Pathogenicity of Listeria monocytogenes grown on crabmeat

The pathogenicity of Listeria monocytogenes as influenced by growth on crabmeat at 5 and 10 degrees C was studied. Crabmeat was inoculated with L. monocytogenes V7 (ca. 10(4) CFU/g) and incubated for up to 14 days at 5 and 10 degrees C. At selected incubation times, L. monocytogenes was removed from crabmeat by washing with 0.1 M potassium phosphate buffer (pH 7.0), and populations were determined by surface plating on LiCl-phenylethanol-moxalactam agar. Buffered suspensions were then centrifuged, and the resulting pellets were suspended in phosphate buffer containing 10% glycerol and stored at -18 degrees C. Thawed, diluted suspensions of cells were tested for pathogenicity by intraperitoneal injection into immunocompromised and nonimmunocompromised mice. L. monocytogenes cells recovered from crabmeat and then recultured in tryptose phosphate broth (TPB), as well as cells which had not been passed through crabmeat but had been cultured in TPB, were likewise harvested, suspended in buffered 10% glycerol, frozen, thawed, diluted, and tested for pathogenicity by intraperitoneal injection. Growth on crabmeat at 5 and 10 degrees C did not have a significant effect on pathogenicity. The population of L. monocytogenes necessary to kill about 50% of the immunocompromised mice in each test set within 7 days was about 10(4) CFU, and this result was not significantly affected by storage temperature of the crabmeat or type of substrate, i.e., crabmeat or TPB, on which it had grown.     

Pathogenicity of Listeria monocytogenes grown on crabmeat.

The pathogenicity of Listeria monocytogenes as influenced by growth on crabmeat at 5 and 10 degrees C was studied. Crabmeat was inoculated with L. monocytogenes V7 (ca. 10(4) CFU/g) and incubated for up to 14 days at 5 and 10 degrees C. At selected incubation times, L. monocytogenes was removed from crabmeat by washing with 0.1 M potassium phosphate buffer (pH 7.0), and populations were determined by surface plating on LiCl-phenylethanol-moxalactam agar. Buffered suspensions were then centrifuged, and the resulting pellets were suspended in phosphate buffer containing 10% glycerol and stored at -18 degrees C. Thawed, diluted suspensions of cells were tested for pathogenicity by intraperitoneal injection into immunocompromised and nonimmunocompromised mice. L. monocytogenes cells recovered from crabmeat and then recultured in tryptose phosphate broth (TPB), as well as cells which had not been passed through crabmeat but had been cultured in TPB, were likewise harvested, suspended in buffered 10% glycerol, frozen, thawed, diluted, and tested for pathogenicity by intraperitoneal injection. Growth on crabmeat at 5 and 10 degrees C did not have a significant effect on pathogenicity. The population of L. monocytogenes necessary to kill about 50% of the immunocompromised mice in each test set within 7 days was about 10(4) CFU, and this result was not significantly affected by storage temperature of the crabmeat or type of substrate, i.e., crabmeat or TPB, on which it had grown.     

Inhibition of Listeria monocytogenes by piscicolin 126 in milk and Camembert cheese manufactured with a thermophilic starter

The effect of bacteriocin, piscicolin 126, on the growth of Listeria monocytogenes and cheese starter bacteria was investigated in milk and in Camembert cheese manufactured from milk challenged with 10(2) cfu ml(-1) L. monocytogenes. In milk incubated at 30 degrees C, piscicolin 126 added in the range of 512-2,048 AU ml(-1) effectively inhibited growth of L. monocytogenes for more than 20 d when challenged with approximately 10(2) cfu ml(-1) L. monocytogenes. At higher challenge levels (10(4) and 10(6) cfu ml(-1)), piscicolin 126 reduced the viable count of L. monocytogenes by 4-5 log units immediately after addition of the bacteriocin; however, growth of Listeria occurred within 24 h. The minimum inhibitory concentration (MIC) of piscicolin 126 against lactic acid cheese starter bacteria was generally greater than 204,800 AU ml(-1) , and the viable count and acid production of these starter cultures in milk were not affected by the addition of 2,048 AU ml(-1) piscicolin 126. Camembert cheeses made from milk challenged with L. monocytogenes and with added piscicolin 126 showed a viable count of L. monocytogenes 3-4 log units lower than those without piscicolin 126. Inactivation of piscicolin 126 by proteolytic enzymes from cheese starter bacteria and mould together with the emergence of piscicolin 126-resistant isolates was responsible for the recovery of L. monocytogenes in the cheeses during ripening.     

