Association between sidedness and survival among chemotherapy refractory metastatic colorectal cancer patients treated with trifluridine/tipiracil or regorafenib.

BACKGROUND: The impact of sidedness on survival of later-line treatment in patients with metastatic colorectal cancer (mCRC) is undetermined. This study aimed to investigate the association between sidedness and survival among chemotherapy refractory patients with mCRC treated with trifluridine/tipiracil (TAS-102) or regorafenib or both. PATIENTS AND METHODS: Patients with mCRC treated with TAS-102 or regorafenib between 2015 and 2020 was retrospectively collected. Patients were stratified into TAS-102 first and regorafenib first, then subdivided into TAS-102 followed by regorafenib (T-R) and regorafenib followed by TAS-102 (R-T) groups. The oncologic outcomes were presented with time-to-treatment failure (TTF) and overall survival (OS). RESULTS: After matching, 376 TAS-102 patients and 376 regorafenib patients were included for outcomes comparison. TTF had insignificant differences while OS was significantly different between TAS-102 and regorafenib groups. Median TTF and OS were 1.9 months versus 2.0 months (P = .701) and 9.1 months versus 7.0 months (P = .008) in TAS-102 and regorafenib, respectively. The OS benefits were consistent regardless primary tumor location. Subgroup analysis with 174 T-R patients and 174 R-T patients was investigated for treatment sequences. TTF and OS had significant differences in both groups. Median TTF and OS were 8.5 months versus 6.3 months (P = .001) and 14.4 months versus 12.6 months (P = .035) in T-R and R-T groups, respectively. The TTF and OS benefits were persisted regardless primary tumor location. CONCLUSION: TAS-102 first provided a better survival benefit in chemotherapy refractory patients with mCRC across all sidedness. Further prospective studies are warranted to validate our conclusions.

Winter is coming-The role of seasonality through the lens of the rodent virome.

Rodent virus communities (viromes) are overrepresented with zoonotic viruses, and as such are a key host system for the study of zoonotic viruses. However, the extent of viral diversity beyond characterized zoonotic viruses, and the factors that modulate the viromes of rodents remain opaque. In this issue of Molecular Ecology, Raghwani et al. (2023) use rodents as a model to understand the role of seasonality in dictating virome abundance and composition-a factor known to play an important role in most animal one-host, one-pathogen systems. These data are not only highly relevant to rodents, but have broad applications across understanding and disentangling animal virome ecology.

Additional single dose GnRH agonist during luteal phase support may improve live birth rate in GnRHa-HRT frozen-thawed embryo transfer cycle: a retrospective cohort study.

BACKGROUND: GnRH agonist (GnRHa) has been reported to have direct effects and functional roles in the endometrium and embryos. Several meta-analyses have shown that GnRHa administration in the luteal phase improved the live birth rate or pregnancy rate in both fresh and frozen embryo transfer (FET) cycles. The aim of this study was to investigate whether luteal GnRHa administration could also improve in vitro fertilization (IVF) outcomes in patients undergoing hormone replacement therapy (HRT) cycles with GnRHa suppression. METHODS: The retrospective cohort study included a total of 350 patients undergoing GnRHa-HRT FET cycles. The study group included 179 patients receiving an additional single dose of GnRHa in the luteal phase following embryo transfer. A total of 171 patients in the control group did not receive luteal GnRHa. The baseline and cycle characteristics and reproductive outcomes were compared between the two groups. RESULTS: Baseline and cycle characteristics were similar between the two groups, except lower AMH levels were found in the luteal GnRHa group than in the control group. The luteal GnRHa group had a significantly higher ongoing pregnancy rate and live birth rate than the control group. The multivariate analysis revealed that luteal GnRHa administration was positively associated with ongoing pregnancy (OR 2.04, 95% CI 1.20-3.47, P = 0.008) and live birth (OR 2.03, 95% CI 1.20-3.45, P = 0.009). When the subgroup of patients with recurrent implantation failure was analyzed, the multivariate analysis also showed that luteal GnRHa administration had beneficial effects on ongoing pregnancy (OR 4.55, 95% CI 1.69-12.30, P = 0.003) and live birth (OR 4.30, 95% CI 1.59-11.65, P = 0.004). CONCLUSIONS: Our data suggest that the addition of one luteal dose of GnRHa may improve the live birth rate in patients undergoing the GnRHa-HRT protocol.

Metatranscriptomic Analysis of Virus Diversity in Urban Wild Birds with Paretic Disease.

