Physiological characterization of spoilage strains of Zygosaccharomyces bailii and Zygosaccharomyces rouxii isolated from high sugar environments

Two isolates of spoilage yeasts Zygosaccharomyces bailii and Zygosaccharomyces rouxii were obtained from a high sugar environment, in a factory producing candied fruit and nougat. Other strains, Z. bailii from other environments and other isolates from high sugar/salt environments were obtained for comparison (Zygosaccharomyces lentus, Candida magnoliae, Candida halophila and Pichia guilliermondii). A full physiological assessment of these isolates was carried out, of preservative and biocide resistance, osmotolerance, ethanol-tolerance, low pH resistance, degree of fermentation, and growth temperature and survival to pasteurisation. Results showed that the strains isolated from high sugar environments did not show extreme physiology. These were robust strains but within the normal parameters expected for the species. One exception to this was that the Z. bailii strains were abnormally able to grow at 37 degrees C. In all strains other than C. magnolia and C. halophila, cells were able to adapt to high levels of sugar. Cultures grown in high glucose concentrations were subsequently able to tolerate higher concentrations of glucose than previously. Similarly, high sugar was found to confer a degree of protection against pasteurisation, enabling survival in what would have otherwise been a lethal treatment. Isolates of Z. bailii showed a high level of resistance to preservatives such as sorbic acid, benzoic acid, acetic acid, cinnamic acid, and ethanol, and also to heat. Unexpectedly Z. bailii isolates were not exceptionally resistant to biocides such as peracetic acid, or hypochlorite. These results indicate that spoilage by yeasts such as Z. bailii may be better prevented by use of biocidal cleaning agents in the factory, rather than treating the food with preservatives.     

The importance of expressing antimicrobial agents on water basis in growth/no growth interface models: a case study for Zygosaccharomyces bailii

In a previous study on Zygosaccharomyces bailii, three growth/no growth models have been developed, predicting growth probability of the yeast at different conditions typical for acidified foods (Dang, T.D.T., Mertens, L., Vermeulen, A., Geeraerd, A.H., Van Impe, J.F., Debevere, J., Devlieghere, F., 2010. Modeling the growth/no growth boundary of Z. bailii in acidic conditions: A contribution to the alternative method to preserve foods without using chemical preservatives. International Journal of Food Microbiology 137, 1-12). In these broth-based models, the variables were pH, water activity and acetic acid, with acetic acid concentration expressed in volume % on the total culture medium (i.e., broth). To continue the previous study, validation experiments were performed for 15 selected combinations of intrinsic factors to assess the performance of the model at 22 °C (60 days) in a real food product (ketchup). Although the majority of experimental results were consistent, some remarkable deviations between prediction and validation were observed, e.g., Z. bailii growth occurred in conditions where almost no growth had been predicted. A thorough investigation revealed that the difference between two ways of expressing acetic acid concentration (i.e., on broth basis and on water basis) is rather significant, particularly for media containing high amounts of dry matter. Consequently, the use of broth-based concentrations in the models was not appropriate. Three models with acetic acid concentration expressed on water basis were established and it was observed that predictions by these models well matched the validation results; therefore a “systematic error” in broth-based models was recognized. In practice, quantities of antimicrobial agents are often calculated based on the water content of food products. Hence, to assure reliable predictions and facilitate the application of models (developed from lab media with high dry matter contents), it is important to express antimicrobial agents' concentrations on a common basis—the water content. Reviews over other published growth/no growth models in literature are carried out and expressions of the stress factors' concentrations (on broth basis) found in these models confirm this finding.     

The antimicrobial properties of chitosan in mayonnaise and mayonnaise-based shrimp salads

