Clinical validation of the PCR-reverse dot blot human papillomavirus genotyping test in cervical lesions from Chinese women in the Fujian province: a hospital-based population study

Pengming Sun; Yiyi Song; Guanyu Ruan; Xiaodan Mao; Yafang Kang; Binhua Dong; Fen Lin

doi:10.3802/jgo.2017.28.e50

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Sun, Song, Ruan, Mao, Kang, Dong, and Lin: Clinical validation of the PCR-reverse dot blot human papillomavirus genotyping test in cervical lesions from Chinese women in the Fujian province: a hospital-based population study

Original Article

J Gynecol Oncol 2017;28(5):e50.

Published online: 4 September 2017

DOI: https://doi.org/10.3802/jgo.2017.28.e50

Clinical validation of the PCR-reverse dot blot human papillomavirus genotyping test in cervical lesions from Chinese women in the Fujian province: a hospital-based population study

Pengming Sun^1,^2,, Yiyi Song², Guanyu Ruan¹, Xiaodan Mao¹, Yafang Kang¹, Binhua Dong¹, Fen Lin¹

¹Laboratory of Gynecologic Oncology, Fujian Provincial Maternity & Children's Health Hospital of Fujian Medical University, Fuzhou, China

²Department of Gynecology, Fujian Provincial Maternity & Children's Health Hospital of Fujian Medical University, Fuzhou, China

Correspondence to Pengming Sun Laboratory of Gynecologic Oncology, Fujian Provincial Maternity & Children's Health Hospital of Fujian Medical University, 18 Daoshan Road, Fuzhou, Fujian 350001, China. E-mail: sunfemy@hotmail.com

Received 6 October 2016 Revised 11 March 2017 Accepted 9 April 2017

This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/by-nc/4.0/) which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited.

Abstract

Objective

To determine the clinical significance of the polymerase chain reaction (PCR)-reverse dot blot (RDB) human papillomavirus (HPV) genotyping assay in cervical cancer screening.

Methods

A total of 10,442 women attending the Fujian Provincial Maternity and Children's Health Hospital were evaluated using the liquid-based cytology (thinprep cytologic test [TCT]) and the PCR-RDB HPV test. Women with HPV infection and/or abnormal cytology were referred for colposcopy and biopsy. For HPV DNA sequencing, 120 specimens were randomly selected. Pathological diagnosis was used as the gold standard.

Results

Using the PCR-RDB HPV test, overall HPV prevalence was 20.57% (2,148/10,442) and that of high-risk (HR)-HPV infection was 18.68% (1,951/10,442). There was 99.2% concordance between HPV PCR-RDB testing and sequencing. In this studied population, the most common HR-HPV types were HPV-16, −52, −58, −18, −53, −33, and −51, rank from high to low. HPV-16, −18, −58, −59, and −33 were the top 5 prevalent genotypes in cervical cancer but HPV-16, −18, −59, −45, and −33 were the top 5 highest risk factors for cancer (odds ratio [OR]=34.964, 7.278, 6.728, 6.101, and 3.658; all p<0.05, respectively). Among 10,442 cases, 1,278 had abnormal cytology results, of which, the HR-HPV positivity rate was 83.02% (1,061/1,278). To screen for cervical cancer by PCR-RDB HPV testing, when using CIN2+, CIN3+, and cancer as observed endpoints, the sensitivity was 90.43%, 92.61%, and 94.78% and the negative predictive value (NPV) was 99.06%, 99.42%, and 99.78%, respectively. PCR-RDB HPV and TCT co-testing achieved the highest sensitivity and NPV.

Conclusion

For cervical cancer screening, the PCR-RDB HPV test can provide a reliable and sensitive clinical reference.

Keywords: Papillomaviridae, Genotype, Cell Biology, Histology, Cancer Screening

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Fig. 1.

Prevalence of different HPV genotype infections. (A) HPV-16 is most prevalent genotype in HPV-positive women. (B) The HR-HPV infection rate showed a bimodal trend, and the rates of different age groups were statistically significant (χ²=272.740; p<0.001). Top 5 most frequent HPV genotypes in (C) NILM patients; (D) LSIL (CIN1) patients; (E) HSIL patients (CIN2 and CIN3); and (F) cervical cancer. Orange bar in (C-F) is the cumulative cases of HPV genotypes. CIN, cervical intraepithelial neoplasia; HPV, human papillomavirus; HR-HPV, high-risk human papillomavirus; HSIL, high-grade squamous intraepithelial lesion; LSIL, low-grade squamous intraepithelial lesion; NILM, negative for intraepithelial lesion or malignancy.

