LSHTM Research Online
Jeronimo, Jose; Bansil, Pooja; Lim, Jeanette; Peck, Roger; Paul, Proma; Amador, Juan Jose;
Mirembe, Florence; Byamugisha, Josaphat; Poli, Usha Rani; Satyanarayana, Labani; +2 more...
Asthana, Smita; START-UP Study Group; (2014) A multicountry evaluation of careHPV testing, visual inspection with acetic acid, and papanicolaou testing for the detection of cervical cancer. International journal of gynecological cancer, 24 (3). pp. 576-585. ISSN 1048-891X DOI:
https://doi.org/10.1097/IGC.0000000000000084
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ORIGINAL STUDY
A Multicountry Evaluation of careHPV Testing, Visual
Inspection With Acetic Acid, and Papanicolaou Testing for
the Detection of Cervical Cancer
Jose Jeronimo, MD,* Pooja Bansil, MPH,* Jeanette Lim, MPH,* Roger Peck, BS,* Proma Paul, MHS,*
Juan Jose Amador, MD, MPH,Þ Florence Mirembe, MD, PhD,þ Josaphat Byamugisha, MD, PhD,§
Usha Rani Poli, MBBSD, DNB, DGO,|| Labani Satyanarayana, PhD,¶ Smita Asthana, MBBS,¶
and the START-UP Study Group
Objective: This study evaluates the feasibility and performance of careHPV, a novel human papillomavirus (HPV) DNA test, when used for screening women for cervical cancer in
low-resource settings.
Methods and Materials: Clinician-collected (cervical) and self-collected (vaginal)
careHPV specimens, visual inspection with acetic acid (VIA), and Papanicolaou test were
evaluated among 16,951 eligible women in India, Nicaragua, and Uganda. Women with
positive screening results received colposcopy and histologic follow-up as indicated. The
positivity of each screening method was calculated overall, by site, and age. In addition, the
clinical performance of each screening test was determined for detection of cervical
intraepithelial neoplasia (CIN) grade 2 (CIN2+) and CIN grade 3.
Results: Moderate or severe dysplasia or cancer (taken together as CIN2+) was diagnosed
in 286 women. The positivity rate ranged between 2.4% to 19.6% for vaginal careHPV,
2.9% to 20.2% for cervical careHPV, 5.5% to 34.4% for VIA, and 2.8% to 51.8% for
Papanicolaou test. Cervical careHPV was the most sensitive for CIN2+ (81.5%; 95%
confidence interval [CI], 76.5Y85.8) and CIN grade 3 (85.3%; 95% CI, 78.6Y90.6) at
all sites, followed by vaginal careHPV (69.6% and 71.3%, respectively). The sensitivity of
VIA ranged from 21.9% to 73.6% and Papanicolaou test from 40.7% to 73.7%. The pooled
specificities of cervical careHPV, vaginal careHPV, VIA, and Papanicolaou test were
91.6%, 90.6%, 84.2%, and 87.7%, respectively.
*Program for Appropriate Technology in Health (PATH), Reproductive Health Global Program, Seattle, WA; †PATH, Reproductive
Health Program, Kampala, Uganda; ‡Department of Obstetrics and
Gynecology, Makerere University, Kampala, Uganda; §Department
of Obstetrics and Gynecology, Mulago Hospital, Kampala, Uganda;
||MNJ Cancer Institute, Oncology and Regional Cancer Center,
Hyderabad, India; and ¶Institute of Cytology and Preventive Oncology, Uttar Pradesh, India.
Address correspondence and reprint requests to Jose Jeronimo,
PATH, Reproductive Health Global Program, 2201 Westlake Ave,
Suite 200, Seattle, WA 98121. E-mail: jjeronimo@path.org.
The START-UP project was funded by a grant from the Bill &
Melinda Gates Foundation.
Supplemental digital content is available for this article. Direct URL
citation appears in the printed text and is provided in the HTML and
PDF versions of this article on the journal’s Web site (www.ijgc.net).
Copyright * 2014 by IGCS and ESGO
ISSN: 1048-891X
DOI: 10.1097/IGC.0000000000000084
576
Philip E. Castle has received commercial HPV tests for research at a
reduced or no cost from Roche, QIAGEN, Norchip, and MTM.
He is a paid consultant for BD, GE Healthcare, Gen-Probe/Hologic,
and Cepheid, and has received a speaker’s honorarium from Roche.
He is a paid consultant for Immunexpress on sepsis diagnostics. He
is compensated as a member of a Merck Data and Safety
Monitoring Board for HPV vaccines. Jose Jeronimo was the
director of the study and received all the tests used in the study as
a donation from the manufacturing company (QIAGEN). Alice
Lytwyn received grants for HPV research from McMaster
University and payment from Bedard Lectureship for lecture on
anal cancer. She also received payment for CD module on HPV
from Merck Frosst and received travel and accommodations
from the College of American Pathologists in related HVP work.
The remaining authors delare no conflicts of interest.
This is an open-access article distributed under the terms of the
Creative Commons Attribution-NonCommercial-NoDerivatives
3.0 License, where it is permissible to download and share the
work provided it is properly cited. The work cannot be changed in
any way or used commercially.
