Canine adenovirus type 1 causing neurological signs in a 5-week-old puppy.

BACKGROUND: Infectious canine hepatitis is a rarely encountered disease, that is caused by Canine Adenovirus-1. Clinical signs can vary dramatically, and neurological signs are rarely seen. Neurological manifestation of this disease is rarely reported in the veterinary literature. CASE PRESENTATION: A 5-week-old, male entire Husky cross puppy presented for a one-day history of abnormal neurological behaviour (circling, ataxia, vocalization and obtund mentation). The puppy was euthanized shortly after presentation due to rapid deterioration. Histopathology raised concerns for Canine Adenovirus 1 (CAdV-1) based on vasculitis in the brain and intranuclear inclusion bodies in endothelial cell and hepatocytes; immunohistochemistry on brain tissue confirmed CAdV-1 infection. CONCLUSIONS: This report discusses possible routes of infection and manifestations of adenovirus infections causing neurologic signs. It also provides a timely reminder that CAdV-1 should be considered a differential in unvaccinated dogs that present with neurological signs. Further studies are required to better understand the neurotrophic tendencies of this virus.

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.

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.

A multiplex PCR method for the simultaneous detection of three viruses associated with canine viral enteric infections.

The aim of this study was to establish a multiplex PCR (mPCR) method that can simultaneously detect canine parvovirus (CPV-2), canine coronavirus (CCoV) and canine adenovirus (CAV), thereby eliminating the need to detect these pathogens individually. Based on conserved regions in the genomes of these three viruses, the VP2 gene of CPV-2, the endoribonuclease nsp15 gene of CCoV, and the 52K gene of CAV were selected for primer design. The specificity of the mPCR results showed no amplification of canine distemper virus (CDV), canine parainfluenza virus (CPIV), or pseudorabies virus (PRV), indicating that the method had good specificity. A sensitivity test showed that the detection limit of the mPCR method was 1 × 104 viral copies. A total of 63 rectal swabs from dogs with diarrheal symptoms were evaluated using mPCR and routine PCR. The ratio of positive samples to total samples for CPV-2, CCoV, and CAV was 55.6% (35/63) for mPCR and 55.6% (35/63) for routine PCR. Thirty-five positive samples were detected by both methods, for a coincidence ratio of 100%. This mPCR method can simultaneously detect CCoV (CCoV-II), CAV (CAV-1, CAV-2) and CPV-2 (CPV-2a, CPV-2b, CPV-2c), which are associated with viral enteritis, thereby providing an efficient, inexpensive, specific, and accurate new tool for clinical diagnosis and laboratory epidemiological investigations.

SURVEILLANCE FOR ANTIBODIES AGAINST SIX CANINE VIRUSES IN WILD RACCOONS (PROCYON LOTOR) IN JAPAN.

Raccoons (Procyon lotor) are found worldwide. They are frequently seen in crowded inner cities as well as in forests or wooded areas, often living in proximity to humans and their pets. We examined sera from 100 wild raccoons in Japan for antibodies to six canine viruses with veterinary significance to assess their potential as reservoirs. We also aimed to understand the distribution of potentially infected wildlife. We found that 7% of samples were seropositive for canine distemper virus (CDV), 10% for canine parvovirus type 2, 2% for canine adenovirus type 1, 6% for canine adenovirus type 2, and 7% for canine coronavirus. No samples were found to be seropositive for canine parainfluenza virus. Seropositivity rates for canine distemper virus and canine parvovirus type 2 were significantly different between areas, and younger raccoons (<1 yr old) were more frequently seropositive than older raccoons. Because raccoons belong to the suborder Caniformia, similar to dogs (Canis lupus familiaris), our results suggest that they can act as reservoirs for some of these important canine viruses and might be involved in viral transmission. Further study should include isolation and analysis of canine viruses in wild raccoons from a wider area.

