Bull World Health Organ, 1994, 72(5), 745 - 9 Cholera in metropolitan Manila: foodborne transmission via street vendors; Lim-Quizon MC et al.; Reported are the results of an unmatched case-control study to determine the risk factors associated with acquisition of cholera in Manila . Cases were patients admitted to the San Lazaro Hospital between July and September 1989 and whose stools yielded Vibrio cholerae O1 on culture . Controls were patients admitted to the same hospital and who had no history of diarrhoea or of having taken antibiotics during the 3 days prior to admission . Of the 158 cases and 158 controls who had bought food from street vendors, cases were more likely to have bought the following items: pansit (rice noodles with shrimp, meat, and vegetables), mussel soup, spaghetti, fish balls, pig blood coagulated with vinegar, and salty brine shrimp with vegetables . Cases were also more likely to lack piped water at home . An unconditional logistic regression analysis indicated that only pansit (OR = 2.15, 95% CI = 1.32-3.51), mussel soup (OR = 2.29, 95% CI = 1.06-4.95), and the absence of piped water at home (OR = 2.70, 95% CI = 1.63-4.46) remained as risk factors . As control measures we recommend stricter implementation of the food sanitation code and the licensing of street food vendors. Trop Geogr Med, 1994, 46(3), 147 - 50 Emergence of a new epidemic strain of Vibrio cholerae in Bangladesh . An epidemiological study; Siddique AK et al.; For decades, epidemic cholera in Bangladesh has produced contrasting pictures of appearance and disappearance of Vibrio cholerae, which until recently, remained confined to the biotypes and to serotypes of V . cholerae O1 . The classical biotype continued to survive and coexisted with El Tor biotype in southern Bangladesh despite its disappearance from the rest of the world during the present pandemic . For the first time in history, during the cholera epidemic in 1993, both biotypes (classical and El Tor) of V . cholerae O1 have disappeared and have been replaced by a new strain of V . cholerae non-O1 (designated as O139 Bengal) . Environmental changes occurring in the Bay of Bengal may have resulted in the emergence of the new epidemic strain of V . cholerae in Bangladesh. Rev Environ Contam Toxicol, 1994, 138, 1 - 20 Antimicrobials in shrimp aquaculture in the United States: regulatory status and safety concerns; Park ED et al.; The consumption of seafood, especially shrimp, increases yearly in the U.S . The U.S . is the second largest importer of shrimp in the world, consuming more than 11% of the total world production . Aquaculture is becoming an increasingly important source of the world's shrimp, currently accounting for approximately 30% of the world's supply . Unfortunately, in this era of international trade deficits, U.S . production of aquacultured shrimp is insignificant (< 0.1%) compared with world production . As shrimp aquaculture expands in the U.S., so does the use of intensive farming techniques . Shrimp aquaculture is like any other animal husbandry industry in that shrimp are subject to disease, especially under intensive farming methods . In penaeid shrimp, the primary diseases associated with mortalities are usually viral or bacterial . The majority of bacterial infections in penaeid shrimp are attributable to Vibrio species, with mortalities ranging from insignificant to 100% . However, the rapid growth of this industry has outpaced efforts by researchers, pharmaceutical companies, and federal regulatory agencies to provide approved therapeutants for shrimp disease management . Approval of drugs and their surveillance for compliance with regulations applicable to seafoods, including aquacultured goods, is the responsibility of the FDA . There are three general areas of concern regarding human health when chemotherapeutants are used in aquaculture: (1) residues of drugs in fish destined for human consumption; (2) development of drug resistance in human pathogenic bacteria; and (3) direct toxic effects to humans from handling of drugs . Currently, there are no antibacterials approved for shrimp aquaculture in the U.S . One of the major obstacles in the development and approval of new drugs for aquaculture is the cost of conducting the required studies . The high cost to pharmaceutical companies discourages investment in shrimp chemotherapeutant research, since the current U.S . market for such products is small . Unfortunately, the U.S . shrimp aquaculture industry will remain small without legal availability of chemotherapeutants . Oxytetracycline (OTC) and Romet-30 are two antibacterials currently approved in the U.S . for catfish and salmonid aquaculture . Shrimp aquaculture facilities outside of the U.S . routinely use these drugs, as well as others, in the treatment of bacterial disease outbreaks . Much of the work required for OTC approval by the FDA for penaeid shrimp has been completed.(ABSTRACT TRUNCATED AT 400 WORDS) Microbiol Immunol, 1994, 38(5), 367 - 71 Regulation of iron on bacterial growth and production of thermostable direct hemolysin by Vibrio parahaemolyticus in intraperitoneal infected mice; Wong HC et al.; Pathogenesis of Vibrio parahaemolyticus is not clearly understood . Effects of iron on the bacterial proliferation and production of thermostable direct hemolysin (TDH) in intraperitoneal infected mice were studied . Injection of bacterial culture in the presence of ferric ammonium citrate (100 micrograms/ml) significantly enhanced the lethality for mice, and simultaneously activated bacterial proliferation in vivo . The iron-limited cultures showed better proliferation than those iron-rich cultures in response to the addition of supplementary iron source . Production of TDH by the hemolytic strains ST550 and D62 was higher in the iron-limited cultures than the iron-rich cultures . Production of TDH by both the iron-limited or iron-rich cultures was inhibited by the addition of iron . In conclusion, the virulence enhancement effect of iron in V . parahaemolyticus was probably by activating bacterial proliferation in vivo and not by stimulating the production of TDH . V . parahaemolyticus precultured in iron-limited condition may be more adaptable to in vivo environment. Microbiol Immunol, 1994, 38(4), 301 - 4 Successful application of enzyme-labeled oligonucleotide probe for rapid and accurate cholera diagnosis in a clinical laboratory; Miyagi K et al.; Two cholera cases were diagnosed using an enzyme-labeled oligonucleotide probe (ELONP) hybridization test for detection of cholera toxin gene (ctx) in a clinical laboratory at Osaka Airport Quarantine Station . The ELONP test with suspicious colonies of Vibrio cholerae O1 grown on TCBS or Vibrio agar plates gave positive result for ctx within 3 hr . We also tried to apply the ELONP test for direct detection of ctx in their stool and their non-selective culture . Specimens from Case #1, which contained 5.9 x 10(5) CFU/g of V . cholerae O1 in the stool, cultured for 7-8 hr or longer in alkaline peptone water or Marine broth at 37C, became positive for ctx . On the other hand, specimens from Case #2, which contained 8.7 x 10(8) CFU/ml (of V . cholerae O1 in the stool), gave positive result in this stool itself without any further culture . These data suggest that the ELONP test provides successfully a more rapid and accurate means of identifying "toxigenic" V . cholerae O1 in a clinical laboratory. Microbiol Immunol, 1994, 38(3), 225 - 7 Pili of a Vibrio cholerae O139; Nakasone N et al.; The pili of a strain of Vibrio cholerae O139 were purified and characterized . They were morphologically, electrophoretically and immunologically indistinguishable from the pili with 16 kDa subunit protein of V . cholerae O1 . All 22 strains of V . cholerae O139 examined possessed the pili . The pili were different in hemagglutination inhibition pattern from V . cholerae O1 16K pili. Infect Immun, 1994 Jan, 62(1), 72 - 8 ToxR regulates virulence gene expression in non-O1 strains of Vibrio cholerae that cause epidemic cholera; Waldor MK et al.; Vibrio cholerae serogroup O1 has historically been thought to be the exclusive cause of epidemic cholera . O139 is a novel serogroup of V . cholerae which emerged on the Indian subcontinent in the last few months of 1992 and is the first non-O1 serogroup of V . cholerae to cause epidemic cholera . We have investigated the expression of some of the known virulence factors of classical and El Tor O1 strains of V . cholerae in clinical isolates of O139 strains . We show that, in contrast to other non-O1 strains, O139 strains express TcpA, the major subunit of the toxin-coregulated pilus found in O1 strains . As in O1 strains, the expression of cholera toxin and TcpA is coordinately regulated by environmental parameters in O139 strains . Derivatives of O139 strains that contain a toxR null mutation were constructed and used to demonstrate that the expression of cholera toxin, TcpA, and the outer membrane protein OmpU in O139 strains, as in O1 strains, is dependent on ToxR . Two kinds of evidence suggest that O139 strains are closely related to El Tor strains of V . cholerae . First, both O139 and El Tor strains share a restriction fragment length polymorphism for tcpA, which distinguishes El Tor from classical strains of V . cholerae . Second, cholera toxin production in O139 strains is greatly enhanced by culture conditions that have been previously shown to promote production of cholera toxin in El Tor strains and not in classical strains of V . cholerae . Although O139 is a novel serotype of V . cholerae, O139 strains conform to a fundamental theme that has evolved from the study of O1 strains: ToxR mediates coordinate regulation of virulence gene expression. Microbiol Immunol, 1994, 38(11), 837 - 42 Thermal stability of an oral killed-cholera-whole-cell vaccine containing recombinant B-subunit of cholera toxin; Ahmed ZU et al.; An oral killed cholera vaccine containing 1 x 10(11) cells of Vibrio cholerae O1 (heat- or formalin-killed) representing the Ogawa and Inaba biotypes and containing 1 mg of B-subunit of cholera toxin (CTB) produced by recombinant DNA technology (the WC/rCTB vaccine) was subjected to temperatures of 4 C, 30 C or 42 C for up to 6 months time . Lipopolysaccharide antigen (LPS) and CTB content of the vaccine samples determined at various times remained unchanged during the study except for the CTB component which decreased by about 50% after 6 months of storage at 42 C . Immunogenicity determined by immunization of rabbits with the vaccine in Freund's complete adjuvant and measuring anti-LPS and anti-CTB antibody titers in the serum by an ELISA was also found to be unaltered . Lyophilization of the vaccine and storage at room temperature for 7 days also did not have any adverse effect on antigen content or immunogenicity as tested above . There was up to one log reduction in serum antibody titers after immunization without using any adjuvant or using Freund's incomplete adjuvant, and up to two logs following oral immunization . Immunization by oral feeding of the vaccine followed by RITARD challenge with a virulent V . cholerae O1 strain showed evidence of protection against severe or lethal diarrhea . The results suggest that the vaccine retains its antigen content and ability to induce antibodies unchanged when maintained at elevated temperatures for relatively long periods of time. DNA Seq, 1994, 5(1), 51 - 5 The accessory colonization factor and toxin-coregulated pilus gene clusters are physically linked on the Vibrio cholerae 0395 chromosome; Everiss KD et al.; A partial nucleotide sequence of the first gene in the Vibrio cholerae accessory colonization factor gene cluster was obtained by sequencing a cloned acfB::TnphoA fusion junction . Analysis of the sequence data revealed that the ATG start codon of the acfB structural gene is located 6 bp downstream of the tcpJ TGA stop codon . This establishes a large region in excess of 20 kbp of the Vibrio cholerae chromosome as a cluster of coordinately regulated virulence genes involved in intestinal colonization . The absence of a characteristic transcription termination site in the tcpJ-acfB intergenic region indicates that acfB may be encoded on the same messenger RNA as tcpJ . The deduced amino acid sequence for the N-terminal 143 amino acids of AcfB reveals a potential leader peptide, that lacks a consensus signal peptidase I recognition sequence . Computer algorithms fail to detect any significant similarities between the partial AcfB amino acid sequence and any entries in the SWISS-PROT database. Acta Microbiol Immunol Hung, 1994, 41(4), 415 - 21 Penaeid prawns and associated luminous bacteria; Jayabalan N et al.; Luminous bacteria associated with the exoskeleton, gill and gut of penaeid prawns, Penaeus indicus H . Milne Edwards and Penaeus monodon Fabricius in Mangalore waters were isolated, identified and quantified . Two species of bacteria, Vibrio harveyi and Vibrio fischeri were recorded, the former was dominant in the exoskeleton and gill of both the penaeids . Gut of prawns supported exclusive and dense populations of V . harveyi suggesting that the species is well adapted to proliferate in this microenvironment especially in the hindgut region. Acta Microbiol Immunol Hung, 1994, 41(4), 403 - 9 Characterization of turbot (Scophthalmus maximus) associated bacteria with inhibitory effects against the fish pathogen Vibrio anguillarum; Westerdahl A et al.; More than 400 isolates from the intestine and the external surface of farmed turbot, as well as from fish food and hatchery water were screened for inhibitory effects against the fish pathogen Vibrio anguillarum HI 11345 and seven other fish pathogens . The bacteria with inhibitory effects were then characterized