The potential for using chitosan glutamate as a natural food preservative in mayonnaise and mayonnaise-based shrimp salad was investigated. Mayonnaise containing 3 g/liter of chitosan combined with acetic acid (0.16%) or lemon juice (1.2 and 2.6%) was inoculated with log 5 to 6 CFU/g of Salmonella Enteritidis, Zygosaccharomyces bailii, or Lactobacillus fructivorans and stored at 5 and 25 degrees C for 8 days. In mayonnaise containing chitosan and 0.16% acetic acid, 5 log CFU/g of L. fructivorans were inactivated, and numbers remained below the sensitivity limit of the plate counting technique for the duration of the experiment. Z. bailii counts were also reduced by approximately 1 to 2 log CFU/g within the first day of incubation at 25 degrees C, but this was followed by growth on subsequent days, giving an overall growth delay of 2 days. No differences in counts of Z. bailii in mayonnaise stored at 5 degrees C or of Salmonella Enteritidis stored at either temperature were observed. In mayonnaise containing lemon juice at both 1.2 and 2.6%, no substantial differences were observed between the controls and the samples containing chitosan. In shrimp salads stored at 5 degrees C, the presence of a coating of chitosan (9 mg/g of shrimp) inhibited growth of the spoilage flora from approximately log 8 CFU/g in the controls to log 4 CFU/g throughout 4 weeks. However, at 25 degrees C, chitosan was ineffective as a preservative. The results demonstrated that chitosan may be useful as a preservative when combined with acetic acid and chill storage in specific food applications.     

A differential medium for the enumeration of the spoilage yeast Zygosaccharomyces bailii in wine

A collection of yeasts, isolated mostly from spoiled wines, was used in order to develop a differential medium for Zygosaccharomyces bailii. The 118 selected strains of 21 species differed in their origin and resistance to preservatives and belonged to the genera Pichia, Torulaspora, Dekkera, Debaryomyces, Saccharomycodes, Issatchenkia, Kluyveromyces, Kloeckera, Lodderomyces, Schizosaccharomyces, Rhodotorula, Saccharomyces, and Zygosaccharomyces. The design of the culture medium was based on the different ability of the various yeast species to grow in a mineral medium with glucose and formic acid (mixed-substrate medium) as the only carbon and energy sources and supplemented with an acid-base indicator. By manipulating the concentration of the acid and the sugar it was possible to select conditions where only Z. bailii strains gave rise to alkalinization, associated with a color change of the medium (positive response). The final composition of the mixed medium was adjusted as a compromise between the percentage of recovery and selectivity for Z. bailii. This was accomplished by the use of pure or mixed cultures of the yeast strains and applying the membrane filtration methodology. The microbiological analysis of two samples of contaminated Vinho Verde showed that the new medium can be considered as a differential medium to distinguish Z. bailii from other contaminating yeasts, having potential application in the microbiological control of wines and probably other beverages and foods.     

Targeted gene deletion in Zygosaccharomyces bailii

Yeasts of the genus Zygosaccharomyces are notable agents of large-scale food spoilage. Despite the economic importance of these organisms, little is known about the stress adaptations whereby they adapt to many of the more severe conditions of food preservation. In this study it was shown that genes of Z. bailii, a yeast notable for its high resistances to food preservatives and ethanol, can be isolated by complementation of the corresponding mutant strains of Saccharomyces cerevisiae. It was also discovered that the acquisition by S. cerevisiae of a single small Z. bailii gene (ZbYME2) was sufficient for the former yeast to acquire the ability to degrade two major food preservatives, benzoic acid and sorbic acid. Using DNA cassettes containing dominant selectable markers and methods originally developed for performing gene deletions in S. cerevisiae, the two copies of ZbYME2 in the Z. bailii genome were sequentially deleted. The resulting Zbyme2/Zbyme2 homozygous deletant strain had lost any ability to utilize benzoate as sole carbon source and was more sensitive to weak acid preservatives during growth on glucose. Thus, ZbYME2, probably the nuclear gene for a mitochondrial mono-oxygenase function, is essential for Z. bailii to degrade food preservatives. This ability to catabolize weak acid preservatives is a significant factor contributing to the preservative resistance of Z. bailii under aerobic conditions. This study is the first to demonstrate that it is possible to delete in Z. bailii genes that are suspected as being important for growth of this organism in preserved foods and beverages. With the construction of further mutant of Z. bailii strains, a clearer picture should emerge of how this yeast adapts to the conditions of food preservation. This information will, in turn, allow the design of new preservation strategies. GenBank Accession Nos: ZbURA3 (AF279259), ZbTIM9 (AF279260), ZbYME2 (AF279261), ZbTRP1 (AF279262), ZbHHT1(AF296170).     

