Effect of NaCl and KCl on fate and growth/no growth interfaces of Listeria monocytogenes Scott A at different pH and nisin concentrations

AIMS: The fate of Listeria monocytogenes Scott A, was studied in broth, at different a(w)s (by adding NaCl or KCl from 0.0 to 1.4 mol l(-1)), pHs (from 4.0 to 7.3 by adding lactic acid), and nisin concentrations (from 0 to 100 IU ml(-1)). METHODS AND RESULTS: Increasing salt and nisin concentrations and decreasing pH resulted in lower growth rates and extended lag phases. At pH 4.5 no growth was observed while in presence of nisin and/or 1 mol l(-1) salts of both kinds, L. monocytogenes Scott A was inactivated. Equal-molar concentrations of NaCl or KCl (similar a(w)), exerted similar effects against L. monocytogenes in terms of lag phase duration, growth or death rate. The growth boundaries of L. monocytogenes Scott A at 5 degrees C were also estimated by growth/no growth turbidity data, modeled by logistic polynomial regression. The concordance of logistic models, were 99.6 and 99.8% for NaCl and KCl, respectively. CONCLUSIONS: The growth interfaces derived by both NaCl and KCl models were almost identical. Hence, NaCl can be replaced by KCl without risking the microbiological safety of the product. Increasing nisin concentrations markedly affected the interface resulting in a more inhibitory environment for L. monocytogenes Scott A. Low to medium salt concentrations (0.3-0.7 mol l(-1) of either NaCl or KCl) provided a protective effect against inhibition of L. monocytogenes Scott A by nisin. SIGNIFICANCE AND IMPACT OF THE STUDY: Modelling the growth boundaries not only contributes to the development of safer food by providing useful data, but can also be used to study interactions between factors affecting initiation of growth of pathogenic micro-organisms.     

Gaseous ozone treatment inactivates Listeria innocua in vitro

AIMS: To investigate the effects of ozone on inactivation of Listeria innocua on solid media. METHODS AND RESULTS: Suspensions of L. innocua ranging from 4.5 x 10(4 )- 6.4 x 10(4) CFU ml(-1) were inoculated onto potato dextrose agar (PDA, pH 5.6 and 6.8) and nutrient agar (NA, pH 6.0 and 6.8), then exposed to gaseous ozone. Variable factors included postinoculation standing time at 20 degrees C before exposure to ozone, ozone concentration, treatment duration and treatment temperature (5 or 20 degrees C). The interaction among ozone concentration, treatment duration, media and temperature in effecting changes in colony-forming units (CFU) was significant. The 100 nl l(-1) ozone treatment for 2 h reduced the microbial populations by 2-3 log CFU ml(-1). Cell viability decreased more rapidly on PDA than on NA. The average time to obtain a 2 log CFU ml(-1) reduction was 1.3 h at 20 degrees C and 2.5 h at 5 degrees C (P < 0.001). CONCLUSIONS: Gaseous ozone effectively inactivates L. innocua at concentrations of 50 and 100 nl l(-1) during short exposure times at both 5 and 20 degrees C. The Gompretz model can be utilized for determining the response of L. innocua to ozone over time. SIGNIFICANCE AND IMPACT OF THE STUDY: This study provides useful information on ozone inactivating Listeria spp., which may be imposed on ensuring quality and safety of horticultural produce and food products.     

