Effect of Challenge Temperature and Solute Type on Heat Tolerance of Salmonella Serovars at Low Water Activity

Salmonella spp. are reported to have an increased heat tolerance at low water activity (a_w; measured by relative vapor pressure [rvp]), achieved either by drying or by incorporating solutes. Much of the published data, however, cover only a narrow treatment range and have been analyzed by assuming first-order death kinetics. In this study, the death of Salmonella enterica serovar Typhimurium DT104 when exposed to 54 combinations of temperature (55 to 80°C) and a_w (rvp 0.65 to 0.90, reduced using glucose-fructose) was investigated. The Weibull model (LogS = −btⁿ) was used to describe microbial inactivation, and surface response models were developed to predict death rates for serovar Typhimurium at all points within the design surface. The models were evaluated with data generated by using six different Salmonella strains in place of serovar Typhimurium DT104 strain 30, two different solutes in place of glucose-fructose to reduce a_w, or six low-a_w foods artificially contaminated with Salmonella in place of the sugar broths. The data demonstrate that, at temperatures of ≥70°C, Salmonella cells at low a_w were more heat tolerant than those at a higher a_w but below 65°C the reverse was true. The same patterns were generated when sucrose (rvp 0.80 compared with 0.90) or NaCl (0.75 compared with 0.90) was used to reduce a_w, but the extent of the protection afforded varied with solute type. The predictions of thermal death rates in the low-a_w foods were usually fail-safe, but the few exceptions highlight the importance of validating models with specific foods that may have additional factors affecting survival.     

Survival and filamentation of Salmonella enterica serovar enteritidis PT4 and Salmonella enterica serovar typhimurium DT104 at low water activity

In this study we investigated the long-term survival of and morphological changes in Salmonella strains at low water activity (a(w)). Salmonella enterica serovar Enteritidis PT4 and Salmonella enterica serovar Typhimurium DT104 survived at low a(w) for long periods, but minimum humectant concentrations of 8% NaCl (a(w), 0. 95), 96% sucrose (a(w), 0.94), and 32% glycerol (a(w), 0.92) were bactericidal under most conditions. Salmonella rpoS mutants were usually more sensitive to bactericidal levels of NaCl, sucrose, and glycerol. At a lethal a(w), incubation at 37 degrees C resulted in more rapid loss of viability than incubation at 21 degrees C. At a(w) values of 0.93 to 0.98, strains of S. enterica serovar Enteritidis and S. enterica serovar Typhimurium formed filaments, some of which were at least 200 microm long. Filamentation was independent of rpoS expression. When the preparations were returned to high-a(w) conditions, the filaments formed septa, and division was complete within approximately 2 to 3 h. The variable survival of Salmonella strains at low a(w) highlights the importance of strain choice when researchers produce modelling data to simulate worst-case scenarios or conduct risk assessments based on laboratory data. The continued increase in Salmonella biomass at low a(w) (without a concomitant increase in microbial count) would not have been detected by traditional microbiological enumeration tests if the tests had been performed immediately after low-a(w) storage. If Salmonella strains form filaments in food products that have low a(w) values (0.92 to 0.98), there are significant implications for public health and for designing methods for microbiological monitoring.     

Identification of novel Salmonella enterica serovar Typhimurium DT104-specific prophage and nonprophage chromosomal sequences among serovar Typhimurium isolates by genomic subtractive hybridization

Genomic subtractive hybridization was performed between Salmonella enterica serovar Typhimurium LT2 and DT104 to search for novel Salmonella serovar Typhimurium DT104-specific sequences. The subtraction resulted mainly in the isolation of DNA fragments with sequence similarity to phages. Two fragments identified were associated with possible virulence factors. One fragment was identical to irsA of Salmonella serovar Typhimurium ATCC 14028, which is suggested to be involved in macrophage survival. The other fragment was homologous to HldD, an Escherichia coli O157:H7 lipopolysaccharide assembly-related protein. Five selected DNA fragments-irsA, the HldD homologue, and three fragments with sequence similarity to prophages-were tested for their presence in 17 Salmonella serovar Typhimurium DT104 isolates and 27 non-DT104 isolates by PCR. All five selected DNA fragments were Salmonella serovar Typhimurium DT104 specific among the serovar Typhimurium isolates tested. These DNA fragments can be useful for better detection and typing of Salmonella serovar Typhimurium DT104.     

