Spatial distribution of canine Leishmania infantum infection in a municipality with endemic human leishmaniasis in Eastern Bahia, Brazil.

Efforts to control a zoonotic disease such as visceral leishmaniasis (VL) caused by Leishmania infantum can be successful if they rely on comprehensive data on animal infection. In Bahia state, Brazil, human VL is endemic, yet some areas have no epidemiological data on canine L. infantum infection and canine leishmaniasis (CanL) to date. We aimed to perform an epidemiological study describing the spatial distribution and characterizing canine L. infantum infection in two districts of the municipality of Muritiba, where human cases have occurred. Brazilian official serodiagnostic protocol (ELISA and immunochromatographic tests), PCR and clinical examination were performed in 351 owned dogs. A seroprevalence of 15.7% (55/351) was found, and L. infantum identified in 88.8% (32/36) of PCR tested samples. Spatial distribution of positive dogs indicated infection in both urban and rural districts. There was no association between seropositivity and sex or breed, but dogs older than 2 years were 3.8 times more likely to be seropositive (95% CI 1.57 - 9.18) than younger dogs. Among seropositive dogs, 80% (44/55) had clinical manifestations of CanL: 75% (33/44) presented dermatopathy, 50% (22/44) emaciation, and 29.5% (13/44) ophthalmopathy. This is the first report on canine seroprevalence and natural L. infantum infection in Muritiba, Bahia.

Risk factors and detection of brucella ovis in naturally infected rams and ewes / Fatores de risco e detecção de brucella ovis em carneiros e ovelhas naturalmente infectados

The objective of this study was to verify the occurrence of ovine brucellosis using Agar Gel Immunodiffusion (AGID) and Polymerase Chain Reaction (PCR) techniques, as well as to identify the main risk factors associated with infection in sheep flocks belonging to municipalities in the microregion from Teresina, PI, Brazil. A total of 100 urine and blood samples were collected from sheep aged 6 months or older. The urine samples were submitted to conventional PCR and the blood samples were examined by the AGID technique. Of the 100 blood samples, 17 (17%) were reactive to the AGID test. In conventional PCR of 100 urine samples, six (6%) were positive. Risk factors associated to infection by B. ovis included the rearing system (OR=0.19), feed management (OR=0.05), presence of dystotic births (OR=4.50), miscarriages (OR=3.75) and source of water offered to the animals (OR=0.19). Thus, it was concluded that it is possible to detect the occurrence of animals with ovine brucellosis since PCR is a reliable method to confirm infection. Furthermore, there are risk factors associated to infection by B. ovis in the municipalities studied.

Objetivou-se verificar a ocorrência da brucelose ovina através das técnicas de Imunodifusão em Gel de Ágar (IDGA) e Reação em Cadeia da Polimerase (PCR), bem como identificar os principais fatores de risco associados à infecção nos rebanhos ovinos pertencentes a municípios da microrregião de Teresina, PI, Brasil. Foram colhidas 100 amostras de urina e de sangue de ovinos com idade superior ou igual a seis meses. As amostras de urina foram submetidas a PCR convencional e as amostras de sangue à técnica de IDGA. Das 100 amostras de sangue 17 (17%) foram reagentes ao teste de IDGA. Já na PCR convencional das 100 amostras de urina, seis (6%) foram positivas. Ressalta-se que três animais foram positivos em ambos os testes. Como fatores associados à infecção por B. ovis, observou-se o tipo de sistema de criação (OR=0,19), o manejo alimentar (OR=0,05), presença de partos distócicos (OR=4,50), abortamentos (OR=3,75) e a fonte de água fornecida aos animais (OR=0,19). Assim, conclui-se que foi possível detectar a ocorrência de animais com brucelose ovina, uma vez que a PCR é um método confirmatório. Além disso, há fatores de risco associados à infecção por B. ovis nos municípios estudados.

