{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,8,18]],"date-time":"2026-08-18T16:05:12Z","timestamp":1787069112407,"version":"build-2736575974"},"reference-count":34,"publisher":"Microbiology Society","issue":"8","license":[{"start":{"date-parts":[[2026,8,18]],"date-time":"2026-08-18T00:00:00Z","timestamp":1787011200000},"content-version":"unspecified","delay-in-days":0,"URL":"http:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"DOI":"10.13039\/100006492","name":"Division of Intramural Research, National Institute of Allergy and Infectious Diseases","doi-asserted-by":"crossref","award":["5K01AI173385"],"award-info":[{"award-number":["5K01AI173385"]}],"id":[{"id":"10.13039\/100006492","id-type":"DOI","asserted-by":"crossref"}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":[],"published-print":{"date-parts":[[2026,8,18]]},"abstract":"\n Background.<\/jats:bold>\n Mycobacterium tuberculosis<\/jats:italic>\n (\n Mtb<\/jats:italic>\n ) genomic epidemiology often relies on culturing patient sputum, a time- and labour-intensive process. Hybrid capture approaches have been successfully used to enrich\n Mtb<\/jats:italic>\n DNA from complex clinical samples, yet the accuracy of variant identification from captured samples has not been systematically evaluated.\n <\/jats:p>\n \n Methods.<\/jats:bold>\n We created artificial strain mixtures of two well-characterized\n Mtb<\/jats:italic>\n isolates such that the minor strain comprised 0\u201350% of\n Mtb<\/jats:italic>\n DNA and serially diluted the\n Mtb<\/jats:italic>\n DNA into human DNA to simulate diagnostic samples with different sputum smear burdens (32\u2009samples). We also prospectively collected paired\n Mtb<\/jats:italic>\n diagnostic cultures and sputum submitted to a national diagnostic laboratory (seven sample pairs). We performed hybrid capture and Illumina whole-genome sequencing for all samples. For the artificial strain mixtures, we measured hybrid capture efficiency, the percentage of total reads mapping to\n Mtb<\/jats:italic>\n and performance of fixed and minority variant identification. For the diagnostic samples, we compared the number, identity and minor allele frequencies of minority variants identified in the cultured and hybrid-captured samples.\n <\/jats:p>\n \n Results.<\/jats:bold>\n In the artificial strain mixture experiment, hybrid capture efficiency was 97% when\n Mtb<\/jats:italic>\n comprised 0.01% of the input DNA. SNP identification via hybrid capture had a sensitivity \u226591%\u2009and precision \u226597% for\n Mtb<\/jats:italic>\n lineages 4.1.2.1 and 4.9, excluding PE\/PPE genes, when\n Mtb<\/jats:italic>\n comprised 0.01% of input DNA. Observed minor allele frequencies were closely correlated (\n r<\/jats:italic>\n =0.62\u2009to\n r<\/jats:italic>\n =0.79,\n P<\/jats:italic>\n <0.001) with input minor allele frequencies across all dilutions. Among paired diagnostic samples, hybrid capture efficiency was high, 95%. However, four of the seven captured sputum samples were overwhelmed with\n Pseudomonas<\/jats:italic>\n contamination, which comprised >25% of sequence reads. We did not detect a significant difference in the number of minority variants identified in cultured (median, 14; range, 10\u201318) and hybrid-captured samples (median: 17 variants, range 9\u201347,\n P<\/jats:italic>\n =0.25) and minor allele frequencies were correlated (\n r<\/jats:italic>\n =0.87,\n P<\/jats:italic>\n <0.001) outside of PE\/PPE genes.\n <\/jats:p>\n \n Conclusions.<\/jats:bold>\n Hybrid capture of diagnostic sputum samples efficiently recovers accurate\n Mtb<\/jats:italic>\n whole-genome sequences. Minority variant detection is robust in controlled mixtures but shows reduced concordance in clinical samples, highlighting the need for further validation in real-world settings.\n <\/jats:p>","DOI":"10.1099\/mgen.0.001806","type":"journal-article","created":{"date-parts":[[2026,8,18]],"date-time":"2026-08-18T15:18:20Z","timestamp":1787066300000},"source":"Crossref","is-referenced-by-count":0,"title":["Optimizing culture-free approaches to recover high-quality Mycobacterium tuberculosis genomic variation"],"prefix":"10.1099","volume":"12","author":[{"ORCID":"https:\/\/orcid.org\/0000-0003-0065-2204","authenticated-orcid":false,"given":"Katharine S.","family":"Walter","sequence":"first","affiliation":[{"name":"Division of Epidemiology, University of Utah, Salt Lake City, UT, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Paulo C\u00e9sar Pereira","family":"Dos Santos","sequence":"additional","affiliation":[{"name":"College of Medicine, Federal University of Mato 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