Frédéric Carlin

The microbiology of minimally processed fresh fruits and vegetables

Minimally processed fresh (MPF) fruits and vegetables are good media for growth of microorganisms. They have been involved in outbreaks because of the consumption of products contaminated by pathogens. They are also sensitive to various spoilage microorganisms such as pectinolytic bacteria, saprophytic Gram-negative bacteria, lactic acid bacteria, and yeasts. Contamination of MPF fruits and vegetables occurs at every stage of the food chain, from cultivation to processing. Polluted environments during cultivation or poor hygienic conditions in processing increase the risk of contamination with foodborne pathogens. Although MPF fruits and vegetables may harbor psychrotrophic microorganisms such as fluorescent pseudomonads or Listeria monocytogenes, good control of refrigeration temperature limits growth of spoilage and pathogenic microorganisms. Modified atmospheres are often efficient to maintain or improve visual organoleptic quality of MPF fruits and vegetables, but their effects on microorganisms are inconsistent. Chemical disinfection can partially reduce the initial bacterial contamination; irradiation seems to be more efficient. The applications of legislations and quality assurance systems to control contamination, survival, and growth of foodborne pathogens in MPF fruits and vegetables are discussed.

Prevalence, characterization and growth of Bacillus cereus in commercial cooked chilled foods containing vegetables.

In cooked‐chilled and pasteurized vegetable products, initial numbers of Bacillus cereus were below 10 cfu g−1. Before the appearance of spoilage, numbers reached 6–8 log cfu g−1 at 20 °C and 4–6 log cfu g−1 at 10 °C. Bacillus cereus was not detected in samples stored at 4 °C. Ten percent of strains isolated from the products were able to grow at 5 °C and 63% at 10 °C. Bacillus cereus strains unable to degrade starch, a feature linked to the production of emetic toxin, did not grow at 10 °C and had a higher heat resistance at 90 °C. Using immunochemical assays, enterotoxin was detected in the culture supernatant fluid of 97·5% of the strains. All culture supernatant fluids were cytotoxic but important variations in the level of activity were found. Psychrotrophic isolates of B. cereus were unable to grow in courgette broth at 7 °C whereas they grew in a rich laboratory medium. At 10 °C, these isolates grew in both media but lag time in courgette broth was 20‐fold longer than in the rich laboratory medium.

Influence of Background Microflora on Listeria monocytogenes on Minimally Processed Fresh Broad-Leaved Endive (Cichorium endivia var. latifolia)

Interactions between Listeria monocytogenes and the background microflora of endive leaves was studied by (i) testing the effect of disinfecting endive leaves on the fate of L. monocytogenes , and (ii) challenging L. monocytogenes along with each of 10 strains of pseudomonads and Enterobacteriaceae isolated from endive leaves on both leaves of endive and in a model sterile medium based on an exudate of endive leaf. There was a higher increase in the population of L. monocytogenes on endive leaves with a background microflora reduced by a chemical disinfection. High numbers (106 to 107 CFU/g) of some strains reduced the growth of L. monocytogenes on endive. None of the 10 strains tested promoted the growth of L. monocytogenes . In the medium made of endive leaf exudate, the 10 strains tested reduced the maximum growth of L. monocytogenes to extents that varied with the strains. A complex bacterial population extracted from endive leaves completely inhibited the growth of L. monocytogenes in the medium.

A retail and consumer phase model for exposure assessment of Bacillus cereus

An exposure assessment is conducted for psychrotrophic and mesophilic Bacillus cereus in a cooked chilled vegetable product. A model is constructed that covers the retail and consumer phase of the food pathway, using the output of a similar model on the industrial process as input. Microbial growth is the predominant process in the model. Variability in time and temperature during transport and storage is included in the model and different domestic refrigerator temperature distributions are compared. As an end point, probable levels of B. cereus colony forming units (cfu) in packages of vegetable purée are predicted at the moment the consumer takes the product from its refrigerator, that is prior to a cooking process. The psychrotrophic strain is predicted to end up above a threshold level of 10(5) cfu/g in 0.9% to 6.3% of the vegetable purée packages, depending on domestic refrigerator temperature. Accounting for spoilage this reduces to 0.3% to 2.4%. Even if the purée is stored at 4 degrees C in the domestic refrigerator and use-by-date (UBD) is respected, the threshold level may be passed. For the mesophilic strain the threshold level is rarely passed, but in contrast to the total viable count, the spore load at the end point is predicted to be higher than in the psychrotrophic strain. Our study illustrates how an exposure assessment model, which may be used in quantitative risk assessment, can integrate expertise in modelling, food processing and microbiology over the food pathway, and thus evaluate food safety, identify gaps in knowledge and compare risk management measures. As important gaps in knowledge, the lack of sporulation and germination models and data, validated non-isothermal growth models and a spoilage model useful for risk assessment are identified. Knowledge of the dose-response relationship is limited and does not allow a full risk assessment. It is shown that exposure can be lowered by lowering domestic refrigerator temperatures, and less so much by monitoring and withdrawing contaminated products at the end of industrial processing.

Ability of Bacillus cereus group strains to cause food poisoning varies according to phylogenetic affiliation (groups I to VII) rather than species affiliation.

