Vibriosis

Trans R Soc Trop Med Hyg, 1995 Sep-Oct, 89(5), 542 - 5
Immunological response to Vibrio cholerae O1 infection and an oral cholera vaccine among Peruvians; Sanchez JL et al.; A 'double-blind', randomized, placebo controlled study of an oral inactivated whole cell plus recombinant B subunit (WC/rBS) cholera vaccine was conducted during February-March 1992 in Peru in 346 military recruits, 307 (89%) of whom received 2 oral doses of vaccine or Escherichia coli K12 placebo, 2 weeks apart . Paired serum samples were obtained from 155 (50%) of the recipients of 2 doses . An epidemic of cholera took place between doses . No difference in cholera attack rates was detected between vaccine and placebo recipients after one dose (8% versus 14%) . Seroconversion (4-fold or higher increase in vibriocidal antibody titres) was detected in 90% and 80% of vaccine and placebo recipients, respectively, with low pre-existing vibriocidal titres (< 0.01) . The anti-cholera toxin seroconversion rate among those with low pre-existing titres was higher in vaccinated subjects (97%) than in placebo recipients (68%) (P < 0.01) . Administration of 2 doses of WC/rBS vaccine concomitantly with natural V . cholerae O1 infection enhanced the serum anti-cholera toxin response . The immune response to the whole cell component of the vaccine was reduced by high pre-existing vibriocidal antibody titres.

Protein Sci, 1995 Sep, 4(9), 1931 - 3
Crystallization and preliminary X-ray crystallographic analysis of arylesterase from Vibrio mimicus; Musayev FN et al.; Single crystals of arylesterase (EC 3.1.1.2) from Vibrio mimicus have been obtained from ammonium sulfate as a precipitant at room temperature for 2 months . The present crystals diffract up to 2.2 A resolution and belong to monoclinic space group P2(1) . The cell dimensions are a = 55.65(1) A, b = 53.46(1) A, c = 65.79(1) A, and beta = 106.54(1) degrees . There are two molecules of molecular weight 22 kDa in an asymmetric unit with a solvent content of 43%.

Arch Microbiol, 1995 Sep, 164(3), 223 - 30
Cell-surface charge and cell-surface hydrophobicity of collagen-binding Aeromonas and Vibrio strains; Ascencio F et al.; Partitioning in aqueous polymer two-phase systems of polyethylene glycol and dextran was used to detect and compare cell-surface charge and cell-surface hydrophobicity of Aeromonas hydrophila, A . caviae, A . sobria, Vibrio cholerae, and V . anguillarum strains . These strains have cell-surface components that bound either native or thermally denatured type I collagen (i.e., a mixture of the alpha1+alpha2 chains) and gelatin immobilized on latex beads . Our goals were: (1) to compare the possible relationship between the cell-surface charge/hydrophobicity and binding to collagen and (2) to evaluate the influence of the culture media on the expression of surface properties . There was no apparent relationship between cell-surface charge, cell-surface hydrophobicity, and binding to collagen . The expression of surface properties was dependent on the culture media . There was no relationship between binding to immobilized collagen and binding to soluble 125I-labeled collagen . Particle-agglutination reactivity differed when using various collagen-coated microbead preparations . There were general differences in the particle-agglutination reactivity when collagen-coated latex beads were prepared using different coating procedures . The negative charge and hydrophobicity of the various collagen-coated microbead preparations were also studied by partitioning in the two-phase system of polyethylene glycol and dextran . Under these conditions, the alpha1+alpha2 collagen-chain mixture covalently immobilized on carboxy-modified latex beads was less hydrophobic and negatively charged than gelatin and native collagen immobilized on the same kind of latex beads . For latex beads passively coated with collagen preparations, the alpha1+alpha2 collagen-chain mixture was more hydrophobic than gelatin and native collagen . We suggest that for screening collagen-binding among Vibrio and Aeromonas strains, a reliable and sensitive particle-agglutination assay should consider the collagen preparation and the coating procedure for the immobilization of collagen onto the latex beads . In this regard, carboxy-modified latex beads coated with an alpha1+alpha2 collagen-chain mixture gave the best results.

J Bacteriol, 1995 Sep, 177(17), 5158 - 60
Isolation of the polar and lateral flagellum-defective mutants in Vibrio alginolyticus and identification of their flagellar driving energy sources; Kawagishi I et al.; Vibrio alginolyticus has two types of flagella (polar and lateral) in one cell . We isolated mutants with only a polar flagellum (Pof+ Laf-) or only lateral flagella (Pof- Laf+) . Using these mutants, we demonstrated that the energy sources of the lateral and polar flagellar motors in V . alginolyticus are H+ and Na+ motive forces, respectively, as in the related species V . parahaemolyticus.

J Infect Dis, 1995 Sep, 172(3), 883 - 6
Initial clinical studies of CVD 112 Vibrio cholerae O139 live oral vaccine: safety and efficacy against experimental challenge; Tacket CO et al.; Since October 1992, epidemics of cholera associated with Vibrio cholerae O group 139 have occurred in India, Bangladesh, and much of the rest of Asia . A volunteer model was used to determine the safety, immunogenicity, and efficacy of an attenuated delta ctxA delta zot delta ace delta cep V . cholerae O139 vaccine strain, designated CVD 112 . Six volunteers received 10(6) cfu and 6 received 10(8) cfu of CVD 112 . No subject who received the 10(6) dose had diarrhea or other severe symptoms after vaccination; 3 vaccinees developed mild diarrhea (mean stool volume, 648 mL) after receiving the higher dose . Five weeks after vaccination, 8 vaccinees and 15 unvaccinated control subjects underwent challenge with 10(6) cfu of wild type V . cholerae O139 AI1837 . One vaccinee (13%) and 12 control subjects (80%) developed diarrhea after challenge (P = .003) . The short-term protective efficacy conferred by vaccine strain CVD 112 was 84% and was remarkably similar to that conferred by primary wild type clinical infection (80%).

J Med Microbiol, 1995 Sep, 43(3), 216 - 20
Prevalence of Vibrio cholerae with heat-stable enterotoxin (NAG-ST) and cholera toxin genes; restriction fragment length polymorphisms of NAG-ST genes among V . cholerae O serogroups from a major shrimp production area in Thailand; Dalsgaard A et al.; A total of 148 Vibrio cholerae isolates from a major shrimp production area in Southern Thailand were examined by colony hybridisation for genes encoding heat-stable enterotoxin (NAG-ST) and cholera toxin (CT) . Only non-O1 V . cholerae strains were found to harbour NAG-ST (14 of 146) whereas no strains hybridised with the CT probe . NAG-ST-positive V . cholerae non-O1 strains were isolated from shrimp farms situated close to urban areas . Five different O serogroups were found among NAG-ST positive non-O1 strains . Southern blot and restriction endonuclease analysis of NAG-ST-positive strains revealed a high degree of genetic divergence . A total of seven classes of enterotoxin gene patterns were found with HindIII and EcoRI restriction endonucleases . Enterotoxin gene patterns correlated with O-antigen expression in 84% of isolates tested . In combination with other molecular techniques Southern blot analysis with an NAG-ST oligonucleotide probe could be useful for studying the molecular epidemiology of V . cholerae non-O1 strains.

Infect Immun, 1995 Sep, 63(9), 3726 - 8
Immunogenicity in Peruvian volunteers of a booster dose of oral cholera vaccine consisting of whole cells plus recombinant B subunit; Begue RE et al.; Forty-nine subjects received two doses of oral cholera vaccine consisting of whole cells plus recombinant B subunit; this was followed by a booster dose one year later . After the primary series, a significant (greater than twofold) increase in the levels of vibriocidal, anti-cholera toxin immunoglobulin G and anti-cholera toxin immunoglobulin A antibodies occurred in 54, 88, and 81% of the subjects, respectively . Within one year, titers decreased to levels close to baseline . A booster dose then induced rises to those which occurred after the initial vaccination . The results suggest that 1-year booster doses may be necessary to maintain immunity against cholera in Latin America.

J Appl Bacteriol, 1995 Sep, 79(3), 264 - 73
Multiplex polymerase chain reaction to detect toxigenic Vibrio cholerae and to biotype Vibrio cholerae O1; Shangkuan YH et al.; A multiplex polymerase chain reaction (PCR) was developed to identify cholera toxin-producing Vibrio cholerae and to biotype V . cholerae O1 . Enterotoxin-producing V . cholerae strains were identified with a primer pair that amplified a fragment of the ctxA2-B gene . Vibrio cholerae O1 strains were simultaneously differentiated into biotypes with three primers specified for the hylA gene in the same reaction . The hlyA amplicon in the multiplex PCR serves as an internal control when testing toxin-producing strains, as hlyA gene sequences exist in all tested V . cholerae strains . Enrichment of V . cholerae present on oysters for 6 h in alkaline peptone water before detection by a nested PCR with internal primers for ctxA2-B gene yielded a detection limit lower than 3 colony-forming units (cfu) per gram of food.

Appl Environ Microbiol, 1995 Sep, 61(9), 3476 - 8
Nested PCR method for rapid and sensitive detection of Vibrio vulnificus in fish, sediments, and water; Arias CR et al.; A nested PCR for the detection of Vibrio vulnificus in fish farms was developed as an alternative to cultural methods by using universal primers flanking the V . vulnificus-specific sequences directed against 23S rRNA genes . This specific assay detected 10 fg of DNA or 12 to 120 cells in artificially inoculated samples without enrichment and within 24 h.

Appl Environ Microbiol, 1995 Sep, 61(9), 3468 - 70
Reexamination of tetrodotoxin production by bacteria; Matsumura K; Vibrio alginolyticus has been reported as a good producer of tetrodotoxin (TTX), but the toxin extracted from this bacterium did not react to the monoclonal antibody against TTX . Surprisingly, chromatographic analyses detected high TTX peaks for polypeptone and yeast extracts used as medium materials, which were, as expected, all negative by the mouse bioassay . These results may require us to revise the bacterial production of TTX.

Appl Environ Microbiol, 1995 Sep, 61(9), 3436 - 42
Application of antisera raised against sulfate-reducing bacteria for indirect immunofluorescent detection of immunoreactive bacteria in sediment from the German Baltic Sea; Lillebaek R; Polyclonal rabbit antisera raised against sulfate-reducing bacteria (SRB) could detect several distinct populations of bacteria in sediment from the German Baltic Sea . The depth distribution of immunoreactive bacteria was determined by an indirect immunofluorescence filter method . Anti-Desulfovibrio desulfuricans DSM 1926 serum showed maximum bacterial numbers at a depth of 18 cm, with a concentration of 60 x 10(6) cells cm-3 . With anti-Desulfovibrio baculatus DSM 2555 serum, counts were highest at the same depth, approaching 0.7 x 10(6) cells cm-3 . Other significantly smaller populations were observed . Anti-SRBStrain 1 (lactate,vibrio) maxima were at 0 to 4 cm and at 17 to 18 cm . Anti-SRBStrain 2 (lactate,vibrio) serum showed several local maxima . Anti-SRBStrain 3 (lactate,oval) serum detected one single peak at a depth of 10 to 12 cm . Also determined were rates of sulfate reduction, total bacterial counts by acridine orange staining, and the viable counts by dilution series on anaerobic lactate medium . The total bacterial counts were highest (180 x 10(6) cells cm-3) at 3 to 4 cm and dropped to 24 x 10(6) cells cm-3 at 10 to 11 cm but showed additional local maxima reaching 140 x 10(6) cells cm-3 at a depth of 17 to 18 cm . Viable counts probable number) were above 10(5) CFU cm-3 at 0 to 3.6 cm but remained below 10(3) CFU at 7.2 to 18 cm . The sulfate reduction rate was maximal (107 nmol cm-3 day-1) at a depth of 1 to 2 cm, dropped to 10 nmol cm-3 day-1 at 12 to 13 cm, and reached 38 nmol cm-3 day-1 at 17 to 18 cm.

Appl Environ Microbiol, 1995 Sep, 61(9), 3379 - 84
Oxidative metabolism in nonculturable Helicobacter pylori and Vibrio vulnificus cells studied by substrate-enhanced tetrazolium reduction and digital image processing; Gribbon LT et al.; Growing and nonculturable cells of Helicobacter pylori and Vibrio vulnificus were studied for the capacity to reduce tetrazolium salts in order to elucidate the possible physiological basis for the proposed "viable but nonculturable" (VNC) state . Initial difficulties in obtaining consistent reduction of rho-iodonitrotetrazolium violet (INT) by H . pylori led us to develop a method for studying the effect of adding exogenous substrates on these reactions . The established procedure provided a profile of substrate enhancement of oxidative activity revealed by INT reduction which was related to both the identity and physiological state of the organism studied . Representation and interpretation of these enhancement profiles were facilitated by digital image processing . Nonculturable cells of H . pylori produced by carbon and nitrogen starvation in air lost all INT-reducing capacity in 24 h when stored at 37 degrees C, while 99% of those produced at 4 degrees C retained oxidative activity for at least 250 days when tested in the presence but not in the absence of succinate, alpha-ketoglutarate, or aspartate . Activity was detected at similar levels in cells with coccoid and spiral shapes . In contrast, only 1% of nonculturable cells of V . vulnificus, produced under conditions previously reported to induce the VNC state in this organism, retained intrinsic INT-reducing capacity; no substrate-enhanced activity occurred in the remainder of the population . Thus, there was no common pattern of oxidative activity indicative of a VNC state in both test organisms . Nonculturable cells of H . pylori can retain several different oxidative enzyme activities; whether these indicate viability or the persistence of cells as "bags of enzymes" remains to be established.

Eur J Biochem, 1995 Sep 1, 232(2), 391 - 6
Structure of the capsular polysaccharide of Vibrio cholerae O139 synonym Bengal containing D-galactose 4,6-cyclophosphate; Knirel YA et al.; The capsular polysaccharide (CPS) of Vibrio cholerae O139 synonym Bengal, which is thought to carry determinants of O-specificity, was isolated by phenol/water extraction followed by delipidation of the contaminating lipopolysaccharide at pH 4.2 and gel-permeation chromatography . The CPS contained D-galactose, 3,6-dideoxy-L-xylo-hexose (colitose, Col), 2-acetamido-2-deoxy-D-glucose, 2-acetamido-2,6-dideoxy-D-glucose (N-acetyl-D-quinovosamine, D-QuiNAc), D-galacturonic acid (D-GalA), and phosphate . The CPS was studied by NMR spectroscopy, methylation analysis, and selective degradations, including partial acid hydrolysis at pH 3.1 and dephosphorylation with aqueous 48% hydrofluoric acid, which both resulted in complete cleavage of Col . It was concluded that the CPS is built up of hexasaccharide repeating units containing inter alia D-galactose 4,6-cyclophosphate and having the following structure {structure: see text} These data basically confirm the structure of the V . cholerae CPS proposed on the basis of an NMR study {L . M . Preston et al . (1995) J . Bacteriol . 177, 835-838} and specify exactly the absolute configurations of the constituent monosaccharides and the position of the cyclic phosphate.

Infect Immun, 1995 Sep, 63(9), 3537 - 42
Cloning and expression of rfb genes from Vibrio anguillarum serotype O2 in Escherichia coli: evidence for cross-reactive epitopes; Amor PA et al.; Vibrio ordalii and Vibrio anguillarum O2 express lipopolysaccharide (LPS) O antigens containing both specific and cross-reactive epitopes . The localization of these epitopes on the O antigen is not known . We have cloned and expressed the rfb gene cluster for O-antigen synthesis from V . anguillarum O2 (rfbVaO2) in Escherichia coli . E . coli DH5 alpha containing the recombinant plasmid pAM86 expressed O antigens which reacted with polyclonal antisera to V . ordalii and to V . anguillarum O2 LPS and with monoclonal antibody (MAb) 7B4, which is specific for V . anguillarum O2 O antigens . The recombinant strains were also protected from bactericidal killing by normal fish serum . Surprisingly, the LPS expressed from the cloned rfbVaO2 genes also reacted with MAb A16, which is specific for V . ordalii O antigens . Western immunoblot analysis revealed that MAb 7B4 reacted with recombinant LPS bearing shorter O-antigen repeat units, while MAb A16 reacted with the longer O antigens . Similar results were obtained when pAM86 was transformed into E . coli CLM4, which has a deletion spanning the sbcB-rfb region, indicating that the changes in antigenic profiles of O antigens from the recombinant strains were not due to genes within the E . coli rfb cluster . These data suggest that the epitope recognized by the MAb A16 is expressed by V . anguillarum O2 strains but it is apparently not accessible to the antibody in the native O polysaccharide . Cloning of the rfbVaO2 gene cluster resulted in expression of a novel O antigen . The modification(s) which leads to the alterations in antigenic profile of these recombinant LPS remains to be determined.

Microbiology, 1995 Sep, 141 ( Pt 9), 2101 - 9
Induction of heat shock response in Vibrio cholerae; JeevanJyot et al.; General properties of the heat shock response in Vibrio cholerae were examined . Enhanced or de novo synthesis of 24 proteins was observed upon heat shock from 30 degrees C to 42 degrees C in cells labelled with {35S}methionine . A similar response could also be induced by a rise in temperature from 30 degrees C to 37 degrees C . Of these heat shock proteins, two were determined to be homologues of GroEL and DnaK, based upon their immunological cross-reactivity with antibodies raised against the Escherichia coli proteins . Three proteins, of molecular sizes 38, 44 and 48 kDa, which were undetectable in the 30 degrees C grown culture, appeared de novo after the heat shock . As in other prokaryotic systems thermal induction of many of the proteins was transient, but both DnaK and GroEL remained induced for at least 28 min after heat shock . DNA hybridization studies revealed that genes analogous not only to dnaK and groEL but also to dnaJ of E . coli exist in V . cholerae . Heat shock induced thermotolerance in V . cholerae but made the cells more sensitive to UV radiation . Unlike in E . coli, however, heat shock had no effect on the progeny virus yield in V . cholerae.

Public Health, 1995 Sep, 109(5), 389 - 95
Outbreak of Vibrio cholerae 01 in Hong Kong related to contaminated fish tank water; Kam KM et al.; An outbreak of 12 cholera cases, caused by Vibrio cholerae eltor inaba, occurred in Hong Kong during a three week period in June-July 1994 . Only adults of both sexes were affected . Epidemiological investigations showed linkage in all cases with consumption of seafood, including shellfish, mantis shrimps and crabs . Microbiological findings demonstrated that contaminated seawater in fish tanks used for keeping alive these seafoods is the most likely vehicle of transmission . Aggressive control measures, promptly instituted, included prohibition of use of contaminated typhoon shelter water in fish tanks, use of seawater with E . coli counts below 610 organisms/100 ml, and the banning of unlicensed food sampans in typhoon shelters . These measures, coupled with public announcements and an active health education campaign on food safety and personal hygiene, abruptly terminated the outbreak . Places which practise the use of seawater, from probable contaminated sources, to keep alive their seafood for human consumption should be alerted to the possibility of transmission of Vibrio cholerae through this route.

Nippon Rinsho, 1995 Sep, 53(9), 2296 - 300
{Detection of bacterial protein toxins by a bead-ELISA}; Kurazono H et al.; A highly sensitive bead-enzyme-linked immunosorbent assay to detect bacterial protein toxins was developed . Fab' of anti-toxin IgG was conjugated with horseradish peroxidase by the maleimide method and tetramethylbenzidine was used as a substrate . As the solid phase, a 6.5 mm diameter polystyrene bead was used and this was coated with the anti-toxin IgG . The sensitivities of the bead-ELISA for various bacterial protein toxins were as follows: less than 40 pg/ml for cholera enterotoxin (CT), less than 20 pg/ml for VT1 and less than 6 pg/ml for VT2 of enterohemorrhagic Escherichia coli . The bead ELISA was evaluated for direct detection of CT from stool specimens of patients with acute secretory diarrhea . Of the 75 stool samples examined, 59 yielded biochemically and serologically confirmed strains of Vibrio cholerae O1 . The bead ELISA was positive for CT in stool supernatants in 50 (84.7%) of the 59 samples from which V . cholerae O1 was isolated . In addition, the bead ELISA was positive for three stool specimens which were negative by culture . These data indicate that the bead ELISA is a sensitive and simple method for direct detection of CT in nonsterile stool samples.

J Mol Biol, 1995 Aug 25, 251(4), 550 - 62
The 2.4 A crystal structure of cholera toxin B subunit pentamer: choleragenoid; Zhang RG et al.; Cholera toxin, a heterohexameric AB5 enterotoxin released by Vibrio cholera, induces a profuse secretory diarrhea in susceptible hosts . Choleragenoid, the B subunit pentamer of cholera toxin, directs the enzymatic A subunit to its target by binding the GM1 gangliosides exposed on the luminal surface of intestinal epithelial cells . The crystal structure of choleragenoid has been independently solved and refined at 2.4 A resolution by combining single isomorphous replacement with non-crystallographic symmetry averaging . The structure of the B subunits, and their pentameric arrangement, closely resembles that reported for the intact holotoxin, choleragen, the heat-labile enterotoxin from Escherichia coli, and for a choleragenoid-GM1 pentasaccharide complex . In the absence of the A subunit the central cavity of the B pentamer is a highly solvated channel . The binding of choleragenoid to the A subunit or to its receptor pentasaccharide modestly affects the local stereochemistry without perceptibly altering the subunit interface.

J Mol Biol, 1995 Aug 25, 251(4), 471 - 6
Immunoglobulin mutant library genetically screened for folding stability exploiting bacterial signal transduction; Kolmar H et al.; A model repertoire of variants of immunoglobulin kappa variable domain REIv with different folding stabilities was generated by oligonucleotide-directed randomization of position 29, a key conserved residue of hypervariable loop 1 . Fused to ToxR', the membrane-anchored cytoplasmic domain of the Vibrio cholerae ToxR transcription activator, different members of the library induce different levels of transcription from the ctx promoter in Escherichia coli . Differences in transcription activation correlate positively with folding stabilities of the corresponding REIv domains . Since conformationally stabilized REIv derivatives elicit a dark red colony phenotype on EMB-lactose indicator plates, this procedure constitutes a genetic screen for immunoglobulin folding stability.

EMBO J, 1995 Aug 15, 14(16), 3895 - 904
Membrane insertion of the bacterial signal transduction protein ToxR and requirements of transcription activation studied by modular replacement of different protein substructures; Kolmar H et al.; The Vibrio cholerae protein ToxR is an integral membrane protein that acts as a transcription activator in response to environmental signals; it controls expression of toxin genes ctxA and ctxB, along with a variety of other genes related to pathogenicity . Here it is shown that: (i) ToxR has a modular architecture and that activation of transcription starting at the ctx promoter depends strictly on dimerization of the periplasmic ToxR domain; (ii) the transmembrane (TM) region of ToxR is sufficient as a topogenic signal but not for stable membrane anchoring of the protein; (iii) the TM region has no special function in signal transduction and (iv) a proline residue located within the TM region minimizes background transcription activation, most plausibly by reducing TM-TM interaction . Possible applications of ToxR as a technical tool for analysing protein-protein interactions between pairs of arbitrary TM domains are discussed.

Biochem Biophys Res Commun, 1995 Aug 15, 213(2), 475 - 83
Vibrio mimicus arylesterase has thioesterase and chymotrypsin-like activity; Chang RC et al.; A Vibrio mimicus serine arylesterase and an Escherichia coli thioesterase/serine protease share 49.4% amino acid identity . The arylesterase has thioesterase activity for benzoyl-CoA and chymotrypsin-like activity for N-carbobenzoxy-L-phenylalanine p-nitrophenyl ester (NBPNPE) . The gene encoding the V . mimicus enzyme is designated etpA . Substituting Ser31 of the V . mimicus enzyme with a glycine or an alanine altered its activity . In comparison with wild type enzyme, the S31A enzyme showed a 5-fold increase and 57% decrease in the catalytic efficiency for benzoyl-CoA and NBPNPE, respectively, and the S31G enzyme showed a 3.6-fold increase and 43% decrease in the catalytic efficiency for benzoyl-CoA and NBPNPE, respectively . For the two mutant enzymes an 8-fold decrease and a 6- to 7-fold increase in Km were seen for benzoyl-CoA and NBPNPE, respectively . The mutagenesis results prove that residue 31 plays an important role in the substrate-specificity.

FEMS Microbiol Lett, 1995 Aug 15, 131(1), 69 - 74
Characterization of phage phi O139, a Vibrio cholerae O139 temperate bacteriophage with cohesive DNA termini; Pajni S et al.; A temperate bacteriophage isolated from Vibrio cholerae O139, the new epidemic strain of cholera, was found to have a polyhedral head 65 nm in diameter and a rigid contractile tail 120 nm in length . The phage chromosome was a double-stranded DNA of 35 kb, with unique cohesive ends and had a G + C content of 58.8% . A restriction map of the phage DNA was constructed using the restriction endonucleases AvaI and BstEII . The phage, whose presence could be detected in nine out of 13 V . cholerae O139 isolates tested, was found to have identical chromosomal integration sites in all the strains . The phage attachment site (attP) was found to be located very close to one end of the genome.

FEBS Lett, 1995 Aug 7, 369(2-3), 173 - 6
The Na(+)-translocating NADH:ubiquinone oxidoreductase from the marine bacterium Vibrio alginolyticus contains FAD but not FMN; Pfenninger-Li XD et al.; The Na(+)-translocating NADH:ubiquinone oxidoreductase from Vibrio alginolyticus was extracted from the bacterial membranes and purified by ion exchange chromatographic procedures . The enzyme catalyzed NADH oxidation by suitable electron acceptors, e.g . menadione, and the Na+ and NADH-dependent reduction of ubiquinone-1 . Four dominant bands and a number of minor bands were visible on SDS-PAGE that could be part of the enzyme complex . Flavin analyses indicated the presence of FAD but no FMN in the purified enzyme . FAD but no FMN were also present in V . alginolyticus membranes . FAD is therefore a prosthetic group of the Na(+)-translocating NADH:ubiquinone oxidoreductase and FMN is not present in the enzyme . The FAD was copurified with the NADH dehydrogenase . The purified enzyme exhibited an absorption spectrum with a maximum at 450 nm that is typical for a flavoprotein . Upon incubation with NADH this absorption disappeared indicating reduction of the enzyme-bound FAD.

J Mol Biol, 1995 Aug 4, 251(1), 50 - 8
High-speed rotation and speed stability of the sodium-driven flagellar motor in Vibrio alginolyticus; Muramoto K et al.; The Na(+)-driven flagellar motor in Vibrio alginolyticus rotates very fast . Rotation of a single flagellum on a stuck cell was measured by laser darkfield microscopy with submillisecond temporal resolution . The rotation rate increased with increasing external concentration of NaCl, and reached 1000 r.p.s . at 300 mM NaCl . The Na+ influx through the motor should determine the rotation period (tau) and affect the speed stability . Fluctuation of the rotation period was analyzed at various rotation rates (from approximately 50 r.p.s . to approximately 1000 r.p.s.), which were changed by changing the external concentration of NaCl and the addition of a protonophore or a specific inhibitor . At high rotation rates (over 400 r.p.s.), the observed rotation was stable, and the standard deviation of tau (sigma tau) ranged from 7% to 16% of the average rotation period (< tau >) . At low rotation rates (under 100 r.p.s), the rotation period tended to fluctuate, and the distributions of tau were non-Gaussian . The value of sigma tau ranged from 10 to 30% of < tau > . However, the observed minimum value of sigma tau at various rotation rates was approximately equal to the calculated standard deviation due to the rotational diffusion of the flagellar filament . These results suggest that the torque was stably generated at various Na+ influxes through the motor . We observed large fluctuations that cannot be explained by rotational diffusion . We discuss the factors that induce the large fluctuation.

Mol Microbiol, 1995 Aug, 17(4), 747 - 56
Antisense RNA regulation of the fatB iron transport protein gene in Vibrio anguillarum; Waldbeser LS et al.; We have recently identified an antisense RNA (RNA alpha) that regulates the expression of the fatA iron transport gene encoding the outer membrane receptor for the iron-anguibactin complex . In this work, we demonstrate that RNA alpha also inhibits the expression of fatB, which encodes a 35 kDa iron transport protein and has domains homologous to other periplasmic transport proteins . The expression of fatA and fatB is repressed under iron-rich conditions, in which RNA alpha is induced . RNA alpha is homologous to two-thirds of the coding region of fatB . By cloning RNA alpha coding sequences immediately downstream of a tet promoter, we were able to obtain constitutive expression of the antisense RNA . The cloned region contains approximately 83% of the 650 nucleotide RNA alpha and is complementary to only 51% of the fatB mRNA but is still capable of causing a repression of the expression of the fatB gene . Our results in this work demonstrate that RNA alpha probably affects the stability of the fatB-specific mRNA.

Microb Pathog, 1995 Aug, 19(2), 65 - 72
Construction and characterization of recombinant Vibrio cholerae strains carrying heterologous genes encoding non-01 antigen or cholera enterotoxin; Smirnova NI et al.; In an attempt to study the effect of heterologous genes on the virulence of Vibrio cholerae 01 and non-01, rfb genes encoding biosynthesis of non-01 antigens were introduced by homologous recombination into the chromosome of V . cholerae 01 strain 569B (serotype Inaba, biotype classical) . Recombinant strains were obtained which were not agglutinated with the diagnostic cholera 01 antiserum and were not sensitive to the cholera diagnostic bacteriophage, but produced as much cholera toxin as 569B and were highly virulent in the infant rabbit intraintestinal injection model . These data indicate that the rfb genes from the studied V . cholerae non-01 did not alter the virulence phenotype of V . cholerae 01 . In contrast, cloned ctxAB genes from V . cholerae 01 encoding cholera toxin introduced into a non-pathogenic strain lead to efficient secretion of cholera toxin but to only low virulence in the infant rabbit model.

Infect Immun, 1995 Aug, 63(8), 3137 - 42
A conserved Aeromonas salmonicida porin provides protective immunity to rainbow trout; Lutwyche P et al.; A protein with an apparent M(r) of 28,000 was isolated from outer membrane preparations of Aeromonas salmonicida A440 . The protein was tested for the ability to form pores, using a planar lipid bilayer model membrane system . The protein appeared to be a monomer with a single-channel conductance in 1.0 M KCl of 1.96 nS and a cation/anion permeability ratio of 2.91 +/- 0.68 . These data show that the porin channel is comparable in size to OmpC and OmpF of Escherichia coli and is relatively nonselective, having some preference for cations over anions . The porin was purified by preparative sodium dodecyl sulfate-polyacrylamide gel electrophoresis, and a polyclonal antibody was raised . Immunoblot analysis showed that an immunologically cross-reactive protein was present in other Aeromonas strains but not in strains of Vibrio or Yersinia . The N-terminal amino acid sequence of the porin was determined and was found to show some homology to an Aeromonas hydrophila outer membrane protein . This is the second porin species of A . salmonicida to be described, and it differs from the other in subunit molecular weight, aggregation properties, peptidoglycan association, pore size, and antigenicity . Rainbow trout (Oncorhynchus mykiss) immunized intraperitoneally with the purified porin protein were significantly protected from experimental A . salmonicida challenge . This is the first report of successful vaccination against A . salmonicida with a purified outer membrane component.

Infect Immun, 1995 Aug, 63(8), 2906 - 11
Capsular polysaccharide-protein conjugate vaccines of carbotype 1 Vibrio vulnificus: construction, immunogenicity, and protective efficacy in a murine model; Devi SJ et al.; Vibrio vulnificus causes septicemia and wound infections in immunocompromised humans . The capsular polysaccharide of Vibrio vulnificus (VvPS) is critical for virulence . We synthesized conjugate vaccines of carbotype 1 VvPS under conditions and in formulations suitable for human use . Purified VvPS was conjugated to tetanus toxoid (TT) or to inactivated V . vulnificus cytolysin or elastase by two different schemes . All conjugates elicited elevated anticapsular immunoglobulin G (IgG) and IgM and antiprotein IgG responses in mice compared with saline placebo . The conjugates prepared through caboxyl activation of VvPS (VvPS-TTa, VvPS-cytolysin, and VvPS-elastase) were more immunogenic than the one prepared through hydroxyl activation (VvPS-TTb) . The protective efficacy of conjugated and unconjugated formulations of VvPS and that of protein carriers were evaluated in a mouse septicemia model . Eighty percent of mice actively immunized with VvPS-TTa vaccine survived challenge with carbotype 1 V . vulnificus, while VvPS-cytolysin and VvPS-elastase conjugates conferred 44 and 40% protection, respectively . Control mice immunized with VvPS, cytolysin, or elastase alone, or saline only, showed 70 to 100% mortality . VvPS-TTa vaccine is nontoxic, immunogenic, and protective in mice.

Gastroenterology, 1995 Aug, 109(2), 381 - 6
Calcium-dependent intestinal chloride secretion by Vibrio parahaemolyticus thermostable direct hemolysin in a rabbit model; Raimondi F et al.; BACKGROUND & AIMS: Vibrio parahaemolyticus is a major agent of seafood gastroenteritis that induces intestinal secretion in the rabbit through its thermostable direct hemolysin . The aim of this study was to characterize the enterotoxicity of purified hemolysin in vitro . METHODS: Rabbit ileum was mounted in Ussing chambers, and changes in potential difference and short-circuit current were monitored after addition of hemolysin . Intracellular calcium concentrations in the nontumoral rat crypt-derived cell line IEC-6 were measured using microspectrofluorometry . RESULTS: In Ussing chamber experiments, mucosal toxin addition up to 50 hemolytic units per milliliter induced a proportional increase of the electrical parameters in normal but not Cl(-)-free Ringer's solution . The response to the toxin was not additive to that of calcium ionophore A23187 and was eliminated by preloading the tissue with 1-2-bis(o-aminophenoxy)ethane N,N,N',N'-tetraacetic acid (BAPTA), a calcium buffer . In IEC-6 cells, a 10-fold increase in intracellular calcium level was found after addition of hemolysin . Such an increase was totally quenched by BAPTA . Finally, preincubation with trisialoganglioside GT1b, but not monosialoganglioside GM1, eliminated toxin-induced increases in potential difference and short-circuit current . CONCLUSIONS: These data support the hypothesis that the thermostable direct hemolysin induces intestinal chloride secretion using GT1b as a putative receptor and Ca2+ as a second messenger.

J Appl Bacteriol, 1995 Aug, 79(2), 135 - 40
The interaction of trimethoprim and quinolones against gram-negative fish pathogens; Inglis V et al.; The effect of combination of trimethoprim with other non-sulphonamide antibacterial agents, in particular oxolinic acid and nalidixic acid, was evaluated against Gram-negative fish pathogens . The species included Aeromonas salmonicida, Yersinia ruckeri, some Vibrio spp . and Escherichia coli as a reference . The extent of synergy found by other workers with these substances against human Gram-negative bacteria was not apparent here . Some positive interaction between trimethoprim and oxolinic acid was found with Aer . salmonicida, Y . ruckeri and E . coli and between trimethoprim and nalidixic acid with Y . ruckeri in double disc diffusion tests but was not supported by fractional inhibitory concentration indices . The combinations were not effective in preventing emergence of resistance in passage on a drug gradient . Trimethoprim-resistant isolates of Aer . salmonicida were inhibited by low levels of oxolinic acid but the converse did not apply.

