Comparative Effects of Chlortetracycline,Freezing, and γ Radiation on Microbial Population of Ocean Perch

The microbial populations in chlortetracycline (CTC)-treated (50, 100, 200, and 500 ppm), frozen (-15 C), and irradiated (0.1 Mrad) ocean perch (Sebastodes alutus) were compared. The control sample spoiled at 7 C, primarily because of the growth of Pseudomonas. Irradiation changed this to Achromobacter-dominated spoilage. Freezing or CTC treatment altered the spoilage pattern very little. CTC was particularly effective against ultraviolet fluorescent Pseudomonas species at the higher concentrations. Freezing and CTC were not effective against “coryneforms.”     

A selective medium for the rapid detection by an impedance technique of Pseudomonasspp. associated with poultry meat

A new medium for detecting and enumerating Pseudomonas spp. associated withpoultry meat spoilage by a rapid impedance technique was developed, after testing potentialgrowth promoters for eight Pseudomonas strains and inhibitors against eight competingstrains (Enterobacteriaceae) able to grow on the medium of Mead and Adams (1977). Four basalmedia (brain heart infusion, brucella broth, Shaedler broth and Whitley impedance broth (WIB))and a synthetic medium were evaluated. Whitley impedance broth was the best basal medium fordetecting variations in impedance in relation to Pseudomonas growth. The efficiency ofWIB was improved by adding compounds which enhanced the growth of Pseudomonas onthe synthetic medium. Among the incubation temperatures tested, 22°C proved to be the bestcompromise between growth of Pseudomonas associated with poultry meat spoilage andinhibition of competitors. Among the 15 inhibitory substances evaluated against Pseudomonas competitors, five were chosen for inclusion in the final medium : metronidazole,carbenicilline, cetrimide, cycloheximide and diamide (MCCCD medium). Preliminary resultsobtained from experiments with beef and pork meat showed that this medium could also be usedwithout diamide and at an incubation temperature of 25°C. The impedance technique usingMCCCD medium was then compared with an official method which uses the medium of Meadand Adams (1977) on 106 samples of poultry neck skin. The linear regression coefficient betweenthe two techniques was approximately r = 0·85. Impedance was able to detect 10³ Pseudomonas g⁻¹ within less than 19 h making it a promisingtechnique for predicting poultry meat spoilage.     

Changes in the Microflora of Irradiated Petrale Sole (Eopsetta jordani) Fillets Stored Aerobically at 0.5 C

The microfloral changes on irradiated petrale sole fillets during aerobic (packaged with oxygen-permeable film), refrigerated storage were determined by the identification of bacterial and yeast isolates to the generic level. The samples were irradiated at 0.0, 0.1, 0.15, 0.2, 0.3, and 0.4 Mrad by use of a cobalt-60 gamma source, were stored at 0.5 C, and were examined periodically for spoilage, total microbial population, and composition. The preirradiation flora of the fresh fillets consisted of coryneforms, Achromobacter, Micrococcus, Flavobacterium, Pseudomonas, and Lactobacillus. Immediately after irradiation, Micrococcus, Achromobacter, coryneforms, and Bacillus were predominant. The flora of the nonirradiated fillets at the time of spoilage consisted of Pseudomonas and Achromobacter. The flora of the irradiated fillets at the time of spoilage consisted of Achromobacter and Trichosporon.     