Fig. 2.

Age-related distribution of HPV infection. Top 5 most frequent HPV genotypes in different age groups of (A) NILM patients; (B) LSIL (CIN1) patients; (C) HSIL patients (CIN2 and CIN3); and (D) cervical cancer. CIN, cervical intraepithelial neoplasia; HPV, human papillomavirus; HSIL, high-grade squamous intraepithelial lesion; LSIL, low-grade squamous intraepithelial lesion; NILM, negative for intraepithelial lesion or malignancy. (continued to the next page)

Fig. 2.

(Continued) Age-related distribution of HPV infection. Top 5 most frequent HPV genotypes in different age groups of (A) NILM patients; (B) LSIL (CIN1) patients; (C) HSIL patients (CIN2 and CIN3); and (D) cervical cancer. CIN, cervical intraepithelial neoplasia; HPV, human papillomavirus; HSIL, high-grade squamous intraepithelial lesion; LSIL, low-grade squamous intraepithelial lesion; NILM, negative for intraepithelial lesion or malignancy.

Fig. 3.

Cumulative occurred risk of HR-HPV genotype for cervical lesions. Cumulative occurred risk of each HPV genotype: in patients with (A) a pathologically diagnosed CIN1; (B) CIN2; (C) CIN3; and (D) invasive cervical cancer. Orange bar represents statistically significant (p<0.05) green bar represents not statistically significant (p>0.05). CIN, cervical intraepithelial neoplasia; HPV, human papillomavirus; HR-HPV, high-risk human papillomavirus.

Table 1.

HPV infection by age related distribution (n=10,442)

Age (yr)	Negative (n=8,294)	Single-type HPV (n=1,449)	Dual-type HPV^* (n=466)	Multi-type HPV^† (n=233)	Total HPV infection (n=2,148)^‡
≤20 (n=89)	63	5	11	10	26 (29.21)
21–25 (n=861)	717	92	30	22	144 (16.72)
26–30 (n=1,749)	1,502	161	56	30	247 (14.12)
31–35 (n=1,693)	1,388	201	72	32	305 (18.02)
36–40 (n=1,765)	1,396	271	80	18	369 (20.91)
41–45 (n=1,853)	1,455	278	84	36	398 (21.48)
46–50 (n=1,335)	1,009	238	60	28	326 (24.42)
51–55 (n=537)	376	113	32	16	161 (29.98)
56–60 (n=284)	186	54	24	20	98 (34.51)
61–65 (n=164)	117	21	11	15	47 (28.66)
≥66 (n=112)	85	15	6	6	27 (24.11)

Values are presented as number (%). Detected by PCR-RDB genotyping, 18 HR-HPV, and 5 LR-HPV subtypes could be analyzed in the population. HPV, human papillomavirus; HR-HPV, high-risk human papillomavirus; LR-HPV, low-risk human papillomavirus; PCR-RDB, polymerase chain reaction-reverse dot blot.

^* Dual-type infection: patients with 2 HPV subtypes infection simultaneously;

^† Multi-type infection: patients with 3 or more HPV subtypes infection simultaneously;

^‡ Represented the percent of HPV-positive patients in different age groups.

Table 2.

Compare HPV infection and TCT with pathological diagnosis (n=10,442)

