Detection of risk areas in dairy powder processes: The development of thermophilic spore forming bacteria taking into account their growth limits.

Anoxybacillus flavithermus, Geobacillus stearothermophilus and Bacillus licheniformis are the main contaminants found in dairy powders. These spore-forming thermophilic bacteria, rarely detected in raw milk, persist, and grow during the milk powder manufacturing process. Moreover, in the form of spores, these species resist and concentrate in the powders during the processes. The aim of this study was to determine the stages of the dairy powder manufacturing processes that are favorable to the growth of such contaminants. A total of 5 strains were selected for each species as a natural contaminant of dairy pipelines in order to determine the minimum and maximum growth enabling values for temperature, pH, and aw and their optimum growth rates in milk. These growth limits were combined with the environmental conditions of temperature, pH and aw encountered at each step of the manufacture of whole milk, skim milk and milk protein concentrate powders to estimate growth capacities using cardinal models and the Gamma concept. These simulations were used to theoretically calculate the population sizes reached for the different strains studied at each stage in between two successive cleaning in place procedures. This approach highlights the stages at which risk occurs for the development of spore-forming thermophilic bacterial species. During the first stages of production, i.e. pre-treatment, pasteurization, standardization and pre-heating before concentration, physico-chemical conditions encountered are suitable for the development and growth of A. flavithermus, G. stearothermophilus and B. licheniformis. During the pre-heating stage and during the first effects in the evaporators, the temperature conditions appear to be the most favorable for the growth of G. stearothermophilus. The temperatures in the evaporator during the last evaporator effects are favorable for the growth of B. licheniformis. In the evaporation stage, low water activity severely limits the development of A. flavithermus.

Development and Evaluation of a Preliminary Screening Assay for Antibiotic Residues in Meat.

A preliminary screening assay based on a microbial chromogenic reaction was developed to detect common antibiotic residues in meat rapidly. The assay comprised two bioassays: one for Escherichia coli and another for Geobacillus stearothermophilus. The assay was optimized and evaluated for the simultaneous screening of 30 antibiotics from five common antibiotic classes (tetracyclines, aminoglycosides, macrolides, ß-lactams, and quinolones) found in meat. Extraction using phosphate-acetonitrile buffer (pH 7.2) and a delipidating treatment using n-hexane resulted in a high extraction efficacy for the five antibiotics, without affecting the microbial color reaction. A carrier, polyvinyl alcohol (0.1 g/mL); a cross-linking agent, boric acid-sodium tetraborate solution (pH 5.5); and a bacterial suspension with an initial optical density of 1.0 were the optimal embedding conditions for stability, microbial activity, and chromogenic efficiency. The assay exhibited a 6-month shelf life, with detection limits of 40-60, 60-140, 60-100, 20-40, and 40-180 µg/kg for tetracyclines, aminoglycosides, macrolides, ß-lactams, and quinolones, respectively, which met the European Commission (37/2010) requirements for antibiotic residue limits. Our assay results were confirmed using LC-MS/MS with 160 samples, revealing a good correlation. This study demonstrates a reliable, easy-to-use, and economical method for preliminary screening of antibiotic residues in meat. This method may find an immediate application in food safety and general testing laboratories.

Combined calorimetric gas- and spore-based biosensor array for online monitoring and sterility assurance of gaseous hydrogen peroxide in aseptic filling machines.

A combined calorimetric gas- and spore-based biosensor array is presented in this work to monitor and evaluate the sterilization efficacy of gaseous hydrogen peroxide in aseptic filling machines. H2O2 has been successfully measured under industrial conditions. Furthermore, the effect of H2O2 on three different spore strains , namely Bacillus atrophaeus, Bacillus subtilis and Geobacillus stearothermophilus, has been investigated by means of SEM, AFM and impedimetric measurements. In addition, the sterilization efficacy of a spore-based biosensor and the functioning principle are addressed and discussed: the sensor array is convenient to be used in aseptic food industry to guarantee sterile packages.

The effect of phosphate on the heat resistance of spores of dairy isolates of Geobacillus stearothermophilus.

