No extraction? No problem. Direct to PCR processing of tongue swabs for diagnosis of tuberculosis disease as an alternative to sputum collection
ABSTRACT Sputum collection and testing for tuberculosis (TB) have been problematic due to the potential for aerosolization, difficulty in generating a quality sample, and complex DNA extraction methods. Tongue swabs are inexpensive, minimally invasive, and a promising alternative to sputum collectio...
| Main Authors: | , , , , , , |
|---|---|
| Format: | Article |
| Language: | English |
| Published: |
American Society for Microbiology
2024-01-01
|
| Series: | Microbiology Spectrum |
| Subjects: | |
| Online Access: | https://journals.asm.org/doi/10.1128/spectrum.03107-23 |
| _version_ | 1827383628835848192 |
|---|---|
| author | Charlotte L. Ahls Duncan Emsweller Seth J. Helfers Xin Niu Douglas Wilson Leah R. Padgett Paul K. Drain |
| author_facet | Charlotte L. Ahls Duncan Emsweller Seth J. Helfers Xin Niu Douglas Wilson Leah R. Padgett Paul K. Drain |
| author_sort | Charlotte L. Ahls |
| collection | DOAJ |
| description | ABSTRACT Sputum collection and testing for tuberculosis (TB) have been problematic due to the potential for aerosolization, difficulty in generating a quality sample, and complex DNA extraction methods. Tongue swabs are inexpensive, minimally invasive, and a promising alternative to sputum collection. We investigated the diagnostic accuracy of tongue swabs for TB diagnosis using the Truenat MTB Plus assay from Molbio Diagnostics with a direct to PCR processing method. Four tongue swabs were collected per participant using two nylon-flocked swabs and two spun polyester swabs. Following tongue swab sample collection, participants also provided two sputum samples, which were tested by Cepheid Xpert MTB/RIF Ultra or culture. Of the 81 participants enrolled, 24 (30%) were positive for TB disease by sputum Ultra. Using the Truenat MTB Plus test, tongue swabs had 54% (52/96) sensitivity and 99% (218/220) specificity compared to sputum Ultra. Crude lysate was also tested using an in-house qPCR assay, which allowed for increased sample input. Using this method, tongue swabs had 70% (67/96) sensitivity and 94% (216/224) specificity. Mycobacterium tuberculosis (MTB) sample quantification using digital PCR yielded between 20 copies (minimum) and 34,000 copies (maximum) of MTB per swab. In addition, serially collected tongue swabs resulted in similar levels of detected MTB, and spun polyester swabs performed equivalently to nylon-flocked swabs. Overall, this study demonstrates that tongue swab samples are compatible with the Truenat MTB testing platform and that a direct to PCR method is a viable diagnostic solution. IMPORTANCE Tuberculosis (TB) remains one of the world’s leading infectious disease killers, despite available treatments. Although highly sensitive molecular diagnostics are available, expensive equipment and poor infrastructure have hindered their implementation in low-resource settings. Furthermore, the collection of sputum poses challenges as it is difficult for patients to produce and creates dangerous aerosols. This manuscript explores tongue swabs as a promising alternative to sputum collection. While previous studies have explored the sensitivity of tongue swabs as compared to sputum, existing literature has not addressed the need to standardize and simplify laboratory processing for easy implementation in high TB burden areas. This manuscript provides the first evidence that detection of TB from a tongue swab is possible without the use of DNA extraction or purification steps. The data provided in this manuscript will improve the collection and testing of tongue swabs for the diagnosis of TB disease. |
| first_indexed | 2024-03-08T14:44:06Z |
| format | Article |
| id | doaj.art-919d7b6d2c8b48dba2c731b4cbddb16c |
| institution | Directory Open Access Journal |
| issn | 2165-0497 |
| language | English |
| last_indexed | 2024-03-08T14:44:06Z |
| publishDate | 2024-01-01 |
| publisher | American Society for Microbiology |
| record_format | Article |
| series | Microbiology Spectrum |
| spelling | doaj.art-919d7b6d2c8b48dba2c731b4cbddb16c2024-01-11T14:04:38ZengAmerican Society for MicrobiologyMicrobiology Spectrum2165-04972024-01-0112110.1128/spectrum.03107-23No extraction? No problem. Direct to PCR processing of tongue swabs for diagnosis of tuberculosis disease as an alternative to sputum collectionCharlotte L. Ahls0Duncan Emsweller1Seth J. Helfers2Xin Niu3Douglas Wilson4Leah R. Padgett5Paul K. Drain6Quantigen LLC , Fishers, Indiana, USAQuantigen LLC , Fishers, Indiana, USAQuantigen LLC , Fishers, Indiana, USADepartment of Global Health, University of Washington , Seattle, Washington, USAUmkhuseli Innovation and Research Management , Pietermaritzburg, South AfricaQuantigen LLC , Fishers, Indiana, USADepartment of Global Health, University of Washington , Seattle, Washington, USAABSTRACT Sputum collection and testing for tuberculosis (TB) have been problematic due to the potential for aerosolization, difficulty