Some virulence properties and characterization of motile Aeromonas species from milk and white cheese

Three hundred and thirty-eight samples of milk and milk products in Ankara were screened for the presence of motile Aeromonas species. The overall frequency of aeromonads in these samples was 27%. As expected, raw milk samples were more contaminated with aeromonads than were other products. Sixty-five (49.2%) of the 132 bulk raw milk samples were contaminated with aeromonads. In 35 of these samples, populations of aeromonads from 1.5 × 10² to 3.0 × 10³ cfu/mL were counted in ampicillin dextrin agar (ADA). Ten (40%) of the 25 raw milk samples sold in the street were contaminated with aeromonads. In eight of these samples, 1.2 × 10²−3.0 × 10³ cfu/mL were counted. Five (16%) of the 31 pasteurized milk and 12 (8%) of the 150 white cheese samples were contaminated with Aeromonas spp., but no countable aeromonads population was noted in ADA. The incidence of A. hydrophila, A. sobria and A. caviae in all samples was found to be 90.2%, 4.3% and 5.4%, respectively. The majority of the strains identified as A. hydrophila and A. sobria were able to produce haemolysin, protease and DNase, while strains identified as A. caviae were only positive for DNase. All isolated Aeromonas species ( A. hydrophila, A. sobria and A. caviae ) were positive for uptake of Congo red dye. Nevertheless, only strains identified as A. hydrophila and A. sobria showed a high rate of positive results when tested for the production of the Voges–Proskauer reaction and lysine decarboxylase. The results of this work show that motile Aeromonas spp., especially A. hydrophila , are frequently found in these samples. As these products are usually commonly consumed in Ankara, they can pose a risk especially for children, the elderly and immunocompromised individuals.     

SHELF-LIFE OF FRESH FILLED PASTA. HAZARD ANALYSIS AND CRITICAL CONTROL POINTS OF THE MANUFACTURING PROCESS AND HOUSEHOLD PRACTICES

Hazard Analysis and Critical Control Point (HACCP) approach was used to assess the safety of a fresh filling pasta (ricotta-filled ravioli) manufacturing plant in La Plata region, Argentina. Household practices like cooking and holding meals before serving were also evaluated. Samples of ricotta, main raw material of the filling, showed colony counts of Enterobacteriaceae total microorganisms, molds and yeasts of (5 × 10³-1 × 10⁵), (1x10⁵−1x10⁸) and 1 × 10⁵ CFU/g, respectively. E. coli was also detected in one out of five samples, suggesting a hazardous condition of this raw material. Salmonella spp. was not isolated from any of the dough samples tested. Ricotta filling showed high colony counts of mesophilic microorganisms (6 × 10⁶ CFU/g), Enterobacteriaceae (1 × 10⁵ - 1 × 10⁶ CFU/g), while E. Coli was detected in 20% of the samples. Counts of B. cereus, S. aureus were less than 1 × 10² CFU/g in the analyzed materials. In the finished product (ricotta-filled ravioli), the colony counts of mesophilic microorganisms and Enterobacteriaceae were 3 × 10⁸ and 8 × 10⁵ CFU/g, respectively. Critical Control Points found were cooking and holding time before serving. The implementation of suggested corrections allowed the microbial quality of the final product (ricotta-filled ravioli) to be improved considerably. The growth of total microbial counts during refrigerated storage (0, 4, 8 and 10C) were measured and the shelf-life of ricotta and ricotta-filled ravioli was calculated.     

Incidence, source and some properties of psychrotrophic Bacillus spp found in raw and pasteurized milk

Spores of psychrotrophic Bacillus spp were isolated from 58% of farm bulk tank milks and about 69% of pasteurized milks. Counts of Bacillus spp in about 10% of raw milk samples reached 1 × 10⁵ cfu/ml and above within seven days at 6°C. Psychrotrophic spore counts in pasteurized milks ranged from <0.5 to 170 spores/litre with an average of about 17/1. There was little correlation between the total bacterial count of the raw milk and presence of psychrotrophic Bacillus spores. There was some evidence that the bulk tank itself may be a source of contamination. The spores in pasteurized milk probably were not the result of postpasteurization contamination. The optimum germination temperature for psychrotrophic Bacillus spores was lower than that for spores of mesophilic strains. About 50% of the psychrotrophic Bacillus strains isolated from milk were capable of growth at 2°C.     

