Mol Microbiol, 2003 Jan, 47(2), 335 - 44 Control of the Salmonella ugd gene by three two-component regulatory systems; Mouslim C et al.; The Salmonella ugd gene is required for the incorporation of 4-aminoarabinose in the lipopolysaccharide and resistance to the antibiotic polymyxin B . Transcription of the ugd gene is induced by Fe3+ via the PmrA-PmrB two-component system and by low Mg2+ in a process that requires the PhoP-PhoQ two-component system, the PhoP-activated PmrD protein and the PmrA-PmrB system . Here, we establish that mutation of the tolB gene promotes ugd transcription independently of both the PhoP-PhoQ and PmrA-PmrB systems . This activation is mediated by the RcsC-YojN-RcsB phosphorelay and the RcsA protein, suggesting a role for ugd in capsule synthesis . Binding sites for the RcsB, PmrA and PhoP proteins were identified in the ugd promoter . Although the PmrA-PmrB and RcsC-YojN-RcsB systems promoted ugd transcription independently of the PhoP-PhoQ system under different environmental conditions, ugd expression inside macrophages was strictly dependent on PhoP-PhoQ, suggesting that low Mg2+ is a cue for the intracellular environment. J Clin Microbiol, 2003 Jan, 41(1), 460 - 2 Characterization of the first extended-spectrum beta-lactamase-producing Salmonella isolate identified in Canada; Mulvey MR et al.; A single Salmonella enterica serovar Typhimurium isolate with an UT2 phage type producing an extended-spectrum beta-lactamase (ESBL) was identified in Canada in 2000 . The isolate harbored two plasmids, one containing a bla(TEM-1) gene and the other containing a bla(SHV-2a) gene . The ESBL gene was located on a 70-kb transferable plasmid which also carried tetracycline and trimethoprim resistance elements. J Clin Microbiol, 2003 Jan, 41(1), 27 - 33 Subtyping of Salmonella enterica serotype enteritidis strains by manual and automated PstI-SphI ribotyping; Clark CG et al.; Salmonella enterica subsp . enterica serotype Enteritidis is not readily subtyped beyond the level of phage type (PT) . A recently developed method for ribotyping of this organism, which uses a mixture of PstI and SphI (PS) for restriction of DNA (PS ribotyping), has proved useful for further subtyping of a number of PTs of this organism, including PT 4 . However, it has not been extensively tested with PT 8 . In the present study the PS ribotyping method was used to investigate outbreaks of both S . enterica serotype Enteritidis PT 4 and PT 8 and provided subtyping data that were consistent with information obtained from epidemiologic investigations . The method proved to be more discriminatory than phage typing and pulsed-field gel electrophoresis (PFGE) combined and was useful for investigating a pseudo-outbreak involving isolates that had identical PTs and PFGE types but that could not be linked epidemiologically . Several PS ribotypes were found within the cluster of isolates indistinguishable by other subtyping methods, confirming the epidemiologic findings . Although the PS ribotyping method proved to have a superior discriminatory ability in resolving clusters, it did not have high enough throughput for use in outbreak investigations . This method has therefore been adapted for use in automated ribotyping with a RiboPrinter, and the results were compared with those obtained by manual ribotyping . Both methods produce equivalent results and are useful for obtaining epidemiologically relevant subtyping data for S . enterica serotype Enteritidis, including PT 8 strains not extensively tested previously. J Agric Food Chem, 2003 Jan 15, 51(2), 345 - 9 Bacteriological and genetic assessment of game meat from Japanese wild boars; Naya Y et al.; Bacterial tests were used to assess bacterial contamination of game meat from Japanese wild boars . The bacterial contamination of wild boar meat was less than that of domestic pork, as determined by aerobic plate counts (APC) and coliform counts . None of the meat examined in this study was contaminated by Salmonella or E . coli O-157 . To detect adulteration by domestic pig meat or European wild boar meat, 46 samples of game meat sold as Japanese wild boar were examined genetically . A total of 17 samples showed genetic haplotypes of European and Asian domestic pigs in the D-loop of mitochondrial DNA (mtDNA), and 16 samples showed nuclear glucosephosphate isomerase-processed pseudogene (GPIP) genotypes of European domestic pigs . The European GPIP genotypes of these samples were confirmed by PCR-RFLP analysis . These results indicate that some game meat sold as Japanese wild boar is adulterated by cross-breeding between pigs and wild boars or by contamination with meat from domestic pigs or European wild boars. J Exp Med, 2003 Jan 6, 197(1), 63 - 75 Critical role of the carboxyl terminus of proline-rich tyrosine kinase (Pyk2) in the activation of human neutrophils by tumor necrosis factor: separation of signals for the respiratory burst and degranulation; Han H et al.; Transduction of Tat-tagged fusion proteins confirmed a hypothesis based on pharmacologic inhibitors (Fuortes, M., M . Melchior, H . Han, G.J . Lyon, and C . Nathan . 1999 . J . Clin . Invest . 104:327-335) that proline-rich tyrosine kinase (Pyk2) plays a critical role in the activation of adherent human neutrophils, and allowed an analysis of individual Pyk2 domains not possible with chemical inhibitors . Acting as a dominant negative, the COOH terminus of Pyk2 fused to a Tat peptide (Tat-CT), but not other regions of Pyk2, specifically inhibited the respiratory burst of cells responding to tumor necrosis factor (TNF), Salmonella, or Listeria, while sparing responses induced by phorbol ester . Tat-CT suppressed TNF-triggered cell spreading and the phosphorylation of endogenous Pyk2 and the associated tyrosine kinase Syk without blocking the ability of neutrophils to degranulate and kill bacteria . Thus, separate signals control the respiratory burst and degranulation, and a normal rate of killing of some bacteria can be sustained by granule products in conjunction with a minimal residual respiratory burst . Inhibition of select inflammatory functions without impairment of antibacterial activity may commend the Pyk2 pathway as a potential target for antiinflammatory therapy. Appl Environ Microbiol, 2003 Jan, 69(1), 668 - 72 Inactivation of Salmonella enterica serovar enteritidis by ultrasonic waves under pressure at different water activities; Alvarez I et al.; The inactivation of Salmonella enterica serovar Enteritidis by ultrasonic waves (20 kHz; 117- microm wavelength) under pressure (175 kPa) at nonlethal temperatures (manosonication {MS}) and lethal temperatures (manothermosonication {MTS}) in media of different water activities has been investigated . Heat decimal reduction time values increased 30 times when the water activity was decreased from nearly 1 to 0.96, but the MS resistance was increased only twofold . The inactivation of Salmonella serovar Enteritidis by ultrasound under pressure at low water activities was a phenomenon of the "all-or-nothing" type . A synergistic lethal effect was observed between heat and ultrasound in media with reduced water activity; the lower the water activity, the greater the synergistic effect . This work could be useful for improving sanitation and preservation treatments of foods, especially those which are sensitive to temperature and those in which components protect microorganisms to heat . It also contributes to our knowledge of microbial inactivation mechanisms by MS and MTS treatments. Appl Environ Microbiol, 2003 Jan, 69(1), 548 - 53 Growth dynamics of Salmonella enterica strains on alfalfa sprouts and in waste seed irrigation water; Howard MB et al.; Alfalfa sprouts and other seed sprouts have been implicated in numerous outbreaks of salmonellosis . The source of these epidemics appears to have been low-level contamination of seeds by Salmonella bacteria that developed into clinically significant populations during the seed germination process . To test the possibility that Salmonella enterica strains carry host range determinants that allow them to grow on alfalfa, strains isolated from alfalfa or other sources were surveyed for their ability to grow on germinating alfalfa seeds . An S . enterica serovar Cubana strain originally isolated from contaminated alfalfa sprouts multiplied most rapidly during the initial 24 h of the seed germination process . Germinating alfalfa seeds supported the multiplication of S . enterica cells prior to the emergence of the root radicle at 72 h . Thereafter, much lower rates of multiplication were apparent . The ability of S . enterica to grow on germinating alfalfa seeds was independent of the serovar, isolation source, or virulence of the strain . Isolates obtained from alfalfa attained population levels similar to those observed for strains isolated from contaminated meat products or stools . Each of the strains could be detected in the waste irrigation water, with populations being strongly correlated with those detected on the germinating alfalfa seeds . The S . enterica strains were capable of utilizing the waste irrigation water as a sole carbon and nitrogen source . S . enterica strains thus appear to grow saprophytically on soluble organics released from seeds during early phases of germination . The ability to detect S . enterica in the waste irrigation water early in the germination process indicates that this method may be used as a simple way to monitor the contamination of sprouts during commercial operations. Appl Environ Microbiol, 2003 Jan, 69(1), 408 - 18 Potential rates of fermentation in digesta from the gastrointestinal tract of pigs: effect of feeding fermented liquid feed; Hojberg O et al.; Microbial catabolic capacity in digesta from the gastrointestinal tract of pigs fed either dry feed or fermented liquid feed (FLF) was determined with the PhenePlate multisubstrate system . The in vitro technique was modified to analyze the kinetics of substrate catabolism mediated by the standing stock of enzymes (potential rates of fermentation), allowing a quantitative evaluation of the dietary effect on the catabolic capacity of the microbiota . In total, the potential rates of fermentation were significantly reduced in digesta from the large intestine (cecum, P < 0.1; colon, P < 0.01; and rectum, P < 0.0001) of pigs fed FLF compared to pigs fed dry feed . No effect of diet was observed in the stomach (P = 0.71) or the distal part of the small intestine (P = 0.97) . The highest rates of fermentation and the most significant effect of diet were observed for readily fermentable carbohydrates like maltose, sucrose, and lactose . Feeding FLF to pigs also led to a reduction in the large intestine of the total counts of anaerobic bacteria in general and lactic acid bacteria specifically, as well as of microbial activity, as determined by the concentration of ATP and short-chain fatty acids . The low-molecular-weight carbohydrates were fermented mainly to lactic acid in the FLF before being fed to the animals . This may have limited microbial nutrient availability in the digesta reaching the large intestine of pigs fed FLF and may have caused the observed reduction in activity and density of the cecal and colonic microbial population . On the other hand, feeding FLF to pigs reduced the viable counts of coliform bacteria (indicator of Escherichia coli and Salmonella spp.) most profoundly in the stomach and the distal part of the small intestine, probably due to the bactericidal effect of lactic acid and low pH . The results presented clearly demonstrate that feeding FLF to pigs had a great impact on the indigenous microbiota, as reflected in bacterial numbers, short-chain fatty acid concentration, and substrate utilization . However, completely different mechanisms may be involved in the proximal and the distal parts of the gastrointestinal tract . The present study illustrates the utility of the PhenePlate system for quantifying the catabolic capacity of the indigenous gastrointestinal tract microbiota. Appl Environ Microbiol, 2003 Jan, 69(1), 290 - 6 Multicenter validation of the analytical accuracy of Salmonella PCR: towards an international standard; Malorny B et al.; As part of a major international project for the validation and standardization of PCR for detection of five major food-borne pathogens, four primer sets specific for Salmonella species were evaluated in-house for their analytical accuracy (selectivity and detection limit) in identifying 43 Salmonella spp . and 47 non-Salmonella strains . The most selective primer set was found to be 139-141 (K . Rahn, S . A . De Grandis, R . C . Clarke, S . A . McEwen, J . E . Galan, C . Ginocchio, R . Curtiss III, and C . L . Gyles, Mol . Cell . Probes 6:271-279, 1992), which targets the invA gene . An extended determination of selectivity by using 364 strains showed that the inclusivity was 99.6% and the exclusivity was 100% for the invA primer set . To indicate possible PCR inhibitors derived from the sample DNA, an internal amplification control (IAC), which was coamplified with the invA target gene, was constructed . In the presence of 300 DNA copies of the IAC, the detection probability for primer set 139-141 was found to be 100% when a cell suspension containing 10(4) CFU/ml was used as the template in the PCR (50 CFU per reaction) . The primer set was further validated in an international collaborative study that included 16 participating laboratories . Analysis with 28 coded ("blind") DNA samples revealed an analytical accuracy of 98% . Thus, a simple PCR assay that is specific for Salmonella spp . and