A Novel Recombinant Canine Adenovirus Type 1 Detected from Acute Lethal Cases of Infectious Canine Hepatitis.

In this study, canine adenoviruses (CAdVs) from two acute fatal cases of infectious canine hepatitis (ICH) were analyzed using molecular detection and sequencing of the pVIII, E3, and fiber protein genes. Pathological findings in affected dogs were typical for CAdV-1 associated disease, characterized by severe centrilobular to panlobular necrohemorrhagic hepatitis and the development of disseminated intravascular coagulation in the terminal stages of disease. Comparison of partial genome sequences revealed that although these newly detected viruses mainly had CAdV-1 genome characteristics, their pVIII gene was more similar to that of CAdV-2. This likely suggests that a recombination has occurred between CAdV-1 and CAdV-2, which possibly explains the cause of vaccine failure or increased virulence of the virus in the observed ICH cases.

Development of a SYBR Green real-time PCR assay with melting curve analysis for simultaneous detection and differentiation of canine adenovirus type 1 and type 2.

Canine adenovirus type 1 (CAdV-1) and canine adenovirus type 2 (CAdV-2) cause infectious canine hepatitis (ICH) and infectious tracheobronchitis (ITB) in dogs, respectively. Cases of ICH have been documented in recent years and recent surveys have demonstrated a wide percentage of asymptomatic CAdV-1 infection in the canine population. Since both CAdV types are detectable in the same biological matrices, and viral coinfection with CAdV-1 and CAdV-2 are reported with high frequency, it is urgent to have available a rapid, highly sensitive and specific assay for the diagnosis of CAdV infection and distinction between CAdV-1 and CAdV-2. In order to detect canine adenovirus in biological samples and to rapidly distinguish the two viral types, a SYBR Green real-time PCR assay was optimized to discriminate CAdV-1 and CAdV-2 via a melting curve analysis. The developed assay showed high sensitivity and reproducibility and was highly efficient and specific in discriminating the two CAdV types. This reliable and rapid technique may represent a simple, useful and economic option for simultaneous CAdV types detection, which would be feasible and attractive for all diagnostic laboratories, both for clinical purposes and for epidemiological investigations.

Molecular analysis of a fragment of gene E1B 19K of canine adenovirus 2 (CAV-2) isolated from dogs with symptoms of cough.

The aim of this study was to perform molecular analysis of canine adenovirus 2 (CAV-2) E1B 19K gene fragment isolated from 20 dogs of various breeds (12 males and 8 females aged 1-9 years), with clinical symptoms of upper respiratory tract infections, from the Lubelszczyzna region. Nasal swabs were taken from dogs. DNA of CAV-2 was detected using the PCR method in 16 swabs. All PCR products were sequenced, and the obtained sequences were compared with each other and with the sequence of the E1B 19K gene of the CAV-2 strain from an online database of NCBI GenBank: AC 000003. Based on analysis of the obtained sequences, three polymorphic variants of CAV-2 (No. 1-3) with homology of 78 - 100% were distinguished. The nucleotide and amino acid sequences of the most frequently represented polymorphic variant, No. 1, differed from the sequences of polymorphic variant No. 2 with one substitution. The nucleotide and amino acid sequence of the E1B 19K gene of CAV-2 AC 000003 differed from the analogous sequences of representatives of variant No. 1 with 44 nucleotide and 19 amino acid substitutions. The small number of nucleotide differences in the E1B 19K CAV-2 gene among the examined own isolates, compared with AC 000003, suggest that the infections in dogs were caused by a relatively genetically stable virus which occurs in eastern

Hemorrhagic and necrotizing hepatitis associated with administration of a modified live canine adenovirus-2 vaccine in a maned wolf (Chrysocyon brachyurus).

