Gut Virome of the World's Highest-Elevation Lizard Species (Phrynocephalus erythrurus and Phrynocephalus theobaldi) Reveals Versatile Commensal Viruses.

The gut virome is a reservoir of diverse symbiotic and pathogenic viruses coevolving with their hosts, and yet limited research has explored the gut viromes of highland-dwelling rare species. Using viral metagenomic analysis, the viral communities of the Phrynocephalus lizards living in the Qinghai-Tibet Plateau were investigated. Phage-encoded functional genes and antibiotic resistance genes (ARGs) were analyzed. The viral communities of different lizard species were all predominated by bacteriophages, especially the Caudovirales order. The virome of Phrynocephalus erythrurus living around the Namtso Lake possessed a unique structure, with the greatest abundance of the Parvoviridae family and the highest number of exclusive viral species. Several vertebrate-infecting viruses were discovered, including caliciviruses, astroviruses, and parvoviruses. Phylogenetic analyses demonstrated that the virus hallmark genes of bacteriophages possessed high genetic diversity. After functional annotation, the majority of phage-associated functional genes were classified in the energy metabolism category. In addition, plenty of ARGs belonging to the multidrug category were discovered, and five ARGs were exclusive to the virome from Phrynocephalus theobaldi. This study provided the first insight into the structure and function of the virome in highland lizards, contributing to the protection of threatened lizard species. Also, our research is of exemplary significance for the gut virome research of lizard species and other cold-blooded and highland-dwelling animals, prompting a better understanding of the interspecific differences and transmission of commensal viruses. IMPORTANCE The Phrynocephalus lizards inhabiting the Qinghai-Tibet Plateau (QTP) are considered to be the highest-altitude lizard species in the world, and they have been added to the IUCN list of threatened species. Living in the QTP with hypoxic, arid, natural conditions, the lizards presented a unique pattern of gut virome, which could provide both positive and negative effects, such as the enrichment of functional genes and the dissemination of antibiotic resistance genes (ARGs). This work provides the foundation for further research on the gut virome in these endangered lizard species and other cold-blooded and highland-dwelling animals, contributing to the maintenance of ecological balance on the plateau.

First Report on Detection and Molecular Characterization of Adenoviruses in the Small Indian Mongoose (Urva auropunctata).

Using a broad-range nested PCR assay targeting the DNA-dependent DNA polymerase (pol) gene, we detected adenoviruses in 17 (20.48%) out of 83 fecal samples from small Indian mongooses (Urva auropunctata) on the Caribbean island of St. Kitts. All 17 PCR amplicons were sequenced for the partial pol gene (~300 bp, hereafter referred to as Mon sequences). Fourteen of the 17 Mon sequences shared maximum homology (98.3-99.6% and 97-98.9% nucleotide (nt) and deduced amino acid (aa) sequence identities, respectively) with that of bovine adenovirus-6 (species Bovine atadenovirus E). Mongoose-associated adenovirus Mon-39 was most closely related (absolute nt and deduced aa identities) to an atadenovirus from a tropical screech owl. Mon-66 shared maximum nt and deduced aa identities of 69% and 71.4% with those of atadenoviruses from a spur-thighed tortoise and a brown anole lizard, respectively. Phylogenetically, Mon-39 and Mon-66 clustered within clades that were predominated by atadenoviruses from reptiles, indicating a reptilian origin of these viruses. Only a single mongoose-associated adenovirus, Mon-34, was related to the genus Mastadenovirus. However, phylogenetically, Mon-34 formed an isolated branch, distinct from other mastadenoviruses. Since the fecal samples were collected from apparently healthy mongooses, we could not determine whether the mongoose-associated adenoviruses infected the host. On the other hand, the phylogenetic clustering patterns of the mongoose-associated atadenoviruses pointed more towards a dietary origin of these viruses. Although the present study was based on partial pol sequences (~90 aa), sequence identities and phylogenetic analysis suggested that Mon-34, Mon-39, and Mon-66 might represent novel adenoviruses. To our knowledge, this is the first report on the detection and molecular characterization of adenoviruses from the mongoose.

Discovery of a phylogenetically distinct poxvirus in diseased Crocodilurus amazonicus (family Teiidae).

