GROWTH OF LISTERIA MONOCYTOGENES, AEROMONAS HYDROPHILA AND YERSINIA ENTEROCOLITICA ON COOKED MUSSEL TISSUE UNDER REFRIGERATION AND MILD TEMPERATURE ABUSE

Strains of the psychrotrophic bacteria Listeria monocytogenes, Aeromonas hydrophila and Yersinia enterocolitica were inoculated onto samples of cooked mussel tissue and incubated at 5 and 10C under both aerobic and anaerobic conditions. These organisms were enumerated over a time course and data were analyzed to give values for the lag and generation times. All species grew under all conditions, and the rate of growth was similar under aerobic and anaerobic incubation. Predicted growth kinetic values from response surface models and measured growth rates for L. monocytogenes and A. hydrophila were in reasonable agreement, but similar comparisons for Y. enterocolitica gave shorter predicted lag and generation times than were measured. A combination of models from different sources would have given more accurate predictions than any one individual model.     

FACTORS AFFECTING GROWTH OR SURVIVAL OF AEROMONAS HYDROPHILA IN FOODS

Recent surveys of retail fresh foods (fish and seafood, poultry, red meat, raw milk and vegetables) have indicated the presence of organisms of the Aeromonas hydrophila group in virtually every sample examined. As part of these surveys, A. hydrophila was observed to increase in number during storage of the food at 5°C. Thus, other measures to control the growth of these organisms in retail fresh foods must be sought and evaluated; in addition, measures to destroy any A. hydrophila introduced into the foods must be investigated. This review discusses the various measures besides temperature (pH, NaCl) to control the growth of A. hydrophila in foods as well as their destruction by heat, irradiation, and sanitizers.     

HIGH HYDROSTATIC PRESSURE EXTRACTION OF SALIDROSIDE FROM RHODIOLA SACHALINENSIS

A nonthermal process, high hydrostatic pressure (HHP), was used as a new extraction technique to obtain salidroside from Rhodiola sachalinensis. The leaching rates of salidroside were measured under different treatment conditions. When the material was treated under 500 MPa of hydrostatic pressure with 50 times of volume 60% (v/v) of ethanol solution as the extracting solvent, the leaching rate of salidroside was up to 0.401% in 3 min, while that of ultrasonic extraction was at 0.288% in 30 min and that of reflux extraction was at 0.302% in 120 min. The HHP extraction technique showed high efficiency in extracting salidroside at room temperature using moderate ethanol concentration.

PRACTICAL APPLICATIONS

Rhodiola is prevalently used as a safe and effective antifatigue drug and as an adaptogen around the world. But the common extraction methods used to yield commercial products are still traditional ones, which could easily lead some thermosensitive ingredients to lose their biologic activities. The nonthermal extraction method is the key point of the realization of herb modernization. The high hydrostatic pressure (HHP) technique can be operated at room temperature and can shorten processing time. The extract produced by the method has proved to be of higher extraction yield than traditional varieties. In addition, the HHP technique only consumes electric energy without exhaust emission, which is an environment-friendly method. In the future, we will cooperate with those far-seeing companies to achieve its industrialization. The annual sale of herbs in the whole world has reached 30 billion. We do not doubt the prospect of this technique.     

EFFECT OF CONCURRENT HIGH HYDROSTATIC PRESSURE, ACIDITY AND HEAT ON THE INJURY AND DESTRUCTION OF LISTERIA MONOCYTOGENES

The effect of concurrent use of high hydrostatic pressure, heat and acidity on Listeria monocytogenes Scott A and CA was investigated. In general, lethality was enhanced when cells were pressurized at higher temperatures or lower pH. Strain CA demonstrated an additional 3-log₁₀ reduction when pressurized at pH 4.0 as compared with pH 6.0 at 353 MPa, 45C for 10 min. Scott A was reduced an additional 1 log₁₀ by increasing the temperature from 25C to 45C with pressurization at 252 MPa, pH 6.0 for 30 min. Exposure to 404 MPa at 45C for 30 min demonstrated complete injury or death of CA cells with an initial concentration of >10⁸ CFU/mL. At least an 8-log₁₀ reduction was observed for both L. monocytogenes strains Scott A and CA when exposed to the combined treatments of 252 MPa, 45C, pH 4.0 for 30 min .     

