Contamination of pasteurized milk by Bacillus cereus in the filling machine

The contamination of pasteurized milk by Bacillus cereus during the filling process was studied in two dairy plants. Samples of pasteurized milk were taken at four different sites along the production line. The samples were stored at 7 degrees C for 7 d, or at 10 degrees C for 5 d, before plate counting and random selection of B. cereus isolates. Isolates of B. cereus were typed by the polymerase chain reaction (PCR)-based method randomly amplified polymorphic DNA (RAPD). Samples taken at three different sites between the pasteurizer and the filling machine were all holding similar low concentrations of B. cereus, while an increase of the B. cereus count was seen in the consumer packages. More B. cereus of different RAPD types was growing in the consumer packages than in samples taken just before the filling machine. Several RAPD types found in the consumer packages were not detected in the samples taken just before the filling machine.     

A survey of the microflora of raw and pasteurized milk and the sources of contamination in a milk processing plant in Addis Ababa, Ethiopia

The microorganisms present in raw and pasteurized milk and the sources of contamination in the milk after it had arrived at the processing plant in Addis Ababa were studied. The lowest count registered for raw milk samples was 4 X 10(7) cfu/ml while the highest was 1 X 10(9) cfu/ml. Pasteurized milk had mesophilic aerobic counts of 7 X 10(5) cfu/ml as it left the pasteurizing unit, but the population increased 2- to 4-fold as a result of subsequent contamination. Of the total counts in raw milk, psychrophilic, thermoduric and thermophilic organisms made up 98.1, 1.4 and 0.5% respectively. In pasteurized milk, the amounts were 53.0, 39.5 and 7.5% respectively. Samples of milk pasteurized in the laboratory contained only 74.5% thermoduric and 25.5% thermophilic organisms. The isolates mostly belonged to the genera Bacillus, Streptococcus, Lactobacillus, Arthrobacter, Alcaligenes, Aeromonas and Pseudomonas. Cocci were more predominant than rod-shaped bacteria. Of the rod-shaped bacteria, 73% were Gram-negative. The utensils holding the raw and pasteurized milk and the plastic sheets used for bagging the pasteurized milk contributed unusually high numbers of bacteria which were either thermoduric or thermophilic. More isolates were obtained from the pasteurized than the raw milk. The keeping quality of the pasteurized milk was found to be much lower than that of the laboratory-pasteurized milk.     

Occurrence of aflatoxins M(1) and M(2) in milk commercialized in Ribeir?o Preto-SP, Brazil.

Aflatoxins are toxic metabolites found in foods and feeds. When ruminants eat foodstuffs containing aflatoxins B(1) and B(2), these toxins are metabolized and excreted as aflatoxin M(1) and M(2) in milk. The aim was to determine the incidence of these aflatoxins in commercial milk collected from supermarkets in Ribeir?o Preto-SP, Brazil, and consisting of 60 ultrahigh temperature (UHT) milk samples and 79 pasteurized milk samples. The milk samples were analysed according to method 986.16 of AOAC International. None of the milk samples analysed were contaminated with aflatoxin M(2), and aflatoxin M(1) was detected in 29 (20.9%) of samples in the range 50-240 ng l(-1). The results show that despite a high occurrence of aflatoxin M(1) in commercial pasteurized and UHT milk sold in Ribeir?o Preto in 1999 and 2000, the contamination level of these toxins could not be considered a serious public health problem according to MERCOSUR Technical Regulations. However, levels in 20.9% of the milk samples exceeded the concentration of 50 ng l(-1) permitted by the European Union. Although it is not necessary to continue monitoring the incidence and levels of aflatoxins M(1) and M(2) in milk samples, surveillance could be appropriate.     