Induction and resuscitation of viable but nonculturable Salmonella enterica serovar typhimurium DT104

Salmonella enterica serovar Typhimurium DT104 11601 was tested for its ability to maintain viability in minimal, chemically defined solutions. Periodic monitoring of growth and survival in microcosms of different ion concentrations, maintained at various temperatures, showed a gradual decline in culturable organisms ( approximately 235 days) at 5 degrees C. Organisms maintained at a higher temperature (21 degrees C) showed continuous, equivalent CFU per milliliter ( approximately 10(6)) up to 400 days after inoculation. Fluorescence microscopy with Baclight revealed that nonculturable cells were actually viable, while observations with scanning electron microscopy showed that the cells had retained their structural integrity. Temperature upshift (56 degrees C +/- 0.5, 15 s) of the nonculturable organisms (5 degrees C) in Trypticase soy broth followed by immediate inoculation onto Trypticase soy agar (TSA) gave evidence of resuscitation. Interestingly, S. enterica serovar Typhimurium DT104 from the microcosms at either 5 degrees C (1 to 200 days) or 21 degrees C (1 to 250 days) did not show enhanced growth after intermittent inoculation onto catalase-supplemented TSA. Furthermore, cells from 21 degrees C microcosms exposed to oxidative and osmotic stress showed greater resistance to stresses over increasing times of exposure than did recently grown cells. It is possible that the exceptional survivability and resilience of this particular strain may in part reflect the growing importance of this multidrug-resistant organism, in general, as a cause of intestinal disease in humans. The fact that S. enterica serovar Typhimurium DT104 11601 is capable of modifying its physiological characteristics, including entry into and recovery from the viable but nonculturable state, suggests the overall possibility that S. enterica serovar Typhimurium DT104 may be able to respond uniquely to various adverse environmental conditions.     

Genomic relationship of Salmonella enterica serovar typhimurium DT104 isolates from Korea and the United States

Salmonella enterica serovar Typhimurium DT104 (Salmonella Typhimurium DT104 or DT104) has been emerging as a common pathogen for human in Korea since 1997. In order to compare the genomic relationship and to search for the dominant strains in Korea, we conducted pulsed-field gel electrophoresis (PFGE) and IS200 fingerprinting of 25 epidemiological unrelated isolates from human and animals from Korea and cattle from America. Two Salmonella Typhimurium DT104 isolates from human in Korea and all 8 isolates from American cattle had indistinguishable patterns from the PFGE and IS200 fingerprinting but multidrug-resistant Salmonella Typhimurium isolates, including DT104, from Korean animals had diverse genetic patterns. The data suggest that a dominant DT104 strain might have circulated between Korean and American cattle and that it had a high level of clonality.     

Habituation of Salmonella spp. at reduced water activity and its effect on heat tolerance

The effect of habituation at reduced water activity (a(w)) on heat tolerance of Salmonella spp. was investigated. Stationary-phase cells were exposed to a(w) 0.95 in broths containing glucose-fructose, sodium chloride, or glycerol at 21 degrees C for up to a week prior to heat challenge at 54 degrees C. In addition, the effects of different a(w)s and heat challenge temperatures were investigated. Habituation at a(w) 0.95 resulted in increased heat tolerance at 54 degrees C with all solutes tested. The extent of the increase and the optimal habituation time depended on the solute used. Exposure to broths containing glucose-fructose (a(w) 0.95) for 12 h resulted in maximal heat tolerance, with more than a fourfold increase in D(54) values. Cells held for more than 72 h in these conditions, however, became as heat sensitive as nonhabituated populations. Habituation in the presence of sodium chloride or glycerol gave rise to less pronounced but still significant increases in heat tolerance at 54 degrees C, and a shorter incubation time was required to maximize tolerance. The increase in heat tolerance following habituation in broths containing glucose-fructose (a(w) 0.95) was RpoS independent. The presence of chloramphenicol or rifampin during habituation and inactivation did not affect the extent of heat tolerance achieved, suggesting that de novo protein synthesis was probably not necessary. These data highlight the importance of cell prehistory prior to heat inactivation and may have implications for food manufacturers using low-a(w) ingredients.     

