Clostridium Sporogenes Spores Research Articles

Utilisation of Lactococcus lactis subsp. lactis (PY91K) and Lactiplantibacillus plantarum (Y48) to control Clostridium sporogenes during Turkish white cheese production and storage

SummaryThis study investigated the inhibition of Clostridium species, isolated from cheeses with late‐blowing defects, by lactic acid bacteria (LAB). Four batches of Turkish white cheeses were produced; control cheese, cheese sample containing Lactococcus lactis subsp. lactis (PY91K) and Lactiplantibacillus plantarum (Y48) as protective cultures (LAB), a cheese sample containing the protective culture and inoculated with Clostridium sporogenes spores (CL‐LAB) and finally a cheese sample containing only Cl. sporogenes spores (CL). White cheese's microbiological and physicochemical properties were also determined throughout the 90‐day storage period at 4 °C. Approximately 1 log reduction in Cl. sporogenes (ZMP‐1‐4) spores was observed in the LAB co‐inoculated Turkish white cheese samples.

Hyperspectral imaging and deep learning for quantification of Clostridium sporogenes spores in food products using 1D- convolutional neural networks and random forest model

Clostridium sporogenes spores are used as surrogates for Clostridium botulinum, to verify thermal exposure and lethality in sterilization regimes by food industries. Conventional methods to detect spores are time-consuming and labour intensive. The objectives of this study were to evaluate the feasibility of using hyperspectral imaging (HSI) and deep learning approaches, firstly to identify dead and live forms of C. sporogenes spores and secondly, to estimate the concentration of spores on culture media plates and ready-to-eat mashed potato (food matrix). C. sporogenes spores were inoculated by either spread plating or drop plating on sheep blood agar (SBA) and tryptic soy agar (TSA) plates and by spread plating on the surface of mashed potato. Reflectance in the spectral range of 547–1701 nm from the region of interest was used for principal component analysis (PCA). PCA was successful in distinguishing dead and live spores and different levels of inoculum (102 to 106 CFU/ml) on both TSA and SBA plates, however, was not efficient on the mashed potato (food matrix). Hence, deep learning classification frameworks namely 1D- convolutional neural networks (CNN) and random forest (RF) model were used. CNN model outperformed the RF model and the accuracy for quantification of spores was improved by 4% and 8% in the presence and absence, respectively of dead spores. The screening system used in this study was a combination of HSI and deep learning modelling, which resulted in an overall accuracy of 90–94% when the dead/inactivated spores were present and absent, respectively. The only discrepancy detected was during the prediction of samples with low inoculum levels (<102 CFU/ml). In summary, it was evident that HSI in combination with a deep learning approach showed immense potential as a tool to detect and quantify spores on nutrient media as well as on specific food matrix (mashed potato). However, the presence of dead spores in any sample is postulated to affect the accuracy and would need replicates and confirmatory assays.

Sterilization of coconut milk in flexible packages via ohmic-assisted thermal sterilizer

Extreme heat treatments of canned coconut milk might reduce overall product quality. Therefore, an ohmic-assisted thermal sterilizer (OATS) may be able to preserve packaged coconut milk as an alternative to an overpressure retort. OATS involves converting electrical energy immediately into heat inside medium, which is then transferred to the food packages. This differs from conventional retorts which relied primarily on heat transfer. To ensure food safety, the OATS was microbiologically validated using Clostridium sporogenes spores. Physicochemical property and aroma compound profile differences in coconut milk sterilized at F0 of 5.2 min using OATS were compared with conventional water spray retort. OATS completely inactivated C. sporogenes spores with more than 5 log CFU/mL at 121.1 °C for 5 min. The total processing time for OATS was 27 min using 32% less energy compared to the 42 min for the retort, indicating the higher process efficiency of OATS. OATS food products surpassed conventional retort properties in appearance, with minor color changes; off-odor, having a lower cooked odor by reducing methyl-ketone and Maillard reaction products; and desired aromas, by preserving contributors such as ethyl octanoate and δ-lactones. Consequently, OATS could potentially produce sterile coconut milk with minimal changes in appearance while decreasing off-odor formation.

