J Agric Food Chem, 2003 Aug 13, 51(17), 5154 - 61 Preparation of antimicrobial reduced lysozyme compatible in food applications; Touch V et al.; The structural and antimicrobial functions of lysozyme reduced with food-compatible reducing agents-cysteine (Cys) and glutathione (GSH)-were investigated . The disulfide bonds were partially reduced by thiol-disulfide exchange reactions under heat-induced denaturing conditions from 55 to 90 degrees C . The results showed that treatment of lysozyme with Cys and GSH resulted in the introduction of new half-cystine residues (2-3 residues/mol of protein) . The released SH groups, in turn, rendered the lysozyme molecule more flexible, being accompanied by a dramatic increase in the surface hydrophobicity and exposure of tryptophan residues . As a consequence, the resulting reduced lysozymes were more capable of binding to lipopolysaccharides (LPS) and permeabilizing the bacterial outer membrane, as evidenced by the liposome leakage experiment, than were native or heated lysozyme . Both reduced lysozymes displayed significantly higher antimicrobial activity than native or heated lysozyme against Salmonella enteritidis (SE) in sodium phosphate buffer (10 mM, pH 7.2) at 30 degrees C for 1 h . Their minimal inhibitory concentrations (MICs) against the tested bacteria were about 150- and 25-fold lower than their respective MICs of native or heated lysozyme . The results suggest that partially reduced lysozyme could be used as a potential antimicrobial agent for prevention of SE attack. Appl Environ Microbiol, 2003 Aug, 69(8), 4556 - 60 Survival of Salmonella enterica in freshwater and sediments and transmission by the aquatic midge Chironomus tentans (Chironomidae: Diptera); Moore BC et al.; Survival of a nalidixic acid-resistant strain of Salmonella enterica serovar Typhimurium mr-DT-104 in water and sediments was tested using artificially contaminated aquaria . Water samples remained culture positive for salmonella for up to 54 days . Sediment samples were culture positive up to 119 days . In addition, potential mechanisms for spreading salmonella in the environments by chironomid larvae and adults were tested . We evaluated the acquisition of mr-DT-104 by chironomids from contaminated aquatic sediments and subsequent spread to uncontaminated sediments . Larval chironomids raised in contaminated sediments became culture positive, and the bacteria were carried over to adults after emergence . Contamination of clean sediments by chironomid larvae was not demonstrated . These findings clearly suggest that mr-DT-104 serovar organisms can survive in aquatic sediments for at least several months . Uptake of salmonellae by chironomid larvae and adults suggests that they are possible vectors of mr-DT-104 in both aquatic and terrestrial environments, although the role of larval defecation in movement of bacteria to new sediments was not demonstrated. Appl Environ Microbiol, 2003 Aug, 69(8), 4352 - 8 Characterization of the RpoS status of clinical isolates of Salmonella enterica; Robbe-Saule V et al.; The stationary-phase-inducible sigma factor, sigma(S) (RpoS), is the master regulator of the general stress response in Salmonella and is required for virulence in mice . rpoS mutants can frequently be isolated from highly passaged laboratory strains of Salmonella: We examined the rpoS status of 116 human clinical isolates of Salmonella, including 41 Salmonella enterica serotype Typhi strains isolated from blood, 38 S . enterica serotype Typhimurium strains isolated from blood, and 37 Salmonella serotype Typhimurium strains isolated from feces . We examined the abilities of these strains to produce the sigma(S) protein, to express RpoS-dependent catalase activity, and to resist to oxidative stress in the stationary phase of growth . We also carried out complementation experiments with a cloned wild-type rpoS gene . Our results showed that 15 of the 41 Salmonella serotype Typhi isolates were defective in RpoS . We sequenced the rpoS allele of 12 strains . This led to identification of small insertions, deletions, and point mutations resulting in premature stop codons or affecting regions 1 and 2 of sigma(S), showing that the rpoS mutations are not clonal . Thus, mutant rpoS alleles can be found in freshly isolated clinical strains of Salmonella serotype Typhi, and they may affect virulence properties . Interestingly however, no rpoS mutants were found among the 75 Salmonella serotype Typhimurium isolates . Strains that differed in catalase activity and resistance to hydrogen peroxide were found, but the differences were not linked to the rpoS status . This suggests that Salmonella serotype Typhimurium rpoS mutants are counterselected because rpoS plays a role in the pathogenesis of Salmonella serotype Typhimurium in humans or in the transmission cycle of the disease. Chem Biol Interact, 2003 Jul 25, 146(1), 19 - 25 Structural activity relationship between Salmonella-mutagenicity and nitro-orientation of nitroazaphenanthrenes; Tokiwa H et al.; Nitroazaphenanthrenes (NAphs) and their N-oxides (NAphOs) were synthesized as derivatives with nitrogen atoms in the 1, 4, and 9 positions of phenanthrene rings, and as nitrated derivatives substituted at the 1, 2, 3, 4, 5, 6, 7, and 8 positions of phenanthrene rings . To determine the structure activity relationship of these derivatives, all 19 isomers were bioassayed with Salmonella tester strains . NAphs substituted at the 4, 6, 7 and 8 positions were mutagenic for TA98, and 1-, 2-, and 3-N-9-AphOs, 6-N-1-AphO and 6-N-4-AphO were mutagenic for TA98 and TA100 without the S9 mix, while 5-N-1-AphO and 5-N-9-AphO were non- or weakly mutagenic . Nitrated derivatives, 6-N-4-Aph, 6-N-9-Aph, 6-N-1-AphO, and 6-N-4-AphO, were powerful mutagens for TA98 and TA100 . Mutagenicity was enhanced by mutant strains producing nitroreductase, such as YG1021 and 1026, and by those producing O-acetyltransferase, such as YG1024 and 1029 . Nitro derivatives substituted at positions 4 and 5 in the phenanthrene rings were perpendicular, while those at positions 2, 3, 6 and 7 were coplanar to the phenanthrene rings . NAphs substituted at the 1 and 8 positions were noncoplanar due to steric hindrance of the aromatic proton at the peri position . On the other hand, 1,5- and 1,8-dinitro-4-azaphenanthrenes showed high mutagenicity for strains TA98 and TA100 in the absence of the S9 mix, and were strongly enhanced by nitroreductase and O-acetyltransferase, over-producing mutants . Therefore, it was found that the mutagenic potency of NAphs and NAphOs was closely associated with the chemical properties and orientation of nitro substitution of aromatic rings. Expert Rev Vaccines, 2003 Feb, 2(1), 31 - 43 Advances in the development of bacterial vector technology; Kochi SK et al.; The demand for new and improved vaccines against human diseases has continued unabated over the past century . While the need continues for traditional vaccines in areas such as infectious diseases, there is an increasing demand for new therapies in nontraditional areas, such as cancer treatment, bioterrorism and food safety . Prompted by these changes, there has been a renewed interest in the application and development of live, attenuated bacteria expressing foreign antigens as vaccines . The application of bacterial vector vaccines to human maladies has been studied most extensively in attenuted strains of Salmonella . Live, attenuated strains of Shigella, Listeria monocytogenes, Mycobacterium bovis-BCG and Vibrio cholerae provide unique alternatives in terms of antigen delivery and immune presentation, however and also show promise as potentially useful bacterial vectors. Prev Vet Med, 2003 Aug 8, 60(2), 155 - 65 Single versus double testing of meat-juice samples for Salmonella antibodies, in the Danish pig-herd surveillance programme; Ekeroth L et al.; In Denmark, a national serological surveillance-and-control programme for Salmonella in pigs has been in operation since 1995 . The programme is based on the Danish mix-ELISA and uses double testing (two ELISA-wells used per sample) of meat-juice samples taken in relation to slaughter . All herds are classified monthly into one of the three levels; the classification is based on the percentage of positive serological results in the previous 3 months . In connection with evaluation of the programme in 2001, we investigated whether single testing (testing in one well only) could be expected to be sufficiently precise compared to double testing . Data from the year 2000 were used, and mathematical modelling . Single testing was simulated by randomised selection of one of the two results in the double testing . A slight increase in the prevalence of Salmonella-positive samples (1.02-1.09 times more through the four quarters of the year 2000) was found in the simulated single testing, as compared to the double testing . Around 0.5% of the herds would be allocated to another herd level in single testing-almost equal numbers one level up and one level down . No herd being seronegative in double testing would be allocated to levels 2 or 3 (herds with >40 or >70%, respectively, serological reactors) in single testing . The prevalence of "false-positive" diagnoses (positive in single testing and negative in double testing) and inversely defined "false-negative" diagnoses varied from 4.2 to 8.7% and from 3.2 to 4.5%, respectively, through the four quarters of the year 2000 . The probability of allocating a herd to a wrong level due to sampling error was on the average 6.2 (varying from 1.66 to over 100) times higher than the probability of allocating a herd to a wrong level due to the test inaccuracy introduced by going from double to single testing . This is, however, an average; a herd with a true prevalence close to one of the level border cut-offs (40 and 70% weighted seroprevalence, respectively) would have a higher risk of being allocated to a wrong level than a herd with a true prevalence far from the level border cut-offs . The results are based on the current Danish sample sizes in the surveillance scheme, which implies that 60, 75 or 100 samples are taken annually in a herd, depending on its size . Other sample sizes would produce other results. Ann Univ Mariae Curie Sklodowska {Med}, 2002, 57(2), 15 - 20 Foodborne infections and intoxications in the Lublin voivodeship in the years 1980-2000 in comparison with their prevalence in the Polish population; Kalinowski P; The aim of the paper is presentation of foodborne infections and intoxications registered in the Lublin voivodeship in comparison with their nationwide prevalence in the whole country in the years 1980-2000 . The analysed material comprised incidence rates of foodborne infections and intoxications, of which those caused by Salmonellas of animal source, registered in Poland and in the Lublin voivodeship in the studied years . In the analysed period in the Lublin voivodeship and throughout Poland there occurred a huge increase in incidence of foodborne infections and intoxications . The epidemics caused by Salmonella strains to the greatest extent influenced the increase in total number of cases of the disease, caused by deterioration in sanitary situation of the country, hygiene of the society and decline in quality of food of animal origin . The epidemiological situation of the disease in the Lublin voievodship is worse than nationwide average. Ann Univ Mariae Curie Sklodowska {Med}, 2002, 57(2), 9 - 14 Prevalence of infections of the Salmonella strains in the Lublin voivodeship and in Poland in the years 1980-2000; Kalinowski P; The aim of the paper is presentation of the prevalence of infections of the Salmonella strains in the Lublin voivodeship in the years 1980-2000 in comparison with their nationwide prevalence in the same period . The analysed material comprised epidemiological data concerning the incidence rates of salmonelloses registered in Poland in the studied period . Since the beginning of the 1980s there was observed a constantly increasing trend in incidence rates of salmonelloses . After 1988 throughout Poland and after 1990 in the Lublin voivodeship there was noted gradual decline in incidence of infections caused by Salmonellas . Administrative decisions and recent improvements in sanitary situation in our country caused appearance of that favourable, decreasing trend of incidence rates of salmonelloses . The statistics concerning extraintestinal infections with Salmonellas are still unreliable, especially in the Lublin voivodeship. Mol Genet Genomics, 2003 Oct, 270(1), 56 - 65 Epub 2003 Jul 30. The gyr genes of Salmonella enterica serovar Typhimurium are repressed by the factor for inversion stimulation, Fis; Keane OM et al.; The DNA sequence of the gyr genes from Salmonella enterica serovar Typhimurium revealed strong similarity between gyrB and its counterpart in Escherichia coli . However, the gyrA gene showed similarity to the E . coli homologue only downstream from the Pribnow box of the promoter, with the sequence upstream diverging markedly . Since this region encompasses the binding sites for the Fis DNA binding protein in E . coli, we investigated the possibility that the gyrA genes in the two species might differ in their responses to this regulatory protein . Fis was found to act as a transcriptional repressor of both gyr genes in S . enterica . In electrophoretic mobility shift assays, Fis was found to bind to both the gyrA and gyrB promoters of S . enterica, despite the strong divergence from the E . coli sequence on the part of the former . The binding sites were mapped by DNase I protection assays, and