Risk factors and detection of brucella ovis in naturally infected rams and ewes / Fatores de risco e detecção de brucella ovis em carneiros e ovelhas naturalmente infectados

The objective of this study was to verify the occurrence of ovine brucellosis using Agar Gel Immunodiffusion (AGID) and Polymerase Chain Reaction (PCR) techniques, as well as to identify the main risk factors associated with infection in sheep flocks belonging to municipalities in the microregion from Teresina, PI, Brazil. A total of 100 urine and blood samples were collected from sheep aged 6 months or older. The urine samples were submitted to conventional PCR and the blood samples were examined by the AGID technique. Of the 100 blood samples, 17 (17%) were reactive to the AGID test. In conventional PCR of 100 urine samples, six (6%) were positive. Risk factors associated to infection by B. ovis included the rearing system (OR=0.19), feed management (OR=0.05), presence of dystotic births (OR=4.50), miscarriages (OR=3.75) and source of water offered to the animals (OR=0.19). Thus, it was concluded that it is possible to detect the occurrence of animals with ovine brucellosis since PCR is a reliable method to confirm infection. Furthermore, there are risk factors associated to infection by B. ovis in the municipalities studied.

Objetivou-se verificar a ocorrência da brucelose ovina através das técnicas de Imunodifusão em Gel de Ágar (IDGA) e Reação em Cadeia da Polimerase (PCR), bem como identificar os principais fatores de risco associados à infecção nos rebanhos ovinos pertencentes a municípios da microrregião de Teresina, PI, Brasil. Foram colhidas 100 amostras de urina e de sangue de ovinos com idade superior ou igual a seis meses. As amostras de urina foram submetidas a PCR convencional e as amostras de sangue à técnica de IDGA. Das 100 amostras de sangue 17 (17%) foram reagentes ao teste de IDGA. Já na PCR convencional das 100 amostras de urina, seis (6%) foram positivas. Ressalta-se que três animais foram positivos em ambos os testes. Como fatores associados à infecção por B. ovis, observou-se o tipo de sistema de criação (OR=0,19), o manejo alimentar (OR=0,05), presença de partos distócicos (OR=4,50), abortamentos (OR=3,75) e a fonte de água fornecida aos animais (OR=0,19). Assim, conclui-se que foi possível detectar a ocorrência de animais com brucelose ovina, uma vez que a PCR é um método confirmatório. Além disso, há fatores de risco associados à infecção por B. ovis nos municípios estudados.

Influence of Polymorphism of Vitamin D Receptor (Fok I) on Hypertension

Abstract Evidence suggests that polymorphisms in the gene encoding a vitamin D receptor might affect blood pressure. The objective of this systematic review was to investigate the association between hypertension and vitamin D receptor (Fok I) gene polymorphism. A literature search was performed according to the PRISMA guidelines using the MEDLINE®/PubMed, Scopus, Cochrane Library CENTRAL, SciELO, and LILACS databases. The quality of case-control or cohort studies and studies based on cross-sectional methodology was evaluated using the Newcastle-Ottawa Scale and the protocol of Loney and coauthors [25], respectively. In this systematic literature search, 215 publications were identified, of which 10 were analyzed, including seven case-control studies, two cross-sectional studies, and one cohort study. The association between Fok I polymorphism and hypertension was reported in 60% of the publications and the risk for hypertension was shown to be related to FF and ff genotypes. In addition, Fok I polymorphism was shown to increase plasma renin activity, which plays an important role in regulating blood pressure. However, no association was observed between Fok I polymorphism and serum vitamin D levels. In conclusion, Fok I polymorphism plays an important role in hypertension.

Association between Insulin-Like Growth Factor-I Levels and the Disease Progression and Anemia in Visceral Leishmaniasis.

We analyzed the association between insulin-like growth factor-I (IGF-I) and the pathogenesis of anemia during active visceral leishmaniasis (VL). Serum levels of IGF-I, IGF-binding protein 3 (IGFBP3), and cytokines were measured in samples from individuals with active VL and cured VL, asymptomatic Leishmania-infected, and noninfected individuals. Then, we extended our analysis to VL dogs to evaluate hematimetric parameters, bone marrow alterations, and cytokine and IGF-I expression. We identified a positive correlation between lower IGF-I and IGFBP3 levels in active VL patients and lower hemoglobin levels. In infected dogs, there was a positive correlation between lower IGF-I expression in the bone marrow and lower peripheral blood hematocrit and hemoglobin levels. There was no correlation between decreased IGF-I level/expression and any measured cytokine serum levels in either host. The data suggest that low IGF-I expression is associated with pathogenesis of anemia in active VL, primarily in severe cases, by mechanisms other than alterations in cytokine production.

Isolation of naturally infecting Leishmania infantum from canine samples in Novy-MacNeal-Nicolle medium prepared with defibrinated blood from different animal species.