Chem Phys Lipids, 1995 Aug 1, 77(1), 41 - 9
Aggregation properties of semisynthetic GD1a ganglioside (IV3Neu5AcII3Neu5AcGgOse4Cer) containing an acetyl group as acyl moiety; Brocca P et al.; GD1a ganglioside containing an acetyl group as acyl moiety, GD1a(acetyl), was synthesized from natural GD1a . The aggregative properties in aqueous solution of GD1a(acetyl) have been studied by static and dynamic laser light-scattering measurements . GD1a(acetyl) spontaneously aggregates as small micelles showing a hydrodynamic radius and molecular mass of 33 A and 96 kDa, respectively . Vibrio cholerae sialidase showed a very high activity on the micelles of GD1a(acetyl), compared to GD1a . This has been explained as a consequence of the high surface curvature of the the small micelles . High resolution proton NMR spectra were recorded from micelles of GD1a(acetyl) in deuterated water . The low overall correlation time of the GD1a(acetyl) micelles was calculated to be about 2 x 10(-8)s, a value one order of magnitude lower than that determined for natural GD1a.

J Clin Microbiol, 1995 Aug, 33(8), 2186 - 7
Evaluation of DNA probes for specific detection of Vibrio cholerae O139 Bengal; Nair GB et al.; Two DNA probes, 2R1 and 2R3, prepared from a region in the chromosome specific for the lipopolysaccharide O side chains of Vibrio cholerae O139 (M.K . Waldor and J.J . Mekalanos, Lancet 343:1366, 1994) were examined for their specificity and sensitivity . Both probes did not hybridize with any strain of V . cholerae belonging to serogroups other than O139 and to any of the other species examined belonging to the family Vibrionaceae . Among the 126 strains of V . cholerae O139 examined, probe 2R1 hybridized with 125 strains while probe 2R3 hybridized with all 126 strains . Both probes were found to be highly specific and sensitive and can be used for the specific identification of V . cholerae O139.

Microbiology, 1995 Aug, 141 ( Pt 8), 1977 - 83
Cholera toxin (CTX) genetic element in Vibrio cholerae O139; Bhadra RK et al.; PFGE analysis of the NotI- and SfiI-digested genome of Vibrio cholerae O139 strains isolated from different epidemic regions of India showed that all the strains are of clonal origin and the genome size is about 2.2 Mb . An analysis of the electrophoretic profiles of the genome of O139 strains, the RFLP of the cholera toxin (ctx) gene and Southern blot hybridization of NotI-digested genomes of classical, El Tor and O139 with a NotI-linking clone of classical strain 569B, suggest that these strains closely resemble V . cholerae O1 biotype El Tor, but are widely different from the classical O1 vibrios . Using restriction enzymes which cleave a single site in either the core region or in the direct repeat sequence (RS) of the CTX genetic element, it has been shown that the genome of most of the O139 strains has two copies of the ctx gene in tandem connected by two RSs . The chromosomal location of the CTX genetic element in the O139 strain is the same as that reported for El Tor vibrios . The organization of the virulence gene cassettes in different O139 strains shows genetic heterogeneity in the population . Whilst most of the epidemic strains have two copies of the CTX genetic element, in some strains the number of elements has been amplified and in at least one strain a single copy of the element has been deleted.

J Biol Chem, 1995 Jul 21, 270(29), 17627 - 32
Purification and properties of periplasmic 3':5'-cyclic nucleotide phosphodiesterase . A novel zinc-containing enzyme from the marine symbiotic bacterium Vibrio fischeri; Callahan SM et al.; The 3':5'-cyclic nucleotide phosphodiesterase (CNP) of Vibrio fischeri, due to its unusual location in the periplasm, allows this symbiotic bacterium to utilize extracellular 3':5'-cyclic nucleotides (e.g . cAMP) as sole sources of carbon and energy, nitrogen, and phosphorus for growth . The enzyme was purified to apparent homogeneity by a four-step procedure: chloroform shock, ammonium sulfate precipitation, and chromotography on DEAE-Sephacel and Cibacron Blue 3GA-agarose . The active enzyme consists of a single polypeptide with a mass of 34 kDa . At 25 degrees C, it has a pH optimum of 8.25, a Km for cAMP of 73 microns, and a Vmax of 3700 mumol of cAMP hydrolyzed/min/mg protein (turnover number of 1.24 x 10(5)/min) . The specific activity of the V . fischeri enzyme is approximately 20-fold greater than that of any previously characterized CNP when comparisons of activity are made at the same assay temperature . Activity increases with temperature up to 60 degrees C . The CNP contains 2 atoms of zinc/monomer, and zinc, copper, magnesium, and calcium can restore activity of the apoenzyme to varying degrees . The exceptional specific activity of the enzyme and its unusual location in the periplasm support proposals that the enzyme enables the bacterium to scavenge 3':5'-cyclic nucleotides in seawater and that the enzyme plays a role in cAMP-mediated host-symbiont interactions.

J Biol Chem, 1995 Jul 14, 270(28), 16813 - 9
Catalytically active forms of the individual subunits of Vibrio harveyi luciferase and their kinetic and binding properties; Choi H et al.; Contradictory findings have recently been reported regarding the (in)abilities of individual subunits of the Vibrio harveyi alpha beta dimeric luciferase to catalyze bioluminescence . We have produced individual alpha and beta subunits separately in Escherichia coli JM109 cells by recombinant DNA techniques . Both subunits were purified to more than 90% homogeneity and found to be catalytically active, with their general catalytic properties and the specific activities similar to those reported earlier (Sinclair, J . F., Waddle, J . J., Waddill, E . F., and Baldwin, T . O . (1993) Biochemistry 32, 5036-5044) . Individual subunits were significantly distinct from the native luciferase with respect to inactivations by trypsin and N-ethylmaleimide, and the stability of the flavin 4a-hydroperoxide intermediate . The active species in isolated alpha and beta samples were each the predominant protein species, corresponding to a 42,000 M(r) alpha monomer and a 67,000 M(r) beta dimer, respectively . These findings clearly indicate that the activities of the individual subunits are not due to trace contaminations of the respective counter subunits . The much reduced specific activities of the individual subunits are, in part, a consequence of diminished abilities to oxidize the aldehyde substrate . Kinetic and equilibrium measurements indicate that alpha and beta 2 each contained a reduced flavin site, an aldehyde substrate site, and an aldehyde inhibitor site . The on and off rates of the decanal inhibitor binding were substantially slower than the bindings of decanal and reduced riboflavin 5'-phosphate substrates . These findings are consistent with a scheme that the aldehyde inhibitor blocks the binding of the reduced flavin substrate.

Wkly Epidemiol Rec, 1995 Jul 14, 70(28), 201 - 8
Cholera in 1994 . Part I; Regulation of angR et al.; Department of Molecular Microbiology and Immunology, Oregon Health Sciences University, Portland 97201, USAAngR and the product(s) encoded in the trans-acting factor (TAF) region are necessary for the full expression of the pJM1 plasmid-mediated anguibactin iron-uptake system in Vibrio anguillarum (Va) . In this report, we analyzed the factors that affect the expression of the angR gene . Northern blot analysis showed that angR encodes a 3.1-kb transcript which is expressed only under iron-limiting conditions . Measurement of steady-state RNA levels show that, under iron-limiting conditions, angR is positively regulated at the transcriptional level by product(s) of the Va TAF region . However, this enhancement of angR expression by TAF does not occur at high levels of the AngR protein, as assessed by using an angR::lacZ fusion in the presence of a construct containing angR under the control of ptac . We also report that repression of angR by iron could possibly be mediated by an endogenous Va antisense RNA beta, which contains a stem-loop structure complementary to the stem-loop structure located at the 5' end of angR.

Int J Syst Bacteriol, 1995 Jul, 45(3), 421 - 8
Classification of fish-pathogenic vibrios based on comparative 16S rRNA analysis; Wiik R et al.; No systematic classification of fish-pathogenic vibrios has been accomplished previously despite the use of serological, physiological, and genetical classification systems . In this study, a comparative 16S rRNA analysis of 34 strains (representing seven species) of fish-pathogenic vibrios was performed . The 16S rRNA sequences were obtained by using reverse transcriptase . Nearly complete sequences were obtained for nine strains . On the basis of the results of this analysis, the remaining strains were investigated by analyzing selected stretches containing a total of 560 nucleotides . With the exception of a few strains, including ATCC 43313 (serovar O9), our comparative 16S rRNA analysis confirmed that strains preliminarily identified as Vibrio anguillarum were phylogenetically closely related . Strains of V . anguillarum could be divided into groups, with the main group containing serotype O1 and O2 strains isolated from Atlantic salmon, rainbow trout, turbot, cod, and saithe . The other distinctive group was represented by type strain NCMB 6 . This strain was nearly indistinguishable from the type strains of Vibrio ordalii and Vibrio damsela on the basis of the 16S rRNA stretches compared . The results of a comparative 16S rRNA analysis justified the status of Vibrio salmonicida as a distinct species . Originally, this species was characterized biochemically as a very homogeneous species . However, two strains, which were isolated from diseased halibut and from the intestines of healthy cod, could not be distinguished from V . salmonicida strains phylogenetically, although they differed from the original species description in several phenotypic traits . Our results indicate that V . salmonicida and Vibrio fischeri form a cluster that is clearly separated from the cluster that includes V . anguillarum.

Microb Pathog, 1995 Jul, 19(1), 39 - 48
Pathogenesis studies on Vibrio alginolyticus in the grouper, Epinephelus malabaricus, Bloch et Schneider; Lee KK; A pathogenic vibrio, strain S3y was isolated from an outbreak of vibriosis in grouper (Epinephelus malabaricus) fry in Kaohsiung, Southern Taiwan in November 1991 . It was identified as Vibrio alginolyticus on the basis of a number of biochemical tests . The bacterium was therefore identified as Vibrio, a Gram-negative, straight rod, motile, oxidase and catalase positive, swarming on TSA plate, fermentative and produced yellow colonies on TCBS agar . It did not utilise sodium citrate, but it was gelatinase positive and sensitive to the vibriostatic agent, 0/129 . It could grow on TSA with up to 11% NaCl . Its G+C ratio was 47% . The minimum lethal dose of strain S3y was 500 cfu/g fish body weight in the grouper (Epinephelus malabaricus) . The same bacterial species could be reisolated from the kidney of moribund fish following bacterial challenge . The extracellular products (ECP) of S3y were lethal to fish with a minimum lethal dose of 0.52 microgram/g fish body weight . A 34 kDa protease was purified from the ECP of S3y and demonstrated to be a toxin by intraperitoneal injection in the grouper . The minimum lethal dose of the purified protease was 0.17 microgram/g fish body weight . Intraperitoneal injection of the bacteria, ECP or the 34 kDa protease all caused slight exophthalmia with corneal opaqueness in moribund and dead fish . The results suggested that the 34 kDa protease might play an important role in the pathology of vibriosis caused by this bacterium.

Microb Pathog, 1995 Jul, 19(1), 11 - 8
Thymine auxotrophy as an attenuating marker in Vibrio cholerae; Attridge SR; Vibrio cholerae CVD102 is a thymine-dependent auxotroph of CVD101, a cholera toxin A-B+ candidate live oral cholera vaccine . Previous clinical experience with these strains suggested that, by restricting intestinal growth, thymine auxotrophy is attenuating for V . cholerae . Studies in the infant mouse cholera model cast doubt upon this conclusion however . Stable thyA mutants selected from each of three pathogenic strains showed unimpaired gut colonization in mixed-infection competition experiments . Similar results were obtained using thyA mutants selected from two atoxigenic strains, including CVD101 . Further studies with CVD102 showed that the reduced colonization potential of this strain could not be compensated by the provision of a functional thyA+ gene in trans . CVD102 shows reduced synthesis of toxin-coregulated pili (TCP) during in vitro growth, suggesting the presence of a second, undefined mutation in this strain . Given the critical role of TCP in intestinal colonization, it seems probable that this previously unrecognized mutation is responsible for the poor in vivo performance of CVD102.

J Infect, 1995 Jul, 31(1), 45 - 7
Epidemiology of Vibrio cholerae O139 with special reference to intrafamilial transmission in Calcutta; Sengupta PG et al.; A total of 27 families of hospitalised patients (index case families) suffering from acute watery diarrhoea caused by Vibrio cholerae O139, and 14 neighbourhood families were bacteriologically screened for 4 consecutive days to determine the extent of V . cholerae O139 infection amongst healthy contacts and other suspected vehicles of transmission at the intrafamilial level . V . cholerae O139 was isolated from faeces of 14.6% of healthy contacts in index case families as compared to none in neighbourhood families (P = 0.002) . The organism could be recovered from 3.7% of handwashings of contacts of index cases and also from stored drinking water (8.0%), open well water (28.6%), flies (3.8%) and pond water (25.0%) used by the index case families and none from neighbourhood families . The large number of asymptomatic infected persons indicate an epidemiological similarity to that of eltor cholera . The organisms may be carried on hands and may act as a potential source of infection to other inmates through contamination of stored drinking water, open wells etc . The results will be useful in formulating strategies for intervention of transmission of V . cholerae O139 at the community level.

Ecotoxicol Environ Saf, 1995 Jul, 31(2), 149 - 52
Toxicity of diphenylamine and some of its nitrated and aminated derivatives to the luminescent bacterium Vibrio fischeri; Drzyzga O et al.; Aqueous samples containing various nitrated and aminated diphenylamine derivatives were subjected to the luminescent bacterium Vibrio fischeri NRRL-B-11177 to determine their ecotoxicological potential . As the most important toxicological parameter, EC50, the concentration needed to reduce bacterial luminescence by 50%, was calculated . All compounds tested must be classified to the category "very toxic to aquatic organisms" using the widely accepted classification scheme of D . Strupp, H.P . Luhr, H . T . Grunder, J . Gerdesmann, and J . Ahlers (1990, UWSF--Z . Umweltchem . Okotox . 2, 151-156) . Only 2, 4-diaminodiphenylamine can be classified as "less toxic to aquatic organisms" . EC50 values after 30, 60, and 90 min of incubation of the test compounds are presented . For many of the compounds tested in this study there are no toxicological data in the literature.

J Infect Dis, 1995 Jul, 172(1), 173 - 9
The emerging diversity of the electrophoretic types of Vibrio cholerae in the Western Hemisphere; Evins GM et al.; Since the Latin American cholera epidemic began in 1991, 447 isolates of Vibrio cholerae O1 from the Western Hemisphere have been assayed by multilocus enzyme electrophoresis (MEE) to determine allelic variation among 16 enzyme-encoding genes . Two electrophoretic types (ETs) were identified among toxigenic isolates from Latin America: 323 were ET 4, the ET associated with the Latin American epidemic, and 29 were ET 3 . Twenty-three of these ET 3 isolates had a distinctive antimicrobial resistance pattern also seen in isolates imported into the United States from Latin America and Southeast Asia . These resistant isolates had an identical ribotype and nearly identical pulsed-field gel electrophoresis (PFGE) patterns . Most nontoxigenic isolates analyzed were not precursors or descendants of toxigenic epidemic strains . MEE provided a population genetic frame-work for the interpretation of PFGE and ribotype data from the isolates in this study . All three methods identified 2 distinct strains of toxigenic V . cholerae O1 currently epidemic in Latin America.

Infect Immun, 1995 Jul, 63(7), 2689 - 96
Heterologous antigen expression in Vibrio cholerae vector strains; Butterton JR et al.; Live attenuated vector strains of Vibrio cholerae were derived from Peru-2, a Peruvian El Tor Inaba strain deleted for the cholera toxin genetic element and attRS1 sequences, which was developed as a live, oral vaccine strain . A promoterless gene encoding the Shiga-like toxin I B subunit (slt-IB) was inserted in the V . cholerae virulence gene irgA by in vivo marker exchange, such that slt-IB was under transcriptional control of the iron-regulated irgA promoter . slt-IB was also placed under transcriptional control of the V . cholerae heat shock promoter, htpGp, and introduced into either the irgA or lacZ locus, or both loci, on the chromosome of Peru-2, generating JRB10, JRB11, or JRB12, respectively . A new technique was used to perform allelic exchange with lacZ . This method uses plasmid p6891MCS, a pBR327 derivative containing cloned V . cholerae lacZ, to insert markers of interest into the V . cholerae chromosome . Recombinants can be detected by simple color screening and antibiotic selection . In vitro measurements of Slt-IB produced by the vector strains suggested that expression of Slt-IB from the irgA and htpG promoters was synergistic and that two copies of the gene for Slt-IB increased expression over a single copy . The V . cholerae vectors colonized the gastrointestinal mucosa of rabbits after oral immunization, as demonstrated by very high serum antibody responses to V . cholerae antigens . Comparison of the serologic responses to the B subunit of cholera toxin (CtxB) following orogastric inoculation either with the wild-type C6709 or with Peru-10, a strain containing ctxB regulated by htpGp, suggested that both the cholera toxin and heat shock promoters were active in vivo, provoking comparable immunologic responses . Orogastric inoculation of rabbits with vector strains evoked serum immunoglobulin G (IgG) responses to Slt-IB in two of the four strains tested; all four strains produced biliary IgA responses . No correlation was observed between the type of promoter expressing slt-IB and the level of serum IgG or biliary IgA response, but the vector strain containing two copies of the gene for slt-IB evoked greater serum IgG responses than strains containing a single copy, consistent with the increased expression of Slt-IB from this strain observed in vitro . A comparison of the serum and biliary antibody responses to Slt-IB expressed from htpGp versus CtxB expressed from the same promoter suggested that CtxB is a more effective orally delivered immunogen.

Klin Lab Diagn, 1995 Jul-Aug, (4), 50 - 3
{Choice of commercial elective nutrient media for the isolation of pathogenic vibrios of different species}; Danilkina EB et al.; Commercial nutrient media and their modifications are assessed, including elective differentiation medium for V . cholerae developed at the Rostov Research Anti-Plague Institute for the isolation of pathogenic vibrios . V . cholerae cholerae P-1 (145), V . cholerae el tor M-878, V . cholerae non 01 P-9741, E . coli 18, P . vulgaris 19, and 48 strains of vibrios belonging to different species were used in the study . All the strains used were characterized as to their nutritive requirements . Alkaline agar manufactured by the I . I . Mechnikov Research Institute of Vaccines and Sera was found to be the optimal medium for the isolation of pathogenic vibrios, for it provided the growth of all the examined strains from the inoculation dose of 100 bacterial cells . Addition of potassium tellurite to this medium in a dose of 30 to 50 mg/ml improved its selective properties . Elective differential medium for V . cholerae, though somewhat inferior to alkaline agar in sensitivity, provided the growth of colonies of the overwhelming majority of the strains and inhibited the growth of associated bacteria . Hence, these two media are recommended as elective media for the diagnosis and selection of pathogenic vibrio . Endo and Ploskirev's media with alkaline agar in 1 to 1 ratio may be recommended as additional ones for the isolation of vibrios from patients and from environmental objects.

Appl Environ Microbiol, 1995 Jul, 61(7), 2624 - 30
Entry into, and resuscitation from, the viable but nonculturable state by Vibrio vulnificus in an estuarine environment; Oliver JD et al.; Using plate counts, total cell counts, and direct viable counts, we examined the fate of cells of Vibrio vulnificus placed into natural estuarine waters during both winter and summer months . Cells inoculated into membrane diffusion chambers and placed into estuarine waters entered into a viable but nonculturable (VBNC) state in January and February, when the water temperatures were low (average, < 15 degrees C) . In contrast, when cells in the VBNC state were placed into the same waters in the warmer months of August through November (average water temperature of ca . 21 degrees C), the cells appeared to undergo a rapid (typically, within 24 h) resuscitation to the fully culturable state . These results were independent of whether the cells were in the logarithmic or stationary phase and whether they were encapsulated or not . This study indicates that the inability to isolate V . vulnificus from cold estuarine sites may be accounted for by entrance of the cells into a VBNC state and that recovery from this state in natural environments may result from a temperature upshift.

Appl Environ Microbiol, 1995 Jul, 61(7), 2620 - 3
In vivo resuscitation, and virulence towards mice, of viable but nonculturable cells of Vibrio vulnificus; Oliver JD et al.; Vibrio vulnificus is an estuarine bacterium responsible for 95% of all seafood-related deaths in the United States . The bacterium occurs naturally in molluscan shellfish, and ingestion of raw oysters is typically the source of human infection . V . vulnificus is also known to enter a viable but nonculturable (VBNC) state, wherein the cells are no longer culturable on routine plating media but can be shown to remain viable . Whether or not this human pathogen remains virulent when entering the VBNC state has not been definitively demonstrated . In this study, the VBNC state was induced through a temperature downshift to 5 degrees C, with cells becoming nonculturable (< 0.1 CFU/ml) within 7 days . As they became nonculturable, virulence was determined by employing an iron overload mouse model . At the point of nonculturability (7 days), injections of the diluted microcosm population resulted in death when < 0.04 CFU was inoculated, although > 10(5) cells in the VBNC state were present in the inoculum . Culturable cells of V . vulnificus, with identification confirmed through PCR, were recovered from the blood and peritoneal cavities of mice which had died from injections of cells present in the VBNC state for at least 3 days . Thus, our data suggest that cells of V . vulnificus remain virulent, at least for some time, when present in the VBNC state and are capable of causing fatal infections following in vivo resuscitation . Our studies also indicate, however, that virulence decreases significantly as cells enter the VBNC state, which may account, at least to some extent, for the decrease in infections caused by this bacterium during winter months.

Appl Environ Microbiol, 1995 Jul, 61(7), 2493 - 8
Analysis of antigens present in the extracellular products and cell surface of Vibrio anguillarum serotypes O1, O2, and O3; Santos Y et al.; Antigens present in the extracellular products (ECP) and cell walls of strains of Vibrio anguillarum of serotypes O1, O2, and O3 isolated from different fish species in distinct geographic areas were characterized . The usefulness of slide agglutination, dot blot assay, and quantitative agglutination for subtyping V . anguillarum serovars was also evaluated . The three serological assays used to establish the serogroups within V . anguillarum isolates demonstrated that serotype O1 constitutes a homogeneous group, whereas within serotypes O2 and O3, two different patterns of serological reactions were detected . Among the three serological methods used, only dot blot and quantitative agglutination assays differentiated subgroups within serotypes O2 and O3 with unabsorbed sera . Electrophoretic analysis and immunoblot assays of cell envelope and ECP components showed that strains belonging to serotype O1 possessed immunologically related lipopolysaccharide (LPS) and proteins, while V . anguillarum isolates grouped in serotypes O2 and O3 exhibited internal heterogeneity in their LPS and protein banding patterns . On the other hand, although the LPS present in the ECP and those obtained from cell envelopes of V . anguillarum strains showed apparently different gel patterns, a strong relationship between both types of LPS was seen by immunoblot assay . From these results, it can be concluded that V . anguillarum strains representative of each of the antigenic groups (O1, O2 alpha, O2 beta, O3A, and O3B) and their ECPs should be included in the formulation of vaccines against vibriosis in areas where the three serotypes coexist.

J Bacteriol, 1995 Jul, 177(14), 4131 - 3
Induction of cold-responsive proteins in Vibrio vulnificus; McGovern VP et al.; We have studied the response of Vibrio vulnificus to temperature shifts (23 to 13 degrees C) within the organism's permissive growth range . Cold shift induced a diminution in protein synthesis . Following a short lag, cells began growth at a new rate . Forty proteins were induced by this downshift.

Lipids, 1995 Jul, 30(7), 677 - 9
Comparison of the fatty acids of the tunicate Botryllus schlosseri from the Black Sea with two associated bacterial strains; Carballeira NM et al.; The fatty acid composition of the tunicate Botryllus schlosseri and of two bacterial strains found within the tunicate, namely Vibrio parahaemolyticus and of an associated but previously unreported gram positive cocci were studied . The polyunsaturated fatty acids 6,9,12-octadecatrienoic acid, 5,8,11,14,17-eicosapentaenoic acid, and 4,7,10,13,16,19-docosahexaenoic acid were particularly abundant in B . schlosseri and were not detected in the two bacterial strains found in the tunicate . The iso/anteiso pair, 13-methyltetradecanoic acid and 12-methyltetradecanoic acid, were the principal fatty acids in the gram positive cocci, and the 9- and 11-hexadecenoic acids were particularly abundant in V . parahaemolyticus . The diunsaturated fatty acid 9,12-octadecadienoic acid was also shown to be present in V . parahaemolyticus . The fatty acid composition of a third bacterial strain, characterized as either a Pseudomonas or an Alteromonas species, and shown to be present only in the sea water from the Black Sea and not in B . schlosseri, is also reported . This is the first investigation on fatty acids from Black Sea bacteria.

Kansenshogaku Zasshi, 1995 Jul, 69(7), 826 - 34
Rapid detection of Vibrio cholerae with a new selective enrichment medium and polymerase chain reaction; Kida N et al.; The inhibitory effect of metallic EDTA compounds on growth of Vibrio cholerae and Escherichia coli was studied . Only Fe-EDTA among the compounds tested showed pH-dependent growth inhibition on E . coli at pH 9.0, but no inhibition of V . cholerae at the same pH . By addition of Fe-EDTA as a selective inhibitor, a novel enrichment broth (tentatively designated as VCF broth) for the selective isolation and cultivation of V . cholerae from other Gram-negative bacilli has been developed, and the selective enrichment capacity of VCF broth for V . cholerae and selective inhibiting activity against E . coli were significantly higher than those of alkaline peptone water . A simple procedure for rapid detection of V . cholerae by selective enrichment for 6 hr with VCF broth and then amplification of the cholera toxin target DNA fragment by the polymerase chain reaction was presented . VCF broth may be a useful tool for the selective enrichment of V . cholerae in bacterial examinations.

New Microbiol, 1995 Jul, 18(3), 289 - 97
First data on the distribution and ecology of Vibrio spp . of the Straits of Magellan (South America); Monticelli LS et al.; During the austral summer of 1991 a study was carried out on the presence and distribution of the genus Vibrio in the Straits of Magellan . Vibrios strains were isolated using membrane filters and Marine Agar 2216 in anaerobiosis . Variations of the populations of total heterotrophic bacteria and vibrios were observed both on the surface and along the column of water . All vibrios are psychrotrophic and were grouped in 4 cluster among which cluster 1, identified as presumed V . anguillarum, seems the most important including 73% of strains . A certain habitat segregation of clusters was noted . Cluster 4 was found only in a deep and permanently colder water mass . The relations between 20 environmental parameters and the bacterial population were also studied . Significant positive correlations were observed between the vibrios population and various fractions of suspended particulate matter.

J Foot Ankle Surg, 1995 Jul-Aug, 34(4), 354 - 7
Lower extremity manifestations of Vibrio vulnificus infection; Laughlin TJ et al.; Vibrio vulnificus is a potentially lethal marine bacterium that has not been previously described in podiatric literature . A review of the microorganism's characteristics, susceptible patient population, and lower extremity manifestations of infection is presented . V . vulnificus is found as part of the normal flora of the Gulf of Mexico, Atlantic, and Pacific coastal waters and is often isolated from the filter feeding shellfish of these regions . Its pathogenicity is generally reserved for the immunocompromised host, and is specifically related to disease states which exhibit high serum iron levels . V . vulnificus infections present in two distinct clinical syndromes: primary sepsis secondary to raw oyster ingestion, or localized infection from wound exposure to V . vulnificus-inhabited salt water . Both syndromes demonstrate characteristic skin lesions of the trunk and extremities that present as hemorrhagic bullae and progress to necrotic ulcerations . Although V . vulnificus infection is rare, its extreme virulence in patients suffering from a chronic disease process and its manifestation of characteristic lower-extremity lesions require the podiatric physician to be able to recognize and treat such a condition.

Mol Gen Mikrobiol Virusol, 1995 Jul-Sep, (3), 30 - 4
{Phenotypic analysis of two morphologically different types of Vibrio cholerae 0139 colonies}; Smirnova NI et al.; A clinical isolate Vibrio cholerae P16064 serogroup 0139 was shown to produce two different kinds of colonies: opaque encapsulated and translucent devoid of capsule . Low virulence of translucent colonies seems to be due to not only loss of capsule which determines serum resistance, but also to decreased expression of genes controlling the motility, and production of protease and mannose-sensitive adhesion pili . Analysis of the lysogenic properties of the two types of colonies permitted us to propose that simultaneous spontaneous alteration of capsule production and the above-mentioned virulence factors in translucent colonies may be caused by a temperate phage in turbid clones of strain P16064.

Mol Microbiol, 1995 Jul, 17(1), 197 - 204
Characterization and regulation of the expression of FatB, an iron transport protein encoded by the pJM1 virulence plasmid; Actis LA et al.; The pJM1-encoded genes fatDCBA are essential for iron acquisition via the siderophore anguibactin . Sequence analysis indicated that the open reading frame corresponding to the fatB gene possesses domains that are characteristic of periplasmic proteins that bind the ferric siderophore . In this work, a monospecific antiserum against an oligopeptide containing the last 27 amino acids of the carboxy-terminal region from this open reading frame was used to demonstrate that fatB encodes a 35 kDa protein that is essential for iron transport . By using this antibody we were able to demonstrate that expression of the fatB gene is negatively regulated by the Fur protein at high iron concentrations . Conversely, its expression was positively regulated by the combined action of the AngR protein and products of the TAF region . FatB, the product of the fatB gene, is isolated with the membrane fraction . In accordance with these findings is the fact that the first 23 amino acid residues of this protein have the properties of a lipoprotein signal sequence . The lipoprotein nature of FatB is supported by the fact that treatment of Vibrio anguillarum cells with globomycin, an inhibitor of the lipoprotein signal peptidase, results in the accumulation of a 38 kDa proFatB precursor protein.

Nippon Saikingaku Zasshi, 1995 Jul, 50(3), 863 - 70
{Causative agent of the so-called "light disease of shrimps" is luminescent Vibrio cholerae non-O1}; Shimada T et al.; A number of luminous fresh-water shrimps were found in a fish preserve in Lake Biwa, Shiga Prefecture, in the middle of July, 1994, and most of them died within several hours after collection (the so-called "light disease of shrimp") . Four luminous organisms were isolated from a dead shrimp . Although the phenotypic properties of these strains were similar to those of V . cholerae or V . mimicus, a representative strain, 838-94, was shown to have a high level (79%) of DNA homology with V . cholerae type strain, ATCC 14035 and a low level (45%) of relatedness to V . mimicus type strain, ATCC 33653 . Therefore, these isolates were identified as V . cholerae . The four strains fell into serogroup O28 of V . cholerae . On the other hand, none of the isolates had CT nor NAG-ST genes . The results obtained herein clearly demonstrate that these organisms isolated from luminous shrimps are luminescent V . cholerae serogroup O28.

Biochim Biophys Acta, 1995 Jun 30, 1230(3), 170 - 6
Three aspartic residues in membrane-spanning regions of Na+/H+ antiporter from Vibrio alginolyticus play a role in the activity of the carrier; Nakamura T et al.; The Na+/H+ antiporter gene from Vibrio alginolyticus restores the growth of an nhaA-defective strain of Escherichia coli, NM81, in a high NaCl medium (Nakamura, T., Komano, Y., Itaya, E., Tsukamoto, K., Tsuchiya, T . and Unemoto, T . (1994) Biochim . Biophys . Acta 1190, 465-468) . This gene, named nhaAv, allowed the nhaA-defective E . coli strains, NM81(delta nhaA) and RS1 (delta nhaA, chaA-), to extrude Na+ at alkaline pH . The extrusion of Na+ occurred against its chemical gradient in the presence of membrane-permeable amine . Thus, the nhaAv gene product is functional as an electrogenic Na+/H+ antiporter in E . coli cells . The NhaAv protein has only four acidic amino acid residues in the putative membrane-spanning regions, that is, Asp-57, Asp-125, Asp-155 and Asp-156, and these Asp residues are conserved in NhaA from E . coli . Asp-111, which is predicted to be in a loop region between the transmembrane segments is also conserved in NhaA . Thus, each conserved Asp residue was replaced with asparagine by a site-directed mutagenesis . E . coli NM81 cells containing a plasmid harboring the nhaAv gene mutated at Asp-125, -155, or -156 could neither grow in a high NaCl medium nor extrude Na+ at alkaline pH against its chemical gradient . These results show that Asp-125, -155, and -156, but not Asp-57 and -111, play a role in the activity of the Na+/H+ antiporter, NhaAv.

J Biol Chem, 1995 Jun 9, 270(23), 13956 - 60
A mutant at position 87 of the GroEL chaperonin is affected in protein binding and ATP hydrolysis; Weiss C et al.; The highly conserved aspartic acid residue at position 87 of the Escherichia coli chaperonin GroEL was mutated to glutamic acid . When expressed in an E . coli groEL mutant strain deficient for phage morphogenesis, plasmid-encoded GroEL mutant D87E restored T4 phage morphogenesis . It did not, however, reactivate the transcription of a recombinant luciferase operon from Vibrio fischeri . In vitro, GroEL mutant D87E was found to be impaired in the ability to bind nonnative proteins and to hydrolyze ATP, resulting in less efficient refolding of urea-denatured ribulose-1,5-bisphosphate carboxylase/oxygenase . Mutant oligomer D87E GroEL14 was able to bind GroES7 as efficiently as wild-type GroEL14 . The conserved aspartic acid residue at position 87 located in the equatorial domain of GroEL (Braig, K., Otwinowski, Z., Hegde, R., Boisvert, D.C., Joachimiak, A., Horwich, A.L., and Sigler, P.B . (1994) Nature 371, 578-586) is thus inferred to have a dual effect on the binding of nonnative proteins to the GroEL14 core chaperonin and on ATP hydrolysis.

Southeast Asian J Trop Med Public Health, 1995 Jun, 26(2), 342 - 6
Widespread emergence of Vibrio cholerae 0139 in India; Sachdeva V et al.; The National Institute of Communicable Diseases (NICD) has been monitoring the incidence of laboratory confirmed cases of cholera in Delhi in collaboration with Infectious Diseases Hospital (IDH) since 1965 . Cholera and cholera-like cases from all hospitals in Delhi are admitted in IDH and the rectal swabs of all such cases are processed for isolation of Vibrio cholerae at NICD laboratory . Since April 1993, there has been isolation of Vibrio cholerae serotype 0139, in increasing numbers (831 out of 2,830, 29.2%) The isolates have been characterized and enterotoxin studies carried out . As a referral laboratory NICD has also confirmed the causative role of Vibrio cholerae 0139 in diarrhea outbreaks from various parts of the country . The implications of establishment of this newer serotype of Vibrio cholerae, as a potential epidemic strain are discussed.

