The FT MTB assay is a new semi-automated assay for detection of MTBC giving results within 3-4 hours. Similar to other semi-automated systems like COBAS TaqMan MTB, different workstations are required including DNA extraction, preparation of PCR mixes and amplification/detection. DNA extraction can be done automated using the GenoXtract instrument requiring only minimal hands-on time. Furthermore, amplification and detection is performed fully-automated in the Fluorocycler instrument; the handling of the instrument is simple and the analysis software designed user-friendly. Including all work steps, hands-on time for 12 specimens adds up to approximately 30 min being comparable to other semi-automated systems . The Fluorocycler system is suitable for low numbers of samples as well as for large series, as up to 8 Fluorocycler instruments (each with 12 positions) can be operated simultaneously by one control computer. In comparison, the Xpert MTB/RIF system enables hands-on time of less than 3 min per specimens being of advantage only when handling a small number of samples.
The aim of the present study was to assess the performance characteristics of the FT MTB assay in a routine setting of a German mycobacteriology laboratory. FT MTB results from 978 respiratory specimens were compared to culture and clinical diagnosis as “gold standard”. The specificity (98.9%) of FT MTB was in the range of that of other NAATs (98.4%-99.7%, mean 99.1%) [2, 4–6, 11, 12]. In particular, no cross-reactivity with 18 specimens growing NTM was observed. Ten positive FT MTB results remained unresolved, even after review of patients` clinical data. It remains elusive whether these false-positive outcomes are caused by residual MTBC DNA despite absence of active disease, as speculated before , or by unspecific amplification. Although reported as positive by the manufacturers’ analysis software, false-positive FT MTB tests seem to give lower MTBC-specific melting curve peaks than true-positive ones. Discrepant positive results in the “zone of low-positivity” are reported also for other NAAT systems [14–16]. A repeat testing is, therefore, generally recommended for specimens with low positivity values before transferring a final laboratory report to the clinical doctor.
The overall sensitivity of FT MTB reached 88.1% being in the upper range (63.2 – 95.0%, mean 84.2%) of levels reported for other commercial NAATs like BD ProbeTec ET (63.2-86.2%) [6, 11], COBAS Taqman MTB (81.1-91.5%) [4, 5], GeneProbe AMTD (95%) , Speed-oligo direct MTB (76%)  or Xpert MTB/RIF (88.0-92.2%) [2, 18].
Considering smear-positive TB specimens only, sensitivity and PPV reached 100%. In comparison, sensitivity levels reported for other commercial assays were 95.5% to 100% [2, 5, 6, 17–19]. A high positive predictive value for smear-positive specimens is particularly important in settings where NTM are common. Particularly in fully industrialized countries, NTM are increasingly isolated from clinical specimens and reported as cause of opportunistic infections mainly in immunocompromised individuals [20–22]. Therefore, modern NAAT assays should rapidly and reliably differentiate between TB and NTM infection, particularly in patients with immunodeficiency or in patients with first smear-positive sputa and unspecific clinical signs of TB. The FT MTB obviously fulfils this requirement.
Considering smear-negative TB specimens only, the sensitivity of FT MTB was 56.3%. Of the smear-negative TB samples which were missed by FT MTB, four exhibited positive culture results only after ≥ 24 days of incubation, indicating a paucibacillary nature of the specimens. This doesn’t mitigate the fact, that we found a sensitivity level for smear-negative samples which was in the lower range of values reported for other NAATs (49.1–86.7%, mean 64.1%) [2, 4, 6, 12, 17, 18]. However, the low absolute number of smear-negative TB samples (n = 16) included in the present study might have biased the outcomes for such samples. Consequently, the sensitivity of FT MTB for paucibacillary specimens should be re-assessed in further, preferably multi-centre studies with higher numbers of smear-negative TB samples.