Safe cured meat using gamma radiation: Effects on spores of Clostridium sporogenes and technological and sensorial characteristics of low nitrite cooked ham

This study evaluated the effect of gamma radiation (0, 1.5, 3, 4.5, 6 KGy) and sodium nitrite (0, 50 and 150 mg/kg) on Clostridium sporogenes spores and on the technological and sensorial characteristics of restructured cooked hams. The number of spores recovered reduced (p < 0.05) with increasing doses of radiation but radiation sensitivity (D-values) was reduced from 1.98 kGy in uncured samples to 1.81 and 1.57 kGy in cured ones. Residual nitrite (RNO2) and pH were not affected (p > 0.05) by irradiation, but RNO2 was reduced (p < 0.05) with lower nitrite additions. CIE color measures and texture profile were affected only by nitrite levels and all sensory attributes of uncured samples were perceived (p < 0.05) as different from the cured ones. Irradiation improved (p > 0.05) the odor scores of cured samples. Irradiation of cooked hams at 3 kGy can be used to improve the sensory quality and ensure the microbiological safety of cooked hams elaborated with 50 mg/kg of nitrite.

Development of Bacterial Spore Pouches as a Tool to Evaluate the Sterilization Efficiency-A Case Study with Microwave Sterilization Using Clostridium sporogenes and Geobacillus stearothermophilus.

In this study, novel spore pouches were developed using mashed potato as a food model inoculated with either Geobacillus stearothermophilus or Clostridium sporogenes spores. These spore pouches were used to evaluate the sterilization efficiency of Coaxially induced microwave pasteurization and sterilization (CiMPAS) as a case study. CiMPAS technology combines microwave energy (915 MHz) along with hot water immersion to sterilize food in polymeric packages. The spore pouches were placed at pre-determined specific locations, especially cold spots in each food tray before being processed using two regimes (R-121 and R-65), which consisted of 121 °C and 65 °C at 12 and 22 kW, respectively, followed by recovery and enumeration of the surviving spores. To identify cold spots or the location for inoculation, mashed potato was spiked with Maillard precursors and processed through CiMPAS, followed by measurement of lightness values (*L-values). Inactivation equivalent to of 1–2 Log CFU/g and >6 Log CFU/g for Geobacillus stearothermophilus and Clostridium sporogenes spores, respectively was obtained on the cold spots using R-121, which comprised of a total processing time of 64.2 min. Whereas, inactivation of <1 and 2–3 Log CFU/g for G. stearothermophilus and C. sporogenes spores, respectively on the cold spots was obtained using R-65 (total processing time of 68.3 min), whereas inactivation of 1–3 Log CFU/g of C. sporogenes spores was obtained on the sides of the tray. The results were reproducible across three processing replicates for each regime and inactivation at the specific locations were clearly distinguishable. The study indicated a strong potential to use spore pouches as a tool for validation studies of microwave-induced sterilization.

Convergence of Bigelow and Arrhenius models over a wide range of heating temperatures

The heat resistance of the bacterial spores of Moorella thermoacetica, Clostridium sporogenes, Geobacillus stearothermophilus and Bacillus coagulans was determined over a wide range of temperatures using the capillary method and thermoresistometer Mastia. The results showed that the two experimental methods gave similar heat resistance values excepted for Geobacillus stearothermophilus.The effect of temperature on thermal resistance was evaluated using the Arrhenius and Bigelow models. The fit of the heat sensitivity parameters of the Arrhenius and Bigelow models on the heat resistance parameter values obtained over a wide temperature range was equally good. Despite the apparent mathematical incompatibility of the two equations, it is recognized that they yield the same goodness of fit. This paper finds a mathematical reason for this convergence and explains why inside a temperature range of at least 100 °C, no significant difference in the quality of fit between these two models can be found.

Sporicidal activities and mechanism of surfactant components against Clostridium sporogenes spores.

