Comparison of immunological and molecular methods for laboratory diagnosis of ocular toxoplasmosis in blood, serum and tears in Brazil.

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.

Occurrence of Toxoplasma gondii and other gastrointestinal parasites in free-roaming cats from the Rio de Janeiro zoo.

This communication aimed to determine the frequency of infection by T. gondii and other gastrointestinal parasites in free-living cats captured in BioParque do Rio (Fundação RioZoo), Brazil. To this end, 58 blood and 51 fecal samples were collected from 68 cats from July 2019 to September 2020. The serum obtained was analyzed by indirect fluorescent antibody test for anti-T. gondii IgG. Fecal samples were examined by microscopic parasitological techniques. Of the total, 10.3% of the animals presented anti-T. gondii IgG. Parasitic structures were observed in 39.2% of the fecal samples. Hookworms were the most detected parasites (19.6%) followed by Cystoisospora sp. (11.7%), Dipylidium caninum (7.8%) and Toxocara cati (3.9%). These results indicated the exposure of this population of stray cats to potentially zoonotic parasites, which in addition to causing possible damage to domestic and wild animal health, also pose risks to public health.

Esta comunicação teve como objetivo determinar a frequência de infecção por T. gondii e outros parasitas gastrointestinais em gatos de vida livre capturados no BioParque do Rio (Fundação RioZoo), Brasil. Para isso, foram coletadas 58 amostras de sangue e 51 amostras fecais de 68 gatos capturados entre julho de 2019 e setembro de 2020. O soro obtido foi submetido à reação de imunofluorescência indireta para IgG anti-T. gondii. As amostras fecais foram submetidas a técnicas parasitológicas microscópicas. Do total, 10,3% dos animais apresentaram IgG anti-T. gondii. Estruturas parasitárias foram observadas em 39,2% das amostras fecais. Os ancilostomídeos foram os parasitas mais detectados (19,6%), seguidos por Cystoisospora sp. (11,7%), Dipylidium caninum (7,8%) e Toxocara cati (3,9%). Esses resultados indicaram a exposição dessa população de gatos de rua a parasitas potencialmente zoonóticos, que, além de causar possíveis danos à saúde dos animais domésticos e selvagens, também representam riscos à saúde pública.

Intestinal parasites and risk factors in dogs and cats from Rio de Janeiro, Brazil.

The urban populations of pets, mainly dogs and cats, have been growing in recent years and, consequently, human-animal relationships have become increasingly closer. However, pets are vulnerable to infection by various parasites, as well are their owners due to the zoonotic potential of these agents. This study aimed to assess the frequency and risk factors of intestinal parasites in dogs and cats attended at the Municipal Institute of Veterinary Medicine Jorge Vaitsman, Rio de Janeiro, Brazil, as well as georeferencing the parasitized animals. A total of 608 animals (400 dogs and 208 cats) were enrolled from August 2017 to November 2018. Fecal samples were obtained from the animal's rectum during clinical examination. In addition, the owners of the enrolled dogs and cats were asked to complete an epidemiological questionnaire with questions about the animals' general information, lifestyle, and handling. Fecal samples were subjected to sedimentation and centrifugal-flotation in sucrose solution techniques. The frequencies of intestinal parasites total in dogs and cats were 11.3% and 24.5%, respectively, with hookworms being the most detected parasites in dogs (8.3%) and Dipylidium caninum in cats (12.5%). Sex was the only variable that showed statistical differences in the canine population males being more often infected than female (p < 0.05), while street access was associated with parasitic infections in cats (p < 0.05). The results obtained in this study demonstrate the need to control parasitic infections in domiciled dogs and cats in different municipalities in the metropolitan region of the state of Rio de Janeiro, including residents in the northern, western and central areas.

Non-invasive sampling in Itatiaia National Park, Brazil: wild mammal parasite detection.