International Journal of Gynecological Cancer
& Volume 24, Number 3, March 2014
Copyright © 2014 by IGCS and ESGO. Unauthorized reproduction of this article is prohibited.
International Journal of Gynecological Cancer
& Volume 24, Number 3, March 2014
Evaluation of Cervical Cancer Detection
Conclusions: careHPV performed well in large multicountry demonstration studies
conducted in resource-limited settings that have not previously been conducted this type of
testing; its sensitivity using cervical samples or vaginal self-collected samples was better
than VIA or Papanicolaou test. The feasibility of using careHPV in self-collected vaginal
samples opens the possibility of increasing coverage and early detection in resourceconstrained areas.
Key Words: Cancer prevention and control, Early detection, Cervical cancer,
Low-resource settings, HPV
Received September 20, 2013, and in revised form December 4, 2013.
Accepted for publication December 5, 2013.
(Int J Gynecol Cancer 2014;24: 576Y585)
cervical cancer is the third leading cancer and
G lobally,
fourth cause of cancer-related mortality in women.
1
There is a high disparity for cervical cancer between higherincome and lower-income regions, with more than 85% of
cervical cancer occurring in low- and middle-income countries.1 This difference is primarily due to the difficulty in
implementing Papanicolaou testYbased screening programs
because of their complexity, need for highly trained providers
for reading the samples, need for close quality control, and
mediocre sensitivity of the test even in optimal conditions.2
Based on the absolute etiologic link between carcinogenic human papillomavirus (HPV) and cervical cancer, 2
new approaches for the prevention of cervical cancer have
emerged, these are as follows: (1) HPV vaccination for preventing incident HPV infection in younger women3,4 and
(2) carcinogenic HPV detection for screening for cervical
precancer and cancer.5Y11 Both vaccine and screening have
demonstrated high degrees of efficacy in prevention of HPV
infection or detection at a treatable stage, with maximum
effectiveness when guided by an understanding of the causal
model and applied in an age-appropriate manner.12 Although
prophylactic HPV vaccination may be the ultimate prevention
strategy, these vaccines do not treat preexisting HPV infections and precancerous conditions.13Y15 Therefore, there
are millions of at-risk women who will not benefit from HPV
vaccination, and robust screening and management programs
developed for low- and middle-income countries are needed
to reduce the burden of cervical cancer. Additionally, because
the HPV vaccines do not protect against all the oncogenic
genotypes of the virus, screening is still required even among
vaccinated cohorts.
Several new screening strategies have emerged as options for areas with limited resources. Visual inspection with
acetic acid (VIA) is a method based on the use of 5% acetic
acid (vinegar) that, when applied to the cervix, makes the
dysplastic epithelium turn white (acetowhitening), becoming
visible on evaluation by the unaided eye. The sensitivity of
VIA is variable; 2 recent meta-analyses have reported sensitivity of 70% to 80% for cervical intraepithelial neoplasia
(CIN) grade 2 (CIN2+) or more severe diagnoses.16,17 One
large randomized clinical trial in India found a 35% reduction
in cervical cancer-related mortality after a single screening
by VIA,18 whereas a second trial did not find a statistically
significant reduction8 in cervical cancer-related mortality
compared with those randomized to the no-intervention arm
of seeking standard services.
Another approach that has recently become available
is a lower-cost DNA test for detection of carcinogenic genotypes of HPV.19 Recent studies have shown that HPV
testing used in a screen-and-treat approach was more effective
than VIA in reducing the prevalence of CIN2+,20 and when
used in a more traditional program (ie, colposcopy and excisional treatment of histologically confirmed CIN2+), it was
more effective in reducing cervical cancer mortality than VIA
and Papanicolaou test.8 In response to the need for more
robust screening tools for low- and middle-income countries,
a public-private collaboration led to the development of
careHPV (QIAGEN, Gaithersburg, MD), a simplified, robust,
and affordable HPV test that could be used in low-resource
settings under a wider range of ambient conditions. The test
can be run in any room because it does not need running water
or air conditioning, and the process is simple and can be
completed by people with limited laboratory training. Preliminary results for careHPV were promising and compared
favorably to the US Food and Drug AdministrationYapproved
Hybrid Capture 2 (hc2; QIAGEN).21
Human papillomavirus DNA testing offers the possibility of using self-collected vaginal samples for primary
screening. The advantages of self-collection are that it does
not require pelvic evaluation; therefore, the sample collection
process could be completed without the need for a speculum
or even a health center facility because the sampling can be
done at the community level. A recently pooled analysis
of data from 5 studies in China found that HPV testing of selfcollected specimens was at least as sensitive for CIN2+
and CIN grade 3 (CIN3+) as liquid-based cytology using
clinician-collected specimens,22 and another review showed
that population coverage can be increased by offering selfsampling.23
Further studies were needed to demonstrate whether
careHPV truly had wide applicability under a variety of
settings and how it compared with the 2 standards, VIA and
* 2014 IGCS and ESGO
Copyright © 2014 by IGCS and ESGO. Unauthorized reproduction of this article is prohibited.
