Molecular evidence of sporadic Coxiella burnetii excretion in sheep milk, central Portugal.

Coxiella burnetii is the etiologic agent of Q fever, a worldwide zoonosis. Cattle, sheep and goats are considered the main reservoirs of the disease. Transmission to humans occurs mainly through the inhalation of infectious aerosols from milk, faeces, urine, and birth products from infected ruminants. In this study, a 2-year longitudinal approach was performed to ascertain the excretion of C. burnetii in bulk tank milk samples of sheep from a mountain plateau in central Portugal, with sampling conducted during the years 2015 and 2016. From a total of 156 bulk tank milk samples tested by qPCR, only one showed to be positive for C. burnetii (1.28% [95%CI: 0.03-6.94]), from 2015, the first year of collection. Bidirectional sequencing and phylogenetic analysis of IS1111 transposase partial region confirmed the presence of C. burnetii DNA. The presence of C. burnetii in raw milk samples highlights the necessity for additional research to determine if raw milk is a potential source for human infection. Animal health surveillance and prevention measures against this zoonotic disease should be considered.

Pulmonary and Gastrointestinal Parasitic Infections in Small Ruminant Autochthonous Breeds from Centre Region of Portugal-A Cross Sectional Study.

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.

Quercus spp. extract as a promising preventive or therapeutic strategy for cancer: A systematic review.

Acorns have traditionally been used in the human diet and for the treatment of specific diseases. Therefore, the present study performed a systematic review of studies which investigated the effects of Quercus spp. extracts in cancer prevention and treatment. A systematic literature search was performed for original records which addressed the anticancer effects of Quercus spp. extract in in vitro and in vivo cancer models. Body composition, food consumption, tumor development and/or toxicity were evaluated in in vivo studies, while cytotoxicity was evaluated in in vitro studies. Few studies and low sample sizes presented a challenge in the drawing of solid conclusions. Overall, the results suggested a positive impact of Quercus spp. extract, by reducing cancer development. Therefore, more studies with different cancer cell lines and animal models to address the efficacy of the acorn extracts in several types of cancer are required. Furthermore, the effects of acorn flour, incorporated in the diet, in an animal model of mammary cancer should be evaluated.

Functional recovery of injured cavernous nerves achieved through endogenous nerve growth factor-containing bioactive fibrous membrane.

Radical prostatectomy is a highly successful treatment for prostate cancer, among the most prevalent manifestations of the illness. Damage of the cavernous nerve (CN) during prostatectomy is the main cause of postoperative erectile dysfunction (ED). In this study, the capability of a personalized bioactive fibrous membrane to regenerate injured CN was investigated. The fibrous membrane bioactivity is conferred by the selectively bound nerve growth factor (NGF) present in the rat urine. In a rat model of bilateral CN crush, the implanted bioactive fibrous membrane induces CN regeneration and restoration of erectile function, showing a significantly increased number of smooth muscle cells and content of endothelial and neuronal nitric oxide synthases (eNOS; nNOS). In addition, the bioactive fibrous membrane promotes nerve regeneration by increasing the number of myelinated axons and nNOS-positive cells, therefore reversing the CN fibrosis found in untreated rats or rats treated with a bare fibrous membrane. Therefore, this personalized regenerative strategy could overcome the recognized drawbacks of currently available treatments for CN injuries. It may constitute an effective treatment for prostate cancer patients suffering from ED after being subject to radical prostatectomy. STATEMENT OF SIGNIFICANCE: The present work introduces a unique strategy to address post-surgical ED resulting from CN injury during pelvic surgery (e.g., radical prostatectomy, radical cystoprostatectomy, abdominoperineal resection). It comprises a bioactive and cell-free fibrous implant, customized to enhance CN recovery. Pre-clinical results in a rat model of bilateral CN crush demonstrated that the bioactive fibrous implant can effectively heal injured CN, and restore penile structure and function. This implant selectively binds NGF from patient fluids (i.e. urine) due to its functionalized surface and high surface area. Moreover, its local implantation reduces adverse side effects. This tailored regenerative approach has the potential to revolutionize the treatment of ED in prostate cancer patients following radical prostatectomy, overcoming current treatment limitations.