Evolution of free amino acid content during ripening of Mahon cheese

Evolution of free amino acids (FAA) during ripening (10–300 days) of five batches of Mahon cheese was studied; three batches were made from raw milk and two from pasteurized milk. Major FAA were GLU, VAL, LEU, LYS and PHE (43–67% of total FAA). Relative THR, SER, GLU and GLY contents significantly increased during ripening whereas LEU, PHE and ORN decreased. GLU, ILE, SER, THR, GLY and PHE varied during ripening according to a zero order reaction (r² > 0.99). Principal component analysis showed that the first principal component could be considered representative of the ripening time. The most ripened cheeses showed high GLU, GLY, SER and THR contents and the less ripened cheeses presented high ORN, LEU, PHE and ALA contents. The second component distinguished between cheeses made with pasteurized milk (with high ASN and GLN contents) and raw milk. Indigenous microflora of raw milk showed a strong influence on the proteolytic activity in Mahon cheese.     

Isolation and characterization of the microbial population of different South African kefir grains

Kefir, a slightly acidic fermented milk, is produced by adding lactic acid bacteria and yeasts, in the form of grains, to milk. The bacteria and yeasts present in the kefir grains are known to vary widely. Selective growth media and morphological and biochemical characteristics were used for the isolation and identification of the microbes present in the grains from eight different sources in South Africa. The kefir grains were activated in milk for only 24 h to prevent any changes in the microbial population of the grains. The microbial numbers varied between 6.4 × 10⁴ and 8.5 × 10 ⁸  cfu/g on the media selective for the bacterial species and between 1.5 × 10 ⁵ and 3.7 × 10 ⁸   cfu/g on the media selective for the yeast species. The bacterial genera that were identified included Lactobacillus , Leuconostoc and Lactococcus and the yeast genera included Zygosaccharomyces , Candida and Saccharomyces . The distribution frequencies of the microbes in the different grains were determined and most of the grains were dominated by two microbial species. No pediococci, acetic acid bacteria or propionibacteria were detected.     

Production of Enterotoxin-B in Cured Meats

SUMMARY— A variety of laboratory cured hams were inoculated with 10³−10⁶ cells of S. aureus strain S-6 and incubated at 10, 22 and 30°C anaerobically for up to 16 weeks. Enterotoxin-B was detected by gel-diffusion in hams with original pH over 5.30, up to 9.2% NaCl (brine) and 0.54 ppm undissociated nitrous acid. There was better toxin production at 30° than at 22° or 10°C. Toxin was detected at 10°C after at least 2 weeks incubation and in most samples after 8 weeks when pH was greater than 5.6. Toxic hams had more than 4 × 10⁶ cells/g. Contaminants were always less than 10⁵/g. Tween 80 inhibited toxin production at 30° but not at 10°C. Toxic hams looked normal even after 2 months incubation at 10°C.     

Microbial flora of technological interest in raw ovine milk during 6°C storage

Changes in the microbial flora of ovine milk of medium hygienic quality (initial mean total plate count 3.3  ×  10⁵ cfu/mL) were studied throughout refrigerated storage at 6°C. Total plate counts after 48 h of refrigerated storage (mean count 4.6  ×  10⁵ cfu/mL) were under the current standards in the European Union (EU) for raw ovine milk to be used for cheesemaking after heat treatment. However, after 96 h, mean total plate counts (1.6  ×  10⁷ cfu/mL) were above the standards. Lactococci were found at levels higher than Pseudomonas spp. in milk freshly drawn (54.5% and 3.5% of total plate count, respectively) and after 96 h at 6°C (47.9% and 33.9% of total plate count, respectively). Lactobacilli, enterococci, coliforms and thermodurics were found at values lower than lactococci and Pseudomonas spp., before and after refrigerated storage ( ≤  0.08% of the total plate count). A significant growth ( P  < 0.001) was detected for mesophiles, Pseudomonas spp. and lactococci after 96 h of refrigeration at 6°C; however, thermodurics, coliforms, lactobacilli and enterococci, showed no significant increase ( P  > 0.05). Presumptive Escherichia coli (β-glucuronidase-positive) underwent a decrease throughout storage at 6°C.     

