- R.P. Betts (1)
- L.R. Beuchat (1)
- M.A.J.S. Boekel van (1)
- F. Bourdichon (1)
- D. Chandra Aryani (1)
- G.B. Dijksterhuis (1)
- C. Dooremalen van (1)
- J. Ellers (1)
- S. Fanning (1)
- W.C. Hazeleger (1)
- S.A.F.T. Hijum van (1)
- L.M. Hornstra (1)
- H.M.L.J. Joosten (1)
- H.J. Kamphuis (1)
- E. Komitopoulou (1)
- B.H. Kuile ter (1)
- L.J.R. Líma (1)
- M.J.R. Nout (1)
- W. Suring (1)
- P.G.M. Teunissen (1)
- W.M. Vos de (1)
- Y.P. Vries de (1)
- A. Wagendorp (1)
- M.H.J. Wells-Bennik (2)
- Y. Xiao (2)
- M.H. Zwietering (3)
Genome-wide transcriptional profiling of Clostridium perfringens SM101 during sporulation extends the core of putative sporulation genes and genes determining spore properties and germination characteristics
Xiao, Y. ; Hijum, S.A.F.T. van; Abee, T. ; Wells-Bennik, M.H.J. - \ 2015
PLoS One 10 (2015)5. - ISSN 1932-6203 - 19 p.
bacillus-subtilis - dipicolinic acid - bacterial-spores - heat-resistance - ser/thr kinase - killing factor - identification - difficile - proteins - sequence
The formation of bacterial spores is a highly regulated process and the ultimate properties of the spores are determined during sporulation and subsequent maturation. A wide variety of genes that are expressed during sporulation determine spore properties such as resistance to heat and other adverse environmental conditions, dormancy and germination responses. In this study we characterized the sporulation phases of C. perfringens enterotoxic strain SM101 based on morphological characteristics, biomass accumulation (OD600), the total viable counts of cells plus spores, the viable count of heat resistant spores alone, the pH of the supernatant, enterotoxin production and dipicolinic acid accumulation. Subsequently, whole-genome expression profiling during key phases of the sporulation process was performed using DNA microarrays, and genes were clustered based on their time-course expression profiles during sporulation. The majority of previously characterized C. perfringens germination genes showed upregulated expression profiles in time during sporulation and belonged to two main clusters of genes. These clusters with up-regulated genes contained a large number of C. perfringens genes which are homologs of Bacillus genes with roles in sporulation and germination; this study therefore suggests that those homologs are functional in C. perfringens. A comprehensive homology search revealed that approximately half of the upregulated genes in the two clusters are conserved within a broad range of sporeforming Firmicutes. Another 30% of upregulated genes in the two clusters were found only in Clostridium species, while the remaining 20% appeared to be specific for C. perfringens. These newly identified genes may add to the repertoire of genes with roles in sporulation and determining spore properties including germination behavior. Their exact roles remain to be elucidated in future studies.
Differential outgrowth potential of Clostridium perfringens food-borne isolates with various cpe-genotypes in vacuum-packed ground beef during storage at 12°C
Xiao, Y. ; Wagendorp, A. ; Abee, T. ; Wells-Bennik, M.H.J. - \ 2015
International Journal of Food Microbiology 194 (2015). - ISSN 0168-1605 - p. 40 - 45.