amplifies a chromosomal DNA fragment detected by gel electrophoresis was established through extensive validation and is proposed as an international standard . This study addresses the increasing demand of quality assurance laboratories for standard diagnostic methods and presents findings that can facilitate the international comparison and exchange of epidemiological data. Appl Environ Microbiol, 2003 Jan, 69(1), 122 - 9 High pH during trisodium phosphate treatment causes membrane damage and destruction of Salmonella enterica serovar enteritidis; Sampathkumar B et al.; Trisodium phosphate (TSP) is now widely used during the processing of poultry and red meats, but the mechanism whereby it inactivates gram-negative bacteria such Salmonella spp . remains unclear . Thus, Salmonella enterica serovar Enteritidis (ATCC 4931) cells were treated with different concentrations of TSP (1.5, 2.0, and 2.5% {wt/vol}) and compared with (i) cells treated with the same pH as the TSP treatments (pH 10.0, 10.5, and 11.0, respectively) and (ii) cells treated with different concentrations of TSP (1.5, 2.0, and 2.5% {wt/vol}) adjusted to a pH of 7.0 +/- 0.2 (mean +/- standard deviation) . Cell viability, loss of membrane integrity, cellular leakage, release of lipopolysaccharides, and cell morphology were accordingly examined and quantified under the above treatment conditions . Exposure of serovar Enteritidis cells to TSP or equivalent alkaline pH resulted in the loss of cell viability and membrane integrity in a TSP concentration- or alkaline pH-dependent manner . In contrast, cells treated with different concentrations of TSP whose pH was adjusted to 7.0 did not show any loss of cell viability or membrane integrity . A 30-min pretreatment with 1.0 mM EDTA significantly enhanced the loss of membrane integrity only when followed by TSP or alkaline pH treatments . Measuring the absorbance at 260 nm, agarose gel electrophoresis, Bradford assay, and Tricine-sodium dodecyl sulfate gel electrophoresis of filtrates of treated cell suspensions revealed considerable release of DNA, proteins, and lipopolysaccharides compared to controls and pH 7.0 TSP treatments . Electron microscopic examination of TSP- or alkaline pH-treated cells showed disfigured cell surface topology and wrinkled appearance and showed evidence of a TSP concentration- and pH-dependent disruption of the cytoplasmic and outer membranes . These results demonstrate that TSP treatment permeabilizes and disrupts the cytoplasmic and outer membranes of serovar Enteritidis cells because of the alkaline pH, which in turn leads to release of intracellular contents and eventual cell death. Acta Microbiol Immunol Hung, 2002, 49(4), 433 - 44 Serotypes, virulence factors, antibiotic sensitivity, beta-lactamase activity and plasmid analysis of Salmonella from children with diarrhea in Tripoli (Libya); el-Ghodban A et al.; A total of 21 Salmonella strains isolated in Libya (16 from children with diarrhea and 5 from healthy controls) were serotyped and studied for their cell invasive ability, production of cytotoxin, antibiotic susceptibility, beta-lactamase activity and plasmid profiles . Eight different serotypes of Salmonella were identified: 6 S . saintpaul, 4 S . wien (1 from control), 2 S . newport, 2 S . muenchen (1 from control), 2 S . typhimurium (1 from control), 2 S . hadar (1 from control), 2 S . reading (1 from control), 1 S . kottbus . Twenty (95%) were positive in the invasiveness assay using HeLa cells, and all (100%) were negative for cytotoxin production in HT29 cells . More than 40% were resistant to ampicillin, cefalexin, cefamandole, cefoperazone, chloramphenicol, gentamicin, mezlocillin and trimethoprimsulphamethoxazole and 100% were susceptible to the new quinolones . Most (67%) of the strains harbored plasmids and 43% produced beta-lactamase . A strong association was observed between the presence of more than one plasmid, beta-lactamase activity, and multiple-resistance to antimicrobial agents and serotypes S . saintpaul and S . wien . Curing experiments with acridine orange showed that 2 plasmids (33 and 1.4 megadaltons) might be responsible for the resistance to chloramphenicol and gentamicin . The present study demonstrated that multiple-resistant salmonellae are widespread in Libya and the resistance is mainly plasmid mediated. J Bacteriol, 2003 Jan, 185(2), 664 - 8 Flagella and Motility in Actinobacillus pleuropneumoniae; Negrete-Abascal E et al.; Actinobacillus pleuropneumoniae has been considered nonmotile and nonflagellate . In this work, it is demonstrated that A . pleuropneumoniae produces flagella composed of a 65-kDa protein with an N-terminal amino acid sequence that shows 100% identity with those of Escherichia coli, Salmonella, and Shigella flagellins . The DNA sequence obtained through PCR of the fliC gene in A . pleuropneumoniae showed considerable identity (93%) in its 5' and 3' ends with the DNA sequences of corresponding genes in E . coli, Salmonella enterica, and Shigella spp . The motility of A . pleuropneumoniae was observed in tryptic soy or brain heart infusion soft agar media, and it is influenced by temperature . Flagella and motility may be involved in the survival and pathogenesis of A . pleuropneumoniae in pigs. J Bacteriol, 2003 Jan, 185(2), 553 - 63 Genomic comparison of Salmonella enterica serovars and Salmonella bongori by use of an S . enterica serovar typhimurium DNA microarray; Chan K et al.; The genus Salmonella consists of over 2,200 serovars that differ in their host range and ability to cause disease despite their close genetic relatedness . The genetic factors that influence each serovar's level of host adaptation, how they evolved or were acquired, their influence on the evolution of each serovar, and the phylogenic relationships between the serovars are of great interest as they provide insight into the mechanisms behind these differences in host range and disease progression . We have used an Salmonella enterica serovar Typhimurium spotted DNA microarray to perform genomic hybridizations of various serovars and strains of both S . enterica (subspecies I and IIIa) and Salmonella bongori to gain insight into the genetic organization of the serovars . Our results are generally consistent with previously published DNA association and multilocus enzyme electrophoresis data . Our findings also reveal novel information . We observe a more distant relationship of serovar Arizona (subspecies IIIa) from the subspecies I serovars than previously measured . We also observe variability in the Arizona SPI-2 pathogenicity island, indicating that it has evolved in a manner distinct from the other serovars . In addition, we identify shared genetic features of S . enterica serovars Typhi, Paratyphi A, and Sendai that parallel their unique ability to cause enteric fever in humans . Therefore, whereas the taxonomic organization of Salmonella into serogroups provides a good first approximation of genetic relatedness, we show that it does not account for genomic changes that contribute to a serovar's degree of host adaptation. J Bacteriol, 2003 Jan, 185(2), 525 - 33 Transcription of the Salmonella invasion gene activator, hilA, requires HilD activation in the absence of negative regulators; Boddicker JD et al.; Salmonella enterica serovar Typhimurium causes human gastroenteritis and a systemic typhoid-like infection in mice . Infection is initiated by entry of the bacteria into intestinal epithelial cells and is mediated by a type III secretion system that is encoded by genes in Salmonella pathogenicity island 1 . The expression of invasion genes is tightly regulated by environmental conditions such as oxygen and osmolarity, as well as by many bacterial factors . The hilA gene encodes an OmpR/ToxR family transcriptional regulator that activates the expression of invasion genes in response to both environmental and genetic regulatory factors . HilD is an AraC/XylS regulator that has been postulated to act as a derepressor of hilA expression that promotes transcription by interfering with repressor binding at the hilA promoter . Our research group has identified four genes (hilE, hha, pag, and ams) that negatively affect hilA transcription . Since the postulated function of HilD at the hilA promoter is to counteract the effects of repressors, we examined this model by measuring hilA::Tn5lacZY expression in strains containing negative regulator mutations in the presence or absence of functional HilD . Single negative regulator mutations caused significant derepression of hilA expression, and two or more negative regulator mutations led to very high level expression of hilA . However, in all strains tested, the absence of hilD resulted in low-level expression of hilA, suggesting that HilD is required for activation of hilA expression, whether or not negative regulators are present . We also observed that deletion of the HilD binding sites in the chromosomal hilA promoter severely decreased hilA expression . In addition, we found that a single point mutation at leucine 289 in the C-terminal domain of the alpha subunit of RNA polymerase leads to very low levels of hilA::Tn5lacZY expression, suggesting that HilD activates transcription of hilA by contacting and recruiting RNA polymerase to the hilA promoter. J Bacteriol, 2003 Jan, 185(2), 432 - 43 Salmonella enterica serovar typhimurium rdoA is growth phase regulated and involved in relaying Cpx-induced signals; Suntharalingam P et al.; The disulfide oxidoreductase, DsbA, mediates disulfide bond formation in proteins as they enter or pass through the periplasm of gram-negative bacteria . Although DsbA function has been well characterized, less is known about the factors that control its expression . Previous studies with Escherichia coli demonstrated that dsbA is part of a two-gene operon that includes an uncharacterized, upstream gene, yihE, that is positively regulated via the Cpx stress response pathway . To clarify the role of the yihE homologue on dsbA expression in Salmonella enterica serovar Typhimurium, the effect of this gene (termed rdoA) on the regulation of dsbA expression was investigated . Transcriptional assays assessing rdoA promoter activity showed growth phase-dependent expression with maximal activity in stationary phase . Significant quantities of rdoA and dsbA transcripts exist in serovar Typhimurium, but only extremely low levels of rdoA-dsbA cotranscript were detected . Activation of the Cpx system in serovar Typhimurium increased synthesis of both rdoA- and dsbA-specific transcripts but did not significantly alter the levels of detectable cotranscript . These results indicate that Cpx-mediated induction of dsbA transcription in serovar Typhimurium does not occur through an rdoA-dsbA cotranscript . A deletion of the rdoA coding region was constructed to definitively test the relevance of the rdoA-dsbA cotranscript to dsbA expression . The absence of RdoA affects DsbA expression levels when the Cpx system is activated, and providing rdoA in trans complements this phenotype, supporting the hypothesis that a bicistronic mechanism is not involved in serovar Typhimurium dsbA regulation . The rdoA null strain was also shown to be altered in flagellar phase variation . First it was found that induction of the Cpx stress response pathway switched flagellar synthesis to primarily phase 2 flagellin, and this effect was then found to be abrogated in the rdoA null strain, suggesting the involvement of RdoA in mediating Cpx-related signaling. Indian J Med Sci, 2002 Jan, 56(1), 1 - 8 Evaluation of minimum inhibitory concentration of quinolones and third generation cephalosporins to Salmonella typhi isolates; Kumar R et al.; A total of 323 Salmonella typhi isolates (261 isolates obtained during 1995-99 from Ludhiana and 62 randomly selected isolates obtained between 1980-99 from Chandigarh) were analyzed for drug resistant pattern . S . typhi isolates prior to 1986 were sensitive to all the antimicrobials tested by disc diffusion method . Most common multidrug resistance pattern noticed was ACCo T i.e . resistance to ampicillin, Chloramphenicol, cotrimoxazole and tetracycline . This study has revealed that withdrawal of chloramphenicol due to high level of resistance during 1990-94, has led to re-emergence of 43-93 percent chloramphenicol sensitive mutants during 1995-99 . Two S . typhi isolates in 1995 and one in 1999 from Ludhiana depicted resistance to ciprofloxacin . Susceptibility of S . typhi isolates to third generation cephalosporins ranged between 87 to 100 per cent . There was a gradual increase with time period in mean minimum inhibitory concentration (MIC) of ciprofloxacin as it increased from 0.066 ug/ml for 1980-83 S . typhi isolates to 0.13 ug/ml for the 1996-99 isolates . Similarly, cefotaxime mean MIC for 1980-83 isolate was 0.172 ug/ml which further increased up to 0.32 ug/ml for S . typhi isolates encountered between 1996-99 . In contrast, mean MIC value of 0.62 ug/ml of Ceftriaxone remained unchanged irrespective of the year of isolation of S . typhi isolates. Vet Immunol Immunopathol, 2003 Jan 10, 91(1), 39 - 44 Effects of live attenuated and killed Salmonella vaccine on T-lymphocyte mediated immunity in laying hens; Babu U et al.; The impact of live and killed Salmonella vaccines on cell-mediated immunity (CMI) was investigated in 18- and 32-week-old White Leghorn chickens, by assessing splenic lymphocyte proliferation, expression of IL-2 mRNA in concanavalin A (Con A) stimulated cells and flow cytometric analysis of cell subpopulations . Con A and Salmonella enteritidis (SE) flagella induced proliferation