Toxicol Lett, 1992 Dec, 64-65 Spec No, 687 - 93 Characterization of the effects of aluminum on luciferase biosensors for the detection of ecotoxicity; Guzzo J et al.; Luciferase-based biosensors are becoming increasingly used for environmental monitoring . A transcriptional fusion of the Vibrio harveyi luxAB genes (encoding bacterial luciferase) to the fliC gene of Escherichia coli was constructed and luminescence shown to be induced (in liquid media) in the presence of 1-10 micrograms/ml aluminum, but not copper, iron or nickel . Moreover, luminescence is markedly increased at pH 5.5, where aluminum is more soluble than at pH 7.0 . However, aluminum also stimulated luciferase activity when the luxAB genes were located in the xyl operon . This suggests that aluminum stimulates luciferase enzyme activity in vivo . These results are specific to E . coli, as no such aluminum stimulation was observed in the luminescent bacterium V . harveyi . These results have important implications in the generalized use of these clones for environmental monitoring, where aluminum can be present at elevated concentrations. Invest Ophthalmol Vis Sci, 1992 Dec, 33(13), 3592 - 600 Guanine nucleotide binding proteins in the dual regulation of lacrimal function; Cripps MM et al.; The purpose of this study was to identify and characterize functional G proteins that couple regulatory peptides with lacrimal secretory functions . Membranes were prepared from isolated rat exorbital lacrimal gland acini, and guanosine 5'-triphosphate (GTP)-dependence of adenylyl cyclase activity, known to be coupled with regulation of secretion, was measured . The guanine nucleotide GTP produced a biphasic response in the activity of membrane-bound adenylyl cyclase during a 10 min incubation with a maximum stimulation at 10(-5) mol/l GTP . Significant inhibition occurred at a dose of 10(-3) mol/l GTP, with cyclic adenosine monophosphate (cAMP) production reduced to less than basal levels . The effect of ADP-ribosylation of membrane proteins by the toxins produced by Vibrio cholera or Bordetella pertussis on lacrimal adenylyl cyclase was assessed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, autoradiography, and laser densitometry . Cholera toxin treatment of membranes resulted in dose-(0.5-100 micrograms/ml) and time-dependent (0-45 min) adenosine diphosphate (ADP)-ribosylation of two membrane proteins with M(r) values of 42,000 and 45,000 . Pertussis toxin treatment resulted in the specific ADP-ribosylation of a single protein that migrates with an M(r) value of 41,000 . This also was dose (0.5-25 micrograms/ml) and time dependent (0-30 min) . Incorporation of 32P into the 45,000 M(r) and 42,000 M(r) proteins in the presence of 50 micrograms/ml cholera toxin was guanine nucleotide dependent, with a two- to threefold increase in labeling when the membranes were incubated with 1 or 0.5 mmol/l GTP . This effect was enhanced in the presence of the nonhydrolyzable GTP analog GTP gamma S.(ABSTRACT TRUNCATED AT 250 WORDS) J Trop Med Hyg, 1992 Dec, 95(6), 365 - 72 Pathogenesis and ecology: the case of cholera; Drasar BS; The way in which the results of various studies of cholera have influenced the understanding of the disease are discussed . Attention is given to the outcomes of medical research, targeted on problems of cholera and biological research focused on understanding Vibrio cholerae . The contrast between these approaches has produced a new understanding of the ecological nature of disease . The importance of the environmental reservoir of V . cholerae as the engine for the generation of diversity is highlighted. Gene, 1992 Dec 1, 122(1), 175 - 80 Cloning and expression in Escherichia coli of the gene encoding an extracellular deoxyribonuclease (DNase) from Aeromonas hydrophila; Chang MC et al.; The gene encoding an extracellular DNase from Aeromonas hydrophila CHC-1 has been cloned and sequenced . Following expression of the dns in Escherichia coli, it was revealed that some of the cloned enzyme was present in the cell-free extracellular supernatant fluid, and there was no cell lysis and concurrent release of cytoplasmic or periplasmic proteins . Therefore, results suggest that E . coli cells were capable of secreting the DNase extracellularly, albeit very inefficiently . The dns is transcribed from its own promoter in E . coli, and expressed as a 25-kDa product, as determined by sodium dodecyl sulfate-polyacrylamide-gel electrophoresis of the culture supernatant preparations followed by a DNA-hydrolysis assay . Nucleotide sequence analysis predicted a single open reading frame of 690 bp encoding a 230-amino acid (aa) polypeptide, with a potential 20-aa signal peptide located at the N terminus of the predicted protein . The deduced aa sequence of the entire protein is highly homologous with that of the DNase of Vibrio cholerae. J Infect Dis, 1992 Dec, 166(6), 1433 - 5 Outbreak of cholera associated with crab brought from an area with epidemic disease; Finelli L et al.; From 31 March through 3 April 1991, 8 New Jersey residents developed severe, watery diarrhea after eating crabmeat brought back in the suitcase of a traveler to Ecuador . Stool cultures yielded toxigenic Vibrio cholerae O1, serotype Inaba, biotype El Tor from 4 persons, and vibriocidal antibody titers were > or = 1:640 in 7 persons, indicating recent infection with Vibrio cholerae O1 . Eating crab was statistically associated with illness (P = .006); however, no leftover crabmeat was available for testing . All 8 patients fully recovered and no cases of secondary transmission were reported . This was the first reported incident of cholera in the continental United States associated with food transported from an area with epidemic disease . Discouraging the transport of perishable souvenir seafood may prevent further outbreaks. J Gen Microbiol, 1992 Dec, 138 ( Pt 12), 2485 - 90 Virulence plasmid pJM1 prevents the conjugal entry of plasmid DNA into the marine fish pathogen Vibrio anguillarum 775; Singer JT et al.; Studies involving the introduction of cloned homologous genes into Vibrio anguillarum revealed that several plasmids could not be conjugally introduced into V . anguillarum 775(pJM1), but were transmissible to the pJM1-cured derivative H775-3 . Recombinant pBR322 plasmids containing V . anguillarum genomic DNA inserts were mobilized from Escherichia coli donors, using pRK2013, into V . anguillarum H775-3 recipients at frequencies of 10(-6) to 10(-5) per recipient . When identical matings were performed with V . anguillarum 775(pJM1) recipients, the infrequent exconjugants recovered carried the pBR322-based plasmid but had lost the large virulence plasmid pJM1 . Similar studies were carried out with plasmid RP4 and with recombinant derivatives of the closely related broad-host-range plasmid pRK290 . While RP4 was transmissible from E . coli to V . anguillarum H775-3 at frequencies of 6.7 x 10(-2) per recipient, transmission to V . anguillarum 775(pJM1) recipients occurred at frequencies of only 2.5 x 10(-7) . When pRK290 contained V . anguillarum DNA inserts, the only exconjugants recovered had lost pJM1, or contained pJM1 and a deletion derivative of the recombinant pRK290 plasmid where all of the DNA insert had been deleted . The use of Dam-, Dcm-, or EcoK- methylation-deficient E . coli donor strains failed to result in appreciable numbers of V . anguillarum 775(pJM1) exconjugants that contained the desired transferred plasmids . Following UV mutagenesis, a derivative of V . anguillarum 775(pJM1) was isolated that would accept conjugally transferred plasmid DNAs at frequencies similar to those observed when using V . anguillarum H775-3 recipients . These data suggest that virulence plasmid pJM1 mediates a restriction system that prevents conjugal transmission of plasmid DNA from E . coli donors into V . anguillarum 775(pJM1) . This putative restriction system appears not to be directed towards Dam-, Dcm-, or EcoK-methylated DNA, and appears not to involve a Type II restriction endonuclease. East Afr Med J, 1992 Dec, 69(12), 707 - 8 Effects of phosphate in the production of hemolysin in Vibrio parahaemolyticus; Kibue MA et al.; Vibrio parahaemolyticus is known to produce several types of hemolysin, the most document is the TDH and TRH . A new type of hemolysin whose expression is repressed by the presence of phosphate in the medium is reported. J Diarrhoeal Dis Res, 1992 Dec, 10(4), 227 - 30 Isolation of enterotoxigenic Vibrio cholerae non-01 from the Buriganga river and two ponds of Dhaka, Bangladesh; Rahim Z et al.; Vibrio cholerae 01 is usually considered the most toxigenic member of the Vibrionaceae and V . cholerae non-01 isolated from the environment is non-toxigenic . In our survey of the pollution of some aquatic environments in and around Dhaka, Bangladesh, we wanted to investigate the toxigenicity of V . cholerae non-01 isolated from water and sediment samples of the Buriganga river and two ponds in Dhaka, in the rabbit ileal loop (RIL) model . Fluid accumulation was induced by 18 of 28 live cultures and five of 18 cell-free culture filtrates in RIL . Seven of ten V . cholerae non-01 which failed to induce fluid in RIL were subjected to repeated passage in rabbit gut . Within two consecutive passages, all the strains could induce fluid in rabbit gut . Both toxigenic and non-toxigenic strains were uniformly sensitive to chloramphenicol and gentamicin but resistant to neomycin, novobiocin, polymyxin-B, streptomycin and vancomycin . Tetracycline sensitivity was found among eight of 17 toxigenic and six of 12 non-toxigenic strains . Sensitivity to trimethoprime-sulfa-methoxazole was noted among seven of 17 toxigenic and six of 12 non-toxigenic strains . Occurrence of enterotoxigenic and drug-resistant V . cholerae non-01 in the surface water is a public health hazard. J Biochem (Tokyo), 1992 Dec, 112(6), 849 - 55 Purification and characterization of monomeric isocitrate dehydrogenase with NADP(+)-specificity from Vibrio parahaemolyticus Y-4; Fukunaga N et al.; NADP(+)-dependent isocitrate dehydrogenase {IDH: EC 1.1.1.42} was purified to electrophoretic homogeneity from Vibrio parahaemolyticus Y-4, and shown to be a monomeric protein of molecular weight 80,000 with a pI of 5.0 . The amino acid composition and partial sequence at the N-terminus resembled those reported for other bacterial monomeric IDHs . Immunotitration with antisera to the monomeric and dimeric enzymes (antisera to IDH-II and -I of Vibrio ABE-1) showed an immunochemical distinction between the monomeric and dimeric IDHs, but there is similarity within the IDHs of each group . The circular dichroism spectra of the native and heat-denatured enzyme are also similar to those of monomeric IDH (IDH-II of Vibrio ABE-1) . These monomeric IDHs are proteins comprising 17-22% helix and 25-35% beta-pleated sheet in the native state. Immunol Cell Biol, 1992 Dec, 70 ( Pt 6), 391 - 5 Serological responses and immunity produced in salmonids by vaccination with Australian strains of Vibrio anguillarum; Munday BL et al.; The Australian populations of rainbow trout (Onchorhynchus mykiss) and Atlantic salmon (Salmo salar) were found to have similar immunological responses to local strains of Vibrio anguillarum as those reported for the more genetically diverse populations of these fish and strains of V . anguillarum found in the Northern Hemisphere . In addition, our studies more precisely defined the respective responses of rainbow trout and Atlantic salmon to immersion vaccination by the bath and dip methods. Eur J Biochem, 1992 Dec 1, 210(2), 491 - 8 The structure of the O-antigenic polysaccharide from lipopolysaccharide of Vibrio cholerae strain H11 (non-O1); Vinogradov EV et al.; After acid degradation of the lipopolysaccharide (LPS) of Vibrio cholerae strain H11 (non-O1), a tetrasaccharide was obtained, the structure of which was determined by quantitative and methylation analyses, periodate oxidation, one- and two-dimensional NMR spectroscopy, and fast-atom-bombardment and four-sector tandem mass spectrometry as beta-D-GalANGro-(1-3)-beta-D-QuiNAc-(1-4)-alpha-D-GalANGr o-(1-4)-NeuAc, in which GalANGro is N-galacturonoyl-2-aminoglycerol and QuiN 2-amino-2,6-dideoxy-glucopyranose . In addition, the trisaccharide beta-D-GalANGro-(1-3)-beta-D-QuiNAc-(1-4)-D-altro-hept ulose and the disaccharide alpha-D-GalANGro-(1-4)-NeuAc were isolated from acid-degraded lipopolysaccharide; the occurrence of sedoheptulose in lipopolysaccharide has not been described before . Based on the result of methylation analysis showing that galacturonic acid was the terminal sugar of the polysaccharide chain, and on the assumption that the tri- and the disaccharide represented the reducing and the non-reducing ends of the polysaccharide, respectively, the chemical structure of the O-specific chain of V . cholerae H11 is proposed as alpha-D-GalANGro-(1-4)-alpha-NeuAc-(2-3)-beta-D-GalANGro-(1- 3)-beta-D-QuiNAc- (1-{4)-alpha-D-GalANGro-(1-4)-alpha-NeuAc-(2-3)-beta-D-GalANGro -(1-3)-beta-D- QuiNAc-(1-}n-(1-4)-D-altro-heptulose . However, other possible structures can not be ruled out since the tri- and the disaccharide could be localised at different positions. Enferm Infecc Microbiol Clin, 1992 Nov, 10(9), 525 - 30 {Features of Vibrio cholerae 0:1 El Tor, Inaba serotype, isolated during the cholera epidemic in Cartagena (Colombia)}; Salim Mattar V; Cholera is an infectious transmissible disease characterized by the development of secretory diarrhea and that presents in epidemic and endemic forms . In Latin-America we are currently seen what could be the eight pandemic in cholera history, due to Vibrio cholerae 0:1 El Tor, Inaba serotype, infection . In the present preliminary study we analyze 300 patients with a clinical picture suggesting