Listeria Monocytogenes In Broth Research Articles

Elucidating antimicrobial mechanism of nisin and grape seed extract against Listeria monocytogenes in broth and on shrimp through NMR-based metabolomics approach

Nisin and grape seed extract (GSE) have been widely used as food preservatives; however, the mechanism against pathogens at molecular level has not been well elucidated. This work aimed to investigate their antimicrobial effect against Listeria monocytogenes and to elucidate the mechanism by NMR-based metabolomics. Nisin exhibited enhanced in vitro antilisterial effect when combined with GSE (4.49 log CFU/mL reduction). Marked change in cell membrane permeability was observed in the combination group using confocal laser scanning microscopy; this was verified by increased leakage of protein and nucleic acid. The underlying antimicrobial mechanism was revealed by NMR coupled with multivariate analysis. Significant decreases in threonine, cysteine, ATP, NADP, adenine were observed, whereas a few of metabolites such as lactic acid and γ-aminobutyric acid (GABA) increased after nisin-GSE treatment (P < 0.05). Pathway analysis further manifested that the nisin-GSE inhibited the survival of L. monocytogenes by blocking the TCA cycle, amino acid biosynthesis and energy-producing pathway. Lastly, nisin and GSE were applied to shrimp and binary combination showed remarkably antilisterial activity (1.79 log CFU/g reduction). GABA shunt and protein degradation from shrimp compensated the unbalanced glycolysis and amino acid metabolism by providing energy and carbon source for L. monocytogenes inoculated on shrimp. Thus, they were more tolerant to nisin and GSE stresses as compared to the broth-grown culture.

Thermal inactivation and sublethal injury kinetics of Salmonella enterica and Listeria monocytogenes in broth versus agar surface

The objective of the present study was to compare the thermal inactivation and sublethal injury kinetics of Salmonella enterica and Listeria monocytogenes in broth (suspended cells) and on solid surface (agar-seeded cells). A 3-strain cocktail of S. enterica or L. monocytogenes inoculated in broth or on agar was subjected to heating in a water bath at various set temperatures (55.0, 57.5 and 60.0°C for S. enterica and 60.0, 62.5 and 65°C for L. monocytogenes). The occurrence of sublethally injured cells was determined by comparing enumerations on nonselective (TSAYE) and selective (XLD or ALOA) media. Results showed that the inactivation curves obtained from selective media were log-linear, and significant shoulders (p<0.05) were observed on some of the inactivation curves from TSAYE media. The D-values derived from the total population were higher than those from the uninjured cells. Generally, cells on agar surface exhibited higher heat resistance than those in broth. For S. enterica, cell injury increased with the exposure time, no difference was observed when treated at temperatures from 55.0 to 60.0°C, while for L. monocytogenes, cell injury increased significantly with heating time and treatment temperature (from 60.0 to 65°C). Moreover, the degree of sublethal injury affected by thermal treatment in broth or on agar surface depended upon the target microorganism. Higher proportions of injured S. enterica cells were observed for treatment in broth than on agar surface, while the opposite was found for L. monocytogenes. The provided information may be used to assess the efficacy of thermal treatment processes on surfaces for inactivation of S. enterica and L. monocytogenes, and it provides insight into the sublethally injured survival state of S. enterica and L. monocytogenes treated in liquid or on solid food.

Growth and inactivation of Salmonella enterica and Listeria monocytogenes in broth and validation in ground pork meat during simulated home storage abusive temperature and home pan-frying.

