Trends Cell Biol, 2005 Jan, 15(1), 2 - 5 Ubiquitination of intracellular bacteria: a new bacteria-sensing system? Veiga E, Cossart P. Ubiquitination is a protein modification generally used by cells to tag proteins that are destined for proteasomal degradation . In a recent article, Perrin et al . reported that the ubiquitination system has a role in the recognition of bacterial pathogens in the cytosol of mammalian cells . They showed that polyubiquitinated proteins accumulate on the surface of cytosolic Salmonella typhimurium . In macrophages, but not epithelial cells, proteasomes become associated with the surface of cytosolic bacteria . The authors proposed that the ubiquitin-proteasome machinery might be implicated indirectly in bacterial clearance. Int Immunopharmacol, 2005 Feb, 5(2), 359 - 68 Effects of Brazilian and Bulgarian propolis on bactericidal activity of macrophages against Salmonella Typhimurium; Orsi RO et al.; Propolis has been used in folk medicine since ancient times due to its many biological properties, such as antimicrobial, antiinflammatory, antioxidant, immunomodulatory activities, among others . Macrophages play an important role in the early phase of Salmonella infection . In this work, macrophages were prestimulated with Brazilian or Bulgarian propolis and subsequently challenged with Salmonella Typhimurium at different macrophage/bacteria ratio . After 60 min of incubation, cells were harvested with Triton-X to lyse the macrophages . To assess the bactericidal activity, the number of colony-forming units (CFU) of S . typhimurium was determined by plating 0.1 mL in Mueller Hinton agar . After 24 h, CFU were counted, and the percentage of bactericidal activity was obtained . Propolis from Brazil and Bulgaria enhanced the bactericidal activity of macrophages, depending on its concentration . Brazilian propolis seemed to be more efficient than that from Bulgaria, because of their different chemical composition . In Bulgaria, bees collect the material mainly from the bud exudate of poplar trees, while in Brazil, Baccharis dracunculifolia DC . was shown to be the main propolis source . Our data also showed that the increased bactericidal activity of macrophages involved the participation of oxygen (H(2)O(2)) and nitrogen (NO) intermediate metabolites. Cancer Biother Radiopharm, 2004 Oct, 19(5), 649 - 57 A Combination of flk1-Based DNA Vaccine and an Immunomodulatory Gene (IL-12) in the Treatment of Murine Cancer; Keke F et al.; Aim: The aim of this study was to investigate the antivasculature effects and the antitumor effects of combining attenuated Salmonella typhimurium vaccine strain encoding murine vascular endothelial growth factor (VEGF) receptor-2 (flk1) with plasmid DNA vector encoding the murine IL-12 (mIL-12) gene . Methods: Mouse models of Gl261 glioblastoma were treated with combining orally given attenuated Salmonella typhimurium vaccine strain encoding flk1 with direct intratumoral injection of a nonviral plasmid DNA vector encoding the murine IL-12 (mIL-12) gene . The volumes of tumors were observed . Cytolytic T lymphocyte (CTL) response was measured by a 4-hour(51) Cr release assay, vessle density and tumorcell proliferation were observed by immunostaining, and tumor apoptosis was determined by TUNELstaining . Results: Compared to mice receiving single agent therapy, received either oral immunization flk1-based vaccine only or the therapeutic gene-IL-12 plasmid DNA only or those in the control group, the combination therapy groups developed a strong CTL response and showed more significantly inhibited tumor growth, apoptosis of tumor cells, and reduced neovascularization and cell proliferation in these mice . Conclusions: The therapy of attenuated Salmonella typhimurium vaccine strain encoding flk1 combined with the interleukin-12 gene has significant synergistic effect against tumors. Ann N Y Acad Sci, 2004 Dec, 1028, 113 - 21 DNA minigene vaccination for adjuvant neuroblastoma therapy; Lode HN et al.; The disruption of self-tolerance against neuroblastoma is the ultimate goal of an effective DNA-vaccine . We demonstrate the induction of protective immunity against syngeneic murine NXS2 neuroblastoma in A/J mice following vaccination with tyrosine hydroxylase (TH)-derived antigens . Oral gene delivery was accomplished using an attenuated strain of Salmonella typhimurium as a carrier harboring vectors encoding for mouse tyrosine hydroxylase (mTH) antigens . Vaccination was effective in protecting animals from a lethal challenge with wild-type NXS2 tumor cells . These findings were extended by comparing efficacy of mTH minigene vaccines with a minigene vaccine comprising three novel epitopes isolated fom NXS2 neuroblastoma cells . For this purpose, MHC class I was immunoprecipitated from NXS2 cell lysates, and peptides were eluted and examined in tandem-mass spectrometry analysis . This led to the identification of three novel natural MHC class I peptide ligands: TEALPVKLI, from ribonucleotide reductase M2; NEYIMSLI, from Ser/Thr protein phosphatase 2A; and FEMVSTLI, of unknown origin . Two minigenes were constructed, one encoding for the three novel epitopes and the second for three known mTH-derived epitopes with high predicted binding affinity to MHC class I, by cloning them into the mammalian expression vector pCMV-3FUB . Immunized mice showed a reduction in primary tumor growth and the absence of spontaneous liver metastasis in the majority of animals . Importantly, there was no significant difference between the two minigenes, suggesting that, compared with tumor peptide isolation, mTH epitope prediction is similarly effective for designing efficient DNA-minigene vaccines . In summary, these findings establish proof of the concept that disruption of self-tolerance against neuroblastoma-associated epitopes may be an effective adjuvant therapeutic strategy. Proc Natl Acad Sci U S A . 2005 Jan 11; {Epub ahead of print} Tumor-targeting bacterial therapy with amino acid auxotrophs of GFP-expressing Salmonella typhimurium; Zhao M et al.; Here we report a genetically modified bacteria strain, Salmonella typhimurium A1, selected for anticancer activity in vivo . The strain grows in tumor xenografts . In sharp contrast, normal tissue is cleared of these bacteria even in immunodeficient athymic mice . S . typhimurium A1 is auxotrophic (Leu/Arg-dependent) but apparently receives sufficient support from the neoplastic tissue to grow locally . Whether additional genetic lesions are present is not known . In in vitro infection, the GFP-expressing bacteria grew in the cytoplasm of PC-3 human prostate cancer cells and caused nuclear destruction . These effects were visualized in cells labeled with GFP in the nucleus and red fluorescent protein in the cytoplasm . In vivo, the bacteria caused tumor inhibition and regression of xenografts visualized by whole-body imaging . The bacteria, introduced i.v . or intratumorally, invaded and replicated intracellularly in PC-3 prostate cancer cells labeled with red fluorescent protein grafted into nude mice . By day 15, S . typhimurium A1 was undetectable in the liver, lung, spleen, and kidney, but it continued to proliferate in the PC-3 tumor, which stopped growing . When the bacteria were injected intratumorally, the tumor completely regressed by day 20 . There were no obvious adverse effects on the host when the bacteria were injected by either route . The S . typhimurium A1 strain grew throughout the tumor, including viable malignant tissue . This result is in marked contrast to bacteria previously tried for cancer therapy that were confined to necrotic areas of the tumor, which may account, in part, for the strain's unique antitumor efficacy. Biochemistry, 2005 Jan 18, 44(2), 675 - 83 Active Site Plasticity of Endonuclease V from Salmonella typhimurium; Feng H et al.; Base deamination is a major type of DNA damage under nitrosative stress . Endonuclease V initiates repair of deaminated base damage by making a nucleolytic incision one nucleotide away from the 3' side of the lesion . Within the endonuclease V family, the substrate specificities are different from one enzyme to another . In this study, we investigated deamination lesion cleavage activities of endonuclease V from the macrophage-residing pathogen, Salmonella typhimurium . Salmonella endonuclease V exhibits limited turnover on cleavage of deoxyinosine- and xanthosine-containing DNA . Binding analysis indicates that this single-turnover property is caused by tight binding to nicked products . The nicking activity is similar between the double-stranded deoxyinosine- and deoxyxanthosine-containing DNA . Cleavage rates are not affected by bases opposite the deoxyinosine or deoxyxanthosine lesions . The enzyme is also active on single-stranded deoxyinosine- and deoxyxanthosine-containing DNA . Unlike endonuclease V from Thermotoga maritima, Salmonella endonucleae V can only turnover deoxyuridine-containing DNA to a limited extent when substrate is in excess . Binding analysis indicates that Salmonella endonuclease V achieves tight binding to deoxyuridine-containing DNA, a property that distinguishes it from Thermotoga endonuclease V . Cleavage analysis on mismatch-containing DNA also indicates that the active site of Salmonella endonuclease V can accommodate pyrimidine-containing mismatches, resulting in more comparable cleavage of pyrimidine- and purine-containing mismatches . This comprehensive DNA cleavage and binding analysis reveals the plastic nature in the active site of Salmonella endonuclease V, which allows the enzyme to enfold both purine and pyrimidine deaminated lesions or base pair mismatches. Zhonghua Shi Yan He Lin Chuang Bing Du Xue Za Zhi, 2004 Sep, 18(3), 238 - 42 {Salmonella typhimurium as DNA delivery vehicle for DNA-mediated immunization.}; He P et al.; BACKGROUND: To study whether the live attenuated AroA-auxotrophic mutant of Salmonella (S.) typhimurium (SL7207) could be used as DNA delivery vehicle to induce more efficient immune response by using the eukaryotic expression plasmid pCMV-beta as report gene . METHODS: Murine peritoneal macrophages were infected with SL7207(pCMV-beta) in vitro, then the expression of the beta-gal were detected by X-gal staining or RT-PCR . After mice were orally immunized with SL7207(pCMV-beta), the expression of beta-gal in the lymphoid tissue were tested by RT-PCR, humoral responses were tested by ELISA, splenic lymphocyte proliferation were tested by 3H-TdR incorporation and cytotoxic T lymphocyte reaction were tested by JAM test . RESULTS: The results indicated that the plasmid pCMV-beta could be delivered by SL7207 into the nucleus of the murine macrophages efficiently and expressed well in vitro; after mice received oral immunizations with attenuated S.typhimurium SL7207 harboring plasmid pCMV-beta mice, the expression of beta-gal could be detected in the spleen, mesenteric lymph nodes and Peyers patches of the mice . Furthermore, the experiments demonstrated that specific humoral immune responses and cell-mediated immune responses were successfully induced in these immunized mice . Compared with the naked DNA vaccination, SL7207 (pCMV-beta) oral immunization were more efficient in inducing cellular immune responses . CONCLUSIONS: Attenuated Salmonella typhimurium SL7207 could be used as DNA delivery vehicle for oral immunization, which have the ability to deliver the antigen-encoding DNA specifically to APC directly for inducing the specific immune response being dominant with cellular immune response. Phytomedicine, 2004 Nov, 11(7-8), 666 - 72 Antimicrobial and cytotoxic activity of 18 prenylated flavonoids isolated from medicinal plants: Morus alba L., Morus mongolica Schneider, Broussnetia papyrifera (L.) Vent, Sophora flavescens Ait and Echinosophora koreensis Nakai; Sohn HY et al.; Antimicrobial activity of the 18 prenylated flavonoids, which were purified from five different medicinal plants, was evaluated by determination of MIC using the broth microdilution methods against four bacterial and two fungal microorganisms (Candida albicans, Saccaromyces cerevisiae, Escherichia coli, Salmonella typhimurium, Staphylococcus epidermis and S . aureus) . Papyriflavonol A, kuraridin, sophoraflavanone D and sophoraisoflavanone A exhibited a good antifungal activity with strong antibacterial activity . Kuwanon C, mulberrofuran G, albanol B, kenusanone A and sophoraflavanone G showed strong antibacterial activity with 5-30 microg/ml of MICs . Morusin, sanggenon B and D, kazinol B, kurarinone, kenusanone C and isosophoranone were effective to only gram positive bacteria, and broussochalcone A was effective to C . albicans . IC50 values of papyriflavonol A, kuraridin, sophoraflavanone D, sophoraisoflavanone A and broussochalcone A in HepG2 cells were 20.9, 37.8, 39.1, 22.1, and 22.0 microg/ml, respectively . These results support the use of prenylated flavonoids in Asian traditional medicine to treat microbial infection and indicate a high potential for prenylated flavonoids as antimicrobial agents as well as anti-inflammatory agents. J Immunol, 2005 Jan 15, 174(2), 1020 - 6 Bacterial Lipoprotein Induces Resistance to Gram-Negative Sepsis in TLR4-Deficient Mice via Enhanced Bacterial Clearance; O'brien GC et al.; TLRs are highly conserved pathogen recognition receptors . As a result, TLR4-deficient C3H/HeJ mice are highly susceptible to Gram-negative sepsis . We have previously demonstrated that tolerance induced by bacterial lipoprotein (BLP) protects wild-type mice against polymicrobial sepsis-induced lethality . In this study, we assessed whether pretreatment of C3H/HeJ mice with BLP could induce resistance to a subsequent Gram-negative Salmonella typhimurium infection . Pretreatment with BLP resulted in a significant survival benefit in TLR4-deficient C3H/HeJ mice (p < 0.0002 vs control C3H/HeJ) after challenge with live S . typhimurium (0.25 x 10(6) CFU/mouse) . This survival benefit was associated with enhanced bacterial clearance from the circulation and in the visceral organs (p < 0.05 vs control C3H/HeJ) . Furthermore, pretreatment with BLP resulted in significant increases in complement receptor type 3 (CR3) and FcgammaIII/IIR expression on polymorphonuclear neutrophils (PMNs) and macrophages (p < 0.05 vs control C3H/HeJ) . There was impaired bacterial recognition and phagocytosis in TLR4-deficient mice compared with wild-type mice . However, a significant augmented uptake, ingestion, and intracellular killing of S . typhimurium by PMNs and peritoneal macrophages was evident in BLP-pretreated C3H/HeJ mice (p < 0.05 vs control C3H/HeJ) . An up-regulation of inducible NO synthase and increased production of intracellular NO were observed in peritoneal macrophages from BLP-pretreated C3H/HeJ mice (p < 0.05 vs control C3H/HeJ) . Depletion of PMNs did not diminish the beneficial effects of BLP with regard to both animal survival and bacterial clearance . These results indicate that BLP, a TLR2 ligand, protects highly susceptible TLR4-deficient mice from Gram-negative sepsis via enhanced bacterial clearance. J Rheumatol, 2005 Jan, 32(1), 86 - 92 Outer membrane protein of salmonella is the major antigenic target in patients with salmonella induced reactive arthritis; Saxena S et al.; OBJECTIVE: We previously reported that Salmonella typhimurium was the triggering agent in one-third of our patients with sporadic enteric reactive arthritis (ReA) and undifferentiated spondyloarthropathy (uSpA) . The antigens recognized by the synovial T cells in Salmonella induced ReA are not known . We investigated the immunodominant antigens in Salmonella ReA . METHODS: Synovial fluid mononuclear cells (SFMC) from 53 patients with ReA/uSpA were cultured with crude lysate of S . typhimurium . In 20 patients, the triggering agent was found to be Salmonella (stimulation index, SI, > 2.5) . For cell fractionation of S . typhimurium, the sonicated crude lysate was separated by ultracentrifugation into a cytoplasmic supernatant (CYT) and membrane pellet (OMP) . The CYT was further separated on SDS-PAGE and blotted onto nitrocellulose membrane for proliferation assays . SFMC from 20 patients with Salmonella ReA/uSpA were stimulated with OMP, CYT, and cytosolic fractions of S . typhimurium, and proliferation was measured by thymidine incorporation . Quantitation of antigen-specific cells in SF was by intracellular interferon-g staining in 7 patients and paired peripheral blood (PB) in 5 patients after stimulation with crude Salmonella lysate, CYT, and OMP . RESULTS: Out of 20 patients with Salmonella ReA/uSpA, the SFMC showed a significant proliferation to OMP in 19 patients and CYT in 17 patients . The median SI of OMP (8.2, range 2.8-52.5) was significantly higher (p < 0.0005) than for the CYT (4.9, 2.7-18.8) . Fifty percent of the patients showed proliferative response to cytosolic fractions of < 60 kDa . The mean antigen-specific T cell frequency was also higher with OMP (0.68% +/- 0.59%) than CYT (0.53% +/- 0.62%) but this was not statistically significant . Compared to PB, the OMP-specific cells were 7.5 times more numerous in the SF (p < 0.05) . CONCLUSION: The cellular immune response in Salmonella ReA/uSpA is directed predominantly against the OMP and low molecular weight proteins in the cytosolic fraction. Biofactors, 2004, 22(1-4), 123 - 5 Antimutagenicity of Japanese traditional herbs, gennoshoko, yomogi, senburi and iwa-tobacco; Hiramatsu N et al.; The multistage induction theory is generally regarded as the mechanism of carcinogenesis . In order to prevent the initiation stage of carcinogenesis, it is meaningful to discover the functional components of edible plants . The objective of this research was to test the antimutagenicity of the functional components of several typical traditional herbs used in Japan . The traditional herbs, gennoshoko (Geranium nepalense var . thunbergii), yomogi (Artemisia vulgaris var . indica), senburi (Swertia japonica), iwa-tobacco (Conandron ramondioides), sarunokoshikake (Elfvingia applanata), kanzo (Glycyeehiza uralensis Fisch) and matatabi (Actinidia polygama) were examined by Ames mutagenesis assay test with Salmonella typhimurium TA98 and TA100 against mutagens, Trp-P-1, Trp-P-2 and B(a)P . The water-soluble components or volatile oil of the herbs were extracted in boiling water . The extracts of gennoshoko showed strong antimutagenicity against B(a)P with S . typhimurium TA98 and TA100, as well as Trp-P-1 and Trp-P-2 with S . typhimurium TA98 . Yomogi, senburi and iwa-tobacco were also proved to have good antimutagenicity against Trp-P-1 and Trp-P-2 with S . typhimurium TA98, but weaker antimutagenicity against B(a)P . Other herbs did not show any obvious antimutagenicity against these mutagens . In addition, the volatile oil of yomogi also had remarkable antimutagenic effect against the mutagens we used with S . typhimurium TA98. Biochem Biophys Res Commun, 1976 Dec 20, 73(4), 1025 - 9 Metabolic epoxidation of trans-4-acetylaminostilbene: a protective mechanism against its activation to a mutagen; Glatt HR et al.; Trans-4-acetylaminostilbene is activated by liver preparations to mutagens for Salmonella typhimurium . Since this compound is metabolized to the trans-alpha,beta-epoxide and since many epoxides are ultimate mutagens, this epoxide was tested for direct mutagenicity . It was, however, found to be non-mutagenic, and, in contrast to the parent compound, the epoxide was no longer activated by liver preparations to mutagens . The same was found for the beta-ketone and for the threo-alpha,beta-dihydrodiol, which are formed metabolically from trans-4-acetylaminostilbene and from its alpha,beta-epoxide . 4-Acetylaminobibenzyl showed a very weak mutagenic activity in the presence of the liver preparation . Thus, it is important to realize that where epoxides are formed from compounds which are known to be metabolized to mutagens, they are not necessarily responsible for the mutagenicity . Epoxidation may even prevent the possibility of bioactivation to mutagens. Natl Toxicol Program Tech Rep Ser, 2004 Sep, (519), 1 - 274 NTP Toxicology and Carcinogensis Studies of STODDARD SOLVENT IIC (CAS NO . 64742-88-7) in B6C3F(1) Mice (Inhalation Studies); National Toxicology Program; Stoddard solvent (white spirit/mineral spirit) is the most widely used solvent in the paint industry . It is used as a dry cleaning agent; as an extraction, cleaning, and degreasing solvent; and as a solvent in aerosols, paints, wood preservatives, asphalt products, lacquers, and varnishes . Stoddard solvent IIC was nominated by the International Union, United Auto Workers, for carcinogenicity testing because of the large volume used in industrial and other settings . Male and female F344/N rats and B6C3F(1) mice were exposed to Stoddard solvent IIC (greater than 99% pure) by inhalation for 2 weeks, 3 months, or 2 years . Genetic toxicology studies were conducted in Salmonella typhimurium and mouse peripheral blood erythrocytes . 2-Week Study in Rats Groups of five male and five female rats were exposed to Stoddard solvent IIC by inhalation at concentrations of 0, 138, 275, 550, 1,100, or 2,200 mg/m(3), 6 hours per day, 5 days per week for 16 days . All rats survived to the end of the study, and mean body weights of all exposed groups were similar to those of the chamber controls . Liver weights of males exposed to 550 mg/m(3) or greater and of females exposed to 275 mg/m(3) or greater were increased . Minimal diffuse cytoplasmic vacuolization of hepatocytes of the liver occurred in all females exposed to 2,200 mg/m(3) . 2-Week Study in Mice Groups of five male and five female mice were exposed to Stoddard solvent IIC by inhalation at concentrations of 0, 138, 275, 550, 1,100, or 2,200 mg/m(3), 6 hours per day, 5 days per week for 17 days . All mice survived to the end of the study, and mean body weights of all exposed groups were similar to those of the chamber controls . Liver weights of males and females exposed to 275 mg/m(3) or greater were significantly increased . Cytomegaly of the liver occurred in all males and females exposed to 2,200 mg/m(3) . 3-Month Study in Rats Groups of 10 male and 10 female rats were exposed to Stoddard solvent IIC by inhalation at concentrations of 0, 138, 275, 550, 1,100, or 2,200 mg/m(3), 6 hours per day, 5 days per week for 14 weeks . All rats survived to the end of the study, and the final mean body weight of females exposed to 275 mg/m(3) was greater than that of the chamber controls . The relative kidney, liver, and testis weights of all exposed groups of males and the absolute kidney weights of males exposed to 550 mg/m(3) or greater were increased . The sperm motility of 550 mg/m(3) or greater males was significantly decreased . The incidences of renal tubule granular casts were significantly increased in males exposed to 550 mg/m(3) or greater, and the severities of renal tubule hyaline droplet accumulation, granular casts, and regeneration increased with increasing exposure concentration in males . The incidences of goblet cell hypertrophy of the nasal respiratory epithelium in males and females exposed to 2,200 mg/m(3) were significantly increased . Sperm motility was decreased in males exposed to 550 mg/m(3) or greater . 3-Month Study in Mice Groups of 10 male and 10 female mice were exposed to Stoddard solvent IIC by inhalation at concentrations of 0, 138, 275, 550, 1,100, or 2,200 mg/m(3), 6 hours per day, 5 days per week for 14 weeks . Mean body weights of exposed groups were similar to those of the chamber controls, but liver weights of males exposed to 2,200 mg/m(3) were significantly increased . The sperm motility of 2,200 mg/m(3) males was significantly decreased . This reduction in sperm motility, while statistically significant, is probably of modest importance as studies in mice have found that fertility is unaffected by motility decreases of less than 40% . The incidences of hematopoietic cell proliferation of the spleen in all exposed groups of females were greater than that in the chamber controls . 2-Year Study in Rats Groups of 50 male and 50 female rats were exposed to Stoddard solvent IIC by inhalation at concentrations of 0, 138 (males), 550, 1,100, or 2,200 (females) mg/m(3), 6 hours per day, 5 days per week for 104 to 105 weeks . Additional groups of 10 males and 10 females were exposed to the same concentrations for 3 months for renal toxicity analyses . Survival in the top exposure concentration groups of males and females was significantly less than that of the chamber controls . Mean body weights of exposed males and females were similar to those of the chamber controls . Cell proliferation analyses were performed in the left kidney of males and females after 3 months of exposure . The mean numbers of labeled cells and the labeling indices in males exposed to 550 and 1,100 mg/m(3) were significantly increased . The amount of alpha2u-globulin in the right kidney of males increased with increasing exposure concentration . Also, the incidences of granular casts and cortical tubule degeneration and regeneration were generally increased in exposed males, as was the severity of hyaline droplets . These effects did not occur in females . At 2 years, the incidences of benign and benign or malignant pheochromocytoma (combined) of the adrenal medulla occurred with positive trends in males, and the incidences in the 550 and 1,100 mg/m(3) groups were significantly increased . Due to increased incidences of renal tubule hyperplasia in males at 2 years, extended kidney evaluations were conducted; a slightly increased incidence of renal tubule adenoma occurred in the 1,100 mg/m(3) group . Nonneoplastic lesions related to Stoddard solvent IIC exposure occurred in the kidney of males . 2-Year Study in Mice Groups of 50 male and 50 female mice were exposed to Stoddard solvent IIC by inhalation at concentrations of 0, 550, 1,100, or 2,200 mg/m(3), 6 hours per day, 5 days per week for 105 weeks . Survival of exposed mice was similar to that of the chamber controls . Mean body weights of exposed females were greater than those of the chamber controls . The incidences of hepatocellular adenoma occurred with a positive trend in females, and the incidence of multiple hepatocellular adenoma in females exposed to 2,200 mg/m(3) was significantly increased . However, the incidences of hepatocellular adenoma or carcinoma (combined) and hepatocellular carcinoma alone in exposed males and females were not significantly increased . Genetic Toxicology Stoddard solvent IIC was tested for mutagenicity in Salmonella typhimurium strains TA97, TA98, TA100, and TA1535, with and without S9 metabolic activation enzymes; all results were negative . In vivo, the frequency of micronucleated erythrocytes was assessed in peripheral blood samples from male and female B6C3F(1) mice after 3 months of inhalation exposure to Stoddard solvent IIC, and results were negative . Conclusions Under the conditions of these 2-year inhalation studies, there was some evidence of carcinogenic activity of Stoddard solvent IIC in male F344/N rats based on increased incidences of adrenal medulla neoplasms; the slightly increased incidences of renal tubule adenoma may have been related to Stoddard solvent IIC exposure . There was no evidence of carcinogenic activity of Stoddard solvent IIC in female F344/N rats exposed to 550, 1,100, or 2,200 mg/m(3) . There was no evidence of carcinogenic activity of Stoddard solvent IIC in male B6C3F(1) mice exposed to 550, 1,100, or 2,200 mg/m(3) . There was equivocal evidence of carcinogenic activity of Stoddard solvent IIC in female B6C3F(1) mice based on increased incidences of hepatocellular adenoma; this slight increase was associated with increased body weight in exposed females . Exposure of male rats to Stoddard solvent IIC resulted in nonneoplastic lesions of the kidney characteristic of alpha2u-globulin accumulation . Synonyms: Medium aliphatic solvent naphtha (petroleum); white spirit. Natl Toxicol Program Tech Rep Ser, 2004 Dec, (516), 1 - 292 NTP Toxicology and Carcinogensis Studies of 2-METHYLIMIDAZOLE (CAS NO . 693-98-1) in B6C3F(1) Mice (Feed Studies); National Toxicology Program; 2-Methylimidazole is used in the manufacture of pharmaceuticals, photographic chemicals, dyes and pigments, agricultural chemicals, and rubber . It has been identified as a by-product in foods and has been detected in mainstream and sidestream tobacco smoke . 