{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"institution":[{"name":"medRxiv"}],"indexed":{"date-parts":[[2026,1,16]],"date-time":"2026-01-16T13:08:48Z","timestamp":1768568928346,"version":"3.49.0"},"posted":{"date-parts":[[2024,6,22]]},"group-title":"Infectious Diseases (except HIV\/AIDS)","reference-count":29,"publisher":"openRxiv","content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":[],"accepted":{"date-parts":[[2024,6,22]]},"abstract":"<jats:title>Abstract<\/jats:title>\n                <jats:p>\n                  Tongue swabs represent a potential alternative to sputum as a sample type for detecting pulmonary tuberculosis (TB) using molecular diagnostic tests. The methods used to process tongue swabs for testing in the WHO-recommended Xpert MTB\/RIF Ultra (Xpert Ultra) assay vary greatly. We aimed to identify the optimal method for processing diagnostic tongue swabs for subsequent testing by Xpert Ultra. We compared four methods for treating dry tongue swabs with Xpert Ultra sample reagent (SR) mixed with various concentrations of Tris-EDTA-Tween (TET), to treatment with SR alone or to a commonly used SR-free heat-inactivation protocol. In each condition, swabs obtained from volunteers without TB were placed into test buffer spiked with known amounts of\n                  <jats:italic>Mycobacterium tuberculosis<\/jats:italic>\n                  (\n                  <jats:italic>Mtb<\/jats:italic>\n                  ) strain H37Rv-mc\n                  <jats:sup>2<\/jats:sup>\n                  6230. Swabs processed with 1:1 diluted SR buffer had the lowest\n                  <jats:italic>Mtb<\/jats:italic>\n                  limit of detection (LOD) at 22.7 CFU\/700 \u00b5l (95% CI 14.2-31.2), followed by 2:1 diluted SR buffer at 30.3 CFU\/700 \u00b5l (95% CI 19.9-40.7), neat SR at 30.9 CFU\/700 \u00b5l (95% CI 21.5-40.3) and SR prefilled in the Xpert Ultra at 57.1 CFU\/700 \u00b5l (95% CI 42.4-71.7). Swabs processed using the heat-based protocol had the highest LOD (77.6 CFU\/700 \u00b5l; 95% CI 51.2\u2013104.0). Similar findings were observed for LOD of RIF-susceptibility. Assay sensitivity using the 2:1 diluted SR buffer did not vary considerably in the presence of sputum matrix or phosphate buffer saline. Further studies are needed to assess the performance of this processing protocol in a clinical setting.\n                <\/jats:p>\n                <jats:sec>\n                  <jats:title>Importance<\/jats:title>\n                  <jats:p>\n                    Xpert MTB\/RIF Ultra (Xpert Ultra) is approved by the World Health Organization for the diagnosis of tuberculosis (TB). This test is typically performed using sputum specimens obtained from people with presumptive TB. In order to inactivate\n                    <jats:italic>Mtb<\/jats:italic>\n                    and aid liquefaction, sputum must be mixed with Xpert SR prior to transfer into the Xpert Ultra. However, some people under evaluation for TB are unable to produce sputum. Alternative sample types for TB diagnosis would therefore be of value. Oral-swabs, including tongue-swabs have shown promise, but there are technical challenges associated with sample processing. In this study, several new tongue swab processing conditions were evaluated, utilizing SR, either neat or diluted in buffer. The ability of Xpert Ultra to detect TB was improved under these conditions compared with the previously published heat-processing method (1\u20133), processing steps were simplified, and technical challenges were overcome.\n                  <\/jats:p>\n                <\/jats:sec>","DOI":"10.1101\/2024.06.20.24309244","type":"posted-content","created":{"date-parts":[[2024,6,22]],"date-time":"2024-06-22T11:50:18Z","timestamp":1719057018000},"source":"Crossref","is-referenced-by-count":0,"title":["Optimal processing of tongue swab samples for Mycobacterium tuberculosis detection by the Xpert MTB\/RIF Ultra assay"],"prefix":"10.64898","author":[{"given":"Gayatri Shankar","family":"Chilambi","sequence":"first","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Robert","family":"Reiss","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Naranjargal","family":"Daivaa","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-5217-6142","authenticated-orcid":false,"given":"Padmapriya","family":"Banada","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Margaretha","family":"De Vos","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Adam","family":"Penn-Nicholson","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-5196-1571","authenticated-orcid":false,"given":"David","family":"Alland","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]}],"member":"54368","reference":[{"key":"2024062415000715000_2024.06.20.24309244v1.1","doi-asserted-by":"crossref","first-page":"e0042122","DOI":"10.1128\/jcm.00421-22","article-title":"Accuracy of Tongue Swab Testing Using Xpert MTB-RIF Ultra for Tuberculosis Diagnosis","volume":"60","year":"2022","journal-title":"J Clin Microbiol"},{"key":"2024062415000715000_2024.06.20.24309244v1.2","doi-asserted-by":"crossref","first-page":"e45","DOI":"10.1016\/S2214-109X(23)00469-2","article-title":"Oral swabs with a rapid molecular diagnostic test for pulmonary tuberculosis in adults and children: a systematic review","volume":"12","year":"2024","journal-title":"Lancet Glob Health"},{"key":"2024062415000715000_2024.06.20.24309244v1.3","doi-asserted-by":"publisher","DOI":"10.1101\/2023.10.10.23296833"},{"key":"2024062415000715000_2024.06.20.24309244v1.4","unstructured":"World Health Organization. Global Tuberculosis Report 2023. https:\/\/www.who.int\/teams\/global-tuberculosis-programme\/tb-reports\/global-tuberculosis-report-2023. Accessed May 2024."},{"key":"2024062415000715000_2024.06.20.24309244v1.5","unstructured":"World Health Organization. WHO consolidated guidelines on tuberculosis. Module 3: diagnosis \u2013 rapid diagnostics for tuberculosis detection, third edition. https:\/\/www.who.int\/publications\/i\/item\/9789240089488. Accessed May 2024."},{"key":"2024062415000715000_2024.06.20.24309244v1.6","doi-asserted-by":"crossref","first-page":"e0236020","DOI":"10.1128\/JCM.02360-20","article-title":"A Look Inside: Oral Sampling for Detection of Non-oral Infectious Diseases","volume":"59","year":"2021","journal-title":"J Clin Microbiol"},{"key":"2024062415000715000_2024.06.20.24309244v1.7","doi-asserted-by":"publisher","DOI":"10.1086\/651490"},{"key":"2024062415000715000_2024.06.20.24309244v1.8","doi-asserted-by":"crossref","unstructured":"Mendelson M . 2007. 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Tongue dorsum swab processing for the detection of tuberculosis with Cepheid Xpert\u00ae MTB\/RIF Ultra. University of Washington."},{"key":"2024062415000715000_2024.06.20.24309244v1.20","doi-asserted-by":"publisher","DOI":"10.1128\/JCM.01463-09"},{"key":"2024062415000715000_2024.06.20.24309244v1.21","doi-asserted-by":"crossref","unstructured":"Cox H , Workman L , Bateman L , Franckling-Smith Z , Prins M , Luiz J , Van Heerden J , Ah Tow Edries L , Africa S , Allen V , Baard C , Zemanay W , Nicol MP , Zar HJ . 2022. Oral Swab Specimens Tested With Xpert MTB\/RIF Ultra Assay for Diagnosis of Pulmonary Tuberculosis in Children: A Diagnostic Accuracy Study. 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