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在通用运输培养基中长期保存后,成功从基于面纱收集的女性生殖分泌物中提取人乳头瘤病毒DNA。

Successful Retrieval of Human Papillomavirus DNA in Veil-Based Collected Female Genital Secretions After Long-Term Storage in Universal Transport Medium.

作者信息

Muwonga Tukisadila Jonathan, Avala Ntsigouaye Juval, Tonen-Wolyec Serge, Mboumba Bouassa Ralph-Sydney, Muwonga Jeremie, Belec Laurent

机构信息

École Doctorale Régionale D'Afrique Centrale en Infectiologie Tropicale, Franceville 876, Gabon.

Laboratoire de Biologie Clinique des Cliniques Universitaires de Kinshasa, Kinshasa 123, Democratic Republic of the Congo.

出版信息

Diagnostics (Basel). 2025 Apr 24;15(9):1079. doi: 10.3390/diagnostics15091079.

Abstract

: The surveillance of viral strain evolution is needed during prophylactic HPV vaccination programs against cervical cancer and necessitates safely archiving and storing cervical samples while maintaining the long-term stability of HPV DNA to carry out molecular diagnosis. The present proof-of-concept study aimed to assess DNA stability for HPV molecular detection from veils resuspended in a universal transport medium (UTM) and conserved at different temperatures after long-term storage. : The detection and quantification of HPV DNA were evaluated in female genital secretions self-collected using veils and conserved in Cyt-All UTM at -30 °C, +4 °C, and +25 °C after long-term 27-month storage. : A slight degradation of the ubiquitous single-copy cellular DNA TOP3 gene was assessed using multiplex real-time PCR (BMRT Human Papillomavirus Genotyping Real Time PCR Kit, Bioperfectus Technologies Co., Ltd., Taizhou, Jiangsu, China) at positive temperatures (+4 °C and +25 °C) but not at a frozen temperature (-30 °C) after 27 months of storage. Nevertheless, HPV DNA preservation was sufficient at the three storage temperatures to detect and quantify HPV DNA, with a similar rate of HPV detection, a similar level of cumulative HPV viral loads, high sensitivity and specificity, and perfect concordance in HPV genotype detection after the long period of 27 months of storage. Finally, the conservation of genital samples for a prolonged period in the Cyt-All medium, even at room temperature, allows for the detection and quantification of any HPV and HR-HPV with high accuracy. : The combination of veil-based self-sampling of female genital secretions and their elution and conservation in UTM may be used in the field to carry out longitudinal molecular epidemiology surveys of circulating HPV.

摘要

在预防宫颈癌的人乳头瘤病毒(HPV)疫苗接种计划期间,需要对病毒株的进化进行监测,这就需要安全地存档和储存宫颈样本,同时保持HPV DNA的长期稳定性以进行分子诊断。本概念验证研究旨在评估从悬浮于通用运输培养基(UTM)中并在长期储存后于不同温度下保存的拭子中进行HPV分子检测的DNA稳定性。:对使用拭子自行采集的女性生殖分泌物中的HPV DNA进行检测和定量,这些分泌物在27个月的长期储存后保存在Cyt-All UTM中,温度分别为-30°C、+4°C和+25°C。:在储存27个月后,使用多重实时PCR(BMRT人乳头瘤病毒基因分型实时PCR试剂盒,江苏泰州博日鸿生物科技有限公司)评估在正温度(+4°C和+25°C)下普遍存在的单拷贝细胞DNA TOP3基因有轻微降解,但在冷冻温度(-30°C)下没有。尽管如此,在这三个储存温度下,HPV DNA的保存足以检测和定量HPV DNA,HPV检测率相似,累积HPV病毒载量水平相似,具有高灵敏度和特异性,并且在27个月的长期储存后HPV基因型检测完全一致。最后,即使在室温下,将生殖样本在Cyt-All培养基中长时间保存也能高精度地检测和定量任何HPV和高危型HPV。:基于拭子的女性生殖分泌物自我采样及其在UTM中的洗脱和保存相结合,可用于现场进行循环HPV的纵向分子流行病学调查。

https://cdn.ncbi.nlm.nih.gov/pmc/blobs/91d6/12071974/e64c881a7f21/diagnostics-15-01079-g001.jpg

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