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To ascertain the clinical features and visual outcome of toxoplasma retinochoroiditis in a large series of cases. Two hundred and thirty subjects diagnosed with active toxoplasma retinochoroiditis were prospectively followed for periods ranging from 269 to 1976 days. All patients presented with active retinochoroiditis and positive IgG T. gondii serology at the beginning of the study and received a standardized drug treatment for toxoplasmosis, both in the first episode and in the subsequent recurrences. The group involved 118 (51.3%) men and 112 (48.7%) women, with ages ranging from 14 to 77 years, mean of 32.4 years (SD = 11.38). Primary retinochoroidal lesions were observed in 52 (22.6%) cases and active retinochoroiditis combined with old scars in 178 (77.4%) subjects at the beginning of the study. A hundred sixty-two recurrent episodes in 104 (45.2%) patients were observed during follow-up. New subclinical retinochoroidal lesions were detected in 23 of 162 (14.2%) recurrences episodes during the follow-up. Posterior segment complications were observed in 73 (31.7%) subjects. Retinochoroidal lesions adjacent to the optic nerve and in the macular area were observed in 27 of 40 (67.5%) cases of severe visual impairment (VA = 20/200 or worse). Toxoplasma retinochoroiditis in this population had a high recurrence rate after an active episode. Severe visual impairment was associated with location of the retinochoroidal scar, recurrences and posterior segment complications. It is crucial to consider the location of the lesion in studies analyzing visual prognosis as a measure for treatment effectiveness and prevention strategies.

Toxoplasma gondii is a highly prevalent zoonotic parasite in Brazil capable of infecting mammals and birds. The increase in the urban populations of pets and the narrowing of the human–animal relationship can facilitate the transmission of important public health zoonoses, such as toxoplasmosis. This study aimed to evaluate the frequency and spatial distribution of T. gondii infection and its risk factors in domiciled dogs and cats attended at the Jorge Vaitsman Institute, Rio de Janeiro. Serum samples from 400 dogs and 272 cats were evaluated by an indirect fluorescent antibody test (IFAT) for IgG anti- T. gondii antibodies. Epidemiological questionnaires were used to interview the animals’ owners to identify risk factors for infection. Of the total, 34% (136/400) of dogs and 8.1% (22/272) of cats had anti- T. gondii antibodies. Breed (OR: 2.10–95%, CI 1.27–3.46) was a risk factor for dogs, while sex (OR: 3.40–95%, CI 1.10–10.52) and homemade food consumption (OR: 8.49–95%, CI 2.48–29.05) were risk factors for cats. Offal consumption was considered a risk factor for both species evaluated (OR: 2.74–95%, CI 1.38–5.43 for dogs; OR: 7.66–95%, CI 1.24–47.29 for cats). The spatial analysis showed that T. gondii seropositive animals were widely distributed in the metropolitan region of Rio de Janeiro state, with a concentration observed mainly in the west and north zones of Rio de Janeiro city. The results emphasize the importance of adopting prophylactic measures to control T. gondii transmission in domiciled dogs and cats in Rio de Janeiro, contributing positively to public health. Toxoplasma gondii est un parasite zoonotique très répandu au Brésil, capable d’infecter les mammifères et les oiseaux. L’augmentation des populations urbaines d’animaux de compagnie et le rétrécissement de la relation homme-animal peuvent faciliter la transmission de zoonoses importantes pour la santé publique, telles que la toxoplasmose. Cette étude visait à évaluer la fréquence et la distribution spatiale de l’infection à T. gondii et ses facteurs de risque chez les chiens et les chats domiciliés qui ont fréquenté l’Institut Jorge Vaitsman de Rio de Janeiro. Des échantillons de sérum de 400 chiens et 272 chats ont été évalués par un test d’immunofluorescence indirecte (IFAT) pour les anticorps IgG anti- T. gondii . Des questionnaires épidémiologiques ont été appliqués aux propriétaires des animaux pour identifier les facteurs de risque d’infection. Sur le total, 34 % (136/400) des chiens et 8,1 % (22/272) des chats avaient des anticorps anti- T. gondii . La race (OR : 2,10-95 %, IC 1,27-3,46) était un facteur de risque pour les chiens, tandis que le sexe (OR : 3,40-95 %, IC 1,10-10,52) et la consommation d’aliments faits maison (OR : 8,49-95 % IC 2,48-29,05) l’étaient pour les chats. La consommation d’abats a été considérée comme un facteur de risque pour les deux espèces évaluées (OR : 2,74-95 %, IC 1,38-5,43 pour les chiens; OR : 7,66-95 %, IC 1,24-47,29 pour les chats). L’analyse spatiale a montré que les animaux séropositifs pour T. gondii étaient largement répartis dans la région métropolitaine de l’État de Rio de Janeiro, avec une concentration observée principalement dans les zones ouest et nord de la ville de Rio de Janeiro. Les résultats soulignent l’importance d’adopter des mesures prophylactiques pour contrôler la transmission de T. gondii chez les chiens et les chats domiciliés à Rio de Janeiro, contribuant positivement à la santé publique.