Antimicrobial activity of Myrtus communis L. water-ethanol extract against meat spoilage strains of Brochothrix thermosphacta and Pseudomonas fragi in vitro and in meat

The antimicrobial activity of hydro-alcoholic extract of Myrtus communis L. was investigate in vitro and in situ against different meat spoilage biotypes of Pseudomonas fragi and Brochothrix thermosphacta. Water-ethanol myrtle extract (WEME) was prepared from myrtle leaves and used to study the antimicrobial activity, Minimum Inhibitory Concentration (MIC) and Minimum Lethal Concentration (MLC). Naturally contaminated ground beef was used to evaluate in situ antibacterial activity of freeze-dried myrtle extract at different concentrations. In vitro antimicrobial activity of WEME was significantly more effective against B. thermosphacta than P. fragi strains. MIC and MLC values were in the range of 12.5–50 and 25–100 mg DM/ml, respectively, for B. thermosphacta and P. fragi strains. In situ results showed that the microbial population, except for Pseudomonas spp., decreased significantly in ground meat with added 5 % of freeze-dried myrtle extract. In conclusion, the findings demonstrate the effectiveness of myrtle extract to control or prevent the proliferation of meat spoilage bacteria.     

Selection of Microbial Population in Chlortetracycline-treated Ocean Perch (Sebastodes alutus)

Microorganisms isolated from chlortetracycline (CTC)-treated ocean perch were individually tested to determine their relative sensitivity to CTC by use of CTC-impregnated paper discs. The CTC sensitivity varied among isolates obtained from untreated fish. The proportion of CTC-resistant species increased with higher CTC concentration and the length of storage at 7 C. Among individual generic groups, "coryneforms" and yeasts were more resistant to CTC than other groups. Pseudomonas, Achromobacter, Flavobacterium, Bacillus, and Lactobacillus all contained species either resistant or sensitive to CTC. More CTC-resistant species in these genera accumulated with the increased CTC concentrations and with the length of storage in the presence of CTC.     

Isolation and characterization of Pseudomonas from processed chicken in Jamaica

Inflation and changes in the poultry industry in Jamaica have necessitated an increased understanding of spoilage flora and the examination of methods for increasing the shelf life of chilled eviscerated poultry. The efficacy of five antibiotics was determined. Despite the widespread use of neooxytetracycline in the treatment of poultry infections, tetracycline concentrations of < 10 μg ml^-1 were found to be highly effective against isolates of the main spoilage organism Pseudomonas.     

Classification of the spoilage flora of raw and pasteurized bovine milk, with special reference to Pseudomonas and Bacillus

Eighty-one bacterial strains isolated from refrigerated raw milk, 124 from pasteurized milk and cream stored at 5°C and 7°C, and 19 type and reference strains of Pseudomonas spp. and Bacillus spp. were characterized by numerical phenotypic analysis. Data were processed with simple matching ( S _SM) and Jaccard ( S _J) coefficients, and UPGMA clustering. Fourteen clusters of Gram-negative bacteria were formed at S _J= 79% ( S _SM= 90%). Raw milk was exclusively spoilt by Gram-negative bacteria, the majority of which were Pseudomonas fluorescens biovar I, Ps. fragi, Ps. lundensis and Ps. fluorescens biovar III. Minor groups in raw milk included Enterobacteriaceae spp. and Acinetobacter spp. Pasteurized milk was spoilt by essentially the same Gram-negative organisms in 65% (5°C) and 50% (7°C) of the cases. The phenotypic characteristics of Gram-negative bacteria are given. Bacillus polymyxa (both temperatures) and B. cereus (only at 7°C) were responsible for 77% of samples spoiled by the Gram-positive organisms. Minor milk spoilage groups included other Bacillus spp. and lactic acid bacteria. All Bacillus spp. grew fermentatively in milk, and most strains denitrified. It is suggested that: (i) industrial recontamination tests of pasteurized milk are directed against Pseudomonas; (ii) milk is stored at 5°C or lower to avoid growth of B. cereus ; and (iii) the significance of gas-producing and nitrate/nitrite-reducing Bacillus strains is recognized in cheese production.     

