Heritability and genome-wide association study of vaccine-induced immune response in Beagles: A pilot study.

Both genetic and non-genetic factors contribute to individual variation in the immune response to vaccination. Understanding how genetic background influences variation in both magnitude and persistence of vaccine-induced immunity is vital for improving vaccine development and identifying possible causes of vaccine failure. Dogs provide a relevant biomedical model for investigating mammalian vaccine genetics; canine breed structure and long linkage disequilibrium simplify genetic studies in this species compared to humans. The objective of this study was to estimate the heritability of the antibody response to vaccination against viral and bacterial pathogens, and to identify genes driving variation of the immune response to vaccination in Beagles. Sixty puppies were immunized following a standard vaccination schedule with an attenuated combination vaccine containing antigens for canine adenovirus type 2, canine distemper virus, canine parainfluenza virus, canine parvovirus, and four strains of Leptospira bacteria. Serum antibody measurements for each viral and bacterial component were measured at multiple time points. Heritability estimations and GWAS were conducted using SNP genotypes at 279,902 markers together with serum antibody titer phenotypes. The heritability estimates were: (1) to Leptospira antigens, ranging from 0.178 to 0.628; and (2) to viral antigens, ranging from 0.199 to 0.588. There was not a significant difference between overall heritability of vaccine-induced immune response to Leptospira antigens compared to viral antigens. Genetic architecture indicates that SNPs of low to high effect contribute to immune response to vaccination. GWAS identified two genetic markers associated with vaccine-induced immune response phenotypes. Collectively, these findings indicate that genetic regulation of the immune response to vaccination is antigen-specific and influenced by multiple genes of small effect.

An Oil-Based Adjuvant Improves Immune Responses Induced by Canine Adenovirus-Vectored Vaccine in Mice.

There is a significant need for highly effective vaccines against emerging and common veterinary infectious diseases. Canine adenovirus type 2 (CAV2) vectors allow rapid development of multiple vaccines and have demonstrated their potential in animal models. In this study, we compared the immunogenicity of a non-replicating CAV2 vector encoding the rabies virus glycoprotein with and without MontanideTM ISA 201 VG, an oil-based adjuvant. All vaccinated mice rapidly achieved rabies seroconversion, which was associated with complete vaccine protection. The adjuvant increased rabies antibody titers without any significant effect on the anti-CAV2 serological responses. An RT2 Profiler™ PCR array was conducted to identify host antiviral genes modulated in the blood samples 24 h after vaccination. Functional analysis of differentially expressed genes revealed the up-regulation of the RIG-I, TLRs, NLRs, and IFNs signaling pathways. These results demonstrate that a water-in-oil-in-water adjuvant can shape the immune responses to an antigen encoded by an adenovirus, thereby enhancing the protection conferred by live recombinant vaccines. The characterization of early vaccine responses provides a better understanding of the mechanisms underlying the efficacy of CAV2-vectored vaccines.

Direct evidence of fiber-protein-directed hemagglutination by canine adenoviruses.

Canine adenoviruses (CAdVs) are divided into two serotypes, CAdV1 and CAdV2, whose members mainly cause infectious hepatitis and laryngotracheitis, respectively, in canids. To gain insight into the molecular basis of viral hemagglutination, we constructed chimeric viruses whose fiber proteins or their knob domains, which play a role in viral attachment to cells, were swapped among CAdV1, CAdV2, and bat adenovirus via reverse genetics. The results revealed that, in each case, viral hemagglutination was specifically mediated by the fiber protein or knob domain, providing direct evidence for fiber-protein-directed receptor-binding characteristics of CAdVs.

Investigation of canine chaphamaparvovirus, canine bufavirus, and canine adenovirus in dogs with diarrhea: First report of novel canine bufavirus in Turkey.

Viral enteritis is a significant cause of death among dogs younger than 6 months. In this study, the presence of canine chaphamaparvovirus (CaChPV), canine bufavirus (CBuV), and canine adenovirus (CAdV) was investigated in 62 diarrheal dogs previously tested for other viral pathogens (canine parvovirus type 2, canine coronavirus, and canine circovirus). CBuV was detected in two dogs (3.22%) and CaChPV in one dog (1.61%). One dog tested positive for three parvoviruses (CPV-2b, CBuV, and CaChPV). All dogs tested negative to CAdV-1/CAdV-2. A long genome fragment of one of the two identified CBuVs and of the CaChPV was obtained and analyzed. New Turkish CBuVs had high identity rates (96%-98% nt; 97%-98% aa) with some Italian CBuV strains (CaBuV/9AS/2005/ITA and CaBuV/35/2016/ITA). The phylogenetic analysis powerfully demonstrated that these viruses belonged to a novel genotype (genotype 2). A part of the genome ChPV-TR-2021-19 revealed high identity rates (> 98% nt and > 99% aa) with some Canadian CaChPV strains (NWT-W88 and NWT-W171) and the Italian CaChPV strain Te/37OVUD/2019/IT. This study is the first report on the detection of CBuV-2 and the concomitant presence of three canine parvoviruses in Turkey. The obtained data will contribute to the molecular epidemiology and the role in the etiology of enteric disease of new parvoviruses.

