Mechanisms of Bacillus subtilis spore inactivation by single- and multi-pulse high hydrostatic pressure (MP-HHP)

Herein we investigate the effect of multi-pulse high hydrostatic pressure (MP-HHP) on the inactivation of Bacillus subtilis spores. B. subtilis spores were subjected to MP-HHP under pressures at 200–500 MPa at temperatures of 40 and 60 °C with 3 pulses (holding time of 3 min) with a total processing time of 10 min and compared it with a single pressurization (S-HHP).Mechanism of spore inactivation by S- or MP-HHP was explored by assessing germination by heat shock treatment, spore susceptibility to lysozyme and hydrogen peroxide (H₂O₂), release of dipicolinic acid (DPA), and the permeability of inner membrane and cortex. Our results presented the highest spore inactivation (5.8 log reduction), when MP-HHP was applied under the highest temperature and pressure. The increased inactivation appears to be largely due to mechanical disruption of spore coat and inner and outer membranes, as evidenced by DPA release, increased susceptibility to lysozyme and H₂O₂ (indicative of breakage of disulfide bonds in the spore coat), and membrane permeability as assessed by spore staining and fluorescence microscopy. No differences were seen in germination between MP-HHP and S-HHP. There was no evidence of any loss of cortex lytic enzymes or degradation of small acid-soluble proteins (SASPs) during both MP-HHP and S-HHP treatments.     

Mechanism of Bacillus subtilis spore inactivation induced by moderate electric fields

Bacterial endospores are the key safety targets for inactivation within low-acid foods. Herein, we investigated the inactivation of Bacillus subtilis CGMCC 1.1087 spores (10⁷ CFU/mL) in sterile distilled water using moderate electric fields (MEF, 300 V/cm) under various temperatures (<30, 55, 65 and 75 °C). MEF treatment at below 30 °C resulted in 0.6-log reduction of spores, while treatments for 60 min without electric fields showed no inactivation. Inactivation induced by MEF in the same treatment time increased to 1.8-, 2.0- and 2.5-log as temperature increased to 55, 65 and 75 °C. Spores treated with MEF at <30, 55, 65 and 75 °C or mild heat (55, 65 and 75 °C) scarcely lost heat resistance, suggesting that spores did not germinate during MEF or mild heat treatment. The viability of MEF-treated spores did not increase by addition of lysozyme (3 μg/mL) in recovery plates, preincubation for 1 h in a 1:1 mixture of 60 mM Ca²⁺ and DPA, or lysozyme treatment in hypertonic medium. Confocal laser scanning microscopy photomicrographs showed that exposure to MEF induced a marked increase in the permeability of inner membrane and cortex. These findings suggested that damage of the cortex and inner membrane, rather than spore nutrient germinant receptors or cortex lytic enzymes, are possible reasons contributing to inactivation of B. subtilis spores by MEF. This study indicates that MEF at mild temperatures (55 to 75 °C) have the potential for spore inactivation.Industrial relevanceLiterature in the past few years has shown that moderate electric fields (MEF), typically associated with ohmic heating, have nonthermal effects on bacterial spores, leading to accelerated inactivation. The current work extends the range of temperatures to those well below thermally lethal conditions, and shows that some spore inactivation occurs under MEF, even when temperatures are sublethal. Little or no germination is observed, and spore inner membranes are increasingly compromised over time. The elucidation of such nonthermal effects would be significant to the food industry as it seeks increasingly nonthermal methods for inactivation of spores.     

