P16

Delamanid (DLM) is an add-on drug for the treatment of multidrug-resistant tuberculosis since 2014 and is currently used in experimental arms of clinical trials. A knowledge gap remains on the correlation of DLM resistance conferring mutations and minimal inhibitory concentration (MIC). We recently initiated DLM MIC testing in the resazurin microtiter assay (REMA) with a tentative critical concentration of 0.06 ug/ml recommended for MGIT. This validation showed the majority of  susceptible reference strains falling out of the expected range (0.002 – 0.06 ug/ml). Thus we investigated several parameters including brand of DLM powder, resazurin concentration, inoculum preparation method and schedules for reading. Higher MICs than expected were still observed across all susceptible reference strains tested irrespective of the powder origin, resazurin concentration (0.01% instead of 0.02%) or reading schedule (24h vs 48h). However, a lower inoculum and slightly different inoculum preparation, 10⁻² McFarland 0.5 suspension instead of a 10⁻¹ McFarland 1.0, showed MICs as expected for DLM susceptible and resistant reference strains. When clinical isolates were tested using this optimised procedure, one third resulted  in invalid MICs due to no growth in the growth controls. Extended plate incubation re-validated the MICs  but would require macroscopic growth observation ideally in round bottom plates for the optimal timepoint to add resazurin. Colour interpretation however is difficult in round bottom plates compared to flat bottom plates. Further efforts aim to optimise DLM REMA testing especially for slow growing clinical isolates.