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  • A Nationwide Seroepidemiologic Study on Q Fever Antibodies in Sheep of Portugal
    Publication . Cruz, Rita; Esteves, Fernando; Nóbrega, Carmen; Santos, Carla; Ferreira, Ana S.; Mega, Cristina; Coelho, Ana C.; Vala, Helena; Mesquita, João
    Introduction: Q fever is an almost global zoonotic disease caused by Coxiella burnetii. Human infections can produce acute and chronic disease that can lead to abortions and stillbirths in pregnant women, usually infected by the inhalation of C. burnetii-contaminated aerosols or through consumption of contaminated products. Sheep are one of the primary animal reservoirs with disease being associated with vast shedding of bacteria in placentas, feces, milk, and birth fluids. Although almost neglected in the past, recent outbreaks of sheep origin have alerted the public and the scientific community. Materials and Methods: An epidemiologic survey to estimate the seroprevalence of Q fever antibodies was performed in a representative number of sheep of all regions of continental Portugal (n = 1068), using a commercial ELISA (ID Screen Q Fever Indirect Multi-species Kit; IDvet, Montpellier, France). Results and Discussion: An anti-C. burnetii seroprevalence of 11.4% (95% confidence interval 9.6–13.5) was found, with a clear distinction between the Center region with highest seroprevalence, and the rest of the territory. Sheep traditional farming is widely present in Portugal and is part of the cultural and gastronomical background of the country. This close proximity to small ruminants may contribute to the zoonotic transfer to humans.
  • Hepatic histopathological lesions in acute controled haemorrhage followed by volume replacement with a crystalloid or colloid solution
    Publication . Vala, Helena; Pina, R.; Cruz, R.; Venâncio, C.; Esteves, Fernando; Silva, A.; Mesquita, João; Ortiz, A. L.; Ferreira, D.
    Introduction: severe hemorrhage remains the major cause of morbidity and mortality in trauma victims or surgical intervened animals, despite of all the advances in the therapeutic approach. The resulting injuries, or even death, are due to the deficit in intravascular volume and subsequent development of hypovolaemic fluid state, leading to poor tissue perfusion and consequent decreased oxygen delivery to the tissues, often with compromise of organ function. Objective: the aim of this work was to evaluate the effect of different intravenous solutions used for volume replacement following acute controlled haemorrhage, one crystalloid (RL) and one colloid (HES130/0.4) in the integrity of the hepatic tissue in a pig animal model. Material and methods: hepatic samples were collected from animals submitted to passive arterial blood bleeding and reperfusion with a crystalloid (RL) (G1) and with a synthetic colloid (HES 130/0.4) (G2). Samples were also collected from animals that were not subjected to acute bleeding nor volume replacement (G3, control group). All procedures were carried out under personal and project licenses approved by the Ethical Committee of the national regulatory office. Samples were collected and fixed in 10% neutral-buffered formalin, for a maximum of 24hours, embedded in paraffin wax and 3µm sections were stained for routine histopathology with haematoxylin and eosin. Results: the histopathological assessment revealed no statistically significant differences between the three groups. However, some lesions were more often expressed in some groups. More severe hepatocellular hydropic degeneration and hepatocellular steatosis was seen in G1, which is the only group in which haemorrhage was observed and within which oedema was not present. Hyperaemia was only observed in G2 and G3. Necrosis was not present in any of the groups. Conclusion: hepatic histopathological lesions following controlled bleeding and intravenous volume replacement with RL or HES130/0.4 were subtle. However, more pronounced hydropic degeneration and hepatocellular steatosis was seen in G1 (RL), which suggests that HES130/04 may be associated with better hepatic perfusion when used for intravenous volume replacement when compared to RL.
  • A 2-Year Longitudinal Seroepidemiological Evaluation of Toxoplasma gondii Antibodies in a Cohort of Autochthonous Sheep from Central Portugal
    Publication . Almeida, Daniela; Quirino, João; Barradas, Patrícia Ferreira; da Silva, Priscilla Gomes; Pereira, Maria; Cruz, Rita; Santos, Carla; Mega, Cristina; Esteves, Fernando; Nóbrega, Carmen; Vala, Helena; Gärtner, Fátima; Amorim, Irina; Mesquita, João R.