2-Methylimidazole was nominated by the National Cancer Institute for a long-term study because of the high potential for human exposure and a lack of carcinogenicity studies in rodents . Male and female F344/N rats and B6C3F(1) mice were exposed to 2-methylimidazole (99.5% pure) in feed for 2 years . Fifteen-day and 14-week toxicity studies of 2-methylimidazole in F344/N rats and B6C3F(1) mice are reported in NTP Toxicity Report No . 67 (NTP, 2004) . Genetic toxicity studies were conducted in Salmonella typhimurium, rat and mouse bone marrow cells, and mouse peripheral blood . 2-YEAR STUDY IN RATS Groups of 60 male and 60 female rats were fed diets containing 0, 300, 1,000, or 3,000 ppm 2-methylimidazole (males) or 0, 1,000, 2,500, or 5,000 ppm 2-methylimidazole (females) (equivalent to average daily doses of approximately 13, 40, or 130 mg 2-methylimidazole/kg body weight to males and 50, 120, or 230 mg/kg to females) for 106 weeks . Ten male and 10 female rats were necropsied at 6 months . Additional groups of 20 male and 20 female special study rats were exposed to the same concentrations for 8 days or 14 weeks and were evaluated for clinical chemistry, liver enzyme activity, and organ weights . Survival of 2,500 ppm females was significantly less than that of the controls . The mean body weights of 3,000 ppm males and 2,500 and 5,000 ppm females were generally less than those of the controls during most of the study . Feed consumption by 5,000 ppm females was less than that by the control group . The hematology results at 6 months indicated that exposure of rats to 2-methylimidazole induced a decreased erythron that was characterized as microcytic, normochromic, and nonresponsive . The thyroid hormone data indicated that rats administered 2-methylimidazole developed alterations in thyroid hormone concentrations; serum thyroxine and triiodothyronine concentrations were decreased, and thyroid stimulating hormone levels were increased . In general, the thyroid hormone effects were most pronounced early in the study and ameliorated with time . The results for the tissue enzyme content analyses of these 2-year feed studies indicated that exposure of rats to 2-methylimidazole induced an increase in total hepatic UDP-glucuronosyltransferase at all time points evaluated through 6 months . The thyroid gland weights of 3,000 ppm males and 2,500 and 5,000 ppm females were significantly increased at 6 months . At 6 months, two 5,000 ppm female rats had a thyroid gland follicular cell adenoma . The incidences of follicular cell adenoma, follicular cell carcinoma, and adenoma or carcinoma (combined) in the thyroid gland of 5,000 ppm females were significantly greater that those in the controls at 2 years . The incidence of follicular cell adenoma or carcinoma (combined) in the thyroid gland occurred with a positive trend in males . The incidences of follicular cell hyperplasia in all exposed groups of rats were significantly increased at 6 months and 2 years . The incidences of follicle mineralization of the thyroid gland in all exposed groups, except 300 ppm males at 6 months and in 1,000 and 5,000 ppm females at 2 years, were significantly greater than those of the controls . In the liver, the incidences of hepatocellular adenoma or carcinoma (combined) in the two highest exposure groups of males and females exceeded the historical ranges for controls, and the incidences of hepatocellular adenoma in females occurred with a positive trend . The incidences of bile duct hyperplasia and granulomatous inflammation were increased in females, as were those of mixed cell focus in males and females . The incidence of granulomatous inflammation of the spleen in 5,000 ppm females was significantly increased . Lower body weights of female rats exposed to 5,000 ppm likely contributed to the decreased incidences of mammary gland fibroadenoma, pituitary gland adenoma, and clitoral gland adenoma in this group . 2-YEAR STUDY IN MICE Groups of 60 male and 60 female mice were fed diets containing 0, 625, 1,250, or 2,500 ppm 2-methylimidazole (equivalent to average daily doses of approximately 75, 150, or 315 mg/kg to males and 80, 150, or 325 mg/kg to females) for 105 weeks . Ten male and 10 female mice were necropsied at 6 months . Additional groups of 20 male and 20 female special study mice were exposed to the same concentrations for 8 days or 14 weeks and were evaluated for clinical chemistry, liver enzyme activity, and organ weights . Survival of all exposed groups of mice was similar to that of the control groups . The mean body weights of 1,250 and 2,500 ppm males and 2,500 ppm females were less than those of the controls during most of the study . Feed consumption by all exposed groups of mice was similar to that by the control groups . The hematology results at 6 months indicated that exposure of mice to 2-methylimidazole induced a decreased erythron that was characterized as macrocytic, normochromic to hypochromic, and responsive . The thyroid gland weights of 2,500 ppm male and female mice and 1,250 ppm females were increased at 6 months . The incidence of follicular cell adenoma in the thyroid gland of 2,500 ppm males was significantly greater than that in the control group at 2 years . Follicular cell hypertrophy of the thyroid gland occurred in most exposed mice at 6 months, and the incidences of this lesion were significantly increased in the 1,250 and 2,500 ppm groups at 2 years; the incidences of follicular cell hyperplasia were significantly increased in 2,500 ppm males and females at 2 years . The liver weights of 2,500 ppm female mice were significantly increased at 6 months . The incidences of hepatocellular adenoma occurred with positive trends in males and females and the incidences were significantly increased in the 2,500 ppm groups . The incidence of hepatocellular carcinoma was significantly increased in 1,250 ppm males and exceeded the historical control range in 2,500 ppm males . The incidences of hepatocellular adenoma or carcinoma (combined) were significantly increased in all exposed groups of males . The incidences of hepatocellular karyomegaly in 2,500 ppm males at 6 months and in 1,250 and 2,500 ppm males at 2 years, of hepatocellular cytoplasmic alteration in 1,250 and 2,500 ppm males at 2 years, and Kupffer cell pigmentation in 2,500 ppm males at 2 years were significantly increased . In the spleen, the incidences of hematopoietic cell proliferation in all exposed groups of males and in 2,500 ppm females were significantly increased at 6 months and 2 years . Pigmentation was present in most 1,250 and 2,500 ppm mice at 6 months, and the incidences of this lesion were significantly increased in all exposed groups of males and in 1,250 and 2,500 ppm females at 2 years . The incidences of bone marrow hyperplasia were significantly increased in 1,250 and 2,500 ppm male mice at 2 years . Renal proximal tubule pigmentation was present in most 2,500 ppm male mice at 6 months and 2 years . The responses in the spleen, bone marrow, and kidney were considered to be related to the responsive anemia . In males, the incidences of chronic active inflammation of the epididymis at 1,250 and 2,500 ppm, sperm granuloma at 2,500 ppm, and of germinal epithelial atrophy of the testis at 1,250 and 2,500 ppm were significantly increased at 2 years . GENETIC TOXICOLOGY 2-Methylimidazole was negative in the S . typhimurium mutation assay when tested in strains TA97, TA98, TA100, and TA1535, with and without S9 activation enzymes . Testing of 2-methylimidazole in vivo for induction of chromosomal damage, as measured by micronucleated erythrocyte frequency, produced mixed results . When administered by intraperitoneal injection three times at 24-hour intervals, 2-methylimidazole produced negative results in bone marrow micronucleus tests in rats and mice . However, in the 14-week study of 2-methylimidazole, a significant exposure-related increase in the frequency of micronucleated normochromatic erythrocytes was noted in peripheral blood of male and female mice . Exposure concentration-related increases in the percentage of micronucleated polychromatic erythrocytes in peripheral blood was also seen in male and female mice in the 14-week study . CONCLUSIONS Under the conditions of this 2-year feed study, there was some evidence of carcinogenic activity of 2-methylimidazole in male F344/N rats based on increased incidences of thyroid gland follicular cell neoplasms . The increased incidences of hepatocellular neoplasms in males may have been related to exposure . There was clear evidence of carcinogenic activity of 2-methylimidazole in female F344/N rats based on increased incidences of thyroid gland follicular cell neoplasms . The increased incidences of hepatocellular adenoma in females may have been related to exposure . There was some evidence of carcinogenic activity in male B6C3F(1) mice based on increased incidences of thyroid gland follicular cell adenoma and hepatocellular neoplasms . There was some evidence of carcinogenic activity in female B6C3F(1) mice based on increased incidences of hepatocellular adenoma . Exposure to 2-methylimidazole resulted in nonneoplastic lesions in the thyroid gland and liver of male rats; the thyroid gland, liver, and spleen of female rats; the thyroid gland, liver, spleen, bone marrow, kidney, epididymis and testes of male mice; and the thyroid gland and spleen of female mice . Synonyms: Imidazole,2-methyl; 2-MeI; 2-methylglyoxaline; 2-MI; 2-MZ. Mutagenesis . 2004 Dec 29; {Epub ahead of print} Evaluation of the potential genotoxicity of the phosphate binder lanthanum carbonate; Damment SJ et al.; Lanthanum was evaluated for potential genotoxicity using a range of in vitro assays (as the carbonate) in the presence and absence of post-mitochondrial fraction (S9) and in vivo in three independent tests for mutagenicity and clastogenicity (as the carbonate and chloride) . The drug was devoid of mutagenic activity in bacterial assays (maximum concentration 5000 microg/plate) using a range of test strains (Salmonella typhimurium TA1535, TA1537, TA1538, TA98, TA100 and TA102 and Escherichia coli WP2 uvrA and WP2 uvrA pkm101) . No effects were seen in the hgprt gene mutation assay in Chinese hamster ovary cells in the presence of S9 . In the absence of S9, sporadic increases in revertant numbers were not dose-related or reproducible in subsequent experiments and hence were concluded to be chance events . In an in vitro chromosome aberration assay using Chinese hamster ovary cells, chromosome damage in the presence and absence of S9 (concentration 200-5000 microg/ml) was attributed to overt cell toxicity . To confirm this, a comprehensive in vivo evaluation of the drug was performed . Negative results were obtained in two independent rodent micronucleus tests . In the first mice were given oral doses (of carbonate) up to 2000 mg/kg, in the second rats were given a single i.v . bolus injection (of chloride) up to 0.1 mg/kg . Negative results were also obtained in a rat liver unscheduled DNA synthesis assay after treatment for 28 days with i.v . bolus injections (of chloride) up to 0.1 mg/kg/day . In these in vivo studies lanthanum plasma concentrations were >3000 times higher than the steady-state peak plasma concentration observed in dialysis patients given therapeutic doses of lanthanum carbonate . It can be concluded that lanthanum is not genotoxic and that lanthanum carbonate is unlikely to present a latent hazard in therapeutic use. J Environ Sci Health B, 2004, 39(5-6), 861 - 70 Salmonella enterica serovar Typhimurium hilA-lacZY fusion gene response to iron chelation or supplementation in rich and minimal media; Rishi P et al.; Virulence expression of Salmonella enterica serovar Typhimurium under iron limited condition was measured by beta-galactosidase (beta-gal) assay using a hilA-lacZY fusion strain and calculated as Miller units . hilA-lacZY beta-galactosidase assays were performed in brain heart infusion (BHI) and minimal media (M9), after iron chelation with 2, 2-dipridyl and iron-supplementation respectively . Before performing virulence assays, concentrations of iron in the media were estimated using ferrozine . Iron content was found to be more in BHI (42.6 microg dL(-1)) as compared to M9 (10.03 microg dL(-1)) . beta-gal activity of Salmonella Typhimurium in BHI was generally less than that observed in M9 . After exposure to various combinations of iron chelator in BHI, hilA-lacZY activity only increased at the highest concentration of chelator (2001 microM) but decreased in M9 media for all iron concentrations when compared to controls with no iron amendment . These results indicate that iron availability may influence S . Typhimurium hilA expression. Drug Metab Pharmacokinet, 2002, 17(1), 1 - 22 Genetically Engineered Bacterial Cells Co-expressing Human Cytochrome P450 with NADPH-cytochrome P450 Reductase: Prediction of Metabolism and Toxicity of Drugs in Humans; Fujita K et al.; Genetically engineered bacterial cells expressing human cytochrome P450 (CYP) have been developed as new tools to predict the metabolism and toxicity of drugs in humans . There are various host cells for the heterologous expression of a form of CYP . Among them, bacterial cells such as Escherichia coli (E . coli) have advantages with regard to ease of use and high yield of protein . CYP protein could be first expressed by the modification of the N-terminal amino acid sequence in E . coli cells in 1991 . Since then, many forms of human CYP have been successfully expressed in E . coli cells . Since the E . coli cells do not possess endogeneous electron transport systems to support the full catalytic activity of CYP, E . coli strains co-expressing both human CYP and NADPH-cytochrome P450 reductase (OR) have been established . Each form of CYP expressed in the E . coli cells efficiently catalyzed the oxidation of a representative substrate at an efficient rate, indicating that the OR was sufficiently expressed to support the catalytic activity of CYP . According to the studies performed so far, the modification of the