{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"institution":[{"name":"medRxiv"}],"indexed":{"date-parts":[[2026,5,30]],"date-time":"2026-05-30T07:03:48Z","timestamp":1780124628516,"version":"3.54.0"},"posted":{"date-parts":[[2026,2,3]]},"group-title":"Infectious Diseases (except HIV\/AIDS)","reference-count":39,"publisher":"openRxiv","license":[{"start":{"date-parts":[[2026,2,3]],"date-time":"2026-02-03T00:00:00Z","timestamp":1770076800000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/www.medrxiv.org\/about\/FAQ#license"}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":[],"accepted":{"date-parts":[[2026,5,27]]},"abstract":"<jats:title>Abstract<\/jats:title>\n                <jats:sec>\n                  <jats:title>Background<\/jats:title>\n                  <jats:p>Stool-based molecular tests are a noninvasive option for pediatric tuberculosis (TB) diagnosis, but have lower sensitivity compared to sputum-based tests. Untargeted metagenomic sequencing (mNGS) on stool could improve sensitivity and identify new gene targets for molecular testing.<\/jats:p>\n                <\/jats:sec>\n                <jats:sec>\n                  <jats:title>Methods<\/jats:title>\n                  <jats:p>\n                    We performed shotgun mNGS on DNA isolated from stool samples of children undergoing assessment for pulmonary TB in Uganda. We defined the performance of mNGS to identify\n                    <jats:italic>Mycobacterium tuberculosis<\/jats:italic>\n                    (\n                    <jats:italic>Mtb<\/jats:italic>\n                    ) against a microbiological reference standard (MRS, TB if sputum Xpert Ultra or culture positive) and a composite reference standard (TB if confirmed or unconfirmed TB). We also compared accuracy of mNGS against the stool-based Xpert Ultra test. Finally, we identified enriched genomic loci among\n                    <jats:italic>Mtb<\/jats:italic>\n                    classified reads.\n                  <\/jats:p>\n                <\/jats:sec>\n                <jats:sec>\n                  <jats:title>Results<\/jats:title>\n                  <jats:p>We analyzed 176 stool samples of children with a median age of 3.6 years (IQR, 1-6 years). Against the MRS, the sensitivities of mNGS with positive TB defined as \u2265 1, 2, or 5 sequence fragments were 35.5% (95% CI 19%-55%), 25.7% (12%-45%), and 19.4% (13%-25%) respectively, and specificities 92.64% (87%-96%), 97% (93%-99%), and 99.3% (96%-100%). Stool Xpert Ultra had similar sensitivity (22.6%) to stool mNGS considering all samples tested. In a head-to-head comparison, stool mNGS had lower sensitivity than stool Xpert Ultra (38.5% vs. 53.8%, difference -15.3%, 95% CI 14-68 to 25-81). mNGS utilized rRNA, virulence proteins and membrane proteins not targeted in current PCR-based platforms.<\/jats:p>\n                <\/jats:sec>\n                <jats:sec>\n                  <jats:title>Conclusions<\/jats:title>\n                  <jats:p>Metagenomic sequencing of stool DNA did not increase sensitivity of TB detection, but identified novel targets for molecular testing that may support development of more sensitive tests.<\/jats:p>\n                <\/jats:sec>","DOI":"10.64898\/2026.01.29.26345155","type":"posted-content","created":{"date-parts":[[2026,2,3]],"date-time":"2026-02-03T15:50:17Z","timestamp":1770133817000},"source":"Crossref","is-referenced-by-count":0,"title":["Evaluating metagenomic sequencing as a stool-based diagnostic in children with presumptive TB in Uganda"],"prefix":"10.64898","author":[{"given":"Carolina","family":"Agudelo","sequence":"first","affiliation":[{"name":"Division of Infectious Disease and Vaccinology, School of Public Health, University of California, Berkeley, Berkeley, CA, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Moses","family":"Nsereko","sequence":"additional","affiliation":[{"name":"World Alliance for Lung and Intensive Care Medicine in Uganda, Kampala, Uganda"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Aggrey","family":"Ainebyona","sequence":"additional","affiliation":[{"name":"World Alliance for Lung and Intensive Care Medicine in Uganda, Kampala, Uganda"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Alfred","family":"Andama","sequence":"additional","affiliation":[{"name":"World Alliance for Lung and Intensive Care Medicine in Uganda, Kampala, Uganda"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Robert","family":"Castro","sequence":"additional","affiliation":[{"name":"Pulmonary and Critical Care Medicine, University of California, San Francisco, San Francisco, CA, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Sarrah Rose","family":"Mikhail Leung","sequence":"additional","affiliation":[{"name":"Division of Infectious Disease and Vaccinology, School of Public Health, University of California, Berkeley, Berkeley, CA, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Jascent","family":"Nakafeero","sequence":"additional","affiliation":[{"name":"World Alliance for Lung and Intensive Care Medicine in Uganda, Kampala, Uganda"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Gertrude","family":"Nannyonga","sequence":"additional","affiliation":[{"name":"World Alliance for Lung and Intensive Care Medicine in Uganda, Kampala, Uganda"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Kevin","family":"Nolan","sequence":"additional","affiliation":[{"name":"Pulmonary and Critical Care Medicine, University of California, San Francisco, San Francisco, CA, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Lucas","family":"Teran","sequence":"additional","affiliation":[{"name":"Division of Infectious Disease and Vaccinology, School of Public Health, University of California, Berkeley, Berkeley, CA, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Peter","family":"Wambi","sequence":"additional","affiliation":[{"name":"World Alliance for Lung and Intensive Care Medicine in Uganda, Kampala, Uganda"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Mark G.","family":"Young","sequence":"additional","affiliation":[{"name":"Division of Infectious Disease and Vaccinology, School of Public Health, University of California, Berkeley, Berkeley, CA, USA"},{"name":"Center for Computational Biology, University of California, Berkeley, Berkeley, CA, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Midori","family":"Kato-Maeda","sequence":"additional","affiliation":[{"name":"Pulmonary and Critical Care Medicine, University of California, San Francisco, San Francisco, CA, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Adithya","family":"Cattamanchi","sequence":"additional","affiliation":[{"name":"Pulmonary Diseases and Critical Care, University of California Irvine, Orange, CA, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Devan","family":"Jaganath","sequence":"additional","affiliation":[{"name":"Pediatric Infectious Disease, University of California, San Francisco, San Francisco, CA, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Eric","family":"Wobudeya","sequence":"additional","affiliation":[{"name":"World Alliance for Lung and Intensive Care Medicine in Uganda, Kampala, Uganda"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Ashley R.","family":"Wolf","sequence":"additional","affiliation":[{"name":"Division of Infectious Disease and Vaccinology, School of Public Health, University of California, Berkeley, Berkeley, CA, USA"},{"name":"Center for Computational Biology, University of California, Berkeley, Berkeley, CA, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"54368","reference":[{"key":"2026053000000854000_2026.01.29.26345155v2.1","unstructured":"WHO Departmental update. 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