Dicrocoeliidae Family: Major Species Causing Veterinary Diseases.

This chapter analyses the taxonomic position of the Dicrocoeliidae family and several of its genera and species. The chapter reviews the biology of major veterinary disease-causing species, including Dicrocoelium dendriticum, Dicrocoelium hospes, Dicrocoelium chinensis, Eurytrema pancreaticum and Platynosomum fastosum. All these species have indirect life cycles with two intermediate hosts: molluscs as the first host and ants, grasshoppers and lizards as the second host. Dicrocoelium dendriticum is a widespread liver fluke found in ruminants across Europe, Asia, North Africa and North America. It can also infect humans. Dicrocoelium hospes is widely distributed in the savanna areas of Africa south of the Sahara, while D. chinensis is mainly found in ruminants in East Asia and some European countries (probably imported from Asia). Eurytrema pancreaticum is a common parasite that lives in the bile ducts, gall bladder, pancreatic ducts and intestines of ruminants. It is found in Europe, Madagascar, Asia and South America. Adults of P. fastosum live in the liver, gall bladder and pancreas of birds and mammals in Europe, Africa, Asia, North, Central and South America. Information on the epidemiology, pathology, clinical aspects, diagnosis, treatment, control, prevention and economic impact of Dicrocoeliosis caused by D. dendriticum, Eurytrematodosis and Platynosomiosis have been included.

Comparison of Intact Allergen Extracts and Allergoids For Subcutaneous Immunotherapy: The Effect of Chemical Modification Differs Both Between Species and Between Individual Allergen Molecules.

BACKGROUND: Allergen products for subcutaneous immunotherapy (SCIT) contain intact allergen extracts or chemically modified allergoids. Chemical modification was introduced to reduce allergenicity while retaining immunogenicity and thereby enable safer and more efficient allergy immunotherapy. METHODS: Experimental allergoids were produced from intact allergen extract for birch, grass, and house dust mite (HDM) to evaluate the effects of chemical modification. Preparations were compared with commercial allergoids and analyzed using SDS-PAGE/immunoblotting, IgE-inhibition assays, and crossed immunoelectrophoresis (CIE). Dermatophagoides pteronyssinus (Der p) vaccines were also tested for protease activity and immunizing capacity in a mouse model. RESULTS: The composition of IgE-binding epitopes in allergoids differed from that of intact allergen vaccines. Birch and grass allergoids produced smears of protein aggregates on SDS-PAGE, whereas intact allergen preparations showed distinct protein bands as expected. Der p allergoid vaccines, however, showed a distinct protein band corresponding to major allergen Der p 1 in both SDS-PAGE and CIE analysis, and commercial Der p allergoid vaccines showed Der p 1-related cysteine protease activity. CONCLUSION: Allergoids and intact allergen preparations differ with respect to the composition of IgE-binding epitopes. However, chemical cross-linking does not affect every allergen molecule to the same degree. Der p 1, for example, remains largely unmodified. Furthermore, the investigational HDM allergoid vaccines showed reduced and delayed immune responses when used for immunization of mice.

Comparison of Intact Allergen Extracts and Allergoids For Subcutaneous Immunotherapy: The Effect of Chemical Modification Differs Both Between Species and Between Individual Allergen Molecules

Background: Allergen products for subcutaneous immunotherapy (SCIT) contain intact allergen extracts or chemically modified allergoids. Chemical modification was introduced to reduce allergenicity while retaining immunogenicity and thereby enable safer and more efficient allergy immunotherapy. Methods: Experimental allergoids were produced from intact allergen extract for birch, grass, and house dust mite (HDM) to evaluate the effects of chemical modification. Preparations were compared with commercial allergoids and analyzed using SDS-PAGE/immunoblotting, IgE-inhibition assays, and crossed immunoelectrophoresis (CIE). Dermatophagoides pteronyssinus (Der p) vaccines were also tested for protease activity and immunizing capacity in a mouse model. Results: The composition of IgE-binding epitopes in allergoids differed from that of intact allergen vaccines. Birch and grass allergoids produced smears of protein aggregates on SDS-PAGE, whereas intact allergen preparations showed distinct protein bands as expected. Der p allergoid vaccines, however, showed a distinct protein band corresponding to major allergen Der p 1 in both SDS-PAGE and CIE analysis, and commercial Der p allergoid vaccines showed Der p 1–related cysteine protease activity. Conclusion: Allergoids and intact allergen preparations differ with respect to the composition of IgE-binding epitopes. However, chemical cross-linking does not affect every allergen molecule to the same degree. Der p 1, for example, remains largely unmodified. Furthermore, the investigational HDM allergoid vaccines showed reduced and delayed immune responses when used for immunization of mice (AU)