of splenocytes were enhanced in the 18- and 32-week-old chickens treated with live vaccine, compared to the corresponding control chickens . Among the killed vaccine treated birds, Con A-mediated response was higher in the 18-week-old chickens compared to the corresponding control birds . Increased proliferation was accompanied by increased CD4 and reduced CD8 and gammadelta T-lymphocytes in the 18-week-old live vaccine treated chickens . Relative expression of IL-2 mRNA in Con A-stimulated splenocytes from 18-week-old birds was not affected by vaccine treatment . Overall, live vaccine was more effective in increasing the lymphocyte proliferation to Con A as well as SE antigen . This enhanced CMI may prove beneficial in protecting chickens against SE infection. J Mol Biol, 2003 Jan 24, 325(4), 795 - 807 Mg2+ sensing by the Mg2+ sensor PhoQ of Salmonella enterica; Chamnongpol S et al.; The PhoP/PhoQ two-component regulatory system governs the adaptation to low Mg(2+) environments and virulence in several Gram-negative species . During growth in low Mg(2+), the sensor PhoQ modifies the activity of the response regulator PhoP promoting gene transcription, whereas growth in high Mg(2+) represses transcription of PhoP-activated genes . The PhoQ protein harbors a periplasmic domain of 146 amino acid residues that binds Mg(2+) in vitro and is required for Mg(2+)-mediated repression in vivo . Here, we identify periplasmic mutants of the Salmonella PhoQ protein that allow transcription of PhoP-activated genes even under high Mg(2+) concentrations . When expressed in a strain harboring a PhoP variant that is phosphorylated from acetyl phosphate, some of the mutants failed to repress PhoP-promoted transcription in high Mg(2+), whereas others displayed a wild-type ability to do so . Mutant PhoQ proteins that allowed expression of PhoP-activated genes in high Mg(2+) displayed a pattern of iron-mediated cleavage in vitro that was different from that displayed by wild-type PhoQ, indicative of altered Mg(2+) binding . A PhoQ protein with the conserved histidine residue (H277) substituted by alanine could not promote transcription of PhoP-activated genes in low Mg(2+) but could turn off expression in response to high Mg(2+) . Our studies demonstrate that residues G93, W97, H120 and T156 are required for a wild-type response to Mg(2+), and suggest that Mg(2+) binding to the periplasmic domain regulates several activities in the PhoQ protein. Zh Mikrobiol Epidemiol Immunobiol, 2002 Nov-Dec, (6), 69 - 72 {Clinical course and treatment regimes of Salmonella infection in children}; Mirismailov MM et al.; Salmonella infection in children caused by polyresistant S . typhimurium strains is characterized by a highly severe course with possible complications . In the treatment of young children with salmonellosis the use of the locally manufactured biologically active additive Bektit-M, possessing adsorption capacity, is recommended . The use of this preparation in complex treatment has made it possible to establish its clinical effect and positive influence on the biochemical characteristics of the blood. Am J Epidemiol, 2003 Jan 1, 157(1), 48 - 57 Epidemiology of salmonellosis in California, 1990-1999: morbidity, mortality, and hospitalization costs; Trevejo RT et al.; Salmonella is a common cause of bacterial foodborne illness in the United States . The epidemiology and costs of nontyphoidal salmonellosis in California from 1990 through 1999 are described using surveillance, hospitalization, and death data . Trends in Salmonella rates and factors associated with prolonged hospitalization were evaluated using Poisson and linear regression models, respectively . There were 56,660 reported cases, 11,102 hospitalizations, and 74 deaths attributed to Samonella . Reported case and hospital discharge rates have decreased since 1996 . Among reported cases, infants had the highest rate (121 cases per 10(5) person-years), followed by children 1-4 years of age (40 cases per 10(5) person-years) . The highest hospitalization rates were among the elderly and young children . Most deaths occurred among persons aged 65 or more years (59%) . Among hospitalizations, gastroenteritis (61%) and septicemia (23%) were the most common Salmonella diagnoses . Salmonella pneumonia patients were the oldest (median age, 55 years) and Salmonella meningitis patients the youngest (median age, 0.3 years) . These two diagnoses were the costliest, approaching 30,000 dollars (median) per hospitalization . Having an acquired immunodeficiency syndrome diagnosis or multiple Salmonella diagnoses was independently associated with prolonged hospitalization . The estimated 10-year hospitalization costs for Salmonella were $200 million . Salmonellosis is a costly disease that disproportionately affects the young and elderly. J Immunol Methods, 2003 Jan 15, 272(1-2), 199 - 210 Expression of recombinant proteins in a lipid A mutant of Escherichia coli BL21 with a strongly reduced capacity to induce dendritic cell activation and maturation; Cognet I et al.; Mutations in the Escherichia coli (E . coli) and Salmonella lpxM gene have been shown to result in strains which grow normally and which produce a non-myristoylated lipopolysaccharide (nmLPS) with strongly reduced endotoxicity . Using homologous recombination, we inactivated the lpxM gene in BL21 (DE3), a strain widely used for the production of recombinant proteins . This led to a derivative unaffected in its capacity to support the production of recombinant proteins . This new strain expresses non-myristoylated LPS that induces markedly less activation and maturation of monocyte-derived dendritic cells (DC), as assessed by nuclear translocation of nuclear factor kappa B (NF-kappaB), production of TNF-alpha and IL-8 or expression of CD86 . Activation of the main signal transducing receptor for extracellular LPS, Toll like receptor (TLR) 4 in conjunction with the soluble accessory protein MD-2 was also markedly decreased.The modified BL21 strain represents a new application of lpxM inactivation for the expression of proteins to be tested on dendritic cells or other LPS sensitive cells/receptor complexes . It is likely to be useful for the identification of new proteins activating the innate immune response and to reducing the risk linked with low level of endotoxin contamination in therapeutic recombinant proteins. Int J Food Microbiol, 2003 Jan 26, 82(1), 13 - 24 Inactivation of Salmonella enteritidis during boiling of eggs; Grijspeerdt K et al.; A series of inactivation curves for Salmonella enteritidis were determined for boiling eggs using different conditions of time and temperature . No significant influence of egg weight could be found on the temperature evolution in the yolk . The inactivation curves consistently showed an initial slow decline in bacterial number at lower temperatures, after which a very rapid inactivation took place . It was not possible to reproduce this behavior using a traditional inactivation model . A pragmatic model existing in two parts was therefore constructed . When the temperature is below a certain threshold, the inactivation follows a second order temperature dependence . Above the temperature threshold, standard Bigelow inactivation kinetics are assumed.This model could describe the data reasonably well, provided that the decimal reduction time in the Bigelow model was assumed to be different for a fast or slow heating process, respectively . The results suggest that the bacteria are more resistant towards a slower heating process, which is confirmed by analyzing the raw data . A fail-safe model can be obtained by using the parameters associated with the slow heating process . The statistical properties of the calibrated model are satisfactory, and a cross-validation shows that it can be used for egg boiling conditions outside its calibration range. Mutat Res, 2003 Jan 10, 534(1-2), 101 - 12 Mutagenic characteristics of river waters flowing through large metropolitan areas in North America; Ohe T et al.; The hanging technique using blue rayon, which specifically adsorbs mutagens with multicyclic planar structures, has the advantages over most conventional methods of not having to bring large volumes of water back to the laboratory for extraction of organic materials . Therefore, for the same effort the hanging blue rayon technique allows for the analysis of more samples from remote sites, although it has a disadvantage of not allowing quantitative analysis . In this study, the blue rayon hanging technique was used to collect organic mutagens in river waters that flow through metropolitan areas in northeastern North America . Monitoring was performed at a total of 21 sites: the Providence River system (4 sites), the Charles River (2 sites), the Potomac River (6 sites), the St . Lawrence River (5 sites), the Hudson River (3 sites), and the East River (1 site) . Mutagenicity was evaluated using the Salmonella assay with strains TA98, TA100, YG1024, YG1041, and YG1042 with and without metabolic activation . The results demonstrated that strains YG1041 and YG1024 were much more sensitive than TA98 with S9 mix . Fifteen samples out of 21 were positive in YG1041 with S9 mix . Six samples gave 5000-18,400 revertants/g blue rayon equivalent . YG1042 was also much more sensitive than TA100 . Eight samples were positive in YG1042 with S9 mix . The highest activity was 10,200 revertants/g blue rayon equivalent . The overall results showed that rivers flowing through major cities in North America contain frameshift-type, aromatic amine-like mutagenic activity . However, the levels of mutagenic activity in these rivers were much lower than expected based on prior analyses and calculated population-to-discharge ratios . Further research, such as detailed chemical analyses and/or simultaneous comparisons of several different adsorbents (e.g . XAD and blue rayon), will be needed to clarify the observed differences between North American blue rayon values and published values for European and Asian river systems. Food Chem Toxicol, 2003 Mar, 41(3), 347 - 50 Lack of in vivo clastogenic activity of grape seed and grape skin extracts in a mouse micronucleus assay; Erexson GL; Meganatural brand grape seed extract (GSE) and grape skin extract (GSKE), containing proanthocyanidin polyphenolic compounds, are intended for use in food as functional ingredients exhibiting antioxidant activity . Proanthocyanidins, as well as the minor constituent phenolic compounds in GSE and GSKE, are present naturally in many foods such as fruits, vegetables, chocolate, tea, etc., and on average people consume 460-1000 mg/day of these combined substances . While some polyphenolic compounds, tested individually, have demonstrated antitumorigenic or antipromotional activity, at least one minor component of GSE and GSKE, quercitin, has exhibited positive activity in Salmonella and other in vitro mutagenicity assays . As part of a program to investigate the safety of GSE and GSKE, these products were tested for in vivo clastogenic activity and/or disruption of the mitotic apparatus by detecting micronuclei in polychromatic erythrocyte (PCE) cells in Crl:CD-1(ICR) BR mouse bone marrow . The appropriate test article was dissolved in 0.5% carboxymethylcellulose and dosed by oral gavage to five males/test article/dose level/harvest time point . Animals were dosed at 500, 1000 and 2000 mg/kg . Five animals dosed with either test article at 500, 1000 and 2000 mg/kg dose levels and five animals dosed with the cyclophosphamide (80 mg/kg) positive control were euthanized approximately 24 h after dosing for extraction of bone marrow . Five animals dosed with either test article at the 2000 mg/kg dose level and five animals dosed with the vehicle control article were euthanized approximately 24 and 48 h after dosing for extraction of bone marrow . At least 2000 PCEs per animal were analyzed for frequency of micronuclei . Cytotoxicity was assessed by scoring the number of PCEs and normochromatic erythrocytes (NCEs) in at least the first 500 erythrocytes for each animal . For both GSE and GSKE, no statistically significant increase in micronucleated PCEs was observed at any dose level or harvest time point . GSE produced indication of cytotoxicity (decreased PCE:NCE ratio) at the 2000 mg/kg dose level for the 48-h harvest time point, confirming that the test article reached the target bone marrow in significant amount . Meganatural GSE and Meganatural GSKE were evaluated as negative in the mouse bone marrow micronucleus assay under the conditions of this assay. J Agric Food Chem, 2003 Jan 1, 51(1), 249 - 53 Eggshell matrix proteins as defense mechanism of avian eggs; Mine Y et al.; This study focused on the role of eggshell matrix proteins as a function of potential natural antimicrobial defenses of avian eggs . The electrophoretic profile of SDS-PAGE showed that the soluble eggshell matrix proteins had three major bands of 15 000, 36 000, and 66 000 and several minor bands comprising 17 000, 25 000, 30 000, and 75 000, while insoluble matrix proteins were consisting of various bands comprising at least 16 distinct migration bands between 10 000 and 200 000 . Three bacteria species, Pseudomonas aureginosa, Bacillus cereus, and Staphylococcus aureus, were found to be inhibited in the presence of soluble eggshell matrix proteins (100 microg/mL) . On the other hand, Escherichia coli and Salmonella enteritidis were weakly inhibited at only an early stage of incubation time (up to 4 h) . Scanning electron microscopy revealed that eggshell matrix proteins might interact and disrupt the