Salmonella typhimurium

Avian Dis, 1995 Jan-Mar, 39(1), 1 - 8
Mitogenic responses of the head-associated lymphoid tissues of the chicken; Maslak DM et al.; A blastogenesis microassay employing 3-(4,5-dimethylthiazol-2-yl)-2,5 diphenyl tetrazolium bromide (MTT) was adapted to measure blastogenic responses of lymphocytes from the chicken's head-associated lymphoid tissues (i.e., the harderian gland and conjunctiva-associated lymphoid tissue) to T- and B-cell mitogens . Lymphocytes isolated from peripheral blood, spleen, and the harderian gland had highly significant (P < 0.01) responses to T- and B-cell mitogens compared with control lymphocytes cultured without mitogens . Cultured lymphocytes obtained from the harderian gland had highly significant mitogenic responses to the T-cell mitogen concanavalin A (25 to 100 micrograms/ml) and to the B-cell mitogen Salmonella typhimurium lipopolysaccharide (1.25 to 5.0 micrograms/ml) compared with the control lymphocytes . Mitogenic responses of cultured lymphocytes obtained from the conjunctiva-associated lymphoid tissue could not be measured within the given parameters of the blastogenesis microassay . This was primarily due to the low yield of lymphocytes, which proved to be a limiting factor . The ability of the MTT blastogenesis microassay to detect blastogenic responses of the harderian gland to mitogens may be indicative of its usefulness for measuring cell-mediated immunity responses to other antigens.

Microbiol Immunol, 1995, 39(2), 95 - 103
Different sensitivity of complement to Salmonella typhimurium accounts for the difference in natural resistance to murine typhoid between A/J and C57BL/6 mice; Nakano A et al.; The difference in natural resistance to Salmonella typhimurium between S . typhimurium-resistant A/J mice and S . typhimurium-susceptible C57BL/6 mice was analyzed . In both strains, the growth of S . typhimurium was controlled in the spleen until 48 hr of infection, while serum C3b levels were increased in A/J mice immediately after infection but not in C57BL/6 mice . Incubation of A/J mouse serum with S . typhimurium or its lipopolysaccharide (LPS) generated sufficient amounts of C3b, but that of C57BL/6 mouse serum with them did not . A/J macrophages had higher intracellular killing activity in vitro than did C57BL/6 cells against S . typhimurium pre-opsonized with each corresponding fresh serum . However, the cells from both mice exhibited a similar level of killing activity against S . typhimurium pre-opsonized with fresh A/J serum or rabbit complement . The resistance of C57BL/6 mice was significantly increased by opsonizing S . typhimurium with fresh A/J serum or rabbit complement before inoculation . The serum level of interferon-gamma (IFN-gamma) in A/J mice was 2.7 times as high as in C57BL/6 mice at 48 hr post-infection . Recombinant murine IFN-gamma enhanced the intracellular killing activity of macrophages from both mice when S . typhimurium was pre-opsonized with fresh A/J serum but not with fresh C57BL/6 serum . These findings suggest that A/J macrophages exhibit maximal killing activity against A/J serum-opsonized S . typhimurium in vivo when the cells are activated with IFN-gamma . Therefore, the rapid and sufficient activation of complement by Salmonella LPS may render A/J mice more resistant against murine typhoid.

Biosci Biotechnol Biochem, 1995 Jan, 59(1), 31 - 4
Purification and characterization of cycloinulooligosaccharide fructanotransferase (CFTase) from Bacillus circulans MCI-2554; Kushibe S et al.; Cycloinulooligosaccharide fructanotransferase (CFTase) that produces cyclofructan from inulin was purified about 69-fold from a culture broth of Bacillus circulans MCI-2554 by column chromatographies on DEAE-Toyopearl, QAE-Toyopearl, hydroxyapatite, and phenyl-Sepharose . The molecular mass of the enzyme was estimated to be 115 kDa by SDS-polyacrylamide gel electrophoresis and gel filtration, indicating a monomer structure . Maximal activity was observed at pH 7.5 and 45 degrees C . The enzyme was active from pH 5.5 to pH 9.5, and at temperatures up to 45 degrees C . The enzyme activity was inhibited by Fe2+ and Cu2+ . A part of the amino acid sequence was identical with that of beta-fructofuranosidases of Zymomonas mobilis, carrot, Salmonella typhimurium, and mung bean.

Plasmid, 1995 Jan, 33(1), 27 - 39
The host range of RK2 minimal replicon copy-up mutants is limited by species-specific differences in the maximum tolerable copy number; Haugan K et al.; The minimal replicon of the broad-host-range plasmid RK2 consists of a gene, trfA (trans-acting replication), encoding a protein required for initiation of plasmid replication . The TrfA protein binds to iterons in the cis-acting origin of vegetative replication (oriV), but the exact mechanism by which TrfA-mediated replication initiation takes place is not known . We report here the isolation and characterization of five mini RK2 trfA mutant plasmids with an elevated plasmid copy number, four in Pseudomonas aeruginosa and one in Azotobacter vinelandii . The mutations are localized between or downstream of previously reported Escherichia coli copy-up mutations in trfA, and one of the mutations has been described earlier as an independent copy-up isolate in E . coli . The five mutant plasmids were all moderately copy up in both E . coli and their host of origin, in spite of the use of isolation procedures which were expected to select efficiently in favor of plasmid mutants specifying high copy numbers . In contrast, previously described high copy-up mutants isolated in E . coli could not be established in P . aeruginosa and A . vinelandii . These high copy-up mutants were shown to induce cell killing in E . coli under conditions where the plasmid copy number was increased as a physiological response to reduced growth rate . We propose that the reason for this killing effect is that the copy number under these conditions exceeds an upper tolerance level specific for E . coli . By assuming that the corresponding tolerance level is lower in P . aeruginosa and A . vinelandii than in E . coli, and that the mechanism of copy number regulation is similar, the model can explain the phenotypes of all tested copy up mutants in these two hosts . Analogous studies were also performed in Salmonella typhimurium and Acetobacter xylinum . The data obtained in these studies indicate that the above model is probably generally true for gram-negative bacteria, and the results also indicate that the maximum tolerable copy number is surprisingly low in some hosts.

Mol Microbiol, 1995 Jan, 15(1), 25 - 38
Functional analysis of the Salmonella typhimurium invasion genes invl and invJ and identification of a target of the protein secretion apparatus encoded in the inv locus; Collazo CM et al.; We have carried out a functional analysis of invl and invJ, two Salmonella typhimurium genes required for this organism to gain access to cultured mammalian cells . These genes are located immediately down-stream of invC, a previously identified gene also required for bacterial invasion . Non-polar mutations in either of these genes rendered S . typhimurium severely defective for entry into cultured epithelial cells, although these mutations did not affect the ability of these organisms to attach to those cells . Nucleotide sequence analysis revealed that the invl and invJ genes encode proteins with molecular weights of 18,077 and 36,415, respectively . Polypeptides of similar sizes were observed when these genes were expressed in a bacteriophage T7 RNA polymerase-based expression system . Comparison of the predicted sequences of invl and invJ with translated sequences in the existing databases indicated that these proteins are identical to the previously identified S . typhimurium SpaM and SpaN proteins . Further analysis of these sequences revealed regions of homology between Invl and the N-terminus of IpaB of Shigella spp . and between InvJ and EaeB of enteropathogenic Escherichia coli . Localization studies by immunoblot analysis indicated that InvJ is secreted to the culture supernatant, a surprising finding since this protein also lacks a typical signal sequence . Mutations in invG and invC, two members of the Salmonella inv locus, effectively prevented the transport of InvJ to the culture supernatant . Thus, InvJ is the first identified target of the protein secretion apparatus encoded in the inv locus and therefore a candidate to have effector functions related to bacterial entry.

J Am Vet Med Assoc, 1995 Jan 1, 206(1), 75 - 6
Septicemic salmonellosis in two llamas; Anderson NV et al.; At necropsy, Salmonella choleraesuis var kunzendorf was recovered from the lungs of a 6-year-old female llama that died within a few days of onset of illness . The llama had had fence-line contact with 40 sows at a farm . Salmonella typhimurium was isolated from a blood sample of a 6-day-old male cria that also died after a short illness.

J Reprod Immunol, 1995 Jan, 28(1), 15 - 30
In vivo inhibition of cell-mediated and humoral immune responses to cellular antigens by SV-IV, a major protein secreted from the rat seminal vesicle epithelium; Romano-Carratelli C et al.; Microgram amounts of protein SV-IV, a major secretory protein produced by adult rat seminal vesicle epithelium, markedly decrease the mouse humoral immune response to cellular xenogeneic or allogeneic antigens (sheep red blood cells (SRBC) or mouse epididymal spermatozoa) . The significant reduction in the total number of splenocytes and their main cell subsets in SRBC-immunized mice, the dramatic decrease in the number of Ia+ splenic T cells and the marked inhibition of splenocyte ability to respond in vitro to polyclonal mitogen stimuli suggest that the macrophage accessory cells are the primary target of the SV-IV immunosuppressive activity in vivo . Moreover, the infection of SV-IV-treated mice with Salmonella typhimurium produced an increased mortality of the experimental animals associated with a marked decrease of the phagocytic and intracellular killing activities of their peritoneal macrophages.

Biol Pharm Bull, 1995 Jan, 18(1), 49 - 52
Activation of N-nitrosodialkylamines to mutagens by a metalloporphyrin/oxidant model system for cytochrome P450; Okochi E et al.; N-Nitrosodialkylamines are environmental alkylating carcinogens which are metabolically activated to alpha-hydroxy nitrosamines by cytochrome P450 . The precise mechanism of their activation is not well understood, and a simplified chemical model for cytochrome P450 as a non-enzymatic system is useful for investigating the mechanism . In the present study, a chemical model was used in a bacterial mutation assay as a substitute of S9 mix, and its ability in the activation of N-nitrosodialkylamines was elucidated . In the presence of tetrakis(pentafluorophenyl)porphyrinatoiron(III) chloride and tert-butyl hydroperoxide, the mutagenicity of N-nitrosodialkylamines {alkyl = methyl (NDM), ethyl (NDE), propyl (NDP) and butyl (NDB)} in Salmonella typhimurium YG7108 was detected . The mutagenicity increased by the pre-incubation and was dependent on the concentration of mutagens . The mutagenic activity of N-nitrosodialkylamines in Salmonella typhimurium YG7108 was in the following order: NDB > NDM > NDE approximately NDP . The formation of aldehydes derived from dealkylation in the present model was exemplified by the formation of acetaldehyde from NDE . These results showed that N-nitrosodialkylamines were activated by the model system, and consequent mutagenicity was observed . The oxidation by the model can mimic the metabolic activation of chemical carcinogens by cytochrome P450, and the biomimetic catalyst is useful in investigating the mechanisms of the metabolic pathway of N-nitrosodialkylamines.

Vet Med (Praha), 1995, 40(1), 29 - 32
{Characteristics of the plasmid profile of inositol- and rhamnose- negative strains of Salmonella typhimurium}; Cizek A et al.; S . typhimurium isolates obtained during a large outbreak of human salmonellosis associated with smoked mackerels in the Czech Republic as well as strains of S . typhimurium isolated from black headed gull (Larus ridibundus) were examined following biotyping, phage typing, plasmid profiling and restriction endonuclease analysis (Eco RI, Hind III and Bam HI) of plasmid DNA . The epidemic strain of S . typhimurium and two isolates from environment of nesting colony black-headed gull were meso-inositol and L-rhamnose negative, phage type 141 . The isolates from human and environment of nesting colony were found to share the same plasmid profile and REA.

Int J Food Microbiol, 1995 Jan, 24(3), 385 - 96
Growth and penetration of Salmonella enteritidis, Salmonella heidelberg and Salmonella typhimurium in eggs; Schoeni JL et al.; Eggs and egg dishes are important vehicles for Salmonella infections . Salmonella enteritidis, Salmonella typhimurium and Salmonella heidelberg, which can be isolated from chicken ovaries and feces, have been implicated in approximately 50% of the foodborne salmonellosis outbreaks in the United States . In this study, the growth of these three organisms, inoculated into yolks and albumen, was compared at 4, 10 and 25 degrees C . Regardless of whether 10(2) cfu/g or 10(4) cfu/g was inoculated into the yolk or albumen, populations of all strains increased 3 logs or more in number in one day when incubated at 25 degrees C . Maximum numbers of Salmonella ranged from 10(8) to 10(10) cfu/g . All strains grew at 10 degrees C, but peak numbers were lower and occurred later than those at 25 degrees C . Populations of the three Salmonella strains inoculated into eggs stored at 4 degrees C grew sporadically; in some test groups populations declined . The potential for Salmonella in contaminated feces to establish in the interior of eggs was examined by monitoring shell penetration . At 25 degrees C, all three Salmonella strains penetrated the shell in 3 days, but at 4 degrees C, only S . typhimurium was found in one membrane sample . When hatchery conditions were simulated by incubating eggs at 35 degrees C for 30 min followed by storage at 4 degrees C, penetration was enhanced . Penetration was observed by day 1-3 when eggs were exposed to 10(4) cfu Salmonella/g feces . Increasing the inoculum to 10(6) cfu/g feces resulted in 50-75% of the contents of eggs to be contaminated by day 1 . All Salmonella-positive samples were detected by enrichment . Results of this study indicate that S . enteritidis, S . typhimurium, or S . heidelberg present in feces can penetrate to the interior of eggs and grow during storage.

J Clin Microbiol, 1995 Jan, 33(1), 173 - 9
Value of plasmid profiling, ribotyping, and detection of IS200 for tracing avian isolates of Salmonella typhimurium and S . enteritidis; Millemann Y et al.; Seventy selected strains of Salmonella typhimurium and S . enteritidis isolated from related poultry flocks in three independent geographical areas were characterized by phenotypic and genotypic methods to compare the usefulness of the methods in epidemiological studies . The 56 S . typhimurium isolates were poorly discriminated by their biotypes, resistance patterns, and plasmid profiles . Nine different ribotypes were obtained after DNA digestion by BglII, PvuII, and SmaI . Seven IS200 types, characterized by six to nine copies of IS200 on the chromosome, were detected after digestion of genomic DNA by PstI . These studies resulted in the definition of 15 clonal lineages distributed in three clusters . The 14 S . enteritidis strains were not discriminated either by ribotyping or by detection of IS200 (IS200 typing), but were separated on the basis of antibiotic resistance and plasmid profiling . The stability of the insertion sequence type was confirmed by inoculation of an S . typhimurium strain to axenic chickens reared for 15 weeks in sterile isolators.

Environ Mol Mutagen, 1995, 26(1), 86 - 93
Role of classical nitroreductase and O-acetyltransferase on the mutagenicity of nifurtimox and eight derivatives in Salmonella typhimurium; Jurado J et al.; This study investigates the mutagenicity of nifurtimox (NFX) and eight analogues in Salmonella typhimurium indicator strains that possess different levels of classical nitroreductase or O-acetyltransferase activities . The NFX analogues tested replace the 3-methyl-4-yl-tetrahydro-1,4-thiazine-1,1-dioxide group of the parent compound with the following other groups: indazol-1-yl (1G); pyrazol-1-yl (1B); benzimidazol-1-yl (1E); 1,2,4-triazol-4-yl (1D); 1-methyl-3-methylthio-1,2,4-triazol-4-yl-5-thione (1I); 3,5-bis(methylthio)-1,2,4-triazol-4-yl (1H); 1-adamantyl (ADA); and 4,6-diphenylpyridin-1-yl-2-one (1K) . In the genetic backgrounds of the standard Ames tester strains TA98 and TA100, these bacteria combine the L-arabinose resistance forward mutation assay (Ara test) with a deficiency or overproduction of either nitroreduction or O-acetylation . The Ara test revealed, in agreement with previous findings, important differences between TA98 and TA100 and demonstrated, moreover, that these genetic differences are of significance in mutagenicity testing with nitrofuran compounds . The Ara test also indicated dissimilarities between the metabolic activation of NFX and its analogues, these compounds being classified in three different groups according to their mutagenicity toward strain BA14 (genetic background of TA98) and its derivatives . The first group included analogues (1G, 1E, 1I, and ADA) that showed similar mutagenic potency in all bacterial strains . These compounds are considered not to be substrates for both classical nitroreductase and O-acetyltransferase . The second group included compounds (analogues 1B and 1K, and the reference drug NFX) with increased mutagenicity toward the strain overproducing the classical nitroreductase, and/or reduced mutagenicity toward the corresponding deficient bacteria . These compounds are considered to be activated by the classical nitroreductase . The third group (analogues 1D and 1H) was activated by bacterial O-acetyltransferase, and consequently showed increased and decreased mutagenicity with the particular overproducer or deficient bacterial strain, as compared to their isogenic parentals . Previous reports have pointed out interest in NFX analogue 1H as a promising candidate for the replacement of NFX . The present study further enhances the putative interest of compound 1H, based on the different metabolic activation pathway exhibited by this analogue as compared to the parental drug, NFX.

Int J Occup Med Environ Health, 1995, 8(1), 41 - 7
Non-mutagenicity of thiadiazole-1,3,4 derivatives in Salmonella typhimurium strains; Kalski A et al.; The Ames mutagenicity assay with Salmonella typhimurium strains (TA97a, TA98, TA100 and TA102) was used to investigate the ability of five dyes (thiadiazole-1,3,4 derivatives) to induce point mutations . The results did not reveal any mutagenic effects of the tested dyes on the S . typhimurium strains . No mutagenic activity was observed in the experimental series with and without exogenous activation.

Acta Vet Hung, 1995, 43(1), 95 - 103
The production of K88 antigen by Escherichia coli and Salmonella typhimurium strains with recombinant DNA; Holoda E et al.; Expression of K88ab antigen by strains with recombinant DNA, differing in molecular weight or promoter, was subjected to investigation . Strains with recombinant DNA produced greater amounts of antigen than did field isolates . Maximal production was recorded for the E . coli C600 strain with 10.85 kb recombinant DNA with promoter P1 of the pBR322 vector . The substitution of promoter Ptac for promoter P1 did not result in an increased expression of K88ab antigen . The production of K88ab antigen by Salmonella typhimurium TM333 with recombinant DNA was on a level comparable to that of E . coli C600 or E . coli HB101 strains with the same recombinant DNA.

Nippon Saikingaku Zasshi, 1995, 50(2), 537 - 45
{Incidence and serotypes of Salmonella in apparently healthy swine at slaughterhouses in Japan: 1975-1989}; Yoshida T et al.; To find healthy Salmonella carriers among swine, isolation of Salmonella from fresh cecum samples was attempted at slaughterhouses in Tokyo, Saitama, and Tochigi during a period of 11 years . Of a total 3,058 samples, 1,341 were collected between 1975 and 1979, and the other 1,717 between 1984 and 1989 . The overall isolation rate of Salmonella was 13.3% (408 pigs) . The rate was 23.1% between 1975 and 1979 and 5.7% between 1984 and 1989 . A total of 1,037 isolates were identified as Salmonella and classified into 28 serotypes . These serotypes involved Salmonella typhimurium (26.1%), S . derby (25.4%), and S . london (9.5%) . However, the detection rate of these serotypes varied from year to year . Two serotypes were detected from each of 37 pigs and three from a pig . Of these 38 pigs, 27 carried the serotypes of S . typhimurium or S . derby or both . The present study revealed that Salmonella carriers were highly frequent among healthy swine in Japan in 1970s, but decreased in 1980s.

DNA Seq, 1995, 5(3), 145 - 52
Nucleotide sequence of the region between crr and cysM in Salmonella typhimurium: five novel ORFs including one encoding a putative transcriptional regulator of the phosphotransferase system; Titgemeyer F et al.; A 4471 bp region between crr and cysM on the Salmonella typhimurium chromosome (49.5 min) has been sequenced . Five ORFs were found within this region, one of which is likely to be the putative regulatory gene, ptsJ, that corresponds in map position to a gene which when mutated allows expression of a cryptic Enzyme I of the phosphotransferase system . The deduced amino acid sequence of the encoded protein is similar to those of several open reading frames (ORFs) including ORFT2 of Rhodobacter spheroides with which it is 28% identical throughout most of its length (comparison score of 21 S.D.) . PtsJ exhibits a putative, N-terminal, helix-turn-helix, DNA binding domain that is similar in sequence to those in members of the GntR family of transcriptional regulators . Analyses of the sequences of the ORFs encoded within this region are presented.

Environ Mol Mutagen, 1995, 25(4), 284 - 7
Mutagenicity in vitro of 3,4-dichloro-5-hydroxy-2(5H)-furanone (mucochloric acid), a chlorine disinfection by-product in drinking water; Jansson K et al.; The mutagenicity of chlorinated humic drinking waters is accounted for mainly by a single contaminant, 3-chloro-4-(dichloromethyl)-5-hydroxy-2(5H)-furanone (MX), as assessed in Salmonella typhimurium strain TA100 . In the present study 3,4-dichloro-5-hydroxy-2(5H)-furanone (mucochloric acid, MA), another drinking water contaminant much less potent as a mutagen in TA100 than MX, was tested in Chinese hamster ovary (CHO) cells for the induction of mutation at the hypoxanthine phosphoribosyl transferase (hprt) locus to 6-thioguanine resistance (TGr) . Unexpectedly, MA induced TGr mutants in CHO cells with a potency comparable to that reported previously for MX . In subsequent experiments with S . typhimurium, the presence of pKM101 plasmid in strain TA100 increased susceptibility to the mutagenicity of MA, but much less than to that of MX, relative to the parental strain TA1535 lacking pKM101 . The difference between the two compounds in TA100 thus appears to be due to a higher enhancement of the mutagenicity of MX than that of MA by pKM101 mediated error-prone DNA repair.

Pharmacogenetics, 1995, 5 Spec No, S103 - 7
Conjugation of carcinogens by theta class glutathione s-transferases: mechanisms and relevance to variations in human risk; Guengerich FP et al.; Conjugation of chemicals with glutathione (GSH) can lead to decreased or increased toxicity . A genetic deficiency in the GSH S-transferase mu class gene M1 has been hypothesized to lead to greater risk of lung cancer in smokers . Recently a gene deletion polymorphism involving the human theta enzyme T1 has been described: the enzyme is present in erythrocytes and can be readily assayed . A rat theta class enzyme, 5-5, has structural and catalytic similarity and the protein was expressed in the Salmonella typhimurium tester strain TA1535 . Expression of the cDNA vector increased the mutagenicity of ethylene dibromide and several methylene dihalides . Mutations resulting from the known GSH S-transferase substrate 1,2-epoxy-3-(4'nitrophenoxy)propane were decreased in the presence of the transferase . Expression of transferase 5-5 increased mutations when 1,2,3,4-diepoxybutane (butadiene diepoxide), 4-bromo-1,2-epoxybutane, or 1,3-dichloracetone were added . The latter compound is a model for the putative 1,2-dibromo-3-chloropropane oxidation product 1-bromo-3-chloroacetone . These genotoxicity and genotyping assays may be of use in further studies of the roles of GSH S-transferase theta enzymes in bioactivation and detoxication and any changes in risk due to polymorphism.

Environ Mol Mutagen, 1995, 26(2), 171 - 7
Genotoxicity of m-phenylenediamine and 2-aminofluorene in Salmonella typhimurium and human lymphocytes with and without plant activation; Plewa MJ et al.; The promutagenic arylamines, m-phenylenediamine (mPDA) and 2-aminofluorene (2-AF), were evaluated for their genotoxicity in Salmonella typhimurium strain YG1024 and in human lymphocytes . These agents were assayed with and without TX1MX plant activation mix . Both arylamines without activation were refractory in S . typhimurium, demonstrating that plant activation was required for the generation of their ultimate mutagenic metabolites . However, using the alkaline single-cell gel/Comet assay, both mPDA and 2-AF directly induced DNA damage in human lymphocytes . This effect was reduced when the human cells were treated with the arylamine plus TX1MX . mPDA with or without plant activation was not toxic to the exposed cells . However, at concentrations over 80 microM, 2-AF was toxic to lymphocytes . This toxic response was eliminated by incubation with TX1MX . mPDA and 2-AF were plant-activated into mutagens for S . typhimurium . However, these plant-activated products had a reduced genotoxic potency in human lymphocytes.

Environ Mol Mutagen, 1995, 26(2), 155 - 62
Relationships among in vitro mutagenicity assays: quantitative vs . qualitative test results; Benigni R et al.; In previous investigations, we studied the relationships between the profiles of the qualitative responses of in vitro short-term tests (mutation in Salmonella typhimurium, chromosomal aberrations in CHO cells, sister chromatid exchanges in CHO cells, and mutation in mouse lymphoma cells) and common sets of chemicals . In this paper, we address the study of the quantitative responses (potency) . We show that two analyses point to similar patterns of relationships: the mutation in mouse lymphoma cells assay is most similar to the CHO sister chromatid exchange assay, and the Salmonella assay is most similar to the CHO chromosomal aberrations assay.

Environ Mol Mutagen, 1995, 25(2), 134 - 47
1-Nitropyrene as a marker for the mutagenicity of diesel exhaust-derived particulate matter in workplace atmospheres; Scheepers PT et al.; The use of 1-nitropyrene (1-NP) as a marker for the occupational exposure to diesel exhaust (DE) mutagens was investigated in workplace atmospheres contaminated with DE from a variety of emission sources, such as power supplies, forklifts, trucks, caterpillar vehicles, trains, ships' engines, and vehicles in city traffic . Total suspended particulate matter was collected by area sampling . The 1-NP content of acetone extracts of these samples as determined by gas chromatography-high resolution mass spectrometry varied from 0.080 to 17 micrograms/g acetone extractable matter, corresponding to air concentrations of 0.012 to 1.2 ng/m3 . A sample collected in a rural area contained 0.0017 ng/m3 1-NP . The mutagenicity of the extracts was tested in the Salmonella typhimurium strains TA98 and TA1538, using the microsuspension assay with and without metabolic activation by an exogeneous metabolizing system (rat liver S9-fraction) . In addition, the S . typhimurium strains YG1021 and YG1024 were used because of their high sensitivity towards the mutagenicity of nitro polycyclic aromatic hydrocarbons . When plotting the mutagenic potency of the air sample extracts as determined in the absence of liver S9 versus the particle-associated 1-NP level, a relatively high correlation (r = 0.80-0.91) was observed in all of the S . typhimurium strains . High correlations (r = 0.80-0.93) were also observed when plotting the results of mutagenicity testing after activation by S9 versus the outcome of chemical analysis . These results show that the 1-NP content of workplace air samples is associated with their mutagenic potency, suggesting that 1-NP may be used as a marker for occupational exposure to DE-derived particle-associated mutagens.

Mutat Res, 1995 Jan, 346(1), 57 - 60
Detection of mutagenicity of diphenyl ether herbicides in Salmonella typhimurium YG1026 and YG1021; Oguri A et al.; Three kinds of diphenyl ether herbicides, 4-nitrophenyl 2,4,6-trichlorophenyl ether (CNP, chlornitrofen), 2,4-dichlorophenyl 3-methoxy-4-nitrophenyl ether (chlomethoxynil) and 2,4-dichlorophenyl 3-methoxycarbonyl-4-nitrophenyl ether (bifenox), were tested for mutagenicity in Salmonella typhimurium YG1026 and YG1021, which have high nitroreductase activity, and also in S . typhimurium TA100 and TA98 . CNP and chlomethoxynil showed mutagenicity in S . typhimurium YG1026, without S9 mix, inducing 50 and 304 revertants per microgram . These mutagenicities were suppressed by the addition of S9 mix . CNP and chlomethoxynil were also mutagenic to YG1021 with and without S9 mix, and their mutagenicities were lower than those to YG1026 . On the other hand, bifenox was mutagenic to YG1026 only with S9 mix, inducing 3.0 revertants per micrograms . These three herbicides showed no mutagenicity in S . typhimurium TA100 and TA98 either with or without S9 mix.

Mutat Res, 1995 Jan, 341(3), 225 - 34
Geographical variations in mutagenicity of blue rayon extracts of Japanese human bile; Mano H et al.; The mutagenicity of human bile was investigated in the Ames test after blue rayon treatment . In the present study, 52 and 59 bile samples were collected from the high and the low risk population for gallbladder cancer (GBC), respectively . The bile mutagenicity was detected only when the blue rayon extracts of bile were assayed with Salmonella typhimurium TA98 in the presence of S9 mix . Thirty-two (61.5%) of 52 samples obtained from the high risk population were mutagenic . In our previous study (Mano et al., 1993), the mutagenicity was observed in 14 (58.3%) of 24 samples . After combining this data with the results of the present study, 46 (60.5%) of 76 samples revealed the mutagenicity . On the other hand, the mutagenicity was detected in only 7 (11.9%) of 59 samples collected from the low risk population . Therefore, we found a significant geographical difference in the bile mutagenicity.

Adv Exp Med Biol, 1995, 371B, 1653 - 7
Assessment of attenuated Salmonella typhimurium strains designed as potential carriers of foreign antigens for mucosal localization and immuno-genicity in a rabbit model; Boedeker EC et al.; Neither wild type nor attenuated S . typhimurium strains induced diarrheal illness in rabbits . All strains localized to the Peyer's patch at higher concentrations than in lumenal contents or adjacent ileum . Wild type S . typhimurium C5 induced a typhoid-like illness in rabbits with severe weight loss, bacteremia, persistent splenic colonization, and serum IgG response . Both attenuated strains were disseminated to spleen (day 3) but produced minimal systemic illness . They induced biliary IgA responses greater than the wild type (day 7), but minimal serum IgG responses . Both mutants of S . typhimurium are suitable for further development as live enteric vaccines to carry foreign antigens since they localize to Peyer's patch after oral inoculation, induce biliary antibody, and produce minimal systemic disease . The attenuated strains tested are systemically disseminated . It remains to be determined whether dissemination (determined by a large virulence plasmid) is necessary for the desired mucosal immune response or acceptable for an oral vaccine strain.

Chirality, 1995, 7(5), 359 - 64
Comparative toxicity of (+)-(R)- and (-)-(S)-1,2-dibromo-3-chloropropane; Kouzi SA et al.; The haloalkane 1,2-dibromo-3-chloropropane (DBCP), an environmental pollutant that was widely used as a soil fumigant, is a carcinogen and a mutagen and displays target-organ toxicity to the testes and the kidneys . Because little is known about effects of stereochemistry on the metabolism and toxicity of halogenated alkyl compounds and because DBCP, which has a chiral center at C-2, may show enantioselectivity in its metabolism and/or toxicities, the optically pure enantiomers of DBCP were tested in vivo in rats for organ toxicity as well as for bacterial mutagenicity . Organ toxicity studies showed that (S)-DBCP was slightly more renal toxic than (R)-DBCP but was not significantly more toxic than the racemate, and that no significant differences were observed in the extents of testicular necrosis and atrophy caused by either enantiomer or the racemate . In contrast, (R)-DBCP was more mutagenic than either (S)-DBCP or the racemate to Salmonella typhimurium (S . typhimurium) strains TA 100 and TA104 . However, there was little or no enantioselectivity in glutathione S-transferase (GST)-catalyzed conjugation reactions of glutathione with DBCP based on the lack of selectivity in the rates of disappearance of the enantiomers of DBCP in the presence of glutathione (GSH) and GSTs as monitored by chiral gas chromatography (GC).

Microbios, 1995, 81(327), 93 - 101
Local skin reaction in mice and guinea pigs induced by a single intradermal inoculation of Fusobacterium necrophorum lipopolysaccharide; Kanoe M et al.; Dermal responses induced by Fusobacterium necrophorum strain VPI 2891 lipopolysaccharide (LPS) were studied using mice and guinea pigs . In ddY mice, the LPS elicited inflammatory, haemorrhagic lesions and an increase in local vascular permeability 24 h postinjection . Of the mouse strains, C3H/HeJ mice were less sensitive . The LPS induced erythema and haemorrhagic responses in guinea pig skin 24 h postinoculation . These responses were dose-dependent . The intensity of dermal inflammation-inducing activity of F . necrophorum LPS was similar to that of Escherichia coli strain 055:B5 LPS, but weaker than that of Salmonella typhimurium LPS . These findings suggest that the fusobacterial LPS may play an important role in contributing to produce the initial lesions in the bacterial infection.

Biochemistry, 1994 Dec 20, 33(50), 14957 - 64
Supramolecular self-assembly of glutamine synthetase: mutagenesis of a novel intermolecular metal binding site required for dodecamer stacking; Dabrowski MJ et al.; Dodecameric glutamine synthetase (GS) from Escherichia coli assembles into highly ordered supramolecular protein tubes in the presence of several divalent metal ions . The molecular mechanism for this metal-induced self-assembly of the E . coli GS has been studied by molecular modeling and site-directed mutagenesis . The X-ray crystal structure of the nearly identical Salmonella typhimurium GS has been used to construct a model of the "stacked" complex between two dodecamers . A complementary fit, based on steric constraints, reveals a possible interaction between the N-terminal helices from adjacent dodecamers . The amino acid side chains of His and Met residues within the helices from each of the subunits of one face of a dodecamer lie within approximately 3.5 A of the analogous side chains in the subunits from the adjacent dodecamer in the stacked complex . His-4, Met-8, and His-12 from adjacent helices provide potential ligands for a binuclear metal binding site . Replacement of each of these surface residues with aliphatic amino acids has negligible effects on the enzymatic activity, the regulation of activity via adenylylation, and gross dodecameric structure . However, the rate and extent of metal ion-mediated self-assembly of GS tubules are reduced to < 2% of the wild-type protein in the single mutants H4A, H12L, and H12D . The M8L mutant demonstrates a 3-fold decrease in the bimolecular rate constant for stacking, but electron microscopy indicates that this mutant does form stacked tubes . The cysteine-containing mutants H4C, M8C, and H12C were also constructed.(ABSTRACT TRUNCATED AT 250 WORDS)

J Immunol, 1994 Dec 15, 153(12), 5634 - 42
Construction, expression, and immunogenicity of multiple tandem copies of the Schistosoma mansoni peptide 115-131 of the P28 glutathione S-transferase expressed as C-terminal fusions to tetanus toxin fragment C in a live aro-attenuated vaccine strain of Salmonella; Khan CM et al.; Genetic fusions have been constructed between the highly immunogenic but atoxic fragment C of tetanus toxin and a guest peptide, aa115-131, from the protective 28-kDa glutathione S-transferase Ag of Schistosoma mansoni . Fusions have been assembled to express one, two, four, and eight tandem copies of the peptide . The recombinant vectors have been electroporated into the nonvirulent aroA strain of Salmonella typhimurium SL3261 . The fusion proteins are soluble and stably expressed in Salmonella as evaluated by Western blotting with fragment C and glutathione S-transferase antisera . Mice have been immunized i.v . with a single dose of the live recombinant salmonellae . The strains are stable in mice and elicit Ab responses directed against fragment C, as determined by enzyme-linked immunosorbent assays . Ab responses were also detected against the guest peptide . The Ab responses improved dramatically toward the aa115-131 peptide with increasing copy number, with the octameric "repitope" fusion displaying the greatest potency . This approach may represent a general strategy for eliciting immune responses against peptides in live bacterial vaccines.

J Bacteriol, 1994 Dec, 176(24), 7630 - 7
Molecular characterization of the Salmonella typhimurium flhB operon and its protein products; Minamino T et al.; The flhB and flhA genes constitute an operon called flhB operon on the Salmonella typhimurium chromosome . Their gene products are required for formation of the rod structure of flagellar apparatus . Furthermore, several lines of evidence suggest that they, together with FliI and FliH, may constitute the export apparatus of flagellin, the component protein of flagellar filament . In this study, we determined the nucleotide sequence of the entire flhB operon from S . typhimurium . It was shown that the flhB and flhA genes encode highly hydrophobic polypeptides with calculated molecular masses of 42,322 and 74,848 Da, respectively . Both proteins have several potential membrane-spanning segments, suggesting that they may be integral membrane proteins . The flhB operon was found to contain an additional open reading frame capable of encoding a polypeptide with a calculated molecular mass of 14,073 Da . We designated this open reading frame flhE . The N-terminal 16 amino acids of FlhE displays a feature of a typical signal sequence . A maxicell labeling experiment enabled us to identify the precursor and mature forms of the flhE gene products . Insertion of a kanamycin-resistant gene cartridge into the chromosomal flhE gene did not affect the motility of the cells, indicating that the flhE gene is not essential for flagellar formation and function . We have overproduced and purified N-terminally truncated FlhB and FlhA proteins and raised antibodies against them . By use of these antibodies, localization of the FlhB and FlhA proteins was analyzed by Western blotting (immunoblotting) with the fractionated cell extracts . The results obtained indicated that both proteins are localized in the cytoplasmic membrane.

J Bacteriol, 1994 Dec, 176(24), 7625 - 9
Isolation and characterization of FliK-independent flagellation mutants from Salmonella typhimurium; Kutsukake K et al.; A flagellum of Salmonella typhimurium and Escherichia coli consists of three structural parts, a basal body, a hook, and a filament . Because the fliK mutants produce elongated hooks, called polyhooks, lacking filament portions, the fliK gene product has been believed to be involved in both the determination of hook length and the initiation of the filament assembly . In the present study, we isolated two mutants from S . typhimurium which can form flagella even in the absence of the fliK gene product . Flagellar structures were fractionated from these suppressor mutants and inspected by electron microscopy . The suppressor mutants produced polyhook-filament complexes in the fliK mutant background, while they formed flagellar structures apparently indistinguishable from those of the wild-type strain in the fliK+ background . Genetic and sequence analyses of the suppressor mutations revealed that they are located near the 3'-end of the flhB gene, which has been believed to be involved in the early process of the basal body assembly . On the basis of these results, we discuss the mechanism of suppression of the fliK defects by the flhB mutations and propose a hypothesis on the export switching machinery of the flagellar proteins.

Carcinogenesis, 1994 Dec, 15(12), 2899 - 903
Mutagenicity of nitric oxide is not caused by deamination of cytosine or 5-methylcytosine in double-stranded DNA; Schmutte C et al.; Several human tumors of diverse histological origin have a high incidence of C:G to T:A transition mutations at methylated CpG sites in tumor suppressor genes . We used a sensitive genetic assay to examine the ability of nitric oxide (NO), a physiological intra- and intercellular messenger molecule, to promote these transitions by deaminating cytosine (C) or methylcytosine (5mC) in double-stranded DNA . Exposure of a test double-stranded plasmid containing C or 5mC at the target site to NO in phosphate-buffered solution at pH 7.4 followed by transformation into Escherichia coli ung- strain to avoid repair of U did not result in a significant increase in reversion frequency . In addition, exposure of E . coli transformed with the target plasmid to an NO-releasing spermine-NO complex during log-phase growth did not result in larger numbers of revertants, whereas Salmonella typhimurium strain TA1535 showed a dose-responsive increase in reversion frequency when treated in the same way . We conclude that genotoxicity of NO is not caused by deamination of C or 5mC to U or T, respectively, in double-stranded DNA . This is supported by the finding that extracts of TA1535 contained high uracil-DNA glycosylase activity, suggesting that the difference in mutagenesis between the strains is not due to a lack of uracil repair . Therefore, mutational hot-spots seen in human tumor tissues at CpG sites are probably not due to the action of NO at 5mC.

Epidemiol Infect, 1994 Dec, 113(3), 425 - 34
Changing epidemiology of human salmonellosis in Hong Kong, 1982-93; Wong SS et al.; A comprehensive analysis of the epidemiology of salmonellosis in a major hospital in Hong Kong from 1982-93 is reported . The trend of salmonella isolations over the past 12 years and changes in the occurrence of individual serotypes are delineated . A total of 5328 isolates were analyzed . Groups B (Salmonella typhimurium and S . derby) and E (S . anatum) were the commonest serogroups isolated from the intestinal tract in all age groups . A significant increase in the isolation of group D salmonellae has been observed since 1989 . This is accounted for by a substantial rise in S . enteritidis isolation as seen in Western countries, despite a concomitant decrease of S . typhi . The extraintestinal isolation index (EII) is proposed as an index of the virulence potential of individual serotypes and serogroups . Group D salmonella was found to be the most invasive serogroup . While group D was the predominant serogroup isolated from extraintestinal sites in patients older than 1 year, group B serotypes (especially S . typhimurium) were more frequently seen in infants younger than 12 months.

J Immunol, 1994 Dec 1, 153(11), 4925 - 33
Phagocytic processing of exogenous particulate antigens by macrophages for presentation by class I MHC molecules; Harding CV et al.; Exogenous Ags that are processed in vacuolar endocytic compartments are generally presented by class II MHC molecules and not class I MHC (MHC-I) molecules, which conventionally present cytoplasmic or endogenous Ags . Accordingly, i.v . immunization of C57BL/6 mice with soluble OVA did not elicit a CD8 T cell response . However, i.v . immunization with OVA coupled to Latex particles (Latex-OVA) elicited an OVA-specific CD8 T cell response in vivo (particles from 59 to 2000 nm diameter were effective) . In vitro, Latex-OVA was processed by H-2b macrophages and presented by Kb at least 100- to 1000-fold more efficiently than was soluble OVA . Inhibition of phagocytosis by cytochalasin D blocked the processing of Latex-OVA, whereas processing was not blocked by Brefeldin A . Latex-OVA was presented directly by H-2b macrophages or after "regurgitation" of processed OVA peptide from viable MHC-disparate macrophages for binding to surface Kb molecules on fixed H-2b macrophages . Peptide regurgitation was observed during processing of both Latex-OVA and Salmonella typhimurium 14028s that express an OVA fusion protein (Crl-OVA) . However, the regurgitation pathway was less efficient than direct processing by viable H-2b macrophages . Thus, macrophages express an alternate pathway that allows MHC-I presentation of vacuolar exogenous particulate Ags, including inert synthetic particles without lipid membranes and intravacuolar bacteria . Peptides from these Ags are released from intracellular compartments to bind to surface MHC-I molecules, but peptide-MHC-I complexes also may be generated within intracellular compartments.