An Antibody-Based Survey of Toxoplasma gondii and Neospora caninum Infection in Client-Owned Cats from Portugal.

Toxoplasma gondii and Neospora caninum are obligate intracellular protozoan parasites infecting a wide range of hosts worldwide. However, information on the epidemiology of toxoplasmosis and neosporosis in cats from Portugal is limited. Thus, this study aims to evaluate anti-T. gondii and anti-N. caninum seroprevalence in client-owned cats from Portugal and to identify risk factors using a panel of well-characterized sera. A total of 183 domestic cats were sampled and screened for antibodies against T. gondii and N. caninum using commercial ELISA assays, and their owners answered an online questionnaire designed to obtain background information. The overall anti-T. gondii and anti-N. caninum seroprevalences were 13.1% (CI: 8.97-18.77) and 3.8% (CI: 1.87-7.68), respectively. Univariate analysis revealed that living strictly indoors was a significant protection factor (cOR: 0.053; CI: 0.005-0.627), and the presence of a chronic disease a significant risk factor (cOR: 3.106; CI: 1.062-9.082) to T. gondii seroprevalence. When performing multivariate analysis, only chronic disease (aOR: 57.527; CI: 1.7-1976.7) and seropositivity to N. caninum (aOR: 7.929; CI:0.8-82.9) were found to be a significant risk factor to anti-T. gondii antibodies. To the best of our knowledge, this is the first report of N. caninum seropositivity in cats from Portugal.

Study on the antineoplastic and toxicological effects of pomegranate (Punica granatum L.) leaf infusion using the K14-HPV16 transgenic mouse model.

Punica granatum L. (pomegranate) has been used in functional foods due to its various health benefits. However, the in vivo biological potential of its leaf remains little known. This study has aimed to characterize the antineoplastic and toxicological properties of using pomegranate leaf infusion (PLI) on transgenic mice carrying human papillomavirus (HPV) type 16 oncogenes. Thirty-eight mice were divided into 3 wild-type (WT) and 3 transgenic (HPV) groups, with exposure to 0.5% PLI, 1.0% PLI, and water. The animals' body weight, drink and food consumption were recorded. Internal organs, skin samples and intracardiac blood were collected to evaluate toxicological parameters, neoplastic lesions and oxidative stress. The results indicated that PLI was safe as no mortality, no behavioural disorders and no significant differences in the levels of microhematocrit, serum biochemical markers, internal organ histology, and oxidative stress was found among the WT groups. Histological analysis revealed that HPV animals that consumed PLI exhibited reduced hepatic, renal and cutaneous lesions compared with the HPV control group. Low-dose PLI consumption significantly diminished renal hydronephrosis lesions and relieved dysplasia and carcinoma lesions in the chest skin. Oxidative stress analysis showed that low-dose PLI consumption may have more benefits than high-dose PLI. These results suggest that oral administration of PLI has the potential to alleviate non-neoplastic and neoplastic lesions against HPV16-induced organ and skin injuries, though this requires further scientific research studies.

TNFR2 as a Potential Biomarker for Early Detection and Progression of CKD.

The inflammatory pathway driven by TNF-α, through its receptors TNFR1 and TNFR2, is a common feature in the pathogenesis of chronic kidney disease (CKD), regardless of the initial disease cause. Evidence correlates the chronic inflammatory status with decreased renal function. Our aim was to evaluate the potential of TNF receptors as biomarkers for CKD diagnosis and staging, as well as their association with the progression of renal lesions, in rat models of early and moderate CKD. We analyzed the circulating levels of inflammatory molecules-tumor necrosis factor-alpha (TNF-α), tumor necrosis factor receptor 1 (TNFR1) and 2 (TNFR2) and tissue inhibitor of metalloproteinase-1 (TIMP-1)-and studied their associations with TNFR1 and TNFR2 renal expression, glomerular and tubulointerstitial lesions, and with biomarkers of renal (dys)function. An increase in all inflammatory markers was observed in moderate CKD, as compared to controls, but only circulating levels of both TNFR1 and TNFR2 were significantly increased in the early disease; TNFR2 serum levels were negatively correlated with eGFR. However, only TNFR2 renal expression increased with CKD severity and showed correlations with the score of mild and advanced tubular lesions. Our findings suggest that renal TNFR2 plays a role in CKD development, and has potential to be used as a biomarker for the early detection and progression of the disease. Still, the potential value of this biomarker in disease progression warrants further investigation.

