Revealing new measles virus transmission routes by use of sequence analysis of phosphoprotein and hemagglutinin genes.

With improved measles virus (MV) control, the genetic variability of the MV-nucleoprotein hypervariable region (NP-HVR) decreases. Thus, it becomes increasingly difficult to determine the origin of a virus using only this part of the genome. During outbreaks in Europe and Africa, we found MV strains with identical NP-HVR sequences. However, these strains showed considerable diversity within a larger sequencing window based on concatenated MV phosphoprotein and hemagglutinin genes (P/H pseudogenes). In Belarus, Germany, Russia, and the Democratic Republic of Congo, the P/H pseudogenes provided insights into chains of transmission, whereas identical NP-HVR provided none. In Russia, for instance, the P/H pseudogene identified temporal clusters rather than geographical clusters, demonstrating the circulation and importation of independent variants rather than large local outbreaks lasting for several years, as suggested by NP-HVR. Thus, by extending the sequencing window for molecular epidemiology, a more refined picture of MV circulation was obtained with more clearly defined links between outbreaks and transmission chains. Our results also suggested that in contrast to the P gene, the H gene acquired fixed substitutions that continued to be found in subsequent outbreaks, possibly with consequences for its antigenicity. Thus, a longer sequencing window has true benefits both for the epidemiological surveillance of measles and for the better monitoring of viral evolution.

Phylogenetic analysis of human parvovirus b19 sequences from eleven different countries confirms the predominance of genotype 1 and suggests the spread of genotype 3b.

Phylogenetic analysis of 166 human parvovirus B19 sequences from 11 different countries attributed 91.57% to genotype 1, 5.42% to genotype 3b, and 3.01% to genotype 3a. Very similar viruses of genotype 1 circulated widely in Europe and Israel. Genotype 3b seems to show an increasing spread outside of Africa.

Genotyping of recent measles virus strains from Russia and Vietnam by nucleotide-specific multiplex PCR.

The nucleoprotein genes of 49 measles virus (MV) strains circulating in Russia between 2000 and 2006 and in Vietnam in 2003 were analyzed by genotype-specific PCR and the results were compared with their sequences. The sequences revealed the presence of genotypes H1 and H2 in the center (Nha Trang) and the north (Hanoi) of Vietnam, respectively. The relative diversity of the H2 strains suggested an endemic circulation of these viruses in the capital. In contrast genotype H1 strains from Nha Trang were homogenous genetically, which may indicate a recent importation. The strains obtained from 12 different regions of the Russian Federation were assigned to the genotypes H1, D4, and D6. Most strains (81.4%) were correctly genotyped by a multiplex PCR method which was sensitive to genotype-specific mutations [Kremer et al. (2004): J Clin Microbiol 42: 3017-3022]. Ambiguous or negative results for some clade H and genotype D6 strains were due to point mutations in the type-specific primer binding sites. After exchanging a single nucleotide in both the clade H- and the genotype D6-specific primers, all strains were assigned correctly to their genotype. A simplified procedure for use in Vietnam was developed to distinguish directly between genotypes H1 and H2 and any non-H genotype. These results demonstrate that our multiplex PCR method can be adapted easily to new sequence variants or specific epidemiological situations, and thus be very useful for rapid genotyping of large number of samples even in laboratories which do not have sequencing facilities.

Antigenic Variation and Some Biological Properties of Wild-Type Measles Virus Strain Buk and Vaccine Strain L-16.

In some measles virus strains protein synthesis is restricted or reduced at higher temperature incubation. But in wild-type Buk strain and L-16 vaccine strain no changes in viral protein synthesis at 40 degrees C were shown. Radiolabelled Buk and L-16 proteins were precipitated with reconvalescent sera, vaccinated children's sera, subacute sclerosing panencephalitis patients' sera and rabbit antisera. There are no differences in the autoradiograms. However, antigenic variations in Buk strain proteins were discovered by using mAbs in radioimmunoprecipitation analysis. Buk strain hemagglutinin (H protein) has low binding ability in epitopes II and III in comparison with L-16 strain. Analogous changes were found in the epitope II of Buk strain phosphoprotein. Nucleocapsid, fusion and membrane proteins of the two strains have the same interactions with mAbs. In order to investigate H protein affinity with antihemagglutinins and receptors for erythrocytes a modified test for HA-inhibiting antibodies was used. But incubation time for sera and tween-ether antigens was varied from 0 min to 1.5 h. In "0 min" incubation time hemagglutinin can bind with antihemagglutinins or receptors for erythrocytes. But H protein binds more readily with antibodies. Buk strain H protein revealed antihemagglutinins in higher titers than L-16 strain antigen.