Bryan D. Dilts

Control of Brochothrix thermosphacta Spoilage of Pork Adipose Tissue Using Bacteriophages

Adipose tissue discs were coinoculated with Brochothrix thermosphacta and homologous bacteriophages (phages) to determine the effects these had on phage multiplication, bacterial growth, and off-odor development during storage at 2 degrees C or under simulated retail display at 6 degrees C. In the presence of about 10(5) bacteria/cm2 and an equivalent number of phages, there was a 3-log increase in phage numbers and a 2-log decrease in bacterial numbers, and objectionable off-odors were suppressed during refrigerated storage. Up to 68% of the surviving bacterial population were resistant to phages. The storage life of adipose tissue could be increased from 4 days in controls to 8 days in phage-treated samples by preventing the development of off-odors associated with the growth of B. thermosphacta. Phages may provide a novel approach to extending the storage quality of chilled meats.

Inability of a bacteriophage pool to control beef spoilage

The biological control of beef spoilage, with a bacteriophage (phage) pool, was evaluated under simulated retail conditions. A pool of seven phages was selected with the potential to lyse 78% of 86 Pseudomonas test strains. Subsequent host range studies with 1023 pseudomonads from three meat species (beef, pork, lamb) and five abattoirs showed that 585 (57.2%) isolates were susceptible to the phage pool. Depending on bacterial origin, bacterial sensitivity to lysis by the phage pool varied from 25 to 72%. When added to ribeye steaks, the phage pool produced a significant reduction in Pseudomonas growth but this was not sufficient to produce any significant effect upon the retail shelf life of beef. The inability of phages to control beef spoilage was not attributed to a loss of phage virulence since sufficient densities (log pfu/cm2 = 5 to 6) of virulent phage could be re-isolated from beef, 14 days after treatment. It was concluded that the efficacy of the current phage pool was limited by a narrow range of specificity.

Lactic acid inhibition of the growth of spoilage bacteria and cold tolerant pathogens on pork

The antibacterial effects of a 3% solution of lactic acid at 55 degrees C were assessed, by examining aerobic bacterial growth on artificially-inoculated pork fat and lean tissue. Discs of fat or lean tissues, each of 10 cm2 surface area, were aseptically excised from pork Longissimus dorsi muscle and inoculated with the cold tolerant pathogens Listeria monocytogenes 4b Scott A no. 3, Yersinia enterocolitica 0:4,32 or Aeromonas hydrophila ATCC 7966, or with the wild type spoilage bacteria Pseudomonas fragi or Brochothrix thermosphacta. After inoculation, each meat disc was immersed in water or lactic acid for 15 s and aerobic bacterial growth followed during 15 days of storage at 4 degrees C. P. fragi and B. thermosphacta grew on both fat and lean, but the pathogens grew on fat tissue only and A. hydrophila did not survive on lean. Lactic acid reduced all test bacteria on fat to below detectable levels within 4 days of treatment and no bacteria could be recovered from acid-treated fat surfaces for the remainder of the 15-day storage interval. Bacteria attached to lean were generally more resistant to lactic acid. In some instances the acid was bacteriostatic (P. fragi, L. monocytogenes) while in others the population declined at a greatly reduced rate as compared with a similar population on fat (B. thermosphacta, Y. enterocolitica). A. hydrophila was equally sensitive to lactic acid on lean and fat. Depending upon the tested strain, tissue type and storage time, maximum reductions in the number of bacteria recovered from acid treated pork ranged from 1 to 8 log cycles. The high bactericidal efficacy of lactic acid applied to pork fat was attributable to a low tissue pH, which varied from 3.49 to 4.41 during the 15 days of aerobic storage.

Effect of growth of selected lactic acid bacteria on storage life of beef stored under vacuum and in air.

The effect of growth of different types of lactic acid bacteria (LAB) on the storage life of normal pH beef was determined anaerobically (under vacuum) and aerobically. Four LAB from meat were inoculated separately onto sterile slices of lean beef. Inoculated samples were stored anaerobically at 2 degrees C for 10 weeks or stored aerobically in an oxygen permeable film at 7 degrees C for 10 days, with and without previous storage under vacuum at 2 degrees C. The LAB strains used were Carnobacterium maltaromicus (previously C. piscicola) LV17 and UAL26, Leuconostoc gelidum UAL187-22 and Lactobacillus sake Lb706. Storage life was determined by sensory panel evaluation of colour and odour. Under anaerobic conditions Lb. sake Lb706, inoculated at log 2 CFU/cm2, grew rapidly to reach maximum population within three weeks of storage. L. gelidum UAL187-22 also grew on the meat but at a slower rate. In contrast, growth of C. maltaromicus LV17 and UAL26 was unpredictable, achieving maximum population after 2 to 8 weeks. None of the test strains caused spoilage of the meat within the 10-week storage period under vacuum. When the test organisms were inoculated at an initial level of log 4 CFU/cm2, C. maltaromicus LV17 and UAL26 produced off-odours after 8 weeks of storage under vacuum at 2 degrees C. Under aerobic conditions at 7 degrees C, all four of the strains grew well on the beef samples. C. maltaromicus LV17 and UAL26 and Lb. sake Lb706 all caused off-odours and discoloration. The rate of aerobic deterioration in meat quality was faster with increased time of storage under vacuum. L. gelidum UAL187-22 could be a suitable antagonistic strain with the potential to extend the storage life of beef, stored anaerobically and packaged aerobically for retail sale, without producing undesirable sensory changes.