EFFECTS OF RAW MATERIALS AND PROCESS VARIABLES ON THE HEAT PENETRATION TIMES, FIRMNESS, AND PECTIC ENZYME ACTIVITY OF DICED TOMATOES (HALLEY BOS 3155 CV)

The effects of raw materials and process variables on the heat penetration times into diced tomatoes (Halley Bos 3155 cv) were evaluated. Variables included dice size (1.27 and 2.54 cm), maturity at harvest (red and red+2 weeks), and processing temperature (88 and 92C). Heat penetration times between dice sizes were significantly different, but not between maturities or processing temperatures. Tomatoes were also evaluated for firmness, pectin-methylesterase (PME) and polygalacturonase (PG) activities. Half-inch size diced tomatoes were processed at 88 and 92C, and evaluated for firmness using the shear-compression method. Firmness decreased to 60% of the initial raw firmness from 8.8 × 10⁵ to 5.3 × 10⁵ g-mm after 15 s at 88C, and to 50% from 8.8 × 10⁵ to 4.4 × 10⁵ g-mm after 15 s at 92C. Diced tomato firmness showed a slight firming trend after 150 s at both temperatures. PME was inactivated after 45 s, while 5% residual PG activity remained after 3 min.     

Rapid dye reduction tests for the determination of microbiological quality of meat

Rapid dye reduction tests have been developed to determine the quality of meat. Three chemical indicators, resazurin and two tetrazolium compounds, were used to correlate the microbial numbers and reduction times in meat samples. Twenty-five surface samples from sheep carcasses were subjected to each reduction test. Total viable counts given were obtained at 37°C. Resazurin reduction time was 90–120 min when the bacterial counts ranged from 1.5×10⁶ to 7.7×10⁶/cm². Samples showing bacterial counts between 1.5×10⁶ and 6.0×10⁶/cm² reduced tetrazolium (NBT) in 360–390 min whereas samples containing bacterial counts of 2.1×10⁶/cm² took 420–450 min to reduce iodophenyl nitrophenyl tetrazolium (INT) dye. Regression equations relating the number of organisms per cm² and reduction time were applied to predict the microbiological quality of meat samples from reduction time data. Among the three dyes, resazurin gave the lowest reduction time.     

MOLD GROWTH AND PRESENCE OF AFLATOXIN IN SOME TURKISH CHEESES

Twenty-five random fresh market samples of Van herby cheese and pickled white cheese were examined for molds and aflatoxins. The mean total mold count in Van herby cheese was 2.50 × 10⁵/g; in pickled white cheese it was 4.95 × 10⁴/g. The mycoflora on the cheeses were determined. In all cheeses, over 65% of molds were Penicillium species . Aspergillus made up 0 to 1.6 % and 2.6 % to 4.0 % of the mold on pickled white cheese and Van herby cheese, respectively. Other isolated molds belonged to Mucor, Geotrichum and Trichoderma genera. None of the samples contained aflatoxins and none of the 6 Aspergillus isolates was an aflatoxin producer .     

Viscosity changes during rennet coagulation of Murciano-Granadina goat milk

The viscosity changes during coagulation of raw and pasteurized Murciano-Granadina goat milk with 10 rennets and coagulant enzymes were studied using a cylindrical rotational viscometer. The effect of three shear rates (7.32, 14.68, 73.42 s^-1) was analysed in reconstituted milk using 60% chymosin rennet and different enzyme concentrations. The shear rate selected (14.68 s^-1) corresponded to that in which the coagulation time was not modified. At a sheer rate of 73.42 s^-1 there was a significant difference (p < .05) in the milk clotting time determined compared with the shear rates of 7.32 and 14.68 s^-1. At the beginning of the coagulation process there was a decrease in the viscosity value, which was higher in raw milk (22%) than in the two heat treated samples (17% in milk pasteurized at 65°C for 30 min, low temperature long time, and 19% in milk pasteurized at 75°C for 15 s, high temperature short time). After this period the viscosity rose rapidly and reached a value 2.89 times greater than at time 0. A highly significant Pearson correlation (0.952-0.994) was found between the coagulation time determined by the Berridge method and viscometric measurement in raw and pasteurized milk.     