Wild birds are major natural reservoirs and potential dispersers of a variety of infectious diseases. As such, it is important to determine the diversity of viruses they carry and use this information to help understand the potential risks of spillover to humans, domestic animals, and other wildlife. We investigated the potential viral causes of paresis in long-standing, but undiagnosed, disease syndromes in wild Australian birds. RNA from diseased birds was extracted and pooled based on tissue type, host species, and clinical manifestation for metagenomic sequencing. Using a bulk and unbiased metatranscriptomic approach, combined with clinical investigation and histopathology, we identified a number of novel viruses from the families Astroviridae, Adenoviridae, Picornaviridae, Polyomaviridae, Paramyxoviridae, Parvoviridae, and Circoviridae in common urban wild birds, including Australian magpies, magpie larks, pied currawongs, Australian ravens, and rainbow lorikeets. In each case, the presence of the virus was confirmed by reverse transcription (RT)-PCR. These data revealed a number of candidate viral pathogens that may contribute to coronary, skeletal muscle, vascular, and neuropathology in birds of the Corvidae and Artamidae families and neuropathology in members of the Psittaculidae The existence of such a diverse virome in urban avian species highlights the importance and challenges in elucidating the etiology and ecology of wildlife pathogens in urban environments. This information will be increasingly important for managing disease risks and conducting surveillance for potential viral threats to wildlife, livestock, and human health.IMPORTANCE Wildlife naturally harbor a diverse array of infectious microorganisms and can be a source of novel diseases in domestic animals and human populations. Using unbiased RNA sequencing, we identified highly diverse viruses in native birds from Australian urban environments presenting with paresis. This research included the clinical investigation and description of poorly understood recurring syndromes of unknown etiology: clenched claw syndrome and black and white bird disease. As well as identifying a range of potentially disease-causing viral pathogens, this study describes methods that can effectively and efficiently characterize emergent disease syndromes in free-ranging wildlife and promotes further surveillance for specific pathogens of potential conservation and zoonotic concern.

Functional analysis of the short isoform of orf virus protein OV20.0.

UNLABELLED: Orf virus (ORFV) OV20.0L is an ortholog of vaccinia virus (VACV) gene E3L. The function of VACV E3 protein as a virulence factor is well studied, but OV20.0 has received less attention. Here we show that like VACV E3L, OV20.0L encodes two proteins, a full-length protein and a shorter form (sh20). The shorter sh20 is an N-terminally truncated OV20.0 isoform generated when a downstream AUG codon is used for initiating translation. These isoforms differed in cellular localization, with full-length OV20.0 and sh20 found throughout the cell and predominantly in the cytoplasm, respectively. Nonetheless, both OV20.0 isoforms were able to bind double-stranded RNA (dsRNA)-activated protein kinase (PKR) and dsRNA. Moreover, both isoforms strongly inhibited PKR activation as shown by decreased phosphorylation of the translation initiation factor eIF2α subunit and protection of Sindbis virus infection against the activity of interferon (IFN). In spite of this apparent conservation of function in vitro, a recombinant ORFV that was able to express only the sh20 isoform was attenuated in a mouse model. IMPORTANCE: The OV20.0 protein of orf virus (ORFV) has two isoforms and contributes to virulence, but the roles of the two forms are not known. This study shows that the shorter isoform (sh20) arises due to use of a downstream initiation codon and is amino-terminally truncated. The sh20 form also differs in expression kinetics and cellular localization from full-length OV20.0. Similar to the full-length isoform, sh20 is able to bind dsRNA and PKR, inactivate PKR, and thus act as an antagonist of the interferon response in vitro. In vivo, however, wild-type OV20.0 could not be replaced with sh20 alone without a loss of virulence, suggesting that the functions of the isoforms are not simply redundant.

Molecular cloning and sequencing analysis of the interferon ß from Coturnix.

One pair of primers was designed according to Gallus and Meleagris gallopavo interferon ß (IFN-ß) sequences published in GenBank. The primers and RNA extraction from the spleen of Coturnix were used to amplify Coturnix IFN-ß cDNA by real-time polymerase chain reaction (RT-PCR). The product was cloned into pEasy-T1 vector. Evaluating recombinant plasmid by PCR and restriction enzyme digestion. Sequence the cloning sequences, comparing the sequencing results by NCBI. We successfully got a Coturnix IFN-ß partial sequence. The sequence was subtyped and put to homologous analysis. The results suggested the homology of IFN-ß gene of Coturnix and gene of Coturnix and chicken (88.7%), the homology of IFN-ß gene of Coturnix and chicken (88.7%), the homology of IFN-ß gene of Coturnix and Anas platyrhynchos (72.5%), the homology of IFN-ß sequence registered in GenBank. The analysis of the genetic tree showed that the relationship of Coturnix and chicken IFN-ß had a high homology. It can be seen that in this study we successfully got a partial sequence of IFN-ß of quail.