Canine Adenovirus 1 Isolation Bioinformatics Analysis of the Fiber.

Canine adenovirus type 1 (CAdV-1) is a double-stranded DNA virus, which is the causative agent of fox encephalitis. The Fiber protein is one of the structural proteins in CAdV-1, which mediates virion binding to the coxsackievirus and adenovirus receptor on host cells. The suspected virus was cultured in the MDCK cells, and it was determined through the cytopathic effects, sequencing and electron microscopy. The informatics analysis of the Fiber was done using online bioinformatics servers. The CAdV-1-JL2021 strain was isolated successfully, and were most similar to the CAdV-1 strain circulating in Italy. The occurrence of negative selection and recombination were found in the CAdV-1-JL2021 and CAdV-2-AC_000020.1. Host cell membrane was its subcellular localization. The CAdV-1-JL2021 Fiber (ON164651) had 6 glycosylation sites and 107 phosphorylation sites, exerted adhesion receptor-mediated virion attachment to host cell, which was the same as CAdV-2-AC_000020.1 Fiber. The Fiber tertiary structure of the CAdV-1-JL2021 and CAdV-2-AC_000020.1 was different, but they had the same coxsackievirus and adenovirus receptor. "VATTSPTLTFAYPLIKNNNH" were predicted to be the potential CAdV-1 B cell linear epitope. The MHC-I binding peptide "KLGVKPTTY" were both presented in the CAdV-1-JL2021 and CAdV-2-AC_000020.1 Fiber and it is useful to design the canine adenovirus vaccine.

Difference Analysis Between Canine Adenovirus Types 1 And 2.

Canine adenoviruses (CAdVs) include type 1 (CAdV-1, virulent strain) and type 2 (CAdV-2, attenuated strain). In recent years, the incidences of CAdV infections are increasing. However, they are difficult to distinguish when the symptoms are untypical. It is pivotal to find the differences between the two virus types for scientific, epidemiological, and specific treatment. CAdV-1 (virulent strain) and CAdV-2 (attenuated strain) induced canine hepatitis (ICH) and tracheobronchitis (ITB), respectively, but the clinical symptom is not obvious. CAdV-1 and CAdV-2 have the same genome structure, diameter, morphological features, and cytopathic features, but the same character hinder the diagnose time of the serotypes. CAdV-1 and CAdV-2 have a difference in the genome sequence, coding proteins, viral activity, hemagglutination patterns. After infection, pathogenicity and transmission route are different between the two serotypes. Sequence alignment, PCR, Real time-PCR assay are useful methods to distinguish the two serotypes. The attenuated live CAdV-2 vaccine is currently used to protect against CAdV-1, but it also has a risk. The further research should focus on the pathogenicity mechanism and the useful vaccine for the two serotypes of canine adenovirus.

Co-circulation of five species of dog parvoviruses and canine adenovirus type 1 among gray wolves (Canis lupus) in northern Canada.

