Influence of different types of mineral fertilizers on reproduction of Listeria monocytogenes in soil

Influence of mineral fertilizers on the reproduction of Listeria monocytogenes in soil ecosystems under different temperatures (4-6 degrees C and 20-22 degrees C) was studied. Application of the fertilizers favored preservation and active reproduction of L. monocytogenes in brown podzolic soil and brown forest soil. Duration of application of the mineral fertilizers positively correlated with preservation and reproduction of L. monocytogenes in different types of soil.     

Listeria monocytogenes Scott A: cell surface charge, hydrophobicity, and electron donor and acceptor characteristics under different environmental growth conditions

We determined the variations in the surface physicochemical properties of Listeria monocytogenes Scott A cells that occurred under various environmental conditions. The surface charges, the hydrophobicities, and the electron donor and acceptor characteristics of L. monocytogenes Scott A cells were compared after the organism was grown in different growth media and at different temperatures; to do this, we used microelectrophoresis and the microbial adhesion to solvents method. Supplementing the growth media with glucose or lactic acid affected the electrical, hydrophobic, and electron donor and acceptor properties of the cells, whereas the growth temperature (37, 20, 15, or 8 degrees C) primarily affected the electrical and electron donor and acceptor properties. The nonlinear effects of the growth temperature on the physicochemical properties of the cells were similar for cells cultivated in two different growth media, but bacteria cultivated in Trypticase soy broth supplemented with 6 g of yeast extract per liter (TSYE) were slightly more hydrophobic than cells cultivated in brain heart infusion medium (P < 0.05). Adhesion experiments conducted with L. monocytogenes Scott A cells cultivated in TSYE at 37, 20, 15, and 8 degrees C and then suspended in a sodium chloride solution (1.5 x 10(-1) or 1.5 x 10(-3) M NaCl) confirmed that the cell surface charge and the electron donor and acceptor properties of the cells had an influence on their attachment to stainless steel.     

Effect of cultivation temperature on nucleic acid level in bacteria Listeria monocytogenes and Yersinia pseudotuberculosis

The relationship between the multiplication of bacteria, the content of nucleic acid and the specific rate of their growth during their batch cultivation in nutrient broth and mineral medium at temperatures of 37 degrees C and 4-6 degrees C was studied in the causative agents of saprozoonotic infections with L. monocytogenes and Y. pseudotuberculosis used as typical representatives of such bacteria. The content of DNA was shown to remain practically unchanged after the alteration of cultivation temperature and the conditions of nutrition. The linear relationship between the content of RNA and specific growth rate was registered both at 37 degrees C and 4-6 degrees C. However a higher content of RNA at low temperatures was found to correspond to one and the same specific growth rate, which was linked with the additional synthesis of this nucleic acid.     

Mild heat treatment of lettuce enhances growth of Listeria monocytogenes during subsequent storage at 5 degrees C or 15 degrees C

AIMS: The objective of this study was to determine the influence of mild heat treatment, storage temperature and storage time on the survival and growth of Listeria monocytogenes inoculated onto cut iceberg lettuce leaves. METHODS AND RESULTS: Before or after inoculation with L. monocytogenes, cut iceberg lettuce leaves were dipped in water (20 or 50 degrees C) containing or not 20 mg l(-1) chlorine, for 90 s, then stored at 5 degrees C for up to 18 days or 15 degrees C for up to 7 days. The presence of 20 mg l(-1) chlorine in the treatment water did not significantly (alpha=0.05) affect populations of the pathogen, regardless of other test parameters. The population of L. monocytogenes on lettuce treated at 50 degrees C steadily increased throughout storage at 5 degrees C for up to 18 days. At day 10 and thereafter, populations were 1.7-2.3 log10 cfu g(-1) higher on lettuce treated at 50 degrees C after inoculation compared with untreated lettuce or lettuce treated at 20 degrees C, regardless of chlorine treatment. The population of L. monocytogenes increased rapidly on lettuce stored at 15 degrees C. At 2 and 4 days, significantly higher populations were detected on lettuce that had been treated at 50 degrees C, compared with respective samples that had been treated at 20 degrees C, regardless of inoculation before or after treatment, or the presence of 20 mg l(-1) chlorine in the treatment water. CONCLUSIONS: The results clearly demonstrated that mild heat treatment of cut lettuce leaves enhances the growth of L. monocytogenes during subsequent storage at 5 or 15 degrees C. SIGNIFICANCE AND IMPACT OF THE STUDY: Mild heat treatment of cut lettuce may result in a prolonged shelf life as a result of delaying the development of brown discoloration. However, heat treatment also facilitates the growth of L. monocytogenes during storage at refrigeration temperature, thereby increasing the potential risk of causing listeriosis.     

