Necropsies of green anaconda juveniles and sample collection. In this article, we report this unprecedented cluster of fatal infections associated with a paramyxovirus in snakes and the characterization of this novel paramyxovirus, which we propose to be named anaconda paramyxovirus (AnaPV). Further viral studies and complete genomic analyses of five isolates revealed a novel paramyxovirus closely related to FDLV. To test this hypothesis, we carried out a molecular study with samples collected from 13 of these green anaconda juveniles that died. Since FDLV has been shown to be associated with fatalities in snakes, we hypothesized that a FDLV-like virus could be the cause of the cluster of infections. These green anaconda juveniles were born in July 2011 from a new reptile in Ocean Park Hong Kong that had been purchased from Japan in June 2011. In addition, there are a number of paramyxoviruses of the subfamily Paramyxovirinae that are still not classified into any genus these include Beilong virus, J virus, and Tailam virus, which probably belong to the same genus ( 2, 13, 19), and tupaia paramyxovirus ( 10).īeginning in July 2011, 31 green anaconda ( Eunectes murinus) juveniles from Ocean Park of the Hong Kong Special Administrative Region (HKSAR) died over a 12-month period. Recently, two new genera, i.e., Aquaparamyxovirus, which consists of Atlantic salmon paramyxovirus, and Ferlavirus, which consists of Fer-de-Lance virus (FDLV) discovered in the common lancehead snake ( Bothrops atrox) ( 17, 18) ( ), were proposed. Traditionally, there are five genera within the subfamily Paramyxovirinae, namely, Respirovirus, Rubulavirus, Morbillivirus, Henipavirus, and Avulavirus. Examples include Tuhoko virus 1, 2, and 3 from fruit bats ( 1), Tailam virus from Sikkim rats ( 2), Beilong virus from brown rats and black rats ( 3, 4), feline morbillivirus from domestic cats ( 5), porcine parainfluenza virus 1 from swine ( 6), Menangle virus from pigs ( 7), Tioman virus from fruit bats ( 8), Cedar virus from Australian bats ( 9), tupaia paramyxovirus from tree shrews ( 10), Salem virus from horses ( 11), Mossman virus from rodents ( 12), J virus from mice ( 13), Tursiops truncatus parainfluenza virus type 1 from dolphins ( 14), Sunshine virus from snakes ( 15), and >60 new paramyxoviruses from bats and rodents ( 16). In the past decade, a number of paramyxoviruses in the subfamily Paramyxovirinae, as well as their natural hosts, have been discovered. Paramyxoviruses are enveloped, negative-stranded RNA viruses that are divided into two subfamilies, Paramyxovirinae and Pneumovirinae. Epidemiological and genomic analyses suggested that the anaconda juveniles acquired the virus perinatally from the anaconda mother rather than from other reptiles in the park, with subsequent interanaconda juvenile transmission. Comparative genomic and phylogenetic analyses and mRNA editing experiments revealed a novel paramyxovirus in the genus Ferlavirus, named anaconda paramyxovirus, with a typical Ferlavirus genomic organization of 3′-N-U-P/V/I-M-F-HN-L-5′. Complete genome sequencing of five isolates confirmed that the infections originated from the same clone.
Electron microscopy with cell culture material revealed enveloped virus with the typical “herringbone” appearance of helical nucleocapsids in paramyxoviruses. The virus could also replicate in 25 of 27 cell lines of various origins, in line with its capability for infecting various organs. The virus was isolated from BHK21 cells, causing cytopathic effects with syncytial formation. None of the other snakes in the park was moribund, and RT-PCR results for surveillance samples collected from other snakes were negative. Reverse transcription (RT)-PCR results were positive for paramyxovirus (viral loads of 2.33 × 10 4 to 1.05 × 10 8 copies/mg tissue) in specimens from anaconda juveniles that died but negative in specimens from the two anaconda juveniles and anaconda mother that survived. Electron microscopic examination of primary tissues revealed intralesional accumulations of viral nucleocapsids with diameters of 10 to 14 nm, typical of paramyxoviruses. Histopathological examination revealed severe necrotizing inflammation in various organs, most prominently the kidneys. Necropsy revealed at least two of the following features in 23 necropsies: dermatitis, severe pan-nephritis, and/or severe systemic multiorgan necrotizing inflammation.
Beginning in July 2011, 31 green anaconda ( Eunectes murinus) juveniles from an oceanarium in Hong Kong died over a 12-month period.