N-terminal amino acid sequence of CYP did not seem to affect the catalytic properties of CYP . The human CYP expressed in the E . coli cells were applicable for studies to determine a metabolic pathway(s) of drugs and to estimate kinetic parameters of drug metabolism by human CYP . Drug-drug interactions caused by inhibition of the metabolism of drugs by human CYP could also be examined by in vitro inhibition studies with CYP expressed in the E . coli cells . Recently, human CYP was co-expressed with the OR in Salmonella typhimurium (S . typhimurium) cells used for mutation assay (Ames test) by applying the technology for the expression of human CYP and the OR in E . coli cells, to evaluate whether chemicals including drugs are metabolically activated by human CYP and show mutagenicity . These strains of bacteria are considered as useful tools to study the metabolism and the toxicity of drugs in humans. Vet Immunol Immunopathol, 1980 Aug, 1(3), 277 - 86 Intestinal immunity following a single intraperitoneal immunisation in lambs; Husband AJ; A single intraperitoneal injection of ovalbumin in oil adjuvant in young lambs has been shown to result in the appearance in the intestinal lamina propria of antibody-containing cells, most of which contained antibody of IgA specificity . Intraperitoneal immunisation of lambs with a Salmonella typhimurium vaccine during the suckling period provided protection against postweaning challenge with live organisms . This response was shown to be associated with specific IgA antibody in intestinal secretion. J Environ Monit, 2005 Jan, 7(1), 60 - 6 Epub 2004 Dec 08. In vitro genotoxicity of exhaust emissions of diesel and gasoline engine vehicles operated on a unified driving cycle; Liu YQ et al.; Acetone extracts of engine exhaust particulate matter (PM) and of vapor-phase semi-volatile organic compounds (SVOCs) collected from a set of 1998-2000 model year normal emitter diesel engine automobile or light trucks and from a set of 1982-1996 normal emitter gasoline engine automobiles or light trucks operated on the California Unified Driving Cycle at 22 {degree}C were assayed for in vitro genotoxic activities . Gasoline and diesel PM were comparably positive mutagens for Salmonella typhimurium strains YG1024 and YG1029 on a mass of PM extract basis with diesel higher on a mileage basis; gasoline SVOC was more active than diesel on an extracted-mass basis, with diesel SVOC more active on a mileage basis . For chromosomal damage indicated by micronucleus induction in Chinese hamster lung fibroblasts (V79 cells), diesel PM expressed about one-tenth that of gasoline PM on a mass of extract basis, but was comparably active on a mileage basis; diesel SVOC was inactive . For DNA damage in V79 cells indicated by the single cell gel electrophoresis (SCGE) assay, gasoline PM was positive while diesel PM was active at the higher doses; gasoline SVOC was active with toxicity preventing measurement at high doses, while diesel SVOC was inactive at all but the highest dose. J Agric Food Chem, 2004 Dec 29, 52(26), 8255 - 60 Chemical compositions and antibacterial effects of essential oils of Turkish oregano (Origanum minutiflorum), bay laurel (Laurus nobilis), Spanish lavender (Lavandula stoechas L.), and fennel (Foeniculum vulgare) on common foodborne pathogens; Dadalioglu I et al.; Chemical compositions and inhibitory effects of essential oils of Turkish oregano (Origanum minutiflorum O . Schwarz & P . H . Davis), bay laurel (Laurus nobilis L.), Spanish lavender (Lavandula stoechas subsp . stoechas L.), and fennel (Foeniculum vulgare Mill.) on Escherichia coli O157:H7, Listeria monocytogenes, Salmonella typhimurium, and Staphylococcus aureus were determined . After the essential oils were applied on the foodborne pathogens at doses of 0 (control), 5, 10, 20, 30, 40, 50, and 80 microL/mL, the resultant numbers of cells surviving were counted . Results revealed that all essential oils exhibited a very strong antibacterial activity against the tested bacteria (P < 0.05) . Gas chromatography-mass spectrophotometry analyses revealed that carvacrol (68.23%), 1,8-cineole (60.72%), fenchone (55.79%), and trans-anethole (85.63%) were the predominant constituents in Turkish oregano, bay laurel, Spanish lavender, and fennel essential oils, respectively. Chin J Dig Dis, 2004, 5(2), 72 - 5 Therapeutic vaccination against Helicobacter pylori infection with attenuated recombinant Salmonella typhimurium urease B subunit and catalase in mice; Li GQ et al.; OBJECTIVE: To investigate the effects of oral immunization with attenuated recombinant Salmonella typhimurium urease B subunit and catalase vaccines in the treatment of Helicobacter pylori infection in a H . pylori infected mouse model . METHODS: Thirty C57BL/6 mice were randomized into three groups and challenged twice with oral administration of H . pylori within 3 days . Four weeks after the second challenge, the mice were immunized by oral administration of attenuated recombinant S . typhimurium urease B subunit (group A), attenuated recombinant S . typhimurium catalase (group B) or saline (group C), and all mice were killed 4 weeks later . The stomachs were collected for a rapid urease test, modified Giemsa staining and quantitative culture to observe the density of H . pylori, hematoxylin-eosin staining was performed to assess the presence of inflammation and lymphocytes from the spleen were used for the lymphoproliferation assay . RESULTS: The gastric H . pylori density of groups A, B and C was 1.58 x 10(5) c.f.u./g, 4.88 x 10(5) c.f.u./g and 1.92 x 10(6) c.f.u./g, respectively . The H . pylori density was significantly decreased in the therapeutic groups (P < 0.05) . No significant inflammation was found in any group of mice . The lymphoproliferation assays of groups A and B were positive . CONCLUSION: Immunization with oral attenuated recombinant S . typhimurium urease B subunit and catalase vaccines is effective in reducing the density of H . pylori colonization. Wei Sheng Yan Jiu, 2004 Sep, 33(5), 591 - 4 {Fluoroquinolone resistance mutations in topisomerase genes of Salmonella typhimurium isolates}; Guo Y et al.; OBJECTIVE: Mutations in topisomerase genes were main cause of the resistence of Salmonella typhimurium to fluoroquinolone . METHODS: The MICs of three Salmonella typhimurium isolates X2, X7, X11 to ciprofloxacin were above 32 microg/ml, 0.38 microg/ml and 0.023 microg/ml, respectively . The genetic alterations in four topisomerase genes, gyrA, gyrB, parC, and parE were detected by multiplex PCR amplimer conformation analysis in these three strains . RESULTS: X2 isolate showed both gyrA mutations (Ser83-->Phe, Asp87-->Asn) and parC mutation (Ser80-->Arg) . X7 isolate showed a single gyrA mutation (Ser83-->Phe) and X11 isolate had no changes in all of the four quinolone resistance genes, gyrA, gyrB, parC, and parE . X7 isolate with a single gyrA mutation was less resistant to ciprofloxacin than X2 with double gyrA mutations and an additional parC mutation . CONCLUSION: GyrA and parC genes play important role of the resistence of Salmonella typhimurium to ciprofloxacin. World J Gastroenterol, 2005 Jan 7, 11(1), 114 - 7 Construction of a recombinant attenuated Salmonella typhimurium DNA vaccine carrying Helicobacter pylori hpaA; Xu C et al.; AIM: To construct a recombinant attenuated Salmonella typhimurium DNA vaccine carrying Helicobacter pylori hpaA gene and to detect its immunogenicity . METHODS: Genomic DNA of the standard H pylori strain 17 874 was isolated as the template, hpaA gene fragment was amplified by polymerase chain reaction (PCR) and cloned into pUCmT vector . DNA sequence of the amplified hpaA gene was assayed, then cloned into the eukaryotic expression vector pIRES through enzyme digestion and ligation reactions . The recombinant plasmid was used to transform competent Escherichia coli DH5alpha, and the positive clones were screened by PCR and restriction enzyme digestion . Then, the recombinant pIRES-hpaA was used to transform LB5000 and the recombinant plasmid isolated from LB5000 was finally used to transform SL7207 . After that, the recombinant strain was grown in vitro repeatedly . In order to identify the immunogenicity of the vaccine in vitro, the recombinant pIRES-hpaA was transfected to COS-7 cells using Lipofectamine2000, the immunogenicity of expressed HpaA protein was detected with SDS-PAGE and Western blot . RESULTS: The 750-base pair hpaA gene fragment was amplified from the genomic DNA and was consistent with the sequence of H pylori hpaA by sequence analysis . It was confirmed by PCR and restriction enzyme digestion that H pylori hpaA gene was inserted into the eukaryotic expression vector pIRES and a stable recombinant live attenuated Salmonella typhimurium DNA vaccine carrying H pylori hpaA gene was successfully constructed and the specific strip of HpaA expressed by pIRES-hpaA was detected through Western blot . CONCLUSION: The recombinant attenuated Salmonella typhimurium DNA vaccine strain expressing HpaA protein with immunogenicity can be constructed and it may be helpful for further investigating the immune action of DNA vaccine in vivo. Histochem Cell Biol . 2004 Dec 18; {Epub ahead of print} Salmonella Typhimurium infection in the porcine intestine: evidence for caspase-3-dependent and -independent programmed cell death; Schauser K et al.; The normal intestinal epithelium is renewed with a turnover rate of 3-5 days . During Salmonella infection increased cell loss is observed, possibly as a result of programmed cell death (PCD) . We have, therefore, studied the effects of Salmonella Typhimurium infection on three elements involved in PCD: caspase-3 activation, c-Jun phosphorylation on serine 63 (both detected by immunocytochemistry), and DNA fragmentation (detected by TUNEL reaction), using a pig jejunal loop model . Additionally, we used nuclear staining for detecting signs of classical apoptosis . Activated caspase-3 was detected in scattered epithelial cells and the number of positive cells increased with increasing times of exposure to Salmonella (P<0.0001) . An increase in phospho-c-Jun in epithelial cells was already detectable 5 min after infection and often occurred in cells that appeared not to be invaded by the organism . Changes in caspase-3 activation and c-Jun phosphorylation were most marked in the proximal region of the jejunum . Although rarely observed in the epithelium, proper TUNEL-positive cells were frequently found in the intestinal lumen . Some, but not all, TUNEL-positive cells were also positive for caspase-3, indicating that both caspase-3-dependent and -independent pathways of PCD increased upon infection. J Mol Biol, 2005 Jan 28, 345(4), 879 - 92 The structure of the oligopeptide-binding protein, AppA, from Bacillus subtilis in complex with a nonapeptide; Levdikov VM et al.; Besides their role as a source of amino acids for Bacillus subtilis, exogenous peptides play important roles in the signalling pathways leading to the development of competence and sporulation . B.subtilis has three peptide transport systems all belonging to the ATP-binding cassette family, a dipeptide permease (Dpp) and two oligopeptide permeases (Opp and App) with overlapping specificity . These comprise a membrane-spanning channel through which the peptide passes, a pair of ATPases which couple ATP hydrolysis to peptide translocation and a lipid-modified, membrane-anchored extracellular "binding-protein" that serves as the receptor for the system . Here, we present the crystal structure of a soluble form of the peptide-binding protein AppA, which has been solved to 1.6 A spacing by anomalous scattering and molecular replacement methods . The structure reveals a protein made of two distinct lobes with a topology similar to those of DppA from Escherichia coli and OppA from Salmonella typhimurium . Examination of the interlobe region reveals an enlarged pocket, containing electron density defining a nonapeptide ligand . The main-chain of the peptide is well defined and makes a series of polar contacts with the protein including salt-bridges at both its termini . The side-chain density is ambiguous in places, consistent with the interpretation that a population of peptides is bound, whose average electron density resembles the amino acid sequence N-VDSKNTSSW-C. J Endotoxin Res, 2004, 10(6), 439 - 44 Deacylation and palmitoylation of lipid A by Salmonellae outer membrane enzymes modulate host signaling through Toll-like receptor 4; Kawasaki K et al.; The Salmonella typhimurium virulence gene products, PhoP/PhoQ sense host micro-environments to regulate the expression of a lipid A 3-O-deacylase, PagL, and a lipid A palmitoyltransferase, PagP . Therefore, deacylation and/or palmitoylation of lipid A could occur in Salmonellae adapted to host environments . The acylation state of lipid A can alter host recognition and signaling by Toll-like receptor (TLR) 4, and may play an important role in host defenses against Salmonellae infection . Deacylated lipid A, deacylated and palmitoylated lipid A, palmitoylated lipid A, and unmodified lipid A species were purified, and the activity was examined using cell lines expressing recombinant human or mouse TLR4 . Compared with unmodified lipid A, the modified lipid A species are 10-100-fold less active . These results suggest that PagL and PagP modify lipid A to reduce TLR4-signaling as part of Salmonellae adaptation to the host environment. J Huazhong Univ Sci Technolog Med Sci, 2004, 24(4), 389 - 91 Anti-angiogenesis effect on glioma of attenuated Salmonella typhimurium vaccine strain with flk-1 gene; Feng K et al.; To investigate the anti-vasculature effects and the anti-glioma effects of attenuated Salmonella typhimurium vaccine strain expressing VEGFR2 (flk-1) gene, plasmid pcDNA3.1-flk1 was constructed and electro-transfected into live attenuated Salmonella typhimurium strain SL7207 . Mouse models of intracranial G1261 glioblastoma were treated with an orally administered attenuated Salmonella typhimurium expressing flk-1 gene . The survival period was recorded and vessel density was observed by immunofluorescence . CTLs activity was measured by MTT assay . Our results showed that attenuated Salmonella typhimurium vaccine strain expressing flk-1 gene could significantly inhibit glioblastoma growth, reduce vessel density, prolong the survival period and improve the survival rate in these mice . The flk-1 specific CTLs activity was increased obviously after the vaccination . Our study showed that attenuated Salmonella typhimurium vaccine strain expressing flk-1 gene could break peripheral immune tolerance a in glioma gainst this self-antigen and kill endothelial cells by the orally administered vaccine and can be used for both prophylactic and therapeutic purposes. J Biol Chem . 