Investigation, management and control of a maedi outbreak in Norway in 2019-2020.

BACKGROUND: Visna-maedi is a notifiable disease in Norway, and eliminating the disease is a national goal. The import of sheep into Norway is very limited, and strict regulations apply to the movement of small ruminants between flocks and within defined geographical regions. Several outbreaks have occurred in the last 50 years, and the most recent before 2019 occurred in Trøndelag county in Central Norway in 2002. A national surveillance programme for small ruminant lentivirus infection exists since 2003. RESULTS: In 2019, the national surveillance programme detected seropositive animals for small ruminant lentivirus in a sheep flock in Trøndelag. Based on the result of polymerase chain reaction analysis and histopathological findings, the Norwegian Food Safety Authority concluded the diagnosis of maedi. Further investigations detected maedi in eight additional sheep flocks in the same county. The flocks were placed under restrictions, and the authorities also imposed restrictions on 82 contact flocks. Sequencing of partial gag genes indicated that the virus in the current outbreak was related to the small ruminant lentivirus detected in the same area between 2002 and 2005. CONCLUSIONS: The outbreak investigation shows the need for sensitive and specific diagnostic methods, and an improved and more targeted surveillance strategy. It also demonstrates the risk of disease spreading between flocks through animal movements, and highlights the importance of biosecurity and structured livestock trade. In addition to allowing livestock trade only from flocks documented free from maedi, it may be necessary to monitor sheep flocks over many years, when aiming to eliminate maedi from the Norwegian sheep population.

Identification of Oocyst-Driven Toxoplasma gondii Infections in Humans and Animals through Stage-Specific Serology-Current Status and Future Perspectives.

The apicomplexan zoonotic parasite Toxoplasma gondii has three infective stages: sporozoites in sporulated oocysts, which are shed in unsporulated form into the environment by infected felids; tissue cysts containing bradyzoites, and fast replicating tachyzoites that are responsible for acute toxoplasmosis. The contribution of oocysts to infections in both humans and animals is understudied despite being highly relevant. Only a few diagnostic antigens have been described to be capable of discriminating which parasite stage has caused an infection. Here we provide an extensive overview of the antigens and serological assays used to detect oocyst-driven infections in humans and animals according to the literature. In addition, we critically discuss the possibility to exploit the increasing knowledge of the T. gondii genome and the various 'omics datasets available, by applying predictive algorithms, for the identification of new oocyst-specific proteins for diagnostic purposes. Finally, we propose a workflow for how such antigens and assays based on them should be evaluated to ensure reproducible and robust results.

Seroprevalence of Hepatitis E Virus in Moose (Alces alces), Reindeer (Rangifer tarandus), Red Deer (Cervus elaphus), Roe Deer (Capreolus capreolus), and Muskoxen (Ovibos moschatus) from Norway.

Hepatitis E virus (HEV), a major cause of viral hepatitis worldwide, is considered an emerging foodborne zoonosis in Europe. Pigs (Sus scrofa domestica) and wild boars (S. scrofa) are recognized as important HEV reservoirs. Additionally, HEV infection and exposure have been described in cervids. In Norway, HEV has been identified in pigs and humans; however, little is known regarding its presence in wild ungulates in the country. We used a species-independent double-antigen sandwich ELISA to detect antibodies against HEV in the sera of 715 wild ungulates from Norway, including 164 moose (Alces alces), 186 wild Eurasian tundra reindeer (Rangifer tarandus tarandus), 177 red deer (Cervus elaphus), 86 European roe deer (Capreolus capreolus), and 102 muskoxen (Ovibos moschatus). The overall seroprevalence was 12.3% (88/715). Wild reindeer had the highest seropositivity (23.1%, 43/186), followed by moose (19.5%, 32/164), muskoxen (5.9%, 6/102), and red deer (4%, 7/177). All roe deer were negative. According to our results, HEV is circulating in wild ungulates in Norway. The high seroprevalence observed in wild reindeer and moose indicates that these species may be potential reservoirs of HEV. To the authors' knowledge, this is the first report of HEV exposure in reindeer from Europe and in muskoxen worldwide.

Toxoplasma gondii infection in two captive Patagonian maras.

Toxoplasma gondii infection was diagnosed in 2 captive Patagonian maras (Dolichotis patagonum). One animal developed fatal systemic toxoplasmosis and had concurrent localized bacterial and fungal infections; its daughter remained clinically healthy. Microscopic findings included acute, coagulative necrosis, lymphohistiocytic inflammatory infiltrates, and extra- and intracellular parasites in the liver, myocardium, urinary bladder, and adrenal glands of the diseased animal. PCR and subsequent genotyping of parasites from fresh tissue from both cases revealed infection with T. gondii genotype II. Direct agglutination testing of blood from the healthy individual revealed high levels of T. gondii IgG antibodies. T. gondii is a potential cause of disease and lethality in captive and wild Patagonian maras, and toxoplasmosis should be considered when managing and providing veterinary care for this species.

A geographic cluster of malignant catarrhal fever in moose (Alces alces) in Norway.