The sporicidal activities of seven kinds of antimicrobial agent were investigated in order to screen for novel inactivation agents to apply to Clostridium sporogenes spores. Antimicrobial agents based on surfactant components, as poly-l-lysine, thiamine dilaurylsulfate, and torilin, were more effective than other agents. The degree of spore reduction with 1-2% surfactant components was 1.5-2.5logCFU/mL. The HLB value (hydrophile-lipophile balance) related to denature protein of spores coat on surfactants with sporicidal activity was ranged from 6 to 16. Average HLB value and spore killing effect was inversely correlated. The proteins on spore structures seemed to be disorganized due to binding between polar groups of coats and hydrophilic and hydrophobic groups of surfactant components, resulting in killing of spores. The components that were effective to inactivate C. sporogenes spores had a chemical structure containing CH3, OH, COOH, sulfate groups, and a double bond. Furthermore, hydrophobic surfactants were more effective than hydrophilic surfactants in inactivating spores. This was likely due to the type of hydrophobic surfactant and to the involvement of hydrophobic interactions on coat of spores.

Benchmarking the minimum Electron Beam (eBeam) dose required for the sterilization of space foods

As manned space missions extend in length, the safety, nutrition, acceptability, and shelf life of space foods are of paramount importance to NASA. Since food and mealtimes play a key role in reducing stress and boredom of prolonged missions, the quality of food in terms of appearance, flavor, texture, and aroma can have significant psychological ramifications on astronaut performance. The FDA, which oversees space foods, currently requires a minimum dose of 44kGy for irradiated space foods. The underlying hypothesis was that commercial sterility of space foods could be achieved at a significantly lower dose, and this lowered dose would positively affect the shelf life of the product. Electron beam processed beef fajitas were used as an example NASA space food to benchmark the minimum eBeam dose required for sterility. A 15kGy dose was able to achieve an approximately 10log reduction in Shiga-toxin-producing Escherichia coli bacteria, and a 5log reduction in Clostridium sporogenes spores. Furthermore, accelerated shelf life testing (ASLT) to determine sensory and quality characteristics under various conditions was conducted. Using Multidimensional gas-chromatography–olfactometry–mass spectrometry (MDGC-O-MS), numerous volatiles were shown to be dependent on the dose applied to the product. Furthermore, concentrations of off –flavor aroma compounds such as dimethyl sulfide were decreased at the reduced 15kGy dose. The results suggest that the combination of conventional cooking combined with eBeam processing (15kGy) can achieve the safety and shelf-life objectives needed for long duration space-foods.

Ability of cleaning-disinfecting wipes to remove bacteria from medical device surfaces

Nosocomial infections are a serious problem in health care facilities. Bacteria can be transferred from patient to patient via contaminated reusable medical devices and equipment. An anesthesia machine and objects representative of smooth and ridged machine knobs were contaminated with Staphylococcus aureus, Bacillus atrophaeus spores, and Clostridium sporogenes spores. The ability of 5 commercially available cleaning-disinfecting wipes to remove bacteria was compared with gauze soaked with water or bleach. Gauze soaked with water was used to determine the optimal wetness for bacteria removal, which was then used to evaluate the efficacy of the wipe ingredients. All of the wipes cleaned the device surfaces significantly better than the no wipe control. Some wipes performed equally well as gauze with water, whereas others performed worse. Overall, the wipe containing sodium hypochlorite was the most effective at removing bacteria. When the wipe ingredients were re-evaluated using the determined optimal wipe wetness on gauze, their effectiveness at cleaning S aureus, but not spores, significantly improved. Physically removing bacteria from device surfaces with water was often as effective as the cleaning-disinfecting wipes. Of the wipe active ingredients evaluated, sodium hypochlorite was the most effective overall. The wetness of the wipes may also play a role in their effectiveness.

Three-dimensional tumor spheroids for in vitro analysis of bacteria as gene delivery vectors in tumor therapy.