Infect Immun, 1994 Dec, 62(12), 5519 - 27
Development and evaluation of an experimental vaccination program using a live avirulent Salmonella typhimurium strain to protect immunized chickens against challenge with homologous and heterologous Salmonella serotypes; Hassan JO et al.; A stable live avirulent, genetically modified delta cya delta crp Salmonella typhimurium vaccine strain, chi 3985, was used in several vaccination strategies to evaluate its use in the control of Salmonella infection in chickens . Oral vaccination of chickens at 1 and at 14 days of age with 10(8) CFU of chi 3985 protected against invasion of spleen, ovary, and bursa of Fabricius and colonization of the ileum and cecum in chickens challenged with 10(6) CFU of virulent homologous Salmonella strains from group B . Chickens challenged with heterologous Salmonella strains from groups C, D, and E were protected against visceral invasion of spleen and ovary, while invasion of the bursa of Fabricius and colonization of ileum and cecum was reduced in vaccinated chickens . Oral vaccination at 2 and at 4 weeks of age induced an excellent protection against challenge with virulent group B Salmonella serotypes and very good protection against challenge with group D or E Salmonella serotypes, while protection against challenge with group C Salmonella serotypes was marginal but significant . Vaccination at 2 and at 4 weeks of age also protected vaccinated chickens against challenge with 10(8) CFU of highly invasive S . typhimurium or S . enteritidis strains . The protection of chickens vaccinated with chi 3985 against challenge with homologous and heterologous Salmonella serotypes is outstanding, and the complete protection against ovarian invasion in chickens challenged with 10(8) CFU of highly invasive S . typhimurium or S . enteritidis strains suggests that vaccination of chickens with chi 3985 can complement the present hygiene- and sanitation-based Salmonella control measures . This paper reports a breakthrough in the use of live avirulent vaccine to control Salmonella carriers in chickens.

Immunology, 1994 Dec, 83(4), 670 - 4
Antigen expressed by Salmonella typhimurium is processed for class I major histocompatibility complex presentation by macrophages but not infected epithelial cells; Harding CV et al.; Macrophages were shown to mediate class I major histocompatibility complex (MHC-1) presentation of a fusion protein (Crl-OVA) expressed in Salmonella typhimurium, a bacterium which fails to escape from vacuolar compartments after phagocytosis or penetration into host cells . Salmonella typhimurium also penetrates into non-phagocytic intestinal epithelial cells, a portal of entry for systemic infection . We tested the ability of infected epithelial cells to process antigen expressed by S . typhimurium for presentation by MHC-I molecules to CD8+ T cells . CMT-93 murine adenocarcinoma cells expressed Kb and effectively presented the OVA 257-264 peptide to CD8 OVA T-hybridoma cells, but infected CMT-93 cells failed to process Crl-OVA expressed in S . typhimurium . Therapeutically useful MHC-I-restricted cytotoxic T-lymphocyte (CTL) responses may be generated by macrophage presentation of Salmonella antigens or recombinant antigens expressed in Salmonella vaccine vectors . Our data suggest that an inability of epithelial cells to present these antigens may limit the utility of CTL in epithelial immunity in salmonellosis, but studies of additional epithelial cell systems are needed.

Epidemiol Mikrobiol Imunol, 1994 Dec, 43(4), 177 - 9
{Distribution of Salmonella typhimurium phage types isolated from various sources in the Slovak Republic 1992-1993}; Majtanova L; The author assessed phagotypes of Salmonella typhimurium strains isolated in 1992 and 1993 from patients with salmonellosis (n = 1,099) from foods (mainly home pig-slaughtering) (n = 10), surface water (n = 1) and one case of a nosocomial infection . In the reference laboratory for phagotyping of Salmonellae a total of 1,111 strains were examined -376 in 1992 and 735 in 1993 . During this period phagotypes 2b dominated (27.81%), 1 (26.28%) and 1 variant (25.02%), whereby in 1993 among the examined strains phagotypes 1 and 1 variant predominated (37% and 36.8% resp.) . All strains isolated from foodstuffs in 1993 belonged to these two phagotypes . Types of 12.8% of the total number of examined strains could not be identified.

Arzneimittelforschung, 1994 Dec, 44(12), 1366 - 8
Mutagenicity studies of benzalazine; Herzog R et al.; Benzalazine (2-hydroxy-5-{(4-carboxyphenyl) azo} benzoic acid, CAS 64896-26-0), a new agent for the treatment of ulcerative colitis and Crohn's disease of the large intestine, was studied for genotoxic effects by using the following short-term in vitro and in vivo test: 1 . reverse mutation test (Ames method) on Salmonella typhimurium, 2 . HGPRT (hypoxanthine-guanine phosphoribosyl transferase) point mutation test on V79 hamster cells, 3 . in vivo cytogenetic test on Chinese hamster cells, and 4 . in vivo sister chromatid exchange test on Chinese hamster cells . Benzalazine did not show any positive response in the reverse mutation test, HGPRT-point mutation test, in vivo cytogenetic and sister chromatid exchange test.

Indian J Med Res, 1994 Dec, 100, 266 - 7
An outbreak of multidrug resistant Salmonella typhimurium in Delhi (India); Wattal C et al.; A total of 85 patients with multidrug resistant S . typhimurium were isolated between May and September 1991 at the Sir Ganga Ram Hospital, New Delhi, India . Fifty eight (72.5%) patients out of 80 stool culture positives suffered from enteritis and 23 (39.6%) of them settled with oral rehydration therapy alone . All strains were sensitive to 4 aminoquinolones (oflaxcin) but five were resistant to third generation cephalosporin (Cefotaxime; MIC between 50-75 micrograms/ml) whereas 88-96 per cent isolated were resistant to most of the other antibiotics . The convalescent carrier rate was prolonged with the use of antibiotics . The phage type of S . typhimurium isolated from the index and other cases was 178 and multidrug resistance strains had seven plasmids (1.2 to 16 kb) . Barrier nursing and sodium hypochlorite disinfection helped in limiting the outbreak.

Comput Chem, 1994 Dec, 18(4), 391 - 6
Artificial neural networks applied to classification of mutagenic activity of nitro-substituted polycyclic aromatic hydrocarbons; Song XH et al.; Three-layer artificial neural networks (ANN) with back-propagation of error have been applied to classification of nitro-substituted polycyclic aromatic hydrocarbons (NPAH) based on the regularity that the structure difference of NPAH compounds leads to different mutagenic activity towards Salmonella typhimurium . The network's architecture and parameters were optimized to give maximum correct classification rate of 93.8% for two different classes of NPAHs: weakly active and strongly active ones . The results compared favourably with those obtained by nonlinear mapping pattern recognition method . Considering that the most important factor for NPAH's mutagenicity might be the electron effect of the substituted nitro-groups, an electrotopological state index of nitrogen atom was introduced additionally as one of network's inputs, and the correct classification rate was consequently raised to 97.5% . The network's prediction ability for untrained samples was also satisfactory.

Appl Environ Microbiol, 1994 Dec, 60(12), 4345 - 50
Flow cytometric analysis of the cellular DNA content of Salmonella typhimurium and Alteromonas haloplanktis during starvation and recovery in seawater; Lebaron P et al.; Flow cytometry was used to investigate the heterogeneity of the DNA content of Salmonella typhimurium and Alteromonas haloplanktis cells that were starved and allowed to recover in seawater . Hoechst 33342 (bisbenzimide) was used as a DNA-specific dye to discriminate between DNA subpopulations . The DNA contents of both strains were heterogeneous during starvation . S . typhimurium cells contained one or two genomes, and A . haloplanktis cells contained up to six genomes . S . typhimurium genomes were fully replicated at the onset of starvation . Each replication cycle was completed in the early stage of starvation for A . haloplanktis by stopping cells in the partition step of the cell cycle prior to division . Multigenomic marine cells can undergo rapid cell division without DNA synthesis upon recovery, resulting in large fluctuations in the DNA contents of individual cells . In contrast, the heterogeneity of the DNA distribution of S . typhimurium cells was preserved during recovery . The fluctuations in the DNA fluorescence of this strain seem to be due to topological changes in DNA . Flow cytometry may provide a new approach to understanding dynamic and physiological changes in bacteria by detecting cellular heterogeneity in response to different growth conditions.

Appl Environ Microbiol, 1994 Dec, 60(12), 4263 - 7
Reduction and mutagenic activation of nitroaromatic compounds by a Mycobacterium sp; Rafii F et al.; Mycobacterium sp . strain Pyr-1 cells, which were grown to the stationary phase in media with and without pyrene, were centrifuged and resuspended in a medium containing 1-nitropyrene . Cells that had been grown with pyrene oxidized up to 20% of the added 1-nitropyrene to 1-nitropyrene-cis-9,10- and 4,5-dihydrodiols . However, cells that had been grown without pyrene reduced up to 70% of the 1-nitropyrene to 1-aminopyrene but did not produce dihydrodiols . The nitroreductase activity was oxygen insensitive, intracellular, and inducible by nitro compounds . Nitroreductase activity was inhibited by p-chlorobenzoic acid, o-iodosobenzoic acid, menadione, dicumarol, and antimycin A . Extracts from cells that had been grown without pyrene activated 1-nitropyrene, 1-amino-7-nitrofluorene, 2,7-dinitro-9-fluorenone, 1,3-dinitropyrene, 1,6-dinitropyrene, and 6-nitrochrysene to DNA-damaging products, as shown in Salmonella typhimurium tester strains by the reversion assay and by induction of the umuC gene . Activation of nitro compounds, as shown by the umu test, was enhanced by NADPH . This study shows that Mycobacterium sp . strain Pyr-1 metabolizes nitroaromatic compounds by both oxidative and reductive pathways . During reduction, it generates products that are mutagenic.

Appl Environ Microbiol, 1994 Dec, 60(12), 4255 - 62
Detection of Salmonella typhimurium in dairy products with flow cytometry and monoclonal antibodies; McClelland RG et al.; Flow cytometry, combined with fluorescently labelled monoclonal antibodies, offers advantages of speed and sensitivity for the detection of specific pathogenic bacteria in foods . We investigated the detection of Salmonella typhimurium in eggs and milk . Using a sample clearing procedure, we determined that the detection limit was on the order of 10(3) cells per ml after a total analysis time of 40 min . After 6 h of nonselective enrichment, the detection limits were 10 cells per ml for milk and 1 cell per ml for eggs, even in the presence of a 10,000-fold excess of Escherichia coli cells.

Behring Inst Mitt, 1994 Dec, (95), 57 - 66
Synthesis and secretion of bacterial antigens by attenuated Salmonella via the Escherichia coli hemolysin secretion system; Gentschev I et al.; We describe a plasmid system which allows the secretion of foreign antigens in attenuated Salmonella aroA strains by the secretion apparatus of E . coli hemolysin . The gene (or gene fragment) encoding the antigen is inserted in frame into a residual position of the hlyA gene, encoding the HlyA secretion signal (HlyAs) . Generally, the fused gene is efficiently expressed and the synthesized antigen is in part secreted into the culture supernatant and in part exposed on the surface of the producing Salmonella strain . The successful use of this approach is demonstrated with two antigens of Salmonella typhimurium, PagC and SlyA, both of which are potent virulence factors but produced only in small amounts under in vitro culture conditions and two virulence proteins of Listeria monocytogenes, p60 and listeriolysin . Interestingly the listeriolysin fusion protein proved to be cytolytically active and allowed, when expressed in Salmonella, the escape of these bacteria into the cytoplasm of infected macrophages.

Microb Pathog, 1994 Dec, 17(6), 409 - 23
Comparison of Salmonella typhi and Salmonella typhimurium invasion, intracellular growth and localization in cultured human epithelial cells; Mills SD et al.; Invasion of the cultured epithelial cell lines HeLa, Henle-407, and Caco-2 (polarized and nonpolarized) by Salmonella typhi and Salmonella typhimurium were compared using conventional gentamicin invasion assays . Additionally, the mechanisms of invasion and intracellular trafficking by S . typhi and S . typhimurium were compared in HeLa cells using indirect immunofluorescence microscopy . S . typhi strain Ty2 was invasive in all human cell lines tested, including apical uptake into polarized Caco-2 cell monolayers . This strain also replicated at levels similar to S . typhimurium strain SL1344 inside HeLa and Henle-407 cells . Indirect immunofluorescence microscopy confirmed that S . typhi, like S . typhimurium, induced membrane ruffles and cytoskeletal rearrangements upon contact with HeLa cell surfaces . Ruffling induced by S . typhi and S . typhimurium was accompanied by macropinocytosis of the fluid phase endocytic marker fluorescein-dextran-sulphate and by aggregation of cell surface class I MHC heavy chain . Intracellular lysosomal trafficking of S . typhi and S . typhimurium in HeLa cells was also studied . The lysosomal membrane glycoprotein marker h-lamp-2 colocalized with S . typhi-containing vacuoles, as previously shown for S . typhimurium . The soluble lysosomal enzyme marker cathepsin D also was found within S . typhi-containing vacuoles to the same extent as previously published for S . typhimurium . The results from this study suggest that S . typhi and S . typhimurium use similar mechanisms for invasion and intracellular trafficking in cultured human epithelial cells.

Hum Exp Toxicol, 1994 Dec, 13(12), 861 - 5
Metabolism and genotoxicity of the halogenated alkyl compound tris(2,3-dibromopropyl)phosphate; van Beerendonk GJ et al.; 1 . The genotoxicity of the flame retardant tris(2,3-dibromopropyl)phosphate (Tris-BP) was studied in vivo . Results showed that Tris-BP was highly clastogenic, but it could only initiate a low number of preneoplastic foci in the rat liver in vivo . In Drosophila, Tris-BP could be classified as a cross-linking agent, because it was more clastogenic than mutagenic . The use of completely deuterated Tris-BP as a metabolic probe revealed that cytochrome P450 and most likely the formation of 2-bromoacrolein (2BA) from Tris-BP is important for the observed genotoxic effects . 2 . In contrast to the high mutagenicity of Tris-BP and 2BA in Salmonella typhimurium, we were unable to detect an increase in mutation frequency of 2BA on the hprt locus of human TK6 cell line . In another system, using a shuttle vector modified with 2BA:DNA-adducts, also no increase in mutation frequency could be detected in human cells . This low mutagenicity of 2BA corresponds with its low mutagenicity in Drosophila and its low induction of preneoplastic foci in the rat liver . 3 . Several DNA adducts of 2BA have been identified, including an unstable 3-(bromooxypropyl)thymidine adduct which has the potential to form cross-links and a cyclic 3,N4-(bromo)propeno-deoxycytidine adduct which can possibly be involved in the clastogenicity of Tris-BP . 4 . Taken together, these data indicate that Tris-BP and 2BA may not effectively induce gene mutations in eukaryotic systems, but rather be potent clastogens . Risk assessment of these and related compounds should therefore be based on the knowledge of clastogens rather than mutagens.

Tierarztl Prax, 1994 Dec, 22(6), 529 - 31
{The effectiveness in calves of subcutaneous vaccination with the Salmonella vaccine Murivac}; Steinbach G et al.; Subcutaneous administration of a Salmonella typhimurium bacterin, given twice in an interval of two weeks, protected calves against the oral infection with a Salmonella wild strain.

Int J Food Microbiol, 1994 Dec, 24(1-2), 137 - 45
Growth and survival of Yersinia enterocolitica, Salmonella and Bacillus cereus in Brie stored at 4, 8 and 20 degrees C; Little CL et al.; The potential of stabilised Brie to support growth of the food-borne pathogens, Yersinia enterocolitica serotypes 0:3 and 0:9, Salmonella typhimurium, S . dublin (both dairy isolates), S . thompson, and Bacillus cereus (3 dairy isolates), after contamination on opening the cheese package was evaluated . Growth kinetics of the different pathogens was determined in relation to inoculum size and storage temperature (4 degrees C, 8 degrees C and 20 degrees C) . Only Y . enterocolitica was found to grow on the surfaces (outer and exposed) of Brie at all three storage temperatures . Growth of this pathogen during refrigerated storage must be avoided to ensure safety . The numbers of B . cereus and Salmonella increased at 20 degrees C but declined at a slow rate during storage at 4 degrees C and 8 degrees C . However, survival of these two pathogens for extended periods at abuse temperatures could pose a health hazard . Predictions from a predictive modelling program (MFS model) and a modelling database (Food Micromodel) were compared to observed growth values in Brie . Although accurate in the case of B . cereus at 20 degrees C, predicted generation times were in general found to be considerably shorter than the observed values, i.e., overall they were 'fail safe' . Predicted lag times, however, were generally longer compared to observed values in the case of Y . enterocolitica and at low inocula for Salmonella, and would 'fail dangerous' if used for predictive purposes.

Int J Food Microbiol, 1994 Dec, 24(1-2), 125 - 36
Development of antibiotic-resistant strains for the enumeration of foodborne pathogenic bacteria in stored foods; Blackburn CD et al.; Strains of Aeromonas spp., Salmonella enteritidis phage type 4, Salmonella typhimurium, verotoxigenic Escherichia coli O157:H7 (VTEC) and Yersinia enterocolitica resistant to streptomycin, nalidixic acid and a combination of both antibiotics were selected . When compared with the parent strains, most of the antibiotic-resistant strains had slightly slower growth rates at their optimum incubation temperature but the difference was reduced progressively when the temperature was lowered . Some antibiotic-resistant strains had considerably slower growth rates in the presence of the relevant antibiotic and these were not used further . Several agar and impedance media with added streptomycin and nalidixic acid were assessed for the enumeration of the antibiotic-resistant strains in artificially contaminated stored foods . Differential/selective media were required to enumerate low numbers of antibiotic-resistant strains in certain foods . The following agar and impedance media were selected: Aeromonas Agar (Ryan) for Aeromonas spp., Xylose Lysine Agar and Lysine Iron Cysteine Neutral Red Medium for Salmonella, Eosin Methylene Blue Agar and Coliform Medium for VTEC, and Yersinia Selective Agar without selective agents for Yersinia enterocolitica . The agar and impedance media have been used successfully to enumerate antibiotic-resistant strains inoculated into foods and stored at different temperatures.

Biomed Environ Sci, 1994 Dec, 7(4), 346 - 56
Mutagenicity and induction of drug-metabolizing enzyme activity by LPG combustion particulates in rats; Yin XJ et al.; Methylene chloride extracts of particulates from liquefied petroleum gas (LPG) combustion appliance were studied by using Ames test, micronucleus test and inducibility of pulmonary and hepatic aryl hydrocarbon hydroxylase (AHH) and glutathione S-transferase (GST) in rats . The extracts showed mutagenicity for Salmonella typhimurium strain TA98 and its derivatives TA98NR and TA98/1,8-DNP6 with or without S9 mix . The revertants in strains TA98NR and TA98/1,8-DNP6 were less than 40% and 50% of that in strain TA98 without S9 mix, respectively . Positive results were obtained in mouse bone marrow micronucleus assay . Intratracheal instillation of the extracts led to increase in pulmonary (but not hepatic) AHH and GST activities in rats . It was seen that AHH was more sensitive than GST to induction by the extracts.

Cancer Lett, 1994 Nov 25, 87(1), 25 - 32
Suppressive effects of the extracts of Japanese edible seaweeds on mutagen-induced umu C gene expression in Salmonella typhimurium (TA 1535/pSK 1002) and tumor promotor-dependent ornithine decarboxylase induction in BALB/c 3T3 fibroblast cells; Okai Y et al.; Some of epidemiological data indicated that ubiquitous consumption of seaweeds in Japan may be a possible protective factor against some types of tumor . To analyse this problem, the authors studied the antimutagenic and antitumor promotion activities in methanol-soluble extracts of typical edible seaweeds which showed suppressive effects on 3-amino-1,4-dimethyl-5H-pyrido{4,3-b}indol (Trp-P-1)-induced umu C gene expression in SOS response of Salmonella typhimurium (TA 1535/pSK 1002) and 12-O-tetradecanoylphorbol-13-acetate (TPA)-dependent ornithine decarboxylase induction in BALB/c 3T3 fibroblast cells . Although eight varieties of edible seaweeds including chlorophyta, Phaenophyta and Rhodophyta showed significant antimutagenic and antipromotion activities, they expressed the activities different from each other . Among these seaweeds, Enteromorpha prolifera ('Sujiaonori' in Japanese) and Porphyra tenera ('Asakusanori') showed relatively strong suppressive activities in both antimutagenic and antipromotion assays compared with other seaweeds . These seaweeds contained considerable amounts of beta-carotene as a possible active principle with anticarcinogenic activity . This compound was partially associated with the antimutagenic activity in the seaweed extract, but did not contribute to the antipromotion activity of seaweed extract under our experimental conditions . These results strongly suggest that Japanese edible seaweeds have possible antimutagenic and antipromotion activities probably associated with antitumor activity.

Proc Natl Acad Sci U S A, 1994 Nov 22, 91(24), 11606 - 10
Detection and classification of mutagens: a set of base-specific Salmonella tester strains; Gee P et al.; A detection and classification system for mutagens has been developed that identifies the six possible base-pair substitution mutations . A set of six Salmonella typhimurium (TA7001 to TA7006) strains has been constructed, each of which carries a unique missense mutation in the histidine biosynthetic operon . In addition to the his mutation, these strains carry different auxiliary features that enhance the mutability of the target his mutation . These include the R factor pKM101, which has the SOS-inducible mucAB system; a deletion of the uvrB component of excision repair; and rfa mutations to increase the accessibility of bulky chemicals to the bacteria . Another set of strains (TA7041 to TA7046) contain a wild-type rfa gene . Reversion via the base substitution unique to each strain was verified by sequence analyses of > 800 revertants obtained from different types of mutagens . The strains have considerably lower spontaneous reversion frequencies and detect a variety of mutagens at a sensitivity comparable to the Salmonella tester strains TA100, TA102, and TA104 . The low spontaneous frequency of reversion of a mixture of the six tester strains (approximately 10 revertants per plate) enables a single mutation assay with the mixture that is followed by classification of the type of mutation with the individual strains.

Proc Natl Acad Sci U S A, 1994 Nov 8, 91(23), 11261 - 5
Construction, expression, and immunogenicity of the Schistosoma mansoni P28 glutathione S-transferase as a genetic fusion to tetanus toxin fragment C in a live Aro attenuated vaccine strain of Salmonella; Khan CM et al.; A vector has been constructed to allow genetic fusions of guest antigens via a hinge domain to the C terminus of the highly immunogenic C fragment of tetanus toxin . A fusion has been constructed with the gene encoding the protective 28-kDa glutathione S-transferase (EC 2.5.1.18) from Schistosoma mansoni . The recombinant vector has been electroporated into the nonvirulent Salmonella typhimurium aroA live vaccine strain SL3261 . The corresponding chimeric protein is stably expressed in a soluble form in Salmonella as evaluated by Western blotting with fragment C and glutathione S-transferase antisera . Mice immunized intravenously with a single dose of the live recombinant bacteria elicit antibodies to both fragment C and glutathione S-transferase as detected by enzyme-linked immunosorbent assays . Furthermore, all of the mice were solidly protected when challenged with lethal doses of either tetanus toxin or the virulent Salmonella typhimurium strain C5 . Mice have also elicited antibodies to fragment C and glutathione S-transferase after oral immunization . It may be that a live trivalent vaccine against typhoid, tetanus, and schistosomiasis is feasible.

J Bacteriol, 1994 Nov, 176(22), 6852 - 60
The Salmonella typhimurium katF (rpoS) gene: cloning, nucleotide sequence, and regulation of spvR and spvABCD virulence plasmid genes; Kowarz L et al.; The spv region of Salmonella virulence plasmids is essential for the development of a systemic infection in mice . Transcriptional activation of the spvABCD operon occurs during stationary growth phase and is mediated by the regulatory gene product SpvR . We have previously shown that expression of a spvRAB'-cat fusion in Escherichia coli was dependent on the katF (rpoS) locus which encodes an alternative sigma factor (sigma S) . The katF gene from Salmonella typhimurium has been cloned, sequenced, and used to construct Salmonella katF mutants by allelic replacement . Using these mutants, we demonstrated by mRNA and gene fusion analyses that sigma S, in conjunction with SpvR, controls the transcription of the regulatory gene spvR . In a second series of experiments, we sought to clarify the relationship between sigma S and SpvR in the control of spvABCD transcription . It was shown that expression of a transcriptional spvAB'-lacZ fusion could be restored in E . coli and Salmonella katF mutants when spvR was expressed in trans from an exogenous promoter . Moreover, identical spvA mRNA startpoints were detected in katF+ and katF strains . These results indicate that the reduction of spvABCD transcription in katF mutants is mainly due to decreased expression of spvR . Finally, mouse inoculation studies with S . typhimurium katF mutants of both wild-type and virulence plasmid-cured strains suggest that katF contributes to Salmonella virulence via the regulation of chromosomal genes in addition to that of spv genes.

J Bacteriol, 1994 Nov, 176(21), 6688 - 96
Analysis of promoters controlled by the putative sigma factor AlgU regulating conversion to mucoidy in Pseudomonas aeruginosa: relationship to sigma E and stress response; Martin DW et al.; Alginate overproducition by mucoid Pseudomonas aeruginosa is a critical pathogenic determinant expressed by this organism during chronic infections in cystic fibrosis . Conversion to mucoidy and a subsequent loss of mucoid character can occur via different mutations in the algU mucA mucB gene cluster . The algU gene encodes a 22.2-kDa putative alternative sigma factor required for expression of the critical alginate biosynthetic gene algD . In this work, algU transcription was studied by S1 nuclease protection analysis . Transcription from the promoter proximal to the algU coding region was found to be dependent on AlgU . The -35 and -10 sequences of this newly mapped promoter showed strong similarity ot the promoters of two other critical alg genes: algD and algR . The proximal promoter of algR was also shown to depend on algU . Interestingly, the putative -35 and -10 regions of all three promoters displayed striking similarity to the consensus sequence of the sigma E-dependent promoters in Escherichia coli and Salmonella typhimurium . This 24-kDa sigma factor, controlling genes participating in resistance to high temperatures and oxidative stress, has been previously biochemically characterized, but the gene for sigma E remained unidentified . To examine whether AlgU is related to sigma E, the effect of algU inactivation on the sensitivity of P . aeruginosa to killing by heat and reactive oxygen intermediates was tested . Two isogenic pairs of algU+ and algU mutant strains were compared . The algU mutants, irrespective of the mucoid status of the parental strains, displayed increased sensitivity to killing by paraquat, known to generate intracellular superoxide radicals, and heat . Further lgobal homology searches revealed the presence of a previously unrecognized E . coli gene with the predicted gene product showing a striking 66% identity to AlgU . The corresponding gene from S . typhimurium was cloned and sequenced, and it is displayed one amino acid substitution relative to its E . coli equivalent . AlgU and its close homologs in E . coli and S . typhimurium may be functionally related.

Carcinogenesis, 1994 Nov, 15(11), 2605 - 11
Substance-dependent sex differences in the activation of benzylic alcohols to mutagens by hepatic sulfotransferases of the rat; Glatt H et al.; Six primary and 10 secondary benzylic alcohols derived from polycyclic aromatic hydrocarbons were tested for mutagenicity in Salmonella typhimurium TA98 in the presence of varying amounts of hepatic cytosol from adult male and female rats and 3'-phosphoadenosine-5'-phosphosulfate, the cofactor for sulfotransferases . With the exception of 1-(9-anthryl)ethanol, 4H-cyclopenta{def}-phenanthren-4-ol and 10-hydroxy-7,8,9,10-tetrahydrobenzo{a}pyrene, all the benzylic alcohols were activated to mutagens . For 1-(1-pyrenyl)ethanol (1-HEP), 1-(2-pyrenyl)ethanol (2-HEP), 6-hydroxymethylanthanthrene (6-HMAA), 2-hydroxymethylpyrene (2-HMP), 10H-indeno{1,2,7,7a-bcd}pyren-10-ol (OH-IP), 3-hydroxy-3,4-dihydrocyclopenta{cd}pyrene (3-OH-H2-CPcdP) and 1-(6-benzo{a}pyrenyl)ethanol (6-HEBP), this is the first observation of a mutagenic activity . The primary alcohols 1-hydroxymethylpyrene, 2-HMP, 9-hydroxymethylanthracene, 7-hydroxymethyl-12-methylbenz{a}anthracene and 6-hydroxymethylbenzo{a}pyrene, as well as the secondary alcohols 1-HEP and 3-OH-H2-CPcdP, were more efficiently activated by hepatic cytosol from females than by preparations from males (2.6- to 8-fold) . A further compound, 6-HEBP showed significant, but relatively weak, effects in the presence of cytosol from females, whereas it was inactive in the presence of hepatic cytosol from males . The reverse sex difference was observed in the activation of 4H-cyclo-penta{def}chrysen-4-ol, the activity of cytosol from males amounting to about four times that from females . Four other compounds, 2-HEP, 7-hydroxy-7,8,9,10-tetrahydrobenzo{a}pyrene, 6-HMAA and OH-IP, were activated with similar efficiency by hepatic cytosol from both sexes (< two-fold differences) . The study indicates that different sulfotransferases are involved in the bioactivation of benzylic alcohols, including forms preferentially expressed in females as well as forms preferentially expressed in males, and that these enzymes qualitatively differ in their substrate tolerance for benzylic alcohols.

Carcinogenesis, 1994 Nov, 15(11), 2523 - 9
Activation of procarcinogens by human cytochrome P450 enzymes expressed in Escherichia coli . Simplified bacterial systems for genotoxicity assays; Shimada T et al.; Bacterial assays were used to examine the activation of 14 known procarcinogens by cytochrome P450 (P450) enzymes . Human P450s 1A1, 1A2 and 3A4 were expressed in Escherichia coli with slight modification of their N-terminal sequences . Genotoxicity was measured by the induction of the SOS response in Salmonella typhimurium NM2009 (TA1535/pSK1002/pNM12), which contains a umuC regulatory sequence attached to the lacZ reporter gene . Conditions for analysis were examined using E . coli membranes and purified enzymes . Membrane fractions, fortified with NADPH-P450 reductase, were found to be useful preparations for measuring activation of the procarcinogens . Conditions of linearity were established for these assays and the systems were applied to several particular problems related to bioactivation of procarcinogens by P450s . The patterns of activation of the 14 individual chemicals were consistent with the literature developed using human liver microsomes, purified liver P450s and other approaches . The P450s expressed in bacterial membranes could be inhibited by antibodies . 7,8-Benzoflavone inhibited P450s 1A1 and 1A2 and stimulated P450 3A4 in the membranes . The contributions of P450s 1A1 and 1A2 were distinguished with some of the arylamines and 7,8-dihydroxy-7,8-dihydrobenzo{a}pyrene . Recombinant P450 3A4 was found to be more active than P450 1A2 in the activation of aflatoxin B1 at all substrate concentrations examined.

Carcinogenesis, 1994 Nov, 15(11), 2507 - 16
Synthesis and mutagenicity of the diastereomeric fjord-region 11,12-dihydrodiol 13,14-epoxides of dibenzo{a,l}pyrene; Luch A et al.; Extensive tumorigenicity studies in rodents revealed that dibenzo{a,l}pyrene (DB{a,l}P) is the most potent carcinogen among all polycyclic aromatic hydrocarbons (PAHs) tested so far . The structure of the genotoxic metabolite(s) responsible for this exceptional carcinogenicity is unknown . The fjord-region syn- and anti-DB{a,l}P-11,12-dihydrodiol 13,14-epoxides (syn- and anti-DB{a,l}PDE) were synthesized to clarify their role as possible ultimate mutagenic and carcinogenic metabolites of DB{a,l}P.9-Formyl-11,12-dimethoxybenzo{g} chrysene was prepared from 9-phenanthrylacetic acid by a photochemical route . After reaction of the aldehyde with trimethylsulfonium iodide to generate an oxiranyl side-chain, treatment with boron trifluoride produced the key intermediate 11,12-dimethoxy-DB{a,l}P in 14% overall yield . From 11,12-dimethoxy-DB{a,l}P the syn- and anti-DB{a,l}PDE were stereoselectively prepared via the trans-11,12-dihydrodiol . The mutagenicity of the syn- and anti-DB{a,l}PDE was examined in four his- strains of Salmonella typhimurium and in Chinese hamster V79 cells . In all five test systems, the new dihydrodiolepoxides were more potent than any of the previously investigated dihydrodiolepoxides . The specific mutagenicity observed with anti-DB{a,l}PDE in strain TA104 exhibited the highest value ever found with any compound in any his- strains of S.typhimurium . The same appears to be true for the activity observed with this compound in V79 cells . In all five systems, syn-DB{a,l}PDE was only moderately less active than its anti-diastereomer (approximately 2-fold) . The exceptional mutagenic activities of these dihydrodiolepoxides may be one of the reasons for the exceptional carcinogenic activity of DB{a,l}P.

Infect Immun, 1994 Nov, 62(11), 5095 - 101
Further characterization of the PhoP regulon: identification of new PhoP-activated virulence loci; Belden WJ et al.; Salmonella typhimurium survival within macrophages is an essential virulence property necessary to enteric fever pathogenesis . This survival requires coordinate transcriptional activation of virulence genes within acidified macrophage phagosomes . Virulence gene transcription is regulated by a two-component system comprising the PhoP (transcriptional activator) and PhoQ (sensor-kinase) proteins . Thirteen new PhoP-activated loci (designated pagD to pagP) encoding membrane or secreted proteins have been identified by use of the transposon TnphoA . Three of these loci have a chromosomal location that was linked to the previously identified pagC locus . Strains with TnphoA insertions in pagD, pagJ, pagK, and pagM were significantly attenuated for mouse virulence (50% lethal dose greater than 1,000 times that of wild-type bacteria) . No strains with pag::TnphoA insertions were found to have altered sensitivity to the cationic antimicrobial peptide NP-1 defensin . PhoP and PhoQ are pleotropic regulators of membrane or secreted proteins, suggesting that the ability to effect a global change in the expression of these proteins is required for S . typhimurium survival within acidified macrophage phagosomes.

Infect Immun, 1994 Nov, 62(11), 4739 - 46
Invasiveness and persistence of Salmonella enteritidis, Salmonella typhimurium, and a genetically defined S . enteritidis aroA strain in young chickens; Cooper GL et al.; Newly hatched chicks were dosed orally with a Salmonella typhimurium wild-type strain, an S . enteritidis wild-type strain, and a genetically defined S . enteritidis aroA vaccine candidate, strain CVL30 . The S . typhimurium strain, 2391 Nalr, was virulent in newly hatched chicks and caused deaths in 7 of 20 chicks after an oral dose of 10(5) organisms . The S . enteritidis wild-type strain, LA5, caused death in 1 of 25 chicks and gross pathology including pericarditis and perihepatitis in 6 of the 24 survivors after an oral dose of 10(9) organisms . S . enteritidis aroA CVL30, attenuated by ca . 6.5 log10 in BALB/c mice, was nonvirulent when administered orally to chicks and did not cause morbidity . When newly hatched chicks were dosed, the pattern of invasion and colonization of the reticuloendothelial system by strain CVL30 was similar to that of its parent strain, LA5, irrespective of the dose . Oral inoculation of newly hatched chicks with < 10 organisms of S . enteritidis LA5 or CVL30 was followed by multiplication in the cecal contents . Within 3 days of hatching, the pH of the cecal contents was reduced from ca . 7 to 5 . Samples of gut contents were inoculated in vitro . The S . enteritidis strains multiplied in samples taken from the ileum and duodenum irrespective of age but multiplied in the cecal samples from newly hatched chicks only . Invasion from the gut by S . enteritidis LA5 and CVL30 was both age and dose dependent.

J Toxicol Sci, 1994 Nov, 19(4), 235 - 8
Evidence of partial involvement of P-450 2D in mutagenic activation of benzo(a)pyrene in liver S-9 fraction from untreated rats; Masuda M et al.; In order to clarify which species of cytochrome P-450 is involved in activation of benzo(a)pyrene (BP) in untreated rat liver, strain and sex differences in the ability of rat liver 9000 g supernatant (S-9) to mutagenically activate BP was investigated using Ames test . The numbers of histidine revertants in Ames test after pre-incubation of TA 98 strain of Salmonella typhimurium and BP with liver S-9 from male rats were markedly higher than those obtained using female rats . In addition, a marked strain difference (Wistar > DA) in the ability of liver S-9 from Wistar and DA rats to activate BP was observed . Antibody against cytochrome P-450 2D inhibited up to 50% of the revertant formation by the activation of BP with liver S-9 from male Wistar rats . These results indicate the partial involvement of cytochrome P-450 2D subfamily as well as cytochrome P-450 species specific to male rats in activation of BP to ultimate mutagen in untreated rat liver.

Mutagenesis, 1994 Nov, 9(6), 553 - 7
Activation of benzylic alcohols to mutagens by human hepatic sulphotransferases; Glatt H et al.; Four primary and five secondary benzylic alcohols derived from polycyclic aromatic hydrocarbons were tested for mutagenicity in Salmonella typhimurium TA98 in the presence of 3'-phosphoadenosine-5'-phosphosulphate, the cofactor for sulphotransferases, and varying amounts of hepatic cytosol from three or four different human subjects, a 3-year-old child, an adult female, an adult male and one unknown . All compounds except one, 4H-cyclopenta{def}phenanthren-4-ol, were activated to mutagens . The interindividual variation in the activities was at most 3-fold and the individual activities towards the different substrates were correlated with each other . The same compounds had previously been tested in the presence of hepatic cytosol from rats and all compounds activated in one species were also activated in the other species . However, there were marked quantitative differences, which were further complicated by the observation of a substantial sex difference in the rat . Male and female rat liver cytosol showed higher sulphotransferase activities towards 1-hydroxymethylpyrene, 9-hydroxymethylanthracene, 7-hydroxymethyl-12-methylbenz{a}anthracene and 4H-cyclopenta{def}chrysen-4-ol than human liver cytosol . The largest difference in activity was seen with 7-hydroxymethyl-12-methylbenz{a}anthracene, reaching a factor of approximately 100 between human and female rat . However, with other benzylic alcohols, the activity in human liver cytosol was in the range of that found in the less active sex of rat (3-hydroxy-3,4-dihydrocyclopenta{cd}pyrene, 2-hydroxymethylpyrene) or the more active sex of rat {1-(1-pyrenyl)ethanol}.(ABSTRACT TRUNCATED AT 250 WORDS)

Mutagenesis, 1994 Nov, 9(6), 547 - 51
9-Methoxytariacuripyrone, a naturally occurring nitro-aromatic compound with strong mutagenicity in Salmonella typhimurium; Schimmer O et al.; 9-Methoxytariacuripyrone, a nitro-aromatic compound isolated from Aristolochia brevipes showed strong mutagenic activity in strain TA98, TA100 and some YG strains of Salmonella typhimurium with and without S9 mix . Incubation with cytosol resulted in a heavy increase in mutagenicity . When incubated with microsomes the activity was dramatically decreased . The results are discussed in view of the enzymes possibly involved in activation and detoxification of the compound . The role of the basic structure on the mutagenicity mediated through the nitro group was also considered.

Mutagenesis, 1994 Nov, 9(6), 523 - 6
Moderate inhibition of mutagenicity and carcinogenicity of benzo{a}pyrene, 1,6-dinitropyrene and 3,9-dinitrofluoranthene by Chinese medicinal herbs; Horikawa K et al.; The activity of six Chinese medicinal herbs against the environmental mutagens and carcinogens benzo{a}pyrene (B{a}P), 1,6-dinitropyrene (1,6-diNP) and 3,9-dinitrofluoranthene (3,9-diNF) was determined . Samples of Prunella spica, Rheum palmatum, Polygonum multiflorum, Agrimonia pilosa, Ephedra sinica and Teitoutou were tested in an in vitro system . Antimutagenic activity against B{a}P was marked in the presence of extracts (boiled for 2 h in a water bath) whereas that against 1,6-diNP and 3,9-diNF varied from 20 to 86% . The differences in inhibition might be due to inactivation of metabolic enzymes . An extract of P . multiflorum was divided into ether, ethyl acetate and water soluble fractions, which were tested for antimutagenic activity against B{a}P . The antimutagenic action of the ethyl acetate soluble fraction was substantial and dose-dependent . Tannins and related compounds were the major components of the extract, of which epigallocatechin, epigallocatechin gallate, epicatechin gallate and tannic acid strongly inhibited the mutagenicity of B{a}P (2.5 micrograms/plate) in Salmonella typhimurium TA98 with S9 mix . To confirm the results of the in vitro test system, F344/DuCrj male rats were given a subcutaneous injection of B{a}P . Thereafter, they received water extracts of the six Chinese medicinal herbs for 50 weeks and were examined for tumors . The P . multiflorum extract significantly reduced the tumor incidence.

