Antibacterial activity of chitosan against Aeromonas hydrophila

The effect of chitosan on growth and production of haemolysin by Aeromonas hydrophila was investigated as well as the effects of temperature, pH, salts and irradiation on the antibacterial activity of chitosan. It was found that chitosan affected growth and haemolysin production of A. hydrophila in varying degrees compared to the control. Growth and haemolysin production were clearly suppressed at 0.04% of chitosan. Suppression was more effective at pH 6.0 than at pH 7.0. The bactericidal effects of chitosan increased with increasing temperature and decreasing pH. Divalent cations at concentrations of 10 and 25 mM reduced the antibacterial activity of chitosan, in the order of Ba2+ > Ca2+ > Mg2+. Sodium ions at concentrations of 10 and 25 mM also reduced chitosan's activity. Irradiation of chitosan at 150 kGy under dry condition was effective in slightly increasing its activity.     

Tolerance and starvation induced cross-protection against different stresses in Aeromonas hydrophila

Aeromonas hydrophila is sometimes considered as a controversial human pathogen and reported to be susceptible to food processing procedures and environmental stresses. In this study, we have shown that early stationary phase cells of A. hydrophila were readily killed during up shifts in temperature (in the range 50-70 degrees C), the course of drying (at relative humidity, temperature and brightness of the laboratory) and after 5 min exposure to 20%, 30% and 40% v/v ethanol. However, this bacterium was found moderately susceptible to down shift to 4 degrees C in nutrient poor water, sodium chloride stresses (1.5 and 2 M) and to 12% and 15% v/v ethanol stresses. Tolerance against 1 M NaCl and 10% v/v ethanol was observed. At ambient temperature (24.5 degrees C), this microorganism exhibited a starvation survival state, which was largely independent of the initial cell concentrations (8.82, 7.71 and 6.76 log units). The cross-protection experiments showed that cells starved for short (1 day) or prolonged (50 days) periods developed increased resistance to down shift at 4 degrees C and ethanol stress. This may be of concern to the food-processing industry from the public health perspective.     

Hemolytic and proteolytic activities of Aeromonas hydrophila and Aeromonas veronii biovar sobria in broth and salmon extract at different temperatures

Expression of hemolytic and proteolytic activities throughout the growth cycle was investigated with two enterotoxic aeromonad strains assigned to the species Aeromonas hydrophila and Aeromonas veronii biovar sobria. Although growth kinetic data were dependent on strain, temperature, and substrate, maximum populations attained were higher than 9 log CFU/ml in aerated tryptone soya broth plus yeast extract (TSBYE) and salmon extract within the range 4 to 28 degrees C. For both strains in TSBYE, variable amounts of hemolytic activity were first detected at any temperature when aeromonad counts were over 9 log CFU/ml. Afterwards, this activity increased up to similar levels (109 to 112 hemolytic units per ml) without a significant increase in populations. Salmon extract supported hemolysin synthesis at 28 but not 4 degrees C. Proteolytic activity of the A. hydrophila strain was only expressed in salmon extract at 28 degrees C, whereas A. veronii biovar sobria did at 28 degrees C in both substrates and at 10 degrees C in TSBYE.     

Growth of and toxin production by Aeromonas hydrophila and Aeromonas sobria at low temperatures

The effects of different temperatures on the growth and toxin production of Aeromonas hydrophila and Aeromonas sobria were studied. The results showed that these Aeromonas species are not only able to grow at low temperatures (e.g. at 4 and 10 degrees C) but may also produce cytotoxin, hemolysin and enterotoxin under suitable growth conditions.     

Inhibitory Effect of Food Preservatives on Protease Secretion by Aeromonas hydrophila

Butylated hydroxyanisole (BHA), propylhydroxy parabenzoate (paraben), and sodium tiipolyphosphate (TPP) were found to inhibit protease secretion by resting cells of Aeromonas hydrophila at lower concentrations than those required for inhibiting growth. Incorporation of the above compounds in calcium caseinate agar resulted in colonies surrounded by smaller areas of clear zones indicating inhibition of protease secretion. The results are discussed with respect to the protective influence of the above compounds against spoilage of flesh foods by microbial proteases.     

Carnosine Increases Irradiation Resistance of Aeromonas hydrophila in Minced Turkey Meat

The effect of carnosine on irradiation inactivation kinetics of Aeromonas hydrophilawas studied. In phosphate buffer, carnosine had no effect on survival of the pathogen, whereas in turkey meat supplemented with carnosine the adverse effect of irradiation on A. hydrophila was reduced. Peroxy radicals were toxic to A. hydrophila , and this effect was counteracted by carnosine. Accordingly, in a kinetic test carnosine was capable of scavenging peroxy radicals. These results indicate that a free radical scavenger antioxidant such as carnosine can increase the radiation resistance of A. hydrophila in meat due to the inhibition of lipid peroxidation caused by radiolysis.     