Appl Environ Microbiol, 1995 Jun, 61(6), 2292 - 6
Evidence for the existence of distinct populations of Vibrio anguillarum serogroup O1 based on plasmid contents and ribotypes; Pedersen K et al.; A total of 103 Vibrio anguillarum serogroup O1 strains displaying 15 different plasmid profiles were characterized with respect to biochemical properties and ribotypes . The results confirmed that V . anguillarum O1 is a biochemically homogeneous group . The 103 strains could be allocated to three main clusters with high similarity coefficients . None of the biochemical properties were connected with the presence of plasmids . In total, 12 different ribotypes were demonstrated, with HindIII being used as the restriction enzyme . Forty of the strains were isolated from the same Danish fish farm, some from the kidneys of diseased fish and some from the environment, and some strains were isolated from the mucus, gills, and feces of healthy fish . Nineteen of these isolates possessed the 67-kb virulence plasmid alone or in combination with other plasmids, while 21 had no plasmids . All strains isolated from the kidneys of diseased fish on this farm had plasmids . Irrespective of their origin (kidneys, gills, or mucus), all 19 strains carrying the 67-kb virulence plasmid had the same ribotype, profile 1, while isolates without plasmids belonged to five different profiles, all different from profile 1 . These results suggest that pathogenic V . anguillarum O1 strains possessing a virulence plasmid and nonpathogenic strains without plasmids from a small geographical area and even from the same fish may constitute two essentially distinct populations . Thus, it may be suggested that an exchange of virulence plasmids among strains is unlikely to occur in vivo.

J Med Microbiol, 1995 Jun, 42(6), 399 - 403
Presence of lysogenic phage in the outbreak strains of Vibrio cholerae O139; Mitra SN et al.; Four outbreak strains of Vibrio cholerae O139 from endemic areas of India and Bangladesh were found to carry lysogenic phage(s) . All of these phage(s) produced turbid plaques characteristic of lysogeny on V . cholerae MAK 757 (El Tor, Ogawa) cells as well as on their VcA-1 lysogens but were unable to infect V . cholerae 154 (classical) cells, the universal host for all classical phages . Colonies in the turbid plaques were O139 lysogens and these developed an auxotrophic requirement, mainly for purines suggesting the integration of the prophage into the host chromosome . The immunity profile of the O139 phage(s) was similar to that of phage alpha but differed in the sensitivity of the phage lysogen of V . cholerae MAK 757 to subsequent infection by phage beta.

FEMS Microbiol Lett, 1995 Jun 1, 129(1), 97 - 101
Intragenic suppression of a luxR mutation: characterization of an autoinducer-independent LuxR; Poellinger KA et al.; The Vibrio fischeri luminescence genes are activated by the LuxR protein and a diffusible signal termed the autoinducer . LuxR consists of two domains, a C-terminal transcriptional activator domain, and an N-terminal autoinducer-binding domain, which serves to regulate the function of the C-terminal domain . We have isolated and characterized an intragenic suppressor of a mutation that maps to the N-terminal domain and blocks autoinducer binding . The suppressor changes an alanine residue at position-221 in the C-terminal domain to a valine . In Escherichia coli, the suppressor allows partial activation of the V . fischeri luminescence genes although E . coli containing this protein remains unable to bind autoinducer . To further analyze the influence of the second-site mutation on luxR function, we constructed a luxR gene that coded for a protein with a wild-type N-terminal domain and with the ala-221 to val substitution in the C-terminal domain . This protein activated the luminescence genes in the presence or absence of autoinducer, and it bound autoinducer at levels comparable to the wild-type LuxR protein . Apparently, the alanine to valine substitution at position-221 allows activity of the C-terminal domain in a fashion independent of whether autoinducer is bound to the N-terminal domain.

Dig Dis Sci, 1995 Jun, 40(6), 1257 - 60
Fatal Vibrio parahemolyticus septicemia in a patient with cirrhosis . A case report and review of the literature; Hally RJ et al.; Vibrio parahemolyticus has been well documented to cause outbreaks of infectious diarrhea, usually related to poor food handling; only rarely has it been reported to cause fetal septicemia . In contrast, Vibrio vulnificus is a well-known cause of septicemia, especially in patients with cirrhosis . A 31-year-old woman with cirrhosis who developed fatal V . parahemolyticus sepsis after ingesting raw seafood is described . We review the clinical syndromes associated with sepsis caused by these two organisms . Leg pain and bullous skin lesions may be a clue to the diagnosis . Febrile patients with cirrhosis should be questioned regarding recent seafood ingestion, and appropriate antibiotics chosen if this history is obtained . Physicians should inform patients at risk to avoid raw seafood in an attempt to prevent this potentially lethal syndrome.

J Pediatr, 1995 Jun, 126(6), 882 - 6
Clinical characteristics and risk factors for Vibrio cholerae infection in children; Fukuda JM et al.; Surveillance was conducted during February and March 1991 in the pediatric emergency department of Cayetano Heredia Hospital, Lima, Peru, to contrast the characteristics of children with epidemic cholera with those of children with noncholera-associated diarrhea . Among 626 patients 14 years of age or younger, Vibrio cholerae O1 was isolated from stool specimens of 310 patients (49%), more commonly from children older than 24 months of age (66%; p < 0.0001) than from younger children . Cholera was clinically characterized by a more sudden onset; watery diarrhea; and associated abdominal pain, muscle cramps, and vomiting, which led to more severe dehydration and hospitalization more often than in noncholera cases . Only one patient with cholera died, for a case-fatality rate of 3.2 deaths per 1000 persons . Nonpotable water and uncooked foods were identified as probable vehicles for V . cholerae . The frequency of diarrhea among relatives of patients with cholera suggested intrafamily transmission . This study of epidemic cholera describes the clinical features and the risk factors for acquisition of the infection, and points out the low case-fatality rate with prompt and appropriate treatment.

Curr Microbiol, 1995 Jun, 30(6), 331 - 6
Isolation of bacteriophage infectious for Vibrio vulnificus; Pelon W et al.; Nine phage isolates infectious for Vibrio vulnificus and falling into four morphological groups were isolated from estuarine waters collected in Louisiana . Of the 60 V . vulnificus strains tested, 87% were susceptible to one or more of the isolates . With the exception of V . fluvialis, Vibrio species other than vulnificus were resistant to infection . A spectrum of enteric bacterial strains were similarly resistant . Susceptibility differences were seen between opaque (virulent) V . vulnificus strains and those with translucent (nonvirulent) colony types, with the former being more susceptible . Susceptibility patterns to infection by the nine phage isolates among the V . vulnificus test strains suggest that the latter may fall into several groups . Other aspects relating to the phage isolates are presented.

J Infect Dis, 1995 Jun, 171(6), 1653 - 6
Impact of infection by Helicobacter pylori on the risk and severity of endemic cholera; Clemens J et al.; To evaluate the relationship between Helicobacter pylori infection and the subsequent risk and severity of endemic Vibrio cholerae O1 diarrhea among rural Bangladeshis, 285 children and adults with cholera (cases) and 881 contemporaneously selected community controls were studied . Cases and controls were contrasted for H . pylori infection, as manifested by serum IgG anti-H . pylori antibodies . Although the overall risk of cholera was not significantly increased among H . pylori-infected subjects, the risk of cholera of life-threatening severity was significantly elevated (relative risk {RR} = 1.61; 95% confidence interval {CI} = 1.07-2.42) . A significant increase in the risk of severe cholera was seen in subjects who lacked natural serum vibriocidal antibodies (RR = 2.88; 95% CI = 1.28-6.48) but not in those with such antibodies . Thus, H . pylori infection was associated with a significant increase in the risk of life-threatening cholera, but only among persons lacking natural vibriocidal immunity.

Infect Immun, 1995 Jun, 63(6), 2356 - 60
Construction of nontoxic derivatives of cholera toxin and characterization of the immunological response against the A subunit; Fontana MR et al.; Using computer modelling, we have identified some of the residues of the A subunit of cholera toxin (CT) and heat-labile toxin that are involved in NAD binding, catalysis, and toxicity . Here we describe the site-directed mutagenesis of the CT gene and the construction of CT mutants . Nine mutations of the A subunit gene were generated . Six of them encoded proteins that were fully assembled in the AB5 structure and were nontoxic; these proteins were CT-D53 (Val-53-->Asp), CT-K63 (Ser-63-->Lys), CT-K97 (Val-97-->Lys), CT-K104 (Tyr-104-->Lys), CT-S106 (Pro-106-->Ser), and the double mutant CT-D53/K63 (Val-53-->Asp, Ser-63-->Lys) . Two of the mutations encoded proteins that were assembled into the AB5 structure but were still toxic; these proteins were CT-H54 (Arg-54-->His) and CT-N107 (His-107-->Asn) . Finally, one of the mutant proteins, CT-E114 (Ser-114-->Glu), was unable to assemble the A and the B subunits and produced only the B oligomer . The six nontoxic mutants were purified from the culture supernatants of recombinant Vibrio cholerae strains and further characterized . The CT-K63 mutant, which was the most efficient in assembly of the AB5 structure, was used to immunize rabbits and was shown to be able to induce neutralizing antibodies against both the A and B subunits . This molecule may be useful for the construction of improved vaccines against cholera.

Int J Food Microbiol, 1995 Jun, 26(1), 77 - 91
Culture media for the isolation and enumeration of pathogenic Vibrio species in foods and environmental samples; Donovan TJ et al.; The genus Vibrio now includes a large number of species . Clear evidence is only available for the aetiological role of V . cholerae, V . vulnificus and V . parahaemolyticus in foodborne diseases . Until recently, V . cholerae serogroup 0:1 was accepted as the cause of epidemic cholera . However, the designation of outbreaks of diarrhoeal diseases caused by V . cholerae 0:139 as clinical cholera has lead to renewed interest in Non 0:1 serogroups of V . cholerae . A wide range of enrichment and selective media for the isolation of vibrios has been developed . These media are reviewed with respect to their ability to recover and differentiate the target vibrios . Alkaline peptone water (APW) remains the recommended enrichment medium for vibrios in parallel with either salt polymyxin broth (SPB) or glucose teepol (or sodium dodecylsulphate) salt broth (GTSB) when tests for V . parahaemolyticus are required . Thiosulphate citrate bile salt agar (TCBS) in parallel with polymyxin mannose tellurite (PMT) or sodium dodecylsulphate polymyxin sucrose agar (SPS) are the recommended selective plating media.

J Trop Pediatr, 1995 Jun, 41(3), 139 - 42
Epidemiology of cholera in Delhi--1992; Singh J et al.; Cholera is endemic in Delhi and is a highly seasonal disease . Suspected cholera cases are referred to Infectious Diseases Hospital, Delhi . Rectal swabs from 2783 cases were bacteriologically examined during 1992, out of which 1075 were found to be positive for Vibrio cholerae O1 biotype El Tor . First isolation was made on 3 April and the last on 14 December . About 87 per cent isolations were made between May and September, which are summer and monsoon months in Delhi . Detailed epidemiological information was collected for about 198 cases of diarrhoea out of which 103 were confirmed cases of cholera . Half of these cases occurred in children below 10 years of age . The other major group affected was adult females, especially housewives . All the cholera cases occurred in those who were illiterate or educated up to primary level . Important risk factors were: contact with person having similar illness, storage of water in wide-mouthed containers, use of glass or mug to draw water from containers, absence of sanitary latrines and habit of washing hands with water alone after defecation, before cooking and eating food . About 30 percent cases had access to piped water supply which was found safe in Delhi during 1992 . The findings suggest that the hygienic practices were more important than contaminated water sources for transmission of cholera in Delhi during the year 1992.

Kansenshogaku Zasshi, 1995 Jun, 69(6), 637 - 41
{Studies on Vibrio cholerae non-O1 isolated from diarrheal patients arrived from overseas}; Tsutsumi H et al.; Between the period January 1991 to June 1993, there were 23,976,238 travellers who arrived from overseas to Narita Airport, of which 20,501 stool specimens were collected from diarrheal patients for bacteriological examination, and infectious agents were detected from 2,751 cases (13.4%) including 250 cases (1.2%) of Vibrio cholerae non-O1 . Countries suspected of infection of these patients were Thailand, the most in number, and followed by Indonesia, India and so on these mostly distributed in South-east or South Asia . About fifty percent of the patients were associated with abdominal pain and some with vomiting or fever . Diarrhea was mostly mild except in 16 patients who had severe diarrhea of more than ten times a day . 237 of the 250 isolated V . cholerae non-O1 strains were classified into 48 serogroups . There were 2 rough strains and 11 other strains which were out of the confirmed serogroups . Positive rate of these strains to haemolysin, cholera toxin (CT) and NAG-ST tests were 87.2, 0.4 and 0.8% respectively.

J Appl Bacteriol, 1995 Jun, 78(6), 621 - 9
The effect of incubation temperature and sodium chloride concentration on the growth kinetics of Vibrio anguillarum and Vibrio anguillarum-related organisms; Guerin-Faublee V et al.; The effect of temperature and NaCl concentration on the growth kinetics of Vibrio anguillarum and V . anguillarum-related (VAR) strains was studied . For one wild VAR strain, NaCl concentration interfered with growth temperature parameters, in particular, with the maximum growth temperature but also with the optimum temperature (defined as the temperature at which mumax equals its maximal value muopt), and with muopt itself . For the same strain, optimal growth required the adding of NaCl to the medium to a final concentration of 1.5% . These results were not confirmed by tests on a V . anguillarum collection strain . When the NaCl concentration in the culture media was 1.5%, the optimum temperature for the nine strains studied ranged from 29.7 degrees C to 34 degrees C whereas the maximum temperature ranged between 35.3 degrees C and 38.5 degrees C . Hence, antibiotic susceptibility testing as well as biochemical identification might be carried out at 30 degrees C in the presence of 1.5% NaCl, which corresponded to a suboptimal growth.

J Diarrhoeal Dis Res, 1995 Jun, 13(2), 127 - 9
Vibriocidal activities of some local herbs; Akinsinde KA et al.; Four of the seven tested medicinal plants exhibited antimicrobial activity against Vibrio cholerae . These 7 plants are: Ficus capensis, Mitragyna stipulosa, Entada africana, Piliostigma reticulatum, Terminalia avicennoides, Mimosa pudica, and Lannea acida . Of them Terminalia avicennoides showed higher antimocrobial activity than others . Potentials of these herbs in the control of cholera need to be determined.

J Diarrhoeal Dis Res, 1995 Jun, 13(2), 118 - 21
Plasmid profiles and antimicrobial susceptibility patterns of Vibrio cholerae O1 strain isolated during a recent outbreak in Nigeria; Olukoya DK et al.; In a study on the outbreak of cholera in Nigeria in 1992, 86 strains of Vibrio cholerae O1 (79 Ogawa serotype and 7 Inaba serotype) were isolated . Antimicrobial susceptibility testing and plasmid profile analysis of the strains were done . Most isolates were highly sensitive to ciprofloxacin, cefotaxime, chloramphenicol, gentamicin, erythromycin, nalidixic acid, and nitrofurantoin, and less sensitive to ampicillin, penicillin, cloxacillin, cotrimoxazole, streptomycin, and tetracycline . The strains showed 13 resistant patterns; the commonest resistant patterns were Apr, Smr, and ApTcr . A total of 41 (47.6%) strains contained one or more plasmid(s) with sizes ranging from 4.5 kilobase to 150 kilobase . Ten isolates were able to transfer resistant plasmids to Escherichia coli K-12 by conjugation . Antibiogram patterns distinguished more isolates than in plasmid profile analysis . Plasmids specifying resistance to ampicillin, tetracycline, and trimethoprim were found . The differing patterns of antibiogram and plasmid profiles indicated that many circulating strains were responsible for the last outbreak in the country.

J Diarrhoeal Dis Res, 1995 Jun, 13(2), 113 - 7
Molecular epidemiology of Vibrio cholerae O1 isolated in Nepal by southern hybridization with a cholera toxin gene probe; Yamamoto K et al.; A cholera epidemic broke out in 1992 due to Vibrio cholerae O1 biotype El Tor in the eastern and southern belt of Nepal mainly among the Bhutanese refugees . Restriction fragment profiles (RFP) of DNA fragments of V . cholerae O1 isolates hybridized with an enzyme-labelled oligonucleotide probe for cholera toxin gene (ctx) by Southern Hybridization were compared . The probe hybridized with the 13- and 8-kb fragments of PstI-digested total DNA in all isolates observed in the epidemic . This RFP in the Nepalese strain was not observed in the strains isolated during other epidemics but was observed in the strains isolated from the exported marine products from Taiwan and Thailand.

Hybridoma, 1995 Jun, 14(3), 271 - 8
Development of highly specific monoclonal antibodies for the diagnosis of Vibrio cholerae 01; Castillo L et al.; We report here the development of two monoclonal antibodies, termed 5G8 and 5C12, belonging to the IgM and IgG1 class, respectively, suitable for the identification of Vibrio cholerae 01 in clinical and environmental samples . The specificities of the monoclonals were evaluated by ELISA and indirect immunofluorescent microscopy of microorganisms normally present in stool samples and with two bacterial panels . One panel included 72 potentially antigenically related bacterial strains and the second panel included 20 pathogenic bacterial strains involved in diarrhea cases . The results of these extensive analyses indicate that monoclonal antibodies 5G8 and 5C12 are highly specific and suitable for the clinical diagnosis of Vibrio cholerae 01 in human stool samples by indirect immunofluorescent microscopy . Although the antigenic sites recognized by these antibodies were not identified in this study, the observation of Western blot patterns suggested that 5G8 and 5C12 monoclonal antibodies bind to LPS epitopes, a good structural marker for the detection of V . cholerae 01 because it is present in all bacterial cell walls.

Pigment Cell Res, 1995 Jun, 8(3), 147 - 52
Characterization of the melanogenic system in Vibrio cholerae, ATCC 14035; Ruzafa C et al.; The nature of the pigment formed by Vibrio cholerae and the characterization of its biosynthetic pathway is shown . This microorganism is able to synthesize melanin-like pigment when cultured in the presence of L-tyrosine . Other phenolic chemicals related to L-tyrosine do not lead to pigment production . The microorganism has no tyrosine hydroxylase activity, and the levels of dopa oxidase activity are very low, making the existence of a tyrosinase very unlikely . However, Vibrio cholerae contained transaminases that transforms L-tyrosine into p-hydroxyphenylpyruvate . Moreover, Vibrio cholerae is able to go further in the catabolic pathway, releasing a great amount of homogentisic acid . This acid can spontaneously be oxidized to its p-quinone form, which subsequently polymerizes leading to pigment formation . It is concluded that the pigment formed by Vibrio cholerae is not synthesized by the Raper-Mason pathway, but by a L-tyrosine catabolism pathway leading to homogentisic acid . Some simple properties of that melanin are compared to model eu- and pheomelanin, but no clear distinction could be stated, indicating the similarity between all these pigments.

Clin Infect Dis, 1995 Jun, 20(6), 1485 - 90
Ciprofloxacin for the treatment of cholera: a randomized, double-blind, controlled clinical trial of a single daily dose in Peruvian adults; Gotuzzo E et al.; We conducted a randomized, double-blind clinical trial to compare ciprofloxacin (250 mg once a day for 3 days) with tetracycline (500 mg four times a day for 3 days) in terms of efficacy and safety in the treatment of moderate-to-severe cholera in Peruvian adults . The baseline characteristics of the groups were similar . A total of 202 patients (102 in the tetracycline group and 100 in the ciprofloxacin group) were included in the efficacy analysis . The clinical and bacteriologic efficacies of the two regimens were similar . The study drugs were well tolerated . We conclude that ciprofloxacin given once a day is as effective as the standard tetracycline regimen for the treatment of cholera in adults . The ciprofloxacin regimen may represent an alternative to the standard treatment in areas where Vibrio cholerae O1 strains that are resistant to commonly used antimicrobials are prevalent.

Clin Infect Dis, 1995 Jun, 20(6), 1480 - 4
Efficacy and tolerability of ciprofloxacin prophylaxis in adult household contacts of patients with cholera; Echevarria J et al.; We conducted a randomized double-blinded study in Lima, Peru, to assess the tolerability and efficacy of a single 250-mg dose of ciprofloxacin in preventing diarrhea and Vibrio cholerae O1 infection among household contacts of bacteriologically confirmed index cases . Adult household contacts with negative baseline stool cultures were included . A total of 213 household contacts were evaluable . The study drugs were well tolerated in both groups . Ciprofloxacin did not prevent the acquisition of V . cholerae O1 infection nor the development of diarrhea . However, in a subgroup of 30 household contacts with positive baseline stool cultures a reduction in the bacterial load and a trend toward prevention of diarrhea were observed among ciprofloxacin recipients . When all household contacts were evaluated, a trend toward prevention of diarrhea was observed with the prophylactic regimen . Ciprofloxacin failed to prevent V . cholerae O1 infections during a period of low transmissibility.

Surg Endosc, 1995 Jun, 9(6), 730 - 2
Acute acalculous cholecystitis due to Vibrio cholerae; Gomez NA et al.; The case of a 57-year-old woman admitted with symptoms and signs suggesting an intestinal infection caused by Vibrio cholerae, and who also developed a clinical picture compatible with acute cholecystitis, is presented . Cholera was diagnosed by examining a fresh sample of stools and cultures . An abdominal sonogram disclosed signs of acute acalculous cholecystitis . She underwent cholecystectomy, and cultures of a clear fluid and a "milky" sediment found within the gallbladder were also positive for V . cholerae . This microorganism was seen at the gallbladder mucosa microscopically . The strain was serotyped V . cholerae 01 (El Tor) Ogawa and was the etiology of the acute acalculous cholecystitis in this patient.

Ugeskr Laeger, 1995 May 29, 157(22), 3202 - 4
{Severe systemic infection with Vibrio vulnificus}; Hansen LN et al.; Infections with Vibrio vulnificus are not common in Denmark, but in 1994 several cases were identified, probably due to the very hot weather conditions, with seawater temperatures above 20 degrees C . Two cases of infection with V . vulnificus are presented.

Gene, 1995 May 26, 158(1), 1 - 7
Putative O-antigen transport genes within the rfb region of Vibrio cholerae O1 are homologous to those for capsule transport; Manning PA et al.; The nucleotide sequence of that part of the Vibrio cholerae (Vc) O1 rfb region encompassing rfbG, rfbH and rfbI is presented . Expression of these genes has enabled the products for rfbG and rfbI to be confirmed, but the rfbH product has not been detected . Comparisons with the sequences of known proteins reveals that RfbH and RfbI are likely to be involved in the export of lipopolysaccharide (LPS) . RfbH shows considerable homology to a number of integral membrane proteins, some of which have been identified as possibly having a role as an export channel for capsular polysaccharides . RfbI corresponds to an ATP-binding protein usually found linked to the membrane protein and is thought to be required for energizing this export process . Thus, we propose that RfbH and RfbI form a complex for the export of Vc O1 LPS . The function of RfbG is unknown, but it would appear to be a relatively hydrophilic protein and we can only speculate that it may be either a specific transferase or possibly the O-antigen polymerase.

Biochemistry, 1995 May 23, 34(20), 6581 - 6
Three-dimensional structure of bacterial luciferase from Vibrio harveyi at 2.4 A resolution; Fisher AJ et al.; Luciferases are a class of enzymes that generate light in the visible spectrum . Luciferase from luminous marine bacteria is an alpha-beta heterodimer monooxygenase that catalyzes the oxidation of FMNH2 and a long-chain aliphatic aldehyde . The X-ray crystal structure of bacterial luciferase from Vibrio harveyi has been determined to 2.4 A resolution . The structure was solved by a combination of multiple isomorphous replacement and molecular averaging between the two heterodimers in the asymmetric unit . Each subunit folds into a (beta/alpha)8 barrel motif, and dimerization is mediated through a parallel four-helix bundle centered on a pseudo 2-fold axis that relates the structurally similar subunits . The vicinity of the active site has been identified on the alpha subunit by correlations with similar protein motifs and previous biochemical studies . The structure presented here represents the first molecular model of a bioluminescent enzyme.

FEMS Microbiol Lett, 1995 May 15, 128(3), 265 - 9
Cloning and sequencing of a novel hemolysis gene of Vibrio cholerae; Nagamune K et al.; A hemolysis gene (hlx) which lyses sheep erythrocytes on blood agar plates when expressed in Escherichia coli was cloned from Vibrio cholerae . The cloned gene is predicted to encode a polypeptide of 92 amino acid residues with a deduced molecular mass of 10,451 . E . coli transformed with this gene lysed sheep, goose, horse and chicken erythrocytes but not those of guinea pig and human . The hlx gene was observed in classical- and El Tor-biotype V . cholerae O1, V . cholerae non-O1, and V . mimicus, but not in V . parahaemolyticus.

Klin Lab Diagn, 1995 May-Jun, (3), 8 - 11
{Vibrios pathogenic to humans and laboratory diagnosis of diseases caused by them}; Smolikova LM et al.; Present-day taxonomic status of vibrios pathogenic for humans is discussed, as are the phenotypical characteristics of some species, factors of virulence, and clinical manifestations of diseases caused by these agents . A scheme of isolation and identification of vibrios pathogenic for humans is offered.

Med Microbiol Immunol (Berl), 1995 May, 184(1), 37 - 44
Characterization of Vibrio cholerae El Tor cytolysin as an oligomerizing pore-forming toxin; Zitzer A et al.; V . cholerae El Tor cytolysin is a secreted, water-soluble protein of M(r) 60,000 that may be relevant to the pathogenesis of acute diarrhea . In this communication, we demonstrate that the toxin binds to and oligomerizes in target membranes to form SDS-stable aggregates of M(r) 200,000-250,000 that generate small transmembrane pores . Pores formed in erythrocytes were approximately 0.7 nm in size, as demonstrated by osmotic protection experiments . Binding was shown to occur in a temperature-independent manner preceding the temperature-dependent oligomerization step . Pores were also shown to be formed in L929 and HEp-2 cells, human fibroblasts and keratinocytes, albeit with highly varying efficacy . At neutral pH and in the presence of serum, human fibroblasts were able to repair a limited number of lesions . The collective data identify V . cholerae El Tor cytolysin as an oligomerizing toxin that damages cells by creating small transmembrane pores.

FEMS Microbiol Lett, 1995 May 1, 128(2), 195 - 200
Utilization of hemin and hemoglobin as iron sources by Vibrio parahaemolyticus and identification of an iron-repressible hemin-binding protein; Yamamoto S et al.; Several clinical isolates of Vibrio parahaemolyticus were examined for their ability to utilize either hemin or hemoglobin as a sole source of iron . Both compounds appeared to be equally good iron sources . Maximum growth was obtained at 5 microM hemin or 1.25 microM hemoglobin under the conditions tested . Using a hemin-agarose batch affinity method, the hemin-binding protein was isolated from crude total membranes of a hemin-utilizing strain, WP1, grown under iron-deficient but not under iron-sufficient conditions . This protein was identical to the 83 kDa outer membrane protein which was expressed in response to iron limitation . The protein was susceptible to proteinase K cleavage in whole cells, indicating its exposure at the cell surface . Hemin and hemoglobin, but not protoporphyrin IX, inhibited binding of the protein to hemin-agarose.

South Med J, 1995 May, 88(5), 531 - 3
Vibrio vulnificus wound infections from the Mississippi Gulf coastal waters: June to August 1993; Penman AD et al.; Vibrio vulnificus, part of the normal marine flora of the Gulf of Mexico, is being increasingly recognized as an important human pathogen . V vulnificus contamination of superficial wounds can cause a severe, rapidly progressive, necrotizing cellulitis with bullous skin lesions that may require surgical debridement and is occasionally fatal . We summarize information about six cases of V vulnificus wound infection reported to the Mississippi State Department of Health from June to August 1993 . Five of the six patients required hospitalization for intravenous antibiotic treatment and, in two cases, surgery . Two patients died from septicemia, despite aggressive antibiotic treatment; both had preexisting medical conditions that could have contributed to immune compromise and fulminant infection . This report underscores the virulence of this organism and the need for awareness by both the clinician and diagnostic laboratory personnel when dealing with superficial wounds occupationally or recreationally exposed to seawater.

Infect Immun, 1995 May, 63(5), 1987 - 92
Purification and characterization of a cell-associated hemagglutinin of Vibrio parahaemolyticus; Nagayama K et al.; We found a positive correlation between cell-associated mannose-sensitive hemagglutination and adherence of Vibrio parahaemolyticus to rabbit enterocytes by investigating 35 strains of V . parahaemolyticus for cell-associated hemagglutinin (cHA) and for the ability to adhere to the enterocytes . We purified a mannose-sensitive cHA from a Kanagawa phenomenon-positive clinical strain of V . parahaemolyticus that exhibited a high level of mannose-sensitive hemagglutination and strongly adhered to the enterocytes . The purified cHA is a heat-labile, tetrameric protein consisting of four identical subunits of approximately 26 kDa each . The adherence to rabbit enterocytes was inhibited in a dose-dependent manner by pretreatment of the bacterial cells with D-mannose and with the Fab fraction of immunoglobulin G against the purified cHA . Furthermore, pretreatment of the enterocytes with the purified cHA inhibited the adherence of V . parahaemolyticus . Immunogold electron microscopy revealed that the cHA is located on the bacterial cell surface and is not associated with pili . These results suggest that cHA is involved in the adherence mechanisms of V . parahaemolyticus to the enterocytes and that the receptors for cHA on the enterocyte appear to be a D-mannose-containing compound.

J Exp Med, 1995 May 1, 181(5), 1693 - 703
Trypanosoma cruzi trans-sialidase: enhancement of virulence in a murine model of Chagas' disease; Chuenkova M et al.; Trypanosoma cruzi, the etiological agent of Chagas' disease, expresses a trans-sialidase at highest levels in infective trypomastigotes, where it attaches to the plasma membrane by a glycophosphoinositol linkage . Bound enzyme sheds into the extracellular milieu in a soluble form . Experiments performed in vitro suggest that the trans-sialidase participates in several parameters of T . cruzi-host interactions, like cell adhesion and complement resistance . However, the role that membrane-bound and soluble trans-sialidase plays in the infection of mammals is not understood . To begin to study the role the enzyme may play in vivo, T . cruzi trypomastigotes were inoculated subcutaneously into mice that had been sensitized for various times with the purified protein . A single dose of either endogenous or recombinant trans-sialidase injected into the connective tissues of BALB/c mice greatly enhanced parasitemia and mortality . Maximum enhancement was achieved with 1-2-h priming . Injection of the enzyme after the parasites had been established in the inoculation site had little, if any, consequence in modifying virulence . The enhancement did not seem to be through a direct effect of the enzyme on trypomastigote-host cell interactions because it occurred when the sites of trans-sialidase sensitization and parasite inoculation were physically separate . Rather, virulence enhancement seemed to depend on inflammatory cells, since priming with trans-sialidase had no significant effect in severe combined immunodeficiency mice, which lack functional T and B lymphocytes . However, antibody response to T . cruzi in the trans-sialidase-primed BALB/c mice was the same as in the control animals . Virulence enhancement was specific for the trans-sialidase because it did not occur in mice primed with Newcastle virus sialidase, which has the same substrate specificity as the T . cruzi enzyme, or with the sialidase from the bacterium Vibrio cholerae, whose substrate specificity is broader than the trypanosome sialidase . Furthermore, no enhancement of virulence occurred after sensitization with another adhesion protein (penetrin) purified from T . cruzi trypomastigotes and engineered bacteria, nor with bacterial lipopolysaccharide . The virulence-promoting activity of soluble trans-sialidase in the mouse model may be physiologically relevant because it was achieved with tiny doses, approximately 1-2 microgram/kg, raising the possibility that neutralization of the enzyme with specific probes could impair the development of Chagas' disease . In fact, a monoclonal antibody specific for the tandem repeat in the trans-sialidase COOH terminus enhanced infection of BALB/c mice, in agreement with earlier experiments in vitro, whereas antibodies against an amino acid sequence in the Cys region had the opposite effect.

Vaccine, 1995 May, 13(7), 691 - 4
Community-based assessment of safety and immunogenicity of the whole cell plus recombinant B subunit (WC/rBS) oral cholera vaccine in Peru; Begue RE et al.; Every year since its introduction in 1991, there have been epidemics of cholera in Lima, Peru . Vaccination is one approach to the control of cholera . A pilot study was conducted to assess the safety and immunogenicity of a whole cell plus recombinant B subunit (WC/rBS) cholrea vaccine in Lima, Peru . Five hundred and forty-one volunteers aged 2-65 years received two doses two weeks apart of WC/rBS vaccine or Escherichia coli K12 placebo administered in bicarbonate buffered water . Symptoms were monitored on all subjects and blood was collected from 102 persons before the first dose and two weeks after the second dose . Mild post-vaccination gastrointestinal symptoms were reported with equal frequency for both the vaccine and placebo recipients . Among 51 vaccines, 49% had a twofold or greater increase in serum vibriocidal titers (GMT = 78; range < 1:10 to 1:5120); and 92% and 82% developed a twofold or greater serum anti-cholera toxin IgG and IgA response, respectively . Persons with elevated prevaccination vibriocidal titers had a decreased response to the WC/rBS . Age and blood group did not affect the immune response . The WC/rBS vaccine was safe and immunogenic in a group of native Peruvians.

Toxicon, 1995 May, 33(5), 651 - 7
The binding of Vibrio parahaemolyticus 125I-labeled thermostable directhemolysin to erythrocytes; Yoh M et al.; Thermostable direct hemolysin (TDH) produced by Vibrio parahaemolyticus was iodinated using chloramine T . The 125I-labeled TDH retained up to 80% of the activity of intact toxin . The binding of 125I-TDH to rabbit erythrocytes was inhibited by addition of nonlabeled TDH . The binding of 125I-TDH to rabbit erythrocytes was completed in the 1st or 2nd min of incubation at 37 degrees C in contrast to that at 4 degrees C . 125I-TDH, which cannot lyse horse erythrocytes as does intact TDH, bound to horse erythrocytes as to those of rabbit . The dissociation constants (KD) derived Scatchard plots were 2.85, 4.39, 4.33 and 5.35 x 10-8M for rabbit, horse, human and sheep erythrocytes, respectively . The lytic sensitivity of various erythrocytes to TDH showed no relationship to the binding capacity.