Characterization of germination of spores of Clostridium estertheticum,the primary causative agent of blown pack spoilage of vacuum packaged beef

The aim of this study was to investigate the effect of various factors on the germination of Clostridium estertheticum endospores (spores) in relation to beef. The effect of heat on germination was determined by recovering C. estertheticum on Columbia agar from spore suspensions not heated or heated at 63, 70 or 80 °C for various times. The effects of pH, temperature and oxygen were determined, by enumeration of remaining ungerminated spores during incubation in Meat Juice medium (MJM). Amino acids and lactate were tested for their ability to trigger germination of C. estertheticum spores by monitoring dipicolinic acid (DPA) release. Heat treatment of spores at 80 °C for ≤ 20 min significantly (p < 0.05) increased the numbers of spores recovered on blood agar. Neither incubation temperature nor oxygen affected germination in MJM. The optimal pH for germination was 7.0 to 7.5. Incubation with leucine or aspartic acid caused a 1.3% release of DPA, the highest among all amino acids tested. Incubation with lactate resulted in a 4.1% release of DPA, which was significantly (p < 0.05) higher than those from incubation with amino acids. The DPA release from incubation with lactate, lactate with amino acids, or MJM was similar (p > 0.05).     

Comparison of High Hydrostatic Pressure,High-PressureCarbon Dioxide and High-Temperature Short-Time Processing on Quality of Mulberry Juice

Changes of microbial, physicochemical and sensory properties of mulberry juice processed by high hydrostatic pressure (HHP) (500 MPa/5 min), high-pressure carbon dioxide (HPCD) (15 MPa/55 °C/10 min), and high-temperature short time (HTST) (110 °C/8.6 s) during 28 days of storage at 4 °C and 25 °C were investigated. Total aerobic bacteria (TAB) and yeast and mold (Y&M) were not detected in HHP-treated and HTST-treated mulberry juices for 28 days at 4 °C and 25 °C, but were detected more than 2 log₁₀ CFU/ml in HPCD-treated mulberry juice for 21 days at 4 °C and 14 days at 25 °C, respectively. Total anthocyanins were retained after HHP and reduced by 4 % after HTST while increased by 11 % after HPCD. Total phenols were retained by HHP, while increased by 4 % after HTST and 16 % after HPCD. The antioxidant capacity was retained by HTST and HHP and increased by HPCD. Both total phenols and antioxidant capacity were decreased during the initial 14 days but then increased up to 28 days regardless of storage temperature. The value of polymeric color and browning index decreased and a* increased in HHP-treated and HPCD-treated mulberry juices, while HTST-treated mulberry juice had a reverse result. The viscosity of mulberry juice increased in HHP-treated and HPCD-treated juices, while decreased in HTST-treated juice. During storage, total anthocyanins, total phenols, and antioxidant capacity and color in all mulberry juices decreased more largely at 25 °C than that at 4 °C. Better quality was observed in HHP- and HPCD-treated mulberry juices, and a longer shelf life was observed in HHP-treated samples compared to HPCD-treated ones.     

Comparative study on cloudy apple juice qualities from apple slices treated by high pressure carbon dioxide and mild heat

Qualities of cloudy apple juices from apple slices treated by high pressure carbon dioxide (HPCD) and mild heat (MH) were evaluated. Temperatures were from 25 to 65 °C, time 20 min, and pressure 20 MPa. Polyphenol oxidase (PPO) was completely inactivated by HPCD and its minimal residual activity (RA) by MH at 65 °C was 38.6%. RA of pectin methylesterase (PME) with HPCD was significantly lower than MH and its minimum was 18%. L value of cloudy apple juice from HPCD-treated apple slices was significantly greater than that from MH-treated apple slices, however, b value, browning degree (BD) and turbidity were lower. And no differences in a value, total soluble solids, pH and conductivity were observed. After 7-day storage at 4 °C, HPCD caused no BD alteration but a significant turbidity loss. MH increased BD at 55 and 65 °C, and led to turbidity loss from 35 to 65 °C. The turbidity was not well related to RA of PME.Industrial relevanceCloudy apple juice is one of the popular fruit juices, and it requires strict processing treatment conditions to protect its quality, especially to prevent enzymatic browning and cloud loss. HPCD is one promising novel non-thermal technique and is likely to replace or partially substitute thermal processes. This study analyzed the effect of HPCD as a pretreatment means on qualities of cloudy apple juice, including inactivating enzymes which are crucial to quality control. Available data provided in this study will benefit the fruit juice industry.     

