Identification and Characterization of a Second Novel Human Erythrovirus Variant, A6

Abstract

Parvovirus B19 (B19), currently the only accepted member of the Erythrovirus genus, is the only parvovirus known to be pathogenic in humans. Recently a viral sequence, tentatively termed V9 which showed 11% variability from the published B19 sequences, was described from a patient with aplastic crisis. To search for additional parvovirus variants, we used the new NS1/7.5EC PCR assay whose primers were designed from a conserved region of the B19/V9 sequence and encompasses an MfeI restriction enzyme site that would allow differentiation between B19- and V9-like sequences. Screening of 225 serum and bone marrow samples and 62 plasma pools identified one new atypical parvovirus sequence, A6, from an anemic HIV-positive patient. A6 exhibited 88% similarity to B19 and 92% to V9, compared to >98% correspondence between reported B19 isolates. Based on the genome similarity to B19, an RT-PCR for A6 capsid transcripts was developed and used to test for A6 infectivity of UT7/Epo/S1 cells. Despite high viral titers, A6 viral transcripts were not detected. Thus, although the prevalence of B19 variants probably is low, the true clinical significance remains unknown. Current PCR analyses are unlikely to detect novel variants without the design of specific primers to the A6/V9/B19 common sequences.

Detection and Differentiation of Human Parvovirus Variants by Commercial Quantitative Real-Time PCR Tests

“…The first B19 variant was "V9," discovered by conventional PCR in the serum of a patient with transient aplastic anemia (19). Recently, other B19 virus variants (K71 and A6) were reported by us and others (13,21,25), and a nomenclature based on three genotypes has been suggested (25). Due to DNA sequence divergence, most B19 PCR methods are unlikely to detect these variants.…”

“…Indeed, LaLi-like sequences were found in two sera and V9-like sequences in nine samples (five serum, three bone marrow, and one blood sample), including subtype D91.1 detected in serum. By the same method, Nguyen et al (21) examined 149 serum and 18 bone marrow samples. Of the 29 PCR-positive sera, only one (A6) was of genotype 2, and all of the others of genotype 1.…”

“…In order to identify such high-titer units, a quantitative DNA detection method is required. On the other hand, new variants of the B19 virus have been discovered (13,19,20,21,25), leading to segregation of the species into three distinct genotypes (25) diverging from each other in sequence by Ͼ10%. In consequence, B19 PCR methods might fail to detect such variants due to sequence divergence between primers and viral target DNA.…”

Clinical features and laboratory findings of human parvovirus B19 in human immunodeficiency virus-infected patients

“…Briefly, viral DNA was extracted from serum samples of 88 HIV-positive patients using a QIAamp DNA Blood Mini Kit (QIAGEN, Hilden, Germany) according to the manufacturer’s instructions. For the screening of B19V DNA, PCR was performed using the primers E1905F (nt 1905-1923) and E1987R (nt 2007-1987), which amplify a 102-bp fragment of the NS1 gene as previously described by Nguyen et al (2002), with some modifications (de Azevedo et al 2012). This PCR can routinely detect as few as 20 copies of the B19V DNA (Nguyen et al 2012).…”

“…Parvovirus B19 (B19V), a member of the genus Erythrovirus within the Parvoviridae family, has been grouped into three distinct genotypes: (i) genotype 1, with subtypes 1a (the prototypic virus) and 1b, (ii) genotype 2 (A6 and LaLi strains) and (iii) genotype 3, with subtypes 3a (V9 strains) and 3b (D91.1 strains) (Nguyen et al 1999, 2002, Servant et al 2002, Fauquet et al 2005). …”