Fate of Listeria spp. on parsley leaves grown in laboratory and field cultures

AIMS: To investigate the population dynamics of Listeria monocytogenes and Listeria innocua on the aerial surfaces of parsley. METHODS AND RESULTS: Under 100% relative humidity (RH) in laboratory and regardless of the inoculum tested (10(3)-10(8) CFU per leaf), counts of L. monocytogenes EGDe, LO28, LmP60 and L. innocua CIP 80-12 tended towards approx. 10(5) CFU per leaf. Under low RH, Listeria spp. populations declined regardless to the inoculum size (10(4)-10(8) CFU per leaf). L. innocua CIP 80-12 survived slightly better than L. monocytogenes in the laboratory and was used in field cultures. Under field cultures, counts of L. innocua decreased more rapidly than in the laboratory, representing a decrease of 9 log(10) in 2 days in field conditions compared to a decrease of 4.5 log(10) in 8 days in the laboratory. Counts of L. innocua on tunnel parsley cultures were always higher (at least by 100 times) than those on unprotected parsley culture. CONCLUSIONS: Even with a high inoculum and under protected conditions (i.e. plastic tunnels), population of L. monocytogenes on the surface of parsley on the field would decrease by several log(10) scales within 2 days. SIGNIFICANCE AND IMPACT OF THE STUDY: Direct contamination of aerial surfaces of parsley with L. monocytogenes (i.e. through contaminated irrigation water) will not lead to contaminated produce unless it occurs very shortly before harvest.     

Inhibition of Listeria monocytogenes by enterocin 4 during the manufacture andripening of Manchego cheese

The inhibitory effect of enterocin 4, a bacteriocin produced by Enterococcus faecalis INIA 4, on Listeria monocytogenes strains Ohio and Scott A during themanufacture and ripening of Manchego cheese was investigated. Raw ewe's milk wasinoculated with ca 10⁵ cfu ml⁻¹ of L.monocytogenes and with 1% of a commercial lactic starter, 1% of an Ent. faecalis INIA 4 culture, or 1% of each culture. Manchego cheeses were manufactured according tousual procedures. Listeria monocytogenes Ohio counts decreased by 3 log units after8 h and by 6 log units after 7 d in cheese made from milk inoculated with Ent. faecalis INIA 4 or with both cultures, whereas no inhibition was recorded after 60 d in cheese made frommilk inoculated with commercial lactic starter. Listeria monocytogenes Scott A wasnot inhibited by enterocin 4 during cheese manufacture, but decreases of 1 log unit after 7 d andof 2 log units after 60 d were achieved in cheese made from milk inoculated with bothcommercial lactic starter and Ent. faecalis INIA 4.     

INFLUENCE OF GLUCOSE OXIDASE ON THE GROWTH OF ESCHERICHIA COLI 0157:H7, LISTERIA MONOCYTOGENES, AND SALMONELLA TYPHIMURIUM IN UNIVERSAL PREENRICHMENT BROTH

Glucose oxidase (GO), a food-grade enzyme, was compared with Oxyrase^TM oxygen reducing membrane fraction in Universal Preenrichment Broth (UPB) for enhancement of the growth of the facultatively anaerobic pathogens Escherichia coli O157:H7, Salmonella typhimurium, and Listeria monocytogenes Scott A. Oxidation-reduction potential (ORP) and pH changes in UPB following the addition of GO (4 units/ml) or Oxyrase^TM (0.5 units/ml) were measured. Microbial growth was evaluated at 0, 3, 6, 18, and 24 h of incubation using spiral plating. Nonenzyme supplemented UPB served as the control. Oxyrase^TM provided a higher oxygen scavenging action in terms of ORP decrease during the initial 6 h of incubation. However, no difference occurred in Eh between Oxyrase^TM and GO by 18 h, with both enzyme systems effectively reducing the Eh compared to that of the control. A 1.0 pH unit reduction was observed in GO-supplemented UPB after 18 h, indicating production of gluconic acid. The pH decrease in Oxyrase^TM - supplemented media was 0.2 units. By 6 h, the E. coli O157:H7 population was enhanced by 0.6 and 1.4 log CFU/ml in Oxyrase^TM -supplemented media, compared to the control and GO-supplemented media, respectively. By 18 h, 0.4 and 0.9 log CFU/ml growth enhancements of the E. coli O157:H7 populations were seen in GO- and Oxyrase^TM -supplemented media, respectively, compared to the control. By the end of 18 h, counts of S. typhimurium and L. monocytogenes increased by 0.6 and 0.2 log units, respectively, in GO-supplemented media compared to the control.     

