més temps, sinó que també és laboriós i consumeix molts recursos. A més, pot ser limitat pel que fa a la detecció de bacteris fisiològicament alterats i/o estressats durant l'emmagatzematge i la conservació. En aquest treball s'ha desenvolupat un protocol senzill i ràpid per a la detecció simultània d' E. coli, L. monocytogenes, S. aureus i S. enterica en aliments, mitjançant la combinació d' una etapa de co-cultiu en medi líquid i la detecció per PCR múltiple. S'ha avaluat l'eficiència de diversos mitjans d'enriquiment i s'ha seleccionat l'aigua de peptona tamponada com el medi òptim per al co-cultiu dels quatre bacteris diana. També es van optimitzar les condicions de PCR múltiple i es van aplicar tant a co-cultius com a mostres d'aliments inoculats artificialment, enciam orgànic i carn picada. Després de l'optimització, la PCR múltiple desenvolupada va ser capaç de detectar els quatre bacteris simultàniament, fins a una inoculació inicial de 10^0 UFC/mL. En presència d' ambdues matrius alimentàries inoculades, després l' etapa de co-cultiu, la PCR múltiple va poder detectar simultàniament els 3 bacteris: E. coli, S. enterica i L. monocytogenes, mentre que S. aureus s' ha detectat per PCR simple, a partir del mateix extracte d' ADN del co-cultiu. Els resultats obtinguts permeten concloure que l' ús d' un pas de co-cultiu en Aigua de peptona tamponada, abans de la detecció per PCR simple i múltiple, pot facilitar la detecció simultània dels quatre bacteris potencialment presents en les matrius alimentàries. La presència o absència del bacteri diana en els aliments es confirma en unes 30 hores, la qual cosa redueix el temps requerit per a la detecció en comparació amb el temps mínim de 7 dies per mètode cultural. Així mateix, permet reduir el nombre de mitjans de cultiu i reactius, per a l' aïllament i identificació de bacteris que no són detectats per PCR i que no estan presents en les matrius alimentàries, la qual cosa suposa un important estalvi econòmic. / [EN] Food safety is a priority for the population and is nowadays more important than ever due to certain dietary trends such as the consumption of raw foods and the widespread distribution of organic foods, which may be the cause of foodborne diseases. To ensure food safety, the detection of these microorganisms must be done quickly and efficiently. Although, the microbiological culture method is considered to be the official method for the detection of these food-borne pathogens, it suffers from significant drawbacks, such as time-consuming, laborious and expensive, in addition it may be limited regarding the detection of physiologically altered and/or stressed bacteria, during storage and preservation. In this work has been developed a simple and rapid protocol for the simultaneous detection of E. coli, L. monocytogenes, S. aureus and S. enterica in food, by combining a liquid co-culture step and detection by multiplex PCR. The efficiency of several enrichment media was evaluated and buffered peptone water was chosen as the optimal medium for the co-culture of the four target bacteria. Then, optimized multiplex PCR conditions were applied to both the co-cultures and the samples of artificially inoculated foods, organic lettuce and ground meat. After optimization, the developed multiplex PCR was able to simultaneously detect the four bacteria, up to an initial inoculation of 10^0 CFU/mL. In the presence of the two inoculated food matrices, after a co-culture step, the multiplex PCR could simultaneously detect the 3 bacteria: E. coli, S. enterica and L. monocytogenes, whereas, S. aureus has been detected by simplex PCR, from the same co-culture DNA template. The results obtained allow conclusion that the use of a co-culture step in Buffered Peptone Water, before detection by simplex and multiplex PCR, can facilitate the simultaneous detection of the four bacteria potentially present in the food matrices. The presence or the absence of the target bacteria in food is confirmed in approximately 30 hours, which reduce the time required for the detection compared to the minimum time of 7 days by cultural method. Also, it allows to reduce the number of culture media and reagents, for the isolation and identification of bacteria that are not detected by PCR and which are not initially present in the food matrices, which represents a significant economic savings. / Boukharouba, A. (2022). Isolation and Identification of Foodborne Pathogens of Special Interest in Food Safety [Tesis doctoral]. Universitat Politècnica de València. https://doi.org/10.4995/Thesis/10251/182828