with regards to their sites of colonization, especially the intestinal regions and sites within each region . No correlations between the different biochemical phena, site of colonization and inhibitory effect could be found . The potential of seven of these strains for adhesion to intestinal mucus from turbot was studied . Rapid growth of V . anguillarum in intestinal mucus was measured, hence it is feasible that the intestinal tract is a site for V . anguillarum multiplication . Strains isolated from the intestine showed greater capacity for in vitro adhesion to and growth in fish intestinal mucus than did the pathogen and skin mucus isolates . Two of the strains isolated from the intestine were studied for their inhibitory kinetics and one strain for the potential of in vivo colonization . The molecular weight of the inhibitory component was below 1000 dalton in 65% of the strains isolated . For the other 35% the inhibitory component ranged between 1000 D-6000 D in dialysis cut off experiments . One week after oral administration, one such isolate still accounted for 45% of the total c.f.u . in the intestinal mucus of 5 g turbot . In preliminary experiments we demonstrated that it is possible to detect the pathogen in intestinal mucus with a fluorescently labelled oligonucleotide probe directed against a specific part of the intracellular rRNA of V . anguillarum. Microbiol Immunol, 1994, 38(9), 687 - 93 Siderophore-mediated utilization of iron bound to transferrin by Vibrio parahaemolyticus; Yamamoto S et al.; Vibrio parahaemolyticus produces a structurally novel type of siderophore, termed vibrioferrin, in response to iron-limitation . This study was performed to examine whether vibrioferrin can assimilate iron from human iron-binding proteins for growth . Comparison of the growth rates between V . parahaemolyticus AQ 3354 and its spontaneously arising, vibrioferrin-deficient mutant revealed that vibrioferrin was able to sequester iron from 30% iron-saturated human transferrin for growth, but not from human lactoferrin even if fully saturated with iron . In both strains, iron limitation induced two high-molecular-weight outer membrane proteins with apparent molecular masses of approximately 78 and 83 kDa . Since only the outer membrane fraction including these proteins showed a binding capacity to ferric vibrioferrin complex, either of them may function as its cell surface receptor . These results suggested that the organism might utilize such a source of host iron through the action of vibrioferrin during in vivo survival and proliferation, although its importance in pathogenesis is unknown. Bull Soc Pathol Exot, 1994, 87(3), 143 - 7 {Seroepidemiological survey of El Tor cholera in an endemic region of Algeria}; Bendib A et al.; Between the 26th of April and the 8th of july 1987 a seroepidemiological survey was conducted in the district of Chlef located in an endemic cholera area of Algeria . The purpose of this work was to evaluate the seroprevalence of vibriocidal antibodies and to study some epidemiological characteristics of cholera . This survey concerns the representative samples of patients and asymptomatic carriers such as they were declared during the epidemic periods of 1982 and 1986 and a sample of contacts of these latter . A bacteriology of stools practised on all the subjects did not reveal a carrying of vibrios . The seroprevalence was 37.6% and it concerned all ages . It was linked meaningfully to the age category and not to the sex . This seroprevalence was not significantly different between the cases, the carriers and the contacts . Thus this study reveals that Algeria has a seroprevalence rate less important than others countries. Chin J Biotechnol, 1994, 10(4), 225 - 32 Construction of an engineered bivalent vaccine strain consisting of Vibrio cholerae CT-B and LPS-O antigens; Yu X et al.; In this study, the engineered E . coli strain 1046 containing V . cholerae LPS-O and CTB bivalent antigen genes has been successfully obtained by using DNA recombinant techniques . E . coli 1046 (pMG305) could not only express CT-B antigen but also secret CTB into medium as shown by GM1-ELISA . Meanwhile, whole cell O-antigen-ELISA, bacterial agglutination test and hemagglutination inhibition assay demonstrated that LPS-O antigen could be expressed on the cell surface by E . coli 1046 (pMG305) and shown LPS band specific for V . cholerae by SDS-PAGE assay . Mouse intraperitoneal immunization and challenge trial indicated that the E . coli 1046 (pMG305) provided good protection against virulent V . cholerae . The engineered vaccine strain reported here is expected to be a live oral candidate vaccine for V . cholerae. Rev Sanid Hig Publica (Madr), 1994 Jan-Feb, 68(1), 187 - 96 {Environmental isolation of Vibrio cholerae 01 in continental waters of the Province of Seville}; Jimenez Madrazo C et al.; BACKGROUND: Many bibliographical references suggest reserves of Vibrio cholerae in the aquatic world, strains of both the serovariety no 01 and 01 having been isolated in different parts of the globe in fresh and extensive salt waters . METHODS: Samples were taken monthly from January to May and from October to December, and every fortnight from June to September . The method of isolation applied is in accordance with the standard set down by the Institute of Health Carlos III, with some specific modifications . RESULTS: During the period 90/91, routine analyses carried out by the Andalusian Service of Health of waters from the area of the Continental Basin in the province of Seville showed the presence of Vibrio cholerae 01 . In the same way, during the complete annual cycle Vibrio cholerae no 01 was also isolated . CONCLUSIONS: These isolations did not coincide with outbreaks or individual cases of cholera, so the authors, in agreement with what is described in the bibliography, consider that Vibrio cholerae no 01 forms a part of certain aquatic environments, which is in direct relation to environmental isolations of the serovariety 01. Antonie Van Leeuwenhoek, 1994, 66(4), 295 - 302 Isolation and study of two strains of Leptospirillum-like bacteria from a natural mixed population cultured on a cobaltiferous pyrite substrate; Battaglia F et al.; Two strains of Leptospirillum-like bacteria, L6 and L8, have been isolated from a mixed inoculum, also containing Thiobacillus ferrooxidans and T . thiooxidans, cultured for one year with a colbaltiferous pyrite as energy substrate in a 100 l continuous bioleaching laboratory unit . Several physiological properties of the strains are described . The vibrio-shaped microorganisms grew at pH values lower than 1.3 . Their growth rate was maximum between 2.5 and 8.0 g l1 ferrous iron . The optimal growth temperature was 37.5 degrees C . Ferric iron had a stimulative effect on bacterial development up to 8 g l-1, and growth was as rapid at 14 g l-1 ferric iron as at 8 g l-1 . The negative influence of cobalt on the final cell concentration was observed at 0.5 g l-1, but the growth rate was not affected up to 2 g l-1 . The G + C content of strains L8 is 55.6 mol%. Acta Vet Scand, 1994, 35(4), 355 - 62 Characterization of Vibrio anguillarum strains isolated from diseased fish in Finland; Tiainen T et al.; Characterization of V . anguillarum strains (n = 109) isolated from diseased salmonids was performed . Eight O serovars were found among the strains . Serovar O1 was predominant (90%), while serovars O2, O3, O5, O8, O9, and a new serovar Va NT2, were represented by 1 or 2 strains . Two strains remained non-typeable . One of these was cross-reactive with several antisera, but had a LPS profile identical to that of serovar O8 . All serovars showed specific LPS profiles . All but 1 of the O1 strains had a plasmid comparable in size to the pJM1 virulence plasmid, while plasmids of different sizes were found in O2, Va NT2 and the non-typeable strains . Apart from a single strain resistant to tetracycline, all the strains were sensitive to oxolinic acid, tetracycline, and trimethoprim-sulfonamides . By their biochemical and antigenic properties strains causing vibriosis among salmonids in Finland closely resemble Scandinavian strains . Predominance of the serovars O1 and O2 suggests that commercial vaccines containing these serovars should afford sufficient protection against vibriosis in Finland. Med Dosw Mikrobiol, 1994, 46(4), 313 - 21 {Vibrio cholerae strains resistant to vibriostatic factor 0/129 isolated from a patient with cholera in Poland}; Stypulkowska-Misiurewicz H et al.; The Vibrio cholerae strain of biotype El Tor and serotype Ol Ogawa was isolated from a 45-year old man after his return from India, hospitalized in Glogow (WSSE Legnica) with symptoms of food poisoning and considerable dehydration . From culture on alkaline agar, the Vibrio cholerae strain was isolated and manifested typical biochemical and serological properties except for resistance to the vibriostatic factor 0/129 (2,4-diamino-6,7-diisopropyl-pteridine) . In addition, the strain was resistant to sulfamethoxazole, trimethoprim, cotrimoxazole and colistine, and susceptible to ampicillin, chloramphenicol, tetracycline, furazolidone and nalidixic acid . In Poland this is the first case isolation of Vibrio cholerae resistant to the vibriostatic factor . Most striking is the fact that Vibrio cholerae resistant to 0/129 is spreading geographically and it isolation is more and more frequent. Microbios, 1994, 77(311), 101 - 9 The 16S rRNA sequence and genome sizing of tributyltin resistant marine bacterium, strain M-1; Suzuki S et al.; The 16S rRNA of the tributyltin resistant marine bacterium, strain M-1, was partly sequenced to confirm the taxonomic status . The results indicated that this bacterium should be classified under the genus Alteromonas, instead of a previous report in which this strain was identified as a Vibrio . The genome size of this strain was also measured by pulsed field gel electrophoresis (PFGE) using a contoureclamped homogeneous electric field . The strain was found to contain a genome size of 2,240 kilo base pairs, whereas Alteromonas nigrifaciens and Shewanella putrefaciens had 2,040 and 2,383 kilo base pairs, respectively . This is the first report of the genome sizing of the genus Alteromonas by PFGE. J Clin Microbiol, 1994 Jan, 32(1), 235 - 7 Modified CAMP test for biogrouping Vibrio cholerae O1 strains and distinguishing them from strains of V . cholerae non-O1; Lesmana M et al.; A modified CAMP test was used to identify 973 Vibrio cholerae isolates by phenotype . Eltor and non-O1 strains were CAMP positive; classical strains were CAMP negative . Sausage-shaped zones of hemolysis of eltor strains were easily distinguished from narrower bands of non-O1 isolates . For O1 isolates, there was 100% agreement between the CAMP test and inhibition by polymyxin B. Clin Diagn Lab Immunol, 1994 Jan, 1(1), 51 - 4 Production, characterization, and application of monoclonal antibodies to Vibrio cholerae O139 synonym Bengal; Qadri F et al.; Mouse monoclonal antibodies (MAbs) were derived against acetone-treated whole cells of the newly recognized Vibrio cholerae O139 serogroup which is causing epidemics of cholera-like disease in India and Bangladesh . Four MAbs specifically recognized the lipopolysaccharide antigens of V . cholerae O139 . MAbs ICL9 and ICL13 were of the immunoglobulin M (IgM) isotype, ICL11 was of the IgG3 isotype, and ICL12 was of the Ig2b isotype . A fifth MAb, ICL10, of the IgG2b isotype cross-reacted with V . cholerae O91 . All five MAbs recognized V . cholerae O139 in an enzyme-linked immunosorbent assay, slide agglutination test, motility inhibition test, and indirect immunofluorescence test . During a 1-month evaluation of these MAbs in our clinical laboratory, all 86 cases diagnosed as V . cholerae O139 by a rabbit polyclonal antiserum were also detected by these MAbs, establishing their utility as highly sensitive and specific diagnostic reagents . With these MAbs, it should now be possible to screen for the V . cholerae O139 serogroup in epidemic and endemic diarrhea cases and in environmental and food samples. Biochemistry, 1993 Dec 28, 32(51), 14183 - 6 Spectral detection of an intermediate preceding the excited state in the bacterial luciferase reaction; Macheroux P et al.; The bioluminescent reaction of luciferase isolated from Vibrio harveyi, strain M17, was initiated by mixing the luciferase-bound flavin 4a-hydroperoxide intermediate, purified in advance, with a long-chain aldehyde (dodecanal or octanal) at -4 degree C . Measurements of absorbance changes from 300 to 600 nm during the course of the reaction revealed the existence of three sets of isosbestic points and three kinetic phases, the second of which parallels kinetically the decay of bioluminescence, measured concurrently . The absorbance changes in this second step and the decay of light emission exhibited similar deuterium isotope effects; this is postulated to be the step giving rise to the excited state and the enzyme-bound flavin 4a-hydroxide . The first step of the reaction, however, did not show an isotope effect; the intermediate thereby formed, observed here for the first time, is postulated to correspond to the luciferase-bound flavin 4a-peroxyhemiacetal. Int J Cancer, 1993 Dec 2, 55(6), 1029 - 35 Differences in cell surface carbohydrates, and in laminin and fibronectin synthesis, between adherent and non-adherent Ehrlich ascites tumor cells; Song Z et al.; Differences in cell surface carbohydrates and in laminin and fibronectin synthesis between 2 Ehrlich ascites tumor (EAT) cell lines, the adherent and non-adherent EAT cells, have been studied . The adherent EAT (a-EAT) cells grow in monolayer in vitro in the presence of fetal bovine serum . The classical, or non-adherent EAT (na-EAT), cells grow in suspension in ascites form in the peritoneal cavity of mice, and they do not adhere when cultured in vitro . Both EAT cell lines express surface glycoproteins reactive with Maackia amurensis lectin (MAL) and Griffonia simplicifolia isolectin GS I-B4 . However, only a-EAT cells react with elderberry (Sambucus nigra) bark lectin (SNA), suggesting that there are some differences in the sialylation of cell surface carbohydrate moieties between these 2 EAT cell lines . Removal of cell surface sialic acid by treating a-EAT cells with Vibrio cholerae neuraminidase did not abolish the ability of these cells to adhere to laminin- or fibronectin-coated plates, indicating that the sialic acid of the cell surface glycoproteins is not essential for their adhesion to these extracellular matrices . Therefore, the difference in sialylation of cell surface glycoproteins is not responsible for the difference in cell adhesion between these 2 lines of EAT cells . Both EAT cell lines express detectable amounts of laminin but not fibronectin on their surfaces; they both secrete fibronectin and entactin into the medium . The na-EAT cells (but not the a-EAT cells) also secrete laminin A chain into the culture medium; however, no B chain was detected in the culture medium of either cell line . The laminin isolated from the cell surface of na-EAT cells reacts with GS I and MAL lectins, but not with SNA, whereas the laminin isolated from a-EAT cells reacts with SNA, as well as GS I and MAL. J Gen Virol, 1993 Dec, 74 ( Pt 12), 2749 - 52 Vibriophage D10 contains non-permuted DNA with cohesive ends; Chakrabarti BK et al.; Phage D10, a Vibrio cholerae O-1 El Tor group X phage, is one of the five newly isolated phages used in the phage typing scheme developed for V . cholerae O-1 biotype El Tor and belongs to the Myoviridae family . From electron microscopic studies it is shown that phage D10 has a DNA genome of 32 +/- 0.2 kb . This is the first report where it has been shown by the construction of a partial denaturation map that this vibriophage genome is nonpermuted and has cohesive ends . The location of the ends of the DNA in the phage head has also been inferred. Eur J Biochem, 1993 Dec 1, 218(2), 543 - 54 The structure of the carbohydrate backbone of the core-lipid-A region of the lipopolysaccharide from Vibrio cholerae strain H11 (non-O1); Vinogradov EV et al.; Lipopolysaccharide from Vibrio cholerae strain H11 (non-O1) was de-O-acylated, dephosphorylated, reduced, de-N-acylated, N-acetylated, and the products were separated by high-performance anion-exchange chromatography (HPAE) . A decasaccharide, 1, was isolated as the major product, representing the core oligosaccharide attached to the reduced GlcN-disaccharide lipid A backbone . Its structure was established by compositional and methylation analyses, and extensive NMR investigations including 1H,1H correlation spectroscopy (COSY), total correlation spectroscopy (TOCSY), and nuclear Overhauser enhancement spectroscopy (NOESY), as well as heteronuclear 13C,1H COSY . In another reaction sequence the lipopolysaccharide was hydrolysed with dilute acetic acid and reduced with NaBH4 . The resulting core fractions were separated by HPAE giving seven individual octasaccharides differing at the reducing 3-deoxy-D-manno-octulosonic acid (Kdo) residue . A major product, 2, was isolated and investigated by the same methods as described for the decasaccharide 1 . The following structures are proposed for compounds 1 and 2: alpha-D-GlcNAcp-(1-7)-{beta-D-Galp-(1-3)-}-alpha-Hepp-(1-2)- alpha-Hepp- (1-3)-{beta-D-Glcp-(1-4)-}- {alpha-D-Glcp-(1-6)-}-alpha-Hepp-(1-5)-R, where R is alpha-Kdop-(2-6)-beta-D-GlcNAcp-(1-6)-D-GlcNAcol in 1 and 4,8-anhydro-Kdool in 2, and Hep is L-glycero-D-manno-heptose . In lipopolysaccharide, the terminal residue of alpha-D-glucosamine possessed a free amino group, as proved by deamination with nitrous acid and the 1H-NMR spectrum of de-O-acylated lipopolysaccharide . The conformational preferences of the terminal core heptasaccharide was assessed by Monte Carlo simulations combined with restrained calculations of side chains based on experimentally determined proton-coupling constants . These calculations, confirmed by NOE data, displayed several long-range interactions, which resulted in a well-defined three-dimensional structure of the core oligosaccharide. Am J Epidemiol, 1993 Dec 1, 138(11), 988 - 93 Immunologic response to oral cholera vaccination in a crossover study: a novel placebo effect; Wasserman SS et al.; The authors conducted a two-period crossover study of the reactogenicity and immunogenicity of live oral cholera vaccine CVD 103-HgR among US university students . Subjects ingested 5 x 10(8) colony forming units of either killed Escherichia coli K12 placebo or vaccine, followed by the opposite treatment one week later . Surprisingly, the dynamics of the immunologic response were influenced by prior ingestion of placebo . Subjects who received placebo first showed stronger vibriocidal antibody responses 2 weeks after vaccination compared with subjects who received vaccine first; this same pattern was seen for antitoxin titers . The authors suggest that ingestion of E . coli K12 one week prior to immunization boosts the immunologic response to vaccine by an unknown mechanism . Future crossover studies that examine immunologic outcomes might be designed to explore the ubiquity of such an effect. J Infect Dis, 1993 Dec, 168(6), 1536 - 40 Safety and immunogenicity of live oral cholera vaccine candidate CVD 110, a delta ctxA delta zot delta ace derivative of El Tor Ogawa Vibrio cholerae; Tacket CO et al.; The current pandemic of cholera is caused primarily by Vibrio cholerae O1 of the El Tor biotype . Live attenuated classical biotype V . cholerae vaccine strains prevent severe and moderate cholera due to either biotype in challenged volunteers but may provide less protection against mild cholera due to El Tor organisms . CVD 110, a new ctxA-deleted vaccine strain derived from an El Tor Ogawa parent, lacks zona occludens toxin (Zot), accessory cholera enterotoxin (Ace), and hemolysin/enterotoxin . Ten healthy adult volunteers were given 10(8) cfu of CVD 110 with buffer; 7 developed diarrhea (mean stool volume, 861 mL) . Vaccine organisms were shed in stool by all vaccines and were recovered from duodenal fluid in three-quarters of vaccinees . After vaccination, the geometric mean peak reciprocal vibriocidal titer among vaccinees was 17,829 . CVD 110 is a powerful immunogen but insufficiently attenuated despite the absence of known potential enterotoxins of V . cholerae . Another unrecognized toxin or colonization alone may be responsible for diarrhea after ingestion of this strain. Infect Immun, 1993 Dec, 61(12), 5279 - 85 Analysis of the roles of antilipopolysaccharide and anti-cholera toxin immunoglobulin A (IgA) antibodies in protection against Vibrio cholerae and cholera toxin by use of monoclonal IgA antibodies in vivo; Apter FM et al.; Secretory immunoglobulin A (IgA) antibodies (sIgA) directed against cholera toxin (CT) and surface components of Vibrio cholerae are associated with protection against cholera, but the relative importance of specific sIgAs in protection is unknown . A monoclonal IgA directed against the V . cholerae lipopolysaccharide (LPS), secreted into the intestines of neonatal mice bearing hybridoma tumors, was previously shown to provide protection against a lethal oral dose of 10(7) V . cholerae cells . We show here that a single oral dose of 5 to 50 micrograms of the monoclonal anti-LPS IgA, given within 2 h before V . cholerae challenge, protected neonatal mice against challenge . In contrast, an oral dose of 80 micrograms of monoclonal IgA directed against CT B subunit (CTB) failed to protect against V . cholerae challenge . A total of 80 micrograms of monoclonal anti-CTB IgA given orally protected neonatal mice from a lethal (5-micrograms) oral dose of CT . Secretion of the same anti-CTB IgA antibodies into the intestines of mice bearing IgA hybridoma backpack tumors, however, failed to protect against lethal oral doses of either CT (5 micrograms) or V . cholerae (10(7) cells) . Furthermore, monoclonal anti-CTB IgA, either delivered orally or secreted onto mucosal surfaces in mice bearing hybridoma tumors, did not significantly enhance protection over that provided by oral anti-LPS IgA alone . These results demonstrate that anti-LPS sIgA is much more effective than anti-CT IgA in prevention of V . cholerae-induced diarrheal disease. FEMS Immunol Med Microbiol, 1993 Dec, 7(4), 297 - 301 Analysis of some factors involved in the virulence mechanism of Vibrio cholerae non-01; Rodrigues DP et al.; A study was carried out to evaluate the potential intestinal toxicity of 188 samples of Vibrio cholerae non-01 isolated from seawater found along the beaches of Rio de Janeiro city . Three different assays were carried out involving: (a) detection of vascular permeability factor (PF) in guinea pigs (together with assessment of two culture media for production of the toxin); (b) intestinal fluid accumulation (FA) in suckling mice; and (c) detection of haemolysin . The results demonstrated that both culture media gave a similar level of performance . In the animal assays, 43% of the samples induced PF in guinea pigs, 28.7% caused intestinal fluid accumulation in suckling mice, and 63.28% contained haemolysin . Only 4.25% of the samples gave positive results in all three tests. J Gen Microbiol, 1993 Dec, 139 ( Pt 12), 3225 - 31 Rapid and specific detection of the thermostable direct haemolysin gene in Vibrio parahaemolyticus by the polymerase chain reaction; Lee C et al.; Synthetic oligonucleotide primers derived from a sequence of the thermostable direct haemolysin (tdh) gene were used in a polymerase chain reaction (PCR) amplification technique to detect this gene in strains of Vibrio parahaemolyticus . A total of 36 TDH-producing, and 89 TDH-negative Vibrio parahaemolyticus strains and 46 other vibrios and enteric pathogens were studied . In all, 36 strains of Vibrio parahaemolyticus from which the tdh gene could be successfully amplified by PCR were found to be TDH-positive in TDH haemolysin assay . No amplification products were obtained from Vibrio parahaemolyticus strains that were TDH-negative in the haemolysin assay or from other vibrios and enteric pathogens, with the exception of two strains . The PCR results were consistent with DNA hybridization tests . The detection limit for the tdh gene by PCR amplification was 40 pg of total DNA, or broth culture containing 1000 viable cells . Amplification products were confirmed by restriction enzyme digestion and Southern blot hybridization . The PCR method could detect the tdh sequences in stool samples from patients with gastroenteritis caused by V . parahaemolyticus . This PCR protocol clearly identified TDH-producing strains of V . parahaemolyticus and provides an alternative to conventional methods for TDH detection by research laboratories, clinical laboratories, regulatory agencies, and the seafood industry. Gig Sanit, 1993 Dec, (12), 14 - 7 {A study of the biological properties of cholera phages and strains of El-Tor vibrions isolated from the external environment in Donetsk}; Kudriakova TA et al.; Choleric phages were identified in 2 out of 44 samples of Donetsk sewage . Biological properties of phages were characterized. Indian J Exp Biol, 1993 Dec, 31(12), 955 - 62 Electron microscopic study of phages and aberrant structures produced by induction of prophage kappa in Vibrio cholerae el tor cells; Pal AK et al.; Prophage kappa in V . cholerae el tor strain SLH22(J) could be induced spontaneously or by treatment with nitrofurantoin, though the efficiency of induction was very low (not more than 0.8%) . V . cholerae el tor cells were found to release many different aberrant structures of the temperate phage, kappa . These aberrant structures were characterized by density gradient centrifugation and electron microscopy. Acta Paediatr Jpn, 1993 Dec, 35(6), 583 - 90 Cooperative studies on diarrheal diseases; Takeda T et al.; Enteric infection still causes the highest global morbidity and mortality in children under 5 years old . While there have been impressive developments in the early treatment of diarrhea, adequate diagnostic techniques are not yet available . An international collaboration was conducted with the National Institute of Cholera and Enteric Diseases, Calcutta on the rapid diagnosis and prevention of diarrheal diseases . Forty to fifty per cent of the 780-bed Infectious Diseases Hospital is occupied by cases of diarrhea . Vibrio cholerae O1 continues to occupy a prominent position, and the prevalence in children up to 2 years is 30.9% . Accordingly, immuno-enzymatic detection of cholera toxin in stool was achieved . Bead enzyme-linked immunosorbent assay (ELISA) could successfully detect 40 pg/mL toxin within 4 h, but the detection rate of culture, bead ELISA and polymerase chain reaction (PCR) method was 72.2, 71.1 and 95.9%, respectively, indicating that PCR provides the most sensitive and specific assay for diagnosis of cholera directly from the stool of patients . Heat-stable enterotoxin (STa) is another virulence factor of gastroenteritis . Various monoclonal antibodies were established against it, and developed a competitive ELISA for the detection of STa producing strains . A DNA probe was also prepared, and genus vibrio was monitored . V . mimicus were found to be the reservoirs of STa among clinical vibrios . Various monoclonal antibodies against STa were also useful for the epitope mapping, providing information on the topology of toxin-host interaction. Asian Pac J Allergy Immunol, 1993 Dec, 11(2), 155 - 65 Localization of Vibrio cholerae O1 