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.

The highly virulent Toxoplasma gondii RH strain is maintained through successive passages in mice, but there is still a lack of studies that refine these procedures from a 3Rs perspective, where humanitarian ideals aim to minimize the stress, pain, or suffering of the animals used in the research without the loss of results. The aim of this study was to establish humane endpoints in Swiss Webster mice inoculated with the T. gondii RH strain. A total of 52 mice were infected with 5 × 106 tachyzoites/mL and monitored for periods of up to 5 days. The parameters body weight; hair condition; higher than normal body temperature; hypothermia; respiratory function; pain; soft stools or diarrhea; bloody diarrhea; tense, nervous, or in distress during handling; and ascites were recorded daily in score tables. The results showed that prominent piloerection, respiratory function, pain parameters, and ascites are important clinical signs to be used as a cut-off point for implementing euthanasia. The application of this refinement method helped to avoid animal suffering and pain without compromising the number of parasites recovered. We therefore suggest adopting these parameters in research protocols that require the maintenance of the T. gondii RH strain in murine models to avoid and reduce animal suffering.

Balantioides coli is a ciliated protist that can cause dysentery in humans, pigs and nonhuman primates and may have the potential for zoonotic transmission. Its diagnosis is routinely performed through conventional parasitological techniques, and few studies have used culturing techniques to isolate it, applying molecular tools for the characterization of this protozoan. Thus, the objective of this study was to confirm B. coli diagnosis using molecular tools and to characterize the genetic variants of this parasite isolated from pigs kept on family farms in Brazil using three different culture media that differed in the serum added. Fecal samples from pigs were inoculated in Pavlova medium plus coconut water (PC), fetal bovine serum (PB) and horse serum (PH). Of the 127 samples positive for forms compatible with the phylum Ciliophora, 31 were selected for isolation. The most successful medium for isolation was PB 19/31 (61.3%), followed by PH 18/31 (58.1%) and PC 11/31 (35.5%). Of the nucleotide sequences generated, 20 were classified as genetic variant type B0, two as A1 and 15 as A0. The results indicated that PC, despite having allowed the isolation of B. coli for a short period, was not an adequate medium for the maintenance of this parasite in vitro, therefore requiring improvement.

Hepatitis E virus (HEV) infection has been demonstrated in various animal species; those recognized as potential zoonotic reservoirs pose a considerable risk to public health. In Brazil, HEV-3 is the only genotype identified in humans and swine nationwide, in a colony-breeding cynomolgus monkey and, recently, in bovines and capybara. There is no information regarding HEV exposure in the equine population in Brazil. This study aimed to investigate anti-HEV antibodies and viral RNA in serum samples from horses slaughtered for meat export and those bred for sport/reproduction purposes. We used a commercially available ELISA kit modified to detect species-specific anti-HEV, using an anti-horse IgG-peroxidase conjugate and evaluating different cutoff formulas and assay precision. Serum samples (n = 257) were tested for anti-HEV IgG and HEV RNA by nested RT-PCR and RT-qPCR. The overall anti-HEV seroprevalence was 26.5% (68/257) without the detection of HEV RNA. Most municipalities (53.3%) and farms (58.8%) had positive horses. Animals slaughtered for human consumption had higher risk of HEV exposure (45.5%) than those bred for sports or reproduction (6.4%) (p < 0.0001). The statistical analysis revealed sex and breeding system as possible risk-associated factors. The first serological evidence of HEV circulation in Brazilian equines reinforces the need for the surveillance of HEV host expansion in a one-health approach.