In this study, we show that phosphate decreases the spore heat resistance by accelerating the rate of loss of cations from spores. Heat resistance of spores of Geobacillus stearothermophilus A1, D1, P3 and ATCC 12980 were determined in distilled water containing varying concentrations (0.1, 1 and 2% w/v) of di­sodium phosphate. The average decimal reduction times (D value) for strains A1, D1, P3 and ATCC 12980 in distilled water were 5.8, 6.8, 5.7 and 9 min at 110 °C respectively. On the addition of 0.1, 1 and 2% w/v of di­sodium phosphate, the average D110 values of all the strains in distilled water were lowered by 50, 61 and 70% respectively. Addition of 0.05% w/v of Na-EDTA to distilled water resulted in lowering of the average D110 value of all the strains by 55%. Heat resistance of spores of A1, D1, P3 and ATCC 12980 was found to be associated with the Dipicolinic Acid (DPA) content whose concentrations were 0.25, 0.30, 0.27 and 1.6 pg per spore respectively. Analysis by atomic absorption spectroscopy revealed that the phosphate present in the heating medium causes excess release of calcium from spores with 2% w/v phosphate being highly effective, thus confirming the chelating effect of phosphate. This study provides insight into the heat resistance and the increased heat sensitivity of spores of G. stearothermophilus A1, D1 and P3 in the presence of phosphate, which can be used in the design of Cleaning in Place (CIP) systems involving phosphate based cleaning agents to combat biofilms and spores in the dairy industry.

Aspartic acid racemization constrains long-term viability and longevity of endospores.

Certain microorganisms survive long periods of time as endospores to cope with adverse conditions. Since endospores are metabolically inactive, the extent of aspartic acid (Asp) racemization will increase over time and might kill the spores by preventing their germination. Therefore, understanding the relationship between endospore survivability and Asp racemization is important for constraining the long-term survivability and global dispersion of spore-forming bacteria in nature. Geobacillus stearothermophilus was selected as a model organism to investigate racemization kinetics and survivability of its endospores at 65°C, 75°C and 98°C. This study found that the Asp racemization rates of spores and autoclaved spores were similar at all temperatures. The Asp racemization rate of spores was not significantly different from that of vegetative cells at 65°C. The Asp racemization rate of G. stearothermophilus spores was not significantly different from that of Bacillus subtilis spores at 98°C. The viability of spores and vegetative cells decreased dramatically over time, and the mortality of spores correlated exponentially with the degree of racemization (R2 = 0.9). This latter correlation predicts spore half-lives on the order of hundreds of years for temperatures typical of shallow marine sediments, a result consistent with studies about the survivability of thermophilic spores found in these environments.

Influence and characteristics of Bacillus stearothermophilus in ammonia reduction during layer manure composting.

Ammonia emissions is an important issue during composting because it can cause secondary pollution and a significant of nitrogen loss. Based on research adding Bacillus stearothermophilus can reduce ammonia emissions during composting because it can use sugar in organic matter fermentation to produce organic acids over 50 °C. This study conducted the batch experiments by adding different concentrations of Bacillus stearothermophilus to reduce the ammonia emissions and find out its characteristic during layer manure composting by using an aerobic composting reactor with sawdust as a bulking agent. The results show that the application of Bacillus stearothermophilus can accelerate the rate of temperature and significantly decrease pH, the warming period was 2 days in the treatment with Bacillus stearothermophilus, while it was 4 days in the treatment without Bacillus stearothermophilus. Ammonia emissions were mainly occurred in warming and high temperature period during composting. The ammonia emissions in the treatment with 8.00 g/kg initial Bacillus stearothermophilus were significantly lower than the other lower Bacillus stearothermophilus treatment and control during composting (p < 0.05), and it can significantly increase ammonium-nitrogen and nitrate-nitrogen concentration, reduce pH (p < 0.05), but the average number of Bacillus stearothermophilus copies in treatment with different initial Bacillus stearothermophilus concentration had no significant difference (p > 0.05). MiSeq System Sequencing results find that the addition of Bacillus stearothermophilus changed the bacterial community structure under warming and high-temperature periods during composting, increased the relative abundance of lactic acid bacillus and nitrification bacteria. Therefore, the reason for the low ammonia emission in 8.00 g/kg initial Bacillus stearothermophilus treatments might be not only due to the Bacillus stearothermophilus itself, but also Bacillus stearothermophilus can change the indigenous microorganism community, including increase the relative content of lactic acid Bacillus and nitrification bacteria, thus reducing the pH and promoting nitrification, and reducing ammonia emissions.