in the intestinal tissue; Sincharoenkul R et al.; Colonization of V . cholerae O1 in vivo is known to be a non-invasive type which the vibrios are confined only to the intestinal tissues . The pathway by which the vibrio antigens reach the lymphoid cells and subsequently give rise to the immune responses is not entirely clear . Thus, experiments were performed in experimental rats by inoculating live V . cholerae O1 into the ligated ileal loops . The fate of the vibrios in the intestinal tissues was then studied by transmission electron microscopy at different times after the inoculation . It was concluded that live V . cholerae O1 were initially taken up by the M cells which overlay Peyer's patches and which subsequently delivered the intact vibrios to phagocytic cells in the Peyer's patches . These phagocytic cells processed (digested) the vibrios while the lymphocytes and plasma cells infiltrated around them . During the late period of infection (12-15 hours after inoculation of the vibrios), vibrios were also found passing through the loose intercellular spaces between the absorptive epithelial cells into the underlying intestinal tissues. Microb Pathog, 1993 Dec, 15(6), 421 - 31 The role of toxin-coregulated pili in the pathogenesis of Vibrio cholerae O1 El Tor; Attridge SR et al.; Studies in the infant mouse cholera model have evaluated the significance of toxin-coregulated pili (TCP) in the pathogenesis of Vibrio cholerae strains of El Tor biotype . Four El Tor strains--two which produce TCP during in vitro growth and two which do not--were mutated by the insertion of an antibiotic-resistance cartridge into the tcpA gene (encoding the pilin monomer) . The resulting mutants were otherwise indistinguishable from wild-type and in particular were unaltered in their sensitivity to antibody-dependent, complement-mediated bacteriolysis . All were dramatically attenuated and showed a marked impairment in terms of in vivo persistence in mixed-infection competition experiments . Virulence was restored by provision of a functional tcp operon in trans, confirming that the pathogenic potential of El Tor strains is critically dependent upon product(s) of this operon. Biotechnology (N Y), 1993 Dec, 11(13), 1574 - 8 Large-scale production of Vibrio cholerae toxin B subunit for use in oral vaccines; Lebens M et al.; By systematically manipulating promoter and ribosome binding structures, plasmid copy number and the structure of the cholera toxin B (CTB) subunit gene, we were able to develop a plasmid expression system that, when used in conjunction with an optimized growth medium, provided yields of CTB approaching one gram per liter . The CTB protein which was secreted to > 95%, could readily be purified from the growth medium of a V . cholerae production strain and was shown to be immunologically indistinguishable from previously used vaccine preparations of native or recombinant CTB. Infect Immun, 1993 Dec, 61(12), 5398 - 400 Purification and characterization of pili of a Vibrio cholerae non-O1 strain; Yamashiro T et al.; Pili of the Vibrio cholerae non-O1 strain V10 were purified and characterized . The V10 pili were physicochemically and immunologically different from those of the previously reported V . cholerae non-O1 strain S7, although the pili of the two strains had homologous N-terminal amino acid sequences . V10 plus antigen was detected only in V . cholerae non-O1 strains. Infect Immun, 1993 Dec, 61(12), 5271 - 8 Monoclonal immunoglobulin A antibodies directed against cholera toxin prevent the toxin-induced chloride secretory response and block toxin binding to intestinal epithelial cells in vitro; Apter FM et al.; Secretory immunoglobulin A (IgA) antibodies directed against cholera toxin (CT) are thought to be important in resistance to oral challenge with virulent Vibrio cholerae, although alternative mechanisms for protection of intestinal epithelia against CT-induced fluid secretion have been proposed . The ability of anti-CT IgA to block the effects of CT on human enterocytes has not been directly tested because of the lack of a well-defined in vitro intestinal epithelial cell system to directly measure toxin action and the limited availability of purified anti-CT IgA antibodies . We have generated hybridomas that produce monoclonal IgA and IgG antibodies directed against CT by fusion of Peyer's patch cells with mouse myeloma cells after oral-systemic immunization of mice with CT and CT B-subunit protein . All of the anti-CT antibodies recognized the B subunit . Three clones (designated anti-CTB IgA-1, IgA-2, and IgA-3) which produced IgA antibodies in dimeric and polymeric forms were selected . Checkerboard immunoblotting demonstrated that IgA-1 recognized an epitope distinct from that recognized by IgA-2 and IgA-3 and that none of the antibodies were directed against the binding site of GM1, the intestinal cell membrane toxin receptor . The protective capacity of these IgAs was tested in vitro with human T84 colon carcinoma cells grown on permeable supports as confluent monolayers of polarized enterocytes . When each anti-CTB IgA was mixed with 10 nM CT and applied to the apical surfaces of T84 cell monolayers, all three IgAs blocked CT-induced Cl- secretion in a dose-dependent manner and completely inhibited binding of rhodamine-labelled CT to apical cell membranes . Thus, monoclonal anti-CTB IgA antibodies are sufficient to protect human enterocytes in vitro against CT binding and action. Biochem Biophys Res Commun, 1993 Nov 15, 196(3), 1309 - 15 O-antigenic lipopolysaccharide of Vibrio cholerae O139 Bengal, a new epidemic strain for recent cholera in the Indian subcontinent; Hisatsune K et al.; Lipopolysaccharide (LPS) from Vibrio cholerae O139 Bengal contained colitose (3,6-dideoxy-L-galactose) in addition to glucose, L-glycero-D-manno-heptose, fructose, glucosamine and quinovosamine in its polysaccharide and only glucosamine in lipid A, while perosamine, a characteristic component sugar of V . cholerae O1 LPS, was absent . 3-Hydroxydodecanoic, tetradecanoic and hexadecanoic acids as ester-bound fatty acids and 3-hydroxytetradecanoic acid as amide-bound fatty acid were identified in the lipid A . A very high serological specificity of O139 LPS distinct from that of O1 V . cholerae was demonstrated by passive hemolysis and passive hemolysis inhibition tests by using the LPS either as antigen for sensitizing sheep red blood cells or as inhibitor in the latter test. J Am Acad Dermatol, 1993 Nov, 29(5 Pt 2), 909 - 12 Fatal septicemia and bullae caused by non-01 Vibrio cholerae; Newman C et al.; Bullous lesions associated with non-01 Vibrio cholerae developed in a patient with hepatic cirrhosis who had recently ingested raw oysters . He died of overwhelming sepsis despite 5 days of aggressive antibiotic therapy . Non-01 V . cholerae was isolated from blood, peritoneal fluid, and bullae . The organism produced a cytotoxic factor that destroyed Chinese hamster ovary cells . Although septicemia caused by non-01 V . cholerae is uncommon, cutaneous manifestations of this organism are even rarer . Our patient represents the first reported case of bullous lesions associated with non-01 V . cholerae septicemia. Zentralbl Bakteriol, 1993 Nov, 279(4), 458 - 65 Contribution to some phenotypical characteristics of Vibrio cincinnatiensis . Studies in one strain of a diarrhoeic human patient and in two isolates from aborted bovine fetuses; Wuthe HH et al.; Until now, only a few strains of V . cincinnatiensis have been isolated . This study describes a further three isolates which originated in one case from a stool specimen of an immunocompromised elder patient suffering from enteritis and in two cases from the rennin stomachs of aborted bovine fetuses . These strains grew on TCBS, CIN, MacConkey and XLD plates . Their biochemical activities were dependent on NaCl concentration, in particular the formation of indole, lysine and ornithine decarboxylases, arginine dihydrolase, the reduction of nitrate and behaviour in the Voges-Proskauer test . Moreover, lysine decarboxylase and nitrate reductase were temperature-dependent . The knowledge of these hitherto unknown phenotypical characteristics may facilitate the diagnosis of the pathogen. Mol Gen Mikrobiol Virusol, 1993 Nov-Dec, (6), 18 - 22 {A possible mechanism for the endemicity of modern cholera (the role of noncultivated forms of Vibrio cholerae 01)}; Chetina EV et al.; The surface water sources of some CIS territories have been screened for cholera toxin genes by the polymerase chain reaction . The vct-genes have been found in the majority of water samples indicating the presence of noncultivated vibrio cholerae cells of an epidemiologic significance . The bacteriological methods failed to isolate the active causative agent of cholera . Additional criteria are proposed for epidemiological typing of territories for cholera . The absence of long deletions or insertions in the vct-genes of noncultivated cholera vibrios has been shown in comparison with analogous gene of the active causative agent of cholera. Appl Environ Microbiol, 1993 Nov, 59(11), 3750 - 6 Cloning and sequencing of agaA, a unique agarase 0107 gene from a marine bacterium, Vibrio sp . strain JT0107; Sugano Y et al.; An agarase gene (agaA) was cloned from genomic DNA of Vibrio sp . strain JT0107 . An open reading frame of 2,985 nucleotides gave a primary translation product composed of the mature protein, agarase 0107 (975 amino acid residues, with a molecular weight of 105,271) and a signal peptide of 20 amino acid residues at the N terminus . Comparison of the deduced amino acid sequence of agarase 0107 with those of Streptomyces coelicolor and Pseudomonas atlantica suggests that these enzymes share two regions in common . The AgaA protein which was expressed in Escherichia coli had the agarase activity . Agarase 0107 hydrolyzes not only agarose but also neoagarotetraose {O-3,6-anhydro-alpha-L-galactopyranosyl (1-->3)-O-beta-D-galactopyranosyl(1-->4)-O-3,6-anhydro-alpha-L-galact opy ranosyl (1-->3)-D-galactose} to yield neoagarobiose {O-3,6-anhydro-alpha-L-galactopyranosyl(1-->3)-D-galactose} . This is a quite unique characteristic for a beta-agarase. Appl Environ Microbiol, 1993 Nov, 59(11), 3519 - 24 Direct plating procedure for enumerating Vibrio vulnificus in oysters (Crassostrea virginica); Miceli GA et al.; A procedure for enumerating and identifying Vibrio vulnificus in oysters was developed and evaluated . This method consists of growth on a direct plating medium (VVE medium) for isolating the organism from shellfish tissues, followed by biochemical tests for differentiating and identifying presumptively positive isolates . Densities of V . vulnificus are reliably obtained in 2 to 4 days, and as few as 10 culturable cells per 100 g can be identified . The procedure was evaluated by using a DNA probe technique specific for the cytotoxin-hemolysin gene of V . vulnificus and gas chromatographic analysis of the fatty acid contents of positive isolates . Only 3.2 and 0.4% of the isolates gave false-positive and false-negative results, respectively . The average level of recovery on VVE medium for 33 strains, including both clinical and environmental isolates, was 92% of the level of recovery obtained with brain heart infusion agar supplemented with 1% NaCl . The densities of V . vulnificus in oyster homogenates and individual oysters harvested from gulf and Atlantic coastal waters revealed that seasonally high levels occurred . The VVE medium procedure facilitated enumeration of this pathogen in molluscan shellfish and had a distinct advantage over the widely used most-probable-number procedure for V . vulnificus enumeration, which requires 5 to 7 days and often gives improbable and imprecise results. Bol Med Hosp Infant Mex, 1993 Nov, 50(11), 789 - 96 {Cholera in children . A report of 8 cases}; Lezama-Basulto LA et al.; Cholera is an acute intestinal infection caused by Vibrio cholerae 01 . When an infected person presents severe dehydration and is not adequately treated, he or she will develop hypovolemic shock and eventually could died . There is scarce information concerning this disease in the Pediatric group . Herein we report on eight cases of Pediatric cholera, in children 17 month to four years of age . Seven patients out of eight were admitted presenting dehydration . Four presenting mild or moderate dehydration and three presenting hypovolemic shock . These three patients were rehydrated by intravenous route and thereafter the hydration was maintained by oral therapy . The outcome was uneventful in six patients . One patient developed abdominal distention probably due to hypopotassemia, and another patient presented hyponatremia and seizures . All the patients recovered within five days after admission. Kansenshogaku Zasshi, 1993 Nov, 67(11), 1045 - 51 {Differentiation of cholera-enterotoxin producing Vibrio strains by polymerase chain reaction}; Kobayashi K et al.; The pathogenic factor of Vibrio cholerae that induces a severe watery diarrhea in humans is cholera enterotoxin (CT) . We have earlier reported on the use of a specific polymerase chain reaction method (PCR) for confirmation of CT-production . In our results, a few CT-producing V . mimicus strains were detected by the method . Here we report on the PCR method using 2-primer sets in the same tube for differentiation of toxigenic V . cholerae (O1 and non-O1) and toxigenic V . mimicus . One primer pair is for CT-gene (ctx), and the other pair is for the toxR gene which regulates the ctx gene of V . cholerae . ToxR genes were detected in all CT-producing V . cholerae (both O1 and non-O1) . There were no isolates of the ctx gene positive and toxR gene negative in all V . cholerae strains . On the other hand, V . mimicus strain