CANINE DISTEMPER IN CARNIVORES

KAI FROLICH

Tierpark Arche Warder e.V., Warder, Germany

Canine distemper (CD), also called La maladie de Carre, is an acute or subacute highly contagious disease that may manifest with signs of generalized infection, respiratory disease, hyperkeratosis, central nervous system (CNS) disease or a combination of these, and can be accompanied by severe immunosuppression.

Aetiology

Canine distemper virus (CDV) is an enveloped, single­stranded, negative RNA virus of the family Paramyxoviri- dae, genus Morbillivirus. The virus is relatively fragile and quickly inactivated in the environment by ultraviolet light and by heat and drying. Temperatures greater then 50°C rapidly destroy the virus, and it is inactivated in several hours in tissues at 20—37° C. It may remain stable for weeks at 4 ° C and for years when frozen at —65°C. Common disinfectants readily inactivate CDV. The haemagglutinin (H) protein is a key component of the virus, responsible for the first virus—host cell interaction and initial virus entry. An adequate host immune response against the H protein may prevent CDV infection. The fusion (F) protein is an integral membrane protein-binding cell surface receptor of target cells. CDV is thought to have a single serotype, but is classified into up to nine lineages, e.g. America-1 and -2, Asia-1 and -2, European and Arctic. However, the distribution of the major lineages through­out the world is currently not clear. The sequence of path­ogenic events depends on the strain. Certain isolates, such as Snyder Hill, A75/17 and R252 strain are highly virulent and neurotropic^(6,7). A small number of specific residues are important in the adaptive evolution of CDV through positive selection, owing to their interactions with cellular receptors or with the host immune system.

EPIDEMIOLOGY

CDV has a broad host range. Members of nine families of the order Carnivora are susceptible to CDV infection, i.e. Ailuridae, Ailuropodidae, Canidae, Hyaenidae, Musteli- dae, Procyonidae, Viveridae, Felidae and Phocidae. Addi­tionally, in the order of even-toed ungulates CDV affects members of the family Tayassuidae, and in primates members of the family Cercopithecidae.

In European wildlife, infection and identification of CDV has been reported in several free-ranging carnivore species (Table 7.1).

CDV affects animals of all ages. Morbidity and mortal­ity rates vary among species and age groups (usually higher mortality in young animals).

Observations on the epidemiology of CD in dogs may assist in understanding the disease in free-ranging species. In endemic areas where dog populations are high, clinical

^a In the UK, all blood samples from an intensively studied population of wild badgers were negative for CDV antibodies^{t 34)}

disease is mostly seen in pups following loss of maternal antibody at 3—6 months.

I n Germany, a higher prevalence of CDV antibodies was detected in wild canids in urban and suburban areas and no positive samples were detected in animals from rural areas. Phylogenetic analyses revealed a distinct relat­edness to domesticated canine CDV isolates. Thus, the data support the concept of transmission of CDV between domestic dogs and wild carnivores¹-²⁵).

In Bavaria, a severe outbreak of canine distemper encephalitis was observed in several wildlife species (red fox ( Vulpes vulpes), Eurasian badger (Meles meles) and stone marten (Martes foina). The H genes of several representa­tive virus samples during this outbreak had a Y549H amino acid substitution in the H protein. The data support the hypothesis that pathogen evolution at residue 549 in the SLAM- binding region of the H protein of CDV is associated, directly or indirectly, with disease emergence in novel host species(¹⁵).

In France, it was postulated that the high prevalence of CDV antibody in five mustelid species was possibly related to the fact that mustelids live in close proximity to humans, and are therefore more likely to come into direct or indi­rect contact with CDV-infected domestic dogs(⁹).

Although CDV is thought to be maintained in domestic dog populations worldwide, the virus has a very broad, and apparently expanding, host range. Transmission to wolves could occur through predation or scavenging of a domestic carnivore (dog or cat) carcass⁽²⁴⁾.

In Spain, the genome of CDV was detected in a feral cat, showing evidence of interspecific transmission of CDV⁽²⁶⁾. It has been proposed that CDV- infected foxes could be a source of infection to other less abundant species that live sympatrically(²⁷).

Since 2006, the Alpine region of North-East Italy has been experiencing a severe and widespread outbreak of CDV, affecting the wild carnivore population. Captured red foxes and Eurasian badgers showed CD-like signs (e.g. prostration, altered behaviour and conjunctivitis). CDV infection was confirmed by quantitative reverse transcrip­tion polymerase chain reaction (RT-PCR) of pooled organs, and sequence analysis of the H gene indicated that the fox strains were highly related to each other and to the strains identified in foxes in Bavaria, as well as to a canine strain identified in Hungary¹-²⁸). In Austria, strains isolated from badger and stone marten were assigned to the Euro­pean wildlife groups(¹⁶). Also the emergence and spread of a novel genetic cluster of CDV in wildlife populations of the Alps has been described¹-¹⁷). This novel genetic group, denoted as WE/06—09 cluster, consists almost exclusively of viruses identified from wild carnivores (red fox, Euro­pean badger, stone marten).