J Toxicol Sci, 1994 Nov, 19 Suppl 3, 487 - 97
{Mutagenicity studies of lactitol (NS-4)}; Iwakura K et al.; Lactitol (NS-4), a hepatic encephalopathy drug, was examined for mutagenicity in the reverse mutation test in bacteria, the chromosome aberration test with cultured mammalian cells, and the micronucleus test in mice . 1 . In the reverse mutation test using Salmonella typhimurium (TA1535, TA100, TA1537, and TA98) and Escherichia coli (WP2uvrA), the drug did not significantly increase revertant colonies in any of the test strains with or without metabolic activation system (S-9mix) . 2 . In the chromosome aberration test with cultured Chinese hamster lung cells (CHL/IU), the drug did not significantly increase aberrant cells in the direct method or in the metabolic activation method . 3 . In the micronucleus test with Slc:ddY male mice, the drug did not significantly increase micronucleated polychromatic erythrocytes in the bone marrows . These results suggest that lactitol has no mutagenicity in vitro or in vivo.

Lett Appl Microbiol, 1994 Nov, 19(5), 294 - 8
Evaluation of different DNA extraction procedures for the detection of Salmonella from chicken products by polymerase chain reaction; Soumet C et al.; Polymerase chain reaction (PCR) was used after a short pre-enrichment culture to detect Salmonella subspecies in chicken fillets . A direct PCR assay performed with chicken meat inoculated with Salmonella Typhimurium produced no PCR products . Six different DNA extraction protocols were tested to recover efficiently Salmonella DNA after a short incubation period . Three of them gave results that were reliable, rapid and sensitive . Successful protocols used Proteinase K and/or a centrifugation step to concentrate the samples . For reliable detection of Salmonella subspecies, a few thousand bacterial cells per ml must be present . To obtain this number of bacterial cells with an inoculation of about one cell in 25 g of ionized food products, it was necessary to incubate samples for at least 10 h before PCR . A larger inoculum of approximately 10 cells in 25 g of ionized food products, required 8 h in culture broth to give positive results by PCR-based assay.

Chem Res Toxicol, 1994 Nov-Dec, 7(6), 779 - 83
Interactive chlorine-by-bromine and hydrogen-by-hydroxyl group replacement effects in 2(5H)-furanone mutagenicity; LaLonde RT et al.; Both bromine- and chlorine-substituted 2(5H)-furanones are produced by the chlorination of ligno-humic waters containing bromide ion . The molar mutagenicities of four bromine- and chlorine-substituted 2(5H)-furanones were determined by the Salmonella typhimurium (TA100) assay to explore Cl-by-Br and H-by-OH replacement effects on mutagenicity . Each of these two replacements was expected to enhance mutagenicity based on earlier work showing that lower LUMO energy levels and greater radical anion stability correlated with elevated TA100 mutagenicity . The four compounds investigated were the following: 2,3-dibromo-5-hydroxy-2(5H)-furanone (mucobromic acid, MBA); 2,3-dibromo-2(5H)-furanone (reduced mucobraomic acid, RMBA); 2,3-dichloro-5-hydroxy-2(5H)-furanone (mucochloric acid, MCA); and 2,3-dichloro-2(5H)-furanone (reduced mucochloric acid, RMCA) . Mean molar mutagenicities were found to be 5.54, 1.18, 2.92 and 0.105 revertants/nmol for the four compounds in the order named . Mutagenicity enhancements resulting from Cl-by-Br and H-by-OH replacements were analyzed by simple ratios of mean molar mutagenicity and by multiple regression analysis . The effect of the Cl-by-Br replacement on mutagenicity amounted to a 1.9-fold enhancement in the presence of C-5 OH, but an 11-fold enhancement in the presence of C-5 H . This demonstrated that the two replacement effects were interactive . Higher mutagenicity values corresponded to lower AM1 computed LUMO energy levels and greater radical anion stabilities.

Mutat Res, 1994 Nov 1, 311(1), 9 - 20
Mutational spectrum of revertants in the hisD3052 allele of Salmonella typhimurium induced by hydrogen peroxide-activated benzidine; Wallace RE et al.; Benzidine is mutagenic in a modified Ames (Salmonella typhimurium) assay, which uses hydrogen peroxide-dependent peroxidative activation . The mutational specificity of benzidine was investigated in tester strain TA98, which reverts by frameshifts of the hisD3052 allele . The most frequently observed mutation is a deletion of two bases from a (CG)4 run . This deletion was elevated in frequency among benzidine-induced revertants, relative to spontaneous revertants . Many other mutations were also observed, including additions, deletions, and complex events . Only small frameshifts were observed among the benzidine-induced revertants, whereas some larger deletions were observed among the spontaneous revertants.

Mutat Res, 1994 Nov 1, 311(1), 149 - 56
Structure-mutagenicity relationships in series of 11H-indolo{3,2-c}quinoline-1,4-diones, tetrahydro-11H-indolo{3,2-c}quinoline-1,4-diones and 11H-pyrido{3',4':4,5}pyrrolo{3,2-c}quinoline-1,4-diones with leukemia cytotoxic properties . Relations with topoisomerase I inhibiting properties; Callais F et al.; Six heterocyclic quinones with topoisomerase I inhibiting properties and cytotoxic activities on L1210 leukemia cells were studied for their mutagenicity in four strains of Salmonella typhimurium . The tested compounds are 3-methoxyindolo{3,2-c}quinoline-1,4-diones and their derivatives in which the common pyrroloquinoline nucleus is annelated either with a benzene or a cyclohexane on a pyridine ring . Almost all quinones were found to be direct-acting mutagens at different levels in all strains, mainly TA97a and TA98 . Relations were established between their structure and their mutagenic activities . The mutagenicity was found to be influenced (i) by the nature of the fourth nucleus: the pyridinic compounds were the most active, the non-aromatic ones were practically inactive; (ii) by the presence of a methyl group in the 6-position that decreased the mutagenicity . Then, the mutagenic properties were compared with the topoisomerase I inhibiting property that is one of the possible mechanisms of action for these cytotoxic quinones . The results indicated a correlation between mutagenicity and enzyme inhibiting properties.

Mutat Res, 1994 Nov, 341(1), 71 - 5
Antimutagenicity of lemon grass (Cymbopogon citratus Stapf) to various known mutagens in salmonella mutation assay; Vinitketkumnuen U et al.; Lemon grass (Cymbopogon citratus Stapf) was extracted with 80% ethanol . The extract was not found to be mutagenic in the Salmonella mutation test with or without metabolic activation . However, the extract was found to possess antimutagenic properties towards chemical-induced mutation in Salmonella typhimurium strains TA98 and TA100 . Mutagenicity of AFB1, Trp-P-1, Trp-P-2, Glu-P-1, Glu-P-2, IQ, MNNG and AF-2, was inhibited by the extract of lemon grass in a dose-dependent manner, but no effect was found on the mutagenic activity of benzo{a}pyrene.

Mutat Res, 1994 Nov, 341(1), 1 - 15
Study of the genotoxic activity of five chlorinated propanones using the SOS chromotest, the Ames-fluctuation test and the newt micronucleus test; Le Curieux F et al.; Three short-term assays (the SOS chromotest, the Ames-fluctuation test and the newt micronucleus test) were carried out to evaluate the genotoxicity of five chlorinated propanones identified in several chlorinated waters (monochloropropanone, 1,1-dichloropropanone, 1,3-dichloropropanone, 1,1,1-trichloropropanone and 1,1,3-trichloropropanone) . In the SOS chromotest, all the compounds except monochloropropanone were found to induce primary DNA damage in Escherichia coli . With the fluctuation test, all five chloropropanones showed mutagenic activity on Salmonella typhimurium strain TA100 . The newt micronucleus assay detected a clastogenic effect on the peripheral blood erythrocytes of Pleurodeles waltl larvae only for 1,3-dichloropropanone and 1,1,3-trichloropropanone . Moreover, two structure-activity relationships are noticeable: (1) chloropropanones with chlorine substituents on both carbon positions (1,3-DCP and 1,1,3-TCP) are by far more genotoxic than chloropropanones substituted only on one carbon position (1,1-DCP and 1,1,1-TCP); (2) the increase of the number of chlorine substituents decreases the mutagenic activity (fluctuation test) of the chlorinated propanones studied.

Mutat Res, 1994 Nov, 325(2-3), 65 - 74
Genotoxicity of dithiocarbamates and their metabolites; Franekic J et al.; Dithiocarbamate fungicides are widely used in agriculture for protection of vegetable crops and seeds . The mutagenicity spectra of ziram, thiram, zineb S-65 and ETU were determined by employing a battery of test systems included the bacterium Salmonella typhimurium (strains TA98, TA100, TA102, TA104, TA1535, TA1538), the yeast Saccharomyces cerevisiae (strain D61.M) and the shallot Allium ascalonicum somatic cells . Plate incorporation assay with S . typhimurium demonstrated direct mutagenicity of ziram in TA100 and thiram in TA100 and TA98 whereas zineb S-65 and ETU were ineffective . Tests for mitotic chromosome malsegregation in S . cerevisiae D61.M gave positive results with thiram, zineb S-69 and ETU . In shallot somatic root-tip cells ziram, thiram and ETU induced different genetic damages e.g . mitotic disturbance, polyploidy and micronuclei.

Infect Immun, 1994 Nov, 62(11), 4969 - 74
Salmonella typhimurium invasion of epithelial cells: role of induced host cell tyrosine protein phosphorylation; Rosenshine I et al.; Salmonella typhimurium invades nonphagocytic epithelial and fibroblast cells via a process resembling phagocytosis . We have compared some phenotypes that are involved in S . typhimurium invasion by using different host cell lines, including HeLa, Henle-407, and A431 . Infection with either wild-type S . typhimurium, bacterial culture supernatant, or the noninvasive invA mutant was associated with induction of tyrosine phosphorylation of host cell mitogenic activating protein kinase . However, we did not detect induction of tyrosine phosphorylation of the epidermal growth factor receptor in S . typhimurium-infected cells . Treatment with the tyrosine protein kinase inhibitor genistein did not reduce S . typhimurium invasion into any of these cell lines . These results suggest that S . typhimurium invasion is independent of host cell epidermal growth factor receptor activation.

Biochem Biophys Res Commun, 1994 Oct 28, 204(2), 937 - 43
Hepatic metabolism of chloral hydrate to free radical(s) and induction of lipid peroxidation; Ni YC et al.; Metabolism of chloral hydrate by male B6C3F1 mouse liver microsomes generates free radical intermediate(s) as evidenced by electron spin resonance spectroscopic analysis . The subsequent induction of endogenous lipid peroxidation was shown by analysis of the resulting products with high-pressure liquid chromatography . Chloral hydrate was found mutagenic in Salmonella typhimurium strain TA104 . Both lipid peroxidation and mutagenicity were efficiently inhibited by free radical scavengers, alpha-tocopherol and menadione.

J Biol Chem, 1994 Oct 28, 269(43), 26591 - 3
Structural and kinetic analysis of a channel-impaired mutant of tryptophan synthase; Schlichting I et al.; Substrate channeling is a process by which two sequential enzymes interact to transfer a metabolite (or intermediate) from one enzyme active site to the next without allowing free diffusion of the metabolite . Channeling is thought to play an important role in metabolic regulation and cellular modulation of enzymatic activities . Although there are numerous examples of sequential enzyme pairs in glycolysis and other biosynthetic pathways that are thought to exhibit channeling, this is as yet controversial . Tryptophan synthase is considered the best example for an enzyme displaying channeling behavior between subunits . Tryptophan synthase is an alpha 2 beta 2 tetrameric enzyme complex, which catalyzes the last two steps in the biosynthesis of L-tryptophan . The alpha subunit catalyzes the cleavage of indole-3-glycerol phosphate to indole and glyceraldehyde-3-phosphate; the beta subunit catalyzes the condensation of indole with serine to form tryptophan, in a reaction mediated by pyridoxal phosphate . The inability to trap free indole in the steady-state reaction and analysis of the kinetics of single turnover reactions have led to the postulate that indole may pass directly from the alpha to the beta site without diffusion through solution (Demoss, J . A . (1962) Biochim . Biophys . Acta 62, 279-293; Matchett, W . M . (1974) J . Biol . Chem . 249, 4041-4049) . The crystal structure of tryptophan synthase from Salmonella typhimurium has provided additional support for substrate channeling by elucidating a 25-A hydrophobic tunnel connecting the two catalytic sites (Hyde, C . C., Ahmed, S . A., Padlan, E . A., Miles, E . W., and Davies, D . R . (1988) J . Biol . Chem . 263, 17857-17871) . The structure suggests that mutation of a residue lining the tunnel to a more bulky residue might impede or block the passage of indole during catalysis thus enabling detection of indole during a single enzyme turnover . A mutant of tryptophan synthase has been prepared in which one of the residues lining the tunnel, beta Cys-170, has been replaced with a bulkier tryptophan residue . Kinetic and structural analyses of the beta C170W mutant by rapid chemical quench methods and x-ray crystallographic analysis show both the transient formation of indole and the obstruction of the tunnel, thus providing direct evidence for the substrate channeling mechanism.

J Biol Chem, 1994 Oct 21, 269(42), 26503 - 11
The cobC gene of Salmonella typhimurium codes for a novel phosphatase involved in the assembly of the nucleotide loop of cobalamin; O'Toole GA et al.; We report the identification of a new locus, designated cobC, involved in the assembly of the nucleotide loop of cobalamin in Salmonella typhimurium . The cobC gene has been mapped, cloned, and sequenced . DNA sequence analysis suggested that cobC is divergently transcribed from the adjacent cobD gene and suggests that the regulatory region of these genes overlap . The cobC gene codes for a predicted polypeptide of 26 kDa with striking homology to phosphoglycerate mutase, fructose-2,6-bisphosphatase, and acid phosphatase enzymes . In vitro experiments demonstrated that CobC dephosphorylated the cobalamin biosynthetic intermediate N1-(5-phospho-alpha-D-ribosyl)-5,6-dimethylbenzimidazole to generate N1-alpha-D-ribosyl-5,6-dimethylbenzimidazole . In vivo data showed that the lack of cobC function blocks the synthesis of cobalamin from its precursors cobinamide and 5,6-dimethylbenzimidazole, i.e . it prevents the assembly of the nucleotide loop of cobalamin . Additionally, exogenous N1-alpha-D-ribosyl-5,6-dimethylbenzimidazole rescues the defect of a cobC mutant . We propose that cobC codes for a novel phosphatase whose primary role is in cobalamin biosynthesis . A model for the sequence of biosynthetic steps that assemble the nucleotide loop of cobalamin in S . typhimurium is presented.

J Biol Chem, 1994 Oct 21, 269(42), 26358 - 62
Signal transduction in chemotaxis . A propagating conformation change upon phosphorylation of CheY; Lowry DF et al.; The CheY protein from Escherichia coli and Salmonella typhimurium are among the best characterized proteins of the receiver domain family of two component signal transduction systems in bacteria . Phosphorylation of CheY plays a central role in bacterial chemotaxis . However, it is not entirely clear how its state of phosphorylation contributes to its function . Genetic evidence suggests that CheY changes its conformation upon phosphorylation . We present evidence for this conformation change by comparing the NMR 15N-1H correlation spectra of CheY.Mg2+ complex and phospho-CheY in the presence of magnesium . Large changes in chemical shift are used as indicators of chemical changes and probable structural changes in the protein backbone . Our observations suggest that significant structural changes occur in CheY upon phosphorylation and that these changes are distinct from the changes produced by magnesium ion binding . In addition to residues Asn-59 and Gly-65 that are immediately adjacent to the site of phosphorylation at Asp-57, a large number of other residues show significant chemical shift changes as a result of phosphorylation . These include Met-17, Val-21, Asn-23, Gly-39, Met-60, Met-63, Asp-64, Leu-66, Glu-67, Leu-68, Leu-69, Met-85, Val-86, Thr-87, Ala-88, Asn-94, Val-107, Lys-109, Thr-112, Ala-113, Ala-114, and Asn-121 . These results appear inconsistent with the recent suggestion that phosphorylation produces the same structural changes as magnesium binding (Bellsolell, L., Prieto, J., Serrano, L., and Coll, M . (1994) J . Mol . Biol . 238, 489-495) . We find that some regions change overlap with a genetically defined motor binding face . We therefore propose that the conformation switch modulates the interaction of CheY with its target, the flagellar motor . Other regions also change, possibly reflecting the many different functions of CheY homologues.

Commun Dis Rep CDR Rev, 1994 Oct 14, 4(11), R136 - 40
Associations between human and farm animal infections with Salmonella typhimurium DT104 in Herefordshire; Fone DL et al.; Reports of human infection with Salmonella typhimurium definitive type (DT) 104 have generated considerable interest . We undertook a descriptive study of infections with S . typhimurium DT 104 infection in humans and farm animals in Herefordshire between 1991 and 1993 . Laboratory reports of human salmonellosis, sent to the consultant in communicable disease control, were compared with cases identified using Statutory Incident Reports of salmonella in animals, birds and their products, received from the Ministry of Agriculture, Fisheries and Food . Six separate associations of infection between farming families and their livestock were identified . Nine out of 23 human cases, including three family outbreaks, were associated with animal infection . This study suggests that occupationally acquired infection in farmers and their families may be contributing to the national increase in cases, and shows the value of drawing together data from human and animal sources for the surveillance, investigation, and control of human infection with S . typhimurium DT104.

Commun Dis Rep CDR Rev, 1994 Oct 14, 4(11), R130 - 5
A case control study of infection with an epidemic strain of multiresistant Salmonella typhimurium DT104 in England and Wales; Wall PG et al.; Laboratory reports of a multiresistant strain of Salmonella typhimurium definitive type (DT) 104 rose in 1993; this led the Public Health Laboratory Service to investigate cases and identify possible risk factors for infection . Information derived from questionnaires, and details of previous isolations of S . typhimurium DT104 from food and animals, were used to design an unmatched case control study . Eighty-three cases whose isolations were of the same plasmid profile type (the 'epidemic strain') and 235 controls were included in the analysis . Illness was independently associated with the consumption of several food items and contact with animals, particularly ill farm animals . The number of isolations of this organism continues to rise, and control measures may include reducing infection in animals used for food, reducing the risk of contamination at all stages of the food chain, and raising awareness of measures to prevent food poisoning among food handlers and the general public.

Gene, 1994 Oct 11, 148(1), 173 - 4
The methyl viologen-resistance-encoding gene smvA of Salmonella typhimurium; Hongo E et al.; The complete nucleotide sequence of the gene encoding the Salmonella typhimurium methyl viologen-resistant protein, SmvA, similar to QacA (resistance to quaternary ammonium ion) of Staphylococcus aureus, and the surrounding sequences were determined . This indicated that the gene arrangement of S . typhimurium is different from that of Escherichia coli in this region.

EMBO J, 1994 Oct 3, 13(19), 4568 - 76
Genetic and molecular analyses of the interaction between the flagellum-specific sigma and anti-sigma factors in Salmonella typhimurium; Kutsukake K et al.; More than 50 genes are required for flagellar formation and function in Salmonella typhimurium . According to the cascade model of flagellar regulon, the flagellar operons are divided into three classes, 1, 2, and 3, with respect to transcriptional hierarchy . FliA is an alternative sigma factor specific for transcription of the class 3 operons, while FlgM is an anti-sigma factor which binds to FliA and prevents its association with RNA polymerase core enzyme . In the present study, we isolated a number of fliA mutants in which the altered FliA proteins become insensitive to inhibition by FlgM . Sequence analysis of their mutation sites revealed that most of them caused the amino acid substitutions in region 4 of the conserved amino acid sequences of sigma factors which lies near the C-terminal end of FliA . Using a set of fliA deletion mutants in a high-expression plasmid, we demonstrated that polypeptides containing the C-terminal portion of FliA could titrate the intracellular FlgM protein resulting in derepression of the class 3 operons . This result indicates that the C-terminal region of FliA contains the FlgM-binding domain . This was confirmed by a chemical cross-linking experiment with FlgM and truncated FliA proteins.

Chem Biol Interact, 1994 Oct, 93(1), 73 - 84
Mutagenic activity of halogenated propanes and propenes: effect of bromine and chlorine positioning; Lag M et al.; A series of halogenated propanes and propenes were studied for mutagenic effects in Salmonella typhimurium TA100 in the absence or presence of NADPH plus liver microsomes from phenobarbital-induced rats as an exogenous metabolism system . The cytotoxic and mutagenic effects of the halogenated propane 1,2-dibromo-3-chloropropane (DBCP) has previously been studied in our laboratories . These studies showed that metabolic activation of DBCP was required to exert its detrimental effects . All of the trihalogenated propane analogues were mutagenic when the microsomal activation system was included . The highest mutagenic activity was obtained with 1,2,3-tribromopropane, with approximately 50-fold higher activity than the least mutagenic trihalogenated propane, 1,2,3-trichloropropane . The order of mutagenicity was as follows: 1,2,3-tribromopropane > or = 1,2-dibromo- 3-chloropropane > 1,3-dibromo-2-chloropropane > or = 1,3-dichloro-2-bromopropane >> 1-bromo-2,3-dichloropropane > 1,2,3-trichloropropane . Compared to DBCP, the dihalogenated propanes were substantially less mutagenic . Only 1,2-dibromopropane was mutagenic and its mutagenic potential was approximately 1/30 of that of DBCP . In contrast to DBCP, 1,2-dibromopropane showed similar mutagenic activity with and without the addition of an activation system . The halogenated propenes 2,3-dibromopropene and 2-bromo-3-chloropropene were mutagenic to the bacteria both in the absence and presence of the activation system, whereas 2,3-dichloropropene did not show any mutagenic effect . The large differences in mutagenic potential between the various halogenated propanes and propenes are proposed to be due to the formation of different possible proximate and ultimate mutagenic metabolites resulting from the microsomal metabolism of the various halogenated propanes and propenes, and to differences in the rate of formation of the metabolites . Pathways are proposed for the formation of genotoxic metabolites of di- and trihalogenated propanes and dihalogenated propenes.

Appl Environ Microbiol, 1994 Oct, 60(10), 3901 - 2
Evaluation of alternariol and alternariol methyl ether for mutagenic activity in Salmonella typhimurium; Davis VM et al.; Alternariol and alternariol methyl ether were tested in the Ames Salmonella typhimurium assay, and both were shown, with and without metabolic activation, to be nonmutagenic to strains TA98 and TA100 . The finding of other investigators that alternariol methyl ether is weakly mutagenic to TA98 without metabolic activation could have resulted from the presence of a small amount of one of the highly mutagenic altertoxins in the alternariol methyl ether originally tested.

Ceska Slov Farm, 1994 Oct, 43(5), 240 - 2
{Mutagenic activity of copper phenoxyacetate complexes}; Blahova M et al.; The mutagenic activity of phenoxyacetatocopper (II) complexes of the composition Cu(R-COO)2Ln-R = phenoxymethyl, L represents H2O (n = 3), phenazone (1), 4-nitropyridine N-oxide (1)--as well free phenoxyacetic acid and uncoordinated 4-nitropyridine N-oxide was tested against strains of Salmonella typhimurium (TA 100, TA 97 and TA 102) with or without metabolic activation . Using Ames methods in vitro, of the above-mentioned compounds only the 4-nitropyridine N-oxide complex in a concentration of 75 micrograms/dish, with 27.7% content of this neutral ligand) and this molecule ligand alone (in a concentration of 12.5 micrograms/dish) exhibited genotoxic effects . The relationship between the coordination-chemical properties and the biological effects of the compounds tested is discussed.

Carcinogenesis, 1994 Oct, 15(10), 2375 - 7
5-Sulfooxymethylfurfural as a possible ultimate mutagenic and carcinogenic metabolite of the Maillard reaction product, 5-hydroxymethylfurfural; Surh YJ et al.; Heat treatment of foods containing reducing sugars and amino acids during cooking or sterilization triggers a sequence of non-enzymatic reactions collectively known as the Maillard reaction . 5-Hydroxymethylfurfural (HMF), one of the major intermediate products in the Maillard reaction, has been found to possess cytotoxic, genotoxic and tumorigenic activities, but the mechanisms of its toxic actions remain unclear . Formation of an electrophilic allylic ester bearing a good leaving group such as sulfate has been proposed as a possible metabolic activation pathway for HMF . In order to further test this possibility, we compared the mutagenic and carcinogenic activities of HMF and its chemically synthesized sulfuric acid ester, 5-sulfooxymethylfurfural (SMF) . SMF induced dose-dependent increases in the number of His+ revertants in Salmonella typhimurium TA100 . This intrinsic mutagenicity of SMF was significantly inhibited by ascorbic acid added to the assay media . In the presence of chloride ions, the bacterial mutagenicity of the highly polar sulfuric acid ester of HMF may also be mediated by formation of a lipophilic chloromethyl derivative . When topically applied to mouse skin, both sulfooxymethyl and chloromethyl derivatives of HMF exhibited higher skin tumor initiating activity than the parent hydroxymethyl compound . 5-Chloromethylfurfural was found to be a strong hepatocarcinogen in infant male B6C3F1 mice.

Carcinogenesis, 1994 Oct, 15(10), 2287 - 90
2,6-Dimethylaniline--hemoglobin adducts from lidocaine in humans; Bryant MS et al.; Lidocaine (xylocaine) is utilized for the treatment of ventricular arrhythmias which occur during cardiac surgery or myocardial infarction and as a local anesthetic . Recent data from the National Toxicology Program reported that a principal metabolite in man, 2,6-dimethylaniline, is carcinogenic in rats . In addition, the putative metabolite N-hydroxy-2,6-dimethylaniline has been reported to be mutagenic in Salmonella typhimurium TA100 . N-Hydroxy metabolites of aromatic amines may be oxidized by hemoglobin to the corresponding nitroso metabolites and the nitroso may covalently bind to cysteine groups in hemoglobin as the corresponding sulfinic acid amide . Since hemoglobin binding is an indirect measure of the formation of the N-hydroxy metabolite, we have examined the possibility that lidocaine or a metabolite may similarly covalently bind to hemoglobin in rats and humans . Using a previously developed gas chromatographic-mas spectrometric assay, hemoglobin adducts of 2,6-dimethylaniline were detected covalently bound to rat hemoglobin after administration of either 2,6-dimethylaniline or lidocaine . Consistent with previously reported observations, low levels of 2,6-dimethylaniline-hemoglobin adducts were also observed in human subjects before lidocaine administration . Following administration of lidocaine, all patients had much higher levels of 2,6-dimethylaniline-hemoglobin adducts . Differences in adduct levels in patients treated with lidocaine (70-3760 mg) ranged from 93 to 636 ng/g hemoglobin . These data indicate that N-hydroxy-2,6-dimethylaniline is a metabolite of lidocaine in man.

J Bacteriol, 1994 Oct, 176(20), 6324 - 33
The accumulation of glutamate is necessary for optimal growth of Salmonella typhimurium in media of high osmolality but not induction of the proU operon; Csonka LN et al.; Synthesis of glutamate can be limited in bacterial strains carrying mutations to loss of function of glutamate synthase (2-oxoglutarate:glutamine aminotransferase) by using low concentrations of NH4+ in the growth medium . By using such gltB/D mutant strains of Salmonella typhimurium, we demonstrated that: (i) a large glutamate pool, previously observed to correlate with growth at high external osmolality, is actually required for optimal growth under these conditions; (ii) the osmoprotectant glycine betaine (N,N,N-trimethylglycine) apparently cannot substitute for glutamate; and (iii) accumulation of glutamate is not necessary for high levels of induction of the proU operon in vivo . Expression of the proU operon, which encodes a transport system for the osmoprotectants proline and glycine betaine, is induced > 100-fold in the wild-type strain under conditions of high external osmolality . Ramirez et al . (R . M . Ramirez, W . S . Prince, E . Bremer, and M . Villarejo, Proc . Natl . Acad . Sci . USA 86:1153-1157, 1989) observed and we confirmed that in vitro expression of the lacZ gene from the wild-type proU promoter is stimulated by 0.2 to 0.3 M K glutamate . However, we observed a very similar stimulation for lacZ expressed from the lacUV5 promoter and from the proU promoter when an important negative regulatory element downstream of this promoter (the silencer) was deleted . Since the lacUV5 promoter is not osmotically regulated in vivo and osmotic regulation of the proU promoter is largely lost as a result of deletion of the silencer, we conclude that stimulation of proU expression by K glutamate in vitro is not a specific osmoregulatory response but probably a manifestation of the optimization of in vitro transcription-translation at high concentrations of this solute . Our in vitro and in vivo results demonstrate that glutamate is not an obligatory component of the transcriptional regulation of the proU operon.

J Bacteriol, 1994 Oct, 176(20), 6221 - 8
Involvement of Escherichia coli DNA polymerase II in response to oxidative damage and adaptive mutation; Escarceller M et al.; DNA polymerase II (Pol II) is regulated as part of the SOS response to DNA damage in Escherichia coli . We examined the participation of Pol II in the response to oxidative damage, adaptive mutation, and recombination . Cells lacking Pol II activity (polB delta 1 mutants) exhibited 5- to 10-fold-greater sensitivity to mode 1 killing by H2O2 compared with isogenic polB+ cells . Survival decreased by about 15-fold when polB mutants containing defective superoxide dismutase genes, sodA and sodB, were compared with polB+ sodA sodB mutants . Resistance to peroxide killing was restored following P1 transduction of polB cells to polB+ or by conjugation of polB cells with an F' plasmid carrying a copy of polB+ . The rate at which Lac+ mutations arose in Lac- cells subjected to selection for lactose utilization, a phenomenon known as adaptive mutation, was increased threefold in polB backgrounds and returned to wild-type rates when polB cells were transduced to polB+ . Following multiple passages of polB cells or prolonged starvation, a progressive loss of sensitivity to killing by peroxide was observed, suggesting that second-site suppressor mutations may be occurring with relatively high frequencies . The presence of suppressor mutations may account for the apparent lack of a mutant phenotype in earlier studies . A well-established polB strain, a dinA Mu d(Apr lac) fusion (GW1010), exhibited wild-type (Pol II+) sensitivity to killing by peroxide, consistent with the accumulation of second-site suppressor mutations . A high titer anti-Pol II polyclonal antibody was used to screen for the presence of Pol II in other bacteria and in the yeast Saccharomyces cerevisiae . Cross-reacting material was found in all gram-negative strains tested but was not detected in gram-positive strains or in S . cerevisiae . Induction of Pol II by nalidixic acid was observed in E . coli K-12, B, and C, in Shigella flexneri, and in Salmonella typhimurium.

Infect Immun, 1994 Oct, 62(10), 4641 - 5
Salmonella invasion of nonphagocytic cells induces formation of macropinosomes in the host cell; Garcia-del Portillo F et al.; Salmonella typhimurium induced massive uptake of extracellular fluid in epithelial cells in the form of macropinosomes . The appearance of macropinosomes in the infected cell was related to the induction of membrane ruffling during bacterial invasion . A noninvasive S . typhimurium invA mutant did not trigger such effects in the host cell . Similarly, S . typhimurium invA mutants that invaded via the invasin protein from Yersinia pseudotuberculosis or adhered to the host cell via the afimbrial AFA-I adhesin from Escherichia coli did not trigger formation of macropinosomes . In contrast to the formation of macropinosomes in macrophages, the appearance of macropinosomes in S . typhimurium-infected epithelial cells did not require microtubules . These data suggest that massive uptake of extracellular fluid in S . typhimurium-infected epithelial cells is an event related to the invasion mechanisms used by this pathogen.

Infect Immun, 1994 Oct, 62(10), 4310 - 9
Genetic control of antibody responses induced against an antigen delivered by recombinant attenuated Salmonella typhimurium; Fayolle C et al.; Recombinant derivatives of nonpathogenic bacteria such as attenuated Salmonella typhi have the potential to be used for delivery of heterologous antigens to the immune system . Genetic factors may modulate the immune responses to these live attenuated organisms and could therefore modify the immunogenicity of future human vaccines . In the present study, we compared the antibody responses of Ity or H-2 congenic strains of mice to a foreign antigen expressed by the murine attenuated aroA S . typhimurium strain . Our results demonstrate that the Ity gene may modulate the antibody responses to the foreign antigen but that the major genetic influence is exerted by H-2 genes, which control the capacity of mice to respond to the antigen expressed by recombinant attenuated Salmonella cells . This genetic control is related to differences in responsiveness of different strains of mice to low doses of antigen . Increasing the amount of foreign antigen expressed by recombinant Salmonella cells overcame the genetic restriction of these responses . These findings are potentially of great importance for the design of live vaccines for humans and show that care must be taken to optimize the amount of foreign antigen delivered to the immune system.

Gastroenterology, 1994 Oct, 107(4), 915 - 23
Monoclonal immunoglobulin A prevents adherence and invasion of polarized epithelial cell monolayers by Salmonella typhimurium; Michetti P et al.; BACKGROUND/AIMS: Invasion of the intestinal epithelium is considered a critical step in Salmonella pathogenesis . Infection by Salmonella of cultured monolayers of polarized Madin-Darby canine kidney (MDCK) cells has been established as a simple in vitro system that mimics the invasion of intestinal enterocytes in vivo . This study analyzes the protective role of secretory immunoglobulin (Ig) A antibodies against epithelial invasion . METHODS: Salmonella typhimurium was applied to MDCK cell monolayers in the presence or absence of a monoclonal, polymeric IgA antibody (Sal4) directed against an antigenic determinant exposed on the surface of wild-type S . typhimurium . RESULTS: In the presence of Sal4 IgA, confluent monolayers of MDCK cells were protected against apical invasion by wild-type S . typhimurium but not against a mutant strain that lacks the Sal4 epitope . Protection was Sal4-specific, dependent on the concentration of Sal4 in the apical medium, and occurred at IgA concentrations at which agglutination of IgA-bacterial complexes was observed . When MDCK cell monolayers were formaldehyde-fixed before incubation with Salmonella to prevent bacterial invasion, adhesion of Salmonella occurred in the absence of IgA and in the presence of control IgA but not in the presence of Sal4 IgA . CONCLUSIONS: IgA alone can prevent bacterial adherence and invasion of epithelial cells in the absence of other immune or nonimmune protective mechanisms.

Environ Health Perspect, 1994 Oct, 102 Suppl 6, 83 - 9
N-hydroxyarylamine O-acetyltransferase of Salmonella typhimurium: proposal for a common catalytic mechanism of arylamine acetyltransferase enzymes; Watanabe M et al.; Acetyl-CoA:N-hydroxyarylamine O-acetyltransferase is an enzyme involved in the metabolic activation of N-hydroxyarylamines derived from mutagenic and carcinogenic aromatic amines and nitroarenes . The O-acetyltransferase gene of Salmonella typhimurium has been cloned, and new Ames tester substrains highly sensitive to mutagenic aromatic amines and nitroarenes have been established in our laboratory . The nucleotide sequence of the O-acetyltransferase gene was determined . There was an open reading frame of 843 nucleotides coding for a protein with a calculated molecular weight of 32,177, which was close to the molecular weight of the O-acetyltransferase protein determined by using the maxicell technique . Only the residue of Cys69 in O-acetyltransferase of S . typhimurium and its corresponding residue (Cys68) in N-acetyltransferase of higher organisms were conserved in all acetyltransferase enzymes sequenced so far . The amino acid sequence Arg-Gly-Gly-X-Cys, including the Cys69, was highly conserved . A mutant O-acetyltransferase of S . typhimurium, which contained Ala69 instead of Cys69, no longer showed the activities of O- and N-acetyltransferase . These results suggest that the Cys69 of S . typhimurium and the corresponding cysteine residues of the higher organisms are essential for the enzyme activities as an acetyl-CoA binding site . We propose a new catalytic model of acetyltransferase for S . typhimurium and the higher organisms.

Environ Health Perspect, 1994 Oct, 102 Suppl 6, 195 - 200
Pathways for the mutagenesis of 1-nitropyrene and dinitropyrenes in the human hepatoma cell line HepG2; Silvers KJ et al.; The mutagenicity, metabolism, DNA adduction and induction of unscheduled DNA synthesis (UDS) of 1-nitropyrene and 1,8-dinitropyrene were investigated in the human hepatoma cell line HepG2 . Previous results had demonstrated that 1-nitropyrene was both mutagenic at the hgprt locus and induced UDS in these cells . In the present study, we find that the dinitropyrenes, although highly mutagenic in Salmonella typhimurium, are not mutagenic and do not induce UDS in the HepG2 . Although the rate of 1,8-dinitropyrene nitroreduction was less than that of 1-nitropyrene nitroreduction, this did not explain the lack of mutagenicity and UDS induction by the dinitropyrenes . Therefore, it is proposed that the arylhydroxylamine O-esterificase is not expressed in these cells . Since cytochrome P450-mediated C-oxidation is the predominant metabolic pathway in vivo, we sought to determine if an increase in the ratio of cytochrome P450-mediated C-oxidation over nitroreduction would result in increased or decreased DNA adducts in the HepG2 . The administration of 2.5 microM 3-methylcholanthrene to the HepG2 increased the ratio of C-oxidation/nitroreduction from 2.8 +/- 1.9 to 50.4 +/- 46.1 . This was accompanied by a decrease in the C8-guanyl adduct of 1-nitropyrene (via nitroreduction) from 18.7 +/- 7.0 to 4.8 +/- 1.7 fmoles/micrograms DNA, without any further increase in other 1-nitropyrene DNA adducts . These results suggest that the cytochrome P450-mediated metabolism of 1-nitropyrene to epoxides, phenols, and dihydrodiols is not an activation pathway in the HepG2 cells, and may explain the weak carcinogenicity of 1-nitropyrene in vivo, where cytochrome P450-mediated C-oxidation predominates.

FEMS Immunol Med Microbiol, 1994 Oct, 9(4), 325 - 32
Soluble plasma antigen in experimental Salmonella typhimurium infection in mice; Butler T et al.; To detect and characterize Salmonella antigen in blood, outbred CF-1 female mice were inoculated intraperitoneally with S . typhimurium LT-2 and blood was assayed by ELISA for Salmonella common structural antigen . Plasma antigen was detectable early in the course of infection and increased in quantity later in the course of illness when animals showed high grade bacteremia and high counts of splenic bacteria . Antigen was associated with a cell-free plasma fraction of blood, passed through filters with cut-offs of 0.2 mu and molecular mass of 1000 kDa, and was enhanced in detectability after heating to 100 degrees C for 15 min . Antigen was concentrated by diluting plasma 1:4 in 0.1 M EDTA, heating to 100 degrees C, and concentrating the supernate with an ultrafiltration membrane with a molecular mass cut-off of 15 kDa . By gel filtration, antigen was associated with a peak at about molecular mass 300 kDa in heated plasma and a peak at about 380 kDa in unheated plasma . These results indicate that murine typhoid infection results in circulating soluble plasma antigen, which is heat-stable with a molecular mass of approximately 300 kDa.

J Am Coll Nutr, 1994 Oct, 13(5), 424 - 8
Magnesium transport systems: genetics and protein structure (a review); Roof SK et al.; Magnesium is unique among biological cations . Its volume change from hydrated cation to atomic ion is over an order of magnitude larger than that of any other biological cation . This volume change presents particular problems for a magnesium transport system and suggests that these systems may be significantly different from other classes of ion transport systems . Detailed study of Mg2+ transport in complex organisms is limited by severe technical problems . However, molecular genetic techniques have enabled the isolation of three Mg2+ transport systems from the Gram-negative bacterium Salmonella typhimurium . The MgtA and MgtB transport systems are members of the P-type ATPase superfamily of transporters but possess unique characteristics among members of this family . The CorA transport protein is the first member of an entirely new class of transport proteins . In addition, another completely new family of Mg2+ transport proteins have been identified that is present in both Gram-negative and Gram-positive bacteria . Characterization of these transporters should provide substantial insight into Mg2+ transport and cellular Mg2+ homeostasis.

Immunology, 1994 Oct, 83(2), 245 - 9
Influence of mouse genotype and bacterial virulence in the generation of interferon-gamma-producing cells during the early phase of Salmonella typhimurium infection; Benbernou N et al.; Interferon-gamma (IFN-gamma) is known to play a major role in resistance to Salmonella typhimurium infection . In this study, the IFN-gamma production in spleens of mice infected with S . typhimurium was analysed at the single cell level using an ELISPOT method . The in vivo IFN-gamma production during the early phase of infection with virulent and avirulent S . typhimurium strains was examined in four mouse strains . Data show that infection with a virulent strain of S . typhimurium caused a much greater enhancement in the frequency of IFN-gamma-producing cells in innately resistant (ltyr) mice (CBA and DBA/2) than in susceptible (ltys) mice (C57BL/6 and BALB/c) . In contrast, infection with an avirulent strain of S . typhimurium induced a clear increase in the number of IFN-gamma-producing cells in susceptible mice which was even greater than in resistant ones . These results indicate that both the host genetic background and bacterial virulence play a critical role in the regulation of IFN-gamma production during the early phase of S . typhimurium infection.