has not recognized toxR genes except one strain which is similar to the character of V . cholerae . These results indicates that the CT-producing V . cholerae strains are regulated by the toxR gene, but the ctx gene of V . mimicus is controlled by another different genome from toxR of V . cholerae. J Clin Microbiol, 1993 Nov, 31(11), 3068 - 70 Detection of cholera toxin gene in stool specimens by polymerase chain reaction: comparison with bead enzyme-linked immunosorbent assay and culture method for laboratory diagnosis of cholera; Ramamurthy T et al.; Stool specimens obtained from 123 hospitalized patients with acute secretory diarrhea admitted to the Infectious Diseases Hospital, Calcutta, India, were examined for isolation of Vibrio cholerae O1 by direct or enrichment plating on selective media for cholera toxin (CT) by bead enzyme-linked immunosorbent assay (bead-ELISA) and for the CT gene by polymerase chain reaction (PCR) . V . cholerae O1 was isolated either by direct culture or by enrichment culture from 70 stool specimens, all of which gave positive results by PCR . Eleven specimens which were culture negative and bead-ELISA positive also gave positive results by PCR . In addition, 13 more specimens which were negative by both the culture method and bead-ELISA, were positive by PCR . With the combined results of both the culture method and the CT bead-ELISA, a confirmed laboratory diagnosis of cholera could be made from 81 stool specimens, while the combined results of the three methods, including PCR, yielded a positive result for 94 specimens examined . From these data, we conclude that PCR provides a more sensitive and specific assay for rapid diagnosis of cholera than currently available methods. Am J Surg, 1993 Nov, 166(5), 563 - 7 Surgical hazards posed by marine and freshwater animals in Florida; Howard RJ et al.; Marine and freshwater animals can cause injury to humans by biting, stinging, being poisonous to eat, and causing infections . Biting aquatic animals in Florida include sharks, barracudas, alligators, and moray eels . Devitalized tissue should be debrided, and vascular, neurologic, and tendinous injuries should be repaired . Radiographs should be obtained to examine the injury sit for fractures and retained foreign bodies (teeth) . The spines of stingrays and marine catfish can cause soft tissue injury and infection . The spine has a recurved, serrated shape that may cause further injury and break if it is pulled out . The venom may cause local tissue necrosis requiring debridement . Soft tissue infections with marine Vibrio bacteria can occur after eating raw oysters or receiving even minor injuries from marine animals . Thirty-one individuals developed soft tissue infections, 49 developed sepsis, and 23 developed both sepsis and soft tissue infection with marine Vibrio species during a 12-year period . Sixteen patients developed necrotizing soft tissue infections . Treatment is with antibiotics and debridement when necrosis occurs. J Infect Dis, 1993 Nov, 168(5), 1169 - 76 Safety, immunogenicity, and transmissibility of single-dose live oral cholera vaccine strain CVD 103-HgR in 24- to 59-month-old Indonesian children; Simanjuntak CH et al.; Recombinant A-B+ Vibrio cholerae O1 strain CVD 103-HgR is a safe, highly immunogenic, single-dose live oral vaccine in adults in industrialized countries . Safety, excretion, immunogenicity, vaccine transmissibility, and environmental introduction of CVD 103-HgR were investigated among 24- to 59-month-old children in Jakarta . In 81 households, 1 child was randomly allocated a single dose of vaccine (5 x 10(9) cfu) and another, placebo . Additionally, 139 unpaired children were randomly allocated vaccine or placebo . During 9 days of follow-up, diarrhea or vomiting did not occur more often among vaccines than controls . Vaccine was minimally excreted and was isolated from no controls and from 1 (0.6%) of 177 unvaccinated family contacts . A 4-fold or higher rise in serum vibriocidal antibody was observed in 75% of vaccines (10-fold rise in geometric mean titer over baseline) . Of 135 paired placebo recipients or household contacts, 5 had vibriocidal seroconversions . Moore swabs placed in sewers and latrines near 97 households failed to detect vaccine . These observations pave the way for a large-scale field trial of efficacy. J Bacteriol, 1993 Nov, 175(22), 7307 - 12 The Vibrio fischeri luminescence gene activator LuxR is a membrane-associated protein; Kolibachuk D et al.; The Vibrio fischeri luminescence (lux) genes are activated at sufficiently high culture densities by the transcriptional activator LuxR in combination with a diffusible signal compound termed autoinducer . We have used antibodies directed against LuxR in immunoprecipitation experiments to study the subcellular location of this transcription factor . The LuxR polypeptide was detected in membranes and not in the soluble pool of cytoplasmic proteins from V . fischeri . LuxR was not released from the membranes by 0.6 M KCl or by the nonionic detergents Nonidet P-40, N-octyl-beta-D-glucopyranoside, and Triton X-100 . LuxR and a number of other V . fischeri proteins were released from the membranes by EDTA . The autoinducer had no detectable influence on the subcellular location of LuxR . In spheroplasts, neither the abundance nor the molecular mass of the LuxR antigen was influenced by treatment with proteinase K . Together with other information, these results indicate that LuxR is an amphipathic protein that is associated with the cytoplasmic membrane of V . fischeri. J Bacteriol, 1993 Nov, 175(21), 6873 - 80 Genes encoding two isocitrate dehydrogenase isozymes of a psychrophilic bacterium, Vibrio sp . strain ABE-1; Ishii A et al.; The genes coding for two structurally different isocitrate dehydrogenase isozymes (IDH-I and IDH-II) of a psychrophilic bacterium, Vibrio sp . strain ABE-1, were cloned and sequenced . Open reading frames of the genes (icdI and icdII) are 1,248 and 2,229 bp in length, respectively . The amino acid sequences predicted from the open reading frames of icdI and icdII corresponded to the N-terminal amino acid sequences of the purified IDH-I and IDH-II, respectively . No homology was found between the deduced amino acid sequences of the isozymes; however, the IDH-I, a dimeric enzyme, had a high amino acid sequence identity (74.3%) to the Escherichia coli IDH . The deduced amino acid sequence of the IDH-II, a monomeric enzyme, was not related to any known sequence . However, the IDH-II had an amino acid sequence homologous to that of a cyanogen bromide-cleaved peptide containing a putative active-site methionyl residue of the monomeric IDH of Azotobacter vinelandii . The two genes (icdlI and icdII) were found to be tandemly located in the same orientation . Northern (RNA) blot analyses showed that the two genes are transcribed independently . Primer extension experiments located single transcriptional start sites 39 and 96 bp upstream of the start codons of icdI and icdII, respectively . The amount of icdI transcript but not icdII increased when Vibrio sp . strain ABE-1 cells were cultured in acetate minimal medium. Mol Microbiol, 1993 Nov, 10(4), 891 - 901 The ompS gene of Vibrio cholerae encodes a growth-phase-dependent maltoporin; Lang H et al.; The outer membrane of Vibrio cholerae contains a maltose-inducible major protein, OmpS (43 kDa), that is common to different isolates . Nucleotide sequence analysis of the corresponding structural gene, ompS, revealed an open reading frame encoding a 412-amino-acid polypeptide . The amino acid sequence of OmpS is similar to that of LamB, the Escherichia coli maltoporin, and to ScrY or Klebsiella pneumoniae, although the antigenic determinants of these proteins are different . The cloned ompS gene complemented an ompS mutation of V . cholerae and the corresponding polypeptide could function as a maltoporin in a LamB- mutant of E . coli . The promoter region of ompS is highly homologous to the malK-lamB promoter of E . coli and the ompS gene is controlled by MalT in E . coli . This indicates that the same kind of regulatory mechanism is used to activate the ompS expression in V . cholerae and malK-lamB expression in E . coli . An ompS-lacZ transcriptional fusion was used to demonstrate a dual control in ompS expression; the ompS gene is responsive to the inducers maltose and trehalose but in their absence it is also expressed in response to growth-phase . These different modes of induction might be of importance during different stages of V . cholerae infection. Appl Microbiol Biotechnol, 1993 Nov, 40(2-3), 356 - 60 Saturation mutagenesis in Escherichia coli of a cloned Xanthomonas campestris DNA fragment with the lux transposon Tn4431 using the delivery plasmid pDS1, thermosensitive in replication; Steinmann D et al.; A system allowing transposon mutagenesis of cloned DNA fragments in Escherichia coli with Tn4431, which carries the promotorless luciferase (lux) operon of Vibrio fischeri, has been developed . The transposon delivery plasmid, pDS1, based on an IncF replicon, is thermosensitive in replication and mobilizable to many Gram-negative bacteria . We used pDS1 for Tn4431-saturation mutagenesis of a 10-kb DNA fragment of Xanthomonas campestris pv . campestris (X.c.c.) in E . coli and showed that the expression of the lux operon was dependent on orientation and location of the transposon . Transfer of a specific Tn4431 insertion to X.c.c . allowed the determination of the bioluminescence phenotype in planta. Zentralbl Bakteriol, 1993 Nov, 279(4), 494 - 504 Entero-cytolysin (EC) from Vibrio cholerae non-O1 (some properties and pore-forming activity); Zitzer AO et al.; A thermolabile extracellular entero-cytolysin (EC) from Vibrio cholerae non-O1 was purified by ammonium sulphate fractionation, DE-52 cellulose ion exchange chromatography, gel-filtration on Ultrogel AcA-44 and high performance liquid chromatography on a Mono Q . The purified EC had a molecular weight of 63 kD and an isoelectric point of 6.2 . It was not inactivated by cholesterol or 5,5'-dithio-bis(2-nitrobenzoic acid), nor activated by dithiothreitol . EC had no immunological cross-reactivity with cholera toxin . The EC caused fluid accumulation in the intestines of infant rabbits, death of mice by intravenous injection, and increased vascular permeability in the paw oedema test in mice . V . cholerae non-O1 EC lysed erythrocytes from various species and cultured cells (CHO, L-929, L-41, HEp-2, Vero, MDCK and BHK-21) . In contrast to cholera toxin, EC caused crude destruction of target cells . The EC caused hemolysis by a colloid-osmotic mechanism due to the formation of hydrophilic pores of 1.8-2.0 nm diameter in the cell membrane . This EC also was able to open pores in lipid membranes . The induced channels were anion-selective and had a diameter of 1.8-2.0 nm. Appl Environ Microbiol, 1993 Nov, 59(11), 3863 - 70 Ribotypes and plasmid contents of Vibrio anguillarum strains in relation to serovar; Olsen JE et al.; Eighty-six strains of the 10 major agglutination types of Vibrio anguillarum (serovars O1 to O10) and 6 nontypeable strains of V . anguillarum have been characterized by ribotyping with a probe complementary to 16S and 23S rRNA of Escherichia coli and by plasmid profile analysis . Forty-four different ribotypes were observed with the restriction enzyme HindIII . Ribotype similarity was compared by using the Dice coefficient (Sd), and three significantly different levels of homogeneity within the V . anguillarum serovars were observed (serovars O1, O3A, O7, and O9, Sds of > 90%; serovars O2B, O4, and O10, Sds of 80 to 90%; serovars O2A, O3B, O5, and O8, Sds between 46 and 70%) . None of the ribotype patterns of V . anguillarum strains were observed among 20 other Vibrio strains typed for comparison . By cluster analysis, the V . anguillarum strains were divided into a main cluster containing 83 strains, while all strains of serovar O3B, one strain (each) of serovars O2A, O5, and O8, and a nontypeable strain were separated from this cluster by at least 15% difference in similarity coefficients . Plasmids were demonstrated in only six strains other than serovar O1 . In serovar O1, a 67- to 70-kilobase-pair (kb) plasmid molecule was present in 17 of 19 strains tested; of the two remaining strains, one strain harbored two plasmids (45 and 6.5 kb) and one strain had no plasmids. Tidsskr Nor Laegeforen, 1993 Oct 10, 113(24), 3017 - 8 {Severe gastroenteritis after domestic infection with Vibrio cholerae non-O1}; Henriksen AZ et al.; Vibrio cholerae non-O1 has previously been isolated only occasionally in Norway from patients infected abroad . This report describes the clinical course in two patients who were domestically infected with V . cholerae non-O1 . In one case, contaminated seafood, i.e . crab, was the probable source of infection . Both patients displayed a cholera-like type of illness, with severe diarrhoea and electrolyte disturbances . It is recommended that Norwegian laboratories be prepared to isolate vibrios also in domestically infected cases when watery diarrhoea is present. Carbohydr Res, 1993 Oct 4, 248, 199 - 211 Structural studies of the Vibrio cholerae O:5 O-antigen polysaccharide; Hermansson K et al.; The O-polysaccharide from Vibrio cholerae O:5 has been investigated, using NMR spectroscopy as the main method . Fast atom bombardment mass spectrometry (FABMS) studies of fragments obtained on treatment with anhydrous hydrogen fluoride or methanolic hydrogen chloride gave further structural information . Some structural features were also determined by comparison of nuclear Overhauser enhancement (NOE) contacts with calculated H-H distance in different oligosaccharide models . It is concluded that the O-polysaccharide has the following structure, in which D-Qui p NAc4NAc is 2,4-diacetamido-2,4,6-trideoxy-D-glucose, and D-Fuc p 3NX is 3-amino-3,6-dideoxy-D-galactose acylated with a (R,R)-3-hydroxy-3-methyl-5-oxoproline group . {formula: see text} Biochim Biophys Acta, 1993 Oct 3, 1158(2), 137 - 45 Subunit interactions and the role of the luxA polypeptide in controlling thermal stability and catalytic properties in recombinant luciferase hybrids; Li Z et al.; Bacterial luciferases with over 70% sequence identity from the terrestrial species, Xenorhabdus luminescens, and the marine species, Vibrio harveyi, exhibit large differences in thermal stability (Szittner and Meighen, 1990, J . Biol . Chem . 