Moreover, this outbreak has reached wildlife (red fox and European badger) in Switzerland and Liechtenstein. Interestingly, there was evidence for the presence of mor- billivirus infection in Swiss wildlife before this outbreak. Seroprevalence of about 25% was recorded in red fox and Eurasian lynx (Lynx lynx), indicating that exposure to a morbillivirus was apparently not uncommon in wild carnivores⁽¹⁹⁾.

Transmission of CDV is primarily by aerosol or contact with oral, respiratory and ocular fluids and exudates con­taining the virus. Close association between affected and susceptible animals is necessary owing to the relative fragil­ity of CDV in the environment, because the virus is quickly inactivated by ultraviolet light, heat and drying. Viral shedding occurs even if animals are subclinically infected, and virus may be shed for up to 90 days after infection.

Pathogenesis, pathology

AND IMMUNITY

As we have seen, the natural route of distemper infection is by inhalation of airborne virus or contact with oral, respiratory and ocular fluids and exudates containing the virus.

Virus replication occurs first in cells of lymphatic tissues in the upper respiratory tract. Within a few days, CDV multiplies in macrophages and spreads to tonsils and regional lymph nodes. After about 1 week, the virus may be found in systemic lymphoid tissues, lamina propria of the digestive system, and Kupffer cells in the liver. The systemic spread of virus corresponds to the development of fever and leucopenia; the latter is due to viral-associated loss of T and B lymphocytes. Destruction of cells of the immune system results in immunosuppression.

Between 1 and 2 weeks after infection, the host’s immune response determines the outcome of infection. If there is a strong antibody response, no clinical illness develops and the virus is cleared from the tissues. If the animal mounts a weak antibody response, illness may ensue, but approxi­mately 3 weeks after infection the virus is cleared from most of the body, with the possible exception of the lungs, skin and CNS. The virus may be shed by these animals for several months. If the animal recovers, it may shed virus for 2—3 months¹-³⁵). Pathological changes ofCD have been described in a variety of carnivores and are, with some exceptions, similar to those described in domestic dogs. They include interstitial pneumonia or broncho-pneumonia, as well as conjunctivitis, rhinitis and inflammation of the tracheo­bronchial tree. Catarrhal to haemorrhagic enteritis, and hyperkeratosis of the nose, lips, eyelids, ears, anus and foot­pads may be present. Older or more immunocompetent animals tend to develop leucoencephalomyelitis, with a predominance of lesions in the caudal brainstem and spinal cord. Absolute lymphopenia is common. One major differ­ence compared with domestic dogs is the jaundice associ­ated with CDV in raccoons (Procyon lotor), ranch-raised foxes and ferrets (Mustela putorius).

On histologic examination, CDV inclusions are most commonly cytoplasmic and acidophilic-staining.

Immunity to CDV infection involves both humoral and cell-mediated mechanisms. Viral proteins are expressed on the cell surface; thus immune-mediated cytolysis plays a role in clearing the virus from infected animals(³⁵). Neutral­izing antibodies in domestic dogs develop 10—14 days post-infection (pi) and titres peak after 4 weeks; they may persist for 2 years after exposure. The titre levels at which antibodies are protective in wild carnivore species is unknown; neutralizing antibody titres between 20 and 100 probably provide protection¹-³⁶). Wild animals that survive CD probably have lifelong immunity to subse­quent infection by CDV.

CLINICAL SIGNS AND TREATMENT

The incubation period ranges from about 1 week to 1 month or longer. The classic clinical signs are depression and mucopurulent oculo-nasal exudates. Fever, anorexia, vomiting and diarrhoea frequently occur. CNS signs may be concurrent or follow systemic disease. Neurological signs depend on the area of brain affected and include abnormal behaviour, convulsions, cerebellar and vestibular signs, paresis or paralysis and incoordination. Animals that recover from clinical CD or that have a prolonged illness may be in poor body condition. Regardless of the ultimate disease outcome, the infection is characterized by a dra­matic decrease in white blood cells and an inhibition of lymphocyte proliferation in the first weeks after infection.

In canids and mink, juveniles appear to be most suscep­tible. In mustelids, the clinical signs first recognized are serous oculo-nasal exudate, photophobia, and hyperemia and thickening of the eyelids, lips and anus. Mortality is variable in adult mink (20—90%) and is approximately 90% in mink kits.

CD in procyonids is clinically similar to that described in canids. Raccoons with CNS manifestations are fre­quently reported as behaving abnormally. Hyperkeratosis of the foot pads, with marked thickening and deep cracks, is relatively common in raccoons that have a prolonged clinical course. Mortality in experimentally infected rac­coons varied from 50 to 100%⁽³⁵⁾.

