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Changes in viability and macromolecular content of long-term batch cultures of Salmonella typhimurium measured by flow cytometry

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JOURNAL OF APPLIED MICROBIOLOGY 2000年第1期89卷 90-99页

作者： Turner, K Porter, J Pickup, R Edwards, C Univ Liverpool Sch Biol Sci Liverpool L69 7ZB Merseyside England Inst Freshwater Ecol Windermere England

Exposure of many Gram-negative bacteria to prolonged starvation induces alternative programmes of gene expression, along with a transition into a dormant condition sometimes referred to as a viable non-culturable (VBNC) slate. Knowledge of how pathogenic species respond to nutrient limitation is therefore important for their detection and dissemination. This study used flow cytometry, coupled with fluorescent dyes for viability and macromolecular content, to study the responses of the pathogen Salmonella typhimurium to prolonged batch culture. Statistical analysis of the flow cytometric data, together with total and culturable cell counts, failed to demonstrate a VBNC state in this pathogen, contrary to reports from other workers. Analysis of rRNA and protein content identified a small proportion of cells in 110 day-old cultures that represented an active sub-population. This observation may provide an explanation for the long-term survival properties of this organism during prolonged exposure to nutrient limitation, as well as the high degree of heterogeneity observed in labelled cells.

关键词：细菌蛋白质类/代谢集落计数微生物培养基流式细胞术荧光染料模型生物学模型统计学普罗匹定 RNA 细菌/代谢 RNA 核糖体/代谢鼠伤寒沙门菌/生长和发育鼠伤寒沙门菌/代谢鼠伤寒沙门菌/生理学

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Effects of simulated solar disinfection of water on infectivity of Salmonella typhimurium

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LETTERS IN APPLIED MICROBIOLOGY 2000年第4期31卷 284-288页

作者： Smith, RJ Kehoe, SC McGuigan, KG Barer, MR Med Sch Newcastle Upon Tyne Dept Microbiol & Immunol Newcastle Upon Tyne NE2 4HH Tyne & Wear England Royal Coll Surgeons Ireland Dept Phys Dublin 2 Ireland

To determine whether cells of Salmonella typhimurium rendered nonculturable by simulated salar disinfection retain infectivity for mice. Bacteria suspended in water were exposed to UVA irradiation for up to 8 h. Culturability, determined by colony forming unit and Most Probable Number counts, fell by six log(10) units, while cellular activity determined by the Kogure cell elongation test was retained by approximately 5% of the cells present after 8 h. Intraperitoneal doses of nonculturable cells and active but nonculturable (ABNC) cells exceeding the LD50 Of the test organism and BALB/c mouse host, respectively by 4 and 3 orders of magnitude failed to produce detectable infections, Culturable cells that had been irradiated for 1.5 h were less infective (virulent) than their nonirradiated counterparts. Nonculturable and ABNC cells of Salm. typhimurium produced by UVA irradiation do not retain infectivity for mice. Although ABNC cells could be produced by low cost solar disinfection systems, they do not appear to pose a potential infection hazard.

关键词：集落计数微生物培养基消毒/方法小鼠近交BALBC 沙门菌感染动物/微生物学鼠伤寒沙门菌/致病力鼠伤寒沙门菌/生理学鼠伤寒沙门菌/辐射效应日光紫外线毒力水微生物学水净化/方法动物女(雌)性小鼠

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Modulation of Salmonella infection by the lectins of Canavalia ensiformis (Con A) and Galanthus nivalis (GNA) in a rat model in vivo

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JOURNAL OF APPLIED MICROBIOLOGY 2000年第4期88卷 720-727页

作者： Naughton, PJ Grant, G Bardocz, S Pusztai, A Rowett Res Inst Aberdeen Scotland

The plant lectins, Concanavalin A (Con A) and Galanthus nivalis agglutinin (GNA) have been prefed to rats for 3 d pre- and 6 d postinfection with Salmonella typhimurium S986 or Salm. enteritidis 857. Con A significantly increased numbers of Salm. typhimurium S986 in the large intestine and in faeces, and severely impaired growth of the rats, more severely than is the case of infection with Salmonella typhimurium alone. Con A had much less effect on rats infected with Salm. enteritidis 857 only showing a significant increase in numbers in the colon, accompanied by intermittent increases of Salmonella in the faeces during the study. GNA significantly reduced pathogen numbers in the lower part of the small bowel and the large intestine of rats infected with Salm. typhimurium S986 and significantly improved rat growth. GNA had little effect on infection by Salm. enteritidis 857 with slight decreases in Salmonella numbers in the small intestine and large intestine and transient increases in the faeces.

