Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:3.1.30.2 (endonuclease)
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Four simple methods, i.e., (i) UV absorption spectrophotometry, (ii) hydroxyapatite chromatography, (iii) fluorescence analysis of ethidium bromide bound to DNA and (iv) assay of S1 endonuclease action, were used in parallel for the estimation of furazolidone-induced inter-strand cross-links in Vibrio cholerae DNA. The data produced by the four methods were in reasonable agreement with each other and provided similar linear dose-response relations, the correlation (between dose and response) coefficient being in any case numerically greater than 0.98. When the data obtained by four independent methods were plotted in a single graph, the resulting dose-response relation could be described by the equation log NR = 1.41 - 0.54 log D, where NR is the % non-reversible DNA remaining in the cells treated by furazolidone at dose D micrograms/ml x h. The correlation coefficient in this plot was -0.98 and significant to a level better than 0.1%. This study thus brings out that any one of these four methods can be used with reasonable confidence for the diagnosis and assay of inter-strand cross-links in DNA.
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PMID:Inter-strand cross-linking of Vibrio cholerae DNA induced by furazolidone: a quantitative assay by four simple methods. 787 98

Isolates of Vibrio cholerae O1 El Tor from two well-defined cholera outbreaks in Malaysia were analyzed by using pulsed-field gel electrophoresis (PFGE). Isolates from sporadic cases occurring during the same time period were also studied. Digestion of chromosomal DNA from these isolates of V. cholerae O1 with restriction endonucleases NotI (5'-GCGGCCGC-3') and SfiI (5'-GGCCNNNN-3'), followed by PFGE, produced restriction endonuclease analysis (REA) patterns consisting of 13 to 24 bands (ranging in size from 46 to 398 kbp). Analysis of the REA patterns generated by PFGE after digestion with NotI and SfiI suggested the clonal nature and close genetic identity of the isolates obtained during each of the two outbreaks (Dice coefficient, 0.93 to 1.0). Although they had very similar REA patterns, the two outbreak clones were not identical. Isolates of V. cholerae O1 from sporadic cases, on the other hand, appeared to be much more heterogeneous (five different REA patterns detected in the five isolates tested; Dice coefficient, 0.31 to 0.81) than those obtained during the two outbreaks. We conclude that PFGE of V. cholerae O1 chromosomal DNA digested with infrequently cutting restriction endonucleases is a useful method for molecular typing of V. cholerae isolates for epidemiological purposes.
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PMID:Molecular epidemiologic analysis of Vibrio cholerae O1 isolates by pulsed-field gel electrophoresis. 788 85

The conventional phage typing scheme proposed by S. Basu and S. Mukerjee (Experientia 24:299-300, 1968) has been used routinely for identification of the strains at the Vibrio Phage Reference Laboratory since 1968. However, because of limitations of this scheme, a new phage typing scheme using five newly isolated phages was incorporated into the conventional scheme. A different definition of routine test dilution (almost confluent lysis) was found to be more useful than the one previously used (confluent lysis). The 1,000 strains tested could be clustered into 27 types with the five new phages. With the new scheme of 10 phages (5 new phages and 5 phages of Basu and Mukerjee), the 1,000 strains could be grouped into 146 types. The new phages were different from each other and also from those of Basu and Mukerjee, as revealed by lytic pattern, electron microscopy, restriction endonuclease digestion, sodium dodecyl sulfate-polyacrylamide gel electrophoresis, and antiphage antiserum studies. With the new typing scheme, 99.6% of the strains were typeable. Phage type 115 was the most common and includes 119 (11.9%) of the 1,000 strains tested. Next most common were phage types 142 (9.4%), 143 (7.0%), 104 and 116 (both 5.4%), 3 (5.3%), 5 (4.1%), 4 (3.9%), 24 (2.1%), and 100 (1.7%). The larger number of types would be useful for further classification of the strains for epidemiological purposes. This newly developed scheme is highly applicable to, and could be widely adopted for, phage typing of Vibrio cholerae O1 biotype El Tor strains.
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PMID:New phage typing scheme for Vibrio cholerae O1 biotype El Tor strains. 831