2004 Dec 6; {Epub ahead of print} A cis-spreading nucleoprotein filament is responsible for the gene silencing activity found in the promoter relay mechanism; Chen CC et al.; Transcription-generated DNA supercoiling plays a decisive role in a promoter relay mechanism for the coordinated expression of genes in Salmonella typhimurium ilvIH-leuO-leuABCD gene cluster . A similar mechanism also operates to control expression of the genes in the Escherichia coli ilvIH-leuO-leuABCD gene cluster . However, the mechanism underlying the DNA supercoiling effect remained elusive . A bacterial gene silencer AT8 was found to be important for the repression state of the leuO gene as part of the promoter relay mechanism . In this communication, we demonstrated that the gene silencer AT8 is a nucleation site for recruiting histone-like nucleoid structuring protein (H-NS) to form a cis-spreading nucleoprotein filament that is responsible for silencing of the leuO gene . With a DNA geometric similarity rather than a DNA sequence specificity, the E . coli gene silencer EAT6 was capable of replacing the H-NS nucleation function of the S . typhimurium gene silencer AT8 for the leuO gene silencing . The interchangeability between DNA geometrical elements for supporting the silencing activity in the region is consistent with a previous finding that a neighboring transcription activity determines the outcome of the gene silencing activity . The geometric requirement, which was revealed for this silencing activity explains the decisive role of transcription-generated DNA supercoiling found in the promoter relay mechanism. Toxicol In Vitro, 2005 Feb, 19(1), 91 - 97 The amoebicidal aqueous extract from Castela texana possesses antigenotoxic and antimutagenic properties; Reyes-Lopez M et al.; Due to long-term treatment toxicity and clinical resistance to drugs commonly used against E . histolytica, new drugs against amoebiasis are urgently needed . Castela texana ("chaparro amargo") is a shrub taken traditionally in teas and capsules of dry plant to treat intestinal amoebic infections . An aqueous extract was prepared and its mutagenic, genotoxic and cytotoxicity properties were evaluated in prokaryotic and eukaryotic systems . This extract was neither mutagenic when evaluated with the Ames test in Salmonella typhimurium strains TA98, TA100 and TA102, nor genotoxic in unscheduled DNA synthesis in hepatocyte cultures, even at the highest concentrations tested . In fact, C . texana extract showed antimutagenic activity on S . typhimurium strains TA98 and TA100 in the Ames test . Furthermore, it was capable of protecting liver cell cultures against unscheduled DNA synthesis induced by 2-acetylaminofluorene at a concentration of 6.77 mug/ml . A free-radical scavenging test was used in order to explore the antioxidant capacity of C . texana extract with S . typhimurium strain TA102 pretreated with norfloxacin, a free radical producer . This extract showed a free radical withdrawal effect . The effective chemoprotective activity of this extract could be due to the antioxidant capacity of the C . texana extract components . In this paper it is shown that the antiamoebic natural product, C . texana, is also antimutagenic and protects against induction of preneoplastic lesions in rat liver . These results justify further studies to extend it use to human beings. FEBS Lett, 2004 Dec 3, 578(1-2), 128 - 34 Enhancing the first enzymatic step in the histidine biosynthesis pathway increases the free histidine pool and nickel tolerance in Arabidopsis thaliana; Wycisk K et al.; Naturally selected nickel (Ni) tolerance in Alyssum lesbiacum has been proposed to involve constitutively high levels of endogenous free histidine . Transgenic Arabidopsis thaliana expressing a Salmonella typhimurium ATP phosphoribosyl transferase enzyme (StHisG) resistant to feedback inhibition by histidine contained approximately 2-fold higher histidine concentrations than wild type plants . Under exposure to a toxic Ni concentration, biomass production in StHisG expressing lines was between 14- and 40-fold higher than in wild-type plants . This suggested that enhancing the first step in the histidine biosynthesis pathway is sufficient to increase the endogenous free histidine pool and Ni tolerance in A . thaliana. Med Sci (Paris), 2004 Dec, 20(12), 1119 - 24 {Protagonists of innate immunity during infection with Salmonella}; Salez L et al.; Salmonella are facultative intracellular Gram-negative bacteria that are found ubiquitously in nature and have the ability to infect a wide range of hosts including humans, domesticated, wild mammals, and birds . The principal clinical manifestations associated with Salmonella infection in humans are enteric fever (typhoid and paratyphoid) and a self-limiting gastroenteritis (salmonellosis) . Additionally, silent carriage of this bacterium is frequent and contributes to disease dissemination . Typhoid fever still represents a major public health problem in many developing countries . On the other hand, industrialized countries experience an increased incidence of nontyphoidal Salmonella infections with most cases tracing back to food contamination . Studies using mouse model of infection with a highly virulent Salmonella typhimurium serotype have provided important insight into the complexity of the innate immune response to infection . The players are numerous but emphasis was placed on the genes that were discovered using genetic approaches and in vivo assay with live pathogen and include positional cloning of mouse mutations and manipulation of genes in the context of whole animal either by transgenesis or knockout technologies . Some of the critical genes include those known to play a role in the detection of the bacteria (Cd14, Lbp, Tlr4 and Tlr5) and in microbicidal activity (Slc11a1, Nos2, NADPH oxidase and cryptdins) . These discoveries have already initiated the search for the contribution of particular genetic pathways in the innate immune response of humans to infection with Salmonella and other intracellular microorganisms. Genetics, 2004 Nov, 168(3), 1119 - 30 Experimental adaptation of Salmonella typhimurium to mice; Nilsson AI et al.; Experimental evolution is a powerful approach to study the dynamics and mechanisms of bacterial niche specialization . By serial passage in mice, we evolved 18 independent lineages of Salmonella typhimurium LT2 and examined the rate and extent of adaptation to a mainly reticuloendothelial host environment . Bacterial mutation rates and population sizes were varied by using wild-type and DNA repair-defective mutator (mutS) strains with normal and high mutation rates, respectively, and by varying the number of bacteria intraperitoneally injected into mice . After <200 generations of adaptation all lineages showed an increased fitness as measured by a faster growth rate in mice (selection coefficients 0.11-0.58) . Using a generally applicable mathematical model we calculated the adaptive mutation rate for the wild-type bacterium to be >10(-6)/cell/generation, suggesting that the majority of adaptive mutations are not simple point mutations . For the mutator lineages, adaptation to mice was associated with a loss of fitness in secondary environments as seen by a reduced metabolic capability . During adaptation there was no indication that a high mutation rate was counterselected . These data show that S . typhimurium can rapidly and extensively increase its fitness in mice but this niche specialization is, at least in mutators, associated with a cost. Biol Pharm Bull, 2004 Dec, 27(12), 2010 - 3 Experimental analysis of antimicrobial action of dicyclomine hydrochloride; Karaka P et al.; Dicyclomine hydrochloride is an antispasmodic agent . The MIC of dicyclomine against standard strains of Gram positive and Gram negative bacteria were performed by NCCLS broth dilution technique . These drugs showed a rapid killing action on Gram positive bacteria, Staphylococcus aureus NCTC 6571, 8530 and several other reference strains . The killing effect against Gram negative bacteria, Shigella boydii 8 NCTC 254/66 and Salmonella typhimurium NCTC 74 showed that the drug was bacteriostatic with respect to these strains . High rate of killing was achieved for most strains of Gram positive bacteria within 2 h . When administered to Swiss strain of white mice at doses of 30 and 60 microg/g of mouse, the drug could significantly protect the animals challenged with 50 MLD of Salmonella typhimurium NCTC 74 . According to chi2 test, the in vivo data were highly significant (p<0.001) . Since dicyclomine showed a remarkable inhibitory action against several pathogenic bacteria, in the course of time, it may be developed as a potent antimicrobial agent for many bacterial infections. Mutat Res, 2004 Dec 31, 565(1), 23 - 34 Influence of extraction parameters on the mutagenicity of soil samples; Courty B et al.; The aim of this study was to investigate the influence of four extraction parameters (type of solvent, temperature, duration of extraction, and soil mass/solvent volume ratio) on the mutagenicity of soil extracts . Four urban soil samples were submitted to the micro-method adaptation of the Ames test on Salmonella typhimurium according to the following sequence: identification of the most sensitive strain (TA98 or TA100), the best solvent(s), the optimum extraction temperature and extraction time, and finally the optimal soil/solvent ratio . Extraction was thus performed using eight different solvents (distilled water, dichloromethane, acetonitrile, acetone, cyclohexane, methanol, hexane, or ethanol), two temperatures (room temperature or 37 degrees C), two durations (4 or 24 h), and two soil mass/solvent volume ratios (1:2 or 1:10) . The results show that strain TA98 was more sensitive than strain TA100, and the observed mutagenicity was expressed as number of TA98 revertants per mg of soil equivalent . No mutagenicity was induced by the distilled water extracts, whereas most of the organic solvent extracts induced a significant mutagenic response . A dichloromethane/acetone mixture appeared to be the best compromise for extraction of mutagens from the urban soils tested . Moreover, the present study showed that a higher mutagenic activity was generally obtained with a temperature of 37 degrees C (compared to room temperature), with an extraction time of 24 h (compared to 4 h), and with a soil mass/solvent volume ratio of 1:10 (compared to 1:2). Food Chem Toxicol, 2004 Dec, 42(12), 2029 - 35 Mutagenic activity of sweepings and pigments from a household-wax factory assayed with Salmonella typhimurium; Varella SD et al.; The mutagenic activity of garbage originating from a household wax industry was determined by the Salmonella/microsome assay, using the bacterial strains TA100, TA98 and YG1024 . The garbage was obtained by sweeping the floor of the factory at the end of the work shift . Organic compounds were extracted by ultrasound for 30 min in dichloromethane or 70% ethanol . After evaporation of solvent, these extracts (HFS: household-wax factory sweepings) were dissolved in DMSO, and were tested for the mutagenic activity at varying concentrations (HFS-ET: 0.08-0.68 mg/plate, HFS-DCM: 0.60-7.31 mg/plate) . The colouring agents (pigments) used in the production of the wax were also dissolved in DMSO and tested with the assay . The concentrations tested for each pigment were: Amaranth: 0.46-3.65 mg/plate, Auramine: 0.15-1.2 mg/plate and Rhodamine B: 0.22-1.82 mg/plate . Both ET and DCM organic extracts had mutagenic activity, especially in the YG1024 strain . The pigments behaved in a similar way, demonstrating that YG1024 was the most sensitive strain for the detection of mutagenicity, and that metabolization increased the activity . Human exposure (occupational and non-occupational) to industrial residues generated during the household-wax manufacturing and packaging process should be monitored, since this type of garbage is normally deposited in the environment without any control. J Nat Prod, 2004 Nov, 67(11), 1876 - 8 Biotransformation of nobiletin by Aspergillus niger and the antimutagenic activity of a metabolite, 4'-hydroxy-5,6,7,8,3'-pentamethoxyflavone; Okuno Y et al.; Biotransformation of nobiletin (1) by Aspergillus niger has been investigated, and the product obtained was determined as 4'-hydroxy-5,6,7,8,3'-pentamethoxyflavone (2) . Antimutagenic activity of compound 2 was found, which showed suppressive effects on umu gene expression of the SOS response to DNA damage in Salmonella typhimurium TA1535/pSK1002, induced by the chemical mutagens furylfuramide, MeIQ, and Trp-P-1. Zh Mikrobiol Epidemiol Immunobiol, 2004 Sep-Oct, (5), 16 - 9 {State of Salmonella typhimurium population in water environment under the influence of temperature}; The impact of prebiotics and salmonellosis on apparent nutrient digestibility and Salmonella typhimurium var . Copenhagen excretion in adult pigeons (Columba livia domestica); Laboratory of Animal Nutrition, Ghent University, Heidestraat 19, 9820 Merelbeke, Belgium . Geert.Janssens@Ugent.be The effects of lactose or fructo-oligosaccharide (FOS) supplementation on the excretion of salmonellae, apparent digestibilities and excreta consistency were studied . Thirty-two male pigeons (Columba livia domestica) were randomly divided into 4 equal groups: 3 of 4 groups were orally infected with 10(9) Salmonella Typhimurium var . Copenhagen, after being offered a drinking water supplement of 2% FOS, 2% lactose, or no supplement, respectively, for 2 wk . Pigeons in the fourth group were not challenged with S . Typhimurium and remained unsupplemented . Initially, FOS increased water intake, resulting in more watery excreta . After infection, supplementation showed no major effects on S . Typhimurium excretion, nitrogen retention, or apparent nutrient digestibilities, although lactose--and to a lesser extent FOS--improved apparent fiber digestibility during recovery from the S . Typhimurium infection . The excreta consistency of all pigeons returned to normal when recovering from the Salmonella infection . In this trial, neither FOS nor lactose was successful in tempering the negative aspects of Salmonella infection in pigeons . Nevertheless, it should be stated that future investigations should clarify the importance of duration and level of prebiotic supplementation and infection level. J Food Prot, 2004 Nov, 67(11), 2403 - 9 A mathematical model for the transmission of Salmonella Typhimurium within a grower-finisher pig herd in Great Britain; Ivanek R et al.; In a study of pigs slaughtered at British abattoirs, approximately 23% carried Salmonella in their cecal (large intestine) contents . The most frequent serotype was Salmonella Typhimurium (STM), which was the second most common cause of human salmonellosis in Great Britain . A pig industry-monitoring program was developed to reduce Salmonella infection on British farms . The control of STM infection on the farm requires an understanding of STM transmission dynamics within the herd, and a mathematical model has been developed for an infected grower-finisher farm . The model estimates the probability of a random pig being infected with STM . There are three broad categories of STM infection in pigs: pigs that are infected but unable to transmit the infection (latent); pigs that are infectious, i.e., able to transmit the infection (shedders); and pigs that have stopped shedding but harbor STM in their internal organs (carriers) . The model estimates that 21.0% (5th and 95th percentiles, 0.05 to 77.5%) of slaughter-age pigs on an infected farm are likely to be shedding STM . Although this range is wide, it is biologically plausible . Sensitivity analysis of the total number of infected pigs revealed that the most significant input parameters are the probability of effective contact between a specific infectious and susceptible pig and the duration of shedding . The model predicted that 11.5% of pigs would be shedding STM at slaughter age . This value is close to the estimate obtained from a British abattoir survey that 11 . 1% of pigs carried STM in their ceca, indicating that the model has reasonable validity. Vaccine, 2004 Dec 16, 23(5), 595 - 603 Oral vaccination of pigs with an invasive gyrA-cpxA-rpoB Salmonella Typhimurium mutant; Roesler U et al.; The potency to protect pigs against colonization and against clinical salmonellosis was evaluated after oral immunization with a live gyrA-cpxA-rpoB Salmonella (S.) Typhimurium mutant (S . Tm . Nal2/Rif9/Rtt) . Twenty 4-week-old male hybrid piglets were immunized orally, a control group received a placebo . Three weeks postimmunization, all pigs were challenged orally with a highly virulent S . Typhimurium DT104 strain . Clinical investigation revealed that immunization prevented the vaccinated pigs from clinical symptoms of salmonellosis . While all placebo-treated animals showed a 2-4-day episode of moderate to severe clinical symptoms, 90% of immunized pigs did not show any clinical signs at all . The bacteriological results showed a marked beneficial effect of the oral immunization . Vaccinated pigs showed a significantly decreased rate of colonization of the inner organs (42.5% versus 87.5%) when compared to the placebo-treated animals . Furthermore, in comparison to the non-immunized pigs, the vaccines developed a higher specific immunoglobulin (Ig)A antibody activity, but a significant lower IgM antibody activity in serum . The findings underline the ability of an attenuated oral live S . Typhimurium mutant to prevent clinical symptoms of salmonellosis in pigs and to significantly reduce the colonization of tissues and inner organs, as well as the shedding of S . Typhimurium. Arch Biochem Biophys, 2004 Dec 15, 432(2), 233 - 43 The reaction of indole with the aminoacrylate intermediate of Salmonella typhimurium tryptophan synthase: observation of a primary kinetic isotope effect with 3-{(2)H}indole; Cash MT et al.; The bacterial tryptophan synthase alpha(2)beta(2) complex catalyzes the final reactions in the biosynthesis of L-tryptophan . Indole is produced at the active site of the alpha-subunit and is transferred through a 25-30 A tunnel to the beta-active site, where it reacts with an aminoacrylate intermediate . Lane and Kirschner proposed a two-step nucleophilic addition-tautomerization mechanism for the reaction of indole with the aminoacrylate intermediate, based on the absence of an observed kinetic isotope effect (KIE) when 3-{(2)H}indole reacts with the aminoacrylate intermediate . We have now observed a KIE of 1.4-2.0 in the reaction of 3-{(2)H}indole with the aminoacrylate intermediate in the presence of monovalent cations, but not when an alpha-subunit ligand, disodium alpha-glycerophosphate (Na(2)GP), is present . Rapid-scanning stopped flow kinetic studies were performed of the reaction of indole and 3-{(2)H}indole with tryptophan synthase preincubated with L-serine, following the decay of the aminoacrylate intermediate at 350 nm, the formation of the quinonoid intermediate at 476 nm, and the formation of the L-Trp external aldimine at 423 nm . The addition of Na(2)GP dramatically slows the rate of reaction of indole with the alpha-aminoacrylate intermediate . A primary KIE is not observed in the reaction of 3-{(2)H}indole with the aminoacrylate complex of tryptophan synthase in the presence of Na(2)GP, suggesting binding of indole with tryptophan synthase is rate limiting under these conditions . The reaction of 2-methylindole does not show a KIE, either in the presence of Na(+) or Na(2)GP . These results support the previously proposed mechanism for the beta-reaction of tryptophan synthase, but suggest that the rate limiting step in quinonoid intermediate formation from indole and the aminoacrylate intermediate is deprotonation. J Biol Chem . 2004 Nov 10; {Epub ahead of print} Structures of dCTP deaminase from Escherichia coli with bound substrate and product . Reaction mechanism and determinants of mono- and bifunctionality for a family of enzymes; Johansson E et al.; dCTP deaminase (EC 3.5.4.13) catalyzes the deamination of dCTP forming dUTP that via dUTPase is the main pathway providing substrate for thymidylate synthase in Escherichia coli and Salmonella typhimurium . dCTP deaminase is unique among nucleoside and nucleotide deaminases as it functions without aid from a catalytic metal ion that facilitates preparation of a water molecule for nucleophilic attack on the substrate . Two active site amino acid residues, Arg115 and Glu138, were identified by mutational analysis as important for activity in E . coli dCTP deaminase . None of the mutant enzymes R115A, E138A or E138Q had any detectable activity but circular dichroism spectra for all mutant enzymes were similar to wild type suggesting that the overall structure was not changed . The crystal structures of wild type E . coli dCTP deaminase and the E138A mutant enzyme have been determined in complex with dUTP and Mg2+, and the mutant enzyme also with the substrate dCTP and Mg2+ . The enzyme is a third member of the family of the structurally related trimeric dUTPases and the bifunctional dCTP deaminase-dUTPase from Methanocaldococcus jannaschii . However, the C-terminal fold is completely different from dUTPases resulting in an active site built from residues from two of the trimer subunits, and not from three subunits as in dUTPases . The nucleotides are well defined as well as Mg2+ that is tridentately coordinated to the nucleotide phosphate chains . We suggest a catalytic mechanism for the dCTP deaminase and identify structural differences to dUTPases that prevent hydrolysis of the dCTP triphosphate. Avian Dis, 2004 Sep, 48(3), 595 - 605 Expression of Escherichia coli antigens in Salmonella typhimurium as a vaccine to prevent airsacculitis in chickens; Roland K et al.; Avian pathogenic Escherichia coli strains are associated with a variety of extraintestinal poultry diseases, including airsacculitis, colisepticemia, and cellulitis . A number of E . coli serotypes are associated with these diseases, although the most prevalent serotype is O78 . Fimbrial proteins expressed by these strains appear to be important virulence factors, including type 1 fimbriae, P fimbriae, and curli . We have been working to develop an effective vaccine to protect chickens against these diseases . We have previously shown that an attenuated Salmonella typhimurium strain expressing O78 lipopolysaccharide provides protection against challenge with an O78 avian pathogenic E . coli strain . In this work, we have constructed an attenuated S . typhimurium that expresses both the O78 lipopolysaccharide and E . coli-derived type 1 fimbriae . In these studies, chickens were vaccinated at day of hatch and again at 2 wk of age . Birds were challenged at 4 wk of age . We found that the vaccine candidate provided significant protection against airsacculitis as compared to untreated controls or birds vaccinated with an attenuated S . typhimurium that did not express any E . coli antigens . In a separate experiment, challenged vaccinates showed significant weight gain compared to challenged nonvaccinates . We were not able to demonstrate protection against E . coli O1 or O2 serotype challenge, nor against challenge with wild-type S . typhimurium. Environ Mol Mutagen, 2004, 44(5), 387 - 93 Antimutagenic activity of spearmint; Yu TW et al.; The antimutagenic activity of spearmint (Mentha spicata), a popular food flavoring agent, was studied in the Salmonella assay . Spearmint leaves were brewed in hot water for 5 min at concentrations up to 5% (w/v), and the water extracts were tested against the direct-acting mutagens 4-nitro-1,2-phenylenediamine (NPD) and 2-hydroxyamino-3-methyl-3H-imidazo{4,5-f}quinoline (N-OH-IQ) using Salmonella typhimurium strain TA98 . Nontoxic concentrations of spearmint extract inhibited the mutagenic activity of N-OH-IQ in a concentration-dependent fashion, but had no effect against NPD . These experiments by design focused on the water extract consumed commonly as an herbal tea, but chloroform and methanol extracts of spearmint also possessed antimutagenic activity against N-OH-IQ . Water extract of spearmint inhibited the mutagenic activity of the parent compound, 2-amino-3-methyl-3H-imidazo{4,5-f}quinoline (IQ), in the presence of rat liver S9; however, the concentration for 50% inhibition (IC(50)) against IQ was approximately 10-fold higher than in assays with N-OH-IQ minus S9 . At concentrations similar to those used in the Salmonella assays, spearmint extract inhibited two of the major enzymes that play a role in the metabolic activation of IQ, namely, cytochromes P4501A1 and 1A2, based on ethoxyresorufin O-deethylase and methoxyresorufin O-demethylase assays in vitro . In vivo, rats were given spearmint water extract (2%; w/v) as the sole source of drinking fluid before, during, and after 2-week treatment with IQ; colonic aberrant crypt foci were inhibited significantly at 8 weeks (P < 0.05, compared with rats given IQ alone) . Collectively, these findings suggest that spearmint tea protects against IQ and possibly other heterocyclic amines through inhibition of carcinogen activation and via direct effects on the activated metabolite(s) . Environ . Mol . Mutagen., 2004 . (c) 2004 Wiley-Liss, Inc. Science, 2004 Nov 5, 306(5698), 1040 - 2 Structural insights into the assembly of the type III secretion needle complex; Marlovits TC et al.; Type III secretion systems (TTSSs) mediate translocation of virulence factors into host cells . We report the 17-angstrom resolution structures of a central component of Salmonella typhimurium TTSS, the needle complex, and its assembly precursor, the bacterial envelope-anchored base . Both the base and the fully assembled needle complex adopted multiple oligomeric states in vivo, and needle assembly was accompanied by recruitment of the protein PrgJ as a structural component of the base . Moreover, conformational changes during needle assembly created scaffolds for anchoring both PrgJ and the needle substructure and may provide the basis for substrate-specificity switching during type III secretion. J Vet Med B Infect Dis Vet Public Health, 2004 Oct-Nov, 51(8-9), 389 - 92 Resistance of Salmonella isolates in Germany; Schroeter A et al.; During 2000-2002 the National Veterinary Reference Laboratory for Salmonella (NRL-Salm) in Germany typed 11,911 isolates from animals, food, feed and the environment . All of them were tested for their susceptibility to 17 anti-microbial agents . Sixty-three per cent of all isolates were resistant and 40% were multiresistant (resistant against more than one anti-microbial) . This general resistance level was strongly influenced by those specific serotypes which dominate the Salmonella epidemiology in Germany . Salmonella Typhimurium DT104 isolates from pig and cattle, and their resulting food products, were multiresistant in 98 and 94% of the cases respectively . During the period 2000-2003 an increasing quinolone resistance especially in Salmonella isolates from poultry and poultry meat (to 26%) and in S . Paratyphi B D-tartrate positive isolates (to 64%) could be observed . This increase was accompanied by a shift towards higher minimal inhibitory concentrations for ciprofloxacin. J Biol Chem, 2005 Jan 7, 280(1), 355 - 60 Epub 2004 Nov 01. Viability of Escherichia coli topA Mutants Lacking DNA Topoisomerase I; Stupina VA et al.; The viability of the topA mutants lacking DNA topoisomerase I was thought to depend on the presence of compensatory mutations in Escherichia coli but not Salmonella typhimurium or Shigella flexneri . This apparent discrepancy in topA requirements in different bacteria prompted us to reexamine the topA requirements in E . coli . We find that E . coli strains bearing topA mutations, introduced into the strains by DNA-mediated gene replacement, are viable at 37 or 42 degrees C without any compensatory mutations . These topA(-) cells exhibit cold sensitivity in their growth, however, and this cold sensitivity phenotype appears to be caused by excessive negative supercoiling of intracellular DNA . In agreement with previous results (Zhu, Q., Pongpech, P., and DiGate, R . J . (2001) Proc . Natl . Acad . Sci . U . S . A . 