Use of human lysozyme transgenic goat milk in cheese making: effects on lactic acid bacteria performance

Genetically engineered goats expressing elevated levels of the antimicrobial enzyme lysozyme in their milk were developed to improve udder health, product shelf life, and consumer well-being. The purpose of this study was to evaluate the effect of lysozyme on the development of lactic acid bacteria (LAB) throughout the cheese-making process. Raw and pasteurized milk from 7 lysozyme transgenic goats and 7 breed-, age-, and parity-matched nontransgenic controls was transformed into cheeses by using industry methods, and their microbiological load was evaluated. The numbers of colony-forming units of LAB were determined for raw and pasteurized goat milk, whey, and curd at d 2 and at d 6 or 7 of production. Selective plating media were used to enumerate lactococcal species separately from total LAB. Although differences in the mean number of colony-forming units between transgenic and control samples in raw milk, whey, and cheese curd were non-significant for both total LAB and lactococcal species from d 2 of production, a significant decrease was observed in both types of LAB among d 6 transgenic raw milk cheese samples. In pasteurized milk trials, a significant decrease in LAB was observed only in the raw milk of transgenic animals. These results indicate that lysozyme transgenic goat milk is not detrimental to LAB growth during the cheese-making process.     

Occurrence of aflatoxin M1 in raw and market milk commercialized in Greece

From December 1999 to May 2000, 114 samples of pasteurized, ultrahigh temperature-treated (UHT) and concentrated milk were collected in supermarkets, whereas 52 raw milk samples from cow, sheep and goat were obtained from different milk producers all over Greece. Sample collection was repeated from December 2000 to May 2001 and concerned 54 samples of pasteurized milk, 23 samples of bulk-tank raw milk and 55 raw milk samples from cow, sheep and goat. The total number of samples analysed for aflatoxin M₁ (AFM₁) contamination by immunoaffinity column extraction and liquid chromatography was 297. In the first sampling, the incidence rates of AFM 1 contamination in pasteurized, UHT, concentrated and cow, sheep and goat raw milk were 85.4, 82.3, 93.3, 73.3, 66.7 and 40%, respectively, with only one cow raw milk and two concentrated milk samples exceeding the EU limit of 50 ng l^-1. In the second sampling, the incidence rates of AFM 1 contamination in pasteurized, bulk-tank and cow, sheep and goat raw milk were 79.6, 78.3, 64.3, 73.3 and 66.7%, respectively, with only one cow and one sheep raw milk samples exceeding the limit of 50 ng l^-1. The results suggest that the current regulatory status in Greece is effective.     

Drug resistance and lecithinase activity of Yersinia enterocolitica isolated from buffalo milk

Two-hundred-and-seven samples of raw buffalo milk and 60 samples of pasteurized buffalo milk were screened for presence of Yersinia enterocolitica. The prevalence of Y. enterocolitica was found to be 24.1% in raw milk, however, no isolation could be made from the pasteurized milk samples. Cold enrichment in trypticase soy broth and alkali treatment methods were followed in this study. The majority of the isolates (62%) were found sensitive to all the antibiotics used and only a few (16%) were resistant to two or more than two antibiotics. The incidence of Y. enterocolitica showed seasonal variations. Incidence was much higher (25-50%) during the winter season as compared to the summer (0-17%). The incidence of lecithinase production was high (40-50%) in Yersinia isolates resistant to one or two antibiotics.     

Occurrence of aflatoxin M1 in Korean dairy products determined by ELISA and HPLC

The occurrence of aflatoxin M₁ (AFM₁)in pasteurized milk and dairy products was investigated by using direct competitive enzyme-linked immunosorbent assay (ELISA) and high-performance liquid chromatography (HPLC). The recoveries of AFM₁ from the samples spiked at levels between 5 and 500 pg/ml were 88.0-106.5% for pasteurized milk and 84.0-94.0% for yoghurt by ELISA. By HPLC, the recoveries were 103-120% for pasteurized milk and 87.0-93.0% for yoghurt. The limits of detection were found to be 2 pg/ml by ELISA and 10 pg/ml by HPLC. Among a total of 180 samples collected in Seoul, Korea, the incidence of AFM₁ in pasteurized milk, infant formula, powdered milk and yoghurt was 76, 85, 75, and 83% , respectively, with a mean concentration of 18, 46, 200, and 29 pg/g, respectively, when determined by ELISA. These results obtained by ELISA were closely related to those by HPLC for AFM₁ (r² = 0.9783).     

