Assessment of risk factors affecting the microbial safety of leafy greens in Spain = Evaluación del riesgo microbiano en la producción y el procesado de hortalizas de hoja

Resumen La presente tesis esta destinada a proporcionar una vision general de los principales riesgos microbiologicos asociados a las hortalizas de hoja durante su produccion, cosecha y procesado, asi como los posibles microorganismos indicadores capaces de predecir o dar una estimacion de la posible contaminacion con microorganismos patogenos en hortalizas de hoja. La primera parte de esta tesis se centra en los principales factores que afectan a la seguridad microbiana de las hortalizas de hoja a nivel de produccion primaria. En el Capitulo 3 se evaluo el efecto de los factores agricolas y las condiciones meteorologicas en la seguridad microbiana de hortalizas de hoja en Espana. En este trabajo, se identificaron el suelo y el agua de riego como los factores mas importantes que afectan a la seguridad microbiana de la 'espinaca baby'. Respecto a los factores climaticos, se observo que la temperatura ambiente afectaba la probabilidad de la contaminacion de la 'espinaca baby'. En el Capitulo 4 se estudio el efecto de las precipitaciones extremas sobre la seguridad microbiana de hortalizas de hoja. En este sentido, se encontro que las muestras de suelo, el agua de riego y hortalizas de hoja tomadas de campos afectados por la inundacion estaban contaminados con altos niveles de E. coli y patogenos. Este estudio corroboro estudios previos que identificaban las inundaciones como importantes factores de riesgo para la contaminacion microbiana de hortalizas de hoja. Por otro lado, se puso de manifiesto que las condiciones climaticas, sobre todo, la radiacion solar juegan un papel importante reduccion de la supervivencia microbiana en el campo. Con los resultados obtenidos, en el Capitulo 5 se desarrollo un modelo de analisis cuantitativo de riesgos microbiologicos para la evaluacion de la exposicion microbiana de E. coli en 'espinaca baby' al nivel de produccion primaria en Espana. Este modelo se uso para evaluar el impacto potencial de factores climaticos y practicas agricolas en la distribucion de E. coli en el momento de la cosecha. Conjuntamente, los resultados confirmaron el impacto tanto de los factores climaticos como de las practicas agricolas en la contaminacion de las hortalizas de hoja con E. coli. Adicionalmente, se puso de manifiesto que la aplicacion de medidas preventivas dirigidas a dichos factores y practicas de riesgo podria reducir considerablemente el riesgo de contaminacion. Por lo tanto, el modelo resulto un metodo util para evaluar el impacto potencial de los diferentes factores de riesgo y las estrategias de intervencion que afectan a las concentraciones de E. coli en el campo. El Capitulo 6 se centra en los riesgos microbiologicos asociados al procesado de hortalizas de hoja. En este estudio, se obtuvo una vision global de la contaminacion microbiana en todas las operaciones que implica el procesado de hortalizas de hoja. A este respecto, se encontro que las muestras de agua obtenidas de la centrifuga eran positivas tanto para E. coli generica como para microorganismos patogenos, sugiriendo el posible origen de la contaminacion en el producto sometido a centrifugacion o en el agua de lavado. El Capitulo 7, se centra en el ultimo paso de la cadena del 'campo a la mesa': el consumidor. Primero, se evaluo la validez de los datos existentes referentes al consumo de frutas y hortalizas frescas en Espana y Belgica, llegando a la conclusion de que la informacion disponible sobre el consumo real y detallado de diferentes tipos de productos frescos asi como las practicas de manipulacion de los mismos era deficiente. Por lo tanto, se proporcionaron datos estandarizados para el consumo de productos frescos destinados a ser consumidos crudos o minimamente procesados. Estos datos podrian ser de gran utilidad en futuros estudios de evaluacion de la exposicion a diferentes riesgos tanto quimicos como microbiologicos con productos frescos listos para el consumo (por ejemplo, hortalizas de hoja). En el Capitulo 8, se discuten los principales resultados de esta tesis y se plantean nuevas perspectivas para futuras investigaciones. Summary The current thesis intended to provide an overview of the prevalence of the main microbial hazards commonly linked to leafy greens during production, harvesting and processing, as well as potential indicator microorganisms able to predict faecal contamination in leafy greens. In order to introduce the research area, a literature review of the relevant aspects and the latest publications related to safety of leafy greens from the cultivation field to consumption was presented in Chapter 1. The first part of the present thesis is focused on the main factors affecting the microbial safety of leafy greens at primary production level. Firstly, Chapter 3 evaluated the effect of agricultural factors and meteorological conditions on the microbial safety of leafy greens in Spain. In this study, soil and irrigation water were found to be the most important factors affecting the microbial safety of baby spinach. Concerning weather factors, it was suggested that ambient temperature affected the likelihood and extent the contamination of baby spinach. In relation to specific weather factors, Chapter 4 studied the effects of extreme rainfall on the microbial safety of leafy greens together with the influence of subsequent weather parameters on the microbial persistence in soil, water and leafy greens. It was found that soil, irrigation and leafy greens samples taken from the floodplains after the event were contaminated with high levels of E. coli and pathogens. The main findings obtained in this Chapter confirmed previous research, which described flooding as a main risk factor for the microbial contamination of leafy greens and highlighted the need for mitigation strategies to protect production areas from incidents capable of releasing microbial contamination. It was also shown that climatic conditions, especially, solar radiation played an important role reducing the microbial survival on the crop. In Chapter 5, mayor findings from chapters 3 and 4 together with a literature review of relevant publications were used to develop a quantitative microbial exposure assessment model of generic E. coli in baby spinach at primary production level in Spain. Once the model was built, it was used to evaluate the potential impact of weather and agricultural practices on the distribution of generic E. coli levels on leafy greens at harvest. Jointly, our results confirmed that weather factors and management practices influenced the likelihood of leafy greens contamination with E. coli and what intervention strategies aimed at such factors and practices could reduce the risk of pre-harvest contamination. Thus, QMEM was a useful approach to assess the potential impact of different risk factors and intervention strategies affecting E. coli concentrations at the field. Once the main risk factors for microbial contamination at field level were recognized, the next step was taken towards processing level. In this sense, Chapter 6 was focused on microbial risks associated to the processing of leafy greens. The main purpose was to gain insight into the bacterial contamination throughout the operations involved in leafy green processing in order to identify critical sampling points and their relation with the microbial safety of end products. Samples of centrifuge water were found to be positive for both, pathogens and generic E. coli, which suggested that the origin of the contamination could be the produce subjected to centrifugation or the remaining process wash water. In Chapter 7, the focus was made on the last step of the farm-to-fork chain: consumers' level. Firstly, the suitability of exiting data on the consumption of fresh fruit and vegetables was evaluated and it was established that detailed information about actual consumption of different types of fresh produce and the corresponding handling practices used by consumers at home was lacking. Therefore, this chapter was intended to provide standardized data for fresh produce consumed raw or minimally processed that could be used in future exposure assessments related to fresh produce. In Chapter 8, the main results of this thesis together with the future needs and perspectives resulting from this work were discussed.