The most commonly used culture medium for the in vitro isolation of Leishmania spp. from canine biological samples is biphasic Novy-MacNeal-Nicolle (NNN) medium, whose solid phase is prepared using rabbit blood. Leishmania infantum parasites from natural infections are highly sensitive and demanding for growth in axenic conditions when firstly obtained from the dog's body. The objective of this study was to evaluate whether NNN medium (NNN-test) prepared with chicken blood (NNN-C), ox blood (NNN-O), horse blood (NNN-H) or sheep blood (NNN-S) was viable for the isolation of parasites from naturally infected dogs, in an endemic area for visceral leishmaniasis caused by L. infantum. Spleen aspirates from six dogs previously diagnosed as infected by parasitological methods were simultaneously inoculated in each NNN-test medium, including the conventional medium prepared with rabbit blood (NNN-R), and the cultures were examined for three weeks under optic microscopy. Spleen samples were also analyzed for parasite loads by quantitative PCR (qPCR). Cultures from three of the six dogs (50%) were positive in at least one of the NNN-test media: one sample presented the highest spleen parasite load by qPCR (1.19 × 104 parasites/mL) and was positive in all test media; the second sample presented parasitic isolation in the first week of culture in all inoculated media, of which the NNN-C medium had the highest mean parasite count (NNN-C = 23.5 × 104/mL vs. NNN-R = 3.25 × 104/mL); the third sample was positive only in the NNN-S medium besides the conventional control NNN-R. Cultures from the three remaining dogs were negative in all NNN media, including the control and test media; of those three dogs, two presented the lowest spleen parasitic loads according to qPCR. Blood from chicken, ox, horse and sheep shown to be viable for the preparation of NNN culture medium for the primary isolation of L. infantum from samples of naturally infected dogs and can be considered as an alternative to rabbit blood when necessary.

Scoring clinical signs can help diagnose canine visceral leishmaniasis in a highly endemic area in Brazil.

Canine visceral leishmaniasis (CVL) diagnosis is still a challenge in endemic areas with limited diagnostic resources. This study proposes a score with the potential to distinguish positive CVL cases from negative ones. We studied 265 dogs that tested positive for CVL on ELISA and parasitological tests. A score ranging between 0 and 19 was recorded on the basis of clinical signs. Dogs with CVL had an overall higher positivity of the majority of clinical signs than did dogs without CVL or with ehrlichiosis. Clinical signs such as enlarged lymph nodes (83.93%), muzzle/ear lesions (55.36%), nutritional status (51.79%), bristle condition (57.14%), pale mucosal colour (48.21%), onychogryphosis (58.93%), skin lesion (39.28%), bleeding (12.50%), muzzle depigmentation (41.07%), alopecia (39.29%), blepharitis (21.43%), and keratoconjunctivitis (42.86%) were more frequent in dogs with CVL than in dogs with ehrlichiosis or without CVL. Moreover, the clinical score increased according to the positivity of all diagnostic tests (ELISA, p < 0.001; parasite culture, p = 0.0021; and smear, p = 0.0003). Onychogryphosis (long nails) [odds ratio (OR): 3.529; 95% confidence interval (CI): 1.832-6.796; p < 0.001], muzzle depigmentation (OR: 4.651; 95% CI: 2.218-9.750; p < 0.001), and keratoconjunctivitis (OR: 5.400; 95% CI: 2.549-11.441; p < 0.001) were highly associated with CVL. Interestingly, a score cut-off value ≥ 6 had an area under the curve of 0.717 (p < 0.0001), sensitivity of 60.71%, and specificity of 73.64% for CVL diagnosis. The clinical sign-based score for CVL diagnosis suggested herein can help veterinarians reliably identify dogs with CVL in endemic areas with limited diagnostic resources.

Scoring clinical signs can help diagnose canine visceral leishmaniasis in a highly endemic area in Brazil