Mikrobiol Z, 1995 May-Jun, 57(3), 56 - 64
{The reasons for the resistance of Vibrio cholerae to diagnostic phages}; Kudriakova TA et al.; Phage resistance of 225 strains of cholera germs of O1 group obtained from different countries in 1988-1992 has been analyzed . Change of sensitivity to diagnostic phages was mostly connected with the decrease or loss of agglutinability in cholera sera . Phage resistance is rather conditioned by the change of the surface structures of the cell and by further change of phage reception zones . The increase in the number of strains sensitive to diagnostic phages after 6-12 months of storage evidenced for stabilization of cell wall structures and increase of their viability under relatively favourable conditions of storage.

J Heart Lung Transplant, 1995 May-Jun, 14(3), 598 - 600
Vibrio vulnificus sepsis in solid organ transplantation: a medical nemesis; Ali A et al.; We report two cases of Vibrio vulnificus wound infection leading to fulminant sepsis syndrome in immunocompromised solid organ transplant recipients . Features of clinical presentation in each of these cases suggest that host immune factors are of great importance in the virulence of this organism and that immunocompromised recipients of solid organ transplants are particularly vulnerable to life-threatening consequences from infection with Vibrio vulnificus . Prompt institution of antibiotic therapy and early consideration for surgical wound debridement are the mainstay of successful management . Heart and other organ transplant recipients should be educated and warned about the hazards associated with raw oysters and shellfish consumption and asked to exercise caution when exposed to a salt water environment.

Zhonghua Yu Fang Yi Xue Za Zhi, 1995 May, 29(3), 138 - 40
{Studies on diagnostic bacteriophage of Vibrio fluvialis}; Chen K et al.; Species of Vibrio fluvialis are identified by their biochemical characteristics so far, with a lot of items to be determined, overelaborate procedure and expensiveness . Inaccordance with the fact that bacteriophage is a specific parasite inhabited in bacteria and has been used in identifying other bacteria with high specificity, some of Vibrio fluvialis bacteriophage isolated from natural environment were selected to make diagnostic preparation for Vibrio fluvialis identification . Diagnostic positivity of Vibrio fluvialis averaged 84.27%, and 87.84% for those of human source . Cross-lysis rates both for the same Vibrio genus and that of different families in the same genus were less then 3%, and no cross-lysis was found for the other enteric bacteria in different genera . There was no significant difference between diagnostic bacteriophage and biochemical tests for Vibrio of unknown species . This method was highly specific, sensitive, rapid, simple and inexpensive, and could be used in diagnosis of Vibrio fluvialis.

J Clin Microbiol, 1995 May, 33(5), 1339 - 40
Isolation of sucrose late-fermenting and nonfermenting variants of Vibrio cholerae O139 Bengal: implications for diagnosis of cholera; Ansaruzzaman M et al.; The sucrose-containing selective medium thiosulfate-citrate-bile salt-sucrose agar missed a sucrose nonfermenting and four sucrose late-fermenting variant strains of Vibrio cholerae O139 Bengal from diarrheal stools . These strains were, however, correctly identified as V . cholerae O139 on a sucrose-deficient selective medium, taurocholate-tellurite-gelatin agar.

Kansenshogaku Zasshi, 1995 May, 69(5), 501 - 5
{Molecular epidemiological study on Vibrio cholerae O139}; Gyobu Y et al.; Genomic DNA from 56 Vibrio cholerae O139 strains isolated in various countries was digested with Sfi I or Not I and analyzed by pulsed-field gel electrophoresis (PFGE) . Eight different PFGE patterns were identified . Although the patterns of a large majority of CT-gene-positive epidemic strains isolated in India, Bangladesh and Thailand were the same or similar, but were slightly different from those of two CT-positive strains from India and Nepal . On the other hand, the patterns of CT-negative three strains from Argentine, Sri Lanka and Bangladesh were apparently different not only from each other, but also from those of epidemic CT-positive strains . The pattern of one V . cholerae O1 E1 Tor strain isolated in India and the pattern of epidemic V . cholerae O139 resemble each other in many points.

Indian J Med Res, 1995 May, 101, 175 - 8
Haemagglutinating property & cell surface hydrophobicity of Vibrio cholerae 0139; Pal S et al.; Cell-associated haemagglutinating activity was detected in all the epidemic strains of V . cholerae 0139 and 01, isolated in different parts of the country, with erythrocytes from rabbit, rat, chicken and guinea pig . Sheep erythrocytes were unresponsive to both groups of strains . While D-mannose, alpha-methyl-D-mannoside, glucosamine, N-acetyl-D-glucosamine, thyroglobuline were effective inhibitors of the haemagglutinating activity of the V . cholerae 0139 and 01 strains, galactose and N-acetyl-D-galactosamine sensitive haemagglutinins (HAs) were detected in the V . cholerae 01 but not in the V . cholerae 0139 strains . The best medium for expression of HAs in V . cholerae 01 and 0139 strains were observed to be penassay broth and tryptic soy broth respectively . The expression of HA in V . cholerae 01 was stimulated by Ca++ and Fe in the growth medium unlike that of V . cholerae 0139 which remained unaffected by these ions.

Plasmid, 1995 May, 33(3), 180 - 90
Iron transport genes of the pJM1-mediated iron uptake system of Vibrio anguillarum are included in a transposonlike structure; Tolmasky ME et al.; The pJM1 genes encoding the proteins involved in iron transport in the anguibactin iron uptake system were found to be flanked by insertion sequences in a composite transposonlike structure . These Vibrio anguillarum insertion sequences, ISV-A1 and ISV-A2, are related to IS903, IS102, and the ISVs found in Vibrio parahaemoliticus, Vibrio mimicus, and non-O1 Vibrio cholerae flanking various tdh (thermostable direct hemolysin) genes . The inverted repeats at the ends of ISV-A1 and ISV-A2 have no more than three mismatches when compared to the inverted repeats of the other ISVs or IS903 and IS102 . ISV-A1 and ISV-A2 are flanked by 9-bp direct repeats, which is the number of bases that are duplicated upon IS903 or IS102 transposition . The similarities found between the V . anguillarum ISVs and the other ISVs as well as IS903 and IS102 suggest that they derive from a common ancestral insertion sequence . At the end of ISV-A1 there is a -35 sequence region followed by a -10 sequence found in the pJM1 sequence immediately outside the ISV . This promoter region is followed by an open reading frame with the potential to encode a polypeptide of 26,985 Da whose function is still unknown . The functionality of this promoter has been demonstrated and expression analysis showed that the promoter is regulated by the iron concentration of the media.

Mol Microbiol, 1995 May, 16(3), 425 - 39
Organization of tcp, acf, and toxT genes within a ToxT-dependent operon; Brown RC et al.; The toxin coregulated pilus (TCP) is required for Vibrio cholerae to colonize the human intestine . The expression of the pilin gene, tcpA, is dependent upon ToxR and upon ToxT . The toxT gene was recently mapped within the TCP biogenesis gene cluster and shown to be capable of activating a tcpA::TnphoA fusion when cloned in Escherichia coli . In this study, we determined that ToxR/ToxT activation occurs at the level of tcpA transcription . ToxT expressed in E . coli could activate a tcp operon fusion, while ToxR, ToxR with ToxS, or a ToxR-PhoA fusion failed to activate the tcp operon fusion and we could not demonstrate binding of a ToxR extract to the tcpA promoter region in DNA mobility-shift assays . The start site for the regulated promoter was shown by primer extension to lie 75 bp upstream of the first codon of tcpA . An 800-base tcpA message was identified, by Northern analysis, that correlates by size to the distance between the transcriptional start and a hairpin-loop sequence between tcpA and tcpB . The more-sensitive assay of RNase protection analysis demonstrated that a regulated transcript probably extends through the rest of the downstream tcp genes, including toxT and the adjacent accessory colonization factor (acf) genes . An in-frame tcpA deletion, but not a polar tcpA::TnphoA fusion, could be complemented for pilus surface expression by providing tcpA in trans . This evidence suggests that the tcp genes, including toxT, are organized in an operon directly activated by ToxT in a ToxR-dependent manner . Most of the toxT expression under induced conditions requires transcription of the tcpA promoter . Further investigation of how tcp::TnphoA insertions that are polar on toxT expression retain regulation showed that a low basal level of toxT expression is present in toxR and tcp::TnphoA strains . Overall, these observations support the ToxR/ToxT cascade of regulation for tcp . Once induced, toxT expression becomes autoregulatory via the tcp promoter, linking tcp expression to that of additional colonization factors, exotoxin production, and genes of unknown function in cholera pathogenesis.

Mol Microbiol, 1995 May, 16(4), 801 - 11
Characterization of heat-shock response of the marine bacterium Vibrio harveyi; Klein G et al.; We have investigated heat-shock response in a marine bacterium Vibrio harveyi . We have found that 39 degrees C was the highest temperature at which V . harveyi was able to grow steadily . A shift from 30 degrees C to 39 degrees C caused increased synthesis of at least 10 proteins, as judged by SDS-PAGE, with molecular masses of 90, 70, 58, 41, 31, 27, 22, 15, 14.5 and 14kDa . The 70, 58, 41 and 14.5 kDa proteins were immunologically homologous to DnaK, GroEL, DnaJ and GroES heat-shock proteins of Escherichia coli, respectively . V . harveyi GroES protein had a lower molecular mass (14.5 kDa) than E . coli GroES, migrating in SDS-PAGE as 15kDa protein . We showed that a protein of approximately 43 kDa, immunologically reactive with antiserum against E . coli sigma 32 subunit (sigma 32) of RNA polymerase, was induced by heat-shock and co-purified with V . harveyi RNA polymerase . These results suggest that the 43 kDa protein is a heat-shock sigma protein of V . harveyi . Preparation containing the V . harveyi sigma 32 homologue, supplemented with core RNA polymerase of E . coli, was able to transcribe heat-shock promoters of E . coli in vitro.

Carbohydr Res, 1995 Apr 30, 270(2), 115 - 22
Circular dichroism of the O-specific polysaccharide of Vibrio cholerae O1 and some related derivatives; Bystricky S et al.; The O-specific polysaccharide (O-SP) of Vibrio cholerae O1 is a homopolymer of alpha-(1 --> 2)-linked 4-amino-4, 6-dideoxy-D-mannopyranose whose amino group is acylated with 3-deoxy-L-glycero-tetronic acid {N-(3-deoxy-L-glycero- tetronyl)-alpha-D-perosamine} . The circular dichroism (CD) of the O-SP as well as of a number of N-acyl (formyl, acetyl, 4-hydroxybutyl, 3-deoxy-L-and D-glycero-tetronyl) derivatives of methyl alpha-glycosides of 4-amino-4,6-dideoxy-D-mannopyranose (methyl alpha-D-perosaminide) has been studied for solutions in water, acetonitrile and 1,1,1-trifluoroethanol . The strong solvent dependence of the sign and intensity of the CD observed for the monosaccharide amides bearing achiral acyl groups is explained by solvent-mediated change of the orientation of the amido group relative to the proximal hydroxyl group at C-3 . A change in the population of the nonplanar conformers with a pyramidal arrangement of bonds at the amido nitrogen has also been considered . The effect of solvents upon the CD spectra of compounds bearing chiral N-acyl substituents is less pronounced than that of their counterparts bearing achiral N-acyl substituents . The sign of the CD for the O-SP was found negative in all solvents used . This result is in agreement with the negative sign of the CD of the n --> pi electron transition observed, independent of the solvent, for the monosaccharide derivative containing the L-glycero-3-deoxytetronamido group, and the positive sign found for its D-glycero-counterpart.

EMBO J, 1995 Apr 18, 14(8), 1664 - 73
Interaction between the autokinase EpsE and EpsL in the cytoplasmic membrane is required for extracellular secretion in Vibrio cholerae; Sandkvist M et al.; Vibrio cholerae secretes a number of proteins important for virulence, including cholera toxin . This process requires the products of the eps genes which have homologues in genera such as Aeromonas, Klebsiella and Pseudomonas and are thought to form a membrane-associated multiprotein complex . Here we show that the putative nucleotide-binding protein EpsE is associated with and stabilized by the cytoplasmic membrane via interaction with EpsL . Analysis of fusion proteins between EpsE and the homologous ExeE from Aeromonas hydrophila demonstrates that the N-terminus of EpsE contains the EpsL binding domain and determines species specificity . An intact Walker A box, commonly found in ATP-binding proteins, is required for activity of EpsE in vivo and for autophosphorylation of purified EpsE in vitro . These results indicate that both the kinase activity of EpsE as well as its ability to interact with the putative cytoplasmic membrane protein EpsL are required for translocation of toxin across the outer membrane in Vibrio cholerae.

FEBS Lett, 1995 Apr 17, 363(1-2), 75 - 7
Sequencing and the alignment of structural genes in the nqr operon encoding the Na(+)-translocating NADH-quinone reductase from Vibrio alginolyticus; Hayashi M et al.; We previously cloned a part of nqr operon encoding the Na(+)-translocating NADH-quinone reductase (NQR) from the marine Vibrio alginolyticus {Hayashi et al., FEBS Lett . 356 (1994) 330-332} . From its nucleotide sequences, four consecutive open reading frames (ORF) encoding the gamma-subunit (27.7 kDa), two unidentified ORFs of 22.6 kDa and 21.5 kDa, and the beta-subunit (45.3 kDa) were recognized . The gene encoding the alpha-subunit was located upstream, and together with the recent report by Beattie et al . {FEBS Lett . 356 (1994) 333-338}, the nqr operon was found to be constructed from six consecutive structural genes, where nqr1, nqr3 and nqr6 correspond to the alpha-, gamma-, and beta-subunits, respectively, of the NQR complex.

Lancet, 1995 Apr 15, 345(8955), 949 - 52
Safety, immunogenicity, and efficacy of live attenuated Vibrio cholerae O139 vaccine prototype; Coster TS et al.; New vaccines are needed to prevent cholera caused by Vibrio cholerae O139 . Attenuated V cholerae O139 vaccines were made by deleting multiple copies of the cholera-toxin genetic element from two virulent strains of the organism, MO10 and AI4456 . The deletion mutants were further modified by insertion of a construct that encoded the B subunit of cholera toxin, thus generating strains Bengal-3 and VRI-16 . A stable spontaneous non-motile derivative of Bengal-3 was isolated and designated Bengal-15; VRI-16 is naturally non-motile . Bengal-3, Bengal-15, and VRI-16 were evaluated as oral single-dose cholera vaccine candidates in 4 volunteers each, and MO10 was given to 3 volunteers . 1 of 4 volunteers who received Bengal-3 and all 3 who received MO10 had diarrhoea . VRI-16 caused no significant symptoms but was not immunogenic . Bengal-15 produced few symptoms and was nearly as immunogenic as MO10 . Subsequently, Bengal-15 was given to 10 volunteers at a dose of 10(8) colony-forming units . No volunteers had diarrhoea, and other subjective symptoms were as common in vaccinees as in 3 buffer recipients . 1 month after vaccination, 7 vaccinees, the 3 buffer recipients, and 3 unimmunised subjects were challenged with 5 x 10(6) colony-forming units of V cholerae O139 . 5 of 6 controls had cholera-like diarrhoea . By contrast, 1 of 7 vaccinees had diarrhoea, which was mild and had a long incubation period . Vaccine protective efficacy was 83% . Our results indicate the Bengal-15 is a safe live attenuated vaccine candidate for cholera caused by the O139 serogroup.

Eur J Biochem, 1995 Apr 15, 229(2), 583 - 8
Immunochemistry of group A and Inaba C antigen factors constituting the O antigen of O1 Vibrio cholerae; Isshiki Y et al.; Serological cross-reactivity among intact lipopolysaccharides (LPS) from O1 Vibrio cholerae Inaba O-form (Inaba), Yersinia enterocolitica O9 (O9), non-O1 V . cholerae serogroup Hakata (Hakata) and Vibrio bio-serogroup 1875 Variant (1875 Variant) (all of which share Inaba antigen factor C), as well as a total of six kinds of chemically modified LPS (three from O9 and three from Inaba) was demonstrated by passive hemolysis and passive hemolysis inhibition by using these LPS as antigen for sensitizing sheep red blood cells and as inhibitor . These intact as well as chemically modified LPS contained, in their O polysaccharide chain, alpha(1-->2)-linked linear perosamine (4-amino-4,6-dideoxy-D-manno-pyranose) homopolymers with different N-acyl groups: their acyl groups comprise 3-deoxy-L-glycero-tetronyl (Inaba LPS), formyl (O9 LPS), 3-hydroxypropionyl (1875 Variant LPS), acetyl (Hakata LPS and artificially introduced into Inaba and O9 LPS), propionyl and butyryl (both artificially introduced into Inaba and O9 LPS) groups . N-Deacylation of the alpha(1-->2)-linked N-(3-deoxy-L-glycero-tetronyl)perosamine homopolymer of Inaba and the N-formyl one of O9 LPS resulted in virtual elimination of their serological reactivity with both homologous and heterologous antisera . Furthermore, when the resultant NH2 groups of the N-deacylated perosamine homopolymers of both LPS were N-acylated with acetyl, propionyl or butyryl groups, they markedly recovered both of their serological reactivities . These results are compatible with the interpretation that the Inaba antigen factor C possessed by the four bacteria is substantially related to the common presence of N-acyl groups, regardless of their identity, residing in the perosamine residues constituting the O polysaccharide chain of their LPS . It was also indicated that the group antigen factor A of O1 V . cholerae is substantially related to the 3-deoxy-L-glycero-tetronyl groups residing in the perosamine homopolymer of Inaba LPS.

Gene, 1995 Apr 14, 156(1), 59 - 61
Isolation and characterization of the Vibrio cholerae acfA gene, required for efficient intestinal colonization; Hughes KJ et al.; The nucleotide sequence of the Vibrio cholerae acfA gene (encoding an accessory colonization factor) has been determined . Sequence analysis revealed the presence of an open reading frame of 215 amino acids with a characteristic signal peptidase I (SPI) cleavage site at the N terminus . Electrophoretic analysis of proteins synthesized by Escherichia coli cells, following T7 promoter/RNA polymerase-directed expression of acfA, revealed a 23-kDa protein corresponding to the mature form of AcfA . The T7 expression system also showed that, in the presence of known SPI inhibitors, a 25-kDa unprocessed form of AcfA is produced.

Lett Appl Microbiol, 1995 Apr, 20(4), 225 - 7
Effect of chill and freezing temperatures on survival of Vibrio parahaemolyticus inoculated in homogenates of oyster meat; Muntada-Garriga JM et al.; Survival of Vibrio parahaemolyticus was determined in oyster meat homogenates at various temperatures . (4 degrees C, 0 degrees C, -18 degrees C and -24 degrees C) and bacterial levels (10(2), 10(4), 10(5) and 10(7) ml-1) . In all cases, the numbers of V . parahaemolyticus were a logarithmic function of log time . This study indicates that high numbers of V . parahaemolyticus can be inactivated at low temperatures . The time of total inactivation depends on the initial number of micro-organisms and incubation temperature . It is possible to use this information to determine the storage time necessary to reduce V . parahaemolyticus hazards in fish.

Appl Environ Microbiol, 1995 Apr, 61(4), 1620 - 2
Homogentisic acid is the primary precursor of melanin synthesis in Vibrio cholerae, a Hyphomonas strain, and Shewanella colwelliana; Kotob SI et al.; The enzyme p-hydroxyphenylpyruvate hydroxylase (HPPH) is involved in pigmentation (pyomelanin) via homogentisic acid (HGA) . Pyomelanin formation is correlated with HGA production and expression of HPPH in three disparate marine species: Vibrio cholerae, a Hyphomonas strain, and Shewanella colwelliana . Induction of pigmentation in V . cholerae 569B by nutrient limitation also correlated with production of HGA.

Appl Environ Microbiol, 1995 Apr, 61(4), 1311 - 7
Sequence of a cloned pR72H fragment and its use for detection of Vibrio parahaemolyticus in shellfish with the PCR; Lee CY et al.; The nucleotide sequence of pR72H cloned from Vibrio parahaemolyticus 93 was determined . We examined all V . parahaemolyticus gene sequences published in the GenBank-EMBL databases for homology and found that no other DNA sequence of V . parahaemolyticus was highly homologous to the sequence reported in this study . A pair of primers, VP33-VP32, derived from a pR72H fragment were selected to detect V . parahaemolyticus . The sensitivity of PCR detection for a pure culture of V . parahaemolyticus was 10 cells from crude bacterial lysates . Furthermore, a detection level of 2.6 fg, equivalent to 1 cell, was obtained by using purified chromosomal DNA as the template . The expected PCR products were obtained from all V . parahaemolyticus strains tested (n = 124), while no PCR amplicons were found in other vibrios or related genera (n = 50) . High levels (10(6) to 10(10) CFU/ml) of Escherichia coli cells did not affect the PCR assay sensitivity . The presence of 10(8) V . parahaemolyticus cells or 10(9) E . coli cells in the PCR mixtures completely inhibited the PCR . When oyster samples were inoculated with V . parahaemolyticus 93 and cultured in tryptic soy broth containing 3% NaCl for 3 h at 35 degrees C, an initial sample inoculum level of 9.3 CFU/g was detected in a PCR assay with crude bacterial lysates . The PCR assay with enrichment culturing in salt polymyxin broth was compared with the conventional method for naturally contaminated shellfish and fish samples . We conclude that this PCR assay with enrichment culturing is a good alternative method for the detection of V . parahaemolyticus.

J Appl Bacteriol, 1995 Apr, 78(4), 387 - 93
Possible interference of lactose-fermenting marine vibrios in coliform beta-D-galactosidase assays; Davies CM et al.; An investigation into possible interferences in beta-D-galactosidase-based assays for coliform bacteria in marine waters was carried out . A rapid instrumental fluorescence assay for beta-D-galactosidase activity, using 4-methylumbelliferyl-beta-D-galactoside as a substrate, was used to investigate activities of this enzyme in non-coliform bacterial isolates from coastal waters . Only 2% of isolates showed slight enzyme activity after a 1-h incubation period at 44.5 degrees C . At a lower incubation temperature of 20 degrees C, 51% and 94% of the isolates showed some enzyme activity within 6 h and 48 h, respectively . Fifty-nine out of 67 of these isolates were identified as Vibrio species . A lac+ strain of Vibrio vulnificus was found to produce beta-D-galactosidase which caused significant false-positive reactions in the Colilert-Marine Water assay when present at concentrations of 10 cfu ml-1 or greater . This interference could be overcome by addition of the vibriostatic agent O/129 . The high fluorescence of this reagent, however, precluded the simultaneous determination of Escherichia coli in the Colilert test and also its use in instrumental fluorescence assays . It was concluded that in assays employing high temperatures and short incubation times, Vibrio species are unlikely to cause significant interferences.

J Med Microbiol, 1995 Apr, 42(4), 251 - 7
A comparative study of the properties of Vibrio cholerae O139, O1 and other non-O1 strains; Nandy RK et al.; Vibrio cholerae O139 organisms isolated from different parts of India and from Bangladesh were characterised with respect to their haemagglutination (HA) activity, plasmid content, cholera toxin (CT) production, cell surface protein and lipopolysaccharide (LPS) profiles, and antigenic properties . Of 28 V . cholerae O139 isolates tested, 14 (50%) were shown to agglutinate chicken erythrocytes; the HA activity was sensitive to D-mannose 0.1% . In parallel experiments, 12 (92.3%) of 13 V . cholerae O1 (El Tor) and 12 (75%) of 16 non-O1, non-O139 strains agglutinated chicken erythrocytes . Plasmid analysis of 32 O139 isolates showed that 12 (37.5%) carried one or more plasmids of 35.8-2.6 MDa . Plasmids were not detected in any of the V . cholerae O1 strains, although plasmids were demonstrable in 35% of the non-O1, non-O139 strains tested . V . cholerae O139 isolates showed an ability to produce CT that depended on media composition and other cultural conditions . A comparison of envelope and outer-membrane protein profiles between O1 and O139 isolates failed to show any significant differences . LPS analysis of O139 isolates revealed that these organisms were devoid of long "O" side-chain polysaccharides . Some of the non-O1, non-O139 strains also showed similar LPS profiles whereas others showed the presence of long repetitive "O" side-chain polysaccharides similar to those seen in O1 organisms . An antiserum raised against V . cholerae O1 strain O395 did not show any significant reactivity towards O139 and non-O1, non-O139 strains although it reacted with other O1 strains . Furthermore, the anti-O1 serum induced marked protection against challenge with an O1 strain but not with an O139 strain in passive protection experiments.(ABSTRACT TRUNCATED AT 250 WORDS)

J Infect Dis, 1995 Apr, 171(4), 903 - 8
Clinical and immunologic characteristics of Vibrio cholerae O139 Bengal infection in North American volunteers; Morris JG Jr et al.; Vibrio cholerae O139 Bengal has recently emerged as a cause of epidemic cholera in Asia . To evaluate clinical and immunologic responses to infection, V . cholerae O139 Bengal AI1837 was administered to healthy adult North American volunteers . Two of 4 persons ingesting 10(4) cfu became ill (incubation period, 48 h; mean diarrheal stool, 1873 g), as did 7 of 9 persons receiving 10(6) cfu (incubation period, 28 h; mean diarrheal stool, 4548 g) . Ill volunteers did not demonstrate a vibriocidal antibody response to the challenge strain or other V . cholerae . Three months later, volunteers were rechallenged with the homologous O139 Bengal strain . Only 1 of 6 persons who had been ill on initial challenge had diarrhea, compared with 11 of 13 controls (P = .01; protective efficacy = 80%) . V . cholerae O139 Bengal can cause severe diarrhea typical of cholera, with clinical characteristics and a dose-response similar to those seen with V . cholerae O1 El Tor . A moderately high level of protection against subsequent disease is provided by initial clinical infection.

J Med Assoc Thai, 1995 Apr, 78(4), 204 - 9
Pathologic changes of gut in non-01 Vibrio cholerae infection; Shuangshoti S et al.; A 14-year-old girl who had beta-thalassemia hemoglobin E disease was infected by bacteriologically proven non-01 Vibrio cholerae at 2 months postsplenectomy and died 37 hours after onset of the malady . Postmortem examination disclosed congestion, edema, and hemorrhagic foci of the mucosa of the small and large intestines . The gut mucosa was focally eroded . The gut wall was infiltrated by leucocytes, especially neutrophils, in all coats representing acute purulent and hemorrhagic enterocolitis . There was hyperplasia of lymphoid follicles in the gut mucosa and lymph nodes . It is suggested that morphologic change of the gut in non-01 Vibrio cholerae infection is more severe than in infection caused by Vibrio cholerae.

FEMS Immunol Med Microbiol, 1995 Apr, 11(2), 87 - 90
Vibrio cholerae O139 produces a protease which is indistinguishable from the haemagglutinin/protease of Vibrio cholerae O1 and non-O1; Naka A et al.; Haemaglutinin/protease (HA/P) is one of the virulence factors of Vibrio cholerae O1 and pathogenic strains of V . cholerae non-O1 . In this study, we examined protease activity of a new serogroup of Vibrio cholerae recently designated as O139 synonym Bengal . The protease activity was produced by all eight isolates of V . cholerae O139 from Bangladeshi patients . Purification and partial characterization of the protease from V . cholerae O139 demonstrated the purified protease (O139-P) was indistinguishable from that previously reported for HA/P of V . cholerae non-O1 (NAG-HA/P) and V . cholerae O1 (Vc-HA/P) . These results prove that V . cholerae O139 produces a protease belonging to HA/P, and suggest that the protease is another virulence factor found in newly emerged V . cholerae O139, as in V . cholerae O1.

FEMS Immunol Med Microbiol, 1995 Apr, 11(2), 131 - 6
Emergence of tetracycline resistance due to a multiple drug resistance plasmid in Vibrio cholerae O139; Yamamoto T et al.; Of the 173 clinical strains of Vibrio cholerae O139 isolated from India, Bangladesh, and Thailand tested, six strains from India were resistant to tetracycline, ampicillin, chloramphenicol, kanamycin, and gentamicin . These six strains harbored a self-transmissible plasmid that mediated resistance to tetracycline, ampicillin, chloramphenicol, kanamycin, gentamicin, sulfamethoxazole, trimethoprim, and O/129 . The multiple drug resistance plasmids were 200 kb in size and belonged to the incompatibility group C . Although a majority of the O139 strains (94.8%) were highly resistant to streptomycin, sulfamethoxazole, trimethoprim, and O/129, the tetracycline-susceptible strains so far tested were plasmid-negative . The data suggest the existence of two distinct multiple antimicrobial agent resistance (MAR) patterns in V . cholerae O139.

J Pak Med Assoc, 1995 Apr, 45(4), 91 - 3
Cholera admissions in adults 1989-1994: a hospital based study; Abbas Z et al.; In order to gain insight into the distribution of cholera over the years and proportion of monthly admissions under our adult medical services, we scrutinized our records of hospital discharges between 1989 and 1994 . Only culture positive cases were included . Each year most of the cases of cholera are admitted between May and November with almost disease free interval from December to April . In 1992 admission rate was 4.24/1000 medical admissions which increased to 12.65 in 1993 and 13.73 in 1994 . Though the Vibrio cholerae 01 Ogawa was the major isolate upto May, 1993, Vibrio cholerae non-01 serogroup 0139 dominated between June and August, 1993 . Ogawa strain re-established itself in October, 1993 . In August, 1994, non-01 strain reappeared and became the major isolate in September . Cholera has caused multiple epidemics throughout the Indian subcontinent . Since 1800, there have been seven pandemics of cholera . The seventh pandemic originated in Indonesia and continues today.

Mol Cell Probes, 1995 Apr, 9(2), 75 - 81
Effect of dilution, incubation time, and temperature of enrichment on cultural and PCR detection of Vibrio cholerae obtained from the oyster Crassostrea virginica; DePaola A et al.; The recovery of Vibrio cholerae 01 by culture from the oyster Crassostrea virginica and detection of the cholera toxin gene by polymerase chain reaction were evaluated using various enrichment procedures in alkaline peptone water . The effects of dilutions (1:10 and 1:100), incubation times (6-8 and 18-21 h), and incubation temperatures (35 and 42 degree) were determined . Recovery of V.cholerae was significantly greater (P<0.05) from oyster homogenates diluted 1:100 in alkaline peptone water and incubated at 42 degree C for 18-21 h . This enrichment procedure also provided the best detection of the cholera toxin gene by polymerase chain reaction.

J Bacteriol, 1995 Apr, 177(8), 2138 - 42
Differential expression in Escherichia coli of the Vibrio sp . strain ABE-1 icdI and icdII genes encoding structurally different isocitrate dehydrogenase isozymes; Suzuki M et al.; The expression of two structurally different isocitrate dehydrogenase isozymes of Vibrio sp . strain ABE-1 in Escherichia coli was examined . At a low temperature (15 degrees C), a thermolabile and monomeric type isozyme (IDH-II), which is quite different in amino acid sequence from the E . coli isocitrate dehydrogenase, was expressed and conferred glutamate prototrophic ability on an E . coli mutant defective in isocitrate dehydrogenase . The ability of IDH-II to confer restoration of the E . coli mutant to glutamate prototrophy was similar to that of IDH-I, which is a dimeric enzyme homologous to the E . coli isocitrate dehydrogenase . At a high temperature (37 degrees C), no functional IDH-II was expressed . Transcription of icdI and icdII genes, which encode IDH-I and IDH-II, respectively, was regulated differently by different environmental conditions . The level of icdII mRNA was increased by lowering the growth temperature for E . coli transformants, while the level of icdI mRNA was increased when E . coli transformants were cultured in acetate minimal medium . Similar patterns of transcriptional regulation of the two icd gene were observed also in Vibrio sp . strain ABE-1 . However, activity of isocitrate dehydrogenase kinase, which can phosphorylate IDH-I and consequently inactivate the enzymatic activity, was detected in cell lysates of E . coli but not of Vibrio sp . strain ABE-1.

J Bacteriol, 1995 Apr, 177(8), 2080 - 6
Bacterial luciferase as a reporter of circadian gene expression in cyanobacteria; Liu Y et al.; To allow continuous monitoring of the circadian clock in cyanobacteria, we previously created a reporter strain (AMC149) of Synechococcus sp . strain PCC 7942 in which the promoter of the psbAI gene was fused to Vibrio harveyi luciferase structural genes (luxAB) and integrated into the chromosome . Northern (RNA) hybridization and immunoblot analyses were performed to examine changes in abundance of the luxAB mRNA, the native psbAI mRNA, and the luciferase protein to determine whether bioluminescence is an accurate reporter of psbAI promoter activity in AMC149 . Under constant light conditions, the mRNA abundances of both luxAB and psbAI oscillated with a period of approximately 24 h for at least 2 days . The expression of these two genes following the same pattern: both mRNAs peaked in the subjective morning, and their troughs occurred near the end of the subjective night . The amount of luciferase protein also oscillated with a period of approximately 24 h, and the protein rhythm is in phase with the bioluminescence rhythm . The rhythm of the luciferase mRNA phase-leads the rhythms of luciferase protein and in vivo bioluminescence by several hours . Comparable results were obtained with a short-period mutant of AMC149 . Together, these results indicate that the bioluminescence rhythm in AMC149 is due primarily to circadian oscillation of psbAI promoter activity in this cyanobacterium.

Biochem Biophys Res Commun, 1995 Mar 28, 208(3), 927 - 34
Sequence alignment and structural modelling of the LamB glycoporin family; Lang H et al.; lamB gene segments were obtained from Yersinia enterocolitica and Vibrio parahaemolyticus by the PCR and the DNA sequence determined . The deduced polypeptide sequences showed high similarity to six other LamB-related proteins and all contained typical signature sequences present in all members of the family but not other proteins . The aligned amino acid sequences permitted derivation of a model of LamB folding across the bacterial outer membrane using an approach successfully applied in the identification of structural features in other porins (Ferenci,T . (1994) Mol . Microbiol . 14:188-189) . The alignment-based model differs from previous LamB structure predictions and is also more complex than that found for OmpF-related porins; more than 16 conserved stretches of amino acid sequence potentially corresponded to membrane-spanning segments.

MMWR Morb Mortal Wkly Rep, 1995 Mar 24, 44(11), 215 - 9
Update: Vibrio cholerae O1--Western Hemisphere, 1991-1994, and V . cholerae O139--Asia, 1994; In Vibrio cholerae serogroup O1 et al.; Department of Microbiology and Immunology, University of Adelaide, AustraliaThe rfaD gene of Escherichia coli encodes ADP-L-glycero-D-mannoheptose-6- epimerase, an enzyme required for the biosynthesis of the lipopolysaccharide (LPS) precursor ADP-L-glycero-D- mannoheptose, associated with production of the core oligosaccharide . We have identified an rfaD homologue in Vibrio cholerae O1 . This gene maps adjacent to the rfb region encoding O-antigen biosynthesis, but is transcribed divergently . The complete nucleotide sequence of rfaD and the flanking DNA has been determined, and rfaD would appear to be the only gene homologous to known LPS core biosynthesis genes in this region . Comparison with the E . coli rfaD shows many similar structural features such as the ADP-binding beta alpha beta fold at the N terminus, as well as a high degree of homology of both the nucleotide and amino-acid sequences . Based on homology, rfaD of V . cholerae may be transcribed using both sigma 70- and sigma 54-dependent promoters.