Influence of pH, water activity and acetic acid concentration on Listeria monocytogenes at 7 degrees C: data collection for the development of a growth/no growth model

Growth/no growth models can be used to determine the chance that microorganisms will grow in specific environmental conditions. As a consequence, these models are of interest in the assessment of the safety of foods which can be contaminated with food pathogens. In this paper, growth/no growth data for Listeria monocytogenes (in a monoculture and in a mixed strain culture) are presented. The data were gathered at 7 degrees C in Nutrient Broth with different combinations of environmental factors pH (5.0-6.0, six levels), water activity (0.960-0.990, six levels) and acetic acid concentration (0-0.8% (w/w), five levels). This combination of environmental factors for the development of a growth/no growth model was based on the characteristics of sauces and mayonnaise based salads. The strains used were chosen from screening experiments in which the pH, water activity and acetic acid resistance of 26 L. monocytogenes strains (LFMFP culture collection) was determined at 30 degrees C in Brain Heart Infusion broth. The screening showed that most L. monocytogenes strains were not able to grow at a(w)<0.930, pH<4.3 or a total acetic acid concentration >0.4% (w/w). Among these strains, the ones chosen were the most resistant to one of these factors in the hope that, if the resulting model predicted no growth at certain conditions for those more resistant strains, then these predictions would also be valid for the less resistant strains. A mixed strain culture was also examined to combine the strains that were most resistant to one of the factors. A full factorial design with the selected strains was tested. The experiments were performed in microtiter plates and the growth was followed by optical density measurements at 380 nm. The plates were inoculated with 6 log CFU/ml and twenty replicates were made for each treatment combination. These data were used (1) to determine the growth/no growth boundary and (2) to estimate the influence of the environmental conditions on the time to detection. From the monoculture and mixed strain data, the growth boundary of L. monocytogenes is shown not to be a straight cut-off but a rather narrow transition zone. The experiments also showed that in the studied region, a(w) did not have a pronounced influence on the position of the growth/no growth boundary while a low concentration of acetic acid (0.2% (w/w)) and a pH decrease from 6.0 to 5.8 was sufficient to significantly reduce the possibility of growth. The determination of the time to detection showed a significant increase at the combinations of environmental conditions near the 'no growth zone'. For example, at 0.2% (w/w) acetic acid, there was an increase from +/-10 days to 30 days by lowering pH from 5.8 to 5.6 at a(w) values of 0.985 and 0.979, while at pH 5.4 less than 50% growth occurred for all a(w) values.     

Predicting growth rates and growth boundary of Listeria monocytogenes — An international validation study with focus on processed and ready-to-eat meat and seafood

The performance of six predictive models for Listeria monocytogenes was evaluated using 1014 growth responses of the pathogen in meat, seafood, poultry and dairy products. The performance of the growth models was closely related to their complexity i.e. the number of environmental parameters they take into account. The most complex model included the effect of nine environmental parameters and it performed better than the other less complex models both for prediction of maximum specific growth rates (µ_max values) and for the growth boundary of L. monocytogenes. For this model bias and accuracy factors for growth rate predictions were 1.0 and 1.5, respectively, and 89% of the growth/no-growth responses were correctly predicted. The performance of three other models, including the effect of five to seven environmental parameters, was considered acceptable with bias factors of 1.2 to 1.3. These models all included the effect of acetic acid/diacetate and lactic acid, one of the models also included the effect of CO₂ and nitrite but none of these models included the effect of smoke components. Less complex models that did not include the effect of acetic acid/diacetate and lactic acid were unable to accurately predict growth responses of L. monocytogenes in the wide range of food evaluated in the present study. When complexity of L. monocytogenes growth models matches the complexity of foods of interest, i.e. the number of hurdles to microbial growth, then predicted growth responses of the pathogen can be accurate. The successfully validated models are useful for assessment and management of L. monocytogenes in processed and ready-to-eat (RTE) foods.     