Factors affecting the susceptibility of meatborne pathogens and spoilage bacteria to organic acids

Abstract The bactericidal effects of lactic and acetic acids were assessed on artificially inoculated lean beef. The test bacteria included cold tolerant pathogens ( Listeria monocytogenes, Yersinia enterocolitica ), mesophilic pathogens ( Salmonella typhimurium, Escherichia coli, Campylobacter jejuni and Staphylococcus aureus ) and spoilage bacteria ( Pseudomonas fragi and Brochothrix thermosphacta ). Acid temperature (20°C, 55°C), acid concentration (1%, 3%) and initial numbers of contaminating bacteria (log CFU/cm 2 of 3–6) were the variables studied. The bactericidal efficacy of organic acids was influenced by two- and three-way interactive effects that were often distinct for each organism. The magnitude of the reductions in bacterial numbers on acid-treated beef was similar with lactic and acetic acids. Bacterial numbers were maximally reduced with 3% acid at 55°C. The variations in the reductions in the numbers of bacteria on acid-treated beef were significantly affected by initial levels of bacterial contamination, but this effect followed no consistent pattern and varied with bacterial species. Of the pathogens tested, S. aureus was the most susceptible to acid (1·4 log cycle reduction) and S. typhimurium the least susceptible (0·4 log cycle reduction). The spoilage bacteria P. fragi (2·3 log cycle reduction) and B. thermosphacta (3·5 log cycle reduction) were more sensitive to the lethal effects of acid treatment than the pathogens.

Bacteriology and Retail Case Life of Pork After Storage in Carbon Dioxide

The effects of prolonged, anoxic storage, under CO2 at -1.5°C, upon the bacteriology and case life of pork on its subsequent transfer to the aerobic conditions of simulated retail display at 8°C was examined. Brochothrix thermosphacta , lactic acid bacteria, enterics, and pseudomonads were enumerated. Panel scores for odor and appearance acceptability were used to quantify retail case life. Lactic acid bacteria were the only bacteria found during loin storage in CO2 for up to 24 weeks. Those organisms reached maximum number of 107 CFU/cm2 within 9 weeks. The number of lactic acid bacteria initially found on the freshly cut surfaces of loin chops increased linearly during the first 9 weeks of loin storage in CO2. Thereafter, they continued to grow on the chops and dominated the spoilage flora during retail display. The pseudomonads grew rapidly and emerged as the next most numerous organism, while B. thermosphacta and enterics showed only limited aerobic growth. The acceptability of pork chop appearance and odor was adversely affected by loin storage time. Each 6-week interval of loin storage produced a 1 d reduction in case life. Should controlled atmospheres be a practicable means of meat distribution to the retail marketplace, efforts will be necessary to assure a maximum case life after their removal from preservative packagings.

Bacterial contamination of recirculating brine used in the commercial production of moisture-enhanced pork.

In a commercial process for the production of moisture-enhanced pork, boneless pork loins were conveyed through a recirculating injection apparatus, and brine (sodium phosphate, sodium chloride, and lemon juice solids) was pumped into the meat through banks of needles inserted automatically into the upper surfaces of cuts. Brine samples were collected at intervals during the production process and analyzed to determine the total plate count and the numbers of lactic acid bacteria, pseudomonads, Brochothrix thermosphacta, and Enterobacteriaceae. Listeria monocytogenes numbers in the brine were determined using a PCR with primers for the hemolysin gene in combination with a most probable numbers determination. Maximum numbers of bacteria (log CFU/ml) recovered from the brine after 2.5 h of recirculation were as follows: total plate count, 4.50; lactic acid bacteria, 2.99; pseudomonads, 3.95; B. thermosphacta, 2.79; and enterics, 3.01. There was an increase in the number of L. monocytogenes in the recirculating brine with time, reaching a maximum of 2.34 log CFU/100 ml after 2.5 h of moisture-enhanced pork production. Thus, recirculating brines can harbor large populations of spoilage bacteria and L. monocytogenes and are an important source of contamination for moisture-enhanced pork.

The aerobic growth of Aeromonas hydrophila and Listeria monocytogenes in broths and on pork.