cholerae . In 250 cases, Vibrio cholerae 0:1 strains were isolated . The identification was made using mainly the following test and procedures: 1) growth in TCBS (saccharose fermentation), 2) Reactivity with sodium desoxycholate, 3) Oxidase-cytochrome test and 4) agglutination with polyvalent antisera . All isolated Vibrio cholerae strains 0:1 were sensitive to tetracycline, trimethoprim-sulfamethoxazole, norfloxacin, ciprofloxacin and gentamicin, but were resistant to ampicillin . This report summarizes the main features of Vibrio cholerae 0:1 El Tor, Inaba serotype, isolated in Colombian cholera epidemic. Can J Microbiol, 1992 Nov, 38(11), 1175 - 80 The thermostable direct hemolysin of Vibrio parahaemolyticus is a pore-forming toxin; Honda T et al.; The hemolytic mechanism of thermostable direct hemolysin (TDH), a possible virulence factor of Vibrio parahaemolyticus, was studied . We demonstrated that TDH acts as a "pore-forming toxin" in temperature-dependent and -independent steps . The first temperature-dependent step requires only about 1-2 min incubation at 37 degrees C and makes a "pore" with a functional diameter of approximately 2 nm . The pore size was deduced from the molecular diameter of the colloidal inhibitory polysaccharides . The formation of the pores on TDH-treated erythrocyte membranes was also demonstrated by electron microscopic examination . The second step, which is a temperature-independent lytic step, causes the erythrocytes to swell owing to a colloidal osmotic influx of water via the "pores" into cells, resulting in erythrocyte lysis (or rupture) owing to increased intracellular pressure. Clin Infect Dis, 1992 Nov, 15 Suppl 1, S241 - 3 Evaluation of new anti-infective drugs for the treatment of cholera . Infectious Diseases Society of America and the Food and Drug Administration; Corrado ML et al.; Cholera is an acute gastrointestinal infection caused by Vibrio cholerae . It is characterized by watery diarrhea that may lead to massive fluid loss, which in turn may result in hypotension, shock, and death within hours . Key to the treatment of cholera is fluid replacement . Anti-infective therapy decreases the severity and duration of diarrhea and the duration of shedding of V . cholerae . Enrolled patients should have diarrhea that is moderate to severe and a culture that ultimately yields V . cholerae . A prospective, randomized, active-controlled clinical trial is preferred . Studies should be double-blinded or evaluator-blinded . The rapidity with which the organism is eliminated from stool may be assessed . Both clinical and microbiological outcome should be determined . Assessment of microbiological eradication is paramount, since fluid replacement may suffice for treatment of signs and symptoms. FEMS Microbiol Lett, 1992 Nov 1, 77(1-3), 217 - 24 N-acetyl-D-glucosamine-specific lectin purified from Vibrio cholerae 01; Sasmal D et al.; An N-acetyl-D-glucosamine-specific cell associated hemagglutinin (HA) was isolated and purified from a strain of Vibrio cholerae 01 by chitin affinity chromatography followed by separation on Bio Gel P-150 . A single stained protein band of 47 kDa in sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) was observed with the purified HA . HA-antisera produced a single precipitin band against the purified HA in an immunodiffusion test without exhibiting any reactivity towards purified lipopolysaccharide (LPS) . Purified HA, used as solid-phase antigen in an enzyme-linked immunosorbent assay (ELISA), reacted strongly with HA-antisera but cross-reacted negligibly with antisera raised against purified LPS . Hemagglutinating activity of the purified HA was highly sensitive to N-acetyl-D-glucosamine . The immunogold-labelling method using HA-antisera confirmed the location of the HA on the surface of the bacterial cells . The HA-antisera reacted with a protein component of the homologous outer membrane preparation . A significant inhibition was observed in the adhesive capability of the V . cholerae 01 strain to isolated rabbit intestinal epithelial cells (RIEC) in vitro when the later were pre-treated with the purified HA. FEMS Microbiol Lett, 1992 Nov 1, 77(1-3), 197 - 200 Characterization of two forms of hemagglutinin/protease produced by Vibrio cholerae non-O1; Naka A et al.; Two forms (34 kDa and 32 kDa) of hemagglutinin/protease produced by Vibrio cholerae non-O1 were characterized . The hemagglutinin/protease purified by immunoaffinity column chromatography using a monoclonal antibody was essentially a 34-kDa form . By incubation of the purified 34-kDa form at 37 degrees C, it was processed (autodigested) to the 32-kDa form . The N-terminal 20 amino acid sequences of both the 34- and 32-kDa forms were identical, suggesting that proteolytic processing at the C-terminal region of the 34-kDa hemagglutinin/protease resulted in the 32-kDa form . With this shift, protease activity increased, but hemagglutinating activity decreased, suggesting that the C-terminal region of the hemagglutinin/protease is related to hemagglutinating activity. Anal Biochem, 1992 Nov 1, 206(2), 419 - 29 The mapping by high-pH anion-exchange chromatography with pulsed amperometric detection and capillary electrophoresis of the carbohydrate moieties of human plasma alpha 1-acid glycoprotein; Hermentin P et al.; The reducing oligosaccharides released from alpha 1-acid glycoprotein (AGP) by conventional hydrazinolysis have been analyzed by two different mapping techniques, using high-pH anion-exchange chromatography with pulsed amperometric detection (HPAE-PAD) and capillary electrophoresis (CE) with uv detection at 190 nm . The CE measurements proved about 4000 times more sensitive than the measurements by HPAE-PAD . The N-glycan pool was fractionated by Mono Q anion-exchange chromatography, and individual fractions so obtained were desialylated using Vibrio cholerae neuraminidase . The resulting asialo-N-glycans were further analyzed by HPAE-PAD, revealing 2 major, 4 intermediate, and 4 small peaks and at least 3 spikes, which counted for at least 13 different asialo-N-glycans . The carbohydrate structures were tentatively assigned by comparison of the Mono Q-separated N-glycans with the known AGP carbohydrate structures and known structures contained in a mapping database that allows structural assignment of N-glycans by mere comparison of retention times . In addition to the hitherto known AGP carbohydrate structures, we have tentatively identified a number of sulfated N-glycans that are currently being analyzed in more detail . We have also compared the glycan pools recovered from AGP using hydrazinolysis and glycopeptidase F (PNGase F) . Approximately 40 distinct peaks could be detected in the hydrazinolysis-derived N-glycan pool by either technique (HPAE-PAD and CE), while about 30 distinct peaks were detected in the N-glycan pool derived by PNGase F digestion of the tryptic AGP digest of the same batch of AGP . These differences were attributed to an increased desialylation (approximately 3 mol%) during hydrazinolysis, based on the detection by HPAE-PAD and CE of free sialic acid and monosialylated oligosaccharides in the glycan pool derived by conventional hydrazinolysis . The integrity of the N-glycans' chitobiose core was examined by 500-MHz 1H NMR spectoscopy . The hydrazinolysis procedure could be optimized such that the hydrazinolysis-derived N-glycan pool was chromatographically essentially identical to the PNGase F-derived N-glycan pool . Hydrazinolysis proved best, with practically no loss of N-acetlylneuraminic acid and the closest resemblance to the PNGase F-derived N-glycan pool, using an automated apparatus . Notably, it was recognized that, in our hands, PNGase F digestion in the presence of sodium dodecyl sulfate resulted in partial desialylation of the liberated N-glycans. J Bacteriol, 1992 Nov, 174(22), 7235 - 44 Cloning of a metalloprotease gene involved in the virulence mechanism of Vibrio anguillarum; Milton DL et al.; Genetic evidence has previously suggested that a zinc metalloprotease is involved in the invasive mechanism of the fish pathogen Vibrio anguillarum NB10 . In this study, the metalloprotease gene was cloned and sequenced . The sequence encodes a polypeptide (611 amino acids) that contains a putative signal sequence followed by a large leader sequence and the mature protein (44.6 kDa) . Since the purified protein has a molecular mass of 36 kDa instead of the predicted 44.6 kDa, the mature protein is most likely processed a third time . Comparative analyses of the protein sequence showed high homologies to other bacterial metalloproteases within the zinc-binding and active-site regions . The Vibrio cholerae hemagglutinin/protease and the Pseudomonas aeruginosa elastase were exceptions in that the homology extended throughout the entire putative preproprotein . A chromosomal metalloprotease mutant was made via the integration of foreign DNA into the protease gene . This mutant did not secrete the metalloprotease, as determined by sodium dodecyl sulfate (SDS)-polyacrylamide protein analysis and by growth on gelatin agar . Transcomplementation of the chromosomal mutation revived the secretion of the metalloprotease and its activity on gelatin agar . Interestingly, when supernatant proteins were analyzed by gelatin-SDS-polyacrylamide electrophoresis, two different proteases (75 and 30 kDa) were detected in the mutant strain but not in the transcomplemented strain or the wild-type strain . Moreover, fish infection studies were done, and implications for the role of the metalloprotease in the virulence mechanism of V . anguillarum are discussed. J Bacteriol, 1992 Nov, 174(22), 7138 - 43 Formation of the LuxR protein in the Vibrio fischeri lux system is controlled by HtpR through the GroESL proteins; Adar YY et al.; The transcription of the luminescence (lux) system of Vibrio fischeri is regulated by the LuxR protein and an autoinducer . We previously showed that apart from these regulatory elements, the transcription of the lux system is negatively controlled by the LexA protein and positively controlled by the HtpR protein (sigma 32) . This study was conducted in order to elucidate the mode of action of the HtpR protein . Using luxR-lacZ fused genes, we showed that the HtpR protein is essential for the maximum expression of beta-galactosidase activity in Escherichia coli lac mutant cells . Using this construct, we also demonstrated that luxR is preferentially expressed toward the end of the logarithmic phase of growth . Starvation and addition of ethanol significantly advanced the appearance of beta-galactosidase activity in htpR+ cells . The luminescence system of E . coli htpR+ cells harboring the pChv1 plasmid with a deletion in the luxI gene is induced in the presence of low and constant concentrations (150 pg/ml) of the inducer only at a late stage of the logarithmic phase of growth . When the cellular LuxR content is reduced, following 23 generations of exponential growth in Luria broth, a mid-log-phase culture does not respond to the inducer (150 pg/ml) . On the basis of the above observations we suggest that the HtpR protein controls the formation of V . fischeri LuxR protein . Preliminary findings indicate that the HtpR protein acts through the chaperonins GroESL . E . coli htpR/pChv1 cells retained their full level of in vivo and in vitro luciferase activities in the presence of multiple copies of groESL genes . The possibility that GroESL proteins stabilize the native form of LuxR protein is discussed. J Infect Dis, 1992 Nov, 166(5), 1029 - 34 Evidence that inactivated oral cholera vaccines both prevent and mitigate Vibrio cholerae O1 infections in a cholera-endemic area; Clemens JD et al.; In a randomized, placebo-controlled field trial of B subunit-killed whole cell (BS-WC) and killed whole cell only (WC) inactivated oral cholera vaccines in rural Bangladesh, active surveillance of selected neighborhoods during the first year after vaccination identified 127 Vibrio cholerae O1 infections among 3285 three-dose recipients . For each vaccine, protective efficacy was greater against symptomatic (57%, P < .05 for BS-WC; 58%, P < .05 for WC) than against asymptomatic infections (46%, P < .05 for BS-WC; 32%, P = .09 for WC), and protection against each grade of infection was demonstrable for both the classical and El Tor biotypes . Although vaccine protection against symptomatic infections was evident in both young children and older persons, only persons vaccinated at age > 5 years were protected against asymptomatic infections . These results suggest that the inactivated oral vaccines acted both to protect against intestinal colonization by V . cholerae O1 and to interrupt the pathogenic sequence of established infections. J Bacteriol, 1992 Nov, 174(21), 6974 - 80 The virulence gene activator ToxT from Vibrio cholerae is a member of the AraC family of transcriptional activators; Higgins DE et al.; Virulence gene expression in Vibrio cholerae is postulated to involve ToxR-dependent activation of the toxT gene followed by ToxT activation of virulence genes, including several of those involved in biogenesis of the toxin-coregulated pilus . ToxR is a transmembrane, DNA-binding protein which is a member of the OmpR subclass of two-component activator systems in bacteria . Data presented in this report demonstrate that ToxT is similar to the AraC family of transcriptional activators identified in a variety of gram-negative bacteria . The toxT open reading frame begins approximately 200 nucleotides from the end of the tcpF gene, which is part of a cluster of genes responsible for production of the toxin-coregulated pilus . Accumulation of toxT specific mRNA is ToxR dependent and is modulated by environmental conditions that modulate expression