Effect of various gas atmospheres on the growth and survival of Campylobacter jejuni on beef

Pieces of fresh beef were inoculated with three strains of Campylobacter jejuni. The meat was then allocated to three treatments: (a) vacuum packaged, (b) packaged in an atmosphere of 20% CO2 + 80% N2, and (c) packaged into sterile Petri dishes in anaerobic cultivation boxes, which were filled with a gas mixture of 5% O2 + 10% CO2 + 85% N2. The packaging material in the first two treatments was PA 80/PE 100-PE 100/PA 80/PE 100. The survival of Campylobacter cells was followed at 37 degrees C, 20 degrees C and 4 degrees C for 48 h, 4 days and 25 days, respectively. At 37 degrees C the counts of two Campylobacter strains increased in each package treatment for 48 h. At 20 degrees C and at 4 degrees C the counts of the same two strains decreased by 1 to 2 log units and 0.5 to 1 log unit, respectively, during storage. The survival of the two strains was about the same in all package treatments. The third strain was the most sensitive of the strains studied. At 37 degrees C its numbers increased only in the optimal gas atmosphere; at 20 degrees C the strain was not detectable after 24 to 48 h storage and at 4 degrees C after 4 days storage. The aerobic plate counts were determined for all samples at the same time as Campylobacter counts. The high indigenous bacterial numbers of the meat samples did not appear to have a great effect on the survival or growth of campylobacters.     

Effects of gaseous environment and temperature on the storage behaviour of Listeria monocytogenes on chicken breast meat

Portions of skinless chicken breast meat (pH 5.8) were inoculated with a strain of Listeria monocytogenes and stored at 1, 6 or 15 degrees C in (1) aerobic conditions; (2) 30% CO2 + air; (3) 30% CO2 + N2; and (4) 100% CO2. When samples were held at 1 degree C the organism failed to grow under any of the test conditions, despite marked differences between treatments in spoilage rate and ultimate microflora. At 6 degrees C counts of L. monocytogenes increased ca 10-fold in aerobic conditions before spoilage of the meat, but only when the inoculum culture was incubated at 1 degree C rather than 37 degrees C. In CO2 atmospheres growth of L. monocytogenes was inhibited on meat held at 6 degrees C, especially under 100% CO2. By contrast, storage at 15 degrees C led to spoilage of the meat within 2 d, in all gaseous environments, and listeria levels increased up to 100-fold. Differences in the behaviour of L. monocytogenes on poultry and red meats are discussed.     

Campylobacter jejuni survival in chicken meat as a function of temperature.

Recognition of Campylobacter fetus subsp. jejuni (referred to hereafter as C. jejuni) as an important human pathogen and its isolation from meat products indicate the need for knowledge of its survival characteristics in meats. Thermal death times (D-values) for a single strain and a five-strain composite were determined in 1% peptone and autoclaved ground chicken meat at temperatures ranging from 49 to 57 degrees C. Survival was determined for these strains in chicken meat at 4, 23, 37, and 43 degrees C. Survival was also determined on raw chicken drumsticks stored at 4 degrees C in either an ambient or a CO2 atmosphere. D-values were greater in chicken meat than in peptone in all cases. D-values in peptone for strain H-840 at 49, 51, 53, 55, and 57 degrees C were 15.2, 4.90, 1.71, 0,64, and 0.25 min, respectively. The corresponding D-values in ground chicken meat were 20.5, 8.77, 4.85, 2.12, and 0.79 min, respectively. Similar results were obtained with a composite of five strains. When sterile ground chicken meat was inoculated with approximately 10(6) to 10(7) C. jejuni cells per g and stored at 37 degrees C in an ambient atmosphere, a 1-to 2-log count increase occurred during the first 4 days, followed by a gradual decline of about 1 log during the remainder of the 17-day storage period. No growth was observed among similarly inoculated samples that were stored at 4, 23, and 43 degrees C but counts declined by about 1 to 2 logs at 4 degrees C (17 day), by 2.5 to 5 logs at 23 degrees C (17 days), and to undetectable levels at 43 degrees C (between 10 and 16 days). Survival on raw chicken drumsticks stored at 4 degrees C in CO2 and in an ambient atmosphere declined by about 1.5 and 2.0 logs, respectively, during 21 days of storage. The effect of temperature on the survival of C. jejuni in chicken meat was similar to that reported in other natural and laboratory milieus. Ordinary cooking procedures that destroy salmonellae would be expected to destroy C. jejuni.     