    Background: Toxoplasma gondii is an important zoonosis and one of the major causes of abortion in sheep worldwide. (2) Methods: We performed a 2-year longitudinal serological antiT. gondii IgG screening on a cohort of a spatially confined population of a Portuguese autochthonous sheep breed in central Portugal. (3) Results: From the screening of the 2015 and 2016 sera, an increase of seroprevalence was observed (57.7% (95% CI: 49.9–65.3%) versus 69.1% (95% CI: 61.5–75.9), from 2015 and 2016, respectively) (p = 0.031). (4) Conclusions The present study is the first to provide prospective data on the anti-T. gondii serological status of a sheep cohort in Portugal, showing an increase in the occurrence of T. gondii. There is a need to provide a clearer understanding of T. gondii epidemiology in Portugal, ideally by implementing monitoring programs on sentinel herds, not only due to the high impact of T. gondii on animal health but also for it being a zoonosis.
  • A Questionnaire-Based Survey on the Long-Term Management of Canine Leishmaniosis by Veterinary Practitioners
    Publication . Pereira, Maria; Santos, Rute; Nóbrega, Carmen; Mega, Cristina; Cruz, Rita; Esteves, Fernando; Santos, Carla; Mesquita, João R.; Vala, Helena; Santos-Gomes, Gabriela
    Canine Leishmaniosis (CanL) is a chronic and potentially fatal disease. In economically disadvantaged regions, costs associated with long-term patient monitoring may determine that some owners decline veterinary follow-up of their dogs. This online, questionnaire-based survey aimed to assess how Portuguese veterinary practitioners perform long-term patient monitoring and recognize relapses. More than 50% of respondents reported that 50–100% of dog owners declared financial restraints. Hence, in these circumstances, most veterinary practitioners only performed clinical examination and serology. However, when owners did not declare financial restriction, other tests were additionally performed, such as renal and hepatic profiles, hemogram, serum protein electrophoresis and urine protein creatinine ratio. The mean number of exams performed when owners presented financial restraints was significantly lower than the number of exams performed without economic limitations. Most veterinary practitioners prescribed allopurinol ad aeternum oruntil disease remission and domperidone. CanL relapses were recognized by more than half of respondents “Always”, through the reappearance or worsening of clinical signs, whereas about a quarter detected an increase in anti-Leishmania antibody levels and identified abnormalities in the serum protein electrophoresis profile. The relapse rate was higher in the Lisbon Metropolitan Area and north, the most economically favored regions of Portugal. This study confirms that owner financial restraints negatively influence veterinary follow-up and relapse recognition, ultimately compromising clinical decision making and favoring the maintenance of Leishmania infantum infection endemicity.
  • Crimean-Congo hemorrhagic fever virus circulating among sheep of Portugal: a nationwide serosurvey assessment
    Publication . Mesquita, João; Cruz, Rita; Esteves, Fernando; Santos, Carla; Pousa, Humberto; Coelho, Catarina; Mega, Cristina; Nóbrega, Carmen; Vala, Helena; Peyrefitte, Christophe Nicolas; Nascimento, Maria São José; Barradas, Patrícia Ferreira
    Crimean-Congo hemorrhagic fever virus (CCHFV) is a widespread zoonotic pathogen that can cause mild to severe hemorrhagic disease in humans. CCHFV may be transmitted through direct contact with tissue or blood of viremic animals; however, the primary transmission route is through infected tick bites. CCHFV RNA has been detected in ticks feeding on domestic and wild animals in western Spain, suggesting an established circulation of CCHFV in Western Europe. Ruminants have been recognized as important CCHFV reservoirs and have been linked to human cases in endemic regions. Given the emergence of CCHF in neighboring Spain, and a report of two CCHFV seropositive humans in southern Portugal in 1985, we investigated the potential circulation of this virus in the country by performing a nationwide anti-CCHFV IgG serosurvey in sentinel sheep of Portugal. Sera (n = 459) randomly selected from widely distributed farms (n = 20) of Portugal were tested using a commercial double-antigen enzyme-linked immunosorbent assay, yielding an overall seroprevalence of 0.4% (95% confidence interval [CI] 0.04-1.56%). Positive sheep were from the southern region of Portugal (Alentejo region), which raise the seroprevalence of this region to 0.74% (95% CI 0.09-2.66%). This is the first study reporting the presence of CCHFV antibodies in sheep of Portugal, thus suggesting a geographical expansion of CCHFV to this country. It seems likely that CCHFV may exist focally in southern Portugal.