of the regulon . Within the intergenic region between tcpF and toxT is a potential stem-loop structure of an unusual nature which may play a role in regulating expression of toxT mRNA . Experiments with tcpF and toxT cloned behind a strong, constitutive promoter suggest that the two genes can be cotranscribed, but Northern (RNA) blot analysis of V . cholerae suggests that if they are, steady-state levels of their messages may be controlled by a posttranscriptional mechanism . Possible mechanisms for ToxR-dependent expression of toxT are discussed. J Bacteriol, 1992 Nov, 174(21), 6807 - 14 ToxR proteins with substitutions in residues conserved with OmpR fail to activate transcription from the cholera toxin promoter; Ottemann KM et al.; The ToxR protein of Vibrio cholerae is an integral membrane protein that coordinately regulates the expression of virulence genes required for successful infection . ToxR has been shown to bind directly to and activate transcription of the cholera toxin (ctx) promoter . Within the amino-terminal cytoplasmic region of ToxR, several amino acids are strictly conserved among ToxR, OmpR, and the other members of a family of bacterial regulatory proteins . To better understand the function of this region, two approaches were taken: conserved residues were changed by site-directed mutagenesis, and random mutations that eliminated ToxR-mediated transcriptional activation were isolated . Several classes of mutations were identified: those that abolish promoter DNA binding and transcriptional activation (toxR R96K, toxR R68K, and toxR R68L), those that abolish transcriptional activation but retain the ability to bind promoter DNA (toxR R96L), and those that have an intermediate phenotype (toxR R77L, toxR E51K, and toxR E51D) . The toxR E51K allele had reduced activity in both Escherichia coli and V . cholerae but also exerted a dominant-negative effect over wild-type ToxR when assayed in V . cholerae . This result provides additional evidence that ToxR acts as an oligomer in the transcriptional activation process . From this mutational analysis of conserved amino acid residues within the OmpR-homologous region of ToxR, we conclude that this region is essential for transcriptional activation at the level of DNA binding and other steps that lead to activation of the ctx promoter. Infect Immun, 1992 Nov, 60(11), 4915 - 24 Fusion proteins containing the A2 domain of cholera toxin assemble with B polypeptides of cholera toxin to form immunoreactive and functional holotoxin-like chimeras; Jobling MG et al.; Cholera enterotoxin (CT) is produced by Vibrio cholerae and excreted into the culture medium as an extracellular protein . CT consists of one A polypeptide and five B polypeptides associated by noncovalent bonds, and CT-B interacts with CT-A primarily via the A2 domain . Treatment of CT with trypsin cleaves CT-A into A1 and A2 fragments that are linked by a disulfide bond . CT-B binds to ganglioside GM1, which functions as the plasma membrane receptor for CT, and the enzymatic activity of A1 causes the toxic effects of CT on target cells . We constructed translational fusions that joined foreign proteins via their carboxyl termini to the A2 domain of CT-A, and we studied the interactions of the fusion proteins with CT-B . The A2 domain was necessary and sufficient to enable bacterial alkaline phosphatase (BAP), maltose-binding protein (MBP) or beta-lactamase (BLA) to associate with CT-B to form stable, immunoreactive, holotoxin-like chimeras . Each holotoxin-like chimera was able to bind to ganglioside GM1 . Holotoxin-like chimeras containing the BAP-A2 and BLA-A2 fusion proteins had BAP activity and BLA activity, respectively . We constructed BAP-A2 mutants with altered carboxyl-terminal sequences and tested their ability to assemble into holotoxin-like chimeras . Although the carboxyl-terminal QDEL sequence of the BAP-A2 fusion protein was not required for interaction with CT-B, most BAP-A2 mutants with altered carboxyl termini did not form holotoxin-like chimeras . When holotoxin-like chimeras containing BAP-A2, MBP-A2, or BLA-A2 were synthesized in V . cholerae, they were found predominantly in the periplasm . The toxin secretory apparatus of V . cholerae was not able, therefore, to translocate these holotoxin-like chimeras across the outer membrane. Infect Immun, 1992 Nov, 60(11), 4848 - 55 Major outer membrane proteins of Vibrio cholerae and their role in induction of protective immunity through inhibition of intestinal colonization; Sengupta DK et al.; Vibrio cholerae O1 organisms belonging to different biotypes and serotypes were shown to express major outer membrane proteins (MOMPs) with subunit molecular masses of 48 to 50, 40 to 43, 35 to 36, 27 to 28, and 20 kDa . Antisera raised against individual MOMPs of a V . cholerae O1 strain recognized MOMPs of corresponding molecular masses in other O1 and non-O1 strains . Serological data also suggested possible differences in the cell surface exposition of these MOMPs . However, no marked differences between V . cholerae cells grown in vitro and in vivo could be noted in respect to the expression or surface exposition of these MOMPs . Of five MOMPs studied in this work, 40- to 43- and 20-kDa cell surface proteins were shown to be of considerable importance, as antisera to these proteins induced significant protection against V . cholerae challenge in the suckling mouse model . Similar protection, although to a lesser extent, was demonstrable with the antiserum to the 27- to 28-kDa protein . These results were corroborated with the Fab (immunoglobulin G) {Fab(IgG)} fragments of the antisera, thereby suggesting that the observed protection induced by anti-MOMP antibodies did not arise as a result of bacterial clumping . Subsequent studies demonstrated that these antisera as well as their Fab (IgG) fragments induced significant inhibition of intestinal colonization of V . cholerae . The 40- to 43- and 27- to 28-kDa proteins appeared to be porinlike, while the 20-kDa protein was found to be antigenically related to TcpA (subunit A of toxin-coregulated pilus) . All these results demonstrate the involvement of more than one cell surface antigen of V . cholerae in the induction of protective immunity through inhibition of intestinal colonization of vibrios. J Bacteriol, 1992 Nov, 174(22), 7490 - 3 The luxR gene product of Vibrio harveyi is a transcriptional activator of the lux promoter; Swartzman E et al.; Expression of the lux operon from the marine bacterium Vibrio harveyi is dependent on cell density and requires an unlinked regulatory gene, luxR, and other cofactors for autoregulation . Escherichia coli transformed with the lux operon emits very low levels of light, and this deficiency can be partially alleviated by coexpression of luxR in trans . The V . harveyi lux promoter was analyzed in vivo by primer extension mapping to examine the function of luxR . RNA isolated from E . coli transformed with the Vibrio harveyi lux operon was shown to have a start site at 123 bp upstream of the first ATG codon of luxC . This is in sharp contrast to the start site found for lux RNA isolated from V . harveyi, at 26 bp upstream of the luxC initiation codon . However, when E . coli was cotransformed with both the lux operon and luxR, the start site of the lux mRNA shifted from -123 to -26 . Furthermore, expression of the luxR gene caused a 350-fold increase in lux mRNA levels . The results suggest that LuxR of V . harveyi is a transcriptional activator stimulating initiation at the -26 lux promoter. J Bacteriol, 1992 Nov, 174(21), 6780 - 8 Ribosomes exist in large excess over the apparent demand for protein synthesis during carbon starvation in marine Vibrio sp . strain CCUG 15956; Flardh K et al.; Carbon starvation induces the development of a starvation- and stress-resistant cell state in marine Vibrio sp . strain S14 (CCUG 15956) . The starved cells remain highly responsive to nutrients during prolonged starvation and exhibit instantaneous severalfold increases in the rates of protein synthesis and RNA synthesis when substrate is added . In order to elucidate the physiological basis for the survival of cells that are starved for a long time, as well as the capacity of these cells for rapid and efficient recovery, we analyzed the ribosome content of carbon-starved Vibrio sp . strain S14 cells . By using direct chemical measurements of the amounts of ribosomal particles in carbon-starved cultures, we demonstrated that ribosomes were lost relatively slowly (half life, 79 h) and that they existed in large excess over the apparent demand for protein synthesis . After 24 h of starvation the total rate of protein synthesis was 2.3% of the rate during growth, and after 3 days this rate was 0.7% of the rate during growth; the relative amounts of ribosomal particles at these times were 81 and 52%, respectively . The ribosome population consisted of 90% 70S monoribosomes, and no polyribosomes were detected in the starved cells . The 70S monoribosomes were responsible for the bulk of the protein synthesis during carbon starvation; some activity was also detected in the polyribosome size region on sucrose density gradients . We suggest that nongrowing carbon-starved Vibrio sp . strain S14 cells possess an excess protein synthesis capacity, which may be essential for their ability to immediately initiate an upshift program when substrate is added. APMIS, 1992 Nov, 100(11), 1027 - 32 Enzyme-linked immunosorbent assay for determination of antibodies to Vibrio cholerae toxin-coregulated pili; Osek J et al.; An ELISA for determination of antibodies to V . cholerae TCP was developed . Since purified TCP preparations contained detectable amounts of LPS (as shown by ELISA and immunoelectron microscopy with anti-LPS polyclonal serum), a capture ELISA was used . In this test the plate was coated with anti-TCP monoclonal antibody followed by incubation with TCP fimbriae . By this procedure no LPS bound to the solid phase as shown by the loss of reactivity with anti-LPS serum . The capture ELISA allowed sensitive and specific determination of TCP antibodies in sera of rabbits immunized with classical but not El Tor V . cholerae strains . There was good agreement between results in the TCP ELISA and reactivity with the TcpA band in immunoblot analyses when antisera raised against classical and El Tor vibrios were studied. Infect Immun, 1992 Nov, 60(11), 4961 - 4 Protection against Vibrio cholerae El Tor infection by specific antibodies against mannose-binding hemagglutinin pili; Osek J et al.; Both specific polyclonal antiserum and monoclonal antibodies against mannose-binding hemagglutinin fimbriae of Vibrio cholerae (mannose-sensitive hemagglutinin {MSHA}) were shown to protect against experimental cholera caused by vibrios of the El Tor biotype in the infant mouse and in the rabbit intestinal loop models . MSHA-specific Fab immunoglobulin fragments were also protective . No protective effect was observed against challenge with V . cholerae O1 of the classical biotype . These results suggest that MSHA pili play an important role in the pathogenesis of cholera caused by the El Tor biotype of V . cholerae and that induction of intestinal anti-MSHA immunity may be a worthwhile additional objective in the development of oral cholera vaccines. Plasmid, 1992 Nov, 28(3), 267 - 71 A stable Escherichia coli-Mycobacterium smegmatis plasmid shuttle vector containing the mycobacteriophage D29 origin; David M et al.; A plasmid shuttle vector for Escherichia coli and mycobacteria was constructed from an E . coli plasmid containing the ColE1 origin, a 2.6-kb PstI fragment from bacteriophage D29 that grows in numerous mycobacterial species, and the kanamycin resistance gene either of Tn903 or of Tn5 . The resultant plasmid is 7.63 kb and can be introduced via transformation into Mycobacterium smegmatis with high efficiency . In M . smegmatis the plasmid is stable and apparently present in multiple copies . Bioluminescence (luxA and luxB of Vibrio harveyi and fischeri) has been expressed in M . smegmatis from the aminoglycoside transferase promoter of Tn5 . The D29 fragment should carry an origin of replication and some associated genes that act on it since various mutations destroy the ability of this fragment to replicate in M . smegmatis . The fragment was localized on the D29 genome map. J Bacteriol, 1992 Nov, 174(21), 6743 - 51 Catabolite repression of the H(+)-translocating ATPase in Vibrio parahaemolyticus; Sakai-Tomita Y et al.; Cells of Vibrio parahaemolyticus grown in the presence of glucose showed reduced (by about 40%) oxidative phosphorylation . With this observation as a basis, we examined the effect of glucose on the level of H(+)-translocating ATPase . The addition of glucose to the growth medium reduced the specific activity and the amount of the H(+)-translocating ATPase in membrane vesicles of V . parahaemolyticus . These reductions were reversed by adding cyclic AMP (cAMP) to the growth medium . We cloned some parts of the unc genes encoding subunits of the H(+)-translocating ATPase of V . parahaemolyticus by means of the polymerase chain reaction . Using an amplified DNA fragment, we carried out Northern (RNA) blot analysis and found that glucose reduced the mRNA level of the H(+)-translocating ATPase gene by about 40% and that cAMP restored it . We determined the DNA sequence of the unc promoter region of V . parahaemolyticus and found a consensus sequence for the cAMP receptor protein-cAMP-binding site . Such a sequence was also found in the promoter region of the unc operon of Vibrio alginolyticus but not in its counterpart in Escherichia coli . We observed a similar reduction in the level of ATPase due to glucose in V . alginolyticus . In E . coli, however, reductions