Experimental Infection of Peromyscus Species Rodents with Sin Nombre Virus

We demonstrate that 6 distinct Peromyscus rodent species are permissive to experimental infection with Sin Nombre orthohantavirus (SNV). Viral RNA and SNV antibodies were detected in members of all 6 species. P. leucopus mice demonstrated markedly higher viral and antibody titers than P. maniculatus mice, the established primary hosts for SNV.

We demonstrate that 6 distinct Peromyscus rodent species are permissive to experimental infection with Sin Nombre orthohantavirus (SNV). Viral RNA and SNV antibodies were detected in members of all 6 species. P. leucopus mice demonstrated markedly higher viral and antibody titers than P. maniculatus mice, the established primary hosts for SNV. trapping locations. Median viral RNA peaked at 21 dpi and were sustained through 42 dpi; however, only serum was positive at 42 dpi in P. maniculatus rufinus mice. P. maniculatus bairdii mice founders were trapped in southeastern Michigan ( Figure), whereas P. maniculatus rufinus mice descended from founders from central and northwestern New Mexico (Figure). P. leucopus mice appear to be highly permissive to SNV based on detection of SNV RNA. We observed high levels of viral RNA in serum, heart, kidney, lung, and spleen samples at 21 dpi and 42 dpi. Further studies are required to confirm whether this finding translates to persistent infection, viral shedding, and possible transmission between animals. Although sex differences have been noted to have little effect on SNV infection in P. maniculatus rufinus mice (10), differences may exist in other rodent species. The numbers in our study are small; however, the data suggest sex-related differences might occur according to species. For example, male P. maniculatus bairdii mice have higher viral RNA titers at 42 dpi, whereas in P. maniculatus sonoriensis mice at 21 dpi, almost all positive animals are female. Meanwhile, P. leucopus mice showed indiscriminate viral RNA detection between sexes across all timepoints (Appendix Figure, https://wwwnc.cdc.gov/EID/ article/28/9/22-0509-App1.pdf). Future studies are required to shed light on this aspect. P. maniculatus mice were established early on as the primary reservoir for SNV (4). However, studies of this virus-host relationship rarely report the host taxon beyond the species level. Moreover, although levels of viral RNA tended to be negligible in the early stages of infection, an observation consistent with previous studies in susceptible animals (9,10,13), the detection of viral RNA at later timepoints probably means members of these species are nonetheless susceptible to SNV infection. That animals were positive at 42 dpi hints at the possibility of persistent infection, although longer-term studies would be needed to confirm.
Antibodies against the SNV nucleoprotein were detected in most serum samples collected from days 21 and 42 from the peromyscine rodents (Table). The lone exception was P. californicus mice, for which only a single animal had detectable SNV antibodies. Rates of seroconversion broadly mirrored the rates of detection by quantitative reverse transcription PCR. The development of antibodies reactive to the SNV nucleoprotein appeared to be delayed in male mice from 2 species (P. maniculatus bairdii and P. maniculatus sonoriensis), and only 1 of 6 of these rodents demonstrated positive serum samples at 21 dpi.

Conclusions
Overall, our data support recent observations from Goodfellow et al. (8) that rodents other than P. maniculatus mice are capable of carrying SNV without showing signs of disease. In that study, wild-caught members of P. boylii mice, Mus musculus mice, and Tamias minimus chipmunks trapped at 2 sites had detectable SNV RNA in their lung tissues. SNV sequences from these rodents were more similar to each other than to previously reported sequences, suggesting circulation of the virus within these rodent populations.
In our study, we experimentally infected 6 Peromyscus mouse species, including 3 that fall under the P. maniculatus species complex, and whose founders originate from locations across North America, with a strain of SNV originating from a single wild-caught animal. Although replicating SNV was detected in animals of all 6 species, permissiveness of each species to SNV infection varied. Susceptibility to SNV infection appears to be multifactorial and is not fully explained by characteristics relating to geographic proximity or genetic relatedness. Given the variability in the patterns of infection between subspecies within P. maniculatus, future studies should consider reporting the subspecies when studying this virus-host relationship. Furthermore, other Peromyscus mouse species may play an important role in the molecular evolution and transmission of SNV. Host-switching events are thought to give rise to SNV variants and even new hantaviral species (1,2,14). That SNV was capable of replicating to high levels in P. leucopus mice for sustained periods is notable. The geographic distributions of P. maniculatus and P. leucopus mice overlap greatly, presenting opportunities for these 2 populations and their respective viruses to come into contact. Together, these regions cover most of North America, including nearly the entirety of the contiguous United States.
These findings demonstrate the importance of broadening our understanding of the SNV-P. maniculatus virus-host relationship and highlight the benefit of identifying infected and infectious rodent reservoirs at the subspecies level to help elucidate epizootics and spillover events to humans. Furthermore, although distinct eastern and western patterns of genetic evolution have been documented in P. maniculatus rodents and associated strains of SNV in North America (1,2), our study suggests that these patterns might not necessarily prevent western lineages of SNV from emerging into eastern populations of peromyscine rodents.