FEMS Microbiol Lett, 1995 Mar 15, 127(1-2), 111 - 5
Detection of heat-stable enterotoxin genes among Australian Vibrio cholerae O1 strains; Mallard KE et al.; DNA probes derived from the heat-stable enterotoxin gene of Vibrio cholerae non-O1 (stn), and the cholera toxin gene (ctx), were used to screen 199 strains of V . cholerae O1, which were isolated within Australia from 1977-1986 . 13 environmental strains isolated from the riverine environment in Southeast Queensland in 1980 and 1981, hybridized with the stn and ctx DNA probes . The concentrated supernatant of 6 of these strains elicited fluid accumulation in the infant mouse assay both before and after heating at 100 degrees C for 5 min . Genetic relationships among the 13 stn+ strains were studied by a comparison of the rRNA-RFLPs (ribotyping) and by Southern blot analysis with a stn gene probe . The results indicate that there is a clonal relationship among the Australian stn+ strains and that there is an environmental reservoir of stn genes among Australian V . cholerae O1 isolates.

Braz J Med Biol Res, 1995 Mar, 28(3), 323 - 5
Protective effect of Saccharomyces boulardii against the cholera toxin in rats; Dias RS et al.; The effect of orogastric administration of Saccharomyces boulardii on the anatomopathological aspect of the jejunal villi was studied in male Fischer rats (weighing about 40 g) orogastrically infected with a culture of Vibrio cholerae . Experimental and control groups received lyophilized S . boulardii (25 mg suspended in 0.5 ml saline) or 0.5 ml saline, respectively, three times a day for 10 days by gastric intubation . On day 5 of treatment, 0.5 ml of a culture of V . cholerae containing 10(8) viable cells was inoculated by gastric intubation into both groups . Histopathological examination of the jejunal mucosa showed extensive lesions of the superficial epithelium of the villi from the control group whereas few lesions of this superficial epithelium were observed in the experimental group . These data show that the inhibition of the action of the cholera toxin on enterocytes by S . boulardii suggested by recent results in vitro can be demonstrated in vivo.

Mutat Res, 1995 Mar, 327(1-2), 5 - 15
Inter-strand cross-linking of Vibrio cholerae DNA induced by furazolidone: a quantitative assay by four simple methods; Basak J; Four simple methods, i.e., (i) UV absorption spectrophotometry, (ii) hydroxyapatite chromatography, (iii) fluorescence analysis of ethidium bromide bound to DNA and (iv) assay of S1 endonuclease action, were used in parallel for the estimation of furazolidone-induced inter-strand cross-links in Vibrio cholerae DNA . The data produced by the four methods were in reasonable agreement with each other and provided similar linear dose-response relations, the correlation (between dose and response) coefficient being in any case numerically greater than 0.98 . When the data obtained by four independent methods were plotted in a single graph, the resulting dose-response relation could be described by the equation log NR = 1.41 - 0.54 log D, where NR is the % non-reversible DNA remaining in the cells treated by furazolidone at dose D micrograms/ml x h . The correlation coefficient in this plot was -0.98 and significant to a level better than 0.1% . This study thus brings out that any one of these four methods can be used with reasonable confidence for the diagnosis and assay of inter-strand cross-links in DNA.

Appl Environ Microbiol, 1995 Mar, 61(3), 1163 - 8
Multiple Vibrio vulnificus strains in oysters as demonstrated by clamped homogeneous electric field gel electrophoresis; Buchrieser C et al.; Clamped homogeneous electric field gel electrophoresis and a computer program for managing electrophoresis banding patterns (ELBAMAP) were used to analyze genomic DNA of 118 Vibrio vulnificus strains, isolated from three oysters by direct plating . Analysis with SfiI resulted in 60 restriction endonuclease digestion profiles (REDP), while analysis with SrfI produced 53 different REDP . Similarities between REDP ranged from 7 to 93% . Principal-component analysis showed that the strains were heterogeneous.

Appl Environ Microbiol, 1995 Mar, 61(3), 1133 - 7
Evidence that water transmits Vibrio vulnificus biotype 2 infections to eels; Amaro C et al.; Vibrio vulnificus biotype 2 is classically considered an obligate eel pathogen . However, it has recently been associated with one human septicemic case . In this paper, the opportunistic behavior of this pathogen is discussed . The bacterium can survive alone in brackish water or attached to eel surfaces for at least 14 days . It is able to spread through water and infect healthy eels by using skin as a portal of entry . These results suggest that water and infected eels may act as reservoirs of infection . A capsule seems to be essential for waterborne infectivity, which would explain why cells recovered from naturally diseased eels give rise to pure cultures of opaque colonies . The spread of the disease is dependent on temperature and water salinity, thus suggesting a method to reduce the risk of epizootics and that of infection for humans.

Trans R Soc Trop Med Hyg, 1995 Mar-Apr, 89(2), 175 - 7
Effect of iron and pH on the survival of Vibrio cholerae in water; Patel M et al.; Many physicochemical factors affect the survival of Vibrio cholerae in the aquatic environment . An attempt was made to study the combined effect of pH and iron on the survival of V . cholerae in water in a laboratory environment . None of the 6 strains of V . cholerae used survived at pH 5.0; survival of all strains increased with increasing pH . The effect of ferric oxide on survival was significant for V . cholerae O1 only, not for non O1 strains . The longest survival of V . cholerae non O1 was 82 d, of El Tor V . cholerae 68 d, and of classical V . cholerae 56 d.

Zh Mikrobiol Epidemiol Immunobiol, 1995 Mar-Apr, Suppl 2, 97 - 101
{A case of the importation onto the territory of Russia of cholera caused by a new serovar}; Lomov IuM et al.; Materials on the import of rarely occurring Vibrio cholerae, not belonging to group O1 of serovar O139, to the territory of Russia are presented . The clinical picture of a cholera case is described and the biological properties of V . cholerae, serovar O139, are presented . A suggestion has been made concerning the appearance of a new V . cholerae serovar, capable of ousting V . cholerae eltor, the cause of the seventh pandemic.

Zh Mikrobiol Epidemiol Immunobiol, 1995 Mar-Apr, Suppl 2, 90 - 1
{The isolation of the Vibrio cholerae dermotoxin and the characterization of its biological properties}; Atarova GT et al.; The dermonecrotic factor (dermotoxin) inducing skin necrosis in rabbits has been isolated from V . cholerae strain B-53-2-38 and partially purified . Dermotoxin has a molecular weight of about 110 kD and possesses pronounced cytotoxic and general toxic action, differing from that of enterotoxin . The introduction of this factor into the blood and peritoneum of laboratory animals causes their death.

Zh Mikrobiol Epidemiol Immunobiol, 1995 Mar-Apr, Suppl 2, 86 - 9
{The outer membranes of Vibrio cholerae as a potential component in a chemical vaccine}; Markov EIu et al.; The results of the study of the preparation of V . cholerae eltor membrane, obtained by the lysis and inactivation of microbial cells with urea and the subsequent differential centrifugation and nuclease treatment . As revealed in this study, the outer membrane preparation, when introduced parenterally and orally to mice, induced pronounced immunity to experimental cholera infection and the production of vibriocidal antibodies in high titers . The treatment of V . cholerae eltor membranes with trypsin led to further increase of the immunogenic potency of the preparation . The protective action of V . cholerae eltor outer membranes considerably exceeded the protective effect of currently used whole-cell eltor vaccine . This opens prospects for using the above-mentioned preparation for the improvement of chemical vaccine as a component ensuring the formation of antibacterial immunity.

Zh Mikrobiol Epidemiol Immunobiol, 1995 Mar-Apr, Suppl 2, 84 - 6
{The properties of the Vibrio cholerae strains isolated in large districts in western Dagestan in 1994}; Cherepakhina IIa et al.; 190 V . cholerae cultures isolated by the specialized antiepidemic brigade of the Rostov-on-Don Research Institute for Plague Control in the Khasavyurt, Babayurt and Novolaksk regions of Daghestan in August-October 1994 . All isolated strains were typical with respect to their morphological and cultural properties and could be agglutinated (with the exception of one strain) to the titer or half-titer with diagnostic cholera serum and Ogawa serum . 4 strains had signs of RO-dissociation, 4 strains were agglutinated with Inaba serum in a low titer . All strains were resistant to diagnostic bacteriophages . Cyproxin and doxicycline proved to be the most active agents for the treatment of patients . Agglutinins, vibriocidins and antidermonectrotic antibodies in diagnostic titers were detected in the sera of all patients and Vibrio carriers.

Zh Mikrobiol Epidemiol Immunobiol, 1995 Mar-Apr, Suppl 2, 80 - 3
{The preservation of the causative agent of cholera in the water supplies of the central regions of Dagestan (experimental data)}; Golubev BP et al.; The acidic pH of water of surface water reservoirs in Izberbash and two adjoining regions, including sea water, seems to be unfavorable for the prolonged preservation of Vibrio cholerae eltor, but additional ecological investigations are necessary to study the possibility for infection to take root at this territory . Water from the Zam-Zam spring, if contaminated with V . cholerae, may serve as a transmission factor, but the duration of its action is limited by the survival term of V . cholerae . The water route of transmission did not play any essential role in the spread of cholera in the central regions of Daghestan.

Zh Mikrobiol Epidemiol Immunobiol, 1995 Mar-Apr, Suppl 2, 78 - 80
{The bacteriophage lysis-susceptibility properties of Vibrio cholerae strains isolated in separate regions of Dagestan in 1994}; Kazakova ES et al.; The study of the properties of V . cholerae strains isolated in June-September 1994 in the Daghestan revealed that they belonged, according to their specific properties, to typical representatives of V . eltor, serovar Ogawa, but a great part of them (67.2%) was not lysed by diagnostic cholera bacteriophages . Experiments with different batches of diagnostic cholera bacteriophages showed the necessity of their further improvement.

Zh Mikrobiol Epidemiol Immunobiol, 1995 Mar-Apr, Suppl 2, 74 - 8
{The characteristics of Vibrio cholerae isolated in Dagestan in 1994}; Podosinnikova LS et al.; A high degree of resistance to cholera diagnostic phages and carriership of prophages characteristic of V . cholerae eltor strains vct+ were shown to be the specific features V . cholerae isolated in Daghestan during the period of June-October 1994 . Among the strains under study, isolated respectively in 12 and 18 out of 19 regions of Daghestan, a high proportion was found to have resistance to tetracycline (65%) and chloramphenicol (28.6%) . Moreover, some strains were found to be resistant to furagin and erythromycin . Out of 242 strains resistant to antibacterial preparations, 163 strains were found to have multiple resistance . Gentamicin, cipropfloxacin and doxicycline were shown to have high in vitro activity with respect to the strains under study.

Zh Mikrobiol Epidemiol Immunobiol, 1995 Mar-Apr, Suppl 2, 71 - 4
{The characteristics of the antibacterial therapy for cholera in Dagestan}; Karbyshev GL et al.; Wide circulation of antibiotic-resistant Vibrio cholerae strains again gives prominence to the problem of etiotropic therapy . The results of the treatment of 428 persons infected with V.cholerae (237 cholera patients and 191 Vibrio carriers) in different regions of Daghestan during the outbreak of epidemic in 1994 are presented . The main criterion of the effectiveness of antibacterial therapy was the determination of the percentage of bacterial relapses . The sensitivity of 118 V.cholerae strains to different antibacterial preparations was studied by the method of serial dilutions . After the clinical use of chloramphenicol 29.7% of bacterial relapses were registered, the in vitro resistance of V . cholerae being 32-64 mkg/ml . After the use of tetracycline 16.5% bacterial relapses were registered with in vitro resistance being the same . The use of the combination of these preparations gave 15% of bacterial relapses . Furazolidone gave 4.3% of bacteria relapses, while after the use of ciprofloxacin 2.8% of bacterial relapses were registered with in vitro sensitivity equal to 0.25-0.5 mkg/ml . Ciprofloxacin was recommended for the treatment of cholera patients and furazolidone, for the treatment of Vibrio carriers.

Zh Mikrobiol Epidemiol Immunobiol, 1995 Mar-Apr, Suppl 2, 64 - 9
{The cholera epidemic in mountainous regions of Dagestan in relation to the probable role of the water factor in its spread}; Onishchenko GG et al.; The follow-up of dynamics of cholera epidemic in 1994 was made in 4 mountain regions of Daghestan with shared river system . There were 537 infection cases in these regions, which was equal to 1/4 from the total number of cholera patients in Daghestan . The probability of cholera distribution by water way has been shown, which can be related to massive dissemination with V . cholerae in river, drink and waste water both from active, and from local (intrahospital) epidemic sources . Vibrio's exit from cholera hospital became possible after dissemination intrahospital V . cholerae of waste water, after contamination of river water and after formation of secondary sources and cholera waves, which resulted in the prolongation up to 40 days of this infection in mountain regions of Daghestan.

Zh Mikrobiol Epidemiol Immunobiol, 1995 Mar-Apr, Suppl 2, 60 - 4
{The epidemiology of cholera in Derbent District . The characteristics of the epidemic process}; Onishchenko GG et al.; Cases of cholera in the Derbent District of the Daghestan took predominantly mild and moderate clinical forms . The ratio of patients to Vibrio carriers was 1:1.25 . The etiological agent of the epidemic was V . eltor, serovar Ogawa . Cholera patients were of greater epidemiological importance as the sources of infection in comparison with Vibrio carriers . Carrier state after convalescence was observed in 1.1% of cases . The population was infected with cholera, as a rule, through everyday contacts and in more rare cases by the water route . The highest rate of contamination with V . cholerae was registered in the age group of 0-2 years.

Zh Mikrobiol Epidemiol Immunobiol, 1995 Mar-Apr, Suppl 2, 56 - 9
{The epidemiology of cholera in Derbent District, Dagestan . The routes of the importation and spread of cholera}; Onishchenko GG et al.; The epidemic of cholera in the Derbent District of the Daghestan was imported into the region . 172 cholera patients and 204 Vibrio carriers were registered in 23 settlements of the region during the period of June 26 to October 12, 1994 . The wide spread of this infection was facilitated by high migration activity of the population and the belated introduction of measures for limiting migration . The most severe outbreaks of cholera were observed in Derbent and in the villages of Mamedkala and Morskoe.

Zh Mikrobiol Epidemiol Immunobiol, 1995 Mar-Apr, Suppl 2, 50 - 2
{The routes of the spread of cholera in the Shamil'skiĭ Mountain District of Dagestan}; Onishchenko GG et al.; The routes of the spread of cholera were analyzed in 273 patients and Vibrio carriers during the outbreak of cholera in a mountainous region of Daghestan during the period of July 18 to September 4, 1994 . Cholera was found to spread mainly after funeral repasts and condolence visits accompanied by the dispensation of foodstuffs, transmission being realized through alimentary and contact routes . Under the conditions of the absence of the centralized water supply system in mountain villages and the contamination of water in open reservoirs it was found to be expedient to use, in addition to the recommended complex of antiepidemic measures, small automatic filtration units.

Zh Mikrobiol Epidemiol Immunobiol, 1995 Mar-Apr, Suppl 2, 46 - 50
{Cholera in Makhachkala during the 1994 epidemic}; Mamaeva SM et al.; During the period of epidemic in the Daghestan 51 patients and 27 Vibrio carriers were detected in Makhachkala . A considerable proportion (30.7%) of cholera cases caused by infection imported from regions, unfavorable with respect to cholera, in the presence of pronounced migration of the population was registered . The role of different transmission routes in cases of cholera was as follows: day contacts were responsible for 43.3%, the alimentary route for 28.4% and the water route for 14.9% of cases . The epidemic situation was characterized by a mild and prolonged type of the epidemic process . Mass diseases were prevented by a complex of cholera control measures, among them the emphasis was made on various limitations and prophylactic measures aimed at the rupture of the transmission routes of V . cholerae.

Zh Mikrobiol Epidemiol Immunobiol, 1995 Mar-Apr, Suppl 2, 40 - 3
{Cholera in a remote region of Dagestan}; Arutiunov IuI et al.; The epidemic situation in a remote region of Daghestan at the period of the cholera outbreak in the republic is considered with the use of concrete examples . The analysis of cases of cholera, as well as Vibrio carriership, at the period of August 5 to October 5, 1994, is presented . The territorial and temporal separation of different cases of the disease and Vibrio carriership were indicative of the periodic import of the causative agent to the territory of the town and the region from other places, unfavorable with respect to cholera, without the involvement of the water factor of infection transmission . Timely and complete antiepidemic measures prevented this infection from acquiring the character of developed epidemic.

Zh Mikrobiol Epidemiol Immunobiol, 1995 Mar-Apr, Suppl 2, 30 - 2
{The use of a cholera coagulating diagnostic agent under the cholera epidemic conditions in Dagestan}; Orlova GM et al.; Dried cholera diagnosticum for the slide coagglutination test was obtained . The diagnosticum, found to be highly active and specific, permitted the detection of Vibrio cholerae in the analyzed material at a concentration of 10(6)-10(8) microbial cells/ml . The diagnosticum was used during cholera epidemic in Daghestan for the detection and rapid identification of cholera vibrios . In all cases the positive results of the coagglutination test were confirmed by other investigation methods (no cases of hyperdiagnosis was registered) . The production of these diagnostica in small batches on the basis of specialized research laboratories was recommended.

Zh Mikrobiol Epidemiol Immunobiol, 1995 Mar-Apr, Suppl 2, 3 - 8
{Cholera in the Republic of Dagestan}; Onishchenko GG et al.; Cases of cholera were registered in Daghestan in all pandemics with the exception of the fourth one . For the first time the import this infection by pilgrims returning from their hajj by motor transport and traveling through the countries of southwest Asia was registered . 184 settlements of 27 regions, 8 towns and 1 housing estate were involved in the epidemic process, the number of registered cholera cases and Vibrio carriers being 2,327 . High contamination rates were detected in regions situated in different geographical zones . Everyday contacts and the alimentary route were the main routes of transmission during this epidemic, while the role of the water route was considerably less important.

Zh Mikrobiol Epidemiol Immunobiol, 1995 Mar-Apr, Suppl 2, 22 - 7
{An operative analysis of the cholera epidemic in the Republic of Dagestan using a computer database}; Moskvitina EA et al.; The epidemiological information on cholera epidemic in Daghestan during the period of June 6 to October 21, 1994, based on the data obtained from 2,327 patients and Vibrio carriers in 184 settlements of 27 regions, 8 towns and 1 housing estate, was collected, systematized and analyzed with the use of the data base (DB) . The use of DB made it possible to carry out the surveillance of the level of and dynamics of morbidity and infections rates, to determine the territories of risk, the age and social groups of risk, the active routes and factors of transmission . DB may be used for analyzing of operative and current epidemiological information in cholera and other infections.

J Clin Microbiol, 1995 Mar, 33(3), 732 - 4
Evaluation of the monoclonal antibody-based kit Bengal SMART for rapid detection of Vibrio cholerae O139 synonym Bengal in stool samples; Qadri F et al.; A monoclonal antibody-based test, Bengal SMART, was developed for rapid detection of Vibrio cholerae O139 synonym Bengal directly from stool specimens . The test, which takes about 15 min to complete, was used to screen 189 diarrheal stool specimens . The results were compared with those of a monoclonal antibody-based coagglutination test (COAT) and the conventional culture methods used as the "gold standard" for detection of V . cholerae O139 . The Bengal SMART test showed a sensitivity of 100% and a specificity of 97% in comparison with the gold standard . It also fared better than COAT, which had a sensitivity of 96% for rapid detection of V . cholerae O139 synonym Bengal . These results show that Bengal SMART is suitable for use in field settings for rapid diagnosis of cholera caused by V . cholerae O139.

Indian J Med Res, 1995 Mar, 101, 94 - 7
A retrospective analysis of the Madras epidemic of non-01 Vibrio cholerae new serogroup 0139 Bengal; Dhamodaran S et al.; During the recent epidemic of cholera in Madras from October-December 1992, a total of 11,100 patients with acute secretory diarrhoea have been admitted to the Communicable Diseases Hospital, Madras, when compared to a total of 2,440 patients admitted during the pre-epidemic period studied between January - September 1992 . A novel strain of non-01 V . cholerae was found to be the most predominant agent during the epidemic period . A representative sample of 84 non-01 strains isolated during the epidemic period were confirmed as V . cholerae non-01 and newly designated as serogroup 0139 at National Institute of Cholera and Enteric Diseases, Calcutta . All 84 strains of non-01 were found to elaborate cholera toxin (CT) . Fourteen strains were also confirmed as serogroup 0139 from our non-01 stock recovered during the pre-epidemic period . Of the 17,540 patients admitted during the post epidemic period, studied from January 1993 - June 1994, 3.8 and 37.8 per cent patients were infected by 01 and non-01 serogroup 0139 respectively . The new serogroup 0139 continued to be the predominant isolate even during the post epidemic period.

Can J Microbiol, 1995 Mar, 41(3), 209 - 16
The plasmid profiles of fish pathogenic isolates of Aeromonas salmonicida, Vibrio anguillarum, and Vibrio ordalii from the Atlantic and Pacific coasts of Canada; Giles JS et al.; The plasmid profiles of oxytetracycline- and streptomycin-resistant isolates of Aeromonas salmonicida, Vibrio anguillarum, and Vibrio ordalii were examined by agarose gel electrophoresis . Bacterial isolates were from disease outbreaks in fish on the Atlantic and Pacific coasts . Resistant isolates were examined when grown in the presence and absence of antibiotic . Alkaline lysis methods were used for plasmid isolation . Vibrio spp . were predominantly plasmidless, except for a 47-kilobase (kb) plasmid . Atlantic coast isolates of A . salmonicida possessed four or six plasmids, with four smaller plasmids ranging in size from 4.3 to 8.1 kb being consistently observed . The plasmid profiles of antibiotic-sensitive ATCC strains were identical . The plasmid profiles of the Pacific coast isolates of A . salmonicida varied slightly from those of the Atlantic coast isolates with six plasmids observed, ranging in size from 4.2 to 8.9 kb . Resistance to the antibiotics was not altered following plasmid curing experiments and resistance was not transferable to Escherichia coli . Thus, resistance to oxytetracycline and streptomycin did not appear to be plasmid mediated.

Infect Agents Dis, 1995 Mar, 4(1), 41 - 6
Vibrio cholerae O139 Bengal: emergence of a new epidemic strain of cholera; Morris JG Jr; In October 1992, a new strain of cholera, subsequently designated Vibrio cholerae O139 Bengal, was detected in Madras, India . This strain spread rapidly through the Indian subcontinent and has now been reported in many parts of Asia, with additional cases identified in travelers to North American and the Middle East . Phylogenetically, V . cholerae O139 Bengal is very closely related to "standard" V . cholerae O1 El Tor strains; it produces cholera toxin and causes an illness identical that seen with V . cholerae O1 . However, prior immunity to V . cholerae O1 El Tor does not appear to protect against illness caused by V . cholerae O139 Bengal . O139 Bengal strains have a short, "semi-rough" O side chain and are encapsulated, changes that are likely to have accounted for their ability to cause disease in persons with prior exposure to cholera . These changes in surface structures appear to have resulted from a limited number of genetic modifications . The appearance of V . cholerae O139 Bengal may well herald the beginning of the eighth pandemic of cholera--and underscores the tremendous potential within nature for creation of new strains of "old" pathogens.

Microbiology, 1995 Mar, 141 ( Pt 3), 597 - 604
Identification of a 29 kDa flagellar sheath protein in Helicobacter pylori using a murine monoclonal antibody; Luke CJ et al.; The membrane-like flagellar sheath of Helicobacter pylori is of unknown function and little is known of its composition . A murine monoclonal antibody to H . pylori, designated GF6, which reacts by immunoblot with a polypeptide with an apparent molecular mass of 29 kDa was shown by immunogold-electron microscopy to label specifically the flagellar sheath structure . The antigen was detected by immunoblot using the monoclonal antibody in all 11 strains, of diverse geographic origin, so far tested . The antibody also reacted weakly with polypeptides with apparent molecular masses of 65 kDa in Vibrio cholerae and Vibrio parahaemolyticus . The antigen was shown by one- and two-dimensional electrophoretic analysis and immunoblotting to be distinct from the abundant urease subunit UreA, of similar molecular mass . Identification of this flagellar sheath polypeptide will facilitate investigation of the structure and function of the flagellar sheath of this important gastric pathogen.

Clin Diagn Lab Immunol, 1995 Mar, 2(2), 177 - 81
A novel method to chemically immobilize antibody on nylon and its application to the rapid and differential detection of two Vibrio parahaemolyticus toxins in a modified enzyme-linked immunosorbent assay; Honda T et al.; A new method of chemically immobilizing antibody on nylon was developed . The method consists of serial treatments with HCl, polyethylene imine, and maleic anhydride methylvinyl ether copolymer, which resulted in the stable immobilization of sufficient amounts of antibodies on nylon . This principle was used to differentially detect two immunologically related but nonidentical hemolysins (thermostable direct hemolysin {TDH} and TDH-related hemolysin {TRH}) of Vibrio parahaemolyticus in a modified enzyme-linked immunosorbent assay with antibodies immobilized on nylon slips (NSIT) . The results (dark purple color on nylon slips) were easily evaluated by the naked eye . The results with NSIT were compatible with those obtained by using DNA probes or a conventional bacterial culture test, not only with cultured specimens but also with clinical specimens (diarrheal stool samples) . Furthermore, the NSIT differentially detected TDH and TRH in a single test . The antibody immobilization method developed here is applicable to various immunological detection methods and may improve their sensitivity and specificity.

Zh Mikrobiol Epidemiol Immunobiol, 1995 Mar-Apr, (2), 22 - 6
{The interrelationships between Vibrio cholerae and the infusorian Tetrahymena pyriformis}; Pogorelov VI et al.; The results of the study of interaction between V . cholerae of different virulence and T . pyriformis are presented . The study has revealed the heterogeneity of V . cholerae population: alongside easily phagocytized vibrios, there are vibrios resistant to the digestive action of T . pyriformis . An increase in the number of V . cholerae in association with T . pyriformis has been evaluated, taking into account the selective multiplication of vibrios resistant to phagocytosis . The data on changes in the agglutinative, phagolytic and virulent properties of V . cholerae cultivated together with T . pyriformis are presented . The suggestion has been made that protozoa can function as hosts of pathogenic vibrios supporting their existence in water.

Microb Pathog, 1995 Mar, 18(3), 231 - 5
Distribution of genes encoding cholera toxin, zonula occludens toxin, accessory cholera toxin, and El Tor hemolysin in Vibrio cholerae of diverse origins; Kurazono H et al.; A large collection of 1154 strains of Vibrio cholerae of diverse origins including serogroups 01 and 0139 and those belonging to the non-01 and non-0139 (non-01:non-0139) serogroups were examined with a battery of DNA probes specific for cholera toxin (CT), zonula occludens toxin (ZOT), accessory cholera toxin (ACE) and El Tor hemolysin (HLY) to determine the distribution of genes among wild strains and to understand the importance of these factors in the pathogenesis of the disease cholera . Among the 01 clinical isolates, the majority of the strains had an intact core region (ctx, zot, ace) and also possessed the hlyA gene . Although rare, strains of 01 with natural deletions of the ctx, zot and/or ace genes were also detected . The absence of the virulence genes comprising the core region and the presence of the hlyA gene dominated the 01 environment, food isolates and the clinical and environmental non-01: non-0139 strains of V . cholerae . All the 0139 strains examined in this study possessed genes located in the core region and the hlyA gene . Among all the virulence-associated genes examined, the hlyA gene was the most conserved genetic element in V . cholerae independent of biotypes and serogroups.

Carbohydr Res, 1995 Mar 1, 268(1), 73 - 84
Synthesis of specifically deoxygenated analogues of the methyl alpha-glycoside of the intracatenary monosaccharide repeating unit of the O-polysaccharide of Vibrio cholerae O:1; Gotoh M et al.; Treatment of methyl alpha-D-perosaminide (1) with gamma-butyrolactone gave the 2'-deoxy analogue of methyl 4,6-dideoxy-4-(3-deoxy-L-glycero-tetronamido)-alpha-D-mannopyranos ide (13), the methyl alpha-glycoside of the intracatenary monosaccharide repeating unit of the O-polysaccharide of Vibrio cholerae O:1 . The analogous 4'-deoxy derivative was obtained by hydrogenolysis of a 4'-chlorodeoxy precursor, obtained by chlorination of methyl 2,3-di-O-benzyl-4-(2-O-benzyl-3-deoxy-L-glycero-tetronamido)-4,6-d ideoxy-alpha- D-mannopyranoside with methanesulfonyl chloride in DMF . To obtain the 3-deoxy analogue of 13, methyl 4-amino-2-O-benzyl-4,6-dideoxy-3-O-p-methoxy-benzyl-alpha-D-mannopyranos ide was converted into methyl 2-O-benzyl-4,6-dideoxy-4-(2,4-di-O-benzyl-3-deoxy-L-glycero-tetronami do)- alpha-D-mannopyranoside, which was deoxygenated via the corresponding 3-O-(imidazol-1-ylthiocarbonyl) derivative . Subsequent catalytic debenzylation gave the deoxy compound (24) . In an alternative synthesis, which is also generally useful for the preparation of 4-N-acyl-3-deoxy derivatives of 1, methyl 4-azido-4,6-dideoxy-alpha-D-mannopyranoside was converted through a series of transformations into methyl 4-amino-2-O-benzyl-3,4,6-tri-deoxy-alpha-D-mannopyranoside . Subsequent reaction with 2-O-benzyl-3-deoxy-L-glycero-tetronolactone, followed by hydrogenolysis of the formed tetronamido derivative, gave 24.

Gene, 1995 Feb 27, 154(1), 55 - 9
The periplasmic endonuclease I of Escherichia coli has amino-acid sequence homology to the extracellular DNases of Vibrio cholerae and Aeromonas hydrophila; Jekel M et al.; The gene endA, encoding the periplasmic endonuclease I (EndoI) of Escherichia coli, was identified on a cloned chromosomal 1.5-kb HindIII fragment . The nucleotide sequence of the fragment revealed an open reading frame (ORF) coding for a polypeptide of 235 amino acids (aa) . The ORF preceeded by a region with two possible promoter sites displays promoter activity when cloned into an expression vector . On the C-terminal side, two sequences with putative transcription termination function are present . The predicted aa sequence suggests the presence of a signal peptide of 22 aa and a signal peptide cleavage site . A cold-shock supernatant from cells harbouring a multicopy endA+ plasmid contained an approx . tenfold higher amount than wild-type cells of the DNA double-strand- and single-strand-cleavage activities characteristic of EndoI . The growth rate and viability of the cells was not affected . The predicted aa sequence of the ORF is 60 and 54% identical to the sequence of extracellular DNases from Vibrio cholerae and Aeromonas hydrophila, respectively.

J Biol Chem, 1995 Feb 17, 270(7), 2914 - 20
A 25-kDa beta-lactam-induced outer membrane protein of Vibrio cholerae . Purification and characterization; Deb A et al.; A 25-kDa outer membrane protein, induced following treatment of Vibrio cholerae cells with beta-lactam antibiotics and constituting about 8-10% of the total outer membrane proteins of beta-lactam-resistant mutants, has been purified to homogeneity . It is a basic (pI 8.5) protein rich in beta-sheet structure and is a homodimer, the monomers being held together by hydrophobic interactions . The effective hydrophobicity of the protein is low, and a large part of the protein is exposed on the surface of the outer membrane . The protein does not have beta-lactamase or autolytic activity and is not a penicillin-binding protein . The Stoke's radius of the 25-kDa protein (26 A) is comparable to the pore size of the V . cholerae OmpF-like porin . Proteoliposome swelling assay showed that the 25-kDa protein might block the pores of OmpF through which beta-lactam antibiotics normally enter the cells . Twenty-two amino acid residues from the N-terminal end of the 25-kDa protein have been sequenced, and a 32-mer oligonucleotide probe was synthesized using the amino acid residues 2-12 . This probe was used to identify the gene encoding the 25-kDa protein . The beta-lactam-resistant cells are insensitive to changes in the osmolarity of the growth medium in contrast to the wild type cells which exhibit osmoregulation of OmpF and OmpC synthesis . All beta-lactam-resistant mutants examined are resistant to novobiocin.

Lancet, 1995 Feb 11, 345(8946), 359 - 61
Why treatment centres failed to prevent cholera deaths among Rwandan refugees in Goma, Zaire; Siddique AK et al.; In July, 1994, in one of the worst cholera outbreaks in recent times, an estimated 12,000 Rwandan refugees died in Goma in eastern Zaire . The Vibrio cholerae strains were resistant to tetracycline and doxycycline, the commonly used drugs for cholera treatment . Despite the efforts of international organisations, which provided medical relief by establishing treatment centres in Goma, mortality from the disease was much higher than expected . In the area of Muganga camp, which had the largest concentration of refugees and where most of the medical aid organisations were active, the highest reported case-fatality ratio for a single day was 48% . The slow rate of rehydration, inadequate use of oral rehydration therapy, use of inappropriate intravenous fluids, and inadequate experience of health workers in management of severe cholera are thought to be some of the factors associated with the failure to prevent so many deaths during the epidemic . In one of the temporary treatment centres with the worst case-fatality record, our team showed that improvement of these factors could increase the odds of survival of cholera patients even in a disaster setting.

J Biol Chem, 1995 Feb 10, 270(6), 2588 - 94
Enzyme assembly after de novo synthesis in rabbit reticulocyte lysate involves molecular chaperones and immunophilins; Kruse M et al.; The folding kinetics of two luciferases were studied after synthesis in reticulocyte lysates to investigate whether molecular chaperones and/or folding catalysts are involved in the folding reactions . Two bacterial luciferases were used as model proteins: heterodimeric Vibrio harveyi luciferase (LuxAB), and a monomeric luciferase fusion protein (Fab2) . Data indicate that folding of these enzymes to the native state occurs in the translation system, and that the extent of folding can be quantified . It was found that (i) folding of LuxAB and Fab2 can clearly be separated in time from synthesis, (ii) folding of Fab2 and LuxAB is slow because it involves either transient (Fab2) or permanent (LuxAB) interaction of polypeptides, (iii) preservation of the assembly competent state of LuxA and/or LuxB and folding of Fab2 depend on ATP-hydrolysis, (iv) folding of Fab2 and LuxAB is partially sensitive to cyclosporin A (CsA) and FK506, i.e . inhibitors of two distinct peptidylprolyl cis/trans-isomerases . Thus, bacterial luciferases provide a unique system for direct measurement of the effects of ATP-dependent molecular chaperones on protein folding and enzyme assembly in reticulocyte lysates . Furthermore, these two luciferases provide the first direct evidence documenting the involvement of peptidylprolyl cis/trans-isomerases in protein biogenesis in a eukaryotic cytosol.

