Human papillomavirus (HPV) infection is one of the most common sexually transmitted infections in the United States. Cervical infection with certain types of HPV is a major risk factor for cervical cancer in women. The “high-risk” types of HPV (e.g., HPV 16, 18) are associated with cervical cancer as well as other anogenital cancers and other types (e.g., HPV 6, 11) with genital warts. No national surveillance system exists to measure the full burden of HPV infection, and no reliable national population estimate of HPV exists. NHANES offers a unique opportunity to assess the prevalence of HPV infection in the general population.
Reducing the prevalence of HPV infection is a Developmental Healthy People 2010 objective: “Reducing the number of new HPV cases can help minimize the overall number of cases of high risk subtypes associated with cervical cancer in females...” Detection and typing of HPV DNA in vaginal swabs will allow evaluation of trends in prevalence of type-specific HPV infection by age, sexual behavior, and race/ethnicity. Three HPV vaccines (Gardasil, Gardasil 9, and Cervarix) are licensed and recommended for use in girls and women. Two vaccines are licensed and recommended for use in males (Gardasil and Gardasil 9). In mid-2006, the Advisory Committee on Immunizations (ACIP), recommended routine vaccination of females aged 11 or 12 years and for those 13-26 years not previously vaccinated. In December, 2011 ACIP recommended routine vaccination of males aged 11 or 12 years and for those aged 13 through 21 years not previously vaccinated. As vaccine becomes more widely used, the national prevalence of HPV infection will be critical for evaluating vaccination strategies in the United States.
Examined female participants aged 14-59 years and older were eligible. The public data file includes data for persons aged 18-59 years. Please see Analytic Notes about the release of data for adolescents aged 14-17 years.
Description of Laboratory Methodology
Roche Cobas HPV DNA Test
LBXHP2C (Cobas HPV Swab High Risk Result)
DNA extraction is performed with a modified protocol and the commercial QIAamp kit (Qiagen, Valencia, CA). Presence of high-risk HPV in the extracted patient DNA is determined with the Cobas Human Papillomavirus (HPV) Test. This qualitative in-vitro diagnostics test uses oligonucleotide probes labeled with four different fluorescent dyes. The primers target a DNA region of approximately 200 nucleotides within the polymorphic L1 region of the HPV genome to detect 14 high-risk HPV types (16, 18, 31, 33, 35, 39, 45, 51, 52, 56, 58, 59, 66 and 68) in a single analysis. The test reports concurrently the presence of one or more of these types at clinically relevant infection levels, however the test has been modified from the FDA approved format (sample type and extraction) and results cannot be used clinically.
Roche Linear Array Assay
This assay uses Roche Linear Array HPV Genotyping test that is based on HPV L1 consensus polymerase chain reaction (PCR) with biotinylated PGMY09/11 primer sets. It also includes biotinylated β-globin primers as an internal control for sample amplification. The primer mix amplifies essentially all HPV types found in the genital tract along with the human β-globin gene. After amplification the samples are typed by hybridization to the typing strips followed by colorimetric detection. The strip is a linear array of probes specific for 37 HPV types (6, 11, 16, 18, 26, 31, 33, 35, 39, 40, 42, 45, 51, 52, 53, 54, 55, 56, 58, 59, 61, 62, 64, 66, 67, 68, 69, 70, 71, 72, 73, 81, 82, 83, 84, 89, and IS39) and for the positive β-globin control as well. Types are read by comparing the reaction pattern to the typing template. Samples that are negative for HPV and the β-globin control indicate lack of a suitable sample and are considered inadequate for interpretation.
Laboratory Method Files
HPV Vaginal Swab Linear Array Laboratory Procedure Manual
HPV Vaginal Swab Cobas High Risk Laboratory Procedure Manual
Laboratory Quality Assurance and Monitoring
Vaginal swab specimens are processed, stored, and shipped to Chronic Viral Diseases Branch, Division of High-Consequence Pathogens and Pathology, National Center for Emerging and Zoonotic Infectious Diseases, Centers for Disease Control and Prevention, Atlanta, GA for analysis.
Detailed instructions on specimen collection and processing are discussed in the NHANES Laboratory Procedures Manual (LPM). Swabs are stored at room temperature until they are shipped to Chronic Viral Diseases Branch, Division of High-Consequence Pathogens and Pathology for analysis.
