Lactic acid concentration as an indicator of acceptability in refrigerated or freeze-thawed ground beef

Lactic acid concentrations increased in refrigerated and freeze-thawed anaerobically stored ground beef. Bacterial counts were higher in refrigerated samples, but the ratios of gram-positive bacteria in refrigerated and freeze-thawed samples were the same. No differences in appearance or odor between refrigerated and freeze-thawed samples were noted after 2 days of aerobic storage. Initial lactic acid concentration can be used to predict the shelf life of frozen beef.     

Relationship between lactic acid concentration and bacterial spoilage in ground beef.

Lactic acid concentration correlated with organoleptic spoilage of refrigerated, coarsely ground beef stored in casings with low oxygen permeability. The samples were assayed over time for lactic acid concentration, total aerobic plate count, percentage of gram-positive organisms, and pH. Lactic acid increased in all samples, as did the bacterial counts and percentage of gram-positive organisms in the total microflora, the latter representing an increase in the lactic acid-producing bacteria. pH was found to decrease in all samples, with the smallest decrease in pH being observed in the meat sample which maintained the lowest proportion of gram-positive organisms. With samples evaluated by a sensory panel, lactic acid levels were found to correlate inversely with odor acceptability.     

Isolation and characterization of Lactococcus piscium strains from vacuum-packaged refrigerated beef

AIMS: To characterize gram-positive, catalase-negative, psychrotrophic, lactic acid-homofermentative, non-motile cocci isolated from vacuum-packaged refrigerated beef using phenotypic and genotypic methods. METHODS AND RESULTS: A total of 89 strains was isolated at 2 and 6 weeks as one of the predominant microflora of five samples of vacuum-packaged beef stored at 2 degrees C. The strains were compared with reference strains of some gram-positive, catalase-negative cocci using SDS-PAGE whole-cell protein pattern analysis, biochemical characterization and 16S rDNA sequencing. The biochemical and physiological characteristics of the isolates resembled those of Lactococcus piscium GTC 552(T). Numerical analysis of the SDS-PAGE whole-cell protein patterns resulted in close clustering of the strains with L. piscium GTC 552(T) (r > 0.68). Other Lactococcus and Leuconostoc species could be distinguished from the isolates using SDS-PAGE whole-cell protein patterns (r < 0.58) and biochemical characteristics. The 16S rDNA sequencing of four randomly selected strains showed that the strains differed from L. piscium GTC 552(T) by two to three bases in the highly variable region of the sequence. This is the first report on the isolation of L. piscium from vacuum-packaged beef. CONCLUSIONS: The gram-positive catalase-negative cocci isolated from vacuum-packaged refrigerated beef have been identified as L. piscium. SIGNIFICANCE AND IMPACT OF THE STUDY: The findings of this work contribute to the knowledge of the microflora of vacuum-packaged refrigerated beef.     

ESTIMATION OF MICROBIOLOGICAL QUALITY OF CHILLED BEEF USING AUTOMATED TURBIDIMETRY

Total bacterial counts on chilled beef samples were estimated by the standard plate count method and by an automated turbidimetric system. The latter method is based on product-specific calibration curves constructed by correlating growth curve parameters calculated for the turbidimeter to the log CFU values obtained by plate counts. A total of 74 beef samples was used to construct the calibration curves. Correlation analysis between turbidimetric parameters and plate count values showed that detection time was the best predictor to estimate microbial loads on fresh (r=0.91) and aged beef (r=0.94). Microbial loads for a different set of aged beef samples (n = 37) refrigerated for 7, 9, 10, 17 and 45 days were compared by turbidimetric measurements and plate counts. Mean total viable counts were log 5.92 ± 1.17 and log 5.54 ± 1.28 CFU/mL, respectively. Results showed that total bacterial counts on chilled beef could be estimated accurately from turbidimetric parameters. Furthermore, setting a cut-off value of log 6 CFU/mL allowed to accepting/rejecting samples according to their microbial condition in shorter periods of time compared to the traditional plate count method.     