265, 16581-16587) . The origin of these differences was investigated with genetically constructed hybrids containing one subunit from X . luminescens and the other from V . harveyi . While no activity was detected with the single (alpha and beta) subunits both in vitro and in vivo, the recombinant hybrid luciferases (alpha Xl beta Vh and alpha Vh beta Xh) were highly active and could be purified to homogeneity . The kinetic properties of the hybrid enzymes including aldehyde specificity, flavin binding and luminescence decay rates, were found to be nearly identical to those of the native luciferases (alpha Xl beta Xl or alpha Vh beta Vh) containing the same alpha subunit . In addition, the rate of thermal inactivation and temperature dependent quenching of the intrinsic fluorescence by flavin were found to be independent of the nature of the beta subunit, quite opposite to previous reports that the thermal stability is controlled by the beta subunit . Thus, the alpha subunit appears primarily responsible for controlling both the catalytic and structural properties of luciferase. J Med Microbiol, 1993 Oct, 39(4), 310 - 7 Virulence patterns of Vibrio cholerae non-O1 strains isolated from hospitalised patients with acute diarrhoea in Calcutta, India; Ramamurthy T et al.; A collection of 28 strains of Vibrio cholerae non-O1 isolated during a 3-year period (1989-1991) from hospitalised patients with acute diarrhoea in Calcutta, India, were examined with regard to virulence-associated factors . Of the 28 isolates (each representing a case), 18 were isolated as the sole infecting agent; the remaining 10 were recovered as co-cultures from cases infected with V . cholerae O1 . Of the strains isolated in this study, 82% could be serotyped, with serovars O5 (32.1%), O11 and O34 (14.3% each) predominant . Serovars O7, O14, O34, O39 and O97 were associated exclusively with sole infections . Two strains of V . cholerae non-O1 produced anti-cholera toxin IgG-absorbable cholera toxin (CT) . Both CT-producing V . cholerae non-O1 strains hybridised with the DNA probe specific for the zonula occludens toxin (ZOT) but none of the remaining 26 strains hybridised with the ZOT probe . The majority of the strains were cytotoxic for CHO, HeLa and Vero cells, with end-point titres of 4-512 . Fewer strains produced a cytotonic effect, with end-point titres of 2-16 . Of the 28 strains of V . cholerae non-O1 examined, 75%, 75%, 25% and 14.3% produced haemolysin that was active against erythrocytes of rabbit, sheep (Eltor haemolysin), chicken and man, respectively . Strains that produced a haemolysin active against both rabbit and sheep erythrocytes were dominant (35.7%) . Ten (35.7%) of the 28 strains examined showed cell-associated haemagglutinating activity on human blood . Of the 10 strains, nine were isolated as sole pathogen and only one strain was associated with mixed infection.(ABSTRACT TRUNCATED AT 250 WORDS) Infect Immun, 1993 Oct, 61(10), 4462 - 8 CVD110, an attenuated Vibrio cholerae O1 El Tor live oral vaccine strain; Michalski J et al.; The recent expansion of the seventh cholera pandemic into South America emphasizes the need for a safe, long-lasting, protective, and nonreactogenic vaccine for this disease . Since the predominant Vibrio cholerae O1 strains in the world today are of the El Tor biotype, a bivalent vaccine containing both classical and El Tor biotypes may be desirable . We have constructed a new oral vaccine candidate, V . cholerae CVD110 El Tor, Ogawa, from which all toxin genes so far identified in V . cholerae have been deleted . Three of these genes, those encoding cholera toxin (ctx), zonula occludens toxin (zot), and accessory cholera enterotoxin (ace), are located on a 4.5-kb virulence cassette flanked by repetitive sequences (RS1 elements) . Homologous recombination between these RS1 elements resulted in the deletion of this virulence cassette to yield V . cholerae CVD109 . Insertion of genes encoding mercury resistance (mer) and the cholera toxin B subunit (ctxB) into the hemolysin locus (hlyA) produced CVD110 . This insertion serves three purpose . (i) It genetically tags the vaccine strain so as to distinguish it from wild-type V . cholerae O1 . (ii) It produces cholera toxin B subunit in order to elicit antitoxic immunity . (iii) It inactivates the hemolysin gene, rendering the strain nonhemolytic on sheep erythrocyte plates . Supernatants from V . cholerae CVD110 cultures are nonreactogenic when assayed in Ussing chambers. Infect Immun, 1993 Oct, 61(10), 4326 - 32 Cation flux studies of the lesion induced in human erythrocyte membranes by the thermostable direct hemolysin of Vibrio parahaemolyticus; Huntley JS et al.; Vibrio parahaemolyticus, an important agent of seafood-borne gastroenteritis, expresses several putative virulence factors that could account for the disease symptoms of infected humans, namely, diarrhea, nausea, and abdominal cramps . The pathogenicity of V . parahaemolyticus correlates well with the Kanagawa phenomenon (the hemolytic ability of strains grown on Wagatsuma blood agar), implicating the thermostable direct hemolysin (TDH) as the predominant toxin responsible for pathogenicity . TDH-induced hemolysis could be inhibited by the addition of the osmolyte sorbitol to the extracellular solution, supporting the hypothesis that hemolysis occurs through colloid osmosis secondary to an increase in the cation permeability of the membrane . The effect of TDH on cation permeability was investigated by measuring K+ (congener, 86Rb+) influx into human erythrocytes in which the endogenous cation transporters had been blocked (by use of ouabain, bumetanide, and nitrendipine) . TDH increased K+ influx into these cells; this increase was rapid in onset and constant in magnitude, suggesting a direct action by TDH on the membrane . The kinetics of leak generation were examined; the relationship between counts accumulated and hematocrit indicated that the TDH-induced lesion is multihit in nature . TDH-induced K+ influx was sensitive to Zn2+ . Time courses of hemolysis in isosmotic solutions of monovalent cation chlorides were used to obtain the selectivity series for the TDH-induced leak: Cs+ > Li+ > K+ > Rb+ > Na+ . Both the Zn2+ sensitivity and this selectivity series were obtained for crude culture supernatants, suggesting that TDH is the predominant leak-inducing agent . Thus, we have identified several features of the TDH-induced leak likely to be important in the diarrhetic action of V . parahaemolyticus in the human intestine. Microb Pathog, 1993 Oct, 15(4), 269 - 82 Cloning and characterization of three hemolysin genes from Aeromonas salmonicida; Hirono I et al.; Two hemolysin genes (ASH3 and ASH4) of Aeromonas salmonicida strain 17-2 and one (ASH1) of A . salmonicida ATCC14174 were cloned into the cosmid vector charomid 9-36 in Escherichia coli DH1 . The overall amino acid sequence of the ASH3 was similar to that of the aerolysins of Aeromonas hydrophila and Aeromonas sobria, and hemolysins AHH3, AHH4, and AHH5 of A . hydrophila, and hemolysin ASA1 of A . sobria . The sequence of ASH4 was similar to that of the AHH1 hemolysin of A . hydrophila . The ASH4 hemolysin contains some homologous sequence regions of the Vibrio vulnificus and Vibrio cholerae cytolysin-hemolysin . Both ASH3 and ASH4 DNA probes reacted with all 104 strains of A . salmonicida, whereas the ASH1 DNA probe did not hybridize with any of the 104 strains studied except strain ATCC14174 . ASH1 and ASH3 were broad spectrum hemolysins with most activity against rabbit and horse erythrocytes, respectively, whereas ASH4 hemolysin did not lyse bovine and horse erythrocytes . ASH3 and ASH4, but not ASH1, were activated by trypsin. Indian J Exp Biol, 1993 Oct, 31(10), 808 - 12 Effects of nitrofurantoin on viability, DNA synthesis and morphology of Vibrio cholerae cells; Mukherjee U et al.; Nitrofurantoin caused a dose dependent inhibition of growth and decrease in viability of V . cholerae cells, the 10% (D10) and 37% (D37) survival doses being 50 and 19 micrograms/ml respectively . The drug at a concentration of 60 micrograms/ml caused 86% inhibition of DNA synthesis . Both light and electron microscopic observations revealed that treatment with nitrofurantoin (60 micrograms/ml for 1 hr at 37 degrees C) led to a significant filamentation of the V . cholerae cells, ultrastructure of the cell cytoplasm, plasma membrane and cell wall however remaining unaltered from those of untreated cells . The results are discussed in relation to DNA lesions produced by and the carcinogenic potential of the drug. Hawaii Med J, 1993 Oct, 52(10), 274 - 5 Vibrio in stinging seaweed: potential infection; Sims JK et al.; Toxic strains of the finely filamentous, velvety, dark-olive green to black algal organism, Microcolus Lyngbyaceus, (formerly Lyngbya majuscula Gomont, or "lyngbya") have been recognized as etiologic agent of "stinging seaweed" dermatitis (one of several forms of "swimmer's itch") in Hawaii since the late 1950s as reviewed . Lymphadenopathy, pustular folliculitus, and local infections have been reported in some persons. Anal Biochem, 1993 Oct, 214(1), 106 - 15 Capsular polysaccharide structure of a clinical isolate of Vibrio vulnificus strain BO62316 determined by heteronuclear NMR spectroscopy and high-performance anion-exchange chromatography; Reddy GP et al.; Virulence of Vibrio vulnificus has been strongly associated with encapsulation . Capsular polysaccharide was purified from a virulent strain of Vibrio vulnificus BO62316, a clinical isolate, by dialysis, centrifugation, enzymatic digestion, and phenol-chloroform extraction . This polysaccharide shows partial reactivity with antibodies to the capsular polysaccharide of a related pathogenic strain of V . vulnificus (MO6-24) whose structure was recently reported . Nuclear magnetic resonance spectroscopic analysis of the purified polysaccharide from strain BO62316 showed that the polymer is composed of a repeating structure with four sugar residues per subunit: a residue of 2-acetamido-2,6-dideoxy-hexo pyranose in the alpha-gluco configuration (QuiNAc), a residue of 2-acetamido-2,6-dideoxy hexopyranose in the alpha-galacto configuration (FucNAc), a residue of 2-acetamido-2,6-dideoxy hexopyranose in the alpha-manno configuration (RhaNAc), and a residue of 2-acetamido-2,6-dideoxy hexouronate in the alpha-galacto configuration (GalNAcA) . The complete carbohydrate structure of the capsular polysaccharide was determined by heteronuclear nuclear magnetic resonance spectroscopy and by high-performance anion-exchange chromatography . The 1H and 13C spectra were completely assigned, and vicinal coupling relationships were used to establish the stereochemistry of each sugar residue, its anomeric configuration, and the position of the glycosidic linkages . The complete structure is Formular; {see text} . This is the first reported occurrence of RhaNAc in a capsular polysaccharide. FEMS Microbiol Lett, 1993 Oct 1, 113(1), 67 - 70 Cell-associated hemagglutinin of classical vibrio cholerae O1 with reference to intestinal adhesion; Nakasone N et al.; Vibrio cholerae O1 86B3 biovar cholerae has at least two types of cell-associated hemagglutinin . One is cell wall-associated and L-fucose sensitive, whereas the other is pili-associated and D-mannose sensitive . A pilus rich variant of 86B3 and a poorly piliated parent strain adhered equally to the rabbit intestine . This adhesion was inhibited by L-fucose but not by D-mannose . A Fab fraction prepared from anti-pilus antibody did not inhibit the adhesion . These results suggest that not the pili but a colonization factor located in the outer membrane of the organisms plays a role in intestinal adhesion. J Appl Bacteriol, 1993 Oct, 75(4), 350 - 9 Construction and detection of bioluminescent strains of Bacillus subtilis; Cook N et al.; Bioluminescence (lux) genes from Vibrio fischeri and V . harveyi were introduced into Bacillus subtilis on a plasmid vector and by chromosomal integration . The plasmid-bearing strain was highly luminescent and stable under antibiotic selection, but luminescence was lost in the absence of selection and following sporulation and germination . The chromosomally marked strains emitted less light but were found to be stable without the requirement for antibiotic selection and following sporulation and germination . Individual luminescing colonies of both B . subtilis strains could be detected against a high background of non-bioluminescent indigenous soil microbial colonies on agar plates using a charge-coupled device camera . These bioluminescent Gram-positive strains could be of value in studies concerning the survival and spread of genetically-modified micro-organisms in soil environments. Mem Inst Oswaldo Cruz, 1993 Oct-Dec, 88(4), 593 - 7 Laboratory evaluation on pathogenic potentialities of Vibrio furnissii; Magalhaes V et al.; Sixteen strains of Vibrio furnissii recovered from 16 Brazilian patients with diarrhea were screened for virulence-associated factors . All strains were non-invasive, non-fimbriated, and did not produce either