The NHANES quality assurance and quality control (QA/QC) protocols meet the 1988 Clinical Laboratory Improvement Act mandates. Detailed QA/QC instructions are discussed in the NHANES LPM.
Mobile Examination Centers (MECs)
Laboratory team performance is monitored using several techniques. NCHS and contract consultants use a structured quality assurance evaluation during unscheduled visits to evaluate both the quality of the laboratory work and the quality-control procedures. Each laboratory staff member is observed for equipment operation, specimen collection and preparation; testing procedures and constructive feedback are given to each staff member. Formal retraining sessions are conducted annually to ensure that required skill levels were maintained.
NHANES uses several methods to monitor the quality of the analyses performed by the contract laboratories. In the MEC, these methods include performing blind split samples collected on “dry run” sessions. In addition, contract laboratories randomly perform repeat testing on 2% of all specimens.
NCHS developed and distributed a quality control protocol for all the contract laboratories which outlined the use of Westgard rules when running NHANES specimens. Progress reports containing any problems encountered during shipping or receipt of specimens, summary statistics for each control pool, QC graphs, instrument calibration, reagents, and any special considerations are submitted to NCHS quarterly. The reports are reviewed for trends or shifts in the data. The laboratories are required to explain any identified areas of concern.
Data Processing and Editing
The data were reviewed. Incomplete data or improbable values were sent to the performing laboratory for confirmation.
Refer to the 2013 - 2014 Laboratory Data Overview for general information on NHANES laboratory data.
MEC exam sample weights should be used for analyses.
NHANES Demographic and Other Related Variables
The analysis of NHANES laboratory data must be conducted using the appropriate survey design and demographic variables. The NHANES 2013-2014 Demographics File contains demographic data, health indicators, and other related information collected during household interviews as well as the sample design variables. The recommended procedure for variance estimation requires use of stratum and PSU variables (SDMVSTRA and SDMVPSU, respectively) in the demographic data file.
This laboratory data file can be linked to the other NHANES data files using the unique survey participant identifier (i.e., SEQN).
The Questionnaire data files contain socio-economic data, health indicators, and other related information collected during household interviews. Certain sensitive data on respondents under 18 years of age (e.g. HPV typing results, sexual behavior variables) are not included in the public use files. These data may be requested as described in the NHANES guidelines.
Roche Cobas HPV DNA Test
The Roche Cobas HPV test is qualitative and only determines the presence or absence of high-risk HPV. If any analyte HPV 16, HPV 18 or Other High-risk is indicated as positive (POS) in the Cobas result file the result for the sample ID will be recorded as positive. If all of the HPV analytes are negative (NEG) in the Cobas result file the result for the sample ID will be recorded as negative. If any of the analytes are indicated as invalid in the Cobas result file, the DNA from that sample will be re-tested one time to obtain valid results. If the repeated result are still invalid the final result will be recorded as invalid.
HPV PCR Assay
HPV Summary Variable
The HPV PCR Summary variable (LBDRPCR) indicates if at least one type is positive (LBDRPCR=1), the sample is negative (LBDRPCR=2), the sample is inadequate (LBDRPCR=3), or the sample is missing (LBDRPCR=.).
If beta-globin is not present (both LBDRHP and LBDRLP are negative) in the sample and no HPV type is detected, the sample is coded as inadequate.
If any of the types on the strips (LBDR06-LBDRPI) are positive, the sample is coded as positive. If all of the types on the strip are coded as negative, and beta-globin is detected (either LBDRHP or LBDRLP is positive) the sample is coded as negative.
Variables LBDRHP through LBDRPI are from the RUO Roche Linear Array HPV typing assay, however LBDR52 also includes information from a type-specific assay for HPV 52. The Linear Array typing strip includes an XR probe that hybridizes with HPV 52 as well as HPV types 33, 35 and 58. Samples positive for the XR probe and 33, 35, or 58 require specific testing to confirm the presence of HPV 52.
Since this data is reported as qualitative data the use of lower LLODs is not applicable.
The public release data file includes HPV data for participants aged 18–59. Data for youth aged 14–17 years are available through the NCHS Research Data Center (RDC).