In this study, we determined whether HPV could be detected in dry swab samples with sensitivities similar to those for STM samples. Using cell line cell pellets, we showed that there was no significant DNA degradation (both β-actin and HPV) in dry cell pellet samples stored at room temperature for up to 1 month regardless of HPV copy numbers. Our data on clinical samples showed that dry swab samples were more likely to be insufficient for HPV detection than the STM swab samples. The presence and number of HPV types detected in STM samples were somewhat greater than in paired dry swab samples, although these differences did not reach statistical significance. The type-specific high-risk HPV positive agreement was 60.7% between dry swab samples and STM samples (kappa = 0.69; 95% CI, 0.53 to 0.82). Importantly, positive agreement for the detection of any high-risk HPV type was better (borderline statistical significance) for samples from women with ASC-US+ (kappa+ = 0.74; 95% CI, 0.52 to 0.94) than for samples from those with negative cytological findings (kappa+ = 0.46; 95% CI, 0.16 to 0.74). Using real-time TaqMan assays, we reported that the degree of genomic DNA degradation in dry versus STM samples was greater than the degree of HPV DNA degradation in dry versus STM samples. Consistent with these observations, we showed that the sensitivities of high-risk HPV DNA detection appeared to be similar for dry and wet swab samples when the patient had a clinically significant cervical lesion (HSIL and cancer) (Table ).
While a few studies have reported using dry sample collection for HPV detection (including one large national survey of women in the United States) (5
), only Shah et al. compared dry swab samples directly to wet swab samples. In that study, three types of clinician-collected samples were compared for HPV detection: vaginal dry swabs, vaginal swabs collected in STM, and cervical swabs collected in STM (13
). HPV was detected and genotyped by dot blot hybridization. The positive agreement of detection of any HPV in vaginal dry swab and vaginal STM swab samples was 68.9% (62/90). The frequency and types of HPV detected in vaginal dry swabs were similar to those found in vaginal STM swab samples.
Several studies investigated the collection of dry cervical exfoliated cell samples using filter papers. With this method, swab samples can be smeared directly onto sterile filter paper, which is then easily air dried and stored at room temperature for up to 1 year. A small piece of the paper smear can then be punched out, boiled in water, and used directly for PCR amplification. Using this method, Kailash et al. showed previously that HPV-16 was correctly identified in 50 cervical paper scrape samples (8
). Similarly, using chemically treated Flinders Technology Associates (FTA) elute microcards, Gustavsson et al. showed previously that the agreement in HPV positivity for 50 cervical samples detected by a real-time PCR assay between cytobrush and FTA samples was 94% (kappa = 0.88; 95% CI, 0.75 to 1) (7
). On the other hand, Banura et al. compared the detection of 25 HPV types in paper and PBS samples from 111 young women. HPV was genotyped by using the SPF10 line probe assay (LiPA). The prevalence of any HPV types was 82.9% for PBS samples compared to 32.4% for paper samples. In addition, fewer HPV types and fewer multiple HPV infections were detected in paper samples (2
). In summary, these studies further support the feasibility of dry sample collection.
To estimate the extent of DNA degradation in dry swab samples, we quantitated β-actin as well as HPV DNA in paired samples using real-time TaqMan assays. Unlike what we observed for the cancer cell line experiment, where no DNA degradation was observed for both β-actin and HPV DNA in dry cell pellet samples, in clinical samples, both β-actin and HPV were significantly degraded in dry swab samples, although genomic DNA degradation was greater than HPV DNA degradation. We hypothesize that the degradation of genomic DNA is likely due to the presence of additional nuclease activity in dry clinical samples and that HPV DNA present predominantly as episomal copies in these samples was less affected. It would be interesting to determine whether integrated HPV DNA, such as that present in cervical cancer samples, would be degraded to an extent similar to that of genomic DNA in dry swab samples.
In our study, since wet swab samples were stored in STM, we used STM to rehydrate the dry swab samples to simplify the comparison. Because dry swab samples are immediately digested during rehydration, it is unlikely to introduce significant degradation by rehydrating dry swab samples in PBS instead of STM. Although our dry swab samples were stored refrigerated before shipment, we do not see any problems storing them at room temperature, since they were transported from clinics (Guayaquil, Ecuador) to the laboratory (Seattle, WA) at ambient temperature. Future studies should investigate the effect of rehydration in PBS and storage at room temperature to reduce the cost of sample collection.
In summary, our study provides evidence that HPV detection can be performed by using dry swab samples stored at room temperature although with an increase in the number of insufficient samples and with marginally less sensitivity for HPV detection. Despite the finding that genomic DNA was significantly degraded in dry swab samples versus wet samples, very little effect was observed on HPV DNA if it was present as episomal copies. Future studies should focus on identifying methods to inactivate nuclease activities in clinical samples and preserve genomic DNA in dry swab samples.