A novel poxvirus was discovered in Crocodilurus amazonicus (Teiidae) presenting with a debilitating skin disease. The generated first genome sequence of a reptilian poxvirus revealed the closest phylogenetic relationship to avipoxviruses, highlighting potential virus exchanges between avian and reptilian species.

Identification of a reptile lyssavirus in Anolis allogus provided novel insights into lyssavirus evolution.

Lyssaviruses (genus Lyssavirus) are negative-strand RNA viruses belonging to the family Rhabdoviridae. Although a lyssa-like virus (frog lyssa-like virus 1 [FLLV-1]), which is distantly related to lyssaviruses, was recently identified in frogs, a large phylogenetic gap exists between those viruses, and thus the evolution of lyssaviruses is unclear. In this study, we detected a lyssa-like virus from publicly available RNA-seq data obtained using the brain and skin of Anolis allogus (Spanish flag anole), which was designated anole lyssa-like virus 1 (ALLV-1), and determined its complete coding sequence. Via mapping analysis, we demonstrated that ALLV-1 was actively replicating in the original brain and skin samples. Phylogenetic analyses revealed that ALLV-1 is more closely related to lyssaviruses than FLLV-1. Overall, the topology of the tree is compatible with that of hosts, suggesting the long-term co-divergence of lyssa-like and lyssaviruses and vertebrates. The ψ region, which is a long 3' untranslated region of unknown origin present in the G mRNA of lyssaviruses (approximately 400-700 nucleotides), is also present in the genome of ALLV-1, but it is much shorter (approximately 180 nucleotides) than those of lyssaviruses. Interestingly, FLLV-1 lacks the ψ region, suggesting that the ψ region was acquired after the divergence of the FLLV-1 and ALLV-1/lyssavirus lineages. To the best of our knowledge, this is the first report to identify a lyssa-like virus in reptiles, and thus, our findings provide novel insights into the evolution of lyssaviruses.

Serpentovirus (Nidovirus) and Orthoreovirus Coinfection in Captive Veiled Chameleons (Chamaeleo calyptratus) with Respiratory Disease.

Serpentoviruses are an emerging group of nidoviruses known to cause respiratory disease in snakes and have been associated with disease in other non-avian reptile species (lizards and turtles). This study describes multiple episodes of respiratory disease-associated mortalities in a collection of juvenile veiled chameleons (Chamaeleo calyptratus). Histopathologic lesions included rhinitis and interstitial pneumonia with epithelial proliferation and abundant mucus. Metagenomic sequencing detected coinfection with two novel serpentoviruses and a novel orthoreovirus. Veiled chameleon serpentoviruses are most closely related to serpentoviruses identified in snakes, lizards, and turtles (approximately 40-50% nucleotide and amino acid identity of ORF1b). Veiled chameleon orthoreovirus is most closely related to reptilian orthoreoviruses identified in snakes (approximately 80-90% nucleotide and amino acid identity of the RNA-dependent RNA polymerase). A high prevalence of serpentovirus infection (>80%) was found in clinically healthy subadult and adult veiled chameleons, suggesting the potential for chronic subclinical carriers. Juvenile veiled chameleons typically exhibited a more rapid progression compared to subadults and adults, indicating a possible age association with morbidity and mortality. This is the first description of a serpentovirus infection in any chameleon species. A causal relationship between serpentovirus infection and respiratory disease in chameleons is suspected. The significance of orthoreovirus coinfection remains unknown.

Meta-Transcriptomic Discovery of a Divergent Circovirus and a Chaphamaparvovirus in Captive Reptiles with Proliferative Respiratory Syndrome.

Viral pathogens are being increasingly described in association with mass morbidity and mortality events in reptiles. However, our knowledge of reptile viruses remains limited. Herein, we describe the meta-transcriptomic investigation of a mass morbidity and mortality event in a colony of central bearded dragons (Pogona vitticeps) in 2014. Severe, extensive proliferation of the respiratory epithelium was consistently found in affected dragons. Similar proliferative lung lesions were identified in bearded dragons from the same colony in 2020 in association with increased intermittent mortality. Total RNA sequencing identified two divergent DNA viruses: a reptile-infecting circovirus, denoted bearded dragon circovirus (BDCV), and the first exogeneous reptilian chaphamaparvovirus-bearded dragon chaphamaparvovirus (BDchPV). Phylogenetic analysis revealed that BDCV was most closely related to bat-associated circoviruses, exhibiting 70% amino acid sequence identity in the Replicase (Rep) protein. In contrast, in the nonstructural (NS) protein, the newly discovered BDchPV showed approximately 31%-35% identity to parvoviruses obtained from tilapia fish and crocodiles in China. Subsequent specific PCR assays revealed BDCV and BDchPV in both diseased and apparently normal captive reptiles, although only BDCV was found in those animals with proliferative pulmonary lesions and respiratory disease. This study expands our understanding of viral diversity in captive reptiles.

