Rapid identification of Mycobacteria to the species level using INNO-LiPA Mycobacteria, a reverse hybridization assay

INNO-LiPA Mycobacteria (LiPA; Innogenetics, Zwijnaarde, Belgium) is a kit f or the simultaneous detection and identification of Mycobacterium species i n culture and identifies the Mycobacterium tuberculosis complex, the M. avi um complex (MAC), and the following Mycobacterium species: M. kansasii, M. avium, M. intracellulare, M. scrofulaceum, M. gordonae, M. xenopi, and the M. chelonae-M. abscessus complex. The assay, which targets the 16S-23S rRNA spacer region, was evaluated on 157 mycobacterial strains that had been id entified by conventional techniques and PCR-restriction enzyme analysis of the hsp65 gene (PRA). Forty-seven reference strains consisting of 37 differ ent species and 110 human clinical isolates were submitted to the test, and all were hybridized with the Mycobacterium genus probe (MYC) on the LiPA s trip (100% sensitivity). Ninety-four isolates hybridized to their correspon ding species- or complex-specific probes; only one isolate phenotypically i dentified as M. gordonae did not react with its specific probe (99.4% accur acy). Thirty-seven MAC strains were phenotypically identified to the comple x level and to the species level by LiPA as M. avium (n = 18) or M. intrace llulare (n = 7) or as belonging to the M. avium-M. intracellulare-M. scrofu laceum complex (n = 12). Of the last 12 strains, 10 had M. avium PRA patter ns and 2 had M. intracellulare PRA patterns. Three isolates that had been i dentified as a single species by conventional identification were proven to be mixed cultures by the LiPA assay. The whole procedure can be performed in 1 working day, starting with the supernatant of a small amount of bacter ial mass that had been treated by freezing and then boiling.