Influence of probiotic Lactobacillus acidophilus and L. helveticus on proteolysis, organic acid profiles, and ACE-inhibitory activity of cheddar cheeses ripened at 4, 8, and 12 degrees C

ABSTRACT:  The influence of adjunct bacteria on composition of cheeses, organic acid profiles, proteolysis, and ACE-inhibitory activity during ripening at 4, 8, and 12 °C for 24 wk was investigated. cheddar cheeses were made with starter lactococci (control), Lactobacillus acidophilus L10, and starter lactococci (L10), and L. acidophilus L10, L. helveticus H100, and starter lactococci (H100). The counts of L. acidophilus in L10 cheeses remained at >10⁶ colony forming units (CFU)/g after 24 wk of ripening at 4, 8, and 12 °C. Concentrations of lactic, acetic, and propionic acids of the L10 and H100 cheeses were significantly higher than those of the control cheeses after 24 wk of ripening ( P < 0.05). Proteolysis of the cheeses was improved as the ripening temperature increased. Water-soluble nitrogen, trichloroacetic acid soluble nitrogen, and phosphotungstic acid soluble nitrogen of L10 and H100 cheeses were significantly higher than those of the control cheeses ( P < 0.05). Increase in ripening temperature from 4 °C to 8 and 12 °C increased the percentage of ACE inhibition. The IC₅₀ value among cheeses ripened at 4, 8, and 12 °C, however, was not significantly different ( P > 0.05). Hence, probiotic L. acidophilus L10 can be added into cheddar cheeses to improve proteolysis and ACE-inhibitory activity.     

THE EFFECT OF AEROBIC MESOPHILIC MICROFLORAL LEVELS ON THE ISOLATION OF INOCULATED LISTERIA MONOCYTOGENES STRAIN LM82 FROM SELECTED FOODS

The effect of aerobic mesophilic microfloral concentration on the isolation of Listeria monocytogenes LM82 was studied in 31 (18 cheeses and 7 noncheese) retail foods having standard plate counts of 10¹ to 10⁸ colony forming units (CFU)/g. Foods were spiked with L. monocytogenes and enriched at 30°C for 24 h in a selective enrichment broth used in a U.S. Food and Drug Administration method. Inoculum levels for isolation on modified McBride agar ranged from 0.1 to > 5 × 10³ with a geometric mean value of 5 inoculated CFU/g or 1.4 CFU/g. Pure Enterococcus (Streptococcus) faecalis ( 0 to 6 × 10⁶ inoculated CFU/mL ) in the absence of food matrix had no effect on the enrichment of L. monocytogenes. Ease of isolation of LM82 was independent of the food microflora concentration both generally and in the specific food type of 9 Brie cheeses. Competition, when it occurs, therefore, may be due to specific bacterial competitors rather than bacterial numbers .     

Biogenic Amines Content of Four Types of “Pecorino” Cheese Manufactured in Tuscany

Biogenic amines content of four types of Tuscan ewes’ milk “pecorino” cheese was evaluated using HPLC-UV analysis. All cheeses were manufactured in the same dairy factory with different combinations of milk (raw or pasteurized) and type of ripening. Total biogenic amines and tyramine levels of a raw milk “pecorino” ripened five months, partly in a traditional cave, were significantly higher than those of a pasteurized milk “pecorino” with a similar ripening; and of a two months raw milk “pecorino” ripened in the dairy plant. No statistical significant difference was found when comparing total biogenic amines and tyramine contents of the same five month ripened raw milk “pecorino” with a pasteurized milk “pecorino” ripened six months, partly in a traditional “fossa.” In raw milk cave-ripened and “fossa”-ripened cheeses, total biogenic amines exceeded 1000 mg/kg. In cheeses manufactured with raw milk and/or in particular ripening environments, specific hygienic cares are needed to limit biogenic amines formation.     