J Toxicol Sci, 1994 Oct, 19 Suppl 2, 263 - 80
{Mutagenicity tests of tazobactam/piperacillin, tazobactam and piperacillin}; Ohuchida A et al.; As a part of safety tests of tazobactam/piperacillin (TAZ/PIPC), the reverse mutation tests using bacteria, the chromosomal aberration tests using cultured cells and the micronucleus tests using male mice were conducted in order to evaluate the in vitro and in vivo mutagenicity of TAZ, PIPC, TAZ/PIPC . 1 . The reverse mutation tests were carried out on TAZ, PIPC and TAZ/PIPC at dose ranges, where few antibacterial effects could be detected, using Salmonella typhimurium strains TA100, TA1535, TA98 and TA1537, and Escherichia coli WP2uvrA . All of three test articles showed that no significant increases were observed in the number of colonies in all tester strains in both systems, with and without mammalian metabolic activation (S9 Mix), as compared with solvent controls . 2 . The chromosomal aberration tests were carried out on these test articles using cultured Chinese hamster lung cells (CHL) . The cells were treated with TAZ, PIPC or TAZ/PIPC at the doses of 2.5, 5.0 and 10 mM with and without S9 Mix . In the test of PIPC with S9 Mix, the dose of 1.25 mM was set in addition to the three doses . The incidences of structural- and numeral-aberration were 0-3% in the absence or presence of mammalian metabolic activation system, and no significant increases were observed in the incidence of chromosomal aberrations as compared with solvent controls . 3 . The micronucleus tests were carried out at doses of 625-5000 mg/kg of TAZ or TAZ/PIPC, or at 625-2500 mg/kg of PIPC . The femoral marrow cells were 48 h after administering intravenously to CD-1 male mice . The frequencies of polychromatic erythrocyte with micronuclei were 0.02-0.17%, 0.02-0.10% and 0.03-0.07% in the groups treated with TAZ, PIPC and TAZ/PIPC, respectively, and no significant increases were observed with dose dependence . The results indicated that these test articles were negative in the assessment standard using the background data . 4 . The present study indicates that TAZ, PIPC and TAZ/PIPC have no in vitro and in vivo mutagenic potential.

Protein Expr Purif, 1994 Oct, 5(5), 518 - 26
Recombinant cholera toxin B subunit in Escherichia coli: high-level secretion, purification, and characterization; Slos P et al.; The gene coding for cholera toxin subunit B (CT-B) was fused to a modified ompA signal sequence and subsequently cloned into a high expression vector based on the regulatory signals of the arabinose operon of Salmonella typhimurium . Upon induction of gene expression in Escherichia coli, a product of the expected size for CT-B monomer was detected at a level of approximately 60% of total periplasmic protein . At pilot scale, batch cultivation in a 20-liter bioreactor allowed a production level of 1 g/liter of recombinant CT-B (rCT-B), the majority of which was released into the culture medium . The latter phenomenon was dependent on the medium selected for cultivation . A simple and inexpensive purification scheme was developed which enabled the recovery of 81% of rCT-B from the culture supernatant . Comparing amino acid composition, amino-terminal sequence, mass spectrum, pentamerisation, and GM1-binding, rCT-B is indistinguishable from natural CT-B produced by Vibrio cholerae . This rCT-B overproducing E . coli strain represents an interesting alternative to overexpressing systems developed in V . cholerae.

Poult Sci, 1994 Oct, 73(10), 1511 - 6
The effect of microaerosolized hydrogen peroxide on bacterial and viral poultry pathogens; Neighbor NK et al.; The effect of microaerosolized H2O2 on bacterial and viral poultry pathogens was investigated . Bacterial cultures and viruses were dried on sterile glass Petri dishes and subjected to direct and indirect 5% (H2O2) microaerosol mist . In the trials using Escherichia coli and Staphylococcus aureus, there was complete inactivation following exposure to H2O2 . Using Salmonella typhimurium, indirect exposure resulted in only partial inactivation whereas direct exposure to H2O2 gave complete inactivation . For the viruses studied, 5% H2O2 microaerosol mist completely inactivated infectious laryngotracheitis virus . Newcastle disease virus, infectious bronchitis virus, and avian influenza virus showed reduced infectivity but were not completely inactivated . Avian reovirus susceptibility varied with the method of exposure and infectious bursal disease virus was highly resistant . The use of 10% H2O2 mist, however, resulted in total inactivation of infectious bursal disease virus . The effect of 10% H2O2 on equipment and selected materials representative of a hatcher or poultry house was investigated . A solar cell calculator, a thermostat containing a microswitch, and samples of uncoated steel, galvanized steel, and uncoated aluminum were subjected to 10 fumigation cycles . No damage was detected in the calculator and the thermostat . Both the uncoated steel and the galvanized steel showed signs of oxidation . The aluminum did not show signs of oxidation.

Immunobiology, 1994 Oct, 191(4-5), 493 - 502
Macrophage nitric oxide mediates immunosuppression in infectious inflammation; Eisenstein TK et al.; A vaccine strain of live, attenuated Salmonella typhimurium induces profound immunosuppression in inoculated mice 7 days after injection . Immunosuppression to mitogens and inability to mount plaque-forming responses to sheep red blood cells occurs in spite of many parameters of upregulated macrophage function and protection against challenge with virulent Salmonella . Studies show that macrophage nitric oxide mediates the immunosuppression and presumably also the early-onset protective capacity of the vaccine . A model of "bystander lymphocyte autotoxicity" is presented to explain the mechanism of immunosuppression . The model proposes that Salmonella-activated macrophages generate nitric oxide which inactivates lymphocytes in the vicinity, so they become dysfunctional . Inhibition of nitric oxide by NG-monomethyl-L-arginine reverses immunosuppression . Evidence is presented that supports a relationship between the microbial burden in the spleen, the degree of nitric oxide produced, and the extent of immunosuppression . It is proposed that this model of microbial immunosuppression mediated by nitric oxide is generalizable for understanding immunosuppression and loss of delayed-type hypersensitivity induced by other microbes, such as Mycobacteria and measles virus . The model could account for anergy during mycobacterial infections, particularly when the burden of acid-fast bacilli is high, as well as loss of skin test reactivity to tuberculin during measles infection.

Singapore Med J, 1994 Oct, 35(5), 525 - 6
Fatal haemoptysis in Salmonella typhimurium septicaemia--a cautionary tale; Tan HC et al.; Septicaemia and mycotic aneurysm may occur in a patient who is an enteric-carrier of Salmonella typhimurium . We report such a case of an elderly man who presented with chest pain and fatal massive haemoptysis from a likely mycotic thoracic aneurysm . This report underscores the importance of increased awareness of the disease which may allow more frequent and earlier diagnosis.

Environ Health Perspect, 1994 Oct, 102 Suppl 6, 69 - 74
Cytosolic activation of aromatic and heterocyclic amines . Inhibition by dicoumarol and enhancement in viral hepatitis B; De Flora S et al.; The aromatic amines 2-aminofluorene (2AF), 2-acetylaminofluorene, and 2-aminoanthracene, and the heterocyclic amines 2-amino-3-methylimidazo{4,5-f}quinoline (IQ), 2-amino-3,4-dimethylimidazo{4,5-f}quinoline, and 3-amino-1-methyl-SH-pyrido{4,3-b}indole (Trp-P-2) were activated by rat liver cytosolic fractions to form mutagenic metabolites in Salmonella typhimurium strains TA98, TA98NR, and TA98/1,8-DNP6 . In the case of the Trp-P-2, the cytosolic activation was even more potent than the microsomal activation, which is classically ascribed to N-hydroxylation and subsequent esterification . The cytosolic activation was a) NADPH-dependent, b) induced by pretreatment of rats with 3-methylcholanthrene and especially Aroclor 1254 but not by phenobarbital, and c) inhibited by dicoumarol . The hypothesis is that, following a preliminary oxidative step in the cytosol (pure cytosolic activation) or in microsomes via prostaglandin H synthase (mixed microsomal-cytosolic activation), an oxidized intermediate of amino compounds may serve as substrate for DT diaphorase activity and bielectronically reduced to the corresponding N-hydroxyamino derivative . Purified DT diaphorase, in the presence of either NADPH or NADH as electron donor, produced mutagenic derivatives from IQ and Trp-P-2 . An NADPH-dependent activation of Trp-P-2 also occurred in the liver cytosol of woodchucks (Marmota monax), but was not inhibited by dicoumarol . As previously demonstrated with liver S-12 fractions in both humans and woodchucks, the cytosolic activation of Trp-P-2 was enhanced in animals affected by hepatitis B virus infection . This enhanced metabolism, which persisted even after appearance of primary hepatocellular carcinoma in virus carriers, is likely to be ascribed to mechanisms other than DT diaphorase induction, such as glutathione depletion.

Mutat Res, 1994 Oct, 322(4), 329 - 39
Ubiquitous presence of mutagenic and antimutagenic components in air-borne particulates of two Japanese cities; Iwado H et al.; Previous studies on several samples of urban air-borne particulates showed that the long-chain fatty acids present in these samples can interfere with the measurement of mutagenicity of the particulates with the Salmonella assay . To explore whether this phenomenon is a general, fatty acid contents and the mutagenicity (with Salmonella typhimurium TA98 without S9) were measured for 34 particulate samples collected in the cities of Okayama and Tokyo over a period of 1 year . Palmitic, stearic, oleic and linoleic acids were found in all these samples in this order of amount, and their interference on mutagenicity measurement was eminent, particularly at high doses of the sample . With the use of blue cotton extraction, the mutagenic components can be freed from most of these antimutagenic factors . Significant correlation was found between the number of particulates and the mutagenicity per unit volume of the air . Eight polycyclic aromatic hydrocarbon compounds, including benzo{alpha}pyrene were quantified for these 34 particulate samples . Their contents were too small to account for the observed mutagenicity, suggesting that other polycyclic compounds, possibly involving nitro aromatics, were responsible for the mutagenicity observed . No remarkable differences were noted between Okayama and Tokyo in fatty acid contents, mutagenicity or polycyclic aromatic-hydrocarbon contents of the samples.

Mutat Res, 1994 Oct, 322(4), 321 - 8
Genotoxicity of 2-halosubstituted enals and 2-chloroacrylonitrile in the Ames test and the SOS-chromotest; Eder E et al.; 2-Chloroacrolein and 2-bromoacrolein are very potent direct mutagens not requiring metabolic activation in Salmonella typhimurium strains TA 100 and TA 1535 . Mutagenic activities decrease with increasing degree of methyl substitution at carbon atom C-3 of the acrolein moiety from 2-chloroacrolein via 2-chlorocrotonaldehyde to 2-chloro-3,3-dimethylacrolein . With 2-chloroacrylonitrile equivocal results are obtained in strain TA 100 without S9-mix and unequivocal with S9-mix . In the SOS-chromotest the 2-chloroenals are also very strong genotoxins and the structure-activity relationships found in the Ames test are clearly confirmed . 2-Chloroacrylonitrile is not positive in the SOS-chromotest . The mutagenic mechanisms are discussed, and indications are provided that genotoxicity/mutagenicity depends on formation of DNA adducts, e.g., 1,N2-cyclic deoxyguanosine adducts.

Mutat Res, 1994 Oct, 322(4), 221 - 32
Mutagenicity of mono- and dinitropyrenes in the Salmonella typhimurium TM677 forward mutation assay; Busby WF Jr et al.; Nitropyrenes are a group of widespread environmental pollutants, some of which are highly potent as bacterial and mammalian cell mutagens and as animal carcinogens . A quantitative bacterial forward mutation assay, based on resistance to 8-azaguanine (8-AG) in Salmonella typhimurium TM677, was employed as an alternative to reversion assays to reexamine the mutagenicity of 1-, 2-, and 4-nitropyrene (1-, 2-, and 4-NP) and 1,3-, 1,6-, and 1,8-dinitropyrene (1,3-, 1,6-, and 1,8-DNP) in the presence and absence of rat liver postmitochondrial supernatant (PMS) . The major finding is that 2-NP, reported as a potent mutagen in the absence of PMS in bacterial reversion assays, was inactive in the absence of PMS in this assay . However, 2-NP was mutagenic in the presence of PMS . The implications of this observation with respect to sample purity and the metabolism of 2-NP are discussed . Without PMS the following minimum detectable mutagen concentration (MDMC) potency series expressed as nmol/ml was obtained: 1,8-DNP (0.5 x 10(-3)), 1,6-DNP (1.2 x 10(-3)), 1,3-DNP (2.3 x 10(-3)), 4-NP (0.2), 1-NP (0.2), 2-NP (> 1200), pyrene (> 1500) . With PMS the potency series was: 1,6-DNP (0.7), 1,8-DNP (2.1), 4-NP (2.2), 2-NP (2.6), 1,3-DNP (3.7), 1-NP (4.6), pyrene (> 1500) . With the exception of 2-NP, all the nitropyrenes were more mutagenic without PMS than with PMS . The greatest difference was observed with the dinitropyrenes, which were three orders of magnitude less potent in the presence of PMS . Pyrene, often reported as a bacterial mutagen in the presence of PMS, was nonmutagenic in this assay when a purified sample was tested.

Infect Immun, 1994 Oct, 62(10), 4542 - 8
Parameters that influence the efficiency of processing antigenic epitopes expressed in Salmonella typhimurium; Wick MJ et al.; We investigated parameters that affect the efficiency with which antigenic epitopes from Salmonella typhimurium are processed for presentation to T lymphocytes . As a model system, the hen egg white lysozyme 52-61 {HEL(52-61)} epitope, which binds the murine major histocompatibility complex class II (MHC-II) molecule I-Ak, was expressed in soluble fusion proteins in S . typhimurium . Murine peritoneal macrophages mediated phagocytic processing of viable S . typhimurium expressing fusion proteins of the HEL epitope for presentation via I-Ak regardless of the bacterial compartment in which the epitope was contained (i.e., surface exposed, facing the periplasmic space, or in the cytoplasm) . Minor differences in processing efficiency observed with different epitope compartmentalizations could be overcome by altering the relative expression level, indicating that epitope abundance is an important factor for efficient processing of epitopes from S . typhimurium . This processing pathway required phagocytosis of bacteria followed by passage through an acidic compartment, suggesting a pathway involving phagolysosomal degradation of the bacteria to liberate epitopes that bind MHC-II . HEL(52-61) was processed more efficiently from heat-killed S . typhimurium than from viable bacteria, and in addition, the HEL epitope was processed more efficiently from a rough lipopolysaccharide (LPS) strain than from its isogenic smooth LPS counterpart, most likely because of enhanced phagocytosis of the rough LPS strain . These data suggest that the efficiency of epitope processing from S . typhimurium for presentation via MHC-II is affected by bacterial viability, epitope abundance, and LPS phenotype, factors which may be important to consider in development of recombinant S . typhimurium vaccine strains.

Biochemistry, 1994 Sep 20, 33(37), 11184 - 8
Interactions of nucleotides with fully unadenylylated glutamine synthetase from Salmonella typhimurium; Liaw SH et al.; Glutamine synthetase (GS) catalyzes the ATP-dependent biosynthesis of glutamine from glutamate and ammonia in the presence of divalent cations . To gain insight into the structural basis of the feedback inhibition of GS by AMP, we have studied crystal structures of GS complexes with AMP and the related molecules: AMPPNP (a less hydrolyzable ATP analog), ADP, GDP, adenosine, and adenine . AMP is a feedback inhibitor of GS; ATP and ADP are cofactors, and AMPPNP, GDP, adenosine, and adenine are also GS inhibitors . GS used in this study is from Salmonella typhimurium and is free of covalent modification by adenylylation . All of the crystals examined contain two bound MN2+ ions per GS subunit . The X-ray structures show that all nucleotides bind at the same site, the cofactor ATP binding site, as do adenosine and adenine . Thus from X-ray structures, AMP, adenosine, adenine, and GDP would be expected to inhibit GS-Mn by competing with the substrate ATP for the active site . This suggestion from the crystal structures that AMP is competitive with respect to ATP is supported by kinetic measurements using the biosynthetic assay.

J Biol Chem, 1994 Sep 16, 269(37), 23051 - 8
The histidine-binding protein undergoes conformational changes in the absence of ligand as analyzed with conformation-specific monoclonal antibodies; Wolf A et al.; The periplasmic histidine-binding protein, HisJ, and the lysine-, arginine-, ornithine-binding protein (LAO) are receptors for histidine transport via the histidine permease of Salmonella typhimurium . The receptors have similar structures, being composed of two lobes held together by two peptide segments, with the ligand-binding site located in a cleft between the lobes . The two lobes are far apart in the unliganded structure (open conformation) and are drawn close together in the liganded structure (closed conformation) . The tight binding of the ligand via protein side chains as well as the peptide backbone from both lobes stabilizes the closed conformation (Oh, B.-H., Pandit, J., Kang, C.-H., Nikaido, K., Gokcen, S., Ames, G . F.-L., and Kim, S.-H . (1993) J . Biol . Chem . 268, 11348-11353; Oh, B.-H., Kang, C.-H., De Bondt, H., Kim, S.-H., Nikaido, K., Joshi, A., and Ames, G . F.-L . (1994) J . Biol . Chem . 269, 4135-4143) . In this study two conformation-specific monoclonal antibodies (mAbs) that trap the protein in the closed empty form have been characterized and used to provide evidence that HisJ can assume the closed empty form in the absence of ligand . Several pieces of evidence were provided to demonstrate that these mAbs are specific for HisJ in the closed form . Histidine improves the interaction of these mAbs with immobilized HisJ . The mAbs inhibit both the exchange and the dissociation of histidine from HisJ, indicating that they are able to trap the protein in the closed liganded form . The characterization of the epitopes of the conformation-specific mAbs shows that they include residues that are located in both lobes and that are far apart in the open form but close to each other in the closed form, so that the mAbs must be sensitive to their spatial orientation . Two mAbs that are not conformation-specific according to these criteria were also identified.

Biochim Biophys Acta, 1994 Sep 13, 1219(1), 198 - 200
Cloning and sequencing of the gene encoding the RpoS (KatF) sigma factor from Salmonella typhimurium 14028s; Prince RW et al.; The gene encoding the alternative sigma factor RpoS in Salmonella typhimurium was cloned by its ability to complement acid susceptibility in rpoS mutant Escherichia coli . Sequence determination and comparison with rpoS from E . coli demonstrates a high degree of conservation, although significant differences are found within the extragenic regulatory regions.

J Bacteriol, 1994 Sep, 176(18), 5729 - 34
High-resolution restriction map for a 240-kilobase region spanning 91 to 96 minutes on the Salmonella typhimurium LT2 chromosome; Wong KK et al.; A hierarchical approach allows the completion of contiguous sets of overlapping clones for small regions of a genome, one at a time rather than tackling the whole genome at once . On the basis of the BlnI restriction map for Salmonella typhimurium LT2, we dissected the chromosome into 21 different fragments by using a Tn5 transposon carrying a BlnI site . Dissected chromosomal fragments were purified by pulsed-field gel electrophoresis and used as probes for sorting a lambda DASHII genomic library of 2,304 primary clones . A total of 129 clones identified as spanning the region from 91 min to 98 min were partly ordered on the basis of the intensity of hybridization with mitomycin-induced Mud-P22 phage DNAs from insertions with pac sites in opposite orientations at 93 min used as probes . Decreased signal intensity with the Mud-P22 probes corresponded to the increased distance of the clone from the site of Mud-P22 insertion and allowed the clones to be placed in two groups from 91 min to 93 min and from 93 min to 98 min and into four intensity categories within the two groups . A member of each category was used to generate a riboprobe from the T3 promoter flanking the insert . This probe identified overlapping clones among the 129 clones . This subchromosomal library was then screened again with riboprobes from nonoverlapping clones . After four cycles of this strategy, a minimal contiguous sequence of 19 partly overlapping clones was selected for restriction mapping . A detailed map of 378 sites for eight restriction enzymes is presented for a region of about 240 kb . Working clockwise, the following genes were placed on this physical map on the basis of their restriction maps: malFEK, lamB, malM, lexA, qor, dnaB, alr, uvrA, proP, pmrB, pmrA, melA, melB, phoN, amiB, mutL, and miaA.

EMBO J, 1994 Sep 1, 13(17), 3964 - 72
A Salmonella protein that is required for resistance to antimicrobial peptides and transport of potassium; Parra-Lopez C et al.; The ability of invading pathogens to proliferate within host tissues requires the capacity to resist the killing effects of a wide variety of host defense molecules . sap mutants of the facultative intracellular parasite Salmonella typhimurium exhibit hypersensitivity to antimicrobial peptides, cannot survive within macrophages in vitro and are attenuated for mouse virulence in vivo . We conducted a molecular genetic analysis of the sapG locus and showed that it encodes a product that is 99% identical to the NAD+ binding protein TrkA, a component of a low-affinity K+ uptake system in Escherichia coli . SapG exhibits similarity with other E . coli proteins implicated in K+ transport including KefC, a glutathione-regulated efflux protein, and Kch, a putative transporter similar to eukaryotic K+ channel proteins, sapG mutants were killed by the antimicrobial peptide protamine in the presence of both high and low K+, indicating that protamine hypersensitivity is not due to K+ starvation . Strains with mutations in sapG and either sapJ or the sapABCDF operon were as susceptible as sapG single mutants, suggesting that the proteins encoded by these loci participate in the same resistance pathway . SapG may modulate the activities of SapABCDF and SapJ to mediate the transport of peptides and potassium.

J Bacteriol, 1994 Sep, 176(17), 5547 - 9
Arsenate arrests flagellar rotation in cytoplasm-free envelopes of bacteria; Margolin Y et al.; The effect of arsenate on flagellar rotation in cytoplasm-free flagellated envelopes of Escherichia coli and Salmonella typhimurium was investigated . Flagellar rotation ceased as soon as the envelopes were exposed to arsenate . Inclusion of phosphate intracellularly (but not extracellular) prevented the inhibition by arsenate . In a parallel experiment, the rotation was not affected by inclusion of an ATP trap (hexokinase and glucose) within the envelopes . It is concluded that arsenate affects the motor in a way other than reversible deenergization . This may be an irreversible damage to the cell or direct inhibition of the motor by arsenate . The latter possibility suggests that a process of phosphorylation or phosphate binding is involved in the motor function.

J Bacteriol, 1994 Sep, 176(17), 5474 - 82
The control region of the pdu/cob regulon in Salmonella typhimurium; Chen P et al.; The pdu operon encodes proteins for the catabolism of 1,2-propanediol; the nearby cob operon encodes enzymes for the biosynthesis of adenosyl-cobalamin (vitamin B12), a cofactor required for the use of propanediol . These operons are transcribed divergently from distinct promoters separated by several kilobases . The regulation of the two operons is tightly integrated in that both require the positive activator protein PocR and both are subject to global control by the Crp and ArcA proteins . We have determined the DNA nucleotide sequences of the promoter-proximal portion of the pdu operon and the region between the pdu and cob operons . Four open reading frames have been identified, pduB, pduA, pduF, and pocR . The pduA and pduB genes are the first two genes of the pdu operon (transcribed clockwise) . The pduA gene encodes a hydrophobic protein with 56% amino acid identity to a 10.9-kDa protein which serves as a component of the carboxysomes of several photosynthetic bacteria . The pduF gene encodes a hydrophobic protein with a strong similarity to the GlpF protein of Escherichia coli, which facilitates the diffusion of glycerol . The N-terminal end of the PduF protein includes a motif for a membrane lipoprotein-lipid attachment site as well as a motif characteristic of the MIP (major intrinsic protein) family of transmembrane channel proteins . We presume that the PduF protein facilitates the diffusion of propanediol . The pocR gene encodes the positive regulatory protein of the cob and pdu operons and shares the helix-turn-helix DNA binding motif of the AraC family of regulatory proteins . The mutations cobR4 and cobR58 cause constitutive, pocR-independent expression of the cob operon under both aerobic and anaerobic conditions . Evidence that each mutation is a deletion creating a new promoter near the normal promoter site of the cob operon is presented.

J Bacteriol, 1994 Sep, 176(17), 5439 - 49
Roles of FliK and FlhB in determination of flagellar hook length in Salmonella typhimurium; Hirano T et al.; The length of flagellar hooks isolated from wild-type and mutant cells with various hook lengths were measured on electron micrographs . The length of the wild-type hook showed a narrow distribution with a peak (+/- standard deviation) at 55.0 +/- 5.9 nm, whereas fliK mutants (so-called polyhook mutants) showed a broad distribution of hook lengths ranging from 40 to 900 nm, strongly indicating that FliK is involved in hook length determination . Among pseudorevertants isolated from such polyhook mutants, fliK intragenic suppressors gave rise to polyhook filaments . However, intergenic suppressors mapping to flhB also gave rise to hooks of abnormal length, albeit they were much shorter than polyhooks . Furthermore, double mutations of flhB and flgK (the structural gene for hook-associated protein 1; HAP1) resulted in polyhooks, suggesting another way in which hook length can be affected . The roles of FliK, FlhB, and HAP1 in hook length determination are discussed.

Infect Immun, 1994 Sep, 62(9), 3984 - 93
Characterization of defined ompR mutants of Salmonella typhi: ompR is involved in the regulation of Vi polysaccharide expression; Pickard D et al.; The ompB operon, comprising the ompR and envZ genes, was cloned from a Salmonella typhi Ty2 cosmid bank and characterized by DNA sequence analysis . The S . typhi ompR and envZ genes contained open reading frames encoding proteins of 240 and 451 amino acids, respectively . Comparison with the Salmonella typhimurium OmpB protein sequences revealed 99.5% homology . The DNA sequence data were used to identify appropriate restriction sites for generating a defined deletion of 517 bp within the open reading frame of the ompR gene . This deletion was introduced by homologous recombination into the chromosomes of two S . typhi strains which already harbored defined deletions in both the aroC and aroD genes . The presence of the deletions within ompR was confirmed by Southern hybridization and sequencing of the DNA fragments surrounding the deleted regions by PCR . The S . typhi ompR mutants displayed a marked decrease in OmpC and OmpF porin expression as demonstrated by examination of outer membrane preparations . It was also found that S . typhi strains harboring the defined ompR deletions no longer agglutinated with Vi antiserum . However, when a functional ompB operon was introduced back into the S . typhi ompR mutants, either on a multicopy plasmid or as a single-copy chromosomal replacement, the Vi+ phenotype was restored . The levels of Vi synthesis were also found to be sensitive to different concentrations of sodium chloride present in the growth medium, although the levels of sensitivity varied between different isolates of S . typhi . It is therefore concluded that the ompR-envZ two component regulatory system plays an important role in the regulation of Vi polysaccharide synthesis in S . typhi and that one of the environmental signals for this regulation may be osmolarity.

Infect Immun, 1994 Sep, 62(9), 3947 - 56
Binding of Legionella pneumophila to macrophages increases cellular cytokine mRNA; Yamamoto Y et al.; Infection of macrophages with Legionella pneumophila induces formation of interleukin 1 beta (IL-1 beta), but the molecular basis of this is not understood . Binding of bacteria to macrophage surfaces is the first step in an infection process . Therefore, we examined whether this step was sufficient to increase the cellular level of mRNAs for IL-1 beta and other cytokines . To assess the effect of binding of L . pneumophila on the steady-state levels of cytokine mRNAs, cultures of thioglycolate-elicited macrophages from L . pneumophila-susceptible A/J mice were treated with cytochalasin D and infected with L . pneumophila and the total RNA was extracted for analysis by reverse transcription-PCR with primers for IL-1 alpha, IL-1 beta, IL-6, tumor necrosis factor alpha, granulocyte macrophage colony-stimulating factor, and beta interferon (IFN-beta) . L . pneumophila treatment increased the cellular steady-state mRNA levels of all cytokines except IFN-beta . To determine the specificity of this effect, macrophage cultures were treated with cytochalasin D and either bacterial lipopolysaccharide, bovine serum albumin-sensitized latex, Salmonella typhimurium, or Escherichia coli . Lipopolysaccharide treatment increased all mRNAs, bovine serum albumin-sensitized latex had no significant effect, and treatment with S . typhimurium or E . coli increased all mRNAs except that of IFN-beta . These results suggested that the binding of gram-negative bacteria to the macrophage surface was sufficient to induce a unique pattern of cytokine mRNAs . Additional studies that examined the characteristics of the bacterial ligands involved indicated involvement of both heat-labile and heat-stable surface ligands.

Infect Immun, 1994 Sep, 62(9), 3745 - 52
Identification and characterization of a Salmonella typhimurium oxygen-regulated gene required for bacterial internalization; Jones BD et al.; Growth of Salmonella typhimurium in a low-oxygen environment induces the ability of these bacteria to enter mammalian cells . We have carried out a search for invasion genes that are expressed under low-oxygen conditions by using Tn5lacZY transcriptional fusions . Several noninvasive oxygen-regulated lacZY insertion strains have been identified . The invasion defect in one of these noninvasive S . typhimurium strains, BJ66, has been complemented by introduction of a cosmid (pBDJ125) from an S . typhimurium SL1344 gene bank . A 1.9-kb EcoRV DNA fragment subcloned from this cosmid, containing a single open reading frame (orgA), restores the ability of BJ66 to invade mammalian cells . Comparative searches of the GenBank and EMBL sequence data banks with the nucleotide sequence of the gene and deduced amino acid sequence of the protein reveal no significant similarities . Interestingly, hybridization of an orgA gene probe with a P22 chromosomal mapping library demonstrated that the orgA gene maps to a region on the chromosome between 57.5 and 60 min where other Salmonella invasion genes have been mapped . Other enteroinvasive bacteria (Shigella flexneri, Escherichia coli, Yersinia spp., and Listeria monocytogenes) lack sequences which cross hybridize to the probe . We have compared the virulence of S . typhimurium SL1344 and an isogenic orgA mutant in a mouse model of typhoid fever . The orgA mutant was as virulent as the wild-type strain was when inoculated intraperitoneally but is significantly reduced (> 60-fold) in its ability to cause disease by an oral route of infection.

Genetics, 1994 Sep, 138(1), 227 - 34
Selection intensity for codon bias; Hartl DL et al.; The patterns of nonrandom usage of synonymous codons (codon bias) in enteric bacteria were analyzed . Poisson random field (PRF) theory was used to derive the expected distribution of frequencies of nucleotides differing from the ancestral state at aligned sites in a set of DNA sequences . This distribution was applied to synonymous nucleotide polymorphisms and amino acid polymorphisms in the gnd and putP genes of Escherichia coli . For the gnd gene, the average intensity of selection against disfavored synonymous codons was estimated as approximately 7.3 x 10(-9); this value is significantly smaller than the estimated selection intensity against selectively disfavored amino acids in observed polymorphisms (2.0 x 10(-8)), but it is approximately of the same order of magnitude . The selection coefficients for optimal synonymous codons estimated from PRF theory were consistent with independent estimates based on codon usage for threonine and glycine . Across 118 genes in E . coli and Salmonella typhimurium, the distribution of estimated selection coefficients, expressed as multiples of the effective population size, has a mean and standard deviation of 0.5 +/- 0.4 . No significant differences were found in the degree of codon bias between conserved positions and replacement positions, suggesting that translational misincorporation is not an important selective constraint among synonymous polymorphic codons in enteric bacteria . However, across the first 100 codons of the genes, conserved amino acids with identical codons have significantly greater codon bias than that of either synonymous or nonidentical codons, suggesting that there are unique selective constraints, perhaps including mRNA secondary structures, in this part of the coding region.

Genetics, 1994 Sep, 138(1), 11 - 28
Suppressors of recB mutations in Salmonella typhimurium; Benson NR et al.; Using a screen that directly assesses transductional proficiency, we have isolated suppressors of recB mutations in Salmonella typhimurium . The alleles of sbcB reported here are phenotypically distinct from those isolated in Escherichia coli in that they restore recombination proficiency (Rec+), resistance to ultraviolet light (UVR), and mitomycin C resistance (MCR) in the absence of an accompanying sbcCD mutation . In addition the sbcB alleles reported here are co-dominant to sbcB+ . We have also isolated insertion and deletion mutants of the sbcB locus . These null mutations suppress only the UVS phenotype of recB mutants . We have also isolated sbcCD mutations, which map near proC . These sbcCD mutations increase the viability, recombination proficiency and MCR of both the transductional recombination suppressors (sbcB1 & sbcB6) and the sbcB null mutations . S . typhimurium recB sbcB1 sbcCD8 strains are 15-fold more recombination proficient than wild-type strains . The increase in transductants in these strains is accompanied by a loss of abortive transductants suggesting that these fragments are accessible to the mutant recombination apparatus . Using tandem duplications, we have constructed sbcB merodiploids and found that, in a recB mutant sbcCD+ genetic background, the sbcB+ allele is dominant to sbcB1 for transductional recombination but co-dominant for UVR and MCR . However, in a recB sbcCD8 genetic background, the sbcB1 mutation is co-dominant to sbcB+ for all phenotypes . Our results lead us to suggest that the SbcB and SbcCD proteins have roles in RecBCD-dependent recombination.

Free Radic Biol Med, 1994 Sep, 17(3), 273 - 7
Biphenyl compounds are hydroxyl radical scavengers: their effective inhibition for UV-induced mutation in Salmonella typhimurium TA102; Fujita S et al.; In our previous study, we found several hydroxylated biphenyl compounds have a great scavenging effect for hydroxyl radicals (.OH) . In this study, to elucidate the relationship between generation of the .OH and photo-mutagenesis, six biphenyl compounds such as dehydrodieugenol, dehydrodivanillyl alcohol, dehydrodidihydroeugenol, dehydrodicreosol, magnolol and honokiol, respectively, were examined for their ability to inhibit UV-induced mutation in Salmonella typhimurium TA102 . The relative mutagenic activities (RMA, %) indicated the mutation frequency of treated cells divided by the mutation frequency of control cells times 100% . The RMA (%) are as follows: 28 +/- 1, 31 +/- 1, 33 +/- 1, 41 +/- 2, 62 +/- 1, and 62 +/- 4 at concentrations of 5 micrograms per plate for dehydrodieugenol, dehydrodivanillyl alcohol, dehydrodidihydroeugenol, dehydrodicreosol, magnolol, and honokiol, respectively . These values indicate that low concentrations of these biphenyl compounds effectively suppress UV-induced mutagenesis . Also, these compounds acted as effective antimutagens in a dose-dependent manner (0.00005-5 micrograms per plate) . These compounds are effective .OH scavengers . Consequently, the results obtained above suggest that these compounds could inhibit against UV-induced mutations by scavenging of .OH generated by UV irradiation . The results also suggest that .OH are associated with UV-induced mutation in Salmonella typhimurium TA102.

APMIS, 1994 Sep, 102(9), 703 - 10
Infection with human cytomegalovirus enhances bacterial adhesiveness and invasiveness in permissive and semipermissive cells; Holberg-Petersen M et al.; The effect of human cytomegalovirus (HCMV) infection on adhesiveness and invasiveness of Salmonella typhimurium was examined in cells permissive (human embryo fibroblasts (HE)), semipermissive (A549) and nonpermissive (HEp-2) for the virus . Preinfection of the cells with HCMV induced enhanced adhesiveness and invasiveness of bacteria in the permissive HE cells . In the semipermissive A549 cells, where HCMV immediate-early (IE) mRNA transcripts and IE proteins were detected, a significant effect on the initial phase of invasiveness, the adherence phase, was demonstrated . HCMV had no effect on invasiveness of S . typhimurium in nonpermissive HEp-2 cells . Neither HCMV IE transcripts nor IE proteins could be detected in these cells.

J Commun Dis, 1994 Sep, 26(3), 147 - 50
Antibacterial activity of tea (Camellia sinensis) and coffee (Coffee arabica) with special reference to Salmonella typhimurium; Shetty M et al.; Extracts of Black tea, Japanese green tea, China tea or Coffee inhibited the growth of various bacteria causing diarrhoeal diseases . Tea or coffee also showed bactericidal activity against Vibrio cholerae, Salmonella typhimurium and Salmonella typhi.

Res Microbiol, 1994 Sep, 145(7), 543 - 52
Preferential interaction of Salmonella typhimurium with mouse Peyer's patch M cells; Clark MA et al.; We have used a mouse Peyer's patch gut loop model to investigate the role of the intestinal membranous epithelial (M) cells in the pathogenesis of Salmonella typhimurium . These specialized antigen sampling cells are located in the follicle-associated epithelium (FAE) overlying the isolated and aggregated lymphoid follicles in the small and large intestines . Our studies have demonstrated that S . typhimurium adheres more frequently to the Peyer's patch FAE cells than to the villous enterocytes and that, within the FAE, this bacterium preferentially interacts with the M cells . Quantitative light microscopic studies, using the lectin Ulex europaeus 1 (UEA1) to identify M cells, revealed that 34-fold more bacteria bound per unit area of M cells than per unit area of enterocyte . Within a 30-min incubation period, some M cells had clearly been invaded by the Salmonella . We therefore propose that M cells are a major route by which S . typhimurium penetrates the intestinal epithelial barrier . Bacterial adhesion to M cells occurred in a non-uniform pattern, suggesting the existence of M-cell subtypes . The interaction of S . typhimurium with mouse Peyer's patch M cells was accompanied by membrane ruffle formation and polymerized actin redistribution similar to that observed in cultured cell lines infected by this bacterium . This study emphasizes the suitability of Salmonella as an oral vaccine delivery system since, by preferentially interacting with the M cells, these bacteria are targeted to sites where cells of the immune system are concentrated.

Anticancer Res, 1994 Sep-Oct, 14(5A), 1775 - 8
A correlative approach for the identification of antimutagens that demonstrate chemopreventive activity; Shamon LA et al.; Seventy natural and synthetic compounds were tested for potential to inhibit mutation induced by 7,12-dimethylbenz(a)anthracene (DMBA) in Salmonella typhimurium strain TM677 . Results were compared with their ability to inhibit DMBA-induced preneoplastic lesions in a mouse mammary gland organ culture system . The response mediated by fifty-five of the test compounds was either positive or negative in both test systems, indicating that the combined use of these assays should aid in the discovery of antimutagenic agents that have cancer chemopreventive potential.

Mutagenesis, 1994 Sep, 9(5), 483 - 8
Antimutagenicity of Maillard reaction products from amino acid/sugar model systems against 2-amino-3-methylimidazo-{4,5-f}quinoline: the role of pyrazines; Jenq SN et al.; The antimutagenicity of dichloromethane extracts from eight amino acid/sugar model systems was determined using Salmonella typhimurium TA98 against 2-amino-3-methyl-imidazo{4,5-f}quinoline (IQ) in the presence of Aroclor 1254-induced rat hepatic S9 . The Maillard reaction products in the dichloromethane extracts were then quantified and qualified by capillary gas chromatography and gas chromatography-mass spectrometry, respectively . Pyrazines and furans were found to be the major Maillard reaction products yielded in the extracts . Moreover, the antimutagenicity of dichloromethane extracts correlated positively with the total amounts of pyrazines and furans . To elucidate the mechanism of antimutagenicity of dichloromethane extracts, the inhibitory effect of pyrazines on ethoxycoumarin deethylase activity in Aroclor 1254-induced hepatic microsomes was examined . We also studied the effects of pyrazines on IQ metabolism by Aroclor 1254-induced microsomes using high-performance liquid chromatography . The antimutagenicity of pyrazines correlated positively with both the inhibition of cytochrome P-450 IA2-linked ethoxycoumarin deethylase in hepatic microsomes and the inhibition of N-hydroxy-IQ formation from IQ metabolism by hepatic microsomes . Thus we concluded that pyrazines in dichloromethane extracts from eight amino acid/sugar model systems play an important role in the antimutagenicity of IQ . Moreover, we concluded that the modifying effect of pyrazines on the mutagenicity of IQ is mediated through interaction with microsomal activating enzymes to inhibit the major active metabolite in N-hydroxy-IQ formation.

Mutagenesis, 1994 Sep, 9(5), 473 - 6
Genotoxicity of 2-halocinnamaldehydes in two bacterial assays . Induction of SOS repair and frame-shift mutation; Eder E et al.; 2-Chlorocinnamaldehyde and 2-bromocinnamaldehyde, compounds of practical interest, for example, as bacteriocides and fungicides or for utilization in light sensitive layers, were tested in the Ames preincubation test with various Salmonella typhimurium strains, and in the SOS chromotest with Escherichia coli PQ 37.2-Chlorocinnamaldehyde was clearly mutagenic in strain TA 100 (6081 revertants/mumol) and in strain TA 98 (3050 revertants/mumol) without S9 mix, and was clearly positive in the SOS chromotest (SOSIP = 0.181) . 2-Bromocinnamaldehyde was a strong mutagen in strain TA 100 (105, 500 revertants/mumol), in strain TA98 (41567 revertants/mumol) and in strain TA 1538 (15825 revertants/mumol), and also unambiguously mutagenic in strain TA 1535 (2110 revertants/mumol) without S9 mix . The SOSIP in the SOS chromotest was 1.5 . Addition of S9 mix led to a marked decrease in the mutagenic activity of 2-bromocinnamaldehyde in all strains tested . In the case of strain TA 1535, mutagenic activity was abolished or not significant in the presence of S9 mix . The possible primary mechanisms underlying these mutagenic effects are discussed . Frame-shift activity of these halocinnamaldehydes can be explained by their planar structure.