Antecedentes: Los productos de alérgenos para inmunoterapia subcutánea (SCIT) contienen extractos de alérgenos intactos o alergoides modificados químicamente. En este trabajo se ha hecho una modificación química para reducir la alergenicidad a la vez que se conservaba la inmunogenicidad, y por lo tanto, permitir una inmunoterapia más segura y eficiente. Métodos: Se produjeron alergoides experimentales a partir de extracto de alérgeno intacto para abedul, hierba y ácaros del polvo doméstico (HDM) y se evaluaron los efectos de la modificación química realizada. Las preparaciones se compararon con alergoides comerciales y se analizaron mediante SDS-PAGE/inmunotransferencia, ensayos de inhibición de IgE e inmunoelectroforesis cruzada (CIE). Las vacunas de Dermatophagoides pteronyssinus (Der p) también se probaron para determinar la actividad de la proteasa y la capacidad de inmunización en un modelo de ratón. Resultados: La composición de los epítopos de unión a IgE en los alergoides difería de las vacunas de alérgenos intactas. Los alergoides de hierba y abedul produjeron manchas de agregados de proteínas en el SDS-PAGE, mientras que las preparaciones de alérgenos intactos mostraron distintas bandas de proteínas como se esperaba. Las vacunas alergoides Der p, sin embargo, mostraron una banda de proteína distinta de la correspondiente al alérgeno principal Der p 1 en los análisis SDS-PAGE y CIE. Las vacunas alergoides comerciales Der p mostraron actividad de cisteína proteasa relacionada con Der p 1.Conclusión: Los alergoides y las preparaciones de alérgenos intactos difieren con respecto a la composición de los epítopos de unión a IgE; sin embargo, el entrecruzamiento químico no afecta a todas las moléculas de alérgenos de un modo similar. Der p 1, por ejemplo, permanece prácticamente sin modificar. Además, las vacunas alergoides de HDM produjeron respuestas inmunitarias reducidas y tardías cuando se usaron para la inmunización de ratones (AU)

Effects of Paratuberculosis Vaccination at Different Ages in a Dairy Goat Herd: A 2-Year Follow-Up.

Vaccination could be considered as an effective method for paratuberculosis control, although controversial, with a need for investigation in some aspects. The objective of this study was to evaluate the effect of vaccination, depending on the age of the animals, on their immune response, the reduction of paratuberculosis cases, mortality and culled animals in a commercial dairy herd. Goats from three different ages were immunized with the inactivated Gudair® vaccine. Peripheral antibody and IFN-γ output were evaluated for 21 months post-vaccination (mpv) and intradermal skin tests (IDSTs) for tuberculosis, with avian- and bovine-purified protein derivatives (PPD), were carried out at 6 and at 18 mpv to evaluate the humoral and cellular immune peripheral responses, respectively. The number of dead or culled animals, regardless of the reason, was also monitored and the causes of death determined by pathological examination. A significant increase in the production of IFN-γ was observed in all the vaccinated groups when the blood samples were stimulated with avian PPD, from 3 mpv to 18 mpv, and with bovine PPD, between 3 and 21 mpv. Moreover, serum antibody levels increased between 3 and 21 mpv in all vaccinated groups. The highest levels were found in animals vaccinated at 5 months, and the lowest in adult individuals. No positive reactants to tuberculosis were found by intradermal skin test. No animal losses associated with clinical paratuberculosis were detected in any of the groups. The number of total culled animals was significantly lower in the vaccinated than in the unvaccinated groups, especially on 1.5-month-old vaccinated kids. These results suggest that vaccination of paratuberculosis, especially in young animals, could induce heterologous protection.

Peripheral IFN-É£ Production after Blood Stimulation with Different Mycobacterial Antigens in Goats Vaccinated against Paratuberculosis.