The effect of citric acid and pH on growth and metabolism of anaerobic Saccharomyces cerevisiae and Zygosaccharomyces bailii cultures

The effects of citric acid at pH values of 3.0, 4.0, and 4.5 on growth and metabolism of anaerobic Saccharomyces cerevisiae and Zygosaccharomyces bailii cultures were investigated. S. cerevisiae and Z. bailii exhibited similar tolerances to citric acid, as determined by growth measurements, at all three pH values investigated. The citric-acid-induced growth inhibition of both yeast species increased with increasing pH values, indicating that the antimicrobial mechanism of citric acid differs from that of classical weak-acid preservatives. In S. cerevisiae, citric acid shifted the primary energy metabolism towards lower ethanol production and higher glycerol production, thus resulting in lower ATP production. These metabolic changes in S. cerevisiae were pH-dependent; i.e. the higher the pH, the lower the ATP production, and they may explain why growth of S. cerevisiae is more inhibited by citric acid at higher pH values. In Z. bailii, citric acid also caused an increased glycerol production, although to a lesser extent than in S. cerevisiae, but it caused virtually no changes in ethanol and ATP production.     

Sorbic acid resistance: the inoculum effect

Zygosaccharomyces is a genus associated with the more extreme spoilage yeasts. Zygosaccharomyces spoilage yeasts are osmotolerant, fructophiles (preferring fructose), highly-fermentative and extremely preservative-resistant. Zygosaccharomyces bailii can grow in the presence of commonly-used food preservatives, benzoic, acetic or sorbic acids, at concentrations far higher than are legally permitted or organolepically acceptable in foods. An inoculum effect has been described for many micro-organisms and antimicrobial agents. The minimum inhibitory concentration (MIC) increases with the size of the inoculum; large inocula at high cell density therefore require considerably higher concentrations of inhibitors to prevent growth than do dilute cell suspensions. A substantial inoculum effect was found using sorbic acid against the spoilage yeast Zygosaccharomyces bailii NCYC 1766. The inoculum effect was not caused by yeasts metabolizing or adsorbing sorbic acid, thereby lowering the effective concentration; was not due to absence of cell-cell signals in dilute cell suspensions; and was not an artefact, generated by insufficient time for small inocula to grow. The inoculum effect appeared to be caused by diversity in the populations of yeast cells, with higher probability of sorbic acid-resistant cells being present in large inocula. It was found that individual cells of Zygosaccharomyces bailii populations, grown as single cells in microtitre plate wells, were very diverse, varying enormously in resistance to sorbic acid. 26S ribosomal DNA sequencing did not detect differences between the small fraction of resistant 'super cells' and the average population. Re-inoculation of the 'super cells' after overnight growth on YEPD showed a normal distribution of resistance to sorbic acid, similar to that of the original population. The resistance phenotype was therefore not heritable and not caused by a genetically distinct subpopulation. It was concluded that resistance of the spoilage yeast Zygosaccharomyces bailii to sorbic acid was due to the presence of small numbers of phenotypically resistant cells in the population.     

Validation of predictive models describing the growth of Listeria monocytogenes

In this study, predictions for growth rate of Listeria on food products were evaluated by both general applicable models and specific growth models. Literature values, obtained from a large number of publications, for growth rates in/on a variety of foods were compared by graphical and mathematical analysis with predictions given by various models. Apart for the great advantage of being generally applicable, the general models performed best. However, only small differences between the various models were observed. Model predictions were accurate within a factor of about two to four, depending on the type of product. The predictions should therefore not be considered as absolute; it is important to understand the limitations of the performance of models. All results and all assumptions should be criticised, but in many cases the accuracy will be sufficient to use these types of models as a tool in management decisions.     

Acidification Requirements for Home Canned Combinations of Tomatoes and Low-Acid Ingredients

Alternative methods of estimating acidification requirements for home canned high-acid foods comprising combinations of tomatoes with low-acid ingredients were compared with forty representative products. Acidification estimates were based on recipe specifications, titration data, predictions made with a regression equation derived from a quadratic model, and predictions derived from a worst case analysis of the model. The last method resulted in satisfactory pH reductions (0.2-0.3 unit) even with products having pH values close to 4.6. Acidification recommendations based on the worst case analysis are given for eight categories of high-acid combination products.     

