Poxviruses as a group can infect a large number of animals. However, at the level of individual viruses, even closely related poxviruses display highly diverse host ranges and virulence. For example, variola virus, the causative agent of smallpox, is human-specific and highly virulent only to humans, whereas related cowpox viruses naturally infect a broad spectrum of animals and only cause relatively mild disease in humans. The successful replication of poxviruses depends on their effective manipulation of the host antiviral responses, at the cellular-, tissue- and species-specific levels, which constitutes a molecular basis for differences in poxvirus host range and virulence. A number of poxvirus genes have been identified that possess host range function in experimental settings, and many of these host range genes target specific antiviral host pathways. Herein, we review the biology of poxviruses with a focus on host range, zoonotic infections, virulence, genomics and host range genes as well as the current knowledge about the function of poxvirus host range factors and how their interaction with the host innate immune system contributes to poxvirus host range and virulence. We further discuss the evolution of host range and virulence in poxviruses as well as host switches and potential poxvirus threats for human and animal health.

Chikungunya virus (CHIKV) is a reemerging arbovirus capable of causing explosive outbreaks of febrile illness, polyarthritis, and polyarthralgia, inflicting severe morbidity on affected populations. CHIKV can be genetically classified into 3 major lineages: West African (WA); East, Central, and South African (ECSA); Indian Ocean (IOL); and Asian. Additionally, the Indian Ocean (IOL) sublineage emerged within the ECSA clade and the Asian/American sublineage emerged within the Asian clade. While differences in epidemiological and pathological characteristics among outbreaks involving different CHIKV lineages and sublineages have been suggested, few targeted investigations comparing lineage virulence levels have been reported. We compared the virulence levels of CHIKV isolates representing all major lineages and sublineages in the type I interferon receptor-knockout A129 mouse model and found lineage-specific differences in virulence. We also evaluated the cross-protective efficacy of the IOL-derived, live-attenuated vaccine strain CHIKV/IRESv1 against the Asian/American CHIKV isolate YO123223 in both murine and nonhuman primate models, as well as the WA strain SH2830 in a murine model. The CHIKV/IRES vaccine provided protection both in mice and in nonhuman primate cohorts against Caribbean strain challenge and protected mice against WA challenge. Taken together, our data suggest that Asian/American CHIKV strains are less virulent than those in the Asian, ECSA, and WA lineages and that despite differences in virulence, IOL-based vaccine strains offer robust cross-protection against strains from other lineages. Further research is needed to elucidate the genetic basis for variation in CHIKV virulence in the A129 mouse model and to corroborate this variation with human pathogenicity.

Myxoma virus (MYXV) is a rabbit-specific poxvirus, which is highly virulent in European rabbits. The attenuation of MYXV and the increased resistance of rabbits following the release of MYXV in Australia is one of the best-documented examples of host-pathogen coevolution. To elucidate the molecular mechanisms that contribute to the restriction of MYXV infection to rabbits and MYXV attenuation in the field, we have studied the interaction of the MYXV protein M156 with the host antiviral protein kinase R (PKR). In yeast and cell-culture transfection assays, M156 only inhibited rabbit PKR but not PKR from other tested mammalian species. Infection assays with human HeLa PKR knock-down cells, which were stably transfected with human or rabbit PKR, revealed that only human but not rabbit PKR was able to restrict MYXV infection, whereas both PKRs were able to restrict replication of a vaccinia virus (VACV) strain that lacks the PKR inhibitors E3 and K3. Inactivation of M156R led to MYXV virus attenuation in rabbit cells, which was rescued by the ectopic expression of VACV E3 and K3. We further show that a mutation in the M156 encoding gene that was identified in more than 50% of MYXV field isolates from Australia resulted in an M156 variant that lost its ability to inhibit rabbit PKR and led to virus attenuation. The species-specific inhibition of rabbit PKR by M156 and the M156 loss-of-function in Australian MYXV field isolates might thus contribute to the species specificity of MYXV and to the attenuation in the field, respectively.oxviruses are large double-stranded DNA viruses that exclusively replicate in the cytoplasm of infected cells. Members of the Poxviridae family can productively infect a wide variety of animal hosts. Interestingly, the binding and entry of poxviruses into cells is largely independent of host species after which virus replication is initiated. The successful completion of virus replication, however, depends on the effective subversion of the host cell's innate immune responses (1). Even closely related poxviruses can exhibit drastic differences in their host ranges. Whereas some poxviruses have only one host species, such as variola virus, the causative agent of smallpox, which is restricted to humans, others, such as cowpox and monkeypox viruses, can infect many different species and thus display very broad host ranges (2). A number of poxvirus genes have been discovered that influence the host range and cell tropism of poxviruses and have therefore been termed "host range genes" (3, 4). Although the molecular mechanisms responsible for their host range functions have not been elucidated in detail, it is clear that most poxviral host range proteins interact with components of the host immune system and that host species-specific interactions likely play a major role.Myxoma virus (MYXV) is a poxvirus that belongs to the genus leporipoxvirus and shows a restricted host range infecting only leporids (rabbits and hares). MYXV is highly lethal to European (E.) rabbits causing case fatality rate...