Vaccination can be an efficient method for the control of paratuberculosis in ruminants. However, the official tuberculosis control tests cross-interfere with the animals vaccinated against paratuberculosis. In order to test and compare new antigens that could solve this problem, the production of interferon-gamma (IFN-γ) in peripheral blood at different post-vaccination days in experimental kids and adult goats, in field conditions, using the avian and bovine purified protein derivative (PPD), the johnin, two peptide cocktails of Mycobacterium bovis (PC-EC and PC-HP) and the antigens VK 055 and VK 067 of Mycobacterium avium subspecies paratuberculosis (Map) has been analyzed in vitro. The non-specific production of IFN-γ was observed after blood stimulation with the PC-EC and PC-HP cocktail in any sample from vaccinated animals, whereas it was detected when bovine PPD was used. These results support the possible use of these new Mycobacterium bovis antigens in the in the differentiation of animals vaccinated against paratuberculosis or infected with tuberculosis by improving the specificity of bovine PPD. In contrast, the two Map antigens tested in this study did not improve the sensitivity of johnin or avian PPD in the detection of vaccinated or Map-infected goats.

Case Report: Multiple Cavernous Pericardial Lymphangioma (Pericardial Lymphangiomatosis) in a Captive Peregrine Falcon (Falco peregrinus brookei).

A 12-year-old female peregrine falcon (Falco peregrinus brookei) from a private raptor breeding facility that presented a good body condition, died suddenly without showing previous symptoms. At necropsy, in the coelomic cavity, multiple cystic structures demarcated by a thin transparent wall and filled with a serous content were observed. They were firmly adhered to the cranial part of the epicardium and adjacent tissues and occupied the entire thoracic area of the coelomic cavity. Microscopically, emerging simultaneously from several areas the epicardium, multiple irregular channels and cystic spaces, lined by a single endothelial cell layer and separated by fibrovascular septa containing smooth muscle tissue, were observed. Immunohistochemical examination revealed that the neoplastic endothelial cells positively immunolabelled for the pan-endothelial marker factor VIII-related antigen but immunostained negative for cytokeratins (PCK26) while strong positivity for sarcomeric α-smooth muscle actin (α-SMA) was detected in the cystic walls. Based on the morphological and immunohistochemical findings, lesions were determined as consistent with a multiple cavernous pericardial lymphangioma, or pericardial lymphangiomatosis, a rare vascular neoplasm. The animal also showed a diffuse chronic perihepatitis, a necrotic area in the liver and foci of cartilaginous metaplasia and calcification in the aorta and vena cava. Literature review, particularly on the epidemiology of lymphangioma, demonstrated the rarity of this tumor in the different animal species and in this location, particularly in birds, being the first report of this type of tumor in a peregrine falcon.

Pathological Study of Facial Eczema (Pithomycotoxicosis) in Sheep.

Facial eczema (FE) is a secondary photosensitization disease of farm ruminants caused by the sporidesmin A, produced in the spores of the saprophytic fungus Pithomyces chartarum. This study communicates an outbreak of ovine FE in Asturias (Spain) and characterizes the serum biochemical pattern and the immune response that may contribute to liver damage, favoring cholestasis and the progression to fibrosis and cirrhosis. Animals showed clinical signs of photosensitivity, with decrease of daily weight gain and loss of wool and crusting for at least 6 months after the FE outbreak. Serum activity of γ-glutamyltransferase and alkaline phosphatase were significantly increased in sheep with skin lesions. In the acute phase, edematous skin lesions in the head, hepatocytic and canalicular cholestasis in centrilobular regions, presence of neutrophils in small clumps surrounding deposits of bile pigment, ductular proliferation, as well as cholemic nephrosis, were observed. Macrophages, stained positively for MAC387, were found in areas of canalicular cholestasis. In the chronic phase, areas of alopecia and crusting were seen in the head, and the liver was atrophic with large regeneration nodules and gallstones. Fibrosis around dilated bile ducts, "typical" and "atypical" ductular reaction and an inflammatory infiltrate composed of lymphocytes and pigmented macrophages, with iron deposits and lipofuscin, were found. The surviving parenchyma persisted with a jigsaw pattern characteristic of biliary cirrhosis. Concentric and eccentric myointimal proliferation was found in arteries near damaged bile ducts. In cirrhotic livers, stellated cells, ductular reaction, ectatic bile ducts and presence of M2 macrophages and lymphocytes, were observed in areas of bile ductular reaction.

Fasciola hepatica co-infection enhances Th1 immune response in the adventitial layer of non-fertile Echinococcus granulosus cysts.