Ground pork meat with natural microbiota and inoculated with low initial densities (1–10 or 10–100 CFU/g) of Salmonella enterica or Listeria monocytogenes was stored under abusive temperature at 10°C and thermally treated by a simulated home pan-frying procedure. The growth and inactivation characteristics were also evaluated in broth. In ground pork meat, the population of S. enterica increased by less than one log after 12-days of storage at 10°C, whereas L. monocytogenes increased by 2.3 to 2.8 log units. No unusual intrinsic heat resistance of the pathogens was noted when tested in broth at 60°C although shoulders were observed on the inactivation curves of L. monocytogenes. After growth of S. enterica and L. monocytogenes at 10°C for 5 days to levels of 1.95 log CFU/g and 3.10 log CFU/g, respectively, in ground pork meat, their inactivation in the burger subjected to a simulated home pan-frying was studied. After thermal treatment S. enterica was undetectable but L. monocytogenes was recovered in three out of six of the 25 g burger samples. Overall, the present study shows that data on growth and inactivation of broths are indicative but may underestimate as well as overestimate behavior of pathogens and thus need confirmation in food matrix conditions to assess food safety in reasonably foreseen abusive conditions of storage and usual home pan-frying of meat burgers in Belgium.

The effect of water soluble fat replacers and fat reduction on the growth of Lactobacillus sakei and Listeria monocytogenes in broth and pork liver paté

The major objective of this study was to evaluate the effect of water soluble fat replacers on the growth of bacteria of importance to processed meats. The first part of the study evaluated the effect of water soluble fat replacers (Beneo™ GR, Beneo™ HPX, STA-LITE WSIII™, Fibersol-2 and Nutriose) on the growth of Lactobacillus sakei and Listeria monocytogenes. Fibersol-2 appears to have the greatest potential for replacing fat without compromising microbiological stability as increase in its water phase concentration resulted in a significant increase in the lag phases of both Lb. sakei and L. monocytogenes. On the other hand, Beneo™ GR stimulated the growth of L. monocytogenes, implying that its use could compromise microbiological safety. In the second part of the study pork liver paté and low fat paté (30% less fat) were challenged with L. monocytogenes. The low fat paté was determined to be microbiologically less stable than the reference paté. This was a result of the low fat paté having significantly higher water activity and pH values than those of the reference paté. The results of this study are of importance to the meat processing industry to make more informed decisions when reducing and/or replacing fat.

Characterization of Mexican Coriander (Eryngium foetidum) Essential Oil and Its Inactivation of Listeria monocytogenes In Vitro and during Mild Thermal Pasteurization of Pineapple Juice

The aim of this work was to characterize the essential oil (EO) of Eryngium foetidum (EfEO) and assess its activity toward Listeria monocytogenes in broth and during thermal inactivation of the pathogen in pineapple juice. In this respect, EfEO was chemically characterized, and its antilisteria potential in broth as a function of pH, cell load, and EfEO concentration was assessed through a central composite design. Furthermore, the inactivation kinetics of L. monocytogenes in the juice were assessed by combining EfEO and low pasteurization temperatures. A total of 81 compounds were identified from EfEO. The reduction of pH and cell load increased EO activity. The use of only 15 ppm of EfEO during pasteurization of pineapple juice at 60°C reduced the time required for a 4-log reduction in L. monocytogenes CFU/ml by 74.9% (i.e., from 8.5 to 2.1 min) compared with treatment without EfEO. It could be concluded that EfEO activity toward L. monocytogenes increases with the reduction of pH and that it can be used at sublethal concentrations in combination with low temperatures in pineapple juice pasteurization. This study demonstrates that EO-assisted pasteurization is a promising strategy for the reduction of thermal impact during juice production. EfEO is easily available and compatible with many juices and is thus promising for industrial application.

The effect of pH and nitrite concentration on the antimicrobial impact of celery juice concentrate compared with conventional sodium nitrite on Listeria monocytogenes

The objectives of this study were to evaluate the impact of pH and nitrite from celery juice concentrate (CJ) on the growth of Listeria monocytogenes in broth and on ham slices, and to evaluate the impact of pH and nitrite from CJ on quality attributes of the ham. The pH of both broth and ham were increased by the addition of CJ. The CJ was less effective than conventional nitrite at 100mg/kg nitrite in broth, but in ham, the CJ treatments at both 100 and 200mg/kg resulted in growth of L. monocytogenes (p>0.05) similar to that of the conventional nitrite at the same concentrations. Reducing the pH of CJ before addition to the ham had greater impact on L. monocytogenes growth at 200mg/kg nitrite than at 100mg/kg. Celery juice concentrate may increase meat product pH which could have implications for the antimicrobial impact of nitrite in some products.

