Detection of sexually transmitted infection and human papillomavirus in negative cytology by multiplex-PCR

Abstract

Background: The aim of this study was to determine the prevalence of human papillomavirus (HPV) and 15 species that cause sexually transmitted infections (STIs) in negative cytology. In addition, we compared the diagnostic performance of multiplex polymerase chain reaction (PCR) with widely available techniques used to detect HPV.

Methods: We recruited 235 women of reproductive age who had negative cytology findings in a liquid-based cervical smear. STIs were identified by multiplex PCR, and HPV genotypes by multiplex PCR, hybrid capture 2, and DNA microaray; discordant results were analyzed by direct sequencing.

Results: Approximately 96.6% of patients with negative cytology results were positive for pathogens that cause STIs. The pathogens most frequently detected were Gardnerella vaginalis, Ureaplasma urealyticum. The incidence of HPV in negative cytology was 23.3%. Low-risk HPV infection was significantly correlated with Chalmaydia trachomatis, and high-risk HPV infection was significantly correlated with Group β streptococcus. The analytical sensitivities of the multiplex PCR and DNA microarray were higher than 80%, and the analytical specificity was nearly 100% for all tests.

Conclusions: Multiplex PCR yielded results that most of patients with negative cytology were positive for pathogens that cause STIs, and were more similar to that of DNA microarray, than that of hybrid capture 2 in terms of analytical sensitivity and prediction value of HPV infection.

Figure 1

Sexually transmitted infection and human papillomavirus (HPV) multiplex PCR. Representative data of multiplex PCR with four multiplex primer mixes: Seeplex^® STI Master Panel 1 (MP1), 2 (MP2), 3 (MP3) and HPV4A ACE Screening. Panel 1 (B31) primers were designed so that amplicons were different sizes: 214 bp (NG), 253 bp (MG), 348 bp (CT1), 435 bp (UU), 502 bp (MH), and 800 bp (internal control). Panel 2 (B32) amplicons were 155 bp (MM), 197 bp (MC), 343 bp (BF), 509 bp (GV), and 647 bp (TV).

Figure 2

ROC curve to detect HPV using multiplex-PCR, HC2, and DNA microarray. ROC curve areas and 95% confidence intervals between each of the three different methods are shown.