Canine visceral leishmaniasis (CVL) diagnosis is still a challenge in endemic areas with limited diagnostic resources. This study proposes a score with the potential to distinguish positive CVL cases from negative ones. We studied 265 dogs that tested positive for CVL on ELISA and parasitological tests. A score ranging between 0 and 19 was recorded on the basis of clinical signs. Dogs with CVL had an overall higher positivity of the majority of clinical signs than did dogs without CVL or with ehrlichiosis. Clinical signs such as enlarged lymph nodes (83.93%), muzzle/ear lesions (55.36%), nutritional status (51.79%), bristle condition (57.14%), pale mucosal colour (48.21%), onychogryphosis (58.93%), skin lesion (39.28%), bleeding (12.50%), muzzle depigmentation (41.07%), alopecia (39.29%), blepharitis (21.43%), and keratoconjunctivitis (42.86%) were more frequent in dogs with CVL than in dogs with ehrlichiosis or without CVL. Moreover, the clinical score increased according to the positivity of all diagnostic tests (ELISA, p < 0.001; parasite culture, p = 0.0021; and smear, p = 0.0003). Onychogryphosis (long nails) [odds ratio (OR): 3.529; 95% confidence interval (CI): 1.832-6.796; p < 0.001], muzzle depigmentation (OR: 4.651; 95% CI: 2.218-9.750; p < 0.001), and keratoconjunctivitis (OR: 5.400; 95% CI: 2.549-11.441; p < 0.001) were highly associated with CVL. Interestingly, a score cut-off value ≥ 6 had an area under the curve of 0.717 (p < 0.0001), sensitivity of 60.71%, and specificity of 73.64% for CVL diagnosis. The clinical sign-based score for CVL diagnosis suggested herein can help veterinarians reliably identify dogs with CVL in endemic areas with limited diagnostic resources.

Classificação das alterações pulmonares na leishmaniose visceral canina / Classification of pulmonary alterations in canine visceral leishmaniasis

O objetivo deste estudo foi identificar e classificar as alterações pulmonares de cães naturalmente infectados por Leishmania (Leishmania) infantum conforme as manifestações clínicas. Após o diagnóstico para LV pelo exame parasitológico, 24 cães infectados foram subdivididos em dois grupos: 10 cães com até quatro sinais clínicos; e 14 cães com cinco ou mais sinais clínicos. Como controle foram avaliados cinco cães sadios. Amostras de tecido pulmonar foram coradas com hematoxilina-eosina (H-E) e Tricrômico de Masson para avaliação histopatológica, e prata-metenamina de Grocott para excluir infecções fúngicas, bem como imunoperoxidase para detecção de Leishmania. A análise microscópica do pulmão revelou espessamento focal do septo interalveolar devido à congestão, edema e infiltrado celular. Além disso, presença de fibrose, exsudato bronquiolar e ainda descamação do epitélio bronquiolar com perda de cílios, presença de hemossiderina e hiperplasia de células caliciformes. O padrão de lesão fibrótico-celular predominou nos cães infectados com cinco ou mais sinais clínicos. Em contrapartida, fibrose estava presente em intensidade maior no grupo nos cães infectados com até quatro sinais clínicos. A análise imunoistoquímica mostrou maior intensidade de antígeno e amastigotas de Leishmania nos cães infectados com cinco ou mais sinais clínicos. A análise semi-quantitativa do antígeno mostrou correlação positiva de acordo com a intensidade da fibrose. Concluiu-se que a presença do parasita e seus antígenos modificam a arquitetura pulmonar evoluindo, em sua maioria, para o padrão fibrótico-celular em associação à intensidade das manifestações clínicas.

The aim of this study was to identify and classify the pulmonary alterations in dogs naturally infected by Leishmania (Leishmania) infantum according to clinical manifestation. After diagnosis to VL by parasitological exam, 24 infected dogs were subdivided in two groups: 10 dogs with until four clinical signs; and 14 dogs with five or more clinical signs. As control were evaluated 5 healthy dogs. Pulmonary tissue samples were stained by hematoxylin-eosin (H-E) and Masson´s Trichrome to histopathological examination, and Grocott's methenamine silver to exclude fungal infections, as well immunoperoxidase to detection of Leishmania. Histopathological examination of the lungs showed a focal thickening of the alveolar septa due to congestion, edema and cellular infiltrate. Moreover, presence of fibrosis, bronchiolar exudate and flaking of the bronchiolar epithelium, presence of hemosiderin and hyperplasia of goblet cell were observed. The pattern fibrotic-cellular lesions predominated in most infected dogs but mainly in infected dogs with five or more signs clinical. However, fibrosis was higher in infected dogs with four signs clinical. Immunoperoxidase staining showed amastigotes and/or antigen of Leishmania in all infected dogs but there was an increase significant in infected dogs with five or more signs clinical. Semi-quantitative analysis of intensities antigen reveled a positive correlation between fibrosis in infected dogs. In conclusion, the presence of the parasite and/or antigens modifies the lung architecture due for fibrotic-cellular pattern in infected dogs with severe manifestation clinical.