Effect of Lactobacillus-protective cultures with bacteriocin-like inhibitory substances' producing ability on microbiological, chemical and sensory changes during storage of refrigerated vacuum-packaged sliced beef

AIMS: To investigate the effect of applying two different Lactobacillus-protective cultures, with bacteriocin-like inhibitory substances' (BLIS) producing ability, individually or in combination, on microbiological, chemical and sensory changes during storage of refrigerated vacuum-packaged sliced beef meat. METHODS AND RESULTS: Lactobacillus sakei CECT 4808 and Lactobacillus curvatus CECT 904(T), which were shown to be producers of BLIS, were inoculated individually or in combination on slices of beef M. semitendinosus. The samples were vacuum packaged and stored at 4 +/- 1 degrees C and were assessed during a 28-day storage period for microbiological [Enterobacteriaceae, Pseudomonas spp., lactic acid bacteria (LAB), Brochothrix thermosphacta and yeasts and moulds], chemical (pH, protein hydrolysis degree, lipid oxidation), sensory (abnormal odour) parameters and instrumental colour. Samples inoculated with the Lact. sakei strain and samples inoculated with the combination of the two strains had significantly (P < 0.05) lower spoilage microbial counts than those inoculated with the Lact. curvatus strain alone or the controls, while both chemical parameters (including lipid oxidation) and abnormal odour scores were also significantly (P < 0.05) improved by the former. Moreover, Lact. sakei alone showed a better preserving effect (P < 0.05) than the combination of both strains in the majority of the parameters tested. Instrumental colour measurements changed with storage time, but no treatment effects (P >or= 0.05) were observed during the whole 28-day storage period. CONCLUSIONS: The BLIS producer Lact. sakei CECT 4808 strain may be used for improving preservation of vacuum-packaged beef slices, as regards spoilage microbial counts and the chemical parameters tested in this study. SIGNIFICANCE AND IMPACT OF THE STUDY: Inoculation with the BLIS producer Lact. sakei CECT 4808 strain would provide an additional hurdle to improve storage life of refrigerated vacuum-packaged sliced beef. Furthermore, this strain demonstrated limited antioxidative ability, which could make a contribution to the prevention of lipid oxidation in meat and meat products.     

冷鲜牛肉中蜂房哈夫尼亚菌(Hafnia alvei)的分离与鉴定

在出口冷鲜牛肉的沙门氏菌检验过程中,从HE琼脂平板上分离到1株蜂房哈夫尼亚菌,该菌三糖铁斜面产碱,底层产酸,不产生H2S,不产气,革兰阴性。经BBL Crystal微生物半自动检测仪检测,同时结合伯杰细菌鉴定手册的生化试验结果鉴定为蜂房哈夫尼亚菌(Hafnia alvei)。     

The Limulus Lysate Endotoxin Assay as a Test of Microbial Quality of Ground Beef

The Limulus lysate test (LLT) for endotoxin assay has been found to be an excellent, simple and rapid test of microbial quality of refrigerated ground beef. In fresh ground beef held at 5°C for 7–12 d, LLT titres increased from 10²–10⁵ and correlated very highly with extract-release volume (ERV) data and total viable Gram negative counts at both 5° and 30°C. The LLT was negative for fresh beef containing low numbers of bacteria and on aged beef in the absence of increasing numbers of Gram negative bacteria. Of 14 Gram negative meat isolates, all gave a positive LLT while none of eight miscellaneous Gram positive bacteria did. The use of this test provides objective information on the microbial quality of fresh refrigerated ground meats in 1 h. Based upon this study, it is suggested that a 0·1 ml inoculum from a 10³ dilution of good quality ground beef should produce a negative lysate test and thus serve as an additional rapid screening test of meat microbial quality.     

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.     

Inactivation of Escherichia coli O157:H7, salmonellae, and Campylobacter jejuni in raw ground beef by gamma irradiation.