Changes in the spoilage-related microbiota of beef during refrigerated storage under different packaging conditions

The microbial spoilage of beef was monitored during storage at 5 degrees C under three different conditions of modified-atmosphere packaging (MAP): (i) air (MAP1), (ii) 60% O2 and 40% CO2 (MAP2), and (iii) 20% O2 and 40% CO2 (MAP3). Pseudomonas, Enterobacteriaceae, Brochothrix thermosphacta, and lactic acid bacteria were monitored by viable counts and PCR-denaturing gradient gel electrophoresis (DGGE) analysis during 14 days of storage. Moreover, headspace gas composition, weight loss, and beef color change were also determined at each sampling time. Overall, MAP2 was shown to have the best protective effect, keeping the microbial loads and color change to acceptable levels in the first 7 days of refrigerated storage. The microbial colonies from the plate counts of each microbial group were identified by PCR-DGGE of the variable V6-V8 region of the 16S rRNA gene. Thirteen different genera and at least 17 different species were identified after sequencing of DGGE fragments that showed a wide diversity of spoilage-related bacteria taking turns during beef storage in the function of the packaging conditions. The countable species for each spoilage-related microbial group were different according to packaging conditions and times of storage. In fact, the DGGE profiles displayed significant changes during time and depending on the initial atmosphere used. The spoilage occurred between 7 and 14 days of storage, and the microbial species found in the spoiled meat varied according to the packaging conditions. Rahnella aquatilis, Rahnella spp., Pseudomonas spp., and Carnobacterium divergens were identified as acting during beef storage in air (MAP1). Pseudomonas spp. and Lactobacillus sakei were found in beef stored under MAP conditions with high oxygen content (MAP2), while Rahnella spp. and L. sakei were the main species found during storage using MAP3. The identification of the spoilage-related microbiota by molecular methods can help in the effective establishment of storage conditions for fresh meat.     

Effect of temperature on the microbial flora of herring fillets stored in air or carbon dioxide

The microbial development on fillets of herring from the Baltic Sea was studied at temperatures from 0-15 C in air or 100% carbon dioxide (96-100% CO2). The shelf-life of the fillets , defined as the time for the 'total aerobic count' to reach 1 X 10(7) c.f.u./g increased with decreased temperature from 1 d at 15 degrees C to 7 d at 0 degrees C (air). The corresponding values in CO2 were 3 d and 33 d, respectively. The initial flora of the herring fillets was dominated by Alteromonas putrefaciens and Pseudomonas spp. and so was the spoilage flora after storage in air (together 62-95% of the flora: all temperatures). Alteromonas putrefaciens predominated slightly at 2 degrees C to 15 degrees C, while Pseudomonas spp. dominated at 0 degrees C. The Pseudomonas flora was mainly split between Ps. fragi , Ps. fluorescens and a heterogenous group of unidentified Pseudomonas spp. The proportions were not influenced by temperature. In 100% CO2 at the time of spoilage the flora consisted of a significant number of Lactobacillus spp. Below 4 C the domination was almost complete while at 10 degrees C and 15 degrees C. Enterobacteriaceae, Vibrionaceae and Alt. putrefaciens was also found. It was concluded that the microbiological shelf-life of herring fillets is improved by refrigeration storage in 100% CO2 but for good results the temperature should not exceed 2 degrees C.     

Enumeration of respiring Pseudomonas spp. in milk within 6 hours by fluorescence in situ hybridization following formazan reduction

Respiring Pseudomonas spp. in milk were quantified within 6 h by fluorescence in situ hybridization (FISH) with vital staining. FISH with an oligonucleotide probe based on 16S rRNA sequences was used for the specific detection of Pseudomonas spp. at the single cell level. 5-cyano-2,3-ditolyl tetrazolium chloride (CTC) was used to estimate bacterial respiratory activity. The numbers of respiring Pseudomonas cells as determined by FISH with CTC staining (CTC-FISH) were almost the same or higher than the numbers of CFU as determined by the conventional culture method.     

Cytotoxic activities of normal cultured human T cells.

