ABSTRACT
AIM: Selection of perspective targets as a base for development of test systems for the detection of legionella DNA in study material by PCR with electrophoresis and hybridization-fluorescent accounting of the results. MATERIALS AND METHODS: 22 Legionella pneumophila, 3 Legionella spp. strains and 30 cultures ofheterologic microorganisms, clinical material and environmental object samples were used in the study. Genome analysis was carried out by using Mega 3.1 program. Primer selection was conducted by using Primer Express program and BLAST algorithm and TaqMan format probes on the website www.genscript.com. RESULTS: Analysis of 712 legionella nucleotide sequences for the presence of novel species-specific and conservative for L. pneumophila loci was carried out. Fragments of life-support genes were selected for the analysis: fliC, mompS, ftsZ, dotA, dnaX, trpS, rpoB, rnp, proA, gspA. The most perspective DNA targets were established to be ftsZ and mompS genes. Based on the selected loci, PCR test systems were constructed for the detection of DNA oflegionella causative agent in biological material and environment objects, their diagnostic value was characterized. CONCLUSION: The studies carried out have shown the perspective of use of life-support ftsZ and mompS genes for the construction of novel preparations for legionellosis genetic diagnostics.
Subject(s)
DNA, Bacterial/genetics , Genes, Bacterial , Legionella pneumophila/genetics , Legionnaires' Disease/diagnosis , Legionnaires' Disease/genetics , Real-Time Polymerase Chain Reaction/methods , Female , Humans , Male , Sensitivity and SpecificityABSTRACT
Using developed scheme, complex study of protective properties of avirulent recombinant strain Vibrio cholerae El Tor Inaba KM 184 was performed. Necessity for broadening of standard procedure of testing of cholera vaccines protective properties by using of quantitative methods of assessment of morphological changes and state of biomodel's functional systems, which could increase the information value of assessment of studied vaccines, was experimentally substantiated.
Subject(s)
Cholera Vaccines/administration & dosage , Cholera/prevention & control , Cholera/physiopathology , Evaluation Studies as Topic , Vaccination , Vibrio cholerae O1/immunology , Administration, Oral , Animals , Cholera Toxin/biosynthesis , Cholera Toxin/genetics , Peptide Fragments/biosynthesis , Peptide Fragments/genetics , Rabbits , Vaccines, Attenuated/administration & dosage , Vaccines, Synthetic/administration & dosage , Vibrio cholerae O1/metabolism , Viscera/physiopathologyABSTRACT
Information on V. cholerae eltor isolated in the focus of cholera in Kazan in 2001 at different periods of the outbreak is presented. The identity of strains isolated from patients, vibriocarriers and environmental objects, including their antibioticograms (sensitivity to cyprofloxacin and resistance to trimethoprim--sulfamethoxazole, streptomycin, furazolidone and nalidixic acid, which may be regarded as markers), is shown. Variable tandem repetitions in the DNA of 30 isolates strains of different origin have been determined. The results of this determination make it possible to classify all these strains as one genotype, which confirms the suggestion on the circulation of one subclone of the infective agent of cholera in the focus. As revealed in this investigation, the isolated strains are labile with respect to diagnostic phage eltor, while ctx+ strains are resistant to phage eltor ctx+.
Subject(s)
Cholera/metabolism , Vibrio cholerae/genetics , Vibrio cholerae/metabolism , Anti-Bacterial Agents/metabolism , Anti-Bacterial Agents/therapeutic use , Biomarkers , Cholera/drug therapy , Cholera/epidemiology , Ciprofloxacin/therapeutic use , Drug Resistance, Microbial/genetics , Furazolidone/metabolism , Genotype , Humans , Nalidixic Acid/metabolism , Russia/epidemiology , Streptomycin/metabolism , Sulfamethoxazole/metabolism , Trimethoprim Resistance/genetics , Vibrio cholerae/classification , Vibrio cholerae/drug effects , Vibrio cholerae/isolation & purificationABSTRACT
V. cholerae infection with clearly pronounced diarrhea was reproduced in adult rabbits with the use of the RITARD system. The state of the APUD system of the intestine of the animals and morphological changes in internal organs in experimental cholera were described. As noted in this study, the manifestation of changes in the intestine and other organs, as well as the reaction of apudocytes to V. cholerae infection, were linked with the intensity of diarrhea and the time of the death of the animals.
Subject(s)
APUD Cells/physiology , Cholera/physiopathology , Intestines/physiopathology , APUD Cells/pathology , Animals , Cell Count , Cholera/pathology , Diarrhea/pathology , Diarrhea/physiopathology , Disease Models, Animal , Female , Intestines/pathology , Male , Rabbits , Time FactorsABSTRACT
Experiments were conducted on animals. Following immunization with cholera vaccine there were revealed vibriocidal antibodies, sometimes -- agglutinating, but no vibriolysins in the cells of the mucosal epithelium of the small intestine, of the liver, the spleen and in the fractions obtained from the cells with the aid of differential centrifugation. This corresponds to the high titre of vibriocidal antibodies in the serum and a relatively low titre of the other types of antibodies. In parenteral immunization with various doses of cholera vaccine the concentration of vibriocidal antibodies reached 10(1)-10(7) per 1 mg of protein, which was millions of times less than in the blood serum.