The use of enterocin CCM 4231 in soy milk to control the growth of Listeria monocytogenes and Staphylococcus aureus

The inhibitory effect of enterocin CCM 4231 (concentration 3200 AU ml-1) was used to control the growth of Listeria monocytogenes Ohio and Staphylococcus aureus in soy milk. The growth and bacteriocin (enterocin) production of producer strain CCM 4231 in soy milk was also checked. Bacteriocin production by CCM 4231 strain in soy milk was first detected after 2 h from the beginning of cultivation (100 AU ml-1). The stationary phase for CCM 4231 was reached after 6 h reaching 10.38 cfu ml-1 (log10) with a slight increase up to 24 h (10.43 cfu ml-1, log10), and the maximum bacteriocin production in soy milk (200 AU ml-1) was noted after 8 h of the beginning of cultivation with stability up to 24 h. The addition of enterocin CCM 4231 at 3200 AU ml-1 to a growing indicator strain, L. monocytogenes Ohio, in soy milk resulted in inhibition for 24 h. The high inhibitory effect of enterocin was found after 1 h and 2 h of its addition (in 5 h-6 h of cultivation), the difference between the experimental and the control samples (ES, CS) being 4.96 log cycles at 5 h and 5.15 log cycles at 6 h. Staphylococcus aureus was not fully inhibited, although a difference of 3.55 log cycles was found when ES and CS were compared at the end of cultivation (24 h). The pH was not influenced by enterocin addition. The inhibitory effect of enterocin CCM 4231 against L. monocytogenes Ohio in soy milk was probably bacteriocidal; while Staph. aureus was influenced bacteriostatically. In general, the observed inhibitory activity confirmed the possibility for further application of bacteriocins in food environments as the protective agents. Of course, legislation problems must be solved.     

Microplate fluorescence assay for measurement of the ability of strains of Listeria monocytogenes from meat and meat-processing plants to adhere to abiotic surfaces

Listeria monocytogenes is a significant food-borne pathogen that is capable of adhering to and producing biofilms on processing equipment, making it difficult to eliminate from meat-processing environments and allowing potential contamination of ready-to-eat (RTE) products. We devised a fluorescence-based microplate method for screening isolates of L. monocytogenes for the ability to adhere to abiotic surfaces. Strains of L. monocytogenes were incubated for 2 days at 30 degrees C in 96-well microplates, and the plates were washed in a plate washer. The retained cells were incubated for 15 min at 25 degrees C with 5,6-carboxyfluorescein diacetate and washed again, and then the fluorescence was read with a plate reader. Several enzymatic treatments (protease, lipase, and cellulase) were effective in releasing adherent cells from the microplates, and this process was used for quantitation on microbiological media. Strongly adherent strains of L. monocytogenes were identified that had 15,000-fold-higher levels of fluorescence and 100,000-fold-higher plate counts in attachment assays than weakly adherent strains. Strongly adherent strains of L. monocytogenes adhered equally well to four different substrates (glass, plastic, rubber, and stainless steel); showed high-level attachment on microplates at 10, 20, 30, and 40 degrees C; and showed significant differences from weakly adherent strains when examined by scanning electron microscopy. A greater incidence of strong adherence was observed for strains isolated from RTE meats than for those isolated from environmental surfaces. Analysis of surface adherence among Listeria isolates from processing environments may provide a better understanding of the molecular mechanisms involved in attachment and suggest solutions to eliminate them from food-processing environments.     