Effect of various gas atmospheres on the growth and survival of Campylobacter jejuni on beef

Pieces of fresh beef were inoculated with three strains of Campylobacter jejuni . The meat was then allocated to three treatments: (a) vacuum packaged, (b) packaged in an atmosphere of 20% CO₂+ 80% N₂, and (c) packaged into sterile Petri dishes in anaerobic cultivation boxes, which were filled with a gas mixture of 5% O₂+ 10% CO₂+ 85% N₂. The packaging material in the first two treatments was PA 80/PE 100–PE 100/PA 80/PE 100. The survival of Campylobacter cells was followed at 37°C, 20°C and 4°C for 48 h, 4 days and 25 days, respectively. At 37°C the counts of two Campylobacter strains increased in each package treatment for 48 h. At 20°C and at 4°C the counts of the same two strains decreased by 1 to 2 log units and 0.5 to 1 log unit, respectively, during storage. The survival of the two strains was about the same in all package treatments. The third strain was the most sensitive of the strains studied. At 37°C its numbers increased only in the optimal gas atmosphere; at 20°C the strain was not detectable after 24 to 48 h storage and at 4°C after 4 days storage. The aerobic plate counts were determined for all samples at the same time as Campylobacter counts. The high indigenous bacterial numbers of the meat samples did not appear to have a great effect on the survival or growth of campylobacters.     

A study of the growth kinetics of Yersinia enterocolitica serotype O:3 in pure and meat culture systems

The growth kinetics of a virulence plasmid-bearing (P+) and a plasmid-cured (P−) strain of Yersinia enterocolitica serotype O:3 in pure and meat culture were investigated. Growth studies were carried out at 25 and 37 °C in supplemented phosphate-buffered saline, buffered peptone water , cefsulodin-irgasan-novobiocin broth base or supplemented broth base (CIN). The lag phase durations and growth rates under these conditions were determined by linear regression analysis. In pure culture, under most sets of equivalent conditions, P+ and P− strains had similar lag phase durations. However, under one set of conditions, i.e. CIN broth at 37 °C, the lag phase duration of the P+ strain was significantly longer than P−. In all but the most selective medium, P+ strains had slower growth rates than P− strains at 37 °C, probably due to the increased metabolic burden entailed in the maintenance of the virulence plasmid. In the most selective medium, i.e. CIN broth, P+ strains grew significantly faster than P−. This finding suggests that possession of virulence plasmid confers an enhanced ability to grow in the presence of selective agents. In meat cultures, both strains had longer lag phases than in equivalent pure cultures, with longer lag phases noted at 37 than at 25 °C. No significant differences were observed between the length of lag phases of P+ and P− strains in meat culture. Both strains of Y. enterocolitica displayed faster growth rates in meat cultures than in pure cultures, indicating that one or more components of meat enhanced the growth of this organism. The effects and interaction of incubation temperature, enrichment broth and meat on the growth kinetics of plasmid-bearing and plasmid-cured Y. enterocolitica strains are discussed.     

Isolation of Campylobacter spp. from pigeon feces by a combined enrichment-filtration technique.

A technique combining enrichment in Preston enrichment broth and direct filtration onto chocolate agar was used to isolate Campylobacter species from pigeon feces. Campylobacter jejuni was isolated from 106 of 200 samples tested; 105 strains were isolated by enrichment-filtration, and 84 strains were isolated by direct plating. Most of the strains grew after 48 h at 37 degrees C.     

Effect of temperature, sodium chloride, and pH on growth of Listeria monocytogenes in cabbage juice.