The fate of Bacillus cereus and Geobacillus stearothermophilus during alkalization of cocoa as affected by alkali concentration and use of pre-roasted nibs.

Alkalization is a step of cocoa processing and consists of the use of alkali and high temperature to improve the sensorial and technological qualities of cocoa. Intense food processing can select spores, which can compromise safety and quality of the final product. Thus, the aim of this study was to evaluate the fate of B. cereus and G. stearothermophilus spores during the alkalization of pre-roasted (Pr) nibs (held at 120 °C) and unroasted (Ur) nibs (held at 90 °C) using potassium carbonate (0, 2, 4 and 6% w/w). In all conditions, log-linear inactivation kinetics with a tail was observed. The inactivation rate (kmax) for B. cereus varied from 0.065 to 1.67 min-1, whereas the kmax for G. stearothermophilus varied from 0.012 to 0.063 min-1. For both microorganisms, the lowest kmax values were observed during Ur nibs alkalization. The carbonate concentration increase promoted kmax values reduction. The highest tail values were observed for G. stearothermophilus in Ur nibs alkalization, reaching 3.04 log spores/g. Tail formation and low kmax values indicated that cocoa alkalization does not cause significant reductions on bacterial spore population. Therefore, the microbiological control should be primarily ensured by the raw material quality and by avoiding recontamination in the cocoa chain.

A quantitative method to assess the role of indoor air decontamination to simultaneously reduce contamination of environmental surfaces: testing with vegetative and spore-forming bacteria.

Indoor air can spread pathogens, which can be removed/inactivated by a variety of means in healthcare and other settings. We quantitatively assessed if air decontamination could also simultaneously reduce environmental surface contamination in the same setting. Two types of vegetative bacteria (Staphylococcus aureus and Acinetobacter baumannii), and a bacterial spore-former (Geobacillus stearothermophilus) were tested as representative airborne bacteria. They were separately aerosolized with a Collison nebulizer into a 24-m3 aerobiology chamber and air samples collected with a programmable slit-to-agar sampler. Settling airborne particles were collected on culture plates placed at, and collected from, five different locations on the floor of the chamber with a custom-built remote plate-placement and -retriever system. Experimentally contaminated air in the chamber was decontaminated for 45 min with a device based on HEPA filtration and UV light. The plates were incubated and CFU counted. The device reduced the viability levels of all tested bacteria in the air by >3 log10 (>99·9%) in 45 min. Based on two separate tests, the average reductions in surface contamination for S. aureus, A. baumannii and G. stearothermophilus were respectively, 97, 87 and 97%. We thus showed that air decontamination could substantially and simultaneously reduce the levels of surface contamination in the same setting irrespective of the type of pathogen present. SIGNIFICANCE AND IMPACT OF THE STUDY: The innovative and generic test protocol described can quantitatively assess the reduction in environmental surface contamination from microbial decontamination of indoor air in the same setting. This added advantage from air decontamination has implications for infection prevention and control in healthcare and other settings without the need for additional expense or effort. Continuous operation of an air decontamination device, such as the one tested here, can lead to ongoing reductions in pathogens in air and on environmental surfaces.

Mapping the risk of evaporated milk spoilage in the Mediterranean region based on the effect of temperature conditions on Geobacillus stearothermophilus growth.