关键词：伴刀豆球蛋白A/投药和剂量伴刀豆球蛋白A/代谢粪便/微生物学雪花莲属外源凝集素类/投药和剂量外源凝集素类/代谢甘露糖结合凝集素类植物凝集素类沙门菌感染动物/微生物学肠炎沙门菌/生长和发育肠炎沙门菌/代谢肠炎沙门菌/生理学鼠伤寒沙门菌/生长和发育鼠伤寒沙门菌/代谢鼠伤寒沙门菌/生理学体重增长动物人类男(雄)性大鼠

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Physiological effects of high hydrostatic pressure treatments on Listeria monocytogenes and Salmonella typhimurium

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JOURNAL OF APPLIED MICROBIOLOGY 2000年第2期88卷 202-212页

作者： Tholozan, JL Ritz, M Jugiau, F Federighi, M Tissier, JP Inst Natl Rech Agron Lab Proc & Technol Alimentaire Villeneuve Dascq France Ecole Natl Vet Nantes Unite Associee INRA Hyg Alimentaire Nantes France

The effect of a high hydrostatic pressure treatment on the Gram-positive Listeria monocytogenes strain Scott A and the Gram-negative Salmonella typhimurium strain Mutton (ATCC13 311) has been determined in stationary phase cell suspensions. Pressure treatments were done at room temperature for 10 min in sodium citrate (pH 5.6) and sodium phosphate (pH 7.0) suspension buffers. Increasing pressure treatments resulted in an exponential decrease of cell counts. Salmonella typhimurium suspended at low pH was more sensitive to pressure treatments. Progressive morphological changes were evident with the pressure increase. Cell lysis only appeared with the highest pressure treatments. Cell volume was not affected by pressure treatment. A progressive decrease of delta pH (pH(in) - pH(out)), intracellular potassium and ATP contents was demonstrated with the pressure increase. A parallel lowering of membrane potentials was measured.

关键词：腺苷三磷酸/代谢集落计数微生物培养基氢离子浓度流体静力压单核细胞增生利斯特菌/代谢单核细胞增生利斯特菌/生理学单核细胞增生利斯特菌/超微结构膜电位显微镜检查电子扫描钾/代谢鼠伤寒沙门菌/代谢鼠伤寒沙门菌/生理学鼠伤寒沙门菌/超微结构

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Heat inactivation of Salmonella typhimurium DT104 in beef as affected by fat content

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LETTERS IN APPLIED MICROBIOLOGY 2000年第6期30卷 461-467页

作者： Juneja, VK Eblen, BS ARS USDA Eastern Reg Res Ctr Wyndmoor PA 19038 USA

The heat resistance of an eight-strain cocktail of Salmonella typhimurium DT104 was determined at 58-65 degrees C in beef containing 7, 12, 18 or 24% fat. Inoculated beef was packaged in bags completely immersed in a circulating water bath and held at 58, 60, 62 . 5 and 65 degrees C for a predetermined length of time. The surviving cell population was enumerated by spiral plating heat-treated samples onto tryptic soy agar supplemented with 0 . 6% yeast extract and 1% sodium pyruvate. Preliminary studies on thermal inactivation of the Salmonellae isolates in chicken broth indicated no correlation between heat resistance and origin of the isolates. While linear survival curves were observed in chicken broth, inactivation kinetics in beef showed deviations from the first order kinetics, represented by an initial lag period or shoulder before any death occurred with time. Overall, increased fat levels in beef resulted in longer lag periods and lower D-values, suggesting that the lag periods must be taken into account and added to the D-values for calculating the time required at a specific temperature for achieving a specific lethality for Salm. typhimurium DT104 in beef. Thermal death times from this study will assist the retail food industry to design cooking regimes that ensure safety of beef contaminated with Salm. typhimurium DT104.

关键词：鸡烹饪/标准培养基脂肪类/分析食品微生物学/标准热温度肉制品/分析肉制品/微生物学鼠伤寒沙门菌/遗传学鼠伤寒沙门菌/生理学时间因素动物牛

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Infection by bacterial pathogens expressing type III secretion decreases luciferase activity: Ramifications for reporter gene studies

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BIOTECHNIQUES 2000年第3期29卷 514-+页

作者： Savkovic, SD Koutsouris, AS Wu, G Hecht, G Univ Illinois W Side VA Med Ctr Chicago IL 60612 USA Univ Penn Sch Med Philadelphia PA 19104 USA