Occurrence of aflatoxins in peanuts, milk, and animal feed in Trinidad

The prevalence of aflatoxin B1 (AFB1) in 186 peanut products (140 peanuts, 32 peanut butter, and 14 nut cakes) from supermarkets, road vendors, and sale outlets, and 40 feed samples from dairy farmers was determined using the radioimmunoassay method (Charm II) test for aflatoxins. The frequency of aflatoxin M1 (AFM1) was also determined in 175 raw milk samples from milk collection centers and 37 pasteurized milk samples obtained from supermarkets and sale outlets. Overall, from a total of 438 samples tested, 18 (4.1%) were positive for aflatoxin comprising 5 (2.2%) of 226 peanut products and feeds positive for AFB1, and 13 (6.1%) of 212 milk samples positive for AFM1. All 186 peanuts and peanut products were negative (0.0%) for AFB1 while 5 (7.4%) of 40 dairy feed samples were positive. Of the 175 raw milk samples tested, 13 (7.4%) were contaminated with AFM1 while all pasteurized milk samples were negative. The detection of AFB1 in feed and AFM1 in milk is of public health importance considering the practice of raw milk consumption by the farmers and their families in the country.     

PREVALENCE OF ANTIMICROBIAL RESIDUES IN PREPROCESSED AND PROCESSED COWS' MILK IN TRINIDAD

The prevalences of antimicrobial residues were determined in cows' bulk raw milk from 16 collection centers, in pasteurized milk obtained from sale outlets, and in raw, pasteurized and sterilized milk in the processing line of a plant in Trinidad. The Delvotest SP 5 Pack test kits were used to detect the antimicrobial residues in milk. Thirty-one (10.3%) of 300 bulk milk samples contained antimicrobial residues with penicillin responsible for 90.3% of all the positive samples which originated from 8 (50.0%) of the 16 collection centers. The prevalence of residues in pasteurized milk was 21.3% compared to 8.3% detected in sterilized milk. The difference was not statistically significant (P ± 0.05; X²). Pasteurized milk obtained from sale outlets but originating from two other processing plants had antimicrobial residue prevalence of 0% and 8.3%, respectively. It is concluded that the presence of antimicrobial residues in raw and processed milk in Trinidad could be of public health and economic significance .     

电阻抗法快速测定食品中菌落总数的应用研究

为更好地提高现行食品中的菌落总数的检测效率,快速、准确地对食品中的菌落总数出评估,将电阻抗法应用到120份巴氏消毒奶和114份瓶装饮用纯净水中的菌落总数的测定,并与现行国标法测定结果作比较。二种方法检测结果经统计学处理均无显性差异。巴氏消毒奶和饮用纯净水的检测结果相符率分别为95%和94.7%,电阻抗法巴氏消毒奶的假阳性率为1.7%,假阴性率3.3%,纯净水假阳性率为1.8%,假阴性率为3.6%。对检出时间的分析表明,巴氏消毒奶的细菌指标合格的时间为4.2h,不合格时间为2.3h,纯净水的细菌指标合格的时间为14.5h,不合格时间为11.3h。以上结果表明,应用电阻抗技术测定巴氏消毒奶和纯净水的落总数,可以将检测时间从常规法的48h缩短至14.5h,是一种省时、可靠的检测方法。     

Chemiluminescent determination of xanthine oxidase activity in milk.

