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.

Influence of killer immunoglobulin-like receptors genes on the recurrence rate of ocular toxoplasmosis in Brazil.

BACKGROUND: Recurrence is a hallmark of ocular toxoplasmosis (OT), and conditions that influence its occurrence remain a challenge. Natural killer cells (NK) are effectors cells whose primary is cytotoxic function against many parasites, including Toxoplasma gondii. Among the NK cell receptors, immunoglobulin-like receptors (KIR) deserve attention due to their high polymorphism. OBJECTIVES: This study aimed to analyse the influence of KIR gene polymorphism in the course of OT infection and its association with recurrences after an active episode. METHODS: Ninety-six patients from the Ophthalmologic Clinic of the National Institute of Infectology Evandro Chagas were followed for up to five years. After DNA extraction, genotyping of the patients was performed by polymerase chain reaction sequence-specific oligonucleotide (PCR-SSO) utilising Luminex equipment for reading. During follow-up, 60.4% had a recurrence. FINDINGS: We identified 25 KIR genotypes and found a higher frequency of genotype 1 (31.7%) with worldwide distribution. We note that the KIR2DL2 inhibitor gene and the gene activator KIR2DS2 were more frequent in patients without recurrence. Additionally, we observed that individuals who carry these genes progressed recurrence episodes slowly compared to individuals who do not carry these genes. MAIN CONCLUSIONS: The KIR2DL2 and KIR2DS2 are associated as possible protection markers against ocular toxoplasmosis recurrence (OTR).

Manejo da coroidite interna punctata complicada por membrana neovascular sub-retiniana / Management of punctate inner choroidopathy complicated by subretinal neovascular membrane

RESUMO Este trabalho visou evidenciar a importância da detecção precoce da coroidite interna punctata e destacar sua fisiopatologia inflamatória e possíveis diagnósticos diferenciais dentro das white dot syndromes. O destaque foi dado principalmente à coroidite multifocal e à panuveíte, ao se demonstrar sua epidemiologia peculiar em mulheres jovens, caracterizar sua apresentação clínica típica na fundoscopia e explorar as vantagens e as desvantagens de realizar os exames complementares que fazem parte da análise multimodal útil para o diagnóstico (especialmente a angiografia fluoresceínica, a tomografia de coerência óptica e a indocianina verde). Descreve-se o caso de uma mulher de 28 anos diagnosticada com coroidite interna punctata com membrana neovascular coroidal em olho direito. O tratamento foi realizado com injeção intravítrea de aflibercepte e corticoterapia sistêmica 1mg/kg ao dia. Este relato é importante por permitir debater o manejo da coroidite interna punctata durante a gestação e a decisão de realizar o tratamento mediante uma diversidade de opções terapêuticas.

ABSTRACT This work aimed to demonstrate the importance of early detection of punctate inner choroidopathy, highlighting the pathophysiology of inflammation and the differential diagnoses among white dot syndromes. Special attention was given to multifocal choroiditis and panuveitis, by demonstrating the peculiar epidemiology in young women, characterizing the typical clinical presentation in ophthalmoscopy, and exploring the advantages and disadvantages of performing the complementary examinations, which are part of the multimodal analysis useful for diagnosis (particularly fluorescein angiography, optical coherence tomography, and indocyanine green). We report the case of a 28-year-old female, diagnosed as punctate inner choroidopathy with choroidal [N.T. no título aparece subretinal = subrretiniana] neovascular membrane in the right eye. She was treated with intravitreal injection of aflibercept and systemic corticosteroid 1 mg/kg/day. This case report is important for addressing the management of punctate inner choroidopathy during pregnancy, and the decision to carry out treatment considering diverse therapeutic options.

Ocular Sporotrichosis: 26 Cases with Bulbar Involvement in a Hyperendemic Area of Zoonotic Transmission.

