Impact of multiple hurdles on Listeria monocytogenes dispersion of survivors.

Pathogen exposure to multiple hurdles could result in variation in the number of survivors, which needs to be carefully considered using appropriate regression models for dealing with survivor dispersion. The aim of this study was to evaluate the impact of the hurdles on the random component of the measured variation and on its unexplained part (over or under-dispersion) representing the departure from randomness, i.e. non-randomness, in survivors of a multi-strain mixture of L. monocytogenes. The pathogen inactivation curves were fitted to the Weibull model within the Conway-Maxwell-Poisson process. In all the 20 hurdle combinations, the surviving cells, whether they showed an upward curvature or linear kinetics, displayed the randomness revealed by the degree of dispersion of the inactivation parameters (-b and p). In 15 combinations, a significant dispersion coefficient (c0), which reflected the non-random component of variation was evident, denoting either over-dispersion (c0 > 0 in 13 combinations) or under-dispersion (c0 < 0 in 2 combinations). The observed dependence of the under- and over-dispersion conditions on the inactivation rate was confirmed by a Monte Carlo simulation based on the inactivation parameter -b. Including both randomness and non-randomness provides a more accurate estimation of survivors, which certainly impacts on intervention practices.

Involvement of alanine racemase in germination of Bacillus cereus spores lacking an intact exosporium.

The L-alanine mediated germination of food isolated Bacillus cereus DSA 1 spores, which lacked an intact exosporium, increased in the presence of D-cycloserine (DCS), which is an alanine racemase (Alr) inhibitor, reflecting the activity of the Alr enzyme, capable of converting L-alanine to the germination inhibitor D-alanine. Proteomic analysis of the alkaline extracts of the spore proteins, which include exosporium and coat proteins, confirmed that Alr was present in the B. cereus DSA 1 spores and matched to that encoded by B. cereus ATCC 14579, whose spore germination was strongly affected by the block of conversion of L- to D-alanine. Unlike ATCC 14579 spores, L-alanine germination of B. cereus DSA 1 spores was not affected by the preincubation with DCS, suggesting a lack of restriction in the reactant accessibility.

Survival strategies of Bacillus spores in food.

Control of bacterial spores is one of the major problem in the food preservation. Spores of Bacillus genus are commonly present in different environments, including soil and the gut of insects and animals and, as a result, they can be spread to all kind of foods. Due to their high resistance properties, their complete inactivation in food is often impossible without changing the product characteristics. Surviving spores can germinate and grow out to vegetative cells, with the consequent great risk of food spoilage and food poisoning after consumption. Spores have evolved various mechanisms, including phenotypic variability, to protect themselves from a wide range of damage resulting from food preservation treatments. Even if the phenotypic heterogeneity contributes to increase the chances of survival of Bacillus spore to conventional preservation treatments, in some specific instances, an homogeneous response could be the result of a strategy adopted by the spores to increase resistance to those treatments.

A simplified approach for modelling the bacterial growth/no growth boundary.

A simplified growth/no growth (G/NG) model, conceptually derived from the Gamma model and making direct and explicit use of growth limits of bacteria through a normalization constant (η), was proposed. The η value, which quantifies the product of the cardinal optimal distances for growth probability, is a species-independent constant. This is of importance when experimental data is missing or insufficient. The simplified G/NG model was developed including the effect of temperature, pH and water activity, and was expanded incorporating the preservative effects. As a practical application, the model was investigated for its ability to describe published data. The successful validation of the simplified G/NG model is discussed in regard to its potential applicability as a first estimate method for the development of safe food products.

The COM-Poisson Process for Stochastic Modeling of Osmotic Inactivation Dynamics of Listeria monocytogenes.

Controlling harmful microorganisms, such as Listeria monocytogenes, can require reliable inactivation steps, including those providing conditions (e.g., using high salt content) in which the pathogen could be progressively inactivated. Exposure to osmotic stress could result, however, in variation in the number of survivors, which needs to be carefully considered through appropriate dispersion measures for its impact on intervention practices. Variation in the experimental observations is due to uncertainty and biological variability in the microbial response. The Poisson distribution is suitable for modeling the variation of equi-dispersed count data when the naturally occurring randomness in bacterial numbers it is assumed. However, violation of equi-dispersion is quite often evident, leading to over-dispersion, i.e., non-randomness. This article proposes a statistical modeling approach for describing variation in osmotic inactivation of L. monocytogenes Scott A at different initial cell levels. The change of survivors over inactivation time was described as an exponential function in both the Poisson and in the Conway-Maxwell Poisson (COM-Poisson) processes, with the latter dealing with over-dispersion through a dispersion parameter. This parameter was modeled to describe the occurrence of non-randomness in the population distribution, even the one emerging with the osmotic treatment. The results revealed that the contribution of randomness to the total variance was dominant only on the lower-count survivors, while at higher counts the non-randomness contribution to the variance was shown to increase the total variance above the Poisson distribution. When the inactivation model was compared with random numbers generated in computer simulation, a good concordance between the experimental and the modeled data was obtained in the COM-Poisson process.

Changes of physicochemical, microbiological, and textural properties during ripening of Italian low-acid sausages. Proteolysis, sensory and volatile profiles.

