Search

Background: Very young children, and those with disabilities and extensive oral pathology, who cannot be treated in the dental chair, require deep sedation or general anesthesia for dental treatment. Objective: The aim of this study is to describe and compare the oral health status in healthy and SHCN children and the treatments performed under deep sedation on an outpatient basis with a minimal intervention approach, and their impact on quality of life. Methods: A retrospective study between 2006 and 2018 was made. A total of 230 medical records of healthy and SHCN children were included. The data extracted were age, sex, systemic health status, reason for sedation, oral health status before sedation, treatments administered during sedation, and follow-up. The quality of life after deep sedation of 85 children was studied through parental questionnaires. Descriptive and inferential analyses were made. Results: Of the 230 children, 47.4% were healthy and 52.6% were SHCN. The median age was 7.10 ± 3.40 years (5.04 ± 2.42 in healthy children and 8.95 ± 3.09 in SHCN children). The main reason for sedation was poor handling in the dental chair (99.5%). The most frequent pathologies were caries (90.9%) and pulp pathology (67.8%). Healthy children had more teeth affected by decay and with pulp involvement. Patients aged < 6 years received more pulpectomies and pulpotomies. After treatment, parents stated that children were more rested and less irascible, ate better, increased in weight, and had improved dental aesthetics. Conclusions: Differences in treatments carried out did not depend on the general health status or the failure rate but on age, with more pulp treatments in healthy children who were younger, and more extractions near to the age of physiological turnover in children with SHCN who were older. Intervention under deep sedation with a minimally invasive treatments approach met the expectations of parents and guardians, as it improved the children’s quality of life.

Consumers’ demands for ready-to-eat, fresh-like products are on the rise during the last years. This type of products have minimal processing conditions that can enable the survival and replication of pathogenic microorganisms. Among them, Listeria monocytogenes is of special concern, due to its relatively high mortality rate and its ability to replicate under refrigeration conditions. Previous research works have shown that nanoemulsified essential oils in combination with thermal treatments are effective for inactivating L. monocytogenes. However, previous research works were limited to isothermal conditions, whereas actual processing conditions in industry are dynamic. Under dynamic conditions, microorganism can respond unexpectedly to the thermal stress (e.g. adaptation, acclimation or increased sensitivity). In this work, we assess the combination of nanoemulsified D-limonene with thermal treatments under isothermal and dynamic conditions. The nanoemulsion was prepared following an innovative methodology using soya lecithin, a natural compound as well as the essential oil. Under isothermal heating conditions, the addition of the antimicrobial enables a reduction of the treatment time by a factor of 25. For time-varying treatments, dynamic effects were relevant. Treatments with a high heating rate (20 °C/min) are more effective than those with a slow heating rate (1 °C/min). This investigation demonstrates that the addition of nanoemulsified D-limonene can greatly reduce the intensity of the thermal treatments currently applied in the food industry. Hence, it can improve the product quality without impacting its safety.

The effects of high hydrostatic pressure (HHP) compared to thermal pasteurization (TP) were studied in healthy citrus-maqui beverages. The impact of the processing technologies on the microbiological and phytochemical profile was assessed by applying two HHP treatments at 450 and 600 MPa for 180 s and TP at 85 °C for 15 s. The shelf life under refrigeration (4 °C) and room temperature (20 °C) was monitored for 90 days. All treatments ensured microbiological stability at both storage temperatures. Aside from that, the physicochemical parameters were not significantly different after processing or throughout the storage period. Regarding color parameters, an increase in the reddish coloration was observed during storage for those beverages treated by HHP. In general, phenolic compounds were little affected by the processing technique, even when treatment under HHP was more stable than by TP during storage. On the other hand, vitamin C showed great degradation after processing under any condition. It can be concluded that HHP is an effective alternative to thermal treatments, achieving effective microbial inactivation and extending the shelf life of the juices by contributing to a better preservation of color and bioactive compounds.

Cooking vegetables in microwave bags is becoming a popular domestic cooking method, being relevant to know how this cooking method affects health-promoting phytochemicals of staples such as broccoli. The aim of this work was to study the effect of microwave bag cooking versus conventional microwaving on bioactive compound content (glucosinolates and hydroxycinnamic acid derivatives) and other quality parameters (such as antioxidant capacity, mineral content and microbial load) of broccoli florets. The influence of cooking time on bioactive compounds content was also evaluated. The study was carried out in two independent experiments; using intact broccoli and broccoli preprocessed in industry. Microwave bag cooked broccoli for 5 min (following label recommendation) showed higher glucosinolate content retention compared to conventional microwaving. Results suggest that volatilization could be an important phenomenon in reduction of glucosinolates during microwave cooking of broccoli florets. Glucosinolate profile did not change after cooking, regardless of cooking method applied. Furthermore, microwave bag cooked broccoli presented higher antioxidant capacity (by DPPH assay) than conventional microwaved broccoli. Hydroxycinnamic acid derivatives content was reduced in microwave cooking, regardless of method applied. Altogether, the use of microwave bags for microwaving is a novel method that retains main bioactive components of broccoli. This option is a fast, easy and considerably clean cooking option to fulfill modern consumer needs., This work was supported by Comision ´ Sectorial de Investigacion ´ Científica, Universidad de la República. Authors are indebted to Agencia Nacional de Innovacion ´ e Investigacion ´ for granting Erika Paulsen a PhD scholarship and an internship at Phytochemistry and Healthy Foods Laboratory of the Department of Food Science and Technology at CEBAS-CSIC (scholarship reference: POS_EMHE_2018_1_1007740)

Cruciferous sprouts (e.g. broccoli and red radish) are rich source of health-promoting phytochemicals that are more concentrated than in the adult plant edible organs; however, these tiny microgreens need cold storage conditions to preserve their quality to reach the consumers in microbiologically safe conditions, maintaining their composition and acceptability. In this work, the microbiological status and phytochemical composition of broccoli and radish sprouts were evaluated at harvest (Day 0), and after seven and fourteen days of storage at 5 and 10 °C. Pathogenic microorganisms were absent during shelf-life; nevertheless, the slight growth of Enterobacteriaceae organisms, aerobic mesophilic and psychotropic bacteria, molds and yeasts was assessed. The storage temperature influenced the quality and content of bioactives in the sprouts, and for practical applications, storage at 5 °C is the most suitable option. Moreover, these fresh crucifers remain acceptable for consumers after 14 d storage period, being an interesting option for consuming fresh and naturally-functional foods.