Survival of bacteria in less than thorough cooked, brine-injected steaks

Bacteria that may include enteric pathogens are introduced into muscle tissues when they are injected with brine. Consequently, steaks of such non-intact, non-comminuted tissues should be cooked to temperatures sufficient to render them microbiologically safe. Studies were conducted to identify the minimum safe temperature for cooking those meats. Suspensions of Escherichia coli and Listeria innocua at numbers about 8 log cfu/ml, in broth or in brines containing 2% or 5% of each of NaCl and sodium tripolyphosphate that were not supplemented or were supplemented with 2% soy protein or 2% emulsified sunflower oil were injected into the centres of 3 cm thick steaks. Groups of four steaks injected with each type of suspension were cooked to temperatures at the centres of 50, 55, 60, 65 or 70 °C. Bacteria were recovered from samples from the centres of steaks, on agars that did or did not allow resuscitation of injured cells of each organism. The results indicated that E. coli but not L. innocua was injured by the brines; that supplementing brines with protein or oil did not protect the organisms from injury or inactivation by heating; and that cooking to a central temperature > 60 ⩽ 65 °C was sufficient to inactivate all bacteria in the meat. The centres of steaks were subsequently injected with 0.1 ml portions of five strain cocktails of E. coli O157:H7 or Listeria monocytogenes in broth containing a dye and the bacteria at numbers >8 log cfu/ml. Steaks were cooked to temperatures of 63, 64 or 65 °C, with holding for 0, 1 or 2 min after cooking before excision of all the dyed tissue from each steak and enumeration of bacteria from the tissue on resuscitating media. The results indicated that cooking to 65 °C without holding would be sufficient to reduce numbers of E. coli O157:H7 or L. monocytogenes in non-intact, non-comminuted steaks by ⩾7 log units.

Determination of the effect of plant essential oils obtained by supercritical fluid extraction on the growth and viability of Listeria monocytogenes in broth and food systems using flow cytometry

The antimicrobial properties of oregano, rosemary and laurel extracts obtained by supercritical fluid extraction were investigated by examining their influence on the growth and viability of Listeria monocytogenes in laboratory medium and broccoli juice at 30 and 8 °C. Important decreases in the L. monocytogenes population were shown in presence of all the extracts obtained from rosemary and one oregano extract. The counts were reduced below the level of detection after 4 h of exposure at 30 °C in laboratory medium. A bactericidal effect was observed also when L. monocytogenes was exposed to rosemary at 30 °C and 8 °C in broccoli juice. Significant reductions in growth rate and an increase in lag phase of L. monocytogenes were observed in presence of some of the laurel and oregano extracts at both temperatures. Flow cytometry was used as a rapid method to determine the antibacterial effect of supercritical extracts and the physiological state of L. monocytogenes. Bacterial viability performed by dual staining of L. monocytogenes with SYTO 9 and propidium iodide revealed three different cell populations, specifically, living, dead and compromised cells. Live cell percentage decreased with the time of exposure, whereas the percentage of compromised cells remained constant and the dead cells increased in the same period.

