RESUMO
S5 (ATCC No. 51352-B2), a Vibrio cholerae O1 ElTor typing phage was characterized. The growth characteristics and inactivation kinetics (thermal, UV and pH) of this lytic phage were investigated. Phage morphology was examined by electron microscopy and was classified as belonging to the family Podoviridae. The S5 phage genome is shown to be a linear double-stranded 39-kb-long DNA as determined by electron microscopy and restriction digestion. Partial denaturation maps were constructed and were used to show that the DNA is non-permuted and terminally redundant. The replication origin of this T7-like phage was visualized by electron microscopy. The polarity of packaging of S5 DNA in the phage head was determined. SDS-PAGE of phage S5 shows two major structural polypeptides of 50 and 42 kDa. A 3D structure of the phage head was reconstructed at a resolution of 37 A using Cryo-EM and a single-particle reconstruction technique.
Assuntos
Bacteriófagos , Vibrio cholerae O1/virologia , Tipagem de Bacteriófagos , Bacteriófagos/química , Bacteriófagos/classificação , Bacteriófagos/genética , Bacteriófagos/fisiologia , Bacteriófagos/ultraestrutura , Microscopia Crioeletrônica , Enzimas de Restrição do DNA/metabolismo , DNA Viral/análise , DNA Viral/ultraestrutura , Genes Virais , Temperatura Alta , Concentração de Íons de Hidrogênio , Processamento de Imagem Assistida por Computador , Imageamento Tridimensional , Modelos Moleculares , Peso Molecular , Podoviridae/química , Podoviridae/classificação , Podoviridae/genética , Podoviridae/fisiologia , Podoviridae/ultraestrutura , Raios Ultravioleta , Vibrio cholerae O1/classificação , Proteínas Virais/análise , Proteínas Virais/química , Virologia/métodos , Inativação de VírusRESUMO
Comparative analysis of CTXphi prophage genome of 366 V. cholerae El Tor strains isolated from infected people and water was carried out using the polymerase chain reaction. Four groups of vibrios, which carry different combinations of ctxA, zot, and ace genes from core region of CTXphi prophage coding key (cholera enterotoxin) and accessory (Zot and Ace toxins) pathogenicity factors, were determined: ctxA(+) zot(-) ace(+), ctxA(-) zot(+) ace(+), ctxA(-) zot(+) ace(-), ctxA(-) zot(-) ace(+). Vibrios that had lost all tested genes were also revealed. Genomic rearrangements occurring in water environment in virulent V. cholerae strains, which acquired foreign pathogenicity genes necessary for their existence in human organism, were proposed as one of the mechanisms of formation of clones with an incomplete or no prophage. Infection process in model animals challenged with wild and isogenic strains of V. cholerae differing in the set of the phage genes (ctxA, zot, and ace) was comparatively analyzed. It was shown that variability of CTXphi prophage genome was an important factor of modification of cholera vibrios virulent characteristics. Obtained data point to usefulness of ctxA, zot, and ace phage genes detection in wild V. cholerae isolates as it could permit evaluation of their virulent potential determining the severity of the infection.
Assuntos
Bacteriófagos/genética , Cólera/microbiologia , Genoma Viral , Prófagos/genética , Vibrio cholerae O1/virologia , Microbiologia da Água , Animais , Toxina da Cólera/genética , Endotoxinas , Variação Genética , Ilhas Genômicas/genética , Humanos , Reação em Cadeia da Polimerase , Coelhos , Vibrio cholerae O1/genética , Vibrio cholerae O1/patogenicidade , Proteínas do Core Viral/genética , VirulênciaRESUMO
Phage N5 is one of the phages of Vibrio cholerae serovar O1 biotype El Tor (Ghosh, A. N., Ansari, M. Q., and Dutta, G. C. Isolation and morphological characterization of El Tor cholera phages. J. Gen. Virol. 70: 2241-2243, 1989). In the present communication the growth curve, molecular weight and confirmation of the genome, partial denaturation map and restriction endonuclease digestion pattern have been determined. Partial denaturation map indicates that the genome has non-permuted / invariant sequence. Presence of cohesive ends has also been documented.