Effect of immersion solutions containing enterocin AS-48 on Listeria monocytogenes in vegetable foods

The effect of immersion solutions containing enterocin AS-48 alone or in combination with chemical preservatives on survival and proliferation of Listeria monocytogenes CECT 4032 inoculated on fresh alfalfa sprouts, soybean sprouts, and green asparagus was tested. Immersion treatments (5 min at room temperature) with AS-48 solutions (25 microg/ml) reduced listeria counts of artificially contaminated alfalfa and soybean sprouts by approximately 2.0 to 2.4 log CFU/g compared to a control immersion treatment in distilled water. The same bacteriocin immersion treatment applied on green asparagus had a very limited effect. During storage of vegetable samples treated with immersion solutions of 12.5 and 25 microg of AS-48/ml, viable listeria counts were reduced below detection limits at days 1 to 7 for alfalfa and soybean sprouts at 6 and 15 degrees C, as well as green asparagus at 15 degrees C. Only a limited inhibition of listeria proliferation was detected during storage of bacteriocin-treated alfalfa sprouts and green asparagus at 22 degrees C. Treatment with solutions containing AS-48 plus lactic acid, sodium lactate, sodium nitrite, sodium nitrate, trisodium phosphate, trisodium trimetaphosphate, sodium thiosulphate, n-propyl p-hydroxybenzoate, p-hydoxybenzoic acid methyl ester, hexadecylpyridinium chloride, peracetic acid, or sodium hypochlorite reduced viable counts of listeria below detection limits (by approximately 2.6 to 2.7 log CFU/g) upon application of the immersion treatment and/or further storage for 24 h, depending of the chemical preservative concentration. Significant increases of antimicrobial activity were also detected for AS-48 plus potassium permanganate and in some combinations with acetic acid, citric acid, sodium propionate, and potassium sorbate.     

Behavior of Listeria monocytogenes in wiener exudates in the presence of Pediococcus acidilactici H or pediocin AcH during storage at 4 or 25 degrees C.

Exudative fluids were collected from packages of five brands of all-beef wieners and inoculated to contain 10(4) to 10(5) CFU of a three-strain (Scott A, V7, and 101M) mixture of Listeria monocytogenes per ml. Listeriae were inactivated (decrease of 0.61 to 3.8 log10 CFU/ml) in all five exudates held at 4 degrees C for 29 days. L. monocytogenes grew (increase of 1.7 to 3.6 log10 CFU/ml) in two of five exudates held at 25 degrees C for 6 days. Exudate was inoculated with a derivative of Pediococcus acidilactici H (designated JBL1095) or treated with pediocin AcH (a bacteriocin) as a novel approach to control the growth of L. monocytogenes in wiener exudates. Initially, pediocin AcH caused rapid death (decrease of 0.74 log10 CFU/ml in 2 h) of L. monocytogenes in exudate held at 4 degrees C, but thereafter the inactivation was similar to that in control exudate (L. monocytogenes only) or exudate containing L. monocytogenes plus JBL1095. At 25 degrees C, L. monocytogenes grew in the presence of JBL1095 during the first 64 h of incubation, but thereafter the numbers of the pathogen decreased appreciably (5.84 log10 CFU/ml in 3 days). In the presence of pediocin AcH, there was a gradual decrease in numbers of L. monocytogenes throughout the storage period at 25 degrees C. These data indicate that added biopreservatives can potentiate and amplify the intrinsic listeriostatic or listericidal activity of wiener exudate.     

Behavior of Listeria monocytogenes in wiener exudates in the presence of Pediococcus acidilactici H or pediocin AcH during storage at 4 or 25 degrees C

Exudative fluids were collected from packages of five brands of all-beef wieners and inoculated to contain 10(4) to 10(5) CFU of a three-strain (Scott A, V7, and 101M) mixture of Listeria monocytogenes per ml. Listeriae were inactivated (decrease of 0.61 to 3.8 log10 CFU/ml) in all five exudates held at 4 degrees C for 29 days. L. monocytogenes grew (increase of 1.7 to 3.6 log10 CFU/ml) in two of five exudates held at 25 degrees C for 6 days. Exudate was inoculated with a derivative of Pediococcus acidilactici H (designated JBL1095) or treated with pediocin AcH (a bacteriocin) as a novel approach to control the growth of L. monocytogenes in wiener exudates. Initially, pediocin AcH caused rapid death (decrease of 0.74 log10 CFU/ml in 2 h) of L. monocytogenes in exudate held at 4 degrees C, but thereafter the inactivation was similar to that in control exudate (L. monocytogenes only) or exudate containing L. monocytogenes plus JBL1095. At 25 degrees C, L. monocytogenes grew in the presence of JBL1095 during the first 64 h of incubation, but thereafter the numbers of the pathogen decreased appreciably (5.84 log10 CFU/ml in 3 days). In the presence of pediocin AcH, there was a gradual decrease in numbers of L. monocytogenes throughout the storage period at 25 degrees C. These data indicate that added biopreservatives can potentiate and amplify the intrinsic listeriostatic or listericidal activity of wiener exudate.     