BACKGROUND: Non-invasive sampling through faecal collection is one of the most cost-effective alternatives for monitoring of free-living wild mammals, as it provides information on animal taxonomy as well as the dynamics of the gastrointestinal parasites that potentially infect these animals. In this context, this study aimed to perform an epidemiological survey of gastrointestinal parasites using non-invasive faecal samples from carnivores and artiodactyls identified by stool macroscopy, guard hair morphology and DNA sequencing in Itatiaia National Park. Between 2017 and 2018, faeces from carnivores and artiodactyls were collected along trails in the park. The host species were identified through macroscopic and trichological examinations and molecular biology. To investigate the parasites, the Faust, Lutz and modified Ritchie and Sheather techniques and enzyme immunoassays to detect Cryptosporidium sp. antigens were used. RESULTS: A total of 244 stool samples were collected. The species identified were Chrysocyon brachyurus, Leopardus guttulus, Canis familiaris, Cerdocyon thous, Puma yagouaroundi, Leopardus pardalis, Puma concolor and Sus scrofa. There were 81.1% samples that were positive for parasites distributed mainly in the high part of the park. Helminths, especially eggs of the family Ascarididae, were more frequently detected in carnivore faeces (70.9%). Protozoa, especially Cryptosporidium sp., represented the highest frequency of infection in artiodactyl faeces (87.1%). This zoonotic protozoon was detected in eight mammalian species, including in a wild boar. High values of structural richness and Shannon and Simpson diversity indices were observed for the parasites, especially in the faeces of C. brachyurus. Significant differences in parasite diversity were observed between wild and domestic animals, such as C. brachyurus and C. familiaris, respectively, and between taxonomically distant species, such as C. brachyurus and S. scrofa. The highest values for parasite similarity were found among the species that frequented similar areas of the park, such as C. brachyurus and L. guttulus. CONCLUSIONS: The animals and parasite infections were identified through the combination of three techniques. High frequency parasite structures were diagnosed. Zoonotic protozoa were found and mainly occurred in samples from introduced species.

Oncicola venezuelensis (Marteau, 1977) (Acanthocephala: Oligacanthorhynchidae) in Puma concolor in Rio de Janeiro, Brazil.

Specimens of Oncicola venezuelensis (Marteau, 1977) were recovered from fragments of intestinal tissue of a female Puma concolar (Linn, 1771) found dead in Petrópolis, Rio de Janeiro in 2017. A total of 140 helminths were recovered. Five males and 5 females of the helminths were analyzed morphologically as well as 50 parasite eggs recovered in intestinal contents. Morphologically, these helminths were compatible with the genus Oncicola, because of the size and shape of the proboscis, the size and disposition of the lemnisci and the morphometry of the eggs, in which the external membrane of the shell was delicate and clear. From histopathology, the helminths were deeply embeded in the mucosa reaching up to the muscle layer. One specimen was also identified molecularly with universal primers that amplified the eukaryote region ITS1-5.8S-ITS2. The helminth showed 99% identity with the gene sequence of O. venezuelensis deposited in GenBank. It is important to emphasize, this parasite has been very little reported in the literature, which reinforces the importance of this report.

Comparison of Four Parasitological Techniques for Laboratory Diagnosis of Eggs from Spirometra spp. in Wild Mammal Fecal Samples.

PURPOSE: This study aimed to compare the performance of four different microscopic coproparasitological techniques in relation to egg recovering and the frequency of alterations in the eggs observed through each technique. METHODS: A total of 213 fecal samples from free-living carnivorous mammals were collected between 2017 and 2018 in Itatiaia National Park, RJ, Brazil. Faust and modified Sheather floatation techniques as well as Lutz and modified Ritchie sedimentation techniques were applied. RESULTS: The total positivity rate for Spirometra spp. eggs was 24.4%, and these were detected mainly through Lutz and modified Ritchie sedimentation techniques, with substantial agreement (kappa = 0.73; p = 0.00 < 0.05; McNemar p value = 1.0; Fisher's exact test p = 0.616). Faust and modified Sheather flotation techniques did not present good egg recovery, with frequencies of 6.6% and 7.5%, respectively. Eggs with morphological alterations were mostly observed through Faust (17.3%) and modified Sheather (13.5%). Both flotation techniques presented statistically significant frequencies of deformed eggs, in comparison with the sedimentation techniques (p = 0.00). Low frequencies of deformed eggs were observed when the samples were analyzed through modified Ritchie and Lutz sedimentation techniques. CONCLUSIONS: From these results, sedimentation techniques such as modified Ritchie and Lutz methods were more efficient for diagnosing the eggs of this helminth in fecal material from free-living carnivores and should always be used when analyzing fecal samples from hosts of different species.