Normal human T cells grown in continued cultures in medium containing conditioned medium (CM) from PHA-stimulated lymphocytes were studied for their ability to manifest three known forms of cell-mediated cytotoxicity: lectin-induced cellular cytotoxicity (LICC), natural killer cell (NK) activity, and antibody-dependent cellular cytotoxicity (ADCC). The cultured T cells (CTC) were very effective mediators of LICC, being cytotoxic even at very low attacker-target cell ratios in the presence of different lectins, and against different types of targets. When tested without the addition of lectin, the CTC demonstrated a low degree of spontaneous cytotoxicity. This spontaneous cytotoxicity might not be due to conventional NK cells however, since the CTC failed to show significant numbers of cells with Fc receptors (FcR) for IgG, and had no detectable ADCC activity. CTC could represent a population enriched in polyclonal activated T cells with low spontaneous cytotoxicity against a variety of allogeneic target cells, which is greatly enhanced by the addition of lectins dur ing the 51Cr release assay.     

碳源及温度对食源假单胞菌群体感应信号分子产生的影响

许多革兰氏阴性菌通过产生N-酰基-高丝氨酸内酯（AHLs）类信号分子来调控某些性状的表达,即群体感应（quorum sensing）。假单胞菌是一种导致食品腐败的重要腐败细菌,也产生AHLs。本文研究了不同温度及碳源对食源假单胞菌AHLs产生的影响。结果表明,该假单胞菌在25℃条件下,产生两种AHL信号分子,而在4℃条件下,所产生的短链AHL分子消失,主要产生长链AHL分子。而且在不同碳源（葡萄糖,果糖,木糖,麦芽糖等）的培养基中生长,所产生的AHLs分子种类也不同。同时发现当pH>7.5时,AHLs的稳定性下降。由此得出,在不同的环境条件(碳源及温度)下假单胞菌所产生的AHLs种类不同。为进一步研究群体感应现象在食品腐败中的作用以及开发基于干扰腐败菌群体感应的新型食品防腐技术提供研究基础。     

Isolation from food sources, of lactic acid bacteria that produced antimicrobials

The potential of lactic acid bacteria, isolated from a variety of foods, to inhibit indicators representative of spoilage and pathogenic bacteria associated with food products was examined. Fruit and vegetables were a poor source of lactic acid bacteria but large numbers were readily isolated on MRS agar from cheese, milk and meat samples. Approximately 1000 isolates from each of the food samples were examined by the deferred antagonism procedure to determine their ability to inhibit Staphylococcus aureus, Listeria innocua and Pseudomonas fragi. Listeria innocua was the bacterium predominantly inhibited by isolates from the cheese, milk and meats, but antagonism was also observed to a lesser extent against the other indicators. The only inhibition observed for isolates from vegetable material was directed against Staph. aureus. The majority of inhibitor producers were effective against only one of the indicators but a small number were isolated which inhibited two or three.     

玉米须提取物对食品腐败菌及致病菌抑制作用的研究

首次以玉米须提取物对7种常见的食品腐败菌及致病菌进行抑菌试验,发现玉米须的乙醇提取物的效果最好,其最低抑菌浓度（MIC）为3．0g/100g,此外,对食品加工条件（如杀菌方式等）及食品介质（如PH）对玉米须提取物抑菌活性的也作了研究,结果表明,玉米须提取物在常规食品杀菌条件（UHT）及中酸到酸性环境下抑菌活性稳定,因而可作为潜在的食品防腐剂。     

