A novel group of avian astroviruses from Neotropical passerine birds broaden the diversity and host range of Astroviridae

Metagenomics is helping to expand the known diversity of viruses, especially of those with poorly studied hosts in remote areas. The Neotropical region harbors a considerable diversity of avian species that may play a role as both host and short-distance vectors of unknown viruses. Viral metagenomics of cloacal swabs from 50 Neotropical birds collected in French Guiana revealed the presence of four complete astrovirus genomes. They constitute an early diverging novel monophyletic clade within the Avastrovirus phylogeny, representing a putative new astrovirus species (provisionally designated as Avastrovirus 5) according to the International Committee on Taxonomy of Viruses (ICTV) classifcation criteria. Their genomic organization shares some characteristics with Avastrovirus but also with Mamastrovirus. The pan-astrovirus RT-PCR analysis of the cloacal samples of 406 wild Neotropical birds showed a community-level prevalence of 4.9% (5.1% in passerines, the highest described so far in this order of birds). By screening birds of a remote region, we expanded the known host range of astroviruses to the avian families Cardinalidae, Conopophagidae, Furnariidae, Thamnophilidae, Turdidae and Tyrannidae. Our results provide important frst insights into the unexplored viral communities, the ecology, epidemiology and features of host-pathogen interactions that shape the evolution of avastroviruses in a remote Neotropical rainforest

A Novel and Divergent Gyrovirus with Unusual Genomic Features Detected in Wild Passerine Birds from a Remote Rainforest in French Guiana

Sequence-independent amplification techniques have become important tools for virus discovery, metagenomics, and exploration of viral diversity at the global scale, especially in remote areas. Here, we describe the detection and genetic characterization of a novel gyrovirus, named GyV11, present in cloacal, oral, and blood samples from neotropical wild birds in French Guiana. The molecular epidemiology revealed the presence of GyV11 only in passerine birds from three different species at a low prevalence (0.73%). This is the first characterization and prevalence study of a gyrovirus carried out in resident wild bird populations in a remote region, and provides evidence of the fecal–oral route transmission and local circulation of the virus. The molecular phylogeny of gyroviruses reveals the existence of two distinct gyrovirus lineages in which GyV11 is phylogenetically distinct from previously reported gyroviruses. Furthermore, GyV11 is placed basal in the gyrovirus phylogeny, likely owing to its ancestral origin and marked divergence. This study also provides important insights into the ecology, epidemiology, and genomic features of gyroviruses in a remote neotropical rainforest. The pathogenesis of this virus in avian species or whether GyV11 can infect humans and/or chickens needs to be further investigated

A Review on the Prevalence of Poxvirus Disease in Free-Living and Captive Wild Birds

Avian pox is a widespread infection in birds caused by genus Avipoxvirus pathogens. It is a noteworthy, potentially lethal disease to wild and domestic hosts. It can produce two different conditions: cutaneous pox, and diphtheritic pox. Here, we carry out an exhaustive review of all cases of avian pox reported from wild birds to analyze the effect and distribution in different avian species. Avian poxvirus strains have been detected in at least 374 wild bird species, a 60% increase on a 1999 review on avian pox hosts. We also analyze epizootic cases and if this disease contributes to wild bird population declines. We frequently observe very high prevalence in wild birds in remote island groups, e.g., Hawaii, Galapagos, etc., representing a major risk for the conservation of their unique endemic avifauna. However, the difference in prevalence between islands and continents is not significant given the few available studies. Morbidity and mortality can also be very high in captive birds, due to high population densities. However, despite the importance of the disease, the current detection rate of new Avipoxvirus strains suggests that diversity is incomplete for this group, and more research is needed to clarify its real extent, particularly in wild birds

Turnip Mosaic Virus Nanoparticles: A Versatile Tool in Biotechnology

15 Pág.Within plant molecular farming (PMF), the use of viruses and virus-like particles (VLPs) is increasingly gaining momentum due to the vast array of possibilities they offer. In addition to the wide application of viruses as vectors of genes for their transient expression in plants, viral particles are being exploited as natural nanoparticles amenable to production in plants and functionalization with very different purposes. One important group of plant viruses exploited in this context is formed by viruses with flexuous elongated virions of a high aspect ratio. One of these viruses is turnip mosaic virus (TuMV), a potyvirus. TuMV virions and VLPs have been produced in plants in different functionalized manners for an ample range of applications. They have also been chemically functionalized “in vitro” after purification of their natural unmodified forms. The chapter describes and discusses the work carried out so far for the development and applications of TuMV in PMF nanobiotechnology.Work on TuMV nanobiotechnological functionalization has benefted from different grants obtained along the years. These are P2018/BAA-4574, COV20/00114, and PanGreen-CM from the Comunidad de Madrid; RTA2015-00017-00-00 from INIA; and ARIMNet-2 618127, an ERANet project. Daniel A. Truchado is a postdoctoral associate funded by a contract of the “Margarita Salas” program of the Spanish Ministry of Universities. Sara Rincón is funded by the P2018/BAA-4574 grant.Peer reviewe

Isopeptide Bonding In Planta Allows Functionalization of Elongated Flexuous Proteinaceous Viral Nanoparticles, including Non-Viable Constructs by Other Means