A predictive model for the effect of storage temperature on the growth of Geobacillus stearothermophilus was applied in order to assess the risk of evaporated milk spoilage in the markets of the Mediterranean region. The growth of G. stearothermophilus in evaporated milk was evaluated during a shelf life of one year based on historical temperature profiles (hourly) covering 23 Mediterranean capitals for five years over the period 2012-2016 obtained from the Weather Underground database (http://www.wunderground.com/). In total, 115 scenarios were tested simulating the distribution and storage conditions of evaporated milk in the Mediterranean region. The highest growth of G. stearothermophilus was predicted for Marrakech, Damascus and Cairo over the period 2012-2016 with mean values of 7.2, 7.4 and 5.5 log CFU/ml, respectively, followed by Tunis, Podgorica and Tripoli with mean growth of 2.8, 2.4 and 2.3 log CFU/ml, respectively. For the rest 17 capitals the mean growth of the spoiler was <1.5 log CFU/ml. The capitals Podgorica, Cairo, Tunis and Ankara showed the highest variability in the growth during the 5 years examined with standard deviation values for growth of 2.01, 1.79, 1.77 and 1.25 log CFU/ml, respectively. The predicted extent and the variability of growth during the shelf life were used to assess the risk of spoilage which was visualised in a geographical risk map. The growth model of G. stearothermophilus was also used to evaluate adjustments of the evaporated milk expiration date which can reduce the risk of spoilage. The quantitative data provided in the present study can assist the food industry to effectively evaluate the microbiological stability of these products throughout distribution and storage at a reduced cost (by reducing sampling quality control) and assess whether and under which conditions (e.g. expiration date) will be able to export a product to a country without spoilage problems. This decision support may lead to a significant benefit for both the competitiveness of the food industry and the consumer.

Scrubbing technique for needleless connectors to minimize contamination risk.

This study aimed to investigate the appropriate scrubbing technique for needleless connectors to minimize contamination risk. To demonstrate a highly effective scrubbing technique to physically eliminate bacteria, needleless connectors were contaminated with Geobacillus stearothermophilus spores and then scrubbed. The study showed that the highest bacterial elimination rate was achieved by scrubbing an access port in a straight line with an alcohol cotton swab, applying a force that was almost equal to an arterial compression haemostasis to the access port, and repeating this procedure once using a new alcohol cotton swab.

Bacteria, mould and yeast spore inactivation studies by scanning electron microscope observations.

Spores are the most resistant form of microbial cells, thus difficult to inactivate. The pathogenic or food spoilage effects of certain spore-forming microorganisms have been the primary basis of sterilization and pasteurization processes. Thermal sterilization is the most common method to inactivate spores present on medical equipment and foods. High pressure processing (HPP) is an emerging and commercial non-thermal food pasteurization technique. Although previous studies demonstrated the effectiveness of thermal and non-thermal spore inactivation, the in-depth mechanisms of spore inactivation are as yet unclear. Live and dead forms of two food spoilage bacteria, a mould and a yeast were examined using scanning electron microscopy before and after the inactivation treatment. Alicyclobacillus acidoterrestris and Geobacillus stearothermophilus bacteria are indicators of acidic foods pasteurization and sterilization processes, respectively. Neosartorya fischeri is a phyto-pathogenic mould attacking fruits. Saccharomyces cerevisiae is a yeast with various applications for winemaking, brewing, baking and the production of biofuel from crops (e.g. sugar cane). Spores of the four microbial species were thermally inactivated. Spores of S. cerevisiae were observed in the ascus and free form after thermal and HPP treatments. Different forms of damage and cell destruction were observed for each microbial spore. Thermal treatment inactivated bacterial spores of A. acidoterrestris and G. stearothermophilus by attacking the inner core of the spore. The heat first altered the membrane permeability allowing the release of intracellular components. Subsequently, hydration of spores, physicochemical modifications of proteins, flattening and formation of indentations occurred, with subsequent spore death. Regarding N. fischeri, thermal inactivation caused cell destruction and leakage of intracellular components. Both thermal and HPP treatments of S. cerevisiae free spores attacked the inner membrane, altering its permeability, and allowing in final stages the transfer of intracellular components to the outside. The spore destruction caused by thermal treatment was more severe than HPP, as HPP had less effect on the spore core. All injured spores have undergone irreversible volume and shape changes. While some of the leakage of spore contents is visible around the deformed but fully shaped spore, other spores exhibited large indentations and were completely deformed, apparently without any contents inside. This current study contributed to the understanding of spore inactivation by thermal and non-thermal processes.

Effect of storage temperature on the lag time of Geobacillus stearothermophilus individual spores.