Microwave processing can be a valid alternative to conventional heating for different types of products. It enables a more efficient heat transfer in the food matrix, resulting in higher quality products. However, for many food products a uniform temperature distribution is not possible because of heterogeneities in their physical properties and non-uniformtiy in the electric field pattern. Hence, the effectiveness of microwave inactivation treatments is influenced by both intrinsic (differences between cells) and extrinsic variability (non-uniform temperature). Interpreting the results of the process and considering its impact on microbial inactivation is essential to ensure effective and efficient processing. In this work, we quantified the variability in microbial inactivation attained in a microwave pasteurization treatment with a tunnel configuration at pilot-plant scale. The configuration of the equipment makes it impossible to measure the product temperature during treatment. For that reason, variability in microbial counts was measured using Biological Inactivation Indicators (BIIs) based on spherical particles of alginate inoculated with spores of Bacillus spp. The stability of the BIIs and the uncertainty associated to them was assessed using preliminary experiments in a thermoresistometer. Then, they were introduced in the food product to analyse the microbial inactivation in different points of the products during the microwave treatment. Experiments were made in a vegetable soup and a fish-based animal by-product (F-BP). The results show that the variation in the microbial counts was higher than expected based on the biological variability estimated in the thermoresistometer and the uncertainty of the BIIs. This is due to heterogeneities in the temperature field (measured using a thermographic camera), which were higher in the F-BP than in the vegetable soup. Therefore, for the process studied, extrinsic variability was more relevant than intrinsic variability. The methodology presented in this work can be a valid method to evaluate pasteurization treatments of foods processed by heating, providing valuable information of the microbial inactivation achieved. It can contribute to design microwave processes for different types of products and for product optimization.

In the present study, it was evaluated how two different thermal treatments (Mild and Severe) may affect the anthocyanin content, antioxidant capacity (ABTS(+), DPPH, and FRAP), quality (CIELAB colour parameters), and microbiological safety of a new isotonic drink made of lemon and maqui berry over a commercial storage simulation using a shelf life of 56days at two preservation temperature (7°C and 37°C). Both heat treatments did not affect drastically the anthocyanins content and their percentage of retention. The antioxidant capacity, probably because of the short time, was also not affected. The CIELAB colour parameters were affected by the heat, although the isotonic drinks remained with attractive red colour during shelf life. From a microbiological point of view, the Mild heat treatment with storage at 7°C is the ideal for the preservation of microbial growth, being useful for keeping the quality and safety of beverages in commercial life.

The application of oily antimicrobials in form of nanoemulsion has been proved to solve the problem of their immiscibility in aqueous media, still preventing microbial growth and even improving the antimicrobial effect observed when applied directly. At present, only a few documented studies have evaluated the combined effect of nanoemulsions with other factors of stress for the microorganism. The present research shows very promising results on the combination of nanoemulsified d -limonene with thermal treatments on the inactivation of Listeria monocytogenes . The thermal resistance of L . monocytogenes was reduced two to five times when 0.5 mM d -limonene was added directly to the heating medium. However, when the same concentration of d -limonene was present in the heating medium in form of nanoemulsion, the heat resistance was reduced by one hundred times at all heating temperatures tested. The addition of nanoemulsified antimicrobials would allow to reduce greatly the intensity of the thermal treatments currently applied in the food processing industry.

The present work evaluated the antibacterial effect of nanoemulsions from natural compounds d-limonene and nisin against Listeria monocytogenes in tryptic soy broth growth medium, chicken broth, and vegetable cream. Experiments were performed by means of optical density growth curves and studies of viability in culture media and foods. Stability of nanoemulsions was evaluated by a Mastersizer 2000 equipment. Results showed greater effectiveness when applying d-limonene in form of nanoemulsion than when applying it directly, and when using both compounds together, both directly and in the form of nanoemulsion. Concentration of L. monocytogenes decreased at least in three log cycles in all the culture media and foods used within the first 90 min after the addition of the natural antimicrobials combined in form of nanoemulsion. Moreover, the growth of L. monocytogenes was inhibited with the combination of antimicrobials in the four weeks that the experiment lasted. Nanoemulsion technology would solve present problems of solubility and stability of oily antimicrobials in the food industry.

In this paper, a novel technique to achieve precise temperatures in food sterilization has been proposed. An accurate temperature profile is needed in order to reach a commitment between the total removal of pathogens inside the product and the preservation of nutritional and organoleptic characteristics. The minimal variation of the target temperature in the sample by means of a monitoring and control software platform, allowing temperature stabilization over 100 °C, is the main goal of this work. A cylindrical microwave oven, under pressure conditions and continuous control of the microwave supply power as function of the final temperature inside the sample, has been designed and developed with conditions of single-mode resonance. The uniform heating in the product is achieved by means of sample movement and the self-regulated power control using the measured temperature. Finally, for testing the sterilization of food with this technology, specific biological validation based on Bacillus cereus as a biosensor of heat inactivation has been incorporated as a distribution along the sample in the experimental process to measure the colony-forming units (CFUs) for different food samples (laboratory medium, soup, or fish-based animal by-products). The obtained results allow the validation of this new technology for food sterilization with precise control of the microwave system to ensure the uniform elimination of pathogens using high temperatures. This work was supported by the Ministerio de Economía y Competitividad under the project with reference DPI2014-61857-EXP.