BackgroundSeveral studies in animal models demonstrated that obligate and facultative anaerobic bacteria of the genera Bifidobacterium, Salmonella, or Clostridium specifically colonize solid tumors. Consequently, these and other bacteria are discussed as live vectors to deliver therapeutic genes to inhibit tumor growth. Therapeutic approaches for cancer treatment using anaerobic bacteria have been investigated in different mouse models. In the present study, solid three-dimensional (3D) multicellular tumor spheroids (MCTS) of the colorectal adenocarcinoma cell line HT-29 were generated and tested for their potential to study prodrug-converting enzyme therapies using bacterial vectors in vitro.ResultsHT-29 MCTS resembled solid tumors displaying all relevant features with an outer zone of proliferating cells and hypoxic and apoptotic regions in the core. Upon incubation with HT-29 MCTS, Bifidobacterium bifidum S17 and Salmonella typhimurium YB1 selectively localized, survived and replicated in hypoxic areas inside MCTS. Furthermore, spores of the obligate anaerobe Clostridium sporogenes germinated in these hypoxic areas. To further evaluate the potential of MCTS to investigate therapeutic approaches using bacteria as gene delivery vectors, recombinant bifidobacteria expressing prodrug-converting enzymes were used. Expression of a secreted cytosine deaminase in combination with 5-fluorocytosine had no effect on growth of MCTS due to an intrinsic resistance of HT-29 cells to 5-fluorouracil, i.e. the converted drug. However, a combination of the prodrug CB1954 and a strain expressing a secreted chromate reductase effectively inhibited MCTS growth.ConclusionsCollectively, the presented results indicate that MCTS are a suitable and reliable model to investigate live bacteria as gene delivery vectors for cancer therapy in vitro.Electronic supplementary materialThe online version of this article (doi:10.1186/s12934-015-0383-5) contains supplementary material, which is available to authorized users.

Effect of Nisin and Thermal Treatments on the Heat Resistance of Clostridium sporogenes Spores

The aim of this research was to evaluate the effect of thermal treatments (isothermal or nonisothermal) combined with nisin, a natural antimicrobial, on the survival and recovery of Clostridium sporogenes spores. The addition of nisin to the heating medium at concentrations up to 0.1 mg liter-1 did not reduce the heat resistance of C. sporogenes. Without a thermal treatment, nisin added at concentrations up to 0.1 mg liter-1 did not reduce the viable counts of C. sporogenes when added to the recovery medium, but inactivation of more than 4 log cycles was achieved after only 3 s at 100°C. At 100°C, the time needed to reduce viable counts by more than 3 log cycles was nine times shorter when 0.01 mg liter-1 nisin was added to the recovery medium than without it. The heat resistance values calculated under isothermal conditions were used to predict the survival in the nonisothermal experiments, and the predicted values accurately fit the experimental data. The combination of nisin with a thermal treatment can help control C. sporogenes.

Modelling the combined effects of salt, sorbic acid and nisin on the probability of growth of Clostridium sporogenes in a controlled environment (nutrient broth)

The safe preservation of new generation foods (high moisture, low salt, high pH and shelf stable under ambient conditions) is microbiologically challenging. The growth of Clostridium botulinum in low acid foods is a hazard for consumers. In this study the combined effect of salt (sodium chloride) (0–4% w/v), potassium sorbate (0–4% w/v) and nisin (0–300 ppm) at two different pHs (5.5 and 7) on the probability of growth of Clostridium sporogenes spores, as a non-toxigenic surrogate of C. botulinum, was evaluated in nutrient broth. Nutrient broth was used as it can easily and accurately be adjusted and controlled in terms of composition, and allows more rapid growth than is observed in food. The aims of this study were to develop probability of growth models in which the logit is expressed as a function of the concentrations of the selected preservatives. The developed models fit the data adequately. The c-values models were close to 1, indicating good predictive power. This indicates the performance of models would be satisfactory. The results of this study indicated that salt, potassium sorbate and nisin had significant inhibitory effects on the growth of C. sporogenes in high moisture (>95%) and high pH conditions (pH > 4.5). Combinations of the selected preservatives were more effective than any of them individually. The inhibitory effects of all three preservatives in the current study were pH dependent. Less stringent combinations of preservative concentrations were required experimentally to stop growth at pH 5.5 compared with pH 7.