in the ATPase and the unc mRNA levels were not observed . Thus, the unc operon is controlled by cAMP-regulated catabolite repression in V . parahaemolyticus and V . alginolyticus but not in E . coli . Catabolite repression of the unc operon in V . parahaemolyticus is not severe. Antibiot Khimioter, 1992 Nov, 37(11), 17 - 21 {Dynamics of drug resistance of Vibrio cholerae isolated from surface water reservoirs in Siberia and the Soviet Far East in 1976-1990}; Ganin VS et al.; In the study on antibiotic resistance 1383 strains of El Tor Vibrio cholerae isolated from surface water reservoirs in 12 administrative territories of the Siberia and Far East within a period of 15 years were tested . The following antibiotics were used: ampicillin, streptomycin, monomycin, polymyxin, tetracycline, chloramphenicol, rifampicin and nalidixic acid . The resistance was unstable and its pattern was wave-like according to annual changes in the biological cycle . It was especially evident in regard to ampicillin, streptomycin, monomycin and polymyxin . The highest numbers of the strains were resistant to polymyxin, ampicillin and streptomycin (up to 100 per cent in some years) . The lowest numbers of the strains were resistant to chloramphenicol (0.4 per cent) and tetracycline (1.9 per cent) . No strains resistant to rifampicin and nalidixic acid were isolated . In some cases the antibiotic resistance level depended on the geographical zone where the strain was isolated . A direct quantitative dependence of the resistance level on the MIC was observed: the lower the MIC of the drug was, the lower the number of the strains resistant to it was . Within the 15-year period there was no general tendency to increase the resistance in V . cholerae to the antibiotics used. Eur J Epidemiol, 1992 Nov, 8(6), 861 - 4 A survey of urease-positive Vibrio parahaemolyticus strains isolated from traveller's diarrhea, sea water and imported frozen sea foods; Honda S et al.; The frequency of urease-positive Vibrio parahaemolyticus among isolates from patients, imported frozen sea foods and the environment (sea water) was studied . The highest isolation frequency of urease-positive V . parahaemolyticus was found in clinical isolates (11.2% out of 204 strains examined) . Urease-positive V . parahaemolyticus was found in 5.7% of 88 frozen sea food-isolates examined, but no strains isolated from sea water were urease-positive . The isolates were further examined for the production of thermostable direct hemolysin (Vp-TDH) and its related hemolysin (Vp-TRH) . Both are possible pathogenic toxins produced by mostly clinical isolates of V . parahaemolyticus . Urease-positive strains have a tendency to associate with clinical isolates producing both or neither Vp-TDH and Vp-TRH . Rabbit ligated ileal loops test was performed with several strains of urease-positive and -negative clinical isolates, and we found that some strains producing urease, even those which do not produce Vp-TDH or Vp-TRH, caused intestinal fluid accumulation. Mikrobiol Zh, 1992 Nov-Dec, 54(6), 45 - 9 {The lysogeny of Vibrio parahaemolyticus of serovar O4:K12}; Kudriakova TA et al.; Lysogeny has been first established in strains of parahemolytic vibrios of serovar O4:K12 . Moderate phages belonged to morphological group IV by home A . S . Tikhonenko's classification and were presented by one serological type . No correlation has been revealed between sensitivity to moderate phages of parahemolytic vibrios and specificity of "O"- or "K"-serotypes. Indian J Med Res, 1992 Nov, 95, 294 - 6 Role of iron in the virulence of Vibrio vulnificus isolated from Cuddalore coastal waters (India); Jayalakshmi S et al.; V . vulnificus strains isolated from different sources of Cuddalore coastal waters were tested for their virulence activity through their LD50 values in mice . As infections of V . vulnificus have been correlated with pre-existing liver disease and hemochromatosis, the role of iron on virulence was determined using . iron overloaded mice . The LD50 was in the range of 10(4)-10(7) cells in normal mice but 10(1)-10(2) cells in iron-injected mice, thus providing evidence that iron may play a major role in the pathogenesis of V . vulnificus. Indian J Med Res, 1992 Nov, 95, 288 - 93 Effect of Vibrio cholerae-L-asparaginase on surface structure of splenic lymphocytes; Dey SK et al.; The surface ultrastructure of splenic lymphocytes and rosetting properties of lymphocytes of Swiss mice were studied under scanning electron microscope following transplantation of Dalton's lymphoma and ascites fibrosarcoma tumour cells and administration of Vibrio cholerae-L-asparaginase . The results were compared to those obtained with the standard Escherichia coli-L-asparaginase . The surface structure of the lymphocytes (T cells) following L-asparaginase administration was not so different from that of normal lymphocytes . V . cholerae-L-asparaginase did not cause higher number of rosette formation in T cells as compared to the normal group . The study thus revealed that V . cholerae-L-asparaginase did not have a significant stimulatory or non-immunosuppressive effect on the lymphocyte functions. Microb Pathog, 1992 Nov, 13(5), 391 - 7 Reversal of hypotension induced by Vibrio vulnificus lipopolysaccharide in the rat by inhibition of nitric oxide synthase; Elmore SP et al.; Intravenous infusion of Vibrio vulnificus lipopolysaccharide (LPS) (1 mg/kg body wt) in rats caused a dramatic drop in mean arterial pressure within 10 min and a further decline in mean arterial pressure and heart rate which lead to death between 25 and 70 min . Rats treated with LPS followed 10 min later by the intravenous infusion of NG-monomethyl-L-arginine (L-NMMA, 20 mg/kg body wt) showed an initial drop in mean arterial pressure owing to the LPS infusion, followed by a transient rise in mean arterial pressure which lasted for approximately 40 min after the infusion of L-NMMA . The pressure values then remained level for at least 150 min post-LPS infusion . Control rats treated with equivalent volumes of saline infusion showed stable values of mean arterial pressure and heart rate . Additional control rats receiving L-NMMA alone showed the transient rise in mean arterial pressure, followed by a return to the baseline values . The results indicate that the symptoms of endotoxic shock resulting from V . vulnificus LPS may result in part from the stimulation of the activity of nitric oxide synthase . Inhibition of nitric oxide synthase by L-NMMA is a possible treatment for toxic shock induced by V . vulnificus. Mikrobiol Zh, 1992 Nov-Dec, 54(6), 50 - 4 {The agglutinability of cholera O-monoclonal immunoglobulins}; Alekseeva LP et al.; A set of 10 monoclonal antibodies specific for vibrio species and of 5 monoclonal antibodies specific for serovar (Ogawa) was studied . These monoclonal antibodies were directed toward V . cholerae O1 antigen in agglutination reaction and on slide plates . Monoclonal antibodies agglutinating typical strains of cholera vibrios with titration range from 1: 6000 to 1: 25,000 were selected . MA were revealed to interact with cholera vibrio strains with reduced agglutinability . The set of agglutinating O monoclonal immunoglobulins was developed for laboratory identification of cholera O1 vibrios. Mol Microbiol, 1992 Nov, 6(22), 3355 - 63 Purification of Aeromonas hydrophila major outer-membrane proteins: N-terminal sequence analysis and channel-forming properties; Jeanteur D et al.; Four outer-membrane proteins of Aeromonas hydrophila were purified and their N-terminal sequences and channel-forming properties were determined . Three could be matched with proteins from other species . One was a maltoporin, as its level increased when cells were grown in maltose-containing media, and the channel it formed was blocked by maltose . Another was like OmpF and OmpC of Escherichia coli, except that its channel fluctuated much more rapidly . The third protein, which was produced in low-phosphate medium, exhibited several properties of the general anion porin PhoE . The fourth showed no similarity to any known proteins . It had a unique N-terminus and it formed small sharply-defined cation-selective channels . Two other proteins which corresponded to OmpW of Vibrio cholerae and E . coli OmpA were partly characterized. Appl Environ Microbiol, 1992 Nov, 58(11), 3567 - 73 Cell surface characteristics of environmental and clinical isolates of Vibrio cholerae non-O1; Chaudhuri K et al.; The cell surfaces of several toxigenic and nontoxigenic environmental and clinical isolates of Vibrio cholerae non-O1 have been examined . The environmental strains, irrespective of toxigenicity, are significantly more resistant to antibiotics and detergents than are V . cholerae O1 strains . The clinical isolates of non-O1 vibrios are as sensitive to a wide variety of chemicals as the O1 vibrios . The environmental non-O1 strains are also less susceptible to lysis when treated with protein denaturants or neutral and anionic detergents than are O1 vibrios and the clinical non-O1 strains . In contrast to O1 vibrios, the environmental non-O1 vibrios do not have exposed phospholipids in their outer membranes . These features of the cell surfaces of environmental non-O1 vibrios might have a role in the better survival of these organisms under environmental fluctuations. Biochim Biophys Acta, 1992 Oct 19, 1111(1), 7 - 16 The mode of action of Vibrio cholerae cytolysin . The influences on both erythrocytes and planar lipid bilayers; Krasilnikov OV et al.; The interaction with erythrocytes of cholera cytolysin (CC) obtained from a non-01 Vibrio cholerae strain results in the osmotic rupture of target cells upon formation by CC of the waterfilled pores in their membranes . The aggregation of several toxin monomers is required for the formation of one CC channel with a radius of 0.9-1.0 nm . The investigations using planar bilayer lipid membranes suggest that the CC-induced pore is an interprotein anion selective channel carrying a fixed positive charge . The role of the charge was supported by the influence of pH on the selectivity, single conductance and voltage gating of the CC channels . The ability of the CC to modify both model and natural membranes has a maximum at pH 6.0-7.0 . It was found that CC channels insert into the membrane asymmetrically . The effect of proteolytic treatment of the channel by papain also indicates that the two entrances of the channel protrude from the plane of the membrane into the solution for different distances . It is proposed that the biological effects of the non-01 V . cholera cytolysin are based on its channel-forming activity. FEMS Microbiol Lett, 1992 Oct 15, 76(3), 215 - 9 Development and evaluation of a rapid, simple, sensitive, monoclonal antibody-based co-agglutination test for direct detection of Vibrio cholerae 01; Colwell RR et al.; Cholera epidemics caused by Vibrio cholerae 01 continue to represent a major public health concern in many developing countries . A rapid and simple test kit for the detection of V . cholerae 01 has been developed . The kit, CholeraScreen is a monoclonal antibody-based, co-agglutination test and is used directly with stool specimens . It does not include culturing the specimen and is performed without the need for sophisticated laboratory equipment . Specificity of the test was demonstrated, using 118 reference cultures, to which cross-reactions were not observed . Preliminary results of field trials carried out in Guatemala and Bangladesh demonstrated that the test is equally sensitive as conventional culture methods in detecting V . cholerae and, in many cases, more sensitive . The CholeraScreen test is simple, specific, and does not require culturing procedures, making it suitable for direct detection of cells of V . cholerae in clinical specimens, even in the field . Also, the test requires less than five minutes to complete. J Biol Chem, 1992 Oct 15, 267(29), 21139 - 45 A membrane-bound monoheme cytochrome c551 of a novel type is the immediate electron donor to P840 of the Chlorobium vibrioforme photosynthetic reaction center complex; Okkels JS et al.; A photosynthetic reaction center complex has been isolated from the green sulfur bacterium Chlorobium vibrioforme . Sodium dodecyl sulfate-polyacrylamide gel electrophoresis reveals polypeptides with apparent molecular masses of 80, 40, 18, 15, 9, and 6 kDa . Only the 18-kDa polypeptide is stained with 3,3',5,5'-tetramethylbenzidine, a heme-specific reagent . Oxidized minus reduced difference spectra show the presence of approximately one heme/P840 and the presence of a cytochrome c551 . Flash photolysis of P840 was followed by rereduction of P840+ and oxidation of cytochrome c551, both with a biphasic kinetic with t1/2 values of 7 and 50 microseconds . Using oligonucleotide probes derived from an N-terminal amino acid sequence of the 18-kDa polypeptide, a genomic clone was isolated . The sequence of the gene, which we designate cycA, predicts a single heme binding site (Cys-Asn-Lys-Cys-His) . The 621-base pair open reading frame encodes an apoprotein of 22,858 Da with three predicted membrane-spanning alpha-helices . No extensive sequence similarity is found to other cytochromes . Northern blotting indicates that the cycA gene is transcribed as a monocistronic mRNA . Southern blotting shows the presence of only one cycA gene in the C . vibrioforme and Chlorobium tepidum genomes . The unique membrane-bound monoheme cytochrome c551 of C . vibriforme is assigned to a new class of c-type cytochromes . The implications for the current view of evolution of photosynthetic reaction center complexes are discussed. J Infect Dis, 1992 Oct, 166(4), 837 - 41 Onset and duration of protective immunity in challenged volunteers