Comparative GRA6 and GRA7 for their Utility as Genetic Markers in the Genotyping of Cerebral Toxoplasmosis in Cerebrospinal Fluid.

INTRODUCTION: Cerebral toxoplasmosis is a severe symptom of Toxoplasma gondii (T. gondii) infection that often affects individuals with Human Immunodeficiency Virus (HIV) infection and can be fatal. T. gondii exhibits diverse strains with varied virulence, such as cerebral toxoplasmosis, which is connected with a specific strain. Molecular methods were used to investigate the genotype of the parasite. Some researchers have used genetic markers, such as the dense granule proteins GRA6 and GRA7, in order to identify T. gondii genotype. This study aimed to evaluate the applicability of GRA6 and GRA7 as genetic markers for determining T. gondii strain from cerebrospinal fluid of AIDS patients with toxoplasmic encephalitis. METHOD: 160 serum and cerebrospinal fluid (CSF) samples were collected from 2013 to 2022. The serum samples were initially tested using ELISA anti Toxoplasma IgG, and the CSF was subsequently PCR of 5'SAG2 gene for those positive IgG. A total of 69 CSF successfully positive on PCR of 5'SAG2 were included for analysis of GRA6 and GRA7 by performing PCR, sequencing and phylogenetic analysis for determination of T. gondii type. RESULT: The findings of this study indicate that the use of GRA7 is better than GRA6 when using direct clinical samples. Out of the 69 samples analyzed, total of 36 samples (52.17%) were positive for GRA7. The cases can be classified as type I: 86,1% (31/36), type III: 2,7% (1/36) and atypical: 11,1% (4/36). CONCLUSION: Comparison results between GRA6 and GRA7 for genotype determination shows good results on GRA7. GRA7 can be used as a genetic marker to find out the genotype of T. gondii in direct clinical samples where GRA6 cannot be used.

Toxoplasma gondii SAG2 type III in an atypical presentation of ocular toxoplasmosis in Indonesia.

OBJECTIVE: This study aimed to perform genotyping of Toxoplasma gondii strain or variant causing atypical toxoplasmic uveitis in Indonesian patients. METHODS: Ocular fluid samples originating from 46 uveitis patients with non-specific ocular manifestations were analysed for Toxoplasma infection by PCR of the B1 locus. The clonal type was determined by amplification, sequencing and phylogenetic analysis of SAG2 and GRA6 loci in B1-positive samples. Clinical data were obtained from the medical records. RESULTS: Pan uveitis was the most frequent manifestation (65.2%) and mostly unilateral (76.1%). PCR of the B1 locus identified eight positive subjects (12.5%); six had panuveitis and two of these had diabetes mellitus. Phylogenetic analysis with maximum likelihood of the SAG2 locus in the B1-positive samples resulted in Toxoplasma gondii SAG2 type III allele. No positive result was obtained from the PCR of GRA6 locus. CONCLUSION: Toxoplasma gondii SAG2-type III allele was identified in an atypical presentation of toxoplasmic uveitis in Indonesia.