A chemiluminescent method for determining xanthine oxidase (XOD) activity was developed and applied to the assay of milk enzyme activity using a photomultiplier luminometer. Various kinds of milk and cream samples were analysed for XOD content. In pasteurized milk, XOD activity depended on the fat content and in UHT milk it disappeared owing to the heat treatment. Milk sample preparation was very simple, requiring only homogenization at 40 degrees C followed by a 1:10 dilution with UHT ('XOD-free') milk. The assay was carried out at 25 degrees C. The response obtained from XOD standard solutions in milk was linear from 0.1 to 500 enzyme units (U) l-1, but for the actual milk samples values ranged only from 1 to 135 U l-1. The detection limit at 2 SD was 0.1 U l-1 in milk, while in buffer it was 100 times lower. The intra-assay and interassay CV for XOD activity in milk were 6-12%.     

Thermal inactivation of foot-and-mouth disease virus in milk using high-temperature, short-time pasteurization

Previous studies of laboratory simulation of high temperature, short time pasteurization (HTST) to eliminate foot-and-mouth disease virus (FMDV) in milk have shown that the virus is not completely inactivated at the legal pasteurization minimum (71.7°C/15 s) but is inactivated in a flow apparatus at 148°C with holding times of 2 to 3 s. It was the intent of this study to determine whether HTST pasteurization conducted in a continuous-flow pasteurizer that simulates commercial operation would enhance FMDV inactivation in milk. Cows were inoculated in the mammary gland with the field strain of FMDV (01/UK). Infected raw whole milk and 2% milk were then pasteurized using an Arm-field pilot-scale, continuous-flow HTST pasteurizer equipped with a plate-and-frame heat exchanger and a holding tube. The milk samples, containing FMDV at levels of up to 10⁴ plaque-forming units/mL, were pasteurized at temperatures ranging from 72 to 95°C at holding times of either 18.6 or 36 s. Pasteurization decreased virus infectivity by 4 log₁₀ to undetectable levels in tissue culture. However, residual infectivity was still detectable for selected pasteurized milk samples, as shown by intramuscular and intradermal inoculation of milk into naïve steers. Although HTST pasteurization did not completely inactivate viral infectivity in whole and 2% milk, possibly because a fraction of the virus was protected by the milk fat and the casein proteins, it greatly reduced the risk of natural transmission of FMDV by milk.     

Genetic diversity of thermoduric spoilage microorganisms of milk from Brazilian dairy farms

When correctly pasteurized, packaged, and stored, milk with low total bacterial counts (TBC) has a longer shelf life. Therefore, microorganisms that resist heat treatments are especially important in the deterioration of pasteurized milk and in its shelf life. The aim of this work was to quantify the thermoduric microorganisms after the pasteurization of refrigerated raw milk samples with low TBC and to identify the diversity of these isolates with proteolytic or lipolytic potential by RFLP analysis. Twenty samples of raw milk were collected in bulk milk tanks shortly after milking in different Brazilian dairy farms and pasteurized. The mean thermoduric count was 3.2 (±4.7) × 10² cfu/mL (2.1% of the TBC). Of the 310 colonies obtained, 44.2% showed milk spoilage potential, 32.6% were proteolytic and lipolytic simultaneously, 31% were exclusively proteolytic, and 48 (36.4%) were only lipolytic. Regarding the diversity, 8 genera were observed (Bacillus, Brachybacterium, Enterococcus, Streptococcus, Micrococcus, Kocuria, Paenibacillus, and Macrococcus); there was a predominance of endospore-forming bacteria (50%), and Bacillus licheniformis was the most common (34.1%) species. Considering the RFLP types, it was observed that the possible clonal populations make up the microbiota of different milk samples, but the same milk samples contain microorganisms of a single species with different RFLP types. Thus, even in milk with a high microbiological quality, it is necessary to control the potential milk-deteriorating thermoduric microorganisms to avoid the risk of compromising the shelf life and technological potential of pasteurized milk.     

Effect of phage on survival of Salmonella enteritidis during manufacture and storage of cheddar cheese made from raw and pasteurized milk.