in generating a quality sample, and complex DNA extraction methods. Tongue swabs are inexpensive, minimally invasive, and a promising alternative to sputum collection. We investigated the diagnostic accuracy of tongue swabs for TB diagnosis using the Truenat MTB Plus assay from Molbio Diagnostics with a direct to PCR processing method. Four tongue swabs were collected per participant using two nylon-flocked swabs and two spun polyester swabs. Following tongue swab sample collection, participants also provided two sputum samples, which were tested by Cepheid Xpert MTB/RIF Ultra or culture. Of the 81 participants enrolled, 24 (30%) were positive for TB disease by sputum Ultra. Using the Truenat MTB Plus test, tongue swabs had 54% (52/96) sensitivity and 99% (218/220) specificity compared to sputum Ultra. Crude lysate was also tested using an in-house qPCR assay, which allowed for increased sample input. Using this method, tongue swabs had 70% (67/96) sensitivity and 94% (216/224) specificity. Mycobacterium tuberculosis (MTB) sample quantification using digital PCR yielded between 20 copies (minimum) and 34,000 copies (maximum) of MTB per swab. In addition, serially collected tongue swabs resulted in similar levels of detected MTB, and spun polyester swabs performed equivalently to nylon-flocked swabs. Overall, this study demonstrates that tongue swab samples are compatible with the Truenat MTB testing platform and that a direct to PCR method is a viable diagnostic solution. IMPORTANCE Tuberculosis (TB) remains one of the world’s leading infectious disease killers, despite available treatments. Although highly sensitive molecular diagnostics are available, expensive equipment and poor infrastructure have hindered their implementation in low-resource settings. Furthermore, the collection of sputum poses challenges as it is difficult for patients to produce and creates dangerous aerosols. This manuscript explores tongue swabs as a promising alternative to sputum collection. While previous studies have explored the sensitivity of tongue swabs as compared to sputum, existing literature has not addressed the need to standardize and simplify laboratory processing for easy implementation in high TB burden areas. This manuscript provides the first evidence that detection of TB from a tongue swab is possible without the use of DNA extraction or purification steps. The data provided in this manuscript will improve the collection and testing of tongue swabs for the diagnosis of TB disease.https://journals.asm.org/doi/10.1128/spectrum.03107-23Mycobacterium tuberculosistongue swabqPCRdirect lysis |
| spellingShingle | Charlotte L. Ahls Duncan Emsweller Seth J. Helfers Xin Niu Douglas Wilson Leah R. Padgett Paul K. Drain No extraction? No problem. Direct to PCR processing of tongue swabs for diagnosis of tuberculosis disease as an alternative to sputum collection Microbiology Spectrum Mycobacterium tuberculosis tongue swab qPCR direct lysis |
| title | No extraction? No problem. Direct to PCR processing of tongue swabs for diagnosis of tuberculosis disease as an alternative to sputum collection |
| title_full | No extraction? No problem. Direct to PCR processing of tongue swabs for diagnosis of tuberculosis disease as an alternative to sputum collection |
| title_fullStr | No extraction? No problem. Direct to PCR processing of tongue swabs for diagnosis of tuberculosis disease as an alternative to sputum collection |
| title_full_unstemmed | No extraction? No problem. Direct to PCR processing of tongue swabs for diagnosis of tuberculosis disease as an alternative to sputum collection |
| title_short | No extraction? No problem. Direct to PCR processing of tongue swabs for diagnosis of tuberculosis disease as an alternative to sputum collection |
| title_sort | no extraction no problem direct to pcr processing of tongue swabs for diagnosis of tuberculosis disease as an alternative to sputum collection |
| topic | Mycobacterium tuberculosis tongue swab qPCR direct lysis |
| url | https://journals.asm.org/doi/10.1128/spectrum.03107-23 |
| work_keys_str_mv | AT charlottelahls noextractionnoproblemdirecttopcrprocessingoftongueswabsfordiagnosisoftuberculosisdiseaseasanalternativetosputumcollection AT duncanemsweller noextractionnoproblemdirecttopcrprocessingoftongueswabsfordiagnosisoftuberculosisdiseaseasanalternativetosputumcollection AT sethjhelfers noextractionnoproblemdirecttopcrprocessingoftongueswabsfordiagnosisoftuberculosisdiseaseasanalternativetosputumcollection AT xinniu noextractionnoproblemdirecttopcrprocessingoftongueswabsfordiagnosisoftuberculosisdiseaseasanalternativetosputumcollection AT douglaswilson noextractionnoproblemdirecttopcrprocessingoftongueswabsfordiagnosisoftuberculosisdiseaseasanalternativetosputumcollection AT leahrpadgett noextractionnoproblemdirecttopcrprocessingoftongueswabsfordiagnosisoftuberculosisdiseaseasanalternativetosputumcollection AT paulkdrain noextractionnoproblemdirecttopcrprocessingoftongueswabsfordiagnosisoftuberculosisdiseaseasanalternativetosputumcollection |