The lag times (λ) of Geobacillus stearothermophilus single spores were studied at different storage temperatures ranging from 45 to 59 °C using the Bioscreen C method. A significant variability of λ was observed among individual spores at all temperatures tested. The storage temperature affected both the position and the spread of the λ distributions. The minimum mean value of λ (i.e. 10.87 h) was observed at 55 °C, while moving away from this temperature resulted in an increase for both the mean and standard deviation of λ. A Cardinal Model with Inflection (CMI) was fitted to the reverse mean λ, and the estimated values for the cardinal parameters Tmin, Tmax, Topt and the optimum mean λ of G. stearothermophilus were found to be 38.1, 64.2, 53.6 °C and 10.3 h, respectively. To interpret the observations, a probabilistic growth model for G. stearothermophilus individual spores, taking into account λ variability, was developed. The model describes the growth of a population, initially consisting of N0 spores, over time as the sum of cells in each of the N0 imminent subpopulations originating from a single spore. Growth simulations for different initial contamination levels showed that for low N0 the number of cells in the population at any time is highly variable. An increase in N0 to levels exceeding 100 spores results in a significant decrease of the above variability and a shorter λ of the population. Considering that the number of G. stearothermophilus surviving spores in the final product is usually very low, the data provided in this work can be used to evaluate the probability distribution of the time-to-spoilage and enable decision-making based on the "acceptable level of risk".

Walking dead: Permeabilization of heat-treated Geobacillus stearothermophilus ATCC 12980 spores under growth-preventing conditions.

Although heat treatment is probably the oldest and the most common method used to inactivate spores in food processes, the specific mechanism of heat killing of spores is still not fully understood. The purpose of this study is to investigate the evolution of the permeabilization and the viability of heat-treated spores during storage under growth-preventing conditions. Geobacillus stearothermophilus spores were heat-treated under various conditions of temperature and pH, and then stored under conditions of temperature and pH that prevent growth. Spore survival was evaluated by count plating immediately after heat treatment, and then during storage over a period of months. Flow cytometry analyses were performed to investigate the Syto 9 permeability of heat-treated spores. Sub-lethally heat-treated spores of G. stearothermophilus were physically committed to permeabilization after heat treatment. However, prolonged heat treatment may abolish the spore permeabilization and block heat-treated spores in the refractive state. However, viability loss and permeabilization during heat treatment seem to be two different mechanisms that occur independently, and the loss of permeabilization properties takes place at a much slower rate than spore killing. Under growth-preventing conditions, viable heat-treated spores presumably lose their viability due to the permeabilization phenomena, which makes them more susceptible to the action of adverse conditions precluding growth.

Spore-forming bacteria responsible for food spoilage.

This review explores the main spore-forming bacteria involved in the spoilage of various processed foods. Bakery products are specifically spoiled by Bacillus species, the dominant one being Bacillus amyloliquefaciens, while different Clostridium species classically contaminate refrigerated vacuum-packed meats. These two genera have also been isolated from milk products, even when milk is pasteurized, sterilized, dehydrated or fermented, according to heat treatment and storage temperature. Finally, the most heat-resistant microorganisms are isolated in low-acid canned foods, the three predominant species being Geobacillus stearothermophilus, Moorella thermoacetica and Thermoanaerobacterium spp.

Development and application of Geobacillus stearothermophilus growth model for predicting spoilage of evaporated milk.

The presence of Geobacillus stearothermophilus spores in evaporated milk constitutes an important quality problem for the milk industry. This study was undertaken to provide an approach in modelling the effect of temperature on G. stearothermophilus ATCC 7953 growth and in predicting spoilage of evaporated milk. The growth of G. stearothermophilus was monitored in tryptone soy broth at isothermal conditions (35-67 °C). The data derived were used to model the effect of temperature on G. stearothermophilus growth with a cardinal type model. The cardinal values of the model for the maximum specific growth rate were Tmin = 33.76 °C, Tmax = 68.14 °C, Topt = 61.82 °C and µopt = 2.068/h. The growth of G. stearothermophilus was assessed in evaporated milk at Topt in order to adjust the model to milk. The efficiency of the model in predicting G. stearothermophilus growth at non-isothermal conditions was evaluated by comparing predictions with observed growth under dynamic conditions and the results showed a good performance of the model. The model was further used to predict the time-to-spoilage (tts) of evaporated milk. The spoilage of this product caused by acid coagulation when the pH approached a level around 5.2, eight generations after G. stearothermophilus reached the maximum population density (Nmax). Based on the above, the tts was predicted from the growth model as the sum of the time required for the microorganism to multiply from the initial to the maximum level ( [Formula: see text] ), plus the time required after the [Formula: see text] to complete eight generations. The observed tts was very close to the predicted one indicating that the model is able to describe satisfactorily the growth of G. stearothermophilus and to provide realistic predictions for evaporated milk spoilage.