Pathogenic microbes influence gene regulation in eukaryotic hosts. Reporter gene studies can define the roles of promoter regulatory sequences. The effect of pathogenic bacteria on reporter genes has not been examined. The aim of this study was to identify which reporter genes are reliable in studies concerning host gene regulation by bacterial pathogens expressing type III secrerory systems. Human intestinal epithelial cells, T-84. Caco-2 and HT-29, were transfected with plasmids containing luciferase (luc), chloramphenicol acetyltransferase (CAT) or beta-galactosidase (beta-gal) as reporter genes driven by the inducible interleukin-8 (IL-8) or constitutively active simian virus 40 (SV40) promoter. Cells were infected with enteropathogenic E. coli or Salmonella typhimurium, and the reporter activity was assessed. Luc activity significantly decreased following infection, regardless of the promoter. The activity of recombinant luc was nearly ablated by incubation with either EPEC or Salmonella in a cell-free system. Activity was partially preserved by protease inhibitors, and immunoblot analysis showed a decreased amount and molecular weight of recombinant luc, suggesting protein degradation. Neither beta-gal nor CAT activity was altered by infection. Disruption of type III secretion prevented the loss of luc activity. We conclude that CAT or beta-gal, but not luc, can be used as reliable reporter genes to assess the impact of pathogenic microbes, especially those expressing type III secretion on host cell gene regulation.

关键词：细胞系氯霉素O-乙酰转移酶/遗传学氯霉素O-乙酰转移酶/代谢上皮细胞/代谢上皮细胞/微生物学大肠杆菌/生理学基因报告肠/代谢肠/微生物学萤光素酶类/遗传学萤光素酶类/代谢质粒/遗传学重组蛋白质类/代谢鼠伤寒沙门菌/生理学转染 β半乳糖苷酶类/遗传学 β半乳糖苷酶类/代谢人类

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Comparison of the attachment of Escherichia coli O157:H7, Listeria monocytogenes, Salmonella typhimurium, and Pseudomonas fluorescens to lettuce leaves

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JOURNAL OF FOOD PROTECTION 2000年第10期63卷 1433-1437页

作者： Takeuchi, K Matute, CM Hassan, AN Frank, JF Univ Georgia Dept Food Sci & Technol Ctr Food Safety & Qual Enhancement Athens GA 30602 USA

Attachment of Escherichia coli O157:H7, Listeria monocytogenes, Salmonella Typhimurium, and Pseudomonas fluorescens on iceberg lettuce was evaluated by plate count and confocal scanning laser microscopy (CSLM). Attachment of each microorganism (similar to 10(8) CFU/ml) on the surface and the cut edge of lettuce leaves was determined. E. coli O157:H7 and L. monocytogenes attached preferentially to cut edges, while P. fluorescens attached preferentially to the intact surfaces. Differences in attachment at the two sites were greatest with L. monocytogenes. Salmonella Typhimurium attached equally to the two sites. At the surface, P. fluorescens attached in greatest number, followed by E. coli O157:H7, L. monocytogenes, and Salmonella Typhimurium. Attached microorganisms on lettuce were stained with fluorescein isothiocyanate and visualized by CSLM. Images at the surface and the cut edge of lettuce confirmed the plate count data. In addition, microcolony formation by P. fluorescens was observed on the lettuce surface. Some cells of each microorganism at the cut edge were located within the lettuce tissues, indicating that penetration occurred from the cut edge surface. The results of this study indicate that different species of microorganisms attach differently to lettuce structures, and CSLM can be successfully used to detect these differences.

关键词：细菌黏附/生理学细菌学技术集落计数微生物大肠杆菌O157/生理学莴苣/微生物学单核细胞增生利斯特菌/生理学显微镜检查共焦/方法荧光假单胞菌/生理学鼠伤寒沙门菌/生理学

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Molecular basis for the enterocyte tropism exhibited by Salmonella typhimurium type 1 fimbriae

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JOURNAL OF BIOLOGICAL CHEMISTRY 1999年第9期274卷 5797-5809页

作者： Thankavel, K Shah, AH Cohen, MS Ikeda, T Lorenz, RG Curtiss, R Abraham, SN Duke Univ Med Ctr Dept Pathol & Microbiol Durham NC 27710 USA Washington Univ Dept Biol St Louis MO 63130 USA Azabu Univ Sch Vet Med Dept Vet Med Kanagawa 229 Japan Washington Univ Sch Med Dept Pathol St Louis MO 63110 USA Washington Univ Sch Med Dept Med St Louis MO 63110 USA