Growth kinetics and predictive model of Aeromonas hydrophila in a broth-based system

This study developed a predictive model of Aermonas hydrophila in tryptic soy broth for any combination of temperatures (5 to 40°C), pH (6 to 8), and NaCl (0 to 5%) using a response surface model. A. hydrophila tended to grow within a pH range of 6.0 to 8.0 and could not tolerate NaCl up to 5.0%. The interaction of pH and NaCl did not affect the specific growth rates (SGR). The primary model to obtain the SGR showed a good fit (R²≥0.980). A secondary model was obtained by non-linear regression analysis and calculated as: SGR= 0.4577+0.0529X₁−0.1641X₂−0.1493X₃−0.0016X₁X₂−0.0001X₁X₃+0.0115X₂X₃0.0006X₁ ²+0.0114X₂ ²+0.0150X₃ ² (X₁=temperature, X₂=pH, X₃=NaCl). The appropriateness of the polynomial model was verified by the mean square error (0.0023), bias factor (0.922), accuracy factor (1.343), and coefficient of determination (0.937). The newly secondary model of SGR for A. hydrophila could be incorporated into the tertiary model to predict the growth of A. hydrophila.     

Combined effects of Lactobacillus casei inoculum, modified atmosphere packaging and storage temperature in controlling Aeromonas hydrophila in ready-to-use vegetables

Carbon dioxide concentration, Lactobacillus casei inoculum size and storage temperature were varied according to a Central Composite Design in order to assess the effects of these variables and their interactions on the growth of Aeromonas hydrophila and lactic acid bacteria in ready-to-use mixed salad vegetables packaged under modified atmosphere. The growth curve parameters, modelled according to the Gompertz equation, were analysed to generate polynomial equations. The model obtained emphasized the role of L. casei inoculum size in controlling A. hydrophila and permitted identification of appropriate combinations of the selected variables to reduce A. hydrophila survival. The use of these hurdles may increase the shelf-life and the microbiological safety of ready-to-use vegetables.     

Experimental evidence for toxin production by Aeromonas hydrophila and Aeromonas sobria in a meat extract at low temperatures

The ability of enterotoxigenic strains of Aeromonas hydrophila and Aeromonas sobria to produce exotoxins (enterotoxin and haemolysin) in a meat extract at low temperatures (5 and 12 degrees C) was investigated. All three strains incubated at 12 degrees C were enterotoxigenic and haemolytic in the meat extract after 5 days. At 5 degrees C, five of the six strains tested were able to produce these exotoxins after 8 days incubation while one strain was neither enterotoxigenic nor haemolytic after 5, 8 and 11 days. The possible involvement of performed toxin(s) in Aeromonas gastroenteritis is also discussed.     

Effects of refrigeration and alcohol on the load of Aeromonas hydrophila in oysters

Members of the bacterial genus Aeromonas are widely distributed throughout the environment and are readily cultured from a variety of foods. One member of this genus, Aeromonas hydrophila, has been reputed to be a significant cause of gastrointestinal disease. In this study, we examined the effects of refrigeration and alcohol on the level of A. hydrophila in oysters. Specifically, vodka was examined because it is used by the food service industry in preparation of Oysters Romanoff. One set of oysters was shucked on receipt, whereas others were refrigerated intact for 7 days at 5 degrees C. The oysters were blended and the numbers of A. hydrophila present determined using starch ampicillin agar. Oysters were also shucked and placed on the half shell with 5 ml of vodka for 10 min. The oysters were then washed and presumptive A. hydrophila levels determined in both the washate and homogenate. On the day of purchase, the average number of presumptive A. hydrophila found was 7.6 x 10(4) CFU/g of oyster meat. After 7 days of refrigeration, the average number had increased to 3.2 x 10(5) CFU/g of oyster meat. In the oysters treated with vodka, the average number of A. hydrophila present internally was 9.9 x 10(4) with high numbers (10(3) to 10(4)) isolated from the oyster surface. From these data, it is clear that refrigeration and alcohol treatment are not sufficient to reduce loads of A. hydrophila in or on oysters.     

Enumeration and confirmation of Aeromonas hydrophila, Aeromonas caviae, and Aeromonas sobria isolated from raw milk and other milk products in Northern Greece.