超高(静)压处理对果蔬理化性质的影响

超高（静）压（以下简称超高压）技术用于乳制品、果蔬汁等的报道已有很多，而超高压用于固态食品是近几年研究的新课题。综述了超高压对果蔬质构的影响。超高压可以引起果蔬变软、褐变。但是超高压比热烫对果蔬的质构影响小。     

POTENTIAL SOURCES OF AEROMONAS HYDROPHILA

Aeromonas species are associated with marine and fresh water environments, including natural mineral springs, chlorinated and unchlorinated domestic supplies, and polluted water sheds. Most studies attribute water supplied as the principal source for A. hydrophila. Recently, Aeromonas has been recognized as a potential cause of food-associated gastroenteritis outbreaks. Its actual involvement in large foodborne outbreaks is presently unknown. Aeromonas can be readily isolated from a variety of food sources such as seafoods, meats, dairy products, and poultry. Another potential source is asymptomatic and immuno-compromised individuals, particularly, food-handling workers.     

HIGH HYDROSTATIC PRESSURE PROCESSING OF PEACH PUREE WITH AND WITHOUT ANTIBROWNING AGENTS

High hydrostatic pressure (HHP) (103 to 517 MPa) was applied for selected times (5 to 25 min) on peach puree with and without the addition of antibrowning agents (1000 ppm ascorbic acid or 300 ppm cysteine). Polyphenoloxidase (PPO) activity was assessed before and after HHP processing. Peach purees were stored at 3C for 13 days and then at 22C for 15 additional days. A z_p value of 224 MPa, in a range from 207 to 517 MPa of pressure, was obtained for peach puree without the addition of antibrowning agents. PPO activation at 103, 207 and 310 MPa or inhibition at 414 and 517 MPa was observed in peach purees containing added ascorbic acid. More than 50% inactivation of PPO was observed after adding cysteine to the peach puree and 99% inhibition was observed after 517 MPa of pressure at the experimental times. The higher the pressure treatment and the longer the treatment time for inactivating PPO, the less the discoloration of the peach puree with and without antibrowning agents during storage at low temperatures.     

EFFECT OF POLYPHOSPHATES AND NaCl ON AEROMONAS HYDROPHILA K144

Polyphosphates are multifunctional ingredients added to many foods, particularly meat products. In addition to their moisture-binding properties, polyphosphates have been reported to inhibit various bacteria. In the current study, four food-grade polyphosphates were evaluated for their effects on the growth of Aeromonas hydrophila K144 in both a model system and a food system. Since polyphosphates can interact with other food factors, effect of these compounds were studied at different NaCl levels and temperatures. The model system was BHI Broth modified by the addition of polyphosphates (sodium pyrophosphate, sodium tripolyphosphate, Hexaphos, or Sodaphos) and NaCl; incubation was aerobic at 5 or 28C. The food system was ground pork with NaCl and polyphosphate added and inoculated with A. hydrophila. Individually, the polyphosphates were relatively noninhibitory in both BHI broth and ground pork, but NaCl and temperature interacted with the polyphosphates in the model system to increase generation and lag times of A. hydrophila. In BHI broth, a combination of 2% of any of the polyphosphates tested and 3.5% NaCl inactivated the bacterium; this inactivation was temperature-dependent. By both a plating system and electron microscopy, the polyphosphate-NaCl combination was shown to injure the bacterium. In the ground pork, the polyphosphate-NaCl combination limited growth of the bacterium during refrigerated storage. These results suggest that polyphosphates could be useful to control the presence of A. hydrophila in certain foods .     