Effects on microbial inactivation and quality attributes in frozen lychee juice treated by supercritical carbon dioxide

In this paper, we described the use of high-pressure carbon dioxide (HPCD) for the inactivation of natural microbes in lychee juice and evaluated its effects on lychee juice quality, compared to a conventional high-temperature, short-time (HTST) method. The HPCD treatments were carried out using a HPCD unit (8 MPa, 36 °C, 2 min), and the HTST was performed at 90 °C for 60 s. The results showed that five log reduction for yeasts and molds and total aerobic microorganisms occurred at 8 MPa for 2 min. And effects of the treatments on pH and concentrations of microbes, organic acids, titratable acidity (TA), total soluble solid (TSS), sugars, polyphenols, color, and free amino acids were also investigated. HPCD could efficiently maintain the concentration of polyphenols and original color at 8 MPa, 36 °C for 2 min. Insignificant differences in colors were observed between unprocessed and HPCD juices, while significant differences were observed between unprocessed and HTST juices. Furthermore, HTST decreased the total free amino acids, whereas HPCD caused a significant increase (increased by 45.92% at 8 MPa) (p < 0.05). The increase in total amino acids induced by HPCD treatment is beneficial for nutritional value of commercial ready-to-drink lychee juice. In general, HPCD treatment had less influence on the measured quality parameters of lychee juice than HTST treatment. Therefore, HPCD treatment could be a useful alternative to traditional heat treatment.     

Effect of high pressure carbon dioxide on alkaline phosphatase activity and quality characteristics of raw bovine milk

Novel food processing techniques would always be pursuits of researchers and food industry to avoid unfavorable thermal effects, especially in dairy and milk processing. In this study, effects of high pressure carbon dioxide (HPCD) on the activity of alkaline phosphatase (ALP) and main quality indices of raw bovine milk at 20 MPa using a batch system were investigated. A complete inactivation of ALP activity as exposure to HPCD treatment at 50 °C and 20 MPa for 50 min was observed. The protein and lactose content of HPCD-treated bovine milk hold steady, while pH value and total solids content decreased, turbidity and average particle size increased significantly (p < 0.05). Although a significant decrease of viscosity (p < 0.05) was observed, the Newtonian flow behavior of raw bovine milk did not alter. More obvious change of quality characteristics of raw bovine milk were observed as subjected to HPCD treatment at higher temperature or treated for longer period. Therefore, a compromise between controlling endogenous enzymes (and/or spoilage and pathogenic microorganisms) and retention of original/fresh like quality of foods should be introduced due to the nature of HPCD processing. It's suggested to keep raw bovine milk with low ALP activity and great quality treated with a batch HPCD apparatus at 20 MPa and 50 °C for 20 min.     

High pressure thermal processing for the inactivation of Clostridium perfringens spores in beef slurry

The spores of Clostridium perfringens can survive and grow in cooked/pasteurized meat, especially during the cooling of large portions. In this study, 600 MPa high pressure thermal processing (HPTP) at 75 °C for the inactivation of C. perfringens spores was compared with 75 °C thermal processing alone. The HPTP enhanced the inactivation of C. perfringens spores in beef slurry, resulting in 2.2 log reductions for HPTP vs. no reductions for thermal processing after 20 min. Then, the HPTP resistance of two C. perfringens spore strains in beef slurry at 600 MPa was compared and modeled, and the effect of temperature investigated. The NZRM 898 and NZRM 2621 exhibited similar resistance, and Weibull modeled well the log spore survivor curves. The spore inactivation increased when HPTP temperature was raised from 38 to 75 °C. The results confirm the advantage of high pressure technology to increase the thermal inactivation of C. perfringens spores in beef slurry.Industrial relevanceC. perfringens spores may cause food/meat poisoning as a result of improperly handled and prepared foods in industrial kitchens. Thermal processes at 100 °C or higher are generally carried out to ensure the elimination of these pathogenic spores. High pressure processing (HPP) is a food pasteurization technique which would help to maintain the sensorial and nutritional properties of food. Preservation of foods with HPP in conjunction with mild heat (HPTP) would enhance the spore inactivation compared to thermal processing alone at the same temperature, due to a known germination–inactivation mechanism. This technology, together with the application of Good Manufacturing Practices, including rapid cooling, is a good alternative to the traditional methods for producing safe processed meat and poultry products with enhanced sensory and nutritional quality.     

