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Accuracy of urinary human papillomavirus testing for presence of cervical HPV: systematic review and meta-analysis

BMJ 2014; 349 doi: https://doi.org/10.1136/bmj.g5264 (Published 16 September 2014) Cite this as: BMJ 2014;349:g5264

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Dear Editor,
It is with great interest that we have read the paper: Accuracy of urinary human papillomavirus testing for presence of cervical HPV and the linked Editorial: Urine testing for HPV.[1,2]
We agree with Kitchener and Owens’ editorial that before possible implementation of urine HPV testing in cervical cancer screening programs further evaluation is needed. However, before launching major prospective studies comparing urine sampling with vaginal self-sampling, the heterogeneity of urine sampling and testing protocols, as reported by the authors, should be resolved. In addition to preventing sample degradation, such protocols should take into account the rationale and evidence for using first void urine for HPV testing.
The hypothesis for finding HPV DNA in urine of women with a cervical HPV infection is that, during urination, urine gets contaminated by debris and impurities lining the urethra opening, including mucus and debris of exfoliated cells from the vagina, cervix and uterus. It hence follows that the initial flow of urine collects most of this debris, which explains why the first collected part of a urine void contains more HPV DNA than subsequent parts, as concluded by the authors of the meta-analysis.[1] Indeed, our recently published research also confirms these findings: results of quantitative PCR showed significantly more HPV (as well as human) DNA in the first void than in subsequent urine fractions.[3]
Researchers, as well as women providing samples, should be made aware of how HPV enters in the first void urine. One of the papers examined in the published meta-analysis describes cleaning the external genital area twice with alcohol swab before urine was collected.[4] Although first void urine was used, cleaning the genitals may have had an important impact on the amount of HPV DNA recovered.
There is also some remaining confusion regarding the definition of first void urine, which should refer to the initial flush of urine, but is sometimes mis-referred to as the first urine of the day. Definitions aside, however, the timing of collection may indeed impact the amount of viral DNA in the urine. In an ongoing trial, women had more HPV DNA detected in their first void urine collected in the morning than in their first void urine samples collected later in the day (Vorsters, unpublished data). This finding strengthens the concept that more HPV DNA is present when the interval between two urinations increases, as more excreted mucus and debris has the time to accumulate.
The volume of collected first void urine is an additional source of variation. There is no defined optimal volume to provide the most concentrated sample, but it can be challenging to interrupt the urine flow immediately after starting urination.
Convinced of the importance of the first void urine for HPV detection, a collection device has been developed that ensures immediately mixing of the first void urine with a preservation medium, whilst allowing the rest of the urine to exit the device unhindered (avoiding the need to interrupt the flow). This device is currently being used in studies of early impact of national HPV vaccination programmes in Bhutan and Rwanda, co-ordinated by the International Agency for Research on Cancer, and has been shown to have excellent acceptability in 1,000 young women in each of the two settings (Gary Clifford, unpublished data).
In summary, as emphasized in the editorial by Kitchener and Owens, standard criteria for volume, collection, storage and extraction are essential to maximise HPV detection from urine, and the collection of the first void is a key step.[2]

References
1. Pathak N, Dodds J, Zamora J, et al. Accuracy of urinary human papillomavirus testing for presence of cervical HPV: systematic review and meta-analysis. BMJ 2014;349:g5264.
2. Kitchener HC, Owens GL. Urine testing for HPV. BMJ 2014;349:g5542.
3. Vorsters A, Van den Bergh J, Micalessi I, et al. Optimization of HPV DNA detection in urine by improving collection, storage, and extraction. Eur J Clin Microbiol Infect Dis 2014.
4. Song ES, Lee HJ, Hwang TS. Clinical efficacy of human papillomavirus DNA detection in urine from patients with various cervical lesions. J Korean Med Sci 2007;22(1):99-104.

Competing interests: AV and PVD are co-founders of Novosanis, a spin-off company of the University of Antwerp, responsible for valorisation of the urine collection device. GC declares no conflicts of interest.

30 September 2014
Alex Vorsters
researcher
Van Damme Pierre, University of Antwerp; Clifford Gary, International Agency for Research on Cancer Lyon, France
University of Antwerp; Centre for the Evaluation of Vaccination; Vaccine & Infectious Disease Institute