after vaccination with live oral cholera vaccine CVD 103-HgR; Tacket CO et al.; CVD 103-HgR is a live oral cholera vaccine that, in phase I and II studies to date, has been well tolerated and immunogenic . In challenge studies of US volunteers conducted 4-5 weeks after vaccination, CVD 103-HgR provided significant protection against experimental cholera due to classical and El Tor Vibrio cholerae O1 . To determine the onset and duration of protection, two volunteer challenge studies were conducted: the first, 6 months after vaccination and the second, 8 days after vaccination . In both studies, CVD 103-HgR was 100% protective against diarrhea and significantly reduced the rate of shedding of vibrios after challenge with V . cholerae classical Inaba strain 569B, the virulent parent strain of CVD 103-HgR . Previously vaccinated subjects were less likely than naive controls to develop rises in titer of vibriocidal antibodies after challenge (P = .002), and the mean peak titer of vibriocidal antibodies was less than among controls . CVD 103-HgR can provide homologous protective immunity as soon as 8 days after vaccination and protection can persist for at least 6 months. Biokhimiia, 1992 Oct, 57(10), 1499 - 507 {Membrane ATPase of Vibrio alginolyticus . Ion transport activity and homology with F0F1-ATPase from E . coli}; Dmitriev OIu et al.; F0F1-ATPase has been isolated from the marine alkali-resistant bacterium Vibrio alginolyticus . The enzyme subunits cross-reacted with antibodies against subunits alpha, beta, gamma, epsilon, and b of E . coli ATPase . The purified ATPase was reconstituted into liposomes effecting an ATP-dependent uptake of H+ . Proton transport was inhibited by the ATPase blockers DCCD, triphenyltin, and venturicidin . Na+ ions had no effect on ATP-dependent proton transport . No ATP-dependent transport of Na+ was detected in proteoliposomes. Appl Environ Microbiol, 1992 Oct, 58(10), 3419 - 22 Use of an oligonucleotide probe to detect Vibrio parahaemolyticus in artificially contaminated oysters; Lee C et al.; A 26-mer oligonucleotide specific to Vibrio parahaemolyticus was synthesized from a 1,275-bp thermostable direct hemolysin (tdh) gene . This oligonucleotide probe specifically reacted with DNA from 89 of 95 V . parahaemolyticus isolates but not with DNA from other vibrios or other enteric and nonenteric organisms (n = 48) . The probe hybridized with Southern blots of 0.5-kb HindIII-restricted chromosomal DNA fragments from all but five V . parahaemolyticus test isolates . The probe could be used to directly identify V . parahaemolyticus in artificially contaminated food without an isolation step. Appl Environ Microbiol, 1992 Oct, 58(10), 3410 - 2 Occurrence of tributyltin-tolerant bacteria in tributyltin- or cadmium-containing seawater; Suzuki S et al.; Tributyltin chloride (TBTCl)-tolerant bacteria accounted for 90% of the flora in natural seawater to which TBTCl was added . These tolerant bacteria were insensitive to 250 nmol of TBTCl per disc, and all were Vibrio species . Total counts of viable bacteria did not decrease upon storage of the TBTCl-treated seawater, indicating that enrichment of tolerant strains took place . Addition of CdSO4 to seawater resulted in the occurrence of TBTCl-tolerant bacteria as well as Cd-tolerant bacteria, suggesting some correlation of Cd tolerance and TBTCl tolerance. Appl Environ Microbiol, 1992 Oct, 58(10), 3257 - 62 Seasonal incidence of Vibrio vulnificus in the Great Bay estuary of New Hampshire and Maine; O'Neill KR et al.; Vibrio vulnificus, a normal bacterial inhabitant of estuaries, is of concern because it can be a potent human pathogen, causing septicemia, wound infections, and gastrointestinal disease in susceptible hosts . From May 1989 through December 1990, oysters and/or water were obtained from six areas in the Great Bay estuary of New Hampshire and Maine . Water was also sampled from three freshwater sites that lead into these areas . V . vulnificus was first detected in the estuary in early July and remained present through September . V . vulnificus was isolated routinely during this period from oysters and water of the Squamscott, Piscataqua, and Oyster Rivers but was only isolated twice from the oysters or water of the Great Bay itself . This study determined that there was a strong correlation (by analysis of variance) between temperature, salinity, and the presence of V . vulnificus in water and oysters . However, other unidentified factors appear to influence its presence in certain areas of the estuary. J Vet Med Sci, 1992 Oct, 54(5), 851 - 5 Chemotactic activity of hemocytes derived from a brackish-water clam, Corbicula japonica, to Vibrio parahaemolyticus and Escherichia coli strains; Kumazawa NH et al.; Hemocytes from adult and juvenile specimens of a brackish-water clam, Corbicula japonica, were attracted chemotactically to live cells of Vibrio parahaemolyticus and Escherichia coli strains in a balanced salt solution, which was enhanced significantly in the presence of respective C . japonica plasma . Chemotactic attractions of adult's and juvenile's hemocytes were seen also in artificial seawater at a similar level to those in the balanced salt solution . Chemotactic attractions of juvenile's hemocytes to these strains were lower in level than those of adult's hemocytes . C . japonica plasma seems to facilitate for C . japonica hemocytes to recognize these organisms. J Clin Microbiol, 1992 Oct, 30(10), 2730 - 2 Vibrio-associated gastroenteritis in the lower Cross-River Basin of Nigeria; Ndon JA et al.; A total of 120 Vibrio species were isolated from 588 patients with acute diarrheal disease during an outbreak of gastrointestinal tract infections at different locations in the lower Cross River Basin of Nigeria . Vibrio cholerae O1, biotype El Tor, serotype Ogawa, was the prominent organism isolated from the Vibrio-associated diarrheal cases . During the 3 months of study, V . cholerae non-O1 was recovered from 10 patients while Vibrio parahaemolyticus was recovered from 19 patients . The significance of this study is the recognition that there is an ecological niche which supports V . cholerae non-O1 and V . parahaemolyticus in the Cross River Basin of Nigeria. J Bacteriol, 1992 Oct, 174(19), 6221 - 9 A 14-kilodalton inner membrane protein of Vibrio cholerae biotype e1 tor confers resistance to group IV choleraphage infection to classical vibrios; Biswas SK et al.; Choleraphage phi 149 differentiates the two biotypes, classical and el tor, of Vibrio cholerae . This phage cannot replicate in V . cholerae biotype el tor cells because the concatemeric DNA intermediates produced are unstable and cannot be chased to mature phage DNA . A V . cholerae biotype el tor gene coding for a 14,000-Da inner membrane protein which destabilizes the concatemeric DNA intermediates by hindering their binding to the cell membrane has been identified . Presumably, a 22,000-Da V . cholerae biotype el tor protein might also have a role in conferring phage phi 149 resistance to cells belonging to the biotype el tor . A nucleotide sequence homologous to the 1.2-kb V . cholerae biotype el tor DNA coding for both the 14,000- and 22,000-Da proteins is present in all strains of classical vibrios but is not transcribed . The nucleotide sequence of the gene coding for the 14,000-Da protein has been determined. FEMS Microbiol Lett, 1992 Oct 1, 76(1-2), 179 - 84 TCP pilus biosynthesis in Vibrio cholerae O1: gene sequence of tcpC encoding an outer membrane lipoprotein; Ogierman MA et al.; The nucleotide sequence of the tcpC gene has been determined . It encodes a 53995-Da protein precursor with a signal sequence and cleavage site typical of a number of outer membrane lipoproteins, which are cleaved by the equivalent of signal peptidase II (Lsp) of Escherichia coli . The location of the tcpC gene is such that it is predicted to be translationally coupled to the 5' and 3' flanking genes, tcpY and tcpD, respectively, indicating that it forms part of an operon . Together with the lipoprotein signal sequence and the several hydrophobic domains it seems likely that TcpC is a surface-anchored trans-outer membrane lipoprotein. Infect Immun, 1992 Oct, 60(10), 4278 - 84 Analysis of expression of toxin-coregulated pili in classical and El Tor Vibrio cholerae O1 in vitro and in vivo; Jonson G et al.; The expression of toxin-coregulated pili (TCP) and their structural subunit TcpA was compared in 20 strains of Vibrio cholerae of the classical and El Tor biotypes . Bacteria were isolated from the intestines of rabbits with experimental cholera and compared with the same strains grown under optimal TCP expression conditions in vitro . Immunoblotting revealed that TcpA production was induced in both biotypes after vibrios entered the intestinal milieu; TcpA-negative inocula gave rise to TcpA-positive vibrios after multiplication in the gut . The levels of TcpA expressed during growth in the intestine were, for most strains, comparable to those attained under optimal growth conditions in vitro . Of 11 classical strains tested, 10 expressed TCP antigen on the bacterial surface at levels comparable to or exceeding those seen after growth in vitro as determined by an inhibition enzyme-linked immunosorbent assay . In contrast, only one of the nine El Tor strains studied produced detectable amounts of TCP surface antigen in vivo and no fimbriae or surface antigen reacting with anti-TCP serum was found on El Tor vibrios from human cholera stools . Distinct TCP fimbriae were observed by immunoelectron microscopy on classical-biotype vibrios grown either in rabbit intestines or in vitro but were not detected on El Tor vibrios . The results show that TCP is expressed on V . cholerae O1 of the classical biotype but not on V . cholerae O1 of the El Tor biotype in the intestines of rabbits with experimental cholera infection. J Gen Microbiol, 1992 Oct, 138 ( Pt 10), 2075 - 82 An analysis of the effect of changes in growth temperature on proteolysis in vivo in the psychrophilic bacterium Vibrio sp . strain ANT-300; Araki T; In the psychrophilic bacterium Vibrio sp . strain ANT-300, the rate of protein degradation in vivo, measured at fixed temperatures, increased with elevation of the growth temperature . A shift in growth temperature induced a marked increase in this rate . Dialysed cell-free extracts hydrolysed exogenous insulin, globin and casein (in decreasing order of activity) but did not hydrolyse exogenous cytochrome c . Cells contained at least seven protease separated by DEAE-Sephacel chromatography, one of which was an ATP-dependent serine protease . The ATP-dependent proteolytic activity in extracts of cells incubated for 3 h at 16 degrees C after a shift-up from 0 degrees C increased to a level 36% and 17% higher than that of cells grown at 0 degrees C and 13 degrees C, respectively . A shift-down to 0 degrees C from 13 degrees C induced only a slight increase in the proteolytic activity . Extracts of all cells, whether exposed to temperature shifts or not, showed the same temperature dependence with respect to both ATP-dependent and ATP-independent protease activity . In all the extracts these proteases also exhibited the same heat lability . The ATP-dependent protease was inactivated by incubation at temperatures above 25 degrees C . There was an increase in ATP-independent protease activity during incubation at temperatures between 25 and 30 degrees C, but a decrease at 35 degrees C and higher . These results suggest that the marked increases in proteolysis in vivo, caused by a shift in temperature, may result not only from increases in levels of ATP-dependent serine protease(s) but also from increases in the susceptibility of proteins to degradation. Eur J Cell Biol, 1992 Oct, 59(1), 12 - 20 Interaction of rat peritoneal macrophages with homologous sialidase-treated thrombocytes in vitro: biochemical and morphological studies . Detection of N-(O-acetyl)glycoloylneuraminic acid; Kluge A et al.; Sialidase treatment of rat thrombocytes led to an increased binding of these cells to homologous peritoneal macrophages, but had no significant effect on the rate of phagocytosis during the experimental time . As revealed by electron microscopy, the partially desialylated thrombocytes adhere to macrophages predominantly via a small part of the membrane in a way that the discoidal cells adopt a vertical position with regard to the macrophage surface . One adherent macrophage was able to bind up to 55 sialidase-treated thrombocytes . Maximum binding was already reached after release of 13% of sialic acids . This interaction could be inhibited by free D-galactose and compounds with terminal D-galactose residues . Bound thrombocytes were released from the macrophages by treatment with lactose or EDTA . These experiments suggest that the interaction is mediated by a galactose-specific receptor on the macrophage surface and that galactose on thrombocytes is not recognized if it is masked by terminal sialic acid residues . The total sialic acid amount of the thrombocytes studied was about 70 micrograms sialic acid/10(10) cells being composed of 78% N-glycoloylneuraminic acid, 17% N-acetylneuraminic acid and 5% of the novel sialic acid N-(O-acetyl)glycoloylneuraminic acid, which was identified by mass spectrometry . Sixty-two percent of these sialic acids were susceptible to enzymic hydrolysis with Vibrio cholerae sialidase. J Mol Biol, 1992 Sep 20, 227(2), 572 - 4 Expression, purification and crystallization of the Vibrio harveyi acyltransferase; Swenson L et al.; We have obtained X-ray quality single crystals of Vibrio harveyi acyltransferase . The protein was obtained from V . harveyi by a gene mobilization expression system . The crystals are monoclinic (space group P2(1), a = 89.9 A, b = 83.6 A, c = 47.1 A, beta = 97.3 degrees) with two molecules related by a pronounced non-crystallographic dyad in the asymmetric unit, with a solvent content of approximately 50% . The diffraction pattern from fresh crystals extends beyond 2 A resolution using sealed tube CuK alpha radiation . The elucidation of the three-dimensional structure of this enzyme, believed to contain a proteinase-like catalytic triad, which resembles in many ways other eukaryotic fatty acid chain terminating enzymes, may have important consequences for our understanding of the molecular basis of the final stages of the synthesis of fatty acids. Lancet, 1992 Sep 19, 340(8821), 689 - 94 Safety and immunogenicity of single-dose live oral cholera vaccine CVD 103-HgR in 5-9-year-old Indonesian children; Suharyono et al.; Oral vaccines offer great promise as public-health measures to prevent disease in less-developed countries . CVD 103-HgR, a genetically engineered, attenuated, Vibrio cholerae O1 strain has proved effective in industrialised countries . We have assessed the safety, immunogenicity, and excretion of this live cholera vaccine in children in north Jakarta, Indonesia . 