Aim. The construction of recombinant attenuated Bordetella pertussis bacteria of ptxP3 genotype and its genetic and biological stability characteristics. Materials and methods. During construction of recombinant attenuated bacteria of ptxP3 genotype virulent Bordetella pertussis bacteria of ptxP1 genotype were used as a recipient. PtxP1 genotype bacteria are used for whole cell pertussis (wP) vaccine production in Russia. Mutant bacteria B. pertussis 475 were received by crossing-over between chromosome comprising native copy of target sequence and its mutant copy in recombinant suicide plasmid transferred in recipient bacteria by conjugation. Genetically engineered construction of recombinant plasmids was conducted. The structure of modified chromosome locus of attenuated bacteria was determined by PCR and amplification fragments sequence. The stability of structure and characteristics of attenuated bacteria was defined after 15 passages of strains on culture medium and 5 passages in mice. Results. Isogenic attenuated ptxP1 B. pertussis 4M and ptxP3 B. pertussis 4MKS were constructed. These bacteria produce non-toxic pertussis toxin (PT) and do not produce dermonecrotic toxin (DNT). The promoter region of ptx operon contains mutation, typical for «new» genotype of circulating virulent bacteria and increasing PT production. The structure of modified DNA fragments and characteristics of attenuated bacteria did not change while storing and after passages on culture medium and in mice. Conclusion. Recombinant attenuated bacteria B. pertussis 4MKS of «new» ptxP3 genotype are constructed. Application perspectiveness of genetically engineered modification of isogenic B.pertussis bacteria for pertussis vaccines development is shown.
Introduction. The increase in the incidence of whooping cough in children and adults of different age groups justifies the need for their revaccination and the development of new, acceptable for these purposes. This work is devoted to substantiating the design of a clinical trial and describing the results of a comparative study of the safety and tolerability of the drug "GamLPV" with two-fold intranasal administration to healthy adult volunteers using two methods. The choice of the scheme and method of administration of the drug is justified. The serological structure of the population of adults aged 18–40 years living in Moscow and the Moscow region is characterized.Aim. Determination of the safety and tolerability of the drug with a double intranasal administration of the vaccine by drip method and spraying through an actuator.Materials and methods. A randomized placebo-controlled trial included 50 healthy male and female volunteers aged 18 to 40 years who met the inclusion criteria. The volunteers were divided into 2 groups of 25 people: a drip method of administration and spraying through an "actuator". By both methods, the drug was administered twice with an interval of 60 days.Results and discussion. Serological analysis of the population of healthy volunteers at the prescreening stage justified the inclusion in the study of volunteers containing anti-pertussis antibodies in the blood (IgG ≤40 Ed/ml). A comparison of the results of preclinical studies on an experimental model of non-human monkeys and the first phase of a clinical study of GamLPV allowed us to propose two methods of double administration of the drug as a promising vaccination scheme for volunteers. A comparative randomized study shows the safety of using the proposed scheme for vaccination of adult volunteers.Conclusions. Both proposed methods of double administration can be used to plan a multicenter study to research the immunogenicity and protective activity of GamLPV.
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