Raw ground beef patties inoculated with stationary-phase cells of Escherichia coli O157:H7, salmonellae, or Campylobacter jejuni were subjected to gamma irradiation (60Co) treatment, with doses ranging from 0 to 2.52 kGy. The influence of two levels of fat (8 to 14% [low fat] and 27 to 28% [high fat]) and temperature (frozen [-17 to -15 degrees C] and refrigerated [3 to 5 degrees C]) on the inactivation of each pathogen by irradiation was investigated. In ascending order of irradiation resistance, the D10 values ranged from 0.175 to 0.235 kGy (C. jejuni), from 0.241 to 0.307 kGy (E. coli O157:H7), and from 0.618 to 0.800 kGy (salmonellae). Statistical analysis revealed that E. coli O157:H7 had a significantly (P < 0.05) higher D10 value when irradiated at -17 to -15 degrees C than when irradiated at 3 to 5 degrees C. Regardless of the temperature during irradiation, the level of fat did not have a significant effect on the D10 value. Salmonellae behaved like E. coli O157:H7 in low-fat beef, but temperature did not have a significant effect when the pathogen was irradiated in high-fat ground beef. Significantly higher D10 values were calculated for C. jejuni irradiated in frozen than in refrigerated low-fat beef. C. jejuni was more resistant to irradiation in low-fat beef than in high-fat beef when treatment was at -17 to -15 degrees C. Regardless of the fat level and temperature during inactivation, these pathogens were highly sensitive to gamma irradiation.(ABSTRACT TRUNCATED AT 250 WORDS)     

Microbial Spoilage of Fresh Refrigerated Beef Liver

The types and numbers of micro-organisms involved in the spoilage of refrigerated beef liver were studied together with pH, hydration and organoleptic changes of the material. Fresh liver harboured a mixed population ( c . 1 × 10⁵ organisms/g) of Gram positive cocci, chromogens and non-chromogens, sporeformers, presumptive coliforms and Gram negative rods. When samples were rejected organoleptically, after 7–10 days at 5°, the contamination attained levels of c . 7–8 × 10⁷ organisms/g. Spoilage was due to souring; the pH fell from 6·3 (fresh liver) to c . 5·9. Lactic acid bacteria were predominant and Gram negative bacteria did not exceed 1·0 × 10⁶ organisms/g. This type of spoilage is explained by the carbohydrate content of c . 5% in liver. The value of pH appears to be a reliable indicator of liver freshness, with a pH of 6·1 indicating incipient spoilage.     

Taxonomic studies on a psychrophilic Clostridium from vacuum-packed beef: Description of Clostridium estertheticum sp. nov.

Abstract Taxonomic studies were performed on an anaerobic Gram-positive, spore-forming, psychrophilic bacterium originally isolated from spoiled vacuum-packed refrigerated beef. Based on the present finding it is proposed that this unknown psychrophilic bacterium be classified as a new species of the genus Clostridium , as Clostridium estertheticum sp. nov. The type strain is NCIMB 12511.     

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.     

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.     

Characteristics of the bacteriocin produced by <Emphasis Type="Italic">Lactococcus lactis</Emphasis> subsp. <Emphasis Type="Italic">cremoris</Emphasis> CTC 204 and the effect of this compound on the mesophilic bacteria associated with raw beef

Summary >Screening for the bacteriocin production of strains of lactic acid bacteria from various meat and meat products resulted in the detection of a bacteriocin-producing Lactococcus lactis subsp. cremoris CTC 204, isolated from chicken. The bacteriocin inhibited not only closely related lactic acid bacteria (Lactobacillus helveticus), but also pathogenic microorganisms (Staphylococcus aureus, Listeria monocytogenes, Bacillus cereus, and Clostridium perfringens). It was inactivated by α-chymotrypsin, ficin, papain, and pronase E, but not by lipase or pepsin. This compound was heat stable even at autoclaving temperature (121°C for 10min) and was produced during refrigerated storage. It was also active over a wide pH range (2–10), but the highest activity was observed in the lower pH range. The results indicated that dipping raw beef in the bacteriocin produced by strain CTC 204 could contribute to the extension of the shelf life of refrigerated bovine meat.     

Changes in the spoilage-related microbiota of beef during refrigerated storage under different packaging conditions