Development of a surface adhesion immunofluorescent technique for the rapid isolation of Listeria monocytogenes and Listeria innocua from meat

The use of a novel surface adhesion technique to isolate Listeria monocytogenes and Listeria innocua from an enrichment meat system was developed. Minced beef samples inoculated with L. monocytogenes (10 cfu g(-1)) were incubated at 30 degrees C for 14-18 h in a suitable enrichment broth. Listeria monocytogenes cells were isolated from the enriched meat sample by surface adhesion onto a polycarbonate membrane which was attached to a glass microscope slide. The Listeria cells on the membrane were subsequently visualized using an immunofluorescent microscopy procedure. The antibody used in this technique reacts with L. monocytogenes and L. innocua. The technique was demonstrated to have a detection level of log10 3.11 cfu ml(-1). There was excellent correlation (r2 = 0.98) between the counts obtained by this surface adhesion immunofluorescent (SAIF) technique and counts obtained using traditional methods, i.e. plate counts on PALCAM. When the regression equation relating the rapid and standard methods was validated using the data from 50 retail beef mince samples, an rsd value of +/- 0.25 was obtained. No false-negative or false-positive results were recorded for L. monocytogenes or L. innocua species using the SAIF technique.     

Efficacy of lactic acid against Listeria monocytogenes attached to poultry skin during refrigerated storage

AIMS: The aim of this study was to evaluate the effect of lactic acid washing on the growth of Listeria monocytogenes on poultry legs stored at 4 degrees C for 7 days. METHODS AND RESULTS: Fresh inoculated chicken legs were dipped into either a 0.11, 0.22 mol l(-1) or 0.55 mol l(-1) lactic acid solution for 5 min or distilled water (control). Surface pH values, sensorial characteristics and L. monocytogenes, mesophiles and pychrotrophs counts were evaluated after treatment (day 0) and after 1, 3, 5 and 7 days of storage at 4 degrees C. Legs washed with 0.55 mol l(-1) lactic acid for 5 min showed a significant (P < 0.05) inhibitory effect on L. monocytogenes compared with control legs, being about 1.74 log units lower in the first ones than in control legs after 7 days of storage. Sensory quality was not adversely affected by lactic acid, with the exception of colour. CONCLUSIONS: Treatments with 0.55 mol l(-1) lactic acid reduced bacterial growth and preserved reasonable sensorial quality after storage at 4 degrees C for 7 days. However, it was observed a reduction in the colour score within 1 day post-treatment with 0.55 mol l(-1) lactic. SIGNIFICANCE AND IMPACT OF THE STUDY: This study demonstrates that, while lactic acid did reduce populations of L. monocytogenes on poultry, it did not completely inactivate the pathogen. The application of lactic acid may be used as an additional hurdle contributing to extend the shelf-life of raw poultry.     

Behavior of Listeria monocytogenes during fabrication and storage of experimentally contaminated smoked salmon

Experiments were carried out to examine the behavior of Listeria monocytogenes in the course of fabrication and storage of smoked salmon. In three trials, raw salmon fillets were surface inoculated with L. monocytogenes, marinated, smoked at 26 to 30 degrees C, and stored at 4 or 10 degrees C for up to 30 days. At different times during the fabrication and storage, samples were taken and, by means of the three-tube most probable number (MPN) method, quantitatively analyzed for the concentration of L. monocytogenes. The initial Listeria levels in the raw fillets were 10(4) MPN/g in trial 1, 10(1) MPN/g in trial 2, and 10(2) MPN/g in trial 3. During the fabrication, neither an increase nor a decrease of the inoculated quantities was observed. During the storage, however, a significant growth was measured in two of three trials; in trial 1, a 2.5 log10 MPN/g increase and in trial 3, an increase of even 4.5 log10 MPN/g. In the second trial, the Listeria level remained about the same. The results indicate the importance of preventing pre- and postprocessing contamination of L. monocytogenes in raw and smoked salmon. Because a significant increase of L. monocytogenes was measured during storage, there might be an increasing risk of infection for the consumer by storing such fish for a long time.     

Behavior of Listeria monocytogenes during fabrication and storage of experimentally contaminated smoked salmon.