PURPOSE: To describe 26 cases of bulbar conjunctival sporotrichosis. METHODS: Review of clinical records of patients with bulbar conjunctivitis due to culture-proven Sporothrix spp. infection, in Rio de Janeiro, from 2007 to 2017. RESULTS: Twenty-six patients were identified. Median age was 25 years. Adults were more affected (53.8%), followed by adolescents (26.9%). There was a predominance of women (73.1%). Twenty-four patients (96%) reported contact with cats with sporotrichosis. Twenty-one patients (80.8%) presented a primary ocular sporotrichosis. Five patients presented associated eyelid lesions, and 21 (80.8%) tarsal conjunctivitis. Parinaud oculoglandular syndrome was observed in 17 (81%) patients. Eight patients (36.4%) reported the use of steroid drops before diagnosis was made. All patients but one were treated with oral itraconazole. Twenty-three patients (88.5%) were completely cured and three (11.5%) were lost to follow-up. Eight patients (34.8%) developed ocular sequelae. CONCLUSION: Bulbar conjunctivitis is an important clinical presentation of ocular sporotrichosis. It can lead to ocular sequelae. Sporotrichosis should be considered in the differential diagnosis of ophthalmic external diseases, especially in patients with cat contact history.

Toxoplasmic retinochoroiditis: The influence of age, number of retinochoroidal lesions and genetic polymorphism for IFN-γ +874 T/A as risk factors for recurrence in a survival analysis.

PURPOSE: To analyze risk factors for recurrent toxoplasmic retinochoroiditis. DESIGN: Single center prospective case series. POPULATION AND METHODS: A total of 230 patients with toxoplasmic retinochoroiditis were prospectively followed to assess recurrences. All patients were treated with a specific drug regime for toxoplasmosis in each episode of active retinochoroiditis. Individuals with chronic diseases and pregnant women were excluded. Survival analysis by extended Cox regression model (Prentice-Williams-Peterson counting process model) was performed to evaluate the time between recurrences according to some potential risk factors: age, number of retinochoroidal lesions at initial evaluation, sex and interferon gamma +874 T/A gene polymorphism. Hazard Ratios (HR) and 95% confidence intervals (CI) were provided to interpret the risk effects. RESULTS: One hundred sixty-two recurrence episodes were observed in 104 (45.2%) patients during follow-up that lasted from 269 to 1976 days. Mean age at presentation was 32.8 years (Standard deviation = 11.38). The risk of recurrence during follow up was influenced by age (HR = 1.02, 95% CI = 1.01-1.04) and number of retinochoroidal lesions at the beginning of the study (HR = 1.60, 95% CI = 1.07-2.40). Heterozygosis for IFN-γ gene polymorphism at position +874 T/A was also associated with recurrence (HR = 1.49, 95% CI = 1.04-2.14). CONCLUSION: The risk of ocular toxoplasmosis recurrence after an active episode increased with age and was significantly higher in individuals with primary lesions, which suggests that individuals with this characteristic and the elderly could benefit from recurrence prophylactic strategies with antimicrobials. Results suggest an association between IFN-γ gene polymorphism at position +874T/A and recurrence.

Toxoplasmic Retinochoroiditis: Clinical Characteristics and Visual Outcome in a Prospective Study.

PURPOSE: To ascertain the clinical features and visual outcome of toxoplasma retinochoroiditis in a large series of cases. SUBJECTS AND METHODS: 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. RESULTS: 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). CONCLUSION: 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.

The IFN-gamma +874T/A gene polymorphism is associated with retinochoroiditis toxoplasmosis susceptibility.

Toxoplasmosis is a worldwide zoonosis that generally produces an asymptomatic infection. In some cases, however, toxoplasmosis infection can lead to ocular damage. The immune system has a crucial role in both the course of the infection and in the evolution of toxoplasmosis disease. In particular, IFN-gamma plays an important role in resistance to toxoplasmosis. Polymorphisms in genes encoding cytokines have been shown to have an association with susceptibility to parasitic diseases. The aim of this work was to analyse the occurrence of polymorphisms in the gene encoding IFN-gamma (+874T/A) among Toxoplasma gondii seropositive individuals, including those with ocular lesions caused by the parasite, from a rural population of Santa Rita de Cássia, Barra Mansa, state of Rio de Janeiro, Brazil. Further, we verified which of these polymorphisms could be related to susceptibility to the development of ocular toxoplasmosis. This study included 34 individuals with ocular toxoplasmosis (ocular group) and 134 without ocular lesions (control group). The differences between A and T allele distributions were not statistically significant between the two groups. However, we observed that a higher frequency of individuals from the ocular group possessed the A/A genotype, when compared with the control group, suggesting that homozygocity for the A allele could enhance susceptibility to ocular toxoplasmosis in T. gondii infection.