ABSTRACT Cytotoxic activity levels of culture filtrates and toxin distributions varied according to the phylogenetic group (I to VII) within the B acillus cereus group, suggesting that these groups are of different clinical significance and are more suitable than species affiliations for determining food poisoning risk. A first-line, simple online tool (https://www.tools.symprevius.org/Bcereus/english.php ) to assign strains to the different phylogenetic groups is presented.

Survival of Bacillus cereus spores and vegetative cells in acid media simulating human stomach

Aims:  To determine the fate of Bacillus cereus spores or vegetative cells in simulated gastric medium.

Research on factors allowing a risk assessment of spore-forming pathogenic bacteria in cooked chilled foods containing vegetables: a FAIR collaborative project

Vegetables are frequent ingredients of cooked chilled foods and are frequently contaminated with spore-forming bacteria (SFB). Therefore, risk assessment studies have been carried out, including the following: hazard identification and characterisation--from an extensive literature review and expertise of the participants, B. cereus and C. botulinum were identified as the main hazards; exposure assessment--consisting of determination of the prevalence of hazardous SFB in cooked chilled foods containing vegetables and in unprocessed vegetables, and identification of SFB representative of the bacterial community in cooked chilled foods containing vegetables, determination of heat-resistance parameters and factors affecting heat resistance of SFB, determination of the growth kinetics of SFB in vegetable substrate and of the influence of controlling factors, validation of previous work in complex food systems and by challenge testing and information about process and storage conditions of cooked chilled foods containing vegetables. The paper illustrates some original results obtained in the course of the project. The results and information collected from scientific literature or from the expertise of the participants are integrated into the microbial risk assessment, using both a Bayesian belief network approach and a process risk model approach, previously applied to other foodborne hazards.

Origin of bacterial spores contaminating foods

Bacterial spores (=endospores) are common contaminants in foods. Sources of contamination in the food chain may include soil, faeces, animal feeds and food ingredients and processing chain themselves. Sporulation may occur in very diverse environments. The environment of sporulation has a strong influence on spore properties relevant for food quality and safety.

Estimation of Microbial Contamination of Food from Prevalence and Concentration Data: Application to Listeria monocytogenes in Fresh Vegetables

ABSTRACT A normal distribution and a mixture model of two normal distributions in a Bayesian approach using prevalence and concentration data were used to establish the distribution of contamination of the food-borne pathogenic bacteria Listeria monocytogenes in unprocessed and minimally processed fresh vegetables. A total of 165 prevalence studies, including 15 studies with concentration data, were taken from the scientific literature and from technical reports and used for statistical analysis. The predicted mean of the normal distribution of the logarithms of viable L. monocytogenes per gram of fresh vegetables was −2.63 log viable L. monocytogenes organisms/g, and its standard deviation was 1.48 log viable L. monocytogenes organisms/g. These values were determined by considering one contaminated sample in prevalence studies in which samples are in fact negative. This deliberate overestimation is necessary to complete calculations. With the mixture model, the predicted mean of the distribution of the logarithm of viable L. monocytogenes per gram of fresh vegetables was −3.38 log viable L. monocytogenes organisms/g and its standard deviation was 1.46 log viable L. monocytogenes organisms/g. The probabilities of fresh unprocessed and minimally processed vegetables being contaminated with concentrations higher than 1, 2, and 3 log viable L. monocytogenes organisms/g were 1.44, 0.63, and 0.17%, respectively. Introducing a sensitivity rate of 80 or 95% in the mixture model had a small effect on the estimation of the contamination. In contrast, introducing a low sensitivity rate (40%) resulted in marked differences, especially for high percentiles. There was a significantly lower estimation of contamination in the papers and reports of 2000 to 2005 than in those of 1988 to 1999 and a lower estimation of contamination of leafy salads than that of sprouts and other vegetables. The interest of the mixture model for the estimation of microbial contamination is discussed.

Effects of carbon dioxide on the fate of Listeria monocytogenes, of aerobic bacteria and on the development of spoilage in minimally processed fresh endive

Minimally processed fresh broad-leaved endive (Cichorium endivia L.) were stored at 3 and 10 degrees C in modified atmospheres containing air, 10% CO2/10% O2, 30% CO2/10% O2, and 50% CO2/10% O2. The effects of these modified atmospheres on the fate of both aerobic bacteria and three strains of Listeria monocytogenes, was investigated. Increases in CO2 concentrations significantly reduced the growth of the aerobic microflora. The best preservation of the visual quality occurred on endive leaves stored in 10% CO2/10% O2, whereas leaves stored in 30% CO2/10% O2 and 50% CO2/10% O2, and to a lesser extent in air, showed extensive spoilage after storage. Listeria monocytogenes was slightly affected at 3 degrees C by the modified atmospheres, as compared to air. At 10 degrees C, results varied between replicate experiments, but L. monocytogenes generally grew better as the CO2 concentration was increased. The three test strains behaved in a similar way. In conclusion, among the modified atmospheres tested, a modified atmosphere containing 10% CO2/10% O2 resulted in improved visual quality of minimally processed fresh endive, without a marked effect on the growth of the aerobic microflora or of L. monocytogenes.