The present work studies the effect of acid shock on cell viability of Bacillus cereus and Bacillus weihenstephanensis. Experiments were performed by means of flow cytometry (FCM) combined with fluorescent labelling. Propidium iodide, for membrane evaluation, and carboxyfluorescein diacetate, for esterase activity were used for differential staining of both strains. pH values of 3.4, 3.8 and 4.2 were selected on the basis of previous screening tests. FCM analysis showed to clearly discriminate between different populations, viable and damage cells, leading to successful assessment of acid shock effect on B. cereus and B. weihenstephanensis vegetative cell viability. Cellular viability decreased at lower pH values and longer exposure times, being B. weihenstephanensis more acid resistant than B. cereus. Reliability of FCM analysis for detection of B. cereus and B. weihenstephanensis vegetative cells was compared with classical viable plate count techniques (VPC). Although FCM and VPC data were not directly correlated and classical analytical methods show higher reliability, FCM analysis provides high-speed information at real time on damage at single cell level, whereas VPC only gives an indication of cells able to grow at a certain time. These data could help to establish more accurately the potential risk associated to foods or processing conditions where pH is involved, taking damage into consideration.

Yeasts are tolerant to acid pH values, are able to grow in anaerobic media and have minimum nutrition requirements. These capabilities enable them to survive and even grow in foods prepared from acid fruits or vegetables. Among yeasts, Candida is one of the genus most frequently isolated from fruit juices. Bacteriocins and essential oils from spices and aromatic herbs are an alternative to preservatives and other technological treatments and have the advantage that their natural origins do not lead to consumer rejection. However, before the food industry uses them on a large scale, it is necessary to know their effects on microorganisms. The objective of this research was to study the effect of different concentrations of nisin, thymol, carvacrol and cymene on the growth of Candida lusitaniae in pH 5 broth at 25 ℃, and their potential uses as food preservatives. The addition of nisin at the concentrations tested (up to 3 µmol L−1) did not affect the yeast growth. Thymol, carvacrol and cymene completely inhibited the yeast growth at concentrations over 1 mM for at least 21 days at 25 ℃. Below this concentration, inhibitions on yeast growth were observed at increasing concentrations. The effect of thymol was also proved in tomato juice. This study indicates the potential use of essential oils for preservation of minimally processed foods.

The use of non-thermal methods for food preservation is due to consumer demands for microbiological safe products, without changes in the sensory and nutritional qualities of the product. High hydrostatic pressure (HHP) has emerged as an alternative to traditional thermal processing methods for foods. Listeria monocytogenes CECT 5672 was treated under HHPs (350, 400 and 450 MPa) for 3, 16 and 23 min. The effects of pH (5, 6 and 7) and sodium chloride concentration (0, 0.5 and 1.0) of the recovery medium were studied on L. monocytogenes . The kinetic parameters were estimated by the method described by Metris, George, and Baranyi (2006) . From results obtained, histograms of the lag phase were generated and distributions were fitted. The duration of the lag phase of HHP damaged cells increased with the application of additional stresses. Histograms showed a shift to longer lag phases and an increase in variability with high stress levels in the recovery medium. Using a primary model together with Monte Carlo simulation, predictions of time to growth (100 cfu/g) of L. monocytogenes were established and they were compared with deterministic predictions. It was evidenced that deterministic predictions do not give a good indication of the probability of a certain level of growth.

Heat resistance of microorganisms can be affected by different influencing factors. Although, the effect of heating rates has been scarcely explored by the scientific community, recent researches have unraveled its important effect on the thermal resistance of different species of vegetative bacteria. Typically heating rates described in the literature ranged from 1 to 20°C/min but the impact of much higher heating rates is unclear. The aim of this research was to explore the effect of different heating rates, such as those currently achieved in the heat exchangers used in the food industry, on the heat resistance of Escherichia coli. A pilot plant tubular heat exchanger and a thermoresistometer Mastia were used for this purpose. Results showed that fast heating rates had a deep impact on the thermal resistance of E. coli. Heating rates between 20 and 50°C/min were achieved in the heat exchanger, which were much slower than those around 20°C/s achieved in the thermoresistometer. In all cases, these high heating rates led to higher inactivation than expected: in the heat exchanger, for all the experiments performed, when the observed inactivation had reached about seven log cycles, the predictions estimated about 1 log cycle of inactivation; in the thermoresistometer these differences between observed and predicted values were even more than 10 times higher, from 4.07 log cycles observed to 0.34 predicted at a flow rate of 70 mL/min and a maximum heating rate of 14.7°C/s. A quantification of the impact of the heating rates on the level of inactivation achieved was established. These results point out the important effect that the heating rate has on the thermal resistance of E. coli, with high heating rates resulting in an additional sensitization to heat and therefore an effective food safety strategy in terms of food processing. This research was financially supported by the Ministry of Economy and Competitiveness of the Spanish Government and European Regional Development Fund (ERDF) through project AGL2013-48993-C2-1-R

Submitted by Biblioteca Suporte PUCRS (biblioteca.suporte@pucrs.br) on 2022-10-05T13:09:08Z No. of bitstreams: 2 Onestep_global_parameter_estimation_of_kinetic_inactivation_parameters_for_Bacillus_sporothermodurans_spores_under.pdf: 476629 bytes, checksum: 65b89c194e93a6e7933c0ab027da722e (MD5) Onestep_global_parameter_estimation_of_kinetic_inactivation_parameters_for_Bacillus_sporothermodurans_spores_under.pdf: 476629 bytes, checksum: 65b89c194e93a6e7933c0ab027da722e (MD5) Made available in DSpace on 2022-10-05T13:09:08Z (GMT). No. of bitstreams: 2 Onestep_global_parameter_estimation_of_kinetic_inactivation_parameters_for_Bacillus_sporothermodurans_spores_under.pdf: 476629 bytes, checksum: 65b89c194e93a6e7933c0ab027da722e (MD5) Onestep_global_parameter_estimation_of_kinetic_inactivation_parameters_for_Bacillus_sporothermodurans_spores_under.pdf: 476629 bytes, checksum: 65b89c194e93a6e7933c0ab027da722e (MD5) Previous issue date: 2016