Cystic echinococcosis is a widespread zoonosis caused by the larval stage of the tapeworm Echinococcus granulosus. In intermediary hosts, two types of echinococcal cysts can be found: fertile, which produce protoscoleces, the infective form of the parasite to dogs; and infertile, that do not present protoscoleces and therefore are not able to continue with the parasite life cycle. The adventitial layer, the local immune response against the cyst, plays an important role in cyst fertility. Grazing cattle can often feature Fasciola hepatica co-infection, a parasite known to modulate the host systemic immune response. In this work the cellular Th1/Th2 immune profiles were evaluated in the adventitial layer of fertile and non-fertile cysts with and without co-infection with Fasciola hepatica. Measuring with immunohistochemistry and qPCR in adventitial layer, we report that non-fertile cysts present higher levels of Th1 cytokines (IFN-γ (P < 0.0001) and TNF-α (P < 0.05)), and fertile cysts have higher levels of Th2 cytokines (IL-4 (P < 0.001)). Co-infection with Fasciola hepatica is associated with a decrease in the expression of IL-4 (P < 0.05) and an increase in the expression of IFN-γ (P < 0.0001) in the adventitial layer of fertile cysts. Non-fertile cysts were associated with higher levels of Th1 cytokines in the adventitial layer, with IFN-γ expression enhanced by F. hepatica co-infection (P < 0.0001), confirming that polyparasitism should be considered in the treatment and control of naturally infected cattle.

Assessment of Acute-Phase Protein Response Associated with the Different Pathological Forms of Bovine Paratuberculosis.

In this study, the concentrations of two acute-phase proteins (APPs), haptoglobin (Hp) and serum amyloid A (SAA), were quantitatively assessed in serum samples from cattle naturally infected with paratuberculosis (PTB). APP profiles were compared across 190 animals classified according to the different pathological forms associated with infection: uninfected (n = 59), with focal lesions (n = 73), multifocal lesions (n = 19), and diffuse paucibacillary (n = 11) and diffuse multibacillary lesions (n = 28). Our results showed a significant increase in both APPs in infected animals compared to the control group, with differences depending on the type of lesion. Hp and SAA levels were increased significantly in all infected animals, except in cows with diffuse multibacillary lesions that showed similar values to non-infected animals. The expression pattern of both APPs was similar and negatively correlated with the antibody levels against PTB. These results indicate that the release of Hp and SAA is related to the presence of PTB lesions associated with a high cell-mediated immune response and a lower bacterial load, suggesting that the pro-inflammatory cytokines that are associated with these forms are the main stimulus for their synthesis. These molecules could show some potential to be used as putative biomarkers of PTB infection, particularly for the identification of subclinical animals showing pathological forms related to latency or resistance to the development of advanced lesions.

Macrophages and T Lymphocytes in the Ovine Placenta After Experimental Infection With Toxoplasma gondii.

Early abortion in ovine toxoplasmosis has had limited investigation. This study evaluated the immune response in the placenta of sheep orally infected with Toxoplasma gondii and euthanized between 2 and 4 weeks postinfection. Toxoplasma infection of the placenta was only found at 4 weeks after infection. Parasitic debris in foci of necrosis were immunolabeled in the maternal caruncle, whereas well-preserved intracellular parasitic vacuole-like structures were found in trophoblasts of fetal cotyledon. Early abortions had increased macrophages in caruncular septa, whereas in later abortions the placentas containing the parasite had an increase of T lymphocytes and macrophages mainly in the fetal cotyledons. This study suggests that the immune response in both the fetal and maternal compartments of the placenta may contribute to the pathogenesis of ovine toxoplasmosis and that these responses differ between early and late presentations of the disease.

Local assessment of WC1+ γδ T lymphocyte subset in the different types of lesions associated with bovine paratuberculosis.

The local expression of WC1+ γδ T lymphocytes subset has been evaluated by immunohistochemical methods at the different types of lesions present in cows naturally infected with Mycobacterium avium subsp. paratuberculosis (Map) and in non-infected control animals. Infected cattle were either in the latent/subclinical (focal lesions) or clinical (diffuse paucibacillary and multibacillary forms) stage of paratuberculosis. To assess the cell distribution, a differential cell count was carried out at the lamina propria, gut-associated lymphoid tissue and submucosa. A significant increase in the number of WC1+ γδ T cells was observed in all the infected animals, regardless of the type of lesion. Cows with focal lesions showed higher number of labeled cells than those with diffuse forms, where no differences were found between the two types. This increase in the number of positively immunolabelled lymphocytes in infected animals was seen in the lamina propria, with higher values in those with focal lesions. While in the lymphoid tissue no differences in the numbers were observed, in animals with focal lesions, WC1+ γδ T cells tended to be located at the periphery of the granulomas. These findings suggest a proinflammatory action of WC1+ γδ T lymphocytes in bovine paratuberculosis, which might play an important role in the containment of the Map-infection in the focal granulomas located in the lymphoid tissue, helping to prevent the progression toward diffuse forms responsible for the clinical signs.

