The seroprevalence of antibodies to HEV was determined in three different Italian populations (volunteer blood donors, normal healthy individuals, and intravenous drug users [IVDUs]). The seroprevalence in the populations ranged from 0.74% to 1.94%, with higher rates among IVDUs and individuals over 40 years of age. None of 35 patients diagnosed with acute hepatitis A but 10 of 153 (6.5%) patients with acute nonA, nonB, nonC hepatitis were found to be positive for anti-HEV antibody. One of these antibody positive cases was linked to travel, while the remaining 9 were not associated with travel to endemic areas. These data suggest a possible low circulation of HEV in Italy.

Hepatitis C virus (HCV) and human pegivirus (HPgV), formerly GBV-C, are the only known human viruses in the Hepacivirus and Pegivirus genera, respectively, of the family Flaviviridae. We present the discovery of a second pegivirus, provisionally designated human pegivirus 2 (HPgV-2), by next-generation sequencing of plasma from an HCV-infected patient with multiple bloodborne exposures who died from sepsis of unknown etiology. HPgV-2 is highly divergent, situated on a deep phylogenetic branch in a clade that includes rodent and bat pegiviruses, with which it shares <32% amino acid identity. Molecular and serological tools were developed and validated for high-throughput screening of plasma samples, and a panel of 3 independent serological markers strongly correlated antibody responses with viral RNA positivity (99.9% negative predictive value). Discovery of 11 additional RNA-positive samples from a total of 2440 screened (0.45%) revealed 93-94% nucleotide identity between HPgV-2 strains. All 12 HPgV-2 RNA-positive cases were identified in individuals also testing positive for HCV RNA (12 of 983; 1.22%), including 2 samples co-infected with HIV, but HPgV-2 RNA was not detected in non-HCV-infected individuals (p<0.0001), including those singly infected by HIV (p = 0.0075) or HBV (p = 0.0077), nor in volunteer blood donors (p = 0.0082). Nine of the 12 (75%) HPgV-2 RNA positive samples were reactive for antibodies to viral serologic markers, whereas only 28 of 2,429 (1.15%) HPgV-2 RNA negative samples were seropositive. Longitudinal sampling in two individuals revealed that active HPgV-2 infection can persist in blood for at least 7 weeks, despite the presence of virus-specific antibodies. One individual harboring both HPgV-2 and HCV RNA was found to be seronegative for both viruses, suggesting a high likelihood of simultaneous acquisition of HCV and HPgV-2 infection from an acute co-transmission event. Taken together, our PLOS Pathogens |

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Sequences from the putative 5 nontranslated region of GB virus A were isolated from mystax, owl monkeys, and tamarins. Though sequences of isolates from each animal species are virtually identical at the nucleotide level (95%), isolates from different species are dramatically different (52 to 79% identical) and genetically cluster on this basis. The GB hepatitis agent was initially characterized in nonhuman primates inoculated with the blood serum of a 34-yearold surgeon experiencing acute clinical hepatitis (2). Following 12 serial animal passages, two distinct viral genomes were cloned from the serum of an animal with acute hepatitis (9). On the basis of genome size and organization, GB virus A (GBV-A) and GBV-B appear to be unique members of the Flaviviridae, most closely related to, but distinct from, the hepatitis C virus (HCV) group (4). Subsequent studies have demonstrated that GBV-B, in the absence of GBV-A, can induce hepatitis in the tamarin animal model (7). However, no significant elevations in serum liver enzyme levels were noted in tamarins singly inoculated with GBV-A, despite persistence of the virus in serum (7). Additionally, several animals were found to have a variant of GBV-A present in their serum prior to inoculation. Sequences within the putative 5Ј nontranslated region (NTR) of this variant were virtually identical to one another, though they were only 79% identical to those in the original GBV-A isolate (8). In the present study, we have identified GBV-A variants in mystax (GBV-A mx), owl monkeys (GBV-A tri), and tamarins (GBV-A lab) not known to have been inoculated with an infectious agent. These sequences are 52 to 79% identical at the nucleotide level to GBV-A and genetically cluster on the basis of the primate species from which they were isolated. Blood sera from Saguinus labiatus (tamarins), Saguinus mystax (mystax monkeys), and Aotus trivirgatus (owl monkeys) were used as source material for nucleic acids. Briefly, 25-to 50-l volumes were extracted with the DNA/RNA isolation kit (U.S. Biochemicals, Cleveland, Ohio) or the QIAamp HCV kit (Qiagen Inc., Chatsworth, Calif.). Random hexamer-primed nucleic acids were reversed transcribed as directed by the manufacturer with the RNA PCR kit (Perkin-Elmer, Foster City, Calif.) in a volume equal to that of the initial serum. Five microliters of the reverse transcription reaction mixture was used in a 25-l PCR mixture with 2.0 mM MgCl 2 and 0.5 M (each) oligonucleotide primer (8): 5ЈvA-s (5Ј-AGGGTTCGT AGGTGGTAAATCCC-3Ј) and 5ЈvA-a (5Ј-TGCCACCAGG GGTCACCCGAAG-3Ј). Amplification was performed by "touchdown" PCR (6) for 43 cycles (94ЊC