Experiments were carried out to examine the behavior of Listeria monocytogenes in the course of fabrication and storage of smoked salmon. In three trials, raw salmon fillets were surface inoculated with L. monocytogenes, marinated, smoked at 26 to 30 degrees C, and stored at 4 or 10 degrees C for up to 30 days. At different times during the fabrication and storage, samples were taken and, by means of the three-tube most probable number (MPN) method, quantitatively analyzed for the concentration of L. monocytogenes. The initial Listeria levels in the raw fillets were 10(4) MPN/g in trial 1, 10(1) MPN/g in trial 2, and 10(2) MPN/g in trial 3. During the fabrication, neither an increase nor a decrease of the inoculated quantities was observed. During the storage, however, a significant growth was measured in two of three trials; in trial 1, a 2.5 log10 MPN/g increase and in trial 3, an increase of even 4.5 log10 MPN/g. In the second trial, the Listeria level remained about the same. The results indicate the importance of preventing pre- and postprocessing contamination of L. monocytogenes in raw and smoked salmon. Because a significant increase of L. monocytogenes was measured during storage, there might be an increasing risk of infection for the consumer by storing such fish for a long time.     

Control of Listeria spp. by competitive-exclusion bacteria in floor drains of a poultry processing plant

In previous studies workers determined that two lactic acid bacterium isolates, Lactococcus lactis subsp. lactis C-1-92 and Enterococcus durans 152 (competitive-exclusion bacteria [CE]), which were originally obtained from biofilms in floor drains, are bactericidal to Listeria monocytogenes or inhibit the growth of L. monocytogenes both in vitro and in biofilms at 4 to 37 degrees C. We evaluated the efficacy of these isolates for reducing Listeria spp. contamination of floor drains of a plant in which fresh poultry is processed. Baseline assays revealed that the mean numbers of Listeria sp. cells in floor drains sampled on six different dates (at approximately biweekly intervals) were 7.5 log(10) CFU/100 cm(2) for drain 8, 4.9 log(10) CFU/100 cm(2) for drain 3, 4.4 log(10) CFU/100 cm(2) for drain 2, 4.1 log(10) CFU/100 cm(2) for drain 4, 3.7 log(10) CFU/100 cm(2) for drain 1, and 3.6 log(10) CFU/100 cm(2) for drain 6. The drains were then treated with 10(7) CE/ml in an enzyme-foam-based cleaning agent four times in 1 week and twice a week for the following 3 weeks. In samples collected 1 week after CE treatments were applied Listeria sp. cells were not detectable (samples were negative as determined by selective enrichment culture) for drains 4 and 6 (reductions of 4.1 and 3.6 log(10) CFU/100 cm(2), respectively), and the mean numbers of Listeria sp. cells were 3.7 log(10) CFU/100 cm(2) for drain 8 (a reduction of 3.8 log(10) CFU/100 cm(2)), <1.7 log(10) CFU/100 cm(2) for drain 1 (detectable only by selective enrichment culture; a reduction of 3.3 log(10) CFU/100 cm(2)), and 2.6 log(10) CFU/100 cm(2) for drain 3 (a reduction of 2.3 log(10) CFU/100 cm(2)). However, the aerobic plate counts for samples collected from floor drains before, during, and after CE treatment remained approximately the same. The results indicate that application of the two CE can greatly reduce the number of Listeria sp. cells in floor drains at 3 to 26 degrees C in a facility in which fresh poultry is processed.     

A 20–24 H MICROCOLONY-IMMUNOBLOT TECHNIQUE to DIRECTLY DETECT and ENUMERATE LISTERIA MONOCYTOGENES INOCULATED INTO FOODS

A microcolony-immunoblot technique (MCIBI) was developed to directly enumerate, in less than 24 h, very low numbers of Listeria monocytogenes (8–12 colony forming units: CFU/g or mL) inoculated into foods. Four meat and poultry and two dairy products were artificially inoculated with L. monocytogenes V7 diluted and plated on Oxford agar medium. Each plate was overlaid with an Immobilon-P membrane and incubated for 18–20 h at 37C. Blot-transferred colonies on these membranes were probed with C11E9 monoclonal antibody (MAb) and developed using peroxidase conjugated goat antimo use Ig G and a water insoluble substrate (3,3-diaminobenzidin tetrahydmchloride; (DAB-HCI), Nickel chloride and H₂O₂). the MCIBT gave L. monocytogenes counts that were not significantly lower than direct colony counts on selective agars. This technique allowed the recovery of 94–100% of L. monocytogenes cells inoculated into foods containing natural background flora counts of 3 × 10⁴ to 8 × 10⁶ CFU/g or mL. Using a 2 h resuscitation period on nonselective agar before overlay with Oxford media, the MCIBT allowed detection of sublethally heat injured cells of strain V7.     