Management of post-harvest fruit spoilage fungi by some potential spice extracts

The experiment was carried out to evaluate the antifungal potential of aqueous and ethanolic extracts of four spices (Allium sativum, Zingiber officinale, Cinnamomum zeylanicum and Capsicum annuum) against the important post-harvest spoilage fungi isolated from diseased fruits. In total, 276 isolates of post-harvest spoilage fungi were isolated from four different fruit types (Persea americana, Musa acuminata, Citrus sinensis and Lycopersicon esculentum), out of which 183 isolates were identified while 93 isolates were remain unidentified. The most dominant post-harvest spoilage fungus was Rhizopus sp. (26.45%), followed by Penicillium sp. (19.93%), Aspergillus sp. (10.86%) and Fusarium sp. (9.06%). Results of disc diffusion assay showed that ethanolic extract of C. zeylanicum was found to be most effective against Penicillium sp., followed by aqueous extracts of A. sativum. The ethanolic extract of C. zeylanicum in agar amended assay and minimal inhibitory concentrations were found to be very efficient (100% inhibition) against all the tested fungi. Results of in vivo study showed that pre-inoculated C. zeylanicum ethanolic extract and A. sativum aqueous extract were found effective in reducing the disease severity (6.24–13.67%) and (7.91–13.15%) against the Penicillium, Rhizopus, Aspergillus and Fusarium sp.     

Disinfectant test against monoculture and mixed-culture biofilms composed of technological, spoilage and pathogenic bacteria: bactericidal effect of essential oil and hydrosol of Satureja thymbra and comparison with standard acid-base sanitizers

Aims: To assess the antimicrobial action of three natural‐derived products (essential oil, decoction and hydrosol of Satureja thymbra) against biofilms, composed of useful, spoilage and pathogenic bacteria (formed as monoculture or/and mixed‐culture), and to compare their efficiency with three standard acid and alkaline chemical disinfectants. Methods and Results: Two acids (hydrochloric and lactic, pH 3), one alkali (sodium hydroxide, pH 11), the essential oil of S. thymbra (1% v/v) and the two by‐products of the essential oil purification procedure (the decoction and the hydrosol fraction of essential oil, 100%), were tested against biofilms formed by five bacterial species, either as monospecies, or as mixed‐culture of all species. The tested bacterial species were Staphylococcus simulans and Lactobacillus fermentum (useful technological bacteria), Pseudomonas putida (spoilage bacterium), Salmonella enterica and Listeria monocytogenes (pathogenic bacteria). Biofilms were left to be formed on stainless steel coupons for 5 days at 16°C, before the application of disinfection treatments, for 60 and 180 min. The disinfection efficiency was evaluated by detaching the remaining viable biofilm cells and enumerating them by agar plating, as well as by automated conductance measurements (using Rapid Automated Bacterial Impedance Technique). Both these methods revealed that the essential oil and the hydrosol of S. thymbra exhibited a strong antimicrobial action against both monospecies and mixed‐culture biofilms. Surprisingly, the efficiency of the other three acid–base disinfectants was not adequate, although a long antimicrobial treatment was applied (180 min). Conclusions: The essential oil of S. thymbra (1%), as well as its hydrosol fraction (100%), presents sufficient bactericidal effect on bacterial biofilms formed on stainless steel. Significance and Impact of the Study: Use of natural antimicrobial agents could provide alternative or supplemented ways for the disinfection of microbial‐contaminated industrial surfaces.     

复合乳酸菌对冷藏海鲈鱼块的保鲜效果

【目的】研究复合乳酸菌对冷藏海鲈鱼块的保鲜效果。【方法】以冷藏海鲈鱼块为对象,筛选出3株能够明显抑制其优势腐败菌(草莓假单胞菌Pseudomonas fragi,腐败希瓦氏菌Shewanella putrefacens)生长的单一乳酸菌,同时也筛选出对其优势腐败菌具有最显著抑制效果的一组复合乳酸菌,再将该复合乳酸菌接种到海鲈鱼块上,在4°C冷藏过程中,通过感官评定、挥发性盐基氮(TVB-N值)的测定和优势腐败菌的计数来评价复合乳酸菌对冷藏海鲈鱼块的保鲜效果。【结果】单一乳酸菌(干酪乳杆菌LC1、植物乳杆菌LP1和乳酸菌L3)对2株冷藏海鲈鱼优势腐败菌的抑制效果明显;复合乳酸菌(干酪乳杆菌LC1+植物乳杆菌LP1+乳酸菌L3)的抑菌效果最为显著;在4°C冷藏过程中,复合乳酸菌能使冷藏海鲈鱼块发生感官变化延缓6 d、使TVB-N值的升高延缓2 d,同时显著抑制优势腐败菌的生长。【结论】复合乳酸菌对冷藏海鲈鱼块具有良好的保鲜作用,能有效延长其货架期。     