MICROBIOLOGICAL QUALITY OF TRADITIONAL MARKET CURED FISH IN TANZANIA

The microbiological quality of six varieties of retail market traditionally cured fish in Morogoro, Tanzania was investigated over a five-month period. The fish were contaminated with bacteria and molds at levels of: total aerobes, 10⁶ - 1.7 × 10⁷ c.f.u/g; faecal coliforms, 1.1 × 10¹ - 2.5 × 10³ MPN/g; faecal streptococci, 1.4 × 10¹ - 1.3 × 10³ MPN/g; Staphylococcus aureus, 1.3 × 10³ - 8.6 × 10³ c.f.u/g; Aspergillus flavus group, 2.1 × 10¹ - 2 × 10² c.f.u/g of fish. Of faecal coliform, 45% of the isolates were Escherichia coli. Twenty-five percent of the S. aureus isolates were coagulase positive. Sixteen percent of A. flavus isolates were aflatoxigenic. Aflatoxin contamination ranged from O to 18.5 μg/kg of fish. Insect infestation by Dermestes spp. and mites was observed. The results of this preliminary study emphasize the importance of proper processing and handling offish in the tropics in order to safeguard public health .     

Biochemical composition and storage stability of a yogurt-like product from African yam bean (Sphenostylis stenocarpa)

The biochemical and storage properties of African yam bean ( Sphenostylis stenocarpa ) yoghurt-like product were evaluated. Milk extracted from the bean flour using hot water was supplemented with 4% milk protein concentrate (MPC), 0.15% dairy calcium (DC) and 0.5% gelatine (G) (singly or in combination). The milk was fermented with Streptococcus thermophilus and Lactobacillus delbrueckii spp bulgaricus at 43 °C for 3–5 h. Supplemented African yam bean (AYB) yoghurts had a gel-like consistency unlike the control (unsupplemented) sample. Total solids, protein, fat, ash, lactic acid bacterial count, lightness (L-value) and viscosity were improved by supplementation. Riboflavin and antioxidants were reduced; macroelements and thiamine were increased by 0.71–15.6% and 16.7%, respectively, because of fermentation. The stability of the product during storage at 4 °C was improved by supplementation and stirring. The pH of the supplemented product ranged from 4.45 to 4.54 and microbial counts from 2.6 × 10⁶ to 1.6 × 10⁸ during storage for not more than 3 weeks.     

Singlet Oxygen Oxidation Rates of ∝-, γ-, and δ-Tocopherols

ABSTRACT:  The reaction rate constants of 5 × 10⁻⁴ M, 10 × 10⁻⁴ M, and 20 × 10⁻⁴ M α-, γ-, and δ-tocopherols with singlet oxygen in methylene chloride containing 1 × 10⁻⁵ M chlorophyll under light at 25 °C for 60 min were studied. The oxidation of tocopherols determined by a spectrophotometric method showed that the losses of 20 × 10⁻⁴ M α-, γ-, and δ-tocopherols after 60 min under light were 21%, 16%, and 9%, respectively. The degradation of α-, γ-, and δ-tocopherols was undetectable in the absence of chlorophyll under light or in the presence of chlorophyll in dark. The losses of tocopherols under light were mainly due to singlet oxygen oxidation. The degradation rates of 20 × 10⁻⁴ M α-, γ-, and δ-tocopherols were 6.6 ×10⁻⁶ M/min, 5.0 × 10⁻⁶ M/min, and 2.9 × 10⁻⁶ M/min, respectively. The reaction rates between α-, γ-, or δ-tocopherol and singlet oxygen were 4.1 ×10⁶/M s, 3.3 × 10⁶/M s, and 1.4 × 10⁶/M s, respectively. The singlet oxygen oxidation rate of δ-tocopherol was significantly lower than α- or γ-tocopherol at α= 0.05. As the electron density in the chromanol ring of tocopherol increased, the singlet oxygen oxidation was increased.     

