Chemical and Microbiological Changes during Storage of Vacuum Packed Sliced Bacon

Summary: The micro-ecology of block cured, sliced, vacuum packed bacon has been studied during storage at 20 and 30°; high salt (8–12% NaCl, based on the aqueous phase) and low salt (5–7% NaCl) bacon were used. The dominant flora of all samples during the first 9 days' storage comprised catalase positive cocci. They persisted in the high salt bacon but, under lower salt conditions, group D streptococci and motile lactic acid bacteria became dominant.
Of the chemical changes studied, proteolysis, lipolysis and reduction of nitrate and nitrite were due largely to coagulase negative subgroup II staphylococci. Micrococci which predominated at 20° were capable of reducing nitrate to nitrite but of only slightly increasing free amino or fatty acids.
Inoculation of low count bacon with subgroup II staphylococci confirmed that this group was responsible for putrefaction in low salt bacon stored at temperatures above 20°. Similarly micrococci which tend to delay spoilage did not contribute much to the off odours and it was thought that the heterofermentative catalase negative organisms could contribute to typical sour spoilage.     

A numerical taxonomic study of lactic acid bacteria from vacuum-packed beef, pork, lamb and bacon

S haw B. G. & H arding C harmaigne D. 1984. A numerical taxonomic study of lactic acid bacteria from vacuum packed beef, pork, lamb and bacon. Journal of Applied Bacteriology 56 , 25–40.
A numerical taxonomic study using 79 unit characters has been performed on 100 isolates of lactic acid bacteria from refrigerated vacuum-packed beef, pork, lamb and bacon. Three clusters were observed at 78% S which contained all the strains apart from three unidentifiable streptobacteria, one Leuconostoc , and one strain of Pediococcus pentosaceus . One cluster (III) consisted of only one strain of Leuc. paramesenteroides and six unidentifiable Leuconostoc strains. The two largest clusters (I and II) were both composed entirely of streptobacteria. Cluster I contained 31 strains (G + C content 33–2–36–9 moles %) which were not identifiable with any described species. Cluster II contained 57 strains (G + C content 40–7–43–7 moles %) which were provisionally identified with Lactobacillus sake or Lact. bavaricus according to the lactic acid isomer produced. The division of nearly all the streptobacteria into two clearly defined clusters has resolved problems which have existed in the classification of lactic acid bacteria from vacuum-packed meat.     

The D(-)lactic acid and acetoin/diacetyl as potential indicators of the microbial quality of vacuum-packed pork and meat products

The suitability of D(-)lactic acid and/or acetoin/diacetyl as indicators of spoilage of vacuum-packed meat and meat products has been studied. When pork was vacuum-packed, these substances reached measurable amounts after storage for only about 10 days. Although microbial counts stabilized from the 20th day of storage, the D(-)lactic acid and acetoin/diacetyl concentrations increased progressively. These substances could therefore be potential indicators of the storage time of vacuum-packed pork. From a survey carried out with several vacuum-packed meat products from the market, it was concluded that the D(-)lactic acid content could be used as an indicator of the storage time of these products. No consistent results were obtained with acetoin/diacetyl.     

Production of histamine and tyramine by lactic acid bacteria isolated from vacuum-packed sugar-salted fish

The incidence of histamine- or tyramine-producing lactic acid bacteria was examined in several products of vacuum-packed sugar-salted fish (salmon, halibut, mackerel). No histamine-producing isolates were observed, whereas the majority of tyramine-producing isolates were identified as Carnobacterium spp. These organisms were shown to be important members of the microbial flora during storage of vacuum-packed sugar-salted salmon at 5°C. The amount of tyramine produced was reduced by lowering the temperature from 9°C to 4°C for all of five strains of carnobacteria or lactobacilli. The majority of tyramine was produced during the exponential growth phase for Carnobacterium piscicola N 5 and Lactobacillus viridescens N 69. The ability of these bacteria to produce tyramine may be used as an index of microbial quality/acceptability of stored vacuum-packed sugar-salted fish.     