Adaptive ultrastructural changes in soil-resident Yersinia pseudotuberculosis bacteria

After the prolonged residence (from 1 month to 2 years) in flow soil columns at 6 - 8 degrees C and 18 - 20 degrees C a complex of ultrastructural changes was detected in Y. pseudotuberculosis bacteria, depending on temperature, the duration of residence in the soil and, to a definite extent, on the strain. They were manifested in the form of cell-wall changes, the formation of the capsule and intercellular slime, changes in the ribosomal saturation of cytoplasm and the conformation state of DNA in the nucleoid zone. As the result of adaptation to nutritional deficit storage substances were accumulated in the form of electron-dense inclusions (polyphosphates) and pseudovacuoles (poly-beta-oxibutyric acid). Temperature influenced the process of mitotic division and the state of chromatin in bacteria. The described ultrastructural changes Y. pseudotuberculosis may be regarded as natural adaptive reaction to the altered conditions of their residence.     

Effect of the lactoperoxidase system on Listeria monocytogenes behavior in raw milk at refrigeration temperatures.

Activity of raw milk lactoperoxidase-thiocyanate-hydrogen peroxide (LP) system on four Listeria monocytogenes strains at refrigeration temperatures after addition of 0.25 mM sodium thiocyanate and 0.25 mM hydrogen peroxide was studied. The LP system exhibited a bactericidal activity against L. monocytogenes at 4 and 8 degrees C; the activity was dependent on temperature, length of incubation, and strain of L. monocytogenes tested. D values in activated-LP system milk for the four strains tested ranged from 4.1 to 11.2 days at 4 degrees C and from 4.4 to 9.7 days at 8 degrees C. The lactoperoxidase level in raw milk declined during a 7-day incubation, the decrease being more pronounced at 8 degrees C than at 4 degrees C and in control milk than in activated-LP system milk. The thiocyanate concentration decreased considerably in activated-LP system milk at both temperatures during the first 8 h of incubation. LP system activation was shown to be a feasible procedure for controlling development of L. monocytogenes in raw milk at refrigeration temperatures.     

Minimum growth temperatures of Listeria monocytogenes and non-haemolytic Listeria

Minimum growth temperatures and those of decreased growth were determined for 100 strains of listerias. The ability of 78 strains of Listeria monocytogenes isolated from animals and 22 non-haemolytic strains to grow at low temperatures was studied, using a flooding technique, in a plate-type continuous temperature gradient incubator at temperatures between -1.6 and 14.5 degrees C. The mean minimum temperature for L. monocytogenes was +1.7 +/- 0.5 degrees C. The growth of non-haemolytic listerias was unobservable at +1.7 +/- 0.5 degrees C. The L. monocytogenes strains grew at about 0.6 degrees C lower than the non-pathogenic strains. No differences in growth temperatures were observed among L. monocytogenes strains isolated from different sources. The serovars with the OI antigen grew at lower temperatures (+1.0 +/- 0.3 degrees C) than the other common serovar 4b (+1.3 +/- 0.4 degrees C). The results indicate that L. monocytogenes grows better than non-haemolytic strains under cold conditions. The possible role of haemolysins as growth factors is also discussed.     

Effect of the lactoperoxidase system on Listeria monocytogenes behavior in raw milk at refrigeration temperatures.