enterotoxins or cholera-like toxin . In contrast, most were hemolytic on blood agar and their broth-culture supernatants damaged HeLa cell monolayers . These cytolysins, as accepted for other enteropathogenic members of the family Vibrionaceae, might be determinants of pathogenicity in V . furnissii-mediated enteritis. Mem Inst Oswaldo Cruz, 1993 Oct-Dec, 88(4), 589 - 92 Evaluation of virulence factors in environmental isolates of Vibrio species; Rodrigues DP et al.; Strains of Vibrio parahaemolyticus, Vibrio fluvialis and Vibrio mimicus isolated from seafood and seawater were examined for characteristics related to infectivity, such as enzymatic activity and animal assays . All strains hydrolysed DNA, starch, gelatin and chitin . Variable results were obtained with the haemolysin, chondroitin, collagen, elastin and lecithin tests . Production of thermostable direct haemolysin by V . parahaemolyticus was detected in 7.1% strains derived from seafood and 2% from seawater . In the animal assays, strains of V . fluvialis showed positive results at skin PF (75%), mouse lethality (100%), but no fluid accumulation in the suckling mice model was noted . Concerning V . mimicus, results showed skin PF (100%), mouse lethality (100%) and fluid accumulation in suckling mice (66.6%). Microb Pathog, 1993 Oct, 15(4), 255 - 68 In vitro production of toxin-coregulated pili by Vibrio cholerae El Tor; Voss E et al.; Toxin-coregulated pili (TCP) have been shown to be a virulence determinant and protective antigen for Vibrio cholerae strains of classical biotype, but their role in infection by strains of the alternative El Tor biotype remains uncertain . In an attempt to demonstrate TCP production by El Tor vibrios we have over-expressed the El Tor tcpA gene in Escherichia coli, in order to prepare a biotype-specific anti-TcpA serum . This reagent proved to be a very sensitive indicator of TcpA production in immunoblotting studies, but failed to detect polymerized pili on the bacterial surface by immuno-electron microscopy (IEM) . However, results with an analogous reagent which detects classical TcpA suggested that antisera to unprocessed TcpA do not efficiently recognize epitopes on native proteins . Accordingly we prepared a serum against a cell envelope fraction rich in processed El Tor TcpA . After extensive absorption this reagent reacted almost exclusively with TcpA by immunoblotting; when used in IEM, it allowed visualization of typical TCP bundles on the surfaces of each of five El Tor strains known to produce TcpA in vitro . We have previously reported that the El Tor strain O17 does not synthesize TcpA during growth in vitro, but that an O17 clone carrying a cosmid of classical origin expresses surface TCP . Using the biotype-specific anti-TcpA reagents in immunoblotting studies it has been possible to detect the product of the host chromosomal tcpA gene in such constructs; transcription of this gene was confirmed using biotype-specific tcpA probes . IEM revealed that El Tor TcpA was present in the TCP bundles associated with the O17 cosmid clones . Further studies suggest that regulation of the genes encoding TcpA and cholera toxin varies between different strains of El Tor biotype. J Med Assoc Thai, 1993 Oct, 76 Suppl 2, 42 - 8 Comparison of efficacy of peptilose-base ORS (ORALNU) and WHO-ORS; Simakachorn N et al.; We examined whether replacing glucose with Peptilose into standard ORS would be advantageous over WHO-ORS . A study was carried out on 134 diarrheal children with mild to moderate dehydration . They received either WHO-ORS or Peptilose-ORS by randomized selection . In only two cases in each group, diarrhea was caused by Vibrio cholerae non 0-1 . Significant per cent weight gain was observed in patients with Peptilose-ORS compared to those treated with WHO-ORS (P = 0.046) . The patients could voluntarily take a higher amount of Peptilose-ORS and had significantly less stool output in the combined mildly and moderately dehydrated patients . It is concluded that Peptilose-ORS is more advantageous and acceptable than the standard WHO glucose-ORS for treatment of non cholera and 2 cases of cholera dehydrating diarrhea in children. Bioorg Khim, 1993 Oct, 19(10), 989 - 1000 {Structure of the repeating unit of O-specific polysaccharide from Vibrio fluvialis}; Nazarenko EL et al.; O-Specific polysaccharide of Vibrio fluvialis, strain AQ-0002B, is built up to pentasaccharide repeating units contained of D-mannose, 2-acetamido-2-deoxy-D-glucose, 2-acetamido-2-deoxy-D-galactose, D-galacturonic acid and 3-0-{(R)-1'-carboxyethyl}-L-rhamnose (rhamnoactylic acid, Rha3Lac) residues . On the basis of methylation studies, solvolysis with HF, Smith degradation, 1H and 13C NMR-spectroscopy including homo- and heteronuclear correlation spectroscopy and NOE experiments, the following structure was suggested for the polysaccharide repeating unit: {formula: see text} Gene, 1993 Sep 30, 132(1), 101 - 6 Genes required for extracellular secretion of enterotoxin are clustered in Vibrio cholerae; Overbye LJ et al.; Pleiotropic transposon insertion mutants of Vibrio cholerae that are unable to secrete enterotoxin, HA/protease and chitinase through the outer membrane have been isolated . The gene, epsM, responsible for complementation of two of the Tn5 insertion mutations was sequenced . It encodes a putative cytoplasmic membrane protein of 18.5 kDa that exhibits similarity to proteins required for extracellular secretion of pullulanase, pectate lyase or elastase in other Gram-bacteria . It is present on a 15-kb DNA fragment from the V . cholerae genome, containing the epsE gene that was previously shown to be required for secretion of cholera toxin {Sandkvist et al., Gene 123 (1993) 81-86} . Partial reading frames flanking epsM also demonstrated similarity to genes required for extracellular secretion of pullulanase in Klebsiella oxytoca. Biochem J, 1993 Sep 15, 294 ( Pt 3), 653 - 6 A kinetic-isotope-effect study of catalysis by Vibrio cholerae neuraminidase; Guo X et al.; Michaelis-Menten parameters for hydrolysis of seven aryl N-acetyl alpha-D-neuraminides by Vibrio cholerae neuraminidase at pH 5.0 correlate well with the leaving-group pKa (delta pK 3.0; beta 1g (V/K) = -0.73, r = -0.93; beta 1g (V) = -0.25; r = -0.95) . The beta-deuterium kinetic-isotope effect, beta D2(V), for the p-nitrophenyl glycoside is the same at the optimum pH of 5.0 (1.059 +/- 0.010) as at pH 8.0 (1.053 +/- 0.010), suggesting that isotope effects are fully expressed with this substrate at the optimum pH . For this substrate at pH 5.0, leaving group 18O effects are 18(V) = 1.040 +/- 0.016 and 18(V/K) = 1.046 +/- 0.015, and individual secondary deuterium effects are beta proRD(V) = 1.037 +/- 0.014, beta proSD(V) = 1.018 +/- 0.015, beta proRD(V/K) = 1.030 +/- 0.017, beta proSD(V/K) = 1.030 +/- 0.017 . All isotope effects, and the beta 1g(V/K) value are in accord with the first chemical step being both the first irreversible and the rate-determining step in enzyme turnover, with a transition state in which there is little proton donation to the leaving group, the C-O bond is largely cleaved, there is significant nucleophilic participation, and the sugar ring is in a conformation derived from the ground-state 2C5 chair . The apparent conflict between the beta 1g (V) value of -0.25 with all the kinetic-isotope-effect data can be resolved by the postulation of an interaction between the pi system of the aglycone ring and an anionic or nucleophilic group on the enzyme. J Clin Microbiol, 1993 Sep, 31(9), 2529 - 30 Severe human infections caused by Vibrio metschnikovii; Hansen W et al.; Vibrio metschnikovii is largely distributed in the aquatic environment; human infections are rarely observed . A fatal case of septicemia in a patient with liver cirrhosis, renal insufficiency, and diabetes is described . A second case in a 82-year-old woman with septicemia, respiratory problems, and infected leg lesions is reported; she was successfully treated. J Clin Microbiol, 1993 Sep, 31(9), 2251 - 4 Construction of a bioluminescent mycobacterium and its use for assay of antimycobacterial agents; Andrew PW et al.; To show, as a model system, that mycobacteria can express heterologous luciferase genes and that bioluminescence can be a rapid method of measuring antimycobacterial activity, a bioluminescent form of Mycobacterium smegmatis was made by transformation with a Mycobacterium-Escherichia coli shuttle vector containing the luxAB genes from Vibrio harveyi . The antimycobacterial effects of antibiotics and biocides could be assayed in real time by using bioluminescent M . smegmatis. J Bacteriol, 1993 Sep, 175(17), 5488 - 504 High-molecular-weight protein 2 of Yersinia enterocolitica is homologous to AngR of Vibrio anguillarum and belongs to a family of proteins involved in nonribosomal peptide synthesis; Guilvout I et al.; The iron-regulated irp2 gene is specific for the highly pathogenic Yersinia species and encodes high-molecular-weight protein 2 (HMWP2) . Despite the established correlation between the presence of HMWP2 and virulence, the role of this protein is still unknown . To gain insight into the function of HMWP2, the entire coding sequence and the promoter of irp2 were sequenced . Two putative -35 and -10 promoter sequences were identified upstream of a large open reading frame, and two potential Fur-binding sites were found overlapping the second -35 box . The large open reading frame is composed of 6,126 nucleotides and may encode a protein of 2,035 amino acids (ca . 228 kDa) with a pI of 5.81 . A signal sequence was not present at the N terminus of the protein . Despite the existence of 30 cysteine residues, carboxymethylation prevented the formation of most if not all disulfide bonds that otherwise occurred when the cells were sonicated . The protein was composed of three main domains: a central region of ca . 850 residues, bordered on each side by a repeat of 550 residues . A high degree of identity (44.5%) was found between HMWP2 and the protein AngR of Vibrio anguillarum . The central part of HMWP2 (after removal of a loop of 337 residues) also displayed significant homology with proteins belonging to the superfamily of adenylate-forming enzymes and, like them, possessed a putative ATP-binding motif that is also present in AngR . In addition, HMWP2 shared with the group of antibiotic and enterochelin synthetases a potential amino acid-binding site . Six consensus sequences defining the superfamily and four defining the family of synthetases were derived from the multiple alignment of the 30 sequences of proteins or repeated domains . A phylogenetic tree that was constructed showed that HMWP2 and AngR are in a family composed of Lys2, EntF, and the tyrocidine, gramicidin, and Beta-lactam synthetases . This finding suggests that HMWP2 may participate in the nonribosomal synthesis of small biologically active peptides. Infect Immun, 1993 Sep, 61(9), 3994 - 7 Safety, immunogenicity, and excretion pattern of single-dose live oral cholera vaccine CVD 103-HgR in Peruvian adults of high and low socioeconomic levels; Gotuzzo E et al.; Groups of 122 Peruvian adults of low socioeconomic level (SEL) and 125 of high SEL received a randomly allocated 5 x 10(9)- or 5 x 10(8)-CFU dose of CVD 103-HgR live oral cholera vaccine or a placebo . The vaccine was well tolerated . Vibriocidal seroconversions occurred in 78% of high-SEL and 72% of low-SEL subjects who ingested the high dose and in 78 and 49%, respectively, of those who received the low dose. J Infect Dis, 1993 Sep, 168(3), 758 - 62 Capsular types of Vibrio vulnificus: an analysis of strains from clinical and environmental sources; Hayat U et al.; Vibrio vulnificus produces a capsular polysaccharide (CPS) that is essential for virulence . CPS from V . vulnificus clinical strain MO6-24 has been purified and the structure determined . In preliminary screening with antisera raised to MO6-24 CPS, 4 (19%) of 21 clinical isolates (including MO6-24), but none of 67 environmental V . vulnificus isolates, agglutinated with anti-MO6-24 antisera (P = .003) . CPS was isolated from a subset of 12 clinical and 7 environmental isolates and analyzed by high-performance anion-exchange chromatography and one-dimensional nuclear magnetic resonance . MO6-24 and 1 other serologically positive strain had identical CPS structures; the other 2 serologically positive strains had substitutions in two of four sugar residues . Thirteen other capsular types were identified among the remaining 15 strains from which CPS was extracted. J Burn Care Rehabil, 1993 Sep-Oct, 14(5), 544 - 51 Preliminary evaluation of vibriolysin, a novel proteolytic enzyme composition suitable for the debridement of burn wound eschar; Durham DR et al.; Vibriolysin, a proteolytic enzyme secreted by the marine microorganism Vibrio proteolyticus, was evaluated for its efficacy as an enzymatic debridement agent . Initial in vitro experiments revealed that the protease readily hydrolyzed proteinaceous components of eschar (e.g., fibrin, elastin, and in particular, collagen) . Enzyme selectivity towards the digestion of denatured proteins but not viable elements was demonstrated in vitro and in vivo by treatment of full-thickness burn wounds with a porcine model . The full-thickness wound eschar was rapidly hydrolyzed by a hydrophilic vibriolysin composition with a resultant wound bed that appeared pink and viable . Vibriolysin exhibited desirable properties heretofore not described for the enzymatic debridement agents, in particular, its selective hydrolysis of dead but not viable tissue, debridement in the absence of bleeding, compatibility with adjunct therapies, and its unique shelf-life stability in a hydrophilic composition at ambient temperature. Appl Environ Microbiol, 