Destruction of Salmonella typhimurium, Escherichia coli O157:H7 and Listeria monocytogenes in chicken manure by drying and/or gassing with ammonia

Escherichia coli O157:H7 and Listeria monocytogenes were able to grow for a period of 2 days in fresh chicken manure at 20 degrees C with a resulting 1-2 log units increase in CFU; Salmonella typhimurium remained stable. Prolongation of the storage time to 6 days resulted in a 1-2 log decreases of S. typhimurium compared to the initial count and a 3-4 log decrease of E. coli O157:H7; the number of L. monocytogenes did not decrease below the initial. These changes were accompanied by an increase in pH and accumulation of ammonia in the manure. The destruction of the three microorganisms was greatly increased by drying the manure to a moisture content of 10% followed by exposure to ammonia gas in an amount of 1% of the manure wet weight; S. typhimurium and E. coli O157:H7 were reduced by 8 log units, L. monocytogenes by 4.     

Bacteriological Survey of the Blue Crab Industry

During sanitation inspections of 46 crabmeat processing plants on the Atlantic and Gulf Coasts, 487 samples of whole crabs immediately after cooking, cooked crabs after cooling, backed or washed (or both) crab bodies and whole crab claws, as well as 1,506 retail units of finished product were collected and analyzed bacteriologically. The 1,506 retail units (1-lb [373.24-g] cans) included 518 cans of regular (special) meat, 487 cans of claw meat, and 501 cans of lump meat. Statistical analyses showed that crabmeat from plants in Mississippi, Louisiana, and Texas had higher counts in 19 of 24 cases for the four bacteriological indices than crabmeat from plants located along the Atlantic Coast and the Gulf Coast of Florida. Aerobic plate counts of retail units collected from a previous day's production were significantly higher than those collected on the day of inspection. Regular crabmeat had consistently higher aerobic plate counts than claw or lump meat. When the product was handled expeditiously under good sanitary conditions, the bacteriological results were significantly better than the results from plants operating under poor sanitary conditions. Crabmeat produced in plants operating under good sanitary conditions had the following bacteriological content: (i) coliform organisms average most-probable-number values (geometric) of less than 20 per g; (ii) no Escherichia coli; (iii) coagulase-positive staphylococci average most-probable-number values (geometric) of less than 30 per g in 93% of the plants; (iv) aerobic plate count average values (geometric) of less than 100,000 per g in 93% of the plants, with the counts from 85% of these plants below 50,000 per g.     

Treatment of sanitary-important bacteria by bacteriocin substance V24 in cattle dung water

Quantification of sanitary-important bacteria (e.g. Enterobacteriaceae), as well as indicators of environmental contamination, was assessed in samples of cattle dung from 25 cattle farms in 15 north-eastern Slovakia districts. The inhibitory effect of crude bacteriocin extract CBE V24 from Enterococcus faecalis V24 against Listeria monocytogenes Ohio and Yersinia enterocolitica YE85 was examined in cattle dung water with the aim of finding a new way of eliminating the health risk of the animal slurry. The following bacterial groups were quantified: Salmonella spp., Shigella-like spp. , Proteus spp., Enterobacter spp., Citrobacter spp., Pseudomonas spp. , Escherichia coli, Listeria spp., staphylococci, streptococci and enterococci (the average count ranged from 102 up to 104 cfu ml-1). Antagonistic effect of the crude bacteriocin from Enterococcus faecalis V24 in the range of 100-600 Arbitrary units per ml (AU ml-1) was shown against the following bacteria: Enterobacter cloacae, Ent. asburiae, Proteus spp., Salmonella spp., Acinetobacter lwoffi, L. monocytogenes as well as Y. enterocolitica YE85. During tests performed to study the inhibitory effect of the crude bacteriocin CBE V24 (concentration 800, 1600 AU ml-1) against L. monocytogenes Ohio and Y. enterocolitica YE85 in experimentally contaminated cattle dung, a reduction of 2.03 and 1.44 log cfu ml-1, respectively, was already noted after 1 h after crude bacteriocin CBE V24 addition.