The microbial spoilage of beef was monitored during storage at 5 degrees C under three different conditions of modified-atmosphere packaging (MAP): (i) air (MAP1), (ii) 60% O2 and 40% CO2 (MAP2), and (iii) 20% O2 and 40% CO2 (MAP3). Pseudomonas, Enterobacteriaceae, Brochothrix thermosphacta, and lactic acid bacteria were monitored by viable counts and PCR-denaturing gradient gel electrophoresis (DGGE) analysis during 14 days of storage. Moreover, headspace gas composition, weight loss, and beef color change were also determined at each sampling time. Overall, MAP2 was shown to have the best protective effect, keeping the microbial loads and color change to acceptable levels in the first 7 days of refrigerated storage. The microbial colonies from the plate counts of each microbial group were identified by PCR-DGGE of the variable V6-V8 region of the 16S rRNA gene. Thirteen different genera and at least 17 different species were identified after sequencing of DGGE fragments that showed a wide diversity of spoilage-related bacteria taking turns during beef storage in the function of the packaging conditions. The countable species for each spoilage-related microbial group were different according to packaging conditions and times of storage. In fact, the DGGE profiles displayed significant changes during time and depending on the initial atmosphere used. The spoilage occurred between 7 and 14 days of storage, and the microbial species found in the spoiled meat varied according to the packaging conditions. Rahnella aquatilis, Rahnella spp., Pseudomonas spp., and Carnobacterium divergens were identified as acting during beef storage in air (MAP1). Pseudomonas spp. and Lactobacillus sakei were found in beef stored under MAP conditions with high oxygen content (MAP2), while Rahnella spp. and L. sakei were the main species found during storage using MAP3. The identification of the spoilage-related microbiota by molecular methods can help in the effective establishment of storage conditions for fresh meat.     

Increased beef consumption increases apolipoprotein A-I but not serum cholesterol of mildly hypercholesterolemic men with different levels of habitual beef intake

The objective of this research was to compare the effects of a lean beef enriched in oleic acid to a beef that is typical of the commercial beef consumed in the United States. Ten mildly hypercholesterolemic men, ages 34-58 years old, were selected from the Texas A&M University faculty and staff. Subjects were randomly assigned to one of two diets for a 6-week duration followed by a crossover after a 4-week habitual diet washout period. Diets were consumed daily for a 6-week study period. Participants substituted lean beef obtained from Wagyu bullocks or commercial beef for the meat typically consumed. Total cholesterol, apolipoproteins A-I and B, triacylglycerols, and low-density lipoprotein (LDL) and high-density lipoprotein (HDL) cholesterol were measured in serum samples collected weekly. Beef type had no effect on any measured variable. There were no significant differences between baseline HDL or LDL cholesterol concentrations after the consumption of the beef test diets. Apolipoprotein A-I, serum glucose, and uric acid concentrations were elevated by the additional dietary beef. Analysis of records of customary diets indicated that one group consumed 160 g of beef daily, whereas the other group consumed only 26 g of beef daily. Therefore, post hoc analyses tested the habitual beef intake x treatment time interaction. LDL cholesterol concentration was markedly higher in the group with low habitual beef intake (180 vs 144 mg/dl), and HDL cholesterol was slightly higher (44 vs 40 mg/dl; post-test values) than for the group with high habitual beef intake, but there were no habitual intake x time interactions for LDL or HDL cholesterol. Creatinine and blood urea nitrogen concentrations also were greater in the individuals habitually consuming less beef. This study had three important findings: i) a lean beef source enriched with oleic acid was no different from commercial beef in its effect on lipoprotein fractions; ii) neither previous level of beef intake nor baseline LDL cholesterol concentration influenced the serum cholesterol response to added dietary beef, which was negative; and iii) apolipoprotein A-I, but not HDL or LDL cholesterol, was sensitive to the additional dietary beef.     

Evaluation of the Extract-Release Volume Phenomenon as a Rapid Test for Detecting Spoilage in Beef

Ground beef of several grades, obtained fresh and held refrigerated until spoiled, was presented to a test panel for scoring on color, odor, and tactile response (tackiness) as to degree of acceptance. Panel scores were correlated with total bacterial counts, ninhydrin-reactive substances, and ERV (extract-release volume) on the same meat. ERV correlated highest with bacterial counts the largest number of times; tackiness, odor, ninhydrin, and color followed in that order. Correlation between bacterial numbers and organoleptic qualities was best, with tackiness followed closely by odor, and then by color. Correlation between tackiness and odor was high. The degree of correlation between bacterial numbers, tackiness, and ERV was high enough to warrant the use of the ERV phenomenon as a rapid test of microbial quality of beef. An ERV value of 25 under the conditions of the test was supported as a divider between acceptable and unacceptable ground beef.     