Plasma Decontamination: A Case Study on Kill Efficacy of Geobacillus stearothermophilus Spores on Different Carrier Materials.

A new technology to the pharmaceutical field is presented: surface decontamination by plasmas The technology is comparable to established barrier systems like e-beam, volatile hydrogen peroxide, or radiation inactivation of microbiological contaminations. This plasma technology is part of a fully automated and validated syringe filling line at a major pharmaceutical company and is in production operation. Incoming pre-sterilized syringe containers ("tubs") are processed by plasma, solely on the outside, and passed into the aseptic filling isolator upon successful decontamination. The objective of this article is to present the operating principles and develop and establish a validation routine on the basis of standard commercial biological indicators. Their decontamination efficacies are determined and correlated to the actual inactivation efficacy on the pharmaceutical packaging material.The reference setup is explained in detail and a short presentation of the cycle development and the relevant plasma control parameters is given, with a special focus on the in-process monitor determining the cycle validity. Different microbial inactivation mechanisms are also discussed and evaluated for their contribution and interaction to enhance plasma decontamination. A material-dependent inactivation behavior was observed. In order to be able to correlate the tub surface inactivation of Geobacillus stearothermophilus endospores to metallic biological indicators, a comparative study was performed. Through consistently demonstrating the linear inactivation behavior between the different materials, it becomes possible to develop an effective and time-saving validation scheme. LAY ABSTRACT: The challenge in new decontamination systems lies in a thorough validation of the inactivation efficacy under different operating regimes. With plasma, as an ionized gas, a new barrier concept is introduced into pharmaceutical aseptic processing of syringes. The presented system operates in vacuum and only decontaminates the outer surface of pre-sterilized syringe containers ("tubs"), before they are transferred into the aseptic area. The plasma does not penetrate into the tub. This article discusses the phase from development and test germ selection, across the identified sporicidal mechanisms, to a proposal for a validation scheme on the basis of commercially available biological indicators. A special focus is placed on an extensive investigation to establish a link between the tub surface microbial kill (polystyrene and Tyvek(and (2)) ) and biological indicator inactivation (stainless steel). Additionally, a rationale is developed on how an optical in-process monitor can be applied to establish a validatable limit on the base of the predetermined inactivation data of Geobacillus stearothermophilus endospores.

A sterilization system using ultraviolet photochemical reactions based on nitrous oxide and oxygen gases.

Active oxygen species (AOS) generated under ultraviolet (UV) lamps can be applied for various industrial processes owing to extremely strong oxidative abilities. We have already reported on an application of the AOS for a sterilization process of microorganisms. Here, a sterilization method using active oxygen generated under ultraviolet (UV) lamps introducing nitrous oxide (N2O) and oxygen gases into a vacuum chamber was investigated. Nitrogen dioxide (NO2) gas was readily produced from N2O by UV photochemical reactions under the low-pressure mercury lamp and then used to sterilize medical devices. We compared the ability of the N2O gas to sterilize Geobacillus stearothermophilus spores with those of conventional methods. Successful sterilization of spores on various biological indicators was achieved within 60 min, not only in sterilization bags but also in a lumen device.

Effect of heating rate on highly heat-resistant spore-forming microorganisms.