[Frequency of lesions suggestive of ocular toxoplasmosis among a rural population in the State of Rio de Janeiro]. / Frequência de lesões sugestivas de toxoplasmose ocular em uma população rural do Estado do Rio de Janeiro.

To determine the prevalence of ocular toxoplasmosis among the general population of the district of Santa Rita de Cassia, Barra Mansa, State of Rio de Janeiro, a cross-sectional study on 1,071 individuals was performed. These subjects underwent serological tests (anti-Toxoplasma IgG and IgM) and physical and ophthalmological examinations. The diagnosis of presumed ocular toxoplasmosis was based on clinical and serological criteria and the appearance of the retinochoroidal lesion. The lesions were classified into three morphological types: 1. Limits marked with a halo of hyperpigmentation and an area of central chorioretinal atrophy; 2. Hypopigmented halo and hyperpigmented central area; and 3. Hyperpigmented or hypopigmented. The prevalence of healed lesions compatible with ocular toxoplasmosis was 3.8% among the general population and 5.8% among individuals who were seropositive for Toxoplasma gondii (65.9% of the individuals evaluated). Type-1 lesions (41.5%), female sex (68.3%), peripheral lesions (58.5%) and lesions smaller than three disc diameters predominated.

The IFN-γ +874T/A polymorphism is associated with retinochoroiditis toxoplasmosis susceptibility

Toxoplasmosis is a worldwide zoonosis that generally produces an asymptomatic infection. In some cases, however, toxoplasmosis infection can lead to ocular damage. The immune system has a crucial role in both the course of the infection and in the evolution of toxoplasmosis disease. In particular, IFN-γ plays an important role in resistance to toxoplasmosis. Polymorphisms in genes encoding cytokines have been shown to have an association with susceptibility to parasitic diseases. The aim of this work was to analyse the occurrence of polymorphisms in the gene encoding IFN-γ (+874T/A) among Toxoplasma gondii seropositive individuals, including those with ocular lesions caused by the parasite, from a rural population of Santa Rita de Cássia, Barra Mansa, state of Rio de Janeiro, Brazil. Further, we verified which of these polymorphisms could be related to susceptibility to the development of ocular toxoplasmosis. This study included 34 individuals with ocular toxoplasmosis (ocular group) and 134 without ocular lesions (control group). The differences between A and T allele distributions were not statistically significant between the two groups. However, we observed that a higher frequency of individuals from the ocular group possessed the A/A genotype, when compared with the control group, suggesting that homozygocity for the A allele could enhance susceptibility to ocular toxoplasmosis in T. gondii infection.

Frequência de lesões sugestivas de toxoplasmose ocular em uma população rural do Estado do Rio de Janeiro / Frequency of lesions suggestive of ocular toxoplasmosis among a rural population in the State of Rio de Janeiro

Para determinar a prevalência da toxoplasmose ocular na população em geral do bairro de Santa Rita de Cássia, Barra Mansa, RJ, foi realizado um estudo seccional no qual 1.071 indivíduos foram submetidos a testes sorológicos (IgG e IgM anti-Toxoplasma) e a exame físico e oftalmológico. O diagnóstico da toxoplasmose ocular presumida foi baseado em critérios clínicos, sorológicos e aspecto da lesão retinocoroidiana. As lesões foram classificadas em três tipos morfológicos: 1. Limites marcados com halo de hiperpigmentação e área de atrofia coriorretiniana central. 2. Halo hipopigmentado e área central hiperpigmentada e 3. Hiperpigmentadas ou hipopigmentadas. A prevalência de lesões cicatrizadas compatíveis com toxoplasmose ocular foi de 3,8 por cento na população em geral e 5,8 por cento entre os indivíduos com sorologia positiva para Toxoplasma gondii (65,9 por cento dos indivíduos analisados), com predominância de: lesões do tipo 1 (41,5 por cento), sexo feminino (68,3 por cento), periféricas (58,5 por cento) e menores que 3 diâmetros de disco (87,8 por cento).