A total of 138 raw cow's and 57 raw ewe's milk samples; 80 pasteurized cow's milk samples; 39 Anthotyros cheese, 36 Manouri cheese, and 23 Feta cheese samples; and 15 rice pudding samples were examined for the presence and any countable population of Aeromonas species. Twenty-two (15.9%) of the 138 cow's milk samples analyzed were contaminated with A. hydrophila. In 13 of these samples, populations of 3.0x10(2) to 5.0x10(3) CFU/ml were counted in starch ampicillin agar (SAA). Eighteen cow's milk samples (13.0%) were contaminated with A. caviae, and in eight of these samples, populations of 2.0x10(2) to 3.0x10(3) CFU/ml were counted in SAA. Five cow's milk samples (3.6%) were contaminated with A. sobria, and in two of these samples, populations of 2.5x10(3) and 5.0x10(3) CFU/ml were counted in SAA. Eleven cow's milk samples (7.9%) were contaminated with other Aeromonas spp. not classified. Eight (14.0%) of the 57 ewe's milk samples analyzed were contaminated with A. hydrophila. In these samples, populations of 5.0x10(2) to 5.0x10(3) CFU/ml were counted in SAA. Six ewe's milk samples (10.5%) were contaminated with A. caviae, and populations of 1.5x10(2) to 1.0x10(3) CFU/ ml were counted in SAA. Two ewe's milk samples (3.5%) were contaminated with A. sobria, and populations counted in SAA were 5.0x10(2) and 1.0x10(3) CFU/ml. Four samples (7.0%) were contaminated with other Aeromonas spp. not classified. A. hydrophila was recovered in 4 (10.2%) and 3 (8.3%) of the Anthotyros and Manouri cheese samples analyzed, respectively, but no countable populations were noted in SAA. None of the pasteurized milk, Feta cheese, and rice pudding samples yielded Aeromonas spp. The results of this work indicate that motile Aeromonas are common in raw milk in Greece. Also, the presence of A. hydrophila in the whey cheeses Anthotyros and Manouri indicates that postprocessing contaminations of these products with motile Aeromonas may occur during production.     

Determination of viability of Aeromonas hydrophila in increasing concentrations of sodium chloride at different temperatures by flow cytometry and plate count technique

Aeromonads in waters and foods can represent a risk to human health. Factors such as sodium chloride concentration and temperature can affect growth and viability of several food and water-borne pathogens. The behaviour of an Aeromonas hydrophila strain in the presence of 1.7%, 3.4% and 6% NaCl concentrations at 24 degrees C and 4 degrees C was studied over a 188 day period. Viability and membrane potential were assessed by flow cytometry; growth was evaluated by plate count technique. Flow cytometry evidenced that A. hydrophila retained viability over the period although varying according to temperature and salt concentrations. Colony Forming Units were generally lower in number than viable cells especially in the presence of 6% NaCl, indicating the occurrence of stressed cells which maintain metabolic activity yet are not able to grow on agar plates. In conclusion, A. hydrophila showed a long-term halotolerance even at elevated (6%) NaCl concentrations and a lesser sensitivity to salt at low temperature; therefore, low temperature and salt, which are two important factors limiting bacterial growth, do not assure safety in the case of high initial contamination. Finally, cytometry appears a valid tool for the rapid detection of the viability of pathogenic bacteria in food and environmental matrices to control and prevent health risks.     

Partial purification and characterization of an extracellular proteinase from Aeromonas hydrophila strain A4

An extracellular metalloproteinase from Aeromonas hydrophila strain A4, isolated from milk, was purified by a factor of 300 by chromatography on DEAE-cellulose and Sephadex G-150. The enzyme had a mol. wt of 43,000 and contained 2 g atom Ca/mol. It was active over a pH range 4.8-9.5 and had optimum activity on casein at pH 7.0 with Km = 0.17 mM. It was strongly inactivated by metal chelators and the apoenzyme was fully reactivated with Ca2+, Mn2+ or Co2+. Heavy metal ions such as Ag+, Hg2+, Fe2+, Zn2+, Cd2+, Ni2+ and Cu2+ totally or partly inactivated the enzymic activity at 5 mM concentration. The enzyme was not inactivated by diisopropylfluorophosphate, soyabean trypsin inhibitor or sulphydryl group reagents. It was optimally active at 45 degrees C; above 50 degrees C activity declined rapidly, but significant activity persisted at 4 degrees C. It was heat labile in phosphate or Tris-maleate buffer but exogenous Ca2+ afforded protection.     