Mutagenesis, 1994 Sep, 9(5), 459 - 65
Photomutagenicity assays in bacteria: factors affecting assay design and assessment of photomutagenic potential of para-aminobenzoic acid; Henderson L et al.; Photomutagenicity assays are required for regulatory submissions of some chemicals . As yet there are no well-validated protocols available for these assays . Critical factors which may contribute to the ability of a bacterial assay to detect photomutagens (e.g . dose of UV and test chemical, exposure conditions, light source, bacterial strains) were investigated using two known photomutagens, chlorpromazine and 8-methoxypsoralen . Salmonella typhimurium strains TA98, TA102 and TA1537 and Escherichia coli strains WP2 and WP2(pKM101) were used and differences in the responsiveness of these strains were observed with these substances . Both chemicals were detected using either UV exposure in suspension or on the agar plates . On the basis of these observations and on other results reported in the literature, recommendations are made on protocol aspects for assessing photomutagenic potential in routine screening tests . Using these recommendations the sunscreen para-aminobenzoic acid was tested in S.typhimurium strains TA98, TA100, TA1535, TA1537 and E . coli strains WP2 and WP2 (pKM101), using both plate irradiation and suspension exposure conditions . No evidence of mutagenic potential was detected.

Mutagenesis, 1994 Sep, 9(5), 401 - 5
A reverse mutagenicity assay for alkylating agents based on a point mutation in the beta-lactamase gene at the active site serine codon; Lee CC et al.; The serine at the active site of beta-lactamase is responsible for the ester link to the acyl group of beta-lactam during hydrolysis of the substrate to its acid derivatives . A construct was made from a plasmid in which the active-site serine of beta-lactamase was substituted by glycine by site-directed mutation . This mutation results in the loss of beta-lactamase activity . This plasmid was used to transform Salmonella typhimurium TA1535 . When the new strain JK947 was treated with a mutagen such as N-methyl-N'-nitro-nitrosoguanidine (MNNG), the bacteria could be recovered as they became ampicillin resistant . The sequence of the active-site serine codon in these revertants was mutated from GGC to AGC . Based upon these findings, we developed a model reverse mutagenicity assay . In this procedure, we treated JK947 with a test chemical, such as N-methyl-nitrosourea (MNU), dimethyl sulphate (DMS) or methylmethane sulphonate (MMS), for 30 min, and then scored the revertants on agar plates containing 50 micrograms/ml ampicillin after incubation at 37 degrees C for 16 h . MNU and MNNG were more potent than DMS and MMS in this assay . Treatment with MNU and MNNG resulted in larger colony numbers in our test than in the Ames test . However, our test was less sensitive to DMS and MMS than the Ames test.

Genomics, 1994 Sep 1, 23(1), 51 - 61
Haplotype mapping and sequence analysis of the mouse Nramp gene predict susceptibility to infection with intracellular parasites; Malo D et al.; The mouse chromosome 1 locus Bcg (Ity, Lsh) controls the capacity of the tissue macrophage to restrict the replication of antigenically unrelated intracellular parasites and therefore determines the natural resistance (BCG-R, dominant) or susceptibility (BCG-S, recessive) of inbred mouse strains to infection with diverse pathogens, including several Mycobacterium species, Salmonella typhimurium, and Leishmania donovani . We have used a positional cloning strategy based on genetic and physical mapping, YAC cloning, and exon trapping to isolate a candidate gene for Bcg (Nramp) that encodes a predicted macrophage-specific transport protein . We have analyzed a total of 27 inbred mouse strains of BCG-R and BCG-S phenotypes for the presence of nucleotide sequence variations within the coding portion of Nramp and have carried out haplotype typing of the corresponding chromosome 1 region in these mice, using 11 additional polymorphic markers mapping in the immediate vicinity of Nramp . cDNA cloning and nucleotide sequencing identified 5 nucleotide sequence variations within Nramp in the inbred strains; while 4 of these represented silent sequence polymorphisms, one G to A substitution at nucleotide position 783 resulted in the non-conservative replacement of Gly105 to Asp105 within the second predicted transmembrane domain (TM2) of the Nramp protein . An absolute association of this allelic variation and Bcg phenotype was observed in the 20 BCG-R strains (Gly105) and 7 BCG-S strains (Asp105) tested . Moreover, sequence analysis of the corresponding region of the Nramp gene from distantly related species indicated strong amino acid sequence conservation of TM2, including an invariant glycine at position 105 . Haplotype mapping using sequence polymorphism identified within Nramp and additional RFLPs and SSLPs from the region revealed that although the 20 BCG-R strains analyzed showed diverse allelic combinations for these markers, the 7 BCG-S strains tested share a conserved core haplotype of 2.2 Mb overlapping and including Nramp . Taken together, these results suggest that (1) Gly105 is the wildtype form of Nramp and that the nonconservative substitution to Asp105 underlies the BCG-S phenotype, and (2) Bcg8 alleles carry the same Gly105-->Asp105 mutation and are identical by descent.

Mol Microbiol, 1994 Sep, 13(5), 797 - 805
Residue threonine-149 of the Salmonella typhimurium CysB transcription activator: mutations causing constitutive expression of positively regulated genes of the cysteine regulon; Colyer TE et al.; In both Salmonella typhimurium and Escherichia coli, CysB is a LysR family transcriptional activator, which regulates genes of the cysteine regulon . Transcription activation of cys genes also requires an inducer, N-acetyl-L-serine, and cysB mutants that do not require inducer are termed constitutive, i.e . cysBc . After finding that two independently isolated cysBc mutants are substituted at amino acid residue threonine-149 (T149), we isolated the other 17 single-amino-acid substitutions by site-directed mutagenesis . Of the 19 mutant alleles, 11 supported normal growth on sulphate, and nine of these were cysBc . Four other mutants were 'leaky' cysB+, and four were cysB- . Insertions of up to 14 amino acids were also tolerated at T149, and two of three such mutants were cysBc . An allele containing a TAG translation terminator at codon 149 had no detectable function in a delta cysB strain, but gave a constitutive phenotype when introduced into either wild-type S . typhimurium or the E . coli strain NK1, which contains a cysB- mutation in a predicted helix-turn-helix region that interferes with specific binding of CysB to DNA and with autoregulation of cysB . The peptide encoded by the T149ter allele is proposed to interact with the wild-type CysB peptide or with the NK1 mutant peptide to form hetero-oligomers that do not require N-acetyl-L-serine for cys gene activation.

J Clin Microbiol, 1994 Sep, 32(9), 2327 - 30
Demonstration of persistence of Salmonella typhimurium in an AIDS patient by molecular methods; Fica AE et al.; We document microbiological persistence of the same Salmonella typhimurium strain in an AIDS patient during 7 months of clinical observation despite prolonged quinolone therapy . Persistence was demonstrated by phage types that closely resembled one another, similar antibiotic resistance patterns, conserved restriction fragment length polymorphism of chromosomal DNA digested with different DNA restriction enzymes, identical ribotypes, and IS200 types in four characterized sequential isolates of S . typhimurium.

Poult Sci, 1994 Sep, 73(9), 1409 - 16
Effect of cecal cultures lyophilized in skim milk or reagent 20 on Salmonella colonization in broiler chicks; Hollister AG et al.; Mixed cultures of cecal bacteria that were grown under continuous flow anaerobic conditions were prepared as lyophilized powder in skim milk or Reagent 20 (R-20; a mixture containing sucrose and bovine serum albumin fraction V) and compared with broth cultures for reduction of Salmonella typhimurium enteric colonization . Day old broiler chicks were provided a standard corn-soybean diet with: 1) no culture, (control); 2) broth culture administered by crop gavage; 3) broth culture added to the drinking water; 4) culture lyophilized in skim milk and added to drinking water; 5) culture lyophilized in skim milk in gelatin capsules and force-fed; 6) culture lyophilized in R-20 and added to drinking water; and 7) culture lyophilized in R-20 in gelatin capsules and force-fed . All groups were challenged on Day 3 with 10(4) cfu of S . typhimurium per chick . Culture by crop gavage, culture in the drinking water, skim milk powder in capsules, and R-20 powder in the water and in capsules significantly (P < .05) reduced mean Salmonella colony-forming units in cecal contents by 3.21 to 5.26 log10 units at 10 d of age . Likewise, the number of cecal-culture-positive chicks in the same groups was significantly less than controls with reductions of 27 to 67% . The numbers of Salmonella per gram of cecal contents and the percentage of cecal-culture-positive chicks in the skim milk powder in the drinking water group were not different from control chicks in one of two experiments.(ABSTRACT TRUNCATED AT 250 WORDS)

J Biomed Mater Res, 1994 Sep, 28(9), 1061 - 7
Mutagenicity of dentin bonding agents; Schweikl H et al.; Aside from the considerable number of reports on the physical and chemical properties of dental bonding agents, information concerning their biologic effects is sparse . Three dentin bonding agents (Prisma Universal Bond, Pertac Universal Bond, and Syntac) and the ingredients methylmethacrylate, 2-hydroxyethyl-methacrylate, and glutaraldehyde were investigated in the Salmonella typhimurium mutagenicity test system using five different bacterial strains (TA97a, TA98, TA100, TA102, and TA104) . The materials as well as the ingredients were eluted in both dimethyl sulfoxide and physiologic saline, and serially diluted eluates were used in the plate incorporation test . Pertac Universal Bond and Prisma Universal Bond did not elicit any mutagenic effects in any of the bacterial strains . In contrast, Syntac adhesive showed clear mutagenicity in S . typhimurium strain TA102 . Dimethyl sulfoxide eluates, as well as physiologic saline eluates of the Syntac bonding agent, caused numbers of revertants that were about 6 times higher than control values . Reversion rates with other strains were moderately enhanced . Glutaraldehyde, an ingredient of Syntac adhesives, caused mutagenicity in a manner similar to Syntac adhesive eluates . Neither 2-hydroxyethyl-methacrylate nor methylmethacrylate monomer was found to be mutagenic over a broad concentration range.

Mutat Res, 1994 Sep, 325(1), 7 - 10
Mutagenic evaluation of primaquine, pentaquine and pamaquine in the Salmonella/mammalian microsome assay; Ono T et al.; The 8-aminoquinolines, primaquine, pentaquine and pamaquine, were investigated for mutagenic activity in Salmonella typhimurium strains TA100, TA98, TA97 and TA102 in the rat liver microsomal activation system . Primaquine and pentaquine induced mutations in TA97 in the presence and absence of S9 mix . Pamaquine was mutagenic to TA98 only in the absence of S9 mix.

Mutat Res, 1994 Sep 1, 309(2), 201 - 10
Antimutagenicity of three isomers of aminobenzoic acid in Salmonella typhimurium; Gichner T et al.; The m-, o- and p-isomers of aminobenzoic acid (ABA) repressed the mutagenicity of N-methyl-N'-nitro-N-nitrosoguanidine (MNNG) in Salmonella typhimurium TA100 . Their antimutagenic potency was in the order of o-ABA > m-ABA > p-ABA . The mechanism of this antimutagenicity is ascribed mainly to the decomposition of MNNG induced by the aminobenzoic acid isomers outside or within the bacterial cells . The inhibition of plant cell peroxidases and bacterial acetyltransferases that are required for the plant activation of 2-aminofluorene (2-AF) to mutagenic product(s) may participate in the repression of 2-AF mutagenesis by the aminobenzoic acids in S . typhimurium strain YG1024 . The aminobenzoic acid isomers exhibited no inhibitory effects towards the direct-acting agent 2-acetoxy-2-acetylaminofluorene, the stable diacetylated metabolic product of 2-AF.

J Exp Med, 1994 Sep 1, 180(3), 1037 - 46
Immunity to malaria elicited by hybrid hepatitis B virus core particles carrying circumsporozoite protein epitopes; Schodel F et al.; The hepatitis B virus (HBV) nucleocapsid antigen (HBcAg) was investigated as a carrier moiety for the immunodominant circumsporozoite (CS) protein repeat epitopes of Plasmodium falciparum and the rodent malaria agent P . berghei . For this purpose hybrid genes coding for {NANP}4 (C75CS2) or {DP4NPN}2 (C75CS1) as internal inserts in HBcAg (between amino acids 75 and 81) were constructed and expressed in recombinant Salmonella typhimurium . The resulting hybrid HBcAg-CS polypeptides purified from S . typhimurium were particulate and displayed CS and HBc antigenicity, however, the HBc antigenicity was reduced compared to native recombinant HBcAg . Immunization of several mouse strains with HBcAg-CS1 and HBcAg-CS2 particles resulted in high titer, P.berghei- or P.falciparum-specific anti-CS antibodies representing all murine immunoglobulin G isotypes . The possible influence of carrier-specific immunosuppression was examined, and preexisting immunity to HBcAg did not significantly affect the immunogenicity of the CS epitopes within HBcAg-CS1 particles . Similarly, the choice of adjuvant did not significantly alter the immunogenicity of HBcAg-CS hybrid particles . Immunization in complete or incomplete Freund's adjuvant or alum resulted in equivalent anti-HBc and anti-CS humoral responses . Examination of T cell recognition of HBcAg-CS particles revealed that HBcAg-specific T cells were universally primed and CS-specific T cells were primed if the insert contained a CS-specific T cell recognition site . This indicates that the internal site in HBcAg is permissive for the inclusion of heterologous pathogen-specific T as well as B cell epitopes . Most importantly, 90 and 100% of BALB/c mice immunized with HBcAg-CS1 particles were protected against a P . berghei challenge infection in two independent experiments . Therefore, hybrid HBcAg-CS particles may represent a useful approach for future malaria vaccine development.

Proc Natl Acad Sci U S A, 1994 Aug 30, 91(18), 8631 - 5
Dimorphic transition in Escherichia coli and Salmonella typhimurium: surface-induced differentiation into hyperflagellate swarmer cells; Harshey RM et al.; We describe a new behavioral response in Escherichia coli and Salmonella typhimurium in which the bacteria differentiate into filamentous, multinucleate, hyperflagellate cells that navigate the surface of solid media by means of coordinated swarming motility . The cue for differentiation into swarmer cells is provided by the concentration and composition of the agar . Examination of the behavior of various mutants shows that the flagellar apparatus used for swimming motility and the chemotaxis system are indispensable for swarming motility.

Gene, 1994 Aug 19, 146(1), 15 - 21
Alternative patterns of his operon transcription and mRNA processing generated by metabolic perturbation; Alifano P et al.; Previous studies have shown that the expression of the his operon of Salmonella typhimurium is regulated at the level of transcription initiation, transcription elongation and RNA processing . We have analyzed his RNA in both prototrophic strains or strains harboring regulatory and auxotrophic mutations grown under a variety of metabolic conditions that lead to differential expression of the operon . Under some of these conditions, there is an increase in the amount of prematurely released his-specific RNA, resulting in modulation of the relative amount of full-length transcripts . Under the same metabolic conditions, there is also a modulation of RNA processing events that generate a very stable RNA species comprising the five distal cistrons . These effects appear to be due to perturbation of the translation process caused by alterations in the intracellular pool of initiator transfer RNA.

Proc Natl Acad Sci U S A, 1994 Aug 16, 91(17), 8092 - 6
A purified selenophosphate-dependent enzyme from Salmonella typhimurium catalyzes the replacement of sulfur in 2-thiouridine residues in tRNAs with selenium; Veres Z et al.; A tRNA-modifying enzyme tentatively termed tRNA 2-selenouridine synthase was purified by a five-step procedure that resulted in 50-60% pure preparations . This enzyme catalyzes the conversion of a 5-methylaminomethyl-2-thiouridine residue in the tRNA substrate to 5-methylaminomethyl-2-selenouridine . The selenium donor substrate for this reaction is shown to be selenophosphate which is formed from ATP and selenide by selenophosphate synthetase . Replacement of sulfur with selenium in tRNAs catalyzed by tRNA 2-selenouridine synthase occurs in the absence of ATP . The dependence of reaction velocity on selenophosphate concentration obeys Michaelis-Menten kinetics indicating an apparent Km value of 17.1 microM . Bulk thio-tRNA preparations from Escherichia coli and Salmonella typhimurium are equally effective as substrates for the selenium incorporation reaction . An intact 3' end of the tRNA molecule does not seem to be essential for selenium incorporation . Identity of the product of the reaction was confirmed by HPLC analysis of digests of {75Se}seleno-tRNAs labeled by incubation with the purified enzyme . A labeled compound in the nucleoside mixture was coeluted with authentic 5-methylaminomethyl-2-selenouridine.

J Biol Chem, 1994 Aug 12, 269(32), 20456 - 61
Nucleotide-induced conformational changes of MalK, a bacterial ATP binding cassette transporter protein; Schneider E et al.; Nucleotide-induced structural rearrangements of MalK, the ATP-hydrolyzing component of the ATP binding cassette transporter for maltose from Salmonella typhimurium were investigated by means of analysis of intrinsic tryptophan fluorescence and limited proteolysis . ATP was found to decrease the tryptophan fluorescence of purified MalK by 37 +/- 1% . ADP or adenosine 5'-O-(3-(thio)triphosphate) (ATP gamma S) caused similar quenching while AMP was rather ineffective . Mg2+ ions were not required . Exposure of MalK to increasing concentrations of trypsin and subsequent analysis by SDS-polyacrylamide gel electrophoresis and immunoblotting revealed the formation of three major transiently stable peptide fragments of 24 (T2), 23 (T3), and 20 kDa (T4), respectively . In addition, a minor rapidly degraded fragment of 33 kDa (T1) was observed . However, in the presence of MgATP, fragment T1 as well as a substantial fraction of native MalK were strongly protected against proteolytic attack . Similar protection against the protease was observed in the presence of MgGTP or, to a lesser extent, MgCTP . In contrast, MgADP, ATP in the presence of EDTA, CaATP or nonhydrolyzable nucleotides such as MgATP gamma S or MgAMP-PNP (beta, gamma-imidoadenosine-5'-triphosphate) failed to significantly affect the susceptibility of MalK to the protease . MgATP similarly affected the tryptic digestion pattern of a mutant protein (MalKK42R) that exhibits only a much reduced ATPase activity but has retained the capability to bind nucleotides . N-terminal protein sequence analysis of the peptides revealed cleavage by trypsin at Arg66 (T1), Arg146 (T2), Arg153 (T3), and Arg185 (T4), respectively . These results indicate that (i) nucleotide binding to MalK is accompanied by a global conformational change of the protein; (ii) a very specific interaction occurs with substrates of the MalK-ATPase, resulting in structural changes that involve the helical domain from Arg66 to Arg146; and (iii) the C-terminal half of MalK is rather resistant to proteolysis.

Biochim Biophys Acta, 1994 Aug 2, 1218(3), 443 - 6
Nucleotide sequence of the flgD gene of Salmonella typhimurium which is essential for flagellar hook formation; Kutsukake K et al.; We determined the complete nucleotide sequence of the flgD gene of Salmonella typhimurium which is essential for flagellar hook formation . The sequence predicts a protein of 232 amino acids and a calculated molecular mass of 23,987 Da . However, the N-terminal 86 amino acids of FlgD were found sufficient to complement all the flgD mutations examined . The predicted secondary structure suggested that FlgD has a high content of beta structure.

FEMS Microbiol Lett, 1994 Aug 1, 121(1), 99 - 105
Transcription of the Salmonella typhimurium spv virulence locus is regulated negatively by the nucleoid-associated protein H-NS; O'Byrne CP et al.; The possibility that the pleiotropic transcriptional regulator H-NS might play a role in regulating expression of the spv virulence locus of Salmonella typhimurium was investigated . A transposon insertion mutation in hns, the gene encoding H-NS, resulted in enhanced transcription of the spvR regulatory gene and the spvB structural gene in stationary phase cultures . Enhanced transcription was not detected prior to stationary phase, indicating that H-NS makes a negative contribution that is growth phase-specific to the control of spv transcription . When H-NS was over-expressed from a multicopy plasmid, the normal stationary phase induction of spv transcription seen in wild-type cells was abolished . spv transcription was also found to be modulated by growth medium osmolarity, a feature common to many H-NS-regulated genes . In addition, transcription of the spv genes was reduced in mutants with abnormal levels of DNA supercoiling.

Carcinogenesis, 1994 Aug, 15(8), 1749 - 51
Iron-dependent formation of 8-hydroxydeoxyguanosine in isolated DNA and mutagenicity in Salmonella typhimurium TA102 induced by crocidolite; Faux SP et al.; Treatment of isolated DNA with crocidolite asbestos significantly increased the concentration of 8-hydroxydeoxyguanosine (8-OHdG) above background . Furthermore, incubating DNA with H2O2 and crocidolite potentiated the formation of 8-OHdG above levels observed with crocidolite alone . In the presence of desferrioxamine, desferrioxamine and ferrozine, dimethylsulphoxide (DMSO) or o-phenanthroline, crocidolite-induced DNA oxidation was reduced by 36, 73, 74 and 70% respectively . Crocidolite, but not chrysotile asbestos, enhanced background revertants in Salmonella typhimurium TA102, at sub-cytotoxic concentrations in a dose-dependent manner . The mutagenic effects of crocidolite were quite small and this indicates that crocidolite was a weak mutagen in this study . The number of revertants was reduced to the spontaneous rate for this strain after the fibres had been pretreated with desferrioxamine before assaying for genotoxicity in this oxygen radical-sensitive strain . These results help to explain a mechanistic role for iron in crocidolite-induced DNA oxidation and mutagenicity in TA102.

Am J Gastroenterol, 1994 Aug, 89(8), 1246 - 8
Intussusception associated with Salmonella typhimurium enterocolitis; Matsushita M et al.; Intussusception in the adult is unusual, and its etiology is mostly neoplastic . In most cases of intussusception, a polypoid tumor located in the terminal ileum invaginates within the cecum . We described an unusual case of adult intussusception associated with Salmonella typhimurium enterocolitis . Preoperative diagnosis of intussusception in the ileocecal region was made by ultrasonography, and the ileocecal region containing the mass in the cecum, causing the intussusception, was resected . Histological features of the mass included an edematous focal wall thickening . Later, the diagnosis of S . typhimurium enterocolitis was made by the preoperative stool cultures . This case demonstrated intussusception as a complication of S . typhimurium enterocolitis . It is worth noting that S . typhimurium enterocolitis is in fact a potential cause of intussusception.

J Bacteriol, 1994 Aug, 176(16), 5001 - 4
A change in a single gene of Salmonella typhimurium can dramatically change its buoyant density; Baldwin WW et al.; The growth rates and buoyant densities of a Salmonella typhimurium mutant, TL126 (proB74A+), with enhanced osmotolerance caused by proline overproduction were measured and compared with the growth rates and buoyant densities of an isogenic (wild-type) strain, TL128 (proB+ A+), with normal control of proline production . Growth rates were determined in a rich medium (Luria broth) with added NaCl to produce various osmotic strengths ranging from 300 to 2,000 mosM . At low concentrations of NaCl, there was little variation in doubling times between the two strains . However, as the osmotic strength of the medium approached and exceeded 1,300 mosM, the doubling times of TL126 (osmotolerant) were 1.5 to 2 times faster than those of TL128 (wild type), confirming the osmotolerance of TL126 . Buoyant densities were determined by equilibrium sedimentation in a Percoll gradient of osmotic strength equal to that of the growth medium . The osmolarity of the Percoll gradient was adjusted by the addition of NaCl . At low osmolarities (300 to 500 mosM), the buoyant density of TL126 (osmotolerant) was slightly but consistently lower than that of TL128 (wild type) . As the osmotic strength was increased, the buoyant density of TL126 (osmotolerant) increased in proportion to the osmotic strength . In contrast, the buoyant density of strain TL128 (wild type) did not increase as much . At high osmolarities (1,600 to 2,000 mosM), the buoyant density of TL126 (osmotolerant) was consistently higher than that of TL128 (wild type) . These results suggest that the intracellular accumulation of proline by TL126, the osmotolerant strain, increases both the growth rates and buoyant densities at osmolarities of 1,300 mosM and above.

J Bacteriol, 1994 Aug, 176(15), 4610 - 6
RpoS is necessary for both the positive and negative regulation of starvation survival genes during phosphate, carbon, and nitrogen starvation in Salmonella typhimurium; O'Neal CR et al.; The starvation stress response of Salmonella typhimurium encompasses the genetic and physiologic changes that occur when this bacterium is starved for an essential nutrient such as phosphate (P), carbon (C), or nitrogen (N) . The responses to the limitation of each of these nutrients involve both unique and overlapping sets of proteins important for starvation survival and virulence . The role of the alternative sigma factor RpoS in the regulation of the starvation survival loci, stiA, stiB, and stiC, has been characterized . RpoS (sigma S) was found to be required for the P, C, and N starvation induction of stiA and stiC . In contrast, RpoS was found to be required for the negative regulation of stiB during P and C starvation-induced stationary phase but not during logarithmic phase . This role was independent of the relA gene (previously found to be needed for stiB induction) . The role of RpoS alone and in combination with one or more sti mutations in the starvation survival of the organism was also investigated . The results clearly demonstrate that RpoS is an integral component of the complex interconnected regulatory systems involved in S . typhimurium's response to nutrient deprivation . However, differential responses of various sti genes indicate that additional signals and regulatory proteins are also involved.

J Bacteriol, 1994 Aug, 176(15), 4534 - 42
YscU, a Yersinia enterocolitica inner membrane protein involved in Yop secretion; Allaoui A et al.; Pathogenic yersiniae secrete antihost Yop proteins by a recently discovered secretion pathway which is also encountered in several animal and plant pathogens . The components of the export machinery are encoded by the virA (lcrA), virB (lcrB), and virC (lcrC) loci of the 70-kb pYV plasmid . In the present paper we describe yscU, the last gene of the virB locus . We determined the DNA sequence and mutated the gene on the pYV plasmid . After inactivation of yscU, the mutant strain was unable to secrete Yop proteins . The topology of YscU was investigated by the analysis of YscU-PhoA translational fusions generated by TnphoA transposition . This showed that the 40.3-kDa yscU product contains four transmembrane segments anchoring a large cytoplasmic carboxyl-terminal domain to the inner membrane . YscU is related to Spa40 from Shigella flexneri, to SpaS from Salmonella typhimurium, to FlhB from Bacillus subtilis, and to HrpN from Pseudomonas solanacearum.

J Bacteriol, 1994 Aug, 176(15), 4501 - 10
Molecular and functional characterization of the Salmonella typhimurium invasion genes invB and invC: homology of InvC to the F0F1 ATPase family of proteins; Eichelberg K et al.; Entry into intestinal epithelial cells is an essential step in the pathogenesis of Salmonella infections . Our laboratory has previously identified a genetic locus, inv, that is necessary for efficient entry of Salmonella typhimurium into cultured epithelial cells . We have carried out a molecular and functional analysis of invB and invC, two members of this locus . The nucleotide sequence of these genes indicated that invB and invC encode polypeptides with molecular masses of 15 and 47 kDa, respectively . Polypeptides with the predicted sizes were observed when these genes were expressed under the control of a T7 promoter . Strains carrying nonpolar mutations in these genes were constructed, and their phenotypes were examined in a variety of assays . A mutation in invC rendered S . typhimurium defective in their ability to enter cultured epithelial cells, while mutations in invB did not . Comparison of the predicted sequences of InvB and InvC with translated sequences in GenBank revealed that these polypeptides are similar to the Shigella spp . proteins Spa15 and Spa47, which are involved in the surface presentation of the invasion protein antigens (Ipa) of these organisms . In addition, InvC showed significant similarity to a protein family which shares sequence homology with the catalytic beta subunit of the F0F1 ATPase from a number of microorganisms . Consistent with this finding, purified preparations of InvC showed significant ATPase activity . Site-directed mutagenesis of a residue essential for the catalytical function of this family of proteins resulted in a protein devoid of ATPase activity and unable to complement an invC mutant of S . typhimurium . These results suggest that InvC may energize the protein export apparatus encoded in the inv locus which is required for the surface presentation of determinants needed for the entry of Salmonella species into mammalian cells . The role of InvB in this process remains uncertain.

J Bacteriol, 1994 Aug, 176(15), 4492 - 500
Identification of flagellar synthesis regulatory and structural genes in a sigma D-dependent operon of Bacillus subtilis; Mirel DB et al.; The sigma D form of RNA polymerase from Bacillus subtilis has been shown previously to direct the synthesis of several transcription units bearing genes for flagellin, motility proteins, and autolysins . In this report, we describe an operon of genes transcribed from the sigma D-dependent promoter PD-1 . We have identified three complete open reading frames and one partial one downstream of this promoter; immediately upstream is the previously identified comF locus . The PD-1 operon encodes the presumptive B . subtilis homologs of two Salmonella typhimurium late flagellar genes, flgM and flgK . Also present in this operon are two genes of unknown function, orf139 and orf160, whose products show similarities to the eukaryotic cytoskeletal proteins myosin and vimentin, respectively . orf139 and orf160 may encode proteins that form extended alpha-helical secondary structures and coiled-coil quaternary structures which may be filamentous components of the gram-positive bacterial flagellum . We have characterized the B . subtilis flgM gene further by constructing an in-frame deletion mutation, flgM delta 80, and creating strains of B . subtilis in which this allele has replaced the wild-type copy . By primer extension analysis of cellular RNA, we have shown that the flgM delta 80 mutation relieves the block to transcription of two other sigma D-dependent operons imposed by an unlinked mutation in a gene directing early flagellar synthesis . We conclude that, as in the case of S . typhimurium, early flagellar synthesis in B . subtilis is coupled to late flagellar synthesis through repression of sigma D-dependent transcription by the flgM gene product.

Infect Immun, 1994 Aug, 62(8), 3162 - 71
Oral immunization with recombinant Salmonella typhimurium expressing surface protein antigen A of Streptococcus sobrinus: persistence and induction of humoral responses in rats; Redman TK et al.; Recombinant Salmonella typhimurium has been used as an oral vaccine for various microbial pathogens . Here we report immune responses in Fischer rats orally immunized with a recombinant S . typhimurium strain encoding surface protein antigen A (SpaA) of Streptococcus sobrinus . The attenuated S . typhimurium chi 4072 delta cya delta crp delta asd mutant used in this study contains the Asd+ plasmid pYA2905 expressing a fragment of the SpaA protein . Salmonella cells were cleared from spleens by 7 days and from Peyer's patches by 14 days in rats receiving a single oral immunization of 10(9) CFU of chi 4072 . In animals receiving multiple (i.e., days 0 and 7 or days 0, 7, and 21) immunizations, Salmonella cells were cleared from the Peyer's patches by 25 days following the initial immunization . Antigen-specific systemic and mucosal antibody responses were greater in rats receiving multiple immunizations than in those receiving a single immunization . Serum anti-Salmonella activity was potentiated following boosting on day 21 . Mucosal immunoglobulin A antibody responses were also greater in rats receiving multiple immunizations than in rats receiving a single immunization . Anti-Salmonella and anti-Streptococcus immunoglobulin A activity persisted longer in rats boosted on day 21 than in rats immunized on days 0 and 7 . These data indicate that oral immunization of rats with the recombinant S . typhimurium chi 4072(pYA2905) vaccine induces systemic as well as mucosal antibody responses specific to the Salmonella cells and to the cloned SpaA protein . This is the first report of the use of an attenuated mutant of the murine pathogen S . typhimurium as an oral vaccine in rats.

Arch Biochem Biophys, 1994 Aug 1, 312(2), 493 - 500
Steady-state kinetics of cabbage histidinol dehydrogenase; Kheirolomoom A et al.; Cabbage histidinol dehydrogenase (HDH) oxidizes L-histidinol to L-histidine through two sequential NAD(+)-linked reactions via an alkaline-labile, L-histidinaldehyde intermediate . The kinetic mechanism of the overall reaction as well as the partial reactions involved in the overall catalysis were investigated at pH 7.2 using L-histidinaldehyde as a substrate . Product inhibition patterns conformed to a Bi Uni Uni Bi Ping Pong mechanism as reported for the HDH from Salmonella typhimurium . Thus, the reaction scheme is ordered with the binding of histidinol first and NAD+ second, and histidine is the last product to be released . The intermediate, L-histidinaldehyde, could be a substrate for both the oxidation and the reduction reactions to produce histidine and histidinol, respectively . L-Histidine was not enzymatically reduced in the presence of NADH, indicating that the reaction to oxidize histidinaldehyde is apparently irreversible . L-Histidinaldehyde exhibited a three times greater binding rate constant than histidinol with a considerably small dissociation constant . These results were in agreement with the observation that histidinaldehyde was not released during the overall reaction . The rate of the reduction of histidinaldehyde to histidinol was almost same as that of the overall oxidation reaction . The overall oxidation from histidinol to histidine proceeded about three times slower than the partial oxidation from histidinaldehyde to histidine, suggesting that the first-half forward reaction is the rate-determining step in the total reaction of cabbage HDH.

Mol Microbiol, 1994 Aug, 13(4), 641 - 53
Cloning and primary structure of Staphylococcus aureus DNA topoisomerase IV: a primary target of fluoroquinolones; Ferrero L et al.; A 4.6 kb Staphylococcus aureus DNA fragment containing DNA gyrase-like genes (grlA and grlB) was cloned and sequenced . The proteins GrlA and GrlB exhibit more than 30% identity with E . coli DNA topoisomerase IV subunits and with the gyrase subunits from S . aureus and Escherichia coli . The combined E . coli cell extracts of GrlA and GrlB overproducing strains catalysed ATP-dependent relaxation and decatenation specific to DNA topoisomerase IV . The temperature-sensitive phenotype of Salmonella typhimurium parC and parE mutants was complemented by the S . aureus grlA and grlB genes, when the two genes were co-expressed . These results show that GrlA and GrlB are the subunits of S . aureus DNA topoisomerase IV . The GyrA subunit of DNA gyrase has been previously defined as a primary target of quinolones based on genetic and biochemical experiments essentially carried out in E . coli . Single-point mutations occurring in the 'quinolone resistance-determining region' (QRDR) of GyrA were found in bacteria exhibiting quinolone resistance, the most common mutation being a substitution of Ser-83 on the E . coli GyrA sequence . We analysed eight S . aureus fluoroquinolone-resistant clinical isolates and observed that mutations in the QRDR of GyrA are not present in the low-quinolone-resistant isolates . In contrast, Ser-80 of GrlA, which corresponds to Ser-83 of E . coli GyrA, is substituted to Phe or Tyr in both high- and low-quinolone-resistant isolates . We propose that DNA topoisomerase IV is a primary target of fluoroquinolones in S . aureus.

Mol Microbiol, 1994 Aug, 13(4), 555 - 68
The Salmonella typhimurium invasion genes invF and invG encode homologues of the AraC and PulD family of proteins; Kaniga K et al.; We have identified two novel Salmonella typhimurium genes, invF and invG, which are required for the efficient entry of these organisms into cultured epithelial cells . invF and invG are located immediately upstream of invE, a previously identified gene also required for Salmonella entry . Non-polar mutations in these genes rendered S . typhimurium severely deficient for entry into cultured epithelial cells . The nucleotide sequences of invF and invG indicated that these genes encode polypeptides with predicted molecular weights of 24,373 and 62,275, respectively . Proteins of similar sizes were observed when invF and invG were expressed in a bacteriophage T7 RNA polymerase-based expression system . Comparison of the predicted sequence of InvF with translated sequences in the existing databases indicated that this protein is homologous to members of the AraC family of prokaryotic transcription regulators . However, mutations in invF did not significantly affect the expression of other members of the inv locus . InvG was found to be homologous to members of the PulD family of specialized translocases . This homology suggests that InvG may be necessary for the export of invasion-related determinants or involved in the assembly of a supramolecular structure that promotes entry.

Mol Microbiol, 1994 Aug, 13(3), 389 - 94
Bees aren't the only ones: swarming in gram-negative bacteria; Harshey RM; Swarming is a form of active surface motility that is widespread among flagellated, Gram-negative bacteria . In the laboratory, growth of the bacteria on certain agar surfaces leads to induction of the differentiated swarmer-cell state . Swarmer cells are generally long and multinucleate, always hyperflagellated, and can move rapidly over the agar surface in a coordinated manner . Some swarm colonies exude large amounts of 'slime', which could be essential for promoting intimate cell-cell contacts during swarming . There is evidence that the differentiated swarmer-cell stage facilitates pathogenic associations with host tissue . Almost nothing is known about the molecular signalling mechanism of surface sensing . Increased viscosity appears to be sensed by several bacteria, but other environmental cues, specific to each bacterium, are also important . In organisms in which swarming motility has been studied in some detail, the chemotaxis system has been shown to play an important role . The recent discovery of swarming motility in two genetically well-characterized organisms--Escherichia coli and Salmonella typhimurium--should lead to rapid progress in understanding this process.

Yeast, 1994 Aug, 10(8), 1031 - 44
Phosphoribosylpyrophosphate synthetase (PRS): a new gene family in Saccharomyces cerevisiae; Carter AT et al.; Saccharomyces cerevisiae contains at least four PRS genes, all of which have been cloned and sequenced . Each of the four derived amino acid sequences have more than 60% similarity to the corresponding polypeptides of man, rat, Escherichia coli and Salmonella typhimurium . The PRS1 gene maps on chromosome XI, PRS2 on chromosome V, PRS3 on chromosome VIII and PRS4 on chromosome II . One member of this gene family, PRS1, contains a region of non-homology (NHR) shown by cDNA cloning and sequencing not to be an intron . The results presented here suggest that the presence of this NHR is not detrimental to the function of the gene . To date the possibility of protein splicing can be neither proven nor disputed.

Vaccine, 1994 Aug, 12(11), 1004 - 11
Construction and evaluation of an expression vector allowing the stable expression of foreign antigens in a Salmonella typhimurium vaccine strain; Tijhaar EJ et al.; Salmonella strains have great potential as live carriers of heterologous antigens to induce immunity against a variety of infectious diseases . However, the amount of heterologous antigen required to induce an adequate immune response may be toxic for the bacterium and result in cell death, overattenuation or loss of expression of the heterologous antigen . To solve this problem an expression vector was developed with a strong promoter located on a DNA fragment which is inverted at random . Antigen is only expressed in one particular orientation of the promoter . Thus a bacterial population harbouring the plasmid will consist of a subpopulation which does not produce heterologous antigen, and is therefore not affected in growth, persistence and dissemination within the host . Further, this non-producing population will continuously segregate antigen-producing bacteria . To evaluate the system, CtxB was used as a model antigen . Analysis of the plasmid DNA isolated from Salmonella revealed a selection against the promoter orientation that directs transcription of the ctxB gene . In spite of this, the vector was stably maintained in vivo and induced CtxB-specific IgA and IgG in mice . These results indicate that this kind of expression vector may offer a solution to the problem of unstable expression of foreign antigens in live bacterial vaccine strains.

Poult Sci, 1994 Aug, 73(8), 1327 - 33
The use of ultraviolet radiation to reduce Salmonella and psychrotrophic bacterial contamination on poultry carcasses; Wallner-Pendleton EA et al.; Broiler carcasses were subjected to ultraviolet (UV) energy (doses range from 82,560 to 86,400 muWs/cm2) at wavelength of 253.7 nm to evaluate the potential of this treatment for improving the microbiological quality of broiler carcasses . Broiler chicken halves were inoculated with a marker strain of Salmonella typhimurium 5 min prior to treatment . A 61% reduction in viable S . typhimurium was observed in UV-treated chicken halves as compared with untreated halves . The UV energy treatment had no deleterious effects on color (Hunter L, aL, or bL) or 2-thiobarbituric acid (TBA) values . After 10 d of storage at 7 C, TBA values of thigh meat were 1.3 mg malonaldehyde/kg meat compared with 1.7 for controls . Psychrotrophic bacteria populations were not appreciably altered by UV treatment when their numbers were compared with bacterial counts obtained from untreated chicken halves held for 10 d at 7 C . This study suggests that UV radiation can reduce Salmonella surface contamination without negatively affecting carcass color or increasing rancidity of the meat.

Poult Sci, 1994 Aug, 73(8), 1241 - 8
Effect of ochratoxin A on Salmonella-challenged broiler chicks; Elissalde MH et al.; Poultry products represent a significant reservoir of Salmonella typhimurium . Ochratoxin A, a mycotoxin and natural contaminant of poultry feedstuffs, produces detrimental effects on the immune and other systems of the broiler chick . Because poultry products are possible sources of S . typhimurium contamination that can potentially infect humans, there is a need to know whether ochratoxin A can alter the growth of Salmonella in poultry . We investigated the pathological alterations of young male broiler chicks by S . typhimurium in the presence (3.0 mg/kg) or absence of ochratoxin A in the diet . Ochratoxin A alone in the diet decreased the body weight and increased the relative organ weights of the liver, kidney, gizzard, spleen, pancreas, and proventriculus . It did not affect the heart and bursa of Fabricius . The mycotoxin altered the serum concentrations of proteins, enzymes, calcium and phosphate salts, normal tissue constituents, and catabolic metabolites in a pattern that would suggest damage to skeletal muscle, liver, kidney, pancreas, and bone . Birds fed diets containing ochratoxin A had microcytic and hypochromic erythrocytes and a decrease in phytohemagglutin- and concanavalin A-stimulated blastogenesis . Salmonella typhimurium alone had no affect on the variables measured except for a decrease in body weight . With the exception of an increase in mortality (13.2%, a significant synergistic interaction) and decrease in body weight, Salmonella in combination with ochratoxin A did not alter the values of the remaining variables measured from those measured in the ochratoxin A diet alone . Cecal colony count of S . typhimurium was not affected by treatment with ochratoxin A.