Molecular cloning and sequencing analysis of the interferon receptor (IFNAR-1) from Columba livia.

OBJECTIVE: Partial sequence cloning of interferon receptor (IFNAR-1) of Columba livia. MATERIAL AND METHODS: In order to obtain a certain length (630 bp) of gene, a pair of primers was designed according to the conserved nucleotide sequence of Gallus (EU477527.1) and Taeniopygia guttata (XM_002189232.1) IFNAR-1 gene fragment that was published by GenBank. Special primers were designed by the Race method to amplify the 3'terminal cDNA. RESULTS: The Columba livia IFNAR-1 displayed 88.5%, 80.5% and 73.8% nucleotide identity to Falco peregrinus, Gallus and Taeniopygia guttata, respectively. Phylogenetic analysis of the IFNAR1 gene showed that the relationship of Columba livia, Falco peregrinus and chicken had high homology. CONCLUSIONS: We successfully obtained a Columba livia IFNAR-1 gene partial sequence. Analysis of the genetic tree showed that the relationship of Columba livia and Falco peregrinus IFNAR-1 had high homology. This result can be used as reference for further research and practical application.

Genetic and antigenic analysis of mink's immunoglobulin G Fc region.

Mink's immunoglobulin (Ig) G Fc gene was cloned, the gene was analyzed by phylogenetic analysis, and western blot was done to prove that the detection of distemper and canine parvovirus in dogs and minks can be universal. In order to get the certain length of Fc segment gene, a pair of primers is designed, which according to the Fc segment gene sequences of mink's IgG (L07789) published by GenBank, extracted total RNA from the spleen of minks and amplified it by RT-PCR. The results showed that the Fc segment gene contained 606 bp. Then it was sequenced after the amplified fragments were cloned into the vector PEasy-T1. Then the genetic evolution was analyzed. An antibody hybridization test was done through western blot. The results showed that nucleotide sequence homologies between minks and canines were 85%, and amino acid sequence homologies between minks and canines were 80.5%. Mink IgG heavy chain can effectively combine to anti-dog IgG by western blot. It was concluded that mink's and dog's IgG Fc had the closest relationship in mammalian through the analysis of the genetic evolution. Based on the above analysis and related literature, we concluded that we could detect mink diseases with a dog diagnosis reagent, or treat mink diseases with dog antiserum.

Distribution of the type I interferon in different organs of chicken digestive system.

OBJECTIVE: Distribution of the type I interferon in different organs of the chicken digestive system. MATERIAL AND METHODS: In order to obtain a certain length (274 bp) of a fragment, a pair of primers was designed according to the conserved nucleotide sequence of gallus IFNAR-1 (EU477527.1) fragment that was published by the GenBank. The fragment was cloned by pEASY-T1 and amplified by relative fluorescence quantitative PCR with SYBR Green I; according to the results, we made a standard curve. The experimental group took interferon orally, while the control group took equivalent physiological saline orally, then we used relative fluorescence quantitative PCR to detect the copies of the IFNAR-1 gene of the palate, tongue, esophagus, craw, glandular stomach, duodenum and rectum of the experimental group and control group. Copies of the IFNAR-1 gene of those organs were calculated by Ct value. Finally, all the chickens were infected with the Newcastle Disease Virus after 48 hours. RESULTS: The results showed that the IFNAR-1 gene had the most expression in the esophagus. In the experiment of interferon antiviral activity detection, the chickens which took interferon orally were healthier than the other group. CONCLUSIONS: It is confirmed that the interferon receptor did exist in the digestive organs. However, according to the physical and chemical properties of interferon, interferon is easily inactivated in the acid and alkali environment, by pepsin and trypsin, so the absorption site for interferon exists in organs above the craw, especially in the esophagus and tongue.

The New South Wales Mouse Plague 2020-2021: A One Health description.