Prevalence of Toxoplasma gondii Antibodies and Risk Factor Investigation in Portuguese Veterinarians: A Matched Case-Control Study.

(1) Background: Toxoplasma gondii is a widespread zoonotic agent that greatly impacts Public Health, being responsible for one of the most important parasitic zoonosis worldwide. T. gondii has a heteroxenous life cycle, with cats being the definitive hosts and all warm-blooded animals, including humans, being intermediate hosts. Veterinary practitioners (VP) may be at a higher risk than the general population for T. gondii infection, as they have direct and daily contact with many animal species. The aim of the present study was to ascertain if VP were more likely to be anti-T. gondii IgG seropositive than the general population, as well as to understand if age, accidents with blood-contaminated sharps (cross-blood contamination), gender, working years, and geographic regions play a role as risk factors for T. gondii infection. For this purpose, a case−control study using archived samples was performed. (2) Methods: A total of 350 veterinary practitioners were tested using a commercial semiquantitative enzyme immunoassay for anti-T. gondii IgG. From the general population, 175 anonymous volunteers (matched with cases by region, age, and gender) were studied for anti-T. gondii IgG. (3) Results: There was no statistical difference found between the presence of anti-T. gondii IgG in practitioners (26%; CI = 21.40−30.60%) and the general population (33.14%; CI = 26.17−40.12%) (p = 0.108). Univariate and multivariate analysis showed that only age (older groups) was found to be associated with a higher prevalence of anti-T. gondii IgG, with significant p values (p < 0.05) for both univariate and multivariate analysis. (4) Conclusions: To the best of our knowledge, this is the first case−control study fully focused on the prevalence of anti-T. gondii IgG in VP in Portugal, showing that there was no significant risk for T. gondii infection in veterinarians exposed daily and repeatedly to different species of animals.

Seroprevalence of SARS-CoV-2 in Client-Owned Cats from Portugal.

The close contact between humans and domestic cats raises concerns about the potential risks of SARS-CoV-2 transmission. Thus, this study aims to investigate anti-SARS-CoV-2 seroprevalence in client-owned cats from Portugal and evaluate the infection risk of cats that maintain contact with human COVID-19 cases. A total of 176 cats, belonging to 94 households, were sampled. Cat owners answered an online questionnaire, and cats were screened for antibodies against SARS-CoV-2 using a commercial ELISA. Twenty (21.3%) households reported at least one confirmed human COVID-19 case. Forty cats (22.7%) belonged to a COVID-19-positive and 136 (77.3%) to a COVID-19-negative household. The seroprevalences of cats from COVID-19-positive and -negative households were 5.0% (2/40) and 0.7% (1/136). The two SARS-CoV-2-seropositive cats from COVID-19-positive households had an indoor lifestyle, and their owners stated that they maintained a close and frequent contact with them, even after being diagnosed with COVID-19, pointing towards human-to-cat transmission. The SARS-CoV-2-seropositive cat from the COVID-19-negative household had a mixed indoor/outdoor lifestyle and chronic diseases. Owners of the three SARS-CoV-2-seropositive cats did not notice clinical signs or behavior changes. This study highlights the low risk of SARS-CoV-2 transmission from COVID-19-positive human household members to domestic cats, even in a context of close and frequent human-animal contact.

Crimean-Congo hemorrhagic fever virus circulating among sheep of Portugal: a nationwide serosurvey assessment.

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.

Rickettsia massiliae circulation in sheep and attached Rhipicephalus sanguineus in Central Portugal.

Rickettsiosis is considered an emerging/re-emerging vector-borne disease that causes significant public health threats. Ticks are reservoirs and vectors of Rickettsia having a significant 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 identified 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 findings 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.