Survival and Growth of Listeria monocytogenes in Broth as a Function of Temperature, pH, and Potassium Lactate and Sodium Diacetate Concentrations

The objective of this study was to determine the antimicrobial effect of a combination of potassium lactate and sodium diacetate (0, 1.8, 3, and 4.5%; PURASAL P Opti.Form 4, 60% solution) on the survival and growth of Listeria monocytogenes Scott A in pH-adjusted broth (5.5, 6.0, 6.5, and 7.0) stored at 4, 10, 17, 24, 30, and 37°C. Appropriate dilutions of broth were enumerated by spiral plating on tryptose agar and counted with an automated colony counter. Growth data were iteratively fit, using nonlinear regression analysis to a three-phase linear model, using GraphPad PRISM. At pH 5.5, the combination of lactate-diacetate fully inhibited (P < 0.001) the growth of L. monocytogenes at all four levels and six temperatures. At pH 6.0, addition of 1.8% lactate-diacetate reduced (P < 0.001) the specific growth rate of L. monocytogenes and increased lag time; however, 3 and 4.5% completely inhibited the growth at the six temperatures studied. Efficacy of the lactate-diacetate mixture was decreased as pH increased and incubation temperature increased. Thus, at pH 6.5, at least 3% was required to retard (P < 0.001) the growth of L. monocytogenes in broth. There was a limited effect of the lactate-diacetate level on the specific growth rate of the pathogen at pH 7.0. However, 1.8 and 3% significantly lengthened the lag time at 4 and 10°C. These results suggest that 1.8% of lactate-diacetate mixture can be used as a substantial hurdle to the growth of L. monocytogenes when refrigerated temperatures are maintained for products with pH less than 6.5.

The contribution of bacteriocin to inhibition of Listeria monocytogenes by Carnobacterium piscicola strains in cold-smoked salmon systems.

To study the importance of bacteriocin production for the antilisterial effect of a bacteriocinogenic Carnobacterium piscicola strain A9b on growth of Listeria monocytogenes in broth and cold-smoked salmon systems. Acriflavin treatment of strain A9b resulted in loss of bacteriocin production and of immunity to carnobacteriocin B2. Two plasmids present in the wild-type were lost in the variant that was also more sensitive to bavaricin and leucocin A than the wild-type indicating cross-resistance to class IIa bacteriocins. The growth rate of the bac- mutant was higher than that of the wild-type at 5 and 37 degrees C but not at 25 or 30 degrees C. In salmon juice the maximum cell density of L. monocytogenes was suppressed 3 and 6 log by co-culture with C. piscicola A9b bac- and bac+, respectively, as compared with the control. Sterile filtered cultures of C. piscicola A9b bac- caused a limited suppression of the maximum cell density of L. monocytogenes similar to that observed when sterile buffer was added in equal amounts. Semi-purified carnobacteriocin B2 caused a 3.5 log decline in viable cell count after 6 day of incubation in cold-smoked salmon juice at 5 degrees C. High resistance level to carnobacteriocin B2 was observed for L. monocytogenes cells exposed to semi-purified and in situ produced carnobacteriocin B2. The presence of bacteriocin production in C. piscicola enhances its inhibition of L. monocytogenes. Due to the emergence of resistance, a bacteriocin negative lactic acid bacteria may be more suited for practical use as a bioprotective agent against L. monocytogenes in ready-to-eat foods.

Relevance of microbial interactions to predictive microbiology

Microbial interaction can be ignored in predictive microbiology under most conditions. We show that interactions are only important at high population densities, using published data on inhibition of growth of Listeria monocytogenes in broth. Our analysis using growth models from predictive microbiology indicated that interactions only occur at population densities of ∼10 8 cfu/ml of the protective cultures. Spoilage is evident at these levels, except for fermented foods. In bacterial colonies, diffusion limitation acts as a constraint to growth. We have shown that these constraints only become important after large outgrowth of colonies (in the order of 5-log growth in Lactobacillus curvatus colonies), which depends on the initial inoculation density. Intra-colony interactions play an important role under these conditions. There is no large outgrowth of colonies when the initial inoculation densities are high and broth culture growth can be used to approximate colony growth.