Flasks of tryptic soy broth (TSB), unacidified (pH 7.2) or acidified with HCl or lactic acid to pH 6.3 or 5.5, and samples of sterile pork fat or muscle tissue, were inoculated with logarithmic phase cultures of a strain of Aeromonas hydrophila or a strain of Listeria monocytogenes. The broth cultures were incubated at temperatures between 0 and 25 degrees C, and growth rates were determined from optical density increases. The tissue samples were incubated at temperatures between -2.4 and 25.2 degrees C, and growth rates were determined from viable count increases. Both organisms grew without lag in all broths at temperatures greater than 10 degrees C. A hydrophila did not grow at 5 degrees C in TSB acidified with lactic acid to pH 5.5, and grew in other broths at that temperature after a lag of about 10 h. The organism did not grow in either broth of pH 5.5 at 2 degrees C, but grew in other broths at that temperature after a lag of about 40 h. A hydrophila did not grow in any broth at 0 degrees C. L. monocytogenes grew in all broths at 5 degrees C only after a lag of about 60 h, and did not grow in any broth at 2 degrees C. For both organisms, the rates of growth, at any temperature, were lower in broths of pH 6.3, and lower again in broths of pH 5.5, than in the unacidified broth. Growth rates in the broths of pH 6.3 were similar, but growth rates were lower in lactic acid acidified broth of pH 5.5 than in HCl acidified broth of that pH. The data for the growth of each organism in each medium were well described by the regression line of the plot of the square roots of growth rates against temperature. A hydrophila grew on fat tissue of pH 6.3 +/- 0.3, without lag, at 1.8 degrees C and higher temperatures at rates greater than the rates of growth in unacidified TSB. Numbers of the organism declined on muscle tissue of pH 5.6 +/0 0.2 at any temperature. L. monocytogenes grew on fat tissue without lag at -0.3 degrees C and higher temperatures, at rates which at lower temperatures were greater than the rates of growth in unacidified TSB. The organism grew on muscle tissue only at temperatures greater than or equal to 15.4 degrees C, at rates which were less than the rates of growth in lactic acid acidified broth of pH 5.5. Models derived from the cultivation of A. hydrophila and L. monocytogenes in commercial broths appear to be highly unreliable guides to the behaviours of those organisms on pork.

Evaluation of the bacteriological consequences of the temperature regimes experienced by fresh chilled meat during retail display

Temperature histories were recorded from the surfaces of chilled steaks in a commercial retail case. Steaks were placed at the rear, centre or front of the case, and at 17 or 13 cm below, or 4 cm above the load line. The coldest average temperatures (1·7-4·5°C) were recorded at the rear, and the warmest (6·1–10°C) at the centre of the case. Average temperatures increased with upward location of the steaks, from between 1·7 and 6·1°C at 17 cm below to between 4·5 and 10°C at 4 cm above the load line. The meat surface temperatures did not correlate with the air temperatures at the rack level. The average temperatures for steak surfaces and case air in the vicinity of steaks differed by up to 1°C, but the surface and deep temperatures of steaks were similar. Calculated values for the proliferations of psychrotrophic pseudomonads and Escherichia coli correlated well with the average surface temperatures of the steaks. Apparently, E. coli growth would be negligible with average surface temperatures of ≤4°C. However, a survey of air temperatures in four commercial retail cases indicated that temperatures of ≤4°C cannot be maintained throughout existing retail cabinets. To assure food safety, some means of identifying the maximum average temperature experienced by a product in individual display cases is required, with a specification of the maximum residence time that can be tolerated at any maximum temperature.

Survival of Campylobacter jejuni on beef and pork under vacuum packaged and retail storage conditions: Examination of the role of natural meat microflora on C. jejuni survival

The ability of Campylobacter jejuni ATCC 11168 to survive on beef and pork stored under chilled, vacuum packaged and retail display conditions were examined. In addition, the effect of natural microflora on commercial beef and pork on the survival of C. jejuni under these storage conditions was examined. When sterile cores of beef and pork were inoculated with ∼ 10(5) to 10(6) cfu cm(-2)C. jejuni, and were stored under aerobic or vacuum packaged conditions at -1.5 or 4 °C, its numbers dropped significantly and C. jejuni could not be enumerated by direct plating after 21 d of the 6 wks study. In contrast, survival of C. jejuni on commercial vacuum packaged beef and pork was significantly enhanced, resulting in only 1 log cfu cm(-2) reduction at the end of 6 wks. During 7 d of display in a retail case, numbers of C. jejuni dropped quickly, but could be enumerated by direct plating even after the 7 d. The presence of high numbers of inoculated C. jejuni on beef and pork had no significant effect on the natural microflora numbers compared to uninoculated controls when the meat was stored either in vacuum or in a retail display case. These results show that natural microflora on vacuum packaged meat afford enhanced survival of C. jejuni present on the surfaces of both beef and pork when stored at refrigeration temperatures. Hence, strict hygienic practices or the implementation of decontamination technologies are recommended to ensure safety of meat with respect to this pathogen.