The differential effects of heat‐shocking on the viability of spores from Bacillus anthracis, Bacillus subtilis, and Clostridium sporogenes after treatment with peracetic acid‐ and glutaraldehyde‐based disinfectants

This study investigated (1) the susceptibility of Bacillus anthracis (Ames strain), Bacillus subtilis (ATCC 19659), and Clostridium sporogenes (ATCC 3584) spores to commercially available peracetic acid (PAA)- and glutaraldehyde (GA)-based disinfectants, (2) the effects that heat-shocking spores after treatment with these disinfectants has on spore recovery, and (3) the timing of heat-shocking after disinfectant treatment that promotes the optimal recovery of spores deposited on carriers. Suspension tests were used to obtain inactivation kinetics for the disinfectants against three spore types. The effects of heat-shocking spores after disinfectant treatment were also determined. Generalized linear mixed models were used to estimate 6-log reduction times for each spore type, disinfectant, and heat treatment combination. Reduction times were compared statistically using the delta method. Carrier tests were performed according to AOAC Official Method 966.04 and a modified version that employed immediate heat-shocking after disinfectant treatment. Carrier test results were analyzed using Fisher's exact test. PAA-based disinfectants had significantly shorter 6-log reduction times than the GA-based disinfectant. Heat-shocking B. anthracis spores after PAA treatment resulted in significantly shorter 6-log reduction times. Conversely, heat-shocking B. subtilis spores after PAA treatment resulted in significantly longer 6-log reduction times. Significant interactions were also observed between spore type, disinfectant, and heat treatment combinations. Immediately heat-shocking spore carriers after disinfectant treatment produced greater spore recovery. Sporicidal activities of disinfectants were not consistent across spore species. The effects of heat-shocking spores after disinfectant treatment were dependent on both disinfectant and spore species. Caution must be used when extrapolating sporicidal data of disinfectants from one spore species to another. Heat-shocking provides a more accurate picture of spore survival for only some disinfectant/spore combinations. Collaborative studies should be conducted to further examine a revision of AOAC Official Method 966.04 relative to heat-shocking.

Study of the combined effect of electro-activated solutions and heat treatment on the destruction of spores of Clostridium sporogenes and Geobacillus stearothermophilus in model solution and vegetable puree

The combined effect of heat treatment and electro-activated solution (EAS) on the heat resistance of spores of Clostridium sporogenes and Geobacillus stearothermophilus was assessed under various heating and exposure time combinations. The acid and neutral EAS showed the highest inhibitory activity, indicating that these solutions may be considered as strong sporicidal disinfectants. These EAS were able to cause a reduction of ≥6 log of spores of C. sporogenes at 60 °C in only 1 min of exposition. For G. stearothermophilus spores, a reduction of 4.5 log was observed at 60 °C in 1 min, while in 5 min, ≥7 log CFU/ml reduction was observed. Inoculated puree of pea and corn were used as a food matrix for the determination of the heat resistance of these spores during the treatments in glass capillaries. The inactivation kinetics of the spores was studied in an oil bath. Combined treatment by EAS and temperature demonstrated a significant decrease in the heat resistance of C. sporogenes. The D100°C in pea puree with NaCl solution was 66.86 min while with acid and neutral EAS it was reduced down to 3.97 and 2.19 min, respectively. The spore of G. stearothermophilus displayed higher heat resistance as confirmed by other similar studies. Its D130°C in pea puree showed a decrease from 1.45 min in NaCl solution down to 1.30 and 0.93 min for acid and neutral EAS, respectively. The differences between the spores of these species are attributable to their different sensitivities with respect to pH, Redox potential and oxygen.