Human illness and death have resulted from the consumption of milk, cheese, and cole slaw contaminated with Listeria monocytogenes. Since the effects of temperature, NaCl, and pH on the growth of the organism in cabbage were unknown, a series of experiments was designed to investigate these factors. Two strains (LCDC 81-861 and Scott A, both serotype 4b) were examined. At 30 degrees C, the viable population of the LCDC 81-861 strain increased in sterile unclarified cabbage juice (CJ) containing 0 to 1.5% NaCl; a decrease in the population of both strains occurred in juice containing greater than or equal to 2% NaCl. At 5 degrees C, the population of the Scott A strain in CJ containing up to 5% NaCl was reduced by about 90% over a 70-day period; the LCDC 81-861 strain was more sensitive to refrigeration but remained viable in CJ containing less than or equal to 3.5% NaCl for 70 days. Growth in CJ at 30 degrees C resulted in a decrease in pH from 5.6 to 4.1 within 8 days. Death of L. monocytogenes occurred at 30 degrees C when the organism was inoculated into sterile CJ adjusted to pH less than or equal to 4.6 with lactic acid. No viable cells were detected after 3 days at pH less than or equal to 4.2. At 5 degrees C, the rate of death at pH less than or equal to 4.8 was slower than at 30 degrees C.(ABSTRACT TRUNCATED AT 250 WORDS)     

Isolation of Campylobacter spp. from pigeon feces by a combined enrichment-filtration technique

A technique combining enrichment in Preston enrichment broth and direct filtration onto chocolate agar was used to isolate Campylobacter species from pigeon feces. Campylobacter jejuni was isolated from 106 of 200 samples tested; 105 strains were isolated by enrichment-filtration, and 84 strains were isolated by direct plating. Most of the strains grew after 48 h at 37 degrees C.     

Mycoplasma somnilux sp. nov., Mycoplasma luminosum sp. nov., and Mycoplasma lucivorax sp. nov., new sterol-requiring mollicutes from firefly beetles (Coleoptera: Lampyridae)

Strain PYAN-1T (T = type strain), which was isolated from a pupal gut of the firefly beetle Pyractonema angulata, and strains PIMN-1T and PIPN-2T, which were isolated from guts of adult Photinus marginalis and Photinus pyralis fireflies, respectively, were demonstrated to be sterol-requiring mollicutes. Cells of the three strains were shown by electron and dark-field microscopy to be small, pleomorphic, nonhelical, nonmotile bodies surrounded by single membranes. No evidence of a cell wall was observed, and the organisms were not susceptible to 500 U of penicillin per ml. The three strains grew rapidly in SP-4 broth medium. Strains PIMN-1T and PIPN-2T grew in medium supplemented with bovine serum fraction, but strain PYAN-1T did not. All three strains grew on solid media when the cultures were incubated aerobically, but only strains PYAN-1T and PIPN-2T formed colonies when anaerobic conditions were employed. The three strains catabolized glucose but hydrolyzed neither arginine nor urea. All of the strains grew at temperatures of 18 to 32 degrees C; strains PYAN-1T and PIMN-1T also grew at 10 degrees C. The optimal temperature for growth for strains PYAN-1T and PIPN-2T was 30 degrees C; strain PIMN-1T grew equally well at 30 or 32 degrees C. None of the three strains grew at 37 degrees C. The genome sizes of strains PYAN-1T, PIMN-1T, and PIPN-2T were about 527 (478 to 589), 570 (480 to 630), and 762 (635 to 871) megadaltons, respectively.(ABSTRACT TRUNCATED AT 250 WORDS)     

Fate of Listeria monocytogenes in processed meat products during refrigerated storage.

The fate of Listeria monocytogenes during refrigerated storage was determined on several processed meat products, including ham, bologna, wieners, sliced chicken, sliced turkey, fermented semidried sausage, bratwurst, and cooked roast beef. The meats were surface inoculated with a five-strain mixture of less than or equal to 200 or ca. 10(5) L. monocytogenes cells per package, vacuum packaged, and stored at 4.4 degrees C. Survival or growth of listeriae was determined for up to 12 weeks of storage or until the product was spoiled. The organism survived but did not grow on summer sausage, grew only slightly on cooked roast beef, grew well on some wiener products but not on others, grew well (10(3) to 10(5) CFU/g increase within 4 weeks) on ham, bologna, and bratwurst, and grew exceptionally well (10(3) to 10(5) CFU/g increase within 4 weeks) on sliced chicken and turkey. The rate of growth depended largely upon the type of product and the pH of the product. Growth was most prolific on processed poultry products. The organism generally grew well on meats near or above pH 6 and poorly or not at all on products near or below pH 5. These results indicate the importance of preventing postprocessing contamination of L. monocytogenes in a variety of ready-to-eat meat products.     