Infected animals with upper respiratory signs should be kept in environments that are clean, warm and free of draughts. Pneumonia is frequently complicated by second­ary bacterial infection, which usually requires a broad­spectrum antibiotic therapy. Parenteral therapy is essential when gastrointestinal signs are present. Food, water and oral medications or fluids should be discontinued if vomit­ing and diarrhoea are present. Therapy for neurologic dis­turbances is less rewarding. In general, animals should not be euthanized unless the neurologic disturbances are pro­gressive or incompatible with life.

In severe clinical cases passive immunization over several days with a hyperimmune serum (e.g. Stagloba®) in is recommended and should be repeated after 14 days.

DIAGNOSIS

An important diagnostic feature of CD is the presence of intracytoplasmatic and intranuclear eosinophilic inclusion bodies in epithelia, neurons and astroglia. Inclusion bodies are also often present in gastric mucosa, enterocytes and pancreatic and biliary duct epithelium, as well as epithe­lium of the respiratory and urogenital tract.

Immunohistochemistry is useful for detection of CDV antigen in formalin-fixed, paraffin-embedded tissues. The use of direct and fast viral nucleic acid detection methods such as RT-PCR is an extremely sensitive assay to detect CDV in various tissues. Viral RNA can be demonstrated in brain, lung, spleen, liver, kidney, urinary bladder and colon samples as well as in buffy coat cells. In addition, they can be applied to phylogenetic analyses, which are often based on partial sequences of the F-gene(³7).

A virus neutralization test is the standard serological test for antibodies against CDV.

MANAGEMENT, CONTROL AND REGULATIONS

On more than one occasion, wild carnivores have been translocated or brought into captivity while incubating CD. Thus, CD must be considered in management plans and during conservation efforts.

Vaccination against CDV in non- domestic carnivores has been problematic. Currently there is no safe and effica­cious commercially available CDV vaccine for use in non­domestic species. Live-virus-vaccine-induced CD has occurred in domestic ferrets (Mustelaputorius furo), black­footed ferrets (Mustela nigripes), European mink and several other wild carnivores(³⁸). Most important is the interference between attenuated live vaccines and maternal antibodies, resulting in insufficient protection of offspring from vac­cinated females(³⁹). Additionally, the attenuated live vac­cines are associated with a risk of reversion to virulence, and various wildlife species have suffered from fatal infections caused by live vaccines. Hence, safer alternatives such as inactivated virus vaccines, subunit vaccines or recombinant vaccines are recommended. DNA vaccines are being inves­tigated as an alternative vaccination strategy against canine distemper to overcome some of the limitations of attenu­ated live vaccines(⁴⁰). The efficacy and safety of three differ­ent CDV vaccines in the Eurasian otter (Lutra lutra), the Asian small-clawed otter (Aonyx cinereus) and the North American river otter (Lutra canadensis) have been evalu- ated(⁴¹). Most of the animals vaccinated with inactivated CDV vaccine showed no seroconversion. Experimental CDV immunostimulating complexes (CDV-ISCOM) induced a variable immune response that depended on the otter species. High serum antibody titres were observed in almost all animals vaccinated with commercially available modified life vaccines (MLV). Taken together, the results indicate a poor immune response to inactivated distemper vaccine, especially in Eurasian otters.

Recently, a chimaeric virus combining the replication complex of a measles vaccine strain with the envelope of a recent CDV wild-type isolate was produced¹-⁴²). The resulting virus did not cause disease or immunosuppres­sion in ferrets and conferred protection from challenge with a lethal wild-type strain, demonstrating its potential value for wildlife conservation efforts.

In conclusion, adequately controlling CDV infection in wildlife in Europe is difficult. It requires concerted tran­snational actions, including effective surveillance and prompt gathering and dissemination of information¹-²⁸).

There are no established EU regulations for wildlife.

PUBLIC HEALTH CONCERN

A suggestion has been made that Paget’s disease, an inflam­matory bone disorder in people, might be related to CDV acquired from exposure to dogs(⁶). The presence of CDV has been shown in Pagetic bone samples and CDV can infect and replicate in human osteoclast precursors in a dose-dependent manner(⁴³). This is the first evidence that CDV should be considered as a potential zoonosis.

SIGNIFICANCE AND IMPLICATIONS

FOR ANIMAL HEALTH

CD is one of the most important infectious diseases in carnivores. It has shown significant impact on free-ranging populations of highly susceptible species such as black­footed ferrets (Mustela nigripes)⁴⁴ and gray foxes ( Urocyon cinereoargenteus)⁴⁵ in North America. In Europe, CD may be a potential risk for e.g. the highly endangered Iberian lynx (Lynx pardinus)⁶⁵)'²⁶⁵. The impact on populations of other susceptible species within Europe is less clear.