Prevalence of Toxoplasma gondii Antibodies in Individuals Occupationally Exposed to Livestock in Portugal.

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 anti-T. 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.

A Questionnaire-Based Survey on the Long-Term Management of Canine Leishmaniosis by Veterinary Practitioners.

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 or until 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.

Occurrence and Distribution of Fasciolosis in a Cohort of Ovine Livestock Restricted to a Mountain Plateau in Central Portugal.

Fasciola hepatica is a parasite that is widespread in Europe, having been reported in ruminants of several European countries and causing an important economic impact. This study ascertained the occurrence and distribution of fasciolosis in Portuguese ovine livestock by assessing F. hepatica IgG antibodies in a cohort of confined sheep from a high-altitude region of central Portugal in a 2-year period. Positive animals were found in most locations and in both years, with 18 of the 92 animals (19.6% [95% confidence interval CI: 12.03-19.15]) and 17 of the same 92 animals (18.5% [95% CI: 11.15-27.93]) showing to be seropositive in the first year and second year, respectively (p = 0.85). Pasture contamination by F. hepatica eggs could be reduced by thorough anthelmintic treatments.

A 2-Year Longitudinal Seroepidemiological Evaluation of Toxoplasma gondii Antibodies in a Cohort of Autochthonous Sheep from Central Portugal.

(1) 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 anti-T. 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.

Prognostic Factors and Life Expectancy in Canine Leishmaniosis.

Canine leishmaniosis (CanL) is a chronic and potentially fatal disease. The prognosis of CanL depends on the severity of the clinical signs and clinicopathological abnormalities presented by the dog at the time of diagnosis. This study aims to estimate the survival time of dogs with CanL, determining the prognostic value of different clinical and clinicopathological parameters. Medical records of 99 dogs diagnosed with CanL in five veterinary centers of the Alentejo region (Portugal) were examined retrospectively. The majority of dogs presented hyperproteinemia, moderate normocytic normochromic anemia, normal blood urea and creatinine levels and were classified as stage 1 according to the International Interest Society (IRIS) guidelines at the time of diagnosis. The severity of anemia, presence of concomitant infectious diseases at the time of diagnosis and the anti-Leishmania therapy were correlated with the survival time. The influence of renal dysfunction was evaluated by Receiver Operating Characteristic (ROC) curve and survival analysis. Survival analysis demonstrated that patients classified as IRIS 1 at the time of diagnosis survived more than four years, in contrast with dogs classified as IRIS 2 that survived around two and half years and dogs classified as IRIS 3-4 that survived around one month. IRIS stage deteriorated during the course of CanL in one third of the dogs and was the principal cause of death or euthanasia in a high proportion of animals. In some cases, dogs did not receive anti-Leishmania treatment or abandoned the veterinary follow-ups, which may have considerable repercussions for animal wellbeing and public health. This study reinforces the value of blood urea and creatinine levels as prognostic factors in CanL.

Occupationally Acquired Q Fever in Shepherds and Sheep Milk Cheesemakers.

Q fever is a zoonosis caused by Coxiella burnetii, and transmission to humans is often associated with contact with ovine and caprine livestock. Those exposed to sheep are particularly at high risk of infection. Recent studies show that Q fever is increasing in sheep farms in Portugal raising alerts on spillover to humans. We detected anti-C. burnetii IgG in shepherds and sheep milk cheesemakers (27 [28.1%] in a total of 96; 95% confidence interval [CI] 19.4-38.2%) and in controls (21 [8.1%] in a total of 260; 95% CI 5.1-12.1%), pointing to an increased risk of C. burnetii infection (P = 0.0001), with an odds ratio for anti-C. burnetii of 4.45 (95% CI 2.4-8.4%; P = 0.0001), in individuals with occupational contact with sheep in Portugal.

Immunohistochemical Studies of Cytokeratins and Differentiation Markers in Bovine Ocular Squamous Cell Carcinoma.