Efficacy of media and methods for detecting and enumerating Campylobacter jejuni in refrigerated chicken meat.

A study was undertaken to compare several enrichment and direct isolation media for their suitability to detect and enumerate five strains of Campylobacter jejuni in refrigerated (5 degrees C) chicken meat. The influence of CO2 on survival at 5 degrees C was also investigated. Selective enrichment media evaluated included Preston broth (PB), selective semisolid brucella medium (SSBM), Campylobacter enrichment broth (CEB), VTP brucella-FBP broth (VTP), Rosef and Kapperud Campylobacter enrichment broth (RKCEB), and Doyle and Roman enrichment broth (DREB). Direct isolation agars included Campy brucella agar (CBAP), blood-free Campylobacter medium (BFCM) and modified Butzler agar (MBA). Comminuted chicken meat was inoculated with C. jejuni, sealed under atmospheric gas or CO2, and stored at 5 degrees C for up to 21 days. Viable population was determined by the most-probable-number technique (PB, SSBM, CEB, VTP, and RKCEB, followed by plating on CBAP, BFCM, and MBA), enrichment on DREB, followed by plating on CBAP, BFCM, and MBA, and direct isolation on CBAP, BFCM, and MBA. Without exception, direct plating of samples was superior to the most-probable-number technique for enumerating C. jejuni; MBA was inferior to CBAP and BFCM, and DREB performed at least as well as other enrichment media evaluated. Carbon dioxide afforded protection against death of three of the five strains of C. jejuni tested.     

Effect of temperature, sodium chloride, and pH on growth of Listeria monocytogenes in cabbage juice

Human illness and death have resulted from the consumption of milk, cheese, and cole slaw contaminated with Listeria monocytogenes. Since the effects of temperature, NaCl, and pH on the growth of the organism in cabbage were unknown, a series of experiments was designed to investigate these factors. Two strains (LCDC 81-861 and Scott A, both serotype 4b) were examined. At 30 degrees C, the viable population of the LCDC 81-861 strain increased in sterile unclarified cabbage juice (CJ) containing 0 to 1.5% NaCl; a decrease in the population of both strains occurred in juice containing greater than or equal to 2% NaCl. At 5 degrees C, the population of the Scott A strain in CJ containing up to 5% NaCl was reduced by about 90% over a 70-day period; the LCDC 81-861 strain was more sensitive to refrigeration but remained viable in CJ containing less than or equal to 3.5% NaCl for 70 days. Growth in CJ at 30 degrees C resulted in a decrease in pH from 5.6 to 4.1 within 8 days. Death of L. monocytogenes occurred at 30 degrees C when the organism was inoculated into sterile CJ adjusted to pH less than or equal to 4.6 with lactic acid. No viable cells were detected after 3 days at pH less than or equal to 4.2. At 5 degrees C, the rate of death at pH less than or equal to 4.8 was slower than at 30 degrees C.(ABSTRACT TRUNCATED AT 250 WORDS)     

Enteropathogenic bacteria in frozen chicken.

Eighty-two samples of frozen chicken from retail stores were examined for the presence of Campylobacter, Yersinia enterocolitica, and salmonellae. Aerobic plate counts and numbers of coliform bacteria at 37 degrees C were determined. Campylobacter fetus subsp. jejuni was found in 22% of the samples, Y. enterocolitica was found in 24.5% and Salmonella typhimurium was found in one sample (1.2%). The isolated strains of Y. enterocolitica belonged to serotypes 4, 5b, 6, and 8. Aerobic plate counts and numbers of coliform bacteria at 37 degrees C were not found to be noticeably higher in samples containing pathogens than in pathogen-free samples. This investigation showed that chicken does contain other pathogenic bacteria than salmonellae. Campylobacter and Y. enterocolitica were isolated in much higher frequencies than Salmonella.     