Arboviruses maintain high mutation rates due to lack of proofreading ability of their viral polymerases, in some cases facilitating adaptive evolution and emergence. Here we show that, just before its 2013 spread to the Americas, Zika virus (ZIKV) underwent an envelope protein V473M substitution (E-V473M) that increased neurovirulence, maternal-to-fetal transmission, and viremia to facilitate urban transmission. A preepidemic Asian ZIKV strain (FSS13025 isolated in Cambodia in 2010) engineered with the V473M substitution significantly increased neurovirulence in neonatal mice and produced higher viral loads in the placenta and fetal heads in pregnant mice. Conversely, an epidemic ZIKV strain (PRVABC59 isolated in Puerto Rico in 2015) engineered with the inverse M473V substitution reversed the pathogenic phenotypes. Although E-V473M did not affect oral infection of Aedes aegypti mosquitoes, competition experiments in cynomolgus macaques showed that this mutation increased its fitness for viremia generation, suggesting adaptive evolution for human viremia and hence transmission. Mechanistically, the V473M mutation, located at the second transmembrane helix of the E protein, enhances virion morphogenesis. Overall, our study revealed E-V473M as a critical determinant for enhanced ZIKV virulence, intrauterine transmission during pregnancy, and viremia to facilitate urban transmission.

Mayaro virus (MAYV) causes an acute febrile illness similar to that produced by chikungunya virus (CHIKV), an evolutionary relative in the Semliki Forest virus complex of alphaviruses. MAYV emergence is typically sporadic, but recent isolations and outbreaks indicate that the virus remains a public health concern. Given the close phylogenetic and antigenic relationship between CHIKV and MAYV, and widespread distribution of CHIKV, we hypothesized that prior CHIKV immunity may affect MAYV pathogenesis and/or influence its emergence potential. We pre-exposed immunocompetent C57BL/6 and immunocompromised A129 or IFNAR mice to wild-type CHIKV, two CHIKV vaccines, or a live-attenuated MAYV vaccine, and challenged with MAYV. We observed strong cross-protection against MAYV for mice pre-exposed to wild-type CHIKV, and moderately but significantly reduced cross-protection from CHIKV-vaccinated animals. Immunity to other alphavirus or flavivirus controls provided no protection against MAYV disease or viremia. Mechanistic studies suggested that neutralizing antibodies alone can mediate this protection, with T-cells having no significant effect on diminishing disease. Finally, human sera obtained from naturally acquired CHIKV infection cross-neutralized MAYV at high titers in vitro. Altogether, our data suggest that CHIKV infection can confer cross-protective effects against MAYV, and the resultant reduction in viremia may limit the emergence potential of MAYV.