Three cases of lethal sheep-associated malignant catarrhal fever (SA-MCF) in free-ranging moose (Alces alces) were diagnosed in Lesja, Norway, December 2008-February 2010. The diagnosis was based on PCR identification of ovine herpesvirus 2 (OvHV-2) DNA (n = 3) and typical histopathologic lesions (n = 1). To study the possibility of subclinical or latent MCF virus (MCFV) infection in this moose population and in red deer (Cervus elaphus), we examined clinically normal animals sampled during hunting in Lesja 2010 by serology and PCR. Sera from 63 moose and 33 red deer were tested for antibodies against MCFV by competitive-inhibition enzyme-linked immunosorbent assay. To test for MCFVs, a consensus PCR for herpesviral DNA was run on spleen samples from 23 moose and 17 red deer. All samples were antibody and PCR negative. Thus, there is no evidence of previous exposure, subclinical infection, or latent infection in this sample. This seasonal cluster of SA-MCF cases (2008-10) may be attributable to exposure of moose to lambs when OvHV-2 shedding is presumed to be maximal, compounded by an unusual extended grazing period by sheep in the autumn.

Malignant catarrhal fever virus identified in free-ranging musk ox (Ovibos moschatus) in Norway.

To study the epizootiology of malignant catarrhal fever viruses (MCFV), serum and spleen samples collected in 2004-2011 from a free-ranging musk ox (Ovibos moschatus) population in Dovrefjell, Norway, were examined. Sera were tested for antibodies against MCFV by competitive enzyme-linked immunosorbent assay and spleen samples were examined by a consensus polymerase chain reaction (PCR) for herpesviral DNA and sequencing identification. The study included 101 musk oxen, of which 61 were examined with both tests. Antibodies against MCFV were found in 65 of 72 musk oxen (90%). Antibody prevalence increased with age from 67% in calves to 96% in adults. We detected MCFV DNA by consensus PCR in 67 of 90 spleen samples tested (74%). The prevalence of PCR-positive musk oxen increased with age from 60% in calves to 81% in adults. Fifty (82%) of the 61 animals subjected to both PCR and serology were positive in both tests. Sequencing analysis showed that all PCR-positive animals were infected with a MCFV previously identified in musk oxen in North America. The results suggest that MCFV-Muskox is enzootic in the Dovrefjell musk ox population and that many calves seem to be infected early in life. This is the first report of MCFV-Muskox infection in free-ranging musk ox outside North America.

Natural killer cells act as early responders in an experimental infection with Neospora caninum in calves.

The intracellular protozoan parasite Neospora caninum is a cause of abortion and congenital disease in cattle worldwide. We have previously shown that natural killer (NK) cells produce IFN-gamma in response to N. caninum tachyzoites in vitro. This study aimed to investigate the role of NK cells and other cellular immune responses in an experimental N. caninum infection model in calves. Phenotyping of peripheral blood lymphocytes showed a drop in the percentage of NK cells at days 4-6 after i.v. inoculation, followed by an increase in the percentage of both NK cells and CD8+ T cells which peaked at days 11-15. A whole blood flow cytometric assay showed that CD4+ T cells were the major IFN-gamma producing cells, but in the early stages of the infection both NK cells and CD8+ T cells contributed to IFN-gamma production. We also compared the ability of two different N. caninum antigen preparations--sonicated soluble antigens and intact heat-inactivated parasites--to induce proliferation and IFN-gamma production in various cell types. Heat-inactivated tachyzoites induced a 3.7 times greater increase in the number of IFN-gamma producing NK cells compared with sonicated soluble antigens. This indicated the presence of some NK cell-stimulating antigens in the intact tachyzoite that were absent from the sonicated soluble antigens. The heat-inactivated whole tachyzoites also inhibited gammadelta T cell proliferation while the soluble antigens from N. caninum did not. We believe this is the first time NK cells have been demonstrated to be early responders in N. caninum infection in calves.

The protozoan Neospora caninum directly triggers bovine NK cells to produce gamma interferon and to kill infected fibroblasts.

Natural killer (NK) cells are considered to be key players in the early innate responses to protozoan infections, primarily indirectly by producing gamma interferon (IFN-gamma) in response to cytokines, like interleukin 12 (IL-12). We demonstrate that live, as well as heat-inactivated, tachyzoites of Neospora caninum, a Toxoplasma-like protozoan, directly trigger production of IFN-gamma from purified, IL-2-activated bovine NK cells. This response occurred independently of IL-12 but was increased by the addition of the cytokine. A similar IFN-gamma response was measured in cocultures of NK cells and N. caninum-infected autologous fibroblasts. However, no NK cell-derived IFN-gamma response was detected when cells were cultured with soluble antigens from the organism, indicating that intact tachyzoites or nonsoluble components are necessary for NK cell triggering. Furthermore, N. caninum-infected autologous fibroblasts had increased susceptibility to NK cell cytotoxicity compared to uninfected fibroblasts. This cytotoxicity was largely mediated by a perforin-mediated mechanism. The activating receptor NKp46 was involved in cytotoxicity against fibroblasts but could not explain the increased cytotoxicity against infected targets. Interestingly, N. caninum tachyzoites were able to infect cultured NK cells, in which tachyzoites proliferated inside parasitophorous vacuoles. Together, these findings underscore the role of NK cells as primary responders during a protozoan infection, describe intracellular protozoan infection of NK cells in vitro for the first time, and represent the first functional study of purified bovine NK cells in response to infection.