A 15-yr-old, female, maned wolf (Chrysocyon brachyurus) was euthanized after presenting semicomatose with severe, uncontrolled frank hemorrhage from her rectum 6 days following a routine physical examination and vaccination. Histopathology indicated severe hemorrhagic and necrotizing hepatitis with intranuclear basophilic inclusion bodies in the liver that were thought to be consistent with adenoviral infection. Further classification by polymerase chain reaction, immunohistochemical staining, virus isolation, and electron microscopy confirmed the etiologic agent to be canine adenovirus-2. A representative sample of the vaccine that had been used was submitted and sequenced along with the virus isolated from the maned wolf. The sequencing of the etiologic agent that had been isolated from the maned wolf was determined to be the same as the strain of virus used in the production of the modified live vaccine that had been administered 6 days prior to death. From this information, the diagnosis of vaccine-induced adenoviral hepatitis was made. This is the first confirmed case of vaccine-induced canine adenoviral hepatitis in a maned wolf.

Characterization of a canine homolog of hepatitis C virus.

An estimated 3% of the world's population is chronically infected with hepatitis C virus (HCV). Although HCV was discovered more than 20 y ago, its origin remains obscure largely because no closely related animal virus homolog has been identified; furthermore, efforts to understand HCV pathogenesis have been hampered by the absence of animal models other than chimpanzees for human disease. Here we report the identification in domestic dogs of a nonprimate hepacivirus. Comparative phylogenetic analysis of the canine hepacivirus (CHV) confirmed it to be the most genetically similar animal virus homolog of HCV. Bayesian Markov chains Monte Carlo and associated time to most recent common ancestor analyses suggest a mean recent divergence time of CHV and HCV clades within the past 500-1,000 y, well after the domestication of canines. The discovery of CHV may provide new insights into the origin and evolution of HCV and a tractable model system with which to probe the pathogenesis, prevention, and treatment of diseases caused by hepacivirus infection.

Análisis de la población canina en el Distrito Capital, 2005 / Analysis of the canine population in the Capital District, 2005

Estudio realizado en Bogotá, Colombia, el primer trimestre de 2005. Se analizaron variables relacionadas con la dinámica de la población canina, igualmente se estimó la población felina, se evaluó el impacto de las estrategias para controlar la población y actividades de prevención de la Rabia Canina. Es un estudio epidemiológico transversal, de tipo probabilística para la ciudad, se compara con un estudio similar realizado en 1999. Los resultados mostraron que la relación perro: hombre se ha mantenido alrededor de 1:10, gracias al programa de recolección y esterilización; relación más estrecha en localidades donde predominan estratos bajos; la problemática de perros callejeros es grave siendo difícil generar una conciencia ciudadana con relación a la tenencia de mascotas. La tasa anual de renovación canina es cercana al 10%, lo cual obliga a realizar vacunación antirrábica de mantenimiento en forma sostenida para no romper las coberturas útiles de vacunación. En los felinos existe una relación de 1 gato por cada 50 personas. Las autoridades sanitarias deben ampliar las metas de esterilización, para bajar el crecimiento poblacional canino; y, buscar la modificación de hábitos con relación a la tenencia de mascotas, a través de actividades educativas y persuasivas.Las autoridades sanitarias deben ampliar las metas de esterilización, para bajar el crecimiento poblacional canino; y, buscar la modificación de hábitos con relación a la tenencia de mascotas, a través de actividades educativas y persuasivas.Para mejorar las acciones de prevención y control de zoonosis Se debe implementar un registro de mascotas urbanas, revisar la normatividad y generar un sistema de vigilancia epidemiológico.