Microbiological quality of retail cheeses made from raw, thermized or pasteurized milk in the UK

Two studies of retail fresh, ripened and semi-hard cheeses made from raw, thermized or pasteurized milk were undertaken in the UK during 2004 and 2005 to determine the microbiological quality of these products. Using microbiological criteria in European Commission Recommendations 2004/24/EC and 2005/175/EC, 2% of both raw, thermized (37/1819 samples) and pasteurized (51/2618 samples) milk cheeses were of unsatisfactory quality. Raw or thermized milk cheeses were of unsatisfactory quality due to levels of Staphylococcus aureus at 10(4)cfu g(-1), Escherichia coli at 10(5)cfu g(-1), and/or Listeria monocytogenes at 10(2)cfu g(-1), whereas pasteurized milk cheeses were of unsatisfactory quality due to S. aureus at 10(3)cfu g(-1) and/or E. coli at 10(3)cfu g(-1). Salmonella was not detected in any samples. Cheeses were of unsatisfactory quality more frequently when sampled from premises rated as having little or no confidence in management and control systems, and stored/displayed at above 8 degrees C. Raw or thermized milk cheeses were also more likely to be of unsatisfactory quality when they were unripened types, and pasteurized milk cheeses when they were: semi-hard types; from specialist cheese shops or delicatessens; cut to order. These results emphasize the need for applying and maintaining good hygiene practices throughout the food chain to prevent contamination and/or bacterial growth. Labelling of cheeses with clear information on whether the cheese was prepared from raw milk also requires improvement.     

Influence of pre-treatments on microbial load of stored dehydrated onion slices

Onions slices were pre-treated in potassium meta-bisulphite (KMS) and sodium chloride with different concentration levels to study the microbial load of tray and greenhouse-dried onion slices up to 6 months of storage. Data were analysed as per procedure of one-way classified anova using DMRT of AgRes statistical package for bacteria, yeast, fungi and Lactobacilli . Results revealed that in almost all samples, permissible levels of bacteria [18.33 × 10¹ colony-forming units (CFU) per gram], yeast (ND), fungi (0.5 × 10¹ CFU g⁻¹) and Lactobacilli (0.25 × 10¹ CFU g⁻¹) were observed after 5 months of storage. All the samples were also found to be free from Escherichia coli and no Mac Conkey growth was noticed. Onion slices pre-treated in 0.25% and 0.50% KMS and dried in tray and greenhouse, respectively, were found best after 6 months of storage period.     

Enhancement of Fermentation Process in Pu-Erh Tea by Tea-Leaf Extract

ABSTRACT:  Pu-erh tea is known as a fermented tea and longer storage enhances its flavor and taste. Recently,  Aspergillu s,  Blastobotrys , and  Streptomyces  are found to play important roles in nutritional enhancement of Pu-erh tea by fermentation. Since water and temperature affect the microbial growth, we therefore explored the factors that might enhance the Pu-erh tea fermentation. The results showed that the addition of fresh tea-leaf extract (TLE) enhanced the withered tea fermentation (at 37 °C, 80 to 85% RH) as compared with the water only. Contents of statin, GABA, gallic acid, DPPH scavenging and polyphenol oxidase (PPO) activities were increased, whereas polyphenols and caffeine were decreased over 6 mo. TLE dose-dependently enhanced some of the qualities (that is, statin, PPO) of Pu-erh tea significantly as compared with the water only. The effect was related to the increase population of  A. niger  and  A. carbonarius  at 6 mo (from 7.6 ± 1.2 × 10¹ and 3.2 ± 1.3 × 10¹ to 3.1 ± 1.2 × 10⁶ and 2.4 ± 1.1 × 10⁵ colony forming units [CFU]/g, respectively). After drying process (90 °C, 30 min), the total microbial count from these samples returned to background level (3 ± 0.5 × 10² CFU/g). None of ochratoxin and fumonisin, toxins from  Aspergillus , was detected in the final products. The flavor and taste were also enhanced by treatment with TLE. The inoculation with  S. cinereus  Y11 with 2% TLE further enhanced these functional contents (about 2-fold increase of statin level) in the experimental Pu-erh tea. Therefore, this result may add a new process for Pu-erh tea manufacture.