Microbial Progression in Sliced Vacuum Packaged Bacon at Refrigeration Temperatures

S ummary . Three hundred and forty eight strains of micro-organisms were isolated from packaged bacon which had been cured by one or other of 2 methods (A and B) and stored at 0–10° for 7 weeks. Before storage, Micrococcus sub-group 7 (47%) and coryneform bacteria (33%) were the principal contaminants of bacon cured by method A and Micrococcus subgroup 1 (20%) and Microbacterium thermosphactum (21%), in bacon cured by method B. After 7 days at 0°, coagulase negative staphylococci accounted for 10 and 33% of the microflora in bacon A and B, respectively: other organisms were micrococci, 60% (A) and 25% (B); corynebacteria, 20% (A) and 12·5% (B), yeasts ( Torulopsis candida and T. dattila ) 5% (A) and 29% (B). In the following month at 10°, Micrococcus sub-group 5 was the dominant (43–88%) contaminant of bacon B; the incidence of Streptococcus Group N ranged from 27–36% and that of unidentified lactic acid bacteria from 23–32%. Towards the end of storage, the order of dominance was Micrococcus sub-group 3 (38–58%) and atypical streptobacteria (38%) in A; Streptococcus Group N (42%) and Gram positive rods (5–20%) in B. Staphylococci tended to die out during storage of bacons cured by the 2 methods and coagulase positive staphylococci were not isolated.     

Effect of Sodium Nitrite and Sodium Chloride on Growth of Lactic Acid Bacteria

The effect of NaNO2 and NaCl on the growth of 24 lactic acid bacteria strains isolated from vacuum-packed cooked ring sausages were examined by analyzing different growth parameters with Bioscreen. NaNO2 had a very limited effect on the growth of lactic acid bacteria at 50 and 100 mg/l but at 400 mg/l a more pronounced inhibitory effect was found. Bacterial growth was enhanced by 1–2% (w/v) of added NaCl, while NaCl concentrations above 3% (w/v) had a clear inhibitory effect. Leuconostoc isolates seemed to be more sensitive to sodium nitrite and sodium chloride than homofermentative lactobacilli strains. Among homofermentative lactobacilli, the strains resembling Lactobacillus curvatus were more sensitive to NaCl than those resembling Lactobacillus sake.     

Effect of Sodium Nitrite on Toxin Production by Clostridium botulinum in bacon

Pork bellies were formulated to 0, 30, 60, 120, 170, or 340 μg of nitrite per g of meat and inoculated with Clostridium botulinum via pickle or after processing and slicing. Processed bacon was stored at 7 or 27 C and assayed for nitrite, nitrate, and botulinal toxin at different intervals. Nitrite levels declined during processing and storage. The rate of decrease was more rapid at 27 than at 7 C. Although not added to the system, nitrate was detected in samples during processing and storage at 7 and 27 C. The amount of nitrate found was related to formulated nitrite levels. No toxin was found in samples incubated at 7 C throughout the 84-day test period. At 27 C, via pickle, inoculated samples with low inoculum (210 C. botulinum per g before processing and 52 per g after processing) became toxic if formulated with 120 μg of nitrite per g of meat or less. Toxin was not detected in bacon formulated with 170 or 340 μg of nitrite per g of meat under these same conditions. Toxin was detected at all formulated nitrite levels in bacon inoculated via the pickle with 19,000 C. botulinum per g (4,300 per g after processing) and in samples inoculated after slicing. However, increased levels of formulated nitrite decreased the probability of botulinal toxin formation in bacon inoculated by both methods.     

Evaluation of the extent and type of bacterial contamination at different stages of processing of cooked ham

In an attempt to determine the composition and origin of the spoilage flora of refrigerated vacuum-packed cooked ham, the changes in microbial numbers and types were followed along the processing line. Results revealed Lactobacillus sake and Leuconostoc mesenteroides ssp. mesenteroides as the major causative agents of spoilage of sliced ham stored at 4 °C and 12 °C, due to recontamination in the cutting room. On the contrary, the progressive deterioration of whole ham under the same storage conditions was associated with a non-identifiable group of leuconostoc-like bacteria. Except for lactic acid bacteria, no other organism grew in vacuum packs of either sliced or whole ham. Although atypical leuconostocs could not be detected among isolates recovered from freshly produced whole ham, they appeared to survive cooking and proliferate during storage. Neither these organisms however, nor Lact. sake and Leuc. mesenteroides were important in curing and tumbling as carnobacteria, mainly Carnobacterium divergens, and Brochothrix thermosphacta dominated at this stage. A progressive inversion of the ham microflora from mostly Gram-negative at the beginning of processing to highly Gram-positive prior to cooking was noted. Listeria monocytogenes cross-contaminated ham during tumbling. However, the pathogen was always absent from the vacuum-packed product provided that heating to a core temperature of 70 °C occurred and recontamination during slicing and packing was prevented. The percentage distribution of different species of lactic acid bacteria as well as the uncommon phenotypic characteristics of some strains were discussed.     