The ability of Salmonella Enteritidis to survive in the presence of phage, SJ2, during manufacture, ripening, and storage of Cheddar cheese produced from raw and pasteurized milk was investigated. Raw milk and pasteurized milk were inoculated to contain 10(4) CFU/ml of a luminescent strain of Salmonella Enteritidis (lux) and 10(8) PFU/ml SJ2 phage. The milks were processed into Cheddar cheese following standard procedures. Cheese samples were examined for Salmonella Enteritidis (lux), lactic acid bacteria, molds and yeasts, coliforms, and total counts, while moisture, fat, salt, and pH values were also measured. Salmonella Enteritidis (lux) was enumerated in duplicate samples by surface plating on MacConkey novobiocin agar. Bioluminescent colonies of Salmonella Enteritidis were identified in the NightOwl molecular imager. Samples were taken over a period of 99 days. Counts of Salmonella Enteritidis (lux) decreased by 1 to 2 log cycles in raw and pasteurized milk cheeses made from milk containing phage. In cheeses made from milks to which phage was not added, there was an increase in Salmonella counts of about 1 log cycle. Lower counts of Salmonella Enteritidis (lux) were observed after 24 h in pasteurized milk cheese containing phage compared to Salmonella counts in raw milk cheese with phage. Salmonella Enteritidis (lux) survived in raw milk and pasteurized milk cheese without phage, reaching a final concentration of 10(3) CFU/g after 99 days of storage at 8 degrees C. Salmonella did not survive in pasteurized milk cheese after 89 days in the presence of phage. However, Salmonella counts of approximately 50 CFU/g were observed in raw milk cheese containing phage even after 99 days of storage. In conclusion, this study demonstrates that the addition of phage may be a useful adjunct to reduce the ability of Salmonella to survive in Cheddar cheese made from both raw and pasteurized milk.     

Calibration of infrared milk analyzers: modified milk versus producer milk

Mid-infrared (MIR) milk analyzers are traditionally calibrated using sets of preserved raw individual producer milk samples. The goal of this study was to determine if the use of sets of preserved pasteurized modified milks improved calibration performance of MIR milk analyzers compared with calibration sets of producer milks. The preserved pasteurized modified milk sets exhibited more consistent day-to-day and set-to-set calibration slope and intercept values for all components compared with the preserved raw producer milk calibration sets. Pasteurized modified milk calibration samples achieved smaller confidence interval (CI) around the regression line (i.e., calibration uncertainty). Use of modified milk calibration sets with a larger component range, more even distribution of component concentrations within the ranges, and the lower correlation of fat and protein concentrations than producer milk calibration sets produced a smaller 95% CI for the regression line due to the elimination of moderate and high leverage samples. The CI for the producer calibration sets were about 2 to 12 times greater than the CI for the modified milk calibration sets, depending on the component. Modified milk calibration samples have the potential to produce MIR milk analyzer calibrations that will perform better in validation checks than producer milk-based calibrations by reducing the mean difference and standard deviation of the difference between instrument values and reference chemistry.     

Quantum dot bead-based competitive immunochromatographic assay for enterotoxin aureus A detection in pasteurized milk