the results are consistent with conservation of the mechanism of Fis-mediated repression between the two bacterial species. Can J Microbiol, 2003 May, 49(5), 326 - 35 Prevalence of Escherichia coli O157:H7 and Salmonella spp . in surface waters of southern Alberta and its relation to manure sources; Johnson JY et al.; The Oldman River watershed in southern Alberta, Canada, is an extensively irrigated region in which intensive agricultural practices have flourished . Concern over water quality in the basin has been expressed because of high levels of enteric disease indigenous to the region . To address these concerns, we conducted a 2-year study to estimate the prevalence of Escherichia coli O157:H7 and Salmonella spp . in surface water within the basin . This study is the first of its kind to identify E . coli O157:H7 repeatedly in surface water collected from a Canadian watershed . Prevalence of E . coli O157:H7 and Salmonella spp . in water samples was 0.9% (n = 1,483) and 6.2% (n = 1,429), respectively . While data examined at a regional level show a relationship between high livestock density and high pathogen levels in southern Alberta, statistical analysis of point source data indicates that predicted manure output from bovine, swine, and poultry feeding operations was not directly associated with either Salmonella spp . or E . coli O157:H7 prevalence . However, geography and weather variables, which are likely to influence bacterial runoff, were not considered in this model . We also postulate that variations in time, amount, and frequency of manure application onto agricultural lands may have influenced levels of surface-water contamination with these bacterial pathogens. J Bacteriol, 2003 Aug, 185(16), 4973 - 82 Genomic profiling of iron-responsive genes in Salmonella enterica serovar typhimurium by high-throughput screening of a random promoter library; Bjarnason J et al.; The importance of iron to bacteria is shown by the presence of numerous iron-scavenging and transport systems and by many genes whose expression is tightly regulated by iron availability . We have taken a global approach to gene expression analysis of Salmonella enterica serovar Typhimurium in response to iron by combining efficient, high-throughput methods with sensitive, luminescent reporting of gene expression using a random promoter library . Real-time expression profiles of the library were generated under low- and high-iron conditions to identify iron-regulated promoters, including a number of previously identified genes . Our results indicate that approximately 7% of the genome may be regulated directly or indirectly by iron . Further analysis of these clones using a Fur titration assay revealed three separate classes of genes; two of these classes consist of Fur-regulated genes . A third class was Fur independent and included both negatively and positively iron-responsive genes . These may reflect new iron-dependent regulons . Iron-responsive genes included iron transporters, iron storage and mobility proteins, iron-containing proteins (redox proteins, oxidoreductases, and cytochromes), transcriptional regulators, and the energy transducer tonB . By identifying a wide variety of iron-responsive genes, we extend our understanding of the global effect of iron availability on gene expression in the bacterial cell. J Bacteriol, 2003 Aug, 185(16), 4837 - 43 Residues C123 and D58 of the 2-methylisocitrate lyase (PrpB) enzyme of Salmonella enterica are essential for catalysis; Grimek TL et al.; The prpB gene of Salmonella enterica serovar Typhimurium LT2 encodes a protein with 2-methylisocitrate (2-MIC) lyase activity, which cleaves 2-MIC into pyruvate and succinate during the conversion of propionate to pyruvate via the 2-methylcitric acid cycle . This paper reports the isolation and kinetic characterization of wild-type and five mutant PrpB proteins . Wild-type PrpB protein had a molecular mass of approximately 32 kDa per subunit, and the biologically active enzyme was comprised of four subunits . Optimal 2-MIC lyase activity was measured at pH 7.5 and 50 degrees C, and the reaction required Mg(2+) ions; equimolar concentrations of Mn(2+) ions were a poor substitute for Mg(2+) (28% specific activity) . Dithiothreitol (DTT) or reduced glutathione (GSH) was required for optimal activity; the role of DTT or GSH was apparently not to reduce disulfide bonds, since the disulfide-specific reducing agent Tris(2-carboxyethyl)phosphine hydrochloride failed to substitute for DTT or GSH . The K(m) of PrpB for 2-MIC was measured at 19 micro M, with a k(cat) of 105 s(-1) . Mutations in the prpB gene were introduced by site-directed mutagenesis based on the active-site residues deemed important for catalysis in the closely related phosphoenolpyruvate mutase and isocitrate lyase enzymes . Residues D58, K121, C123, and H125 of PrpB were changed to alanine, and residue R122 was changed to lysine . Nondenaturing polyacrylamide gel electrophoresis indicated that all mutant PrpB proteins retained the same oligomeric state of the wild-type enzyme, which is known to form tetramers . The PrpB(K121A), PrpB(H125A), and PrpB(R122K) mutant proteins formed enzymes that had 1,050-, 750-, and 2-fold decreases in k(cat) for 2-MIC lyase activity, respectively . The PrpB(D58A) and PrpB(C123A) proteins formed tetramers that displayed no detectable 2-MIC lyase activity indicating that both of these residues are essential for catalysis . Based on the proposed mechanism of the closely related isocitrate lyases, PrpB residue C123 is proposed to serve as the active site base, and residue D58 is critical for the coordination of a required Mg(2+) ion. J Bacteriol, 2003 Aug, 185(16), 4748 - 54 DapE can function as an aspartyl peptidase in the presence of Mn2+; Broder DH et al.; Extracts of a multiply peptidase-deficient (pepNABDPQTE iadA iaaA) Salmonella enterica serovar Typhimurium strain contain an aspartyl dipeptidase activity that is dependent on Mn(2+) . Purification of this activity followed by N-terminal sequencing of the protein suggested that the Mn(2+)-dependent peptidase is DapE (N-succinyl-L,L-diaminopimelate desuccinylase) . A dapE chromosomal disruption was constructed and transduced into a multiply peptidase-deficient (MPD) strain . Crude extracts of this strain showed no aspartyl peptidase activity, and the strain failed to utilize Asp-Leu as a leucine source . The dapE gene was cloned into expression vectors in order to overproduce either the native protein (DapE) or a hexahistidine fusion protein (DapE-His(6)) . Extracts of a strain carrying the plasmid overexpresssing native DapE in the MPD dapE background showed a 3,200-fold elevation of Mn(2+)-dependent aspartyl peptidase activity relative to the MPD dapE(+) strain . In addition, purified DapE-His(6) exhibited Mn(2+)-dependent peptidase activity toward aspartyl dipeptides . Growth of the MPD strain carrying a single genomic copy of dapE on Asp-Leu as a Leu source was slow but detectable . Overproduction of DapE in the MPD dapE strain allowed growth on Asp-Leu at a much faster rate . DapE was found to be specific for N-terminal aspartyl dipeptides: no N-terminal Glu, Met, or Leu peptides were hydrolyzed, nor were any peptides containing more than two amino acids . DapE is known to bind two divalent cations: one with high affinity and the other with lower affinity . Our data indicate that the form of DapE active as a peptidase contains Zn(2+) in the high-affinity site and Mn(2+) in the low-affinity site. Eur J Cardiothorac Surg, 2003 Aug, 24(2), 320 - 2 Successful treatment of Salmonella mycotic aneurysm of the descending thoracic aorta; Lin CY et al.; Salmonella mycotic aneurysms of the descending thoracic aorta are exceedingly rare . There are few case reports and even fewer reports of long term survival . The case of a 68-year-old female presenting with a mycotic aneurysm of the descending thoracic aorta caused by Salmonella species is described, which involved successful surgical intervention. Berl Munch Tierarztl Wochenschr, 2003 Jul-Aug, 116(7-8), 293 - 8 {Intracellular survival of Salmonella strains in Caco-2 cells}; Dinjus U et al.; In the in vitro model using Caco-2 cells at different stages of differentiation the invasion and intracellular survival of virulent (predominant infection strains) and less virulent (predominant attenuated mutant strains) Salmonella strains were studied . The statistical evaluation of experimental data has shown that the logarithmized colony forming unit after 18 hours of incubation in differentiated cells (14 days old) is a suitable parameter for the determination of intracellular survival . Using this parameter a relationship between intracellular survival and Salmonella virulence (LD50 mouse) was demonstrated and quantified . The model presented could be suitable for the replacement of animal experiments after further investigations. Berl Munch Tierarztl Wochenschr, 2003 Jul-Aug, 116(7-8), 281 - 7 {Usefulness of serological examinations in the analysis of Salmonella infections in pig herds}; Steinbach G et al.; The development of the antibody concentration against lipopolysaccharide (LPS) of S . Typhimurium und S . Choleraesuis in rearing pigs during the fattening period and in breeding sows of the corresponding age was recorded . The studies revealed the following results . Antibodies of isotypes IgG1 and IgG2 revealed a more pronounced specificity against the according Salmonella serovar than IgM antibodies . The calculated "antibody percent value" based on the total amount of Salmonella antibodies is mainly determined by the IgM antibodies in sera and meat juice, respectively . In fattening pigs a significant increase of antibodies against IgM and total Ig was observed between week 3 and 10 after beginning of the rearing period . In breeding pigs this increase was detectable already earlier . In only 3 out of 10 groups an increase of IgG1 and IgG2 was also seen . The detected significant increase of total Ig and IgM in the other groups might be the result of a less intensive exposure to salmonellas or it might be due to an increase of unspecific antibodies induced by other antigens . Serological investigations represent a valuable tool to record the intensity and development in time of the Salmonella exposure in pigs farms . Examination of total Ig is an appropriate method to detect pig herds with a high level of Salmonella exposure, for detailed epidemiological studies in pig farms the examination of antibody isotypes will give more comprehensive information. FEMS Microbiol Lett, 2003 Jul 29, 224(2), 291 - 7 msDNA-St85, a multicopy single-stranded DNA isolated from Salmonella enterica serovar Typhimurium LT2 with the genomic analysis of its retron; Ahmed AM et al.; Bacterial reverse transcriptase is responsible for the production of a small satellite DNA-RNA complex called multicopy single-stranded DNA (msDNA) that has been found in a wide variety of Gram-negative bacteria . Here we describe the isolation and characterization of a novel msDNA, msDNA-St85, from Salmonella enterica serovar Typhimurium LT2 . We determined the nucleotide sequence of msDNA-St85 and the location of retron-St85 on the chromosome that is responsible for msDNA-St85 production by analyzing the complete genomic sequence of S . typhimurium LT2 . It was found that the G+C content and the codon usage of retron-St85 were significantly different from those of the S . typhimurium genome, indicating that retron-St85 was probably acquired recently in this bacterium . This is the first report for identification of an msDNA in the genus Salmonella with the complete description and analysis of its retron. FEMS Microbiol Lett, 2003 Jul 29, 224(2), 239 - 46 Identification of the CysB-regulated gene, hslJ, related to the Escherichia coli novobiocin resistance phenotype; Lilic M et al.; The cysB gene product is a LysR-type regulatory protein required for expression of the cys regulon . cysB mutants of Escherichia coli and Salmonella, along with being auxotrophs for the cysteine, exhibit increased resistance to the antibiotics novobiocin (Nov) and mecillinam . In this work, by using lambdaplacMu9 insertions creating random lacZ fusions, we identify a gene, hslJ, whose expression appeared to be increased in cysB mutants and needed for Nov resistance . Measurements of the HSLJ::lacZ gene fusion expression demonstrated that the hslJ gene is negatively regulated by CysB . In addition we observe the negative autogenous control of HslJ . When the control imposed by CysB is lifted in the cysB mutant, the elevation of Nov resistance can be achieved only in the presence of wild-type hslJ allele . A double cysB hslJ mutant restores the sensitivity to Nov . Overexpression of the wild-type HslJ protein either in a cysB(+) or a cysB(-) background increases the level of Nov resistance indicating that hslJ product is indeed involved in accomplishing this phenotype . The HSLJ::OmegaKan allele encodes the C-terminaly truncated mutant protein HslJ Q121Ter which is not functional in achieving the Nov resistance but when overexpressed induces the psp operon . Finally, we found that inactivation of hslJ does not affect the increased resistance to mecillinam in cysB mutants. Emerg Infect Dis, 2003 Jul, 9(7), 774 - 80 Salmonella control programs in Denmark; Wegener HC et al.; We describe Salmonella control programs of broiler chickens, layer hens, and pigs in Denmark . Major reductions in the incidence of foodborne human salmonellosis have occurred by integrated control of farms and food processing plants . Disease control has been achieved by monitoring the herds and flocks, eliminating infected animals, and diversifying animals (animals and products are processed differently depending on Salmonella status) and animal food products according to the determined risk . In 2001, the Danish society saved U.S.