Spoilage-related activity of Carnobacterium maltaromaticum strains in air-stored and vacuum-packed meat

One hundred three isolates of Carnobacterium spp. from raw meat were analyzed by random amplification of polymorphic DNA (RAPD) and PCR and were identified by 16S rRNA gene sequencing. Forty-five strains of Carnobacterium maltaromaticum were characterized for their growth capabilities at different temperatures, NaCl concentrations, and pH values and for in vitro lipolytic and proteolytic activities. Moreover, their spoilage potential in meat was investigated by analyzing the release of volatile organic compounds (VOCs) in meat stored in air or vacuum packs. Almost all the strains were able to grow at 4, 10, and 20°C, at pH values of 6 to 9, and in the presence of 2.5% NaCl. The release of VOCs by each strain in beef stored at 4°C in air and vacuum packs was evaluated by headspace solid-phase microextraction (HS-SPME)-gas chromatography-mass spectrometry (GC-MS) analysis. All the meat samples inoculated and stored in air showed higher numbers of VOCs than the vacuum-packed meat samples. Acetoin, 1-octen-3-ol, and butanoic acid were the compounds most frequently found under both storage conditions. The contaminated meat samples were evaluated by a sensory panel; the results indicated that for all sensory odors, no effect of strain was significant (P > 0.05). The storage conditions significantly affected (P < 0.05) the perception of dairy, spoiled-meat, and mozzarella cheese odors, which were more intense in meat stored in air than in vacuum packs but were never very intense. In conclusion, different strains of C. maltaromaticum can grow efficiently in meat stored at low temperatures both in air and in vacuum packs, producing volatile molecules with low sensory impacts, with a negligible contribution to meat spoilage overall.     

Determinants in microbial colonization of the murine gastrointestinal tract: pH, temperature, and energy-yielding metabolism of Torulopsis pintolopesii.

Torulopsis pintolopesii is an indigenous yeast that colonizes the secreting epithelia in the stomachs of mice and rats. A wild-type strain of this microbe was isolated and identified. To attempt to learn characteristics of the yeast that are advantageous to it in colonizing its natural habitat in vivo, we examined some aspects of its nutrition and energy-yielding metabolism and some environmental conditions that influence its growth in vitro. The yeast appeared to be limited in the compounds it can utilize as carbon and nitrogen sources. It grew best at 37 degrees C and did not grow at 23 or 43 degrees C. It grew optimally at neutral pH but could grow aerobically at pH values as low as 2.0 and anaerobically at pH values as low as 3.4. As assessed by measurements of growth rates and yield coefficients, it grew better aerobically than anaerobically. When grown aerobically, it had a cyanide-sensitive system for taking up O(2) and tested positively for cytochrome c oxidase activity. A petite mutant strain isolated from the wild-type strain had a growth rate and yield coefficient when incubated aerobically that were essentially the same as those of the wild-type parent grown anaerobically. Likewise similar to the wild-type parent grown anaerobically, the petite strain, though incubated aerobically, did not take up O(2). Yeast-free mice associated with either the wild-type or the petite mutant strain were colonized at essentially the same rates and to similar final population levels by both strains. The yeast's capacity to respire may be of little advantage to it in its natural environment. By contrast, its abilities to grow best at 37 degrees C and to grow at low pH values are undoubtedly advantageous characteristics in this respect. The limitations in its carbon and nitrogen nutrition are difficult to evaluate as ecological factors in its colonization of the natural habitat.     

Survival of Campylobacter jejuni on beef trimmings during freezing and frozen storage

AIMS: To investigate the survival of two animal isolates of Campylobacter jejuni on beef trimmings during freezing and frozen storage. METHODS AND RESULTS: Meat packs inoculated with 10(3) or 10(6) cfu g(-1) of either strain of C. jejuni were frozen to -18 degrees C, and sampled at regular intervals over 112 d storage to determine Campylobacter numbers and sublethal injury. For both strains and inoculation levels the numbers of Campylobacter decreased in the first 7 d of storage by ca. 0.6-2.2 log cfu g(-1) and then remaining constant over the remainder of the storage trial, with neither isolate exhibiting sublethal injury. CONCLUSIONS: Despite an initially significant decrease in number, these pathogens were able to survive standard freezing conditions in meat, but did not exhibit sublethal injury. SIGNIFICANCE AND IMPACT OF THE STUDY: Strict hygiene and/or the implementation of decontamination technologies are recommended as a means to assure the safety of meat with respect to this pathogen.     