EFFECTS OF ORGANIC ACIDS, NISIN, LYZOZYME AND EDTA ON THE SURVIVAL OF YERSINIA ENTEROCOLITICA POPULATION IN INOCULATED ORANGE BEVERAGES

Inactivation of a Yersinia enterocolitica strain by 3.4% ascorbic acid (AA), 0.5% citric acid (CA), 0.2% lactic acid, 100 IU/mL nisin, 2,400 IU/mL lysozyme and 20 mM ethylenediaminetetraacetic acid disodium salt (EDTA ), alone and combined, was studied at different temperatures (4 and 25C) in commercial and laboratory-prepared orange beverages. In laboratory-prepared juice, highest reductions (between 3.06 and 4.07 log units) were obtained with acid mixtures at 25C after 20 min of incubation. At 24 h of incubation, EDTA was bactericidal and the mixtures with nisin or lysozyme resulted in count reductions of 6.03 and 5.98 log units, respectively. In commercial orange beverage, AA and CA gave reductions of 5.43 and 4.26 log units, respectively. The three acid mixtures were bactericidal within the first 10 min. EDTA alone or mixed completely inhibited Y. enterocolitica strain at 6 and 24 h of incubation. At 4C, all the results were significantly lower than at 25C.

PRACTICAL APPLICATIONS

Yersinia enterocolitica is a common cause of gastrointestinal disorders and may lead to sequelae. This pathogen can maintain its virulence in a wide temperature range and under adverse conditions such as pH lower than 4. There are reports describing the isolation of Y. enterocolitica from water and ice in Brazil and Latin America. The sale of unpasteurized drinks is a common practice in our region and their consumption has increased over the past years. In the present work, chemical compounds and biological inhibitors were studied. Some of these compounds showed to be efficient for reducing Y. enterocolitica populations. Furthermore, they did not modify the organoleptic characteristics of the beverages and they are not toxic for human beings.     

THERMAL RESISTANCE OF YERSINIA ENTEROCOLITICA AND LISTERIA MONOCYTOGENES IN MEAT AND POTATO SUBSTRATES

The heat resistance of a wild type and nalidixic acid resistant strain of Yersinia enterocolitica, and Listeria monocytogenes, was measured in meat (minced beef and minced beef homogenate) and potato substrates over the temperature range 50–60C. Comparisons of heat resistance were determined using D-values calculated using a linear survival model. The results showed that the wild-type strain of Y. enterocolitica was more heat resistant than the mutant (p<0.05). Under most conditions, the use of a nonselective/overlay recovery medium resulted in higher D-values compared to a selective recovery medium (p<0.05). Analysis of the data using a nonlinear survival model (D₁ and D₂ -values) suggested the presence of heat resistant subpopulations and was particularly evident for the mutant strain, and in potatoes compared to minced beef.     

TOXICOLOGY AND HISTOPATHOLOGY OF THE HEAT-STABLE ENTEROTOXIN OF YERSINIA ENTEROCOLITICA IN INFANT AND ADULT MICE

Yersinia enterocolitica heat-stable enterotoxin (YST) was partially purified by ion exchange, gel filtration and Fast Protein Liquid chromatography. Infant mice given YST orally (p.o.) exhibited diarrhea 4 h after administration; a few infants died by 6.5 h. Pups dosed intraperitoneally (i.p.) and adults dosed p.o., i.p. and intravenously showed no symptoms during the 7 days of observations. Tissues of adult mice orally challenged with YST showed no pathological changes by light microscopy. Livers of infant mice orally dosed with YST showed evidence of severe glycogen depletion. The intestinal tracts of YST-dosed (p.o.) infant mice exhibited severe pathological changes including: epithelial cell and villous tip necrosis, villous edema, and capillary and lacteal engorgement. Kidneys of YST-dosed (p.o.) infant mice exhibited devacuolization of the proximal convoluted tubule cells which indicates metabolic inactivity and a probable decrease in the resorption of glomerular filtrate (protein, carbohydrate, electrolytes and water).     