$25.5 million by controlling Salmonella . The total annual Salmonella control costs in year 2001 were U.S.$14.1 million (U.S.$0.075/kg of pork and U.S.$0.02/kg of broiler or egg) . These costs are paid almost exclusively by the industry . The control principles described are applicable to most industrialized countries with modern intensive farming systems. Can J Vet Res, 2003 Jul, 67(3), 219 - 24 Comparison of bacterial enriched-broth culture, enzyme linked immunosorbent assay, and broth culture-polymerase chain reaction techniques for identifying asymptomatic infections with Salmonella in swine; Sibley J et al.; A polymerase chain reaction (PCR) assay was combined with a broth-culture enrichment system to detect Salmonella shed in feces from subclinically infected swine . The effectiveness of the broth culture-polymerase chain reaction (BC-PCR) assay to identify pigs shedding Salmonella in feces was compared with a microbiological culture and a commercial enzyme linked immunosorbent assay (ELISA) kit to detect Salmonella-specific serum antibody . A total of 67 pigs were tested by each of the 3 methodologies . Forty-one pigs tested positive for Salmonella by BC-PCR and ELISA identified 6 positives and 23 suspicious samples . It was shown that the BC-PCR assay is a rapid diagnostic tool for detecting of Salmonella shed by asymptomatic swine compared with current diagnostic technologies. Scand J Gastroenterol, 2003 Jul, 38(7), 719 - 26 Luminal salmonella endotoxin affects epithelial and mast cell function in the proximal colon of pigs; Aschenbach JR et al.; BACKGROUND: Salmonellosis and systemic endotoxaemia affect intestinal function . However, little is known about the functional importance of luminal Salmonella (S.) endotoxin during intestinal infection . METHODS: Pigs were either given or not given lipopolysaccharide (LPS, 30 mg day(-1)) of S . Typhimurium DT-104 orally for 14 days . Blood samples were taken weekly . After slaughter (day 14), epithelia of the proximal colon were investigated in Ussing chambers . Bacterial translocations to lung, liver, spleen and several lymph nodes were determined by culture . RESULTS: Endotoxin feeding increased plasma C-reactive protein (CRP) and histamine levels without evoking clinical signs . Postmortem, proximal colonic epithelia of LPS-treated animals showed both a decreased histamine release after mast cell stimulation with A23187 and a smaller increase in short-circuit current after A23187 application . Addition of the nitric oxide donor, sodium nitroprusside (SNP), also elicited lower increases in short-circuit current in the proximal colon of endotoxin-treated pigs . Endotoxin pre-feeding decreased colonic ion conductance, although mannitol and histamine fluxes were high in some epithelia of this group . Luminal Salmonella endotoxin increased bacterial translocation to proximal jejunal lymph nodes . LPS applied to colonic epithelia in vitro had no electrophysiological effects . CONCLUSIONS: Luminal endotoxin elicits an acute phase response and affects intestinal electrolyte transport and mast cell function . Furthermore, LPS induces epithelial spots of increased mannitol permeability that could be identical to spots of enhanced bacterial translocation. Biotechnol Bioeng, 2003 Sep 20, 83(6), 721 - 8 Detection and identification of Escherichia coli, Shigella, and Salmonella by microarrays using the gyrB gene; Kakinuma K et al.; Commonly, 16S ribosome RNA (16S rRNA) sequence analysis has been used for identifying enteric bacteria . However, it may not always be applicable for distinguishing closely related bacteria . Therefore, we selected gyrB genes that encode the subunit B protein of DNA gyrase (a topoisomerase type II protein) as target genes . The molecular evolution rate of gyrB genes is higher than that of 16S rRNA, and gyrB genes are distributed universally among bacterial species . Microarray technology includes the methods of arraying cDNA or oligonucleotides on substrates such as glass slides while acquiring a lot of information simultaneously . Thus, it is possible to identify the enteric bacteria easily using microarray technology . We devised a simple method of rapidly identifying bacterial species through the combined use of gyrB genes and microarrays . Closely related bacteria were not identified at the species level using 16S rRNA sequence analysis, whereas they were identified at the species level based on the reaction patterns of oligonucleotides on our microarrays using gyrB genes . Nucleic Acids Res, 2003 Aug 1, 31(15), 4517 - 22 Transcription increases multiple spontaneous point mutations in Salmonella enterica; Hudson RE et al.; The spontaneous rate of G.C-->A.T mutations and a hotspot T.A-->G.C transversion are known to increase with the frequency of transcription-increases that have been ascribed primarily to processes that affect only these specific mutations . To investigate how transcription induces other spontaneous point mutations, we tested for its effects in repair-proficient Salmonella enterica using reversion assays of chromosomally inserted alleles . Our results indicate that transcription increases rates of all tested point mutations in the induced gene: induction significantly increased the individual rates of an A.T-->T.A transversion, an A.T-->G.C transition and the pooled rates of the three other point mutations assayed . Although the S.enterica genome is thought to have a mutational bias towards G.C base pairs, transitions creating A.T pairs were approximately 10 times more frequent than the reverse mutation, resulting in an overall mutation pressure to lower G+C contents . Transitions occurred at roughly twice the rate of transversions, similar to results from sequence comparisons; however, several individual transversions are more frequent than the least common transition. Avian Dis, 2003 Apr-Jun, 47(2), 387 - 95 A restriction fragment length polymorphism-based polymerase chain reaction as an alternative to serotyping for identifying Salmonella serotypes; Hong Y et al.; The phase 1 (fliC) and phase 2 (fljB) Salmonella flagella genes were analyzed by restriction fragment length polymorphism (RFLP)-polymerase chain reaction (PCR) to aid in the identification of different Salmonella serotypes . Twenty-four phase 1 flagellin and eight phase 2 flagellin genes could be differentiated among each other with restriction endonucleases Sau3A and HhaI in RFLP-PCR analysis . These flagellin genes comprise the major antigenic formulas for 52 serotypes of Salmonella sp., which include the common serotypes found in poultry and other important food animal species . With the knowledge of the O antigen composition determined from conventional O serotyping, 90% of the Salmonella serotypes could be identified by this double restriction enzyme RFLP analysis of fliC and fljB genes . This RFLP-PCR flagellar typing scheme was successfully applied to the identification of serotype for 112 Salmonella isolates obtained from poultry environment . There was a significant correlation between RFLP-PCR and conventional serotyping (chi-square, P < 0.001) . Overall, PCR-RFLP proved to be a fast, accurate, and economical alternative approach to serotyping Salmonella sp. Avian Dis, 2003 Apr-Jun, 47(2), 380 - 6 Rapid detection of Salmonella from poultry by real-time polymerase chain reaction with fluorescent hybridization probes; Eyigor A et al.; Detection of Salmonella by bacteriologic methods is known to be time consuming . Therefore, we have developed a real-time probe-specific polymerase chain reaction (PCR) to rapidly detect Salmonella invA gene-based PCR products from chicken feces and carcasses by a fluorescence resonance energy transfer assay . The sensitivity and the specificity of this system were determined as 3 colony-forming units ml(-1) and 100%, respectively . Overnight tetrathionate broth enrichment cultures of chicken feces and carcass samples were used in template preparation for PCR . Also, a standard bacteriology was performed (National Poultry Improvement Plan-U.S . Department of Agriculture, Bacteriological Analytical Manual-Food and Drug Administration Center for Food Safety and Applied Nutrition) for confirmation . Seventy-two cloacal swab, 147 intestine, and 50 carcass (neck) samples were examined . Thirteen (8.8%) and 25 (17%) of the intestinal samples were found to harbor Salmonella by bacteriology and PCR, respectively . Forty-five of 50 (90%) carcass samples were Salmonella positive by both methods . Salmonella was not detected from cloacal swab samples . Results indicate that this assay has the potential for use in routine monitoring and detection of Salmonella in infected flocks and carcasses. Anim Biotechnol, 2003 May, 14(1), 61 - 76 Candidate gene approach: potentional association of caspase-1, inhibitor of apoptosis protein-1, and prosaposin gene polymorphisms with response to Salmonella enteritidis challenge or vaccination in young chicks; Liu W et al.; Salmonella enteritidis (SE) contamination of poultry products is a major cause of foodborne disease worldwide . Caspase-1 and inhibitor of apoptosis protein-1 (IAP-1) were selected as candidate genes for chicken response to SE because their proteins play critical roles in the apoptotic pathway when intracellular bacteria interact with host cells . Prosaposin (PSAP) was selected as a positional candidate gene based on a previous quantitative trait loci (QTL) linkage study using the same population . The F1 offspring of outbred sires crossed with three diverse, highly inbred dam lines (two major histocompatibility complex-congenic Leghorn lines named G-B1 and G-B2, and one Fayoumi line) were used to define the phenotypes . The F1 birds were involved in either pathogenic SE challenge, in which spleen and cecum content bacterial load were quantified, or SE vaccination, in which plasma antibody level to SE vaccine was evaluated . A polymerase chain reaction-restriction fragment length polymorphisms (PCR-RFLP) assay was developed to identify single-nucleotide polymorphism (SNP) in the three genes . The F1 offspring of heterozygous sires for each gene were genotyped . The sire caspase-1 gene was significantly associated with cecum content bacterial load (P = 0.04) in the three combined dam line crosses, and with spleen bacterial load in the G-B1 cross (P=0.02) . The sire caspase-1 gene was also significantly associated with antibody level to SE vaccine (P=0.03) in F1 males in the three combined dam line crosses . The sire IAP-1 gene was significantly associated with spleen bacterial load (P=0.04) in the three combined dam-line crosses, and interacted with dam-line genetics (P = 0.01) for cecum content bacterial load . The sire PSAP gene significantly interacted with sex for spleen bacterial load (P = 0.004) . This study is the first to demonstrate the association of SNPs for caspase-1, IAP-1, and PSAP genes with SE vaccine and with pathogen challenge response in chickens. Zh Mikrobiol Epidemiol Immunobiol, 2003 May-Jun, (3), 79 - 80 {Detection of lymphocytes binding erythrocytes conjugated with Salmonella antigens}; Denisova TG et al.; Immune reagents for the detection of specific antigen-binding lymphocytes (ABL) with respect to different Salmonells antigens were developed . Rabbits were immunized with killed S . typhi and other salmonellae containing cross-reacting antigens, and the dynamics of the formation of ASL of each specificity was studied . Differences in the time of the appearance of ASL with receptors to thymus-independent (09, 12 or Vi) and thymus-dependent (Hd) antigens were studied . The relative content of ASL, determined with the use of immune reagents prepared from S . typhi antigens, was higher, on the whole, in rabbits immunized with S . typhi than in rabbits immunized with salmonellae containing one of cross-reacting antigens (S . enteritidis--09, 12; S . paratyphi C--Vi; S . virginia--Hd). Mem Inst Oswaldo Cruz, 2003 Apr, 98(3), 419 - 23 Epub 2003 Jul 18. The effect of nitric oxide combined with fluoroquinolones against Salmonella enterica serovar Typhimurium in vitro; Coban AY et al.; Two regulons, soxRS and marRAB, are associated with resistance to quinolones or multiple antibiotic in Salmonella enterica serovar Typhimurium . These regulons are activated by nitric oxide and redox-cycling drugs, such as paraquat and cause on activation of the acrAB-encoded efflux pump . In this study, we investigated the effect of nitric oxide (NO) alone and in combination with ofloxacin, ciprofloxacin, and pefloxacin against S . typhimurium clinical isolates and mutant strains in vitro . We did not observe synergistic effect against clinical isolates and SH5014 (parent strain of acr mutant), while we found synergistic effect against PP120 (soxRS mutant) and SH7616 (an acr mutant) S . typhimurium for all quinolones . Our results suggest that the efficiencies of some antibiotics, including ofloxacin, ciprofloxacin, and pefloxacin are decreased via activation of soxRS and marRAB regulons by NO in S . enterica serovar Typhimurium . Further studies are warranted to establish the interaction of NO with the genes of Salmonella and, with multiple antibiotic resistance. Folia Microbiol (Praha), 2003, 48(3), 339 - 45 Is Pseudobutyrivibrio xylanivorans strain Mz5T suitable as a probiotic? An in vitro study; Cepeljnik T et al.; Rumen bacterium Pseudobutyrivibrio xylanivorans strain Mz5T possessed