412 children aged 5-9 years received single doses of 5 x 10(6), 5 x 10(7), 5 x 10(8), 5 x 10(9), or 1 x 10(10) colony forming units (CFU) of CVD 103-HgR or placebo (5 x 10(8) inactivated Escherichia coli K-12) with buffer . All doses were well tolerated . The 5 x 10(8) CFU dose, which is highly immunogenic in subjects in industrialised countries (greater than 90% seroconversion), elicited seroconversions of vibriocidal antibody in only 16% of Indonesian children . By contrast, a single 5 x 10(9) CFU dose of vaccine resulted in high rates (75% and 87%) of seroconversion with two different batches of vaccine . A batch prepared with a centrifugation step gave significantly higher geometric mean titres (16-fold increase over baseline) than did a batch in which there was a filtration step between fermentation and lyophilisation (10-fold increase over baseline) . At a 5 x 10(9) CFU dose, CVD 103-HgR is well tolerated and highly immunogenic in Indonesian children and should therefore be further investigated for use as a one-dose live oral cholera vaccine in developing countriesPIP: In February-March 1990, health workers administered a single dose of the oral cholera CVD 103 HgR vaccine with 5 x 100 million colony forming units (CFU), 5 x 10 million CFU, or 5 x 1 million CFU or of a placebo to 274 5-9 year old children in a village within the catchment area of the Infectious Diseases Hospital in North Jakarta, Indonesia . In September-October 1990, they gave a dose of the same vaccine containing either 5 x 1 billion CFU or 1 x 10 billion CFU from 1 of 2 different batches or a placebo (5 x 100 million inactivated Escherichia coli K 12) to 140 5-9 year old children . 70 children also received an extra dose of buffer . They conducted these trials to examine the safety, immunogenicity, and excretion of this live cholera vaccine . The higher dose genetically engineered oral cholera vaccine (5 x 1 billion CFU) resulted in higher seroconversion rates than the 5 x 100 million CFU vaccine which has 90% seroconversion rates among North American and European children (16% vs . 75-87% for 2 different batches) . The centrifuged prepared vaccine resulted in significantly greater geometric mean titers (16-fold rise over baseline) than did the filtered prepared vaccine (10-fold rise over baseline) (p=.001) . The extra buffer did not improve immunogenicity of CVD 10 HgR in these children . None of the 124 children who took the 5 x 1 billion or 1 x 10 billion CFU dose excreted the attenuated strain of Vibrio cholerae 01 . Thus this recombinant vaccine strain would unlikely enter the environment or b transmitted to others . The frequency of adverse reactions was basically the same for the vaccine and the placebo . These results showed that the Indonesian children tolerated a single 5 x 1 billion dose of CVD 103 HgR well and induced considerable immunogenicity . Future studies using the same dose in 2-4 year old children are planned . Infect Immun, 1992 Sep, 60(9), 3916 - 7 Safety and immunogenicity of a booster dose of Vibrio cholerae CVD 103-HgR live oral cholera vaccine in Swiss adults; Cryz SJ Jr et al.; Adult volunteers received a booster dose (4 x 10(8) CFU) of attenuated Vibrio cholerae CVD 103-HgR oral vaccine 15 or 24 months after primary immunization . The immune response was modest, presumably due to rapid clearance of the vaccine strain by a primed immune system. Appl Environ Microbiol, 1992 Sep, 58(9), 2776 - 82 Virulence characteristics of clinical and environmental isolates of Vibrio vulnificus; Stelma GN Jr et al.; Twenty-four randomly selected clinical and environmental Vibrio vulnificus isolates were tested for virulence in iron-overloaded mice (250 mg of iron dextran per kg of body weight) . The log10 50% lethal doses of 17 isolates were lower by greater than or equal to 3.5 log10 units in iron-overloaded mice than in control mice . These isolates were classified as virulent . The 50% lethal doses of these virulent isolates were also lower in mice that were immunosuppressed by treatment with cyclophosphamide (150 mg/kg) . Four of the seven isolates initially classified as avirulent were virulent in mice that were simultaneously iron overloaded and immunosuppressed . These isolates were classified as moderately virulent . The remaining three isolates were avirulent under all conditions . The incidence of virulent strains among clinical and environmental isolates did not differ . The virulent isolates produced high titers of hemolysin, were resistant to inactivation by serum complement, produced phenolate siderophore, and utilized transferrin-bound iron . The moderately virulent isolates differed from the virulent isolates only in their increased sensitivity to inactivation by serum complement . The avirulent isolates differed from those of the other two classes in their inability to either produce significant amounts of phenolate siderophore or utilize transferrin-bound iron . A modified agar plate diffusion method for transferrin-bound iron utilization was developed to differentiate the two classes of virulent isolates from the avirulent isolates in vitro. J Clin Microbiol, 1992 Sep, 30(9), 2302 - 6 Development and testing of a nonradioactive DNA oligonucleotide probe that is specific for Vibrio cholerae cholera toxin; Wright AC et al.; An alkaline phosphatase-labeled oligonucleotide DNA probe (CTAP) that was specific for the cholera toxin gene (ctxA) was identified . All cholera toxin-producing strains of Vibrio cholerae, regardless of serotype, hybridized with the CTAP probe, while nontoxigenic strains from either environmental sources or from deletion or substitution mutations did not hybridize . Unlike the whole-gene probes for either ctxA or for the heat-labile toxin or Escherichia coli (eltA), this 23-base sequence did not hybridize with E . coli or with vibrios other than V . cholerae that produce related toxins . By using CTAP to identify colonies grown on nonselective medium, V . cholerae was enumerated at concentrations of 10(3) to 10(7)/g from stool samples of volunteers who had ingested V . cholerae O1 strain 569B . CTAP provides a specific and sensitive tool for diagnosis and environmental monitoring of cholera toxin-producing V . cholerae. J Bacteriol, 1992 Sep, 174(17), 5732 - 9 Behavioral analysis of Vibrio parahaemolyticus variants in high- and low-viscosity microenvironments by use of digital image processing; Lawrence JR et al.; Digital image analysis and light microscopy were used to study and quantify the growth and behavior of two variants and selected flagellar mutants of Vibrio parahaemolyticus in glass flow cells under high- and low-viscosity conditions . The observations showed a series of surface-associated behaviors, including attachment, microcolony formation, migration, chemotactic movements, and aggregation, indicating a substantial degree of adaptive flexibility and multicellular behavior during growth of V . parahaemolyticus at interfaces. Infect Immun, 1992 Sep, 60(9), 3539 - 45 Enterotoxigenicity of Vibrio parahaemolyticus with and without genes encoding thermostable direct hemolysin; Nishibuchi M et al.; Vibrio parahaemolyticus produces a thermostable direct hemolysin (TDH) that has been implicated in the pathogenesis of diarrheal disease caused by this organism . However, previous studies attempting to demonstrate the contribution of the hemolysin to virulence have been inconclusive . We investigated this putative virulence factor by using an isogenic TDH-negative (TDH-) strain constructed by specifically inactivating the two copies of the tdh gene encoding TDH . The enterotoxigenicities of the parent strain (AQ3815) and the mutant strain were tested by adding sterile culture supernatants to rabbit ileal tissue mounted in Ussing chambers . The culture filtrate of the parent strain produced a significant increase in short-circuit current (Isc), compared with the change induced by the TDH- mutant . The capacity of the culture filtrate of AQ3815 to increase the Isc was reduced by neutralization with anti-TDH serum, and the return of the cloned tdh gene to the TDH- mutant restored the ability to increase the Isc . These results were corroborated by rabbit ileal loop assays in which AQ3815 caused fluid accumulation but the TDH- mutant did not . No microscopic damage was seen in mucosal tissues exposed to the culture filtrate of either strain . These results indicate that TDH has an enterotoxigenic effect on rabbit small intestine and could be responsible for the watery diarrhea seen with V . parahaemolyticus. Mikrobiol Zh, 1992 Sep-Oct, 54(5), 105 - 12 {The comparative characteristics of Vibrio cholerae hemolysins}; Telesmanich NR; Biochemical and biological properties of in vivo and in vitro hemolysins of cholera germs isolated by different authors as well as hemolysins of Vibrio parahaemolyticus and Escherichia coli have been comparatively characterized . According to the above data hemolysin of cholera germ El Tor is a thermolabile protein with molecular mass of 60-80 thou.; it is cytotoxic, enteropathogenic and lyses all the species of erythrocytes . Hemolysins of V . eltor and germs are immunologically related . Hemolysin of the cholera germ is thermostable and has molecular mass of 20-40 thou., that makes it similar to hemolysin of V . parahaemolyticus. Appl Environ Microbiol, 1992 Sep, 58(9), 2861 - 5 Induction of melanin biosynthesis in Vibrio cholerae; Coyne VE et al.; Vibrio cholerae synthesized the pigment melanin in response to specific physiological conditions that were stressful to the bacterium . Pigmentation was induced when V . cholerae was subjected to hyperosmotic stress in conjunction with elevated growth temperatures (above 30 degrees C) . The salt concentration tolerated by V . cholerae was lowered by additional abiotic factors such as acidic starting pH of the growth medium and limitation of organic nutrients . Although the amount of toxin detected in the culture supernatant decreased significantly in response to stressful culture conditions, no correlation between the physiological conditions that induced melanogenesis and expression of OmpU or cholera toxin was detected . Since conditions that induce melanin production in V . cholerae occur in both the aquatic environment and the human host, it is possible that melanogenesis has a specific function with respect to the survival of the bacterium in these habitats. J Diarrhoeal Dis Res, 1992 Sep, 10(3), 161 - 3 Acute secretory travellers' diarrhoea caused by Vibrio cholerae non-01 which does not produce cholera-like or heat-stable enterotoxins; Bhattacharya SK et al.; An Australian tourist suffering from severe acute watery diarrhoea and dehydration due to Vibrio cholerae non-01 was studied . The V . cholerae strain isolated from the patient belonged to serovar 05 . The organism did not produce any of the conventional enterotoxins including cholera-toxin (CT) or heat-stable toxins (NAG-ST) that are known to be associated with intestinal secretion . This report suggests that toxin(s) other than CT-like or NAG-ST may be involved in the pathogenesis of diarrhoea by some V . cholerae non-01 strains. Eur J Epidemiol, 1992 Sep, 8(5), 743 - 7 The protease from Vibrio cholerae nicks arginine at position 192 from the N-terminus of the heat-labile enterotoxin a subunit from enterotoxigenic Escherichia coli; Ichinose Y et al.; It was examined where a protease purified from Vibrio cholerae might nick the heat-labile enterotoxin (LT) A subunit from enterotoxigenic Escherichia coli . LT was digested by the protease and contained a fragment which had the same mobility on SDS-PAGE as that of the A1 fragment of LT digested by trypsin . The biological activity of LT by this protease was also identical to that of LT by trypsin . The amino acid sequence of the N-terminus of the A2-like fragment was Thr-Ser-Thr-Gly, which corresponded to the sequence from 193 to 196 of the A subunit . These data suggest that this protease, like trypsin, nicks arginine at position 192 from the N-terminus of the A subunit and that the biological activation of LT by this protease is similar to that by trypsin. J Appl Bacteriol, 1992 Sep, 73(3), 197 - 202 Incidence of toxigenic vibrios in foods available in Taiwan; Wong HC et al.; A total of 1088 vibrios and related species were isolated from seafood and aquacultured foods available in Taiwan . They were identified as Vibrio alginolyticus, V . cholerae, V . fluvialis I, V . fluvialis II, V . parahaemolyticus, V . mimicus, Aeromonas caviae, A . hydrophila, A . sobria and other species . Incidence of these Vibrio and Aeromonas species in these