The Role of microRNAs in the Infection by T. gondii in Humans.

MicroRNAs are molecules belonging to an evolutionarily conserved family of small non-coding RNAs, which act on post-transcriptional gene regulation, causing messenger RNA (mRNA) degradation or inhibiting mRNA translation into proteins. These molecules represent potential biomarkers for diagnosis, non-invasive prognosis, and monitoring the development of the disease. Moreover, they may provide additional information on the pathophysiology of parasitic infections and guide strategies for treatment. The Apicomplexan parasite Toxoplasma gondii modifies the levels of microRNAs and mRNAs in infected host cells by modulating the innate and adaptive immune responses, facilitating its survival within the host. Some studies have shown that microRNAs are promising molecular markers for developing diagnostic tools for human toxoplasmosis. MicroRNAs can be detected in human specimens collected using non-invasive procedures. changes in the circulating host microRNAs have been associated with T. gondii infection in mice and ocular toxoplasmosis in humans. Besides, microRNAs can be amplified from samples using sensitive and molecular-specific approaches such as real-time PCR. This review presents recent findings of the role that microRNAs play during T. gondii infection and discuss their potential use of these small nuclei acid molecules to different approaches such as laboratory diagnosis, modulation of cell and tissue infected as other potential applications in human toxoplasmosis.

A simple method to generate PCR-RFLP typing profiles from DNA sequences in Toxoplasma gondii.

Multilocus polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) has been widely used to genotype microorganisms. Accumulation of data generated by this method has helped to reveal genetic diversity, population structure and transmission of many microbial pathogens. Advances in DNA sequencing technologies have made it possible to identify microorganisms by multilocus sequencing typing (MLST) or whole genome sequence typing (WGST) to reach high resolution of identification. While MLST and WGST are gradually replacing PCR-RFLP for genotyping, invaluable databases generated by the latter may not be easily linked to datasets generated by sequencing based methods. In addition, DNA sequences corresponding to PCR-RFLP markers are often deposited in public domains but not fully explored to infer the RFLP profile. To alleviate this problem, we developed a simple protocol that can generate PCR-RFLP profiles from DNA sequence data, therefore facilitating the integration of data generated by different typing methods. Here we used the protozoan parasite Toxoplasma gondii as an example to bridge different typing methods.

Free-range chickens from Santa Catarina state, southern Brazil, as asymptomatic intermediate hosts for Toxoplasma gondii clonal type I and typical Brazilian genotypes.

Chickens are a host that is very resistant to the development of clinical toxoplasmosis. Free-range chickens have been used to indirectly track environmental contamination with Toxoplasma gondii oocysts because they feed on the ground. This study evaluated the genetic diversity of T. gondii isolates from free-range chickens from Florianópolis island in Santa Catarina state, southern Brazil. Sera from 21 chickens were tested for IgG anti-T. gondii antibodies using the modified agglutination test (MAT). Tissue homogenates from the 11 seropositive birds (MAT titres ≥5) were bioassayed in mice. The four obtained isolates (TgCkBrSC1-4) were genotyped using 11 PCR-RFLP markers and 15 microsatellite markers (MS). Four genotypes were identified, three of which are typical Brazilian genotypes (ToxoDB-RFLP #26 and #53 were previously reported and #278 is new), and the other is the rare clonal type I genotype. This type I isolate was considered a variant according to MS analysis, with two atypical alleles, which emphasizes the genetic diversity of the parasite in Brazil. The genetic variability of T. gondii in South America may be related to the high occurrence of severe ocular and congenital toxoplasmosis in humans in this region. High human seroprevalence and frequency of ocular toxoplasmosis are reported in southern Brazil, but there is limited information on the T. gondii strains that are circulating in this region, so more studies should be conducted to identify the strains in different hosts and in human toxoplasmosis cases.