Gene, 1995 Feb 3, 153(1), 85 - 7
A new mobilizable cosmid vector for use in Vibrio cholerae and other gram-negative bacteria; Connell TD et al.; A new mobilizable cosmid vector, pCOS5, was engineered for use in Vibrio cholerae (Vc) . Plasmid pCOS5 is small in size (7286 bp), contains the oriT from plasmid RK2, and has several unique restriction sites . The complete nucleotide sequence of pCOS5 was deduced from the DNA fragments used in its construction . Biparental matings using Escherichia coli (Ec) SM10 and triparental matings using Ec DH5 alpha{pRK2013} were used to measure the conjugation frequency of pCOS5 and pAJM1, a clone containing a 40-kb insert of chromosomal DNA from Vc ligated into pCOS5 . Transfer of pCOS5 or pAJM1 to Vc occurred at a frequency of between 10(-2)-10(-3) transconjugants per recipient cell . The promiscuous nature of RP4/RK2 transfer functions makes pCOS5 a potentially useful vector for mobilizing large fragments of cloned DNA between different Gram- bacteria that support replication of ColE1 plasmids or as a mobilizable suicide vector in Gram- bacteria where replication of ColE1 plasmids is not supported.

Gene, 1995 Feb 3, 153(1), 81 - 4
Isolation and characterization of a Vibrio cholerae gene (tagA) that encodes a ToxR-regulated lipoprotein; Harkey CW et al.; The Vibrio cholerae (Vc) gene (tagA) coding for the TagA lipoprotein has been isolated . Sequencing of tagA revealed the presence of an open reading frame (ORF) of 568 amino acids with a characteristic signal peptidase II cleavage site at the N terminus . Electrophoretic analysis of proteins synthesized by Escherichia coli (Ec) cells following T7 promoter/RNA polymerase-directed expression of tagA, revealed a closely migrating doublet of proteins corresponding to two species of TagA . Computer-generated alignment algorithms predict that a homology exists between Vc TagA and Ec K99 fimbriae biogenesis determinant FanD.

FEMS Microbiol Lett, 1995 Feb 1, 126(1), 43 - 8
Diversity of DNA sequences among Vibrio cholerae O139 Bengal detected by PCR-based DNA fingerprinting; Makino S et al.; Vibrio cholerae O139, a causative agent of a large epidemic of cholera-like illness, has suddenly emerged and spread widely over several months . To investigate the characteristics unique to O139, traditional typing techniques for V . cholerae, such as biochemical characteristics, antibiotic susceptibility and detection of toxin production, were performed, with the result that 145 O139 strains, except for two O139 strains isolated from Argentina and Germany, were indistinguishable from O1 strains . Thus, in order to clarify the genetical relatedness among O139 strains, and between O139 and O1 strains, the RAPD (random amplified polymorphic DNA) DNA fingerprinting method was undertaken . Although the RAPD arrays in five O139 isolates from Vellore with one arbitrary primer were slightly different from the other O139 strains, the RAPD patterns of the 145 forty-five O139 strains except for two O139 strains from Argentina and Germany were quite similar to each other, but were different from those of O1 strains, indicating that those O139 epidemic strains are closely related to each other regardless of their place of isolation . Furthermore, the RAPD patterns of the O139 strains resembled those of E1 Tor strains rather than classical strain, and a small change in the RAPD pattern of O139 strains occurred during subculture for 200 generations . These results taken together suggested that O139 V . cholerae have emerged from a common origin associated with the E1 Tor strain.

Am J Trop Med Hyg, 1995 Feb, 52(2), 124 - 7
Molecular characterization of Vibrio cholerae O139 isolates from Asia; Echeverria P et al.; In 1992, a serologically novel clone of Vibrio cholerae, designated O139, caused large epidemics of diarrhea in India and Bangladesh . To determine the extent of the spread of V . cholerae O139 worldwide, 484 V . cholerae non-O1 strains isolated from different patients with diarrhea in Thailand, Indonesia, the Philippines, and Peru in 1993 were tested for agglutination in O139 antisera . One hundred fifty-one of these 484 isolates were examined for genes encoding cholera toxin, zonula occlulans toxin, the repetitive sequence 1, and the toxin coregulated pilin A (the V . cholerae virulence gene complex) . Thirty-three percent (122 of 364) of V . cholerae non-O1 strains isolated from different patients with diarrhea in Thailand agglutinated in O139 antisera . Ninety-eight percent (120 of 122) of V . cholerae O139 contained the V . cholerae virulence gene complex . None of the 104 V . cholerae non-O1 strains isolated from patients with diarrhea in Indonesia or the 14 strains from patients with diarrhea in the Philippines were serotype O139 . Four different ribotypes were found in V . cholerae O139 isolated in Asia . Twenty-three (47%) of 49 Thai O139 strains examined were of different ribotypes than isolates from India and Bangladesh; V . cholerae strains that were not O1 or O139 that were isolated from flies and water in Thailand 11 years previously in 1981 contained the same V . cholerae virulence gene complex found in V . cholerae O1 and O139 . This suggests that other unidentified virulence determinants are involved in V . cholerae O139 pathogenesis.

J Med Microbiol, 1995 Feb, 42(2), 83 - 90
Characterisation of a haemolysin related to Vp-TDH produced by a Kanagawa phenomenon-negative clinical isolate of Vibrio parahaemolyticus; Nagayama K et al.; The production of a family of haemolysins--thermostable direct haemolysin (Vp-TDH), Vp-TDH-related haemolysin (Vp-TRH) and Vp-TDH/I--has been reported in clinical isolates of Vibrio parahaemolyticus . This paper describes a fourth type of haemolysin--Vp-TDH/II--produced by a Kanagawa phenomenon-negative clinical isolate of V . parahaemolyticus (O13:K, untypable) . Vp-TDH/II was purified by ammonium sulphate precipitation and successive filtrations on DEAE-cellulose, hydroxyapatite, Sepharose 4B and Mono Q columns . Vp-TDH/II was biophysicochemically and immunologically similar to, but not identical to Vp-TDH, Vp-TRH and Vp-TDH/I . Vp-TDH/II stimulated vascular permeability in rabbit skin and was lethal to mice . Purified Vp-TDH/II and viable cells of the Vp-TDH/II-producing strain both induced fluid accumulation in ligated rabbit intestine . The plasmid-determined structural gene for Vp-TDH/II was cloned and the nucleotide sequence determined . The deduced amino acid sequence of Vp-TDH/II differed from those of Vp-TDH, Vp-TRH and Vp-TDH/I.

Epidemiol Infect, 1995 Feb, 114(1), 71 - 3
Vibrio cholerae O139 in Thailand in 1994; Bodhidatta L et al.; Vibrio cholerae O139 first appeared in India and Bangladesh in 1992 . Surveillance for O139 was started at three hospitals in Thailand in 1993 . By 1994 all three hospitals surveyed in Thailand had experienced an increase in Vibrio cholerae O139 infections.

Epidemiol Infect, 1995 Feb, 114(1), 65 - 70
Distribution and virulence of Vibrio cholerae belonging to serogroups other than O1 and O139: a nationwide survey; Mukhopadhyay AK et al.; The distribution and virulence of Vibrio cholerae serogroups other than O1 and O139 in India before, during and after the advent of O139 serogroup was investigated . A total of 68 strains belonging to 31 different 'O' serogroups were identified during the study period . With the exception of O53, there was no spatial or temporal clustering of any particular non-O1 non-O139 serogroup at any given place . Two of the 68 strains examined produced cholera toxin (CT) which could only be partially absorbed with anti-CT immunoglobulin G . Tissue culture assay revealed that some of the non-O1 non-O139 strains produced factors which evoked either a cell rounding or cell elongation response depending upon the medium used . This study indicates that serogroups other than O1 and O139 should also be continuously monitored.

Epidemiol Infect, 1995 Feb, 114(1), 51 - 63
Molecular characterization and antibiotic susceptibility of Vibrio cholerae non-O1; Dalsgaard A et al.; A collection of 64 clinical and environmental Vibrio cholerae non-O1 strains isolated in Asia and Peru were characterized by molecular methods and antibiotic susceptibility testing . All strains were resistant to at least 1 and 80% were resistant to two or more antibiotics . Several strains showed multiple antibiotic resistance (> or = three antibiotics) . Plasmids most often of low molecular weight were found in 21/64 (33%) strains . The presence of plasmids did not correlate with antibiotic resistance or influence ribotype patterns . In colony hybridization studies 63/64 (98%) V . cholerae non-O1 strains were cholera toxin negative, whereas only strains recovered from patients were heat-stable enterotoxin positive . Forty-seven Bgl I ribotypes were observed . No correlation was shown between ribotype and toxin gene status . Ribotype similarity was compared by cluster analysis and two main groups of 13 and 34 ribotypes was found . Ribotyping is apparently a useful epidemiological tool in investigations of V . cholerae non-O1 infections.

J Bacteriol, 1995 Feb, 177(4), 1053 - 8
Detection and quantification of Vibrio fischeri autoinducer from symbiotic squid light organs; Boettcher KJ et al.; Vibrio fischeri is the specific light organ symbiont of the sepiolid squid species Euprymna scolopes and Euprymna morsei . Both species of squid are luminescent by virtue of their bacterial symbionts, but the natural symbionts of E . scolopes do not produce visible luminescence in laboratory culture . The primary cause of this depressed luminescence by E . scolopes symbionts in culture was found to be the production of relatively low levels of V . fischeri autoinducer, a positive transcriptional coregulator of the lux regulon, identified as N-(3-oxohexanoyl) homoserine lactone . Concentrations of autoinducer activity produced by these symbionts in culture were quantified and found to be at least 10-fold lower than those produced by E . morsei isolates (which are visibly luminous outside the association) and perhaps 10,000-fold lower than those of the brightest V . fischeri strains . Despite the differences in their symbiont strains, the intact light organs of the two species of squid contained comparable amounts of extractable autoinducer activity (between 100 and 200 pg per adult animal) . The chromatographic behavior of this autoinducer activity on reverse-phase high-performance liquid chromatography was consistent with its presumptive identification as V . fischeri autoinducer . Within the 5-microliter volume of the epithelial core of the light organ in which the symbiotic V . fischeri strains are housed, these amounts would result in an effective autoinducer concentration of at least 100 nM . Because these levels are over 40-fold higher than the concentration needed for the induction of luminescence of bacteria in culture, we conclude that the inherent degree of autoinducer production by strains of V . fischeri may not influence their effectiveness as light organ symbionts . Furthermore, this study provides the first direct evidence that the phenomenon of cell density-dependent autoinduction, discovered and described first for laboratory cultures of V . fischeri but believed to be a general phenomenon in many species of host-associated symbionts and pathogens, is in fact a consequence of bacterial colonizations of host tissues.

Am J Public Health, 1995 Feb, 85(2), 168 - 72
Emerging diseases and ecosystem instability: new threats to public health; Epstein PR; Ecologists have begun to describe an environmental distress syndrome, whereby widespread loss of top predators and harsh environmental conditions are encouraging the selection of opportunistic pests and pathogens across a wide taxonomic range of plants and animals . Environmental change and pollutants stress individuals and populations, and this may be reflected in the global resurgence of infectious disease as these stresses cascade through the community assemblages of species . In 1993, the sudden appearance of a virulent, rodent-borne hantavirus in the arid US Southwest accompanied anomalous weather patterns, and a novel Vibrio cholerae variant (O139 Bengal) emerged in Asia where marine ecosystems are experiencing a pandemic of coastal algal blooms, apparently harboring and amplifying the agent . This paper suggests a framework for integrating the surveillance of health outcomes and key reservoir and vector species, with ecological and climatic monitoring.

J Bacteriol, 1995 Feb, 177(3), 835 - 8
Preliminary structure determination of the capsular polysaccharide of Vibrio cholerae O139 Bengal Al1837; Preston LM et al.; Vibrio cholerae O139 Bengal has recently been identified as a cause of epidemic cholera in Asia . In contrast to V . cholerae O1, V . cholerae O139 Bengal has a polysaccharide capsule . As determined by high-performance anion-exchange chromatography and 1H nuclear magnetic resonance analysis, the capsular polysaccharide of V . cholerae O139 Bengal strain Al1837 has six residues in the repeating subunit; this includes one residue each of N-acetylglucosamine, N-acetylquinovosamine (QuiNAc), galacturonic acid (GalA), and galactose and two residues of 3,6-dideoxyxylohexose (Xylhex) . The proposed structure is {formula: see text}

J Bacteriol, 1995 Feb, 177(3), 815 - 7
Evidence that the N-terminal region of the Vibrio fischeri LuxR protein constitutes an autoinducer-binding domain; Hanzelka BL et al.; The Vibrio fischeri luminescence genes are regulated by the LuxR protein and an N-acyl homoserine lactone compound termed the autoinducer . The C-terminal one-third of LuxR contains a domain that can interact with the transcription complex and activate the luminescence genes . On the basis of limited evidence it has been suggested that the N-terminal two-thirds of LuxR constitutes a domain that serves to bind the autoinducer . We show that tritium-labeled autoinducer binds to Escherichia coli cells in which LuxR is overexpressed . We also show that tritium-labeled autoinducer binds to E . coli in which truncated LuxR proteins missing portions of the C-terminal domain are expressed but does not bind to E . coli cells in which truncated LuxR proteins missing portions of the N-terminal region are expressed . Our results provide evidence that the autoinducer binds to LuxR and that in E . coli the N-terminal two-thirds of LuxR can fold into a polypeptide capable of binding the autoinducer in the absence of the C-terminal domain.

J Bacteriol, 1995 Feb, 177(3), 738 - 44
MyfF, an element of the network regulating the synthesis of fibrillae in Yersinia enterocolitica; Iriarte M et al.; The Yersinia enterocolitica surface antigen Myf is a fibrillar structure that resembles CS3 fimbriae . Gene myfA encodes the 21-kDa major subunit of the antigen, while genes myfB and myfC are required for the transport and assembly of pilin subunits at the bacterial cell surface . Here we show that the expression of Myf is regulated at the transcriptional level by temperature and pH . Gene myfA is transcribed at 37 degrees C and in acidic medium . The transcription start is preceded by a putative -10 box for the vegetative RNA polymerase as well as by sequences resembling the consensus sequence recognized by sigma 28 . Thus, myfA could be transcribed either from a classical sigma 70 promoter or from a sigma 28 promoter . Transcription of myfA requires at least two genes, myfF and myfE, situated immediately upstream from myfA . The myfF product does not show similarity to any known regulatory protein . It is an 18.5-kDa protein with no typical helix-turn-helix motif and a unique hydrophobic domain in the NH2-terminal part . T7 expression, osmotic shock, fractionation experiments, and TnphoA fusion analyses carried out in Escherichia coli suggest that MyfF is associated with the inner membrane by means of its hydrophobic domain whereas the hydrophilic part protrudes in the periplasm . These features strikingly evoke ToxS, a protein involved in regulation of Tcp pilus production in Vibrio cholerae . MyfE resembles PsaE, a protein involved in regulation of pH6 antigen in Yersinia pestis . Genes myfF and myfE are presumably part of a whole regulatory network . MyfF could be an element of the signal transducing system.

J Bacteriol, 1995 Feb, 177(3), 654 - 9
Activation of the Pseudomonas aeruginosa lasI gene by LasR and the Pseudomonas autoinducer PAI: an autoinduction regulatory hierarchy; Seed PC et al.; In Pseudomonas aeruginosa, the transcriptional activator LasR and the Pseudomonas autoinducer PAI, are necessary for efficient transcriptional activation of the lasB gene, encoding elastase (L . Passador, J . M . Cook, M.J . Gambello, L . Rust, and B . H . Iglewski, Science 260:1127-1130, 1993) . The transcriptional start points of lasI in Escherichia coli and P . aeruginosa were determined by S1 nuclease mapping . In the presence of both LasR and PAI, the start site, T1, is located at position -25 relative to the ATG translational start codon . A minor transcriptional start, T2, is found at position -13 when lasI is transcribed in the absence of either LasR or PAI in P . aeruginosa and E . coli, respectively . To begin to closely examine the regulation of lasI, whose product is involved in the synthesis of PAI, a lasI-lacZ fusion on a lambda phage was constructed to form monolysogens of E . coli MG4 . Lysogens supplied only with either lasI or lasR via multicopy plasmids demonstrated no significant increase in beta-galactosidase expression compared with control levels . Lysogens in which both lasR and lasI were supplied in multicopy exhibited a 62-fold increase in expression, and a lysogen in which lasR was supplied in trans and which was grown in the presence of exogenous PAI exhibited a 60-fold increase . Thus, LasR and PAI are necessary for the full expression of lasI in E . coli . The interchangeability of the P . aeruginosa and Vibrio fischeri homologs LasR and LuxR and their respective autoinducers, PAI and VAI, as activators of lasI-lacZ was examined . Only the combination of LasR and PAI significantly increased the expression of lasI . The comparison of lasI-lacZ and lasB-lacZ expression lysogens grown in the presence of lasR and PAI revealed that half-maximal expression of lasI required 0.1 nM PAI, in contrast to the 1.0 nM PAI necessary for lasB half-maximal expression . These results suggest an autoinduction regulatory hierarchy in which LasR and low PAI concentrations primarily activate lasI expression in a regulatory loop . With the accumulation of PAI, secondary activation of virulence product genes such as lasB occurs.

Infect Immun, 1995 Feb, 63(2), 724 - 8
Sequencing of the gene encoding the major pilin of pilus colonization factor antigen III (CFA/III) of human enterotoxigenic Escherichia coli and evidence that CFA/III is related to type IV pili; Taniguchi T et al.; The plasmid-encoded structural gene cofA necessary for the production of the major pilin subunit of pilus colonization factor antigen III (CFA/III) of human enterotoxigenic Escherichia coli was identified, and the nucleotide sequence of the gene was determined . cofA consists of 714 nucleotides encoding a 238-amino-acid protein (molecular weight of 25,309) . CofA seems to be a precursor of CFA/III pilin, because the first 23 residues of the N-terminal amino acid sequence of the purified CFA/III pili coincided with the deduced amino acid sequence for residues 32 to 54 of CofA . Western blot (immunoblot) analysis of CofA also indicated its processing to form mature pilin in the presence of the downstream region of cofA . These results suggest that the major pilin of CFA/III pili is produced as a precursor form which is posttranslationally modified to the mature pilin and forms morphological pili after cleavage of the Gly-30-Met-31 junction, probably by a protease encoded by an as-yet-unknown gene located downstream of cofA . Interestingly, the N-terminal 30-amino-acid sequence of mature CFA/III shows the highest identity (76.7%) to TcpA pilin of Vibrio cholerae, which is a type IV class B pilin.

Infect Immun, 1995 Feb, 63(2), 707 - 9
Attenuated live cholera vaccine strain CVD 103-HgR elicits significantly higher serum vibriocidal antibody titers in persons of blood group O; Lagos R et al.; Persons of blood group O are at increased risk of developing cholera gravis . In a randomized, placebo-controlled, double-blind safety-immunogenicity trial of live oral cholera vaccine CVD 103-HgR in 5- to 9-year-old Chilean children, vibriocidal antibody seroconversion (74% overall) did not differ by blood group . However, the reciprocal geometric mean titer (GMT) in blood group O vaccines (GMT = 486) was higher than that in non-O vaccines (GMT = 179) (P < 0.02).

Mol Microbiol, 1995 Feb, 15(4), 719 - 31
Analysis of Vibrio cholerae ToxR function by construction of novel fusion proteins; Ottemann KM et al.; The ToxR protein is a transmembrane protein that regulates the expression of several virulence factors of Vibrio cholerae . Previous analysis of fusion proteins between ToxR and alkaline phosphatase (ToxR-PhoA) suggested that ToxR was active as a dimer . In order to determine whether dimerization of the ToxR periplasmic domain was essential for activity, this domain was replaced by monomeric and dimeric protein domains . Surprisingly, PhoA (dimeric), beta-lactamase (monomeric, ToxR-Bla), or the leucine zipper of GCN4 (dimeric, ToxR-GCN4-M) could substitute functionally for the ToxR periplasmic domain . ToxR-GCN4 fusion proteins, in which the ToxR transmembrane domain was eliminated (ToxR-GCN4-C), were inactive, but an additional fusion protein that contained a heterologous membrane-spanning domain retained activity . Strains containing each of these ToxR fusion proteins were analysed for in vivo colonization properties and response to in vitro growth conditions that are known to affect expression of the ToxR regulon . Strains containing ToxR-GCN4-M and ToxR-Bla responded like wild-type strains to in vitro growth conditions . In the infant-mouse colonization model, strains containing ToxR fusion proteins were all deficient in colonization relative to strains containing wild-type ToxR, and strains containing monomeric ToxR-Bla were most severely outcompeted . These results suggest that, under in vitro conditions, ToxR does not require a dimerized periplasmic domain, but that, under in vivo conditions, the correct conformation of the ToxR periplasmic domain may be more important for function.

FEMS Immunol Med Microbiol, 1995 Feb, 10(3-4), 199 - 205
Studies on adhesion, haemagglutination and other biological properties of Vibrio cholerae O139; Sasmal D et al.; The adhesive capabilities of eight Vibrio cholerae O139 epidemic strains to isolated rabbit intestinal epithelial cells (RIEC) were observed to be high similar to those observed with a Vibrio cholerae O1 strain isolated from patients . Toxin production by the strains, measured by accumulation of fluid in rabbit ileal loop model, was high and the toxin was lethal as the animal expired within 6 h . Culture filtrates of the strains exhibited the presence of vascular permeability factor which produce induration and necrosis in the adult rabbit and guinea pig skin . All the strains showed high to moderate haemagglutinin titres against chicken erythrocytes and produced El Tor-like haemolysin . SDS-PAGE of the outer membrane preparation of the strains showed the presence of major protein component at 38 kDa region . The lethality of the toxin, high adhesive activity, shifting of the major outer membrane protein band and production of thermolabile haemolysin on Wagatsuma agar were the major variations of these epidemic strains from V . cholerae O1 and V . cholerae non-O1 strains isolated previously.

Glycobiology, 1995 Feb, 5(1), 67 - 75
In situ accessibility of murine macrophage gangliosides; Macala LJ et al.; Gangliosides are implicated in cell signal transduction . Prior to investigating this phenomenon in macrophages, the in situ accessibility of gangliosides to macromolecules was assessed for peritoneal macrophages isolated from normal C3H/HeN and endotoxin-hyporesponsive C3H/HeJ mice . C3H/HeJ resident and thioglycolate-elicited macrophage ganglioside patterns are the same as normal strains, and no strain differences in galactose oxidase accessibility for resident or thioglycolate-elicited macrophage gangliosides were found . The only gangliosides accessible to galactose oxidase in resident macrophages are GM1a structures . In thioglycolate-elicited macrophages, an additional ganglioside is accessible . For Escherichia coli-activated macrophages, where ganglioside distribution differs between strains, a difference in galactose oxidase-accessible gangliosides also exists . Escherichia coli-activated C3H/HeN patterns show three triplets absent in C3H/HeJ patterns . There were no differences in ganglioside accessibility to Vibrio cholerae sialidase between the thioglycolate-elicited C3H/HeJ and C3H/HeN macrophages . However, despite differences in sialidase-sensitive ganglioside content between E.coli-activated macrophages of these strains, sialidase accessibility for E.coli-activated macrophages was also similar . Sialidase-susceptible GM3 was cryptic in either strain under all conditions examined . The accessibility of murine macrophage gangliosides to galactose oxidase or sialidase was independent of their sialic acid species and chain length of the ceramide fatty acid . With the exception of GM3, major murine macrophage gangliosides are accessible in situ to macromolecules, especially to exogenous pathogenic bacterial sialidase which can alter macrophage cell surface characteristics . Altered macrophage ganglioside accessibility appears sometimes as a consequence, but not a cause, of C3H/HeJ endotoxin hyporesponsiveness.

Lipids, 1995 Feb, 30(2), 181 - 5
Effect of growth temperature on the positional distribution of eicosapentaenoic acid and trans hexadecenoic acid in the phospholipids of a Vibrio species of bacterium; Henderson RJ et al.; Phosphatidylethanolamine (PE) and phosphatidylglycerol (PG) were isolated from a Vibrio species of bacterium, known to produce eicosapentaenoic acid (20:5n-3) and trans-hexadecenoic acid (16:1n-7), and subjected to phospholipase A2 degradation to determine the positional distribution of component fatty acids . At the two growth temperatures studied (20 and 5 degrees C), both 20:5n-3 and trans 16:1 n-7 were located mainly at position sn-2 in PE . Increases in the proportions of 20:5n-3 and trans 16:1n-7 in position sn-2 with decreasing growth temperature were balanced mainly by decreases in the level of iso-15:0 . In PG, trans 16:1n-7 was located predominantly in position sn-1, although the difference between the two positions was not as great as in PE . Eicosapentaenoic acid was preferentially located in position sn-2 of PG, particularly at 5 degrees C when it comprised 29.9% of the total fatty acids in this position . It is concluded that trans 16:1n-7/20:5n-3 is not a major molecular species of phospholipid in this species of Vibrio and that changes in the levels of molecular species of PE containing iso-15:0 may feature in thermal acclimation.

Acta Paediatr, 1995 Feb, 84(2), 160 - 4
Risk factors for development of dehydration in young children with acute watery diarrhoea: a case-control study; Bhattacharya SK et al.; In a case-control study to understand the risk factors for development of life-threatening dehydration, a total of 379 children comprising 243 cases (moderate or severe dehydration) and 136 controls (non or mild dehydration) up to 2 years of age suffering from acute watery diarrhoea were studied . By univariate analysis, the presence of vibrios in stool, withdrawal of breast feeding during diarrhoea, not giving fluids, including oral rehydration solution (ORS), during diarrhoea, frequent purging ( > 8/day), vomiting ( > 2/day) and undernutrition were identified as risk factors . However, by multivariate analysis after controlling for confounders, withdrawal of breast feeding during diarrhoea (odds ratio (OR) = 6.8, p < 0.00001) and not giving ORS during diarrhoea (OR = 2.1, p < 0.006) were identified as significant risk factors . The confounding variables which also contributed significantly to increasing the risk were age ( < or = 12 months; OR = 2.7, p = 0.001), frequent purging ( > 8/day; OR = 4.1, p < 0.00001), vomiting ( > 2/day; OR = 2.4, p = 0.001) and severe undernutrition (%median < or = 60 weight-for-age of Indian Academy of Paediatrics classification; OR = 3.1, p = 0.001) . We feel that these findings will be useful for Global and National Diarrhoeal Diseases Control Programmes for formulating intervention strategies for preventing death due to diarrhoeal dehydration.

Kansenshogaku Zasshi, 1995 Feb, 69(2), 170 - 4
{Clinical bacteriological analysis of Vibrio vulnificus infection--a report of five case}; Tanabe I et al.; Clinical features in Vibrio infection are generally represented by gastrointestinal involvements such as food poisoning, and its prognosis is usually good . However, Vibrio vulnificus infection not uncommonly causes serious problems including sepsis, necrotizing fasciitis of the extremities, and other conditions, sometimes resulting in fatal outcome . In the present study, we analyzed clinical microbiological aspects of five cases with V . vulnificus infection . All the strains of V . vulnificus isolated in five patients are oxidase-positive Gram negative rods presenting comma-like configuration, which were yielded on TCBS agar forming green colonies; they were grayish-white in color and viscous in texture on 5% sheep blood agar, identification of bacteria were done using VITEK AMS (BioMerieux) . Piperacillin and third-generation cephalosporins were found to have bactericidal activities against these strains . All five cases we experienced have primary ailments, and three cases out of the five had taken perishable sea-food before showing disease symptoms . V . vulnificus has two infection channels; one is external wound and the other is oral intake . The latter is said that it may become serious . This has a rather short period from the starting the symptoms to death, and there is high death rate . For life-saving, it is inevitably necessary to dose an effective antibacterial medicine in the early stage . If we suspect this bacteria in the test laboratory, it is important to report this to the clinical doctor . In other words, this is one of the bacteria that needs prompt treatment and further microbiology testing.

Indian J Med Res, 1995 Feb, 101, 62 - 3
Outbreak due to Vibrio cholerae E1 Tor & serotype O139 in Yavatmal (Maharashtra) during June-July, 1994; Fule RP et al.; A total of 65 strains of V . cholerae were isolated during June-July 1994 at Yavatmal (Maharashtra) . Of the 65 strains isolated, 62 were 01 El Tor Vibrios, while three were non 01 serotype 0139 . The novel epidemic strain designated as 0139 reported during the outbreak in 1993, has been supplanted by the usual El Tor Vibrio during the present outbreak while 0139 serotype has remained sporadic.

Indian J Med Res, 1995 Feb, 101, 57 - 61
Colonization ability & intestinal pathology of rabbits orally fed with Vibrio cholerae O139 Bengal; Koley H et al.; The colonization ability of a representative epidemic strain of V . cholerae O139 Bengal was studied in the oral rabbit colonization model and the nature of colonization in the ileal and jejunal tissues was examined ultrastructurally . Results of the colonization study and ileal loop assay indicated that the strain proliferates and colonizes the small intestine of the rabbit mucosal surface . Further, the electronmicroscopic study revealed the disruptive effect of the strain on the apical membrane of the epithelial cells . The results of this study suggested that apart from colonization, invasion of the bacteria was important in the pathogenesis of V . cholerae O139 mediated infections.

Microbiology, 1995 Feb, 141 ( Pt 2), 377 - 83
Temperature-induced recovery of Vibrio cholerae from the viable but nonculturable state: growth or resuscitation?
Ravel J, Knight IT, Monahan CE, Hill RT, Colwell RR.
Vibrio cholerae cells were incubated at 4 degrees C in nutrient-limited artificial seawater (ASW) microcosms . Plate counts declined from 8 x 10(5) to less than 2 c.f.u . ml-1 in about 23 d . When samples of microcosms were shifted to 30 degrees C, plate counts increased to 2.2 x 10(5) c.f.u . ml-1 in 72 h . An experiment was performed to determine whether culturable cells obtained after temperature upshifts were the result of 'resuscitation', or outgrowth, of nonculturable cells or of cell division and growth of the few culturable cells that remained in samples . Prior to temperature upshift, samples from the microcosms were diluted 10- and 100-fold in filter-sterilized (0.1 microns) ASW from the microcosms . Undiluted, 1/10, and 1/100 diluted samples recovered culturability to about 2.2 x 10(5) c.f.u . ml-1 within 72 h of temperature upshift . If resuscitation of nonculturable cells had occurred, the resultant number of culturable cells in diluted samples would have been 1/10 and 1/100 that of undiluted samples, respectively . In microcosms where plate counts had declined to less than 1 c.f.u . ml-1, 1/100 diluted samples did not regain culturability, i.e . no culturable cells remained from which growth could occur . Our conclusions are that in the experiments reported here, recovery of culturable cells on temperature upshifts resulted from growth and that there were no growth-inhibiting factors in the spent growth medium, supported by the finding that about 10(2) recovered V . cholerae cells ml-1 inoculated into filter-sterilized microcosm ASW grew to about 6.2 x 10(5) c.f.u . ml-1 in 24 h, confirming that V . cholerae is capable of significant growth in ASW.

Toxicon, 1995 Feb, 33(2), 209 - 16
Isolation of mutant toxins of Vibrio parahaemolyticus hemolysin by in vitro mutagenesis; Iida T et al.; Thermostable direct hemolysin produced by Vibrio parahaemolyticus is a major virulence factor of the organism . The hemolysin has a variety of biological activities such as lethality to mice, cytotoxicity to cultured cells, cardiotoxicity, and fluid accumulating activity in rabbit ileal loop test . In this study, we attempted to isolate less hemolytic mutant toxins of the thermostable direct hemolysin to use them for analysis of mode of action of the hemolysin . Six mutant toxins were obtained by in vitro mutagenesis of the cloned gene for the hemolysin . Characterization of the mutant toxins demonstrated that single amino acid substitutions at Gly62, Trp65, Thr67, Gly86, Glu116 and Glu138 resulted in a loss or lowering of the hemolytic activity . Two of the mutant toxins inhibited hemolysis by wild-type toxin on rabbit blood agar plates, while their hemolytic activity was below the detectable level . These mutant toxins would be useful for identifying the as yet unknown receptor for the hemolysin on the target cell membrane.

Appl Environ Microbiol, 1995 Feb, 61(2), 804 - 6
Molecular cloning of the gene which encodes beta-N-acetylglucosaminidase from a marine bacterium, Alteromonas sp . strain O-7; Tsujibo H et al.; The gene encoding the periplasmic beta-N-acetylglucosaminidase (GlcNAcase B) from a marine Alteromonas sp . strain, O-7, was cloned and sequenced . The protein sequence of GlcNAcase B revealed a highly significant homology with Vibrio GlcNAcase and alpha- and beta-chains of human beta-hexosaminidase.

Appl Environ Microbiol, 1995 Feb, 61(2), 476 - 80
Sandwich enzyme-linked immunosorbent assay for Vibrio vulnificus hemolysin to detect V . vulnificus in environmental specimens; Parker RW et al.; Vibrio vulnificus hemolysin, purified by quantitative isoelectric focusing, was used to prepare rabbit and goat anti-hemolysin . The resulting antibodies were used as capture and detector antibody reagents in a sandwich enzyme-linked immunosorbent assay (ELISA) to detect V . vulnificus in environmental samples . By this technique, 4 laboratory-maintained V . vulnificus strains and 33 environmental V . vulnificus isolates were detected . Also, the technique distinguished five other Vibrio species from V . vulnificus, and when it was used in combination with colistin-polymyxin-cellobiose agar, 31 non-V . vulnificus isolated were excluded . This sandwich ELISA compared favorably with the current Food and Drug Administration standard immunoassay in confirming presumptive V . vulnificus colonies from environmental specimens: oysters, sediment, and seawater . Among 340 presumptive V . vulnificus colonies, the sandwich ELISA detected 95% of the confirmed V . vulnificus colonies . Equally important, the technique correctly distinguished 99% of the non-V . vulnificus colonies . The sandwich ELISA offers time-saving and labor-saving advantages over the currently accepted immunoassay.

Vet Microbiol, 1995 Feb, 43(2-3), 167 - 71
A specific oligonucleotide probe based on 5S rRNA sequences for identification of Vibrio anguillarum and Vibrio ordalii; Ito H et al.; An oligonucleotide DNA probe based on 5S rRNA sequence data was constructed for the identification of the fish pathogens, Vibrio anguillarum and Vibrio ordalii . Specificity of the probe was tested in a colony blot hybridization assay . The respective probe was found to be specific for both V . anguillarum and V . ordalii . No cross hybridization was observed against other fish pathogens and the closely related Vibrionaceae genera . This specific probe may be useful for rapid identification of V . anguillarum and V . ordalii.

