Development of Enzymatic Recombinase Amplification Assays for the Rapid Visual Detection of HPV16/18
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Human papillomavirus (HPV) types 16 and 18 are the major causes of cervical lesions and are associated with 71% of cervical cancer cases globally. However, public health infrastructures to support cervical cancer screening may be unavailable to women in low-resource areas. Therefore, sensitive, convenient, and cost-efficient diagnostic methods are required for the detection of HPV16/18. Here, we designed two novel methods, real-time ERA and ERA-LFD, based on enzymatic recombinase amplification (ERA) for quick point-of-care identification of the HPV E6/E7 genes. The entire detection process could be completed within 25 min at a constant low temperature (35–43°C), and the results of the combined methods could be present as the amplification curves or the bands presented on dipsticks and directly interpreted with the naked eye. The ERA assays evaluated using standard plasmids carrying the E6/E7 genes and clinical samples exhibited excellent specificity, as no cross-reaction with other common HPV types was observed. The detection limits of our ERA assays were 100 and 101 copies/μl for HPV16 and 18 respectively, which were comparable to those of the real-time PCR assay. Assessment of the clinical performance of the ERA assays using 114 cervical tissue samples demonstrated that they are highly consistent with real-time PCR, the gold standard for HPV detection. This study demonstrated that ERA-based assays possess excellent sensitivity, specificity, and repeatability for HPV16 and HPV18 detection with great potential to become robust diagnostic tools in local hospitals and field studies.
Abstract
Human papillomavirus (HPV) types 16 and 18 are the major causes of cervical lesions and are associated with 71% of cervical cancer cases globally. However, public health infrastructures to support cervical cancer screening may be unavailable to women in low-resource areas. Therefore, sensitive, convenient, and cost-efficient diagnostic methods are required for the detection of HPV16/18. Here, we designed two novel methods, real-time ERA and ERA-LFD, based on enzymatic recombinase amplification (ERA) for quick point-of-care identification of the HPV E6/E7 genes. The entire detection process could be completed within 25 min at a constant low temperature (35–43°C), and the results of the combined methods could be present as the amplification curves or the bands presented on dipsticks and directly interpreted with the naked eye. The ERA assays evaluated using standard plasmids carrying the E6/E7 genes and clinical samples exhibited excellent specificity, as no cross-reaction with other common HPV types was observed. The detection limits of our ERA assays were 100 and 101 copies/μl for HPV16 and 18 respectively, which were comparable to those of the real-time PCR assay. Assessment of the clinical performance of the ERA assays using 114 cervical tissue samples demonstrated that they are highly consistent with real-time PCR, the gold standard for HPV detection. This study demonstrated that ERA-based assays possess excellent sensitivity, specificity, and repeatability for HPV16 and HPV18 detection with great potential to become robust diagnostic tools in local hospitals and field studies.
참고문헌 (43)
- Sung H, "Global cancer statistics 2020: GLOBOCAN estimates of incidence and mortality worldwide for 36 cancers in 185 countries", CA Cancer J. Clin, 71, pp.209~249 (2021)
- Kombe Kombe AJ, "Epidemiology and burden of human papillomavirus and related diseases, molecular pathogenesis, and vaccine evaluation", Front. Public Health, 8, pp.552028~ (2021)
- Serrano B, "Epidemiology and burden of HPV-related disease", Best Pract. Res. Clin. Obstet. Gynaecol, 47, pp.14~26 (2018)
- Chan CK, "Human Papillomavirus infection and cervical cancer:epidemiology, screening, and vaccination-review of current perspectives", J. Oncol, 2019, pp.3257939~ (2019)
- Boon SS, "Review of the standard and advanced screening, staging systems and treatment modalities for cervical cancer", Cancers (Basel), 14, pp.2913~ (2022)
- Pogoda CS, "Immunizing against anogenital cancer: HPV vaccines", PLoS Pathog, 12, pp.e1005587~ (2016)
- Maver PJ, "Primary HPV-based cervical cancer screening in Europe: implementation status, challenges, and future plans", Clin. Microbiol. Infect, 26, pp.579~583 (2020)
- Brisson M, "Impact of HPV vaccination and cervical screening on cervical cancer elimination: a comparative modelling analysis in 78 low-income and lower-middle income countries", Lancet, 395, pp.575~590 (2020)
- Singh D, "Global estimates of incidence and mortality of cervical cancer in 2020: a baseline analysis of the WHO Global Cervical Cancer Elimination Initiative", Lancet Glob. Health, 11, pp.e197~e206 (2023)
- Bhatla N, "Primary HPV screening for cervical cancer", Best Pract. Res. Clin. Obstet. Gynaecol, 65, pp.98~108 (2020)
- Catarino R, "Cervical cancer screening in developing countries at a crossroad: emerging technologies and policy choices", World J. Clin. Oncol, 6, pp.281~290 (2015)
- Chrysostomou AC, "Cervical cancer screening programs in Europe: the transition towards HPV vaccination and population-based HPV testing", Viruses, 10, pp.729~ (2018)