Subjects	Pathologic diagnosis					χ²^*	p-value	χ²^†	p-value
Subjects	NILM (n=9,309)	CIN1 (n=395)	CIN2 (n=227)	CIN3 (n=292)	Cancer (n=219)	χ²^*	p-value	χ²^†	p-value
TCT (n=10,442)
HPV-positive (n=2,148)	1,155	333	197	262	201
HPV-negative (n=8,294)	8,154	62	30	30	18
Cytological diagnosis
NILM (n=9,164)						743.729	0.001	1,125.568	0.001
HPV-positive (n=1,087)	890	63	52	49	33
HPV-negative (n=8,077)	8,027	14	19	12	5
ASC (n=587)						22.125	0.001	38.252	0.001
HPV-positive (n=437)	168	142	41	57	29
HPV-negative (n=150)	99	36	6	6	3
LSIL (n=312)						0.584	0.540	3.103	0.524
HPV-positive (n=283)	68	118	47	39	11
HPV-negative (n=29)	6	11	4	5	3
≥HSIL (n=317)^‡						10.688	0.011	10.741	0.020
HPV-positive (n=300)	10	4	53	111	122
HPV-negative (n=17)	4	0	1	7	5
≥AGC (n=62)^§						5.866	0.019	9.103	0.038
HPV-positive (n=41)	19	6	4	6	6
HPV-negative (n=21)	18	1	0	0	2

ASC, atypical squamous cells; AGC, atypical glandular cells; CIN, cervical intraepithelial neoplasia; HPV, human papillomavirus; HSIL, high-grade squamous intraepithelial lesion; LSIL, low-grade squamous intraepithelial lesion; NILM, negative for intraepithelial lesion or malignancy; TCT, thinprep cytologic test.

^* χ²: the difference of patients with CIN2+ between the HPV-positive and -negative groups;

^† χ²: the difference between the HPV-positive and -negative groups in patients with different pathologic diagnosis and cytological diagnosis;

^‡ ≥HSIL: HSIL and cervical squamous carcinoma;

^§ ≥AGC: AGC and cervical adenocarcinoma.

Table 3.

Compare cytology and HPV in the different degree of cervical lesion detection (n=10,442)

Variable	Sensitivity	Specificity	PPV	NPV	PLR	NLR
CIN2+^*
HPV	90.43 (88.21–92.65)	84.83 (84.12–85.55)	31.02 (29.05–32.98)	99.06 (98.85–99.27)	5.8977 (5.5919–6.2202)	0.1246 (0.1010–0.1537)
TCT	76.96 (73.93–80.00)	92.73 (92.21–93.25)	44.65 (41.92–47.39)	98.14 (97.86–98.42)	10.5837 (9.7569–11.4807)	0.2484 (0.2177–0.2835)
HPV+TCT	95.26 (93.72–96.79)	83.04 (82.29–83.79)	29.98 (28.12–31.83)	99.57 (99.42–99.71)	5.6162 (5.3588–5.8861)	0.0571 (0.0413–0.0789)
CIN3+^†
HPV	92.61 (89.08–93.64)	83.20 (82.46–83.93)	21.76 (20.00–23.51)	99.42 (99.26–99.58)	5.3918 (5.1188–5.6793)	0.1129 (0.0862–0.1478)
TCT	80.63 (77.20–84.05)	91.32 (90.77–91.87)	32.39 (29.82–34.96)	98.92 (98.71–99.13)	9.2889 (8.6029–10.0296)	0.2122 (0.1777–0.2532)
HPV+TCT	96.67 (95.12–98.23)	81.32 (80.55–82.09)	21.07 (19.75–22.72)	99.79 (99.69–99.89)	5.1747 (4.9514–5.4081)	0.0409 (0.0256–0.0653)
Cervical cancer
HPV	94.78 (91.14–95.42)	81.11 (80.35–81.87)	9.45 (8.27–10.69)	99.78 (99.68–99.88)	4.8581 (4.5915–5.1403)	0.1013 (0.0651–0.1578)
TCT	82.65 (77.63–87.66)	89.30 (88.70–89.90)	14.23 (12.31–16.15)	99.58 (99.45–99.72)	7.7270 (7.1142–8.3925)	0.1943 (0.1455–0.2594)
HPV+TCT	97.72 (95.74–99.70)	79.11 (78.31–79.90)	9.13 (8.65–10.29)	99.94 (99.88–99.99)	4.6772 (4.4810–4.8820)	0.0289 (0.0121–0.0686)

Values are presented as value (95% CI). CI, confidence interval; CIN, cervical intraepithelial neoplasia; HPV, human papillomavirus; NLR, negative likelihood ratio; NPV, negative predictive value; PLR, positive likelihood ratio; PPV, positive predictive value; TCT, thinprep cytologic test.

^* CIN2+: included CIN2, CIN3, and cervical carcinoma;

^† CIN3+: included CIN3 and cervical cancer.

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