Assessment of Mitochondrial and Nuclear Genes for Molecular Detection during Early Trichinella spiralis Infection

Abstract

Early serological diagnosis of trichinellosis is affected by a long-term immunological "silent" period following infection. This emphasises the need for the development of sensitive diagnostic methods to be used when antibodies cannot be detected. In this study, we assessed the usefulness of three DNA sequences as a direct diagnosis method to detect early infection with Trichinella spiralis in peripheral blood by SYBR green real-time PCR using a murine model. Primers were designed from a nuclear repetitive DNA element (Rep), the nuclear ribosomal internal transcribed spacer 2 (ITS2) region, and the mitochondrial large subunit of the ribosomal RNA gene (LSU). CF-1 mice were orally inoculated with 500 muscle larvae of T. spiralis and molecular detection was assessed in blood samples between 1 and 20 days post-inoculation (pi). In addition, antibody detection was evaluated every 5 days by ELISA using excretory-secretory antigens. Results showed that the Rep primers were found to amplify between 5 and 19 days pi and the ITS2 and LSU primers between 6 and 15 days pi, whereas the antibodies were detected at 30 days pi. Therefore, this molecular system could be a useful tool for the detection of early T. spiralis infection, in particularly by using the repetitive DNA element.