Several viruses can infect wild carnivores but their impact on wildlife health is poorly understood. We investigated the presence, diversity and distribution of various DNA viruses in 303 wolves inhabiting a vast area of the Northwest Territories, Canada, over a period of 13 years. We found evidence for the presence of canine bufavirus (CBuV, 42.6%), canine parvovirus 2 (CPV-2, 34.0%), canine bocavirus 2 (CBoV-2, 5.0%), cachavirus (CachaV-1, 2.6%), canine adenovirus 1 (CAdV-1, 1%) and minute virus of canines (MVC, 0.3%). To our knowledge, this is the first detection of CBoV-2, MVC and CachV-1 in wild animals. We also demonstrate that CBuV and CachaV-1 were already circulating among wild animals at least 11 and 10 years, respectively, before their discoveries. Although CBuV prevalence was higher, CPV-2 was the most prevalent virus among juveniles, while CBuV infection was associated with poor nutrition conditions. Even if its prevalence was low, CachaV-1 had the highest multiple infection rate (87.5%). CadV-1 and MVC sequences were highly identical to reference strains, but we observed a high diversity among the other viruses and detected three new variants. One CPV-2 variant and one CBuV variant were endemic since the beginning of the 2000s in the entire investigated region, whereas one CBuV variant and two CBoV-2 variants were found in a more restricted area over multiple years and CachaV-1 was found only in one region. Two CPV-2 variants and one CachaV-1 variant were observed only once, indicating sporadic introductions or limited circulation. Different patterns of endemicity might indicate that viruses were introduced in the wolf population at different timepoints and that mixing between wolf packs may not be constant. Different epidemiological behaviors depend on viral factors like infectivity, transmission routes, pathogenicity and tissue-tropism, and on host factors like proximity to densely populated areas, carnivory and pack density and mixing.

Wolf (Canis lupus) as canine adenovirus type 1 (CAdV-1) sentinel for the endangered cantabrian brown bear (Ursus arctos arctos).

Canine adenovirus type 1 (CAdV-1) causes infectious canine hepatitis (ICH) and has recently been described as a cause of death among endangered populations of European brown bear (Ursus arctos arctos) in the Cantabrian mountain range in Asturias, Spain. Sympatric wild and domestic carnivores can act as reservoirs of the virus and likely spread it into the environment and subsequently transmit it to brown bears. The present work investigates the prevalence and geo-temporal distribution of CAdV-1 among free-ranging wolves (Canis lupus) in Asturias from 2009 to 2018, during which three fatal cases of ICH were reported among brown bears in the region. A total of 149 wolves were analysed in this study, of which 21 (14%) were found to have CAdV-1 DNA based on real-time polymerase chain reaction (RT-PCR) of spleen samples. Prevalence of the virus was similar between males and females. All but one of the 20 CAdV-1-positive animals of estimable age were younger than 2 years, and only one of the 46 adult animals (>2 years) tested positive. Prevalence was highest in the western area of Asturias and during 2010 and 2011. Our results confirm that CAdV-1 is circulating in Asturian free-ranging wolves, supporting their possible role as virus reservoirs and sentinels in the region of this emerging disease in brown bears.

In vitro functional genetic modification of canine adenovirus type 2 genome by CRISPR/Cas9.

Genetically modified oncolytic adenoviruses have been proposed as a vehicle for cancer therapy. However, several concerns, such as toxicity to normal cells and organs, lack of suitable cell surface receptors to allow viral entry to the desired cell type(s), and activation of both innate and adaptive immune systems in patients, restrict the successful clinical application of adenoviral-mediated cancer gene therapy. Successful virotherapy will require efficient transductional and transcriptional targeting to enhance therapeutic efficacy by ensuring targeted adenoviral infection, replication, and/or therapeutic transgene expression. Targeted modification of viral components, such as viral capsid, fiber knob, and the insertion of transgenes for expression, are prerequisites for the necessary transductional and transcriptional targeting of adenovirus. However, the conventional approach to modify the adenoviral genome is complex, time consuming, and expensive. It is dependent on the presence of unique restriction enzyme sites that may or may not be present in the target location. Clustered regularly interspaced short palindromic repeat (CRISPR) along with the RNA-guided nuclease Cas9 (CRISPR/Cas9) is one of the most powerful tools that has been adopted for precise genome editing in a variety of cells and organisms. However, the ability of the CRISPR/Cas9 system to precisely and efficiently make genetic modification, as well as introduce gene replacements, in adenoviral genomes, remains essentially unknown. Herein the ability of in vitro CRISPR/CAS9-mediated editing of the canine adenovirus type 2 (CAV2) genome to promote targeted modification of the viral genome was assessed. To demonstrate the feasibility of this goal, CRISPR/Cas9 has been used to successfully insert the RFP (red fluorescent protein) reporter construct into the CAV2 genome. Initial results demonstrated high efficiency and accuracy for in vitro CRISPR-mediated editing of the large CAV2 genome. Furthermore, this application was expanded, using multiple guide RNAs, to conduct gene replacement in the CAV2 genome by substituting a portion of the E3 gene with a construct designed to express a single chain antibody to canine PD-1. Thus, this work provides a significantly improved and efficient method for targeted editing of adenoviruses to generate altered and potentially therapeutic viral genomes in the shortest possible time.

Isolation and genetic characterization of canine adenovirus type 2 from a domestic dog showing neurological symptoms.