98, 9766-9771), E . coli cells lacking both type IA DNA topoisomerases I and III are found to be nonviable, indicating that the two type IA enzymes share a critical cellular function. Biol Pharm Bull, 2004 Nov, 27(11), 1840 - 3 Limitation of polymyxin B on suppression of endotoxin shock induced by Salmonella infection in mice; Morita H et al.; The protective effects of an antibiotic polymyxin B (PLB), having lipopolysaccharide (LPS)-binding activity, on infection-induced endotoxin shock in mice were investigated . Infection with 10(8) colony forming units of an attenuated Salmonella typhimurium aroA strain caused lethal endotoxin shock to ddY mice . Treatment with PLB 1 h post infection (p.i.) resulted in significant reduction of mortality and bacterial numbers in livers . In addition, treatment with PLB 1 h p.i . resulted in a transient increase at the early stage and gradual decline in plasma LPS levels . Although plasma levels of sCD14 and high mobility group box chromosomal protein-1 (HMGB-1) increased according with progression of infection, increases in plasma levels of sCD14 and HMGB-1 were downregulated by treatment with PLB 1 h p.i . However, the lethal shock was not blocked by treatment with anti-CD14 monoclonal antibody at 3 h and 6 h p.i . Interestingly, administration of PLB 6 h p.i . did not show any protective activities, indicating that a time window for effective PLB action is present. J Toxicol Environ Health A, 2004 Dec, 67(23-24), 2037 - 44 Genotoxicity evaluation of Isaria sinclairii (ISE) extract; Ahn MY et al.; The mutagenic potential Isaria sinclairii, a traditional Chinese medicine composed of the fruiting bodies of I . sinclairii and its parasitic host larva, was evaluated using short-term genotoxicity tests, namely, the Ames test, chromosome aberration (CA), and micronuclei (MN) tests . In a Salmonella typhimurium assay, I . sinclairii extract (ISE) did not produce any mutagenic response in the absence or presence of 59 mix with TA98, TA100, TA1535, and TA1537 . In the chromosome aberration (CA) test, ISE induced no significant effect on Chinese hamster ovary (CHO) cells compared with control . In the MN test, no significant change in the occurrence of micronucleated polychromatic erythrocytes was observed in male ICR mice intraperitoneally administered ISE at doses of 15, 150, or 1500 mg/kg . These results indicate that ISE has no mutagenic potential in these in vitro and in vivo systems. J Environ Pathol Toxicol Oncol, 2004, 23(4), 297 - 302 Mutagenic potential of Mancozeb in Salmonella typhimurium; Shukla Y et al.; Mancozeb, a dithiocarbamate fungicide, was examined for its possible mutagenic activity using Salmonella typhimurium tester strains TA97a, TA98, TA100, and TA102 . We found that Mancozeb exhibited toxic effects at the dose of 40 microg/plate and higher with all tester strains . Mancozeb showed dose-dependent increases in the number of revertants with and without metabolic activation when it was dissolved in DMSO or acetone with strain TA97a; however, the number of revertants at the highest dose was less than two-fold compared to control values . We postulate that the true mutagenic potential of Mancozeb may be masked by its toxic effect to the tester strain used. J Cell Sci, 2004 Nov 15, 117(Pt 24), 5771 - 80 Epub 2004 Oct 26. Salmonella typhimurium transcytoses flagellin via an SPI2-mediated vesicular transport pathway; Lyons S et al.; Apical colonization of polarized epithelia by Salmonella typhimurium results in translocation of flagellin to the basolateral membrane domain, thus enabling activation of toll-like receptor 5 (TLR5)-mediated pro-inflammatory gene expression . Such flagellin transcytosis occurred without a change in epithelial permeability to 40 kDa FITC dextran, did not require bacterial motility and was independent of transepithelial movement of intact bacteria . Flagellin transcytosis was blocked at 20 degrees C, suggesting dependence on vesicular transport consistent with results from confocal microscopy that showed flagellin independent of bacteria inside epithelial cells . Furthermore, vesicles isolated from S . typhimurium-infected epithelia were highly enriched in flagellin . Flagellin transcytosis was dependent upon genes of Salmonella pathogenicity island (SPI)-2, which alter vesicular trafficking, but independent of SPI-1 that mediates bacterial invasion . Furthermore, such SPI-2 mutants were unable to mediate the localization of flagellin into intracellular vesicles consistent with flagellin transcytosis mediated by a S . typhimurium take-over of host vesicle trafficking pathways . As a result of their inability to transcytose flagellin, apical colonization by SPI-2 mutants induced substantially less epithelial IL-8 secretion than wild-type strains suggesting that such SPI-2 mediated transcytosis of flagellin plays a role in the pathogenesis of the mucosal inflammation characteristic of human Salmonellosis. Mutat Res, 2004 Dec 12, 564(2), 149 - 57 Analysis of mutagenic activity of airborne particulate matter, standard reference materials and reference compounds using base pair-specific Salmonella typhimurium tester strains; Erdinger L et al.; The mutagenicity profiles of organic extracts of airborne dust samples from Mannheim, Germany, and two standard reference materials (SRM) as well as eight compounds with different chemical properties were investigated using tester strains Salmonella typhimurium TA700x (Ames II Assay) . Each strain of this series carries a unique missense mutation in the histidine operon and is reverted by only one specific base substitution out of six possible changes . Mutation patterns of eight compounds with different modes of genotoxic action reveal significant differences . Samples of airborne particulate matter (APM) from an industrialized town in Germany (Mannheim) were collected for five consecutive days once a month for 1 year using an automatic high-volume air sampler . Samples taken from Monday to Friday were Soxhlet-extracted and prepared according to standard methods . Although the threshold limit for the least active strains is not triggered by all samples, it can be concluded that mutation patterns of the samples do not vary between different seasons . Standard reference materials (SRMs) were prepared and tested using the same methods . SRMs and APM samples from Mannheim reveal similar mutagenicity profiles in TA700x strains . The comparison of the mutagenicity profiles of air dust extracts from Mannheim and the SRMs, respectively, with reference compounds investigated so far shows some similarities although the patterns do not fit perfectly . Mutagenicity profiles of TA700x-activity of nitro-aromatic compounds published so far are similar to those of APM collected in Mannheim, Germany, as well as to standard reference materials 1648 and 1649. Mutat Res, 2004 Dec 12, 564(2), 103 - 13 Effects of exposure to diesel exhaust particles (DEP) on pulmonary metabolic activation of mutagenic agents; Zhao HW et al.; Exposure of rats to diesel exhaust particles (DEP) or carbon black (CB) has been shown to induce time-dependent changes in CYP1A1and CYP2B1 in the lung . The present study evaluated the role of these metabolic enzymes on the pulmonary bioactivation of mutagens . Male Sprague-Dawley rats were intratracheally instilled with saline (control), DEP or CB (35 mg/kg body weight) and sacrificed at 1, 3, or 7 days post-exposure . Both control and exposed lung S9 increased the mutagenic activity of 2-aminoanthracene (2-AA), 2-aminofluorene (2-AF), 1-nitropyrene (1-NP), and the organic extract of DEP (DEPE) in Ames tests with Salmonella typhimurium YG1024 in a dose-dependent manner . Lung microsomes prepared form control or particle-exposed S9, but not cytosolic protein, activated 2-AA mutagenicity . Compared to saline controls, CB-exposed S9 was a less potent inducer of 2-AA mutagenicity at all time points, whereas DEP-exposed S9 was less potent than control saline at 3 and 7 days but not 1 day post-exposure . At 3 days post-exposure, DEP- or CB-exposed lung S9 did not significantly affect the mutagenicity of DEPE or 1-NP, when compared to the controls . The mutgenicity of 2-AA, 2-AF, 1-NP, and DEPE were significantly decreased in the presence of inhibitors for CYP1A1 (alpha-naphthoflavone) or CYP2B (metyrapone), but markedly enhanced by CYP1A1 or CYP2B1 supersomes with all the cofactors, suggesting that both CYP1A1 and CYP2B1 were responsible for mutagen activation . These results demonstrated that exposure of rats to DEP or CB altered metabolic activity of lung S9 and S9 metabolic activity dependent mutagen activation . The bioactivation of mutagens are metabolic enzyme- and substrate-specific, and both CYP1A1 and CYP2B1 play important roles in pulmonary mutagen activation. J Ethnopharmacol, 2004 Dec, 95(2-3), 437 - 45 Mutagenic and antioxidant activities of Croton lechleri sap in biological systems; Lopes MI et al.; The sap of Croton lechleri Muell.-Arg (Euphorbiaceae), called Dragon's blood, is used in folk medicine as a cicatrizant, anti-inflammatory and to treat cancer . In this research, the antioxidant activity of Croton lechleri sap was evaluated against the yeast Saccharomyces cerevisiae and against maize plantlets treated with the oxidative agents apomorphine and hydrogen peroxide . The mutagenic activity of the sap was also analyzed using the Salmonella/microsome assay (Salmonella typhimurium TA97a, TA98, TA100, TA102, TA1535) and in cells of the yeast Saccharomyces cerevisiae . The results showed that Croton lechleri sap possesses significant antioxidant activity against the oxidative damages induced by apomorphine in Saccharomyces cerevisiae under all the conditions studied . However, in the case of hydrogen peroxide, antioxidant activity of the sap was detected only in cells in the stationary phase of growth . The sap was also able to protect cells of the maize plantlets from the toxic effect of apomorphine . This sap showed mutagenic activity for strain TA1535 of Salmonella typhimurium in the presence of metabolic activation and a weak mutagenic activity for strain TA98 . These strains detect base pair substitutions and frameshift mutations, respectively . Mutagenicity was also observed in a haploid Saccharomyces cerevisiae strain XV185-14c for the lys1-1, his1-7 locus-specific reversion and hom3-10 frameshift mutations. Vet Immunol Immunopathol, 2004 Dec 8, 102(3), 321 - 8 Humoral immune response induced by oral administration of S . typhimurium containing a DNA vaccine against porcine reproductive and respiratory syndrome virus; Jiang P et al.; The ORFs-encoded major envelope glycoprotein (GP5) of porcine reproductive and respiratory syndrome virus (PRRSV) is one of the major structural proteins of this virus . In this report, we described the induction of a PRRSV GP5-specific immune response by oral vaccination of mice with eukaryotic expression vectors containing the GP5 gene of PRRSV (pcDNA3-GP5), delived by attenuated Salmonella typhimurium aroA . It demonstrated that oral administration of the transformants resulted in expression of the GP5 transcript in the intestinal epithelium . The level of serum neutralization antibodies to PRRSV was not significantly different between the mice immunized with the transformants and the naked plasmid DNA . But the neutralizing antibody titres in sera of the mice immunized with SL7207/pcDNA3-GP5' and pcDNA3-GP5' (resides 2-25 deletion mutant of GP5) were significantly lower than those immunized with the complete GP5 gene . These results show that oral inoculation of the transformants can induce humoral immune response to PRRSV . The signal peptide of the GP5 protein of PRRSV is associated with the neutralizing epitope of the protein . The attenuated S . typhimurium may be used as a delivery system for oral DNA vaccines containing PRRSV GP5 glycoprotein. FASEB J, 2005 Jan, 19(1), 158 - 9 Epub 2004 Oct 21. Multidrug resistance-1 (MDR-1): a new target for T cell-based immunotherapy; Niethammer AG et al.; Acquired multidrug resistance (MDR) remains a major challenge in the treatment of cancer with chemotherapeutic drugs . It can be mediated by the up-regulated expression of different proteins within the tumor cell membrane . Here, we used murine multidrug resistance-1 (MDR-1) as a target-antigen for the immunotherapy of cancer . We successfully demonstrated that peripheral T cell tolerance can be broken by oral administration of a DNA vaccine encoding MDR-1 and carried by attenuated Salmonella typhimurium to secondary lymphoid organs . Thus, mice, immunized orally three times at 2-wk intervals and challenged 2 wk thereafter with either MDR-1 expressing CT-26 colon carcinoma cells or MDR-1 expressing Lewis lung carcinoma cells, revealed a significant increase in life span . This was evident, when compared with animals either vaccinated with the empty control vector or challenged with the parental cell lines lacking overexpression of MDR-1 . The immune response induced was antigen-specific and CD8+ T cell-mediated . The presence of the target antigen led to up-regulation of activation markers on CD8+ T cells and resulted in a strong cytotoxic T cell response as well as lysis of tumor target cells in vitro . We furthermore established the vaccine to be an effective treatment for established multi-drug-resistant tumor metastases, resulting in a significantly increased life span of experimental animals . Absence of CD8+ T cells due to in vivo depletion led to abrogation of effectiveness . Taken together, our results demonstrate that T cell tolerance against the MDR-1 self-antigen can be broken . It is anticipated that