Luiz Cláudio Ferreira

Sensitivity and specificity of serological tests, histopathology and immunohistochemistry for detection of Toxoplasma gondii infection in domestic chickens

Since free-range chickens are important for the epidemiology of toxoplasmosis, this study evaluated the sensitivity and specificity of different laboratory techniques for the diagnosis of Toxoplasma gondii in these animals. Serum samples from 135 adult domestic chickens were tested for anti-T. gondii antibodies by the indirect fluorescent antibody test (IFAT), enzyme-linked immunosorbent assay (ELISA), modified agglutination test (MAT), and indirect hemagglutination test (IHAT). Tissue samples from all animals were analyzed by histopathology, immunohistochemistry and mouse bioassay (gold standard). Fifty-four chickens were positive for T. gondii in the bioassay. The sensitivity and specificity of the different tests were, respectively, 85% and 56% for ELISA; 80% and 52% for IFAT; 76% and 68% for MAT; 61% and 80% for IHAT; 7% and 98% for immunohistochemistry, and 6% and 98% for histopathology. The MAT was the most effective method for the diagnosis of T. gondii infection in chickens, followed by ELISA. Histopathology and immunohistochemistry are useful tools for the diagnosis of T. gondii infection in chickens due to their specificity.

Topical application of Acheflan on rat skin injury accelerates wound healing: a histopathological, immunohistochemical and biochemical study

BackgroundDermal wound healing involves a cascade of complex events including angiogenesis and extracellular matrix remodeling. Several groups have focused in the study of the skin wound healing activity of natural products. The phytomedicine Acheflan®, and its main active constituent is the oil from Cordia verbenacea which has known anti-inflammatory, analgesic and antimicrobial activities. To our knowledge, no investigation has evaluated the effect of Acheflan® in an experimental model of skin wound healing.The present study has explored the wound healing property of Acheflan® and has compared it with topical effectiveness of collagenase and fibrinolysin by using Wistar rat cutaneous excision wound model.MethodsAnimals were divided into four groups: untreated animals are negative control (NC), wounds were treated topically every day with Collagenase ointment (TC), with Fibrinolysin ointment (TF) and with cream Acheflan (TAc). Skin samples were collected on zero, 8th and 15th days after wounding. The healing was assessed by hematoxylin-eosin (HE), picrosirius red, hydoxyproline content and immunohistochemical analysis of the vascular endothelial growth factor (VEGF) and matrix metalloprotease-9 (MMP-9). Statistical analysis was done by ANOVA and Student t-test (p < 0.05).ResultsThe histological analysis HE of wound in the TAc group was more efficient because it was possible to observe the complete remodeling of the epidermis indicating the regression of lesions compared with the NC. The evaluation of picrosirius staining has demonstrated a significant increase of collagen distribution in the TC and TAc treatments compared with NC and TF groups. These results are corroborated with hydroxyproline content. Skin TC and TAc treated rats have showed an increase of VEGF and MMP-9 compared with NC and TF groups. All parameters were significant (P < 0.05).ConclusionThe phytomedicine Acheflan® (oil of Cordia verbenacea) and TC possess higher therapeutic properties for wound healing compared with TF. These ointments seem to accelerate wound healing, probably due to their involvement with the increase of angiogenesis and dermal remodeling.

Occurrence of anti-Toxoplasma gondii antibodies in cattle and pigs slaughtered, State of Rio de Janeiro.

This study aimed to estimate the occurrence of anti-Toxoplasma gondii antibodies by means of the Indirect Immunufluorescent Antibody Assay (IFAT) in cattle and pigs from slaughterhouses in two municipalities in the State of Rio de Janeiro. IgG anti-T. gondii antibodies was found in 1.96% (9/459) of the cattle and 7.64% (31/406) of the pigs, without any positive association (p > 0.05) with risk factors for transmission and seropositivity. Taking the cutoff point of IFAT ³ 1:64, titers of 64 and 256 were detected, suggesting chronic infection. Despite the low prevalence rates found for cattle and pigs, these indicated environmental contamination in the municipalities investigated, biosafety failures at the technified pig farm evaluated and the possibility of human and animal infection through eating raw or undercooked meat from these animals.

Prevalência da infecção por Toxoplasma gondii em galinhas criadas extensivamente em Rio Bonito, Rio de Janeiro

The prevalence of T. gondii infection was investigated in free-range chickens for human consumption in the county of Rio Bonito, Rio de Janeiro, Brazil. Thus, a search for antibodies to T. gondii by indirect fluorescent antibody test was done in the serum of 220 chickens from twenty two farms. Titres equal or greater than 16 were considered positive. The prevalence of toxoplasmic infection found in chickens was 27.6% (61/220). There were positive cases in eighteen (77.2%) farms. The more frequent titre was 16 (12.5%) and the highest was 4096 (0.4%). The high prevalence of T. gondii in chickens and the high percentage of farms with positive cases indicate environmental contamination in the studied area showing the possibility of human and animal infection.