enterotoxin gene - multiplex pcr - heat-resistance - cooked beef - a strains - plasmid - growth - spores - sporulation - expression
In the current study, the outgrowth of spores of 15 different food isolates of Clostridium perfringens was evaluated in vacuum-packed ground beef during storage at 12 °C and 25 °C. This included enterotoxic strains carrying the gene encoding the CPE enterotoxin on the chromosome (C-cpe), on a plasmid (P-cpe) and cpe-negative strains. The 15 strains were selected from a larger group of strains that were first evaluated for their ability to sporulate in modified Duncan–Strong sporulating medium. Sporulation ability varied greatly between strains but was not associated with a particular cpe genotype. In line with previous studies, the tested C-cpe strains produced spores with significantly higher heat resistance than the cpe-negative and P-cpe strains (both IS1151 and IS1470-like) with the exception of strain VWA009. Following inoculation of vacuum-packed cooked ground beef with spores, the heat-resistant C-cpe strains showed lower outgrowth potential in this model food stored at 12 °C than the P-cpe and cpe-negative strains, while no significant differences were observed at 25 °C. These results suggest that the latter strains may have a competitive advantage over C-cpe strains at reduced temperatures during storage of foods that support the growth of C. perfringens. While spores of P-cpe strains are readily inactivated by heat processing, post-processing contamination by food handlers who may carry P-cpe strains that have a better growth potential at lower temperatures must be avoided. The varying responses of C. perfringens spores to heat and the differences in outgrowth capacity at different temperatures are factors to be considered in strain selection for challenge tests, and for predictive modelling of C. perfringens.
Quantifying variabiltiy on thermal resistance of Listeria monocytogenes
Chandra Aryani, D. ; Besten, H.M.W. den; Hazeleger, W.C. ; Zwietering, M.H. - \ 2015
International Journal of Food Microbiology 193 (2015). - ISSN 0168-1605 - p. 130 - 138.
heat-resistance - escherichia-coli - salmonella-typhimurium - inactivation parameters - physiological-state - bacillus-cereus - salt stress - shock - growth - acid
Knowledge of the impact of strain variability and growth history on thermal resistance is needed to provide a realistic prediction and an adequate design of thermal treatments. In the present study, apart from quantifying strain variability on thermal resistance of Listeria monocytogenes, also biological variability and experimental variability were determined to prioritize their importance. Experimental variability was defined as the repeatability of parallel experimental replicates and biological variability was defined as the reproducibility of biologically independent reproductions. Furthermore, the effect of growth history was quantified. The thermal inactivation curves of 20 L. monocytogenes strains were fitted using the modified Weibull model, resulting in total 360 D-value estimates. The D-value ranged from 9 to 30 min at 55 °C; from 0.6 to 4 min at 60 °C; and from 0.08 to 0.6 min at 65 °C. The estimated z-values of all strains ranged from 4.4 to 5.7 °C. The strain variability was ten times higher than the experimental variability and four times higher than the biological variability. Furthermore, the effect of growth history on thermal resistance variability was not significantly different from that of strain variability and was mainly determined by the growth phase.
Low-water activity foods: increased concern as vehicles of foodborne pathogens
Beuchat, L.R. ; Komitopoulou, E. ; Beckers, H. ; Betts, R.P. ; Bourdichon, F. ; Fanning, S. ; Joosten, H.M.L.J. ; Kuile, B.H. ter - \ 2013
Journal of Food Protection 76 (2013)1. - ISSN 0362-028X - p. 150 - 172.
escherichia-coli o157-h7 - powdered infant formula - salmonella-agona infection - eat savoury snack - enterobacter-sakazakii - cronobacter spp. - peanut butter - international outbreak - heat-resistance - milk powder
Foods and food ingredients with low water activity (aw) have been implicated with increased frequency in recent years as vehicles for pathogens that have caused outbreaks of illnesses. Some of these foodborne pathogens can survive for several months, even years, in low-aw foods and in dry food processing and preparation environments. Foodborne pathogens in low-aw foods often exhibit an increased tolerance to heat and other treatments that are lethal to cells in high-aw environments. It is virtually impossible to eliminate these pathogens in many dry foods or dry food ingredients without impairing organoleptic quality. Control measures should therefore focus on preventing contamination, which is often a much greater challenge than designing efficient control measures for high-aw foods. The most efficient approaches to prevent contamination are based on hygienic design, zoning, and implementation of efficient cleaning and sanitation procedures in the food processing environment. Methodologies to improve the sensitivity and speed of assays to resuscitate desiccated cells of foodborne pathogens and to detect them when present in dry foods in very low numbers should be developed. The goal should be to advance our knowledge of the behavior of foodborne pathogens in low-aw foods and food ingredients, with the ultimate aim of developing and implementing interventions that will reduce foodborne illness associated with this food category. Presented here are some observations on survival and persistence of foodborne pathogens in low-aw foods, selected outbreaks of illnesses associated with consumption of these foods, and approaches to minimize safety risks
Fatty acid compostion and extreme temperature tolerance following exposure to fluctuating temperatures in a soil arthropod
Dooremalen, C. van; Suring, W. ; Ellers, J. - \ 2011
Journal of Insect Physiology 57 (2011)9. - ISSN 0022-1910 - p. 1267 - 1273.