To determine the prevalence of immunoglobulin G (IgG) and immunoglobulin M (IgM) anti-Toxoplasma gondii antibodies among pregnant and postpartum women attended within the public healthcare system in Niterói, State of Rio de Janeiro,and to detect possible exposure factors associated with T. gondii infection in this population. IgM and IgG anti- T. gondii antibodies were investigated in 276 pregnant and 124 postpartum women by using the indirect immunofluorescence (IFAT) and immunoenzymatic assay (ELISA) techniques. The participants were selected by convenience sampling. All these 400 patients filled out a free and informed consent statement, answered an epidemiological questionnaire and were informed about the disease. Among the 400 samples analyzed, 234 (58.5%) were reactive to IgG anti-T. gondii antibodies, according to the IFAT and/or ELISA assay. One pregnant woman was found to be reactive to IgM anti- T. gondii antibodies, with an intermediate IgG avidity test. Risk factor analysis showed that seropositivity was significantly associated (p<0.05) with age, contact with cats and presence of rodents at home. Through a logistic regression model, these associations were confirmed for age and contact with cats, while education at least of the high school level was found to be a protective factor. The prevalence rate of IgG anti-T. gondii antibodies in the city of Niterói was high and the risk factors for infection detected after multivariate analysis were: age over 30 years, contact with cats and education levels lower than university graduate level.

Ocular toxoplasmosis (OT) is caused by protozoan T . gondii . Ophthalmological examination is considered the gold standard for OT diagnosis, and laboratory tests are used for diagnostic confirmation. However, these tests can present different results, which change depending on their basis, on sample type and on patients’ clinical alteration. Thus, the aim of the present study is to assess immunodiagnostic and molecular techniques applied in blood, serum and tear fluid to diagnose T . gondii infection in patients seen at an Ophthalmology Clinic. In total, 160 patients were included in the study, 40 of them had OT with active lesions (G1); 40 had OT with healed lesions (G2), 40 had non-toxoplasmic uveitis (G3) and 40 had no ocular alterations (G4). Serum samples were subjected to Immunoenzymatic Assay (ELISA) and to Indirect Immunofluorescence Reaction (IFAT) to search for anti- T . gondii IgM and IgG. Tear fluid samples were analyzed through ELISA for IgA research. All blood and tear fluid samples were subjected to conventional polymerase chain reaction (cPCR) and in a Nested PCR model for T . gondii DNA amplification with targets B1, GRA7 and REP 529. IgG and IgM anti- T . gondii was detected in serum samples from 106 and 15 patients, respectively, when combining ELISA and IFAT results. Anti- T . gondii IgA antibodies were detected in 9.2% of the tear material. Nested PCR with GRA7 target showed higher positivity in blood samples (24.4%); Nested PCR with B1 target showed a higher frequency of positivity in tears (15%). Biological samples of patients with active lesions showed the highest positivity frequencies in all immunodiagnostic assays, as well as in most PCR models. The present results highlighted the need of associating techniques with different fundamentals to confirm OT diagnosis. Furthermore, further tear fluid analyses should be performed to validate this biological material as lesser invasive alternative for the more accurate OT diagnosis.

[This corrects the article DOI: 10.1371/journal.pone.0247560.].

In canine leishmaniosis caused by the protozoan Leishmania infantum , little is known about how co-infections with or co-seropositivities for other pathogens can influence aggravation of this disease. Therefore, the objectives of this study were to evaluate the frequency of co-infections with or co-seropositivities for certain pathogens in dogs seropositive for L . infantum and their relationship with clinical signs, histological changes and L . infantum load. Sixty-six L . infantum -seropositive dogs were submitted to clinical examination, collection of blood and bone marrow, culling, and necropsy. Antibodies against Anaplasma spp., Borrelia burgdorferi sensu lato, Ehrlichia spp. and Toxoplasma gondii and Dirofilaria immitis antigens were investigated in serum. Samples from different tissues were submitted to histopathology and immunohistochemistry for the detection of Leishmania spp. and T . gondii . Quantitative real-time PCR was used to assess the L . infantum load in spleen samples. For detection of Coxiella burnetii , conventional PCR and nested PCR were performed using bone marrow samples. All 66 dogs tested positive for L . infantum by qPCR and/or culture. Fifty dogs (76%) were co-seropositive for at least one pathogen: T . gondii (59%), Ehrlichia spp., (41%), and Anaplasma spp. (18%). Clinical signs were observed in 15 (94%) dogs monoinfected with L . infantum and in 45 (90%) dogs co-seropositive for certain pathogens. The L . infantum load in spleen and skin did not differ significantly between monoinfected and co-seropositive dogs. The number of inflammatory cells was higher in the spleen, lung and mammary gland of co-seropositive dogs and in the mitral valve of monoinfected dogs. These results suggest that dogs infected with L . infantum and co-seropositive for certain pathogens are common in the region studied. However, co-seropositivities for certain pathogens did not aggravate clinical signs or L . infantum load, although they were associated with a more intense inflammatory reaction in some organs.