Occurrence of Leishmania infantum in the central nervous system of naturally infected dogs: Parasite load, viability, co-infections and histological alterations

Zoonotic visceral leishmaniasis is caused by the protozoan Leishmania infantum and little is known about the occurrence and pathogenesis of this parasite in the CNS. The aims of this study were to evaluate the occurrence, viability and load of L. infantum in the CNS, and to identify the neurological histological alterations associated with this protozoan and its co-infections in naturally infected dogs. Forty-eight Leishmania-seropositive dogs from which L. infantum was isolated after necropsy were examined. Cerebrospinal fluid (CSF) samples were analyzed by parasitological culture, quantitative real-time PCR (qPCR) and the rapid immunochromatographic Dual Path Platform test. Brain, spinal cord and spleen samples were submitted to parasitological culture, qPCR, and histological techniques. Additionally, anti-Toxoplasma gondii and anti-Ehrlichia canis antibodies in serum and distemper virus antigens in CSF were investigated. None of the dogs showed neurological signs. All dogs tested positive for L. infantum in the CNS. Viable forms of L. infantum were isolated from CSF, brain and spinal cord in 25% of the dogs. Anti-L. infantum antibodies were detected in CSF in 61% of 36 dogs. Inflammatory histological alterations were observed in the CNS of 31% of the animals; of these, 66% were seropositive for E. canis and/or T. gondii. Amastigote forms were associated with granulomatous non-suppurative encephalomyelitis in a dog without evidence of co-infections. The highest frequency of L. infantum DNA was observed in the brain (98%), followed by the spinal cord (96%), spleen (95%), and CSF (50%). The highest L. infantum load in CNS was found in the spinal cord. These results demonstrate that L. infantum can cross the blood-brain barrier, spread through CSF, and cause active infection in the entire CNS of dogs. Additionally, L. infantum can cause inflammation in the CNS that can lead to neurological signs with progression of the disease.

Parinaud's oculoglandular syndrome associated with Sporothrix schenckii

The Parinaud oculoglandular syndrome is a rare eye disease caused by different etiologic agents, including bacteria, viruses and fungi. It is characterized by a granulomatous conjunctivitis, accompanied by adjacent preauricular lymphadenopathy and can bring consequences if not treated promptly. We present a case of Parinaud oculoglandular syndrome caused by Sporothrix schenckii from the sporotrichosis epidemic in its zoonotic form occurring in Rio de Janeiro, Brazil.

Avaliação da confiabilidade entre dois observadores em exames citopatológico e imunocitoquímico de aspirado de medula óssea no diagnóstico da leishmaniose visceral canina

Visceral leishmaniasis is a zoonosis in which the dog appears as the main source of infection in urban areas. Its diagnosis is complex and the cytopathological exam is a fast and cheap alternative to parasite direct visualization and its sensitivity can be increased by immunocytochemistry, though with a higher cost. The accuracy of such methods is dependent on the microscopists experience and therefore, this study evaluated the reliability of such techniques between two observers, from bone marrow aspirates of 50 dogs from an endemic area for the disease. The parasitological culture in Novy-MacNeal-Nicolle medium was used as the reference standard. Among the main findings, the sensitivities obtained by observers I and II were respectively 62.5% and 37.5%, while specificities were 81.1% and 100%. On immunocytochemistry evaluation, the sensitivity was 0% for both evaluators and the specificity 97.3% and 100%. The agreement between evaluators was weak (κ = 0.167) for the cytopathological test and it could not be evaluated for immunocytochemistry, for which there was no detection by the evaluator II. The agreements among the diagnostic methods and the standard reference for the observer I were reasonable (κ = 0.364) for cytopathological examination and bad (κ = -0.041) for immunocytochemistry. For observer II, such agreement could be assessed only for the cytopathological test, being moderate (κ = 0.497). The results point to the possible expertise difference between evaluators, with the evaluator II demonstrating greater experience when interpreting the citopathological test. Although there was the expected sensitivity increase with immunocytochemistry, the technique used in this study was not effective for the diagnosis of infection, regardless of the evaluator.

Gastrointestinal parasites of swine raised in different management systems in the State of Rio de Janeiro, Brazil

De 2012 a 2013 foram pesquisados parasitos gastrintestinais de suinos de granjas localizadas em diferentes municipios do estado do Rio de Janeiro. Amostras fecais de 790 suinos foram coletadas da ampola retal, sendo 88 de propriedades familiares e 702 de granjas com producao industrial. As amostras foram submetidas as tecnicas de Faust et al., Sheather, Ritchie, Lutz e exame direto. A prevalencia estimada foi de 93,1% nas granjas familiares e 59,1% nas granjas industriais. Balantidium coli, coccidios e Entamoeba sp. foram os parasitos que apresentaram as maiores frequencias, sendo as categorias machos e femeas reprodutoras e leitoes de terminacao as mais infectadas (p<0,05). Trofozoitas de B. coli foram mais evidenciados nas amostras fecais de consistencias semi-solida, seguida pelas fezes com consistencias solida e diarreica. Ovos de estrongilideos e Trichuris suis foram detectados exclusivamente em criacoes familiares. Ovos de Ascaris suum e de Strongyloides ransomi apresentaram baixa frequencia. O alto grau de parasitismo, principalmente de protozoarios, indica a necessidade de reavaliacao do manejo dos suinos em ambos os tipos de producao.