membrane-lipid-composition - thermal-reaction norms - chill coma recovery - drosophila-melanogaster - homeoviscous adaptation - cycling temperatures - cold resistance - heat-resistance - plasticity - acclimation
Ectotherms commonly adjust their lipid composition to ambient temperature to counteract detrimental thermal effects on lipid fluidity. However, the extent of lipid remodeling and the associated fitness consequences under continuous temperature fluctuations are not well-described. The objective of this study was to investigate the effect of repeated temperature fluctuations on fatty acid composition and thermal tolerance. We exposed the springtail Orchesella cincta to two constant temperatures of 5 and 20 °C, and a continuously fluctuating treatment between 5 and 20 °C every 2 days. Fatty acid composition differed significantly between constant low and high temperatures. As expected, animals were most cold tolerant in the low temperature treatment, while heat tolerance was highest under high temperature. Under fluctuating temperatures, fatty acid composition changed with temperature initially, but later in the experiment fatty acid composition stabilized and closely resembled that found under constant warm temperatures. Consistent with this, heat tolerance in the fluctuating temperature treatment was comparable to the constant warm treatment. Cold tolerance in the fluctuating temperature treatment was intermediate compared to animals acclimated to constant cold or warmth, despite the fact that fatty acid composition was adjusted to warm conditions. This unexpected finding suggests that in animals acclimated to fluctuating temperatures an additional underlying mechanism is involved in the cold shock response. Other aspects of homeoviscous adaptation may protect animals during extreme cold. This paper forms a next step to fully understand the functioning of ectotherms in more thermally variable environments
Microbiota of cocoa powder with particular reference to aerobic thermoresistant spore-formers
Líma, L.J.R. ; Kamphuis, H.J. ; Nout, M.J.R. ; Zwietering, M.H. - \ 2011
Food Microbiology 28 (2011)3. - ISSN 0740-0020 - p. 573 - 582.
bacillus-subtilis - heat-resistance - sp-nov - aflp - heterogeneity - bacteria - sequence - strains - milk - rna
The microbiological criteria of commercial cocoa powder are defined in guidelines instituted by the cocoa industry. Twenty-five commercial samples were collected with the aim of assessing the compliance with the microbiological quality guidelines and investigating the occurrence and properties of aerobic Thermoresistant Spores (ThrS). Seventeen samples complied with the guidelines, but one was positive for Salmonella, five for Enterobacteriaceae and two had mould levels just exceeding the maximum admissible level. The treatment of the cocoa powder suspensions from 100 °C to 170 °C for 10 min, revealed the presence of ThrS in 36% of the samples. In total 61 ThrS strains were isolated, of which the majority belonged to the Bacillus subtilis complex (65.6%). Strains resporulation and spore crops inactivation at 110 °C for 5 min showed a wide diversity of heat-resistance capacities. Amplified fragment length polymorphism analysis revealed not only a large intraspecies diversity, but also different clusters of heat-resistant spore-forming strains. The heat-resistance of spores of six B. subtilis complex strains was further examined by determination of their D and z-values. We concluded that B. subtilis complex spores, in particular those from strain M112, were the most heat-resistant and these may survive subsequent preservation treatments, being potentially problematic in food products, such as chocolate milk
A systematic approach to determine global thermal inactivation parameters for various food pathogens
Asselt, E.D. van; Zwietering, M.H. - \ 2006
International Journal of Food Microbiology 107 (2006)1. - ISSN 0168-1605 - p. 73 - 82.