Canine adenoviruses (CAVs) are of two types: canine adenovirus type 1 (CAV-1), which causes infectious canine hepatitis, and canine adenovirus type 2 (CAV-2), which is mainly associated with the respiratory type of disease in dogs. Due to the widespread use of modified live vaccines to control canine adenoviral infections and subsequently reduced disease incidence, CAVs are often neglected by clinicians. Although a number of studies are available about CAV-1 prevalence in India, only meagre information is available about CAV-2. This study reports the CAV-2 infection in a vaccinated dog with neurological and respiratory symptoms which was found negative for other canine pathogens like canine distemper virus and canine parvovirus. The virus was successfully isolated from rectal swab in MDCK cells and characterized by immunofluorescence assay and virus neutralization test. On phylogenetic analysis of partial E3 region, the Indian CAV-2 grouped in a separate clade different from established subgroups. An insertion of "G" nucleotide was reported at nucleotide (nt.) position 1077 in the E3 gene of Indian CAV-2 isolates which led to a frameshift in the coding region of E3 gene thereby imparting additional eleven amino acids to its C-terminal end in comparison to isolates from other parts of the world. This may have an implication on the functional role of E3 protein inside the cell. This study reinforces the unique signature insertion in the E3 gene of Indian CAV-2 and is the second study in the world to report the association of CAV-2 with neurological disease in dogs.

The biological characteristics of the canine adenovirus type 1 from fox and the transcriptome analysis of the infected MDCK cell.

Canine adenovirus type 1 (CAdV-1) is the etiologic agent of fox encephalitis, and a virus strain from fox encephalitis is isolated and related research are conducted. In this experiment, the results showed that the F1301 strain was confirmed to be the CAdV-1. The whole genome of the CAdV-1 F1301 strain isolated from fox was 30,535 bp and had higher homology to the other reported CAdV-1 strains. After 0, 12, and 36 h of CAdV-1 infection, the difference gene of the 592 long noncoding RNA and 11,215 microRNA were involved in cell responses to CAdV-1 infection through the PI3K-AKT, Wnt, Herpes simplex, hepatitis C, and Epstein-Barr virus infection pathway in Madin-Darby canine kidney cell line (MDCK). The results indicate that the biological characterization of the CAdV-1 and the MDCK cell-CAdV-1 interaction are clarified.

Establishment of a Simple and Efficient Reverse Genetics System for Canine Adenoviruses Using Bacterial Artificial Chromosomes.

Canine adenoviruses (CAdVs) are divided into pathotypes CAdV1 and CAdV2, which cause infectious hepatitis and laryngotracheitis in canid animals, respectively. They can be the backbones of viral vectors that could be applied in recombinant vaccines or for gene transfer in dogs and in serologically naïve humans. Although conventional plasmid-based reverse genetics systems can be used to construct CAdV vectors, their large genome size creates technical difficulties in gene cloning and manipulation. In this study, we established an improved reverse genetics system for CAdVs using bacterial artificial chromosomes (BACs), in which genetic modifications can be efficiently and simply made through BAC recombineering. To validate the utility of this system, we used it to generate CAdV2 with the early region 1 gene deleted. This mutant was robustly generated and attenuated in cell culture. The results suggest that our established BAC-based reverse genetics system for CAdVs would be a useful and powerful tool for basic and advanced practical studies with these viruses.

Multicentric Molecular and Pathologic Study On Canine Adenovirus Type 1 in Red Foxes (Vulpes vulpes) in Three European Countries.

Canine adenovirus type 1 (CAdV-1) is the agent of infectious canine hepatitis, a severe frequently fatal disease affecting primarily dogs (Canis lupus familiaris). The virus has been detected in many wild carnivore species. Our aim was to evaluate the prevalence and genetic and histopathologic features of CAdV-1 in wild red foxes (Vulpes vulpes). Kidney and liver samples were obtained from 86 subjects, coming from the UK (n=21), Italy (n=36), and Germany (n=29). We used PCR, targeting the viral E3 gene and flanked regions, to detect the presence of the virus; viral E3, fiber, and E4 genes were sequenced and their sequences were compared with published sequences. Kidneys and liver from foxes in Italy and Great Britain (n=57) were prepared for histologic and immunohistochemical examination for CAdV-1. Viral DNA was detected in 22% (19 of 86) kidney samples, with E3 and E4 genes showing reported and unreported single nucleotide changes. No pathologic changes or viral immunopositive signals were detected in the examined tissues. Our study suggests that red foxes could be considered potential shedders of CAdV-1, as they showed a relatively high prevalence without related pathologic changes in the organs examined.