OBSERVATIONS ON THE EFFECT OF ADDED HYPOCHLORITE ON THE NUMBERS AND KINDS OF BACTERIA IN MILK

SUMMARY: In an investigation of the effect of added hypochlorite on the bacterial flora and bacterial numbers of milk no bactericidal action was apparent within 15 min of adding the disinfectant unless at least 500 p/m available chlorine had been added. After 24 hr storage at atmospheric temperature a reduction in numbers was found with 10 p/m available chlorine and this became more pronounced with increased concentrations. Micrococci seemed more susceptible to the bactericidal effect than streptococci, which formed, after 24 hr storage, the predominant residual flora where 250 and 500 p/m available chlorine had been added. Acid production by lactic streptococci was retarded at concentrations greater than 50 p/m available chlorine in a 5 hr incubation period at 30°, but after a 24 hr incubation the decrease in acid production was only apparent when 500 or 1,000 p/m available chlorine was present. Little effect on acid production by lactic streptococci was found when less than 50 p/m available chlorine had been added. The residual chlorine content of rinses of equipment sterilized in hypochlorite solutions was negligible.     

Changes in the Microbiology of Vacuum-packaged Beef

The development of the microbial flora on meat stored in vacuum-bags at 0–2° for up to 9 weeks was studied. Although the proportion of lactic acid bacteria increased relative to the aerobic spoilage organisms, the numbers of the latter continued to increase throughout storage. The initial contamination of the meat before vacuum-packaging was important; meat with a very low initial number had lower numbers of bacteria throughout storage for up to 9 weeks and steaks cut from such meat which had been stored always had 1–2 days' additional aerobic shelf life at 4°. Spoilage of these steaks was due either to slime formation and off-odour associated with high counts of presumptive Pseudomonas spp., or by discoloration and souring (lactic acid bacteria). Extract release volume and pH measurements performed on the vacuum-packaged primal joints were only of value in determining the onset of aerobic spoilage when large numbers of Gram negative organisms were present, whereas the titrimetric method of spoilage evaluation of the vacuum-packaged meat showed a correlation with spoilage due to lactic organisms.     

A note on the microbiology of retail packs of prepared salad vegetables

Retail packs of mixed, prepared salad vegetables from two different manufacturers were stored at 7 degrees C until the end of storage-life (sell-by date plus 1 d), when the microbial flora was examined. The quality of the salads was acceptable at the end of storage life. The oxygen concentrations in packs were lower, and the carbon dioxide concentrations were higher, than those in air. High numbers of bacteria were present, with Pseudomonas spp. and Enterobacter agglomerans predominating in packs of both salads, together with lactic acid bacteria in one of the salads. Significant numbers of pectolytic bacteria including Pseudomonas spp. and Erwinia carotovora were detected. Despite the presence of high numbers of coliform bacteria, Escherichia coli was not detected in batches of one salad, and was detected in relatively low numbers in batches of the other. Yersinia spp., predominantly environmental strains of Yersinia enterocolitica, were isolated by enrichment from all samples tested; Staphylococcus aureus and enterococci were not detected.     

Soil and sediment bacteria capable of aerobic nitrate respiration.

Several laboratory strains of gram-negative bacteria are known to be able to respire nitrate in the presence of oxygen, although the physiological advantage gained from this process is not entirely clear. The contribution that aerobic nitrate respiration makes to the environmental nitrogen cycle has not been studied. As a first step in addressing this question, a strategy which allows for the isolation of organisms capable of reducing nitrate to nitrite following aerobic growth has been developed. Twenty-nine such strains have been isolated from three soils and a freshwater sediment and shown to comprise members of three genera (Pseudomonas, Aeromonas, and Moraxella). All of these strains expressed a nitrate reductase with an active site located in the periplasmic compartment. Twenty-two of the strains showed significant rates of nitrate respiration in the presence of oxygen when assayed with physiological electron donors. Also isolated was one member of the gram-positive genus Arthrobacter, which was likewise able to respire nitrate in the presence of oxygen but appeared to express a different type of nitrate reductase. In the four environments studied, culturable bacteria capable of aerobic nitrate respiration were isolated in significant numbers (10(4) to 10(7) per g of soil or sediment) and in three cases were as abundant as, or more abundant than, culturable bacteria capable of denitrification. Thus, it seems likely that the corespiration of nitrate and oxygen may indeed make a significant contribution to the flux of nitrate to nitrite in the environment.     