Highly heat-resistant spore-forming Bacillus cause nonsterility problems in canned food and reduce the shelf life of many processed foods. The aim of this research was to evaluate the thermal inactivation of Bacillus sporothermodurans IIC65, Bacillus subtilis IC9, and Geobacillus stearothermophilus T26 under isothermal and nonisothermal conditions. The data obtained showed that B. sporothermodurans and B. subtilis were more heat resistant than G. stearothermophilus. The survival curves of B. sporothermodurans and B. subtilis showed shoulders, while the survival curves of G. stearothermophilus showed tails. Under nonisothermal treatment, at heating rates of 1 and 20 ℃/min, time needed to completely inactivate G. stearothermophilus was shorter than that required for B. sporothermodurans and B. subtilis. In complex heat treatments (heating-holding-cooling), the survival curves of B. sporothermodurans and B. subtilis showed the same activation shoulders than those obtained under isothermal treatments and the activation shoulders were again absent in the case of G. stearothermophilus. Predictions fitted quite well the data obtained for B. sporothermodurans. In contrast, the data for B. subtilis showed half a log cycle more survival than expected and in the case of G. stearothermophilus, the survival curve obtained showed much higher inactivation than expected.

Estimation and evaluation of management options to control and/or reduce the risk of not complying with commercial sterility.

In a previous study, a modular process risk model, from the raw material reception to the final product storage, was built to estimate the risk of a UHT-aseptic line of not complying with commercial sterility (Pujol et al., 2015). This present study was focused on demonstrating how the model (updated version with uncertainty and variability separated and 2(nd) order Monte Carlo procedure run) could be used to assess quantitatively the influence of management options. This assessment was done in three steps: pinpoint which process step had the highest influence on the risk, identify which management option(s) could be the most effective to control and/or reduce the risk, and finally evaluate quantitatively the influence of changing process setting(s) on the risk. For Bacillus cereus, it was identified that during post-process storage in an aseptic tank, there was potentially an air re-contamination due to filter efficiency loss (efficiency loss due to successive in-place sterilizations after cleaning operations), followed by B. cereus growth. Two options were then evaluated: i) reducing by one fifth of the number of filter sterilizations before renewing the filters, ii) designing new UHT-aseptic lines without an aseptic tank, i.e. without a storage period after the thermal process and before filling. Considering the uncertainty in the model, it was not possible to confirm whether these options had a significant influence on the risk associated with B. cereus. On the other hand, for Geobacillus stearothermophilus, combinations of heat-treatment time and temperature enabling the control or reduction in risk by a factor of ca. 100 were determined; for ease of operational implementation, they were presented graphically in the form of iso-risk curves. For instance, it was established that a heat treatment of 138°C for 31s (instead of 138°C for 25s) enabled a reduction in risk to 18×10(-8) (95% CI=[10; 34]×10(-8)), instead of 578×10(-8) (95% CI=[429; 754]×10(-8)) initially. In conclusion, a modular risk model, as the one exemplified here with a UHT-aseptic line, is a valuable tool in process design and operation, bringing definitive quantitative elements into the decision making process.

Combined pressure-thermal inactivation effect on spores in lu-wei beef--a traditional Chinese meat product.

AIMS: This study investigated the inactivation effect and kinetics of Bacillus coagulans and Geobacillus stearothermophilus spores suspended in lu-wei beef by combining high pressure (500 and 600 MPa) and moderate heat (70 and 80 °C or 80 and 90 °C). METHODS AND RESULTS: During pressurization, the temperature of pressure-transmitting fluid was tested with a K-type thermocouple, and the number of surviving cells was determined by a plate count method. The pressure come-up time and corresponding inactivation of Bacillus coagulans and G. stearothermophilus spores were considered during the pressure-thermal treatment. For the two types of spores, the results showed a higher inactivation effect in phosphate buffer solution than that in lu-wei beef. Among the bacteria evaluated, G. stearothermophilus spores had a higher resistance than B. coagulans spores during the pressure-thermal processing. One linear model and two nonlinear models (i.e. the Weibull and log-logistic models) were fitted to the survivor data to obtain relevant kinetic parameters, and the performance of these models was compared. The results suggested that the survival curve of the spores could be accurately described utilizing the log-logistic model, which produced the best fit for all inactivation data. CONCLUSIONS: The compression heating characteristics of different pressure-transmitting fluids should be considered when using high pressure to sterilize spores, particularly while the pressure is increasing. Spores can be inactivated by combining high pressure and moderate heat. SIGNIFICANCE AND IMPACT OF THE STUDY: The study demonstrates the synergistic inactivation effect of moderate heat in combination with high pressure in real-life food. The use of mathematical models to predict the inactivation for spores could help the food industry further to develop optimum process conditions.