Isolation of an acetyl-CoA synthetase gene (ZbACS2) from Zygosaccharomyces bailii

A gene homologous to Saccharomyces cerevisiae ACS genes, coding for acetyl-CoA synthetase, has been cloned from the yeast Zygosaccharomyces bailii ISA 1307, by using reverse genetic approaches. A probe obtained by PCR amplification from Z. bailii DNA, using primers derived from two conserved regions of yeast ACS proteins, RIGAIHSVVF (ScAcs1p; 210-219) and RVDDVVNVSG (ScAcs1p; 574-583), was used for screening a Z. bailii genomic library. Nine clones with partially overlapping inserts were isolated. The sequenced DNA fragment contains a complete ORF of 2027 bp (ZbACS2) and the deduced polypeptide shares significant homologies with the products of ACS2 genes from S. cerevisiae and Kluyveromyces lactis (81% and 82% identity and 84% and 89% similarity, respectively). Phylogenetic analysis shows that the sequence of Zbacs2 is more closely related to the sequences from Acs2 than to those from Acs1 proteins. Moreover, this analysis revealed that the gene duplication producing Acs1 and Acs2 proteins has occurred in the common ancestor of S. cerevisiae, K. lactis, Candida albicans, C. glabrata and Debaryomyces hansenii lineages. Additionally, the cloned gene allowed growth of S. cerevisiae Scacs2 null mutant, in medium containing glucose as the only carbon and energy source, indicating that it encodes a functional acetyl-CoA synthetase. Also, S. cerevisiae cells expressing ZbACS2 have a shorter lag time, in medium containing glucose (2%, w/v) plus acetic acid (0.1-0.35%, v/v). No differences in cell response to acetic acid stress were detected both by specific growth and death rates. The mode of regulation of ZbACS2 appears to be different from ScACS2 and KlACS2, being subject to repression by a glucose pulse in acetic acid-grown cells.     

Modeling the Growth/No-Growth Interface of Zygosaccharomyces bailii in Mango Puree

Probabilistic microbial modeling using logistic regression was used to predict critical temperatures to inhibit for at least 35 d Zygosaccharomyces bailii growth in a pH 3.5 mango puree formulated with 1000 ppm of potassium sorbate (KS) or sodium benzoate (NaB) at selected a_w (0.99, 0.98, or 0.97). The probability of growth was calculated, thereby ascertaining the set conditions and critical temperatures required inhibiting yeast growth for different storage times. Using the logistic model, with a growth probability of 0.05, critical temperatures were higher for KS than for NaB. Use of KS to inhibit Z. bailii growth enabled for mango puree 30 d of storage at 6.4 °C.     

Antimicrobial action of synthetic peptides towards wine spoilage yeasts

The antimicrobial action of selected short synthetic peptides against wine spoilage yeasts such as Cryptococcus albidus, Dekkera bruxellensis, Pichia membranifaciens, Saccharomyces cerevisiae, Zygosaccharomyces bailii and Zygosaccharomyces bisporus has been examined. Peptides analyzed include nine sequence-related antifungal hexapeptides (PAFs) previously developed by a combinatorial approach, and two representative lactoferricin B (LfcinB)-derived peptides. Different peptides had distinct activity profiles. In vitro assays identified the peptides PAF26, PAF36, and LfcinB(17-31), as having growth inhibitory properties towards several of the yeasts at low micromolar concentrations. Z. bailii and Z. bisporus were the most sensitive yeasts. In addition to their fungistatic activity, the three peptides showed fungicidal properties towards Z. bailii, Z. bisporus, and S. cerevisiae in laboratory growth medium. Remarkably, only LfcinB(17-31) against Z. bisporus had inhibitory and fungicidal properties in wine at the concentrations assayed, showing that the antimicrobial action of each peptide is dependent on both the food matrix and the target micro-organism. Lack of fungicidal activity of peptides against Z. bailii in wine is related to the presence of salt ions other than divalent cations. On the contrary, fungicidal activity of LfcinB(17-31) towards Z. bisporus was not significantly affected by wine salts. Our data identify a bioactive peptide from natural origin with potential use against the food spoilage yeast Z. bisporus, and indicate that the application of antimicrobial peptides in wine preservation deserves further investigation.     