a potent xylanolytic enzyme system consisting of at least 7 different xylan hydrolases with molar mass 27-145 kDa . Three of them were successfully isolated in active native form . This strain produced butyrate and lactate on different saccharides . cis-9, trans-11-Conjugated linoleic acid was also detected in the culture medium . Bacteriocin-like inhibitory substances of Mz5T were active against some strains of rumen bacteria and against selected Salmonella and E . coli isolates from poultry meat . The strain Mz5T retained viability and xylanolytic activity also under not fully anaerobic conditions; its cells attached to the Caco-2 cells so that its successful association with gut epithelial cells may be expected . These in vitro results confirmed several probiotic traits of the isolate Mz5T and justified further in vivo experiments to test its ability to improve animal health and performance. Arkh Patol, 2003 May-Jun, 65(3), 36 - 8 {Proliferative processes in the epithelium of colon mucosa in patients with Salmonella infection during first 24 hours after the onset of the disease}; Mokretsova EV et al.; DNA synthesis in the epithelium of a distal portion of human colon was studied by radioautography with 3H-thymidine in patients during the first 24 hours of gastrointestinal salmonellosis . All 13 patients aged 18-50 years were admitted to hospital during 9-20 hours after the disease onset . Nobody had a background of gastrointestinal pathology . The control group consisted of samples from sigmoid colon epithelium of 30 healthy individuals . The labelled nuclei index (LNI) was 5.16 +/- 0.27%, the intensity of labelling (IL) was 13.65x +/- 0.82 . Endoscopic and pathomorphological studies of colonic mucosa in salmonellosis by the end of the first 24 hours of the disease showed superficial inflammation . Increased LNI (11.48 +/- 1.07%, p < 0.05) characterized activation of proliferation and increased IL (22.24 +/- 2.1, p < 0.05) showed intensification of DNA synthesis . Additionally, in one third of cases enlargement of proliferation compartment up to 2/3 crypt was noticed . It is unlikely that in this situation immune mechanisms play the key role, it seems that a direct stimulation of proliferative processes by a bacterial agent takes place. Neuroradiology, 2003 Aug, 45(8), 541 - 5 Epub 2003 Jul 16. Extracranial internal carotid artery Salmonella mycotic aneurysm complicated by occlusion of the internal carotid artery: depiction by color Doppler sonography, CT and DSA; Sidiropoulou MS et al.; Mycotic aneurysms of the extracranial carotid artery are rare . Seventy-four cases have been described in the medical literature and only eight secondary to Salmonella infection . To our knowledge, color Doppler sonography, computed tomography (CT), and digital subtraction angiography (DSA) findings relating to the diagnosis and follow-up of extracranial internal carotid artery mycotic aneurysm complicated by occlusion have not previously been described in the literature . We present a report of color Doppler sonography, CT, and DSA findings of a mycotic aneurysm of the right extracranial internal carotid artery due to Salmonella associated with occlusion of the internal carotid artery, promptly diagnosed and followed up using these imaging modalities. Ann Hematol, 2003 Oct, 82(10), 646 - 8 Epub 2003 Jul 22. Suppurative salmonella thyroiditis in a patient with chronic lymphocytic leukemia; Dai MS et al.; We describe an 82-year-old man with undiagnosed chronic lymphocytic leukemia (CLL) who presented with acute swelling of the thyroid goiter . Subsequent thyroid aspirate and blood culture yielded group B Salmonella thyroid abscess with septicemia . Infectious complications are the major cause of morbidity and mortality in patients with CLL since most of them can be timely detected and few can arise from innocent-looking lesions. Int J Food Microbiol, 2003 Aug 25, 85(3), 237 - 48 Invasion of Salmonella enteritidis in avian intestinal epithelial cells in vitro is influenced by short-chain fatty acids; Van Immerseel F et al.; Fermentation reactions in the caeca of chickens, the predominant place for Salmonella colonization, result in high concentrations of short-chain fatty acids (SCFA) . Thus Salmonella bacteria are in close contact with SCFA during their life cycle . A study was carried out to analyse the effects of SCFA on invasion of Salmonella enteritidis in an avian intestinal epithelial cell line . Preincubation of S . enteritidis for 4 h in growth media supplemented with various concentrations of propionate or butyrate resulted in decreased invasion compared to bacteria, preincubated in nonsupplemented media, and to bacteria, preincubated in media supplemented with formate or acetate . Incubation of the S . enteritidis bacteria in media supplemented with mixtures of SCFA mimicking the in vivo caecal concentrations resulted in increased invasion compared with butyrate-exposed bacteria, but equal invasion compared with nonexposed bacteria . Increasing the butyrate concentration in these mixtures did not modify invasion compared with the original mixtures. Am J Pathol, 2003 Aug, 163(2), 711 - 21 An angiogenic switch in macrophages involving synergy between Toll-like receptors 2, 4, 7, and 9 and adenosine A(2A) receptors; Pinhal-Enfield G et al.; Adenosine A(2A) receptor (A(2A)R) agonists synergize with Escherichia coli (E . coli) LPS {toll-like receptor (TLR)4 agonist} to up-regulate vascular endothelial growth factor (VEGF) expression in murine macrophages . Here, we demonstrate that TLR2, TLR7, and TLR9, but not TLR3 and TLR5 agonists, also synergize with A(2A)R agonists and adenosine to up-regulate VEGF, while simultaneously strongly down-regulating TNFalpha expression . In the absence of adenosine or A(2A)R agonists, Porphyromonas gingivalis (P . gingivalis) LPS and PAM(3)CAG (TLR2 agonists), resiquimod (R848) (TLR7 agonist), and non-methylated CpG DNA (TLR9 agonist) strongly up-regulate TNFalpha expression, with no effect on VEGF . In the presence of adenosine or A(2A)R agonists, but not A(1)R agonists, TLR2, 4, 7, and 9 agonists strongly up-regulate VEGF expression, while simultaneously down-regulating TNFalpha . C57BL/10ScN (TLR4 deletion mutant) macrophages produce TNFalpha in response to TLR2, 3, 7, and 9 agonists, but not the TLR4 agonist E . coli LPS . With adenosine or A(2A)R agonists, TLR2, 7, and 9, but not TLR4 agonists, also synergistically up-regulate VEGF, while down-regulating TNFalpha expression . Polyinosinic-polycytidilic acid (poly(I:C)) (TLR3 agonist) stimulates TNFalpha expression in macrophages from both C57BL/10ScSn and C57BL/10ScN mice, but has little effect on VEGF expression in the presence of adenosine or A(2A)R agonists . R-flagellins from Serratia marcescens (S . marcescens) and Salmonella muenchen (S . muenchen) do not stimulate TNFalpha expression in either C57BL/10ScSn or C57BL10/ScN mice, and have no effect on VEGF production in the presence of adenosine or A(2A)R agonists . While adenosine and A(2A)R agonists strongly down-regulate TNFalpha protein expression induced by TLR2, 3, 4, 7, and 9 agonists, TNFalpha mRNA and NF-kappaB activation are not reduced . We propose a novel signaling pathway in murine macrophages involving synergy between TLRs 2, 4, 7, and 9 and A(2A)Rs, that up-regulates VEGF and down-regulates TNFalpha expression, thus acting as an angiogenic switch . This angiogenic switch may play an important role in ischemia when TLR agonists are present, providing an interface between innate immunity and wound healing. Infect Immun, 2003 Aug, 71(8), 4795 - 803 Secreted effector proteins of Salmonella enterica serotype typhimurium elicit host-specific chemokine profiles in animal models of typhoid fever and enterocolitis; Zhang S et al.; Infection of bovine ligated loops with the Salmonella enterica serotype Typhimurium wild type but not a sipA sopABDE2 mutant resulted in fluid accumulation, polymorphonuclear cell infiltration, and expression of CXC chemokines, particularly GRO alpha . None of these sipA sopABDE2-dependent responses was observed in murine-ligated loops . The majority of GRO alpha transcripts localized to bovine intestinal epithelium . Thus, different disease outcomes between mice (i.e., no diarrhea) and calves (i.e., diarrhea) may be due to differences in sipA sopABDE2-dependent CXC chemokine gene expression in epithelial cells. Infect Immun, 2003 Aug, 71(8), 4733 - 41 Induction of antimicrobial pathways during early-phase immune response to Salmonella spp . in murine macrophages: gamma interferon (IFN-gamma) and upregulation of IFN-gamma receptor alpha expression are required for NADPH phagocytic oxidase gp91-stimulated oxidative burst and control of virulent Salmonella spp; Foster N et al.; The effect of gamma interferon (IFN-gamma) on elevation of reactive oxygen species and the viability of virulent wild-type and avirulent mutants of Salmonella enterica serovar Typhimurium and S . enterica serovar Infantis was studied in a murine macrophage cell line (J774.2 cells) . S . enterica serovar Typhimurium 14028 phoP and a rough lipopolysaccharide mutant of S . enterica serovar Infantis 1326/28 (phi(r)) (avirulent mutants) induced NADPH phagocytic oxidase gp91 (gp91(phox)) activity and a significant (P < 0.05) elevation of reactive oxygen species within 12 h without coculture with IFN-gamma . This coincided with reduced survival of S . enterica serovar Typhimurium14028 phoP or stasis of S . enterica serovar Infantis phi(r) . Fluorometric studies indicated that expression of IFN-gamma on infected J774.2 cells was not significantly (P > 0.05) elevated . However, studies with the virulent S . enterica serovar Typhimurium strains showed that a comparable level of control of bacterial numbers could only be achieved by coculture with IFN-gamma . This coincided with significant upregulation of IFN-gamma receptor alpha expression on the surface of J774.2 cells and was completely abolished by N-acetyl-L-cysteine captopril (an inhibitor of reactive oxygen species) . Delay in reactive oxygen species induction due to a requirement for IFN-gamma and upregulation of IFN-gamma receptor alpha in macrophages infected with virulent salmonellae may result in greater dissemination of virulent salmonellae in host tissue. Infect Immun, 2003 Aug, 71(8), 4664 - 73 Construction, characterization, and immunogenicity of an attenuated Salmonella enterica serovar typhimurium pgtE vaccine expressing fimbriae with integrated viral epitopes from the spiC promoter; Chen H et al.; Transmissible gastroenteritis virus (TGEV) is a porcine coronavirus that causes diarrhea, leading to near 100% mortality in neonatal piglets with corresponding devastating economic consequences . For the protection of neonatal and older animals, oral live vaccines present the attractive property of inducing desired mucosal immune responses, including colostral antibodies in sows--an effective means to passively protect suckling piglets . Newly attenuated Salmonella vaccine constructs expressing TGEV S protein epitopes were studied and evaluated for improved humoral immune response to TGEV . The macrophage-inducible Salmonella ssaH and spiC/ssaB promoters were compared for their ability to express the TGEV C and A epitopes in the context of the heterologous 987P fimbriae on Salmonella vaccines . Compared to the ssaH promoter, the Salmonella cya crp vector elicited significantly higher levels of mucosal and systemic antibodies in orally immunized mice when the chimeric fimbriae were expressed from the spiC promoter . The Salmonella spiC promoter construct induced the highest level of chimeric fimbriae after being taken up by the J774A.1 macrophagelike cells . The Salmonella cya crp vaccine vector was shown to incorporate into 987P partially degraded chimeric subunits lacking the TGEV epitopes . In contrast, its isogenic pgtE mutant produced fimbriae consisting exclusively of intact chimeric subunits . Mice immunized orally with the Salmonella pgtE vaccine expressing chimeric fimbriae from the spiC promoter elicited significantly higher systemic and mucosal antibody titers against the TGEV epitopes compared to the parental vaccine . This study indicates that the Salmonella cya crp pgtE vector and the spiC promoter can be used successfully to improve immune responses toward heterologous antigens. Infect Immun, 2003 Aug, 71(8), 4382 - 8 Human alveolar macrophages infected by virulent bacteria expressing SipB are a major source of active interleukin-18; Obregon C et al.; Recent publications have demonstrated that the protease caspase-1 is responsible for the processing of pro-interleukin 18 (IL-18) into the active form . Studies on cell lines and murine macrophages have shown that the bacterial invasion factor SipB activates caspase-1, triggering cell death . Thus, we investigated the role of SipB in the activation and release of IL-18 in human alveolar macrophages (AM), which are the first line of defense against inhaled pathogens . Under steady-state conditions, AM are a more important source of IL-18 than are dendritic cells (DC) and monocytes . Cytokine production by AM and DC was compared after both types of cells had been infected with a virulent strain of Salmonella enterica serovar Typhimurium and an isogenic sipB mutant, which were used as an infection model . Infection with virulent Salmonella led to marked cell death with features of apoptosis while both intracellular activation and release of IL-18 were demonstrated . In contrast, the sipB mutant did not induce such cell death or the release of active IL-18 . The specific caspase-1 inhibitor Ac-YVAD-CMK blocked