Strains belonging to Bacillus cereus Group include six different species, among which are Bacillus thuringiensis, Bacillus weihenstephanensis, and Bacillus cereus sensu stricto, a causative agent of food poisoning. Sequence of the panC-housekeeping gene is used for B. cereus Group affiliation to seven major phylogenetic groups (I-VII) with different ecological niches and variations in thermal growth range and spore heat resistance of B. cereus Group microorganisms varies among phylogenetic groups. We assigned a selection of B. cereus sensu stricto strains related to food poisoning from the Spanish cultivar Collection (Valencia) to Group IV strains based on panC gene sequence. Thermal inactivation assays revealed variability of spore heat resistance within these Group IV strains. Adequate food sanitizing treatments therefore require fast and reliable identification of particular strains. In the present study, feasibility of genotyping via high-resolution melting (HRM) analysis was examined. HRM analysis of amplified polymorphic 16S-23 intergenic spacer region (ISR) region proved to be discriminatory for B. cereus sensu stricto strain typing, while two other polymorphic regions within the bacterial rRNA operon allowed differentiation between Bacillus species, demonstrating its applicability for discrimination on the species and strain level within B. cereus Group.

Stochastic models can be useful to predict the risk of foodborne illness. The presence of Bacillus cereus in liquid egg can pose a serious hazard to the food industry, since a mild heat treatment cannot guarantee its complete inactivation. However, most of the information available in the scientific literature is deterministic, including growth of B. cereus. In this paper, a stochastic approach to evaluate growth of B. cereus cells influenced by different stresses (presence of nisin and lysozyme separately or in combination) was performed, using an individual-based approach of growth through OD measurements. Lag phase duration was derived from the growth curves obtained. From results obtained, histograms of the lag phase were generated and distributions were fitted. Normal and Weibull distributions were ranked as the bestfit distributions in experiments performed at 25 °C. At 16 °C, lag values (obtained in presence of combinations of both antimicrobials) were also fitted by a Gamma distribution. Predictions were compared with growth curves obtained in liquid egg exposed to mild heat, nisin and/or lysozyme to assess their validity.

Spore-forming bacteria with high heat resistance increasingly challenge industrial sterilisation processes in foods. To ensure stability of manufactured foods, generally worst case scenarios are applied often leading to unwanted over processing of foods. This means bigger requirements of energy and larger emissions of CO2 into the atmosphere. A heat-resistant spore-forming bacterium, Bacillus sporothermodurans, was tested for its ability to produce consistently highly heat-resistant spore crops to be used to more precisely determine the minimal thermal process that is compatible with microbial stability, thus avoiding the indicated overprocessing. Isothermal survival curves of B. sporothermodurans IC4 spores were determined in distilled water at temperatures up to 125 °C using the capillary tube technique and a thermoresistometer Mastia. B. sporothermodurans consistently produced crops of heat-resistant spores with kmax values at 121 °C up to 0.46 min−1. After consecutive sporulation cycles, the measured heat resistance of B. sporothermodurans was not reduced. Survival curves showed shoulders that were characterized by means of existing models (Geeraerd, Weibull) that were compared with the classical, log-linear one. Shoulders are a common phenomenon in heat-resistant spore-forming bacteria and may be related to the need of more stringent processing conditions. B. sporothermodurans spores were considered adequate to optimize sterilisation processes of low acid foods under a wide variety of conditions. Establishing precise heat treatments can be a way to guarantee food safety and stability while reducing the use of energy and contaminating emissions.

In general, food engineers are trained to solve engineering problems in the food industry. More specifically, the food engineer must specify the functional requirements, design, and testing of food products, and finally, the evaluation of products to check for overall efficiency, cost, reliability, and most importantly, safety. Food safety must be considered foremost as the overall engineering problem encountered in the food supply chain, and it must be solved from a food safety engineering perspective. This article will show that the food safety engineering perspective is needed in order to produce high quality food products (minimally processed) that are both safe and secure. This multi-disciplinary approach will involve certain engineering components: (i) predictive microbiology as a tool to evaluate and improve food safety in traditional and new processing technologies, (ii) advanced food contaminants detection methods, (iii) advanced processing technologies, (iv) advanced systems for re-contamination control, (v) advanced systems for active and intelligent packaging.

The microbial safety and stability of minimally processed foods are based on the application of combined preservative factors. Since microorganisms are able to develop adaptive networks to survive under conditions of stress, food safety may be affected, and therefore understanding of stress adaptive mechanisms plays a key role in designing safe food processing conditions. In the present study, the viability and the sublethal injury ofListeria monocytogenesexposed to moderate heat (55°C) and/or essential oil compounds (carvacrol and thymol, 0.3 mM) treatments were studied. Synergistic effects were obtained when combining mild heat (55°C) with one or both essential oil compounds, leading to inactivation kinetics values three to four times lower than when using heat alone. All the treatments applied caused some injury in the population. The injury levels ranged from around 20% of the surviving population under the mildest conditions to more than 99.99% under the most stringent conditions. Protein extracts of cells exposed to these treatments were analysed by two-dimensional gel electrophoresis. The results obtained revealed that stressed cells exhibited differential protein expression to control cells. The proteins upregulated under these stressing conditions were implicated, among other functions, in stress response, metabolism, and protein refolding.

A model for prediction of bacterial spore inactivation was developed. The influence of temperature, pH and NaCl on the heat resistance of Bacillus stearothermophilus spores was described using low-complexity, black box models based on artificial neural networks. Literature data were used to build and train the neural network, and new experimental data were used to evaluate it. The neural network models gave better predictions than the classical quadratic response surface model in all the experiments tried. When the neural networks were evaluated using new experimental data, also good predictions were obtained, providing fail-safe predictions of D values in all cases. The weights and biases values of neurons of the neural network that gave the best results are presented, so the reader can use the model for their own purposes. The use of this non-linear modelling technique makes it possible to describe more accurately interacting effects of environmental factors when compared with classical predictive microbial models.