Translational control of transcription factor ATF4 through paired upstream ORFs (uORFs) plays an important role in eukaryotic gene regulation. While it is typically induced by phosphorylation of eIF2α, ATF4 translation can be also induced by expression of a translational inhibitor protein, eIF5-mimic protein 1 (5MP1, also known as BZW2) in mammals. Here we show that the 5MP gene is maintained in eukaryotes under strong purifying selection, but is uniquely missing in two major phyla, nematoda and ascomycota. The common function of 5MP from protozoa, plants, fungi and insects is to control translation by inhibiting eIF2. The affinity of human 5MP1 to eIF2β was measured as being equivalent to the published value of human eIF5 to eIF2β, in agreement with effective competition of 5MP with eIF5 for the main substrate, eIF2. In the red flour beetle, Tribolium castaneum, RNA interference studies indicate that 5MP facilitates expression of GADD34, a downstream target of ATF4. Furthermore, both 5MP and ATF4 are essential for larval development. Finally, 5MP and the paired uORFs allowing ATF4 control are conserved in the entire metazoa except nematoda. Based on these findings, we discuss the phylogenetic and functional linkage between ATF4 regulation and 5MP expression in this group of eukaryotes.

Zika virus (ZIKV) emerged from obscurity in 2013 to spread from Asia to the South Pacific and the Americas, where millions of people were infected, accompanied by severe disease including microcephaly following congenital infections. Phylogenetic studies have shown that ZIKV evolved in Africa and later spread to Asia, and that the Asian lineage is responsible for the recent epidemics in the South Pacific and Americas. However, the reasons for the sudden emergence of ZIKV remain enigmatic. Here we report evolutionary analyses that revealed four mutations, which occurred just before ZIKV introduction to the Americas, represent direct reversions of previous mutations that accompanied earlier spread from Africa to Asia and early circulation there. Our experimental infections of Aedes aegypti mosquitoes, human cells, and mice using ZIKV strains with and without these mutations demonstrate that the original mutations reduced fitness for urban, human-amplifed transmission, while the reversions restored fitness, increasing epidemic risk. These findings include characterization of three transmission-adaptive ZIKV mutations, and demonstration that these and one identified previously restored fitness for epidemic transmission soon before introduction into the Americas. The initial mutations may have followed founder effects and/or drift when the virus was introduced decades ago into Asia.

This paper reports the capture and detection of vaccinia virus particles based on AC dielectrophoresis (DEP) and electrochemical impedance measurements employing an embedded vertically aligned carbon nanofiber (VACNF) nanoelectrode array (NEA) versus a macroscopic indium-tin-oxide (ITO) transparent electrode in a "points-and-lid" configuration. The nano-DEP device was fabricated by bonding two SU-8 covered electrodes patterned using photolithography. The bottom electrode contains a 200 × 200 μm active region on a randomly distributed NEA and the top electrode contains a microfluidic channel in SU-8 spin-coated on ITO to guide the flow of the virus solution. The real-time impedance change was measured during DEP capture and validated with fluorescence microscopy measurements. The NEA was able to capture virus particles with a rather low AC voltage (∼8.0 V peak-to-peak) at 1.0 kHz frequency as the particles were passed through the fluidic channel at high flow velocities (up to 8.0 mm/s). A concentration detection limit as low as ∼2.58 × 10 particles/mL was obtained via impedance measurements after only 54 sec of DEP capture. At the low AC frequencies (50.0 Hz or less), the high electric field at the exposed VACNF tips induced electroporation of the DEP-captured virus particles, which was validated by fluorescence emission from the dyes staining lipophilic membrane and internal nucleic acid, respectively. This study suggests the possibility of integration of a fully functional electronic device for rapid, reversible and label-free capture and detection of pathogenic viruses, with a potential of generating electroporation to the captured the virus particles for further biochemical study.