the early IL-18 release in AM infected with the virulent strain . However, the type of Salmonella infection did not differentially regulate IL-18 gene expression . We concluded that the bacterial virulence factor SipB plays an essential posttranslational role in the intracellular activation of IL-18 and the release of the cytokine in human AM. Poult Sci, 2003 Jul, 82(7), 1170 - 3 Invasion of Salmonella enteritidis in the tissues of reproductive organs in laying Japanese quail: an immunocytochemical study; Takata T et al.; The aim of this study was to determine whether Salmonella enteritidis (SE) inoculated into the peritoneal cavity would colonize tissues of reproductive organs in Japanese quail hens . Quail hens regularly laying were intraperitoneally inoculated with 5 x 10(7) or 5 x 10(8) SE cells, and the ovary, oviduct, kidney, spleen, liver, and large intestine were excised 24 or 48 h after the treatment . Paraffin sections of these organs were immunostained for SE . Invasion of SE was found in the tissues of the ovarian stroma, the follicular wall including superficial and theca layers, and occasionally in the granulosa layer and yolk . The SE immunoreaction product frequently was found in the fibroblast-like and macrophage-like cells in the stroma and surface layer of follicles . The SE immunoreaction products were identified on the mucosal surface, in the mucosal epithelium, and in the stromal tissues in all segments of the oviduct . Many of the bacteria were contained in the cytoplasm of mucosal epithelial cells and stromal cells in those tissues . The SE immunoreactions were also found in the tissues of kidney, spleen, and liver and in the large intestine . These results suggest that SE organisms introduced into the peritoneal cavity can invade and colonize the tissues of ovary and oviduct and may be responsible in the production of contaminated eggs. Vet Res Commun, 2003 May, 27(4), 257 - 73 Evaluation of treatment and prophylaxis with nitrofurans and comparison with alternative antimicrobial agents in experimental Salmonella enterica Serovar enteritidis infection in chicks; Chadfield MS et al.; The ability of the nitrofuran antimicrobial agents furazolidone and furaltadone to prevent, reduce or eliminate Salmonella enterica serovar enteritidis PT4 infection in artificially challenged day-old chicks was evaluated . Treating the birds with the nitrofurans failed to eliminate established infections with either furazolidone-resistant (FzR) or furazolidone-sensitive (FzS) strains . Simultaneous administration of the nitrofurans to day-old chicks challenged with FzS failed to prevent infection but reduced colonization significantly (p<0.05) compared to unmedicated controls . No reduction of colonization occurred with FzR . Challenging birds with FzS and simultaneous dosing with nitrofurans for 1 week, followed by a second week of continued treatment, resulted in an increase in the level of colonization in the second week rather than a decrease . Dosing with the nitrofurans (200 ppm) for 1 week prior to challenge with FzS and continued medication for a further week prevented colonization of the caecum, liver and spleen . However, cessation of dosing at the time of challenge with salmonella resulted in colonization . Chloramphenicol and tetracycline at concentrations of 200 ppm were both independently capable of preventing colonization by salmonella . Sulphadiazine initially reduced colonization but failed to eliminate the infection . Only when furazolidone was combined with chloramphenicol or when sulphadiazine was combined with trimethoprim, and the combined drugs were administered concurrently with the challenge, was colonization prevented. J Basic Microbiol, 2003, 43(4), 328 - 36 Multiplex PCR for the direct detection of Salmonella enterica from chicken, lamb and beef food products; Malkawi HI et al.; Three sets of known Salmonella enterica-specific primers were used collectively, for the first time, to evaluate the use of multiplex polymerase chain reaction (m-PCR) as a diagnostic tool to detect Salmonella enterica in naturally contaminated meat and poultry products . For this purpose a total of 300 samples representing the most frequently used fresh and frozen meat (beef and lamb) and poultry (chicken) products (whole, cut, ground, and processed) were collected from eight locations within Irbid city (Jordan) . After an enrichment step, DNA was extracted directly from each food sample and amplified using six Salmonella enterica specific primers . Samples were also analyzed using conventional microbiological methods for the confirmation of Salmonella enterica presence . Out of 300 samples, 93 samples were positive by m-PCR . On the other hand, 67 samples were positive by conventional microbiological methods and only 26 samples were positive by m-PCR alone.The primer pairs, used here, proved to be highly specific for Salmonella enterica . Using m-PCR, Salmonella enterica detection could be achieved easily and the results had been confirmed effortlessly within a short period of time (24-36 hours) compared to 3-8 days for the conventional microbiological methods . Our findings emphasize the value of using a combination of specific primer pairs instead of a single primer pair in the detection process to minimize any chance for any inherent experimental or natural error. J Food Prot, 2003 Jul, 66(7), 1253 - 9 Industry practices and compliance with U.S . Food and Drug Administration guidelines among California sprout firms; Thomas JL et al.; Since 1995, raw vegetable sprouts have been implicated as the vehicle of infection in 15 foodborne outbreaks involving Salmonella and 2 foodborne outbreaks involving Escherichia coli O157:H7 . To reduce the numbers of sprout-related outbreaks, the U.S . Food and Drug Administration (FDA) published Guidance for Industry: Reducing Microbial Food Safety Hazards for Sprouting Seeds in 1999 . Between October 2000 and April 2001, 61.5% (16 of 26) of the known commercial sprout firms in California were enrolled in a survey to evaluate the industry practices of California sprouting operations and to determine compliance with FDA guidelines . A standardized questionnaire was used to collect data on firm demographics and seed disinfection practices . Additionally, free chlorine levels in seed disinfection solutions were measured, and 48-h spent irrigation water samples were collected from each firm . The irrigation water was screened for Salmonella and E . coli O157:H7 with FDA-recommended test kits . Free chlorine levels in the treatment solutions ranged from 50 to 35,000 mg/liter (ppm), with a median of 14,000 mg/liter (ppm) . Free chlorine levels were higher for firms producing alfalfa sprouts than for those producing only mung bean or soybean sprouts (P=0.03) . Levels of free chlorine tended to be higher for firms using a calcium hypochlorite treatment solution than for firms using a sodium hypochlorite treatment solution (P=0.067) . All 32 irrigation water samples screened for Salmonella tested negative . Of the irrigation water samples tested for E . coil O157:H7, 75% (24 of 32) tested negative, and 25% (8 of 32) tested presumptive positive . The eight presumptive positive samples were found to be negative after further testing . These results indicate that producers of alfalfa sprouts are generally achieving the FDA-recommended calcium hypochlorite level of 20,000 mg/liter (ppm), whereas mung bean sprout producers are not. J Food Prot, 2003 Jul, 66(7), 1158 - 65 Reduction of Escherichia coli O157:H7 and Salmonella on laboratory-inoculated alfalfa seed with commercial citrus-related products; Fett WF et al.; Alfalfa sprouts contaminated with the bacterial pathogens Escherichia coli O157:H7 and Salmonella have been the source of numerous outbreaks of foodborne illness in the United States and in other countries . The seed used for sprouting appears to be the primary source of these pathogens . The aim of this study was to determine whether the efficacy of commercial citrus-related products for sanitizing sprouting seed is similar to that of high levels of chlorine . Five products (Citrex, Pangermex, Citricidal, Citrobio, and Environne) were tested at concentrations of up to 20,000 ppm in sterile tap water and compared with buffered chlorine (at 16,000 ppm) . Alfalfa seeds were inoculated with four-strain cocktails of Salmonella and E . coli O157:H7 to give final initial concentrations of ca . 9.0 and 7.0 CFU/g, respectively . Treatments (10 min) with Citrex, Pangermex, and Citricidal at 20,000 ppm and chlorine at 16,000 ppm produced similar log reductions for alfalfa seed inoculated with four-strain cocktails of E . coli O157:H7 and Salmonella (3.42 to 3.46 log CFU/g and 3.56 to 3.74 log CFU/g, respectively), and all four treatments were significantly (P<0.05) more effective than the control treatment (a buffer wash) . Citrobio at 20,000 ppm was as effective as the other three products and chlorine against Salmonella but not against E . coli O157:H7 . Environne was not more effective (producing reductions of 2.2 to 2.9 log CFU/g) than the control treatment (which produced reductions of 2.1 to 2.3 log CFU/g) against either pathogen . None of the treatments reduced seed germination . In vitro assays, as well as transmission electron microscopy, confirmed the antibacterial nature of the products that were effective against the two pathogens and indicated that they were bactericidal . When used at 20,000 ppm, the effective citrus-related products may be viable alternatives to chlorine for the sanitization of sprouting seed pending regulatory approval. J Food Prot, 2003 Jul, 66(7), 1154 - 7 Prevalence and antibiotic susceptibility of Salmonella isolated from foods in Korea from 1993 to 2001; Chung YH et al.; This study determined the prevalence of Salmonella in foods widely consumed in Korea from 1993 to 2001, along with antimicrobial susceptibility profiles of Salmonella isolates from these foods for 11 antibiotics . Overall, 41 Salmonella isolates, representing 15 serotypes, were obtained from 2.2% (29 of 1,334) of the samples examined, and most of the Salmonella isolates were recovered from broiler carcasses . The most common serotypes were Salmonella Enteritidis (29.3%), Salmonella Virginia (14.6%), and Salmonella Haart (12.2%) . All isolates were screened for antibiotic resistance; 14.6% of the isolates were susceptible to all of the antibiotics, 4.9% were resistant to one antimicrobial agent, 14.6% were resistant to two antimicrobial agents, 22.0% were resistant to three antimicrobial agents, 39.0% were resistant to four antimicrobial agents, and 4.9% were resistant to five antimicrobial agents . Most of the isolates showed resistance or intermediate resistance to streptomycin, ampicillin, carbenicillin, and/or tetracycline. J Food Prot, 2003 Jul, 66(7), 1139 - 45 Evaluation of the safety assurance level for Salmonella spp . throughout the food production chain in Switzerland; Sauli I et al.; In Switzerland . the safeguarding of food is the responsibility of industry, organizations, and governmental authorities . The dispersion of the tasks and the diversity of implemented safety measures among involved stakeholders do not allow a general overview of the national safety assurance level provided . A comprehensive evaluation of the level of safety assurance provided for foodborne pathogens such as Salmonella spp . is therefore lacking, and the prevalence of Salmonella spp . at various points in the food production chain is not known . The objectives of this study were to (i) collect data on safety measures implemented throughout the food production chain in Switzerland regarding Salmonella spp.; (ii) evaluate the safety assurance level for Salmonella spp . at each step of the production chain for chicken meat, pork, beef, and milk and dairy products (bovine origin); and (iii) gather data on the prevalence of the pathogen at each step . Data on implemented safety assurance measures for Salmonella spp . were gathered from the various stakeholders in the food production chain . The data were analyzed by a semiquantitative method that considered the quality and relevance of the implemented safety measures for Salmonella spp . The safety assurance level for Salmonella spp . was evaluated from "no safety assurance" to "very good safety assurance." Available results of testing for Salmonella spp . from 1998 to 2000 were used for calculating the prevalence of the pathogen throughout the food production chain . The results showed a varying safety assurance level for Salmonella spp . throughout the food production chain . Strengths (e.g., feed production for chickens) and weaknesses (e.g., pork production) were observed . These results serve as a basis for a rational optimization of the system. J Food Prot, 2003 Jul, 66(7), 1115 - 25 Tolerance to stress and ability of acid-adapted and non-acid-adapted Salmonella enterica serovar Typhimurium DT104 to invade and survive in mammalian cells in vitro; Fratamico PM; The ability of acid-adapted (AA) and non-acid-adapted (NA) Salmonella enterica serovar Typhimurium definitive type 104 (DT104) strains to invade and multiply in mammalian cells in vitro and to survive stress conditions was examined . DT104 and non-DT104 strains were grown in tryptic soy broth without glucose (NA) or in tryptic soy broth containing 1% glucose (AA) for 18 h at 37 degrees C . The invasiveness of DT104 strains in J774A.1 macrophage and Int407 intestinal cell lines was not more extensive than that of non-DT104 strains . In most cases, AA bacteria were less invasive than NA bacteria in both cell lines . Confocal microscopy showed that both DT104 and non-DT104 strains replicated in the two cell lines . In related studies, the survival levels of three strains of AA and NA DT104 and a non-DT104 (LT2) strain in 150 and 15 mM H2O2, 170 and 43 mM acetic acid, 2.6 M NaCl, 2.6 M NaCl containing 170 mM acetic acid, synthetic gastric fluid (SGF) at pH 2 and pH 3, and apple cider were compared . For all four strains, acid adaptation did not result in increased survival in apple cider . After 15 days of storage at 4 degrees C, reductions ranged from 1.96 to 4.1 log10 CFU/ml for AA bacteria and from 0.48 to 1.34 log10 CFU/ml for NA bacteria from a starting level of ca . 