The antibacterial action of carvacrol and cymene on two Listeria monocytogenes strains (STCC4031 and NCTN4032) was studied. Carvacrol or cymene showed inhibitory effect on the growth of L. monocytogenes during lag and exponential growth phases and was more evident with increasing concentrations in brain heart infusion broth at 30 degrees C. Carvacrol or cymene also decreased the survival of mid-exponential-growth-phase L. monocytogenes STCC4031 cells in potassium-N-2-hydroxy-ethylpiperazine-N-ethanesulfo nic acid, at 30 degrees C. The combination of carvacrol and cymene resulted in an increased antibacterial effect on the growth and a synergistic effect on the viability of L. monocytogenes compared with the natural compounds applied separately. The analysis of survival curves by the Weibull frequency distribution function allowed an accurate prediction of the level of inactivation achieved. Interestingly, an important bactericidal effect (4.7-log reduction) of low concentrations of both antimicrobials combined (0.75 mM) was observed on L. monocytogenes in carrot juice. This study indicates the potential use of carvacrol and cymene applied simultaneously for preservation of minimally processed foods.

The bactericidal action of thymol and cymene on two Bacillus cereus strains (INRA-AVTZ415 and INRA-AVZ421) was studied. Increasing concentrations of thymol (0.2–1.0 mmol l−1) or cymene (0.2–2.0 mmol l−1) showed higher bactericidal effect on exponential growth phase B. cereus cells suspended in HEPES buffer (pH 7), at 30°C. The two strains tested presented different sensitivity to these natural antimicrobials. When thymol and cymene were combined, it resulted in a greater bactericidal effect on B. cereus cells than when these compounds in the essential oil fraction of aromatic plants were applied separately. There was a synergistic effect of both natural antimicrobials on the viability of exponential growth phase B. cereus cells in pH 7 HEPES buffer, at 30°C. Survivor curves did not follow a first-order kinetics, which makes it difficult to establish processing conditions. Experimental data were modeled using a frequency distribution function (Weibull). This model guaranteed a good description of the experimental results and allowed predictions of time to a specific decrease in bacterial populations. This study indicates the potential use of thymol and cymene applied separately or simultaneously for preservation of minimally processed foods, as well as the validity of the Weibull distribution to describe the resulting nonlinear data and establish effective treatment conditions with these compounds.

The antibacterial action of thymol and cymene on two Bacillus cereus strains (INRA-AVTZ415 and INRA-AVZ421) was studied. Thymol and cymene alone showed higher antibacterial effect on the growth of B. cereus in BHI at 30 °C, both in lag and in exponential growth phases, with increasing added concentrations. However, the two strains tested showed different sensitivity to these natural compounds. When thymol and cymene were applied simultaneously, it resulted in a greater antibacterial effect than when the compounds were applied separately. The synergistic behaviour of both compounds was also shown for lag phase B. cereus INRA-AVZ421 cells in food substrates (carrot juice, chicken soup and pumpkin cream) at 30 °C, though a clear food-related effect was observed. This study indicates the potential use of thymol and cymene applied simultaneously for preservation of minimally processed foods.

Different inactivation kinetics data have been used to predict the number of survivors exposed to a heat treatment and, in consequence, to design thermal processes for the food industry. In this work, spores of an acidophilic strain of Bacillus subtilis were heated under isothermal and non-isothermal conditions. Experimental results obtained after isothermal treatments were analysed using the classical two-step linear regression procedure and a one-step non-linear regression method. Data obtained after non-isothermal treatments were analysed using a one-step, non-linear procedure. Kinetic parameters obtained from isothermal heating were close, either using the two-step linear regression (D 100=6.5 min) or the one-step non-linear regression (D 100=6.3 min), although the second method gave smaller 95% confidence intervals. The z values derived from non-isothermal heating were higher than those obtained in isothermal conditions (z=9.3 °C for non-isothermal heating at 1 °C/min versus z=7.7 °C for isothermal heating one step non-linear regression). Results were validated with experimental data obtained after different heat treatments, consisting of a phase of temperature increase at a fixed rate, followed by a holding phase. Non-isothermal methods predicted accurately the number of survivors after the heating ramp, while isothermal methods were more accurate for the holding phase of the treatment. When a temperature profile of a typical heat treatment process applied in the food industry was simulated, all predictions were on the safe side.

A new microbiological predictive model was developed, relating pH, NaCl concentration and temperature (T) with the decimal reduction coefficient D as dependent variable, using data previously reported by [1]. The model was obtained by multiple linear regression using only three predictor variables (T 2, pH2 and NaCl), easily computed from the basic variables pH, NaCl concentration and temperature (T). The fitted model is robust, stable and satisfies all the basic hypotheses of the regression models. It also provides simplicity when compared with previously published models based on multiple linear regression tools.

Inactivation of prions responsible of bovine spongiform encephalytis (BSE) by heat is a major concern for the canned food industry in Europe. Experimental data of heat inactivation of prions obtained from the scientific literature have been modelled. Two models were applied to analyse the inactivation curves, the Weibull distribution model and a two-parameter empirical model. Statistical analysis of available data indicated that the Weibull frequency distribution provided the best description of non-linear survival curves. The effect of the temperature on Weibull model parameters ( a and b) was also studied. The shape parameter, b, indicates deviations from linearity in the inactivation curves, and the scale parameter, a, can be considered as a kinetic constant rate for the inactivation. The values of the b parameter were on the order of 0.15-0.2 which meant that these curves had a strong upper concavity. The a values obtained ranged from 1.8 × 10−3 to 2.3 × 10−6 for temperatures between 100 and 160°C. These results indicate that temperature has a relevant effect on the inactivation of prions, although it remains a considerable fraction of prions with infective capacity. The Weibull frequency distribution model appears as a useful and convenient model because the a parameter could allow a quantitative comparison of the inactivation of prions at different temperatures or under different conditions.