A Divergent Articulavirus in an Australian Gecko Identified Using Meta-Transcriptomics and Protein Structure Comparisons.

The discovery of highly divergent RNA viruses is compromised by their limited sequence similarity to known viruses. Evolutionary information obtained from protein structural modelling offers a powerful approach to detect distantly related viruses based on the conservation of tertiary structures in key proteins such as the RNA-dependent RNA polymerase (RdRp). We utilised a template-based approach for protein structure prediction from amino acid sequences to identify distant evolutionary relationships among viruses detected in meta-transcriptomic sequencing data from Australian wildlife. The best predicted protein structural model was compared with the results of similarity searches against protein databases. Using this combination of meta-transcriptomics and protein structure prediction we identified the RdRp (PB1) gene segment of a divergent negative-sense RNA virus, denoted Lauta virus (LTAV), in a native Australian gecko (Gehyra lauta). The presence of this virus was confirmed by PCR and Sanger sequencing. Phylogenetic analysis revealed that Lauta virus likely represents a newly described genus within the family Amnoonviridae, order Articulavirales, that is most closely related to the fish virus Tilapia tilapinevirus (TiLV). These findings provide important insights into the evolution of negative-sense RNA viruses and structural conservation of the viral replicase among members of the order Articulavirales.

The complete genome sequence of bearded dragon adenovirus 1 harbors three genes encoding proteins of the C-type lectin-like domain superfamily.

Bearded dragon adenovirus 1 (BDAdV-1), also known as agamid adenovirus 1, has been described worldwide as a prevalent infectious agent of the inland bearded dragon (Pogona vitticeps), the most common squamate exotic pet reptile. Previous limited sequence data of the adenoviral DNA polymerase and hexon genes indicated that BDAdV-1 is a member of genus Atadenovirus family Adenoviridae. Atadenoviruses infect ruminants, marsupials, testudine reptiles and birds, yet the genus has been shown to be of squamate reptile origin. Here, we report a screening survey along with the complete genome sequence of BDAdV-1, derived directly from the sample of a deceased juvenile dragon showing central nervous system signs prior to passing. The BDAdV-1 genome is 35,276 bp and contains 32 putative genes. Its genome organization is characteristic of the members of genus Atadenovirus, however, a divergent LH3 gene indicates structural interactions of different nature compared to other genus members such as snake adenovirus 1. We identified five novel open reading frames (ORFs), three of which encode proteins of the C-type lectin-like domain (CTLD) superfamily. ORF3 has a CTLD group II-like domain architecture displaying structural similarity with natural killer cell surface receptors and with an alphaherpesviral virulence factor gene for neurotropism, UL45. ORF4 and 6 are extremely long compared to typical adenoviral right-end genes and possibly encode members of the CTLD superfamily with novel, previously undescribed domain architectures. BDAdV-1 is the hitherto most divergent member of genus Atadenovirus providing new insights on adenoviral diversity, evolution and pathogenesis.

The use of a salmonella bacteriophage in bearded dragons: application, passage time and reisolation.