LACK OF CORRELATION BETWEEN PLASMID-ASSOCIATED PHENOTYPES OF YERSINIA ENTEROCOLITICA AND PATHOGENICITY IN THE MOUSE1

Various phenotypic characteristics have been correlated with the pathogenicity of plasmid-bearing virulent strains of Yersinia enterocolitica. Two transposon Tn801-insertion derivatives (JD193 and JD217) of the virulence plasmid from serotype 0:3 were used to determine the correlation between pathogenicity and plasmid-associated properties of this organism. Both Tn801-inserted derivatives expressed five plasmid-associated phenotypic characteristics at 37C: (1) colony morphology, (2) calcium-dependent growth or low-calcium response, (3) crystal violet binding, (4) autoagglutination, and (5) hydrophobicity. However, for mouse pathogenicity only JD193 was positive whereas, JD217 was avirulent for mice. Thus, it is possible to have a lack of correlation between plasmid-mediated traits and the actual pathogenicity of the organism in the mouse; however, these plasmid-mediated phenotypic characteristics provide simple and efficient techniques to evaluate the virulence potential of wild-type strains isolated from food poisoning outbreaks and clinical cases.     

EXPANDED MODEL FOR THE AEROBIC GROWTH OF AEROMONAS HYDROPHILA

The previously published (Palumbo et al. 1991) response surface model for describing the influence of temperature, pH, NaCl, and sodium nitrite on the aerobic growth of Aeromonas hydrophila K144 in BHI broth has been expanded to incorporate additional data. The effects of the variables on A. hydrophila aerobic growth kinetics were modeled by response surface analysis using quadratic and cubic polynomial models of (1) natural logarithm transformation of both the Gompertz B and M parameters and the lag phase duration (LPD) and generation time (GT), and (2) the square root transformation of B and 1/M calculated from 268 cultures (212 of which supported growth) from 81 variable combinations. In addition, the six models generated also were subjected to backward elimination regression analysis to remove nonsignificant variables. Based on examination of the adjusted R² values of the resulting 12 models, three were selected for further evaluation by comparing their observed and predicted T₁₀₀₀-values (time for a 1000-fold increase in number; this concept incorporates the influence of the variables on both lag and generation times), LPDs and GTs. Using this method of comparison and evaluation, models based on cubic polynomial, natural logarithm transformation of GT and LPD gave the best "first estimates" of the aerobic growth characteristics of A. hydrophila.     

EFFECT OF HIGH HYDROSTATIC PRESSURE ON SPORES OF GEOBACILLUS STEAROTHERMOPHILUS SUSPENDED IN SOYMILK

The inactivation of Geobacillus stearothermophilus spores (ATCC 7953) inoculated in soymilk was investigated using high hydrostatic pressure (550, 585 and 620 MPa) in combination with temperature (70, 80 and 90C) for selected times (2 s to 15 min). Inactivation of spores occurred at all selected treatments. Less than 10 CFU/mL of G. stearothermophilus were observed after 7 min of treatment at 620 MPa and 90C. An increase in the inactivation rate constant, at the highest pressure, was observed, resulting in a decrease in D values at all temperatures. D values were calculated as 10.6, 6.2 and 3.5 min for 70, 80 and 90C, respectively after pressurization at 620 MPa. z_p values decreased as temperature increased with values ranging from 142 to 238 MPa. The activation energy required for inactivation of G. stearothermophilus spores in soymilk, at the selected treatments, was in the range of 37.9–57.4 kJ/mol.     

EFFECTS OF HYDROSTATIC PRESSURE ON ALPHA-MACROGLOBULIN AND SELECTED PROTEASES

The enzymes, chymotrypsin, trypsin, collagenase and cathepsin C, as well as the protease inhibitor, α₂-macroglobulin, have been shown to be susceptible to hydrostatic pressure inactivation. a₂-Macroglobulin lost about 15% activity at a pressure of 2,000 atm and as much as 80% at 3,000 atm pressure applied for 1 h in both cases. Thermal stability studies also indicated the protein to be completely denatured at about 75C. The enzymes were also inactivated to various extents depending on the amount of pressure and the duration of application, The degree of susceptibility to pressures ranging between 2,000–4,000 atm applied for time periods ranging from 5–60 min was found to be as follows: chymotrypsin < cathepsin < trypsin < collagenase. The hydrostatic pressure effects were irreversible when pressure-treated enzymes were stored at room temperature (20–25C), but showed various levels of reactivation when stored at refrigerated temperatures (4–7C).