Disseminated melanized fungal infection due to Cladosporium halotolerans in a dog coinfected with canine adenovirus-1 and canine parvovirus-2.

This report presents the pathologic findings associated with disseminated infection due to Cladosporium halotolerans in a dog that was simultaneously infected with canine adenovirus-1 (CAdV-1) and canine parvovirus-2 (CPV-2). A 12-year-old, mixed breed dog, with a clinical history of neurological manifestations was submitted for routine autopsy due to poor prognosis. The principal pathologic findings were mycotic necrotizing nephritis, hepatitis, and splenitis with embolic dissemination to the brain resulting in mycotic necrotizing meningoencephalitis, ventriculitis, choroid plexitis, and obstructive hydrocephalus associated with intralesional and intravascular septate pigmented fungi. PCR and sequencing of the ITS region of fungi revealed that the intralesional fungal organisms had 82% nucleotide identity with members of the Cladosporium sphaerospermum complex of organisms. However, a PCR assay and sequencing of the beta tubulin gene confirmed that the organism identified in this dog had 100% nucleotide sequence identity with C. halotolerans. Using immunohistochemistry, intralesional antigens of CAdV-1 were identified within the epithelial cells of the liver and lungs; there was positive immunolabeling for CPV-2 antigens in degenerated cardiomyocytes. These findings confirmed the active participation of C. halotolerans in the development of disseminated cladosporiosis in this dog and represent a rare occurrence of concomitant infection with CAdV-1 and CPV-2.

Unique genetic features of canine adenovirus type 1 (CAdV-1) infecting red foxes (Vulpes vulpes) in northern Norway and arctic foxes (Vulpes lagopus) in Svalbard.

Canine adenovirus type 1 (CAdV-1) is the aetiological agent of infectious canine hepatitis (ICH) in domestic dogs (Canis familiaris). In spite of the widespread use of vaccination, CAdV-1 continues to circulate in the dog population. Although a high number of serological screenings have indicated that CAdV-1 is widespread in fox species, little is known about the potential role of foxes as reservoirs of CAdV-1. Furthermore, very little data exist on the molecular features of this virus in foxes. To add to existing knowledge on CAdV-1 circulating in wild carnivores, tissue samples from CAdV-seropositive red foxes (Vulpes vulpes, n = 10) from the northern mainland of Norway and arctic foxes (Vulpes lagopus, n = 10) from the Svalbard archipelago, Norway, were investigated using a molecular approach to detect CAdV-1 DNA and important structural and non-structural genes of the detected viruses were sequenced and analysed. Amplicons characteristic for CAdV-1 were amplified from 14 out of 20 foxes (7 red foxes and 7 arctic foxes) and spleen and lymph node tissues resulted optimal targets for the viral DNA detection. The nucleotide sequences showed unique features that distinguished the viruses detected in this study from the CAdV-1 to date identified in wild carnivores and dogs. Greater attention should be given to genetically different CAdV-1 circulating in wild carnivores that may be transferred to dogs, potentially causing disease and reducing the effectiveness of available vaccines.

Genetic Characterization of Canine Adenovirus Type 1 Detected by Real-Time Polymerase Chain Reaction in an Oral Sample of an Italian Wolf (Canis Lupus).

We report the detection of canine adenovirus type 1 DNA by real-time PCR technique in an oral sample of an Italian wolf (Canis lupus italicus). Genetic characterization of the virus revealed a strict relationship with viruses detected in dogs (Canis lupus familiaris), wolves, and red foxes (Vulpes vulpes), suggesting that transmission between wild animals and dogs had occurred.

Canine adenovirus type 1 (CAdV-1) in free-ranging European brown bear (Ursus arctos arctos): A threat for Cantabrian population?