OBJECTIVE: This study determined the passage time and phage propagation time of a salmonella specific phage, Felix O1, in bearded dragons, based on reisolation from cloacal swabs and faecal samples following oral administration, as a possible tool for reducing the zoonotic risk of salmonella from pet reptiles. An application scheme for this phage in bearded dragons was developed. MATERIAL AND METHODS: Ten healthy bearded dragons (Pogona vitticeps) were used in the study. The pH tolerance of the phage was tested and drugs were used to evaluate their influence on the gastric pH of the reptiles. After pH adjustment, the phage was administered orally for 12 consecutive days. Over 60 days, swabs were taken from the cloaca and examined for the presence of phages using culture and PCR. Furthermore, faecal samples were collected for phage quantification. RESULTS: Felix O1 displayed no activity at pH below 2.8. A calcium- and magnesium carbonate buffer induced an appropriate gastric pH increase for 30 minutes. Phages were reisolated for up to 24 days (mean shedding: 19 days) after last administration. Titres between 105 and 107 plaque forming units/g faeces were detected. The animals did not show any clinical signs related to phage application. CONCLUSION AND CLINICAL RELEVANCE: The study provides first results on oral administration, passage time, and reisolation of a phage in reptiles. It could be shown that the phage was able to replicate in the intestine, and was shed for a prolonged period and therefore could potentially contribute to a reduction of salmonella shedding.

The First Co-Opted Endogenous Foamy Viruses and the Evolutionary History of Reptilian Foamy Viruses.

A recent study reported the discovery of an endogenous reptilian foamy virus (FV), termed ERV-Spuma-Spu, found in the genome of tuatara. Here, we report two novel reptilian foamy viruses also identified as endogenous FVs (EFVs) in the genomes of panther gecko (ERV-Spuma-Ppi) and Schlegel's Japanese gecko (ERV-Spuma-Gja). Their presence indicates that FVs are capable of infecting reptiles in addition to mammals, amphibians, and fish. Numerous copies of full length ERV-Spuma-Spu elements were found in the tuatara genome littered with in-frame stop codons and transposable elements, suggesting that they are indeed endogenous and are not functional. ERV-Spuma-Ppi and ERV-Spuma-Gja, on the other hand, consist solely of a foamy virus-like env gene. Examination of host flanking sequences revealed that they are orthologous, and despite being more than 96 million years old, their env reading frames are fully coding competent with evidence for strong purifying selection to maintain expression and for them likely being transcriptionally active. These make them the oldest EFVs discovered thus far and the first documented EFVs that may have been co-opted for potential cellular functions. Phylogenetic analyses revealed a complex virus-host co-evolutionary history and cross-species transmission routes of ancient FVs.

Detection and Characterization of Invertebrate Iridoviruses Found in Reptiles and Prey Insects in Europe over the Past Two Decades.

Invertebrate iridoviruses (IIVs), while mostly described in a wide range of invertebrate hosts, have also been repeatedly detected in diagnostic samples from poikilothermic vertebrates including reptiles and amphibians. Since iridoviruses from invertebrate and vertebrate hosts differ strongly from one another based not only on host range but also on molecular characteristics, a series of molecular studies and bioassays were performed to characterize and compare IIVs from various hosts and evaluate their ability to infect a vertebrate host. Eight IIV isolates from reptilian and orthopteran hosts collected over a period of six years were partially sequenced. Comparison of eight genome portions (total over 14 kbp) showed that these were all very similar to one another and to an earlier described cricket IIV isolate, thus they were given the collective name lizard-cricket IV (Liz-CrIV). One isolate from a chameleon was also subjected to Illumina sequencing and almost the entire genomic sequence was obtained. Comparison of this longer genome sequence showed several differences to the most closely related IIV, Invertebrateiridovirus6 (IIV6), the type species of the genus Iridovirus, including several deletions and possible recombination sites, as well as insertions of genes of non-iridoviral origin. Three isolates from vertebrate and invertebrate hosts were also used for comparative studies on pathogenicity in crickets (Gryllusbimaculatus) at 20 and 30 °C. Finally, the chameleon isolate used for the genome sequencing studies was also used in a transmission study with bearded dragons. The transmission studies showed large variability in virus replication and pathogenicity of the three tested viruses in crickets at the two temperatures. In the infection study with bearded dragons, lizards inoculated with a Liz-CrIV did not become ill, but the virus was detected in numerous tissues by qPCR and was also isolated in cell culture from several tissues. Highest viral loads were measured in the gastro-intestinal organs and in the skin. These studies demonstrate that Liz-CrIV circulates in the pet trade in Europe. This virus is capable of infecting both invertebrates and poikilothermic vertebrates, although its involvement in disease in the latter has not been proven.

Adenoviruses in free-ranging Australian bearded dragons (Pogona spp.).