J Bacteriol, 1995 Feb, 177(3), 758 - 64
Carbohydrate-dependent binding of the cell-free hemagglutinin of Vibrio cholerae to glycoprotein and glycolipid; Saha N et al.; The carbohydrate-binding specificity of the cell-free hemagglutinin (HA) of Vibrio cholerae (K.K . Banerjee, A.N . Ghose, K . Datta-Roy, S.C . Pal, and A.C . Ghose, Infect . Immun.58:3698-3705, 1990) was studied by using glycoconjugates with defined sugar sequences . The HA was not inhibited by simple sugars including glucobiose, galabiose, and their N-acetylated derivatives . The hemagglutination of rabbit erythrocytes by the HA was inhibited moderately by fetuin, calf thyroglobulin, and human alpha 1-acid glycoprotein, all of which contain multiple asparagine-linked complex-type oligosaccharide units alone or in combination with serine/threonine-linked oligosaccharide units . The inhibitory potencies of the glycoproteins increased approximately 10-fold following removal of the terminal sialic acid and were completely destroyed by exhausative proteolysis . The HA agglutinated phosphatidylcholine liposomes containing GM1-ganglioside or its asialo-derivative in the presence of Ca2+ ions . The association constants of the complexes of the HA with asialofetuin, asialothyroglobulin, GM1-ganglioside, and asialo-GM1-ganglioside were determined by an enzyme-linked immunosorbent assay-based assay and found to be 1.7 x 10(7) M-1, 1.5 x 10(7) M-1, 1.8 x 10(7) M-1, and 2.4 x 10(7) M-1, respectively . Studies using chemically modified glycoproteins and plant lectins with defined sugar specificity revealed that the HA recognized the terminal beta 1-galactosyl moiety of these glycoconjugates . There was no evidence for the presence of an extended carbohydrate-binding domain in the HA molecule or a preference of the HA for a complex, branched oligosaccharide structure . Similar to the mechanisms proposed for the binding of cholera toxin and Shiga toxin to glycolipids and neoglycoproteins, the strong interaction of V . cholerae cell-free HA with glycoconjugates appeared to be a consequence of multiple weak binding to terminal beta1-galactosyl moieties of the glycoproteins or glycolipids.

Ugeskr Laeger, 1995 Jan 16, 157(3), 280 - 3
{Is Vibrio cholerae serotype 0139 a potential cause of a new pandemic?}; Dalsgaard DA et al.; Vibrio cholerae O139, a new V . cholerae serotype that does not react with the 138 so far known antisera, was first isolated in 1992 in Madras, India . Although a V . cholerae non-O1, it behaved quite differently from this particular group of organisms by its clinical appearance in causing epidemic cholera-like disease . The organism possessed the same register of virulence factors as V . cholerae O1 . Biochemical and genetic analyses have shown that V . cholerae O139 was closely related to the El Tor biotype and it has been suggested that it might be a mutant of this biotype . Despite the fact that V . cholerae O139 isolates were very homologous in many respects, ribotyping of Thai isolates showed a certain degree of genetic diversity . From its rapid spread in populations, pattern of infection and ability to survive in aquatic environments, it may be suggested that we are dealing with a more infectious, more virulent and in an ecological sense more robust organism, whose pandemic potential appears significant . Commercial cholera vaccines offer no or only limited protection . Treatment comprises an adequate rehydration and use of effective antibiotics--at present tetracycline, ampicillin, erythromycin and ciprofloxacin . Tourism and perhaps the import of foods from Third World countries may excert a potential risk for the Danish population.

EMBO J, 1995 Jan 16, 14(2), 209 - 16
Genesis of the novel epidemic Vibrio cholerae O139 strain: evidence for horizontal transfer of genes involved in polysaccharide synthesis; Bik EM et al.; Only Vibrio cholerae strains of serotype O1 are known to cause epidemics, while non-O1 strains are associated with sporadic cases of cholera . It was therefore unexpected that the recent cholera epidemic in Asia was caused by a non-O1 strain with the serotype O139 . We provide evidence that O139 arose from a strain closely related to the causative agent of the present cholera pandemic, V . cholerae O1 El Tor, by acquisition of novel DNA which was inserted into, and replaced part of, the O antigen gene cluster of the recipient strain . Part of the novel DNA was sequenced and two open reading frames (otnA and otnB) were observed, the products of which showed homology to proteins involved in capsule and O antigen synthesis, respectively . This suggests that the otnAB DNA determines the distinct antigenic properties of the O139 cell surface . The otnAB DNA was not detected in O1 strains, but was present in two non-O1 V . cholerae strains with serotypes O69 and O141 . In the O69 and O139 strains the otnAB genes were located proximate to the putative insertion sequence (IS) element rfbQRS, which is associated with O antigen synthesis genes in O1 strains, and may have played a role in the insertion of the otnAB DNA in the recipient chromosome . Our results suggest that the O139 strain arose by horizontal gene transfer between a non-O1 and an O1 strain . The acquired DNA has altered the antigenic properties of the recipient O1 strain, providing a selective advantage in a region where a large part of the population is immune to O1 strains.(ABSTRACT TRUNCATED AT 250 WORDS)

FEMS Microbiol Lett, 1995 Jan 15, 125(2-3), 205 - 9
Detection and differentiation of the gene for toxin co-regulated pili (tcpA) in Vibrio cholerae non-O1 using the polymerase chain reaction; Said B et al.; The polymerase chain reaction has been used to differentiate the gene which encodes the toxin co-regulated pili (tcpA) of the El Tor and classical biotypes of Vibrio cholerae O1 . The same PCR primers were applied to strains belonging to non-O1 serogroups that produced cholera toxin . The size of fragment amplified was either identical to the tcpA of biotype El Tor (471 bp) or to the tcpA of biotype classical (617 bp) . All strains belonging to the novel epidemic serogroup O139 generated a 471-bp fragment identical to El Tor tcpA . The present study suggests that there may be an association between non-O1 serogroup and tcpA type.

Gene, 1995 Jan 11, 152(1), 59 - 63
Cloning and characterization of the gene encoding an extracellular alkaline serine protease from Vibrio metschnikovii strain RH530; Kwon YT et al.; The gene vapF, encoding VapT, one of the extracellular sodium dodecyl sulfate (SDS)-resistant alkaline serine proteases (Serp) from the Gram- Vibrio metschnikovii strain RH530 has been cloned in Escherichia coli . The recombinant E . coli produced a protease which co-migrated with VapT on gelatin polyacrylamide gels . The nucleotide (nt) sequence of the cloned vapT revealed a single open reading frame of 1641 bp encoding 547 amino acids (aa) (58,961 Da) . Upon analysis of the N-terminal aa sequence, VapT was shown to be processed properly in recombinant E . coli and to consist of 428 aa (45,626 Da) . The deduced aa sequence of VapT showed significant sequence homology to subtilisin Carlsberg from Bacillus licheniformis, particularly in the regions containing active site residues and calcium-binding sites . VapT had an intervening region of approx . 149 aa between the His and Ser residues of the active site, as compared with other Serp.

Gene, 1995 Jan 11, 152(1), 135 - 6
The deduced Vibrio cholerae RecA amino-acid sequence; Margraf RL et al.; The nucleotide sequence of the recA gene of Vibrio cholerae (Vc) has been determined . The amino acid (aa) sequence of the protein product is very similar to other known RecA aa sequences . However, this sequence does not agree with a previously reported Vc RecA aa sequence {Ghosh et al., Nucleic Acids Res . 20 (1992) 372}.

Arch Med Res, 1995, 26 Spec No, S47 - 53
In vitro growth of Vibrio cholerae in cholera stool fluid leads to differential expression of virulence factors; Sanchez J et al.; We report on the physiological response of Vibrio cholerae upon growth on bacteria-free intestinal fluids prepared from feces of individuals in the acute phase of cholera . Sterilized stool fluids supported growth of V . cholerae to reach 0.3-0.4 O.D . units (600 nm) at 37 degrees C . Scanning electron microscopy showed vibrios to be slender and elongated as compared to bacteria in synthetic media . Growth in stool fluid apparently induced expression of several immunoreactive proteins using cholera convalescent sera . Supernatants of fluid-grown vibrios had undetectable cholera toxin (CT) concentrations . Soluble hemagglutinins and soluble proteases were much less reduced when compared to cultures in Syncase or AKI media while cell-associated mannose-sensitive hemagglutinin (MSHA) was expressed at good levels . Lack of production of CT in fluid devoid of tissue may be due to absence of stimulating elements in intact intestine . Alternatively, culturing V . cholerae in stool fluid might resemble a late proliferation stage where downregulation of toxin might occur . Irrespectively, concomitant production of other virulence factors represents a phenomenon of differential regulation by fluid . Efforts are now underway to determine if this response depends upon factors in stool fluid acting through known genetic regulatory cascades or other . Attempts are also geared to identify fluid-induced proteins and their genes.

Microbiol Immunol, 1995, 39(12), 1003 - 6
Rapid and sensitive PCR detection of Vibrio trachuri pathogenic to Japanese horse mackerel (Trachurus japonicus); Iwamoto Y et al.; A polymerase chain reaction (PCR) method was performed for rapid and sensitive detection of pathogenic Vibrio trachuri isolated from cultured Japanese horse mackerel . A set of primers was selected from the base sequence of the Pst I fragment of T9210 chromosomal DNA and used for PCR detection of T9210 . This PCR specifically amplified the DNAs from V . trachuri T9210, T9213, and T9216 but not of those other bacterial strains . PCR using a Pst I-1 primer set made it possible to detect 100 fg of T9210 DNA . The PCR method reported here may be useful for detection and identification of V . trachuri pathogenic to Japanese horse mackerel.

Microbiol Immunol, 1995, 39(12), 959 - 66
Actions of Vibrio vulnificus metalloprotease on human plasma proteinase-proteinase inhibitor systems: a comparative study of native protease with its derivative modified by polyethylene glycol; Miyoshi S et al.; Vibrio vulnificus, an opportunistic human pathogen causing wound infection and septicemia, produces a metalloprotease (VVP) which is suspected to be a virulent determinant . The interactions of VVP, as well as its derivative (PEG1-VVP) modified with polyethylene glycol, with a variety of human plasma proteins were investigated . We found that native VVP and its derivative were able to act directly on many biologically important human plasma proteins even in the presence of alpha-macroglobulin, the sole plasma inhibitor of native VVP . The activities of both classical and alternative pathways of the complement cascade system were drastically abolished by incubation with either VVP . Furthermore, these proteases rapidly digested the A alpha-chain of human fibrinogen into fragment(s) with no clotting ability . Therefore both VVPs are thought to function as a fibrinogenolytic enzyme, causing delay of the coagulation reaction . VVP and PEG1-VVP were also shown to destroy plasma proteinase inhibitors including alpha 1-proteinase inhibitor, a major inhibitor in human plasma . Because endogenous proteolytic enzymes and their inhibitors are indispensable in maintaining physiological homeostasis, these findings suggest that VVP (and PEG1-VVP) may cause an imbalance of human plasma proteinase-proteinase inhibitor systems, thus eliciting an immunocompromised state in the host and facilitating the development of a systemic V . vulnificus infection such as septicemia.

Antisense Res Dev, 1995 Winter, 5(4), 261 - 9
Nonsequence-specific inhibition of bacterial luminescence by phosphorothioate oligodeoxyribonucleotides; Chrisey LA et al.; To evaluate the effect of synthetic DNA oligomers on regulation of bacterial genes in vivo, we tested 63 oligomers of variable length and chemistry for their ability to selectively suppress light production in the bioluminescent marine organism, Vibrio fischeri . Phosphodiester, phosphorothioate, and mixed backbone oligomers were designed to be lux gene targeted or nontargeted (negative) controls . Although significant suppression of luminescence was observed, most notably with the phosphorothioate oligomers, there was no correlation between inhibitory activity and oligomer sequence . The phosphorothioate oligomer that was most potent for inhibition of luminescence in bacterial culture had no effect on the activity of purified luciferase . Mechanisms other than sequence-specific inhibition of gene expression or direct interaction with luciferase are discussed.

Scand J Infect Dis, 1995, 27(6), 585 - 8
Restriction fragment length polymorphisms (RFLP) of cholera toxin genes in Vibrio cholerae O139 recovered from patients in Thailand, India and Bangladesh; Dalsgaard A et al.; Since its first appearance in 1992, Vibrio cholerae O139 has caused large epidemics of a cholera-like disease in India and Bangladesh and has subsequently spread to several neighboring countries . We characterized and compared 56 V . cholerae O139 isolates recovered from patients in Thailand, India and Bangladesh by analyzing restriction fragment length polymorphisms (RFLP) of their ctx genes . The strains comprised 9 different BglI cleavage patterns of ctx genes (CT genotypes) and contained 1-4 gene copies . A total of 6 different CT genotypes were found among the 52 Thai isolates studied whereas the 2 Indian isolates and 1 isolate from Bangladesh all showed unique CT genotypes . The molecular analysis of ctx genes appeared superior to ribotyping for subspecies differentiation of O139 strains since ribotyping revealed 3 different BgII ribotypes . In addition, no correlation was found between the two methods . The molecular analysis of virulence determinants such as ctx genes in combination with other molecular techniques appears to be promising for the study of genetic changes within V . cholerae O139.

Scand J Infect Dis, 1995, 27(4), 425 - 6
Short-term quinolones for successful eradication of multiply resistant Vibrio cholerae in adult patients; Doganci L et al.; There has been an increasing multiple drug resistance problem in Vibrio cholerae biotype Eltor, the causative agent of 7th pandemic . The aim of this study was to show in vitro and in vivo susceptibility and effectiveness of quinolones in the treatment of endemic cholera cases . Excellent results were obtained in 53 bacteriologically confirmed cholera patients treated with short-term ofloxacin and ciprofloxacin . To our knowledge, there has been no previous report on this subject in the international medical literature . Our results show that quinolones can be an alternative drug for the treatment of multiply resistant V . cholerae infections.

Microbiol Immunol, 1995, 39(11), 891 - 4
Flow cytometric analysis using lipophilic dye PKH-2 for adhesion of Vibrio cholerae to Intestine 407 cells; Taguchi H et al.; A comparative study of indirect and direct flow cytometric analysis for adherence of Vibrio cholerae to Intestine 407 cells was performed . The direct flow cytometric analysis employed the lipophilic dye PKH-2 . It was concluded that direct flow cytometry using the lipophilic dye PKH-2 is useful and convenient for analyzing bacterium-host cell interactions, since it does not require any specific antibody as the first antibody.

Microbiol Immunol, 1995, 39(11), 831 - 37
Vibrio trachuri sp . nov., a new species isolated from diseased Japanese horse mackerel; Iwamoto Y et al.; A new species, Vibrio trachuri sp . nov., was isolated from the cultured Japanese horse mackerel (Trachurus japonicus) . These Vibrio were Gram negative, motile rods and formed yellow colonies on BTB teepol and TCBS plate, turned TSI medium to yellow and was sensitive to 150 microM O/129 (2,4-diamino-6,7-diisopropyl pteridine phosphate) like Listonella anguillarum which has been described as Vibrio anguillarum . However, the results of VP test and decarboxylation of lysine or dihydrolation of arginine suggested that these Vibrio are rather closely related to V . parahaemolyticus . DNA similarity determined by the microplate hybridization technique revealed that these Vibrio are genetically quite distant from Listonella anguillarum or V . parahaemolyticus and rather close to V . harveyi, although there was no Vibrio species which had more than 70% similarity value . From these results we propose to nominate Vibrio trachuri sp . nov . for this new Vibrio species.

Trop Geogr Med, 1995, 47(6), 305 - 7
Shellfish-borne illnesses . A Hong Kong perspective; Chan TY; This article provides an overview of the spectrum of infectious and toxic illnesses that may occur following the consumption of contaminated shellfish in Hong Kong . These include hepatitis A, hepatitis E, infections due to vibrio species, paralytic shellfish poisoning, neurotoxic shellfish poisoning and heavy metal poisoning . Possible preventive measures are discussed.

Microbiol Immunol, 1995, 39(10), 759 - 66
Demonstration of a ferric vibrioferrin-binding protein in the outer membrane of Vibrio parahaemolyticus; Yamamoto S et al.; Under iron-restricted conditions, Vibrio parahaemolyticus produces a siderophore, vibrioferrin, accompanying expression of two major outer membrane proteins of 78 and 83 kDa . Autoradiographic analysis of nondenaturing polyacrylamide gel electrophoregrams of outer membrane preparations previously incubated with {35Fe}ferric vibrioferrin revealed a single radiolabeled band, in which the 78-kDa protein was detected predominantly by sodium dodecyl sulfate-polyacrylamide gel electrophoresis . The antiserum against the purified 78-kDa protein partially inhibited Fe-VF binding to isolated OMPs . The 78-kDa protein was cleaved by the treatment of whole cells with proteinase K, indicating that a portion of this protein is exposed on the surface of the outer membrane . The treated cells lost most of their iron uptake activity mediated by vibrioferrin . These results suggest that the ferric vibrioferrin-binding protein of 78 kDa may function as the receptor for ferric vibrioferrin involved in the initial step of vibrioferrin-mediated iron uptake . Immunoblot analysis using the antiserum against the 78-kDa protein demonstrated that the molecular mass and antigenic properties of the protein were highly conserved among V . parahaemolyticus strains examined . The antiserum also recognized an iron-repressible outer membrane protein of 78 kDa from iron-restricted V . alginolyticus strains, some of which appeared to produce vibrioferrin.

Microbiol Immunol, 1995, 39(10), 741 - 4
The measurement of swimming velocity of Vibrio cholerae and Pseudomonas aeruginosa using the video tracking methods; Shigematsu M et al.; The swimming velocities of two monotrichous flagellated bacteria were measured by a computer-assisted video tracking method . Tracing the moving path of the individual bacterium revealed that the bacterial cell did not swim continuously in a straight direction, but frequently changed swimming direction and velocity . The average swimming velocities calculated from the 3-sec path were 75.4 +/- 9.4 microns/sec in four strains of Vibrio cholerae and 51.3 +/- 8.4 microns/sec in five strains of Pseudomonas aeruginosa . These results suggest that V . cholerae swim faster than P . aeruginosa at 30 C in nutrient broth . This method is useful for a detailed analysis of bacterial movement and moving patterns in different environmental conditions.

Infect Immun, 1995 Jan, 63(1), 301 - 8
Expression and purification of Shiga-like toxin II B subunits; Acheson DW et al.; Shiga-like toxins (SLTs), which are produced by certain strains of Escherichia coli, are composed of enzymatically active A and B subunit multimers responsible for the toxin's binding . We have previously purified large amounts of the SLT-I B subunit by using a hyperexpression vector in Vibrio cholerae under the control of the trc promoter . In this study we examined various expression vectors to maximize yields of the SLT-II B subunit . The SLT-II B subunit has been expressed by using both the T7 promoter and the tac promoter in E . coli . When expressed from a plasmid containing the structural gene for SLT-II B deleted of the leader sequence, SLT-II B was able to form multimers when cross-linked, although SLT-II B production from this plasmid was unreproducible . SLT-II B expressed in all three systems appeared to form unstable multimers, which did not readily bind to a monoclonal antibody which preferentially recognizes B subunit multimers . SLT-II B expression was not increased by moving any of the plasmids into V . cholerae . Polyclonal antibodies raised to SLT-II B in rabbits recognized B subunit in SLT-II holotoxin yet were poorly neutralizing . SLT-II B was also expressed as a fusion protein with maltose-binding protein and could be cleaved from maltose-binding protein with factor Xa . Although the expression vectors were able to make large amounts of SLT-II B, as determined by Western blotting (immunoblotting), the levels of purified SLT-II B subunit were low compared with those obtained previously for SLT-I B subunit, probably because of instability of the multimeric SLT-II B subunit.

Dev Biol Stand, 1995, 84, 237 - 44
Vibrio cholerae CVD103-HgR live oral attenuated vaccine: construction, safety, immunogenicity, excretion and non-target effects; Cryz SJ Jr et al.; In many controlled studies, CVD103-HgR has been shown to be safe and immunogenic and to offer a significant degree of protection against experimental cholera after a single dose . Its minimal excretion and limited ability to compete and survive in various ecosystems indicate that this strain presents little risk to the environment . Furthermore, the potential of CVD103-HgR to regain virulence by acquisition of the CT A or LT A gene is extremely remote even under optimal conditions . Therefore, CVD103-HgR possesses those traits desired in a live oral attenuated vaccine produced by recombinant DNA technology.

Scand J Infect Dis, 1995, 27(1), 81 - 2
First documented case of bacteremia with Vibrio vulnificus in Sweden; Melhus A et al.; A few days after a mild trauma to a toe, a 90-year-old woman presented with fever, malaise and cellulitis . On suspicion of erysipelas the patient was initially treated with benzylpenicillin and cefuroxime . Her general condition improved rapidly, but there was local progression with numerous necrotic areas with surrounding bullae . Vibrio vulnificus was isolated from the blood . After susceptibility testing, the patient was finally treated with ciprofloxacin and pivampicillin, and recovered slowly . To our knowledge, this is the first reported case of bacteremia with V . vulnificus in Sweden.

Rev Latinoam Microbiol, 1995 Jan-Mar, 37(1), 27 - 31
A discrete factor A is detected on Vibrio cholerae Non-O1, O:139 by laser flow cytometry; Alvarado-Aleman FJ et al.; A new serogroup of Vibrio cholerae non-O1, O:139, has been implicated in recent epidemics . It was scanned with a factor A-specific fluoresceine-conjugated monoclonal antibody, searching for antigen determinants by laser flow cytometry . First, a group of gram-negative 4-amine-4, 6 dideoxy-D-mannose antigen-related microorganisms were tested to assess monoclonal antibody cross reactions . Later, a clear recognition of antigen determinants was found with this monoclonal antibody, on V . cholerae non-O1, O:136, Bengal, and MO45 strains, showing no cross reactions with the antigenically related non O1, O:22, and Inaba and Ogawa O1 strains . On the other hand, factor A of V . cholerae O1 strains was recognized by the specific monoclonal antibody and a discrete factor A on V . cholerae non-O1, O:139, Bengal and MO45 strains was detected.

Microbiol Immunol, 1995, 39(2), 87 - 94
Characterization of an enterotoxin produced by Vibrio cholerae O139; Nakashima K et al.; A cholera-like enterotoxin was purified from Vibrio cholerae O139 strain AI-1841 isolated from a diarrheal patient in Bangladesh . Its characteristics were compared with that of cholera toxins (CTs) of classical strain 569B and El Tor strain KT25 . Al-1841 produced as much toxin as O1 strains . The toxins were indistinguishable in terms of their migration profiles in conventional polyacrylamide gel disc electrophoresis, sodium dodecyl sulfate-polyacrylamide gel electrophoresis and isoelectrofocusing as well as their affinity for hydroxyapatite . The skin permeability factor activity and the fluid accumulation induced in rabbit ileal loops of the toxin of AI-1841 were identical to those of the CTs . Three toxins equally reacted against anti-569B CT antiserum in Western blotting, and their B subunits formed a precipitin line against any anti-B subunit antiserum by double gel immunodiffusion . Anti-569B CTB antibody neutralized the three toxins in their PF activities and enterotoxicities . The amino acid sequence of 1841 toxin B subunit was identical with that of KT25 CTB, corresponding to the DNA sequence of ctxB from El Tor strains of the seventh pandemic . We concluded 1841 toxin was identical to CT of the seventh pandemic El Tor vibrios.

Microbiol Immunol, 1995, 39(2), 157 - 9
Evaluation of two assay kits for thermostable direct hemolysin (TDH) as an indicator of TDH-related hemolysin (TRH) produced by Vibrio parahaemolyticus; Yoh M et al.; Reversed passive latex agglutination (RPLA) or enzyme-linked immunosorbent assay kits with beads (Bead-ELISA) are commercially available in Japan to detect the thermostable direct hemolysin (TDH) produced by Vibrio parahaemolyticus isolates . We evaluated whether these kits can be used to assay the pathogenic toxin, TDH-related hemolysin (TRH), produced by some so-called Kanagawa phenomenon-negative V . parahaemolyticus strains isolated from patients with diarrhea . Our results showed that the two kits, RPLA and Bead-ELISA, can detect TRH, although they were originally developed for detection of TDH . This may be due to the use of polyclonal anti-TDH antisera that cross react with TRH . Although the sensitivity for TDH detection by RPLA and Bead-ELISA differed tenfold, that for TRH detection was essentially equal . The minimum concentration of TRH required for detection by the two assay kits was about 10 ng/ml.

Microbiol Immunol, 1995, 39(1), 67 - 70
Production of antigenically related exocellular elastolytic proteases mediating hemagglutination by vibrios; Alam M et al.; Exocellular proteases produced by Vibrio fluvialis, V . furnissii, V . metschnikovii and V . campbellii were characterized and compared to those of V . mimicus protease (VMP) and V . vulnificus protease (VVP) . These proteases possessed both elastolytic and hemagglutinating abilities and were identified, except that of V . metschnikovii, as metalloprotease . Conversely, V . metschnikovii protease failed to exhibit some of the salient features for metalloproteases suggesting the existence of protease(s) other than metalloprotease . However, antibodies against VVP cross-reacted to these proteases and to VMP indicating antigenic relatedness amongst vibrio proteases . This study, thus, demonstrated the prevalent distributions of antigenically related proteases both in pathogenic and non-pathogenic vibrios, bringing their status as a virulence determinant into question.

Microbiol Immunol, 1995, 39(1), 59 - 61
Comparison of a reversed passive latex agglutination and a polymerase chain reaction for identification of cholera toxin producing Vibrio cholerae O1; Honma Y et al.; Production of cholera toxin (CT) in AKI medium and conservation of CT gene (ctx) of 49 strains of Vibrio cholerae O1 were compared by reversed passive latex agglutination (RPLA) and polymerase chain reaction (PCR) . The production of CT agreed with conservation of the ctx in 48 out of the 49 strains . Ten strains were positive, and 38 strains were negative by both methods . Only one strain was negative in RPLA and positive in PCR . This suggested that the combination of AKI-SW and RPLA is comparable to PCR to identify CT-producing V . cholerae O1.

Phytochemistry, 1995 Jan, 38(1), 167 - 9
An antimicrobial abietane from the root of Plectranthus hereroensis; Batista O et al.; A new abietane diterpene, 16-acetoxy-7 alpha, 12-dihydroxy-8,12-abietadiene-11,14-dione, has been isolated from the acetone extract of the root of Plectranthus hereroensis and its structure established by spectroscopic means . This compound showed antibacterial activity against Staphylococcus aureus and Vibrio cholerae, and antiviral activity against Herpes simplex type II.

Trans R Soc Trop Med Hyg, 1995 Jan-Feb, 89(1), 75 - 7
Extension of the volunteer challenge model to study South American cholera in a population of volunteers predominantly with blood group antigen O; Tacket CO et al.; The volunteer challenge model was used to study the virulence of strains of Vibrio cholerae O1 El Tor recently isolated from cases of cholera in South America . Fifteen of the 24 volunteers (62%) were of blood group O, the group most prevalent in South America and the group at increased risk of more severe cholera . Two El T or Inaba strains and 2 El Tor Ogawa strains were given to volunteers at a dose of 1-2 x 10(6) colony-forming units . All 4 strains caused diarrhoea in 67-83% of volunteers . Volunteers with blood group antigen O had an increased attack rate for diarrhoea (P = 0.015) and a marginally increased mean diarrhoeal stool volume (P = 0.08) after challenge . One-third of the volunteers with blood group O, and none of the volunteers with other blood groups, developed severe diarrhoea (> 5 L) (P = 0.01) . This study established a model of South American cholera that can be used to predict field efficacy of candidate vaccines among populations with a high prevalence of blood group antigen O.

Trans R Soc Trop Med Hyg, 1995 Jan-Feb, 89(1), 103 - 6
Comparative trial of five antimicrobial compounds in the treatment of cholera in adults; Khan WA et al.; To compare the efficacy of ciprofloxacin, erythromycin, nalidixic acid and pivmecillinam in the treatment of tetracycline-resistant strains of Vibrio cholerae O1 in adults, a randomized, open, clinical trial was conducted . A tetracycline group was used for comparison . Seventy-five adult men infected with V . cholerae O1 were randomly assigned to receive either 400 mg pivmecillinam or 500 mg of one of each of the other drugs . Ciprofloxacin was given every 12 h and the others every 6 h for 3 d . The mean total stool volume per kg was 155 mL for the ciprofloxacin group, 212 mL for the erythromycin and pivmecillinam groups, 246 mL for nalidixic acid, and 293 mL for tetracycline . The difference between ciprofloxacin and tetracycline was significant (P = 0.045) . After 72 h, diarrhoea had stopped in 14 patients (93%) in the ciprofloxacin group and 12 (80%) in the erythromycin group, compared to 5 (42%) of those receiving tetracycline (P = 0.006 and 0.049, respectively) . Bacteriological clearance was 100% at 24 h in patients treated with ciprofloxacin compared to 20% and 8.3% (P < 0.001 for both comparisons) in the erythromycin and tetracycline groups . Ciprofloxacin in conjunction with appropriate fluid therapy was the most effective treatment for cholera in adults; erythromycin was the next best.

J Med Microbiol, 1995 Jan, 42(1), 20 - 5
Studies on the genesis of Vibrio cholerae O139: identification of probable progenitor strains; Pajni S et al.; Four lines of evidence suggest that the recent outbreak strains of Vibrio cholerae O139 could have emerged from serogroup O1 strains typified by isolates M01 and M0477 described in this paper, which are neither truly classical nor truly E1 Tor in their biotype attributes . Firstly, like all O139 isolates, these O1 strains, isolated in Madras during and before the O139 outbreak, were resistant not only to polymyxin B but also to all biotype-specific choleraphages, i.e . classical phage phi 149 and E1 Tor phages e4 and e5 . Secondly, the restriction fragment pattern (RFP) polymorphism displayed by these strains for the cholera toxin (ctx) gene, were identical with those produced by O139 isolates but were different from those of O1 type strains, namely V . cholerae 569B (classical) and V . cholerae MAK757 (E1 Tor) . Thirdly, all the O139 isolates and the two O1 isolates carried an identical large number of copies of cholera toxin gene in their chromosomes . Finally, the outer-membrane protein profiles of strains M01 and M0477 were identical to those of O139 isolates but were different from those displayed by strains 569B and MAK757.

Br Vet J, 1995 Jan-Feb, 151(1), 45 - 69
Vaccination in European salmonid aquaculture: a review of practices and prospects; Press CM et al.; Disease control by vaccination is widely used in European salmonid aquaculture against vibriosis (Vibrio anguillarum), cold-water vibriosis (Vibrio salmonicida), yersiniosis or enteric redmouth disease (Yersinia ruckeri) and furunculosis (Aeromonas salmonicida subsp . salmonicida) . The vaccines against the Vibrio spp . and Y . ruckeri have proven effective especially when administered by injection . Furunculosis vaccines have been less successful and have relied on combination with potent adjuvants to achieve acceptable protection . Application of modern molecular techniques to furunculosis research has delivered a crop of experimental vaccines that incorporate purified virulence factors and have shown increased protection during challenge . Gene technology has also been used to create a defined, nonreverting mutation in a strain of A . salmonicida, which has enhanced the feasibility of attenuated live vaccines . The development of experimental subunit vaccines against the viral infections and the continued advances in the field of immunostimulants, adjuvants and antigen carriers provide considerable promise for the future development of commercial vaccines for use in salmonid aquaculture.

Clin Microbiol Rev, 1995 Jan, 8(1), 48 - 86
Cholera; Kaper JB et al.; Despite more than a century of study, cholera still presents challenges and surprises to us . Throughout most of the 20th century, cholera was caused by Vibrio cholerae of the O1 serogroup and the disease was largely confined to Asia and Africa . However, the last decade of the 20th century has witnessed two major developments in the history of this disease . In 1991, a massive outbreak of cholera started in South America, the one continent previously untouched by cholera in this century . In 1992, an apparently new pandemic caused by a previously unknown serogroup of V . cholerae (O139) began in India and Bangladesh . The O139 epidemic has been occurring in populations assumed to be largely immune to V . cholerae O1 and has rapidly spread to many countries including the United States . In this review, we discuss all aspects of cholera, including the clinical microbiology, epidemiology, pathogenesis, and clinical features of the disease . Special attention will be paid to the extraordinary advances that have been made in recent years in unravelling the molecular pathogenesis of this infection and in the development of new generations of vaccines to prevent it.

Antimicrob Agents Chemother, 1995 Jan, 39(1), 241 - 4
Survey of in vitro susceptibilities of Vibrio cholerae O1 and O139 to antimicrobial agents; Yamamoto T et al.; Vibrio cholerae O139 (173 strains) and O1 (221 strains) were tested for their in vitro susceptibilities to 39 antimicrobial agents . Both O139 and O1 strains were highly susceptible to azithromycin, cephems, minocycline, penems, and newer fluoroquinolones . O139 strains (94.8%), O1 Indian El Tor strains (97%), and Bangladeshi El Tor strains (50%) were highly resistant to streptomycin, sulfamethoxazole, and trimethoprim and moderately resistant to chloramphenicol and furazolidone, in sharp contrast to O1 Peruvian El Tor and O1 classical strains . Some Bangladeshi El Tor strains (43.3%) showed tetracycline resistance as well.

West Afr J Med, 1995 Jan-Mar, 14(1), 1 - 5
Isolation and characterization of the exo-enterotoxin of Vibrio cholerae strain 110 Cal; Chukwumah EI et al.; Vibrio Cholerae "strain 110 Cal" from Calabar, Nigeria were grown in syncase broth . Exo-enterotoxin secreted into the medium was then isolated purified and characterized . The toxin had enterotoxic activity using the infant mouseassay of World Health Organization . The toxin also had a molecular weight of about 89,000 daltons by gel filtration through sephadex G-150 and 100,000 daltons by polyacrylamide gel electrophoresis (PAGE) . The toxin exhibited two subunits with molecular weights of about 63,000 and 31,000 daltons on SDS-PAGE . The toxin was resolved through sephadex G-150 into three peaks having molecular weights of about 89,000, 44,000 and 14,000 daltons respectively . The three peaks had enterotoxic activity . Each peak contained a single protein band on PAGE and SDS-PAGE showed that peak I consisted of two subunits with molecular weights of 56,000 and 31,000 daltons and peak II and III were monomers.