Canine adenovirus type 1 (CAdV-1) is responsible for infectious canine hepatitis. The disease has been described in captive American black bear (Ursus americanus) and European brown bear (Ursus arctos arctos), with just one recently reported case in a cub of a free-ranging brown bear (Ursus arctos horribilis) from Alaska. The aim of this work is to summarize findings related to presence and associated mortality of CAdV-1 in 21 free-ranging Cantabrian brown bears (Ursus arctos arctos) submitted to necropsy in Asturias and Castilla y León (northwestern Spain) from 1998 to 2018. On the basis of the anatomopathological findings and laboratory results three free-ranging brown bears died due to infectious canine hepatitis, which is to our knowledge the first description of death due to this disease in free-ranging bears in Europe. Gross lesions consisted of petechial haemorrhages and congestion in different internal organs, haemorrhagic fluid in internal cavities, friable and yellowish liver and thickening of gall bladder. Microscopic lesions were observed mainly in liver, kidney and brain and consisted of multifocal necrosis of cells with presence of basophilic intranuclear inclusions. Immunohistochemical (IHC) and real-time polymerase chain reaction (qPCR) techniques were used to assess the presence of CAdV-1 in paraffin-embedded liver samples. Viral antigens were detected by IHC labelling within hepatocytes and Küppfer cells in the three animals. The presence of viral DNA was confirmed by qPCR in one of them. In order to evaluate the circulation of CAdV-1 in brown bears, a retrospective study was performed using both IHC and qPCR techniques in 11 and 12 additional brown bears, respectively. An extra brown bear was found positive by IHC. This study shows that CAdV-1 surveillance of brown bears and sympatric carnivores should be considered as major concern for the monitoring the population evolution throughout time in this endangered species.

Sequential circulation of canine adenoviruses 1 and 2 in captive wild carnivores, France.

Scarce data are currently available about the ecology of canine adenoviruses (CAdVs) in wild carnivores. In this paper, the consecutive circulation of CAdV-1 and CAdV-2 in wild carnivores maintained in a French zoological park is reported. A fatal CAdV-1 infection was observed in a Eurasian wolf (Canis lupus lupus), which displayed gross lesions, histopathological changes and immunohistochemical findings suggestive of CAdV-1 infection. The virus was isolated on cell cultures and its genome was determined through next-generation sequencing, resulting genetically related to a recent Italian CAdV-1 strain detected in an Italian wolf. Subsequently, subclinical circulation of CAdV-2 was demonstrated by molecular methods in wild carnivores maintained in the same zoological park, some of which had been previously vaccinated with a CAdV-2 vaccine. Virus detection at a long distance from vaccination and by unvaccinated animals was suggestive of infection by a CAdV-2 field strain, although no data are available about the extent and duration of shedding of CAdV-2 modified-live virus in wild or domestic carnivores. The present paper provides new insights into the CAdV ecology in wildlife, although future studies are needed to fully understand the pathogenic potential of both CAdVs especially in endangered carnivore species.

Exogenous LRRK2G2019S induces parkinsonian-like pathology in a nonhuman primate.

Parkinson's disease (PD) is the second most prevalent neurodegenerative disease among the elderly. To understand its pathogenesis and to test therapies, animal models that faithfully reproduce key pathological PD hallmarks are needed. As a prelude to developing a model of PD, we tested the tropism, efficacy, biodistribution, and transcriptional effect of canine adenovirus type 2 (CAV-2) vectors in the brain of Microcebus murinus, a nonhuman primate that naturally develops neurodegenerative lesions. We show that introducing helper-dependent (HD) CAV-2 vectors results in long-term, neuron-specific expression at the injection site and in afferent nuclei. Although HD CAV-2 vector injection induced a modest transcriptional response, no significant adaptive immune response was generated. We then generated and tested HD CAV-2 vectors expressing leucine-rich repeat kinase 2 (LRRK2) and LRRK2 carrying a G2019S mutation (LRRK2G2019S), which is linked to sporadic and familial autosomal dominant forms of PD. We show that HD-LRRK2G2019S expression induced parkinsonian-like motor symptoms and histological features in less than 4 months.

A Viral Receptor Complementation Strategy to Overcome CAV-2 Tropism for Efficient Retrograde Targeting of Neurons.

Retrogradely transported neurotropic viruses enable genetic access to neurons based on their long-range projections and have become indispensable tools for linking neural connectivity with function. A major limitation of viral techniques is that they rely on cell-type-specific molecules for uptake and transport. Consequently, viruses fail to infect variable subsets of neurons depending on the complement of surface receptors expressed (viral tropism). We report a receptor complementation strategy to overcome this by potentiating neurons for the infection of the virus of interest-in this case, canine adenovirus type-2 (CAV-2). We designed AAV vectors for expressing the coxsackievirus and adenovirus receptor (CAR) throughout candidate projection neurons. CAR expression greatly increased retrograde-labeling rates, which we demonstrate for several long-range projections, including some resistant to other retrograde-labeling techniques. Our results demonstrate a receptor complementation strategy to abrogate endogenous viral tropism and thereby facilitate efficient retrograde targeting for functional analysis of neural circuits.