Characterization of the genomes of a diverse collection of Salmonella enterica serovar Typhimurium definitive phage type 104

Salmonella enterica serovar Typhimurium definitive phage type 104 (DT104) has caused significant morbidity and mortality in humans and animals for almost three decades. We completed the full DNA sequence of one DT104 strain, NCTC13348, and showed that significant differences between the genome of this isolate and the genome of the previously sequenced strain Salmonella serovar Typhimurium LT2 are due to integrated prophage elements and Salmonella genomic island 1 encoding antibiotic resistance genes. Thirteen isolates of Salmonella serovar Typhimurium DT104 with different pulsed-field gel electrophoresis (PFGE) profiles were analyzed by using multilocus sequence typing (MLST), plasmid profiling, hybridization to a pan-Salmonella DNA microarray, and prophage-based multiplex PCR. All the isolates belonged to a single MLST type, sequence type ST19. Microarray data demonstrated that the gene contents of the 13 DT104 isolates were remarkably conserved. The PFGE DNA fragment size differences in these isolates could be explained to a great extent by differences in the prophage and plasmid contents. Thus, here the nature of variation in different Salmonella serovar Typhimurium DT104 isolates is further defined at the gene and whole-genome levels, illustrating how this phage type evolves over time.     

Identification of Novel Salmonella enterica Serovar Typhimurium DT104-Specific Prophage and Nonprophage Chromosomal Sequences among Serovar Typhimurium Isolates by Genomic Subtractive Hybridization

Genomic subtractive hybridization was performed between Salmonella enterica serovar Typhimurium LT2 and DT104 to search for novel Salmonella serovar Typhimurium DT104-specific sequences. The subtraction resulted mainly in the isolation of DNA fragments with sequence similarity to phages. Two fragments identified were associated with possible virulence factors. One fragment was identical to irsA of Salmonella serovar Typhimurium ATCC 14028, which is suggested to be involved in macrophage survival. The other fragment was homologous to HldD, an Escherichia coli O157:H7 lipopolysaccharide assembly-related protein. Five selected DNA fragments—irsA, the HldD homologue, and three fragments with sequence similarity to prophages—were tested for their presence in 17 Salmonella serovar Typhimurium DT104 isolates and 27 non-DT104 isolates by PCR. All five selected DNA fragments were Salmonella serovar Typhimurium DT104 specific among the serovar Typhimurium isolates tested. These DNA fragments can be useful for better detection and typing of Salmonella serovar Typhimurium DT104.     

Capsular polysaccharide surrounds smooth and rugose types of Salmonella enterica serovar Typhimurium DT104