  • Prevalence of Toxoplasma gondii Antibodies in Individuals Occupationally Exposed to Livestock in Portugal
    Publication . Almeida, Daniela; Quirino, João; Matos, Pedro; Esteves, Fernando; Cruz, Rita; Vala, Helena; Mesquita, João R.
    Toxoplasmosis is a worldwide zoonotic disease with different and complex routes for transmission. Workers occupationally exposed to animals or raw meat and viscera (WOE) may be at more risk than the general population, however conflicting data exist on the risk of developing toxoplasmosis due to this close contact. To add knowledge to this topic, the aim of the present study was to ascertain if WOE were more likely to be anti-T. gondii IgG seropositive than the general population as well as to study risk factors for T. gondii infection such as professional activity, gender, age, years of work and region. For this purpose, a case–control study using archived samples was setup. A total of 114 WOE (including pig slaughterhouse workers, butchers, veterinarians and farmers) and 228 anonymous volunteers (matched with cases by region, age and gender) were studied for anti-T. gondii IgG. A significantly higher anti-T. gondii IgG occurrence (p = 0.0282) was found in WOE when compared with the general population (72.8% [CI = 64.6–81.0%] versus 60.1% [CI = 54.6–65.6%]). Multivariate analysis showed that WOE of more than 50 years of age were more likely to be seropositive for anti-T. gondii IgG (aOR = 16.8; 95% CI 3.6–77.5; p < 0.001) than those aged less than 50 years. To our knowledge, this is the first case–control study on the prevalence of antiT. gondii IgG in WOE in Portugal, also showing an added risk for T. gondii infection in those exposed to animals or their meat and viscera.
  • Rickettsia massiliae circulation in sheep and attached Rhipicephalus sanguineus in Central Portugal
    Publication . Mesquita, João R.; Santos-Silva, Sérgio; de Sousa Moreira, Alícia; Baptista, Maria Beatriz; Cruz, Rita; Esteves, Fernando; Vala, Helena; Barradas, Patrícia F.
    Rickettsiosis is considered an emerging/re-emerging vector-borne disease that causes signifcant public health threats. Ticks are reservoirs and vectors of Rickettsia having a signifcant role in the transmission of rickettsiae. In Portugal, little is known about tick-borne Rickettsia species in sheep. Therefore, this study aimed to investigate rickettsiae infection in ticks and their sheep host from 27 farms in four districts of central Portugal, to clarify the role of the sheep host in the circulation of this zoonotic agent. Between March and May 2021, EDTA blood samples (n=100) of healthy grazing sheep and their ticks (n=100, one tick per animal) were collected during a herd health program in central Portugal. Obtained ticks were identifed as Rhipicephalus sanguineus sensu lato by PCR targeting a partial sequence of 16S rRNA gene followed by sequence analysis. Rhipicephalus sanguineus s.l. and host sheep blood were tested for the presence of Rickettsia spp. by PCR targeting a partial sequence of ompB and ompA genes. From a total of 100 paired R. sanguineus s.l. and host sheep, Rickettsia massiliae was detected in 62 ticks and 35 grazing sheep blood samples, collected in central Portugal, 2021. All 35 positive sheep had attached positive R. sanguineus s.l., with matching nucleotidic sequences. These fndings suggest that sheep may develop rickettsiemia and are likely capable of transmitting and amplifying the infection to uninfected ticks maintaining rickettsiae in circulation in the domestic cycle.
  • Inspecção do sistema nervoso central em pequenos ruminantes
    Publication . Vala, Helena; Mesquita, João; Esteves, Fernando; Pinho, Fátima
    O Scrapie é uma doença neurodegenerativa fatal que afecta ovinos e caprinos e é a forma mais comum das Encefalopatias Espongiformes Transmissíveis (EET's), que incluem a doença de Creutzfeldt – Jakob (CJD), em humanos, e a Encefalopatia Espongiforme Bovina (BSE). O Plano de Vigilância Activa do Scrapie, em aplicação na União Europeia desde 2002, permitiu determinar uma prevalência de Scrapie superior ao esperado. De forma a efectuar uma avaliação preliminar da incidência desta patologia em Portugal, foram recolhidos os troncos cerebrais e cerebelos de 4822 pequenos ruminantes abatidos num matadouro no centro de Portugal e, posteriormente, enviados para pesquisa laboratorial de EETs. Após análise laboratorial, detectaram-se 8 casos de Scrapie atípico em ovinos. A percentagem de casos positivos (0,17%) detectados em pequenos ruminantes com mais de 18 meses, durante o período de realização deste estudo, foi baixa e, segundo os resultados laboratoriais, enquadram-se no perfil de Scrapie atípico.