The growth of Aeromonas hydrophila K144 in ground pork at 5 degrees C

The influence of NaCl, pH, atmosphere, and background microflora on the growth and/or survival of Aeromonas hydrophila K144 was studied in ground pork held at 5 degrees C. In ground pork, A hydrophila was sensitive to pH values below 6.0 in the form of either a low starting pH in the pork itself or induced by lactic acid bacteria action on added glucose. Growth of the organism is inhibited by NaCl levels of 3% (w/w) (approx. 4% brine content). A hydrophila grew in vacuum-packaged ground pork; its growth was diminished by the presence of the naturally occurring meat microflora. Except for pH values below 6.0, conditions which inhibited growth permitted survival of the organisms for extended periods. Data indicate that the growth of A. hydrophila in ground pork can be controlled by factors such as NaCl, pH, and background microflora. In general, measures designed to control other foodborne pathogens appear adequate to limit A. hydrophila.     

Antibiotic resistance of Aeromonas hydrophila isolated from marketed fish and prawn of South India

From February to November 1999, 198 samples of chicken meat for sale in retail outlets and supermarkets in nine provinces of Castilla and León (Spain) were analysed for the prevalence of Salmonella and Campylobacter. Salmonella was isolated from 71 (35.83%) of the samples analysed. The predominant serovars were S. enteritidis (47.88%), S. hadar (25.35%) and serotype 4,12:b:-(II) (19.71%). Other serovars such as S. mbandaka, S. derby, S. virchow and S. paratyphi B were isolated in much lower levels. Thermophilic campylobacters were isolated in 49.50% of the samples studied.     

Fate of Listeria monocytogenes and Aeromonas hydrophila on Catfish Fillets Cooked in a Microwave Oven

Channel catfish fillets were inoculated with SJ lo⁶ cell/cm² each of Listeria monocytogenes and Aeromonas hydrophila and cooked in a microwave oven to internal temperatures of 55, 60, and 70°C. The fillets were either left uncovered or covered with polyvinylidene chloride films during cooking. A. hydrophila populations were reduced to nondetectable levels on covered fillets cooked to 70 °C. At 60 °C, L. monocyrogenes populatoins were reduced by = 4 logs for covered and by 2 logs for uncovered fillets. Covering fillets increased lethality at each temperature; however, some pathogens could survive at < 60 °C.     

Innovative applications of food-related emulsions

Research on oxidative stability of multiple emulsions is very scarce. Given that this is a relevant topic that must be ascertained before the successful application of multiple emulsions in foods (especially when a combination of highly unsaturated oils is used as a lipid phase), this review mainly focuses on various aspects of the multiple emulsions.

Fat replacement in meat products using emulsions is critically discussed along with innovative applications of natural antioxidants in food-based emulsions and multiple emulsions based on bioactive compounds/encapsulation as well as confectionery products.     

Effect of Ether Extracts from Condensed Wood Smokes on the Growth of Aeromonas hydrophila and Staphylococcus aureus

Ether extracts of smoke condensates (“liquid smokes”) from 20 different woods were screened for antimicrobial activity. Testing was done in trypticase soy broth of pH values in the range of 5.78–6.81 against three strains each of Aeromonas hydrophila (ATCC 7965, K1414, BW37) and Staphylococcus aureus (ATCC 12600, LP, S6) incubated at 35°C. There was variation in inhibition of growth among types of liquid smoke, species, and strains of microorganisms tested. Staphylococcus aureus was generally more sensitive to inhibition than A. hydrophila strains. The most inhibitory liquid smoke was derived from the sapwood of douglas fir. Other inhibitory liquid smokes were from birch, southern yellow pine, and aspen. The least inhibitory liquid smokes were derived from lodge pole pine and mesquite woods.     

Toxin production by strains of Aeromonas hydrophila grown in laboratory media and prawn purée

Toxin production by four strains of Aeromonas hydrophila grown at 30 and 37 degrees C in two laboratory media and prawn purée was studied. Three different cell lines were used to test for cytotoxic activity, haemolytic activity was tested against rabbit and guinea pig erythrocytes, proteolytic activity was assayed with azo-casein and enterotoxic activity using the suckling mouse assay. Results showed reduced cytotoxic and haemolytic activities in prawn purée compared with the two media, but in most cases increased proteolytic activity. No enterotoxic activity was observed in prawn purée although it was occasionally detectable in both laboratory media.     

Enterotoxigenicity and drug sensitivity of Aeromonas hydrophila isolated from well water in Sweden: a case study

A large number of Aeromonas spp. have been found in drinking water from a drilled well in Sweden. Isolates identified as A. hydrophila were tested for production of enterotoxin, hemolysin, enzymes and for resistant patterns to different antibiotics. The enterotoxin-producing A. hydrophila could be responsible for the long-term diarrhoeal case of a 1 1/2 year old child who consumed the contaminated water.