J Dairy Sci, 1994 Aug, 77(8), 2272 - 80
Effect of Re-17 mutant Salmonella typhimurium bacterin toxoid on clinical coliform mastitis; McClure AM et al.; The objective of this study was to test the hypothesis that the incidence and severity of clinical coliform mastitis could be decreased by Re-17 mutant Salmonella typhimurium bacterin toxoid . Holstein-Friesian cows from two Arizona dairies were selected for this study based on July through November projected calving dates; peak lactation occurred during the period of highest rainfall and peak environmental stress . The cows were randomly assigned to either a vaccinate or a control group, and 1292 cows were paired by herd, parity, calving date, and milk yield . The 646 vaccinates were injected twice during the third trimester of pregnancy with an Re-17 mutant S . typhimurium bacterin toxoid, and the 646 controls were not vaccinated . Vaccinated cows had significantly fewer clinical cases of coliform mastitis with positive coliform cultures and had lower culling rate from coliform mastitis than control cows during the first 5 mo of lactation . During the same period, the mortality rate from clinical coliform mastitis was 75% less in the vaccinated clinical coliform mastitic group than in the control group . Incidence of mastitis increased with advancing parity . The Re-17 mutant Salmonella typhimurium bacterin toxoid provided cross-protection against coliform mastitis; incidence and severity of clinical coliform mastitis were significantly lowered during the first 5 mo of lactation.

J Appl Bacteriol, 1994 Aug, 77(2), 149 - 54
Thermostability of bacterial luciferase expressed in different microbes; Mackey BM et al.; Bacterial luciferase was used to investigate the relationship between the thermostability of a cytoplasmic reporter molecule and cellular heat resistance . The luciferase activity of Vibrio fischeri was expressed in strains of Escherichia coli, Salmonella typhimurium, Listeria monocytogenes and Brochothrix thermosphacta following transformation with plasmid pSP13 carrying the luxAB genes . The thermostability of intracellular luciferase varied depending on the organism in which it was expressed, but was not related to the cellular heat resistance of the different organisms . Addition of xylitol to the heating medium protected against loss of viability and inactivation of intracellular luciferase . Glycerol also protected against loss of viability but was less effective at preventing thermal denaturation of luciferase.

Hum Exp Toxicol, 1994 Aug, 13(8), 558 - 62
Glutathione deficiency does not elevate susceptibility of bacteria to the mutagenicity of chlorinated humic acids; Ubom GA et al.; 1 . Rat liver 9,000 g supernatant protected against the mutagenic effect of chlorinated hydrophilic macromolecular humic acids (CHMA) in Salmonella typhimurium strain TA100 . 2 . Protection against mutagenicity of CHMA was mediated by glutathione and was partially dependent on glutathione S-transferase activity . 3 . In contrast to the above findings, CHMA showed lower mutagenicity in Salmonella typhimurium and Escherichia coli strains of bacteria that are deficient in glutathione compared to their mutagenicity in parental (glutathione-rich) bacterial strains . 4 . Glutathione-deficient cells do not provide test systems with elevated sensitivity for the detection of mutagenic chlorinated humic substances.

J Mol Evol, 1994 Aug, 39(2), 129 - 33
Genetic variation in IncI1-ColIb plasmids; Ayala FJ et al.; Nucleotide sequences of portions of three plasmid genes (cib, cir, and abi) present in IncI1-ColIb colicin plasmids obtained from strains of Salmonella typhimurium isolated in either 1974 (Barker strains) or between 1935 and 1941 (Murray strains) were examined along with sequences of the chromosomal gene for 6-phosphogluconate dehydrogenase (gnd) . Our principal findings were: (1) The plasmid genes were virtually identical to those in IncI1-ColIb plasmids from E . coli, suggesting that Salmonella and E . coli share overlapping pools of these plasmids . (2) The plasmid genes were much less polymorphic than gnd or any other known chromosomal gene from Salmonella, further suggesting horizontal transfer with rapid transmission and turnover . (3) No characteristic differences were found in either the plasmid genes or the chromosomal gene between the 1974 isolates and the Murray strains, indicating that these plasmids have been stable for at least several decades . (4) There was an excess of amino-acid replacement polymorphisms, relative to synonymous polymorphisms, in the plasmid genes, which is consistent with the hypothesis of diversifying selection among colicin-producing plasmid families . (5) The abi (abortive infection) gene present in each of the plasmids contained two single-nucleotide insertions relative to the published sequence . These result in a putative abi protein of 114 amino acids instead of 89.

Microbiology, 1994 Aug, 140 ( Pt 8), 1977 - 82
Identification and characterization of a putative serine protease expressed in vivo by Mycobacterium avium subsp . paratuberculosis; Cameron RM et al.; A putative serine protease expressed in vivo by Mycobacterium avium subsp . paratuberculosis was isolated from a lambda gt11 genomic expression library by screening with serum from a naturally infected sheep . The gene was contained in two overlapping clones, which were shown by antibody elution to encode a protein of 34 kDa in M . a . paratuberculosis . The clones were sequenced and database searches detected a motif identical to the active serine site in trypsin, and 30% homology to the putative serine proteases (HtrA proteins) of Escherichia coli, Salmonella typhimurium, Brucella abortus and Rochalimaea henselae.

Eur J Epidemiol, 1994 Aug, 10(4), 367 - 71
Salmonella typhimurium and Salmonella enteritidis: changing epidemiology from 1973 until 1992; Le Bacq F et al.; A retrospective analysis, covering a period of 20 years, of 2049 primo-isolates of Salmonella typhimurium and 343 primo-isolates of Salmonella enteritidis was performed at our laboratory . In 1991, S . enteritidis (43.8%) outnumbered S . typhimurium (32.1%) as the most frequently isolated Salmonella . The age group distribution of S . enteritidis yielded peaks in the under-five and above-60 year age groups, whereas S . typhimurium only peaked in the under-five age group . S . enteritidis appeared to be twice as invasive as S . typhimurium . Both serotypes were more invasive in middle and older age groups than in under-fives.

Xenobiotica, 1994 Aug, 24(8), 713 - 27
Biotransformation of furaltadone by pig hepatocytes and Salmonella typhimurium TA 100 bacteria, and the formation of protein-bound metabolites; Hoogenboom LA et al.; 1 . The major metabolite resulting from the biotransformation of furaltadone (5-morpholinomethyl-3-{5-nitrofurfurylidene-amino}-2-oxazoli dinone) by pig hepatocytes was shown to result from the N-oxidation of the tertiary nitrogen in the morpholino-ring, leaving the nitrofuran ring unchanged . 2 . No evidence could be obtained for the formation of an open-chain cyano-metabolite, a minor metabolite in the case of the related nitrofuran drug furazolidone (N-(5-nitro-2-furfurylidene)-3-amino-2-oxazolidinone) . This metabolite was the major metabolite, following incubation of furaltadone and furazolidone with Salmonella typhimurium bacteria . 3 . The N-oxide was not further metabolized by pig hepatocytes or bacteria, and gave negative test results in the Ames-test (TA 100, no S9-mix) at the highest tested dose of 1 microgram/plate . Furaltadone gave a positive result at 10 ng/plate . 4 . The biotransformation of both drugs by pig hepatocytes and bacteria resulted in the formation of protein-bound metabolites, with no clear quantitative differences between the two drugs . The intact 3-amino-2-oxazolidinone (AOZ) and 5-morpholinomethyl-3-amino-2-oxazolidinone (AMOZ) side-chains of furazolidone and furaltadone, respectively, could be released from these metabolites by mild acid treatment . 5 . Hepatocytes incubated with the AMOZ side-chain of furaltadone showed a decreased monoamine oxidase activity at high dose levels (IC50 3.7 mM), whereas exposure to the AOZ side-chain of furazolidone resulted in a clear inhibition at 10,000-fold lower concentrations (IC50 0.5 microM) . In the presence of 1% dimethylsulphoxide (DMSO), the MAO-inhibition by AMOZ and especially AOZ was remarkably reduced . 6 . It is concluded that protein-bound metabolites containing an intact and releasable side-chain might be present in tissues of animals treated with furaltadone . However, these residues might be of less toxicological concern than those of furazolidone.

FEMS Immunol Med Microbiol, 1994 Aug, 9(2), 125 - 33
A cell-free Salmonella typhimurium extract modulates interleukin-2 (IL-2) receptor expression but not IL-2-stimulated responses of murine splenic lymphocytes; Matsui K et al.; In a previous study, we observed that a cell-free Salmonella typhimurium extract induced suppression of mitogen-induced T-cell proliferation and this suppression involved non-responsiveness of T-cells to interleukin-2 (IL-2) . In this study, we found that a cell-free S . typhimurium extract modulated IL-2 receptor (IL-2R) expression on phytohemagglutinin (PHA)-stimulated murine spleen cells and this was a mechanism of T-cell non-responsiveness to IL-2, but did not affect IL-2 binding to IL-2R and the consequent responses . Western blotting using anti-phosphotyrosine antibodies showed that IL-2R-mediated tyrosine phosphorylation of protein substrates in PHA-activated murine splenic T-cells, which express a high-affinity IL-2R (alpha- and beta-chains), was not affected by treatment with the S . typhimurium cell-free extract . Furthermore, PHA-activated spleen T-cells responded to recombinant IL-2 and this was not inhibited by the extract . Surprisingly, IL-2R expression was augmented by treatment with the extract, although this was independent of IL-2 production . These results suggest that the suppression of T-cell proliferation induced by the Salmonella cell-free extract was associated with augmentation of IL-2R expression, rather than down-regulation of the IL-2 response . This may be a mechanism responsible for the Salmonella extract-evoked suppression of mitogen-induced T-cell proliferation.

J Vet Med Sci, 1994 Aug, 56(4), 765 - 6
Serum concentration of a bovine mannan-binding protein reactive with a Ra chemotype strain of Salmonella typhimurium: no significant changes associated with mastitis; Sugii S et al.; The serum concentration of a bovine mannan-binding protein reactive with a Ra chemotype strain of Salmonella Typhimurium in sera from cows with or without mastitis were determined by sandwich enzyme-linked immunosorbent assay . From the results obtained for 10 healthy cows aged 2 to 7 years, mean +/- SD serum concentrations of MBP in bovine sera were 77.5 +/- 33.1 micrograms/ml . Concentrations in 7 healthy heifer calves aged 6 months were 52.2 +/- 10.2 micrograms/ml, whereas those in 7 healthy bullocks 65.8 +/- 21.8 micrograms/ml . Concentrations in 4 cows aged 4 to 7 years with mastitis were slightly lower (34 to 72 micrograms/ml) . After recovery, the serum concentrations rose to the normal concentrations in healthy cows . These findings indicate that the serum concentrations of bovine MBP decrease during mastitis, suggesting that bovine MBP may not be an acute phase reactant.

J Vet Med Sci, 1994 Aug, 56(4), 747 - 51
Identification and carbohydrate specificity of a chicken serum mannan-binding protein reactive with a Ra chemotype strain of Salmonella typhimurium; Sugii S et al.; To compare a chicken serum mannan-binding protein (MBP) with a chicken Ra-reactive factor (RaRF), both serum proteins were isolated by combination of different chromatography . In SDS-PAGE, both purified proteins were resolved into a major protein band with a molecular weight of 34,000 and a few minor protein bands . In Western blotting with both purified proteins, only a single band with a molecular weight of 34,000 was detected with anti-chicken serum MBP antibody, indicating that the major protein band of chicken serum MBP and RaRF are antigenically identical . The combining sites of these two proteins were most reactive with N-acetylmannosamine followed by mannose, N-acetylglucosamine, and L-fucose . These findings suggest that chicken serum MBP may be identical to its corresponding bactericidal factor RaFR in terms of antigenicity and monosaccharide specificity.

Mol Microbiol, 1994 Aug, 13(3), 541 - 53
Vitamin B12 repression of the cob operon in Salmonella typhimurium: translational control of the cbiA gene; Richter-Dahlfors AA et al.; Expression of the cob operon is repressed by B12 via a post-transcriptional control mechanism which requires sequence elements within the leader region of the mRNA and the first gene of the operon, the cbiA gene . Here we show that B12 repression of cbiA gene expression occurs at the level of translation initiation through sequestration of the ribosomal binding site (rbs) in an RNA hairpin . Analysis of mutations that destabilize or restabilize the secondary structure demonstrates that folding of the hairpin is essential for repression . The existence of the hairpin was confirmed by a secondary structure analysis of RNA from the wild type and three mutants . Deletions that remove the upstream part of the leader confer a drastic reduction in translation efficiency . This low-level translation is caused by the hairpin, as indicated by the finding that suppressor mutations that destabilize the hairpin restore efficient translation . Thus, the native upstream RNA functions as a translation enhancer and acts to relieve the hairpin's inhibitory effect on translation initiation . The inhibitory effect of the hairpin was confirmed by a ribosomal toeprinting analysis . We propose that the translational control of the cbiA gene mediates repression of the entire cob operon.

Mol Microbiol, 1994 Aug, 13(3), 525 - 30
Construction and characterization of isogenic O-antigen variants of Salmonella typhi; Hone DM et al.; A 7.5 kb KpnI-generated fragment, from within the rfb cluster of Salmonella typhimurium LT2 that encodes abequose synthase (the rfbJ gene) which is necessary for O4 antigen synthesis, and flanking sequences, was inserted into a suicide vector . Using allelic exchange techniques, these rfb sequences of S . typhimurium were integrated into the rfb clusters of wild-type Salmonella typhi Vi-positive strain ISP 1820 (i.e . serotype O9,12; Vi+; H-d), S . typhi Vinegative strain H400 (i.e . serotype O9,12; Vi-; H-d), and a double aro mutant of S . typhi ISP 1820, strain CVD 906, resulting in the isolation of strains H325, H404 and CVD 906-O4, respectively . Immunoblot analysis of lipopolysaccharide (LPS) purified from H325, H404 and CVD 906-O4 demonstrated that these strains express the O4 antigen (an abequose residue) in place of the O9 antigen (a tyvelose residue) in the LPS molecule . Hence, the serotype of H325 is O4,12; Vi+; H-d and the serotype of H404 is O4,12; Vi-; H-d . DNA hybridization analysis of chromosomal DNA from H325, H404 and CVD 906-O4 confirmed that a precise recombination event within sequences flanking rfbSE of S . typhi (which encodes the enzymes necessary for cytidine diphosphate-tyvelose synthesis) resulted in replacement of rfbSE with rfbJ (which encodes abequose synthase and is necessary for O4 synthesis) of S . typhimurium in strains H325, H404 and CVD 906-O4 . The resistance of each strain to the bactericidal effects of guinea-pig serum (GPC) was assessed . Whereas ISP 1820, H325 and H404 exhibit similar resistance patterns in GPC, strain H400 is sensitive to the bactericidal effects of GPC.(ABSTRACT TRUNCATED AT 250 WORDS)

Chemosphere, 1994 Aug, 29(3), 441 - 50
Sources and seasonal variability of mutagenic agents in the Barcelona City aerosol; Bayona JM et al.; Organic extracts (dichloromethane) isolated from airborne particulate matter, collected in two sampling sites located in the Barcelona City, were mutagenic in the Salmonella typhimurium (TA98 +/-S9) bioassay . The highest direct-acting mutagenicity (69-78 rev m-3) was detected during fall and spring, which corresponds to the highest levels of mutagenic nitroarenes (248 to 350 pg m-3) . On the other hand, the highest level of indirect-acting mutagenicity was obtained in summer, paralleling with the highest concentrations of polycyclic aromatic ketones and polycyclic aromatic quinones . Furthermore, the sources of PAH in the urban particulate matter were estimated from the ratio of the less reactive components (i.e . benzofluranthenes/benzo{e}pyrene, indeno{1,2,3-cd}pyrene/benzo{ghi}perylene, methylphenantherenes/phenanthrene) and reflected a predominance of pyrolytic mobile sources (i.e . vehicular emissions) . Nevertheless, a contribution of stationary sources in winter was also apparent . Finally, the seasonal variability of polycyclic aromatic ketones, quinones, aromatic lactones and aldehydes reflected a major contribution of the atmospheric transformation processes from related PAH rather than a direct emission from combustion sources.

Mutat Res, 1994 Aug 1, 309(1), 17 - 28
Mutation induction by accelerated heavy ions in bacteria; Kozubek S et al.; Induction of lacI- forward mutations in Escherichia coli Ymcl and his(-)-->his+ reversions in Salmonella typhimurium TA102 was investigated after irradiation with heavy ions in the range of Z = 1-36 . Particle specific energies (E) were in the range of 1-600 MeV/u . A strong dependence of the mutation induction cross-section (sigma m) on both particle energy and LETinfinity was observed . The results suggest that two different ranges of LETinfinity can be distinguished . In the range of high LETinfinity (> 100 keV/micron) sigma m increases with increasing specific particle energy if LETinfinity is kept constant (Fe ions as compared with carbon ions or alpha-particles) . In the range of low LETinfinity (< 100 keV/micron) sigma m decreases with increasing energy (Ne ions as compared with He ions).

J Bacteriol, 1994 Aug, 176(16), 4858 - 64
apbA, a new genetic locus involved in thiamine biosynthesis in Salmonella typhimurium; Downs DM et al.; In Salmonella typhimurium, the synthesis of the pyrimidine moiety of thiamine can occur by utilization of the first five steps in de novo purine biosynthesis or independently of the pur genes through the alternative pyrimidine biosynthetic, or APB, pathway (D . M . Downs, J . Bacteriol . 174:1515-1521, 1992) . We have isolated the first mutations defective in the APB pathway . These mutations define the apbA locus and map at 10.5 min on the S . typhimurium chromosome . We have cloned and sequenced the apbA gene and found it to encode a 32-kDa polypeptide whose sequence predicts an NAD/flavin adenine dinucleotide-binding pocket in the protein . The phenotypes of apbA mutants suggest that, under some conditions, the APB pathway is the sole source of the pyrimidine moiety of thiamine in wild-type S . typhimurium, and furthermore, the pur genetic background of the strain influences whether this pathway can function under aerobic and/or anaerobic growth conditions.

Mutat Res, 1994 Aug, 322(2), 133 - 40
Bacterial mutagenicity of eight medicinal herbs from Zimbabwe; Sohni YR et al.; Eight plants traditionally used as medicines in Zimbabwe were evaluated for mutagenicity . The required plant parts were dried, powdered and extracted in a Soxhlet apparatus with distilled water . The extracts were tested using Salmonella typhimurium strains TA97a, TA98, TA100 and TA102 . The results indicate five of these extracts are nonmutagenic in the assay while three of the extracts were found to be mutagenic . The three plant extracts, namely those of Combretum erythrophyllum, Gnidia kraussiana and Barlerii randii, were found to be mutagenic to strain TA102 . Furthermore, the extract of C . erythrophyllum was also mutagenic to strain TA100 . The presence of S9 mix appeared to diminish the mutagenicity of the extracts except in the case of C . erythrophyllum, the mutagenicity of which was enhanced in strain TA100 . These results assume importance in view of the fact that these plants are used as therapeutic agents.

Cancer Lett, 1994 Jul 29, 82(2), 217 - 24
Effects of isothiocyanate alkyl chain-length on hamster liver cytochrome P-450 activity; Hamilton SM et al.; The tobacco-specific nitrosamine 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK) is metabolized by various isozymes of cytochrome P-450 present in microsomes . In this study, we examined the effects of the isothiocyanate homologues, phenyl isothiocyanate (PITC), benzyl isothiocyanate (BITC), phenethyl isothiocyanate (PEITC) and phenylpropyl isothiocyanate (PPITC) on the mutagenicity and in vitro metabolism of NNK by Syrian golden hamster liver microsomes and on the in vitro microsomal metabolism of testosterone . Each isothiocyanate compound inhibited N-oxidation and alpha-hydroxylation reactions of NNK that, except for PITC, correlated with an inhibition of microsomal-mediated mutagenicity of NNK in Salmonella typhimurium TA1535 . Each isothiocyanate also inhibited cytochrome P-450-mediated hydroxylation reactions of the metabolism of testosterone . In general, the inhibitory potency of the isothiocyanates corresponded with the length of the alkyl chain of the compound . Our data support the ability of isothiocyanates to inhibit the activity of a number of isozymes of cytochrome P-450.

Mol Gen Genet, 1994 Jul 25, 244(2), 216 - 8
The gene fimU affects expression of Salmonella typhimurium type 1 fimbriae and is related to the Escherichia coli tRNA gene argU; Swenson DL et al.; The gene fimU, located on a recombinant plasmid carrying the Salmonella typhimurium type 1 fimbrial gene cluster is closely related to the Escherichia coli tRNA gene argU . The fimU gene complements an E . coli argU mutant that is a P2 lysogen, thereby allowing the phage P4 to grow in this strain but preventing the growth of phage lambda . In addition, fimU was shown to be involved in fimbrial expression since transformants of the E . coli argU mutant could produce fimbriae only in the presence of fimU but not in its absence, whereas in an E . coli argU+ strain fimbriation did not require the fimU gene.

Proc Natl Acad Sci U S A, 1994 Jul 19, 91(15), 7017 - 21
Cloning and sequencing of thiol-specific antioxidant from mammalian brain: alkyl hydroperoxide reductase and thiol-specific antioxidant define a large family of antioxidant enzymes; Chae HZ et al.; A cDNA corresponding to a thiol-specific antioxidant enzyme (TSA) was isolated from a rat brain cDNA library with the use of antibodies to bovine TSA . The cDNA clone encoded an open reading frame capable of encoding a 198-residue polypeptide . The rat and yeast TSA proteins show significant sequence homology to the 21-kDa component (AhpC) of Salmonella typhimurium alkyl hydroperoxide reductase, and we have found that AhpC exhibits TSA activity . AhpC and TSA define a family of > 25 different proteins present in organisms from all kingdoms . The similarity among the family members extends over the entire sequence and ranges between 23% and 98% identity . A majority of the members of the AhpC/TSA family contain two conserved cysteines . At least eight of the genes encoding AhpC/TSA-like polypeptides are found in proximity to genes encoding other oxidoreductase activities, and the expression of several of the homologs has been correlated with pathogenicity . We suggest that the AhpC/TSA family represents a widely distributed class of antioxidant enzymes . We also report that a second family of proteins, defined by the 57-kDa component (AhpF) of alkyl hydroperoxide reductase and by thioredoxin reductase, has expanded to include six additional members.

Mutat Res, 1994 Jul 16, 308(2), 215 - 22
The effects of antioxidants and enzymes involved in glutathione metabolism on mutagenesis by glutathione and L-cysteine; Stark AA et al.; The effects of small molecular weight antioxidants and antioxidant enzymes on the mutagenicities of glutathione (GSH) and L-cysteine were studied in Salmonella typhimurium strain TA102 . GSH and cysteine mutagenesis were inhibited by antioxidants and radical scavengers such as alpha-tocopherol, Trolox C, butylated hydroxyanisole (BHA), and retinyl acetate . Superoxide dismutase (SOD) had no effect, but catalase and horseradish peroxidase (HRP) inhibited mutagenesis . The heat-denatured enzymes had no effect on mutagenesis . Cysteine mutagenesis was enhanced by native and by heat-denatured rat-kidney post-mitochondrial supernatant, and by ferric ions . H2O2 and the H2O2-generating system of glucose-glucose oxidase (GOX) were mutagenic in TA102 . Synergistic increases in mutagenesis were obtained in systems containing combinations of GSH or cysteine, with either H2O2 or the H2O2-generating system of glucose-GOX . GSH peroxidase (GPX) had no effect on mutagenesis of GSH or of H2O2, whereas the synergistic increase in mutagenesis by a combination of GSH and H2O2 was effectively inhibited by GPX . The results suggest strongly that, at least in biochemically-defined systems, GSH and cysteine mutagenesis are oxidative in nature, and involve reactive forms of oxygen and/or other radicals.

Mutat Res, 1994 Jul 16, 308(2), 135 - 41
Mutagenicity of K-region oxides and imines of chrysene, benzo{c}phenanthrene and benzo{g}chrysene in Salmonella typhimurium; Glatt H et al.; The K-region oxides and imines of chrysene, benzo{c}phenanthrene and benzo{g}chrysene were investigated for mutagenicity in Salmonella typhimurium TA98 and TA100, using two different exposure media . All six compounds were mutagenic under all four experimental conditions . The imines were 17-3800 times more potent than the corresponding oxides . Omission of KCl (125 mM) from the exposure medium resulted in enhanced mutagenic effects . The enhancement was stronger in strain TA98 (3.1-5.2-fold) than in strain TA100 (1.3-2.5-fold), suggesting an influence on the bacteria rather than on the chemicals.

Science, 1994 Jul 15, 265(5170), 383 - 6
Crystal structure of P22 tailspike protein: interdigitated subunits in a thermostable trimer; Steinbacher S et al.; The tailspike protein (TSP) of Salmonella typhimurium phage P22 is a part of the apparatus by which the phage attaches to the bacterial host and hydrolyzes the O antigen . It has served as a model system for genetic and biochemical analysis of protein folding . The x-ray structure of a shortened TSP (residues 109 to 666) was determined to a 2.0 angstrom resolution . Each subunit of the homotrimer contains a large parallel beta helix . The interdigitation of the polypeptide chains at the carboxyl termini is important to protrimer formation in the folding pathway and to thermostability of the mature protein.

J Immunol, 1994 Jul 1, 153(1), 212 - 9
Treatment of mice with IL-1 before infection increases resistance to a lethal challenge with Salmonella typhimurium . The effect correlates with the resistance allele at the Ity locus; Morrissey PJ et al.; Administration of a single dose of IL-1 alpha to various strains of mice approximately 16 h before a lethal infection with Salmonella typhimurium resulted in a significant augmentation of survival in most Ityr, but in no Itys strains of mice . Lower numbers of bacteria were observed in the liver and spleen in response to IL-1 pretreatment shortly after infection in all Ityr strains of mice tested, including the congenic C.D2-Ityr mice . Treatment with IL-1 alpha after infection had no effect on survival in either Ityr or Itys mice . A combination of IL-1 alpha pretreatment with IL-1 alpha post-treatment did not increase survival over the effect of IL-1 alpha pretreatment alone in Ityr mice and did not increase the survival of the Itys mice . The combination of IL-1 alpha pretreatment with GM-CSF post-treatment was effective in Ityr but not in Itys strains of mice . Thus, IL-1 alpha pretreatment enhances the resistance of Ityr, but not Itys strains of mice to a lethal challenge with S . typhimurium by up-regulating antibacterial mechanisms shortly after infection.

Mol Plant Microbe Interact, 1994 Jul-Aug, 7(4), 488 - 97
Characterization of the hrpJ and hrpU operons of Pseudomonas syringae pv . syringae Pss61: similarity with components of enteric bacteria involved in flagellar biogenesis and demonstration of their role in HarpinPss secretion; Lidell MC et al.; The hrp/hrmA gene cluster of Pseudomonas syringae pv . syringae Pss61 has been shown to form a minimum genetic unit sufficient to enable nonpathogenic bacteria, such as Escherichia coli, to elicit the hypersensitive response associated with disease resistance . The biochemical functions of most of these genes have not been established . The nucleotide sequence of a 4.3-kb SstI-BglII fragment carrying hrp apparent translational units V, VI, and VII revealed one partial open reading frame (ORF) and five complete ORFs producing 35,126-, 48,866-, 17,308-, 20,482-, and 26,364-Da gene products (hrpJ3, J4, J5, U1, U2, respectively) . The production of these proteins was confirmed by using T7 RNA polymerase-directed expression . The partial ORF was found to be identical to the C terminus of HrpJ2 . The absence of apparent transcriptional terminators and promoters between hrpI (hrpJ2), hrpJ3, hrpJ4, and hrpJ5 together with the observation that the HrpL-dependent hrpJ promoter directs expression of hrpJ3-J5 indicates that these genes form a single operon controlled by the HrpL-dependent hrpJ promoter . A second HrpL-dependent promoter consensus sequence was also identified upstream of hrpU1 and demonstrated to function as a HrpL-dependent promoter, thus indicating that hrpU1, hrpU2, and additional downstream genes may be part of a second operon . The deduced product of hrpJ3 exhibits similarity to FliG of Salmonella typhimurium, a cytoplasmic protein that regulates flagellar rotation and biogenesis . HrpJ4 shares extensive similarity with the FliI family of ATPase-like proteins and retains the known functional domains conserved among this family of proteins . HrpJ5 has properties similar to the S . typhimurium FliJ . Neither HrpU1 nor HrpU2 exhibit significant similarity to known proteins . Secretion of HarpinPss by E . coli MC4100 transformants carrying pHIR11::TnphoA derivatives was blocked in hrpJ4, J5, and U2 mutants . In view of the previously reported similarity of HrpJ2 to the LcrD super-family that includes FlhA, these results predict that the gene products of the hrpJ and hrpU operons form an inner membrane complex for translocation of proteins similar to that used by the flagellar biogenesis system of S . typhimurium.

J Bacteriol, 1994 Jul, 176(14), 4260 - 8
Evidence for two NAD kinases in Salmonella typhimurium; Cheng W et al.; The electron-carrying cofactor NADP is formed by phosphorylation of NAD . A strategy for the isolation of NAD kinase mutants revealed two classes of temperature-sensitive mutations, nadF and nadG, mapping at min 13 and 72 of the Salmonella chromosome . Both mutant types grew on nutrient broth at both 30 and 42 degrees C but on minimal medium showed a temperature-sensitive growth defect which was not corrected by any of the single nutritional supplements tested . A nadF deletion mutant grew on nutrient broth but not on minimal medium . A double mutant with the nadF deletion and a nadG(Ts) mutation showed temperature-sensitive growth on all media . We propose that Salmonella typhimurium has two NAD kinases, one encoded by the nadF and one by the nadG gene . This is supported by the fact that temperature-sensitive mutants of both genes produce kinase activity with altered heat stability . Results suggest that either one of two NAD kinases is sufficient for growth on rich medium, but that both are needed for growth on minimal media . Enzyme assays show that the nadF gene is responsible for about 70% of total NAD kinase activity, and that the nadG gene dictates the remaining 30% . While testing nutritional phenotypes of nadF and nadG mutants, we found that the biosynthetic intermediate, quinolinic acid (QA) inhibited growth of nadF mutants on nutrient broth . This suggested that the NadG enzyme might be inhibited by QA . Enzyme assays demonstrated that QA inhibits the NadG but not the NadF enzyme . This suggests the existence of a regulatory mechanism which controls NADP levels.

J Bacteriol, 1994 Jul, 176(14), 4210 - 8
Mutational activation of CheA, the protein kinase in the chemotaxis system of Escherichia coli; Tawa P et al.; In Escherichia coli and Salmonella typhimurium, appropriate changes of cell swimming patterns are mediated by CheA, an autophosphorylating histidine protein kinase whose activity is regulated by receptor/transducer proteins . The molecular mechanism underlying this regulation remains unelucidated but may involve CheA shifting between high-activity and low-activity conformations . We devised an in vivo screen to search for potential hyperkinase variants of CheA and used this screen to identify two cheA point mutations that cause the CheA protein to have elevated autokinase activity . Each point mutation resulted in alteration of proline 337 . In vitro, CheA337PL and CheA337PS autophosphorylated significantly more rapidly than did wild-type CheA . This rate enhancement reflected the higher affinities of the mutant proteins for ATP and an increased rate constant for acquisition by CheA of the gamma-phosphoryl group of ATP within a kinetically defined CheA.ATP complex . In addition, the mutant proteins reacted with ADP more rapidly than did wild-type CheA . We considered the possibility that the mutations served to lock CheA into an activated signaling conformation; however, we found that both mutant proteins were regulated in a normal fashion by the transducer Tsr in the presence of CheW . We exploited the activated properties of one of these mutants to investigate whether the CheA subunits within a CheA dimer make equivalent contributions to the mechanism of trans phosphorylation . Our results indicate that CheA trans phosphorylation may involve active-site residues that are located both in cis and in trans to the autophosphorylation site and that the two protomers of a CheA dimer make nonequivalent contributions in determining the affinity of the ATP-binding site(s) of CheA.

J Bacteriol, 1994 Jul, 176(13), 4092 - 103
Salmonella recD mutations increase recombination in a short sequence transduction assay; Miesel L et al.; We have identified recD mutants of Salmonella typhimurium by their ability to support growth of phage P22 abc (anti-RecBCD) mutants, whose growth is prevented by normal host RecBCD function . As in Escherichia coli, the recD gene of S . typhimurium lies between the recB and argA genes at min 61 of the genetic map . Plasmids carrying the Salmonella recBCD+ genes restore ATP-dependent exonuclease V activity to an E . coli recBCD deletion mutant . The new Salmonella recD mutations (placed on this plasmid) eliminate the exonuclease activity and enable the plasmid-bearing E . coli deletion mutant to support growth of phage T4 gene 2 mutants . The Salmonella recD mutations caused a 3- to 61-fold increase in the ability of a recipient strain to inherit (by transduction) a large inserted element (MudA prophage; 38 kb) . In this cross, recombination events must occur in the short (3-kb) sequences that flank the element in the 44-kb transduced fragment . The effect of the recD mutation depends on the nature of the flanking sequences and is likely to be greatest when those sequences lack a Chi site . The recD mutation appears to minimize fragment degradation and/or cause RecBC-dependent recombination events to occur closer to the ends of the transduced fragment . The effect of a recipient recD mutation was eliminated if the donor P22 phage expressed its Abc (anti-RecBC) function . We hypothesize that in standard (high multiplicity of infection) P22-mediated transduction crosses, recombination is stimulated both by Chi sequences (when present in the transduced fragment) and by the phage-encoded Abc protein which inhibits the host RecBCD exonuclease.

J Bacteriol, 1994 Jul, 176(13), 3911 - 9
Changes of ploidy during the Azotobacter vinelandii growth cycle; Maldonado R et al.; The size of the Azotobacter vinelandii chromosome is approximately 4,700 kb, as calculated by pulsed-field electrophoretic separation of fragments digested with the rarely cutting endonucleases SpeI and SwaI . Surveys of DNA content per cell by flow cytometry indicated the existence of ploidy changes during the A . vinelandii growth cycle in rich medium . Early-exponential-phase cells have a ploidy level similar to that of Escherichia coli or Salmonella typhimurium (probably ca . four chromosomes per cell), but a continuous increase of DNA content per cell is observed during growth . Late-exponential-phase cells may contain > 40 chromosomes per cell, while cells in the early stationary stage may contain > 80 chromosomes per cell . In late-stationary-phase cultures, the DNA content per cell is even higher, probably over 100 chromosome equivalents per cell . A dramatic change is observed in old stationary-phase cultures, when the population of highly polyploid bacteria segregates cells with low ploidy . The DNA content of the latter cells resembles that of cysts, suggesting that the process may reflect the onset of cyst differentiation . Cells with low ploidy are also formed when old stationary-phase cultures are diluted into fresh medium . Addition of rifampin to exponential-phase cultures causes a rapid increase in DNA content, indicating that A . vinelandii initiates multiple rounds of chromosome replication per cell division . Growth in minimal medium does not result in the spectacular changes of ploidy observed during rapid growth; this observation suggests that the polyploidy of A . vinelandii may not exist outside the laboratory.

J Bacteriol, 1994 Jul, 176(13), 3870 - 7
pH dependence of CheA autophosphorylation in Escherichia coli; Conley MP et al.; Chemotaxis by cells of Escherichia coli and Salmonella typhimurium depends upon the ability of chemoreceptors called transducers to communicate with switch components of flagellar motors to modulate swimming behavior . This communication requires an excitatory pathway composed of the cytoplasmic signal transduction proteins, CheAL, CheAS, CheW, CheY, and CheZ . Of these, the autokinase CheAL is most central . Modifications or mutations that affect the rate at which CheAL autophosphorylates result in profound chemotactic defects . Here we demonstrate that pH can affect CheAL autokinase activity in vitro . This activity exhibits a bell-shaped dependence upon pH within the range 6.5 to 10.0, consistent with the notion that two proton dissociation events affect CheAL autophosphorylation kinetics: one characterized by a pKa of about 8.1 and another exhibiting a pKa of about 8.9 . These in vitro results predict a decrease in the rate of CheAL autophosphorylation in response to a reduction in intracellular pH, a decrease that should cause increased counterclockwise flagellar rotation . We observed such a response in vivo for cells containing a partially reconstituted chemotaxis system . Benzoate (10 mM, pH 7.0), a weak acid that when undissociated readily traverses the cytoplasmic membrane, causes a reduction of cytoplasmic pH from 7.6 to 7.3 . In response to this reduction, cells expressing CheAL, CheAS, and CheY, but not transducers, exhibited a small but reproducible increase in the fraction of time that they spun their flagellar motors counterclockwise . The added presence of CheW and the transducers Tar and Trg resulted in a more dramatic response . The significance of our in vitro results, their relationships to regulation of swimming behavior, and the mechanisms by which transducers might affect the pH dependence of CheA autokinase activity are discussed.

J Infect Dis, 1994 Jul, 170(1), 135 - 9
Neutrophil activation by Helicobacter pylori lipopolysaccharides; Nielsen H et al.; Helicobacter pylori-induced release of toxic substances by neutrophils could be of potential importance in the pathogenesis of gastroduodenal inflammatory diseases . Lipopolysaccharide (LPS) has the ability to induce neutrophil activation at very low concentrations . Neutrophil oxidative metabolism and enzyme release were assessed after stimulation of neutrophils with various preparations of LPS from H . pylori and compared with that obtained with Salmonella typhimurium LPS . No direct activation of neutrophils by LPS was observed . Preincubation with LPS showed a significant priming for increased activity on subsequent stimulation, particularly with rough-form LPS . The potency of H . pylori LPS was 10-fold lower than that of S . typhimurium LPS, probably reflecting the unique biochemical structure of H . pylori LPS . Chronic low-grade stimulation by H . pylori LPS in the gastric mucosa may render neutrophils primed for the excessive release of detrimental substances into the tissue on stimulation by other mediators.

J Med Microbiol, 1994 Jul, 41(1), 20 - 8
Differences in the immune responses of mice and sheep to an aromatic-dependent mutant of Salmonella typhimurium; Brennan FR et al.; A live mutant aroA Salmonella serotype Typhimurium ovine strain (S25/1) could be cultured from tissues of mice for up to 90 days after oral infection . Following vaccination, high levels of Salmonella-specific serum IgM, IgG and IgA were produced in addition to high levels of specific intestinal IgA . Moreover, there was also evidence of Salmonella-specific cell-mediated immunity in vaccinated mice in the form of strong delayed-type hypersensitivity and the production of interferon-gamma (IFN-tau) by spleen cells stimulated with Salmonella antigen . The aroA strain was also recovered from the mesenteric lymph nodes and most tissues examined from sheep vaccinated by the oral route . Salmonella-specific IgM was detected in the serum; however, specific IgG responses were very low and there was an absence of specific copro-antibody . Although strong Salmonella-specific lymphocyte proliferative responses were detected, they did not result in the production of IFN-tau and flow cytometric analysis revealed that the proliferating cells were predominantly B lymphocytes . Despite the absence of strong vaccine-specific immune responses in vaccinated sheep compared with those seen in mice, both mice and sheep were protected against challenge with virulent wild-type strain S25/1.

J Exp Med, 1994 Jul 1, 180(1), 15 - 23
Salmonella typhimurium initiates murine infection by penetrating and destroying the specialized epithelial M cells of the Peyer's patches; Jones BD et al.; Salmonella species are known to initiate infection of mammalian hosts by penetrating the intestinal epithelium of the small bowel . These bacteria preferentially interact with Peyer's patches which are collections of lymphoid follicles making up the gut-associated lymphoid tissue . We infected murine ligated intestinal loops with invasive and noninvasive Salmonella typhimurium strains for 30, 60, 120, and 180 min and examined the infected tissue by transmission electron microscopy . Within 30 min, we found that invasive S . typhimurium exclusively entered M cells found within the follicle-associated epithelium (FAE) of the Peyer's patches . Initially, interactions between invasive bacteria and enterocytes adjacent to the M cells were not found . Invasion of M cells was associated with the ability of the bacteria to invade tissue culture cells . S . typhimurium mutants, which were noninvasive for tissue culture cells, could not be found in ligated loops associated with M cells or enterocytes after incubations of 30, 60, 120, or 180 min . At 60 min, internalized invasive S . typhimurium were cytotoxic for the M cells . Destruction of an M cell formed a gap in the FAE which allowed organisms to invade enterocytes adjacent to the dead cell . Later in the infection process (120 and 180 min), the presence of bacteria beneath the FAE correlated with changes in the cytoarchitecture of the lymphoid follicle . In addition, replicating Salmonella began to enter both the apical and basolateral surfaces of enterocytes adjacent to infected M cells.