Salmonella typhimurium exhibits a distinct tropism for mouse enterocytes that is linked to their expression of type 1 fimbriae, The distinct binding traits of Salmonella type 1 fimbriae is also reflected in their binding to selected mannosylated proteins and in their ability to promote secondary bacterial aggregation on enterocyte surfaces. The determinant of binding in Salmonella type I fimbriae is a 35-kDa structurally distinct fimbrial subunit, FimHs, because inactivation of fimHs abolished binding activity in the resulting mutant without any apparent effect on fimbrial expression. Surprisingly, when expressed in the absence of other fimbrial components and as a translational fusion protein with MalE, FimHs failed to demonstrate any specific binding tropism and bound equally to all cells and mannosylated proteins tested. To determine if the binding specificity of Salmonella type 1 fimbriae was determined by the fimbrial shaft that is intimately associated with FimHs, we replaced the amino-terminal half of FimHs with the corresponding sequence from Escherichia coli FimH (FimHE) that contains the receptor binding domain of FimHE, The resulting hybrid fimbriae bearing FimHES on a Salmonella fimbrial shaft exhibited binding traits that resembled that of Salmonella rather than E, coli fimbriae, Apparently, the quaternary constraints imposed by the fimbrial shaft on the adhesin determine the distinct binding traits of S. typhimurium type 1 fimbriae.

关键词：黏着素细菌/化学黏着素细菌/遗传学黏着素细菌/生理学黏着素大肠杆菌氨基酸序列细菌黏附碱基序列细胞系 DNA引物菌毛蛋白质类纤毛细菌/生理学肠/细胞学肠/微生物学肠/超微结构显微镜检查电子/方法分子序列数据诱变定点蛋白质生物合成重组融合蛋白质类/遗传学重组融合蛋白质类/代谢鼠伤寒沙门菌/生理学序列同源性氨基酸向性尿道上皮/细胞学尿道上皮/微生物学动物小鼠

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ISOLATION, CHARACTERIZATION AND COMPLEMENTATION OF SOLMONELLA-TYPHIMURIUM CHEMOTAXIS MUTANTS

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JOURNAL OF MOLECULAR BIOLOGY 1975年第2期97卷 225-&页

作者： ASWAD, D KOSHLAND, DE UNIV CALIF DEPT BIOCHEMBERKELEYCA 94720

A novel technique has been developed for selecting non-chemotactic mutants of Salmonella typhimurium that are defective in transmission of information from the chemoreceptor to the flagella. This technique utilizes a preformed gradient in a vertical column of liquid medium, which separates wild types, non-motile and motile but non-chemotactic into three regions. In the first application of the technique, 71 such mutants were obtained, of which 67 exhibited smooth motility and four had unco-ordinated motility. Complementation analysis, done by abortive transduction of recombination-defective recipients, indicates that there may be as many as six genetically distinct classes of che mutants in Salmonella .

关键词：趋化作用基因显性基因隐性遗传互补测验基因型突变重组遗传鼠伤寒沙门菌/分离和提纯鼠伤寒沙门菌/生理学种特异性

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EVIDENCE FOR AN S-ADENOSYLMETHIONINE REQUIREMENT IN CHEMOTACTIC BEHAVIOR OF SALMONELLA-TYPHIMURIUM

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JOURNAL OF MOLECULAR BIOLOGY 1975年第2期97卷 207-207页

作者： ASWAD, DW KOSHLAND, DE UNIV CALIF DEPT BIOCHEMBERKELEYCA 94720

The following evidence suggests that S -adenosyl- l -methionine is involved in regulating the chemotactic response of Salmonella typhimurium . (a) Methionine analogs that replace methionine as substrates in the S -adenosylmethionine synthetase reaction will also replace methionine in restoring chemotaxis to starved cells, while those that are non-substrates show no replacement activity. (b) Cycloleucine, a methionine analog that inhibits S -adenosylmethionine synthesis, produces the same behavioral change as methionine starvation. (c) S -adenosylmethionine is one of only two metabolites derived from methionine that are rapidly formed and present in high concentrations during incubation in chemotaxis medium. The presence of the other metabolite, spermidine, does not correlate with chemotactic behavior. (d) When methionine-starved, chemotactically wild type cells are given an exogenous supply of methionine, the S -adenosylmethionine pool is replaced in about the same time that spontaneous tumbling reappears. Likewise, the rapid turnover of S -adenosylmethionine correlates well with the rapid loss of tumbling that occurs when the exogenous supply of methionine is removed. A possible model for the role of S -adenosylmethionine is presented, which states that S -adenosylmethionine is necessary for maintaining the normal steady-state level of a hypothetical tumble regulator.

关键词：砷酸盐类/药理学趋化作用/药物作用氯霉素/药理学环亮氨酸/药理学甲硫氨酸/类似物和衍生物甲硫氨酸/代谢甲硫氨酸/药理学甲硫氨酸腺苷转移酶突变蛋白质生物合成/药物作用 S-腺苷甲硫氨酸/代谢 S-腺苷甲硫氨酸/药理学 S-腺苷甲硫氨酸/生理学鼠伤寒沙门菌/药物作用鼠伤寒沙门菌/生理学种特异性

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