7.00 log10 CFU/ml of cider . Neither AA nor NA DT104 strains were more resistant to NaCl, acetic acid, H2O2, or SGF solutions than non-DT104 strain LT2 . The level of AA bacteria was not appreciably reduced after exposure to SGF; however, the level of NA bacteria decreased to nondetectable levels in SGF at pH 2 within 3 h of exposure . These results indicate that the DT104 strains examined were not more invasive, nor did they display increased survival in mammalian cells or increased resistance to food environment stresses compared with non-DT104 strains . However, acid adaptation resulted in increased resistance to a low-pH gastric environment for all strains tested . These data indicate that DT104 strains are likely not more virulent or resistant to stresses relevant to foods than are non-DT104 Salmonella and that procedures used to inactivate or inhibit the growth of Salmonella in foods are likely adequate for DT104 strains. Verh K Acad Geneeskd Belg, 2003, 65(3), 189 - 202 {Fertility and sterility in domestic animals}; de Kruif A; For most of the domestic animals the fertility rate is generally very good . If a uniparous female animal is served during the oestrus period by a male with good sperm quality, the pregnancy rate is 60 to 70% . Among the animals that deliver more than one offspring, in many cases the pregnancy rate even reaches more than 90% . Nevertheless, veterinarians are very frequently consulted for fertility problems in individual animals or in cattle or swine herds . The main causes of subfertility are: Insufficient sperm quality An inseminator with insufficient professional knowledge . The majority of the cows, horses and pigs are inseminated artificially (AI) . The insemination is not always carried out by experts . Not the right time for insemination . It especially occurs in animals with weak oestrus symptoms . Venereal infections . In earlier days these infections would occur very often . Due to the application of AI, most of these infections have been eradicated . The malfunctioning of the female genital system, which can be caused by various factors, such as cystic ovarian follicles, endometritis and anatomic abnormalities . The research on reproduction which has been going on during the past ten years in the department of Obstetrics, Reproduction and Herd Health, has been mainly concerned with: Embryo transplantation, in vitro fertilisation and ovum pick up The evaluation of sperm quality Improved freezing methods for both sperm and embryo's The development of new insemination techniques The composition of a new diluent for fresh sperm Cystic ovarian follicles in cows Subfertility in different species of animals Next to the above mentioned study fields, the department is also involved in the research into swine fever, respiratory diseases in pigs, antibiotic resistance in pigs and cattle, mastitis and metabolic problems in cattle and salmonella infections in pigs. Eur J Biochem, 2003 Aug, 270(15), 3271 - 9 Physicochemical characterization and biological activity of a glycoglycerolipid from Mycoplasma fermentans; Brandenburg K et al.; We report a comprehensive physicochemical characterization of a glycoglycerolipid from Mycoplasma fermentans, MfGl-II, in relation to its bioactivity and compared this with the respective behaviors of phosphatidylcholine (PC) and a bacterial glycolipid, lipopolysaccharide (LPS) from deep rough mutant Salmonella minnesota strain R595 . The beta left arrow over right arrow alpha gel-to-liquid crystalline phase transition behavior of the hydrocarbon chains with Tc = 30 degrees C for MfGl-II as well as for LPS exhibits high similarity between the two glycolipids . A lipopolysaccharide-binding protein (LBP)-mediated incorporation into negatively charged liposomes is observed for both glycolipids . The determination of the supramolecular aggregate structure confirms the existence of a mixed unilamellar/cubic structure for MfGl-II, similar to that observed for the lipid A moiety of LPS . The biological data clearly show that MfGl-II is able to induce cytokines such as tumor necrosis factor-alpha (TNF-alpha) in human mononuclear cells, although to a significantly lower degree than LPS . In contrast, in the Limulus amebocyte lysate test, MfGl-II is completely inactive, and in the CHO reporter cell line it does not indicate any reactivity with the Toll-like receptors TLR-2 and -4, in contrast to control lipopeptides and LPS . These data confirm the applicability of our conformational concept of endotoxicity to nonlipid A structures: an amphiphilic molecule with a nonlamellar cubic aggregate structure corresponding to a conical conformation of the single molecules and a sufficiently high negative charge density in the backbone. Int J Med Microbiol, 2003 Jun, 293(2-3), 219 - 23 Molecular epidemiology of ampicillin-resistant clinical isolates of Salmonella enterica serovar Typhimurium; Biendo M et al.; Thirty-nine multiresistant Salmonella enterica serovar Typhimurium (S . Typhimurium) isolates were obtained from 33 children and 6 adults hospitalized from 1996 to 1999 in the University Hospital of Amiens (France) . S . Typhimurium was cultured from stools (n=36), blood samples (n=2) and peritoneal fluid (n=1) . These isolates were characterized by biotyping, antibiotic susceptibility test, RAPD-PCR, and PFGE typing . Emergence of pentaresistant S . Typhimurium isolates (phenotype ACSSuTe) was observed, and five of them were resistant to nalidixic acid and of intermediate susceptibility to pefloxacin . Genotypic analysis of both RAPD and PFGE results showed that there were 7 different patterns . Thirty-three isolates gave an identical pattern (AI) and were considered as epidemic isolates; the six remaining patterns (each containing one isolate) corresponded to sporadic cases . Antibiotic susceptibility patterns, RAPD and PFGE patterns subdivided the 39 isolates into 9 clonally related groups . One of them (pattern AI and R-pattern a) was implicated in 74% of the cases. J Bacteriol, 2003 Aug, 185(15), 4508 - 18 Lateral flagellar gene system of Vibrio parahaemolyticus; Stewart BJ et al.; Vibrio parahaemolyticus possesses dual flagellar systems adapted for movement under different circumstances . A single polar flagellum propels the bacterium in liquid (i.e., swimming) with a motor that is powered by the sodium motive force . Multiple proton-driven lateral flagella enable translocation over surfaces (i.e., swarming) . The polar flagellum is produced continuously, while production of lateral flagella is induced when the organism is grown on surfaces . This work describes the isolation of mutants with insertions in the structural and regulatory laf genes . A Tn5-based lux transcriptional reporter transposon was constructed and used for mutagenesis and subsequent transcriptional analysis of the laf regulon . Twenty-nine independent insertions were distributed within 16 laf genes . DNA sequence analysis identified 38 laf genes in two loci . Among the mutants isolated, 11 contained surface-induced lux fusions . A hierarchy of laf gene expression was established following characterization of the laf::lux transcriptional fusion strains and by mutational and primer extension analyses of the laf regulon . The laf system is like many enteric systems in that it is a proton-driven, peritrichous flagellar system; however, laf regulation was different from the Salmonella-Escherichia coli paradigm . There is no apparent flhDC counterpart that encodes master regulators known to control flagellar biosynthesis and swarming in many enteric bacteria . A potential sigma(54)-dependent regulator, LafK, was demonstrated to control expression of early genes, and a lateral-specific sigma(28) factor controls late flagellar gene expression . Another notable feature was the discovery of a gene encoding a MotY-like product, which previously had been associated only with the architecture of sodium-type polar flagellar motors. Chemosphere, 2003 Sep, 52(9), 1641 - 6 The mutagenic potentials of tap water samples in Shanghai; Shen L et al.; The tap water samples were collected from the users' ends in several areas of Shanghai, which is located in Taihu Lake basin, Eastern China . Source water samples were also collected from two municipal source water facilities at the same time . Samples were assayed by three different short-term mutagenicity test systems: Salmonella/microsome assay (Ames test), the Arabinose resistance test (Ara test) and the SOS/umu test . The data showed that two source water samples did not display direct mutagenic potentials . Two tap water samples from city north, which were directly from Yangtze River, were also not mutagenic . Water samples from city south and middle which used source water originating from Taihu Lake were proved to be contaminated with mutagenic potentials by three different assay techniques . The boiled water displayed an even stronger mutagenic potential compared to its original tap water . The molecular mechanism of mutagenicity was associated with a reading frame-shifting potential . GC-MS analysis of tap water extracts from city middle and corresponding source water was compared . Qualitatively similar spectra were observed except for the peaks of three chlorinated aromatic hydrocarbon compounds, which existed only in the tap water . Since the water source has been polluted, raw water was heavily chlorinated in order to sterilize . More toxic compounds, including mutagens, might form during the multi-chlorination . Caution about the possibility of elevated cancer risks in the population that consumes heavily chlorinated water should be kept in mind . A cohort study in the residents of Shanghai is required. Indian J Med Res, 2003 Jan, 117, 10 - 2 An unusually high occurrence of Salmonella enterica serotype paratyphi A in patients with enteric fever; Tankhiwale SS et al.; BACKGROUND & OBJECTIVES: Salmonella enterica serotype Paratyphi A has been reported less frequently as a causative agent of enteric fever . Reports on the antimicrobial susceptibility of this pathogen are few and varied . An unusually high occurrence of S . Paratyphi A was noted in a tertiary care hospital at Nagpur, Maharashtra during April 2001-September 2002 . An effort was made to study the antimicrobial susceptibility pattern and phage types of the isolates . METHODS: Blood cultures of patients suspected to have enteric fever admitted to the Indira Gandhi Medical College and Hospital, Nagpur were processed by conventional methods . Antimicrobial susceptibility was tested by Kirby-Bauer disc diffusion method and the minimum inhibitory concentration (MIC) to chloramphenicol was determined . RESULTS: Eighteen (46.15%) of 39 Salmonella isolates were S . Paratyphi A and all were sensitive to ciprofloxacin and cephotaxime . Twelve (66.67%) strains were sensitive to ampicillin and 13 (72.22 %) to chloramphenicol . Two strains (11.11%) were resistant to three drugs (ampicillin, chloramphenicol and cotrimoxazole) simultaneously . The prevalent phage type in the local population was phage type I . INTERPRETATION & CONCLUSION: The high occurrence of S . Paratyphi A found in the present study indicated the emergence of this rare pathogen of enteric fever in the local population . Though some degree of resistance was encountered with ampicillin and chloramphenicol, all the isolates were sensitive to ciprofloxacin, currently a drug of choice for enteric fever . Multidrug resistance was rare. Curr Gastroenterol Rep, 2003 Aug, 5(4), 279 - 86 Current trends in typhoid Fever; Crum NF; Typhoid fever, a systemic infection caused by Salmonella enterica serotype typhi, remains an important worldwide cause of morbidity and mortality . Endemic cases in the United States are unusual, with most following foreign travel to the Indian subcontinent, Africa, Asia, or Latin America . The classic findings of typhoid fever include rose spots, relative bradycardia, and stepwise fevers, but unfortunately these signs are frequently absent . Gastrointestinal manifestations may include diffuse abdominal pain, bleeding, perforation, cholecystitis, and cholangitis . The diagnosis should be suspected after collection of the appropriate clinical and travel history with confirmation by blood or bone marrow culture . Novel methods are in development to establish the diagnosis when cultures are negative or unavailable . Multidrug resistance has increased worldwide, and decisions on antimicrobial therapy must take such resistance into account . The empiric treatment of choice is a fluoroquinolone drug; ceftriaxone and azithromycin are alternatives . Preventive strategies include good sanitation and food handling practices along with vaccination of selected groups. Mol Microbiol, 2003 Aug, 49(3), 685 - 704 Salmonella type III effectors PipB and PipB2 are targeted to detergent-resistant microdomains on internal host cell membranes; Knodler LA et al.; The intracellular pathogen, Salmonella enterica, translocates type III effectors