PCR-RFLP and FTIR-based detection of high-risk human papilloma virus for cervical cancer screening and prevention

Autor: Tanmoy Bhattacharjee, Magda Rogéria Pereira Viana, Breno Pupin, Renata de Azevedo Canevari, Igor Martins Alves Melo
Rok vydání: 2021
Předmět:
Zdroj: Biochemistry and Biophysics Reports
Biochemistry and Biophysics Reports, Vol 26, Iss, Pp 100993-(2021)
ISSN: 2405-5808
DOI: 10.1016/j.bbrep.2021.100993
Popis: Background Approximately 70% of cervical carcinoma cases show the presence of high-risk Human Papilloma Virus (HPV), especially HPV-16 and HPV-18, and can be used to stratify high risk patients from low risk and healthy. Currently, molecular biology techniques such as polymerase chain reaction (PCR) are used to identify the presence of virus in patient samples. While the methodology is highly sensitive, it is labor intensive and time-consuming. Alternative techniques, such as vibrational spectroscopy, has been suggested as a possible rapid alternative. Therefore, in this study, we evaluate the efficiency of cervical fluid Fourier Transform Infrared spectroscopy (FTIR) in patient risk stratification informed by PCR. Methods Cervical fluid samples (n = 91) were obtained from patients who have undergone routine Papanicolaou (Pap) test. Viral genome was identified and classified as high/low-risk by PCR-Restriction Fragment Length Polymorphism (PCR-RFLP). FTIR spectra were acquired from samples identified by PCR-RFLP as No-HPV (n = 10), high-risk HPV (n = 7), and low-risk HPV (n = 7). Results Of the 91 samples, was detected the viral genome by PCR in 36 samples. Of these 36 samples, nine samples were identified to contain high-risk HPV (HR-HPV) and nine samples were found to have low-risk HPV (LR-HPV). The FTIR spectra acquired from No-HPV, LR-HPV, and HR-HPV showed differences in 1069, 1437, 1555, 1647, 2840, 2919, and 3287 cm-1 bands. Principal Component Analysis (PCA) showed distinct clusters for No-HPV and HR-HPV and No-HPV and LR-HPV, but there was significant overlap in the clusters of HR-HPV and LR-HPV. PCA-Linear Discriminant Analysis (PC-LDA) after Leave One Out Cross Validation (LOOCV) classified No-HPV from HR-HPV and No-HPV from LR-HPV with 100% efficiency in the 1400-1800 cm-1 spectral range. LOOCV classifications for LR-HPV and HR-HPV from each other were 71 and 75%, respectively, in the 2800-3400 cm-1 spectral range. Conclusions The results highlight the high sensitivity of PCR-RFLP in HPV identification and show that FTIR can classify samples identified as healthy, low, and high-risk samples by PCR-RFLP. General significance We show the possibility of using FTIR for initial cervical cancer risk stratification followed by detailed PCR-RFLP investigations for suspect cases.
Highlights • Molecular biology showed high sensitivity and specificity in HPV detection compared with cytological analysis. • PCR-RFLP technique identify high-risk variants of HPV can significantly assist in the prevention of cervical cancer. • PC-LDA and LOOCV classified normal from high-risk HPV and normal from low-risk HPV with 100% efficiency. • FTIR can be used for initial cervical cancer risk stratification followed by more detailed investigations by PCR-RFLP.
Databáze: OpenAIRE
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