Hepatitis E virus (HEV) is a human pathogen with zoonotic spread, infecting both domestic and wild animals. About 17% of the Swedish population is immune to HEV, but few cases are reported annually, indicating that most infections are subclinical. However, clinical hepatitis E may also be overlooked. For identified cases, the source of infection is mostly unknown. In order to identify whether HEV may be spread from wild game, the prevalence of markers for past and/or ongoing infection was investigated in sera and stool samples collected from 260 hunted Swedish wild ungulates. HEV markers were found in 43 (17%) of the animals. The most commonly infected animal was moose (Alces alces) with 19 out of 69 animals (28%) showing HEV markers, followed by wild boar (Sus scrofa) with 21 out of 139 animals (15%), roe deer (Capreolus capreolus) with 2 out of 30 animals, red deer (Cervus elaphus) with 1 out of 15 animals, and fallow deer (Dama dama) 0 out of 7 animals. Partial open reading frame 1 (ORF1) of the viral genomes from the animals were sequenced and compared with those from 14 endemic human cases. Phylogenetic analysis revealed that three humans were infected with HEV strains similar to those from wild boar. These results indicate that wild animals may be a source of transmission to humans and could be an unrecognized public health concern.

Genetic recombination has been suggested to occur in mosquito-borne flaviviruses. In contrast, tick-borne flaviviruses have been thought to evolve in a clonal manner, although recent studies suggest that recombination occurs also for these viruses. We re-analyzed the data and found that previous conclusions on wild type recombination were probably falsely drawn due to misalignments of nucleotide sequences, ambiguities in GenBank sequences, or different laboratory culture histories suggestive of recombination events in laboratory. To evaluate if reliable predictions of wild type recombination of tick-borne flaviviruses can be made, we analyzed viral strains sequenced exclusively for this study, and other flavivirus sequences retrieved from GenBank. We detected genetic signals supporting recombination between viruses within the three clades of TBEV-Eu, TBEV-Sib and TBEV-Fe, respectively. Our results suggest that the tick-borne encephalitis viruses may undergo recombination under natural conditions, but that geographic barriers restrict most recombination events to involve only closely genetically related viruses.

Herpes simplex virus type 1 variants selected by virus propagation in cultured cells in the presence of the sulfated oligosaccharide PI-88 were analyzed. Many of these variants were substantially resistant to the presence of PI-88 during their initial infection of cells and/or their cell-to-cell spread. Nucleotide sequence analysis revealed that the deletion of amino acids 33-116 of gC but not lack of gC expression provided the virus with selective advantage to infect cells in the presence of PI-88. Purified gC (Delta33-116) was more resistant to PI-88 than unaltered protein in its binding to cells. Alterations that partly contributed to the virus resistance to PI-88 in its cell-to-cell spread activity were amino acid substitutions Q27R in gD and R770W in gB. These results suggest that PI-88 targets several distinct viral glycoproteins during the course of initial virus infection and cell-to-cell spread.

Adaptation of some viruses to replication in cultured cells selects variants that due to alterations in the viral attachment proteins convert to using heparan sulfate (HS) as initial receptor. We report that the nucleotide sequence of herpes simplex virus type 1 (HSV-1) glycoprotein C (gC), a principal attachment component of the virus, remained unchanged during adaptation of wild-type strains to cultured cells. Likewise, amino acid residues critical for binding of gC to HS were conserved in viral strains that replicated in vivo in different human tissues. Moreover wild-type HSV-1 strains derived directly from clinical specimens were, similar to their cell culture propagated progeny viruses and common laboratory strains, sensitive to heparin and demonstrated impairment in their ability to infect HS/chondroitin sulfate deficient cells. These results demonstrate that the HS-binding ability is a feature of wild-type strains of HSV-1.

b T ick-borne encephalitis (TBE) is a viral zoonosis of increasing incidence in Eurasia, and diagnosis relies mainly on the detection of IgM antibodies in serum. We here present two cases of TBE, of four patients tested, in which we have detected TBE virus (TBEV) RNA in urine with real-time PCR during the encephalitic phase. This observation suggests a new diagnostic opportunity that could be evaluated in a larger cohort of patients.In the first case we report here, 1 week after a tick bite, a 61-yearold woman had high fever, nausea, and fatigue lasting for 5 to 6 days. She was asymptomatic for a week, after which the fever returned along with vomiting and severe headache. After admission, her condition worsened. She was unable to follow instructions, became somnolent and disoriented, and had signs of dysphasia. A cerebrospinal fluid (CSF) sample showed a leukocyte count of 69 ϫ 10 6 /liter (41% polymorphonuclear and 59% mononuclear) and elevated albumin. Specific TBE virus (TBEV) IgM antibodies were detected in serum drawn on day 2 after admission, and the patient seroconverted as regards IgG during the hospital stay. Before nucleic acid extraction using the EasyMag instrument (bioMérieux, Marcy l=Etoile, France), 250-l amounts of serum, CSF, and urine samples were added to 2 ml of lysis buffer and incubated for 10 min at room temperature. TBEV RNA was detected from urine by TaqMan PCR performed as described previously (1) on day 19 after fever onset (Fig. 1). The patient's condition improved, and she was discharged after 10 days of hospitalization.The second case was a 70-year-old woman who reported several tick bites during the preceding weeks and was admitted 2 days after the onset of high fever, nausea, and fatigue. She was on treatment with immunosuppressive drugs (mycophenolate, tacrolimus, and prednisolone) due to kidney transplantation. She developed paresis of the lower extremities and lost the strength in her hands. A CSF sample showed 488 ϫ 10 6 leukocytes/liter (28% polymorphonuclear and 72% mononuclear), elevated albumin and lactate, and a slightly decreased glucose quotient. TBEV IgM antibodies (but not IgG) were detected in serum. On day 8 after admission, the patient became deeply unconscious, had a generalized status epilepticus, and was intubated. Magnetic resonance imaging (MRI) demonstrated high signals in the amygdala and hippocampus. Her neurological condition deteriorated, and she died 11 weeks after admission. TBEV RNA was detected from urine on day 7 and day 16 after onset ( Fig. 1). The RNA quantities allowed sequencing of 10,432 nt from TBEV RNA amplicons, as