Effect of High-pressure CO2 Processing on Bacterial Spores

High-pressure CO₂ (HPCD) is a nonthermal technology that can effectively inactivate the vegetative forms of pathogenic and spoilage bacteria, yeasts, and molds at pressures less than 30 MPa and temperatures in the range of 20°C to 40°C. However, HPCD alone at moderate temperatures (20–40°C) is often insufficient to obtain a substantial reduction in bacterial spore counts because their structures are more complex than those of vegetative cells. In this review, we first thoroughly summarized and discussed the inactivation effect of HPCD treatment on bacterial spores. We then presented and discussed the kinetics by which bacterial spores are inactivated by HPCD treatment. We also summarized hypotheses drawn by different researchers to explain the mechanisms of spore inactivation by HPCD treatment. We then summarized the current research status and future challenges of spore inactivation by HPCD treatment.     

Analysis of Escherichia coli cell damage induced by HPCD using microscopies and fluorescent staining

Cellular damage of Escherichia coli (E. coli) induced by high pressure carbon dioxide (HPCD) at 37-57 °C and 10-30 MPa for 5-75 min was investigated using scanning electronic microscopy (SEM), transmission electronic microscopy (TEM), confocal laser scanning microscopy (CLSM), and fluorospectrophotometer (FSM). HPCD-induced alterations in the morphology and the intracellular organization of E. coli cells was more susceptible to HPCD. A vast majority of HPCD-treated E. coli cells with seemingly intact morphology sustained severe damage in their intracellular organization. CLSM suggested that initial disruption of the outer membrane and later permeabilization of the cytoplasmic membrane of HPCD-treated E. coli cells was a consecutive and progressive process. These results were confirmed by FSM with the probes PI and SYTO 9. The membrane fluidity of HPCD-treated E. coli cells decreased as suggested by increased fluorescence polarization using FSM with the probe 1,6-diphenyl-1,3,5-hexatriene (DPH). The temperatures of 37, 42 and 47 °C alone showed no impact on the outer membrane and membrane fluidity of E. coli cells whereas 57 °C alone had greater impact on them. Combined with HPCD, the temperatures of 37, 42 and 47 °C disrupted the outer membrane of E. coli cells without damage to the cytoplasmic membrane and of 57 °C damaged the cytoplasmic membrane, but all these temperatures decreased the membrane fluidity of E. coli cells. Higher temperature increased HPCD-induced outer membrane disruption and the cytoplasmic membrane damage and decreased the membrane fluidity.     

Effect of high pressure carbon dioxide on the quality of carrot juice

The effect of high pressure carbon dioxide (HPCD) on the quality of carrot juice was investigated. The L-value of HPCD-treated juices increased significantly (P < 0.05) as compared to untreated juices, and the a-value exhibited an increase tendency with increasing the treatment time. However, the b-value of HPCD-treated juices did not change. The browning degree (BD) and pH of HPCD-treated juices decreased, the cloud and titratable acidity (TA) increased significantly, the UV–visible spectra of juices were lower, but the total soluble solid (TSS) and the carotenoids of juices were stable. The particle size of juices treated by HPCD for 15, 30 and 45 min increased significantly (P < 0.05), for 60 min showed a noticeable decrease and was almost close to untreated juice. HPCD treatment could not alter the Newtonian flow behavior of the carrot juice, but caused a significant increase in juice viscosity (P < 0.05).Industrial relevanceCarrot juice is one of the most popular vegetable juices, but it requires severe heat treatment for protection from spoilage due to a higher pH, its heat-sensitive quality is inevitably destructed. In this study, HPCD can avoid the drawbacks of the heat treatment as a novel non-thermal pasteurization, available data are provided for the application and evaluation of HPCD in the juice industry.     

