Blood screening nucleic acid amplification tests for human immunodeficiency virus Type 1 may require two different amplification targets

Abstract

Five cases of human immunodeficiency virus Type 1 (HIV-1) RNA-positive blood donations are described that escaped detection by three different CE-marked nucleic acid amplification technique (NAT) screening assays. These events were associated with two HIV-1 transmissions to recipients of blood components. The implicated NAT assays are monotarget assays and amplify in different viral genome regions (group-specific antigen or long terminal repeat). Investigations into the cause of the false-negative test results were initiated. Plasma specimens of the five NAT false-negative cases were comparatively investigated in 12 CE-marked HIV-1 NAT systems of differing design. The relative amplification efficiency for the HIV-1 variant was determined for each assay. Sequencing of the variants in the region targeted by each false-negative NAT assay allowed comparison with the respective primers and probes. Some of the NAT assays designed in a similar way to false-negative monotarget NATs also revealed deficiencies in detecting the viral variants. In each case sequencing of the assay target region in the variants demonstrated mismatches with primers and probes used by the assays. Some dual-target assays showed decreased amplification efficiency, but not false-negative results. HIV is characterized by its rapid evolution of new viral variants. The evolution of new sequences is unpredictable; NAT screening assays with a single target region appear to be more vulnerable to sequence variations than dual-target assays. Based on this experience with false-negative tests results by monotarget NAT assays, the Paul-Ehrlich-Institut is considering requesting dual-target NAT assays for HIV-1 blood donation screening in Germany.