The biofilms and rugose colony morphology of Salmonella enterica serovar Typhimurium strains are usually associated with at least two different exopolymeric substances (EPS), curli and cellulose. In this study, another EPS, a capsular polysaccharide (CP) synthesized constitutively in S. enterica serovar Typhimurium strain DT104 at 25 and 37 degrees C, has been recognized as a biofilm matrix component as well. Fluorophore-assisted carbohydrate electrophoresis (FACE) analysis indicated that the CP is comprised principally of glucose and mannose, with galactose as a minor constituent. The composition differs from that of known colanic acid-containing CP that is isolated from cells of Escherichia coli and other enteric bacteria grown at 37 degrees C. The reactivity of carbohydrate-specific lectins conjugated to fluorescein isothiocyanate or gold particles with cellular carbohydrates demonstrated the cell surface localization of CP. Further, lectin binding also correlated with the FACE analysis of CP. Immunoelectron microscopy, using specific antibodies against CP, confirmed that CP surrounds the cells. Confocal microscopy of antibody-labeled cells showed greater biofilm formation at 25 degrees C than at 37 degrees C. Since the CP was shown to be produced at both 37 degrees C and 25 degrees C, it does not appear to be significantly involved in attachment during the early formation of the biofilm matrix. Although the attachment of S. enterica serovar Typhimurium DT104 does not appear to be mediated by its CP, the capsule does contribute to the biofilm matrix and may have a role in other features of this organism, such as virulence, as has been shown previously for the capsules of other gram-negative and gram-positive bacteria.     

Identification of specific gene sequences conserved in contemporary epidemic strains of Salmonella enterica

Genetic elements specific to recent and contemporary epidemic strains of Salmonella enterica were identified using comparative genomic analysis. Two epidemic multidrug-resistant (MDR) strains, MDR Salmonella enterica serovar Typhimurium definitive phage type 104 (DT104) and cephalosporin-resistant MDR Salmonella enterica serovar Newport, and an epidemic pansusceptible strain, Salmonella serovar Typhimurium DT160, were subjected to Salmonella gene microarray and suppression subtractive hybridization analyses. Their genome contents were compared with those of coexisting sporadic strains matched by serotype, geographic and temporal distribution, and host species origin. These paired comparisons revealed that epidemic strains of S. enterica had specific genes and gene regions that were shared by isolates of the same subtype. Most of these gene sequences are related to mobile genetic elements, including phages, plasmids, and plasmid-like and transposable elements, and some genes may encode proteins conferring growth or survival advantages. The emergence of epidemic MDR strains may therefore be associated with the presence of fitness-associated genetic factors in addition to their antimicrobial resistance genes.     

Prevalence and genetic properties of Salmonella enterica serovar typhimurium definitive phage type 104 isolated from Rattus norvegicus and Rattus rattus house rats in Yokohama City, Japan

Salmonella enterica serovar Typhimurium was isolated from the intestinal contents of Rattus rattus and Rattus norvegicus house rats captured at two buildings, designated buildings J and YS, in Yokohama City, Japan. From October 1997 to September 1998, 52 of 339 (15.3%) house rats were found to carry Salmonella serovar Typhimurium definitive phage type 104 (DT104). In building J, 26 of 161 (16.1%) house rats carried DT104 over the 1-year study period, compared to 26 of 178 (14.6%) rats in building YS. The isolation rates of DT104 from R. rattus and R. norvegicus were similar in the two buildings. Most DT104 strains from building J (24 of 26) showed resistance to ampicillin, chloramphenicol, streptomycin, sulfisoxazole, and tetracycline and contained both the 1.0- and 1.2-kbp integrons, carrying genes pse1, pasppflo-like, aadA2, sulI, and tet(G). All DT104 strains from building YS were resistant to ampicillin and sulfisoxazole, and had the 1.2-kbp integron carrying pse1 and sulI. Cluster analysis of pulsed-field gel electrophoresis patterns of BlnI-digested DT104 DNAs showed that 22 of 26 DT104 strains from building J and 24 of 26 strains from building YS could be grouped into separate clusters each specific for the building origin. These results indicated that DT104 strains were prevalent in house rat colonies in each building and suggest that house rats may play an important role in the epidemiology of DT104.     

Molecular characterization of the multidrug-resistant phage types Salmonella enterica serovar Typhimurium DT104, DT20A and DT120 strains in the Slovakia

An increase in the number of multidrug-resistant Salmonella enterica serovar Typhimurium strains (definitive phage type DT20a and DT120) as well as the occurrence of DT104 strains during 2003-2005 in Slovakia was documented. Based on the results of the molecular analysis we suggest that multidrug-resistant DT20a and DT120 phage types are more closely related to multidrug-resistant phage type, and that the occurrence is probably due to changes in the phage susceptibility of DT104. Continued surveillance and molecular analysis should be maintained to follow the spread of these new multidrug-resistant DT104 variants in animals and humans.     