Efficacy of media and methods for detecting and enumerating Campylobacter jejuni in refrigerated chicken meat

A study was undertaken to compare several enrichment and direct isolation media for their suitability to detect and enumerate five strains of Campylobacter jejuni in refrigerated (5 degrees C) chicken meat. The influence of CO2 on survival at 5 degrees C was also investigated. Selective enrichment media evaluated included Preston broth (PB), selective semisolid brucella medium (SSBM), Campylobacter enrichment broth (CEB), VTP brucella-FBP broth (VTP), Rosef and Kapperud Campylobacter enrichment broth (RKCEB), and Doyle and Roman enrichment broth (DREB). Direct isolation agars included Campy brucella agar (CBAP), blood-free Campylobacter medium (BFCM) and modified Butzler agar (MBA). Comminuted chicken meat was inoculated with C. jejuni, sealed under atmospheric gas or CO2, and stored at 5 degrees C for up to 21 days. Viable population was determined by the most-probable-number technique (PB, SSBM, CEB, VTP, and RKCEB, followed by plating on CBAP, BFCM, and MBA), enrichment on DREB, followed by plating on CBAP, BFCM, and MBA, and direct isolation on CBAP, BFCM, and MBA. Without exception, direct plating of samples was superior to the most-probable-number technique for enumerating C. jejuni; MBA was inferior to CBAP and BFCM, and DREB performed at least as well as other enrichment media evaluated. Carbon dioxide afforded protection against death of three of the five strains of C. jejuni tested.     

The protective effect of some food ingredients on Staphylococcus aureus MF31

The upper limiting temperature of growth of Staphylococcus aureus MF31 in heart infusion broth (HI) was about 44 degrees C but addition of monosodium glutamate (MSG) and soy sauce permitted the organism to grow above this temperature. This effect is similar to that of NaCl. Tomato ketchup, Worcestershire and HP sauces added to HI did not allow growth at the non-permissive temperature of 46 degrees C but death was delayed. Staphylococcus aureus died in unsupplemented chicken meat slurry at 46 degrees C but grew at 48 degrees C in slurry supplemented with 5.8% NaCl and survived incubation for 18 h at 50 degrees C in slurry supplemented with 5.8% NaCl and 5% MSG. Cultures grown at 37 degrees C had a D60 value of 2 min in 50 mmol/l Tris (pH 7.2) buffer. Cultures grown at 46 degrees C in HI containing 5.8% NaCl had a D60 value of 8 min in Tris buffer. Addition of 5.8% NaCl plus 5% MSG to the buffer increased the D60 by a factor of about 7 for both cultures. In storage experiments at room temperature, the culture grown at 37 degrees C and at 46 degrees C plus 5.8% NaCl died at about the same rate in salami. In milk powder, however, the count of 37 degrees C culture decreased from 10% g to 10(6)/g in 5 weeks while the count of 46 degrees C culture remained unchanged. In cottage cheese, freeze-dried rice and macaroni, the 37 degrees C cultures also died more rapidly. It is suggested that cultures grown at 46 degrees C plus 5.8% NaCl may be suitable for experiments with artificially contaminated foods.     

Studies on the adaptation of influenza virus replicated at low temperature. III. Biochemical studies.

Five strains of influenza viruses A(H3N2) replicated at low temperature passaged in cotton rats were reisolated. The properties of these strains replicated at low temperature were compared before and after passage in susceptible animals to check the stability of some its markers. At the same time original viruses replicated at 37 degrees C--which are different in epidemiological potency--were compared. The following parameters being tested: NA activity, HA titers, heat inactivation NA and Ha, Michaelis constants and optimum pH. We observed some differences between strains both replicated at low temperature after passage in the susceptible animal organism and original viruses from 37 degrees C. Viruses replicated at low temperature from original epidemiostrain are really cold adapted and remained stable after passage in the animals when the others derived from no epidemic strain are not stable.     

The Growth of Microbacterium thermosphactum on Beef

Microbacterium thermosphactum grew aerobically at 5°C on beef when the pH was as low as 5.4. It did not grow anaerobically if the pH of the meat was below 5.8, but grew readily at pH 6.0 or higher. On vacuum-packaged beef of pH 5.8 or lower, growth was dependent not only on pH but also on the oxygen permeability of the packaging film. Above pH 5.9 growth occurred with all the films tested.