ABSTRACT
The genetic typing of measles virus in clinical samples using xMAP technology was applied. The study provided the calculation and application of specific oligonucleotide probes of genotypes D4, D6 and D7 of measles virus. The strain HobO96 genotype A of measles virus as a check sample was used. The technical approaches to the optimization of preparatory work organization and to the process of identification of measles virus genotypes are described. The presence of genotypes D4, D6 and D7 in clinical samples is proved by the sequence analysis. The genetic typing effectiveness of technique of DNA hybridization using xMAP technology on the instrumental base BioPlex (BioRad, USA) is demonstrated.
Subject(s)
DNA, Complementary/analysis , Measles virus/classification , Measles virus/genetics , Branched DNA Signal Amplification Assay/methods , Genotype , Humans , Measles/genetics , Measles/virology , Measles virus/isolation & purification , Molecular Sequence Data , Nucleocapsid Proteins , Nucleoproteins/genetics , Phylogeny , RNA, Viral/isolation & purification , Reverse Transcriptase Polymerase Chain Reaction , Russia , Sequence Analysis, DNA/methods , Viral Proteins/geneticsABSTRACT
BACKGROUND: A number of cases of measles have been reported in the Republic of Belarus despite vaccine coverage of 98%. The absence of information on measles virus genotypes circulating in the Republic of Belarus has made it difficult to asses the situation. OBJECTIVES: The purpose of this study was to isolate and sequence measles virus strains from clinical cases in Minsk, Belarus, and to estimate the role of vaccine failure in those cases. STUDY DESIGN: Between 2001 and 2003 years, 14 measles cases admitted to the Hospital of Infectious Diseases of Minsk were enrolled in our study. Clinical, routine laboratory, as well as serological and virological examinations were carried out. Detection of measles antibodies and IgG avidity testing was performed using commercial test kits. All measles cases were confirmed by RT-PCR and phylogenetically characterized. RESULTS: Only 42.9% of the cases met the WHO laboratory criteria for measles, however, all cases were confirmed by RT-PCR. Most of the measles cases were attributed to secondary vaccine failure (SVF). Phylogenetic analysis revealed the presence of genotype A virus strains in 2001 and 2002 with D6 and D7 genotypes in 2003. CONCLUSIONS: For the first time, MVs were genetically characterized in Belarus. Our results suggest that in a highly vaccinated population, most of measles cases represent vaccine failures and are vaccine-modified. Our results also indicate that confirmation of a clinical diagnosis of vaccine-modified measles requires a combination of serological and virological tests.
Subject(s)
Measles virus/genetics , Measles/virology , Adolescent , Adult , Antibodies, Viral/analysis , Genotype , Humans , Immunoglobulin G/analysis , Immunoglobulin M/analysis , Infant , Measles/epidemiology , Measles/prevention & control , Measles Vaccine/therapeutic use , Measles virus/immunology , Phylogeny , Republic of Belarus/epidemiology , Viral Proteins/geneticsABSTRACT
The plasmid DNA pCDNA3.1-H encoding the N-terminal sequence of the measles hemagglutinin (H) protein was constructed. Virus-specific particles (VSP) containing the plasmid DNA pCDNA3.1-H coated by the spermidine-polyglucin complex. The mice were immunized by VSP. ELISA, HAIT and immunoblot showed a shaping specific humoral response. The sera of immunized animals were proven to neutralize the wild strain of the NOV96 measles virus. The formation of the specific cell immunity was confirmed by erythrocyte proliferation assay and ELISpot. PCR was used to detect the presence of the plasmid DNA in different intestines and tissues of animals after a single immunization. It was not detected at any time interval in the brain, liver, thymus and blood. And it was present on days 7 and 14 in the red bone marrow, spleen, muscular tissue, lungs and fatty tissue. On day 21 the plasmid DNA was not detected in any of the investigated organs.