Inactivation of microorganisms naturally present in raw bovine milk by high‐pressure carbon dioxide

This study assessed the inactivation of microorganisms naturally present in raw bovine milk by high‐pressure carbon dioxide (HPCD) at 10–30 MPa and 20–50 °C for 20–70 min. The log reduction of microorganisms increased as raw bovine milk was exposed to higher pressures and temperatures and longer treatment times. The maximum reduction of aerobic bacteria (AB) was 4.96‐log at 25 MPa and 50 °C for 70 min. At lower temperatures and treatment times, a complete inactivation of yeasts and moulds (Y&M) and coliform bacteria (CB) was obtained at 25 MPa. Changes in microorganisms naturally present in raw bovine milk during storage were also assessed. There were 1.83‐log survival of AB, 0.65‐log survival of Y&M and a complete inactivation of CB in raw bovine milk when subjected to HPCD at 25 MPa and 40 °C for 50 min. Moreover, the AB, Y&M and the CB in raw bovine milk exhibited insignificant alterations during storage at 4 °C for 15 days, indicating a potential capability of HPCD to extend the shelf life of milk.     

Combined high pressure and heat treatment effectively disintegrates spore membranes and inactivates Alicyclobacillus acidoterrestris spores in acidic fruit juice beverage

The commercial potential of high pressure and thermal processing (HPTP) was investigated against Alicyclobacillus acidoterrestris spores in commercial acidic apple juice beverage and in acidified and neutral potassium buffers. With starting spore counts prior to treatments being 6.5 and 7.2 log₁₀ respectively for strains AJA 66 (D_90°C 15.4 min) and ATCC 49025 (D_90°C 8.5 min), HPTP at 600 MPa at 80 °C for 3 min provided an optimal treatment with spore viability reduced below the detection limit for both strains. HPTP at 80 °C for 1 min and HPTP at 70 °C for 3 min achieved 4.1–4.5 log₁₀ CFU/mL reduction. HPTP at 70 °C for 1 min reduced the number of viable spores by 2.0–2.5-log₁₀ CFU/mL. Flow cytometry revealed the presence of membrane-compromised spores among culturable spores following HPTP and heat alone treatments at different temperatures. Electron microscopy clearly showed the efficiency of HPTP with crushed or hollow spores predominating after treatments. No correlation between HPTP susceptibility and genetic diversity was observed for two genotypes of A. acidoterrestris spores. The treatment combination provides a promising option for industrial utility since it requires lower heat and processing time.     

高压CO2处理对枯草杆菌芽孢杀灭及协同作用的影响

选用枯草芽孢杆菌为研究对象,以磷酸盐缓冲液为基质,研究高压(5～25MPa) CO2协同热处理对其芽孢杀灭作用的影响。结果表明：20MPa高压CO2处理前在45℃预热处理30min时枯草芽孢杆菌芽孢的失活率达到了3.84个数量级,单纯20MPa高压CO2处理30min时枯草芽孢杆菌芽孢的失活率达到2.44个数量级。采用连续式和间歇式两种不同的施压方式考查其对枯草芽孢杆菌芽孢的灭活及萌发影响,结果表明：10MPa、50℃连续30min高压CO2处理对枯草芽孢杆菌芽孢的杀灭使其数量下降了0.67个数量级,间歇施压 (低压10MPa施压10min,高压30MPa施压10min)循环1次时枯草芽孢杆菌芽孢的失活率达到2.55个数量级。选择添加3种组合的营养发芽诱导因子考察在不同的压力条件下(10MPa 30℃、10MPa 50℃、30MPa 30℃、30MPa 50℃)与高压CO2处理相结合对芽孢发芽的影响。结果表明：添加的营养发芽诱导因子在低压(10MPa)时的诱导作用不如高压(30MPa)时明显。     

The impact of high pressure and temperature on bacterial spores: inactivation mechanisms of Bacillus subtilis above 500 MPa