A mouse plague occurred in Eastern Australia from spring 2020 to winter 2021, impacting an area of around 180,000 km2. It harmed human physical and psychological health, damaged the natural and built environment, and endangered farmed, domestic and native animals. However, the mouse plague was overshadowed by the COVID-19 pandemic, especially as the end of the plague coincided with the arrival and surge of the COVID-19 delta strain in rural New South Wales (NSW). In this article, we systematically overview the multiple impacts of the plague and highlight their complex interactions. Using a One Health framework, we comprehensively review the i) human, ii) animal and iii) environmental impacts including economic dimensions. Given the damage that the mouse plague caused to infrastructure, we consider the environment from two perspectives: the natural and the built environment. This One Health description of the 2020-2021 mouse plague identifies priorities for preparedness, response and recovery at local, regional land levels to inform response and management of future mouse plague events in Australia. It also highlights the need for ongoing collaboration between researchers and practitioners in the human, animal and environmental health sectors.

Development of a Machine Learning Algorithm to Correlate Lumbar Disc Height on X-rays with Disc Bulging or Herniation.

Lumbar disc bulging or herniation (LDBH) is one of the major causes of spinal stenosis and related nerve compression, and its severity is the major determinant for spine surgery. MRI of the spine is the most important diagnostic tool for evaluating the need for surgical intervention in patients with LDBH. However, MRI utilization is limited by its low accessibility. Spinal X-rays can rapidly provide information on the bony structure of the patient. Our study aimed to identify the factors associated with LDBH, including disc height, and establish a clinical diagnostic tool to support its diagnosis based on lumbar X-ray findings. In this study, a total of 458 patients were used for analysis and 13 clinical and imaging variables were collected. Five machine-learning (ML) methods, including LASSO regression, MARS, decision tree, random forest, and extreme gradient boosting, were applied and integrated to identify important variables for predicting LDBH from lumbar spine X-rays. The results showed L4-5 posterior disc height, age, and L1-2 anterior disc height to be the top predictors, and a decision tree algorithm was constructed to support clinical decision-making. Our study highlights the potential of ML-based decision tools for surgeons and emphasizes the importance of L1-2 disc height in relation to LDBH. Future research will expand on these findings to develop a more comprehensive decision-supporting model.

Helicobacter pylori and Its Treatment Impact on Immune-Mediated Ocular Diseases.

PURPOSE: Helicobacter pylori (HP), which colonizes exclusively in the gastrointestinal tract, has been reported to dysregulate the immune response and gives rise to several extra-gastrointestinal autoimmune disorders. However, the relationship between HP and immune-mediated ocular diseases remains ambiguous. This study aims to clarify the association between immune-mediated ocular diseases and HP infection, as well as the impact of HP treatment on the incidence of immune-mediated ocular diseases. METHODS: This is a retrospective population-based study using National Health Insurance Research Database in Taiwan. Patients with newly diagnosed peptic ulcer disease or HP infection between 2009 and 2015 were identified as HP group and compared to the non-HP group with one-to-one exact matching. Moreover, the incident risk of immune-mediated ocular diseases and its two subgroups (ocular surface and orbital inflammation group, intraocular inflammation group) were compared in HP patients with or without treatment. RESULTS: A total of 1,030,119 subjects in the non-HP group and 1,030,119 patients in the HP group were enrolled. The incidence rate of immune-mediated ocular diseases was significantly higher in the HP group (95% confidence interval (CI): 2.534-2.547). The incident rate ratio was significantly higher in HP with treatment than without treatment (HR: 1.654, 95% CI: 1.641-1.668). The Cox proportional hazards regression model demonstrated a significantly increased HR of immune-mediated ocular diseases in HP treated group (HR: 2.265, 95% CI: 2.024-2.534) and less increased HR in HP non-treated group (HR: 1.427, 95% CI: 1.273-1.598) when comparing to non-HP group. Subgroup analysis demonstrated a significantly higher incidence rate of ocular surface and orbital inflammation as well as intraocular inflammation in the HP group. CONCLUSION: This study illustrated a higher incidence of immune-mediated ocular diseases in HP infection, and a heightened risk following HP eradication.

Gammaretroviruses, novel viruses and pathogenic bacteria in Australian bats with neurological signs, pneumonia and skin lesions.