11 Pág.Plant viral nanoparticles (VNPs) have become an attractive platform for the development of novel nanotools in the last years because of their safety, inexpensive production, and straightforward functionalization. Turnip mosaic virus (TuMV) is one example of a plant-based VNP used as a nanobiotechnological platform either as virions or as virus-like particles (VLPs). Their functionalization mainly consists of coating their surface with the molecules of interest via chemical conjugation or genetic fusion. However, because of their limitations, these two methods sometimes result in non-viable constructs. In this paper, we applied the SpyTag/SpyCatcher technology as an alternative for the functionalization of TuMV VLPs with peptides and proteins. We chose as molecules of interest the green fluorescent protein (GFP) because of its good traceability, as well as the vasoactive intestinal peptide (VIP), given the previous unsuccessful attempts to functionalize TuMV VNPs by other methods. The successful conjugation of VLPs to GFP and VIP using SpyTag/SpyCatcher was confirmed through Western blot and electron microscopy. Moreover, the isopeptide bond between SpyTag and SpyCatcher occurred in vivo in co-agroinfiltrated Nicotiana benthamiana plants. These results demonstrated that SpyTag/SpyCatcher improves TuMV functionalization compared with previous approaches, thus implying the expansion of the application of the technology to elongated flexuous VNPs.This work was conceived as transversal to efforts experimentally aimed to TuMV nanobiotechnological functionalizations. As a consequence, it has benefited from different grants obtained along the years. These are P2018/BAA-4574, COV20/00114, and PanGreen-CM from the Comunidad de Madrid; RTA2015-00017-00-00 from INIA; and ARIMNet-2 618127, an ERANet project. D.A.T. was supported by a Margarita Salas postdoctoral grant funded by the Spanish Ministry of Universities and UCM (CT31/21).Peer reviewe

Antigen-functionalized turnip mosaic virus nanoparticles increase antibody sensing in saliva. A case study with SARS-CoV-2 RBD

10 Pág.Nanoparticles derived from plant viruses play an important role in nanomedicine due to their biocompatibility, self-assembly and easily-modifiable surface. In this study, we developed a novel platform for increasing antibody sensing using viral nanoparticles derived from turnip mosaic virus (TuMV) functionalized with SARS-CoV-2 receptor binding domain (RBD) through three different methods: chemical conjugation, gene fusion and the SpyTag/SpyCatcher technology. Even though gene fusion turned out to be unsuccessful, the other two constructs were proven to significantly increase antibody sensing when tested with saliva of patients with different infection and vaccination status to SARS-CoV-2. Our findings show the high potential of TuMV nanoparticles in the fields of diagnostics and immunodetection, being especially attractive for the development of novel antibody sensing devices.This work was mostly funded by grant COV20/00114 from Comunidad de Madrid to FP. We want to particularly acknowledge the donors and the Biobank Biobanco Hospital Universitario de La Princesa (Madrid) (ISCIII B.0000763) for their collaboration. We also thank Agrenvec S.L (Tres Cantos, Madrid, Spain) for the generous gift of its plant-made RBD. DAT was supported by a Margarita Salas postdoctoral grant funded by the Spanish Ministry of Universities and UCM (CT31/21). SR was funded by a contract under grant P2018/BAA-4574 from the Comunidad de Madrid. This work was part of the Final Grade Thesis of CM for graduation in biotechnology by the UPM. The CBGP was granted ‘Severo Ochoa’ Distinctions of Excellence by the Spanish Ministry of Science and Innovation (SEV- 2016-0672 and CEX2020-000999-S).With funding from the Spanish government through the ‘Severo Ochoa Centre of Excellence’ accreditation (CEX2020‐000999‐S)Peer reviewe

Genomic characterization of the first oral avian papillomavirus in a colony of breeding canaries (Serinus canaria)

Papillomaviruses are non-enveloped, DNA viruses that infect skin and mucosa of a wide variety of vertebrates, causing neoplasias or simply persisting asymptomatically. Avian papillomaviruses, with six fully sequenced genomes, are the second most studied group after mammalian papillomaviruses. In this study, we describe the first oral avian papillomavirus, detected in the tongue of a dead Yorkshire canary (Serinus canaria) and in oral swabs of the same bird and other two live canaries from an aviary in Madrid, Spain. Its genome is 8,071 bp and presents the canonical papillomavirus architecture with six early (E6, E7, E1, E9, E2, E4) and two late open reading frames (L1 and L2) and a long control region between L1 and E6. This new avian papillomavirus L1 gene shares a 64% pairwise identity with FcPV1 L1, so it has been classified as a new species (ScPV1) within the Ethapapillomavirus genus. Although the canary died after showing breathing problems, there is no evidence that the papillomavirus caused those symptoms so it could be part of the oral microbiota of the birds. Hence, future investigations are needed to evaluate the clinical relevance of the virus

Comparative metagenomics of Palearctic and Neotropical avian cloacal viromes reveal geographic bias in virus discovery

Our understanding about viruses carried by wild animals is still scarce. The viral diversity of wildlife may be best described with discovery-driven approaches to the study of viral diversity that broaden research efforts towards non-canonical hosts and remote geographic regions. Birds have been key organisms in the transmission of viruses causing important diseases, and wild birds are threatened by viral spillovers associated with human activities. However, our knowledge of the avian virome may be biased towards poultry and highly pathogenic diseases. We describe and compare the fecal virome of two passerine-dominated bird assemblages sampled in a remote Neotropical rainforest in French Guiana (Nouragues Natural Reserve) and a Mediterranean forest in central Spain (La Herrería). We used metagenomic data to quantify the degree of functional and genetic novelty of viruses recovered by examining if the similarity of the contigs we obtained to reference sequences differed between both locations. In general, contigs from Nouragues were significantly less similar to viruses in databases than contigs from La Herrería using Blastn but not for Blastx, suggesting that pristine regions harbor a yet unknown viral diversity with genetically more singular viruses than more studied areas. Additionally, we describe putative novel viruses of the families Picornaviridae, Reoviridae and Hepeviridae. These results highlight the importance of wild animals and remote regions as sources of novel viruses that substantially broaden the current knowledge of the global diversity of viruses