Apparatus for the simultaneous processing of mesophilic spores by heat-only and by high pressure and heat in a high pressure vessel to investigate synergistic spore inactivation

Abstract We sought to develop a paired sample container system that permits simultaneous processing of two samples in the same high pressure vessel, one pressurized and one protected by a pressure-resistant shell (heat-only), and subsequently compared the effect of pressure (600 MPa) on the inactivation of spores of Bacillus amyloliquefaciens and Clostridium sporogenes at high temperature. Reducing the compression rate to 200 MPa/min saw near-identical thermal profiles ( R 2 0.990) delivered to paired samples, thereby facilitating a means to directly attribute increases in inactivation in pressurised samples to the synergistic effect of pressure and heat. At 115 °C (3.5 min hold time), inactivation of B. amyloliquefaciens spores was around 7 log 10 cfu/mL greater with pressure while the high pressure thermal (HPT) inactivation of C. sporogenes spores was approximately 2 log 10 cfu/mL more than by heat alone after 1–2 min at 100 °C. Using this system we have demonstrated unequivocally that high pressure acts synergistically with heat to inactivate spores of B. amyloliquefaciens and C. sporogenes . Industrial relevance To facilitate commercialization of HPT processing for low-acid foods, a significant improvement in spore inactivation by HPT compared to traditional thermal processing, or HPT synergy, needs to be demonstrated. The development of a system that permits unequivocal assessment of the synergistic effect of pressure and heat on spore inactivation is an important tool to assist validating such processes.

Viability of Clostridium sporogenes spores after CaO hygienization of meat waste.

The occurrence of the pathogenic species C. perfringens and C. botulinum spores in animal by-products poses a potential epidemiological hazard. Strong entero- and neurotoxins produced by these bacteria adversely affect human health. To inactivate pathogens present in animal by-products, waste must be subjected to various methods of sanitization. The aim of the presented study was to estimate the effect of different doses of CaO on the viability of spores Clostridium sporogenes in meat wastes category 3. During the research, two doses of burnt lime were added to the poultry mince meat and meat mixed with swine blood contaminated with Clostridium sporogenes spore suspension. Half of the samples collected for microbiological analyses were buffered to achieve the pH level ~7, the other were examined without pH neutralization. To estimate the spore number, 10-fold dilution series in peptone water was prepared and heat-treated at 80 °C for 10 min. After cooling-down, one milliliter of each dilution was pour-plated onto DRCM medium solidified with agar. Statistical analysis were performed using the Statistica software. Application of 70% CaO caused complete inactivation of Clostridium spores in meat wastes after 48 hours. The highest temperature achieved during the experiment was 67 °C. Rapid alkalization of the biomass resulted in increasing pH to values exceeding 12. The effect of liming was not dependent on the meat wastes composition nor CaO dose. The experiment proved the efficiency of liming as a method of animal by-products sanitization. Application of the obtained results may help reduce the epidemiological risk and ensure safety to people handling meat wastes at each stage of their processing and utilization.

Effect of Microwave Radiation on Microorganisms in Fish Meals

Abstract This study was aimed at testing the effect of microwave radiation on survival of E. coli, Salmonella Enteritidis, Enterococcus spp. and Clostridium spores in two kinds of fish meals. The material used in the study consisted of samples of two kinds of fish meal – salmon and cod. In the experiment samples of both kinds of fish meals were inoculated with suspensions of tested bacteria and spores of Clostridum sporogenes. After inoculation the material was exposed to microwave radiation with a frequency of 2.45 ghz and microwave energy power of 0, 100, 180, 300, 450, 600 and 700 w, respectively, for 2.5 min for bacteria and 11 minutes for spores. then the reisolated microorganisms were counted and theoretical lethal doses of radiation were determined. among the studied vegetative forms of bacteria, the largest decreases in the numbers at the same radiation dose were observed in the rods of E. coli, whereas the smallest in enterococci. spores of Clostridium sporogenes showed a considerably higher resistance to the effect of that factor. the power of dose resulting in the complete inactivation of the studied bacteria should be about 430 kJ×g–1, and in the case of spores – 1 900 kJ×g–1.