Microbiological and Chemical Changes in Lean Wiltshire Bacon During Aerobic Storage

Total microbial growth and chemical changes in aerobically stored bacon lean were found to fall into 2 phases. Phase 1: there was an increase in total microbial load to c . 10⁹/g, during which most of the nitrate was broken down and there was an accumulation of nitrite. Phase 2: the microbial load increased further to c . 10¹⁰/g and during this time most of the accumulated nitrite was broken down to unknown products, so that at the end of the experiments most of the nitrate and nitrite had disappeared from the bacon.
The most important types of micro-organisms isolated from the bacons were Micrococcus , yeasts, Vibrio, Acinetobacter, Alcaligenes and Arthrobacter-Corynebacterium. The latter 3 types appeared to be associated with phase 2 changes.     

Effect of Packaging under Carbon Dioxide, Nitrogen or Air on the Microbial Flora of Pork Stored at 4°C

Changes in the microbial flora of pork packed in laminated plastic bags and stored at 4 °C were studied in an initial atmosphere of carbon dioxide, nitrogen or air. The time needed for the total aerobic count at 28 °C to reach 5 × 10⁶ organisms/cm² was about 7 times longer in carbon dioxide than in air, whilst in nitrogen it was about twice as long.
The predominant organisms on fresh pork taken directly from the processing line were: Acinetobacter calcoaceticus , non-fluorescent Pseudomonas spp. and Flavobacterium spp. After storage in air for 7 d, more than 90% of the flora consisted of non-fluorescent Pseudomonas spp. After storage in nitrogen for 10 d, 70% of the flora consisted of non-fluorescent Pseudomonas spp. with lower levels of fluorescent Pseudomonas spp., Kurthia zopfii, Aeromonas hydrophila and Lactobacillus plantarum. The non-fluorescent Pseudomonas spp. could be divided into three different groups, on proteolytic and lipolytic ability; the distribution of the groups was markedly different between pork loins stored in air and nitrogen.
On pork stored in carbon dioxide for 21 d the flora consisted of L. plantarum together with lower levels of heterofermentative lactic acid bacteria. When the storage time in carbon doxide was prolonged to 35 d, the proportion of heterofermentative lactic acid bacteria increased to about 50% of the flora.     

An investigation into the sources of lactic acid bacteria in grass silage

During the 1983 silage making season an investigation was carried out to determine the contribution made by farm machinery to the numbers of lactic acid bacteria present on grass. Although no lactic acid bacteria were isolated from the standing crop, up to 10³/g were found in the grass as it entered the clamp. Examination of machinery in close contact with the grass revealed lactic acid bacteria on the mower blades and on the forks of the rough-terrain vehicle used to pack the grass into the clamp. During the 1984 season the occurrence of lactic acid bacteria in particular batches of grass from the standing crop through to silage after 2 d in the clamp was studied. There was a gradual increase in numbers of lactic acid bacteria in the grass during the process of mowing and collection. Wilting resulted in a greater initial increase in numbers but non-wilted silage had the highest count after 48 h. The majority of organisms isolated from machinery before silage making were streptococci. After silage making the number and proportions of organisms changed, with numbers increasing in all cases, particularly from the mower with lactobacilli and streptococci predominating which indicates that passage through the machinery served to inoculate the grass.     

Influence of nitrate on fermentation pattern, molar growth yields and synthesis of cytochrome b in Propionibacterium pentosaceum.