Activity of raw milk lactoperoxidase-thiocyanate-hydrogen peroxide (LP) system on four Listeria monocytogenes strains at refrigeration temperatures after addition of 0.25 mM sodium thiocyanate and 0.25 mM hydrogen peroxide was studied. The LP system exhibited a bactericidal activity against L. monocytogenes at 4 and 8 degrees C; the activity was dependent on temperature, length of incubation, and strain of L. monocytogenes tested. D values in activated-LP system milk for the four strains tested ranged from 4.1 to 11.2 days at 4 degrees C and from 4.4 to 9.7 days at 8 degrees C. The lactoperoxidase level in raw milk declined during a 7-day incubation, the decrease being more pronounced at 8 degrees C than at 4 degrees C and in control milk than in activated-LP system milk. The thiocyanate concentration decreased considerably in activated-LP system milk at both temperatures during the first 8 h of incubation. LP system activation was shown to be a feasible procedure for controlling development of L. monocytogenes in raw milk at refrigeration temperatures.     

Thermal inactivation of Listeria monocytogenes during a process simulating temperatures achieved during microwave heating

Conventional heating was used to expose cells of Listeria monocytogenes, either in broth or in situ on chicken skin, to the mean times and temperatures that are achieved during a 28 min period of microwave cooking of a whole chicken. Heating L. monocytogenes by this method in culture broth resulted in a reduction in viable cell numbers by a factor of greater than 10(6) upon reaching 70 degrees C. Simulated microwave cooking of L. monocytogenes in situ, on chicken skin, resulted in more variability in the numbers of survivors. Heating for the full cook time of 28 min, however, resulted in a mean measured temperature of 85 degrees C and no surviving listerias were detected. This indicated a reduction in viable numbers of greater than 10(6). To reduce temperature variation, cells were heated on skin in a submerged system in which exposure to 70 degrees C for 2 min resulted in a reduction in viable cell numbers of all strains of listerias tested of between 10(6) and 10(8). These results show that when a temperature of 70 degrees C is reached and maintained for at least 2 min throughout a food there is a substantial reduction in the numbers of L. monocytogenes. The survival of this organism during microwave heating when temperatures of over 70 degrees C are reported is probably due to uneven heating by microwave ovens resulting in the presence of cold spots in the product. The heat resistance of L. monocytogenes is comparable with that of many other non-sporing mesophilic bacteria.     

Shelf life and safety aspects of chilled cooked and peeled shrimps (Pandalus borealis) in modified atmosphere packaging

AIMS: To evaluate the growth of Listeria monocytogenes and shelf life of cooked and peeled shrimps in modified atmosphere packaging (MAP). METHODS AND RESULTS: Storage trials with naturally contaminated cooked and peeled MAP shrimps (Pandalus borealis) were carried out at 2, 5 and 8 degrees C. Challenge tests at the same conditions were performed after inoculation with Listeria monocytogenes. Both storage trials and challenge tests were repeated after 4 months of frozen storage (-22 degrees C). Brochothrix thermosphacta and Carnobacterium maltaromaticum were responsible for sensory spoilage of cooked and peeled MAP shrimps. In challenge tests, growth of L. monocytogenes was observed at all of the storage temperatures studied. At 5 and 8 degrees C the concentration of L. monocytogenes increased more than a 1000-fold before the product became sensory spoiled whereas this was not observed at 2 degrees C. Frozen storage had only a minor inhibiting effect on growth of L. monocytogenes in the thawed product. CONCLUSIONS: To prevent L. monocytogenes becoming a safety problem, cooked and peeled MAP shrimps should be distributed at 2 degrees C and with a maximum shelf life of 20-21 d. At higher temperatures shelf life is significantly reduced. SIGNIFICANCE AND IMPACT OF THE STUDY: Information is provided to establish shelf life of cooked and peeled MAP shrimps.     

Inhibition of Listeria monocytogenes by fatty acids and monoglycerides.