  • A case of vascular tumour mimicking lymphangioma
    Publication . Vala, Helena; Mega, Cristina; Santos, C.; Cruz, Rita; Santos, M.; Esteves, Fernando; Nóbrega, Carmen; Mesquita, João
    Introduction Sarcomas, particularly vascular tumours, can mimic other neoplastic conditions or even inflammatory diseases. Objectives The authors describe an unusual case of vascular tumour, histologically overlapping with benign lymphangioma and liposarcoma. Medical History A 8 year old Shar Pei male dog was presented with a subcutaneous nodule in the chest, measuring 3x2 cm. Material and Methods The sample was fixed in 10 % buffered formalin solution and processed for routine histopathological diagnosis with Haematoxylin and Eosin. A standard Streptavidin-Biotin Complex technique for immunohistochemical evaluation was also performed, using a Polyclonal Rabbit Anti-Human Von Willebrand Factor. Results Macroscopic examination revealed a well circumscribed nodule with homogeneous and brownish appearance and moderate consistency on a cut surface. Microscopic examination revealed clefts and variably-sized vascular spaces, lined by a single layer of spindle cells, with round-to-oval nuclei, small or indistinct nucleoli, small amount of eosinophilic cytoplasm, with endothelial appearance – areas like hemangiosarcoma, interspersed with areas, where these cells appeared to grow directly on thick bundles of dermal collagen, occasionally with erythrocytes – areas like lymphangioma. The stroma was also interspersed with vacuolar cells – areas like liposarcoma. No mitotic figures were seen, however, the presence of very rich cellular areas, with more prominent cells revealing pleomorphism and anisocariosis, were more favour to malignancy Conclusion The authors suggested the diagnosis of vascular neoplasia with unusual features, based on the histopathological diagnosis and immunohistochemistry results, and submit to the discussion, especially for assessment of histological grade and prognosis.
  • Pulmonary and Gastrointestinal Parasitic Infections in Small Ruminant Autochthonous Breeds from Centre Region of Portugal—A Cross Sectional Study
    Publication . Pereira, Maria; Vila-Viçosa, Maria; Coelho, Catarina; Santos, Carla; Esteves, Fernando; Cruz, Rita; Gomes, Liliana; Henriques, Diogo; Vala, Helena; Nóbrega, Carmen; Mega, Cristina; Melo, Carolina de; Malva, Madalena; Braguez, Joana; Mateus, Teresa Letra
    The production of small ruminant autochthonous breeds in the Centre region of Portugal is practiced in a semi-extensive husbandry system, exposing animals to parasitic infections. The main objective of this study was to estimate the prevalence of lungworm infection and identify risk factors. Fecal samples of 203 goats and 208 sheep from 30 herds were collected per rectum and subjected to the modified Baermann test. The overall prevalence of infection was 57.7%, significantly higher in goats (95.6%) than in sheep (20.7%) (p < 0.001). According to the binary logistic regression model, sheep dewormed with albendazole, mebendazole plus closantel, or ivermectin plus clorsulon presented a risk of Protostrongylidae infection 29.702, 7.426, or 8.720 times higher, respectively, than those dewormed with eprinomectin. Additionally, the presence of gastrointestinal parasites was investigated in 307 fecal samples using Mini-FLOTAC®. The overall prevalence of infection was 86.3%, also significantly higher in goats (93.2%) than in sheep (79.9%) (p < 0.001). Strongyle-type eggs were the most frequently identified, both in sheep (69.8%) and goats (87.8%), followed by Eimeria oocysts (40.3% in sheep and 68.9% in goats). Considering the high prevalence and the burden of lungworm parasitic infection, it is urgent to determine its economic impact and the repercussions in animal health in the Centre region of Portugal to establish appropriate therapeutic guidelines.