the combination of such an approach with chemotherapy could lead to more effective treatments of cancer. Mutat Res, 2004 Nov 22, 556(1-2), 65 - 74 Delineation of antimutagenic activity of catechin, epicatechin and green tea extract; Geetha T et al.; Tea is consumed worldwide as second largest to water in popularity as a beverage . It has been reported that tea extracts have antibacterial, antiviral, antioxidative, antitumor and antimutagenic activities . The protective effect of green tea has been assumed to be due to the powerful scavenging and antioxidative property of high concentrations of unpolymerised catechins and their gallates . In the present proposal green tea extract (GT), (+)-catechin (C) and (-)-epicatechin (EC) were investigated for their antioxidant activity by different in vitro methods like (i) DPPH assay (ii) superoxide anion scavenging and (iii) hydrogen peroxide scavenging activity . Further these agents were also tested against mutagenesis using the well-standardized Ames microsomal test system . The Ames tester strain Salmonella typhimurium TA102, which readily responds to reactive oxygen species, was used and the antimutagenic activity was evaluated against oxidative mutagens tertiary butyl hydroperoxide (ID50-24.41, 29.63 and 113.23 microg for EC, C and GT, respectively) and hydrogen peroxide (ID50-17.3, 18.4 and 88.1 microg for EC, C and GT, respectively) . Ascorbic acid was used as a standard antioxidant in all the experiments . Results indicate that all the three agents possess excellent DPPH free radical scavenging activity (IC50-1.5 microg for EC, 3.45 microg for C and 3.8 microg for GT), good hydrogen peroxide (IC50-11.18 microg for EC, 13.5 microg for C and 11.78 microg for GT) and superoxide anion scavenging (IC50-1.64 microg for EC, 1.74 microg for C and 3.52 microg for GT) activities . Further, they also show antimutagenic activity in the above-mentioned test systems establishing their antioxidant nature to be responsible for such activity . The in vitro antioxidant activity correlates well with the antimutagenic action . (-)-Epicatechin is indicated to be a better agent in comparison to the other two agents (ID50-1.2 times more than C and 5 times more than GT in antimutagenicity studies against t-BOOH and hydrogen peroxide induced mutagenesis) . Ascorbic acid however showed a much less activity (ID50-12.1 mg against t-BOOH and 7.2 mg with hydrogen peroxide induced mutagenesis). Biochemistry, 2004 Oct 26, 43(42), 13370 - 9 Crystal structure of Escherichia coli ArnA (PmrI) decarboxylase domain . A key enzyme for lipid A modification with 4-amino-4-deoxy-L-arabinose and polymyxin resistance; Gatzeva-Topalova PZ et al.; Gram-negative bacteria including Escherichia coli, Salmonella typhimurium, and Pseudomonas aeruginosa can modify the structure of lipid A in their outer membrane with 4-amino-4-deoxy-l-arabinose (Ara4N) . Such modification results in resistance to cationic antimicrobial peptides of the innate immune system and antibiotics such as polymyxin . ArnA is a key enzyme in the lipid A modification pathway, and its deletion abolishes both the Ara4N-lipid A modification and polymyxin resistance . ArnA is a bifunctional enzyme . It can catalyze (i) the NAD(+)-dependent decarboxylation of UDP-glucuronic acid to UDP-4-keto-arabinose and (ii) the N-10-formyltetrahydrofolate-dependent formylation of UDP-4-amino-4-deoxy-l-arabinose . We show that the NAD(+)-dependent decarboxylating activity is contained in the 360 amino acid C-terminal domain of ArnA . This domain is separable from the N-terminal fragment, and its activity is identical to that of the full-length enzyme . The crystal structure of the ArnA decarboxylase domain from E . coli is presented here . The structure confirms that the enzyme belongs to the short-chain dehydrogenase/reductase (SDR) family . On the basis of sequence and structure comparisons of the ArnA decarboxylase domain with other members of the short-chain dehydrogenase/reductase (SDR) family, we propose a binding model for NAD(+) and UDP-glucuronic acid and the involvement of residues T(432), Y(463), K(467), R(619), and S(433) in the mechanism of NAD(+)-dependent oxidation of the 4''-OH of the UDP-glucuronic acid and decarboxylation of the UDP-4-keto-glucuronic acid intermediate. Prev Vet Med, 2004 Oct 14, 65(3-4), 147 - 71 Estimating the number of undetected multi-resistant Salmonella Typhimurium DT104 infected pig herds in Denmark; Rugbjerg H et al.; In Denmark, the detection of multi-resistant Salmonella Typhimurium DT104 (MRDT104)-infected pig herds relies on the national Salmonella surveillance programme at the farm and slaughterhouse levels of production . With the surveillance sampling protocol and the diagnostic methods currently used, some herds might remain undetected . The number of undetected Danish pig herds infected with MRDT104 in the period 1 August 2001-31 July 2002 was estimated and compared with the number of culture-confirmed detected herds . A flow chart was constructed to illustrate where infected herds will go undetected in the surveillance system and Monte Carlo simulation was used to model the actual number of pig herds infected with MRDT1104 . We estimated that 52 (90% CI {28, 178}) finisher herds were infected with MRDT104 compared to 23 (44%) detected . Among sow herds with production of weaners or growers, we estimated that 38 (90% CI {23, 74}) were infected with MRDT104 compared to 7 (18%) actually detected . Among breeder and multiplier herds, we estimated that five (90% CI {3, 8}) herds were infected with MRDT104 compared to three (60%) detected . In total, we estimated that 102 pig herds were infected with MRDT104 from 1 August 2001 till 31 July 2002 (90% CI {63, 228}) . In comparison, 33 (32%) infected herds were detected in this period . The predicted proportion of undetected herds varied considerably with herd type . We infer that the proportion of detected MRDT104 infected herds depended on the intensity of the combined serological and bacteriological testing. Environ Sci Technol, 2004 Sep 15, 38(18), 4713 - 22 Chemical and biological characterization of newly discovered iodoacid drinking water disinfection byproducts; Plewa MJ et al.; Iodoacid drinking water disinfection byproducts (DBPs) were recently uncovered in drinking water samples from source water with a high bromide/iodide concentration that was disinfected with chloramines . The purpose of this paper is to report the analytical chemical identification of iodoacetic acid (IA) and other iodoacids in drinking water samples, to address the cytotoxicity and genotoxicity of IA in Salmonella typhimurium and mammalian cells, and to report a structure-function analysis of IA with its chlorinated and brominated monohalogenated analogues . The iodoacid DBPs were identified as iodoacetic acid, bromoiodoacetic acid, (Z)- and (E)-3-bromo-3-iodopropenoic acid, and (E)-2-iodo-3-methylbutenedioic acid . IA represents a new class (iodoacid DBPs) of highly toxic drinking water contaminants . The cytotoxicity of IA in S . typhimurium was 2.9x and 53.5x higher than bromoacetic acid (BA) and chloroacetic acid (CA), respectively . A similar trend was found with cytotoxicity in Chinese hamster ovary (CHO) cells; IA was 3.2x and 287.5x more potent than BA and CA, respectively . This rank order was also expressed in its genotoxicity with IA being 2.6x and 523.3x more mutagenic in S . typhimurium strain TA100 than BA and CA, respectively . IA was 2.0x more genotoxic than BA and 47.2x more genotoxic than CA in CHO cells . The rank order of the toxicity of these monohalogenated acetic acids is correlated with the electrophilic reactivity of the DBPs . IA is the most toxic and genotoxic DBP in mammalian cells reported in the literature . These data suggest that chloraminated drinking waters that have high bromide and iodide source waters may contain these iodoacids and most likely other iodo-DBPs . Ultimately, it will be important to know the levels at which these iodoacids occur in drinking water in order to assess the potential for adverse environmental and human health risks. Lett Appl Microbiol, 2004, 39(5), 401 - 6 Antimicrobial activity of ultrasound-assisted solvent-extracted spices; Thongson C et al.; AIMS: The objective of this research was to determine the antimicrobial activity of conventional and high-intensity ultrasound-assisted (HI-US) solvent-extracted Thai spices, including ginger (Zingiber officinale Rose), fingerroot (Bosenbergia pandurata Holtt) and turmeric (Curouma longa Linn) . METHODS AND RESULTS: Extracts were obtained using hexane, isopropanol and a 7 : 3 isopropanol : hexane mixture as solvents with and without HI-US . The antimicrobial activity of the extracts was assayed against four strains each of Listeria monocytogenes and Salmonella Typhimurium DT 104 using an agar dilution assay . Application of HI-US did not alter antibacterial activity against S . Typhimurium, but antilisterial activity of some HI-US spice extracts decreased . Solvent type affected antimicrobial efficacy of extracts with hexane producing the least antimicrobial activity . Fingerroot extracted with isopropanol-hexane and without HI-US had the best antilisterial effect while HI-US-isopropanol fingerroot extract had the greatest antimicrobial efficacy against S . Typhimurium . CONCLUSIONS: Application of HI-US reduced time of extraction to 5 min, compared with the 24 h required for conventional extraction and maintained antimicrobial activity against Salmonella but slightly reduced activity against Listeria . SIGNIFICANCE AND IMPACT OF THE STUDY: HI-US in combination with proper solvent selection may offer a new tool to optimize extraction of spice essential oil for use as antimicrobial agents, and reduce processing time and costs. Commun Dis Public Health, 2004 Sep, 7(3), 193 - 9 Antimicrobial resistance and phage types of human and non-human Salmonella enterica isolates in Ireland, 1998-2003; O'Hare C et al.; Between 1998 and 2003, 5,161 isolates (3,182 human) of Salmonella enterica were received by the National Salmonella Reference Laboratory of Ireland . Serotyping, antimicrobial susceptibility testing and phage typing were performed by standard methods . The number of isolates of S . enterica serovar Typhimurium decreased from 579 (80%) in 1998 to 208 (19%) in 2003, while S . enterica serovar Enteritidis increased from 59 (8%) in 1998 to 219 (20%) in 2003 . Definitive (DT) phage types 104 and DT104b accounted for a declining proportion of all Salmonella Typhimurium isolates (from n = 523 {90%} in 1998 to 126 {60%} in 2003) . Numbers of Salmonella Enteritidis phage type 4 declined from 50 (85%) in 1998 to 59 (27%) in 2003 . Twenty-eight isolates of typhoidal Salmonella were received with a history of recent travel in 17 cases . Resistance to multiple (four or more) antimicrobial agents was related to serotype and, where applicable, phage type, and was common in Salmonella Typhimurium . Salmonella Typhimurium predominated among isolates from cattle and pigs (n = 213 {58%}), while Salmonella Livingstone (n = 327) and S . Kentucky (n = 227) were predominant in isolates from poultry (total n = 554 {43%}) . This paper discusses trends, and their implications, in Irish salmonella isolates since the establishment of the Reference Laboratory. J Appl Microbiol, 2004, 97(5), 964 - 72 Premature Salmonella Typhimurium growth inhibition in competition with other Gram-negative organisms is redox potential regulated via RpoS induction; Komitopoulou E et al.; AIMS: To identify the role of oxidation-reduction (redox) potential in the premature growth inhibition and RpoS induction in Salmonella serotype Typhimurium in competitive growth experiments . METHODS AND RESULTS: Oxidation-reduction potential was measured throughout the growth of a minority population of Salm . Typhimurium in mixed cultures with other Gram-negative and Gram-positive organisms . A lux-based reporter was also used to evaluate RpoS activity in Salm . Typhimurium in competitor studies . In a mixed culture, the multiplication of a minority population of Salm . Typhimurium was inhibited when competing Gram-negative organisms entered the stationary phase . This was not seen when the competing flora was Gram-positive . The change in redox potential during growth in mixed cultures was closely linked to the inhibition of Salm . Typhimurium growth by Gram-negative competitors . An artificially induced drop in redox potential earlier during growth in mixed cultures with Gram-negative organisms reduced the time to RpoS induction in Salm . Typhimurium and thus inhibited its multiplication prematurely . In contrast, RpoS induction and growth inhibition were prevented under high redox potential conditions . CONCLUSIONS: This work shows that the inhibitory activity of competitive organisms can be mediated through their effect on redox potential-regulated RpoS induction . SIGNIFICANCE AND IMPACT OF THE STUDY: Redox potential is shown to be an important determinant of Salm . Typhimurium growth, an observation with practical implications both for its control and detection. J Appl Microbiol, 2004, 97(5), 916 - 22 Application of antimicrobial ice for reduction of foodborne pathogens (Escherichia coli O157:H7, Salmonella Typhimurium, Listeria monocytogenes) on the surface of fish; Shin JH et al.; AIMS: The efficacy of antimicrobial ice was evaluated for the reduction of foodborne pathogens on the surface of fish . METHODS AND RESULTS: Antimicrobial ice containing chlorine dioxide (ClO2) was utilized to control foodborne pathogens in laboratory media and on fish skin . Escherichia coli O157:H7, Salmonella serotype Typhimurium and Listeria monocytogenes strains were treated with antimicrobial ice for 30 min on plates of selective agar and for 120 min on fish skin at room temperature, and then incubated for enumeration . After treatment with 100 ppm ClO2 for 30 min, 5.4, 4.4 and 3.2 log10 reduction was obtained with E . coli O157:H7, Salm . Typhimurium and L . monocytogenes on laboratory media, respectively . When antimicrobial ice (100 ppm ClO2) was applied to fish skin for 120 min, total reduction of E . coli O157:H7, Salm . Typhimurium