Canine distemper (CD) is a highly contagious multi-systemic disease caused by canine distemper virus (CDV). The clinical disease can be sub-clinical to fatal, spreading through the bloodstream to multiple organs, with clinical signs that include respiratory distress, anxiety, conjunctivitis, anorexia, diarrhea, lymphopaenia, encephalitis, coughing, rhinorrhoea, ocular discharge and fever (1, 2).

CDV has been detected in a wide range of domestic and wild animal hosts worldwide including canids, felids, mustelids, procyonids, urisds and hyaenids (3–5). It is highly prone to cross-species transmission between domestic and wildlife reservoir hosts, therefore representing a significant conservational and animal health risk around the globe (6, 7). CDV transmission primarily occurs via various body fluids, such as respiratory droplets, ocular and nasal discharge, saliva, urine and feces (8). In Italy, CDV has been reported in both domestic and free-ranging canids, as well as in mustelids. The virus poses a significant threat to wildlife conservation and domestic animal health, making its study crucial for disease management and control. CDV is an enveloped virus with a non-segmented, negative sense, single-stranded RNA genome and belongs to the family Paramyxoviridae, genus Morbillivirus (3). His genome encodes six structural proteins: nucleocapsid (N), matrix (M), fusion (F), hemagglutinin (H), polymerase (L), and phosphoprotein (P) (9). Based on H-gene sequences, CDV strains have been classified into 19 genetic lineages: America-1, America-2, North America-3, South America/North America-4, America-5 (formerly regarded as a sub-genotype of America-2), Asia-1, Asia- 2, Asia-3, Asia-4, Asia-5, Europe Wildlife, Arctic, Africa-1, Africa-2, Europe-1/South America-1, South America-2, South America-3, Rockborn-like, and Asia-6 (5, 10). Recently, Lanszki et al. (11) proposed a distinct CDV classification in 16 genetic lineages based on full-genome sequences. In Italy, the presence of CDV has been reported within the Canidae family, including domestic dogs (12) and free-ranging canids, like foxes and wolves (13, 14), and also within the family Mustelidae (15–17).

In recent decades, three CDV genetic lineages have been reported to circulate in Italy: the Europe/South America-1, EuropeWildlife, and Arctic-like lineages (10). The first to be detected in both domestic dogs and wild carnivores was the Europe/South America-1 lineage, while the Europe Wildlife have been reported in wildlife and only sporadically in domestic dogs (13, 18). Conversely, Arctic-like lineage has been reported mainly in Italian dogs and, less frequently, in wildlife (14, 16, 19). These data indicate the circulation of the three different CDV strains across the domestic/wildlife interface in Italy.

This study aimed at investigating the presence of CDV and other selected viruses (rabies virus, canine coronavirus, canine adenoviruses, canine herpesvirus type 1, rotavirus, protoparvovirus carnivoran 1) from internal organs of wild carnivorans of the family Mustelidae and Canidae and rodents of the family Hystricidae, collected in Southern Italy (Campania region) in 2022–2024CDV was consistently detected in this study and genetic characterization of the identified CDV strains was performed.

Overall, out of 136 wild animals collected in Campania region in a period spanning from 2022 to 2024, 14 (10.3%) tested positive for CDV RNA by RT-qPCR (Table 1) with a cycle threshold (Ct) ranging from 17.8 to 42.5. CDV infections were observed in badgers (6/34, 17.6%), foxes (5/66, 7.6%), wolves (2/11, 18.2%) and martens (1/3, 33.3) while CDV was not retrieved from stone martens, porcupines and otters (Table 2). Six out 44 (13.6%) CDV positive cases (3 foxes, 2 badgers and 1 marten) were identified in 2022, 6/74 (8.1%) (3 badgers, 2 foxes and 1 wolf) in 2023, 2/18 (11.1%) (1 wolf and 1 badger) in 2024. CDV was unevenly identified in the lung, intestine, brain, spleen, liver, heart, muscle and kidney of the positive animals. The 14 CDV positive animals did not present co-infections with other analyzed pathogens. The 6 CDV-positive badgers were 3 males and 3 females and comprised 3 adults, 2 subadults and 1 young. The 5 CDV-positive foxes were 3 males and 2 females and included 3 adults, 1 subadult and 1 young. The 2 wolves were 1 male and 1 female and encompassed 1 adult and 1 subadult. The marten was an adult male. Out of 14 CDV-positive animals, the most frequent clinical observations at necroscopy were congested lungs and possibly also trachea in 4 (28.5%) animals followed by enteritis in 3 (21.4%) animals. The cause of death of the 14 CDV-positive animals were either trauma or infection (6/14, 42.8% for both), or gunshot and bite wounds (1/14, 7.1% for both). All the samples were tested for rabies virus and found to be negative.