Cholera is endemic in Bangladesh, and a regular seasonal pattern of cholera epidemics occurs. We examined the clonal relationships among 103 clinical and environmental Vibrio cholerae isolates belonging to O1, O139, or non-O1 non-O139 serogroups isolated during epidemic and interepidemic periods in Bangladesh and compared them with those of 51 V. cholerae isolates from four countries in Asia and Africa. These studies were done by a computer-assisted numerical analysis of the restriction endonuclease cleavage patterns of rRNA genes (ribotypes). Unweighed pair-group cluster analysis of BglI- and HindIII-generated band patterns revealed 16 clusters. Ribotypes were defined as clusters of strains possessing > 98% similarity. The results showed that 154 isolates could be differentiated into 15 different ribotypes, and strains belonging to 3 of these ribotypes (ribotypes I, V, and VIIIA and VIIIB) were isolated more frequently during the epidemic periods than during interepidemic periods in Bangladesh. Classical vibrios belonged to six different ribotypes (ribotypes I to VI), with a mean similarity coefficient of 0.84, and the El Tor vibrios belonged to five different ribotypes (ribotypes VIIIA and IX to XII), with a mean similarity coefficient of 0.82. A single clone of El Tor vibrios (ribotype XII) was resident in Tanzania, whereas Nigeria, Syria, and India shared toxigenic El Tor strains with Bangladesh. Cholera toxin (CT)-positive O139 vibrios isolated from Bangladesh and India belonged to a single ribotype (ribotype VIIIB) and were > 98% similar to one of the ribotypes of El Tor vibrios (ribotype VIIIA), but a CT-negative O139 vibrio from Argentina (ribotype XIII) was < 75% similar to the same cluster of El Tor vibrios, thus suggesting more than one possible origin for O139 vibrios. Strains belonging to the same ribotypes (ribotypes VIII to X) were isolated from both patients and surface water in Bangladesh, indicating possible transmission through surface water. A clone of a CT-positive environmental isolate of non-O1 V. cholerae (ribotype VII) was found to be closely related (76.3% similarity) to a clone of classical vibrios (ribotype I) and was only between 27.2 and 56.1% similar to clusters of El Tor, O139, and two other non-O1 nontoxigenic clones.
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PMID:Molecular epidemiology of toxigenic Vibrio cholerae in Bangladesh studied by numerical analysis of rRNA gene restriction patterns. 857 28

Previously the heat-stable enterotoxin in Vibrio cholerae and V. mimicus has been detected by suckling mouse assay, a non-specific approach, and by DNA probes, a time-consuming method. This report describes a polymerase chain reaction (PCR) procedure for the detection of the stn (NAG-ST) and sto (O1-ST) gene sequences that is rapid and specific, allowing toxin gene molecular characterisation. A total of 34 V. cholerae and V. mimicus isolates was examined for ST and CT genes. The NAG-ST gene sequence was amplified in 13 of 22 non-O1/non-O139 V. cholerae and three of five V. mimicus strains. A new enterotoxin gene sequence pattern was found with MseI and TaqI restriction endonuclease PCR fragment digestion of two V. cholerae isolates, in addition to the pattern anticipated from the Genbank sequence, and found with the other ST+. These results show that ST-PCR detection is useful for the characterisation of V. cholerae and V. mimicus.
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PMID:Detection of Vibrio cholerae and V. mimicus heat-stable toxin gene sequence by PCR. 915 35

A total of 173 Vibrio cholerae O1 isolates from the recent cholera epidemic in Colombia was analysed by the polymerase chain reaction (PCR) for the genes encoding the A subunit of cholera toxin (ctxA) and the zonula occludens toxin (zot), and by ribotyping. All isolates were positive for ctx A and zot, which was confirmed by hybridisation. Ribotyping with restriction endonuclease Bg/I digestion of total DNA revealed three ribotypes: B5a comprising 165 (96.4%) isolates, and two new designated ribotypes B20 and B21a in six (3.5%) isolates and two (1.1%) isolates, respectively. These findings have significant public health implications.
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PMID:Molecular epidemiology of Vibrio cholerae O1 isolates from Colombia. 923 46

