Comparative evaluation of real-time PCR and ELISA for the detection of human fascioliasis.

Fascioliasis is a zoonotic parasitic infection caused by Fasciola species in humans and animals. Despite significant advances in vaccination and new therapeutic agents, little attention has been paid to validating methods for the diagnosis of fascioliasis in humans. Serological techniques are convenient assays that significantly improves the diagnosis of Fasciola infection. However, a more sensitive method is required. The aim of this study was to compare the Real-Time PCR technique with the indirect-ELISA for the detection of Fasciola hepatica in human. Using a panel of sera from patients infected with Fasciola hepatica (n = 51), other parasitic infections (n = 7), and uninfected controls (n = 12), we optimized an ELISA which employs an excretory-secretory antigens from F. hepatica for the detection of human fascioliasis. After DNA extraction from the samples, molecular analysis was done using Real-Time PCR technique based on the Fasciola ribosomal ITS1 sequence. Of 70 patient serum samples, 44 (62.86%) samples were identified as positive F. hepatica infection using ELISA and Real-Time PCR assays. There was no cross-reaction with other parasitic diseases such as toxoplasmosis, leishmaniasis, taeniasis, hydatidosis, trichinosis, toxocariasis, and strongyloidiasis. The significant difference between the agreement and similarity of the results of patients with indirect ELISA and Real-Time PCR was 94.4% and 99.2%, respectively (Cohen's kappa ≥ 0.7; P = 0.02). Based on the Kappa agreement findings, the significant agreement between the results of ELISA and Real-Time PCR indicates the accuracy and reliability of these tests in the diagnosis of F. hepatica in humans.

Evaluation of Semi-Nested PCR Compared with Indirect-ELISA to Diagnose Human Fasciolosis.

Background: We aimed to compare semi-nested PCR with indirect ELISA to diagnose human fasciolosis. Methods: Overall, 70 serum samples were collected from different areas in Iran suspected for fascioliasis. Individuals were classified based on diagnostic of fascioliasis and habitat in endemic areas. Finally, all serum samples were tested by indirect ELISA (using secretory excretory antigen) and semi-nested PCR (using ITS1 gene). The study was conducted in the School of Publish Health, Tehran University of Medical Sciences, Iran in 2021. Results: Significant differences were found between agreement and similarity of patients' results of indirect ELISA and semi-nested PCR 94.46% and 98.4% respectively (Cohen's kappa ≥0.6; P-value≤0.05). No cross-reactions were observed with other parasitic diseases (toxocariasis, hydatidosis, strongyloidiasis, toxoplasmosis, cutaneous leishmaniasis, taeniasis and trichinosis). 69.84% of samples were positive by both techniques. In addition, the percentage of agreement and similarity between the results of the two techniques based on habitat in endemic areas was 88.9-100% and 97.7-100%, respectively (Cohen's kappa ≥0.6; P-value≤0.05). Conclusion: Semi-nested PCR could be a suitable method for following up on patients' treatment and a confirmatory method for ELISA as for diagnosis of human fascioliasis.

Changes in the expression of miR-103a and miR-21: a functional diagnosis of toxocariasis in rats.

Introduction. Toxocariasis is a zoonotic parasitic disease caused by migrating nematode worms, Toxocara species larvae, within tissues. MicroRNAs (miRNAs) are small RNA molecules that regulate gene expression at a post-transcriptional level.Hypothesis/Gap Statement. miRNA-based diagnostic biomarkers for toxocariasis are emerging, but there is limited information about the role of many miRNAs and a more detailed diagnostic evaluation of miRNA expression patterns is needed to understand their immunobiological function.Aim. We investigated the expression levels of circulating miRNA 21 and miRNA 103a as potential biomarkers for the prediction and diagnosis of toxocariasis in Wistar rats infected with Toxocara canis.Methodology. Thirty Wistar rats were inoculated orally with 2500 T. canis embryonated eggs via gavage. Serum samples were collected from infected animals and were tested against T. canis antigens for 60 days post-infection. The plasma samples were isolated for quantitative real-time PCR (qPCR) assays and qPCR was used to assess transcription levels of miRNA 21 and miRNA 103a.Results. The prevalence of anti-Toxocara IgG was detected in 7/30 (23.3 %) infected rats. Molecular analysis of miRNAs 21 and 103a showed that expression levels of miRNAs in both groups of Toxocara-positive and negative samples were the same without significant association. The ratio of housekeeping gene expression (U6) to gene expression of miRNAs 21 and 103a indicated the rate of change (1/1.38 ≈ 0.75 and 1/0.751 ≈ 1.3, respectively).Conclusion. Our study revealed that miRNAs 21 and 103a might play fundamental roles as biomarkers and diagnostic tools for toxocariasis. However, the changes in expression of these miRNAs were not adequate to be used as biomarkers in diagnosis.

Human Toxocariasis in individuals with blood disorders and cancer patients: the first seroepidemiological study in Iran.

Toxocara is one of the most prevalent nematodes in Iran, which infect humans as an intermediate host. Infection complications result from the larva migration. Human toxocariasis prevalence was various in Iran according to the area of study and population. This study was designed to evaluate the seropositivity of Toxocara IgG in patients with blood disorders and cancer patients in southwest Iran. Moreover, the study of the associated risk factors for this infection. A total of 1122 serum samples, from February 8, 2019 to August 21, 2019, including 600 healthy individuals and 522 individuals with cancer and blood disorders patients were collected. Serum samples were collected for detection of Toxocara IgG by using ELISA (Enzyme-Linked Immunosorbent Assay) kit. Sociodemographic data of all participants were collected and examined to determine their association with the infection. Out of 101 individuals with white blood cell disorders (5.94%), red blood cell disorders (7.48%) and cancer patients (11.06%) were seropositive for Toxocara IgG antibodies. The infection rate among all study population revealed that (10.76%) were positive for Toxocara IgG. This study showed the fundamental role of contact with pets and infection in groups with blood cell disorders (P-value ≤ 0.05%); while in cancer patients the association wasn't significant. Other factors such as age, location of residence, and sex showed that the association with this infection wasn't significant.