More than 70 bat species are found in mainland Australia. While most studies of bat viromes focus on sampling seemingly healthy individuals, little is known about the viruses and bacteria associated with diseased bats. We performed traditional diagnostic techniques and metatranscriptomic sequencing on tissue samples from 43 Australian bats, comprising three flying fox (Pteropodidae) and two microbat species experiencing a range of disease syndromes, including mass mortality, neurological signs, pneumonia and skin lesions. Of note, we identified the recently discovered Hervey pteropid gammaretrovirus in a bat with lymphoid leukemia, with evidence of replication consistent with an exogenous virus. The possible association of Hervey pteropid gammaretrovirus with lymphoid leukemia clearly merits additional investigation. One novel picornavirus and at least three new astroviruses and bat pegiviruses were also identified in a variety of tissue types, as well as a number of likely bacterial pathogens or opportunistic infections, most notably Pseudomonas aeruginosa.

Data-driven, two-stage machine learning algorithm-based prediction scheme for assessing 1-year and 3-year mortality risk in chronic hemodialysis patients.

Life expectancy is likely to be substantially reduced in patients undergoing chronic hemodialysis (CHD). However, machine learning (ML) may predict the risk factors of mortality in patients with CHD by analyzing the serum laboratory data from regular dialysis routine. This study aimed to establish the mortality prediction model of CHD patients by adopting two-stage ML algorithm-based prediction scheme, combined with importance of risk factors identified by different ML methods. This is a retrospective, observational cohort study. We included 800 patients undergoing CHD between December 2006 and December 2012 in Shin-Kong Wu Ho-Su Memorial Hospital. This study analyzed laboratory data including 44 indicators. We used five ML methods, namely, logistic regression (LGR), decision tree (DT), random forest (RF), gradient boosting (GB), and eXtreme gradient boosting (XGB), to develop a two-stage ML algorithm-based prediction scheme and evaluate the important factors that predict CHD mortality. LGR served as a bench method. Regarding the validation and testing datasets from 1- and 3-year mortality prediction model, the RF had better accuracy and area-under-curve results among the five different ML methods. The stepwise RF model, which incorporates the most important factors of CHD mortality risk based on the average rank from DT, RF, GB, and XGB, exhibited superior predictive performance compared to LGR in predicting mortality among CHD patients over both 1-year and 3-year periods. We had developed a two-stage ML algorithm-based prediction scheme by implementing the stepwise RF that demonstrated satisfactory performance in predicting mortality in patients with CHD over 1- and 3-year periods. The findings of this study can offer valuable information to nephrologists, enhancing patient-centered decision-making and increasing awareness about risky laboratory data, particularly for patients with a high short-term mortality risk.

Expanding the range of the respiratory infectome in Australian feedlot cattle with and without respiratory disease using metatranscriptomics.

BACKGROUND: Bovine respiratory disease (BRD) is one of the most common diseases in intensively managed cattle, often resulting in high morbidity and mortality. Although several pathogens have been isolated and extensively studied, the complete infectome of the respiratory complex consists of a more extensive range unrecognised species. Here, we used total RNA sequencing (i.e., metatranscriptomics) of nasal and nasopharyngeal swabs collected from animals with and without BRD from two cattle feedlots in Australia. RESULTS: A high abundance of bovine nidovirus, influenza D, bovine rhinitis A and bovine coronavirus was found in the samples. Additionally, we obtained the complete or near-complete genome of bovine rhinitis B, enterovirus E1, bovine viral diarrhea virus (sub-genotypes 1a and 1c) and bovine respiratory syncytial virus, and partial sequences of other viruses. A new species of paramyxovirus was also identified. Overall, the most abundant RNA virus, was the bovine nidovirus. Characterisation of bacterial species from the transcriptome revealed a high abundance and diversity of Mollicutes in BRD cases and unaffected control animals. Of the non-Mollicutes species, Histophilus somni was detected, whereas there was a low abundance of Mannheimia haemolytica. CONCLUSION: This study highlights the use of untargeted sequencing approaches to study the unrecognised range of microorganisms present in healthy or diseased animals and the need to study previously uncultured viral species that may have an important role in cattle respiratory disease. Video Abstract.

The admixed brushtail possum genome reveals invasion history in New Zealand and novel imprinted genes.

Combining genome assembly with population and functional genomics can provide valuable insights to development and evolution, as well as tools for species management. Here, we present a chromosome-level genome assembly of the common brushtail possum (Trichosurus vulpecula), a model marsupial threatened in parts of their native range in Australia, but also a major introduced pest in New Zealand. Functional genomics reveals post-natal activation of chemosensory and metabolic genes, reflecting unique adaptations to altricial birth and delayed weaning, a hallmark of marsupial development. Nuclear and mitochondrial analyses trace New Zealand possums to distinct Australian subspecies, which have subsequently hybridised. This admixture allowed phasing of parental alleles genome-wide, ultimately revealing at least four genes with imprinted, parent-specific expression not yet detected in other species (MLH1, EPM2AIP1, UBP1 and GPX7). We find that reprogramming of possum germline imprints, and the wider epigenome, is similar to eutherian mammals except onset occurs after birth. Together, this work is useful for genetic-based control and conservation of possums, and contributes to understanding of the evolution of novel mammalian epigenetic traits.