The effect of irradiation with accelerated electrons on Bacillus cereus and Bacillus subtilis spore counts and spore heat resistance was investigated. Spore counts reduced approximately two log cycles for B. cereus and up to five log cycles for B. subtilis after a radiation dose of 7.6 kGy, with D10 values ranging from 1.5 to 3.8 kGy. Radiation had an important heat-sensitising effect on bacterial spores. After a 3.3 kGy treatment DT values were reduced more than three times for both B. cereus and B. subtilis spores. This study indicates the potential use of accelerated electrons, alone or followed by a heat treatment, for food preservation.

The heat-adaptive response of the psychrotrophic spoilage bacterium Bacillus weihenstephanensis DSM11827 is described. It is demonstrated that vegetative cells of B. weihenstephanensis adapts to heat exposure at 47 degrees C by prior exposure to heat at the nonlethal temperature of 38 degrees C. For this adaptive response, protein synthesis is required and maximum adaptation was noted after 15 min to 2 h prior exposure at 38 degrees C. By using two-dimensional gel electrophoresis (2D-E), an overview of the heat-shock proteins (HSPs) of B. weihenstephanensis was obtained and it was shown that the production of 15 proteins increased upon exposure to 38 degrees C. In more detail, the use of specific antibodies revealed induction of the HSPs DnaK, DnaJ, GroEL, ClpC, ClpP and ClpX of B. weihenstephanensis. In addition, also pre-exposure to other stresses than heat, such as exposure to a high salt concentration, low pH, a high ethanol concentration or low temperature, resulted in development of increased heat tolerance of B. weihenstephanensis, and during these conditions, an increased production of some HSPs was noted. This phenomenon of cross-protection might be of substantial importance in relation to the design of safe minimal processing regimes.

To monitor the ability of the food-borne opportunistic pathogen Bacillus cereus to survive during minimal processing of food products, we determined its heat-adaptive response. During pre-exposure to 42°C, B. cereus ATCC 14579 adapts to heat exposure at the lethal temperature of 50°C (maximum protection occurs after 15 min to 1 h of pre-exposure to 42°C). For this heat-adaptive response, de novo protein synthesis is required. By using two-dimensional gel electrophoresis, we observed 31 heat-induced proteins, and we determined the N-terminal sequences of a subset of these proteins. This revealed induction of stress proteins (CspB, CspE, and SodA), proteins involved in sporulation (SpoVG and AldA), metabolic enzymes (FolD and Dra), identified heat-induced proteins in related organisms (DnaK, GroEL, ClpP, RsbV, HSP16.4, YflT, PpiB, and TrxA), and other proteins (MreB, YloH, and YbbT). The upregulation of several stress proteins was confirmed by using antibodies specific for well-characterized heat shock proteins (HSPs) of B. subtilis . These observations indicate that heat adaptation of B. cereus involves proteins that function in a variety of cellular processes. Notably, a 30-min pre-exposure to 4% ethanol, pH 5, or 2.5% NaCl also results in increased thermotolerance. Also, for these adaptation processes, protein synthesis is required, and indeed, some HSPs are induced under these conditions. Collectively, these data show that during mild processing, cross-protection from heating occurs in pathogenic B. cereus , which may result in increased survival in foods.

The influence of mild heat pretreatment on the bactericidal action of nisin, carvacrol and thymol on stationary and exponential phase cells of two Bacillus cereus strains was studied. Carvacrol or thymol concentrations of 0.3 mmol/L had no bactericidal effect on unheated, and a minor effect on heated B. cereus cells either in stationary or exponential growth phases. Nisin (0.15 mg/mL) alone decreased the viable counts in all cases (unheated and heated cells in stationary and exponential phase) although the two strains tested showed different sensitivity to this natural antimicrobial between them. When carvacrol or thymol were combined with nisin, it resulted in a greater loss of viability of heated cells than when nisin was applied alone. There was a synergistic effect of nisin and both essential oils with a mild heat pretreatment on the viability of B. cereus cells. This study pointed out the potential use of nisin alone and in combination with carvacrol or thymol applied after a mild heat pretreatment for preservation of minimally processed foods.

A risk analysis was applied to experimental heat resistance data. This analysis is an approach for processing experimental thermobacteriological data in order to study the variability of D and z values of target microorganisms depending on the deviations range of environmental factors, to determine the critical factors and to specify their critical tolerance. This analysis is based on sets of sensitivity functions applied to a specific case of experimental data related to the thermoresistance of Clostridium sporogenes and Bacillus stearothermophilus spores. The effect of the following factors was analyzed: the type of target microorganism; nature of the heating substrate; pH, temperature; type of acid employed and NaCl concentration. The type of target microorganism to be inactivated, the nature of the substrate (reference or real food) and the heating temperature were identified as critical factors, determining about 90% of the alteration of the microbiological risk. The effect of the type of acid used for the acidification of products and the concentration of NaCl can be assumed to be negligible factors for the purposes of engineering calculations. The critical non-uniformity in temperature during thermobacteriological studies was set as 0.5% and the critical tolerances of pH value and NaCl concentration were 5%. These results are related to a specific case study, for that reason their direct generalization is not correct.

Thermobacteriological studies using Bacillus stearotherrnophilus spores were carried out by heating the spores under isothermal and non-isothermal conditions followed by an isothermal period. Ex perimental data obtained after isothermal heating were analyzed using a two-step linear regression procedure and a one-step nonlinear regression method. Results obtained using both analytical tech niques were close, but the 90% interval of confidence for predictions was lower when the one-step nonlinear regression was used. These results indicated the convenience of using the one-step nonlin ear regression method to obtain thermal kinetic parameters for bacterial spores. Also, the parameters obtained by a non-isothermal program followed by an isothermal heating period were similar to those obtained after isothermal heating. Nevertheless, the joint confidence regions at 90% did not overlap, indicating that there are significant differences. The z values obtained under isothermal (7.0 °C) and non-isothermal (8.5 °C) conditions are close and characteristic for this microorganisms.