membrane integrity of bacteria . The present study clearly indicated that avian eggshell matrix proteins possess a potential of novel antimicrobial defensin mechanism. J Vet Med Sci, 2002 Nov, 64(11), 1079 - 80 Incidence of Salmonella infection in healthy dogs in Gifu Prefecture, Japan; Fukata T et al.; A total of 1,013 feces samples and 8 mesenteric lymphonodus samples obtained from apparently healthy dogs were examined for the incidence of salmonella infection . One strain of S . typhimurium (ST) was isolated from feces of one dog, and S . enteritidis (SE) was isolated from the mesenteric lymphonodus of one dog . Sera obtained from 330 apparently healthy dogs were examined for Salmonella antibodies using an ELISA with heated whole cells of SE and ST . Fifty-one of the 330 serum samples were considered to be positive for salmonella antibodies, including 12 which were SE-positive and 39 which were ST-positive . These results indicate that dogs cause possible environmental problems as Salmonella carriers. J Immunol, 2003 Jan 1, 170(1), 597 - 603 Impaired accumulation and function of memory CD4 T cells in human IL-12 receptor beta 1 deficiency; Cleary AM et al.; Defects in IL-12 production or IL-12 responsiveness result in a vulnerability to infection with non-viral intracellular organisms, but the immunological mechanisms responsible for this susceptibility remain poorly understood . We present an immunological analysis of a patient with disseminated Salmonella enteritidis and a homozygous splice acceptor mutation in the IL-12Rbeta1-chain gene . This mutation resulted in the absence of IL-12Rbeta1 protein on PBMC and an inability of T cells to specifically bind IL-12 or produce IFN-gamma in response to either IL-12 or IL-23 . The accumulation of memory (CD45R0(high)) CD4 T cells that were CCR7(high) (putative central memory cells) was normal or increased for age . Central memory CD4 T cells of the patient and age-matched controls were similar in having a low to undetectable capacity to produce IFN-gamma after polyclonal stimulation . In contrast, the patient had a substantial decrease in the number of CCR7(neg/dull) CD45R0(high) memory CD4 T cells (putative effector memory cells), and these differed from control cells in having a minimal ability to produce IFN-gamma after polyclonal stimulation . Importantly, tetanus toxoid-specific IFN-gamma production by PBMC from the patient was also significantly reduced compared with that in age-matched controls, indicating that signaling via the IL-12Rbeta1-chain is generally necessary for the in vivo accumulation of human memory CD4 T cells with Th1 function . These results are also consistent with a model in which the IL-12Rbeta1 subunit is necessary for the conversion of central memory CD4 T cells into effector memory cells. Infect Immun, 2003 Jan, 71(1), 504 - 9 Cholera toxin induces migration of dendritic cells from the subepithelial dome region to T- and B-cell areas of Peyer's patches; Shreedhar VK et al.; Intestinal M cells deliver macromolecules, particles, and pathogens into the subepithelial dome (SED) region of Peyer's patch mucosa, an area rich in dendritic cells (DCs) . We tested whether uptake of the mucosal adjuvant cholera toxin (CT) or live Salmonella bacteria can induce DC migration within Peyer's patches . Virus-sized, fluorescent polystyrene microparticles were efficiently transported by M cells and ingested by CD11c(+), CD11b(-), and CD8a(-) DCs in the SED region . DCs loaded with microparticles remained in the SED for up to 14 days . CT (but not the CT B subunit) and live attenuated Salmonella enterica serovar Typhimurium bacteria induced migration of the microparticle-loaded DCs from the SED region into underlying B-cell follicles and adjacent parafollicular T-cell zones . Our data provide the first demonstration that DCs move in response to enterotoxin adjuvants and live bacteria that enter the mucosa via M cells. Infect Immun, 2003 Jan, 71(1), 418 - 27 The Salmonella enterica serovar typhimurium translocated effectors SseJ and SifB are targeted to the Salmonella-containing vacuole; Freeman JA et al.; The Salmonella enterica serovar Typhimurium type III secretion system (TTSS) encoded in Salmonella pathogenicity island 2 (SPI-2) promotes replication within host cells and systemic infection of mice . The SPI-2 TTSS is expressed following Salmonella internalization into host cells and translocates effectors across the membrane of the Salmonella-containing vacuole (SCV) . Two effectors with similar amino-terminal domains, SseJ and SifB, localize to the SCV membrane in infected HEp-2 cells and subsequently traffic away from the SCV along Salmonella-induced-filaments (Sifs) . Following infection of RAW cells, SseJ and SifB localize to the SCV as well as LAMP-1-positive, vesicular-appearing structures distant from the SCV . Trafficking of SseJ and SifB away from the SCV requires the SPI-2 effector SifA . Deletion of sseJ, but not sifB, leads to attenuation of Salmonella replication in mice following intraperitoneal inoculation . The contribution of SseJ to in vivo replication is identical in wild-type and sifA deletion backgrounds, suggesting that SseJ trafficking away from the SCV along Sifs is unnecessary for its virulence function. Infect Immun, 2003 Jan, 71(1), 287 - 97 Impact of vector priming on the immunogenicity of recombinant Salmonella vaccines; Vindurampulle CJ et al.; There are conflicting reports concerning the impact of prior vector priming on the immunogenicity of recombinant-Salmonella-based vaccines . A comparison of experimental protocols identified two variables which might account for this inconsistency: the potential of the vector strain to colonize the murine gut-associated lymphoid tissue (GALT) and the nature of the foreign antigen subsequently delivered by the recombinant Salmonella construct . The former was investigated by constructing an aroA mutant of the Salmonella enterica serovar Stanley vector previously used in our laboratory . Although the introduction of an aroA mutation had surprisingly little effect on GALT colonization, it did reduce the strength of antilipopolysaccharide (anti-LPS) antibody responses and the impact of vector priming . Studies were also performed to ascertain the extent to which any observed hyporesponsiveness consequent upon vector priming might be determined by the characteristics of the foreign antigen . S . enterica serovar Stanley was used to deliver either of two Escherichia coli antigens, K88 pilus protein or the LT-B toxin subunit, to vector-primed mice . Both serum immunoglobulin G (IgG) and intestinal IgA responses to K88 were completely abolished, and those to LT-B were significantly reduced, as a consequence of vector priming . When similar experiments were performed with an aroA S . enterica serovar Dublin vector, responses to K88 were significantly reduced but those to LT-B were unaffected by vector priming . Paradoxically, a priming infection with this vector induced stronger anti-LPS antibody responses but was less likely to elicit a state of hyporesponsiveness to subsequently presented foreign antigen . The impact of vector priming thus depends on both the Salmonella strain used and the nature of the foreign antigen, but our present data strengthen concerns that preexisting antivector immunity represents a serious threat to the Salmonella-based vaccine strategy. Infect Immun, 2003 Jan, 71(1), 242 - 53 Identification of substrates and chaperone from the Yersinia enterocolitica 1B Ysa type III secretion system; Foultier B et al.; All pathogenic Yersinia enterocolitica strains carry the pYV plasmid encoding the Ysc-Yop type III secretion (TTS) system, which operates at 37 degrees C . In addition, biovar 1B Y . enterocolitica strains possess a second, chromosomally encoded, TTS system called Ysa, which operates, at least in vitro, under low-temperature and high-salt (LTHS) conditions . Six open reading frames, sycB, yspB, yspC, yspD, yspA, and acpY, neighbor the ysa genes encoding the Ysa TTS apparatus . Here we show that YspA, YspB, YspC, and YspD are secreted by the Ysa TTS system under LTHS conditions . SycB is a chaperone for YspB and YspC and stabilizes YspB . YspB, YspC, and SycB share some similarity with TTS substrates and the chaperone encoded by the Mxi-Spa locus of Shigella flexneri and SPI-1 of Salmonella enterica . In addition, Ysa also secretes the pYV-encoded YopE under LTHS conditions, indicating that YopE is a potential effector of both Y . enterocolitica TTS systems . YspC could also be secreted by S . flexneri, but no functional complementation of ipaC was observed, which indicates that despite their similarity the Ysa and the Mxi-Spa systems are not interchangeable . When expressed from the yopE promoter, YspB and YspC could also be secreted via the Ysc injectisome . However, they could not form detectable pores in eukaryotic target cells and could not substitute for YopB and YopD for translocation of Yop effectors. Infect Immun, 2003 Jan, 71(1), 117 - 25 Effective induction of acquired resistance to Listeria monocytogenes by immunizing mice with in vivo-infected dendritic cells; Sashinami H et al.; Splenic dendritic cells (DCs) obtained from mice at 48 h after Listeria monocytogenes infection exhibited up-regulation of CD80 and produced higher titers of gamma interferon (IFN-gamma) and interleukin-12 (IL-12) than did DCs obtained from uninfected mice . Mice immunized with DCs obtained from mice that had been infected with L . monocytogenes 48 h before acquired host resistance to lethal infection with L . monocytogenes at 4 and 8 weeks . Immunization with DCs from heat-killed L . monocytogenes failed to induce resistance . Acquired antilisterial resistance is specific, since the immunized mice could not be protected from Salmonella enterica serovar Typhimurium infection . Infected DCs stimulated proliferation of naive CD4(+) and CD8(+) cells in vitro, suggesting that in vivo-infected DCs activate CD8(+) T cells, which are critical in acquired antilisterial resistance, as well as CD4(+) T cells . When wild-type mice were immunized with DCs from IFN-gamma-deficient mice, they were protected against a lethal L . monocytogenes challenge . In contrast, when mice were immunized with DCs from anti-IL-12 p40 monoclonal antibody-injected mice, they failed to gain acquired antilisterial resistance . These results suggest that DC-derived IL-12, but not IFN-gamma, may play a critical role in induction of acquired antilisterial resistance . Our present results suggest that splenic DCs obtained from mice infected with L . monocytogenes in vivo may be an effective immunogen with which to induce antigen-specific immunity. Infect Immun, 2003 Jan, 71(1), 30 - 9 Oral immunization with ATP-dependent protease-deficient mutants protects mice against subsequent oral challenge with virulent Salmonella enterica serovar typhimurium; Matsui H et al.; We evaluated the efficacy of mutants with a deletion of the stress response protease gene as candidates for live oral vaccine strains against Salmonella infection through infection studies with mice by using a Salmonella enterica serovar Typhimurium mutant with a disruption of the ClpXP or Lon protease . In vitro, the ClpXP protease regulates flagellum synthesis and the ClpXP-deficient mutant strain exhibits hyperflagellated bacterial cells (T . Tomoyasu et al., J . Bacteriol . 184:645-653, 2002) . On the other hand, the Lon protease negatively regulates the efficacy of invading epithelial cells and the expression of invasion genes (A . Takaya et al., J . Bacteriol . 184:224-232, 2002) . When 5-week-old BALB/c mice were orally administered 5 x 10(8) CFU of the ClpXP- or Lon-deficient strain, bacteria were detected with 10(3) to 10(4) CFU in the spleen, mesenteric lymph nodes, Peyer's patches, and cecum 1 week after inoculation and the bacteria then decreased gradually in each tissue . Significant increases of lipopolysaccharide-specific immunoglobulin G (IgG) and secretory IgA were detected at week 4 and maintained until at least week 12 after inoculation in serum and bile, respectively . Immunization with the ClpXP- or Lon-deficient strain protected mice against oral challenge with the serovar Typhimurium virulent strain . Both the challenged virulent and immunized avirulent salmonellae were completely cleared from the spleen, mesenteric lymph nodes, Peyer's patches, and even cecum 5 days after the challenge . These data indicate that Salmonella with a disruption of the ATP-dependent protease ClpXP or Lon can be useful in developing a live vaccine strain. Microb Pathog, 2002 Dec, 33(6), 279 - 87 Cytopathic effects observed upon expression of a repressed collagenase gene present in Salmonella and related pathogens: mimicry of a cytotoxin from multiple antibiotic-resistant Salmonella enterica serotype Typhimurium phagetype DT104; Wu MT et al.; Recently, we reported that certain strains of Salmonella enterica serovar Typhimurium phagetype DT104 (DT104) secrete a putative cytotoxin . While searching for the gene that encodes this toxin, we noted a previously reported but uncharacterized DNA fragment (clg) in Salmonella that could be potentially relevant to cytotoxin-like activity . Therefore, we cloned and expressed clg in cytotoxin-negative Escherichia coli and Salmonella and subsequently assessed the bioactivity of Clg in vitro and in vivo . Lysates containing Clg from both expression hosts exerted cytopathic effects on murine enterocytes while semi-purified Clg was determined to be cytopathic to HEp-2 cells . Sequence and RT-PCR analyses of the clg gene indicated that a homologue of clg