Abstract: High‐pressure thermal sterilization (HPTS) is an emerging technology to produce shelf stable low acid foods. Pressures below 300 MPa can induce spore germination by triggering germination receptors. Pressures above 500 MPa could directly induce a Ca⁺²‐dipicolinic acid (DPA) release, which triggers the cortex‐lytic enzymes (CLEs). It has been argued that the activated CLEs could be inactivated under HPTS conditions. To test this claim, a wild‐type strain and 2 strains of Bacillus subtilis spores lacking germinant receptors and one of 2 CLEs were treated simultaneously from 550 to 700 MPa and 37 to 80 C (slow compression) and at 60 to 80 C up to 1 GPa (fast compression). Besides, an additional heat treatment to determine the amount of germinated cells, we added TbCl₃ to detect the amount of DPA released from the spore core via fluorescent measurement. After pressure treatment for 120 min at 550 MPa and 37 °C, no inactivation was observed for the wild‐type strain. The amount of released DPA correlated to the amount of germinated spores, but always higher compared to the belonging cell count after pressure treatment. The release of DPA and the increase of heat‐sensitive spores confirm that the inactivation mechanism during HPTS passes through the physiological states: (1) dormancy, (2) activation, and (3) inactivation. As the intensity of treatment increased, inactivation of all spore strains also strongly increased (up to ?5.7 log₁₀), and we found only a slight increase in the inactivation of one of the CLE (sleB). Furthermore, above a certain threshold pressure, temperature became the dominant influence on germination rate. Practical Application: The continuous increase of high‐pressure (HP) research over the last several decades has already generated an impressive number of commercially available HP pasteurized products. Furthermore, research helped to provoke the certification of a pressure‐assisted thermal sterilization process by the U.S. FDA in February 2009. However, this promising sterilization technology has not yet been applied in industrial settings. An improved understanding of spore inactivation mechanisms and the ability to calculate desired inactivation levels will help to make this technology available for pilot studies and commercialization at an industrial scale. Moreover, if the synergy between pressure and elevated temperature on the inactivation rate could be identified, clarification of the underlying inactivation mechanism during HP thermal sterilization could help to further optimize the process of this emerging technology.     

Clostridium sporogenes-ATCC 7955 Spore Destruction Kinetics in Milk Under High Pressure and Elevated Temperature Treatment Conditions

Clostridium sporogenes (ATCC 7955) spores inoculated in milk (2% fat) were subjected to high-pressure (HP) treatments (700–900 MPa) and at elevated temperatures (80–100 °C) for selected times up to 32 min. Samples were sealed in 1-mL plastic vials and placed in a specially constructed insulated chamber to prevent temperature drop during the treatment. Both pressure pulse (with no hold time) and pressure hold techniques were employed for treatment. Pressure pulse resulted in a small, but consistent, destruction (up to 0.5 log kill) of spores. During the pressure hold treatment, the destruction followed a first-order model (R ² > 0.90). The kinetic data were compensated for the small variations in temperature during the treatment. As expected, higher pressures and higher temperatures resulted in a faster rate of spore destruction. Temperature-corrected D values ranged from 13.6 to 2.4 min at 700 MPa and 7.0 to 1.3 min at 900 MPa, respectively, with process temperatures set at 90 and 100 °C. In comparison, thermal treatments gave D values ranging from 156 min at 90 °C to 12.1 min at 100 °C. The temperature sensitivity Z _P values (16.5 to 20.3 °C) under high pressure (700–900 MPa) were higher than under conventional thermal processing (9.0 °C), indicating the spore’s thermal resistance to increase at HP processing conditions. The pressure sensitivity Z _T values varied between 450 and 680 MPa under the elevated temperature (80–100 °C) processing conditions. Overall, C. sporogenes 7955 spores were relatively more sensitive to temperature than pressure.     

Effects of the repair treatment on improving the heat resistance of Lactiplantibacillus plantarum LIP-1