across its vacuolar membrane into host cells . Herein we describe a new Salmonella effector, PipB2, which has sequence similarity to another type III effector, PipB . In phagocytic cells, PipB2 localizes to the Salmonella-containing vacuole (SCV) and tubular extensions from the SCV, Salmonella-induced filaments (Sifs) . We used the specific targeting of PipB2 in macrophages to characterize Sifs in phagocytic cells for the first time . In epithelial cells, PipB2 has a unique localization pattern, localizing to SCVs and Sifs and additionally to vesicles at the periphery of infected cells . We further show that the N-terminal 225-amino-acid residues of PipB2 are sufficient for type III translocation and association with SCVs and Sifs, but not peripheral vesicles . Subcellular fractionation demonstrated that both PipB and PipB2 associate with host cell membranes and resist extraction by high salt, high pH and to a significant extent, non-ionic detergent . Furthermore, PipB and PipB2 are enriched in detergent-resistant microdomains (DRMs), also known as lipid rafts, present on membranes of SCVs and Sifs . The enrichment of Salmonella effectors in DRMs on these intracellular membranes probably permits specific interactions with host cell molecules that are concentrated in these signalling platforms. Cell Microbiol, 2003 Aug, 5(8), 501 - 11 Functions and effectors of the Salmonella pathogenicity island 2 type III secretion system; Waterman SR et al.; Salmonella enterica uses two functionally distinct type III secretion systems encoded on the pathogenicity islands SPI-1 and SPI-2 to transfer effector proteins into host cells . A major function of the SPI-1 secretion system is to enable bacterial invasion of epithelial cells and the principal role of SPI-2 is to facilitate the replication of intracellular bacteria within membrane-bound Salmonella-containing vacuoles (SCVs) . Studies of mutant bacteria defective for SPI-2-dependent secretion have revealed a variety of functions that can be attributed to this secretion system . These include an inhibition of various aspects of endocytic trafficking, an avoidance of NADPH oxidase-dependent killing, the induction of a delayed apoptosis-like host cell death, the control of SCV membrane dynamics, the assembly of a meshwork of F-actin around the SCV, an accumulation of cholesterol around the SCV and interference with the localization of inducible nitric oxide synthase to the SCV . Several effector proteins that are translocated across the vacuolar membrane in a SPI-2-dependent manner have now been identified . These are encoded both within and outside SPI-2 . The characteristics of these effectors, and their relationship to the physiological functions listed above, are the subject of this review . The emerging picture is of a multifunctional system, whose activities are explained in part by effectors that control interactions between the SCV and intracellular membrane compartments. Microb Pathog, 2003 Jul, 35(1), 43 - 8 Intracellular expression of the Salmonella plasmid virulence protein, SpvB, causes apoptotic cell death in eukaryotic cells; Kurita A et al.; The spv genes carried on the Salmonella virulence plasmid are commonly associated with severe systemic infection in experimental animals . The SpvB virulence-associated protein has been shown to ADP-ribosylate actin, and this enzymatic activity is essential for virulence in mice . Here, we present evidence that intracellular expression of SpvB protein induces not only disruption of actin filaments but also apoptotic cell death in eukaryotic cells. J Interferon Cytokine Res, 2003 Jun, 23(6), 319 - 27 Differential regulation of cytokine gene expression by avian heterophils during receptor-mediated phagocytosis of opsonized and nonopsonized Salmonella enteritidis; Kogut MH et al.; Internalization of pathogens by phagocytic cells triggers the innate immune response, which in turn regulates the acquired response . Phagocytes express a variety of receptors that are involved in recognition of pathogens, including (1) pattern recognition receptors (PRR), which recognize conserved motifs, (2) complement receptors (CR), which recognize complement-opsonized pathogens, and (3) Fc receptors (FcR), which recognize antibody-opsonized pathogens . Recognition of microbes is accompanied by the induction of multiple cell processes, including the production of proinflammatory and anti-inflammatory cytokines and chemokines . The objective of the present experiments was to use probes to known avian proinflammatory and anti-inflammatory cytokines and TaqMan technology to ascertain levels of cytokine gene expression in avian heterophils following receptor-mediated phagocytosis of either nonopsonized Salmonella enteritidis (SE), serum-opsonized SE, or IgG-opsonized SE . Expression of interleukin-6 (IL-6) and IL-8, considered in mammals as a proinflammatory chemokine, were upregulated following exposure to the nonopsonized or the opsonized SE . However, mRNA expression for IL-18 and interferon-gamma (IFN-gamma) was downregulated, and the expression of mRNA for the anti-inflammatory cytokine transforming growth factor-beta4 (TGF-beta 4) was upregulated . Interestingly, IL-1beta mRNA expression was significantly upregulated in heterophils that phagocytized either the nonopsonized SE via PRRs or IgG-opsonized SE via FcRs, whereas serum-opsonized SE phagocytized by CRs induced a downregulation of IL-1beta mRNA . These results suggest that signaling interactions initiated by receptor recognition of the microbe surface differentially regulate the induction of inflammatory cytokines in avian heterophils. J Virol, 2003 Aug, 77(15), 8249 - 55 Vaccination of mice with bacteria carrying a cloned herpesvirus genome reconstituted in vivo; Cicin-Sain L et al.; Bacterial delivery systems are gaining increasing interest as potential vaccination vectors to deliver either proteins or nucleic acids for gene expression in the recipient . Bacterial delivery systems for gene expression in vivo usually contain small multicopy plasmids . We have shown before that bacteria containing a herpesvirus bacterial artificial chromosome (BAC) can reconstitute the virus replication cycle after cocultivation with fibroblasts in vitro . In this study we addressed the question of whether bacteria containing a single plasmid with a complete viral genome can also reconstitute the viral replication process in vivo . We used a natural mouse pathogen, the murine cytomegalovirus (MCMV), whose genome has previously been cloned as a BAC in Escherichia coli . In this study, we tested a new application for BAC-cloned herpesvirus genomes . We show that the MCMV BAC can be stably maintained in certain strains of Salmonella enterica serovar Typhimurium as well and that both serovar Typhimurium and E . coli harboring the single-copy MCMV BAC can reconstitute a virus infection upon injection into mice . By this procedure, a productive virus infection is regenerated only in immunocompromised mice . Virus reconstitution in vivo causes elevated titers of specific anti-MCMV antibodies, protection against lethal MCMV challenge, and strong expression of additional genes introduced into the viral genome . Thus, the reconstitution of infectious virus from live attenuated bacteria presents a novel concept for multivalent virus vaccines launched from bacterial vectors. Arch Toxicol, 2003 Aug, 77(8), 477 - 84 Epub 2003 Jul 11. Effect of chrysin, a flavonoid compound, on the mutagenic activity of 2-amino-1-methyl-6-phenylimidazo{4,5- b}pyridine (PhIP) and benzo(a)pyrene (B(a)P) in bacterial and human hepatoma (HepG2) cells; Uhl M et al.; The aim of the present study was to investigate the antimutagenic effects of chrysin (CR), a flavonoid compound contained in many fruits, vegetables and honey . Earlier investigations with bacterial indicators showed that CR is one of the most potent antimutagens among the flavonoids . In the present study, we tested the compound in the Salmonella strains TA98 and TA100 in combination with benzo(a)pyrene (B(a)P) and 2-amino-1-methyl-6-phenylimidazo{4,5- b}pyridine (PhIP) and found pronounced protective activity over a concentration range between 10 and 100 microg/ml . The compound itself was devoid of mutagenic activity at all concentrations tested . In the micronucleus (MN) assay with human-derived HepG2 cells, a different pattern of activity was seen . CR itself caused significant induction of MN at dose levels > or =15 microg/ml; in combination experiments with B(a)P and PhIP, U-shaped dose-response curves were obtained and protection was found only in a narrow dose range (5 - 10 microg/ml) . Our findings indicate that the molecular mechanisms that account for the antimutagenic effects of CR in bacterial cells are different from those responsible for the effects in HepG2 cells . Earlier reports indicate that the antimutagenic effects of CR towards B(a)P and heterocyclic amines in bacterial indicators is due to inhibition of the activity of CYP1A . In contrast to this, we found a significant induction of CYP1A1 activity in HepG2 cells by CR . It can also be excluded that induction of GST, which is involved in the detoxification of polycyclic aromatic hydrocarbons accounts for the protective effects of CR against B(a)P since this enzyme was not significantly induced in the HepG2 cells . In the case of PhIP, induction of UDGPT and/or inhibition of sulfotransferase seen in human derived HepG2 cells after exposure to CR might play a role in the antimutagenic effects . In conclusion, our findings show that data from antimutagenicity studies with bacterial indicators cannot be extrapolated to HepG2 cells, and that CR causes genotoxic effects at higher dose levels in the latter cells . The implications of these observations for human chemoprevention strategies are discussed. Am J Reprod Immunol, 2003 May, 49(5), 297 - 307 MD-1 is a critical part of the mechanism causing Th1-cytokine-triggered murine fetal loss syndrome; Clark DA et al.; PROBLEM: Fetal loss syndrome (abortion/resorption) occurring on or after gestation day (gd) 9.5 in CBA/JxDBA/2 matings is dependent upon presence of TNF-alpha + IFN-gamma, which act by increasing expression of fg12 prothrombinase at the feto-maternal interface . The magnitude by which the abortion rate can be boosted by an injection of these cytokines on gd 7.5 depends on endogenous rate of loss, and appears to depend on microbial flora . Is cytokine-triggered abortion dependent upon a third signaling pathway that senses 'danger'? METHODS: Female CBA/J were mated to DBA/2 males and, C57B1/6 and C57B1/6 TNFalphaR1-/-Mak were mated to C57B1/6 control or TNFalphaR1-/-Mak males . LPS from Escherichia coli and Salmonella enteritidis, or the combination of TNF-alpha + IFN-gamma, was injected to stimulate abortions . The effect of anti-MD-1, which interferes with expression of CD14 and, hence, with signaling by LPS via the CD14-tlr4 complex, on TNF-alpha + IFN-gamma was tested . The presence of MD-1 in the uterus was evaluated by in situ hybridization, and effect of lipopolysaccharide (LPS) on mice lacking TNF-alphaR1 was tested . RESULTS: Anti-MD-1 completely abrogated TNF-alpha + IFN-gamma-induced abortions . MD-1 was expressed on trophoblast and in deciduas on gd 8.5 but LPS could not abort mice that lacked the type 1 receptor for TNF-alpha . Pregnant CBA/J females had classical resorptions (abortions) countable on gd 13.5-14.5 in response to LPS from E . coli or S . enteritidis, but C57B1/6 strain mice resorbed only in response to the latter, and E . coli LPS appeared to induce 'occult' losses . 