Causes of Mortality and Disease in Rabbits and Hares: A Retrospective Study.

In this study we determined the causes of mortality and disease in a total of 325 lagomorphs (rabbits and hares) in northern Spain between 2000 and 2018. Risk factors such as the species, age, sex, time of year and origin were also considered. Clinical signs, gross and histopathological findings and ancillary test results were the basis for the final diagnoses that were reviewed to classify and identify the different disorders. A total of 26 different conditions were identified. A single cause of death or illness was detected in 267 animals. They were grouped into parasitic conditions (n= 65; 24.34%) represented by encephalitozoonosis, hepatic coccidiosis, hepatoperitoneal cysticercosis, intestinal coccidiosis, parasitic gastritis and cutaneous ectoparasitosis; bacterial diseases (n = 56; 20.97%) including pseudotuberculosis, blue breast, skin abscesses, tularemia, pneumonic pasteurellosis and staphylococcal infections; nutritional and metabolic diseases (n = 48; 17.97%) with epizootic rabbit enteropathy, hepatic steatosis and pregnancy toxemia as prominent diseases; viral infections (n= 31; 11.61%) comprising rabbit hemorrhagic disease and myxomatosis and miscellaneous causes (n = 31; 11.61%) where rabbit enteritis complex, renal conditions (nephrosis), heat stroke, and arterial bone metaplasia were included; neoplasms (n = 12; 4.49%) represented by uterine adenocarcinoma, mammary adenocarcinoma, cutaneous fibroma, intestinal lymphoma and hepatic cholangiocarcinoma; toxicoses (n = 11; 4.11%); trauma-related injuries (n = 9; 3.37%) and finally congenital diseases (n = 4; 1.49%). In 58 animals of the study, some of these conditions were presented jointly. We discuss the detection frequency, possible causes or associated factors of the different pathologies as well as the importance of the different variables considered.

Lymphocyte Populations in the Adventitial Layer of Hydatid Cysts in Cattle: Relationship With Cyst Fertility Status and Fasciola Hepatica Co-Infection.

Cystic echinococcosis is a worldwide zoonosis caused by the cestode Echinococcus granulosus. Two types of hydatid cysts occur in intermediate hosts: fertile cysts that generate protoscoleces from the germinal layer of the cyst, and infertile cysts that do not produce protoscoleces and are unable to continue the life cycle of the parasite. The adventitial layer, a host-derived fibrous capsule surrounding the hydatid cyst, is suggested to play an important role in local immune regulation during infection and in fertility of the cysts. Fasciola hepatica, another important parasite of cattle, induces a characteristic Th2-like immune response that could modulate the immune response against E. granulosus. Natural co-infection of both parasites is common in cattle, but no reports describe the local immune response against E. granulosus with F. hepatica infection in the same host. This study analyzed the number and distribution of T and B cells in the adventitial layer of liver and lung cysts and the relationship with cyst fertility and F. hepatica co-infection. T lymphocytes were the predominant cell type in the adventitial layer of infertile hydatid cysts and were more numerous in infertile hydatid cysts. B lymphocyte numbers were not associated with hydatid cyst fertility. Mast cells were infrequent in the adventitial layer. The number of T and B cells was not associated with F. hepatica co-infection. The present study contributes to the understanding of local immune responses in bovine cystic echinococcosis.

Peripheral and placental immune responses in sheep after experimental infection with Toxoplasma gondii at the three terms of gestation.