Bovine Ocular Squamous Cell Carcinoma is considered the most common bovine tumour, causing significant economic losses, mainly by abattoir condemnations. To obtain a better insight into the genesis and neoplastic transformation, 19 samples collected at slaughter from Holstein Friesian cattle and diagnosed as Ocular Squamous Cell Carcinoma were studied. Tumours were histologically classified into three categories: poorly (26.3%), moderately (26.3%), and well differentiated (47.4%). Expression of keratins (MNF116 and LP34) and of cornified envelope precursors (involucrin and profilaggrin) was studied. Expression of MNF116 was observed in all carcinomas. LP34 immunostaining was seen in all but three carcinomas, one from each degree. Involucrin immunoreaction was observed in all but one poorly differentiated carcinoma. Profilaggrin was present in only two moderately differentiated carcinomas, in all but one well differentiated, and in all but one poorly differentiated. MNF116 is a useful marker to confirm the epithelial origin of the tumour and stain most neoplastic cells in these tumours. The expression of involucrin and LP34 demonstrates that, in all tumours, cells have reached the final program of differentiation, regardless of the grade. The expression of profilaggrin could indicate molecular changes during malignant transformation but their expression does not seem to be of diagnostic value.

Increased risk of hepatitis E virus infection in workers occupationally exposed to sheep.

Hepatitis E virus (HEV) is an enteric RNA virus from the family Hepeviridae with five genotypes (genotypes 1-4 and 7) known to infect humans. HEV infection is known to have a zoonotic swine origin in industrialized countries. The role of pigs and wild boars as major reservoirs for human infection is today well-established; however, the list of new animal reservoirs is ever-expanding as new HEV strains are continuously being found in a broad host range. The recent detection of HEV in sheep stools brings concerns on the possibility of HEV transmission from these animals to humans, particularly in those occupationally exposed. The present work investigated the potential occupational risk of HEV infection in shepherds and sheep milk cheesemakers-workers occupationally exposed to ovine (WOEOs; N = 96)-from a region of the Centre of Portugal ('Serra da Estrela') based on the differences of anti-HEV IgG seroprevalence rates between these professionals and the general population (N = 192). The presence of HEV-specific antibodies in sheep (N = 90) from the same region was also evaluated. The HEV seroprevalence in WOEOs (29.3%) was found to be significantly higher (p = .0198) when compared with population controls (16.1%) which suggests an increased risk for HEV infection in these workers. HEV-specific antibodies were also found in 16.6% of the studied sheep showing that HEV circulates in these animals. Further studies are needed to confirm the zoonotic potential of sheep HEV.

Mortality and Morbidity of Beef Calves in Free-Range Farms in Alentejo, Portugal-A Preliminary Study.

Extensive cow-calf beef cattle farms play a significant role in Portuguese livestock production, but records of important production variables, such as calf mortality, are scarce. To assess herd-level calf mortality and its potential economic impact, farmers from the Alentejo region were asked to fill a questionnaire regarding herd size, feeding and reproductive management practices, calf mortality (from birth to weaning), and detection of the main morbidity causes, as well as of sudden, unexplained deaths during the previous 12 months. Farmers were also requested to quantitatively assess the economic impact of calf mortality on their annual revenue. In the majority of farms, the herd size was larger than 100 animals. The median stocking rate was 0.41 adult animals/hectare, and 70% of farmers stated their farm was self-sufficient in forage. The percent of live births that resulted in weaned calves averaged 94.3%, which led to the assumption of a 5.7% calf mortality rate from birth to weaning. In the previous 12 months, 78.8% of the farms identified at least one case of calf diarrhea, 60.7% identified at least one case of respiratory disease, and 82.1% had at least one sudden, unexplained calf death. As expected, farmers that assessed a higher impact of calf losses on their annual economic revenue were also those who reported higher incidences of calf diarrhea, respiratory disease, and sudden, unexplained deaths. One-quarter of the farmers were unable to assess the economic impact of calf mortality on the farm's revenue. Herd size appears to have a predictive value on calf mortality in the first month of life, and from 30 days to weaning. The number of sudden, unexplained calf deaths seems to have a predictive value on overall calf mortality (from birth to weaning), suggesting that the farms' management practices probably play an important role in calf mortality throughout the suckling period. Further studies are needed to fully understand calf mortality in Portuguese extensive rearing systems.