Study realized in Bogotá, Colombia, in the first trimester of 2005, analyzing variables related to the dynamics of the canine population, equally estimated the feline population, evaluated the impact of strategies for controlling the population and activities for the prevention of Canine Rabies. It is a transversal epidemiology study, of the probabilistic type for the city, to be compared with a similar study done in 1999.The results show that the man/dog ratio has remained around 1:10, thanks to the program of recollection and sterilization; the ratio is more stretched in the localities, where the lower class predominates.The problem of stray dogs is grave, it being difficult to generate a civic consciousness with relation to the mascot tendency. The annual rate of canine renewal is around 10%, which obliges a program of anti-rabies vaccinations maintained in a sustained form so as not to destroy the useful covering of vaccinations. Among cats there exists a rate of one cat to every 50 people.The sanitary authorities have to increase the sterilization goals in order to lower canine population growth; and search to modify habits with relation to the mascot tendency, through educative and persuasive activities. To improve zoonosis prevention and control actions it will be necessary to implement a register of urban mascots, revise the normative and generate a system of epidemiological vigilence.

[Construction of recombinant canine adenovirus type 2 expressing Canine coronavirus spike glycoprotein and its immunogenicity].

In order to construct a recombinant Canine adenovirus type 2 (CAV-2) expressing the spike glycoprotein of Canine coronavirus (CCV), the S1 gene fragment of CCV strain DXMV, encoding major antigenic region A, B, C and D of S protein, was amplified by RT-PCR and cloned into pVAX1 vector. The complete S1 expression cassette was subcloned into the shuttle vector pVAXE3, then further cloned into the backbone vector pPoly2-CAV2 containing complete genome of CAV-2. To gain the recombinant Canine adenovirus, the recombinant plasmid pCAV-2-CCV-S1 was linearized by Cla I/Asc I to release recombinant genome, and then transfected into MDCK cell. The recombinant virus CAV-2-S1 was gained through 4 passages in MDCK, which showed classical CPE of CAV-2. The expressed S1 protein of CCV, which was identified by RT-PCR and Western blot, can be specifically recognized by polyclonal antibody against CCV. The immunization in dogs indicated that the recombinant CAV-2 could effectively induce the specific antibodies against CCV and CAV.

Detection and differentiation of CAV-1 and CAV-2 by polymerase chain reaction.

Canine adenovirus type 1 (CAV-1) and type 2 (CAV-2) can be categorized in the laboratory by haemagglutination and neutralization tests, but they are difficult to differentiate from each other in specimens, especially when infection occurs in the digestive tract. The object of this study was to develop a simple method of detecting and differentiating them. One pair of common primers was designed and synthesized according to the sequences of the E3 and flanking regions and a polymerase chain reaction (PCR) assay was established using these two primers to amplify the virus-specific DNA fragment from clinical specimens as well as from cell cultures. After elecctrophoresis, under the same amplification conditions, 508 bp and 1030 bp PCR products were observed for CAV-1 and CAV-2, respectively. These were further shown to be adenovirus specific by dot hybridization and sequencing. As only one pair of primers was involved in the PCR procedure, it was faster and easier to perform than any of the other assays used for detecting canine adenovirus, making it applicable in the rapid confirmation of diagnosis and differentiation of the two types of canine adenoviruses.

Sequence analysis of the canine adenovirus 2 fiber-encoding gene.

The gene coding for the fiber protein (Fip) of canine adenovirus type 2 (CAV-2) has been cloned and sequenced. The putative protein has 80% sequence similarity with the CAV-1 Fip. The observed differences may contribute to the known differences in cell tropism and virulence of the two types of CAV.

Adenoviruses of man and animals.

Adenoviruses are capable of causing natural infections in a wide variety of vertebrates, including amphibia, birds and mammals. Although frequently causing inapparent infections, they may be associated with diseases of varying severity and localizations. The clinico-pathological manifestations most commonly associated with adenovirus infections involve the respiratory tract and the ocular conjunctiva. Involvement of the digestive, renal and nervous systems may also be observed. A state of latency with persistence of infection for prolonged periods has been described in many hosts. Many adenovirus types are oncogemic under experimental conditions but none have been associated with naturally occurring tumours. Although the diseases caused by adenoviruses tend to be mild, this is not invariably the case and these viruses may give rise to serious epidemiological situations and represent a threat to life under certain circumstances in which the use of prophylactic vaccination may be indicated.