'Occult' loss did not require TNF-alphaR1 . CONCLUSIONS: TNF-alpha + IFN-gamma could not induce murine abortions without co-presence of a 'danger' signal such as LPS acting via CD14 on toll receptors, and LPS could not act without co-signaling by TNF-alpha . Classical resorptions/abortions and 'occult' losses have a different mechanism in these models as reflected in type of endotoxin and requirement for TNF-alphaR1 signaling. Ugeskr Laeger, 2003 Jun 16, 165(25), 2577 - 8 {Salmonella infection complicated with acute renal failure}; Thogersen T et al.; Acute renal failure is a known complication to Salmonella gastroenteritis, and patients with chronic renal failure or impaired host defence are at increased risk . In the two presented cases there had been a few days of gastroenteritis before the hospitalisation, but the only symptoms at the admission were fatigue and dyspnoea . In both cases severe uraemia had developed and the patients and their physicians did not expect the episode of gastroenteritis to be the only etiology of acute renal failure . Both patients had normal renal histology and Salmonella was grown in their faeces . Subsequently, their renal function was normalised . In these patients dialysis and renal biopsies would have been unnecessary if the ability of even a moderate Salmonella infection to cause acute renal failure in a healthy subject had been realised and prompt rehydration had been initiated. J Infect Dis, 2003 Jul 15, 188(2), 202 - 8 Epub 2003 Jun 20. Epidemiology of bloodstream infections in a bacille Calmette-Guérin-vaccinated pediatric population in Malawi; Archibald LK et al.; The risk of Mycobacterium bovis bloodstream infection (BSI) in bacille Calmette-Guerin (BCG)-vaccinated children with human immunodeficiency virus (HIV) infection remains uncharacterized . We studied pediatric inpatients during the 1998 dry season in Malawi . After a detailed clinical evaluation, blood was drawn for culture and HIV testing . Of 229 children, 128 (56%) were male, 35 (15.3%) had BSI, and 30% of children aged >1.5 years (median, 2.7 years; range, 1 month-13 years) had HIV infection . The predominant pathogen was non-typhi Salmonella; neither Mycobacterium tuberculosis nor M . bovis was isolated . A diagnosis of malnutrition or sepsis was predictive of BSI; malnutrition alone correlated with both death and BSI . The bloodstream dissemination of M . tuberculosis and M . bovis BCG is uncommon in HIV-infected children vaccinated with BCG . Correlates such as malnutrition or sepsis can provide algorithms for identifying children who need observation or empirical antimicrobial therapy for BSI in the absence of appropriate laboratory testing. Clin Diagn Lab Immunol, 2003 Jul, 10(4), 670 - 9 Immune responses against Salmonella enterica serovar enteritidis infection in virally immunosuppressed chickens; Sheela RR et al.; To understand the role of immune mechanisms in protecting chickens from Salmonella infections, we examined the immune responses of Salmonella enterica serovar Enteritidis-infected chickens and the effect of chicken anemia virus (CAV), a T-cell-targeted virus, on S . enterica serovar Enteritidis-induced immune responses . One-day-old chicks were orally inoculated with S . enterica serovar Enteritidis with or without intramuscular injection of CAV . The bacterial infection, pathology, and immune responses of chickens were evaluated at 14, 28, and 56 days postinoculation . The infection increased the levels of S . enterica serovar Enteritidis-specific mucosal immunoglobulin A (IgA), the number of gut-associated T cells, and the titer of serum IgG specific for S . enterica serovar Enteritidis surface antigens . CAV infection depressed these immune responses, especially the mucosal immune responses, but did not increase the number of S . enterica serovar Enteritidis-infected cells in the intestine . The severity of pathological lesions appeared to be reciprocal to the level of immune responses, but the S . enterica serovar Enteritidis infection persisted . These results suggest that oral infection of S . enterica serovar Enteritidis in chickens induces both mucosal and systemic immune responses, which have a limited effect on the S . enterica serovar Enteritidis infection under conditions designed to mimic the field situation. Clin Diagn Lab Immunol, 2003 Jul, 10(4), 546 - 51 Adrenomedullin expression by gastric epithelial cells in response to infection; Allaker RP et al.; Many surface epithelial cells express adrenomedullin, a multifunctional peptide found in a wide number of body and cell systems . Recently, we and others have proposed that adrenomedullin has an important novel role in host defense . This peptide has many properties in common with other cationic antimicrobial peptides, including the human beta-defensins . Upon exposure of human gastric epithelial cells to viable cells of invasive or noninvasive strains of Helicobacter pylori, Escherichia coli, Salmonella enterica, or Streptococcus bovis, a significant increase in adrenomedullin secretion from these cells was demonstrated . Adrenomedullin gene expression was also increased in response to these microorganisms . Similar observations were noted when these cells were incubated with proinflammatory cytokines such as interleukin 1 alpha (IL-1 alpha), IL-6, tumor necrosis factor alpha and lipopolysaccharide . In cultured cells and an animal infection model, increased adrenomedullin peptide and gene expression was demonstrated when exposed to E . coli or Mycobacterium paratuberculosis, respectively . The data suggest there is a strong association between epithelial infection, inflammation, and adrenomedullin expression, which may have clinical relevance . The regulation of adrenomedullin expression may have therapeutic applications, such as improving or enhancing mucosal immunity. J Toxicol Environ Health A, 2003 Jul 25, 66(14), 1351 - 70 A comparison of two methods for fractionating complex mixtures in preparation for toxicity analysis; Cizmas L et al.; Chemical fractionation is a widely used tool for the chemical and toxicological characterization of complex mixtures . The objective of this research was to compare two frequently employed methods for fractionating a wood preserving waste (WPW) containing polycyclic aromatic hydrocarbons (PAHs) and pentachlorophenol (PCP) . The first method involved fractionation of the WPW into acid, base, and neutral fractions using a liquid-liquid acid/base/neutral (A/B/N) technique . The second method utilized alumina column chromatography to produce two fractions, A1 and A2 . Gas chromatography and mass spectrometry were used to quantify the chemical components in all fractions . The alumina method recovered 473,338 mg of total PAHs (tPAHs) per kilogram crude, while the A/B/N method yielded only 193,379 mg tPAHs/kg crude . In contrast, the A/B/N method recovered 13.7 mg PCP/kg crude while the alumina method yielded only 0.5 mg PCP/kg crude . Three bioassays were used to determine the toxicity of the crude extract and fractions . The neutral and A1 fractions contained the highest levels of tPAHs and benzo{a}pyrene (BaP) but failed to induce a positive response in the Salmonella/microsome assay with concentrations containing as much as 1800 and 2500 ng BaP/plate, respectively . In the Escherichia coli prophage induction assay, the acid fraction, which contained 472 mg PCP/kg fraction, induced a positive response, as did the base fraction, which did not contain detectable PCP . Significant reduction of gap junctional intercellular communication in hepatic cells occurred with the crude extract and acid, base, and neutral fractions . Overall, the results of these bioassays suggest that PCP genotoxicity was expressed in the acid fraction, whereas the cumulative genotoxicity of genotoxic PAHs appeared to be masked in the isolates from either fractionation method . The optimal fractionation method for a mixture of chlorophenols and PAHs may involve a refined hybrid method. Avian Pathol, 2003 Jun, 32(3), 225 - 37 Investigation of the distribution and control of Salmonella enterica serovar Enteritidis PT6 in layer breeding and egg production; Davies R et al.; Investigations were carried out in a layer breeder hatchery, a layer parent rearing farm, a layer parent farm and in a commercial pullet rearing and cage layer farm where Salmonella enterica serovar Enteritidis (S . Enteritidis) PT6 had become established . PT6 was initially found in focal points in the hatchery, such as hatcher ventilation ducting, tray wash areas and waste areas, but improved disinfection was followed by a rapid disappearance of contamination . Several different phage types of S . Enteritidis were found in the hatchery but most of these proved to be genotypically identical with PT6 . Investigations of contaminated layer breeder and rearing sites showed that the terminal disinfection programmes in place were effective in that no carry-over of infection occurred and the organism was rapidly eliminated from the organization . Infection with PT6 originating from chicks was investigated on a commercial pullet rearing farm . After several rounds of treatment with a fluoroquinolone antibiotic and competitive exclusion, no Salmonella was found in faeces or cloacal swabs but was present in dust in one of six houses . Sampling carried out after cleaning and disinfection confirmed clearance of the organism from the site, but infection did become established in a commercial laying house receiving the birds. J Infect, 2003 Jul, 47(1), 33 - 9 Comparison of phenotypic and genotypic characteristics of Salmonella bredeney associated with a poultry-related outbreak of gastroenteritis in Northern Ireland; Moore JE et al.; OBJECTIVES . To employ a combination of phenotypic and genotypic subspecies typing methods to aid in an epidemiological investigation of an outbreak of Salmonella bredeney involving ten persons . METHODS . Isolates were characterised by employing antibiogram typing, in addition to two genotyping techniques, including pulsed field gel electrophoresis (PFGE), random amplification of polymorphic DNA (RAPD) with two oligonucleotide primers . RESULTS . An outbreak of gastroenteritis associated with S . bredeney (serovar O:4 H:Lv 1,7) occurred in Belfast, Northern Ireland in November 1997 . In total, ten cases were confirmed, of which eight had consumed chicken cooked at local butchers and retailed through one of two local bakeries . One of the remaining cases was secondarily infected within her home and the final case had eaten a product other than cooked chicken from one of the bakeries . Food preparation practices were inadequate in one of the bakeries in question and record keeping and possibly cooking procedures were inadequate in the butchers . S . bredeney was isolated from an uncooked chicken supplied to the butchers confirming that improperly cooked chicken was most likely the source of the outbreak . All outbreak clinical isolates were indistinguishable from each other and were similar to the isolate obtained from the uncooked poultry demonstrating that these DNA-based methods were valuable in the molecular characterization of S . bredeney . CONCLUSIONS . This report emphasises the importance and maintenance of an effective hazard analysis critical control point (HACCP) approach to the processing and retailing of foodstuffs containing chicken in order to help eliminate hazards to public health. Toxicol In Vitro, 2003 Aug, 17(4), 403 - 12 Mutagenicity of bitumen and asphalt fumes; Heikkila PR et al.; The mutagenicity of asphalt fumes was tested with the Salmonella bioassays . The aim was to investigate if recycled additives modify the genotoxicity of emissions . Recycling of old asphalt is increasing, and we studied also the mutagenicity of emissions sampled during the re-use of asphalt . The composition of vapours and fumes were analysed by gas chromatography and by liquid chromatography . Bitumens containing coal fly ash (CFA) or waste plastics were heated to the paving temperatures in the laboratory . In the field, bitumen fumes were collected during paving of stone mastic asphalts (lime or CFA as a filler), remixing of stone mastic asphalt (lime or CFA as a filler), and of asphalt concrete . All the lab-generated vapour fractions were non-mutagenic . The particulate fractions were mutagenic with TA98 in the presence of the S9 activation . In addition, the lab-fumes from bitumen containing waste plastics were positive with both strains without S9 . Only particulate fractions sampled in the field were tested . They were mutagenic with and without metabolic activation with both strains . The mutagenic potency of the field samples was higher than that of the lab-generated fumes without S9, and the remixing fumes were more mutagenic than the normal paving and lab-generated fumes with S9 . The use of inorganic additive, CFA, did not change the mutagenicity of the fumes, whereas the organic additive, waste plastics, increased the mutagenicity of the laboratory emissions significantly. Toxicol Lett, 2003 Aug 28, 143(3), 291 - 9 Inhibition of aflatoxin B1 mutagenicity by cyclopiazonic acid in the presence of human liver preparations; Sabater Vilar M et al.; Co-occurrence of cyclopiazonic acid (CPA) and aflatoxin B(1) (AFB(1)) has been reported in different food commodities . Recently, we have shown that CPA reduces AFB(1) mutagenicity in the standard Salmonella-Microsome-Assay using rat S9-mix for metabolic activation (Environ . Toxicol . Pharmacol . 11 (2002) 207) . When using S9-mix prepared from individual liver fractions of human patients, CPA was found to be non-mutagenic, but exerted a significant reduction of the mutagenicity of AFB(1) . Moreover, CPA was shown to inhibit testosterone hydroxylation, but not methoxyresorufin dealkylation (MROD), in human S9 . Thus, the reduction of the AFB(1) mutagenicity by CPA may be attributed to the inhibitory effect of CPA on cytochrome P450 (CYP450) 3A4 activity . These findings might be of relevance to the epidemiology of food-borne mycotoxicosis as similar molar ratios to those investigated here have been reported in food commodities. MMWR Morb Mortal Wkly Rep, 2003 Jul 4, 52(26), 613 - 5 Multistate outbreak of Salmonella serotype typhimurium infections associated with drinking unpasteurized milk--Illinois, Indiana, Ohio, and Tennessee, 2002-2003; Effects of carrageenans on the binding et al.; Division of Microbiology, National Institute of Health Sciences, 1-18-1 Kamiyoga, Setagaya-ku, Tokyo 158-8501, JapanThe effects of carrageenans (CGNs) on the host defense mechanisms of macrophages against Salmonella infection were examined in vitro by using macrophage-like J774.1 cells . Iota-CGN reduced the Salmonella-binding and phagocytotic activities of J774.1 cells, but it increased the killing activity of the cells . Kappa-CGN increased the binding activity, but reduced the killing ability . CGNs would affect the host defense mechanisms by modulating the macrophage functions. J Clin Microbiol, 2003 Jul, 41(7), 3229 - 32 Comparison of Salmonella chromogenic medium with DCLS agar for isolation of Salmonella species from stool specimens; Cassar R et al.; Salmonella chromogenic medium (SCM; Oxoid, Basingstoke, United Kingdom), a new selective chromogenic medium, was compared to DCLS agar (Oxoid) for the detection and presumptive identification of Salmonella species from stool samples . This medium contains two chromogenic substrates, Magenta-cap (5-bromo-6-chloro-3-indolylcaprylate), which is hydrolyzed by Salmonella species to give magenta colonies, and X-Gal (5-bromo-4-chloro-3-indolyl-beta-D-galactopyranoside), which is incorporated to