A Predictive Model for the Growth/No Growth Boundary of Zygosaccharomyces bailii at 7?°C and Conditions Mimicking Acidified Sauces

The spoilage potential of Zygosaccharomyces bailii has been widely recognized within the food industry. However, few data are available on the growth characteristics of this yeast at low temperature and in the absence of chemical preservatives. In this study, the growth/no growth boundary of Z. bailii was defined at refrigeration temperature (7?°C) and at conditions relevant to high-sugar, low-pH foods (such as ketchup and salad dressing), i.e. pH?3.0?C5.0 (five levels), a _w 0.93?C0.97 (five levels) and acetic acid concentration 0?C2.5% (v/v; six levels). Yeast growth was followed during 90?days by optical density measurements, and logistic regression models were used to describe the data. Acetic acid had a significant effect on the relation between a _w and NaCl concentration and this interaction had important consequences for the model development. When data were modelled as a function of a _w or NaCl concentration, a stimulatory effect by acetic acid was observed. In contrast, as a function of (toxic) Na⁺ ions, no evidence was found of such a phenomenon. These results indicate that in cases where the relationship between a _w and solute concentration is not straightforward such as in the presence of acid preservatives, one must be critical towards the interpretation of data and correspondingly, the development of predictive models. On a practical note, the developed models, especially the one incorporating Na⁺ ions, may be used (1) to assess the stability of shelf-stable acidified foods stored under chilled conditions after opening or (2) to formulate new additive-free products intended for storage at 7?°C.     

A peculiar stimulatory effect of acetic and lactic acid on growth and fermentative metabolism of Zygosaccharomyces bailii

Stimulatory or protective effects of organic acids at low concentrations, e.g. acetic and lactic acid, on microorganisms have previously been reported. Especially in case of Zygosaccharomyces bailii, a peculiar growth stimulation by these two acids has recently been noticed. In order to elucidate this interesting phenomenon, growth and fermentative metabolism of Z. bailii was investigated in media with low pH (pH 4.0), high sugar (15% (w/v)) and different acetic and lactic acid concentrations. At both experimental temperatures (7 and 30 °C), a growth stimulation in the presence of 2.5% (v/v) lactic acid was observed. Furthermore at 7 °C, the yeast exhibited another unusual behaviour as it grew much faster in media containing 1.25% (v/v) acetic acid than in the control (without any acid). Production of fermentative metabolites was also increased together with the enhanced growth at both temperatures. These possible stimulatory effects of acetic and lactic acid should be taken into consideration when the acids are used at low doses for food preservative purpose. Presence of the acids may stimulate Z. bailii growth and fermentative metabolism, particularly at refrigeration temperature, consequently resulting in an earlier spoilage.     

A probability of growth model for Escherichia coli O157:H7 as a function of temperature, pH, acetic acid, and salt.

Data accumulated on the growth of Escherichia coli O157:H7 in tryptic soy broth (TSB) were used to develop a logistic regression model describing the growth-no growth interface as a function of temperature, pH, salt, sucrose, and acetic acid. A fractional factorial design with five factors was used at the following levels: temperature (10 to 30 degrees C), acetic acid (0 to 4%), salt (0.5 to 16.5%), sucrose (0 to 8%), and pH (3.5 to 6.0). A total of 1,820 treatment combinations were used to create the model, which correctly predicted 1,802 (99%) of the points, with 10 false positives and 8 false negatives. Concordance was 99.9%, discordance was 0.1%, and the maximum rescaled R2 value was 0.927. Acetic acid was the factor having the most influence on the growth-no growth interface; addition of as little as 0.5% resulted in an increase in the observed minimum pH for growth from 4.0 to 5.5. Increasing the salt concentration also had a significant effect on the interface; at all acetic acid concentrations, increasing salt increased the minimum temperature at which growth was observed. Using two literature data sets (26 conditions), the logistic model failed to predict growth in only one case. The results of this study suggest that the logistic regression model can be used to make conservative predictions of the growth-no growth interface of E. coli O157:H7.     