The effect of different concentrations of sodium chloride (NaCl) in the recovery medium on the apparent thermal resistance of Bacillus stearothermophilus spores (strain ATCC 12980) was studied. Spores were heated at different temperatures (115–125°C) and subcultured in medium with different NaCl concentrations [1, 2, 3 and 3.5% (w/v)]. The results indicate that increasing NaCl levels decreases the apparent heat resistance of the spores, over a temperature range of 115–125°C. No significant difference was found between 3 and 3.5% (w/v) NaCl although both reduced B. stearothermophilus heat resistance considerably. A second-order polynomial equation provided a good description of the spores’ D value, as influenced by the temperature and NaCl concentration. Predictions from the model compared well with published data, and therefore the model is suitable for predicting the effect of NaCl concentration on thermal inactivation of B. stearothermophilus.

The effect of recovering Bacillus stearothermophilus spores under anaerobic conditions on their apparent thermal resistance was studied. Spores were suspended in bidistilled water as a reference medium, heated at 115, 117, 119, 121, 123 and 125°C and recovered under aerobic and anaerobic conditions. D values (decimal reduction time) obtained following recovery under anaerobic conditions were lower than those obtained under aerobic conditions. Reductions of between 31 and 48% were found for all the temperatures studied. When spores were suspended in mushroom extract and recovered under anaerobic conditions the apparent heat resistance was much lower than that obtained under aerobic conditions (D121°C was 4.3 min and 1.7 min, under aerobic and anaerobic conditions, respectively). Heating the spores in mushroom extract and recovering the spores under anaerobic conditions produced an additive effect, decreasing the apparent heat resistance of the B. stearothermophilus spores.

A time temperature integrator (TTI) was developed by immobilizing Bacillus stearothermophilus spores in a cylindrical particle consisting of an alginate-starch-mushroom puree. The particle showed homogeneous spore distribution, and when heated over a temperature range of 121 – 130° C negligible spore leakage was observed after the thermal process. The experimental data on spore survivor levels obtained for each temperature-time combination were compared with theoretical predictions using a mathematical model. The results showed a good correlation between the experimental and theoretical data. All these results provide evidence that this artificial particle could be a very reliable TTI for monitoring the thermal impact on micro-organisms during validating sterilization processes in continuous aseptic systems.

Bacillus weihenstephanensis is a psychrotolerant bacterium belonging to the Bacillus cereus group. Some strains may be cytotoxic although they have not been described as food-poisoning agents so far. The objective of this work is to model the effects of temperature and a w downshifts on the lag time of B. weihenstephanensis and the dependence of μ max on the growth conditions (temperature and a w ). Effects of temperature downshifts were studied on 30 experimental conditions (shifts magnitude ranging from 2 to 20 °C, temperature after shift from 10 to 20 °C and a w ranging from 0.977 to 0.997). Osmotic shifts were studied for 13 conditions (shift magnitude ranging from 0.008 to 0.020 units of a w , temperature from 10 to 30 °C, a w after shift from 0.977 to 0.997). Experimental results show that temperature downshifts were able to induce considerable lag times (up to 20 days) when occurring near the growth limits. At lower temperatures, osmotic shifts had also a significant effect. Validation experiments in food subjected to changing conditions of temperature showed that the model provided valid predictions in diluted creamed pasta but overestimated bacterial growth in carrot soup (fail safe predictions).

Two different effects of calcium were studied, respectively, in plasma membrane vesicles and in protoplasts isolated from roots of control pepper plants (Capsicum annuum L cv. California) or of plants treated with 50 mM NaCl, 10 mM CaCl(2) or 10 mM CaCl(2) + 50 mM NaCl. Under saline conditions, osmotic water permeability (P ( f )) values decreased in protoplasts and plasma membrane vesicles, and the same reduction was observed in the PIP1 aquaporin abundance, indicating inhibitory effects of NaCl on aquaporin functionality and protein abundance. The cytosolic Ca(2+) concentration, [Ca(2+)](cyt), was reduced by salinity, as observed by confocal microscope analysis. Two different actions of Ca(2+) were observed. On the one hand, increase in free cytosolic calcium concentrations associated with stress perception may lead to aquaporin closure. On the other hand, when critical requirements of Ca(2+) were reduced (by salinity), and extra-calcium would lead to an upregulation of aquaporins, indicating that a positive role of calcium at whole plant level combined with an inhibitory mechanism at aquaporin level may work in the regulation of pepper root water transport under salt stress. However, a link between these observations and other cell signalling in relation to water channel gating remains to be established.