Although it is known that gestation could influence the clinical course of ovine toxoplasmosis, the precise effect of the term of gestation when sheep are infected are yet mostly unknown. The aim of this study was to evaluate the peripheral and placental immune responses developed in pregnant sheep after experimental infection with Toxoplasma gondii at different times of gestation. Thirty-six pregnant sheep were allocated in different groups, orally inoculated with sporulated oocysts of T. gondii at early, mid and late gestation and culled within 30 days post-infection. The peripheral humoral and cytokine responses were evaluated, as well as the transcription of cytokines at the placenta. Serological analysis revealed that, regardless the term of gestation when infected, specific IgG against T. gondii were detected from day 8 post-infection and there was an early peripheral release of IFN-γ at the first week post-infection followed by a short peak of IL10 and TNF-α at the second week post-infection. There were no significant differences in this response between infected groups. At the placenta, a similar increase in transcription of IFN-γ, and TNF-α was found at the three terms of gestation, while IL-4 increased mainly at the first and second terms and IL-10 transcription was higher at the last term. While these findings show that both Th1 and Th2 cytokines play a key role in the pathogenesis of ovine toxoplasmosis and that placental and peripheral immune responses do not closely correlate, there seems to be no clear modulation of these responses along the gestation.

Dicrocoeliidae Family: Major Species Causing Veterinary Diseases.

This chapter analyses the taxonomic position of Dicrocoeliidae family and several of its genus and species. The biology of the major species causing veterinary diseases such Dicrocoelium dendriticum, Dicrocoelium hospes, Dicrocoelium chinensis, Eurytrema pancreaticum and Platynosomum fastosum, has been reviewed. All these species have an indirect life cycle, involving two intermediate hosts (molluscs as first and ants, grasshoppers and lizards as second). Dicrocoelium dendriticum is a very widespread hepatic trematode in the ruminants of many countries in Europe, Asia, North Africa and North America, even affecting humans. Dicrocoelium hospes is widely distributed in the savanna areas of Africa south of the Sahara, whilst D. chinensis has mainly been found in ruminants in East Asia and some European countries (probably imported from Asia). Eurytrema pancreaticum is a common parasite whose adults live in ruminant bile ducts, gall bladder, pancreatic ducts and intestines in Europe, Madagascar, Asia and South America. Adult P. fastosum live in the liver, gall bladder and pancreas of birds and mammals in Europe, Africa, Asia, North, Central and South America. Information about the epidemiology, pathology, clinical aspect, diagnosis, treatment, control, prevention and economic impact mainly of Dicrocoeliosis produced by D. dendriticum, as well as of Eurytrematodosis and Platynosomiosis, has been included.

Tissue Tropism of Chicken Anaemia Virus in Naturally Infected Broiler Chickens.

Chicken anaemia virus (CAV) causes chicken infectious anaemia, a severe disease characterized by anaemia and immunosuppression and leading to serious economic losses in the poultry industry worldwide. Although CAV infection has been investigated under experimental conditions, information regarding natural infection is scarce. This report describes an outbreak of CAV infection in 18-day-old broiler chickens and investigates virus tropism in affected birds. Thymic atrophy, pale bone marrow, swelling of the legs and foot ulcers (gangrenous dermatitis) were the most common gross lesions. Severe lymphoid cell depletion in the thymic cortex and presence of intranuclear acidophilic inclusion bodies, depletion of haemopoietic cells in bone marrow and presence of lymphoid infiltrates in several organs were also observed. Immunohistochemical labelling demonstrated the CAV antigens VP1 and VP3 in several organs. The expression of both proteins was similar in the thymic cortex and in the bone marrow, the main target organs of CAV; however, VP3 expression was more abundant in the other organs. Labelling of serial sections showed that CD3+ T lymphocytes might be responsible for the dissemination of the virus from the thymus and bone marrow to other organs and that virus-induced apoptosis, mediated through caspase-3, occurred mainly in the thymus and bone marrow.

Systemic Necrotizing Vasculitis in Sheep Is Associated With Ovine Herpesvirus 2.

Ovine herpesvirus 2 (OvHV-2) is one of the gammaherpesviruses in the genus Macavirus that can cause malignant catarrhal fever (MCF) in ungulates. Sheep are the adapted host for OvHV-2 and it is generally assumed that infection is not associated with disease in this species. However, cases of "polyarteritis nodosa" or idiopathic systemic necrotizing vasculitis reported in sheep are similar to vascular lesions in clinically susceptible species with MCF. Using a recently developed in situ hybridization (ISH) method, we were able to identify OvHV-2 nucleic acids within lesions and correlate the viral distribution with systemic necrotizing vasculitis in 9 sheep, including both naturally and experimentally OvHV-2-infected animals. ISH, combined with polymerase chain reaction and histology, identify OvHV-2 as the likely agent responsible for sporadic, MCF-like vascular disease in sheep.