Phylogenetic Analysis of Rubella Virus Isolated during a Period of Epidemic Transmission in Italy, 1991–1997

Abstract

To study the molecular epidemiology of rubella virus during endemic transmission, phylogenetic analysis of the nucleotide sequence of the E1 gene was done with 31 isolates collected in northern Italy during 1991-1997, a period spanning 3 epidemics. The viruses segregated into distinct genotypic strains. Cocirculation of genotypic strains was detected; however, each epidemic was associated with specific strains, and strain displacement occurred concomitantly with each epidemic. Although most of the viruses from Italy belonged to rubella genotype I and many were related to viruses isolated concurrently in other European countries, 3 viruses belonged to rubella genotype II, which previously had been isolated only in Asia. Thus, intercontinental importation of viruses also occurred.

Novel Replicon-Based Reporter Gene Assay for Detection of Rubella Virus in Clinical Specimens

“…In terms of rapidity of the replicon-based assay, although cells inoculated with clinical specimens were incubated for 6 to 7 days before transfection with RUBrep/GFP-⌬NotI transcripts to duplicate the virus isolation procedures used previously (11), we have found that cultures inoculated with clinical specimens can be transfected 2 days postinoculation with GFP expression detectable 1 day posttransfection, for a total assay time of 3 days (W.-P. Tzeng, preliminary observations). In addition to virus detection, we showed that the novel replicon-based assay described here is also useful for virus genotyping, which remains a necessary part of the surveillance component of vaccination programs to determine virus distribution and transmission patterns (8,11,22,23). The E1 gene has been used for all molecular epidemiology studies done on RUB thus far, although the region of the E1 gene used in these studies has not been standardized.…”

“…The E1 gene has been used for all molecular epidemiology studies done on RUB thus far, although the region of the E1 gene used in these studies has not been standardized. Sensitive RT-PCR assays developed for direct RUB detection in clinical specimens produce amplicons of 125 to 300 nt (3,(14)(15)(16)(17)24), shorter than the 600-to 1,400-nt amplicons generated from virus isolates used for genotyping (8,11,22,23), although recently a nested RT-PCR assay producing an ϳ500-nt amplicon directly from clinical specimens that could potentially be used for both detection and genotyping was reported (6,21). From cultures inoculated with clinical specimens shown to be positive for RUB by GFP expression after RUBrep/GFP-⌬NotI transfection, we RT-PCR amplified an ϳ500-nt window previously reported to be optimal for genotyping (22) that was RUB genome specific since it did not overlap with the replicon sequence.…”

Sequence and Phylogenetic Analysis of Wild Type Rubella virus isolated in Iran

“…The rubella virus genome is ~10,000 nucleotides and encodes five protein products, including three virion proteins: the C or capsid protein and two envelope glycoproteins, E1 and E2. The E1 gene sequence has been used for genotyping and phylogenetic analysis of rubella virus isolates (2)(3)(4)(5)(6). To date Rubella viruses from Europe, Asia, and North America have been shown for the most part to group in a single genotype (Rubella Genotype I "RGI") that has a maximum diversity at the nucleotide level of ~5%.…”

“…To date Rubella viruses from Europe, Asia, and North America have been shown for the most part to group in a single genotype (Rubella Genotype I "RGI") that has a maximum diversity at the nucleotide level of ~5%. However, a limited number of viruses from Asia, Europe and Africa, formed a distant phylogenetic branch, differing from RGI viruses by 8% to 10%, which was designated Rubella Genotype II (RGII) (4,5,7,8). These two genotypes (RGI and RGII) belong to the single rubella virus serotype (7).…”

“…As part of the surveillance component of these efforts, an understanding of the worldwide molecular epidemiology of rubella virus, which is limited, is necessary. Molecular epidemiologic surveillance provides important information on: origin of the virus, virus strains circulating in the country, and, whether these strains have become endemic in the country (2,4,5,9,12,13). After mass vaccination for rubella infection in Iran, seroepidemiological studies have been performed (11,14,15), but there is no molecular data for specific characterization of virus particularly virus genotyping up to now (Fig.…”