Acid resistance variability among isolates of Salmonella enterica serovar Typhimurium DT104

AIMS: Acid resistance could be an indicator of virulence since acid resistant strains are able to better survive the human stomach passage and in macrophages. We studied the acid resistance of several Salmonella Typhimurium DT104 strains isolated from food and humans and identified cellular parameters contributing to the enhanced acid resistance of these isolates. METHODS AND RESULTS: Acid resistance was tested in 37 Salmonella enterica Typhimurium serovar DT104 (S. Typhimurium DT104) strains. Acid adaptation at pH 5 followed by exposure for 2 h at pH 2.5 in the 27 human, nine nonhuman, and in two reference strains, revealed strong variation of acid survival. After 2 h at pH 2.5 six strains of S. Typhimurium DT104 were considered high acid resistant as they displayed a level of survival >10%, 14 strains were considered intermediate acid resistant (level of survival was <10% and >0.01%) and 19 strains were considered low acid resistant (level of survival <0.01%). Six strains were selected for further studies and proteomics revealed a relatively high amount of phase 2 flagellin in an acid-sensitive strain and a relatively high amount of the beta component of the H(+)/ATPase in an acid-resistant strain. Two strains were slightly more heat resistant possibly as the result of increased levels of DnaK or GroEL. CONCLUSIONS: A significant difference could be detected between human and food isolates regarding their acid resistance; all high acid-resistant strains were human isolates. SIGNIFICANCE AND IMPACT OF THE STUDY: S. Typhimurium DT104 is known for two decades and has a great impact on human health causing serious food-borne diseases. Our results suggest the existence of a positive correlation between acid resistance and pathogenicity in S. Typhimurium DT104 as all high acid-resistant strains were isolated from humans.     

Cost-effective application of pulsed-field gel electrophoresis to typing of Salmonella enterica Serovar Typhimurium

Salmonella enterica serovar Typhimurium is frequently isolated from humans and animals. Phage typing is historically the first-line reference typing technique in Europe. It is rapid and convenient for laboratories with appropriate training and experience, and costs of consumables are low. Phage typing and pulsed-field gel electrophoresis (PFGE) were performed on 503 isolates of serovar Typhimurium. Twenty-nine phage types and 53 PFGE patterns were observed. Most isolates of phage types DT104, DT104b, and U310 are not distinguishable from other members of their phage type by PFGE. By contrast, PFGE of isolates of phage types DT193 and U302 shows great heterogeneity. Analysis of experience with PFGE and phage typing can facilitate the selective application of PFGE to maximize the yield of epidemiologically relevant additional information while controlling costs.     

Cost-Effective Application of Pulsed-Field Gel Electrophoresis to Typing of Salmonella enterica Serovar Typhimurium

Salmonella enterica serovar Typhimurium is frequently isolated from humans and animals. Phage typing is historically the first-line reference typing technique in Europe. It is rapid and convenient for laboratories with appropriate training and experience, and costs of consumables are low. Phage typing and pulsed-field gel electrophoresis (PFGE) were performed on 503 isolates of serovar Typhimurium. Twenty-nine phage types and 53 PFGE patterns were observed. Most isolates of phage types DT104, DT104b, and U310 are not distinguishable from other members of their phage type by PFGE. By contrast, PFGE of isolates of phage types DT193 and U302 shows great heterogeneity. Analysis of experience with PFGE and phage typing can facilitate the selective application of PFGE to maximize the yield of epidemiologically relevant additional information while controlling costs.     