Seasonal influence on heat-resistant proteolytic capacity of Pseudomonas lundensis and Pseudomonas fragi, predominant milk spoilers isolated from Belgian raw milk samples

Psychrotolerant bacteria and their heat-resistant proteases play a major role in the spoilage of UHT-processed dairy products. Summer and winter raw milk samples were screened for the presence of such bacteria. One hundred and three proteolytic psychrotolerant bacteria were isolated, characterized by API tests, rep-PCR fingerprint analysis and evaluated for heat-resistant protease production. Twenty-nine strains (representing 79% of the complete collection) were further identified by 16S rRNA gene sequencing, rpoB gene sequencing and DNA–DNA hybridizations. A seasonal inter- and intra-species influence on milk spoilage capacity (e.g. growth rate and/or protease production) was demonstrated. Moreover, this polyphasic approach led to the identification of Pseudomonas fragi and Pseudomonas lundensis (representing 53% of all isolates) as predominant producers of heat-resistant proteases in raw milk. The role of Pseudomonas fluorescens , historically reported as important milk spoiler, could not unequivocally be established. The use of more reliable identification techniques and further revision of the taxonomy of P. fluorescens will probably result in a different perspective on its role in the milk spoilage issue.     

Microbial spoilage of whole sheep livers.

When liver treated with antibiotics to inhibit microbial growth were held at 10 degrees C, the initial high pH (6.4) declined as lactic acid accumulated throughout the storage period of 10 days. The glycogen content also declined, but the glucose concentration in the tissues remained high. When livers were allowed to spoil at 10 degrees C, distinct but variable floras developed within the tissues, in the drip, and on the upper surface. Deep-tissue floras were composed of anaerobic and facultative organisms (Lactobacillus, Enterobacter, Aeromonas); surface floras were generally dominated by strictly aerobic organisms (Pseudomonas, Acinetobacter); drip floras contained variable proportions of organisms of all three types, but the facultatively anaerobic Enterobacter were usually present as a major component. Spoilage occurred after 4 to 6 days with the development of visible discrete colonies on the upper surface without spoilage odors being evident. Changes in tissue and drip composition due to microbial activity could be detected only when spoilage had reached an advanced stage.     

Estimation of bacteriological spoilage of pork cutlets by electronic nose

The utility of chemosensor array (EN) signals of head-space volatiles of aerobically stored pork cutlets as a non-invasive technique for monitoring their microbiological load was studied during storage at 4, 8 and 12 degrees C, respectively. The bacteriological quality of the meat samples was determined by standard total aerobic plate counts (TAPC) and colony count of selectively estimated Pseudomonas (PS) spp., the predominant aerobic spoilage bacteria. Statistical analysis of the electronic nose measurements were principal component analysis (PCA), and canonical discriminant analysis (CDA). Partial least squares (PLS) regression was used to model correlation between microbial loads and EN signal responses, the degree of bacteriological spoilage, independently of the temperature of the refrigerated storage. Sensor selection techniques were applied to reduce the dimensionality and more robust calibration models were computed by determining few individual sensors having the smallest cross correlations and highest correlations with the reference data. Correlations between the predicted and "real" values were given on cross-validated data from both data reduced models and for full calibrations using the 23 sensor elements. At the same time, sensorial quality of the raw cutlets was noted subjectively on faultiness of the odour and colour, and drip formation of the samples. These preliminary studies indicated that the electronic nose technique has a potential to detect bacteriological spoilage earlier or at the same time as olfactory quality deterioration.