Action of divergicin M35, a class IIa bacteriocin, on liposomes and Listeria

AIMS: The mode of action of divergicin M35, a class IIa bacteriocin, was studied against Listeria monocytogenes with sensitive (DivS) and resistant (DivM) phenotypes, as well as on synthetic phospholipid liposomes. METHODS AND RESULTS: Divergicin-induced release of 1,6-diphenyl-1,3,5-hexatriene (DPH) from zwitterionic (DMPC) and anionic (DMPC/DMPG, 4:1) liposomes, divergicin binding to liposomes, intracellular ATP concentration, cation efflux, cell affinity for hydrocarbons and cell lysis were measured and cell damage was visualized by fluorescence imaging and transmission electron microscopy. Divergicin M35 at 5 microg ml(-1) induced DPH efflux from anionic and zwitterionic liposomes at rates of about 2.58% and 1.61% per minute, respectively. DPH efflux rate from anionic liposomes was reduced by about 1.83% and 2.1% per minute in the presence of Li+ and Ca2+, respectively. Binding affinity of divergicin M35 to anionic and zwitterionic liposomes was about 86% and 63%, respectively. Intracellular ATP decreased in the sensitive and the resistant strains by 96.7% and 72.8%, respectively after 20 min of exposure to 5 microg ml(-1) divergicin M35. Lysis of the sensitive strain reached 57% in 18 h at a concentration of 5 microg ml(-1) when compared with the lysis of the divergicin-resistant strain (38.8%). The K+ and Na+ efflux from the divergicin-sensitive strain reached 87% and 80% of the total ion content within 5 min of exposure. This strain also showed higher affinity for hydrocarbons. CONCLUSIONS: The cell death of listerial strains upon addition of divergicin M35 could result from ATP depletion, K+ and Na+ efflux, and bacteriolysis. This triple biological effect was attenuated in the DivM strain. SIGNIFICANCE AND IMPACT OF THE STUDY: This study contributed to the understanding of the mode of action of divergicin M35, a pediocin-like bacteriocin.     

Susceptibility of food-borne bacteria to binary combinations of antimicrobials at selected a(w) and pH

AIM: To evaluate the antibacterial susceptibilities of food-borne bacteria to individual and binary mixtures of a synthetic antimicrobial agent with a natural phenolic compound. METHODS AND RESULTS: Antibacterial susceptibilities of Escherichia coli, Listeria innocua, Salmonella Typhimurium and Staphylococcus aureus to individual and binary mixtures of potassium sorbate with a phenolic compound (thymol, carvacrol, or eugenol) were evaluated, at selected water activity (a(w); 0.99 or 0.97) and pH (5.5 or 4.5). The bacteria studied were susceptible to the action of the antimicrobials individually with minimal inhibitory concentrations that varied from 800-ppm potassium sorbate for Staph. aureus at a(w) 0.99, and pH 5.5 to 100-ppm thymol or carvacrol for the four studied bacteria at a(w) 0.97 and pH 4.5. Several binary mixtures of potassium sorbate with thymol, carvacrol or eugenol inhibited bacterial growth. Antimicrobial agent inhibitory concentrations in the mixture varied among bacteria, additionally depending on the a(w) and the pH tested. CONCLUSIONS: Synergistic binary mixtures with fractional inhibitory concentration index <0.6 include 100- or 200-ppm potassium sorbate with 50- or 100-ppm thymol, carvacrol or eugenol. SIGNIFICANCE AND IMPACT OF THE STUDY: The synergistic combinations could be useful in reducing the amounts of antimicrobials needed to inhibit growth, thus diminishing consumer concerns regarding chemical preservatives.     

Influence of immunomodulation on the development of Listeria monocytogenes infection in aged guinea pigs

We investigated the impact of immunomodulation on the development of listeriosis within an aged population of guinea pigs after an intragastric challenge with Listeria monocytogenes. Supplementation with vitamin E for 35 days significantly increased the level of cytotoxic T cells (CD8(+)), while treatment with cyclosporin A resulted in a 25% decrease of CD8(+) T cells. In the animals receiving the low dose (10(2) CFU) of L. monocytogenes, 50% of the control-group animals became infected. Only 22% of animals receiving the orthomolecular dose of vitamin E became infected, whereas animals that were immunosuppressed had an infection rate of 89%. In the immunosuppressed group three animals (16%) developed listerial infection with a quantifiable bacterial level of 0.3-3 log CFU g(-1) of organ in the spleen and liver. In the high-dose study, the population of L. monocytogenes was consistently 1 log CFU g(-1) lower in the spleen or liver of the vitamin E-supplemented group, compared with the control and cyclosporin A-treated animals. At day 4, a significant increase in the levels of CD8(+) during listerial infection occurred in vitamin E-supplemented animals, suggesting an increased ability to produce CD8(+) T cells. The results suggest that immunomodulation of the host can influence listerial infection within an aged population of guinea pigs.     