Infect Immun, 1994 Jul, 62(7), 2662 - 8
Comparison of the sensitivities of Salmonella typhimurium oxyR and katG mutants to killing by human neutrophils; Papp-Szabo E et al.; The respiratory burst of neutrophils is believed to kill bacteria by generating oxidative species, such as superoxide anion, hydrogen peroxide, and oxidized halogen species . The oxyR gene of Salmonella typhimurium controls a regulon induced by oxidative stress, such as exposure to hydrogen peroxide . Some researchers have suggested that oxyR may play a key role in bacterial survival following phagocytosis . We have tested this possibility by comparing the survival, following exposure to human neutrophils, of isogenic strains bearing different oxyR alleles . Neither inactivation of the oxyR gene nor constitutive overexpression of the oxyR-regulated proteins (oxyR1 allele) greatly alters bacterial resistance to neutrophils . The katG gene, encoding the oxyR-regulated enzyme hydroperoxidase I, was also without effect on survival following exposure to neutrophils . We conclude that the oxyR response does not play a significant role in the resistance of S . typhimurium to phagocytic killing in vitro.

Biol Pharm Bull, 1994 Jul, 17(7), 990 - 2
Ursolic acid inhibits aflatoxin B1-induced mutagenicity in a Salmonella assay system; Young HS et al.; An attempt was made to isolate the active component of Eriobotrya japonica, which inhibits aflatoxin B1-induced mutagenicity in the Salmonella assay system . The number of revertants per plate was significantly decreased when a MeOH extract of Eriobotrya japonica was added to the assay system using Salmonella typhimurium TA100 or TA98 . Furthermore, we examined the effect of each fraction purified from the MeOH extract, and an EtOAc fraction was found to be the most effective . Ursolic acid isolated from the EtOAc fraction markedly and significantly decreased the numbers of Salmonella typhimurium TA100 revertants per plate, thus showing antimutagenic activity.

Mol Microbiol, 1994 Jul, 13(1), 133 - 40
Role of hns in the virulence phenotype of pathogenic salmonellae; Harrison JA et al.; A TnphoA-generated mutant C5060, attenuated for virulence, was derived from the mouse-virulent Salmonella typhimurium strain C5 . This mutation, designated hns-112::TnphoA, harbours the transposon in the 3' end of hns, with the alkaline phosphatase open reading frame in the opposite orientation to that of hns . Bacterial strains harbouring hns-112::TnphoA were mucoid and had altered levels of DNA supercoiling, as monitored using pUC18 as a reporter plasmid . Transduction of hns-112::TnphoA into mouse virulent strains, including S . typhimurium SL1344 and Salmonella enteritidis Se795, resulted in attenuation . When an independent hns mutation, harbouring a kanamycin-resistance cassette inserted into the Kpnl site at base pair 237 of the hns gene, was introduced into S . typhimurium C5, the isolates were also attenuated . S . typhimurium C5 isolates harbouring the multicopy plasmid pGB651, which encodes the Escherichia coli hns gene, were partially attenuated in mice . Transductional analysis, using Tn10 insertions located close to the hns gene, showed that virulence could be restored in genetic crosses that eliminated the resident hns mutations . However, some hns+ transductants were still attenuated, suggesting that secondary attenuating lesions can accumulate in hns-deficient strains . These studies show that the hns locus plays a role in Salmonella virulence.

Chem Res Toxicol, 1994 Jul-Aug, 7(4), 482 - 6
Oxidation and radical intermediates associated with the glutathione conjugation of mucochloric acid; LaLonde RT et al.; The inactivation of the drinking water mutagen mucochloric acid (MCA) by reduced glutathione (GSH) was linked to the formation of an MCA-GSH conjugate, a nonmutagen in the Salmonella typhimurium (TA100) plate incorporation assay . Anaerobic formation of MCA-GSH is found now to be associated with oxidized glutathione (GSSG) and unconverted MCA . The anaerobic reaction of GSH with MCA in the presence of the radical trap 2-methyl-2-nitrosopropane (tNB; "tert-nitrosobutane") gives rise to an electron paramagnetic resonance (EPR) resulting from the overlapping spectra of two radical adducts . The first species exhibited hyperfine coupling constants of aN = 13.65 G and aH beta = 0.73 G . The second radical adduct exhibited a three-line signal of aN = 12.8 G . The first species is assigned to an adduct of the MCA radical because deuteration of MCA (5-deuterio-MCA) caused the beta-hydrogen hyperfine coupling to collapse . The second radical adduct is unaffected by the deuteration of MCA . Thus, the involvement of both GSSG and a carbon-centered MCA radical in the action of MCA on GSH is indicated.

Am J Vet Res, 1994 Jul, 55(7), 921 - 7
Effects of polymyxin B and Salmonella typhimurium antiserum on horses given endotoxin intravenously; Durando MM et al.; Polymyxin B and an antiserum against an Re mutant Salmonella typhimurium were evaluated for protective effect in an equine model endotoxemia . Six 3- to 5-month-old foals were given endotoxin (0.25 micrograms/kg of body weight) IV after no pretreatment, or pretreatment with polymyxin B (6,000 U/kg, IV) or S typhimurium antiserum (1.5 ml/kg, IV) . When given without pretreatment, endotoxin caused transient recumbency and increases in rectal temperature, and heart and respiratory rates . In addition, leukopenia and increases in circulating tumor necrosis factor (TNF) and interleukin 6 (IL-6) activities were detected . Compared with results obtained when endotoxin was given alone, pretreatment with polymyxin B resulted in significantly (P < 0.05) lower maximal plasma TNF and IL-6 activities, and significantly lower rectal temperature and respiratory rate . In contrast, compared with effects of endotoxin given without pretreatment, use of antiserum was associated with significantly (P < 0.05) higher respiratory rate, maximal plasma IL-6 activity, and total TNF response (as determined by areas under curves of plasma TNF vs time) . These results indicate that polymyxin B may have potential as a treatment for equine endotoxemia . Salmonella typhimurium antiserum had no positive effect in this model, and, under certain conditions, may exacerbate the actions of endotoxin.

Genetika, 1994 Jul, 30(7), 898 - 902
{Mutagenic activity of 2,4,6-trinitrotoluene: the role of metabolizing enzymes}; Karamova NS et al.; Mutagenic activity of 2,4,6-trinitrotoluene (2,4,6-TNT) and one of its derivatives, 2,4-diamino-6-nitrotoluene (2,4-DA-6-NT) was studied in the Ames test using the Salmonella typhimurium tester strains TA98, TA100, TA100NR (deficient in classical nitroreductases), and TA100/1, 8DNP (deficient in O-acetyltransferase) . The number of revertants was significantly increased by 2,4,6-TNT, in comparison with that of spontaneous revertants . Base pair substitutions were preferentially induced by 2,4,6-TNT . The mutagenic effect of this compound was virtually altered by in vitro metabolic activation with the S9 fraction of human placenta . Neither 2,4,6-TNT nor 2,4-DA-6-NT exhibited mutagenic activity in the strains TA 100NR and TA 100/1,8DNP . Based on the results obtained, it is concluded that the mutagenic potential of 2,4,6-TNT is determined by its metabolites that form during bacterial metabolism . Bacterial nitroreductases and O-acetyltransferases are the key activators of this compound to form the ultimate mutagens.

Fundam Appl Toxicol, 1994 Jul, 23(1), 9 - 20
Carcinogenicity and mutagenicity studies with fluvastatin, a new, entirely synthetic HMG-CoA reductase inhibitor; Robison RL et al.; The HMG-CoA reductase inhibitors are a new and novel class of cholesterol-lowering agents which are widely used worldwide . Fluvastatin is the first entirely synthetic compound in this class and is structurally distinct from fungal metabolite derivatives which are already marketed . As the liver is the site of some toxic effects for these compounds, it was not entirely unexpected that liver cancer was found in rats and/or mice with the first three marketed compounds, lovastatin, pravastatin, and simvastatin . Four lifetime carcinogenicity studies (two rat and two mouse) did not give any evidence that fluvastatin induced liver tumors in rodents . Fluvastatin induced thyroid neoplasms in rats and forestomach papillomas in rodents, as other compounds in this pharmacologic class have also done . The genotoxic potential of fluvastatin has been assessed in vitro using Salmonella typhimurium, Escherichia coli (gene mutations), V79 Chinese hamster cells (HGPRT gene mutations, chromosomal aberrations), rat hepatocyte primary cultures (DNA repair), and BALB/3T3 cells (malignant transformations) . Fluvastatin was also tested in vivo for clastogenicity using the mouse bone marrow micronucleus test and by performing a cytogenetic analysis in the rat bone marrow after acute and subacute treatment . In all seven assays fluvastatin was found to be free of any genotoxic potential.

J Appl Bacteriol, 1994 Jul, 77(1), 113 - 9
The effect of transient temperatures on the growth of Salmonella typhimurium LT2 . I: Cycling within the growth region; Mitchell GA et al.; The effect of fluctuating temperatures on microbial growth is important in the passage of foods through the food chain . Suspensions of Salmonella typhimurium were subjected to sinusoidally time-varying temperatures of periods from 40 to 480 min within their growth temperature range . The change in the numbers of viable bacteria was measured with time and the experimental growth curves and average generation times compared with predictions based on isothermal growth data . The experimental average generation times exceeded the predictions by less than 30%, although the discrepancy increased with cycle frequency . Instantaneous growth rates obtained for the 480 min cycles were in agreement with those predicted from isothermal behaviour.

Protein Sci, 1994 Jul, 3(7), 1074 - 80
UDP-glucose dehydrogenase from bovine liver: primary structure and relationship to other dehydrogenases; Hempel J et al.; The primary structure of bovine liver UDP-glucose dehydrogenase (UDPGDH), a hexameric, NAD(+)-linked enzyme, has been determined at the protein level . The 52-kDa subunits are composed of 468 amino acid residues, with a free N-terminus and a Ser/Asn microhetergeneity at one position . The sequence shares 29.6% positional identity with GDP-mannose dehydrogenase from Pseudomonas, confirming a similarity earlier noted between active site peptides . This degree of similarity is comparable to the 31.1% identity vs . the UDPGDH from type A Streptococcus . Database searching also revealed similarities to a hypothetical sequence from Salmonella typhimurium and to "UDP-N-acetyl-mannosaminuronic acid dehydrogenase" from Escherichia coli . Pairwise identities between bovine UDPGDH and each of these sequences were all in the range of approximately 26-34% . Multiple alignment of all 5 sequences indicates common ancestry for these 4-electron-transferring enzymes . There are 27 strictly conserved residues, including a cysteine residue at position 275, earlier identified by chemical modification as the expected catalytic residue of the second half-reaction (conversion of UDP-aldehydoglucose to UDP-glucuronic acid), and 2 lysine residues, at positions 219 and 338, one of which may be the expected catalytic residue for the first half-reaction (conversion of UDP-glucose to UDP-aldehydoglucose) . A GXGXXG pattern characteristic of the coenzyme-binding fold is found at positions 11-16, close to the N-terminus as with "short-chain" alcohol dehydrogenases.(ABSTRACT TRUNCATED AT 250 WORDS)

APMIS, 1994 Jul, 102(7), 521 - 5
Salmonella typhimurium phage types from human salmonellosis in Denmark 1988-1993; Wegener HC et al.; A total of 989 isolates of Salmonella enterica ssp . enterica serovar Typhimurium from cases of human salmonellosis were investigated by phage typing . The isolates comprised all isolates recovered during the month of August in each of the years from 1988 to 1993 . Phage typing assigned 82.6% of the strains to 36 different definitive types, 11.9% of the strains belonged to types of unknown lysis pattern (RDNC), and 5.5% could not be typed by the phages used (NT) . Three phage types (12, 66 and 110) made up approximately 50% of the isolates in each of the years investigated . During the period in question these types showed major changes in prevalence: phage type 12 increased from 4.0% in 1988 to a maximum of 55.2% in 1992, and phage type 66 and phage type 110 were reduced from 40.1% and 27.8% to a minimum of 3.9% and 4.8% in 1993 and 1992, respectively . The increasing prevalence of phage type 12 among isolates from human salmonellosis most likely reflects the increasing significance of pork as a source of human salmonellosis . The reduction in phage types 66 and 110 is paralleled by a reduction in S . Typhimurium in Danish poultry . Some phage types were associated with travel, namely 17, 193 and 194 . It is concluded that phage typing, although here performed retrospectively, produces valuable epidemiological information regarding changes in the relative importance of different sources of infection in humans . It is suggested that phage typing be performed prospectively on both human and animal S . Typhimurium isolates in Denmark.

APMIS, 1994 Jul, 102(7), 489 - 94
Immunity to experimental Salmonella typhimurium infections in rats . Transfer of immunity with primed CD4+CD25high and CD4+CD25low T lymphocytes; Thygesen P et al.; The protective effect of primed CD4+ T lymphocytes against a lethal dose of 10(8) viable Salmonella typhimurium was studied in Lewis rats . Primed CD4+ T lymphocytes were obtained by inoculating Lewis rats with a non-lethal dose of 10(6) viable S . typhimurium . Four weeks after the infection, spleen CD4+ T lymphocytes were separated using magnetic microspheres coated with an antibody against the CD4 molecule (W3/25) . Subsequent sorting into activated and non-activated subpopulations using the p55 alpha-chain of the interleukin-2 receptor (CD25) as an activation marker was performed by a fluorescence-activated cell sorter . Untreated Lewis rats were injected with 10(4) different primed CD4+ T-cell populations 24 h prior to the lethal dose of 10(8) viable S . typhimurium . Blood samples were drawn from the orbital plexus 1, 2, 3, and 4 weeks after the infection, and analysed for specific IgM and IgG antibodies . Cell sorting revealed that 2/3 of the primed CD4+ T lymphocytes expressed high levels of CD25 . Cell transfer revealed that both CD25high and CD25low expression populations could induce immunity against a lethal dose of S . typhimurium, whilst antibody analysis revealed that antibody levels were not correlated with protection against S . typhimurium infections, although it showed that a higher and more persistent level of specific IgG antibodies was produced in animals receiving the CD4+CD25high fraction . It is concluded that 10(4) primed CD4+ T lymphocytes can induce immunity in animals challenged with a lethal dose of S . typhimurium and that antibodies do not seem to be correlated with the immunity induced . The CD4+CD25high fraction was, however, associated with a higher and more persistent level of specific IgG antibodies.

Appl Environ Microbiol, 1994 Jul, 60(7), 2568 - 74
Accumulation of glutamate by Salmonella typhimurium in response to osmotic stress; Botsford JL et al.; Salmonella typhimurium accumulates glutamate in response to osmotic stress . Cells in aerobic exponential growth have an intracellular pool of approximately 125 nmol of glutamate mg of protein-1 . When cells were grown in minimal medium with 500 mM NaCl, KCl, or sucrose, 290 to 430 nmol of glutamate was found to accumulate . Values were lower when cells were harvested in stationary phase . Cells were grown in conventional medium, harvested, washed, resuspended in the control medium or in medium with osmolytes, and aerated for 1 h . With aeration, glutamate was found to accumulate at levels comparable to those observed in exponential cultures . Antibiotics inhibiting protein synthesis did not affect glutamate accumulation when cells were aerated . Strains with mutations in glutamate synthase (glt) or in glutamate dehydrogenase (gdh) accumulated nearly normal levels of glutamate under these conditions . A double (gdh glt) mutant accumulated much less glutamate (63.9 nmol mg of protein-1), but a 1.9-fold excess accumulated when cells were aerated with osmotic stress . Methionine sulfone, an inhibitor of glutamate synthase, did not prevent accumulation of glutamate in cells aerated with osmotic stress . Glutamate dehydrogenase is thought to have minimum activity when ammonium is limiting . Resuspending cells with limiting ammonium reduced glutamate production but did not eliminate accumulation of excess glutamate when cells were osmotically stressed . Amino oxyacetic acid, an inhibitor of transamination reactions, did not prevent accumulation of excess glutamate.(ABSTRACT TRUNCATED AT 250 WORDS)

Infect Immun, 1994 Jul, 62(7), 2874 - 84
T lymphocytes in host defense against bacterial translocation from the gastrointestinal tract; Gautreaux MD et al.; Flow cytofluorometric analyses of lymphocytes harvested from the mesenteric lymph node (MLN), mucosal epithelium, and lamina propria of C57BL/6 mice demonstrate that expression of alpha/beta or gamma/delta T-cell receptors (TCR) and CD4 or CD8 molecules by T lymphocytes in the intestinal immune system varies depending upon their anatomic location . The MLN contained equivalent numbers of CD4+ and CD8+ T cells, the vast majority of which were alpha/beta TCR positive (alpha/beta TCR+) . The lamina propria T cells were predominantly CD4+ and alpha/beta TCR+, while the intestinal intraepithelial lymphocytes consisted of equivalent numbers of alpha/beta and gamma/delta T cells, the majority of which were CD8+ . There were no significant changes in these T-cell phenotypic profiles when the mice were antibiotic decontaminated or monoassociated with Escherichia coli . Mice were depleted of CD4+ T cells and/or CD8+ T cells in vivo by intraperitoneal injections of monoclonal antibody GK 1.5 (rat anti-mouse CD4) and/or monoclonal antibody 2.43 (rat anti-mouse CD8) . T-cell depletion was confirmed in the MLN, lamina propria, and the intestinal epithelium by flow cytometry . E . coli C25 translocation from the gastrointestinal (GI) tract to the MLN was significantly increased in mice depleted of CD4+ T cells, CD8+ T cells, or both . T-cell-deficient athymic beige/nude mice also exhibited greater levels of E . coli C25 translocation to the MLN than beige/het euthymic littermates . Salmonella typhimurium translocation also was increased following CD4+ and CD8+ T-cell depletion in mice monoassociated with S . typhimurium . Depletion of CD4+ and/or CD8+ T cells also increased the translocation to the MLN of certain indigenous GI flora bacteria . These results confirm that T-cell-mediated immunity is involved in the host defense against bacterial translocation from the GI tract.

Infect Immun, 1994 Jul, 62(7), 2784 - 91
Protective role for heat shock protein-reactive alpha beta T cells in murine yersiniosis; Noll A et al.; To investigate the role of heat shock proteins (HSP) of Yersinia enterocolitica for the host immune response against this pathogen, we cloned and expressed a 60-kDa HSP of Y . enterocolitica serotype O8 . A fragment of Y . enterocolitica O8 HSP60 encoded by amino acids 90 to 286 was sequenced and showed more than 90% homology with HSP60 of Y . enterocolitica O3 and GroEL of Escherichia coli and 59% homology with HSP65 of Mycobacterium bovis . The arthritogenic T-cell epitope of mycobacterial HSP65 (amino acid residues 180 to 188) was not found on Yersinia HSP60 . To determine whether Yersinia HSP60 is an immunodominant antigen, the immune responses of Yersinia-infected C57BL/6 mice were analyzed . Yersinia-infected mice evolved a significant serum antibody and splenic T-cell response against Yersinia HSP60 . CD4+ alpha beta T-cell clones which were generated from splenic T cells isolated from either Yersinia-infected or Yersinia HSP60-immunized mice, recognized both heat-killed Yersinia serotypes O3 and O8 as well as recombinant Yersinia HSP60 but not heat-killed Yersinia pseudotuberculosis, Salmonella typhimurium, or recombinant HSP65 of Mycobacterium bovis . The adoptive transfer of HSP60-reactive T-cell clones mediated significant protection against a lethal infection with Y . enterocolitica O8 . These results indicate that HSP60 of Y . enterocolitica is an immunodominant antigen which is recognized by both antibodies and CD4+ alpha beta T cells . Moreover, this is the first report providing direct evidence that microbial HSP may elicit a protective immune response which is not associated with autoimmunity.

Infect Immun, 1994 Jul, 62(7), 2702 - 6
Listeria monocytogenes, but not Salmonella typhimurium, elicits a CD18-independent mechanism of neutrophil extravasation into the murine peritoneal cavity; Conlan JW et al.; This study shows that extravasation of neutrophils into the peritoneal cavities of mice in response to intraperitoneal (i.p.) inoculation of wild-type Listeria monocytogenes requires the participation of leukocyte adhesion molecules that are different from those involved in neutrophil recruitment in response to i.p . inoculation of Salmonella typhimurium . In the case of S . typhimurium, extensive neutrophil influx could be essentially abolished by treating mice with either anti-CD11b or anti-CD18 monoclonal antibodies, whereas the same monoclonal antibodies failed to prevent neutrophil accumulation in the peritoneal cavity in response to inoculation of L . monocytogenes . On the other hand, i.p . inoculation of a listeriolysin-negative strain of L . monocytogenes induced a CD11b-dependent neutrophil influx . The possibility that wild-type L . monocytogenes, by virtue of its ability to inhabit the cytosol of the cells it infects, induces the expression of endothelial cell adhesion molecules in the microvasculature of the peritoneal cavity to which neutrophils adhere via leukocyte adhesion molecules distinct from beta-2 integrins is discussed.

Avian Dis, 1994 Jul-Sep, 38(3), 598 - 604
A rapid method for screening for Salmonella typhimurium in a chicken cecal microbial consortium using gene amplification; Pillai SD et al.; A rapid sample processing method has been developed to detect low numbers of Salmonella typhimurium in a chicken cecal microbial consortium . Using phoP-specific primers under stringent amplification conditions and gene probe analysis, fewer than 100 colony-forming units (CFUs) were detectable when pure cultures were employed . When the polymerase chain reaction assay was run on cecal contents from birds infected with S . typhimurium, only positive cecal samples containing as few as 700 CFUs reacted to the assay, and the negative bird samples reacted only when the samples were spiked with S . typhimurium cells . The method employed for sample processing is simple and provides a sensitive means of detecting S . typhimurium-specific sequences in trace amounts in the presence of mixed chicken cecal microbial populations.

Avian Dis, 1994 Jul-Sep, 38(3), 583 - 8
Transmission of Salmonella typhimurium during hatching of broiler chicks; Cason JA et al.; Horizontal spread of Salmonella during hatching of broiler chicks was studied in three experiments . In each experiment, 120 unincubated, fertile hatching eggs were inoculated by immersion for 15 min in a 16 C physiological saline solution containing 1 x 10(8) colony-forming units per ml of a nalidixic-acid-resistant strain of S . typhimurium . When inoculated eggs were transferred to hatchers after 17 to 18 days of incubation, control eggs at the same stage of incubation were added to the same tray and to trays above and below the tray containing inoculated eggs . Fertile inoculated eggs hatched at a rate of 86%, despite the high level of Salmonella contamination, indicating that chicks in eggs contaminated with salmonellae are likely to hatch and may contaminate other chicks in the same hatcher cabinet . Air samples showed a sharp increase in contamination in the hatcher at 20 days of incubation . Approximately 58% of mouth swabs and 90% of chick rinses were Salmonella-positive, in both inoculated and control eggs . In samples from inoculated eggs, Salmonella was detected in the digestive tract of 8% of embryos at transfer from incubator to hatcher and in 55% of chicks at hatch . From control eggs, 44% of digestive tracts of hatched chicks were positive, indicating that Salmonella in a contaminated hatcher can reach the gut of chicks hatching from Salmonella-free eggs before they are removed from the hatcher.

Avian Dis, 1994 Jul-Sep, 38(3), 401 - 8
Altered colonizing ability for the ceca of broiler chicks by lipopolysaccharide-deficient mutants of Salmonella typhimurium; Craven SE; Salmonella typhimurium strain 3333/O was used to assess the role of bacterial lipopolysaccharide (LPS) in intestinal colonization of broiler chicks by salmonellae . LPS-defective TnPhoA mutants of this strain were isolated . The sensitivities of the mutants to smooth and rough phages and LPS banding patterns in sodium dodecyl sulfate-polyacrylamide gel electrophoresis indicated a defect in the polysaccharide side chain of the LPS molecule . Colonization was determined by orally administering 10(8) cells each of the wild-type and/or the mutant strains per chick and counting the colony-forming units (CFU) from the ceca 1 to 3 weeks after gavage . CFU of chicks given the LPS-deficient strains either were not detected or were significantly lower than the CFU from chicks given the wild-type strain . The incidence of the wild-type strain in spleens was higher than incidence of the mutant strains . In vitro binding studies with LPS-deficient mutants derived in this study and from S . typhimurium LT2 suggest that LPS side-chain components may shield the bacterial cell from entrapment in the chicken mucus . The LPS layer appears to enhance persistence of Salmonella in the avian intestinal tract.

J Cell Sci, 1994 Jul, 107 ( Pt 7), 2005 - 20
Salmonella typhimurium induces selective aggregation and internalization of host cell surface proteins during invasion of epithelial cells; Garcia-del Portillo F et al.; Salmonella interact with eucaryotic membranes to trigger internalization into non-phagocytic cells . In this study we examined the distribution of host plasma membrane proteins during S . typhimurium invasion of epithelial cells . Entry of S . typhimurium into HeLa epithelial cells produced extensive aggregation of cell surface class I MHC heavy chain, beta 2-microglobulin, fibronectin-receptor (alpha 5 beta 1 integrin), and hyaluronate receptor (CD-44) . Other cell surface proteins such as transferrin-receptor or Thy-1 were aggregated by S . typhimurium to a much lesser extent . Capping of these plasma membrane proteins was observed in membrane ruffles localized to invading S . typhimurium and in the area surrounding these structures . In contrast, membrane ruffling induced by epidermal growth factor only produced minor aggregations of surface proteins, localized exclusively in the membrane ruffle . This result suggests that extensive redistribution of these proteins requires a signal related to bacterial invasion . This bacteria-induced process was associated with rearrangement of polymerized actin but not microtubules, since preincubation of epithelial cells with cytochalasin D blocked aggregation of these proteins while nocodazole treatment did not . Of the host surface proteins aggregated by S . typhimurium, only class I MHC heavy chain was predominantly present in the bacteria-containing vacuoles . No extensive aggregation of host plasma membrane proteins was detected when HeLa epithelial cells were infected with invasive bacteria that do not induce membrane ruffling, including Yersinia enterocolitica, a bacterium that triggers internalization via binding to beta 1 integrin, and a S . typhimurium invasion mutant that utilizes the Yersinia-internalization route . In contrast to the situation with S . typhimurium, class I MHC heavy chain was not selectively internalized into vacuoles containing these other bacteria . Extensive aggregation of host plasma membrane proteins was also not observed when other S . typhimurium mutants that are defective for invasion were used . The amount of internalized host plasma membrane proteins in the bacteria-containing vacuoles decreased over time with all invasive bacteria examined, indicating that modification of the composition of these vacuoles occurs . Therefore, our data show that S . typhimurium induces selective aggregation and internalization of host plasma membrane proteins, processes associated with the specific invasion strategy used by this bacterium to enter into epithelial cells.

Mutat Res, 1994 Jul, 324(3), 111 - 4
The presence of KCl in the exposure medium strongly influences the mutagenicity of metabolites of polycyclic aromatic hydrocarbons in Escherichia coli; Glatt H et al.; Previous studies demonstrated that the ion composition of the exposure medium may strongly influence the mutagenicity of many compounds in the liquid preincubation modification of the reversion assay with his- Salmonella typhimurium strains . Similar influences were now observed in the reversion assay with trp- Escherichia coli strain WP2 uvrA . The exposure medium was 8 mM sodium phosphate buffer (pH 7.4), containing no other ions or 125 mM KCl . Omission of KCl resulted in an about 10-fold enhancement of the mutagenic activity of 7-methylbenz{a}anthracene 5,6-oxide, but in a strong decrease in the mutagenicity of 1-hydroxymethylpyrene sulphate, close to the limit of detection . The findings with these two representative mutagens are very similar to those previously observed in S . typhimurium, suggesting that unobtrusive medium components may exert strong influences on the results with many test compounds in various bacterial test systems.

Mutat Res, 1994 Jul 1, 308(1), 33 - 42
Effects of deuterium labeling on azido amino acid mutagenicity in Salmonella typhimurium; Mangold JB et al.; The mutagenic effects of azide (N3-) anion in bacterial test systems require the formation of the novel mutagenic metabolite, 3-azido-L-alanine (AZAL) . Although the mechanism of AZAL-induced mutagenicity is unknown, subsequent bioactivation of this metabolite appears likely . Earlier studies have shown that other azide-containing amino acids are mutagenic as well . In fact, the mutagenic potency of the synthetic AZAL homologue, L-2-amino-4-azidobutanoic acid (HomoAZAL), was several times that of AZAL . To gain insight into the biochemical processing and mutagenicity of azido amino acids in Salmonella typhimurium, several specifically deuterium-labeled azido amino acids have been prepared and tested for mutagenic potency . In addition, the effect of (aminooxy)acetic acid (AOA) (a potent inhibitor of pyridoxal-dependent processes) on AZAL-induced mutagenesis was examined . The results showed that 2-deuterium substitution of AZAL resulted in a slight increase in mutagenic potency, while AOA treatment resulted in no change in AZAL potency . Taken together these findings did not support the involvement of pyridoxal-dependent processes in AZAL bioactivation . In contrast, deuterium substitution adjacent to the azide group in HomoAZAL and 5-azido-L-norvaline (N3-Norval) resulted in a large decrease in mutagenic potency when compared to the non-deuterium labeled compounds . These observations are consistent with a bioactivation mechanism involving rate-limiting C-H bond cleavage in the formation of the ultimate mutagen . Moreover, this effect of deuterium labeling points to processing of the azide-containing side chain as a key feature in the mutagenic activation mechanism.

Gene, 1994 Jun 24, 144(1), 81 - 5
Improved synthesis of Salmonella typhimurium enterotoxin using gene fusion expression systems; Chopra AK et al.; Salmonella enterotoxin (Stn) is a virulence factor in S . typhimurium strain Q1 that causes both fluid secretion in ligated intestinal loops of rabbits and elongation of Chinese hamster ovary (CHO) cells . High-level expression systems are needed to provide Stn in soluble form for detailed study of the biological activity of Stn . To maximize the synthesis and solubility of Stn, we systematically compared the production of native Stn synthesized with a T7 RNA polymerase/promoter system to that of two fusion proteins: glutathione S-transferase::Stn (Gst::Stn) and thioredoxin A::Stn (TrxA::Stn) . The latter fusion protein expression systems resulted in a 64-fold increase in Gst::Stn and TrxA::Stn antigen concentration, as measured by specific anti-peptide antibodies in an enzyme-linked immunosorbent assay (ELISA) . Most of the toxin derived using these vector systems was insoluble; however, the solubility of the TrxA::Stn antigen increased by at least 50-fold, with a concomitant increase in CHO cell elongation activity . In addition, stn gene expression was enhanced more than 50-fold by addition of 0.2-0.4 M NaCl to Luria-Bertani medium . The biological activity of Stn also was increased in the high-osmolarity medium . Consequently, the expression of stn may be regulated by DNA supercoiling.

Mol Gen Genet, 1994 Jun 15, 243(6), 674 - 80
Mutations affecting a regulated, membrane-associated esterase in Salmonella typhimurium LT2; Collin-Osdoby P et al.; Mutations at the apeA locus in Salmonella typhimurium lead to loss of a soluble enzyme ("protease I") that hydrolyzes the chromogenic endoprotease substrate N-acetyl phenylalanine beta-naphthyl ester . We have isolated pseudorevertants of S . typhimurium apeA mutations that have regained the ability to hydrolyze this compound . These pseudorevertants contain mutations (apeR) that lead to overproduction of a membrane-bound esterase different from protease I . The apeR locus is phage P1 cotransducible with ilvC (83 map units) and is unlinked to apeA . Mutations at still another locus, apeE, lead to loss of the membrane-associated esterase . The apeE locus is P1 cotransducible with purE (12 map units) . In an apeE-lacZ operon fusion strain, an apeR mutation increases the level of beta-galactosidase approximately 60-fold . We propose that apeR encodes a repressor of apeE . The evidence available suggests that the ApeE protein is not a protease.

Mol Gen Genet, 1994 Jun 15, 243(6), 605 - 12
Excretion of the anti-sigma factor through a flagellar substructure couples flagellar gene expression with flagellar assembly in Salmonella typhimurium; Kutsukake K; More than 50 genes are required for flagellar formation and function in Salmonella typhimurium . According to the cascade model of the flagellar regulon, the flagellar operons are divided into three classes, 1, 2, and 3, with reference to their relative positions in the transcriptional hierarchy . This sequential transcription is coupled to the assembly process of the flagellar structure, that is, genes involved in formation of the hook-basal body complex belong to the class-2 operons, whereas those involved in formation of filament belong to the class-3 operons . The fliA gene encodes an alternative sigma factor specific for transcription of the class-3 operons . A negative regulatory gene, flgM, which is responsible for the coupling of expression of class-3 operons to flagellar assembly, encodes an anti-sigma factor that binds to FliA and prevents its association with RNA polymerase core enzyme . In the present study, we showed that the flgM gene is transcribed from two different promoters: one is its own class-3 promoter and the other is the class-2 promoter for the upstream gene, flgA . Furthermore, we showed that FlgM is excreted into culture medium from cells of the wild-type strain and of class-3 mutants that can produce complete hook-basal body structures . On the other hand, FlgM is not excreted from mutants defective in the hook-basal body genes . These results indicate that FlgM is excreted from the cells through the flagellar substructures that are formed by the function of the hook-basal body genes.(ABSTRACT TRUNCATED AT 250 WORDS)

J Biol Chem, 1994 Jun 10, 269(23), 16486 - 92
Aliphatic alcohols stabilize an alternative conformation of the tryptophan synthase alpha 2 beta 2 complex from Salmonella typhimurium; Ahmed SA et al.; The conformational changes that accompany association of the tryptophan synthase alpha and beta 2 subunits are probed by investigating the effects of solvents on the catalytic and spectroscopic properties of tryptophan synthase . Low concentrations of ethanol, propanol, and butanol inhibit conversion of L-serine and indole to L-tryptophan by the alpha 2 beta 2 complex . Inhibition depends on the concentration, chain length, and hydrophobicity of the alcohol . In contrast, these alcohols increase the rates of conversion of beta-chloro-L-alanine and indole to L-tryptophan and of L-serine to pyruvate . Thus, alcohols alter the substrate specificity and reaction specificity of the alpha 2 beta 2 complex . These altered specificities are similar to those of the free beta 2 subunit in aqueous solution . The spectroscopic properties of enzyme-substrate intermediates formed by the alpha 2 beta 2 complex in 3 M ethanol and 0.5 M 1-butanol also resemble the intermediates formed by the free beta 2 subunit in aqueous solution . Experiments using gel-filtration, membrane ultrafiltration, and limited proteolysis show that the alpha 2 beta 2 complex does not dissociate at low concentrations of alcohols . Our results provide evidence that alcohols can stabilize an alternative conformation of the alpha 2 beta 2 complex which is similar to the open conformation of the free beta 2 subunit in aqueous solution.

Science, 1994 Jun 10, 264(5165), 1578 - 81
The structural basis of sequence-independent peptide binding by OppA protein; Tame JR et al.; Specific protein-ligand interactions are critical for cellular function, and most proteins select their partners with sharp discrimination . However, the oligopeptide-binding protein of Salmonella typhimurium (OppA) binds peptides of two to five amino acid residues without regard to sequence . The crystal structure of OppA reveals a three-domain organization, unlike other periplasmic binding proteins . In OppA-peptide complexes, the ligands are completely enclosed in the protein interior, a mode of binding that normally imposes tight specificity . The protein fulfills the hydrogen bonding and electrostatic potential of the ligand main chain and accommodates the peptide side chains in voluminous hydrated cavities.

Proc Natl Acad Sci U S A, 1994 Jun 7, 91(12), 5485 - 9
Liberation of an interaction domain from the phosphotransfer region of CheA, a signaling kinase of Escherichia coli; Morrison TB et al.; The CheA protein of Escherichia coli is a histidine autokinase that donates its phosphate groups to two target proteins, CheY and CheB, to regulate flagellar rotation and sensory adaptation during chemotactic responses . The amino-terminal third of CheA contains the autophosphorylation site, determinants needed to interact with the catalytic center of the molecule, and determinants needed for specific recognition of its phosphorylation targets . To understand the structural basis for these activities, we examined the domain organization of the CheA phosphotransfer region by using DNA sequence analysis, limited proteolytic digestion, and a genetic technique called domain liberation . Comparison of the functionally interchangeable CheA proteins of E . coli and Salmonella typhimurium revealed two extensively mismatched segments within the phosphotransfer region, 22 and 25 aa long, with sequences characteristic of domain linkers . Both segments were readily susceptible to proteases, implying that they have an extended, flexible structure . In contrast, the intervening segments of the phosphotransfer region, designated P1 and P2 (roughly 140 and 65 aa, respectively), were relatively insensitive, suggesting they correspond to more compactly folded structural domains . Their functional properties were explored by identifying portions of the cheA coding region capable of interfering with chemotactic behavior when "liberated" and expressed as polypeptides . P1 fragments were not inhibitory, but P2 fragments blocked the interaction of CheY with the rotational switch at the flagellar motor, leading to incessant forward swimming . These results suggest that P2 contains CheY-binding determinants which are normally responsible for phosphotransfer specificity . Domain-liberation approaches should prove generally useful for analyzing multidomain proteins and their interaction targets.

J Bacteriol, 1994 Jun, 176(12), 3757 - 64
Topological promoter coupling in Escherichia coli: delta topA-dependent activation of the leu-500 promoter on a plasmid; Chen D et al.; The leu-500 promoter of Salmonella typhimurium is activated in topA mutants . We have previously shown that this promoter can be activated on circular plasmids in a manner that depends on transcription and translation of the tetracycline resistance gene tetA and insertion of its product into the cell membrane . We have suggested that in the absence of enzymatic relaxation by topoisomerase I, the local domain of transcription-induced DNA supercoiling reaches a steady-state level that leads to the activation of the leu-500 promoter . In the present paper, we have shown that the leu-500 promoter may also be activated in Escherichia coli . Comparison of the closely related pair of E . coli strains DM800 (delta topA) and SD108 (topA+) shows that the activation is dependent on the presence of a null mutation in topA . We have also shown that activation of the plasmid-borne leu-500 promoter depends, as in S . typhimurium, on the function of an adjacent tetA gene, suggesting that membrane anchorage of the TetA peptide prevents dissipation of transcription-induced supercoiling by superhelical diffusion . The activity of the leu-500 promoter is boosted by placing a divergent tac promoter on the side opposite to tetA . The topoisomer distributions of these plasmids extracted from the cell have been analyzed . We find that when the parent plasmid pLEU500Tc, containing the leu-500 promoter upstream of the complete tetA gene, is extracted from E . coli DM800 (delta topA), the distribution of linking numbers is bimodal . There is a fraction with a lower level of supercoiling (mean linking difference approximately -0.05) that is constant for all plasmids extracted from either delta topA or topA+ cells . In addition, we observe a second fraction with highly negatively supercoiled DNA (mean linking difference approximately -0.09) only in DNA extracted from delta topA cells . The proportion of the oversupercoiled fraction correlates with the activity of the leu-500 promoter: it is strongly reduced when the tetA promoter is deleted or when translation of TetA is prematurely terminated, while it is increased when the strong tac promoter is present in cis . We suggest that this oversupercoiled fraction represents the proportion of plasmid molecules active in tetA transcription and that it is this supercoiling that activates the leu-500 promoter.

J Bacteriol, 1994 Jun, 176(12), 3683 - 91
Overproduction of the bacterial flagellar switch proteins and their interactions with the MS ring complex in vitro; Oosawa K et al.; The flagellar switch proteins (FliG, FliM, and FliN) of Salmonella typhimurium were overproduced in Escherichia coli and partially purified in soluble form . They were mixed with purified MS ring complexes (which consist of subunits of FliF protein) to examine their interactions in vitro . The degree of interaction was estimated by ultracentrifugation, followed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis . From the band density on the gel, we estimated that FliG bound to the MS ring complex at an approximately 1:1 molar ratio (FliG:FliF), whereas FliM did so only at a 1:5 molar ratio (FliM:FliF) . FliN did not bind to the MS ring complex by itself or in the presence of the other switch proteins . A possible configuration of the switch proteins is discussed.

J Bacteriol, 1994 Jun, 176(12), 3673 - 82
Stoichiometry of binding of CysB to the cysJIH, cysK, and cysP promoter regions of Salmonella typhimurium; Hryniewicz MM et al.; CysB is a member of the LysR family of transcriptional activators and regulates genes of the cysteine regulon in Salmonella typhimurium and Escherichia coli . CysB binds to specific sites just upstream of the -35 regions of the cysJIH, cysK, and cysP promoters, where, in the presence of N-acetyl-L-serine, it stimulates transcription initiation . The cysK and cysP promoters contain additional binding sites, and we have proposed that CysB bends these promoters by binding to adjacent sites . N-Acetyl-L-serine is thought to decrease the magnitude of such bending . Since stoichiometric data bearing on this model have been lacking, we analyzed complexes in gel mobility shift experiments with 35S-labeled CysB and 32P-labeled promoter fragments . CysB was found to bind as a tetramer, and N-acetyl-L-serine increased the electrophoretic mobilities of one-protein complexes of the multibinding site cysK and cysP promoters without changing their stoichiometry, indicating that a single CysB tetramer can bend these promoters and that N-acetyl-L-serine diminishes such bending . Bend angles for both promoters were calculated to be 100 and 50 degrees in the absence and presence of N-acetyl-L-serine . N-Acetyl-L-serine affected neither the stoichiometry nor the electrophoretic mobility of cysJIH promoter complexes, which are not known to contain bent DNA . DNA bending may be a mechanism for sequestering CysB at certain promoter sites by increasing their affinity for this protein in the absence of N-acetyl-L-serine.