To determine the prevalence of ocular toxoplasmosis among the general population of the district of Santa Rita de Cassia, Barra Mansa, State of Rio de Janeiro, a cross-sectional study on 1,071 individuals was performed. These subjects underwent serological tests (anti-Toxoplasma IgG and IgM) and physical and ophthalmological examinations. The diagnosis of presumed ocular toxoplasmosis was based on clinical and serological criteria and the appearance of the retinochoroidal lesion. The lesions were classified into three morphological types: 1. Limits marked with a halo of hyperpigmentation and an area of central chorioretinal atrophy; 2. Hypopigmented halo and hyperpigmented central area; and 3. Hyperpigmented or hypopigmented. The prevalence of healed lesions compatible with ocular toxoplasmosis was 3.8 percent among the general population and 5.8 percent among individuals who were seropositive for Toxoplasma gondii (65.9 percent of the individuals evaluated). Type-1 lesions (41.5 percent), female sex (68.3 percent), peripheral lesions (58.5 percent) and lesions smaller than three disc diameters predominated.

Prevalência da Toxoplasmose Ocular presumida em uma população rural, no bairro de Santa Rita de Cássia, Município de Barra Mansa, RJ / Prevalence of the ocular toxoplasmosis presumed in an agricultural population, in the neighborhood of Santa Rita de Cassia in the town of Barra Mansa, RJ

O T. gondii é o agente etiológico mais frequente das uveítes posteriores no Brasil e no mundo, entretanto, os estudos sobre a prevalência da toxoplasmose ocular na população geral são escassos. Nenhum trabalho neste sentido foi encontrado na literatura no que concerne à região sul do Estado do Rio de Janeiro. Com o objetivo de determinar a prevalência de formas oculares da toxoplasmose no bairro de Santa Rita de Cássia, Barra Mansa - RJ foi realizado um estudo seccional em 1071 indivíduos de uma população estimada em 1300 habitantes. Todos responderam a um questionário epidemiológico, foram submetidos a testes sorológicos específicos para dosagem de IgG e IgM anti-Toxoplasma, e também a exame físico e oftalmológico na busca de evidências da infecção pelo T. gondii. O diagnóstico de toxoplasmose ocular presumida foi feito baseado em critérios clínicos (aspecto da lesão retinocoroidiana à oftalmoscopia indireta sob midríase) e sorológicos; e as lesões foram classificadas em três tipos morfológicos (tipo 1-Limites bem marcados com halo de hiperpigmentação e área de atrofia coriorretiniana central, tipo 2- Lesões com halo hipopigmentado e área central hiperpigmentada e tipo 3- Lesões hiperpigmentadas ou hipopigmentadas compatíveis com hiperplasia ou atrofia do epitélio pigmentar retiniano). A soroprevalência de IgG anti-Toxoplasma foi de 65,92por cento e nenhum caso apresentou IgM reagente. Foram encontrados 41 pacientes com lesões compatíveis com toxoplasmose ocular presumida na face cicatricial, o que corresponde a uma prevalência de 3,83 por cento na população geral e de 5,81 por cento entre os soropositivos. Entre estes pacientes foi encontrada uma predominância de indivíduos do sexo feminino (68,29 por cento) com lesões do tipo 1 (41,46 por cento), periféricas (58,54 por cento) e menores que 3 diâmetros de disco (87,80 por cento). Embora tenha sido encontrada uma alta prevalência de casos de retinocoroidite toxoplasmática presumida em...estudada.