exists in different Gram-negative bacteria although the gene is not expressed in vitro . Although Clg-mediated lesions are similar to those mediated by the DT104 cytotoxin, further investigations are necessary to examine the relationship between these two proteins in DT104 . Nonetheless, we report here a defined Salmonella protein that is capable of inflicting damage on tissue culture cells and murine enterocytes. Avian Dis, 2002 Oct-Dec, 46(4), 1015 - 20 Mucosal humoral immunity to experimental Salmonella enteritidis infection in the chicken crop; Seo KH et al.; In this report, we show that chickens, infected with Salmonella enteritidis (SE) by oral gavage, produce secretory immunoglobulin As (sIgAs) that specifically bind to numerous SE antigens . Chickens infected with SE showed strong sIgA response against flagella in both bile and crop . The optical density values of enzyme-linked immunosorbent assay (ELISA) tests in positive bile and crop were 1.17 and 0.38, respectively, and were significantly different from those of negative samples . Western immunoblotting revealed that approximately 13.5-, approximately 56-, approximately 62-, approximately 80-, and approximately 143-kD polypeptides were immunodominant proteins in bile, whereas approximately 56-, approximately 62-, and approximately 80-kD polypeptides were found to be strong antigens in crop . These results indicate that the crop may function as another site for mucosal immunity, and the SE flagella-based ELISA of crop samples can provide a useful screening test of SE exposure in chickens. Avian Dis, 2002 Oct-Dec, 46(4), 997 - 1000 Investigation of resistance of bacteria from commercial poultry sources to commercial disinfectants; Sander JE et al.; Concern by consumers about food safety has resulted in increased pressure on poultry companies to develop effective sanitation programs . Salmonella isolates in hatcheries are often the same species isolated from processing plants . Resistance develops in bacteria after prolonged exposure to disinfectants . The methods available in published literature to detect the efficacy of disinfectants are labor intensive and do not consider how bacteria behave when adhered to a solid surface . We used a recently developed technique, which utilizes the actual surfaces on which the disinfectant is to be applied, to evaluate the degree of resistance to four commercially available disinfectants of 17 bacterial isolates from poultry hatcheries . We found that bacterial isolates within the same genus and species have different sensitivities to the same disinfectant . In addition, disinfectants with similar but not identical chemical formulations have different efficacies against the same bacteria. Avian Dis, 2002 Oct-Dec, 46(4), 869 - 76 Associations of interferon-gamma genotype and protein level with antibody response kinetics in chickens; Zhou H et al.; Although previous studies have demonstrated an association between interferon-gamma (IFN-gamma) promoter genotype and antibody response kinetics in chickens, the protein levels that may mediate such a gene-trait association have not been determined . The objective of this study, therefore, was to determine the correlation of circulating IFN-gamma levels with both the IFN-gammaIFN-gamma promoter polymorphisms and antibody response in order to evaluate the potential role of IFN-gamma protein in mediating genetic control of antibody response in chickens . Antibody response after Salmonella enteritidis (SE) vaccination at day 10, antibody response to sheep red blood cells (SRBCs) and killed Brucella abortus after immunizations at 19 wk and 22 wk, and serum IFN-gamma protein level were measured in an F2 population derived from inbred lines . A single nucleotide polymorphism in the IFN-gamma promoter region was associated with IFN-gamma protein expression as measured by an enzyme-linked immunosorbent assay after both primary and secondary immunizations . Higher IFN-gamma protein level was correlated with higher antibody level to SE and with increased maximum level and decreased time to reach the maximum secondary antibody response to SRBCs . These results suggest that one of the mechanisms by which promoter polymorphism of IFN-gamma affects antibody production in chickens may involve the circulating level of IFN-gamma protein. J Food Prot, 2002 Dec, 65(12), 1976 - 80 Death of Salmonella, Escherichia coli O157:H7, and Listeria monocytogenes in garlic butter as affected by storage temperature; Adler BB et al.; Garlic is known to have antimicrobial activity against several spoilage and pathogenic bacteria . However, the fate of Salmonella, Escherichia coli O157:H7, and Listeria monocytogenes in garlic butter has not been reported . This study was undertaken to determine the viability of these organisms in garlic butter as affected by the type of raw minced garlic added to the butter, storage temperature, and storage time . Unsalted butter at 40 degrees C was combined with raw minced jumbo, elephant, or small-cloved garlic at a 4:1 butter/garlic ratio (wt/wt), inoculated with mixed-strain suspensions of Salmonella, E . coli O157:H7, or L monocytogenes, and stored at 4.4, 21, or 37 degrees C for up to 48 h . All pathogens retained their viability at 4.4 degrees C, regardless of the presence of garlic . The addition of garlic to butter enhanced the rates of inactivation of all three pathogens at 21 and 37 degrees C . The most rapid decline in pathogen populations was observed at 37 degrees C . The inactivation of L . monocytogenes occurred more slowly than did that of Salmonella or E . coli O157:H7 . The inactivation of Salmonella and L . monocytogenes was more rapid in jumbo garlic butter than in elephant or small-cloved garlic butter . It is concluded that Salmonella, E . coli O157:H7, and L . monocytogenes did not grow in unsalted butter, with or without garlic added (20%, wt/wt), when inoculated products were stored at 4.4, 21, and 37 degrees C for up to 48 h. J Food Prot, 2002 Dec, 65(12), 1909 - 15 Microbial antagonists of foodborne pathogens on fresh, minimally processed vegetables; Schuenzel KM et al.; On many types of raw or minimally processed foods, the bacterial microbiota is often composed of mixed species . The activities of one bacterial species may influence the growth and activities of others that are present . The objective of this project was to evaluate the microbial composition of fresh and minimally processed vegetables to determine if naturally occurring bacteria on produce are competitive with or antagonistic to potentially encountered pathogens . Naturally occurring bacteria were obtained from ready-to-eat salad vegetables on four occasions to allow for seasonal variation . Minimally processed vegetables were sampled at various stages in their processing from raw vegetables to packaged products . Some portions were analyzed microbiologically within 24 h, while other portions were stored refrigerated and analyzed after 72 h . Microbiological analysis was conducted for bacterial enumeration and to obtain isolates . An agar spot method was used to screen isolates for antimicrobial activity against Staphylococcus aureus ATCC 27664, Escherichia coli O157:H7 E009, Listeria monocytogenes LCDC 81-861, and Salmonella Montevideo . Of the 1,180 isolates screened for inhibitory activity, 37 (3.22%) were found to have various degrees of inhibitory activity against at least one test pathogen . Many isolates showed inhibitory activity against all four pathogens . The isolates with the most extensive inhibition were removed from finished lettuce piece shreds . Of the 37 inhibitory isolates, 34 (91.9%) were gram negative . All isolates with inhibitory activity are able to multiply at both 4 and 10 degrees C. J Food Prot, 2002 Dec, 65(12), 1869 - 72 Prevalence of Salmonella in two Botswana abattoir environments; Motsoela C et al.; A 1-year study was carried out to investigate the prevalence of Salmonella in two abattoir environments coded "A" and "B" in Gaborone, Botswana . The total number of environmental samples collected from abattoirs A and B was 250 and 300, respectively . The samples were taken from soils in the corrals, knife blades, saw blades, cattle-drinking water, cattle feces, and feed . Preenrichment, enrichment, and selective/differential media, which enabled the favorable growth of Salmonella, were used in the study . Salmonellae were present in all sampled environments . The most common serotypes found in the environment at abattoir A were E1, C1, C2, and B . Serotypes B, C1, C2, C3, and E1 were common in abattoir B . Antigenic characterization of the salmonellae isolates showed that Salmonella Anatum, Salmonella Azteca, Salmonella Saintpaul, Salmonella Cerro, and Salmonella Westhampton were predominant in abattoir A, whereas Salmonella Anatum, Salmonella Mbandaka, Salmonella Molade, Salmonella Reading, and Salmonella Oranienburg were dominant in abattoir B . Implementing hazard analysis critical control point principles in work procedures would definitely reduce the gross contamination taking place in abattoirs. Pediatr Med Chir, 2002 Sep-Oct, 24(5), 390 - 1 Case report: Salmonella panama osteomyelitis in a Ghanaian child with sickle cell disease; Busetti M et al.; Sickle cell disease is a rare condition in italian patients and even rarer are its complications, in particular Salmonella osteomyelitis . We describe a case of a Ghanaian child with sickle cell disease who developed osteomyelitis due to Salmonella panama, treated successfully with surgical debridement, followed by a prolonged period of specific antibiotic therapy. Science, 2002 Dec 20, 298(5602), 2390 - 2 Sir2-dependent activation of acetyl-CoA synthetase by deacetylation of active lysine; Starai VJ et al.; Acetyl-coenzyme A (CoA) synthetase (Acs) is an enzyme central to metabolism in prokaryotes and eukaryotes . Acs synthesizes acetyl CoA from acetate, adenosine triphosphate, and CoA through an acetyl-adenosine monophosphate (AMP) intermediate . Immunoblotting and mass spectrometry analysis showed that Salmonella enterica Acs enzyme activity is posttranslationally regulated by acetylation of lysine-609 . Acetylation blocks synthesis of the adenylate intermediate but does not affect the thioester-forming activity of the enzyme . Activation of the acetylated enzyme requires the nicotinamide adenine dinucleotide-dependent protein deacetylase activity of the CobB Sir2 protein from S . enterica . We propose that acetylation modulates the activity of all the AMP-forming family of enzymes, including nonribosomal peptide synthetases, luciferase, and aryl- and acyl-CoA synthetases . These findings extend our knowledge of the roles of Sir2 proteins in gene silencing, chromosome stability, and cell aging and imply that lysine acetylation is a common regulatory mechanism in eukaryotes and prokaryotes. Diagn Microbiol Infect Dis, 2002 Nov, 44(3), 313 - 8 Salmonella spp . isolates causing bloodstream infections in Latin America: report of antimicrobial activity from the SENTRY Antimicrobial Surveillance Program (1997-2000); Gales AC et al.; Recent years have seen a dramatic rise in the incidence and severity of cases of human salmonellosis . In addition, multidrug resistant strains have arisen . The objective of this study was to characterize the antimicrobial susceptibility profile of Salmonella spp . clinical isolates collected from Latin American medical centers as part of the SENTRY Antimicrobial Surveillance System . A total of 144 bloodstream Salmonella spp . isolates were collected between 1997 and 2000 . The susceptibility to diverse antimicrobial agents was tested by broth microdilution techniques according to the NCCLS recommendations . The Salmonella spp . strains were more frequently collected from adult patients (67.0%; 21-60 years) and isolated from Chile (28.5%) > Brazil (25.0%) > Mexico (18.8%) > Colombia (11.8%) . Ampicillin (MIC(50), 1 microg/ml) showed good in vitro activity (92.4% susceptibility) . Meropenem (MIC(50), 0.06 microg/ml) and gatifloxacin (MIC(50), 0.03 microg/ml) were the most potent compounds against the Salmonella spp . isolates (100.0% susceptible) followed by piperacillin/tazobactam (99.3%), ceftazidime (98.6%), ceftriaxone (98.6%), and amoxicillin/clavulanic acid (94.4%) . The lowest susceptibility rates were observed for tetracycline (87.5%) and trimethoprim/sulfamethoxazole (91.7%) . Four of 144 (2.8%) Salmonella spp . isolates demonstrated an ESBL phenotype; however, only two (1.4%) were confirmed as an inhibitable ESBL producer . The results of this study show that multidrug resistance, especially fluoroquinolone resistance, does not represent a serious problem among Salmonella isolates collected from the Latin American medical centers as monitored by the SENTRY Program. J Appl Microbiol, 2003, 94(1), 65 - 72 Comparison of phenotypic and genotypic markers for characterization of an outbreak of Salmonella serotype Havana in captive raptors; Reche MP et al.; AIMS: To establish a typing method for tracing the epidemic relationship of 16 strains of Salmonella serotype Havana isolated from captive raptors showing no symptomatology and residing in a wildlife hospital in Spain . METHODS AND RESULTS: Antimicrobial susceptibility testing, ribotyping, pulsed field gel electrophoresis (PFGE) and amplified fragment length polymorphism (AFLP) methodology were applied . Ten unrelated strains of serotype Havana were included as a control group to provide a basis of for the efficiency of the different markers used . All outbreak-related strains were resistant to nalidixic acid and streptomycin and showed the same ripotype, pulsotype and AFLP pattern . CONCLUSIONS: This is the first time that AFLP analysis has been tested with serotype Havana isolates and it has demonstrated to be the most useful epidemiological tool for discriminating between unrelated and outbreak-related strains of this serotype . The results obtained suggest that all the Salmonella serotype Havana isolates represented a common outbreak strain whose origin of contamination could not be established although it is thought that it was the poultry meat used for raptors'diet . SIGNIFICANCE AND IMPACT OF THE STUDY: Our study suggests the importance of microbiological analysis of these products in order to prevent contamination and dissemination of Salmonellae in this kind of Hospital. J Appl Microbiol, 2003, 94(1), 16 - 24 The incidence of antimicrobial-resistant Salmonella spp on freshly processed poultry from US Midwestern processing plants; Logue CM et al.; AIMS: To determine the incidence of antimicrobial-resistant Salmonella spp . on processed poultry (turkey) at Midwestern poultry plants . METHODS AND RESULTS: Two participating plants were visited at monthly intervals for a period of 1 year . Surface swabs were obtained from carcasses at two selected points on the production line, pre- and post-chill . In addition, samples of the chill water from chill tanks were also examined . Isolation and detection of Salmonella spp . from carcass swabs and chill water was carried out using standard enrichment techniques . Immunomagnetic separation was used to enhance the recovery of the pathogen . Salmonella isolates recovered were identified, serotyped and their antimicrobial resistance profiles determined using the National Antimicrobial Resistance Monitoring System . Results from the study indicated that the overall incidence of Salmonella was approx . 