A novel high performing multiplex immunoassay Multi-HTLV for serological confirmation and typing of HTLV infections

Background: Human T-Cell Lymphotropic Viruses (HTLV) type 1 and type 2 account for an estimated 5 to 10 million infections worldwide and are transmitted through breast feeding, sexual contacts and contaminated cellular blood components. HTLV-associated syndromes are considered as neglected diseases for which there are no vaccines or therapies available, making it particularly important to ensure the best possible diagnosis to enable proper counselling of infected persons and avoid secondary transmission. Although high quality antibody screening assays are available, currently available confirmatory tests are costly and have variable performance, with high rates of indeterminate and non-typable results reported in many regions of the world. The objective of this project was to develop and validate a new high-performance multiplex immunoassay for confirmation and discrimination of HTLV-1 and HTLV-2 strains. Methodology/Principal findings: The multiplex platform was used first as a tool to identify suitable antigens and in a second step for assay development. With data generated on over 400 HTLV-positive blood donors sourced from USA and French blood banks, we developed and validated a high-precision interpretation algorithm. The Multi-HTLV assay demonstrated very high performance for confirmation and strain discrimination with 100% sensitivity, 98.1% specificity and 100% of typing accuracy in validation samples. The assay can be interpreted either visually or automatically with a colorimetric image reader and custom algorithm, providing highly reliable results. Conclusions/Significance: The newly developed Multi-HTLV is very competitive with currently used confirmatory assays and reduces considerably the number of indeterminate results. The multiparametric nature of the assay opens new avenues to study specific serological signatures of each patient, follow the evolution of infection, and explore utility for HTLV disease prognosis. Improving HTLV diagnostic testing will be critical to reduce transmission and to improve monitoring of seropositive patients. Author summary: HTLV viruses are responsible for more than 10 million cases of infection worldwide. The infection is considered as a neglected disease due to lack of vaccines and treatments. Accurate diagnosis is crucial for counselling infected persons and prevention of secondary transmissions. In spite of the development of excellent serological screening assays, many cases of indeterminate and untyped results are still regularly reported and their infection status remain uncertain. To address the need of more precise diagnosis, we have developed a new cutting-edge in-vitro diagnostic confirmation test, named Multi-HTLV, which has been validated on a large panel of HTLV samples. The test is a multiplex immunoassay allowing powerful detection of antibodies against HTLV through combination of a set of selective and validated virus-specific antigens in a blood sample. The Multi-HTLV assay increases the reliability of HTLV diagnostics and strain typing thanks to a high precision mathematical algorithm.