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An integrated isothermal nucleic acid amplification test to detect HPV16 and HPV18 DNA in resource-limited settings.

ABSTRACT: High-risk human papillomavirus (HPV) DNA testing is widely acknowledged as the most sensitive cervical cancer screening method but has limited availability in resource-limited settings, where the burden of cervical cancer is highest. Recently, HPV DNA tests have been developed for use in resource-limited settings, but they remain too costly for widespread use and require instruments that are often limited to centralized laboratories. To help meet the global need for low-cost cervical cancer screening, we developed a prototype, sample-to-answer, point-of-care test for HPV16 and HPV18 DNA. Our test relies on isothermal DNA amplification and lateral flow detection, two technologies that reduce the need for complex instrumentation. We integrated all test components into a low-cost, manufacturable platform, and performance of the integrated test was evaluated with synthetic samples, provider-collected clinical samples in a high-resource setting in the United States, and self-collected clinical samples in a low-resource setting in Mozambique. We demonstrated a clinically relevant limit of detection of 1000 HPV16 or HPV18 DNA copies per test. The test requires six user steps, yields results in 45 min, and can be performed using a benchtop instrument and minicentrifuge by minimally trained personnel. The projected per-test cost is <$5, and the projected instrumentation cost is <$1000. These results show the feasibility of a sample-to-answer, point-of-care HPV DNA test. With the inclusion of other HPV types, this test has the potential to fill a critical gap for decentralized and globally accessible cervical cancer screening.

SUBMITTER: Kundrod KA

PROVIDER: S-EPMC10566637 | biostudies-literature | 2023 Jun

REPOSITORIES: biostudies-literature

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An integrated isothermal nucleic acid amplification test to detect HPV16 and HPV18 DNA in resource-limited settings.

Kundrod Kathryn A KA Barra Maria M Wilkinson Alexis A Smith Chelsey A CA Natoli Mary E ME Chang Megan M MM Coole Jackson B JB Santhanaraj Akshaya A Lorenzoni Cesaltina C Mavume Celda C Atif Hira H Montealegre Jane Richards JR Scheurer Michael E ME Castle Philip E PE Schmeler Kathleen M KM Richards-Kortum Rebecca R RR

Science translational medicine 20230621 701

High-risk human papillomavirus (HPV) DNA testing is widely acknowledged as the most sensitive cervical cancer screening method but has limited availability in resource-limited settings, where the burden of cervical cancer is highest. Recently, HPV DNA tests have been developed for use in resource-limited settings, but they remain too costly for widespread use and require instruments that are often limited to centralized laboratories. To help meet the global need for low-cost cervical cancer scre ...[more]

PMID: 37343083

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Project description:IntroductionBuruli ulcer (BU) caused by Mycobacterium ulcerans (MU) is a devastating necrotic skin disease. PCR, recommended for confirmation of BU by WHO, requires an adequately equipped laboratory, therefore often delaying timely diagnosis and treatment of BU patients in remote settings. Loop-mediated isothermal amplification (LAMP) is a PCR-based protocol for isothermal amplification of DNA that has been suggested for diagnosis of BU in low-resource settings.Study aims and methodsThis is an exploratory diagnostic test evaluation study, with an embedded qualitative sub-study. Its aims are two-fold: First, to evaluate a simple rapid syringe-based DNA extraction method (SM) in comparison with a more elaborate conventional DNA extraction method (CM), followed by a LAMP assay targeting IS2404 for the detection of MU, either using a commercially available pocket warmer (pw) or a heat block (hb) for incubation. Second, to complement this by exploring the diagnostic workflow for BU at a community-based health centre in an endemic area in rural Ghana as an example of a potential target setting, using interviews with researchers and health care workers (HCWs). Diagnostic test evaluation results are discussed in relation to the requirements of a target product profile (TPP) for BU diagnosis and the target setting.ResultsA protocol using SM for DNA extraction followed by IS2404 PCR (IS2404 PCRSM) was able to identify MU DNA in 73 out of 83 BU clinical specimens submitted for diagnosis. The sensitivity, specificity, positive predictive value (PPV) and negative predictive value (NPV) of IS2404 PCRSM were 90.12%, 100%, 100% and 65.21% respectively, as compared to the reference standard IS2404 PCR in combination with a standard extraction protocol for mycobacterial DNA. Evaluation of the LAMP assay on 64 SM DNA extracts showed a sensitivity, specificity, PPV and NPV of 83.6%, 100%, 100% and 50%, respectively, using either pocket warmer (pwLAMPSM) or heat block (hbLAMPSM) for incubation of the reaction, as compared to the same reference standard. The limit of detection of pwLAMPSM was found to be 30 copies of the IS2404 target. Interview findings explored barriers to BU diagnosis and treatment, including perceptions of the disease, costs, and availability of transport. Participants confirmed that a diagnosis at the PoC, in addition to screening based on clinical criteria, would be advantageous in order to prevent delays and loss to follow-up.Discussion and conclusionsThe high diagnostic and analytic accuracy of the pwLAMP, evaluated by us in combination with a syringe-based DNA extraction method, supports its potential use for the rapid detection of MU in suspected BU samples at the community or primary health care level without reliable electricity supply. Further optimization needs include a lysis buffer, evaluation directly at the PoC and/or other sites, assessing staff training requirements and quality control.

Project description:BackgroundAlthough laparoscopic surgery has made remarkable progress and become the standard approach for various surgical procedures worldwide over the past 30 years, its establishment in low-resource settings, particularly in public hospitals, has been challenging. The lack of equipment and trained expertise has hindered its widespread adoption in these settings. Cholecystectomy is one of the most commonly performed procedures using laparoscopy world wide AIM: The aim of the study is to determine whether laparoscopic cholecystectomy is feasible in a resource challenged setting METHODS: The research focused on individuals who underwent laparoscopic or open cholecystectomies at Yekatit 12 Hospital in Addis Ababa, Ethiopia, over a one-year period. Comprehensive data collection was conducted prospectively, encompassing both intraoperative and postoperative parameters. Follow-up was carried out via phone calls. The surgical procedures employed innovative techniques, including the reuse of sterilized single-use equipment and the utilization of local resources. The evaluation involved a comparison of demographic information, intraoperative details (such as critical view determination and operative duration), and postoperative complications, including assessments of pain and wound infections RESULTS: From August 2021 to September 2022, 119 patients were assessed. Among these patients, 65 (54.6%) underwent open cholecystectomies, while the remaining 54 (45.4%) underwent laparoscopic cholecystectomies. The average duration of the laparoscopic cholecystectomies was 90.7 min, which is 17.7 min behind the open. Patients in the laparoscopy group had significantly shorter hospital stays than the open group, and 94% were discharged by post operative day 2. The conversion rate from laparoscopic to open surgery was determined to be 3.3% CONCLUSION: To sum up, the safe execution of laparoscopic cholecystectomies is feasible in public hospitals and settings with limited resources, given adequate training and resource distribution. The study findings showcased superior outcomes, including reduced hospitalization duration and fewer complications, while maintaining comparable levels of operative duration and patient satisfaction in both groups.

Dataset Information

An integrated isothermal nucleic acid amplification test to detect HPV16 and HPV18 DNA in resource-limited settings.

Publications

An integrated isothermal nucleic acid amplification test to detect HPV16 and HPV18 DNA in resource-limited settings.

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