OR15

1:KNCV tuberculosis foundation, The Hague, The Netherlands; 2:National Institute for Public Health and the Environment, Bilthoven, The Netherlands; 3:London School of Hygiene & Tropical Medicine, London, United Kingdom; 4:KNCV Tuberculosis Foundation, Bishkek, Kyrgyzstan; 5:National Center for Phthisiology, Bishkek, Kyrgyzstan

The World Health Organization (WHO) “Catalogue of mutations in Mycobacterium tuberculosis complex (MTBC)” was updated in 2023. The limited availability of genomic-/phenotypic data restricts the accuracy of genotypic drug susceptibility prediction for newer anti-tuberculosis drugs. To address this, rules were added to the catalogue to provide guidance for these drugs, including pretomanid (PTM) and delamanid (DLM). However, the interpretation of these rules may be complex. For example, the M. tuberculosis H37rv reference genome contains 2.7 (imperfect) repeats at the end of the fbiC gene. Yet, we encountered MTBC strains with repeat deletions and insertions. Due to variability in the number of repeats, sequencing platform independent mapping issues were encountered for both Nanopore targeted next-generation- and Illumina short read whole genome sequencing. Reads that were too short to read through the 2.7 repeat area were incorrectly mapped to the fbiC reference, triggering the catalogue rule associating loss of function (LoF) of the gene with PTM/DLM resistance. Nevertheless, further analysis revealed that in both single and double repeat deletions, the remaining repeat’s stop codon replaced the fbiC stop codon leaving the coding region unchanged. Thus, variability in the number of repeats at the end of fbiC does not cause LoF nor does it impact susceptibility. Our findings underline the importance of rapid communication of exceptions to the WHO catalogue rules, so data analysis pipelines can be updated and systematic errors eliminated, and the need for external quality control samples or files (allowing digital dissemination), designed to validate the implementation of new rules.