16.7%, with a greater incidence of the pathogen observed on pre-chill than post-chill carcasses . Salmonella isolates recovered displayed resistance to an average of four different antimicrobials . Approximately 15 different serotypes of Salmonella spp . were recovered, with Salmonella serotype Agona, Salmonella serotype Hadar, Salmonella serotype Heidelberg and Salmonella serotype Senftenberg being the most common . CONCLUSIONS: The incidence of Salmonella spp . was relatively low and isolates recovered showed significant degrees of antimicrobial resistance . Factors such as the processing plant examined, the season and farms that were presenting animals for processing influenced the incidence of the pathogen . SIGNIFICANCE AND IMPACT OF THE STUDY: Differences were observed in the serotypes of Salmonella recovered and the types of antimicrobial resistance found at the two plants . The study suggests that the use of antimicrobials at the farm level influences the creation of an environment that promotes the selection of antimicrobial-resistant Salmonella spp . The incidence, isolation and detection of Salmonella spp . on processed poultry are discussed. Mol Microbiol, 2003 Jan, 47(1), 103 - 18 Unravelling the biology of macrophage infection by gene expression profiling of intracellular Salmonella enterica; Eriksson S et al.; For intracellular pathogens such as Salmonellae, Mycobacteriae and Brucellae, infection requires adaptation to the intracellular environment of the phagocytic cell . The transition from extracellular to intravacuolar environment has been expected to involve a global modulation of bacterial gene expression, but the precise events have been difficult to determine . We now report the complete transcriptional profile of intracellular Salmonella enterica sv . Typhimurium following macrophage infection . During replication in murine macrophage-like J774-A.1 cells, 919 of 4451 S . Typhimurium genes showed significant changes in transcription . The expression profile identified alterations in numerous virulence and SOS response genes and revealed unexpected findings concerning the biology of the Salmonella-macrophage interaction . We observed that intracellular Salmonella are not starved for amino acids or iron (Fe2+), and that the intravacuolar environment is low in phosphate and magnesium but high in potassium . S . Typhimurium appears to be using the Entner-Douderoff pathway to use gluconate and related sugars as a carbon source within macrophages . Almost half the in vivo-regulated genes were of unknown function, suggesting that intracellular growth involves novel macrophage-associated functions . This is the first report that identifies the whole set of in vivo-regulated genes for any bacterial pathogen during infection of mammalian cells. Rinsho Byori, 2002 Nov, 50(11), 1041 - 6 {Microbiology--laboratory examinations for bacterias}; Hen R et al.; As it has been required to identify pathogenic microbes in shorter times, simple and rapid methods have been developed and used . Here, we summarized the present situation of rapid diagnostic testing in clinical microbiology in Japan, and also presented our results on PBP2' detection . The rapid test kits available in Japan for E . coli, Helicobacter pylori, Salmonella, Streptococcus and Staphylococcus aureus were described . Rapid examination methods are based mainly on immunologic reactions, which included slide agglutination using latex particle, immunochromatography and ELISA . Times required for the identification are 10 to 15 minutes . Moreover, rapid test kits employing PCR are also marketed . Further, we evaluated MRSA-LA "Seiken" which is a rapid detection kit for PBP2' produced by MRSA . The test was shown to be highly sensitive and specific . For the rapid identification of pathogenic microbes, simple and rapid test kits described here will be used more in clinical diagnosis. Clin Infect Dis, 2003 Jan 1, 36(1), 112 - 5 Epub 2002 Dec 11. Rapid diagnosis of acute Salmonella gastrointestinal infection; Oracz G et al.; Serologic tests for the detection of Salmonella serotype Enteritidis in children may become supplementary to stool culture examination . A total of 190 children were examined with a new 1-step, 2-minute test (TUBEX) that detects anti-Salmonella immunoglobulin M antibodies, which was found to be 92.6% sensitive and 94.8% specific (P<.0001). J Vet Med B Infect Dis Vet Public Health, 2002 Nov, 49(9), 438 - 44 Estimating the prevalence of Salmonella spp . in swine herds--influence of sensitivity and specificity of Salmonella detection; Steinbach G et al.; Information about the proportion of truly Salmonella-free herds is required for an evaluation of the epidemiological situation, the development of control strategies and their implementation . Findings regarding the presence of salmonellas in faeces and intestinal lymph nodes as well as the presence of Salmonella antibodies in meat juice from slaughtered pigs were obtained in the context of a study conducted by a number of institutes . These data were used for an analysis of the validity of data on the prevalence of infected animals within herds and on the prevalence of infected herds . The proportion of batches or herds with exclusively negative individual findings was found to depend not only on the true proportion of truly Salmonella-free animals within herds but quite essentially also on the distribution of the proportion of infected animals within herds, the sensitivity of the methods of examination and sample sizes . When taking into account the existing dependencies, it was found that among the swine, the real numbers of Salmonella carriers were much higher than shown by bacteriological and serological examination . Regarding salmonellosis in swine, also a number of contaminated herds must be expected which is far higher than that shown by the number of herds with positive findings in at least one animal . Even a low contamination of all or almost all herds would result in the numbers of 'negative' batches observed, i.e . batches with exclusively negative individual findings . A rating of the salmonella exposure of herds as high, low, or very low is possible and may, and should be, used for measures of consumer protection, irrespective of the proportion of truly Salmonella-free herds. J Med Microbiol, 2003 Jan, 52(Pt 1), 91 - 9 Lack of flagella disadvantages Salmonella enterica serovar Enteritidis during the early stages of infection in the rat; Robertson JM et al.; The roles of flagella and five fimbriae (SEF14, SEF17, SEF21, pef, lpf) in the early stages (up to 3 days) of Salmonella enterica serovar Enteritidis (S . Enteritidis) infection have been investigated in the rat . Wild-type strains LA5 and S1400 (fim+/fla+) and insertionally inactivated mutants unable to express the five fimbriae (fim-/fla+), flagella (fim+/fla-) or fimbriae and flagella (fim-/fla-) were used . All wild-type and mutant strains were able to colonize the gut and spread to the mesenteric lymph nodes, liver and spleen . There appeared to be little or no difference between the fim-/fla+ and wild-type (fim+/fla+) strains . In contrast, the numbers of aflagellate (fim+/fla- or fim-/fla-) salmonella in the liver and spleen were transiently reduced . In addition, fim+/fla- or fim-/fla- strains were less able to persist in the upper gastrointestinal tract and the inflammatory responses they elicited in the gut were less severe . Thus, expression of SEF14, SEF17, SEF21, pef and lpf did not appear to be a prerequisite for induction of S . Enteritidis infection in the rat . Deletion of flagella did, however, disadvantage the bacterium . This may be due to the inability to produce or release the potent immunomodulating protein flagellin. MMWR Morb Mortal Wkly Rep, 2002 Nov 22, 51(46), 1044 - 7 Multistate outbreaks of Salmonella serotype Poona infections associated with eating cantaloupe from Mexico--United States and Canada, 2000-2002; Spontaneous bacterial peritonitis in patients with hepatic cirrhosis: evaluation of a treatment protocol at specialized units; Dinis-Ribeiro M, Cortez-Pinto H, Marinho R, Valente A, Raimundo M, Salgado MJ, Ramalho F, Alexandrino P, Carneiro-de-Moura M. Liver Unit and Hepatology and Gastroenterology Intensive Care Unit, Department of Medicine II, University Hospital of Santa Maria, Lisbon, PortugalINTRODUCTION: Spontaneous bacterial peritonitis is a common and severe complication in patients with cirrhosis and ascitis . Its prognosis clearly depends on its precocious clinical recognition and efficacious therapy . AIM: To optimize a treatment protocol, after auditing clinical efficacy and describe microorganisms implicated at our institution . MATERIAL AND METHODS: Retrospective study of clinical files of patients with hepatic cirrhosis with positive culture of ascitic fluid (AF) and/or an AF polymorphonuclear (PMN) count of more than 250/mm3, treated at our units between 1st January, 2000 and 31st December, 2001 (n = 38) . Patients showed a median age of 49 years (30-76), 63% of which were male . Forty-eight percent were classified as belonging to Child-Pugh B class, and 52% to C . RESULTS: First, considering cases with PMN > 250/mm3 (n = 29), antibiotics were given to all patients (cefotaxime and ampiciline) . Fifty-two percent had hepatic encephalopathy, 42% had fever, 66% abdominal pain . In 42% a microorganism was isolated . Although 24% of fatal cases (only two related to infection), we noted a 73% clinical and laboratorial response . Five patients (72%) that died, showed renal failure by the time of death . Second, in all cases with positive culture of ascitic fluid (n = 21), 42% of which with PMN > 250/mm3 and 9 monobacterial nonneutrocytic bacterascites' cases, one only agent was found: E . coli in 36%, Streptococci (37%), Staphylococci (14%), and other (14%): Klebsiella oxytoca, n = 1; Salmonella enteritidis, n = 1; Enterococcus faecium, n = 1, Acinectobacter anitratus, n = 1 . Only one of the agents, E . faecium (3%) showed in vitro sensitivity exclusively to ampiciline; all other were cefotaxime sensitivite . CONCLUSIONS: Our protocol will be modified, to treat patients with spontaneous bacterial peritonitis with cefotaxime, as monotherapy . Albumin infusion will also be added to the protocol, as, we found renal failure to be an important negative prognosis factor. J Mol Evol, 2002 Dec, 55(6), 734 - 44 Gene Import or Deletion: A Study of the Different Genes in Escherichia coli Strains K12 and O157:H7; Hooper SD et al.; By comparing two strains of Escherichia coli (K12 and O157:H7) with an outgroup of Salmonella and Klebsiella species and analyzing the sets of genes which are present or absent in either of the three groups, we study the gene history of K12, in particular, since the respective divergences of these bacteria . Furthermore, by using a compositional method based on context bias, we evaluate not only recently imported genes but also deleted genes . In addition, we examine recent gene duplications in the two E . coli strains . It is found that turnover of DNA is high in E . coli and, more importantly, that turnover is highest for genes of low GC content . Although levels of import are high, most of the imported genes seem to be "junk" or have poorly understood functions . Nevertheless, selected genes do persist, and may even define some E . coli strains as pathogenic . Our results support the conclusion that some of the pathogenic islands in O157:H7 are likely to have been imported in recent time. J Bacteriol, 2003 Jan, 185(1), 374 - 6 Membrane topology of the ZntB efflux system of Salmonella enterica serovar Typhimurium; Caldwell AM et al.; The membrane topology of the ZntB Zn(2+) transport protein of Salmonella enterica serovar Typhimurium was determined by constructing deletion derivatives of the protein