新鲜土壤样品储存温度和时间对不同形态氮素的影响

土壤氮素形态及含量具有重要的生态学研究意义,而土壤样品的储存对土壤氮素含量的准确测定有很大影响.为了选择合理的土壤样品储存方法,本研究以福建省建瓯市万木林保护区罗浮栲林土壤为研究对象,测定在不同温度(25、4和-20 ℃)、不同储存时间(0、7和30 d)下土壤铵态氮、硝态氮、总氮、可溶性有机氮、氨基酸氮含量和微生物生物量氮,以及冷冻后常温培养过程中的氮素含量.结果表明: 在7 d的储存时间内,除氨基酸氮以外,常温培养样品下其余的氮素含量均有所增加;与新鲜样品相比,冷藏、冷冻样品的所有氮素含量之间均无显著性差异,且氮素含量变化较常温培养下更加稳定.因低温储存样品有刺激氮矿化的效果,在30 d储存时间内,与新鲜样品相比,除可溶性有机氮外,冷藏、冷冻样品的所有氮素含量均显著升高;两种冷储存方法之间无显著差异.因此,新鲜样品带回实验室后应及时处理;如需要冷储藏,时间不要超过半个月.如果需要较长的储存时间,则需将样品放置于更低的温度(-40或-80 ℃).在对储存土壤样品进行培养试验之前,需要进行预培养处理.在预培养过程中,除硝态氮含量呈现先下降再迅速升高的趋势外,其余氮素均随着培养时间逐渐趋近于新鲜土壤样品含量,在培养一周左右恢复到与新鲜土壤样品氮含量最为接近的状态.结合已有研究,对野外取样和风干样品需要5～14 d的预培养,冷储存样品预培养时间不应少于一周.     

Use of a Titrimetric Method to Assess the Bacterial Spoilage of Fresh Beef

A new method of determining bacterial spoilage in fresh beef is presented. The technique is based upon the fact that as beef undergoes refrigerator spoilage, there is a gradual increase in the production of alkaline substances by the spoilage flora. The level of these substances was measured by titrating meat homogenates to a pH 5.00 end point, employing 0.02 n HCl and an autotitrator. When 23 samples of ground beef from retail stores were tested, an average of 1.32 ml of acid was required for titration of 1 g of fresh beef to pH 5.00, whereas 2.58 ml was required for the same meat at the onset of spoilage. Preliminary data indicate that beef which requires more than 2 ml of 0.02 n HCl/g to lower its pH to 5.00 under the conditions of the test is in some state of incipient spoilage. The statistical correlation between titration values, log bacterial numbers, and extract-release volume was high (P < 0.001). The technique is simple to execute and is highly reproducible, and duplicate samples can be run within 15 min.     

Impact of inoculum preparation and storage conditions on the response of Escherichia coli O157:H7 populations to undercooking and simulated exposure to gastric fluid

This study evaluated the impact of inoculum preparation and storage conditions on the response of Escherichia coli O157:H7 exposed to consumer-induced stresses simulating undercooking and digestion. Lean beef tissue samples were inoculated with E. coli O157:H7 cultures prepared in tryptic soy broth or meat decontamination runoff fluids (WASH) or detached from moist biofilms or dried biofilms formed on stainless steel coupons immersed in inoculated WASH. After inoculation, the samples were left untreated or dipped for 30 s each in hot (75 degrees C) water followed by lactic acid (2%, 55 degrees C), vacuum packaged, stored at 4 (28 days) or 12 degrees C (16 days), and periodically transferred to aerobic storage (7 degrees C for 5 days). During storage, samples were exposed to sequential heat (55 degrees C; 20 min) and simulated gastric fluid (adjusted to pH 1.0 with HCl; 90 min) stresses simulating consumption of undercooked beef. Under the conditions of this study, cells originating from inocula of planktonic cells were, in general, more resistant to heat and acid than cells from cultures grown as biofilms and detached prior to meat inoculation. Heat and acid tolerance of cells on meat stored at 4 degrees C was lower than that of cells on nondecontaminated meat stored at 12 degrees C, where growth occurred during storage. Decontamination of fresh beef resulted in injury that inhibited subsequent growth of surviving cells at 12 degrees C, as well as in decreases in resistance to subsequent heat and acid stresses. The shift of pathogen cells on beef stored under vacuum at 4 degrees C to aerobic storage did not affect cell populations or subsequent survival after sequential exposure to heat and simulated gastric fluid. However, the transfer of meat stored under vacuum at 12 degrees C to aerobic storage resulted in reduction in pathogen counts during aerobic storage and sensitization of survivors to the effects of sequential heat and acid exposure.