Meta-transcriptomic analysis of the virome and microbiome of the invasive Indian myna (Acridotheres tristis) in Australia.

Invasive species exert a serious impact on native fauna and flora and have become the target of eradication and management efforts worldwide. Invasive avian species can also be important pathogen reservoirs, although their viromes and microbiomes have rarely been studied. As one of the top 100 invasive pest species globally, the expansion of Indian mynas (Acridotheres tristis) into peri-urban and rural environments, in conjunction with increasing free-ranging avian agricultural practices, may increase the risk of microbial pathogens jumping species boundaries. Herein, we used a meta-transcriptomic approach to explore the microbes present in brain, liver and large intestine of 16 invasive Indian myna birds in Sydney, Australia. From this, we discovered seven novel viruses from the families Adenoviridae, Caliciviridae, Flaviviridae, Parvoviridae and Picornaviridae. Interestingly, each of the novel viruses identified shared less than 80% genomic similarity with their closest relatives from other avian species, indicative of a lack of detectable virus transmission between invasive mynas to native or domestic species. Of note, we also identified two coccidian protozoa, Isospora superbusi and Isospora greineri, from the liver and gut tissues of mynas. Overall, these data demonstrate that invasive mynas can harbor a diversity of viruses and other microorganisms such that ongoing pathogen surveillance in this species is warranted.

Characterization of the Gut Microbiome and Resistomes of Wild and Zoo-Captive Macaques.

Rhesus macaques (Macaca mulatta) are the most widely distributed species of Old World monkey and are frequently used as animal models to study human health and disease. Their gastrointestinal microbial community likely plays a major role in their physiology, ecology and evolution. Herein, we compared the fecal microbiome and antibiotic resistance genes in 15 free-ranging and 81 zoo-captive rhesus macaques sampled from two zoos in China, using both 16S amplicon sequencing and whole genome shotgun DNA sequencing approaches. Our data revealed similar levels of microbial diversity/richness among the three groups, although the composition of each group differed significantly and were particularly marked between the two zoo-captive and one wild groups. Zoo-captive animals also demonstrated a greater abundance and diversity of antibiotic genes. Through whole genome shotgun sequencing we also identified a mammalian (simian) associated adenovirus. Overall, this study provides a comprehensive analysis of resistomes and microbiomes in zoo-captive and free-ranging monkeys, revealing that semi-captive wildlife might harbor a higher diversity of antimicrobial resistant genes.

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.

Novel hepaci- and pegi-like viruses in native Australian wildlife and non-human primates.

The Flaviviridae family of positive-sense RNA viruses contains important pathogens of humans and other animals, including Zika virus, dengue virus, and hepatitis C virus. The Flaviviridae are currently divided into four genera-Hepacivirus, Pegivirus, Pestivirus, and Flavivirus-each with a diverse host range. Members of the genus Hepacivirus are associated with an array of animal species, including humans, non-human primates, other mammalian species, as well as birds and fish, while the closely related pegiviruses have been identified in a variety of mammalian taxa, also including humans. Using a combination of total RNA and whole-genome sequencing we identified four novel hepaci-like viruses and one novel variant of a known hepacivirus in five species of Australian wildlife. The hosts infected comprised native Australian marsupials and birds, as well as a native gecko (Gehyra lauta). From these data we identified a distinct marsupial clade of hepaci-like viruses that also included an engorged Ixodes holocyclus tick collected while feeding on Australian long-nosed bandicoots (Perameles nasuta). Distinct lineages of hepaci-like viruses associated with geckos and birds were also identified. By mining the SRA database we similarly identified three new hepaci-like viruses from avian and primate hosts, as well as two novel pegi-like viruses associated with primates. The phylogenetic history of the hepaci- and pegi-like viruses as a whole, combined with co-phylogenetic analysis, provided support for virus-host co-divergence over the course of vertebrate evolution, although with frequent cross-species virus transmission. Overall, our work highlights the diversity of the Hepacivirus and Pegivirus genera as well as the uncertain phylogenetic distinction between.