Staphylococcal enterotoxin A (SEA) is an important biotoxin, produced by Staphylococcus aureus under appropriate conditions, and often contaminates milk and dairy products. Herein, an anti-SEA monoclonal antibody (anti-SEA mAb) was prepared by injecting the SEA protein in BALB/c mice, and a novel immunochromatographic assay (ICA) was developed for the rapid and sensitive determination of SEA in pasteurized milk by using highly luminescent quantum dot beads (QB) as signal amplification probe. Given the 1020-fold enhancement in the photoluminescence intensity of QB to the original quantum dot, the proposed QB-ICA exhibits high sensitivity for SEA determination in real milk samples with a limit of detection of 1.89 ng/mL, and shows good dynamic linearity for SEA quantitative detection from 2 to 150 ng/mL within 15 min of test time. The proposed QB-ICA also shows good selectivity to SEA detection with a negligible cross-reaction to common analogs, including staphylococcal enterotoxins B, C, D, and E. In addition, the accuracy and precision of QB-ICA were assessed by analyzing SEA-fortified milk samples. The average recoveries of intra- and interassays range from 85.5 to 128.1%, and the coefficients of variation range from 4.6 to 14.2%, indicating an acceptable accuracy for the quantitative detection of SEA in real milk samples. In summary, this work provides a powerful and rapid analysis tool for the sensitive monitoring of SEA contamination in pasteurized milk samples.     

The effect of extended low-temperature storage of raw milk on the quality of pasteurized and UHT milk

The effect of storage of raw milk at 2 and 6°C on the quality of pasteurized and UHT milks produced from them has been investigated. There was no difference in shelf-lives of pasteurized milks produced from raw milks which had no obvious physical defects, odours and taints after storage at 2 and 6°C. This was true for pasteurized milks in the presence and absence of post-heat-treatment contamination. However, pasteurized milks of good quality could be produced from more than 80% of raw milk samples which had been stored for up to 5 days at 2°C, but this was only possible with raw milks which had been stored at 6°C for 2 days.Failure rates of experimentally-produced UHT milks were much higher in products manufactured from raw milks stored at 6°C for 4 days than those produced from raw milks stored at 2°C for 4 days. The main cause of failure was due to thermostable bacterial protease associated with high levels of bacterial growth in the raw milks. Other causes of failure included spore-forming bacteria, which may have survived UHT processing, and other organisms probably introduced as contaminants on filling.     

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.     

Determination of melamine in milk and dairy products by high performance liquid chromatography

A simple, precise, accurate, and validated reverse-phase HPLC method was developed for the determination of melamine in milk (pasteurized and UHT milk) and dairy products (powdered infant formula, fruit yogurt, soft cheese, and milk powder). Following extraction with acetonitrile:water (50:50, vol/vol), samples were purified by filter (0.45 μm), separated on a Nucleosil C8 column (4.6 mm × 250 mm, 3 μm) with acetonitrile:10 mmol/L sodium L-octane sulfonate (pH 3.1; 15:85, vol/vol) as mobile phase at a flow rate of 1 mL/min, and determined by a photodiode array detector. A linear calibration curve was obtained in the concentration range from 0.05 to 5 mg/kg. Milk and dairy products were fortified with melamine at 4 levels producing average recovery yields of 95 to 109%. The limits of detection and quantification of melamine were 35 to 110 and 105 to 340 μg/kg, respectively. The method was then used to analyze 300 samples of milk and dairy products purchased from major retailers in Turkey. Melamine was not found in infant formulas and pasteurized UHT milk, whereas 2% of cheese, 8% of milk powder, and 44% of yogurt samples contained melamine at the 121, 694±146, and 294±98 μg/kg levels, respectively. These findings were below the limits set by the Codex Alimentarius Commission and European Union legislation. This is the first study to confirm the existence of melamine in milk and dairy products in Turkey. Consumption of foods containing these low levels of melamine does not constitute a health risk for consumers.     

Post-pasteurization contamination and shelf-life of HTST-pasteurized milk when filled in a Liqui-Pak® conventional or Model 820A cartoning machine

Bacterial contamination of pasteurized milk filled in a Liqui-Pak ® Model 820A cartoning machine has been examined. Compared with the conventional piston type, this new machine incorporates a number of design features to reduce or eliminate contamination during filling. The Model 820A did not normally cause post-pasteurization contamination with psychrotrophic Gram-negative rods, the typical spoilage organisms of conventionally filled pasteurized milk. This led to a considerable improvement in the keeping quality of the filled pasteurized milk, especially at a storage temperature of 7® C.