Experimental Reptarenavirus Infection of Boa constrictor and Python regius.

Boid inclusion body disease (BIBD) causes losses in captive snake populations globally. BIBD is associated with the formation of cytoplasmic inclusion bodies (IBs), which mainly comprise reptarenavirus nucleoprotein (NP). In 2017, BIBD was reproduced by cardiac injection of boas and pythons with reptarenaviruses, thus demonstrating a causative link between reptarenavirus infection and the disease. Here, we report experimental infections of Python regius (n = 16) and Boa constrictor (n = 16) with three reptarenavirus isolates. First, we used pythons (n = 8) to test two virus delivery routes: intraperitoneal injection and tracheal instillation. Viral RNAs but no IBs were detected in brains and lungs at 2 weeks postinoculation. Next, we inoculated pythons (n = 8) via the trachea. During the 4 months following infection, snakes showed transient central nervous system (CNS) signs but lacked detectable IBs at the time of euthanasia. One of the snakes developed severe CNS signs; we succeeded in reisolating the virus from the brain of this individual and could demonstrate viral antigen in neurons. In a third attempt, we tested cohousing, vaccination, and sequential infection with multiple reptarenavirus isolates on boas (n = 16). At 10 months postinoculation, all but one snake tested positive for viral RNA in lung, brain, and/or blood, but none exhibited the characteristic IBs. Three of the four vaccinated snakes seemed to sustain challenge with the same reptarenavirus; however, neither of the two snakes rechallenged with different reptarenaviruses remained uninfected. Comparison of the antibody responses in experimentally versus naturally reptarenavirus-infected animals indicated differences in the responses.IMPORTANCE In the present study, we experimentally infected pythons and boas with reptarenavirus via either intraperitoneal injection or tracheal instillation. The aims were to experimentally induce boid inclusion body disease (BIBD) and to develop an animal model for studying disease transmission and pathogenesis. Both virus delivery routes resulted in infection, and infection via the trachea could reflect the natural route of infection. In the experimentally infected snakes, we did not find evidence of inclusion body (IB) formation, characteristic of BIBD, in pythons or boas. Most of the boas (11/12) remained reptarenavirus infected after 10 months, which suggests that they developed a persistent infection that could eventually have led to BIBD. We demonstrated that vaccination using recombinant protein or an inactivated virus preparation prevented infection by a homologous virus in three of four snakes. Comparison of the antibody responses of experimentally and naturally reptarenavirus-infected snakes revealed differences that merit further studies.

Arenavirus Coinfections Are Common in Snakes with Boid Inclusion Body Disease.

Recently, novel arenaviruses were found in snakes with boid inclusion body disease (BIBD); these form the new genus Reptarenavirus within the family Arenaviridae. We used next-generation sequencing and de novo sequence assembly to investigate reptarenavirus isolates from our previous study. Four of the six isolates and all of the samples from snakes with BIBD contained at least two reptarenavirus species. The viruses sequenced comprise four novel reptarenavirus species and a representative of a new arenavirus genus.