and genetically fusing them to blaM or lacZ cassettes . The enzymatic activities of the hybrid proteins indicate that ZntB is a bitopic integral membrane protein consisting largely of two independent domains . The first 266 amino acids form a large, highly charged domain within the cytoplasm, while the remaining 61 residues form a small membrane domain containing two membrane-spanning segments . The overall orientation towards the cytoplasm is consistent with the ability of ZntB to facilitate zinc efflux. J Bacteriol, 2003 Jan, 185(1), 332 - 9 The stm4066 gene product of Salmonella enterica serovar Typhimurium has aminoimidazole riboside (AIRs) kinase activity and allows AIRs to satisfy the thiamine requirement of pur mutant strains; Dougherty M et al.; In bacteria the biosynthetic pathways for purine mononucleotides and the hydroxymethyl pyrimidine moiety of thiamine share five reactions that result in the formation of aminoimidazole ribotide, the last metabolite common to both pathways . Here we describe the characterization of a Salmonella enterica mutant strain that has gained the ability to efficiently use exogenous aminoimidazole riboside (AIRs) as a source of thiamine . The lesion responsible for this phenotype is a null mutation in a transcriptional regulator of the GntR family (encoded by stm4068) . Lack of this protein derepressed transcription of an associated operon (stm4065-4067) that encoded a predicted kinase . The stm4066 gene product was purified and shown to have AIRs kinase activity in vitro . This activity was consistent with the model presented to explain the phenotype caused by the original mutation . This mutation provides a genetic means to isolate the synthesis of the hydroxymethyl pyrimidine moiety of thiamine from the pathway for purine mononucleotide biosynthesis and thus facilitate in vivo analyses. J Bacteriol, 2003 Jan, 185(1), 98 - 106 Lack of the ApbC or ApbE protein results in a defect in Fe-S cluster metabolism in Salmonella enterica serovar Typhimurium; Skovran E et al.; The isc genes function in the assembly of Fe-S clusters and are conserved in many prokaryotic and eukaryotic organisms . In most bacteria studied, the isc operon can be deleted without loss of cell viability, indicating that additional systems for Fe-S cluster assembly must exist . Several laboratories have described nutritional and biochemical defects resulting from mutations in the isc operon . Here we demonstrate that null mutations in two genes of unknown function, apbC and apbE, result in similar cellular deficiencies . Exogenous ferric chloride suppressed these deficiencies in the apbC and apbE mutants, distinguishing them from previously described isc mutants . The deficiencies caused by the apbC and isc mutations were additive, which is consistent with Isc and ApbC's having redundant functions or with Isc and ApbC's functioning in different areas of Fe-S cluster metabolism (e.g., Fe-S cluster assembly and Fe-S cluster repair) . Both the ApbC and ApbE proteins are similar in sequence to proteins that function in metal cofactor assembly . Like the enzymes with sequence similarity to ApbC, purified ApbC protein was able to hydrolyze ATP . The data herein are consistent with the hypothesis that the ApbC and ApbE proteins function in Fe-S cluster metabolism in vivo. Arch Orthop Trauma Surg, 2002 Dec, 122(9-10), 544 - 6 Epub 2002 Nov 05. A rare case of Salmonella osteomyelitis in the humerus as a differential diagnosis to a malignant bone tumor; Bettin D et al.; Salmonella osteomyelitis without predisposing factors is seldom seen and thus difficult to diagnose . We report on a 14-year-old healthy boy with Salmonella osteomyelitis which occurred 2 years after trauma . Radical operative debridement is recommended . Intravenous ciprofloxacin has proved to be effective because of good tissue penetration and sensitivity towards Salmonella. Microbiology, 2002 Dec, 148(Pt 12), 3789 - 99 O-antigen expression in Salmonella enterica serovar Typhi is regulated by nitrogen availability through RpoN-mediated transcriptional control of the rfaH gene; Bittner M et al.; The authors previously reported increased expression of the Salmonella enterica serovar Typhi (S . typhi) rfaH gene when the bacterial cells reach stationary phase . In this study, using a lacZ fusion to the rfaH promoter region, they demonstrate that growth-dependent regulation of rfaH expression occurs at the level of transcription initiation . It was also observed that production of the lipopolysaccharide (LPS) O-antigen by S . typhi Ty2 correlated with the differential expression of rfaH during bacterial growth . This was probably due to the increased cellular levels of RfaH, since expression of the distal gene in the O-antigen gene cluster of S . typhi Ty2, wbaP, was also increased during stationary growth, as demonstrated by RT-PCR analysis . Examination of the sequences upstream of the rfaH coding region revealed homologies to potential binding sites for the RcsB/RcsA dimer of the RcsC/YopJ/RcsB phosphorelay regulatory system and for the RpoN alternative sigma factor . The expression of the rfaH gene in rpoN and rcsB mutants of S . typhi Ty2 was measured . The results indicate that inactivation of rpoN, but not of rcsB, suppresses the growth-phase-dependent induction of rfaH expression . Furthermore, production of beta-galactosidase mediated by the rfaH-lacZ fusion increased approximately fourfold when bacteria were grown in a nitrogen-limited medium . Nitrogen limitation was also shown to increase the expression of the O-antigen by the wild-type S . typhi Ty2, as demonstrated by a similar electrophoretic profile to that observed during the stationary phase of growth in rich media . It is therefore concluded that the relationship between LPS production and nitrogen limitation parallels the pattern of rfaH regulation under the control of RpoN and is consistent with the idea that RpoN modulates LPS formation via its effect on rfaH gene expression during bacterial growth. Food Chem Toxicol, 2003 Feb, 41(2), 275 - 90 Implication of nitro group reduction in the mutagenic and chromosome damaging activities of 22 new 5-nitroisoquinolines by the Salmonella mutagenicity test and the cytokinesis-blocked micronucleus assay; Orsiere T et al.; The mutagenic (MUT) and chromosome-damaging (CHR) activities of 22 potential antimalarial drugs (5-nitroisoquinoline derivatives) were evaluated by the Salmonella test and the cytokinesis-blocked micronucleus assay (CBMN) . The Salmonella mutagenicity test was performed with and without metabolic activation (S9 mix) in S . typhimurium strains TA100 and YG1042 (an overproducing nitroreductase and O-acetyltransferase TA100 strain) . The CBMN was carried out on human lymphocytes without metabolic activation . Four concentrations were tested: 1, 10, 100 and 1000 ng/ml . MUT was expressed as minimal mutagenic concentrations (MMC, microM) and CHR was expressed as minimal chromosome-damaging concentrations (MCDC, nM) to compare both activities . All the 5-nitroisoquinoline compounds were mutagenic in TA100 . MMC ranged from 0.1 to 52.9 microM in TA100 . A statistically significant decrease in MMC was observed in YG1042 (8 x 10(-3) to 3.5 microM), implicating reduction of the nitro group . Modulation of MUT by S9 mix was not significant in TA100 and YG1042 . CHR was detected in 13 products for at least one concentration . Among the chromosome-damaging compounds, the MCDC ranged from 2.9 x 10(-3) to 3.6 nM . No relationship was found between MUT and CHR, suggesting two distinct pathways of DNA damage. Scand J Infect Dis, 2002, 34(10), 770 - 2 Multifocal simultaneous Salmonella typhi osteomyelitis in an immunocompetent adult; Toth F et al.; We report a case of Salmonella typhi osteomyelitis that presented as a pathologic fracture of the left femur and had additional silent manifestations in the right tibia and humerus in an otherwise healthy adult . This is the first report in the literature of multifocal simultaneous Salmonella osteomyelitis in an adult with no underlying medical condition. J AOAC Int, 2002 Nov-Dec, 85(6), 1338 - 40 In-house validation study for salmonella unique testing of juice (modification of AOAC official method 2000.07); Dailianis AE et al.; Following an industry request, a study was undertaken to validate a minor change to the Unique method for testing fruit juice . Twenty foods were tested in the original precollaborative study for TECRA Unique Salmonella test (2000.07) . To validate the modification for juice, both the modified method (42 degrees C module incubation with a 5 h replication step) and the current AOAC Method 2000.07 (37 degrees C incubation with a 4 h replication step) were compared with the U.S . Food and Drug Administration's Bacteriological Analytical Manual (BAM; 8th Ed., 1998) reference method, which uses lactose broth as pre-enrichment medium . Twenty uninoculated replicates, 20 replicates with low-level inoculum (target 1-5 cells/25 g), and 20 replicates with high-level inoculum (target 10-50 cells/25 g) were tested for a single batch of fresh orange juice in accordance with AOAC requirements . There was exact agreement between the 2 Unique methods for all samples and exact agreement between the 2 Unique methods and the BAM method for the uninoculated and high-level inoculum samples . For low-level inoculum, 17 samples were confirmed positive with the new Unique method, 17 with AOAC Method 2000.07, and 14 with the BAM method. Inflamm Res, 2002 Oct, 51(10), 500 - 5 Endotoxin tolerance in rats: influence on LPS-induced changes in excretory liver function; Dominguez Fernandez E et al.; OBJECTIVE AND DESIGN: We investigated in a rat model of endotoxic shock whether endotoxin tolerance (ETT) prevents lipopolysaccharide (LPS) associated lethality and studied the initial function of liver response to LPS . ANIMALS: Male Sprague-Dawley rats . TREATMENT: ETT was induced by i.p . injection of LPS (Salmonella friedenau) intraperitoneally over 5 days . Rats (n = 6 each group) received 1 mg LPS/kg b . w . intravenously (Salmonella friedenau) . The common bile duct was then canalized and bile was collected every 60 min for 6 h . 1 h after LPS-application liver biopsies were taken for determination of TNF-alpha by RT-PCR . Sham operated animals (n = 6 each group) were treated identically but without application of LPS . RESULTS: All ETT animals survived the duration of the experiment whereas non-tolerant animals (NETT) died before the end of the experiment (5/6) . NETT animals showed a continuous decrease in bile flow after 240 min . The amount of bile acids was significantly lower (ANOVA) in NETT animals than in sham operated controls or ETT-animals . Analysis of TNF-alpha mRNA expression in the liver revealed an upregulation 1 h after LPS application, which was significantly lower in LPS-tolerant animals . CONCLUSIONS: Our results show that excretory liver failure and death subsequent to intravenous LPS application can be successfully counteracted by induction of ETT. J Agric Food Chem, 2002 Dec 18, 50(26), 7639 - 44 Green-leaf-derived C6-aroma compounds with potent antibacterial action that act on both Gram-negative and Gram-positive bacteria; Nakamura S et al.; All eight C6-aliphatic alcohol and aldehyde compounds in naturally occurring green leaves showed bacteriostatic effects against Staphylococcus aureus IFO 12732, methicillin-resistant S . aureus, Escherichia coli IFO 3301, E . coli O157:H7, and Salmonella enteritidis, with bacteriostatic activities of less than 12.5 microg mL(-1) . In this study, the susceptibility of Gram-positive bacteria tested was observed to be greater than that of Gram-negative bacteria . The bactericidal action of the aldehyde compounds was found to be much stronger than that of the alcohol compounds under both liquid and gaseous conditions . The most effective compound was (3E)-hexenal at concentrations of 0.1 and 1 microg mL(-1), which killed 2.1 x 10(5) cfu mL(-1) of S . aureus IFO 12732 and 1.4 x 10(5) cfu mL(-1) of E . coli IFO 3301, respectively, by direct contact with the compound . Lethality of (3E)-hexenal against S . aureus IFO 12732 and E . coli IFO 3301 was also observed as a result of gaseous contact at concentrations of 3 and 30 microg mL(-1), respectively . The bactericidal effects of 30 microg mL(-1) (3E)-hexenal were thoroughly maintained throughout periods of 2 days and 1 day against S . aureus IFO 12732 and E . coli IFO 3301, respectively, by a complex formation with alpha-cyclodextrin. Eur J Biochem, 2002 Dec, 269(24), 6184 - 94 Oxidation of propionate to pyruvate in Escherichia coli . Involvement of methylcitrate dehydratase and aconitase; Brock M et al.; The pathway of the oxidation of propionate to pyruvate in Escherichia coli involves five enzymes, only two of which, methylcitrate synthase and 2-methylisocitrate lyase, have been thoroughly characterized . Here we report that the isomerization of (2S,3S)-methylcitrate to (2R,3S)-2-methylisocitrate requires a novel enzyme, methylcitrate dehydratase (PrpD), and the well-known enzyme, aconitase (AcnB), of the tricarboxylic acid cycle . AcnB was purified as 2-methylaconitate hydratase from E . coli cells grown on propionate and identified by its N-terminus . The enzyme has an apparent Km of 210 micro m for (2R,3S)-2-methylisocitrate but shows no activity with (2S,3S)-methylcitrate . On the other hand, PrpD is specific for (2S,3S)-methylcitrate (Km = 440 micro m) and catalyses in addition only the hydration of cis-aconitate at a rate that is five times lower . The product of the dehydration of enzymatically synthesized (2S,3S)-methylcitrate was designated cis-2-methylaconitate because of its ability to form a cyclic anhydride at low pH . Hence, PrpD catalyses an unusual syn elimination, whereas the addition of water to cis-2-methylaconitate occurs in the usual anti manner . The different stereochemistries of the elimination and addition of water may be the reason for the requirement for the novel methylcitrate dehydratase (PrpD), the sequence of which seems not to be related to any other enzyme of known function . Northern-blot experiments showed expression of acnB under all conditions tested, whereas the RNA of enzymes of the prp operon (PrpE, a propionyl-CoA synthetase, and PrpD) was exclusively present during growth on propionate . 