foods was high . Vibrio parahaemolyticus was frequently found in seawater and in foods of freshwater origin . The Vibrio isolates were examined for enzymatic and toxigenic activities . Most of them showed strong lipase or protease activities . Haemolytic activities of V . cholerae, V . fluvialis I and V . fluvialis II isolates were mostly strong . About 49% showed cytotoxic activity and 5% cytotonic activity in Chinese hamster ovary cell culture assay . Nevertheless, only three non-O1 V . cholerae (2.07%) and two V . parahaemolyticus isolates (1.65%) produced cholera toxin and thermostable direct haemolysin activity, respectively . Various toxigenic vibrios may be important food-borne pathogens in this region because of their high incidence in foods. Eur J Immunol, 1992 Sep, 22(9), 2277 - 81 The adjuvant effect of Vibrio cholerae and Escherichia coli heat-labile enterotoxins is linked to their ADP-ribosyltransferase activity; Lycke N et al.; This study addressed the question of whether the mucosal adjuvant property of cholera toxin (CT) and the structurally closely related Escherichia coli heat-labile toxin (LT) requires the enterotoxic and adenylate cyclase/cAMP activating property of these molecules . Therefore, we investigated the cytotoxic and adjuvant abilities of the enterotoxins and compared the results with those obtained with the non-toxic CT and LT derivatives; recombinant CTB (rCTB) and a mutated LT (mLT), which had a single amino acid substitution in position 112 (Glu----Lys) of the A subunit . Detailed functional studies revealed that, in contrast to the enterotoxins, both rCTB and mLT lacked ADP-ribosylating and cAMP-stimulating abilities . However, similar membrane ganglioside GM1-receptor binding ability of all the putative adjuvants was demonstrated . When the probe antigen, keyhole limpet hemocyanin (KLH), was given perorally together with CT or LT strong gut mucosal anti-KLH immune responses were stimulated, whereas no or very low anti-KLH responses were seen in the groups which received antigen admixed with rCTB or the mLT . Moreover, the specific serum antibody responses to the various immunization protocols closely paralleled the local anti-KLH response in the gut . From these results it appears that the adjuvant mechanism of LT, and probably also of CT, is linked to the ability to ADP-ribosylate and to stimulate cAMP formation . However, this study does not unequivocally rule out other possibilities such as interactions by the A1 fragment of CT or LT with other G-proteins than Gs alpha or events that parallel or precede the effects on the adenylate cyclase/cAMP system . Thus, the levels of ADP-ribosylation and cAMP-induction that are required and the key event or target cell that is responsible for the adjuvant effect of CT and LT remain to be elucidated . Studies are underway to address these issues. Appl Environ Microbiol, 1992 Sep, 58(9), 2771 - 5 Effects of temperature abuse on survival of Vibrio vulnificus in oysters; Murphy SK et al.; Opaque and translucent morphotypes of a TnphoA-containing strain of Vibrio vulnificus were fed to oysters, which were subsequently stored at temperatures ranging from 0.5 to 22 degrees C for 10 days . Samples of oysters were homogenized and plated at intervals to determine the cell density of V . vulnificus and total aerobic population of bacteria present . At all temperatures, the numbers of V . vulnificus (both morphotypes) declined over the 10-day study period . The same observation was made with a lower inoculum of V . vulnificus . Identical experiments with shucked oysters showed a more rapid decrease in V . vulnificus . Identical experiments with shucked oysters showed a more rapid decrease in V . vulnificus to levels below limits of detection . Little change in the total bacterial counts was observed in shellstock oysters at any of the test temperatures, whereas incubation at the higher temperatures (17 and 22 degrees C) resulted in large increases in total counts in shucked oysters . These data suggest that temperature abuse of oysters may not be a factor in increasing the public health risk of V . vulnificus through raw oyster consumption . However, the data also suggest that even with proper storage, indigenous levels of V . vulnificus may remain sufficiently higher in shellstock oysters to produce infection in compromised hosts. Acta Gastroenterol Belg, 1992 Sep-Dec, 55(5-6), 430 - 6 Recent progress in cholera vaccination; Pierre PG et al.; Cholera disease remains an important cause of morbidity and mortality in the third world . The parenteral cholera vaccine actually used offers only a 50% protection during 6 months . As Vibrio cholerae and its toxin don't cross the gut wall, the aim of new vaccines is to prevent the colonization and growth of the vibrio in the jejuno-ileum and to inhibit the fixation of cholera toxin (CT) on its enterocyte membrane receptor . This can be afforded by stimulation of the gut local immune system mainly represented by secretory IgA antibodies (Abs) . New vaccines should comprise both bacterial and CT antigens and must be given by the oral route to induce the production of specific secretory IgA Abs in the gut . Four different ways are actually under study to produce an oral cholera vaccine . 1 . Combination of CT-B subunit and killed vibrios . 2 . Live recombinant Vibrio cholerae in which the CT coding gene has been deleted . 3 . Synthetic peptides reproducing some immunodominant CT-epitopes . 4 . Manipulation of the idiotypic network to induce the production of Abs mimicking CT-epitopes . This paper reviews the actual developments and advantages of these four approaches. Appl Environ Microbiol, 1992 Sep, 58(9), 3078 - 82 Vibrio cholerae non-O1 isolated from ayu fish (Plecoglossus altivelis) in Japan; Kiiyukia C et al.; A fish pathogen, Vibrio cholerae non-O1, was isolated from diseased ayu fish (Plecoglossus altivelis) collected from rivers in eight prefectural districts of Japan . This organism was found to have biochemical characteristics similar to those of V . cholerae non-O1, except that our isolates were negative for ornithine decarboxylase . Antiserum against an ayu isolate did not agglutinate with the majority of environmental V . cholerae non-O1 isolates, but a major O antigen was common among the ayu isolates . All strains were hemolytic to sheep erythrocytes, and oral administration of culture supernatants induced fluid accumulation in suckling mice . However, the crude toxin was not lethal to adult mice, and no cholera toxin-like enterotoxins were detected. FEBS Lett, 1992 Aug 24, 308(3), 315 - 9 Activation mechanism of human Hageman factor-plasma kallikrein-kinin system by Vibrio vulnificus metalloprotease; Miyoshi S et al.; Vivrio vulnificus, an opportunistic human pathogen, secretes a metalloprotease (VVP) . The VVP inoculated into a guinea pig is known to generate bradykinin through activation of the Hageman factor-plasma kallikrein-kinin system . VVP was shown to possess the ability to activate the human system through the same mechanism as that clarified in the guinea pig system, namely, VVP converted both human zymogens (Hageman factor and plasma prekallikrein) to active enzymes (activated Hageman factor and plasma kallikrein), and the then generated kallikrein liberated bradykinin from high-molecular-weight kininogen . However, in the presence of plasma alpha 2-macroglobulin (alpha 2M), the VVP action was drastically decreased . This finding suggests that the human system might be activated only at the interstitial-tissue space which contains negligible amounts of alpha 2M or in the bloodstream of the individuals whose plasma alpha 2M level is extremely reduced. J Bacteriol, 1992 Aug, 174(16), 5211 - 8 Structural analysis of the acfA and acfD genes of Vibrio cholerae: effects of DNA topology and transcriptional activators on expression; Parsot C et al.; The Vibrio cholerae acfA, B, C, and D genes are involved in the synthesis of a colonization factor; their expression is under the control of ToxR, the cholera toxin transcriptional activator . By a combination of Southern blot analysis, cloning, and nucleotide sequence analysis, we determined that the acf genes are clustered on a 5-kb region, the acfA and acfD genes are transcribed divergently, and the translation start sites of the two genes are separated by only 173 bp . Expression from the acfA and acfD promoters in V . cholerae was studied by using acfA:phoA translational and acfD-lacZ transcriptional fusions; when carried by the chromosome, the acfA-acfD intergenic region flanked by the two reporter genes was found to contain the cis-acting element(s) necessary for the environmental regulation of the two promoters . However, this regulation was almost completely abolished when the same construction was carried by a low-copy-number plasmid . These results suggested that differences in DNA topology between the plasmid versus the chromosomal constructs might influence the expression of the acfA and acfD promoters . Support for this conclusion was obtained by showing that ToxR-dependent but not basal expression of both promoters was strongly inhibited by nalidixic acid and novobiocin, two DNA gyrase inhibitors, suggesting that the activation of these promoters is affected by changes in DNA supercoiling . Expression of the acfA and acfD promoters was also investigated in the heterologous host Escherichia coli harboring plasmids expressing either ToxR or ToxT, two transcriptional activators of the V . cholerae virulence genes . ToxR activated the acfD promoter 2.5-fold but inhibited the acfA promoter 2-fold . In contrast, the expression of the acfA promoter was activated 10-fold and that of the acfD promoter was activated 3-fold by ToxT, supporting the previously proposed cascade model for organization of the ToxR regulon. Gene, 1992 Aug 1, 117(1), 103 - 6 Isolation and characterization of new restriction endonucleases from Vibrio parahaemolyticus: VpaK32I enzyme with the class-IIS heptanucleotide specificity, GCTCTTCN1/N4; Miyahara M et al.; Six restriction endonucleases (ENases), classified into four different specificities, were found in a screen among 68 reference strains of Vibrio parahaemolyticus of human origin . Five of these ENases are isoschizomers of well-known ENases, while the remaining one, designated VpaK32I, is a novel and highly efficient class-IIS ENase with the hepatanucleotide recognition site, 5'-GCTCTTC(1/4)-3'. Infect Immun, 1992 Aug, 60(8), 3201 - 8 Synthesis, characterization, and some immunological properties of conjugates composed of the detoxified lipopolysaccharide of Vibrio cholerae O1 serotype Inaba bound to cholera toxin; Gupta RK et al.; Protection against cholera has been correlated with the level of serum vibriocidal antibodies . The specificity of these vibriocidal antibodies was mostly to the lipopolysaccharide (LPS) . We synthesized conjugates of detoxified LPS with cholera toxin (CT) and other proteins in order to elicit serum LPS antibodies with vibriocidal activity . Treatment with hydrazine (deacylated LPS) reduced the endotoxic properties of the LPS to clinically acceptable levels and resulted in a molecule larger and more antigenic than the saccharide produced by acid hydrolysis . More immunogenic conjugates resulted from multipoint compared with single-point attachment of the deacylated LPS to the protein . The conjugates containing CT had low levels of pyrogen and no toxic activity upon Chinese hamster ovary cells and elicited booster responses of vibriocidal and CT antibodies when injected subcutaneously as saline solutions into mice; the vibriocidal titers were similar to those elicited by comparable doses of cellular vaccines . We suggest how serum vibriocidal antibodies might prevent cholera. Clin Infect Dis, 1992 Aug, 15(2), 271 - 6 Vibrio vulnificus infection in Taiwan: report of 28 cases and review of clinical manifestations and treatment; Chuang YC et al.; From May 1985 through July 1990, 28 episodes of Vibrio vulnificus infection in 27 patients were encountered in five major hospitals in Taiwan . The ages of patients ranged from 19 to 76 years; the ratio of male to female patients was 2:1 . Eighteen episodes manifested as bacteremia and eight as wound infections alone . One patient each developed gastroenteritis and pneumonia after nearly drowning . Twenty-three patients exhibited skin manifestations . Twenty patients had underlying diseases . All patients were treated with antibiotics, and 14 also underwent some form of surgical treatment (incision and drainage, fasciotomy, debridement, or amputation) . Thirteen of the 28 episodes were preceded by precipitating factors; most were due to ingestion of seafood or exposure of abraded skin to salt water . Ten of the 18 septicemic patients died--most within 48 hours of hospitalization . One patient without bacteremia who had a wound infection died . Results of in vitro susceptibility studies suggested that ampicillin or a third-generation cephalosporin would be effective . Susceptibility to aminoglycosides was observed for greater than 90% of isolates . We recommend combined therapy with a third-generation cephalosporin or ampicillin and an aminoglycoside along with appropriate surgical therapy for the treatment of V . vulnificus infection. Arch Dis Child, 1992 Aug, 67(8), 1027 - 9 Breast feeding and oral rehydration at home during diarrhoea to prevent dehydration; Faruque AS et al.; In a case-control study we evaluated the role of maternal behaviour, as reflected in maintenance of breast feeding and the use of oral rehydration therapy (ORT) at home during acute diarrhoea, in preventing dehydration in infants and young children . A systematic 5% sample was taken of all children aged 1-35 months attending the treatment centre of the International