DNA Seq, 1995, 5(3), 181 - 4
The Vibrio cholerae hlyC gene encodes a protein that is related to lipases of pseudomonas species; Casanova TB et al.; The nucleotide sequence of the Vibrio cholerae N16961 hlyC gene was determined . The hlyC gene encompasses 513 nucleotides that are predicted to encode a 171-amino acid protein with a calculated molecular weight of 18.2 kDa . The predicted HlyC protein contains a region that is 93.5% similar to the substrate-binding/catalytic domain of the Pseudomonas species triacylglycerol acylhydrolase (lipase) . The proposed catalytic serine residue is also conserved in the HlyC protein . The contribution of the putative HlyC lipase to the physiology of V . cholerae is currently under investigation.

Microbiol Immunol, 1995, 39(3), 213 - 5
Nutrient starvation induces cross protection against heat, osmotic, or H2O2 challenge in Vibrio parahaemolyticus; Koga T et al.; Glucose-starved cells of Vibrio parahaemolyticus were compared with non-starved counterparts with respects to heat, osmotic, and oxidative challenges . The starved cells demonstrated greater thermal and oxidative resistance than did the non-starved cells . The starved cells also showed greater resistance against low osmotic challenge than did the non-starved cells although both cells showed a comparable resistance against high osmotic challenge.

Acta Vet Scand, 1995, 36(1), 55 - 64
Antibiotic resistance of Vibrio anguillarum, in relation to serovar and plasmid contents; Pedersen K et al.; A total of 520 Vibrio anguillarum strains, isolated from fish and the environment, were tested for their sensitivity to 20 different antibiotics . Most isolates were of European origin . The results were compared with data on the O-serogroup and plasmid contents . All strains were sensitive to neomycin, spectinomycin, nitrofurantoin, flumequine and oxolinic acid, while most strains were sensitive to streptomycin, oxytetracycline, chloramphenicol, sulphonamides, trimethoprim, sulphonamides with trimethoprim, nalidixan, rifampicin, novobiocin and O/129 . A major part of the strains were resistant to the macrolides, spiramycin and lincomycin . For ampicillin, cephalothin, and colistin marked differences were recorded with respect to O-serogroup . Most O1 strains were resistant to colistin and sensitive to ampicillin and cephalothin, while most O2 strains were sensitive to colistin but resistant to ampicillin and cephalothin . Some antibiotic resistant strains carried plasmids but no conjugation experiments were carried out to detect possible R factors.

Medicina (B Aires), 1995, 55(1), 28 - 32
{The role of food in cholera transmission}; Dobosch D et al.; The spreading of cholera, from Peru to other Latinoamerican countries in 1991, raised questions regarding food safety, food transportation and handling . Control, prevention and risks implied in food import-export were also matters of concern . We deemed it interesting to determine the viability of Vibrio cholerae in wide consumption food locally . Selected food had different intrinsic characteristics such as: acidity (pH), water activity (aw), chemical composition, indigenous flora and other biologic and physic parameters . Twenty food products were contaminated with V . cholerae O1, Ogawa, toxigenic and not toxigenic strains: yoghurt, cream cheese, apricot marmelade, hip rose marmelade, mayonnaise, italian pasta for "empanadas", "dulce de leche", meat sausage, meat and spinach ravioli, margarine, milk dessert (made with cocoa, milk confiture, starch and additives), lettuce, tuna fish, ricotta and sterilized milk . Table I shows the viability of V . cholerae in tested foods, its pH and the reasons why the experiments were ended: 75% of the products studied could tolerate the development of the microorganism for a period ranging from one day (pasta for "empanadas") to ninety days (sterilized milk) . Foods with acredity higher than pH 5.5 did not favor the growth of Vibrio . When pH was neutral or slightly acid, viability persisted independently from aw, microbial antagonisms and other physic, chemical or biologic parameters . Nevertheless, other factors such as: surface adherence, amino acids, magnesium and environmental influences not yet well determined, could eventually modify the persistence of V . cholerae in food . According to this study, most food products could tolerate growth and persistence of the infectant agent, up for three months in some cases.(ABSTRACT TRUNCATED AT 250 WORDS)

J Clin Microbiol, 1995 Jan, 33(1), 114 - 8
New variant of Vibrio cholerae O1 from clinical isolates in Amazonia; Coelho A et al.; A survey of pathogenic Vibrio cholerae O1 strains from the north of Brazil by using arbitrarily primed PCR fingerprints revealed a group of strains with similar fingerprint patterns that are distinct from those of the current El Tor epidemic strain . These strains have been analyzed by in vivo and in vitro techniques and the group has been denominated the Amazonian variant of V . cholerae O1.

Appl Environ Microbiol, 1995 Jan, 61(1), 245 - 51
Application of ribotyping for differentiating Vibrio cholerae non-O1 isolated from shrimp farms in Thailand; Dalsgaard A et al.; A collection of 143 Vibrio cholerae non-O1 strains isolated from shrimp farms in Thailand were characterized and grouped by ribotyping . Sixty-four ribotypes were distinguished following digestion of chromosomal DNA with the restriction enzyme BglI, and the reproducibility of the method was 100% . There was no correlation between specific ribotype distributions and the locations of the shrimp farms . Ribotype similarity was examined by cluster analysis, and two main groups with 10 and 54 ribotypes, respectively, were found . Correlation between ribotype and O-antigen expression was shown to exist among those isolates tested . Ribotyping appears to be a suitable method for differentiating environmental V . cholerae non-O1 strains, and comparison of ribotype patterns showed a high degree of genetic divergence within V . cholerae non-O1.

Indian J Med Res, 1995 Jan, 101, 10 - 2
Monoclonal antibodies against Ogawa specific & Ogawa-Inaba common antigenic determinants of Vibrio cholerae O1 & their diagnostic utility; Ramamurthy T et al.; Monoclonal antibodies to Ogawa-Inaba common antigenic determinant and Ogawa specific antigenic determinant of V . cholerae belonging to the serogroup O1 were generated from BALB/c mice immunized with V . cholerae O1 Eltor Ogawa strain . Reactivity and specificities of the monoclonal antibodies were examined by slide agglutination method . The monoclonal antibodies agglutinated all the V . cholerae O1 strains tested but did not agglutinate with any of the other currently recognized 140 serogroups of V . cholerae non-O1 as well as with a variety of other enteric pathogens . Diagnostic utility of the MAbs produced in this study as compared to polyclonal O1 and monospecific antisera showed that the MAbs were as good as the latter for serological diagnosis of the two serotypes of V . cholerae O1.

Mutat Res, 1995 Jan, 336(1), 79 - 89
Evidence for a weak adaptive response to alkylation damage in Vibrio cholerae; Bhasin N et al.; Wild-type Vibrio cholerae cells, when adapted by a stepwise treatment with sub-lethal concentrations of N-methyl-N'-nitro-N-nitrosoguanidine (MNNG), acquired resistance to killing and mutagenesis by subsequent challenges with higher concentrations of MNNG . This was also seen in the rec isogenic strain indicating that the observed phenomenon was not due to the induction of SOS functions . Further, the adapted cells of both the wild-type and rec strains could reactivate lethally alkylated phages with equal efficiency . Increased resistance of adapted cells correlated with the induction of a 17-kDa DNA methyltransferase, capable of repairing O6-methylguanine lesions in DNA . This induced methyltransferase was found to be antigenically unrelated to the Escherichia coli methyltransferase (Ada protein) as determined by Western blotting with polyclonal antiserum raised against the E . coli protein . Even though no counterpart of the constitutively expressed methyltransferase (Ogt) of E . coli could be detected in V . cholerae, several lines of evidence pointed towards the presence of an E . coli alk A-like gene in the organism.

Infect Immun, 1995 Jan, 63(1), 317 - 23
The capsule and O antigen in Vibrio cholerae O139 Bengal are associated with a genetic region not present in Vibrio cholerae O1; Comstock LE et al.; Vibrio cholerae O139 Bengal, although closely related to V . cholerae O1 El Tor, produces a polysaccharide capsule and has a distinct O antigen . We have identified a chromosomal region of at least 11 kb, as defined by three TnphoA mutations, that is required for the expression of both polysaccharides . Electron microscopy and sodium dodecyl sulfate-polyacrylamide gel electrophoresis show that these TnphoA mutants have lost the abilities both to express capsule and to produce lipopolysaccharide beyond the core oligosaccharide . Reactivity with O139 typing serum and resistance to serum are also lost in the mutants . DNA probes for this region do not hybridize with O1 V . cholerae but do react with other vibrios, implying that the region was recently acquired.

J Infect Dis, 1995 Jan, 171(1), 122 - 7
Molecular subtyping of toxigenic Vibrio cholerae O139 causing epidemic cholera in India and Bangladesh, 1992-1993; Popovic T et al.; Since October 1992, > 150,000 cases of cholera have been reported from India and Bangladesh; the great majority of Vibrio cholerae isolates belong to the newly established serogroup O139 . To better understand the interaction of genetic and epidemiologic factors responsible for their sudden appearance and rapid spread, representative toxigenic V . cholerae O139 isolates were molecularly characterized and compared with a set of toxigenic V . cholerae O1 and non-O1/non-O139 strains . DNA sequences of the cholera toxin B subunit gene and multilocus enzyme electrophoresis markers of V . cholerae O139 strains were identical to those of V . cholerae O1 isolates of the seventh pandemic . Two distinct ribotypes and four pulsed-field gel electrophoretic patterns were observed for O139 strains . V . cholerae O139 strains were very similar to V . cholerae O1 strains of the seventh pandemic but clearly different from the toxigenic V . cholerae strains of serogroups other than O1 and O139.

Gac Med Mex, 1995 Jan-Feb, 131(1), 28 - 35; discussion 36-7
{Mexican phenotype and genotype Vibrio cholerae 01}; Giono S et al.; This paper presents the phenotypical and genotypical characterization of 26922 Vibrio cholerae 01 strains isolated in Mexico from 1991 to 1993 . All strains isolated were El Tor biovar . Strains were sensitive to antibiotics excluding furazolidone, streptomycin and sulfisoxasole to which we found resistance in 97% and we are using this characteristic as epidemiological markers . We detected a marked change in frequency of Inaba serotype from 1991, when it was dominant, with 99.5%, until 1992 when Ogawa serotype turned to be dominant with 95% of isolates . All Vibrio cholerae 01 strains, except one Ogawa strain, were to igenic, and V . choleraeno 01 were not toxigenic by ELISA, PCR and cell culture tests . Dominant ribotype was 5, but we found some strains with 6a pattern and two with ribotype 12 . We are searching for ribotype 2 among hemolytic strains in order to learn if there is any relation to Gulf Coast strains prevalent in the USA, but until now we have not found any V . cholerae ribotype 2 in our isolates . Even if rapid tests are recommended for immediate diagnosis of cholera, it is necessary to continue bacterial isolation in order to have strains for phenotyping and genotyping studies that may support epidemiological analysis.

Przegl Epidemiol, 1995, 49(3), 237 - 43
{Vibrio cholera (Vibrio cholerae non-01) isolated in Poland from the Bug river}; Stypulkowska-Misiurewicz H et al.; Non-01 cholera vibrios for the first time has been isolated from freshwater in Poland . In October 1994 during bacteriological examination of Bug water by a two-step enrichment method in alkaline peptone water at pH 8.6 and TCBS . Repeated examination in November, December and January in the same and other locations among the polish-ukrainian borderline revealed persistence of V . cholerae in the river; 22 strains were isolated . All 22 strains were identical to V . cholerae E1 Tor 01 Ogawa reference strain except to antigens 0 . The strains isolated from water were no agglutinable by standard 01 serum neither by 0139 serum . Determination of their antigens 0 is under investigations . They might be pathogenic for people as it could be judged by their high resistance to bile, proteolytic and haemolytic activity . No cholera-like cases were notified on the polish side of the river.

Proc Natl Acad Sci U S A, 1994 Dec 20, 91(26), 12619 - 23
Synergistic binding of the Vibrio fischeri LuxR transcriptional activator domain and RNA polymerase to the lux promoter region; Stevens AM et al.; LuxR, the Vibrio fischeri luminescence gene (lux) activator, is the best-studied member of a family of bacterial transcription factors required for cell density-dependent expression of specific genes involved in associations with eukaryotic hosts . Neither LuxR nor any other LuxR homolog has been shown to bind DNA directly . We have purified the LuxR C-terminal transcriptional activator domain from extracts of recombinant Escherichia coli in which this polypeptide was expressed . The purified polypeptide by itself binds to lux regulatory DNA upstream of the lux box, a 20-bp palindrome that is required for LuxR activity in vivo, but it does not bind to the lux box . However, the LuxR C-terminal domain together with RNA polymerase protects a region including the lux box and the lux operon promoter from DNase I cleavage . There is very little protection of the lux operon promoter region from DNase I digestion in the presence of RNA polymerase alone . Apparently, there is a synergistic binding of the LuxR C-terminal domain and RNA polymerase to the promoter region . The upstream binding region for the purified polypeptide encompasses a binding site for cAMP receptor protein (CRP) . Under some conditions, CRP binding can block the binding of the LuxR C-terminal domain to the upstream binding region, and it can also block the synergistic binding of the LuxR C-terminal domain and RNA polymerase to the lux box and luminescence gene promoter region . This description of DNA binding by the LuxR C-terminal domain should lead to an understanding of the molecular interactions of the LuxR family of transcriptional activators with regulatory DNA.

FEBS Lett, 1994 Dec 19, 356(2-3), 333 - 8
Cloning and sequencing of four structural genes for the Na(+)-translocating NADH-ubiquinone oxidoreductase of Vibrio alginolyticus; Beattie P et al.; Oligonucleotide probes based on the N-terminal amino acid sequences of the NqrA and NqrC subunits were used to clone genes for the Na(+)-dependent NADH-ubiquinone oxidoreductase complex from Vibrio alginolyticus . Four consecutive ORFs were identified encoding subunit proteins of 48.6, 46.8, 27.7 and 22.6 kDa, respectively (NqrA-D) . A further ORF, showing 71% homology to the BolA protein of Escherichia coli, was located upstream . From sequence comparisons, we conclude that the Na(+)-dependent NADH-ubiquinone oxidoreductase complex of V . alginolyticus is clearly distinct from the corresponding H(+)-dependent enzymes of both prokaryotes and eukaryotes.

FEBS Lett, 1994 Dec 19, 356(2-3), 330 - 2
Cloning of the Na(+)-translocating NADH-quinone reductase gene from the marine bacterium Vibrio alginolyticus and the expression of the beta-subunit in Escherichia coli; Hayashi M et al.; The Na(+)-translocating NADH-quinone reductase purified from the marine bacterium Vibrio alginolyticus is composed of three subunits, alpha, beta and gamma . From the N-terminal amino acid sequences of each subunit and its polypeptide fragment obtained by partial digestion with V8 protease, oligonucleotides corresponding to forward and reverse primers for each gene (NQR A, B and C) encoding the alpha, beta and gamma subunit, respectively, were synthesized . Using these primers, a part of each gene was amplified from the chromosomal DNA of V . alginolyticus by a PCR method, and the PCR products were used for the cloning of the NQR gene in lambda phage . Among the subclones selected by probe C, the expression of the beta-subunit as a gene product was detected in Escherichia coli membranes by activity staining and Western blotting.

Commun Dis Rep CDR Rev, 1994 Dec 9, 4(13), R157 - 64
Cholera: current epidemiology; Crowcroft NS; Cholera remains an important cause of morbidity and mortality worldwide . Its epidemiology has changed in the 1990s, with the spread of the seventh pandemic to the western hemisphere and the emergence of a new serogroup, Vibrio cholerae O139 . The spread of cholera may be rapid and unpredictable because of aeroplane travel, international shipping, and the migration of people due to war or political unrest . Increasing amounts of largely untreated faeces from growing human populations favour cholera's survival . Most of the world has inadequate sanitation, and future prospects are undermined by the impact of international debt on ailing economies . Furthermore, because cholera is difficult to eradicate from water it is likely to remain a serious threat to public health for some time . Progress is being made in the development of oral vaccines against V . cholerae O1 and serogroup O139.

Gene, 1994 Dec 2, 150(1), 17 - 25
Genetic characterization of mannose-sensitive hemagglutinin (MSHA)-negative mutants of Vibrio cholerae derived by Tn5 mutagenesis; Hase CC et al.; El Tor biotype Vibrio cholerae strains express a cell-associated mannose-sensitive hemagglutinin (MSHA) which is a putative attachment factor . Several MSHA-negative mutants from V . cholerae strain JBK70 were previously generated by Tn5 mutagenesis {Finn et al., Infect . Immun . 55 (1987) 942-946} . The chromosomal DNA regions containing the Tn5 insertions were isolated from eight strains for further analysis . Nucleotide sequencing of the insertional junctions and corresponding clones containing the intact chromosomal region from the parental strain revealed the presence of several contiguous ORFs . Only two ORFs of this region had received insertions, and these showed remarkable homology to genes involved in the general secretory pathway found in several Gram- bacterial species . Proteins corresponding to the observed ORFs were visualized with the T7 promoter/RNA polymerase expression system . Marker exchange mutagenesis was used to insert kanamycin-resistance cassettes and TnphoA insertions into different locations of this region in the chromosome of wild-type V . cholerae strains . The phenotypes of these mutants showed that this DNA region is involved in MSHA production, but is not required for general extracellular protein secretion.

Gene, 1994 Dec 2, 150(1), 169 - 74
Expression of luxCD-E in Anabaena sp . can replace the use of exogenous aldehyde for in vivo localization of transcription by luxAB; Fernandez-Pinas F et al.; The genes luxCDABE from four luminescent bacteria suffice for light production in Escherichia coli {Meighen, Microbiol . Rev . 55 (1991) 123-142} . We have inserted these gene clusters between inverted polylinkers, and placed the resulting cassettes as reporters within derivatives of transposon Tn5 . Anabaena sp . strain PCC 7120 was mutagenized with these transposons . The luminescence of all but the most highly self-luminescent resulting derivatives of Anabaena sp . was strongly dependent on exogenously added aldehyde . Thus, luminescence based on luxCDABE is multiplicatively limited by production of luciferase and aldehyde . No toxicity was observed over protracted periods of luminescence . By deletion, new cassettes were derived in which only the aldehyde biosynthetic genes, luxCD-E, remained intact . Transcription was localized at the single-cell level in strains of cyanobacteria bearing constitutively expressed Xenorhabdus luminescens luxCD-E on a plasmid and relatively weakly expressed, developmentally regulated luxAB from Vibrio spp . in the chromosome . The developmentally critical gene, hetR, was thereby shown to remain active in mature heterocysts.

J Bacteriol, 1994 Dec, 176(24), 7558 - 65
Multiple N-acyl-L-homoserine lactone autoinducers of luminescence in the marine symbiotic bacterium Vibrio fischeri; Kuo A et al.; In Vibrio fischeri, the synthesis of N-3-oxohexanoyl-L-homoserine lactone, the autoinducer for population density-responsive induction of the luminescence operon (the lux operon, luxICDABEG), is dependent on the autoinducer synthase gene luxI . Gene replacement mutants of V . fischeri defective in luxI, which had been expected to produce no autoinducer, nonetheless exhibited lux operon transcriptional activation . Mutants released into the medium a compound that, like N-3-oxohexanoyl-L-homoserine lactone, activated expression of the lux system in a dose-dependent manner and was both extractable with ethyl acetate and labile to base . The luxI-independent compound, also like N-3-oxohexanoyl-L-homoserine lactone, was produced by V . fischeri cells in a regulated, population density-responsive manner and required the transcriptional activator LuxR for activity in the lux system . The luxI-independent compound was identified as N-octanoyl-L-homoserine lactone by coelution with the synthetic compound in reversed-phase high-pressure liquid chromatography, by derivatization treatment with 2,4-dinitrophenylhydrazine, by mass spectrometry, and by nuclear magnetic resonance spectroscopy . A locus, ain, necessary and sufficient for Escherichia coli to synthesize N-octanoyl-L-homoserine lactone was cloned from the V . fischeri genome and found to be distinct from luxI by restriction mapping and Southern hybridization . N-Octanoyl-L-homoserine lactone and ain constitute a second, novel autoinduction system for population density-responsive signalling and regulation of lux gene expression, and possibly other genes, in V . fischeri . A third V . fischeri autoinducer, N-hexanoyl-L-homoserine lactone, dependent on luxI for its synthesis, was also identified . The presence of multiple chemically and genetically distinct but cross-acting autoinduction systems in V . fischeri indicates unexpected complexity for autoinduction as a regulatory mechanism in this bacterium.

J Infect Dis, 1994 Dec, 170(6), 1518 - 23
Development of a live, oral, attenuated vaccine against El Tor cholera; Taylor DN et al.; Vibrio cholerae El Tor strains from Peru, Bangladesh, and Bahrain were attenuated by deletion of a genetic element that encodes virulence factors and RS1 . The B subunit of ctx (ctxB) was reintroduced into the recA gene of the deletion mutants, rendering them unable to recombine with exogenous genetic elements and generating Peru-3, Bang-3, and Bah-3 . Fifteen volunteers received one dose of various vaccine strains at 4 x 10(6) to 1 x 10(8) cfu . All strains colonized the gut . A > or = 4-fold rise in vibriocidal titer was observed in 14 volunteers, with titers of > or = 1600 in 13 . Peru-3 was the least reactogenic, but 2 of 6 volunteers had loose stools . Peru-14, a filamentous motility-deficient mutant of Peru-3, was well tolerated and colonized 18 of 21 volunteers at doses of 2 x 10(6) to 1 x 10(9) cfu . Also, when 8 Peru-3 or Peru-5 vaccinees, 5 Peru-14 vaccinees, and 8 controls were challenged with 2 x 10(6) cfu V . cholerae El Tor Inaba (N16961), 11 vaccinees were protected compared with no controls . Peru-14 shows promise as a safe, effective, single-dose oral vaccine against El Tor cholera.

Nature, 1994 Dec 1, 372(6505), 475 - 8
Protein disaggregation mediated by heat-shock protein Hsp104; Parsell DA et al.; The heat-inducible members of the Hsp100 (or Clp) family of proteins share a common function in helping organisms to survive extreme stress, but the basic mechanism through which these proteins function is not understood . Hsp104 protects cells against a variety of stresses, under many physiological conditions, and its function has been evolutionarily conserved, at least from Saccharomyces cerevisiae to Arabidopsis thaliana . Homology with the Escherichia coli ClpA protein suggests that Hsp104 may provide stress tolerance by helping to rid the cell of heat-denatured proteins through proteolysis . But genetic analysis indicates that Hsp104 may function like Hsp70 as a molecular chaperone . Here we investigate the role of Hsp104 in vivo using a temperature-sensitive Vibrio harveyi luciferase-fusion protein as a test substrate . We find that Hsp104 does not protect luciferase from thermal denaturation, nor does it promote proteolysis of luciferase . Rather, Hsp104 functions in a manner not previously described for other heat-shock proteins: it mediates the resolubilization of heat-inactivated luciferase from insoluble aggregates.

J Bacteriol, 1994 Dec, 176(23), 7378 - 82
Characterization of a glucose transport system in Vibrio parahaemolyticus; Sarker RI et al.; Cells of a glucose-PTS (phosphoenolpyruvate:carbohydrate phosphotransferase system)-negative mutant of Vibrio parahaemolyticus transport D-glucose in the presence of Na+ . Maximum stimulation of D-glucose transport was observed at 40 mM NaCl, and Na+ could be replaced partially with Li+ . Addition of D-glucose to the cell suspension under anaerobic conditions elicited Na+ uptake . Thus, we conclude that glucose is transported by a Na+/glucose symport mechanism . Calculated Vmax and Km values for the Na(+)-dependent D-glucose transport were 15 nmol/min/mg of protein and 0.57 mM, respectively, when NaCl was added at 40 mM . Na+ lowered the Km value without affecting the Vmax value . D-Glucose was the best substrate for this transport system, followed by galactose, alpha-D-fucose, and methyl-alpha-glucoside, judging from the inhibition pattern of the glucose transport . D-Glucose itself partly repressed the transport system when cells were grown in its presence.

Infect Immun, 1994 Dec, 62(12), 5624 - 31
Heat shock response and heat shock protein antigens of Vibrio cholerae; Sahu GK et al.; Sixteen heat shock proteins (Hsps) have been identified in the hypertoxinogenic strain 569B of Vibrio cholerae which are synthesized in response to small and large elevations of temperature . The induction of the Hsps is necessary for the cells to survive the deleterious effects of heat . There is no difference in the pattern of induction of the Hsps in V . cholerae strains varying in levels of toxinogenicity . One of the major low-molecular-mass Hsps, a 16-kDa protein, is preferentially degraded following shift down of temperature . This protein is induced at a much lower level at high temperatures in cells maintained in the laboratory for a prolonged period . The only Hsp located in the outer membrane of V . cholerae cells is a 23-kDa protein . Western immunoblot analysis with human immune sera collected from convalescent cholera patients revealed that this protein is markedly immunogenic . The human immune serum also reacted with the 69- and 16-kDa major Hsps and the 88-, 66-, and 46-kDa Hsps but not with the 61-kDa major Hsp identified as the groEL gene product . All major Hsps reacted with rabbit anti-V . cholerae sera . Ethanol stress leads to the induction of four of the major Hsps and three additional proteins.

J Clin Microbiol, 1994 Dec, 32(12), 3091 - 2
Case of Aeromonas veronii (DNA group 10) bacteremia; Abbott SL et al.; We describe the first case report of bacteremia due to Aeromonas veronii biotype veronii . The infection occurred in a 77-year-old man suffering from multiple underlying conditions which included cancer of the sigmoid colon . Because of the unusual biochemical phenotype of this group (ornithine decarboxylase positive), it was originally identified as Vibrio cholerae.

J Clin Microbiol, 1994 Dec, 32(12), 2975 - 9
Molecular epidemiologic analysis of Vibrio cholerae O1 isolates by pulsed-field gel electrophoresis; Mahalingam S et al.; Isolates of Vibrio cholerae O1 El Tor from two well-defined cholera outbreaks in Malaysia were analyzed by using pulsed-field gel electrophoresis (PFGE) . Isolates from sporadic cases occurring during the same time period were also studied . Digestion of chromosomal DNA from these isolates of V . cholerae O1 with restriction endonucleases NotI (5'-GCGGCCGC-3') and SfiI (5'-GGCCNNNN-3'), followed by PFGE, produced restriction endonuclease analysis (REA) patterns consisting of 13 to 24 bands (ranging in size from 46 to 398 kbp) . Analysis of the REA patterns generated by PFGE after digestion with NotI and SfiI suggested the clonal nature and close genetic identity of the isolates obtained during each of the two outbreaks (Dice coefficient, 0.93 to 1.0) . Although they had very similar REA patterns, the two outbreak clones were not identical . Isolates of V . cholerae O1 from sporadic cases, on the other hand, appeared to be much more heterogeneous (five different REA patterns detected in the five isolates tested; Dice coefficient, 0.31 to 0.81) than those obtained during the two outbreaks . We conclude that PFGE of V . cholerae O1 chromosomal DNA digested with infrequently cutting restriction endonucleases is a useful method for molecular typing of V . cholerae isolates for epidemiological purposes.

Microbiology, 1994 Dec, 140 ( Pt 12), 3399 - 406
Cloning and expression in Escherichia coli of the nahA gene from Porphyromonas gingivalis indicates that beta-N-acetylhexosaminidase is an outer-membrane-associated lipoprotein; Lovatt A et al.; Porphyromonas gingivalis has been implicated in human periodontal diseases . It expresses a number of exoglycosidase enzymes capable of hydrolysing host proteoglycan residues . As a first stage to explore the role of these enzymes in periodontal tissue damage, the nahA gene of P . gingivalis W83, which encodes beta-N-acetylhexosaminidase (beta-Nahase), was cloned . The gene was expressed poorly in Escherichia coli, but increased expression was achieved by cloning the nahA gene downstream of the tac promoter . Southern blot analysis revealed that nahA was present as a single copy, and it was found in all the other P . gingivalis strains tested . In contrast, sequences homologous to nahA were not detected in either P . endodontalis or P . asaccharolytica . The nahA gene was 2331 bp long and encoded a beta-Nahase enzyme of 777 amino acids with a predicted molecular mass of 87 kDa . A characteristic signal peptide for an acylated lipoprotein was present at the amino-terminus, suggesting that the mature beta-Nahase is a lipoprotein . The predicted amino acid sequence of the P . gingivalis beta-Nahase shared homology with the catalytic domains of the human beta-Nahase enzyme and the chitinase of Vibrio harveyi, suggesting a common catalytic mechanism.

Indian J Biochem Biophys, 1994 Dec, 31(6), 441 - 8
Cholera vaccine: developmental strategies and problems; Bhadra RK et al.; Over a hundred years have elapsed since Vibrio cholerae, the etiological agent for the disease cholera, was discovered by Robert Koch . Ever since then serious efforts have been made to develop prophylactic measures to combat the disease without much success . Seven pandemics have so far been reported and cholera still remains a public health problem in developing countries . Several strategies have been adopted to develop vaccines against the disease and many of these vaccines have undergone field trials . During the last two decades, an enormous amount of information has accumulated regarding the organism V . cholerae, its virulence factors, including cholera toxin, and the molecular basis of its pathogenicity . In recent years, with the advent of recombinant DNA technology and major breakthroughs in molecular biology and immunology, a new dimension has been given to the design of vaccine strains . The second generation live oral vaccines will perhaps soon replace the long-used first generation parenterally administered killed whole cell vaccines which offered protection for not more than three months . All the recombinant vaccines tested so far produced adverse reactions in volunteers, although they provided varying degrees of protection upto about one year of surveillance . Parallel to the trials of live oral vaccines, combination vaccines comprising killed whole cells and purified B subunit of cholera toxin was also tried . These vaccines had minimal side-effects but the efficacy was not upto expectations . From the failure of each vaccine strain, new information had emerged and improved strategies were adopted.(ABSTRACT TRUNCATED AT 250 WORDS)

Changgeng Yi Xue Za Zhi, 1994 Dec, 17(4), 339 - 46
Vibrio vulnificus infection--report of 8 cases and review of cases in Taiwan; Chang JJ et al.; Vibrio vulnificus infection, which is a rare and fatal disease, can be categorized clinically as either primary septicemia or wound infection . The clinical presentation of patients with primary septicemia can vary from fever alone to a more severe illness including high-grade bullous lesions, hypotension, and shock . Wound infection typically results from either injury to the skin in a marine environment or contact of a preexisting wound with sea water . We reported eight cases with Vibrio vulnificus infection in Chang gung Memorial Hospital and reviewed ten other cases previously reported with details in Taiwan . Fourteen patients presented with primary septicemia, and four with wound infection . Thirteen patients had alcoholism or chronic liver disease, two had peptic ulcer disease, one was steroids abuser, and one patient had thalassemia and chronic liver disease . Overall mortality was 55.6% (ten patients) . Patients with hypotension within 48 hours of admission had higher mortality than normotensive patients (77% vs . 0%, P = 0.007) . Patients with chronic liver disease or liver cirrhosis also had tendency to a higher mortality than not (64% vs . 25%, P = 0.274) . Chronic liver diseases and liver cirrhosis are common disease in Taiwan . They take a high risk for Vibrio vulnificus infection . Clinician should keep in mind of this potentially fatal infection in these patients reporting a history of recent raw oyster consumption and presented with sepsis and characterized skin lesions . Prompt empirical antibiotics treatment and aggressive surgical treatment may be lifesaving for this acute and fatal disease.

Indian J Med Res, 1994 Dec, 100, 262 - 5
Evaluation of potency of inactivated cholera vaccine by mouse protection assay & antibody induction method; Sood A et al.; Twenty one batches of whole cell inactivated cholera vaccine manufactured at Central Research Institute, Kasauli were evaluated for potency by mouse protection assay (MPA) and antibody induction method . In the antibody induction method the sera of immunized mice were screened for the presence of antibodies against Vibrio cholerae by microagglutination (MA) test and IgG ELISA . The number of organisms estimated by MPA were correlated with agglutinating and neutralizing antibodies against individual serotypes by MA and ELISA respectively . Correlation coefficient(r) of 0.692 and 0.815 were observed for the titres evaluated by MA and ELISA when compared with standard MPA method for the serotype Ogawa . Similarly r values of 0.925 and 0.849 were observed for titres evaluated by MA and ELISA when compared with standard MPA method for the serotype Inaba . Antibody induction method can be as an alternative method for determining the potency of inactivated cholera vaccine.

Singapore Med J, 1994 Dec, 35(6), 648 - 9
Non-O1 Vibrio cholerae septicaemia: a case report; Tan KK et al.; Non-O1 vibrio cholerae infections are associated with sporadic cases of gastroenteritis and extraintestinal infections . Septicaemia due to non-O1 vibrio cholerae is rare and are mainly reported in adults, particularly in immunocompromised patients . We report a case of non-O1 vibrio cholerae septicaemia and gastroenteritis in an 8-year-old child . The patient presented with bloody diarrhoea, fever and severe dehydration . Non-O1 vibrio cholerae were isolated from blood and stool cultures . The clinical course was uneventful after starting appropriate rehydration and supportive therapy.

Glycobiology, 1994 Dec, 4(6), 791 - 6
Characterization of ganglioside associated with the thyrotrophin receptor; Kielczynski W et al.; The receptor protein for thyrotrophin (thyroid-stimulating hormone; TSH) is associated with a glycosphingolipid moiety . The protein belongs to the family of receptors that couple to guanine nucleotide binding proteins; the glycosphingolipid contains sialic acid and belongs to the family of gangliosides . This report defines the structure of the receptor ganglioside in the Fisher rat thyroid cell line (FRTL-5) . Receptor protein was purified by TSH affinity chromatography from FRTL-5 cells, biosynthetically labelled with {3H}galactose and {3H}glucosamine, and resolved by SDS-PAGE . A single radiolabelled band of Mr approximately 80 kDa, corresponding to the predicted size of the cloned receptor, contained ganglioside . Gangliosides were extracted from unlabelled receptor protein after SDS-PAGE and probed on TLC plates with 125I-labelled Limax flavus agglutinin or the B subunit of cholera toxin, before and after digestion with Vibrio cholerae sialidase or beta-galactosidase . The TSH receptor (TSH-R) ganglioside belongs to the gangliotetraose family, having sialic acid attached to both galactose molecules . Its sialic acid is devoid of negative charge because of the formation of internal esterlactones . Its structure is lactonized N-acetylneuraminyl-(alpha 2-->3)galactosyl(beta 1-->3)-N-acetylgalactosaminyl(beta 1-->4)-{N-acetylneuraminyl(alpha 2-->3)}galactosyl(beta 1-->4)glucosyl(beta 1-->1)ceramide (GDla-lactone) . Ganglioside lactones have not been previously described as components of thyroid cells . They are highly rigid and are more likely than their parent structures to serve as molecular recognition sites and elicit immunoreactivity . Identification of this unique ganglioside intimately associated with the TSH receptor implies that it has an integral role in receptor structure and function.

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