Impact of selected Lactobacillus and Bifidobacterium species on Listeria monocytogenes infection and the mucosal immune response

The pathogenesis of Listeria monocytogenes depends on its ability to attach to and invade the gastrointestinal epithelium and subsequently withstand the host immune response. Despite a thorough understanding of the intracellular phase of infection, relatively little is known about how the pathogen behaves in the gastrointestinal tract and whether it is affected by the presence of host commensal microbiota. Lactobacillus and Bifidobacterium are two important genera of the human gut microbiota proposed to possess probiotic effects. Here we demonstrate that probiotic bacteria significantly inhibit subsequent listerial infection in an in vitro C2Bbe1 epithelial cell model. In the case of Lactobacilli, inhibition was due to a combination of acid production and secretion of an as yet unidentified protein. In the case of Bifidobacterium, inhibition was attributable to an extracellular proteinaceous secreted compound. In addition, we observed a significant reduction in interleukin-8 and an increase in IL-10 cytokines secreted from epithelial cells following probiotic pretreatment and subsequent infection with Listeria. A reduction in the infection of epithelial cells and an altered mucosal immune response suggests that probiotic bacteria could be of therapeutic benefit against listerial infection. This study infers a role for probiotic bacteria as an antagonist of Li. monocytogenes infection.     

An oral Salmonella vaccine promotes the down-regulation of cell surface Toll-like receptor 4 (TLR4) and TLR2 expression in mice

A single oral immunization with the Lon-protease-deficient Salmonella enterica serovar Typhimurium (strain CS2022) induced protective immunity in mice against a subcutaneous challenge with virulent Listeria monocytogenes as well as virulent Salmonella serovar Typhimurium. The populations of cell surface Toll-like receptor 4 (TLR4) and TLR2 on peritoneal macrophages decreased at week 6 after immunization. This population decrease was not reversed after a challenge with either Salmonella or Listeria. These results suggest that oral immunization with CS2022 induced immune tolerance correlated with the down-regulation of cell surface TLR expression. This down-regulation may in part account for the development of cross-protection against a Listeria challenge by immunization with CS2022.     

熏鸡加工过程中微生物分布分析

研究熏鸡加工过程中大肠菌群数、大肠埃希菌数和常见致病菌的分布情况。通过对生产流程关键控制环节抽样进行大肠菌群计数、大肠埃希菌计数、沙门氏菌检测、金黄色葡萄球菌检测和单核细胞增生李斯特氏菌检测,掌握熏鸡生产过程中的微生物分布。根据实验数据分析和熏鸡生产流程,研究熏鸡加工过程中生产原料环节和熏制环节的关键控制点作用。     

一多重耐药单增李斯特菌株的生长特性及毒力分析

为了解单增李斯特菌株耐药后可能发生的生物学变化,以哈市生肉中分离到的1株对17种抗生素耐受的单增李斯特菌株L.M.B8为研究对象,对其生长及毒力特性进行研究。结果显示,L.M.B8的生长及毒力特性均与标准菌株有明显差异。在NaC l浓度为0.5%~5%、pH值为4.0~10.0及温度为20~45℃范围内,L.M.B8的生长速度均明显高于标准菌株。L.M.B8对高浓度盐的敏感性高于标准菌株,且对温度的适应能力强于标准菌株。从生长曲线看,L.M.B8的对数生长期与稳定期均较标准菌株提前2~3 h,且其稳定期较标准菌株明显缩短。L.M.B8小鼠腹腔注射半数致死量（LD50）较标准菌株明显降低。该研究为进一步探讨单增李斯特菌的耐药性与其他生物学特性的相关性奠定基础。