Adenoviruses are a relatively common infection of reptiles globally and are most often reported in captive central bearded dragons (Pogona vitticeps). We report the first evidence of adenoviruses in bearded dragons in their native habitat in Australia. Oral-cloacal swabs and blood samples were collected from 48 free-ranging bearded dragons from four study populations: western bearded dragons (P. minor minor) from Western Australia (n = 4), central bearded dragons (P. vitticeps) from central Australia (n = 2) and western New South Wales (NSW) (n = 29), and coastal bearded dragons (P. barbata) from south-east Queensland (n = 13). Samples were tested for the presence of adenoviruses using a broadly reactive (pan-adenovirus) PCR and a PCR specific for agamid adenovirus-1. Agamid adenovirus-1 was detected in swabs from eight of the dragons from western NSW and one of the coastal bearded dragons. Lizard atadenovirus A was detected in one of the dragons from western NSW. Adenoviruses were not detected in any blood sample. All bearded dragons, except one, were apparently healthy and so finding these adenoviruses in these animals is consistent with bearded dragons being natural hosts for these viruses.

Pathogenesis of Bohle Iridovirus (Genus Ranavirus) in Experimentally Infected Juvenile Eastern Water Dragons ( Intellagama lesueurii lesueurii).

Juvenile eastern water dragons ( Intellagama lesueurii lesueurii) are highly susceptible to infection with Bohle iridovirus (BIV), a species of ranavirus first isolated from ornate burrowing frogs in Townsville, Australia. To investigate the progression of BIV infection in eastern water dragons, 11 captive-bred juveniles were orally inoculated with a dose of 104.33 TCID50 and euthanized at 3, 6, 8, 10, 12, and 14 days postinfection (dpi). Viral DNA was detected via polymerase chain reaction (PCR) in the liver, kidney, and cloacal swabs at 3 dpi. Mild lymphocytic infiltration was observed in the submucosa and mucosa of the tongue and liver at 3 dpi. Immunohistochemistry (IHC) first identified viral antigen in foci of splenic necrosis and in hepatocytes with intracytoplasmic inclusion or rare single-cell necrosis at 6 dpi. By 14 dpi, positive IHC labeling was found in association with lesions in multiple tissues. Selected tissues from an individual euthanized at 14 dpi were probed using in situ hybridization (ISH). The ISH labeling matched the location and pattern detected by IHC. The progression of BIV infection in eastern water dragons, based on lesion severity and virus detection, appears to start in the spleen, followed by the liver, then other organs such as the kidney, pancreas, oral mucosa, and skin. The early detection of ranaviral DNA in cloacal swabs and liver and kidney tissue samples suggests these to be a reliable source of diagnostic samples in the early stage of disease before the appearance of clinical signs, as well as throughout the infection.

FATAL RANAVIRUS INFECTION IN A GROUP OF ZOO-HOUSED MELLER'S CHAMELEONS (TRIOCEROS MELLERI).

A group of five juvenile Meller's chameleons (Trioceros melleri) experienced 100% mortality over a period of 1 mo due to ranavirus infection. The index case was found dead without premonitory signs. The three subsequent cases presented with nonspecific clinical signs (lethargy, decreased appetite, ocular discharge) and were ultimately euthanatized. The final case died after initially presenting with skin lesions. Postmortem examination revealed thin body condition in all five animals and mild coelomic effusion and petechiae affecting the tongue and kidneys of one animal. Microscopically, all animals had multifocal necrosis of the spleen, liver, and kidney; four of five animals had necrosis of the nasal cavity; and two of five had necrosis of adrenal tissue, bone marrow, and skin. Numerous basophilic intracytoplasmic inclusions were present in the liver of all animals and nasal mucosa of three of the five animals. Consensus polymerase chain reaction for herpesvirus and adenovirus were negative, whereas ranavirus quantitative polymerase chain reaction was positive. Virus isolation followed by whole genome sequencing and Bayesian phylogenetic analysis classified the isolates as a strain of frog virus 3 (FV3) most closely related to an FV3 isolate responsible for a previous outbreak in the zoo's eastern box turtle (Terrapene carolina carolina) group. This case series documents the first known occurrence of ranavirus-associated disease in chameleons and demonstrates the potential for interspecies transmission between chelonian and squamate reptiles.

Pneumonia in a Captive Central Bearded Dragon With Concurrent Detection of Helodermatid Adenovirus 2 and a Novel Mycoplasma Species.