To improve the heat resistance of lactobacilli during spray-drying, we first investigated the effect of heat shock and repair treatment on the heat resistance of Lactiplantibacillus plantarum LIP-1, and the specific cell repair mechanism. Compared with control group, the reduction of the strain after the heat treatment (75 °C for 40 s) decreased from 1.05 to 0.36 log CFU/mL (initial cell counts 9.30 log CFU/mL) by heat shock (44 °C for 10 min). The residuals of the strain after heat-treated increased by 0.90 log CFU/mL by heat shock firstly, and then the repair treatment (30 °C for 10 min).During recovery period, the relative content of unsaturated long-chain fatty acids (C18:1n9 & C18:2n6) induced by heat shock proteins increased by 7.0%, and the amount of DnaK protein anchored on the cell membrane increased by 0.17 pg/mL. Cell membrane damage caused by heat shock was reduced and strain heat resistance was improved.Industrial relevanceOur research found that the repair treatment after heat shock reduced cell membrane damage caused by heat shock, which is a very promising technology for improving the heat resistance of L. plantarum LIP-1. This technology is also expected to be widely used in the preparation of LAB powders by spray-drying technology.     

不同菌体肽聚糖对产气荚膜梭菌芽孢萌发的诱导作用

为了研究不同菌体肽聚糖对芽孢萌发的影响,本研究分别提取了产气荚膜梭菌(Clostridium perfringens,C. perfringens)、枯草芽孢杆菌(Bacillus subtilis,B. subtilis)及金黄色葡萄球菌(Staphylococcus aureus,S. aureus)细胞壁肽聚糖,以C. perfringens芽孢为研究对象,按照1:10、1:100、1:1000比例分别加入提取的三种菌体肽聚糖,并以芽孢浑浊度(OD_(600))、芽孢折光性、2,6-吡啶二羧酸(DPA)的释放量、热抗性损失等指标比较不同菌体肽聚糖对C.perfringens芽孢萌发的效果。结果显示,按照一定百分比分别加入三种菌体肽聚糖,1:10比例添加C. perfringens营养体肽聚糖诱导C. perfringens芽孢萌发效果最佳,在60 min后,其芽孢浑浊度(OD_(600))下降了31.71%,DPA释放率上升可达到38.29%;1:10比例添加B. subtilis营养体肽聚糖诱导C. perfringens芽孢萌发效果次之,其芽孢浑浊度(OD_(600))下降了24.92%,DPA释放率上升可达到30.84%;而不同比例S. aureus营养体肽聚糖对C. perfringens芽孢萌发均无显著影响。本研究可为进一步阐明肽聚糖诱导芽孢萌发的机制提供重要信息。     

Germination and Heat Resistance of Bacillus cereus Spores from Strains Associated with Diarrheal and Emetic Food-Borne Illnesses

Bacillus cereus has been implicated as the cause of both diarrheal and emetic forms of food-borne illness. Spores of eight strains of B. cereus, representing diairheal, emetic and atoxigenic origins, were examined for heat resistance and germination responses. No correlation was observed between heat resistance at 85° or 90°C and origin of the strain. Germination of spores in Trypticase soy broth at 30°C, measured by loss of heat resistance, was more extensive for diarrheal strains than for emetic strains. These data should be useful in evaluating potential hazards from B. cereus in foods.     

Membrane permeabilization and cellular death of Escherichia coli, Listeria monocytogenes and Saccharomyces cerevisiae as induced by high pressure carbon dioxide treatment

In this study, the relationship between (irreversible) membrane permeabilization and loss of viability in Escherichia coli, Listeria monocytogenes and Saccharomyces cerevisiae cells subjected to high pressure carbon dioxide (HPCD) treatment at different process conditions including temperature (35–45 °C), pressure (10.5–21.0 MPa) and treatment time (0–60 min) was examined. Loss of membrane integrity was measured as increased uptake of the fluorescent dye propidium iodide (PI) with spectrofluorometry, while cell inactivation was determined by viable cell count. Uptake of PI by all three strains indicated that membrane damage is involved in the mechanism of HPCD inactivation of vegetative cells. The extent of membrane permeabilization and cellular death increased with the severity of the HPCD treatment. The resistance of the three tested organisms to HPCD treatment changed as a function of treatment time, leading to significant tailing in the survival curves, and was dependent on pressure and temperature. The results in this study also indicated a HPCD-induced damage on nucleic acids during cell inactivation. Transmission electron microscopy showed that HPCD treatment had a profound effect on the intracellular organization of the micro-organisms and influenced the permeability of the bacterial cells by introducing pores in the cell wall.