Tahyna virus genetics, infectivity, and immunogenicity in mice and monkeys

Abstract

Background: Tahyna virus (TAHV) is a human pathogen of the California encephalitis virus (CEV) serogroup (Bunyaviridae) endemic to Europe, Asia, and Africa. TAHV maintains an enzootic life cycle with several species of mosquito vectors and hares, rabbits, hedgehogs, and rodents serving as small mammal amplifying hosts. Human TAHV infection occurs in summer and early fall with symptoms of fever, headache, malaise, conjunctivitis, pharyngitis, and nausea. TAHV disease can progress to CNS involvement, although unlike related La Crosse virus (LACV), fatalities have not been reported. Human infections are frequent with neutralizing antibodies present in 60-80% of the elderly population in endemic areas.

Results: In order to determine the genomic sequence of wild-type TAHV, we chose three TAHV isolates collected over a 26-year period from mosquitoes. Here we present the first complete sequence of the TAHV S, M, and L segments. The three TAHV isolates maintained a highly conserved genome with both nucleotide and amino acid sequence identity greater than 99%. In order to determine the extent of genetic relatedness to other members of the CEV serogroup, we compared protein sequences of TAHV with LACV, Snowshoe Hare virus (SSHV), Jamestown Canyon virus (JCV), and Inkoo virus (INKV). By amino acid comparison, TAHV was most similar to SSHV followed by LACV, JCV, and INKV. The sequence of the GN protein is most conserved followed by L, N, GC, NSS, and NSM. In a weanling Swiss Webster mouse model, all three TAHV isolates were uniformly neurovirulent, but only one virus was neuroinvasive. In rhesus monkeys, the virus was highly immunogenic even in the absence of viremia. Cross neutralization studies utilizing monkey immune serum demonstrated that TAHV is antigenically distinct from North American viruses LACV and JCV.

Conclusions: Here we report the first complete sequence of TAHV and present genetic analysis of new-world viruses, LACV, SSHV, and JCV with old-world viruses, TAHV and INKV. Using immune serum generated in monkeys against TAHV, LACV, and JCV, we have demonstrated cross-neutralization within the CEV serogroup. Such cross reactivity may complicate virus identification, especially following JCV infection which elicited antibodies that cross neutralized both LACV and TAHV. These data also suggest that a single vaccine could generate a cross-neutralizing antibody response which may provide protection against CEV serogroup viruses from a wide geographic range.

Figure 1

Alignment of 3' (A) and 5' (B) untranslated regions of the S, M, and L genome segments of TAHV (cDNA presented). for each genome segment the consensus sequence consists of two or more sequences sharing the same nucleotide at a given position. Underlined sequence indicates region known to be conserved among orthobunyaviruses. Since differences in the 3' or 5' UTR of TAHV/58/CZ-cl(1) or TAHV/58/CZ-cl(2) were not observed, only one sequence is presented.

Figure 2

Growth kinetics of TAHV in Vero and C6/36 cells. Replication of TAHV/58/CZ-cl (1), TAHV/58/CZ-cl (2), TAHV/84/CZ-cl, and TAHV/68/FR-cl in Vero cells or C6/36 cells infected at an MOI of 0.01. All viruses demonstrated rapid growth with a complete replication cycle in both cell types in less than 8 hours.

Figure 3

Alignment of the 3' (A) and 5' (B) untranslated regions of LACV/78-cl, TAHV/58/CZ-cl, and JCV/61/CO-cl. Sequence identical to the consensus indicated with a (.), areas where no censuses exist indicated by (X), gaps indicated by (-).

Figure 4

Alignment of the M polyproteins of TAHV/58/CZ-cl, SSHV/59/MT, LACV/78-cl, JCV/61/CO-cl, and INKV/64/FI. Sequence identical to the consensus indicated with a (.), areas where no consensus exist indicated by (X).

Figure 5

Alignment of the L proteins (aa 1-1200) of TAHV/58/CZ-cl, SSHV/59/MT, LACV/78-cl, JCV/61/CO-cl, and INKV/64/FI. Sequence identical to the consensus indicated with a (.), areas where no consensus exist indicated by (X). RNA-dependent RNA polymerase conserved motifs are underlined in the consensus sequence for L protein.

Figure 6

Alignment of the L proteins (aa 1201-2263) of TAHV/58/CZ-cl, SSHV/59/MT, LACV/78-cl, JCV/61/CO-cl, and INKV/64/FI. Sequence identical to the consensus indicated with a (.), areas where no consensus exist indicated by (X). RNA-dependent RNA polymerase conserved motifs are underlined in the consensus sequence for L protein.

Figure 7

Alignment of the N and NS_S proteins of TAHV/58/CZ-cl, SSHV/59/MT, LACV/78-cl, JCV/61/CO-cl, and INKV/64/FI. Sequence identical to the consensus indicated with a (.), areas where no consensus exist indicated by (X).