escherichia-coli o157-h7 - bacillus-cereus spores - perfringens vegetative cells - in-ground beef - salmonella-typhimurium dt104 - nonproteolytic type-b - heat-resistance - listeria-monocytogenes - clostridium-botulinum - water activity
Thermal inactivation of pathogens has been studied extensively, which has resulted in a wide range of D- and z-values. Estimating the inactivation rate for a specific condition based on these reported values is difficult, since one has to select representative conditions, and data obtained exactly at the required representative conditions are generally not available. Therefore, a first step could be to globally assess a heat treatment taking into account largest effects only. Once the most important parameters are known, a more precise study of inactivation can be performed. Therefore, in this study a large quantity of D-values (n = 4066) was collected from literature for various pathogens and linear regression was applied to obtain average D-values (together with the 95% upper prediction level) and z-values. When comparing these overall data, it can be seen that most factors reported to have an effect on the D-value are smaller than the variability of all published D-values. Even effects of shoulders disappear in the overall analysis. Only a limited number of factors that did have a significant effect (p <0.05) on the D-value were identified: for Salmonella spp., the presence of chocolate ingredients gave protection to the cells, for Listeria monocytogenes the presence of 10% salt (or aw <0.92) resulted in a higher heat resistance, for Bacillus cereus there were significant differences for various strains and in oily products and for Clostridium botulinum there were significant differences in heat resistance between different types of C. botulinum. This does not mean that other effects do not occur, but it shows the main effects that have to be included for a first impression on the performance of a heating process. The obtained 95% upper prediction levels of the D-values can be used as a (conservative) estimate of inactivation and can be used to give order of magnitude values in overall process evaluations
Influence of glutamate on growth, sporulation, and spore properties of Bacillus cereus ATCC 14579 in defined medium
Vries, Y.P. de; Atmadja, R.D. ; Hornstra, L.M. ; Vos, W.M. de; Abee, T. - \ 2005
Applied and Environmental Microbiology 71 (2005)6. - ISSN 0099-2240 - p. 3248 - 3254.
high hydrostatic-pressure - bacterial-spores - different temperatures - dipicolinic acid - heat-resistance - water-content - subtilis - germination - inactivation - initiation
A chemically defined medium in combination with an airlift fermentor system was used to study the growth and sporulation of Bacillus cereus ATCC 14579. The medium contained six amino acids and lactate as the main carbon sources. The amino acids were depleted during exponential growth, while lactate was metabolized mainly during stationary phase. Two concentrations of glutamate were used: high (20 mM; YLHG) and low (2.5 mM; YLLG). Under both conditions, sporulation was complete and synchronous. Sporulation started and was completed while significant amounts of carbon and nitrogen sources were still present in the medium, indicating that starvation was not the trigger for sporulation. Analysis of amino acids and NH4+ in the culture supernatant showed that most of the nitrogen assimilated by the bacteria was taken up during sporulation. The consumption of glutamate depended on the initial concentration; in YLLG, all of the glutamate was used early during exponential growth, while in YLHG, almost all of the glutamate was used during sporulation. In YLLG, but not in YLHG, NH4+ was taken up by the cells during sporulation. The total amount of nitrogen used by the bacteria in YLLG was less than that used by the bacteria in YLHG, although a significant amount of NH4+ was present in the medium throughout sporulation. Despite these differences, growth and temporal expression of key sigma factors involved in sporulation were parallel, indicating that the genetic time frames of sporulation were similar under both conditions. Nevertheless, in YLHG, dipicolinic acid production started later and the spores were released from the mother cells much later than in YLLG. Notably, spores had a higher heat resistance when obtained after growth in YLHG than when obtained after growth in YLLG, and the spores germinated more rapidly and completely in response to inosine, L-alanine, and a combination of these two germinants
Dormant ascospores of Talaromyces macrosporus are activated to germinate after treatment with ultra high pressure
Dijksterhuis, G.B. ; Teunissen, P.G.M. - \ 2004
Journal of Applied Microbiology 96 (2004)1. - ISSN 1364-5072 - p. 162 - 169.
