Serafina Farigu

Maedi-Visna Virus Detection in Ovine Third Eyelids

Maedi-visna is a systemic disease of sheep caused by a lentivirus, maedi-visna virus (MVV), which mainly affects the lungs and central nervous system but may also affect the mammary glands, joints and other tissues. The aim of the present study was to determine whether the third eyelid was affected in cases of systemic infection. Third eyelid and lung samples from sheep naturally infected with maedi were used. Total DNA was extracted from paraffin-wax-embedded tissues, and a nested polymerase chain reaction (PCR) was performed to amplify MVV proviral DNA. The samples were also tested by in-situ PCR and immunohistochemical methods specific for the detection of MVV proviral DNA and p25, respectively. All sheep showed moderate to severe chronic lymphoproliferative inflammation in the third eyelids. Products of the expected size were obtained by PCR from both lung and third eyelid tissue. In the nictitating membrane, MVV proviral DNA was detected in situ within macrophages, and glandular, ductal and surface epithelia. Immunohistochemistry demonstrated that the infection was productive. Taken together, these results indicate that the third eyelid may represent a target for natural MVV infection and may play a role in disease transmission.

Exocyclic DNA adducts in sheep with skeletal fluorosis resident in the proximity of the Portoscuso-Portovesme industrial estate on Sardinia Island, Italy

The mechanisms by which fluoride produces its toxic effects are still not clear. Therefore, we conducted a cross-sectional study to evaluate the fluoride-induced toxicity on randomly selected sheep with skeletal fluorosis resident near the large non-ferrous metallurgy Portoscuso-Portovesme industrial estate and the Carbonia and Gonnessa towns (control district) with respect to animals from a remote site without industrial settlements, on Sardinia Island, Italy. We measured the prevalence of 3-(2-deoxy-β-D-erythro-pentafuranosyl)pyrimido[1,2-α]purin-10(3H)-one deoxyguanosine (M1dG) adducts, a biomarker of oxidative stress and lipid peroxidation, in the soft organs of the study animals using the 32P-postlabelling assay. Then, we analysed the association between M1dG adducts and sheep-farm localization. In addition, cadmium and lead levels were measured in the same matrices by mass spectrometry. The histopathology analysis showed that the sheep resident near the industrial site and in the control district presented typical manifestations of fluorosis. The statistical analyses using log-normal regression models indicated that there was a significant association between exocyclic DNA adducts and skeletal fluorosis. The mean ratios, adjusted for age, of renal and hepatic M1dG for the sheep with fluorosis were 5.09, 95% C.I. 1.67–15.53 and 2.04, 95% C.I. 0.91–4.57, p = 0.009 and p = 0.078, respectively. After stratification for sheep-farm localization, the renal and hepatic levels of M1dG adducts were significantly higher in the sheep resident near the industrial estate as compared to controls. Intermediate amounts of DNA damage were observed in the kidney of the sheep living in the control district. As expected, the levels of cadmium and lead were significantly increased in the sheep with fluorosis as compared to controls. Also, the concentrations of heavy metals were driven from the distance of the sheep-farms to the industrial site. Our results broaden knowledge about the role of exocyclic DNA adducts in the etiology of skeletal fluorosis. Fluoride generated from the non-ferrous metallurgy complexes may increase the intracellular amounts of oxidative stress and ROS within the soft organs. A continuous attack to DNA may contribute to the general decline of cellular functions, ranging from disturbance of DNA metabolism, triggering cell-cycle arrest and apoptosis, to inducing necrosis and cellular degenerative changes.

Map Lesions: In Situ Investigation on Samples of Multibacillar and Paucibacillar Ovine PTBC

Paratuberculosis (PTBC) is a chronic inflammation of the intestines of ruminants but it can also strike other species, including primates. Ovine paratuberculosis lesions have been the subject of authoritative observations (Perez et al ., 1996). The macroscopic picture shows a thickening and wrinkling of the intestinal mucous membrane , regional lymphangitis and lymphandenopathy, principally found in the terminal part of the ileum but also in the colon, jejunum and duodenum. Histomorphological pictures are characterised by chronic granulomatous enterocolitis, whose typical clinical symptom is the loss of weight due to profuse diarrhea. The etiological agent of PTBC is the bacterium Mycobacterium avium subsp. paratuberculosis (MAP) (Marek, 1910); its presence gives rise to a persistent infection of the intestinal macrophages which appear foamy, in a small granuloma within the interfollicular-basal area of the Peyer’s plaques of the ileal tract. In some cases, even in the presence of marked lesions and clear, clinical symptomatology, coloration of the Ziehl-Nielsen (ZN) produces a negative result. DNA analysis of MAP has identified a peculiar insertion sequence, IS900, present in 15–20 pairs per genome (Green et al ., 1989).