Evaluation of the pH-dependent,stationary-phase acid tolerance in Listeria monocytogenes and Salmonella Typhimurium DT104 induced by culturing in media with 1% glucose: a comparative study with Escherichia coli O157:H7

AIMS: To comparatively evaluate the adaptive stationary-phase acid tolerance response (ATR) in food-borne pathogens induced by culturing in glucose-containing media, as affected by strain variability and antibiotic resistance, growth temperature, challenge pH and type of acidulant. METHODS AND RESULTS: Antibiotic resistant or sensitive strains of Listeria monocytogenes, Salmonella including S. Typhimurium DT104, and Escherichia coli O157:H7 were cultured (30 degrees C for 24 h; 10 degrees C for up to 14 days) in trypticase soya broth with yeast extract (TSBYE) with 1% or without glucose to induce or prevent acid adaptation, respectively. Cultures were subsequently exposed to pH 3.5 or 3.7 with lactic or acetic acid at 25 degrees C for 120 min. Acid-adapted cultures were more acid tolerant than nonadapted cultures, particularly those of L. monocytogenes and Salmonella. No consistent, positive or negative, influence of antibiotic resistance on the pH-inducible ATR or acid resistance (AR) was observed. Compared with 30 degrees C cultures, growth and acid adaptation of L. monocytogenes and S. Typhimurium DT104 at 10 degrees C markedly reduced their ATR and AR in stationary phase. E. coli O157:H7 had the greatest AR, relying less on acid adaptation. A 0.2 unit difference in challenge pH (3.5-3.7) caused great variations in survival of acid-adapted and nonadapted cells. CONCLUSIONS: Culturing L. monocytogenes and Salmonella to stationary phase in media with 1% glucose induces a pH-dependent ATR and enhances their survival to organic acids; thus, this method is suitable for producing acid-adapted cultures for use in food challenge studies. SIGNIFICANCE AND IMPACT OF THE STUDY: Bacterial pathogens may become acid-adapted in foods containing glucose or other fermentable carbohydrates. Low storage temperatures may substantially decrease the stationary-phase ATR of L. monocytogenes and S. Typhimurium DT104, but their effect on ATR of E. coli O157:H7 appears to be far less dramatic.     

Identification of Specific Gene Sequences Conserved in Contemporary Epidemic Strains of Salmonella enterica

Genetic elements specific to recent and contemporary epidemic strains of Salmonella enterica were identified using comparative genomic analysis. Two epidemic multidrug-resistant (MDR) strains, MDR Salmonella enterica serovar Typhimurium definitive phage type 104 (DT104) and cephalosporin-resistant MDR Salmonella enterica serovar Newport, and an epidemic pansusceptible strain, Salmonella serovar Typhimurium DT160, were subjected to Salmonella gene microarray and suppression subtractive hybridization analyses. Their genome contents were compared with those of coexisting sporadic strains matched by serotype, geographic and temporal distribution, and host species origin. These paired comparisons revealed that epidemic strains of S. enterica had specific genes and gene regions that were shared by isolates of the same subtype. Most of these gene sequences are related to mobile genetic elements, including phages, plasmids, and plasmid-like and transposable elements, and some genes may encode proteins conferring growth or survival advantages. The emergence of epidemic MDR strains may therefore be associated with the presence of fitness-associated genetic factors in addition to their antimicrobial resistance genes.     

Salmonella enterica serovar Typhimurium DT104 displays a rugose phenotype

Rugose phenotypes, such as those observed in Vibrio cholerae, have increased resistance to chlorine, oxidative stress, and complement-mediated killing. In this study we identified and defined a rugose phenotype in Salmonella enterica serovar Typhimurium DT104 and showed induction only on certain media at 25 degrees C after 3 days of incubation. Incubation at 37 degrees C resulted in the appearance of the smooth phenotype. Observation of the ultrastructure of the rugose form and a stable smooth variant (Stv), which was isolated following a series of passages of the rugose cells, revealed extracellular substances only in cells from the rugose colony. Observation of the extracellular substance by scanning electron microscopy (SEM) was correlated with the appearance of corrugation during development of rugose colony morphology over a 4-day incubation period at 25 degrees C. In addition, the cells also formed a pellicle in liquid broth, which was associated with the appearance of interlacing slime and fibrillar structures, as observed by SEM. The pellicle-forming cells were completely surrounded by capsular material, which bound cationic ferritin, thus indicating the presence of an extracellular anionic component. The rugose cells, in contrast to Stv, showed resistance to low pH and hydrogen peroxide and an ability to form biofilms. Based on these results and analogy to the rugose phenotype in V. cholerae, we propose a possible role for the rugose phenotype in the survival of S. enterica serovar Typhimurium DT104.     