The unprecedented genesis of a novel non-O1 Vibrio cholerae strain belonging to serogroup O139, which caused an epidemic in late 1992 in the Indian subcontinent, and its subsequent displacement by El Tor O1 vibrios after 18 months initiated a renewed investigation of the aspects of the organism that are related to pathogenesis. The reappearance of V. cholerae O139 with altered antibiotic sensitivity compared to O139 Bengal (O139B) in late 1996 has complicated the epidemiological scenario of V. cholerae and has necessitated an examination of possible rearrangements in the genome underlying such rapid changes in the phenotypic traits. With a view to investigating whether the phenotypic changes that have occurred are associated with alteration in the genome, the genome of the resurgent V. cholerae O139 (O139R) strains were examined. Pulsed-field gel electrophoresis analysis of NotI- and SfiI-digested genomic DNA of O139R isolates showed restriction fragment length polymorphism including in the cholera toxin (CTX) genetic element locus and with O139B isolates. Analyses of the organization of the CTX genetic elements in O139R strains showed that in contrast to two copies of the elements connected by two direct-repeat sequences (RS) in most of the genomes of O139B isolates, the genomes of all O139R strains examined, except strain AS192, have three such elements connected by a single RS. While the RS present in the upstream of the CTX genetic elements in the genome of O139R is of O139B origin, the RS connecting the cores of the elements has several new restriction sites and has lost the BglII site which is supposed to be conserved in all O1 strains and O139B. The endonuclease I-CeuI, which has sites only in the rrn operons in the genomes of all organisms examined so far, has 10 sites in the genomes of O139R strains, compared to 9 in the genomes of O139B strains. The recent isolates of V. cholerae O139 have thus gained one rrn operon. This variation in the number of rrn operons within a serogroup has not been reported for any other organism. The results presented in this report suggest that like the pathogenic El Tor O1 strains, the genomes of O139 strains are undergoing rapid alterations.
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PMID:Resurgent Vibrio cholerae O139: rearrangement of cholera toxin genetic elements and amplification of rrn operon. 986 9

A collection of 89 Vibrio cholerae O1 strains, isolated in Romania between 1977 and 1994, and 6 strains from the Republic of Moldavia, was characterized by ribotyping, toxin gene restriction pattern (toxinogenotype) and distribution of cholera toxin gene (ctx), accessory toxin gene (ace) and zonula occludens toxin gene (zot). After Bg/I endonuclease restriction of chromosomal DNA, a total of 18 ribotypes and 21 toxinogenotypes were distinguished. Deletions in the core region of the toxin gene cassette were found in 20% of strains; however, with the exception of one strain, all the isolates contained the ctx gene. Used in association, the three methods of molecular typing provided an accurate characterization of V. cholerae O1 isolates.
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PMID:Molecular characterization of Vibrio cholerae O1 strains isolated in Romania. 992 81

The scenario of cholera that existed previously changed in 1992 and 1993 with the emergence of toxigenic Vibrio cholerae O139 in India. The genesis of the new serogroup formed the impetus to search for O139 phages in and around the country. A total of five newly isolated phages lytic to V. cholerae O139 strains were used for the development of this phage typing scheme. These phages differed from each other and also differed from the existing O1 phages in their lytic patterns, morphologies, restriction endonuclease digestion profiles, and immunological criteria. With this scheme, 500 V. cholerae O139 strains were evaluated for their phage types, and almost all strains were found to be typeable. The strains clustered into 10 different phage types, of which type 1 (38.2%) was the dominant type, followed by type 2 (22.4%) and type 3 (18%). Additionally, a comparative study of phage types in 1993 and 1994 versus those from 1996 to 1998 for O139 strains showed a higher percentage of phage type 1 (40.5%), followed by type 3 (18.8%) during the period between 1993 and 1994, whereas phage type 2 (32. 1%) was the next major type during the period from 1996 to 1998. This scheme comprising five newly isolated phages would be another useful tool in the study of the epidemiology of cholera caused by V. cholerae O139.
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PMID:Development and evaluation of a phage typing scheme for Vibrio cholerae O139. 1061 61

Pasteurella multocida is an important veterinary and opportunistic human pathogen. The species is diverse and complex with respect to antigenic variation, host predeliction and pathogenesis. Certain serological types are the aetiologic agents of severe pasteurellosis, such as fowl cholera in domestic and wild birds, bovine haemorrhagic septicaemia and porcine atrophic rhinitis. The recent application of molecular methods such as the polymerase chain reaction, restriction endonuclease analysis, ribotyping, pulsed-field gel electrophoresis, gene cloning, characterisation and recombinant protein expression, mutagenesis, plasmid and bacteriophage analysis and genomic mapping, have greatly increased our understanding of P. multocida and has provided researchers with a number of molecular tools to study pathogenesis and epidemiology at a molecular level.
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PMID:The molecular biology of pasteurella multocida. 1069 99


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