Arenavirus Coinfections Are Common in Snakes with Boid Inclusion Body Disease

Abstract

e 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. Until very recently, arenaviruses were known as a group of mainly rodent-borne zoonotic viruses (1). The negativesense RNA genome of arenaviruses is divided into two segments, which are designated small (S, approximately 3.5 kb) and large (L, approximately 7 to 7.5 kb); both use an ambisense coding strategy (1). The S segment encodes the glycoprotein precursor (GPC) and the nucleoprotein (NP), whereas the RNA-dependent RNA polymerase (RdRp) and the Z protein (ZP) are encoded in the L segment (1). The discovery of arenaviruses in snakes with boid inclusion body disease (BIBD) by three independent groups (2-4) has expanded the family Arenaviridae by a new group of viruses. In fact, the BIBD-associated arenaviruses (BIBDAVs) have been suggested to form a new arenavirus genus called Reptarenavirus (5). At the same time, the arenavirus study group of the International Committee on Taxonomy of Viruses (ICTV) has suggested that the genus Arenavirus, harboring the "classical" Old and New World arenaviruses, be renamed Mammarenavirus (6). According to a recent proposal, the genus Reptarenavirus would contain three species: alethinophid reptarenaviruses 1 (member virus: Golden Gate virus), 2 (CAS virus, CASV), and 3 (University of Helsinki virus 1 [UHV-1], boa AV NL B3 virus). While in vitro evidence suggests a causal relationship between arenavirus infection and BIBD (4), the in vivo proof is still missing. Also, the reservoir host(s) of the reptarenaviruses has not yet been confirmed; however, our recent study suggests that these viruses preferentially grow in organisms with body temperatures close to 30°C (7).To study the diversity of reptarenaviruses, we applied nextgeneration sequencing (NGS) to characterize certain isolates described in our previous report (4). On the basis of phylogeny, we selected isolates originating from six Boa constrictor snakes and used a continuous B. constrictor kidney cell line, I/1Ki (4), for their propagation. While the virus isolate of snake 1 has already been almost fully sequenced (GenBank accession numbers KF297880.1 and KF297881.1), only partial L segment sequences were available for isolates from snakes 5 (KF564801.1), 8 (KF564796.1), 9 (KF564800.1), 11, and 41 (KF564797.1). We infected clean I/1Ki cultures with tissue homogenates and collected the viruses produced during the first passage by pelleting through a sucrose cushion as previously described (4). Viral RNAs were extracted from the pelleted viruses with the QIAmp Viral RNA minikit (Qiagen) according to the ma...

Snake Deltavirus Utilizes Envelope Proteins of Different Viruses To Generate Infectious Particles

“…Affinity purification served to produce anti-SDAg and anti-NP (reptarenavirus nucleoprotein, the main antigen present in infected cells), which we directly labeled with Alexa Fluor 488 or Alexa Fluor 594 dyes (anti-SDAg-AF488, anti-SDAg-AF594, anti-NP-AF488, and anti-NP-AF594). Our studies show that the anti-NP antibody can be used to detect all reptarenavirus NPs thus far described (30)(31)(32)(33). As demonstrated in Fig.…”

Snake deltavirus utilizes envelope proteins of different viruses to generate infectious particles

“…For superinfection studies we used two reptarenaviruses: University of Helsinki virus-2 (UHV-2 29 ) and University of Giessen virus-1 (UGV-1 33 ), and one hartmanivirus, Haartman Institute Snake virus-1 (HISV-1 29 ).…”

Bioinformatics analysis for L polymerase homology of arenaviruses

“…Mammarenaviruses distributing worldwide and cause febrile diseases for both humans and animals (Buchmeier et al, 2007). Reptarenaviruses are infected snakes (Hepojoki et al, 2015), were the infection in human have not been reported. Arenaviruses have a segmented ambisense single strand RNA genome, which is typically grouped as negative sense RNA viruses (Buchmeier et al, 2007).…”