Combined antimicrobial effect of nisin and a listeriophage against Listeria monocytogenes in broth but not in buffer or on raw beef

The effect of nisin and listeriophage LH7, alone and in combination, on the growth and survival of two strains of Listeria monocytogenes in broth and two model food systems, with appropriate controls, was determined. Growth curves for both bacterial strains in tryptic soy broth incubated at 7 or 30 °C, and with the addition of nisin and/or listeriophage at lag, mid-exponential or early stationary phase, were obtained by measuring absorbance at 550 nm. Numbers of mixed populations of both L. monocytogenes strains in phosphate buffered saline (pH 5.5) and on vacuum-packaged fresh beef, both stored for 4 weeks at 4 °C, and with the addition of nisin and/or listeriophage, were determined. This was achieved by plating appropriately diluted samples on both Tryptic Soy Agar and Modified Oxford Agar to determine both L. monocytogenes numbers and the presence of sub-lethal injury. In broth nisin alone, reduced levels or prevented growth of the two strains under the conditions studied, but regrowth to levels equivalent to those of untreated cells, occurred. Listeriophage LH7 alone, on the other hand, had no effect in broth under the conditions studied. Notably, however, a mixture of nisin and listeriophage displayed a combined effect in broth and reduced levels of cells substantially without regrowth under the conditions studied. In both model food systems only nisin appeared to be active, in a manner consistent with existing literature, and no combined action was apparent. The use of nisin and listeriophage has potential to control L. monocytogenes in foods but a further understanding of the interactions in this complex system needs to be achieved before it could be applied practically.

The combined effect of nisin, leucocin F10, pH, NaCl and EDTA on the survival of Listeria monocytogenes in broth

The combined effect of the bacteriocins nisin (1–2100 IU/ml) and leucocin F10 (1–2100 AU/ml), pH (4.7–6.5), NaCl (0.7–4.5% w/l), ethylene diaminetetraacetic acid disodium salt (EDTA, 0.08–4.72 mmol/l) and inoculum level (10 3–10 8 cfu/ml) on the survival of a pool of three strains of Listeria monocytogenes in broth was evaluated in three factorial experiments. Several factor combinations were found to prevent growth. Logistic regression analysis of the categorical data (survival/no survival) was used to generate predictive models for the probability of survival in 0.01 ml ( P0.01) or 1 ml ( P1). Predicted and observed probabilities of survival were not significantly different in 72% and 68.9% of treatments for P0.01 and P1, respectively. Unsafe predictions were obtained in 9.4% and 14.8% of treatments for P0.01 and P1, respectively. Nisin had a major effect on the probability of survival but the addition of leucocin F10 was necessary to prevent the survival of L. monocytogenes. Lower pH values significantly decreased the probability of survival, while NaCl and EDTA had only a minor effect. Doses of bacteriocins >250 AU/ml, pH <5.6 and EDTA>0.2 mmol/l (0.074 g/l) were needed to reliably prevent survival of Listeria monocytogenes.

Survival and Growth of Listeria monocytogenes in Broth Supplemented with Sodium Chloride and Held at 4 and 13 °C

Growth and survival of Listeria monocytogenes strains Scott A and V7 at 4 and 13°C in tryptose broth containing 0 to 100 g/L NaCl was determined. Both strains grew at all concentrations of NaCl at 13°C; however, 90 and 100 g/L NaCl delayed the onset of growth by strain V7 more than that of strain Scott A. The maximum population achieved by strain V7 in the presence of 100 g/L NaCl was less than that of strain Scott A. When incubation was at 4°C, strain V7 initially grew in the presence of 0 to 60 g/L NaCl, but during extended incubation the population decreased in the presence of 70 to 100 g/L NaCl, with the final population being less than the initial population when the medium contained 90 and 100 g/L NaCl. Strain Scott A, at 4°C, grew at all concentrations of NaCl tested although at 90 and 100 g/L there was an initial decrease in population. After some growth, another decrease in population occurred midway through a 12-d incubation, which was followed by an increase and another decrease in population. A three-way analysis of variance showed that storage time, strain of L. monocytogenes, concentration of NaCl, and their interactions were significant in determining behavior of the bacterium at the two holding temperatures.