In this study low-acid sausages were studied to characterize their physicochemical, microbiological, and textural properties during ripening. The final a(w) was 0.87-0.88, whereas pH values stayed around their initial values during processing. Lactic acid bacteria increased very slowly in number and a small increase of Micrococcaceae was also noticed. Low-acid sausages showed low hardness and cohesiveness, and were easily distinguishable by sensory analysis from other industrial and artisan sausages. Under the conditions of the study, observed volatile compounds were mainly from spices and wine. The respective contribution of muscle and indigenous bacterial enzymes to proteolysis was determined by comparing changes in low-acid sausages to those containing an antibiotic-antimycotic mixture or sugar. A large part of the degradation of myofibrillar proteins appeared due to endogenous enzymes, although bacterial proteinases contributed to the degradation of these proteins. The role of microorganisms in proteolysis was more evident in the degradation of sarcoplasmic proteins.

Praedicere Possumus: An Italian web-based application for predictive microbiology to ensure food safety.

The use of predictive modelling tools, which mainly describe the response of microorganisms to a particular set of environmental conditions, may contribute to a better understanding of microbial behaviour in foods. In this paper, a tertiary model, in the form of a readily available and userfriendly web-based application Praedicere Possumus (PP) is presented with research examples from our laboratories. Through the PP application, users have access to different modules, which apply a set of published models considered reliable for determining the compliance of a food product with EU safety criteria and for optimising processing throughout the identification of critical control points. The application pivots around a growth/no-growth boundary model, coupled with a growth model, and includes thermal and non-thermal inactivation models. Integrated functionalities, such as the fractional contribution of each inhibitory factor to growth probability (f) and the time evolution of the growth probability (Pt), have also been included. The PP application is expected to assist food industry and food safety authorities in their common commitment towards the improvement of food safety.

Glassy state in Bacillus subtilis spores analyzed by differential scanning calorimetry.

Thermal properties of dried spores of Bacillus subtilis, investigated by differential scanning calorimetry (DSC), were studied. A reversible heat capacity shift ascribable to glass-rubber transition was observed at 90-115 degrees C. The transition was found to be a pressure-inhibited volume-activated event. The decoated spores and the extracted peptidoglycan material exhibited glass transition, suggesting that the cortex could be involved in the event. Furthermore, the glass transition was evident when spores were treated with strong acid, and when the isogenic strain PS578 was scanned, indicating that core integrity and core components are not involved in the occurrence of the event. These results suggest that in the dried B. subtilis spores an amorphous biomaterial, possibly the cortex peptidoglycan, is present as a glass.

Growth of Listeria monocytogenes as influenced by viscosity and water activity.

The effects of osmotic stress on Listeria monocytogenes growth parameters was examined in relation to the viscosity of the growth media. In low-viscosity systems, growth of L. monocytogenes in glucose-supplemented media was comparable to growth in sucrose-supplemented media. The relative lag time (RLT: the lag time divided by the generation time) responses were found to increase in the more restrictive water activity conditions. In high-viscosity systems containing polyvinylpyrrolidone (PVP), growth rate was reduced, whereas lag time showed no discernible modification. Osmotic stress in medium- and high-viscosity media supplemented with glucose resulted in approximately exponential increasing of the RLT values. Thus, the biological effects of osmotic stress on L. monocytogenes could be affected by the physical properties of the system, such as viscosity and diffusivity.

Effect of Essential Oils on Aeromonas hydrophila in a Culture Medium and in Cooked Pork.

The inhibitory effects of essential oils of coriander, clove, nutmeg, and pepper towards Aeromonas hydrophila were studied in tryptic soy agar. Essential oils of clove, coriander, and nutmeg at concentrations of 500, 1,250, and 10,000 µg/ml, respectively, were effective in inhibiting the growth of A. hydrophila on tryptic soy agar. Pepper was the least active oil of which 15,000 µg/ml appeared to inhibit only a diluted culture of A. hydrophila . Behavior of A. hydrophila was also evaluated in samples of noncured cooked pork treated either with coriander or clove oil. Meat samples were packaged either under vacuum or air and stored at 2 and 10°C. Both essential oils caused a marked reduction of A. hydrophila number. In the meat samples treated with clove oil and stored at 10°C, the lethal effect was significantly enhanced by the vacuum packaging. These data suggest that essential oils of coriander and clove could be used to control the hazard of A. hydrophila in noncured cooked meat at low and increased temperatures, especially in combination with vacuum packaging.

The Behavior of Yeast Populations in Stracchino Cheese Packaged under Various Conditions.

The growth of yeasts, lactic acid bacteria, pseudomonads, and enterobacteria during the refrigerated storage of an Italian soft cheese (Stracchino) in various packaging conditions (under air, under vacuum, and in the presence of ethanol) was studied. Yeasts were found to play a significant role in cheese spoilage, producing unplesant flavors and odors and causing significant reductions in the shelf life of the paper-wrapped cheeses. Packaging under vacuum decreased the yeast growth rates and reduced the populations attained at the end of the exponential phase of growth, resulting in a shelf-life extension of the Stracchino cheese of over 28 days. A total of 129 yeasts isolates were identified according to conventional methods. The most frequently isolated yeasts were Candida colliculosa , Debaryomyces hansenii , and Candida famata . Other species encountered were Tolurospora delbrueckii , Kluyveromyces marxianus , and Saccharomyces cerevisiae.