Study population.

<p>CIN2: Cervical intraepithelial neoplasia grade 2. CIN3+ includes CIN grade 3-carcinoma <i>in </i><i>situ</i>, invasive squamous cell carcinoma, adenocarcinoma <i>in </i><i>situ</i>, and adenocarcinoma. Controls for the analysis were women without progression to CIN2 or CIN3+ during the two-year follow-up of the prospective study of HPV16/18-positive women. Cases were women with histological diagnosis of HPV16/18-positive CIN2 or CIN3+ at enrolment or during the two-year follow-up of the prospective study. Enrolment of the prospective study: from 2009 to 2010. Additional cases were HPV16-positive CIN2 or CIN3+ cases collected from 2006 to 2008 and 2011 to 2012.</p

Prevalence of HPV 16 and HPV 18 Lineages in Galicia, Spain

<div><p>Genetic variants of human papillomavirus types 16 and 18 (HPV16/18) could differ in their cancer risk. We studied the prevalence and association with high-grade cervical lesions of different HPV16/18 variant lineages in a case-control study including 217 cases (cervical intraepithelial neoplasia grade 2 or grade 3 or worse: CIN2 or CIN3+) and 116 controls (no CIN2 or CIN3+ in two-year follow-up). HPV lineages were determined by sequencing the long control region (LCR) and the E6 gene. Phylogenetic analysis of HPV16 confirmed that isolates clustered into previously described lineages: A (260, 87.5%), B (4, 1.3%), C (8, 2.7%), and D (25, 8.4%). Lineage D/lineage A strains were, respectively, detected in 4/82 control patients, 19/126 CIN3+ cases (OR = 3.1, 95%CI: 1.0–12.9, p = 0.04), 6/1 glandular high-grade lesions (OR = 123, 95%CI: 9.7–5713.6, p<0.0001), and 4/5 invasive lesions (OR = 16.4, 95%CI: 2.2–113.7, p = 0.002). HPV18 clustered in lineages A (32, 88.9%) and B (4, 11.1%). Lineage B/lineage A strains were respectively detected in 1/23 control patients and 2/5 CIN3+ cases (OR = 9.2, 95%CI: 0.4–565.4, p = 0.12). In conclusion, lineages A of HPV16/18 were predominant in Spain. Lineage D of HPV16 was associated with increased risk for CIN3+, glandular high-grade lesions, and invasive lesions compared with lineage A. Lineage B of HPV18 may be associated with increased risk for CIN3+ compared with lineage A, but the association was not significant. Large well-designed studies are needed before the application of HPV lineage detection in clinical settings.</p></div

Phylogenetic tree of the HPV18 isolates based on LCR/E6 sequences.

<p>Phylogenetic analysis confirmed the presence of two lineages <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0104678#pone.0104678-Burk1" target="_blank">[4]</a>: A and B. A maximum likelihood (ML) tree was inferred from an alignment of nine reference sequences and seven study sequences of HPV18 LCR-E6 using RAxML HPC v8 <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0104678#pone.0104678-Stamatakis1" target="_blank">[22]</a>. Highly related sequences (<0.4 differences) from the study were not included in this figure. Reference sequences were denominated as lineage|strain.</p

The distribution of HPV18 lineages in the case-control study.

<p>CIN2: Cervical intraepithelial neoplasia grade 2. CIN3-CIS: CIN grade 3-carcinoma in situ. SCC: Invasive squamous cell carcinoma. AIS: Adenocarcinoma <i>in</i><i>situ</i>. ADCA: Adenocarcinoma.</p><p>CIN3+ includes CIN3-CIS, SCC, AIS, and ADCA.</p><p><i>p</i>, control group as reference.</p

Epidemiological characteristics of some patients included in the case-control study.

<p>Age is given as mean (SD, n) and other as n/N (%).</p><p>FSI: First sexual intercourse.</p><p>HC: Hormonal contraceptives.</p><p>*p<0.05, statistically significant, control group as reference.</p><p>CIN3+ includes cervical intraepithelial neoplasia grade 3-carcinoma in situ, invasive squamous cell carcinoma, adenocarcinoma <i>in</i><i>situ</i> and adenocarcinoma.</p

Nucleotide sequence variations of LCR/E6 among HPV18 isolates.

<p>Position number refers to the HPV18 prototype sequence previously described <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0104678#pone.0104678-Cole1" target="_blank">[18]</a>.</p

Phylogenetic tree of the HPV16 isolates based on LCR/E6 sequences.

<p>Phylogenetic analysis confirmed the presence of the four lineages <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0104678#pone.0104678-Burk1" target="_blank">[4]</a>: A, B, C and D. A maximum likelihood (ML) tree was inferred from an alignment of ten reference sequences and fifteen study sequences of HPV16 LCR-E6 using RAxML HPC v8 <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0104678#pone.0104678-Stamatakis1" target="_blank">[22]</a>. Highly related sequences (<0.4 differences) from the study were not included in this figure. Reference sequences were denominated as lineage|strain.</p

Nucleotide sequence variations of LCR/E6 among HPV16 isolates.

<p>Position number refers to the HPV 16 prototype sequence previously described <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0104678#pone.0104678-Myers1" target="_blank">[19]</a>.</p