J Bacteriol, 1994 Jun, 176(12), 3631 - 7
Genetic and biochemical analysis of the flagellar hook of Treponema phagedenis; Limberger RJ et al.; The periplasmic flagellum of Treponema phagedenis consists of the flagellar filament and hook-basal body . We report here a characterization of the hook gene and flagellar hook of T . phagedenis, and in the process of this analysis we found evidence that the hook polypeptide is likely cross-linked in situ . A T . phagedenis genomic library was screened with a Treponema pallidum antiserum, and the DNA segments from several positive plaques were subcloned and sequenced . DNA sequencing of two overlapping segments revealed a 1,389-nucleotide (nt) open reading frame (ORF) with a deduced amino acid sequence that was 36% identical to that of FlgE, the hook polypeptide of Salmonella typhimurium . This gene was designated T . phagedenis flgE . Beginning at 312 nt downstream from flgE was a partial ORF of 486 nt with a deduced amino acid sequence that was 33% identical to that of MotA of Bacillus subtilis, a polypeptide that enables flagellar rotation . Upstream of flgE, separated by 39 nt, was a partial (291-nt) ORF with a deduced amino acid sequence that was homologous to that of ORF8, a polypeptide of unknown function located in an operon encoding polypeptides involved in motility of B . subtilis . The T . phagedenis flgE gene was cloned into an Escherichia coli protein expression plasmid, and the purified recombinant protein was used to prepare a FlgE antiserum . Western blots (immunoblots) of whole-cell lysates probed with this antiserum revealed a 55-kDa polypeptide and a ladder of polypeptide bands with increasing molecular masses . T . phagedenis hooks were then isolated and purified, and electron microscopic analysis revealed that the morphology of the hooks resembled that in other bacteria . The hooks were slightly curved and had an average length of 69 +/- 8 nm and a diameter of 23 +/- 1 nm . Sodium dodecyl sulfate-polyacrylamide gel electrophoresis and Western blots of purified hook preparations using the FlgE antiserum also revealed a polypeptide ladder, suggesting that the hooks are composed of a covalently cross-linked polypeptide.

J Bacteriol, 1994 Jun, 176(12), 3598 - 605
Role of the FliA-FlgM regulatory system on the transcriptional control of the flagellar regulon and flagellar formation in Salmonella typhimurium; Kutsukake K et al.; In the flagellar regulon of Salmonella typhimurium, the flagellar operons are divided into three classes, 1, 2, and 3, with respect to transcriptional hierarchy . The class 2 operons are controlled positively by the class 1 genes, flhD and flhC . The class 3 operons are controlled positively by fliA and negatively by flgM . It has been shown that FliA is a sigma factor specific for class 3, whereas FlgM is an anti-sigma factor which binds FliA to prevent its association with RNA polymerase core enzyme . Therefore, the FliA-FlgM regulatory system has been believed to control specifically the class 3 operons . In the present study, we showed that the flgM mutation enhanced the expression of class 2 by more than fivefold . When a fliA mutation was present simultaneously, this enhancement was not observed . These results indicate that the FliA-FlgM regulatory system is involved not only in the expression of class 3 but also in that of class 2 . However, though neither flhD nor flhC mutants could express the class 2 operons, the fliA mutants permitted the basal-level expression of those operons . Therefore, FlhD and FlhC are indispensable for the expression of class 2, whereas FliA is required only for its enhancement in the FlgM-depletion condition . Furthermore, we showed that the flgM mutation resulted in a two- to threefold increase in flagellar number . On the basis of these results, we propose that the relative concentration of FliA and FlgM may play an important role in the determination of flagellar numbers produced by a single cell.

J Bacteriol, 1994 Jun, 176(12), 3589 - 97
Isolation and characterization of a gene, pmrD, from Salmonella typhimurium that confers resistance to polymyxin when expressed in multiple copies; Roland KL et al.; We have isolated from Salmonella typhimurium a gene, designated pmrD, that confers resistance to the membrane-damaging drug, polymyxin B when expressed from the medium-copy-number plasmid pHSG576 . The gene maps to 46 min on the standard genetic map, near the menB gene, and is therefore distinct from the previously described pmrA locus . We have mapped the polymyxin resistance activity to a 1.3-kb ClaI-PvuII fragment which contains a small open reading frame that could encode an 85-amino-acid peptide . When an omega-Tet insertion was made into the putative pmrD open reading frame (pmrD2::omega-Tet), the resulting plasmid no longer conferred polymyxin resistance, whereas an omega-Tet insertion into vector sequences had no effect . Maxicell analysis confirmed that a protein of the expected size is made in vivo . The PmrD protein shows no significant homology to any known protein, but it does show limited homology across the active site of the p15 acid protease from Rous sarcoma virus, indicating that the protein may have proteolytic activity . However, changing the aspartic acid residue at the putative active site to alanine reduced but did not eliminate polymyxin resistance . When pmrD2::omega-Tet replaced the chromosomal copy of pmrD, the resulting strain showed wild-type sensitivity to polymyxin and could be complemented to resistance by a plasmid that carried pmrD . The pmrA505 allele confers resistance to polymyxin when present in single copy on the chromosome or when present on a plasmid in pmrA+ pmrD+ cells . In combination with the pmrD(2)::-Tet mutation, the effect o the pmrA505 allele on polymyxin resistance was reduced, whether pmrA505 was present in the chromosome or on a plasmid . Conversely, a strain carrying an insertion in pmrA could be complemented to polymyxin resistance by a plasmid carrying the pmrA505 allele but not by a plasmid carrying pmrD . On the basis of these results, we suggest that polymyxin resistance is mediated by an interaction between PmrA or a PmrA-regulated gene product and PmrD.

J Bacteriol, 1994 Jun, 176(12), 3518 - 26
Quantification of the regulation of glycerol and maltose metabolism by IIAGlc of the phosphoenolpyruvate-dependent glucose phosphotransferase system in Salmonella typhimurium; van der Vlag J et al.; The amount of IIAGlc, one of the proteins of the phosphoenolpyruvate:glucose phosphotransferase system (PTS), was modulated over a broad range with the help of inducible expression plasmids in Salmonella typhimurium . The in vivo effects of different levels of IIAGlc on glycerol and maltose metabolism were studied . The inhibition of glycerol uptake, by the addition of a PTS sugar, was sigmoidally related to the amount of IIAGlc . For complete inhibition of glycerol uptake, a minimal ratio of about 3.6 mol of IIAGlc to 1 mol of glycerol kinase (tetramer) was required . Varying the level of IIAGlc (from 0 to 1,000% of the wild-type level) did not affect the growth rate on glycerol, the rate of glycerol uptake, or the synthesis of glycerol kinase . In contrast, the growth rate on maltose, the rate of maltose uptake, and the synthesis of the maltose-binding protein increased two- to fivefold with increasing levels of IIAGlc . In the presence of cyclic AMP, the maximal levels were obtained at all IIAGlc concentrations . The synthesis of the MalK protein, the target of IIAGlc, was not affected by varying the levels of IIAGlc . The inhibition of maltose uptake was sigmoidally related to the amount of IIAGlc . For complete inhibition of maltose uptake by a PTS sugar, a ratio of about 18 mol of IIAGlc to 1 mol of MalK protein (taken as a dimer) was required.

Toxicol Lett, 1994 Jun, 72(1-3), 13 - 21
Activation of promutagens by endogenous and heterologous sulfotransferases expressed in continuous cell cultures; Glatt H et al.; Various environmental chemicals are metabolised to chemically reactive sulfuric acid esters, which may covalently bind to cellular macromolecules and induce mutations and tumours . This activation pathway is usually not taken into account in external xenobiotic-metabolising systems used in short-term tests . We therefore analysed the abilities of cytosols from mammalian cell lines to activate benzylic alcohols (1-hydroxymethylpyrene and 9-hydroxymethylanthracene) to mutagens detectable in Salmonella typhimurium TA98 . No activation was observed in cell lines which are commonly used in mutagenicity and cell transformation assays, and only low activities were found in epithelial cell lines in culture . We have therefore constructed Chinese hamster V79-derived cell lines which stably express a heterologous sulfotransferase, rat hydroxysteroid sulfotransferase a . Cytosol of these cells effectively activated 1-hydroxymethylpyrene and 9-hydroxymethylanthracene to mutagens detected in S . typhimurium . The hepatocarcinogen 6-hydroxymethylbenzo{a}pyrene induced gene mutations in sulfotransferase-expressing V79-derived cells, whereas it elicited only marginal effects in sulfotransferase-deficient control cells . The new cell lines may allow the detection of novel classes of mutagens, since some externally generated reactive sulfuric acid esters may not readily penetrate target cells due to their short life span and their ionization.

Clin Pediatr (Phila), 1994 Jun, 33(6), 349 - 52
Intravenous immunoglobulin in the treatment of Salmonella typhimurium infections in preterm neonates; Gokalp AS et al.; The purpose of this study was to determine the role of intravenous immunoglobulin (IVIG) administration in preterm neonates with S . typhimurium infection . A randomized trial of 47 preterm neonates with intestinal or extraintestinal S . typhimurium infection was performed . Neonates were randomly divided into two groups: 22 neonates were only given cefoperazone (group 1); 25 neonates were given cefoperazone plus IVIG (group 2) . IVIG was given at a dose of 500 mg/kg on days 1, 2, 3, and 8 after entry into the study . Following treatment, bacteremia, complications, mortality rate, recovery time, and duration of antimicrobial therapy were evaluated in two groups . Bacteremia was found in 31.4% in group 1 and 8% in group 2 (P < .05); complications developed in 81.8% in group 1 and 16% in group 2 (P < 0.01); mortality was 40.9% in group 1 and 12% in group 2 (P < .05) . Recovery took 15 days in group 1 and 8 days in group 2 (P < .01) . The duration of antimicrobial therapy was 20 days in group 1 and 14 days in group 2 (P < .01) . We conclude that IVIG treatment in combination with antibiotics in preterm neonates with S . typhimurium infection reduces the complications, mortality rate, and duration of therapy.

J Parasitol, 1994 Jun, 80(3), 432 - 7
Trypanosoma cruzi affects nitric oxide production by murine peritoneal macrophages; Pakianathan DR et al.; Macrophages from mice that are infected with various intracellular pathogens including Leishmania major, Trypanosoma cruzi, and Salmonella typhimurium are stimulated to produce large quantities of nitric oxide (NO) . Both viable and heat-treated L . major amastigotes have been shown to be effective co-signals for NO production in vitro . NO produced by macrophages has anti-microbial and immunosuppressive functions in an immune response . We have shown previously that NO plays a complicated role in T . cruzi infections since macrophages are important both in mediating an immune response against the parasite as well as in mediating immunosuppression . In this study we examined how T . cruzi affects NO production by macrophages from C3HeB/FeJ and C57BL/6 mice in vitro . We found that live trypomastigotes neither stimulate nor decrease NO production by interferon (IFN)-gamma-activated macrophages . However, heat-treated or glutaraldehyde-fixed trypomastigotes of T . cruzi significantly decrease NO production by IFN-gamma-activated macrophages and as a result decrease macrophage-mediated trypanocidal and immunosuppressive activity . We have determined that this decrease in NO production by T . cruzi is not due to stimulation of transforming growth factor-beta production and involves tumor necrosis factor-alpha only in C3HeB/FeJ macrophages . This study demonstrates the complexity of the T . cruzi-macrophage interaction as well as confirms previously demonstrated differences between macrophages from 2 strains of mice.

J Bacteriol, 1994 Jun, 176(11), 3420 - 7
Cloning and sequencing of the genes from Salmonella typhimurium encoding a new bacterial ribonucleotide reductase; Jordan A et al.; A plasmid library of Salmonella typhimurium was used to complement a temperature-sensitive nrdA mutant of Escherichia coli . Complementation was obtained with two different classes of plasmids, one carrying the E . coli nrdAB-like genes and the second containing an operon encoding a new bacterial ribonucleotide reductase . Plasmids harboring these new reductase genes also enable obligately anaerobic nrdB::Mud1 E . coli mutants to grow in the presence of oxygen . This operon consists of two open reading frames, which have been designated nrdE (2,145 bp) and nrdF (969 bp) . The deduced amino acid sequences of the nrdE and nrdF products include the catalytically important residues conserved in ribonucleotide reductase enzymes of class I and show 25 and 28% overall identity with the R1 and R2 protein, respectively, of the aerobic ribonucleoside diphosphate reductase of E . coli . The 3' end of the sequenced 4.9-kb fragment corresponds to the upstream region of the previously published proU operon of both S . typhimurium and E . coli, indicating that the nrdEF genes are at 57 min on the chromosomal maps of these two bacterial species . Analysis of the nrdEF and proU sequences demonstrates that transcription of the nrdEF genes is in the clockwise direction on the S . typhimurium and E . coli maps.

J Bacteriol, 1994 Jun, 176(11), 3400 - 2
The Escherichia coli efg gene and the Rhodobacter capsulatus adgA gene code for NH3-dependent NAD synthetase; Willison JC et al.; The essential gene efg, which complements ammonia-dependent growth (adgA) mutations in Rhodobacter capsulatus and is located at 38.1 min on the Escherichia coli chromosome, was found to code for NH3-dependent NAD synthetase . Crude extracts from a strain which overproduces the efg gene product contained up to 400 times more activity than crude extracts from the control strain, and the purified Efg protein possessed-NH3-dependent NAD synthetase activity . Glutamine-dependent NAD synthetase activity was found in crude extracts of E . coli but not in the purified enzyme, suggesting that it may be catalyzed by an additional subunit . An R . capsulatus strain carrying an adgA mutation was found to be deficient in NAD synthetase activity, and activity was restored by complementation with the E . coli gene . In accordance with the nomenclature proposed for Salmonella typhimurium (K . T . Hughes, B . M . Olivera, and J . R . Roth, J . Bacteriol . 170:2113-2120, 1988), the efg and adgA genes should now be designated nadE.

J Bacteriol, 1994 Jun, 176(11), 3196 - 203
Cloning, DNA sequence, and complementation analysis of the Salmonella typhimurium hemN gene encoding a putative oxygen-independent coproporphyrinogen III oxidase; Xu K et al.; Coproporphyrinogen oxidation is a last step in heme biosynthesis . The biochemically characterized eukaryotic coproporphyrinogen III oxidases have an obligate requirement for molecular oxygen, and a similar enzyme is encoded by the hemF gene in Salmonella typhimurium . Anaerobic heme synthesis requires an oxygen-independent coproporphyrinogen oxidase, which is probably encoded by the hemN gene in S . typhimurium . The hemN gene has been cloned from an insertion mutant . The nucleotide sequence was obtained and used for PCR amplification of the wild-type gene . A single open reading frame was identified as the hemN gene on the basis of its interruption by the insertion mutation and plasmid complementation studies of hemF hemN double mutants . The predicted HemN protein has 38% amino acid sequence identity to a putative anaerobic Rhodobacter sphaeroides coproporphyrinogen oxidase . The hemN RNA 5' end and the inferred transcription initiation site were mapped by primer extension . The 52.8-kDa HemN protein is expressed from the second ATG codon of the hemN open reading frame . An open reading frame with an unknown function directly upstream of hemN has a striking amino acid sequence, including 11 acidic residues in a row.

Infect Immun, 1994 Jun, 62(6), 2590 - 9
Immune responses to Yersinia enterocolitica in susceptible BALB/c and resistant C57BL/6 mice: an essential role for gamma interferon; Autenrieth IB et al.; Susceptibility of mice to infection with Yersinia enterocolitica has been shown to be related to neither the Ity locus encoding for resistance to Salmonella typhimurium and other pathogens nor the H-2 locus . Recent studies in our laboratory have demonstrated that T-cell-mediated immune responses are required for overcoming primary Yersinia infection . In the present study, we investigated the course of infection with Y . enterocolitica and the resulting immune responses in Yersinia-susceptible BALB/c and Yersinia-resistant C57BL/6 mice . In the early phase of infection, the clearance of the pathogen was comparable in both strains of mice, suggesting similar mechanisms of innate resistance . Splenic T cells from Yersinia-infected C57BL/6 mice exhibited marked proliferative responses and produced gamma interferon (IFN-gamma) upon exposure to heat-killed yersiniae . By contrast, the Yersinia-specific T-cell response in BALB/c mice was weak, and IFN-gamma production could not be detected before day 21 postinfection . T cells isolated from C57BL/6 mice 7 days after infection mediated immunity to Y . enterocolitica but those from BALB/c mice did not, while at 21 days postinfection T cells from both strains mediated protection . Neutralization of IFN-gamma abrogated resistance to yersiniae in C57BL/6 mice but to a far smaller extent in BALB/c mice . Administration of recombinant IFN-gamma or anti-interleukin-4 antibodies rendered BALB/c mice resistant to yersiniae, whereas this treatment did not significantly affect the course of the infection in C57BL/6 mice . These results indicate that the cellular immune response, in particular the production of IFN-gamma by Yersinia-specific T cells, is associated with resistance of mice to Y . enterocolitica.

Infect Immun, 1994 Jun, 62(6), 2285 - 8
Toxicity of lipopolysaccharide and of soluble extracts of Salmonella typhimurium in mice immunized with a live attenuated aroA salmonella vaccine; Mastroeni P et al.; Mice immunized intravenously 10 days earlier (but not those immunized 2 months earlier) with an attenuated Salmonella typhimurium SL3261 aroA live vaccine and tested for delayed-type hypersensitivity by injection of crude Salmonella extracts in the footpad can die within 24 to 48 h of an unexplained allergic reaction . The lethal reaction could be prevented by prior administration of anti-tumor necrosis factor alpha serum . Injection of lipopolysaccharide (LPS) (either purified phenol-water-extracted {Westphal} LPS or protein-rich trichloracetic acid-extracted {Boivin} LPS) was also lethal for mice immunized 10 days before . An LPS-rich crude Salmonella extract was more toxic than one which contained less LPS, suggesting that LPS may have been involved in the lethal reactions to crude antigens . Mild alkaline hydrolysis removes O-linked acyl groups from lipid A and eliminates many toxic effects of LPS; however, both Boivin LPS and Westphal LPS remained toxic for immunized mice after alkaline hydrolysis . In contrast, alkaline hydrolysis of crude whole Salmonella extracts (which caused marked protein degradation) reduced the lethal toxicity of the extracts, especially for an LPS-rich preparation . Mice immunized orally with the live vaccine did not show hypersensitivity to either LPS or crude extracts . The results suggest that the lethal reaction to crude Salmonella antigens in mice immunized 10 days earlier is complex, that tumor necrosis factor alpha is involved, and that allergic reactions to crude antigens (but not to LPS alone) can be reduced by mild alkaline hydrolysis.

Clin Infect Dis, 1994 Jun, 18(6), 873 - 8
Analysis of factors influencing the outcome and development of septic metastasis or relapse in Salmonella bacteremia; Galofre J et al.; One-hundred seventy-two consecutive adult patients with salmonella bacteremia documented by at least one positive blood culture were prospectively evaluated over a 10-year period . Salmonella enteritidis was isolated in 121 cases (70.3%), Salmonella typhimurium in 29 (16.9%), and other Salmonella species in 22 (12.8%) . Twenty-seven patients (15.7%) developed septic metastasis; 21 patients (12.2%) died of bacteremia, and 24 (16.7%) of the 144 patients who survived had at least one relapse . A logistic regression analysis selected three variables as independently influencing outcome: septic shock (P = .005), coma (P = .029), and immunosuppression (P = .04) . By means of the same statistical analysis, leukopenia (a white blood cell count of < 4 x 10(9)/L) was identified as an independent risk factor for relapse (P < .0001) . The possibility of salmonella bacteremia must be considered when immunosuppressed patients have fever and no obvious source of infection . Treatment with a drug active against Salmonella species is essential in this population . Patients with leukopenia should be considered as recipients of prophylaxis for relapse.

Biol Reprod, 1994 Jun, 50(6), 1297 - 302
Effect of Salmonella endotoxin administered to the pregnant sheep at 133-142 days gestation on fetal oxygenation, maternal and fetal adrenocorticotropic hormone and cortisol, and maternal plasma tumor necrosis factor alpha concentrations; Schlafer DH et al.; We studied the effects of maternal exposure to endotoxin on pregnancy outcome, stimulation of the hypothalamo-hypophyseal-adrenal axes (HHAA) of ewes and their fetuses, and maternal production of tumor necrosis factor alpha (TNF alpha), a central mediator in the pathogenesis of bacterial endotoxemia . Either endotoxin (lipopolysaccharide, LPS) from Salmonella typhimurium or saline was administered to pregnant sheep (n = 10) at 137.6 +/- 3.5 days gestational age by slow i.v . infusion at 1 microgram.kg-1 total dose over a 3-h period . Fetuses of ewes treated with LPS became hypoxemic by 2.5 h after initiation of LPS infusion . Maternal ACTH increased and peaked during LPS administration to levels approximately 10-fold greater in treated than in control ewes . In contrast, fetal ACTH levels were maximum at 6-12 h . Maternal and fetal cortisol levels were significantly different from control levels by 1 and 12 h, respectively . Maternal plasma TNF alpha peaked (6-7-fold over baseline) at 1-2 h after initiation of LPS administration and steadily declined over the following 48 h . Four of 5 treated ewes either delivered or their fetuses died within 28 h . In summary, infusion of LPS caused fetal hypoxemia and stimulated both fetal and maternal HHAA followed by preterm labor by 28 h after infusion in 4 of 5 ewes . Three of the fetuses died in utero . Maternal plasma TNF alpha rose rapidly, but the specific role it may play in initiation of preterm labor remains to be elucidated.

Genetics, 1994 Jun, 137(2), 353 - 9
Poisson-like fluctuation patterns of revertants of leucine auxotrophy (leu-500) in Salmonella typhimurium caused by delay in mutant cell division; Dijkmans R et al.; Leu+ mutants from Salmonella typhimurium leu-500 strain MA412 arise at high frequencies and mutant colonies appear over a broad range of time on selective plates . This observation suggested that these Leu+ mutants might be induced or "directed."= If such a mechanism was responsible, mutants should originate on selective plates rather than in the preceding culture in nonselective conditions and should give rise to Poisson-like fluctuation curves upon plating of sister cultures on selective medium . Poisson-like distribution profiles were indeed observed for Leu+ mutants of S . typhimurium MA412 . However, an explanation for the observed Poisson-like fluctuation patterns without a need for selection-induced mutations was found . Microscopical analysis and cell mass/viable count measurements showed that the size of Leu+ mutant cells was often much larger than those of nonmutants . This size difference was a stable characteristic of a large proportion of Leu+ mutants, was observed both in stationary and growing culture and did not measurably affect the division rates of the cells in nutrient broth . As the transition from normal-sized nonmutant to oversized mutant cells during the nonselective culture phase of the fluctuation experiment may have been accompanied by a period with no or few completed cell division cycles, the number of mutant offspring may have been smaller than that of sibling nonmutants . Such underrepresentation of mutants in the final culture is expected to give rise to Poisson-like fluctuation patterns without invoking "directed" mutations.

Berl Munch Tierarztl Wochenschr, 1994 Jun, 107(6), 192 - 8
{The effectiveness of Salmonella immunization of broiler breeders on the Salmonella colonization of the animals and their progeny after experimental oral infection}; Methner U et al.; The immunization of broiler breeder birds with Salmonella typhimurium live or inactivated vaccines (orally and/or parenterally administered) resulted in a considerable antibody response . Maternally produced antibodies were transferred via egg to the progeny . But the transferred antibodies did not increase considerably the resistance of 1 day old chickens to oral Salmonella typhimurium infection . The results of the oral challenge of the broiler breeder birds showed the protective effect induced by the oral immunisation with the Salmonella live vaccine strain . This protection is expressed by a reduced number of salmonella organisms in the caeca and internal organs.

Pharmazie, 1994 Jun, 49(6), 448 - 51
An evaluation of 55 commercial plant extracts in the Ames mutagenicity test; Schimmer O et al.; 55 commercial phytopharmaceuticals (extracts and tinctures) from 44 plant species were evaluated for mutagenic potential in the Salmonella/mammalian microsome mutagenicity test (Ames assay), utilizing tester strains TA98 and TA100 of Salmonella typhimurium with and without S9 mix from induced rat liver microsomes . Weak activities were detected after exposure of the bacteria to Alchemillae tinctura, Centaurii extractum, Hippocastani extractum and Myrtilli extractum . Moderate effects were observed with Crataegi extractum, Echinaceae angustifoliae extractum, Hyperici tinctura, Rutae tinctura and Trifolii fibrini extractum and tinctura . Quercetin was detected by TLC in all extracts with mutagenic activity except in Echinaceae angustifoliae and Centaurii extractum . From this study and earlier results we suggest that quercetin is possibly the main mutagenic principle in the following phytopharmaceuticals: Alchemillae tinctura, Cratagei extractum, Hippocastani extractum, Hyperici tinctura, Myrtilli extractum, Trifolii fibrini extractum and Trifolii fibrini tinctura.

Chem Biol Interact, 1994 Jun, 92(1-3), 305 - 19
1-Hydroxymethylpyrene and its sulfuric acid ester: toxicological effects in vitro and in vivo, and metabolic aspects; Glatt H et al.; 1-Hydroxymethylpyrene (HMP) is activated to a potent mutagen, detectable in Salmonella typhimurium, in the presence of hepatic cytosol, cofactor for sulfotransferases, and chloride anions . The number of induced mutations is linear to the amount of cytosol used over a wide range, allowing for the quantification of this activity . The activity is expressed with high selectivity in certain tissues and cell types . In adult rats, the highest level is found in the liver, the activity in females exceeding that in males about threefold . About half of the activity in the liver of females is provided by hydroxysteroid sulfotransferase a (STa), whereas other enzymes may be more important in males on account of their very low level of STa . The expression of STa is decreased in ATPase-negative, presumably preneoplastic, hepatic foci in female rats . In contrast to its high mutagenicity in bacteria, SMP shows only weak mutagenic activity in mammalian cells (Chinese hamster V79 cells), independently of whether it is externally added, or generated from HMP within the cells by heterologously expressed STa . Sulfation, however, strongly enhances the cytotoxicity of HMP in mammalian cells . The high cytotoxicity and low mutagenicity in mammalian cells in culture have possible correlates in vivo: while HMP is only a weak initiator of ATPase-negative hepatic foci in newborn rats, it shows substantial promoting activity with regard to such foci in female, but not in male rats . We postulate that this promotion results from selective toxification by STa in the normal hepatic parenchyma of female rats, and resistance of ATPase/STa-negative foci.

J Appl Bacteriol, 1994 Jun, 76(6), 626 - 31
Factors that interact with the antibacterial action of thyme essential oil and its active constituents; Juven BJ et al.; The viable counts of Salmonella typhimurium on nutrient agar (NA) decreased upon the addition of either the essential oil of thyme or its constituent thymol, especially under anaerobic conditions . Antagonistic effects of thymol against Staphylococcus aureus were also greater under anaerobic conditions . In contrast to the phenolic constituents of the oil, thymol and carvacrol, the chemically related terpenes p-cymene and gamma-terpinene had no antagonistic effects against Salm . typhimurium . The addition of Desferal to NA counteracted the antibacterial effects of both thyme oil and thymol . No support was obtained, however, for a possible role of iron in the oxygen-related antibacterial action of the thyme oil and thymol or for the observed effect of Desferal . In the presence of thymol, the viable counts of Salm . typhimurium obtained on a minimal medium (MM) were lower than those obtained on NA . Addition of bovine serum albumin (BSA) neutralized the antibacterial action of thymol . It is suggested that the effects of BSA or Desferal are due to their ability to bind phenolic compounds through their amino and hydroxylamine groups, respectively, thus preventing complexation reactions between the oil phenolic constituents and bacterial membrane proteins . This hypothesis is supported by the marked decrease in the viable counts of Salm . typhimurium caused by either thyme oil or thymol when the pH of the medium was changed from 6.5 to 5.5 or the concentration of Tween 80 in the medium was reduced.

Genetika, 1994 Jun, 30(6), 769 - 75
{UV-induction of the LT-toxin operon depending on genes lexA, recA, and umuD}; Tiganova IG et al.; UV induction of the elt operon (the LT-toxin operon in Escherichia coli) was demonstrated in experiments using fusion of elt::lac operons with the help of Mud1(Ap lac) phage . UV induction of the elt operon is lexA-dependent; thus, the possibility of SOS regulation of this process may be assumed . However, UV induction of the elt operon turned out to be recA-independent, which makes it impossible to consider this induction as a typical SOS response . UV induction of the elt operon is also observed in Salmonella typhimurium, which differs from E . coli in the product of umuD, which suggests that the UV induction of the elt operon is umuD independent.

Biol Pharm Bull, 1994 Jun, 17(6), 819 - 22
Formation of chloroamines from styrene under conditions mimicking those of water "chlorination" treatment; Nojima K et al.; The reaction of styrene with sodium hypochlorite in the presence of ammonium ion in 0.3 M phosphate buffer (pH 6.0) at room temperature afforded N-chloro-1-phenyl-2-chloroethylamine and N,N-dichloro-1-phenyl-2-chloroethylamine . 1-Phenyl-2-chloroethylamine hydrochloride, obtained as the reduction product of the two N-chloroamines, was shown to be mutagenic in the Salmonella typhimurium test.

Biomed Environ Sci, 1994 Jun, 7(2), 144 - 9
Reversion of bioluminescent bacteria (Mutatox) to their luminescent state upon exposure to organic compounds, munitions, and metal salts; Arfsten DP et al.; Mutatox is a new genotoxicity bioassay which uses as the endpoint the bioluminescence produced on reversion of a dark strain of the marine bacterium Vibrio fischeri +/- S9 . Reversion can occur by several mechanisms, including base substitution, frame-shift, SOS induction, and DNA intercalation . For screening, Mutatox provides many advantages over the Salmonella typhimurium (Ames) assay: it requires minimal sterility, employs a shorter incubation period, and does not require culture maintenance . Eighteen organic chemicals (phenol, polynuclear aromatic hydrocarbons, nitrotoluenes, others), Na3PO4, and 4 genotoxic metals (Cu2+, Ni2+, As3+, Cd2+) were tested . Most of the organic compounds positive in S . typhimurium assays were positive in Mutatox . None of the metals was genotoxic in V . fischeri, possibly due to poor uptake from the saline medium.

Am J Vet Res, 1994 Jun, 55(6), 785 - 9
Effect of mimicking prepartum concentration of estradiol-17 beta on the inflammatory response to endotoxin in gilts; Magnusson U et al.; The effect of mimicking prepartum concentration of estradiol-17 beta on the inflammatory response to endotoxin in gilts was studied . The study was performed in a split-litter design and comprised 5 pairs of littermates . A catheter was inserted into the jugular vein 2 days prior to the start of the study . In each pair, 1 littermate was treated IM with 2.5 mg of estradiol-17 beta/75 kg of body weight, and the other littermate was given peanut oil IM as a control . The day after treatment, all gilts were challenge-exposed with a Salmonella typhimurium-derived endotoxin (1 microgram/kg, IV) and the inflammatory response to challenge exposure was monitored . There was no effect of estradiol treatment on the transient clinical signs of endotoxemia or on the increase in rectal temperature . The increase in blood concentrations of prostaglandin F2 alpha metabolite and cortisol after endotoxin challenge exposure was not affected by estradiol . Decrease in number of circulating blood mononuclear cells and polymorphonuclear leukocytes was not changed by estradiol treatment . Taken together, mimicking prepartum concentration of estradiol did not affect either the magnitude or the kinetics of the inflammatory response to endotoxin in gilts . Relevance of these findings to development of endotoxin-mediated diseases, such as the postpartum agalactia syndrome, needs further study.

Southeast Asian J Trop Med Public Health, 1994 Jun, 25(2), 328 - 31
Incidence of salmonellae in duck eggs in Thailand; Saitanu K et al.; Detection of salmonellae was performed on egg shells and egg contents of duck eggs . Five hundred and sixty-four tested samples were came from 1,128 eggs, 2 eggs in each sample . Eggs were collected from retail markets in Bangkok, Chon Buri, Chachoengsao, Lop Buri, Ang Thong and Nakhon Ratchasima provinces during January through June 1992 . The percentage of salmonellae contamination on the egg shells only, egg contents only and both shells and contents were 12.4%, 11% and 0.2%, respectively . Twenty three serotypes were identified from the 133 salmonellae isolates . The common serotypes found from duck eggs were Salmonella typhimurium, S . cerro, S . tennessee, S . amsterdam, S . agona and S . infantis accounting for 5.5%, 4.1%, 2.8%, 2.1%, 1.4% and 1.1%, respectively.

Res Microbiol, 1994 Jun-Aug, 145(5-6), 473 - 80
The role of the PhoP/PhoQ regulon in Salmonella virulence; Garcia Vescovi E et al.; Salmonella typhimurium is a facultative intracellular pathogen that is able to survive in a wide variety of inhibitory and nutritionally deprived host environments . The ability to survive under such hostile conditions, which are often encountered during the course of infection, contributes to its pathogenic properties . Some of the virulence determinants of S . typhimurium are under the transcriptional control of the PhoPQ two-component regulatory system . Several virulence phenotypes have been associated with mutations in the phoPQ operon including the inability to survive within macrophages and increased susceptibility to antimicrobial peptides and acid pH . Only 25% of PhoP-modulated genes are involved in virulence and the phoPQ operon is present in both pathogenic and non-pathogenic microbes . These data suggest that PhoP is not exclusively involved in virulence and that it is required for the physiological control of activities common to other bacteria.

Zentralbl Bakteriol, 1994 Jun, 281(1), 8 - 15
Purification and partial characterization of type 3 fimbriae from Salmonella typhimurium var . copenhagen; Stolpe H et al.; Aggregative thin fimbriae from a pigeon pathogen, Salmonella typhimurium var . copenhagen (STMVC) Mo 8 were isolated and purified . These fimbriae remained associated with the cells even after attempts to separate them from blended cells by centrifugation . After purification, fimbriae and little cell fractions were polymerized in formic acid and then analyzed by SDS-PAGE . This pretreatment resulted in the appearance of a main protein band of 17 kDa . The N-terminal amino acid sequence of 19 residues of purified 17 kDa protein showed considerable homology with the N-terminal sequence of thin fimbriae of S . enteritidis . Native fimbriae on whole cells were specifically labelled with immune serum raised to the purified fimbriae . This immune serum also reacted with the denatured 17 kDa protein in Western blots . The polyclonal immune serum did not cross-react with the type-1 fimbriae produced by STMVC.

Tokushima J Exp Med, 1994 Jun, 41(1-2), 57 - 64
The efficiency of solvent extraction of mutagenic compounds in particulates exhausted from a small diesel engine; Tahara I et al.; Organic materials were extracted from particulates exhausted from a small diesel engine (displacement 269 ml) by the ultrasonic extraction method with three different solvent systems, methanol, dichloromethane and a 4:1 (v:v) mixture of benzene and ethanol . These solvent-extracted materials were tested for mutagenic activity by the Ames Salmonella/microsome assay system using Salmonella typhimurium strains TA98, TA100, TA98NR and TA98/1,8-DNP6 . The concentrations of 1-nitropyrene (1-NP) and 1,6-dinitropyrene (1,6-diNP) in these extracted materials were also measured after nitroreduction by high pressure liquid chromatography . The methanol-extracted and benzene-ethanol-extracted materials showed the lowest and the highest mutagenic activity, respectively . The methanol-extracted, dichloromethane-extracted and benzene-ethanol-extracted materials induced 260, 1,570 and 3,240 His+ revertants per plate per mg of extracted materials, respectively, from strain TA98 in the absence of S9 mix . These materials showed decreased mutagenicity for strains TA98NR and TA98/1,8-DNP6, indicating that the particulates in the diesel engine exhaust contained 1-NP and diNPs . Actually, the amount of 1-NP and 1,6-diNP in the methanol-extracted, dichloromethane-extracted and benzene-ethanol-extracted materials were 17.0 and 0.03 ng, 37.5 and 0.97 ng, and 71.3 and 1.03 ng per mg of extracted materials, respectively, accounting for 11.9 and 3.2%, 4.4 and 17.3%, and 4.0 and 8.9%, respectively, of the total mutagenicity of the extracted materials . From these results it is concluded that a mixture of benzene-ethanol (4:1, v/v) is the most suitable solvent for extraction of organic matter containing nitrated polycyclic aromatic hydrocarbons such as NPs from particulates in diesel engine exhaust.

Mutat Res, 1994 Jun, 324(1-2), 43 - 50
Microbial mutagenicity and in vitro chromosome aberration induction by FK973, a new antitumor agent; Hirai O et al.; The genotoxic activity of a new antitumor agent, FK973, was compared with that of mitomycin C (MMC) in eukaryotic and prokaryotic cells . In chromosome aberration tests using Chinese hamster fibroblast Don cells, FK973 induced a dose-related increase of aberrant cells after 6 h-pulse treatments, and the minimum effective concentrations with and without S9 were 0.625 and 0.0625 micrograms/ml, respectively . The compound increased revertant colonies in Salmonella typhimurium TA102 at the dose range of 10-5000 micrograms/plate with S9 . Without S9, FK973 induced a small increase at the dose range of 500-5000 micrograms/plate in two of three independent experiments, but the number of revertant colonies was less than double that of the vehicle control . The compound did not induce any revertant colonies in colonies in S . typhimurium TA100, TA98, TA1535 or TA1537 with or without S9 . MMC was confirmed to increase both chromosome aberrations in Don cells and revertant colonies in TA102 . The minimum clastogenic and mutagenic concentrations without S9 were 0.0156 microgram/ml and 0.005 microgram/plate, respectively . The results indicate that FK973 needs metabolic activation to induce reverse mutation in prokaryotic cells, but caused chromosome aberrations in mammalian cells without added S9.

Mutat Res, 1994 Jun, 321(4), 253 - 64
Mutagenicity of coal-dust and smokeless-tobacco extracts in Salmonella typhimurium strains with differing levels of O-acetyltransferase activities; Stamm SC et al.; Epidemiological studies have indicated an increased incidence of gastric neoplasia in coal miners . Because smokeless tobacco use is prevalent in the mining industry, nitrites or other components of these products may be etiologically associated with these gastric neoplasms . In this study both nitrosated and non-nitrosated coal-dust (from West Virginia and New Mexico) as well as smokeless-tobacco (snuff and chewing tobacco) extracts were examined for the presence of aromatic amines and nitroarenes by comparing the activities of these extracts in the pre-incubation variant of the Ames assay . Salmonella strains with differing O-acetyltransferase activities (TA98 and YG1024) were utilized in this investigation . The results of the examination of the coal-dust extracts indicated positive activity only in the nitrosated extracts . Both nitrosated extracts elicited an increased number of revertants (2-4-fold) on YG1024 without S9 in comparison to TA98, suggesting the presence of nitroarenes in these extracts . Additionally, the nitrosated West Virginia coal extract showed higher levels of activity on YG1024 with S9, indicating the possible presence of aromatic amines in this complex mixture . The non-nitrosated smokeless-tobacco extracts showed activity only on YG1024 in the presence of S9, with the highest amount of activity occurring in the snuff sample . Except for the chewing-tobacco extract on TA98 without S9, positive activity was found in both nitrosated tobacco extracts on YG1024 and TA98 . As with the coal extracts, the presence of nitroarenes was inferred for these nitrosated materials . A comparative study of the non-nitrosated snuff extract across 5 tester strains with varying sensitivities to aromatic amines and nitroarenes (TA98NR, TA98/1,8-DNP6, TA98, YG1021 and YG1024) indicated that aromatic amines were a probable source of the mutagenic activity . The curing process and/or the addition of certain flavorants are potential sources of the mutagenic aromatic amines suggested to be present in the non-nitrosated snuff extract . These findings are consistent with an etiologic role supplementary to the nitroso compounds for mutagenic nitroarenes and aromatic amines in the development of gastric neoplasia in coal miners.

Mutat Res, 1994 Jun, 321(4), 197 - 201
Studies on the mutagenicity of a peptoplast adhesive in Salmonella typhimurium; Leuschner J et al.; A new class of adhesives--peptoplasts--was tested for their mutagenic potential in Salmonella typhimurium . The tested peptoplasts revealed no mutagenic properties under the present test conditions.

Mutat Res, 1994 Jun, 321(4), 187 - 95
Mutagenic activity of newly synthesized sulfa drugs to Salmonella typhimurium; Temcharoen P et al.; The mutagenic activity of seven newly synthesized sulfa drugs was studied in Salmonella typhimurium, using forward mutation to 8-azaguanine (8-AG) resistance and reversion mutation assays (Ames test) both in the absence and presence of Aroclor induced rat liver S9 . In forward mutation assays, N1-methylsulfanilamide, N4-acetyl-N1-methylsulfanilamide and N4-acetyl-N1-diethylsulfanilamide were mutagenic to S . typhimurium TM677 both in the presence and absence of metabolic activation while N4-acetylsulfanilamide, N1-diethylsulfanilamide and 4-nitro-N-2-pyridinylbenzenesulfonamide {2-(p-nitrobenzenesulfonamido)pyridine} were mutagenic only in the presence of metabolic activation . But 2-(N4-acetylsulfanilamido)pyridine was mutagenic in neither the presence nor the absence of metabolic activation . However, none of the seven compounds had any mutagenic effect on S . typhimurium TA98 or TA100 in the absence or presence of metabolic activation, by the Ames test preincubation method . The relationship between the structure of the compounds and their mutagenic activity is also discussed.

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