1993 Sep, 59(9), 3083 - 90 Bioluminescent sensors for detection of bioavailable Hg(II) in the environment; Selifonova O et al.; Biosensors for the detection of pollutants in the environment can complement analytical methods by distinguishing bioavailable from inert, unavailable forms of contaminants . By using fusions of the well-understood Tn21 mercury resistance operon (mer) with promoterless luxCDABE from Vibrio fischeri, we have constructed and tested three biosensors for Hg(II) . Bioluminescence specified by pRB28, carrying merRo/pT, by pOS14, mediating active transport of Hg(II), and by pOS15, containing an intact mer operon, was measured in rich and minimal media . The highest sensitivities were achieved in minimal medium and were 1, 0.5, and 25 nM Hg(II) for pRB28, pOS14, and pOS15, respectively . The utility of the biosensors in natural waters was demonstrated with freshwater, rain, and estuarine samples supplemented with Hg(II) . mer-lux carried by pRB28 and pOS14 responded to Hg(II) in mercury-contaminated water samples collected from a freshwater pond . Semiquantitative analyses based on light emission in samples collected from the inlet (analytically determined total mercury, approximately 20 nM) and outlet (total mercury, approximately 7 nM) of the pond showed bioavailable mercury at approximately 20 and 1 to 2 nM, respectively . Thus, the biosensors described here semiquantitatively detect bioavailable inorganic mercury (at a nanomolar to micromolar concentration range) in contaminated waters. Southeast Asian J Trop Med Public Health, 1993 Sep, 24(3), 449 - 54 Bacteriophage typing of Vibrio fluvialis; Suthienkul O; Six stable bacteriophages of Vibrio fluvialis were isolated from 44 surface water specimens collected in Thailand and Japan . Twelve different phages types were found among 109 V . fluvialis isolated from feces of diarrheal patients and the environment . Seventy-three percent (80/109) of these 109 isolates were typable with these phages . One phage type, designated as A (1) was predominant and accounted for 43% of the V . fluvialis examined . The six bacteriophages used in this typing scheme were stable for at least during a three-month storage at 4 degrees C . This proposed bacteriophage typing scheme may be of valuable aid in tracing sources and routes of infection in outbreaks of V . fluvialis infection in man. FEMS Microbiol Lett, 1993 Sep 1, 112(2), 237 - 42 A 20-kDa pilus protein with haemagglutination and intestinal adherence properties expressed by a clinical isolate of non-01 Vibrio cholerae; Sengupta TK et al.; A clinical isolate of non-01 V . cholerae (10325) was shown to exhibit higher haemagglutination and intestinal adherence activities in vitro when grown in enriched media, such as trypticase soy broth (TSB) as compared to those of cells grown in a synthetic Tris-buffered or 'T'-medium . A comparison of their cell-surface protein and lipopolysaccharide profiles suggested the involvement of a 20-kDa protein in the cellular adherence process . An antiserum, raised specifically against the 20-kDa protein, recognised pilus structures on the surface of TSB grown cells . Further studies showed that the pilus was morphologically as well as antigenically distinct from toxin coregulated pilus (TCP) or other types of pili expressed by both 01 and non-01 organisms . Inhibition data established the involvement of the 20-kDa protein in haemagglutination as well as intestinal tissue adherence activities of the parent organism. Zh Mikrobiol Epidemiol Immunobiol, 1993 Sep-Oct, (5), 55 - 60 {The determination of the optimal inoculation dose of an oral cholera bivalent chemical vaccine in a controlled trial of the vaccination of children and adolescents}; Sumarokov AA et al.; The reactogenicity and immunological effectiveness of different doses of cholera vaccine for oral administration (in tablets), containing choleragen toxoid, Ogawa and Inaba O-antigens and some of Vibrio cholerae exoenzymes, have been tested on 143 children aged 2-17 years . In this investigation the optimum immunization doses of the preparation have been established: two tablets containing 100,000 binding units (BU) of the toxoid and 2,500 units of O-antigen for children 11-14 years; four tablets containing 200,000 BU of the toxoid and 5,000 units of O-antigen for adolescents aged 15-17 years . An essential advantage of the oral vaccine over vaccines intended for parenteral administration lies in its capacity for inducing the accumulation of secretory antibodies (coproantibodies) in practically all vaccinees. J Biolumin Chemilumin, 1993 Sep-Oct, 8(5), 261 - 6 GroESL proteins facilitate binding of externally added inducer by LuxR protein-containing E . coli cells; Adar YY et al.; htpR- (rpoH, sigma 32 minus) strain of E . coli harbouring the whole lux system of Vibrio fischeri is very dim . We have recently shown that GroESL proteins fully recover the expression of the lux system in this strain . This work has been undertaken to study our assumption that the GroESL proteins stabilize the LuxR protein, thus enhancing the formation of LuxR-Inducer complex . E . coli htpR- cells harbouring the luxR gene were unable to bind extracellularly added inducer, while late logarithmically growing htpR+ strain bound small quantities of the inducer . Reduction in the nutrient content of the growth medium resulted in a large increase in the capability of these cells to bind the inducer . htpR+ or htpR- E . coli strains harbouring both the luxR and the groESL genes bound large quantities of the inducer . The molecular and ecological significance of these results is discussed. J Clin Microbiol, 1993 Sep, 31(9), 2513 - 6 Clonal relationships among classical Vibrio cholerae O1 strains isolated between 1961 and 1992 in Bangladesh; Faruque SM et al.; In Bangladesh, the replacement of classical Vibrio cholerae by the E1 Tor biotype in 1968 and the reappearance of the classical biotype and its coexistence with the E1 Tor biotype after 1982 were never adequately explained . We have analyzed 23 classical V . cholerae isolates collected between 1961 and 1968, 14 classical isolates collected between 1982 and 1992 from the capital city, Dhaka, and 6 classical V . cholerae isolates collected from two southern districts of Bangladesh and studied restriction endonuclease cleavage patterns of rRNA genes (ribotypes) to investigate the clonal relationships among the isolates . Southern blots of total DNA digested with restriction enzyme BamHI, BglI, EcoRI, HindIII, or PstI were probed, using a cloned Escherichia coli rRNA operon . While restriction enzymes BamHI, EcoRI, and PstI failed to differentiate the isolates on the basis of ribotyping, BglI and HindIII produced digestion patterns that allowed differentiation . Ribotyping the isolates with BglI and HindIII revealed five different clones (ribotypes IA, IB, IIA, IC, and IIC) of classical vibrios in Bangladesh . Strains belonging to ribotypes IA and IB were isolated in Dhaka before 1968, and one ribotype (IA) was again isolated between 1982 and 1992 . Ribotype IIA was isolated in 1988 and 1989, when both clones (IA and IIA) of classical vibrios coexisted with the EI Tor vibrios . Isolates belonging to ribotypes IC and IIC were collected in the southern districts of Bangladesh and were clearly different from those collected in Dhaka between 1968 and 1992 by ribotyping analysis with BglI . These results support the previous assumption that classical vibrios were never completely replaced in Bangladesh and also demonstrate the existence of more than one genetically different clone of classical V . cholerae. J Clin Microbiol, 1993 Sep, 31(9), 2474 - 82 Epidemiologic application of a standardized ribotype scheme for Vibrio cholerae O1; Popovic T et al.; A standardized scheme of 27 different BglI ribotypes and subtypes of Vibrio cholerae O1 strains is proposed on the basis of data from 214 human and environmental strains isolated in 35 countries and 14 U.S . states over the past 60 years . The ribotype patterns obtained are reproducible and stable over time . Seven different but very similar ribotypes (1a to 1g) were observed among 16 strains of the classical biotype . Twenty ribotypes and subtypes were identified among 198 V . cholerae O1 strains of the El Tor biotype . Six different patterns were found among the strains causing the current seventh pandemic . Strains of ribotype 8 originated only in central African countries, while those of ribotype 3 originated mainly in Asia and the Pacific Islands . The most widely distributed strains were those of ribotype 6, which was subdivided into three very similar but still distinguishable subtypes . The present Latin American epidemic is caused by strains of ribotype 5 . Strains of this ribotype were isolated from several other geographic locations but can be differentiated from the Latin American strains by other molecular methods . Strains associated with two documented environmental reservoirs exhibited three distinct ribotype patterns; those isolated from patients who ate food from the U.S . Gulf waters were all of ribotype 2, while the strains related to the northeast Australian rivers were of ribotypes 9 and 10 . Nontoxigenic V . cholerae O1 strains originating in Latin America and the U.S . Gulf Coast did not form a specific cluster of ribotypes . Ribotyping in combination with other well-defined methods can assist in epidemiologic investigations, helping to trace the movement of strains and to identify their geographic origins. Lancet, 1993 Aug 14, 342(8868), 387 - 90 Large epidemic of cholera-like disease in Bangladesh caused by Vibrio cholerae O139 synonym Bengal . Cholera Working Group, International Centre for Diarrhoeal Diseases Research, Bangladesh; Vibrio damsela . A cause of fulminant septicemia; Section of Infectious Diseases, Hackensack NJ Medical CenterA previously healthy 70-year-old man presented with a rapidly progressive and fulminant infection due to Vibrio damsela after suffering a knife cut while filleting bluefish at the New Jersey Shore . Despite appropriate antibiotic therapy and localized wound exploration, the patient died . To our knowledge, this is the first reported case of V damsela sepsis with simultaneous isolation of the organism from both blood and wound . We are reporting this case to heighten physicians' awareness of this infection and the importance of early management with antibiotics and surgical consultation. J Biol Chem, 1993 Aug 5, 268(22), 16706 - 16 Purification and characterization of a poly(dA-dT) lux-specific DNA-binding protein from Vibrio harveyi and identification as LuxR; Swartzman E et al.; A lux-specific DNA-binding protein was purified to homogeneity from Vibrio harveyi by chromatography on DEAE-Sepharose, DNA-cellulose, Superose 12, and Mono Q . A single polypeptide of M(r) = 23,000 was found on SDS-polyacrylamide gel electrophoresis with an amino-terminal sequence corresponding to that predicted for luxR, a gene that causes a shift in the transcriptional start site from position -123 to -26 base pairs upstream of the initiation codon of luxC in the V . harveyi lux operon and is required for high expression of lux mRNA in recombinant Escherichia coli . Identification of the DNA-binding protein as LuxR was confirmed by showing its absence in V . harveyi luxR-mutants and its synthesis in recombinant E . coli containing V . harveyi luxR . The LuxR protein was shown to bind to two specific (A + T)-rich regions of DNA upstream of the V . harveyi luxC gene: region A, -290 to -253 base pairs, and region B, -170 to -116 base pairs . Synthetic poly(dA-dT) but not poly(dA)-poly(dT) competed with the lux DNA for binding to LuxR suggesting that this protein may be a novel poly(dA-dT)-binding protein in prokaryotes . The LuxR protein inhibited transcription from the -123 promoter in vitro; however, transcription from the -26 promoter was not reconstituted suggesting the possible requirement for other factors in lux gene regulation . LuxR shared sequence identity with two proteins linked to the regulation of enzymes involved in electron transport indicating that it may be a member of a family of regulators of metabolic functions responsible for diverting electrons from the respiratory chain. J Fla Med Assoc, 1993 Aug, 80(8), 536 - 8 Vibrio vulnificus from raw oysters . Leading cause of reported deaths from foodborne illness in Florida; Hlady WG et al.; Seventy-two cases of Vibrio vulnificus infection from raw oysters were reported from 1981-1992; 36 (50%) patients died, making this infection the leading cause of reported deaths from foodborne illness in Florida . The bacterium naturally occurs in coastal waters and may contaminate legally harvested and properly handled shellfish . Infection, usually by ingestion of contaminated raw oysters, can cause severe illness especially in individuals with preexisting liver disease . They are at 80 times greater risk of illness and over 200 times greater risk of death . The case fatality rate (63%) among patients with liver diseases was over 2.5 times the rate (23%) among those without liver disease . Infections usually occurred during the warm weather months and presented as fulminant septicemia, often complicated by necrotizing cutaneous lesions . Early treatment with antibiotics, debridement and amputation when necessary may improve survival . Prevention relies upon educating patients regarding risk and thorough cooking of shellfish. FEMS Microbiol Lett, 1993 Aug 1, 111(2-3), 219 - 24 Very efficient extracellular production of cholera toxin B subunit using Bacillus brevis; Ichikawa Y et al.; We have constructed a very efficient synthesis and secretion system for cholera toxin B subunit (CTB) of Vibrio cholerae 569B using Bacillus-brevis . The constructed expression-secr