A 4-year-old captive male central bearded dragon ( Pogona vitticeps) was presented for recurrent episodic dyspnea and anorexia with occasional expulsion of oral mucoid discharge. Despite empirical antimicrobial therapy and supportive care, the animal died and was submitted for autopsy. Defining histologic features included heterophilic and lymphocytic interstitial pneumonia, with occasional amphophilic intranuclear inclusions and prominent type II pneumocyte hyperplasia. Transmission electron microscopy revealed intranuclear 80-nm, nonenveloped, hexagonal viral particles within pneumocytes. Helodermatid adenovirus 2 (HeAdV2) was determined as the etiologic agent through pan-adenoviral consensus polymerase (PCR) chain reaction and sequencing. Nucleic acid from a novel Mycoplasma sp. (provisionally called Mycoplasma pogonae) was identified by pan-generic PCR targeting the mycoplasma 16S ribosomal RNA gene with sequencing and phylogenetic analysis. As bacteria morphologically consistent with Mycoplasma sp. were not observed by special stains and transmission electron microscopy, the detection of M. pogonae nucleic acid is of indeterminate significance; however, M. pogonae and HeAdV2 coinfection may have exacerbated disease.

Susceptibility of eastern water dragons Intellagama lesueurii lesueurii to Bohle iridovirus.

Ranaviruses infect and have been associated with mass mortality events in fish, amphibians and reptiles and are capable of interclass transmission. Eastern water dragons (EWDs), a semi-aquatic squamate, have an overlapping distribution with several species shown to be susceptible to Bohle iridovirus (BIV). However, this species has not been previously investigated, and no known mass mortalities have occurred in wild populations. Here we report the experimental infection of juvenile EWDs with BIV to investigate a water-dwelling lizards' susceptibility to a ranaviral strain present in northern Queensland, Australia. Lizards were exposed via oral inoculation, intramuscular injection, or cohabitation with orally infected lizards. All exposure methods were effective in establishing an infection as demonstrated by skin lesions and pathological changes in the internal organs. Necrosis, haemorrhage and inflammation were observed histologically in the pancreas, liver, spleen, kidney and submucosa of the gastrointestinal tract of BIV-exposed lizards. Variably sized basophilic intracytoplasmic inclusion bodies were observed in the liver of 6/14 BIV-exposed lizards. Virus was isolated from the liver and kidney of all BIV-infected lizards and confirmed with quantitative PCR (qPCR). The outcome of this study demonstrates that juvenile EWDs are susceptible to BIV, thereby adding Australian lizards to the broad host range of ranaviruses. Furthermore, this study provides additional evidence of BIV's ability to infect different classes of ecothermic vertebrates.

Detection of Papillomavirus DNA in Cutaneous Squamous Cell Carcinoma and Multiple Papillomas in Captive Reptiles.

Papillomaviruses (PVs) are small, non-enveloped DNA viruses that cause mucocutaneous tumours including squamous cell carcinoma (SCC) in man. In animals, evidence supports a causal role for PVs in the development of cutaneous and oral SCC in some species. In reptiles, three cases of papilloma or fibropapilloma have been associated with PV infection, but no association has been reported to date with SCC. Two cases of cutaneous epithelial tumours, multiple papillomas in a spiny-tailed lizard (Uromastyx acanthinura) and SCC in a Dumeril's boa (Acrantophis dumerili), were investigated by polymerase chain reaction. PV DNA was amplified from samples of both lesions. Typical microscopical features suggestive of PV infection (e.g. the presence of koilocytes) were observed in the lesions from the spiny-tailed lizard. This is the first report of an association between PV and SCC in reptiles. Further studies are needed to better clarify the role of PVs in these species and to characterize the PV strains involved.

Discovery and Partial Genomic Characterisation of a Novel Nidovirus Associated with Respiratory Disease in Wild Shingleback Lizards (Tiliqua rugosa).