Fatty acids and monoglycerides were evaluated in brain heart infusion broth and in milk for antimicrobial activity against the Scott A strain of Listeria monocytogenes. C12:0, C18:3, and glyceryl monolaurate (monolaurin) had the strongest activity in brain heart infusion broth and were bactericidal at 10 to 20 micrograms/ml, whereas potassium (K)-conjugated linoleic acids and C18:2 were bactericidal at 50 to 200 micrograms/ml. C14:0, C16:0, C18:0, C18:1, glyceryl monomyristate, and glyceryl monopalmitate were not inhibitory at 200 micrograms/ml. The bactericidal activity in brain heart infusion broth was higher at pH 5 than at pH 6. In whole milk and skim milk, K-conjugated linoleic acid was bacteriostatic and prolonged the lag phase especially at 4 degrees C. Monolaurin inactivated L. monocytogenes in skim milk at 4 degrees C, but was less inhibitory at 23 degrees C. Monolaurin did not inhibit L. monocytogenes in whole milk because of the higher fat content. Other fatty acids tested were not effective in whole or skim milk. Our results suggest that K-conjugated linoleic acids or monolaurin could be used as an inhibitory agent against L. monocytogenes in dairy foods.     

Biology of Cerotoma arcuatus (Coleoptera: Chrysomelidae) and field validation of a laboratory model for temperature requirements

Cerotoma arcuatus Olivier is a polyphagous pest of legumes [soybean, Glycine max (L.); dry beans, Phaseolus vulgaris (L.); and cowpeas, Vigna unguiculata (L.)], all of which are considered important protein sources for humans and domestic animals. Studies on the biology and temperature requirements of C. arcuatus were made under laboratory and field (cage) conditions. In the laboratory, insects were reared on soybean plants in incubators held at 18, 20, 22, 25, 28, 30, or 32 degrees C, 70 +/- 10% RH, and a photoperiod of 14:10 (L:D) h. A degree-days (DD) model developed based on the incubator data were validated in the field based on air and soil temperatures. The duration of the egg, larva-to-adult, and egg-to-adult period was inversely correlated with the temperature within the range of 18-32 degrees C, with the highest viability found from 20 to 30 degrees C. The temperature threshold for development and the thermal constant for the egg phase and the larva-to-adult periods were 13.6 degrees C and 106.7 DD, 8.3 degrees C and 399.4 DD, and 10.7 degrees C and 489.0 DD, respectively. The DD model for the egg-to-adult period, calculated using constant temperatures in the laboratory, was found to be valid for populations of C. arcuatus in the field, based on the fluctuating air and soil temperatures, although air temperatures provided more precise predictions. These data provide support for the rational control of this pest through population predictions based on their temperature requirements.     

Inhibition of Listeria monocytogenes by fatty acids and monoglycerides.

Fatty acids and monoglycerides were evaluated in brain heart infusion broth and in milk for antimicrobial activity against the Scott A strain of Listeria monocytogenes. C12:0, C18:3, and glyceryl monolaurate (monolaurin) had the strongest activity in brain heart infusion broth and were bactericidal at 10 to 20 micrograms/ml, whereas potassium (K)-conjugated linoleic acids and C18:2 were bactericidal at 50 to 200 micrograms/ml. C14:0, C16:0, C18:0, C18:1, glyceryl monomyristate, and glyceryl monopalmitate were not inhibitory at 200 micrograms/ml. The bactericidal activity in brain heart infusion broth was higher at pH 5 than at pH 6. In whole milk and skim milk, K-conjugated linoleic acid was bacteriostatic and prolonged the lag phase especially at 4 degrees C. Monolaurin inactivated L. monocytogenes in skim milk at 4 degrees C, but was less inhibitory at 23 degrees C. Monolaurin did not inhibit L. monocytogenes in whole milk because of the higher fat content. Other fatty acids tested were not effective in whole or skim milk. Our results suggest that K-conjugated linoleic acids or monolaurin could be used as an inhibitory agent against L. monocytogenes in dairy foods.     