Centre for Diarrhoeal Disease Research, Bangladesh, with acute watery diarrhoea of six days or less between August 1988 and September 1989 . There were 285 children with moderate or severe dehydration as cases and 728 with no dehydration as controls in the study . In a multivariate analysis using a logistic regression model we showed that withdrawal of breast feeding during diarrhoea was associated with a five times higher risk of dehydration compared with continuation of breast feeding during diarrhoea at home . Lack of ORT with either complete formula or a salt and sugar solution at home was associated with 57% higher risk of dehydration compared with receipt of a reasonable amount of ORT after controlling for several confounders . The confounding variables--that is, lack of maternal education, history of vomiting, high stool frequency, young age and infection with Vibrio cholerae 01--were also shown to be risk factors of dehydration . Health education programmes should promote continued breast feeding and adequate oral rehydration therapy for infants with acute diarrhoea at home. Appl Environ Microbiol, 1992 Aug, 58(8), 2679 - 82 Development of the immunomagnetic enrichment method selective for Vibrio parahaemolyticus serotype K and its application to food poisoning study; Tomoyasu T; A method using immunomagnetic separation was developed to isolate the specified K serotype of Vibrio parahaemolyticus from a mixture of a large number of bacteria with other K serotypes . This method was applied to food poisoning studies and could recover the V . parahaemolyticus serotype found in the patient from the incriminated foods. Appl Environ Microbiol, 1992 Aug, 58(8), 2485 - 9 Reassessment of the prevalence of heat-stable enterotoxin (NAG-ST) among environmental Vibrio cholerae non-O1 strains isolated from Calcutta, India, by using a NAG-ST DNA probe; Pal A et al.; A collection of 521 environmental isolates of Vibrio cholerae which were previously examined by the suckling mouse assay and found to be negative for the heat-stable enterotoxin NAG-ST were reassessed by a recently developed DNA probe for NAG-ST . A total of 12 (2.3%) of the isolates hybridized with the NAG-ST probe . By using a cholera toxin (CT) DNA probe, the CT gene was detected in six of the strains in the collection, although none of the isolates of V . cholerae non-O1 hybridized with both of the toxin probes . All of the NAG-ST and CT probe-positive strains were hemolysin positive . Thirty-fold-concentrated supernatants of the three representative NAG-ST DNA probe-positive V . cholerae non-O1 strains gave positive fluid accumulation ratios in the suckling mouse assay even after heating (100 degrees C for 5 min) and also inhibited the binding of a NAG-ST monoclonal antibody to the bound NAG-ST in a competitive enzyme-linked immunosorbent assay (ELISA) . Likewise, all six CT probe-positive V . cholerae non-O1 strains produced in vitro CT when examined by the CT bead ELISA . HindIII digest patterns of chromosomal DNA from the representative NAG-ST gene-positive strains were visually indistinguishable . Between the groups of NAG-ST probe-positive strains examined, there was a variation in the hybridizable fragments, with one group of strains exhibiting a hybridizable fragment similar to that of the NRT 36 reference strain; a smaller HindIII fragment hybridized with the NAG-ST probe in the other group of strains.(ABSTRACT TRUNCATED AT 250 WORDS) Appl Environ Microbiol, 1992 Aug, 58(8), 2474 - 8 Two types of bacterial alginate lyases; Kitamikado M et al.; The extracellular alginate lyases were purified from Vibrio harveyi AL-128 and V . alginolyticus ATCC 17749 . The former enzyme appears to be specific for alpha-1,4 bonds involving L-guluronate units in alginate, whereas the latter exhibits specificity for beta-1,4 bonds involving D-mannuronate units . The molecular weights of the enzymes were estimated to be 57,000 and 47,000, and they had isoelectric points of 4.3 and 4.6, respectively . The enzyme from strain AL-128 was most active at NaCl concentrations of 0.3 to 1.0 M . Optimum activity of the enzyme from strain ATCC 17749 was found in the presence of 5 to 10 mM CaCl2. Appl Environ Microbiol, 1992 Aug, 58(8), 2449 - 57 Sequence variation in the thermostable direct hemolysin-related hemolysin (trh) gene of Vibrio parahaemolyticus; Kishishita M et al.; Our previous molecular epidemiologic study with gene probes (H . Shirai, H . Ito, T . Hirayama, Y . Nakamoto, N . Nakabayashi, K . Kumagai, Y . Takeda, and M . Nishibuchi, Infect . Immun . 58:3568-3573, 1990) demonstrated that the gene (trh) encoding a thermostable direct hemolysin-related hemolysin was strongly associated with clinical strains of Vibrio parahaemolyticus . Strain-to-strain variation in the intensities of the hybridization signals observed in the above study also suggested that the trh genes in different strains may have significantly divergent nucleotide sequences . To assess the public health significance of the rare environmental strains which exhibited very weak hybridization signals with the trh gene-specific DNA probe, the trh-like sequence was cloned from one of the environmental strains and the nucleotide sequence was determined in this study . A hemolysin gene (trh2) which was 84% homologous to the trh gene (newly named trh1) and 54.8 to 68.8% homologous to the genes (tdh) encoding thermostable direct hemolysins was detected in the cloned sequence . The trh2 gene product showed a profile of hemolytic activities against various animal erythrocytes different from that of the trh1 gene product . The trh2 gene product was antigenically related (partially identical) to the trh1 and tdh gene products . DNA colony blot and Southern blot hybridization analyses with trh1- and trh2-specific DNA probes showed that the trh1 probe-positive strains exhibiting hybridization signals with varying intensities could be clustered into trh1 and trh2 subgroups . In addition, hybridization analysis with oligonucleotide probes demonstrated significant strain-to-strain variation in the trh1 and trh2 gene sequences.(ABSTRACT TRUNCATED AT 250 WORDS) Anal Biochem, 1992 Aug 1, 204(2), 315 - 23 Use of bacterial and firefly luciferases as reporter genes in DEAE-dextran-mediated transfection of mammalian cells; Pazzagli M et al.; The aim of this study was to compare three different luciferase genes by placing them in a single reporter vector and expressing them in the same mammalian cell type . The luciferase genes investigated were the luc genes from the fireflies Photinus pyralis (PP) and Luciola mingrelica (LM) and the lux AB5 gene, a translational fusion of the two subunits of the bacterial luciferase from Vibrio harveyi (VH) . The chloramphenicol acetyltransferase (CAT) gene was also included in this study for comparison . The performances of the assay methods of the corresponding enzymes were evaluated using reference materials and the results of the expressed enzymes following transfection were calculated using calibration curves . All of the bioluminescent assays possess high reproducibility both within and between the batches (less than 15%) . The comparison of the assay methods shows that firefly luciferases have the highest detection sensitivity (0.05 and 0.08 amol for PP and LM, respectively) whereas the VH bacterial luciferase has 5 amol and CAT 100 amol . On the other hand, the transfection of the various plasmids shows that the content of the expressed enzyme within the cells is much higher for CAT than for the other luciferase genes . VH luciferase is expressed at very low levels in mammalian cells due to the relatively high temperature of growing of the mammalian cells that seems to impair the correct folding of the active enzyme . PP and LM luciferases are both expressed at picomolar level but usually 10 to 70 times less in content with respect to CAT within the transfected cells . On the basis of these results the overall improvement in sensitivity related to the use of firefly luciferases as reporter genes in mammalian cells is about 30 to 50 times with respect to that of CAT. Antimicrob Agents Chemother, 1992 Aug, 36(8), 1738 - 43 Temperature-dependent in vitro antimicrobial activity of four 4-quinolones and oxytetracycline against bacteria pathogenic to fish; Martinsen B et al.; The in vitro antimicrobial activities of oxolinic acid, flumequine, sarafloxacin, enrofloxacin, and oxytetracycline against strains of bacteria pathogenic to fish (Aeromonas salmonicida subsp . salmonicida, atypical A . salmonicida, Vibrio salmonicida, Vibrio anguillarum, and Yersinia ruckeri) were determined at two different incubation temperatures, 4 and 15 degrees C, by a drug microdilution method . The main objective of the study was to examine the effect of incubation temperature on the in vitro activities of 4-quinolones and oxytetracycline against these bacteria . When tested against A . salmonicida subsp . salmonicida, all of the quinolones examined had MICs two- to threefold higher at 4 degrees C than at 15 degrees C . Similarly, 1.5- to 2-fold higher MICs were recorded for all of the quinolones except sarafloxacin at 4 degrees C than at 15 degrees C when the drugs were tested against V . salmonicida . In contrast to those of the quinolones, the MICs of oxytetracycline were two- to eightfold lower at 4 degrees C than at 15 degrees C against all of the bacterial species tested . Of the antimicrobial agents tested against the bacterial species included in the study, enrofloxacin was the most active and oxytetracycline was the least active . Sarafloxacin was slightly more active than flumequine and oxolinic acid, especially against oxolinic acid-resistant A . salmonicida subsp . salmonicida strains. Mol Microbiol, 1992 Aug, 6(16), 2407 - 18 Characterization of a Vibrio cholerae virulence factor homologous to the family of TonB-dependent proteins; Goldberg MB et al.; IrgA is an iron-regulated virulence factor for infection in an animal model with classical Vibrio cholerae strain 0395 . We detected gene sequences hybridizing to irgA at high stringency in clinical isolates in addition to 0395, including another classical strain of V . cholerae, three V . cholerae strains of the El Tor biotype, three non-O1 isolates of V . cholerae, and individual isolates of Vibrio parahaemolyticus, Vibrio fluvialis, and Vibrio alginolyticus . No hybridization to irgA was seen with chromosomal DNA from Vibrio vulnificus or Aeromonas hydrophila . To verify that irgA is the structural gene for the major iron-regulated outer membrane protein of V . cholerae, we determined the amino-terminal sequence of this protein recovered after gel electrophoresis and demonstrated that it corresponds to the amino acid sequence of IrgA deduced from the nucleotide sequence . Gel electrophoresis showed that two El Tor strains of V . cholerae had a major iron-regulated outer membrane protein identical in size and appearance to IrgA in strain 0395, consistent with the findings of DNA hybridization . We have previously suggested that IrgA might be the outer membrane receptor for the V . cholerae siderophore, vibriobactin . Biological data presented here, however, show that a mutation in irgA had no effect on the transport of vibriobactin and produced no defect in the utilization of iron from ferrichrome, ferric citrate, haemin or haemoglobin . The complete deduced amino acid sequence of IrgA demonstrated homology to the entire class of Escherichia coli TonB-dependent proteins, particularly Cir . Unlike the situation with Cir, however, we were unable to demonstrate a role for IrgA as a receptor for catechol-substituted cephalosporins . The role of IrgA in the pathogenesis of V . cholerae infection, its function as an outer membrane receptor, and its potential interaction with a TonB-like protein in V . cholerae remain to be determined. East Afr Med J, 1992 Aug, 69(8), 442 - 4 The fimbriae-like structures on V . cholerae isolated in Kenya; Kibue MA et al.; For Vibrio cholerae 01 to overcome the normal intestinal clearing mechanisms and facilitate colonization in the host gut, fine filamentous structures covering the surface of all the cell must exist . These fimbriae-like structures were investigated in this study . Selection of the K23 V . cholerae 01 biotype el-tor isolated in Kisumu, Kenya, was based on its proteinase activity and ability to agglutinate 0.2M ammonium sulphate . This strain was cultured in modified tryptone broth pH 8.4, at 37 degrees C overnight . Electron microscopy of the strain revealed the presence of fimbriae-like structures which varied in number . Some cells showed more than 200, whereas in other cells only several filamentous structures were detected . It is suggested that these structures could be a possible candidate for a cholera vaccine. J Bacteriol, 1992 Aug, 174(15), 5132 - 5 Evidence that GroEL, not sigma 32, is involved in transcriptional regulation of the Vibrio fischeri luminescence genes in Escherichia coli; Dolan KM et al.; In Escherichia coli, transcription of the inducible Vibrio fischeri luminescence operon, luxICDABE, has been reported to require sigma 32, the product of rpoH . Consistent with previous studies, we report that an E . coli delta rpoH mutant, KY1601 containing luxICDABE and luxR, which codes for the activator of luxICDABE transcription on a plasmid (pJE202), was weakly luminescent . Transformation of this E . coli strain with a plasmid containing rpoH under the control of the tac promoter resulted in high levels of cellular luminescence . However, the level of expression of the pJE202 luxICDABE was also high in E . coli 1603, a delta rpoH mutant with a second-site mutation that resulted in sigma 32-independent overexpression of the groE operon . Apparently, sigma 32 is not directly requi