A respiratory disease syndrome has been observed in large numbers of wild shingleback lizards (Tiliqua rugosa) admitted to wildlife care facilities in the Perth metropolitan region of Western Australia. Mortality rates are reportedly high without supportive treatment and care. Here we used next generation sequencing techniques to screen affected and unaffected individuals admitted to Kanyana Wildlife Rehabilitation Centre in Perth between April and December 2015, with the resultant discovery of a novel nidovirus significantly associated with cases of respiratory disease according to a case definition based on clinical signs. Interestingly this virus was also found in 12% of apparently healthy individuals, which may reflect testing during the incubation period or a carrier status, or it may be that this agent is not causative in the disease process. This is the first report of a nidovirus in lizards globally. In addition to detection of this virus, characterisation of a 23,832 nt segment of the viral genome revealed the presence of characteristic nidoviral genomic elements providing phylogenetic support for the inclusion of this virus in a novel genus alongside Ball Python nidovirus, within the Torovirinae sub-family of the Coronaviridae. This study highlights the importance of next generation sequencing technologies to detect and describe emerging infectious diseases in wildlife species, as well as the importance of rehabilitation centres to enhance early detection mechanisms through passive and targeted health surveillance. Further development of diagnostic tools from these findings will aid in detection and control of this agent across Australia, and potentially in wild lizard populations globally.

Triple infection with agamid adenovirus 1, Encephaliton cuniculi-like microsporidium and enteric coccidia in a bearded dragon (Pogona vitticeps). / Dreifachinfektion mit Agamid-Adenovirus 1, einem Encephalitozoon-cuniculi-ähnlichen Mikrosporidium und Darmkokzidien bei einer Bartagame (Pogona vitticeps).

A 2-month-old juvenile central bearded dragon was presented for anorexia and cachexia. Another specimen from the same cage had died suddenly 2 weeks prior. Fecal analysis revealed a high quantity of Isospora amphiboluri and a few pinworm eggs. Other examinations were not performed and the animal died a few days later despite supportive care. A third individual from the same cage presented with anorexia and a distended cœlom and was euthanized. In this third dragon, histological examination revealed intestinal coccidiosis, basophilic intranuclear inclusions compatible with adenovirus infection, acute hepatic necrosis with intrahepatocytic and intraenteritic organisms typical of microsporidia and renal gout. A PCR confirmed the diagnosis of adenovirosis. Sequencing showed that the PCR product was 100% identical to the corresponding portion of the agamid adenovirus 1 genome. A PCR for the detection of Encephalitozoon (E.) cuniculi was positive. Partial sequencing revealed 100% identity to an E. cuniculi-like organism previously found in bearded dragons. In cases where environmental factors such as poor hygiene or stress can be excluded, the presence of opportunistic pathogens in high numbers can be due to a systemic (viral) infection with temporary immunosuppression.

Detection of SFTS Virus in Ixodes nipponensis and Amblyomma testudinarium (Ixodida: Ixodidae) Collected From Reptiles in the Republic of Korea.

A survey of reptile-associated ticks and their infection status with severe fever with thrombocytopenia syndrome (SFTS) virus was conducted to determine the relative abundance and distribution among lizards, skinks, and snakes in the Republic of Korea (ROK). In total, 132 reptiles, including 49 lizards (two species), 15 skinks (one species), and 68 snakes (eight species) were collected. In total, 84 ixodid ticks belonging to two genera (Ixodes and Amblyomma) were collected from 28/132 (21.2%) lizards, skinks, and snakes. Ixodes nipponensis Kitaoka & Saito was only collected from lizards and skinks, while Amblyomma testudinarium Koch was only collected from snakes. Takydromus wolteri had the highest tick index (0.7; total number ticks/total number collected hosts) among lizards and skinks, while Rhabdophis tigrinus had the highest tick index (2.2) among the snakes. Ixodes nipponensis larvae and nymphs accounted for 11.1% and 88.9%, respectively, of all ticks collected from lizards and skinks, while only A. testudinarium nymphs were collected from snakes. Nymphs of both species of ticks were collected from lizards and skinks from April to October, while I. nipponensis larvae were collected only from September to October. Ixodes nipponensis larvae and nymphs were preferentially attached to the lateral trunk (83.3%) and the foreleg axillae (16.7%) of lizards and skinks. SFTS virus was detected in both I. nipponensis and A. testudinarium collected from lizards and snakes. Phylogenetic analysis of SFTS viruses of ticks collected from two lizards and one snake demonstrated close relationships with SFTS virus strains observed from humans and ticks in the ROK, China, and Japan. These results implicate lizards and snakes as potential hosts of SFTS virus.