The multiplication dynamics of pathogenic bacteria in relation to the trophic and temperature cultivation conditions

The comparative study of the dynamics of multiplication of Listeria monocytogenes and Yersinia pseudotuberculosis in organic and synthetic media and in distilled water at temperatures of 37 degrees C and 6 degrees C was carried out. This study revealed that in organic media the multiplication of bacteria was good at 37 degrees C and 6 degrees C. In mineral media and distilled water their multiplication was observed only at 6 degrees C. Moreover, conditions necessary for the multiplication of pathogenic bacteria in distilled water were shown; these conditions depended on the inoculated dose, the number of autolyzed microbial cells and the state of the culture. Proofs of the multiplication of the bacteria under the conditions of minimum nutrition and low temperature were obtained with the use of labeled 3H-thymidine.     

Effect of water temperature on the ability of Diplostomum spathaceum miracidia to establish in lymnaeid snails

The effects of water temperature on the ability of Diplostomum spathaceum miracida to infect and establish patent infections in Lymnaea peregra and L. stagnalis were investigated. Snails were infected over a range of temperatures (6-20 degrees C) and kept thereafter at 20 degrees C or were infected at 20 degrees C and kept at either 14, 20, or 25 degrees C. Infection success was determined after 8 weeks by either observing cercarial shedding or examining snail viscera for sporocysts. The establishment of miracidia declined at lower water temperatures despite maintenance for 8 weeks at 20 degrees C while exposure of snails to miracidia at 20 degrees C and maintenance at different temperatures had little apparent effect. Infection success under these conditions was related more to the numbers of miracidia to which the snails were exposed. However, under this latter experimental regime, the time taken for the infection to become patent clearly depended upon maintenance temperature.     

Factors affecting the development of Dipylidium caninum in Ctenocephalides felis felis (Bouché, 1835)

Ctenocephalides felis felis larvae were infected with Dipylidium caninum at a range of temperatures from 20 degrees - 35 degrees C at 3 mm Hg saturation deficit (SD) and 30 degrees C at 8 mm Hg SD. Hosts were subsequently dissected at 6, 9 and 12 days after infection. Four replicate experiments were performed and results of development, and host reactions analysed by the Genstat computer programme. These were found to depend on the temperature and saturation deficit of the environment. Unlike previous findings, parasite development and host reaction were found to be independent of host development. Host reaction was more marked and prolonged at 20 degrees - 25 degrees C than at higher temperatures. No perceptible growth of the parasite occurred at 20 degrees C. The development patterns of growth at the higher temperatures were similar but shifted in time so that faster growth occurred at higher temperatures. Rate of growth was fastest at 35 degrees C, despite the fact that this temperature was unfavourable to the hosts, all of which died at the time of pupation.     

The differential adherence capabilities of two Listeria monocytogenes strains in monoculture and multispecies biofilms as a function of temperature

AIMS: To determine the differential adherence capabilities at three different temperatures of Listeria monocytogenes Scott A, a clinical food pathogen, and L. monocytogenes FM876, a persistent strain from a milk-processing environment, to stainless steel. METHODS AND RESULTS: Differential adherence was investigated by submerging stainless steel coupons in both 48-h Listeria monocultures and mixed cultures additionally containing Staphylococcus xylosus DP5H and Pseudomonas fragi ATCC 4973. Immunofluorescent microscopy and image analysis techniques were utilized to identify and quantify the L. monocytogenes cells adhering to the steel at 4 degrees C, 18 degrees C and 30 degrees C. The monoculture biofilms consistently contained greater L. monocytogenes numbers than the multispecies biofilms, with the persistent strain FM876 showing significantly greater adherence than strain Scott A. Optimum adherence occurred at 18 degrees C in monoculture biofilms. CONCLUSION: L. monocytogenes strains exhibit differential, temperature-dependent, adherence to stainless steel. SIGNIFICANCE AND IMPACT OF THE STUDY: These results demonstrate temperature dependent biofilm adherence and support previous findings that persistent strains exhibit increased adherence capability.     