A neural network approach to predict survival/death and growth/no-growth interfaces for Escherichia coli O157:H7

An artificial neural network (ANN) model was developed to predict survival/death and growth/no-growth interfaces for Escherichia coli O157:H7 in a mayonnaise-type system. Temperature, pH, acetic acid, sucrose and salt were the inputs to a three-layer back-propagation neural network. The ANN model was trained using the data-set of McKellar et al. [2002. A probability model describing the interface between survival and death of E. coli O157:H7 in a mayonnaise model system. Food Microbiol. 19, 235-247] that consisted of 1,820 treatment combinations from controlled experiments with a cocktail of five strains of E. coli O157:H7. After training, the model correctly predicted the growth/no-growth in 1,810 combinations (99.5%) with 8 false positives and 2 false negatives, and survival/death in 1,804 combinations (99.1%) with 13 false positives and 3 false negatives. Classification accuracy was validated using additional literature data-sets for growth of E. coli O157:H7 under various environmental conditions. The ANN model accurately predicted the survival/death in 27 of 30 cases (90%) in experimental mayonnaise inoculated with E. coli O157:H7, with 3 fail-positive predictions and all observed growth (100%). Simulations were used to estimate the influence of incubation temperature on survival and growth for specific combinations of acetic acid, salt, pH and sucrose. The ANN model is recommended as an alternative tool for classification of survival and growth conditions in predictive microbiology.     

INHIBITION OF MICROORGANISMS IN SALAD DRESSING BY SUCROSE AND METHYLGLUCOSE FATTY ACID MONOESTERS

The antimicrobial activity of sucrose and methylglucose esters of medium to long chain fatty acids was studied with two microorganisms involved in the spoilage of salad dressings, Zygosaccharomyces bailii and Lactobacillus fructivorans. The microorganisms were inhibited to various degrees by 0.1, 0.5, and 1.0% synthesized sucrose or methylglucose monoesters using a modified broth dilution method. Sucrose monoesters were most inhibitory when the esterified fatty acid was myristic (C₁₄) or palmitic acid (C₁₆). Methylglucose monoesters with lauric (C₁₂) or myristic acid (C₁₄) exhibited greater inhibition than those with longer chain fatty acids. The least inhibition was generally observed with sucrose and methylglucose oleate (C_18:1). Sucrose monoesters were usually more inhibitory than methylglucose monoesters of the same fatty acid, especially for palmitic and stearic (C₁₈) acids. In salad dressing, 1% sucrose monoesters of lauric, myristic, or palmitic acid significantly (P < 0.05) inhibited the growth of Z. bailii and L. fructivorans, and were comparable with or more effective than 0.1% sodium benzoate. Z. bailii growth was nearly completely inhibited by sucrose laurate, myristate and palmitate by 9 days of salad dressing storage. Sucrose monoesters did not delay the lag phase of L.     

Inactivation of food spoilage microorganisms by hydrodynamic cavitation to achieve pasteurization and sterilization of fluid foods

ABSTRACT:  Hydrodynamic cavitation is the formation of gas bubbles in a fluid due to pressure fluctuations induced by mechanical means. Various high-acid (pH ≥ 4.6) fluid foods were processed in a hydrodynamic cavitation reactor to determine if commercial sterility can be achieved at reduced processing temperatures. Sporicidal properties of the process were also tested on a low-acid (pH < 4.6) fluid food. Fluid foods were pumped under pressure into a hydrodynamic cavitation reactor and subjected to 2 rotor speeds and flow rates to achieve 2 designated exit temperatures. Thermal inactivation kinetics were used to determine heat-induced lethality for all organisms. Calcium-fortified apple juice processed at 3000 and 3600 rpm rotor speeds on the reactor went through a transient temperature change from 20 to 65.6 or 76.7 °C and the total process lethality exceeded 5-log reduction of Lactobacillus plantarum and Lactobacillus sakei cells, and Zygosaccharomyces bailii cells and ascospores. Tomato juice inoculated with Bacillus coagulans spores and processed at 3000 and 3600 rpm rotor speeds endured a transient temperature from 37.8 to 93.3 or 104.4 °C with viable CFU reductions of 0.88 and 3.10 log cycles, respectively. Skim milk inoculated with Clostridium sporogenes putrefactive anaerobe 3679 spores and processed at 3000 or 3600 rpm rotor speeds endured a transient temperature from 48.9 to 104.4 or 115.6 °C with CFU reductions of 0.69 and 2.84 log cycles, respectively. Utilizing hydrodynamic cavitation to obtain minimally processed pasteurized low-acid and commercially sterilized high-acid fluid foods is possible with appropriate process considerations for different products.