CDV strain ITA/2022/badger/33340, retrieved from spleen of an adult male badger in 2022 (Ct = was subjected to full genome amplification and ONT sequencing protocol).

On full genome level, the CDV strain retrieved in this study displayed the highest nt identity (99.4%) to the isolate CDV11956/2015 (KX024708). The genomic sequence of the CDV strain ITA/2022/badger/33340 was aligned with cognate CDV sequences available in GenBank. Upon ML phylogenetic tree the CDV strain ITA/2022/badger/33340 segregated with other CDV strains included in the Arctic-like cluster (Figure 1).

The results of this study demonstrated the circulation of CDV in the wildlife in Southern Italy with infection rate equal to nearly 10%. No co-infections with other pathogens (i.e., CCoV, CAdVs type 1 and 2, CaHV-1, RVA and protoparvovirus carnivoran 1) were detected in the CDV-positive animals. In a previous report CDV infection rate reached 16% in tissue samples of foxes and badgers collected in Northern Italy (33). In this study, also, no other pathogens were detected in the animals that tested positive for distemper. In some cases, we can hypothesize, that the absence of other pathogens could also be related to the poor conditions of the carcasses.

About the rabies virus our territory has been declared free since 2013, checks are currently carried out only for constant monitoring, due to the presence of rabies in Eastern Europe. The complete genome sequence of the CDV strain, that we obtained from an adult male badger, phylogenetically characterized as an Arctic-like lineage, clustered along with other CDV strains retrieved from badger and dog from Italy.

The arctic lineages and the EuropeanWildlife have been mostly detected in Central and Southern Italy (14–16) while the Europe/South America-1 lineage is peculiarly spread in Northern Italy (13, 17, 34).

The Arctic CDV strains were first reported in animals of the Arctic ecosystem but this lineage appears common in European territories in both domestic and wildlife carnivores. In Italy, CDV strains belonging to the Arctic lieage were first reported in dogs in the early 21th century (35) and subsequently in the United States (36) and other European countries (2, 12, 37), raising questions on the origin of these unusual strains.

A large distemper epidemic, sustained by an Arctic-lineage (38), occurred in Italy during 2013 and involved primarily the Abruzzi region and neighboring regions, affecting foxes (Vulpes vulpes), badgers (Meles meles), beech martens (Martes foina) and European polecats (Mustela putorius) (14, 34).

Compared to other morbilliviruses, the CDV shows the highest genetic variability (39). The main factors fostering CDV spread, cross-species infection (40, 41) and increasing virulence (17, 42–45) are genetic variability, broad spectrum of hosts and the uncontrolled animal movements of stray and domestic dogs (2, 46–48) and other wild animals (49).

As previously observed in CDV infected animals, traumatic lesions appear a frequent cause of death in wild animals, probably due to vehicular collisions (33). In this study an equal number of CDV-positive animals dying from trauma and infections were observed.

A possible correlation between neurological disease due to either traumatic or infective etiology, and a delay in the animal’s response to external stimuli could be hypothesized (49).

Although the presence of specific clinical signs or anatomopathological lesions in live or dead animals could be related to CDV infection, confirmatory laboratory tests are needed to roule out other diseases. Molecular approaches could help monitoring CDV spread in susceptible wild animal hosts, particularly endangered species, thus supporting global animal welfare.

To date, the presence of CDV in Southern Italy has been described in both domestic dogs (2, 35, 46, 50) and wildlife (51). Our study contributes to extend the knowledge on the epidemiology of CDV in wildlife.

Continuous surveillance of CDV occurrence in wildlife will help depicting a better picture the ecology of CDV in domestic and wildlife animals and safeguarding endangered species.