Polyphosphate Storage during Sporulation in the Gram-Negative Bacterium Acetonema longum

Using electron cryotomography, we show that the Gram-negative sporulating bacterium Acetonema longum synthesizes high-density storage granules at the leading edges of engulfing membranes. The granules appear in the prespore and increase in size and number as engulfment proceeds. Typically, a cluster of 8 to 12 storage granules closely associates with the inner spore membrane and ultimately accounts for ∼7% of the total volume in mature spores. Energy-dispersive X-ray spectroscopy (EDX) analyses show that the granules contain high levels of phosphorus, oxygen, and magnesium and therefore are likely composed of polyphosphate (poly-P). Unlike the Gram-positive Bacilli and Clostridia, A. longum spores retain their outer spore membrane upon germination. To explore the possibility that the granules in A. longum may be involved in this unique process, we imaged purified Bacillus cereus, Bacillus thuringiensis, Bacillus subtilis, and Clostridium sporogenes spores. Even though B. cereus and B. thuringiensis contain the ppk and ppx genes, none of the spores from Gram-positive bacteria had granules. We speculate that poly-P in A. longum may provide either the energy or phosphate metabolites needed for outgrowth while retaining an outer membrane.

Bio-validation of bi-axial rotary thermal processing

In continuous processing of canned particulate liquid foods, biological validation is necessary for lethality verification because of difficulties associated with gathering temperature history at the particles centers. In this study, carrot (d = 1.9 cm) and meat (d = 1.8 cm) alginate spherical particles, inoculated with spores of Clostridium sporogenes and Geobacillus stearothermophilus, respectively, were filled into cans along with a non-Newtonian liquid (carboxymethyl cellulose). Process simulations were used and time temperature profiles at the particles center were predicted for processing at three temperatures (110–125 °C) and two rotation speeds (5–25 rpm). Process times, B, were calculated to achieve an accumulated lethality of 3 and 15 min for carrot and meat alginate fabricated particles, respectively. These ranged from 19.5 to 36.4 min for carrot and from 19.3 to 40.1 min for meat alginate particles. Using spore counts of the particles before and after the given process, the number of log reductions, n, of the bacterial spores and hence the process lethality, Fo, was determined. Results indicated that there was no significant difference (p > 0.05) in Fo values between those obtained from biological validation and from numerical simulation, demonstrating that the standardized bio-validation technique can be effectively used for establishing/verification of thermal processing schedules.

Synergistic effects of high hydrostatic pressure, mild heating, and amino acids on germination and inactivation of Clostridium sporogenes spores.

The synergistic effects of high hydrostatic pressure (HHP), mild heating, and amino acids on the germination of Clostridium sporogenes spores were examined by determining the number of surviving spores that returned to vegetative growth after pasteurization following these treatments. Pressurization at 200 MPa at a temperature higher than 40°C and treatment with some of the 19 l-amino acids at 10 mM or higher synergistically facilitated germination. When one of these factors was omitted, the level of germination was insignificant. Pressures of 100 and 400 MPa were less effective than 200 MPa. The spores were effectively inactivated by between 1.8 and 4.8 logs by pasteurization at 80°C after pressurization at 200 MPa at 45°C for 120 min with one of the amino acids with moderate hydrophobicity, such as Leu, Phe, Cys Met, Ala, Gly, or Ser. However, other amino acids showed poor inactivation effects of less than 0.9 logs. Spores in solutions containing 80 mM of either Leu, Phe, Cys, Met, Ala, Gly, or Ser were successfully inactivated by pasteurization by more than 5.4 logs after pressurization at 200 MPa at 70°C for 15 to 120 min. Ala and Met reduced the spore viability by 2.8 and 1.8 logs, respectively, by pasteurization at a concentration of 1 mM under 200 MPa at 70°C. These results indicate that germination of the spores is facilitated by a combination of high hydrostatic pressure, mild heating, and amino acids.