Natural antimicrobials are an alternative to the use of chemically synthesized preservatives and other technological treatments. They have the advantage of not being rejected by consumers because of their natural origin. However, prior to their usage at a food factory scale, their precise effects on microorganisms have to be known. Nisin, a bacteriocin, and carvacrol and thymol, phenolic compounds present in the essential oil fraction of Oreganum and Thymus plants, have been shown to inhibit growth of different bacteria. This research was conducted to show the effect of a thermal treatment, applied previously, on the growth of B. megaterium cells in a culture medium with and without nisin, carvacrol and thymol. It has been shown that a thermal treatment able to kill 90 % of the spore population of B. megaterium had little effect on the growth of the survivors in nutrient broth at 30 °C. Nisin, in a concentration of 0.05 mM, did not affect greatly the growth of unheated B. megaterium. When the same concentration of nisin was applied to the survivors of the heat treatment, lag phase was further increased and growth rate decreased. Thymol, applied at a concentration of 0.6 mM, increased the lag phase duration almost three times, although it did not change the growth rate. Although carvacrol, at the same concentration (0.6 mM), did reduce the growth rate to half its original value, it did not increase the lag phase duration significantly (only twice), resulting in a quite similar growth curve. The combination of thymol and carvacrol at these high doses (0.6 mM) resulted in a further increase of the lag phase and significant decrease of the growth rate. When carvacrol and/or thymol were combined with a previous thermal treatment (able to kill 90 % of the population), the growth of the survivors was inhibited for at least seven days. Therefore a combination of moderate conditions could be effective to control spoilage by sporeforming bacteria., Prirodna antimikrobna sredstva su alternativa kemijski sintetiziranim konzervansima i drugim tehnološkim postupcima. Njihova je prednost u tome što su zbog svoga prirodnog podrijetla prihvatljivi potrošačima. Prije njihove primjene u prehrambenoj industriji, potrebno je znati kako djeluju na mikroorganizme. Nisin, bakteriocin, te karvakrol i timol, fenolni spojevi prisutni u frakcijama eteričnog ulja biljaka Oreganum i Thymus, inhibiraju rast raznih bakterija. Opisano je istraživanje provedeno da bi se utvrdio učinak prethodno provedene toplinske obrade na rast stanica B. megaterium u podlozi s nisinom, karvakrolom i timolom te bez njih. Pokazalo se da toplinska obrada može usmrtiti 90 % populacije spora B. megaterium, a gotovo nikako ne utječe na rast preživjelih u hranjivoj podlozi pri 30 °C. 22 P.M. PERIAGO et al.: B. megaterium Spore Germination and Growth Inhibition, Food Technol. Biotechnol. 44 (1) 17–23 (2006) Nisin koncentracije od 0,05 mM nije bitno utjecao na rast toplinski neobra|enog B. megaterium. Kada se ista koncentracija nisina primijeni na preživjele spore, produlji se lag faza, a smanji brzina rasta. Timol koncentracije od 0,6 mM produžio je trajanje lag faze skoro 3 puta, a nije promijenio brzinu rasta. Iako je karvakrol iste koncentracije (0,6 mM) smanjio brzinu rasta na polovinu originalne vrijednosti, dužinu lag faze produžio je samo za 2 puta, što je rezultiralo skoro jednakom (0,6 mM) krivuljom rasta. Kombinacijom tih velikih koncentracija timola i karvakrola znatno je produljena lag faza i smanjena brzina rasta. Kada se karvakrol i/ili timol kombiniraju s prethodnom toplinskom obradom (pri čemu je uništeno 90 % populacije), rast preživjelih bio je inhibiran barem 7 dana. Stoga kombinacija jednostavnih uvjeta može biti djelotvorna u kontroli onečišćenja sa sporulirajućim bakterijama

6 pages, 3 tables, 4 figures., Dietary ellagic acid and related polyphenols are metabolized in humans to dibenzopyran-6-one derivatives, and the microbial origin of these metabolites has been suggested. However, this has not been demonstrated so far. Fecal samples donated by six volunteers were incubated under anaerobic conditions, and aliquots were used to evaluate the fecal metabolism of ellagic acid, the ellagitannin punicalagin, and an ellagitannin rich extract from walnuts. The isoflavone daidzein was also incubated with the same fecal samples to follow the production of the microbial metabolites previously reported (dihydrogenistein, O-demethylangolensin, and equol) as a positive control of the system and to evaluate similarities between isoflavone and ellagic acid fecal flora metabolism. After fermentation the metabolite “urolithin A” (3,8-dihydroxy-6H-dibenzo[b,d]pyran-6-one) was produced from ellagic acid, punicalagin, and the ellagitannin extract in all the fecal cultures from different volunteers, but with very different production rates and concentrations. This large variability in the concentration of metabolite and kinetics of metabolite production is consistent with the large variability found in the excretion of these metabolites in urine in vivo after human consumption of ellagitannins, and with differences in the composition of the fecal microflora. No correlation between isoflavone and ellagic acid metabolism by fecal microflora was observed. The present study confirms the microbial origin of the recently reported in vivo generated hydroxy-6H-dibenzo[b,d]pyran-6-one derivatives in humans and is a further step in the study of the bioavailability and metabolism of ellagic acid and ellagitannins., This investigation has been supported by Spanish CICYT, projects AGL2003-02195 and AGL-2004-03989, and by the “Fundación Séneca”, project PB/18/FS/02. B.C. and P.P. have fellowships from the Spanish “Consejo Superior de Investigaciones Científicas” (CSIC) and ESF (I3P Programme).

The isothermal survival curves of the heat resistant spores of Bacillus sporothermodurans IC4, in the range of 117–125 °C, were determined in chicken, mushroom and pea soups by the capillary method. They were all non-linear with a noticeable upper concavity and could be described by the equation log 10 [ N ( t )/ N 0 ]=− b ( T ) t n with a fixed power, n , of the order of 0.7–0.8. The temperature dependence of b ( T ) could be described by the equation b ( T )=log e {1+exp[ k ( T − T c )]}, where T c is the temperature where intensive inactivation starts and k is the slope of b ( T ) at temperatures well above T c . They were 121–123 °C and 0.2–0.4 °C −1 , respectively, depending on the soup. These parameters were used to estimate the survival curves of the spores in two non-isothermal heat treatments using the procedure originally proposed by Peleg and Penchina [Crit. Rev. Food Sci. Nutr. 40 (2000) 159]. The results were compared with experimental survival curves, determined by the direct injection method, in another laboratory. There was a general agreement, although not perfect, between the predicted and experimentally observed survival ratios. Also, the isothermal survival parameters, estimated directly from the non-isothermal inactivation data using the model, were in general agreement with those calculated from the isothermal data. This suggests that the heat inactivation patterns of B. sporothermodurans IC4 spores in soups can be at least roughly estimated using the same general survival model, which has until now only been experimentally validated for vegetative bacterial cells.