Induction and Resuscitation of Viable but Nonculturable Salmonella enterica Serovar Typhimurium DT104†

Salmonella enterica serovar Typhimurium DT104 11601was tested for its ability to maintain viability in minimal, chemically defined solutions. Periodic monitoring of growth and survival in microcosms of different ion concentrations, maintained at various temperatures, showed a gradual decline in culturable organisms (~235 days) at 5°C. Organisms maintained at a higher temperature (21°C) showed continuous, equivalent CFU per milliliter (~10⁶) up to 400 days after inoculation. Fluorescence microscopy with Baclight revealed that nonculturable cells were actually viable, while observations with scanning electron microscopy showed that the cells had retained their structural integrity. Temperature upshift (56°C ± 0.5, 15 s) of the nonculturable organisms (5°C) in Trypticase soy broth followed by immediate inoculation onto Trypticase soy agar (TSA) gave evidence of resuscitation. Interestingly, S. enterica serovar Typhimurium DT104 from the microcosms at either 5°C (1 to 200 days) or 21°C (1 to 250 days) did not show enhanced growth after intermittent inoculation onto catalase-supplemented TSA. Furthermore, cells from 21°C microcosms exposed to oxidative and osmotic stress showed greater resistance to stresses over increasing times of exposure than did recently grown cells. It is possible that the exceptional survivability and resilience of this particular strain may in part reflect the growing importance of this multidrug-resistant organism, in general, as a cause of intestinal disease in humans. The fact that S. enterica serovar Typhimurium DT104 11601 is capable of modifying its physiological characteristics, including entry into and recovery from the viable but nonculturable state, suggests the overall possibility that S. enterica serovar Typhimurium DT104 may be able to respond uniquely to various adverse environmental conditions.     

Effects of storage and the presence of a beef microflora on the thermal resistance of Salmonella Typhimurium DT104 in beef and broth systems

AIMS: To investigate the effects of storage and the presence of a beef microflora on the thermal resistance of Salmonella serotype Typhimurium DT104 on beef surfaces and in a broth system during subsequent heat treatments after extended low-temperature storage (4 degrees C for 14 days) or mild temperature abuse (10 degrees C for 7 days). METHODS AND RESULTS: Surviving Salm. Typhimurium DT104 cells were estimated after heating in a water bath (55 degrees C) by plating beef and broth samples on tryptone soya agar and overlaying with xylose-lysine-deoxycholate agar. In beef and broth systems, D(55) values for Salm. Typhimurium DT104 stored at 4 degrees C or 10 degrees C in the presence or absence of a beef microflora were significantly lower (P < 0.01) than the D values for this organism heat-treated immediately after inoculation. In beef systems, the D(55) values were significantly lower (P < 0.05) in the presence of a beef microflora than the D(55) values obtained in 'pure' culture under all temperature/storage combinations. However, in broth systems, there was no significant difference between the D(55) values obtained in 'pure' culture and the D(55) values obtained from systems containing beef microflora. CONCLUSIONS: Storage of Salm. Typhimurium DT104 significantly reduced the thermal resistance of the pathogen in beef and broth systems. In the presence of high numbers of a Gram-negative beef microflora, the heat sensitivity of the pathogen was further increased on beef surfaces but not in broth. SIGNIFICANCE AND IMPACT OF THE STUDY: Studies investigating the survival of Salm. Typhimurium DT104 in different food systems will help define safe food preservation processes and will aid in the elimination this pathogen from the food production environments.