Under anaerobic conditions, Propionibacterium pentosaceum reduces nitrate to nitrite until nitrate is exhausted from the medium when nitrite is converted into N2 or N2O. In the presence of nitrate, fermentation patterns for lactate, glycerol and pyruvate were different from those obtained during anaerobic growth without an inorganic electron acceptor. In the presence of these substrates, a drastic decrease in propionate formation was observed, some pyruvate accumulated during growth with lactate, and acetate was produced from glycerol. Acetate production from lactate and pyruvate was not influenced by the presence of nitrate. Furthermore, CO2 was produced by citric acid cycle activity. The fermentation pattern during nitrite reduction resembled that of P. pentosaceum grown anaerobically without an inorganic electron acceptor. Nitrits has a toxic effect, since bacteria inoculated into a medium with 9 mM-nitrite failed to grow. The cytochrome spectrum of anaerobically grown P. pentosaceum was similar with and without nitrate. In membrane fractions of bacteria grown anaerobically with nitrate, cytochrome b functioned in the transfer of electrons from lactate, glycerol I-phosphate and NADH to nitrate. Molar growth yeilds were increased in the presence of nitrate, indicating an increased production of ATP. This could be explained by citric acid cycle activity, and by ocidative phosphorylation coupled to nitrate reduction. Assuming that I mol ATP is formed in the electron transfer from lactate or glycerol I-phosphate to nitrate, and that 2 mol ATP are formed in the electron transfer from NADH to nitrate, YATP values (g dry wt bacteria/mol ATP) were obtained of between 5-0 and 12-6. The higher YATP values were similar to those obtained during anaerobic growth without an inorganic electron acceptor. This supports the assumptions about the efficiency of oxidative phosphorylation for electron transport to nitrate. Low YAPT values were found when high concentrations of nitrite (15 to 50 mM) accumulated, and were probably due to the toxic effect of nitrite.     

A SALT-TOLERANT DENITRIFYING BACILLUS STRAIN WHICH 'BLOWS' CANNED BACON

SUMMARY: A Bacillus strain, isolated from a 'blown' can of bacon, is described: it tolerates 15% NaCl and reduces nitrate and nitrite to N₂O and N₂. The pH-salt relationship, the tolerance to nitrite under aerobic and anaerobic conditions, the heat resistance of the spores, and the composition of the gas produced, have been investigated. The characters of the organism are such that, should its spores survive the processing of a cured meat, their subsequent development and the spoilage of the product seem almost certain.     

Classification of some lactic acid bacteria from vacuum-packed meats by direct probe mass spectrometry

The technique of direct probe mass spectrometry (DPMS) has been applied to the classification of 40 strains of lactic acid bacteria from refrigerated vacuum-packed beef, pork, lamb and bacon. Relationships between strains were examined by multi-variate statistical techniques using sets of ions selected for reproducibility and sample discrimination. Five groups were distinguished which corresponded closely to those detected in a previous numerical taxonomic study. Two groups contained all 12 representatives of a cluster of unidentifiable non-aciduric streptobacteria whose sub-division is supported by other taxonomic evidence. All twenty-one strains from a cluster of aciduric streptobacteria provisionally identified with Lacto-bacillus sake were contained in two further groups. The sub-division of these acid-uric strains revealed by DPMS has not been verified by other techniques and requires further investigation. The fifth group contained Leuconostoc strains. The study demonstrates the value of DPMS in confirming and clarifying classification schemes obtained by conventional methods.     

Application of antimicrobial-producing lactic acid bacteria to control pathogens in ready-to-use vegetables

Five psychrotrophic strains of lactic acid bacteria (Lactobacillus casei, Lact. plantarum and Pediococcus spp.) were isolated from 22 samples of commercial salads. These strains were shown to inhibit Aeromonas hydrophila, Listeria monocytogenes, Salmonella typhimurium and Staphylococcus aureus on MRS agar, in salads and in juice prepared from vegetable salads. Lactobacillus casei IMPCLC34 was most effective in reducing total mesophilic bacteria and the coliform group; Aer. hydrophila, Salm. typhimurium and Staph. aureus disappeared after 6 d of storage, while the counts for L. monocytogenes remained constant. The potential application of antimicrobial-producing lactic acid bacteria as biopreservatives of ready-to-use vegetables is suggested.     

Classification of some lactic acid bacteria from vacuum-packed meats by direct probe mass spectrometry

The technique of direct probe mass spectrometry (DPMS) has been applied to the classification of 40 strains of lactic acid bacteria from refrigerated vacuum-packed beef, pork, lamb and bacon. Relationships between strains were examined by multi-variate statistical techniques using sets of ions selected for reproducibility and sample discrimination. Five groups were distinguished which corresponded closely to those detected in a previous numerical taxonomic study. Two groups contained all 12 representatives of a cluster of unidentifiable non-aciduric streptobacteria whose sub-division is supported by other taxonomic evidence. All twenty-one strains from a cluster of aciduric streptobacteria provisionally identified with Lactobacillus sake were contained in two further groups. The sub-division of these aciduric strains revealed by DPMS has not been verified by other techniques and requires further investigation. The fifth group contained Leuconostoc strains. The study demonstrates the value of DPMS in confirming and clarifying classification schemes obtained by conventional methods.