2D gel electrophoresis showed the production of all proteins encoded by the prp operon during growth on propionate as sole carbon and energy source, except PrpE, which seems to be replaced by acetyl-CoA synthetase . This is in good agreement with investigations on Salmonella enterica LT2, in which disruption of the prpE gene showed no visible phenotype. Acta Chir Belg, 2002 Oct, 102(5), 348 - 50 Perforation due to ileocaecal salmonellosis; Bos WT et al.; A 54-year old male patient was admitted with a tentative diagnosis of biliary pancreatitis . After 3 days, he developed an acute abdomen with a pneumoperitoneum . A laparotomy was performed: multiple perforations of the terminal ileum and a necrotic gallbladder were found . A right hemicolectomy with 30 cm of the terminal ileum and a cholecystectomy were carried out . At the same time, positive blood cultures for Salmonella typhi were obtained and this confirmed the diagnosis of ileal perforation due to salmonella typhi infection . This case illustrates that Salmonella typhi infection can be a surgical problem in the western countries. Hum Mol Genet, 2002 Dec 15, 11(26), 3361 - 9 Identification of the gene responsible for the cblB complementation group of vitamin B12-dependent methylmalonic aciduria; Dobson CM et al.; The methylmalonic acidurias are metabolic disorders resulting from deficient methylmalonyl-CoA mutase activity, a vitamin B(12)-dependent enzyme . We have cloned the gene for the cblB complementation group caused by deficient activity of a cob(I)alamin adenosyltransferase . This was accomplished by searching bacterial genomes for genes in close proximity to the methylmalonyl-CoA mutase gene that might encode a protein with the properties of an adenosyltransferase . A candidate was identified in the Archaeoglobus fulgidus genome and was used to probe the human genome database . It yielded a gene on chromosome 12q24 that encodes a predicted protein of 250 amino acids with 45% similarity to PduO in Salmonella enterica, a characterized cob(I)alamin adenosyltransferase . A northern blot revealed an RNA species of 1.1 kb predominating in liver and skeletal muscle . The gene was evaluated for deleterious mutations in cblB patient cell lines . Several mutations were identified including a 5 bp deletion (5del572gggcc576), two splice site mutations (IVS2-1G>T, IVS3-1G>A), andt several point mutations (A135T, R186W, R191W and E193K) . Two additional amino acid substitutions (R19Q and M239K) were found in several patient cell lines but were found to be common polymorphisms (36% and 46%) in control alleles . The R186W mutation, which we suggest is disease-linked, is present in four of the six patient cell lines examined (homoallelic in two) and in 4 of 240 alleles in control samples . These data confirm that the identified gene, MMAB, corresponds to the cblB complementation group and has the appearance of a cob(I)alamin adenosyltransferase, as predicted from biochemical data. Int Immunopharmacol, 2002 Nov, 2(12), 1641 - 5 Effects of intraperitoneal and intranasal application of Lentinan on cellular response in rats; Markova N et al.; Lentinan (Ajinomoto, Japan) was administrated intraperitoneally (i.p.) and intranasally (i.n.) at different doses (1, 5 and 10 mg/kg) to rats . Effectiveness of Lentinan treatment was evaluated by comparative testing of cell activation (establishing the number, glycolytic and acid phosphatase activity, H2O2 production and killing ability against Salmonella enteritidis and Staphylococcus aureus) at two different compartments--peritoneal and broncho-alveolar cavities . The results indicated that Lentinan induced high-grade activation of peritoneal cells (PCs) and especially of broncho-alveolar cells (BACs) with markedly enhanced effector function (killing ability against S . aureus) . Generally, Lentinan, known usually with its parenteral routes of application, can be successful to stimulate the host cell response in the respiratory tract by intranasal route of administration. J Gen Appl Microbiol, 2002 Aug, 48(4), 193 - 9 Effects of ozone treatment on cell growth and ultrastructural changes in bacteria; Thanomsub B et al.; Ozone appeared to inhibit growth and caused the death of gram negative and gram positive tested bacteria: Escherichia coli, Salmonella sp., Staphylococcus aureus and Bacillus subtilis . Bacterial cultures at 10(3), 10(4), 10(5), 10(6), and 10(7) cfu/ml dilution were exposed to 0.167/mg/min/L of ozone at different time intervals (0, 5, 10, 15, 30, 60, 90, 120, and 150 min) . Cell viability was observed in all types of tested bacteria at 10(3), 10(4), 10(3) cfu/ml within 30 min after ozone exposure . However, cell inactivation was not significantly observed at concentrations of 10(6), 10(7) cfu/ml even after an exposure of 150 min . Ultrastructural changes of treated bacteria showed deformation, rough damage and surface destruction revealed by scanning electron microscopy . Some bacterial cells showed collapsed and shrunken patterns within 60 min and severe rupture and cellular lysis after 90 min of ozone treatment . This study supports the proposed mechanism of the bacteria inactivation by ozone that caused cell membrane destruction and finally lysis reaction . Thus, the precaution of using ozone as a biocide should be used to address appropriate concentrations of bacterial contamination in water. Nutr Cancer, 2002, 43(1), 111 - 8 Mutagenicity of tocopheryl quinones: evolutionary advantage of selective accumulation of dietary alpha-tocopherol; Cornwell DG et al.; We have shown that phenolic antioxidant tocopherols are oxidized to nonarylating alpha-tocopheryl quinone (alpha-TQ) and arylating gamma- and delta-TQ electrophiles . The arylating quinones stimulate apoptosis and are highly cytotoxic in mammalian cells . Some xenobiotic phenolic antioxidants are mutagens, and it has been suggested that their arylating quinone metabolites are the active agents in mutagenesis related to carcinogenesis . We found that neither alpha- nor gamma-TQ was directly genotoxic in supercoiled-to-nicked circular DNA conversions, but these agents interacted with the cytomegalovirus reporter-driven plasmid and enhanced luciferase transfection, with gamma-TQ > alpha-TQ . The Ames test, using gamma-TQ and a number of Salmonella strains, showed no evidence of bacterial mutagenesis . gamma-TQ was highly cytotoxic and alpha-TQ slightly cytotoxic in eukaryocyte AS52 cells . A guanosine phosphoribosyltransferase gene assay showed that gamma-TQ was highly mutagenic and alpha-TQ slightly mutagenic in AS52 cells . A review of the literature identified associations where a decrease in dietary gamma-tocopherol (gamma-T) diminishes and an increase in dietary gamma-T and its quinone enhances carcinogenicity . Humans and other omnivores selectively accumulate alpha-tocopherol, even though gamma-T is their principal dietary tocopherol . We suggest that this selectivity confers an evolutionary advantage by limiting tissue gamma-T, a putative precursor of the mutagen gamma-TQ. Am J Forensic Med Pathol, 2002 Dec, 23(4), 382 - 5 Fatal salmonella aortitis with mycotic aneurysm rupture; Salzberger LA et al.; Salmonellae most commonly cause uncomplicated cases of gastroenteritis but have a predilection for damaged blood vessels, especially those damaged by atherosclerosis . The abdominal aorta is most frequently affected . The most serious complication of aortitis is mycotic aneurysm formation with subsequent rupture . The authors present the case of a 61-year-old man who was found unresponsive at home 3 days after discharge from the hospital for treatment of gastroenteritis with bacteremia . Postmortem examination revealed a ruptured mycotic aneurysm with a large retroperitoneal hematoma . Numerous gram-negative rods were embedded in the wall of the aorta and surrounding inflammatory infiltrate, compatible with the patient's previously isolated . Whereas abdominal aortic aneurysm rupture is most commonly associated with atherosclerosis, the isolation of from blood cultures, coupled with radiographic evidence of gas surrounding the aorta, should raise the suspicion of infectious aortitis . Whereas fatal rupture of an aortic aneurysm secondary to atherosclerosis alone or in conjunction with aortitis will not have an impact on the manner of death, infections are reportable and thus have public health implications. Anim Genet, 2002 Dec, 33(6), 407 - 14 Microsatellite markers associated with quantitative trait loci controlling antibody response to Escherichia coli and Salmonella enteritidis in young broilers; Yunis R et al.; A unique resource population was produced to facilitate detection of microsatellite markers associated with quantitative trait loci controlling antibody (Ab) response in broiler chickens . Three F1 males were produced by mating two lines divergently selected on Ab response to Escherichia coli vaccination . Each F1 male was mated with females from four genetic backgrounds: F1, high-Ab line (HH), low-Ab line and commercial line, producing three resource families, each with four progeny types . About 1700 chicks were immunized with E . coli and Salmonella enteritidis vaccines . Selective genotyping was conducted on the individuals with highest or lowest average Ab to E . coli and S . enteritidis within each progeny type in each sire family . Twelve markers were significantly associated with Ab to E . coli and six of them were also associated with Ab to S . enteritidis, mostly exhibiting a similar low effect (approximately 0.35 phenotypic SD) in all progeny types . Four markers exhibited a highly significant and much larger effect (approximately 1.7 SD), but only in progeny of females from the HH, suggesting that a backcross to the high parental line should be preferred over the commonly used F2 population . Results from two markers suggested a quantitative trait locus on chromosome 2 around 400 cM . The marker MCW0083, significant in two sire families, is closely linked to the bone morphogenetic protein 2 (BMP2) gene, known to be associated with the control of T-cell transformation in humans. Cell Microbiol, 2002 Dec, 4(12), 813 - 24 SseF and SseG are translocated effectors of the type III secretion system of Salmonella pathogenicity island 2 that modulate aggregation of endosomal compartments; Kuhle V et al.; The type III secretion system encoded by Salmonella pathogenicity island 2 (SPI 2) is important for intracellular proliferation in infected host cells . Intracellular Salmonella use this system to translocate a set of effector proteins into the host cell . We studied the role of SseF and SseG, two SPI 2-encoded proteins . SseF and SseG are not required for translocation of effector proteins such as SseJ, encoded by genes outside of SPI 2 . Rather, both proteins are translocated and interact with phagosomal membranes after translocation . In infected epithelial cells the formation of Salmonella-induced filaments, endosomal aggregates rich in lysosomal glycoproteins, is dependent on the function of SPI 2 . We observed that, in mutant strains deficient for sseF or sseG, the formation of aggregated endosomes can take place, but the composition of the structures is different from those observed in cells infected with Salmonella wild type . These observations indicate that SseF and SseG modulate the aggregation of host endosomes. J Bone Joint Surg Br, 2002 Nov, 84(8), 1167 - 72 Septic arthritis of the shoulder in children in Malawi . A randomised, prospective study of aspiration versus arthrotomy and washout; Smith SP et al.; We undertook a prospective study of 61 children in Malawi with septic arthritis of the shoulder . They were randomised into two groups, treated by aspiration (group 1, 31 patients) or arthrotomy (group 2, 30 patients) . Both received antibiotics for six weeks . We studied the results of blood tests, microbiology, and the clinical and radiological outcome one year after diagnosis . Only one patient was sickle-cell positive and three were HIV-positive . Non-typhoidal Salmonella species accounted for 86% (19/22) of the positive joint cultures in group 1 and 73% (16/22) in group 2 . Of the 33 radiographs available for review at follow-up at six months, 23 (70%) showed evidence of glenohumeral damage . There was no statistical difference in radiological outcome for the two groups . We devised and validated a scoring system, the Blantyre Septic Joint Score, for the assessment of joints based upon swelling, tenderness, function and range of movement . Despite the radiological changes only one of the 24 joints examined at one year had any deficit in these parameters . There was no statistical difference in the clinical outcome for the two treatment groups at any stage during the period of follow-up. Shock, 2002 Dec, 18(6), 561 - 6 Removal of phosphate from lipid A as a strategy to detoxify lipopolysaccharide; Bentala H et al.; Lipopolysaccharide (LPS) may cause sepsis when