Clonal structure of Yersinia pestis populations in experimental soil ecosystems

Two basic tendencies--formation of latent (uncultivable) form (LF) and hemin storage variability--has been revealed during study of clonal structure dynamics of Y. pestis populations in artificial soil ecosystems in long-term incubation conditions. Y. pestis populations disappeared within 3 - 6 months at 18 - 22 degrees C, whereas at 4 - 8 degrees C a subsequent replacement of vegetative cells on LF, which are capable to prolonged survival (up to 22 months) in soil with ability to reversion in the presence of abundance of nutrients, has been observed. Bacteria of virulent strain retained all determinants of pathogenicity when reverted to LF, whereas bacteria of avirulent strain (defective on plasmid of Ca-dependence), on the contrary, undergo further degradation that resulted in loss of a pgm locus and gradual disappearance of population. LF revertants of highly virulent strain restored properties of initial population and were highly virulent.     

Thermal resistance of intracellular Listeria monocytogenes cells suspended in raw bovine milk.

The thermal resistance of Listeria monocytogenes associated with a milk-borne outbreak of listeriosis was determined in parallel experiments by using freely suspended bacteria and bacteria internalized by phagocytes. The latter inoculum was generated by an in vitro phagocytosis reaction with immune-antigen-elicited murine peritoneal phagocytes. The heat suspension medium was raw whole bovine milk. Both suspensions were heated at temperatures ranging from 52.2 to 71.7 degrees C for various periods of time. Mean D values for each temperature and condition of heated suspension revealed no significant differences. The extrapolated D71.7 degrees C (161 degrees F) value for bacteria internalized by phagocytes was 1.9 s. Combined tube and slug-flow heat exchanger results yielded an estimated D71.7 degrees C value of 1.6 s for freely suspended bacteria. The intracellular position did not protect L. monocytogenes from thermal inactivation.     

Critical role of anteiso-C15:0 fatty acid in the growth of Listeria monocytogenes at low temperatures.

Listeria monocytogenes is a food-borne pathogen capable of growth at refrigeration temperatures. Membrane lipid fatty acids are major determinants of a sufficiently fluid membrane state to allow growth at low temperatures. L. monocytogenes was characterized by a fatty acid profile dominated to an unusual extent (> 95%) by branched-chain fatty acids, with the major fatty acids being anteiso-C15:0, anteiso-C17:0, and iso-C15:0 in cultures grown in complex or defined media at 37 degrees C. Determination of the fatty acid composition of L. monocytogenes 10403S and SLCC 53 grown over the temperature range 45 to 5 degrees C revealed two modes of adaptation of fatty acid composition to lower growth temperatures: (i) shortening of fatty acid chain length and (ii) alteration of branching from iso to anteiso. Two transposon Tn917-induced cold-sensitive mutants incapable of growth at low temperatures had dramatically altered fatty acid compositions with low levels of i-C15:0, a-C15:0, and a-C17:0 and high levels of i-C14:0, C14:0, i-C16:0, and C16:0. The levels of a-C15:0 and a-C17:0 and the ability to grow at low temperatures were restored by supplementing media with 2-methylbutyric acid, presumably because it acted as a precursor of methylbutyryl coenzyme A, the primer for synthesis of anteiso odd-numbered fatty acids. When mid-exponential-phase 10403S cells grown at 37 degrees C were temperature down-shocked to 5 degrees C they were able, for the most part, to reinitiate growth before the membrane fatty acid composition had reset to a composition more typical for low-temperature growth. No obvious evidence was found for a role for fatty acid unsaturation in adaptation of L. monocytogenes to cold temperature. The switch to a fatty acid profile dominated by a-C15:0 at low temperatures and the association of cold sensitivity with deficiency of a-C15:0 focus attention on the critical role of this fatty acid in growth of L. monocytogenes in the cold, presumably through its physical properties and their effects, in maintaining a fluid, liquid-crystalline state of the membrane lipids.