- Arbyn M, "Evidence regarding human papillomavirus testing in secondary prevention of cervical cancer", Vaccine, 30(Suppl 5), pp.F88~F99 (2012)
- Laudadio J, "Human papillomavirus detection: testing methodologies and their clinical utility in cervical cancer screening", Adv. Anat. Pathol, 20, pp.158~167 (2013)
- Loopik DL, "Benefit and burden in the Dutch cytology-based vs highrisk human papillomavirus-based cervical cancer screening program", Am. J. Obstet. Gynecol, 224, pp.200.e1~200.e9 (2021)
- Isidean SD, "Cobas 4800 HPV Test, a real-time polymerase chain reaction assay for the detection of human papillomavirus in cervical specimens", Expert Rev. Mol. Diagn, 14, pp.5~16 (2014)
- Kashir J, "Loop mediated isothermal amplification (LAMP) assays as a rapid diagnostic for COVID-19", Med. Hypotheses, 141, pp.109786~ (2020)
- Mu D, "The fluorescent probe-based recombinase-aided amplification for rapid detection of Escherichia coli O157:H7", Mol. Cell. Probes, 60, pp.101777~ (2021)
- Davi SD, "Recombinase polymerase amplification assay for rapid detection of Monkeypox virus", Diagn. Microbiol. Infect. Dis, 95, pp.41~45 (2019)
- Piepenburg O, "DNA detection using recombination proteins", PLoS Biol, 4, pp.e204~ (2006)
- Liu D, "Establishment and application of ERA-LFD method for rapid detection of feline calicivirus", Appl. Microbiol. Biotechnol, 106, pp.1651~1661 (2022)
- Zhang W, "Enzymatic recombinase amplification coupled with CRISPR-Cas12a for ultrasensitive, rapid, and specific Porcine circovirus 3 detection", Mol. Cell. Probes, 60, pp.101772~ (2021)
- Yang K, "Reverse transcription-enzymatic recombinase amplification coupled with CRISPR-Cas12a for rapid detection and differentiation of PEDV wild-type strains and attenuated vaccine strains", Anal. Bioanal. Chem, 413, pp.7521~7529 (2021)
- Li J, "An isothermal enzymatic recombinase amplification (ERA) assay for rapid and accurate detection of Enterocytozoon hepatopenaei infection in shrimp", J. Invertebr. Pathol, 197, pp.107895~ (2023)
- Xia S, "Single-copy sensitive, field-deployable, and simultaneous dual-gene detection of SARS-CoV-2 RNA via modified RT-RPA", Cell Discov, 6, pp.37~ (2020)
- Ma W, "Recombinase polymerase amplification assay for sensitive and rapid detection of southern rice black-streaked dwarf virus in plants", J. Virol. Methods, 301, pp.114467~ (2022)
- Jang WS, "Development of a multiplex Loop-Mediated Isothermal Amplification (LAMP) assay for on-site diagnosis of SARS CoV-2", PLoS One, 16, pp.e0248042~ (2021)
- Wang J, "Real-time RPA assay for rapid detection and differentiation of wild-type pseudorabies and gE-deleted vaccine viruses", Anal. Biochem, 543, pp.122~127 (2018)
- Wang X, "Real-time reverse transcription recombinase polymerase amplification for rapid detection of murine hepatitis virus", Front. Microbiol, 13, pp.1067694~ (2022)
- Tian XX, "Rapid visual detection of porcine reproductive and respiratory syndrome virus via recombinase polymerase amplification combined with a lateral flow dipstick", Arch. Virol, 167, pp.493~499 (2022)
- Zhou Y, "A rapid detection of tomato yellow leaf curl virus using recombinase polymerase amplification-lateral flow dipstick assay", Lett. Appl. Microbiol, 74, pp.640~646 (2022)
- Lin H, "Rapid visual detection of plasmodium using recombinase-aided amplification with lateral flow dipstick assay", Front. Cell. Infect. Microbiol, 12, pp.922146~ (2022)
- Wang R, "Human papillomavirus vaccine against cervical cancer: Opportunity and challenge", Cancer Lett, 471, pp.88~102 (2020)
- Bordigoni A, "Development and validation of a multiplex qPCR assay for detection and relative quantification of HPV16 and HPV18 E6 and E7 oncogenes", Sci. Rep, 11, pp.4039~ (2021)
- Williams J, "Molecular detection methods in HPV-related cancers", Front. Oncol, 12, pp.864820~ (2022)
- Harari A, "Human papillomavirus genomics: past, present and future", Curr. Probl. Dermatol, 45, pp.1~18 (2014)
- Ma X, "Development of a sensitive and specific nanoparticle-assisted PCR assay for detecting HPV-16 and HPV-18 DNA", J. Med. Virol, 12, pp.92~ (2020)
- Chen Q, "Rapid and simultaneous visual typing of high-risk HPV-16", 18 with use of integrated lateral flow strip platform, 2022, pp.189, 350~
- Kumvongpin R, "High sensitivity, loop-mediated isothermal amplification combined with colorimetric gold-nanoparticle probes for visual detection of high-risk human papillomavirus genotypes 16 and 18", J. Virol. Methods, 234, pp.90~95 (2016)
- Ma B, "A simple and efficient method for potential point-of-care diagnosis of human papillomavirus genotypes: combination of isothermal recombinase polymerase amplification with lateral flow dipstick and reverse dot blot", Anal. Bioanal. Chem, 411, pp.7451~7460 (2019)
- Zheng X, "Rapid detection of HPV16", 18 based on a CRISPR-Cas13a, Anal. Methods, pp.2022, 14[DOI]
- He A, "Development of field-applicable endogenous internally controlled recombinase-aided amplification (EIC-RAA) assays for the detection of human papillomavirus genotypes 6 and 11 using sample releasing agent", Heliyon, 8, pp.e11323~ (2022)
- Wang J, "Internally controlled recombinase-aided amplification (IC-RAA) assays for the detection of human papillomavirus genotypes 16 and 18 using extracted DNA and samples treated with nucleic acid releasing agent", Arch. Virol, 165, pp.2241~2247 (2020)