byssochlamys-nivea ascospores - bacillus-subtilis spores - heat-resistance - phycomyces-blakesleeanus - hydrostatic-pressure - neosartorya-fischeri - flavus - inactivation - fulva
Aims: Ascospores of Talaromyces macrosporus are constitutively dormant and germinate after a strong external shock, classically a heat treatment. This fungus is used as a model system to study heat resistance leading to food spoilage after pasteurization. This study evaluates the effect of high pressure on the germination behaviour of these spores. Methods and Results: Ascospore containing bags were subjected to ultra high pressure and spores were plated out on agar surfaces. Untreated suspensions showed invariably very low germination. Increased germination of ascospores occurred after short treatments at very high pressure (between 400 and 800 MPa). Activation is partial compared with heat activation and did not exceed 6·9% (65 times that of untreated suspensions) of the spore population. Maximum activation was attained shortly (10 s¿3 min) after the pressure was applied and accompanied by cell wall deformations as judged by scanning electron microscopy. The spores observed in this study were harvested from cultures that were 39¿58 days old. The maturity of spores at similar developmental stages was measured by assessing the heat resistance of ascospores. Between 20 and 40 days heat resistance increased 2·4-fold, but only an additional increase of 1·3-fold was observed at later stages (40¿67 days). Conclusions: Our investigations show that high pressure constitutes a second type of shock that can activate heat-resistant ascospores to germinate. Activation is maximal after very short treatments and accompanied with changes in the cell wall structure. High-pressure activation is not the result of immaturity of the ascospores. Significance and Impact of the Study:These observations are relevant for the application of high pressure as a novel pasteurization method.
On the use of the Weibull model to describe thermal inactivation of microbial vegetative cells
Boekel, M.A.J.S. van - \ 2002
International Journal of Food Microbiology 74 (2002)1-2. - ISSN 0168-1605 - p. 139 - 159.
log-logistic model - listeria-monocytogenes - heat-resistance - survival curves - yersinia-enterocolitica - salmonella-enteritidis - predictive models - bovine-milk - temperature - kinetics
This paper evaluates the applicability of the Weibull model to describe thermal inactivation of microbial vegetative cells as an alternative for the classical Bigelow model of first-order kinetics; spores are excluded in this article because of the complications arising due to the activation of dormant spores. The Weibull model takes biological variation, with respect to thermal inactivation, into account and is basically a statistical model of distribution of inactivation times. The model used has two parameters, the scale parameter alpha (time) and the dimensionless shape parameter beta. The model conveniently accounts for the frequently observed nonlinearity of semilogarithmic survivor curves, and the classical first-order approach is a special case of the Weibull model. The shape parameter accounts for upward concavity of a survival curve (beta <1), a linear survival curve (beta = 1), and downward concavity (beta > 1). Although the Weibull model is of an empirical nature, a link can be made with physiological effects. beta <1 indicates that the remaining cells have the ability to adapt to the applied stress, whereas beta > 1 indicates that the remaining cells become increasingly damaged. Fifty-five case studies taken from the literature were analyzed to study the temperature dependence of the two parameters. The logarithm of the scale parameter a depended linearly on temperature, analogous to the classical D value. However, the temperature dependence of the shape parameter beta was not so clear. In only seven cases, the shape parameter seemed to depend on temperature, in a linear way. In all other cases, no statistically significant (linear) relation with temperature could be found. In 39 cases, the shape parameter beta was larger than 1, and in 14 cases, smaller than 1. Only in two cases was the shape parameter b = 1 over the temperature range studied, indicating that the classical first-order kinetics approach is the exception rather than the rule. The conclusion is that the Weibull model can be used to model nonlinear survival curves, and may be helpful to pinpoint relevant physiological effects caused by heating. Most importantly, process calculations show that large discrepancies can be found between the classical first-order approach and the Weibull model. This case study suggests that the Weibull model performs much better than the classical inactivation model and can be of much value in modelling thermal inactivation more realistically, and therefore, in improving food safety and quality. (C) 2002 Elsevier Science B.V. All rights reserved.