Evaluation of high-risk human papillomavirus types PCR detection in paired urine and cervical samples of women with abnormal cytology

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Abstract

Background

During the last decade, increasing efforts have focused on HPV detection in self-obtained samples, to increase the overall proportion of patients participating in cervical cancer screening procedures.

Objectives

A clinical evaluation study of an optimized protocol for PCR detection of high-risk human papillomavirus (HPV) types in urine compared with cervical samples in consecutive women referred to the colposcopy clinic with abnormal cervical cytology.

Study design

Paired urine and cervical specimens were collected from 100 consecutive women referred to the colposcopy clinic with abnormal cervical cytology and normal urine parameters. In-house and a commercial PCR method for the detection of HPV types 16 and 18, and a commercial multiplex PCR for HPV types 6, 11, 16, 18, and 33 were performed. All HPV cervix-positive/urine-negative paired urine samples were spiked with serial dilutions of cell lines infected with HPV 16 or 18 to test the sensitivity of HPV detection in these urine samples.

Results

In all but two cases HPV type 16 was detected. In cancer cases, the urine/cervix HPV detection sensitivity was 88.8%; in cases with high-grade lesions it was 76.5%; and in cases with low-grade lesions it was 45.5%. In all concordant cases the same HPV type was detected in both samples. The urine/cervix HPV detection sensitivity was higher when urine samples contained two or more epithelial cells per field in urine microscopy. HPV detection in 9 cervix-positive but urine-negative urine samples spiked with serial dilutions of HPV-positive cell lines showed that in these cases urine PCR inhibitors did not affect PCR amplification.

Conclusions

A higher urine/cervix HPV detection sensitivity in cancer and high-grade lesions suggests that urine testing could be used to detect HPV mainly when these lesions are present.

Introduction

Cevical cancer is strongly associated with infection by certain human papillomavirus (HPV) types. A yearly pelvic exam and Pap smear are recommended to look for cervical atypia, probably due to infection by high-risk HPV types. However, the majority of HPV infections might be undetectable by Pap test (Meisels, 1983). Also, histological diagnosis of HPV infection has certain limitations and do not correlate completely with viral detection by molecular assays (Salvia et al., 2004).

The drawbacks of Pap testing prompted the need for screening techniques that detect HPV DNA independently of cervical cytology, using cervical sampling and HPV DNA detection. However, this examination is impractical in some settings, considering that a gynaecological examination is required. Moreover, physician-obtained cervical sampling is considered uncomfortable, time-consuming and requires a degree of skill (Mandelblatt et al., 2002). Therefore, screening procedures based on self-sampling of urine, vulvar or vaginal samples for the detection of HPV DNA have been investigated (Brinkman et al., 2004, Sellors et al., 2000).

Urine testing for HPV has been previously examined in women at risk for disease (Brinkman et al., 2004, Forslund et al., 1993, Jacobson et al., 2000; Sellors et al., 2000, Strauss et al., 1999), as well as in women diagnosed with cervical cancer (Stanczuk et al., 2003, Vossler et al., 1995). However, the results vary due to differing methods and perhaps due to differences in studied populations. Recently, an optimization of methods to detect HPV in urine by PCR resulted in a modified protocol (Brinkman et al., 2004). This protocol, was evaluated in urine from women with abnormal cervical cytology.

Section snippets

Study population

Urine and cervical specimens (paired samples) were collected from 100 HIV-negative consecutive women referred with abnormal cervical cytology (atypical squamous epithelial cells of unknown significance [ASCUS] or worse) and normal urine parameters (acidic pH, non-detection of white blood cells, nitrites, proteins, crystals, and blood). All study participants were informed for the research purposes of the sampling by a specialist research nurse and gave their informed consent. The study was

Patient's characteristics

Histological examination revealed invasive cervical cancer in 9, high-grade intraepithelial lesions in 29, low-grade lesions 39, and various non-pre-malignant lesions in 23 women (Table 1).

HPV prevalence in urine and cervical samples

Cervical samples were tested positive for the oncogenic HPV types 16 or 18 in 37 of the 77 (48.1%) (95% CI 36.5–59.7%) women that proved to have a pre-malignant or malignant lesion by histology, while urine samples were positive in 26 (33.8%) (95% CI 23.4–45.4%) of these cases. This difference was not

Discussion

Many HPV types may contribute to the cervical carcinogenesis, with types 16 and 18, being considered among the most important ones (Lukaszuk et al., 2003). This is the reason for focusing on HPV 16 and 18.

In the literature, several studies have tested HPV in urine (Brinkman et al., 2004, Jacobson et al., 2000; Stanczuk et al., 2003, Strauss et al., 1999, Vossler et al., 1995). Urine HPV sampling for carcinogenic types was previously shown to be applicable to women attending a colposcopy clinic

Acknowledgements

The study was sponsored by the Procter & Gamble Papanikolaou award for the year 2004. We thankfully acknowledge the technical assistance of the laboratory technician Ioulia Kristo and the midwife Chrysa Kratsagoni.

References (18)

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