Bacterial Outer Membrane Proteins ; chemistry ; genetics ; physiology ; Bacteriophages ; chemistry ; Humans ; Receptors, Virus ; chemistry ; Vibrio cholerae ; virology

OBJECTIVETo investigate the inherent characteristics of Vibrio cholerae (V. cholerae) and other vibrios and their relationship.

METHODSPolymerase chain reaction (PCR), DNA sequence analysis, randomly amplified polymorphic DNA (RAPD) analysis and average linkage cluster analysis were used to study 3 isolates of V. cholerae strains O139, three isolates O1 biotype El Tor, four isolates O1 biotype classical and 3 other vibrios.

RESULTSV. cholerae O139 contained the genomic sequences of ctx A2-B as well as V. cholerae O1. V. cholerae and others vibrios were divided into 4 groups by fingerprint patterns of RAPD, that is (1) V. cholerae O139 and V. cholerae O1 El Tor; (2) V. cholerae O1 classical; (3) V. paraheamolyticus and V. vulnificus and (4) V. flluvialis. V. cholerae O139 DNA fingerprint of RAPD was consistent with the El Tor biotype: average linkage cluster distance was 0, and slightly different from the classical biotype, with a distance of 2.07. It was much more different from vibrio paraheamolyticus and others, with a distance of 6.76 - 8.54.

CONCLUSIONV. cholerae and other vibrios are polymorphic in inherent characteristics. The inherent characteristics of V. cholerae O139 are the same as El Tor biotype. O139 may have evolved from the El Tor biotype. The inherent characteristics of vibrio paraheamolyticus are the same as vibrio vulnificus.

DNA, Bacterial ; chemistry ; genetics ; Polymorphism, Genetic ; Random Amplified Polymorphic DNA Technique ; Sequence Analysis, DNA ; Species Specificity ; Vibrio ; genetics ; Vibrio cholerae ; genetics

A real time PCR assay for the detection of Vibrio parahaemolyticus in seafood samples was developed using a novel specific target and a competitive internal amplification control (IAC). The specificity of this assay was evaluated using 390 bacterial strains including V. parahaemolyticus, and other strains belonging to Vibrio and non-Vibrio species. The real time PCR assay unambiguously distinguished V. parahaemolyticus with a detection sensitivity of 4.8 fg per PCR with purified genomic DNA or 1 CFU per reaction by counting V. parahaemolyticus colonies. The assays of avoiding interference demonstrated that, even in the presence of 2.1 μg genomic DNA or 10(7) CFU background bacteria, V. parahaemolyticus could still be accurately detected. In addition, the IAC was used to indicate false-negative results, and lower than 94 copies of IAC per reaction had no influence on the detection limit. Ninety-six seafood samples were tested, of which 58 (60.4%) were positive, including 3 false negative results. Consequently, the real time PCR assay is effective for the rapid detection of V. parahaemotyticus contaminants in seafood.
ATP-Binding Cassette Transporters ; genetics ; DNA Primers ; chemistry ; metabolism ; Food Microbiology ; methods ; Genome, Bacterial ; Real-Time Polymerase Chain Reaction ; Seafood ; microbiology ; Vibrio ; genetics ; isolation & purification ; Vibrio parahaemolyticus ; genetics ; isolation & purification

Vibrio vulnificus cytolysin (VVC) has been implicated as one of the important virulence determinants of V. vulnificus that causes serious septicemia and wound infection. An attempt was made to investigate that VVC could act as a ligand which stimulates intracellular signaling systems. Cholesterol dose-dependently blocked VVC hemolytic activity through oli-gomerization of cytolysin. Among cholesterol derivatives including 7-dehydrocholesterol, cholesteryl esters, deoxycholate, and cholestane tested, only 7-dehydrocholesterol induced oligomerization as well as inactivation of VVC. These results show that oligomerization of VVC is completely dependent on three-dimensional structure of cholesterol where specific interaction of cholesterol at oligomerization sites of VVC is very selective. These findings support the idea that cholesterol which constitute many of cellular plasma membrane could be a receptor of VVC on plasma membrane of target cells.
Animals ; Bacterial Toxins/antagonists & inhibitors/chemistry/metabolism ; Cholesterol/chemistry/*pharmacology ; Cytotoxins/antagonists & inhibitors/*chemistry/*metabolism ; Dehydrocholesterols/chemistry/pharmacology ; Dose-Response Relationship, Drug ; Erythrocytes/drug effects ; Hemolysis/drug effects ; Mice ; Molecular Structure ; Signal Transduction ; Substrate Specificity ; Vibrio/*chemistry

OBJECTIVETo isolate novel actinomycetes and to evaluate their antibacterial activity.

METHODSThree soil samples were collected from Vengodu (village) in Kanchipuram district, Tamil Nadu, India. Actinomycetes were isolated using serial dilution and plating method on actinomycetes isolation agar.

RESULTSTotally 35 isolates were obtained on the basis of colony characteristics on actinomycetes isolation agar. All the isolates were screened for antibacterial activity by cross streak method. Medium and optimization of day were done for the potent strains using Nathan's agar well diffusion method. Isolation of bioactive compounds from significant active isolates was done by using different media. The most active isolate VAS 10 was identified as Actinobacterium Loyola PBT VAS 10 (accession No. JF501398) using 16s rRNA sequence method. The hexane, ethyl acetate, dichloromethane and butanol extracts of VAS 10 were tested against bacteria. The maximum antibacterial activity was observed in dichloromethane and ethyl acetate; maximum zones of inhibition were observed against Enterococcus durans. The rRNA secondary structure and the restriction sites of Actinobacterium Loyola VAS 10 were predicted using Genebee and NEBCutter online tools respectively.

CONCLUSIONSThe present study showed that among the isolated actinomycetes, Actinobacterium Loyola PBT VAS 10 (accession No. JF501398) showed good antibacterial activity against the tested bacteria.

Actinobacteria ; chemistry ; isolation & purification ; physiology ; Animals ; Anti-Bacterial Agents ; pharmacology ; Antibiosis ; physiology ; Bacillus subtilis ; drug effects ; Enterobacter aerogenes ; drug effects ; Escherichia coli ; drug effects ; India ; Microbial Sensitivity Tests ; Phylogeny ; RNA, Ribosomal, 16S ; genetics ; Soil Microbiology ; Species Specificity ; Vibrio parahaemolyticus ; drug effects