577
Jeronimo et al
International Journal of Gynecological Cancer
Papanicolaou testing, in a more realistic setting. We therefore
conducted a multisite study in routine public health settings in
India, Nicaragua, and Uganda using existing staff and resources to assess the clinical performance of careHPV with
both clinician-collected and self-collected specimens, VIA,
and Papanicolaou testing for detection of cervical precancer
and cancer.
MATERIALS AND METHODS
Screening Technologies to Advance Rapid TestingYUtility
and Program Planning (START-UP) study is a multisite demonstration project study conducted in India (Hyderabad and rural
Uttar Pradesh), Nicaragua (Masaya Province), and Uganda
(Kampala). Study protocols for each site were similar and were
approved by institutional review boards in all project sites and the
United States.
Efforts to promote enrollment in the study and meet
screening targets (5000 women from each site) used populationbased recruitment strategies that were specifically tailored to
each site. Local community leaders and stakeholders were
contacted in advance to gain their support in the mobilization
and education of the population. Women were then invited
to participate in the study using a variety of methods that
differed by study site (Table 1, Supplemental Digital Content 1,
http://links.lww.com/IGC/A201).
In India, women living in urban slums were invited to
enroll in the study. Screenings occurred at urban health
centers or mobile health clinics in Hyderabad, Andhra
Pradesh, whereas in Uttar Pradesh, women were recruited
from rural areas at the Dadri Tehsil of Gautam Buddha Nagar
District to be screened at primary health centers and/or subcenters. In Nicaragua, women from rural and semirural areas
within Masaya Province were invited to be screened at 1
of the following 3 stationary health centers: Monimbo, La
Concepcion, and Tisma. In Uganda, women were selected
from 5 divisions in Kampala and Wakiso district to attend
screenings at Mulago Hospital, the national referral hospital.
All recruited women were assessed for eligibility. Age
inclusion criteria differed slightly among sites to align and support local health practices and guidelines. In both Hyderabad
and Nicaragua, the age inclusion criteria were 30 to 49 years
old; in rural Uttar Pradesh, 30 to 59 years old; and in Uganda,
25 to 60 years old. Similarly, in both Nicaragua and Uganda,
eligibility was restricted to women who reported having sexual intercourse in the past. However, in the India sites, having
been married was used as a proxy for a history of having sexual
intercourse because it was culturally more appropriate. In all
sites, women were excluded if they had a history of CIN or
cervical cancer, a hysterectomy that removed the cervix, and
any condition that would pose a health risk to the participant
or interfere with the evaluation of the study objectives such as
a history of bleeding or clotting disorders, debilitating physical
and/or mental illness, and inability to provide informed consent. Pregnancy was considered a temporary exclusion until
3 months postdelivery. All participants provided written consent before the screenings began. For women who were not
able to read or write, the staff read the consent aloud to them
and women provided a thumbprint to indicate consent in the
presence of a witness.
578
& Volume 24, Number 3, March 2014
All eligible and consented women were offered 4
screening tests. First, they were instructed (by a female study
staff ) how to obtain a self-collected vaginal sample for
careHPV testing and were invited to provide a sample in a
private room or in the presence of study staff, depending on
their preference. Next, they underwent a pelvic examination
using a speculum during which additional cervical samples
were taken for careHPV and conventional Papanicolaou test.
The careHPV specimen for both vaginal and cervical samples
was collected with careHPV cervical sampler into Digene collection media (DCM). Finally, VIA was performed. Screening
tests were administered per protocol, and results were acquired
using standardized methodology. Before testing, careHPV specimens were stored at room temperature (15-C to 30-C) for
maximum of 14 days, 2-C to 8-C for maximum of 30 days,
or j20-C for maximum of 60 days.
In careHPV test results, represented as a ratio of viral
load expressed in relative light units (RLU), compared with
the mean RLU from a positive control set at 1 pg/mL cutoff
(CO),21 the equipment had been set by the manufacturer to
consider positive if the RLU/CO value was 1.0 and above.
Papanicolaou tests were graded by local pathologists according to the Bethesda classification system, and any smear
with atypical cells of undetermined significance or more
severe changes was considered positive. Lastly, epithelia that
turned white with acetic acid were classified as VIA positive
following standard VIA criteria.24
The careHPV testing was done locally at each study site.
The conditions for testing varied by site; in 1 site (Nicaragua),
the testing was done in a small laboratory in a health center.
In Uganda and Uttar Pradesh, the testing was done in offices,
and in Hyderabad, the testing was done in a storage room. Air
conditioning was not available at the testing facilities except
for in Nicaragua. Most of the required supplies are included
in the test kits; some supplies such as paper blotting towels
were procured locally in each country.
Women who resulted negative on all screening tests
were advised to be screened again in 3 years, and those who
resulted positive on any of the screening tests were referred to
colposcopy and directed biopsy of any abnormal epithelial
area and/or endocervical curettage, as necessary for diagnosis. Women with moderate to severe CIN (CIN2+) were
treated with cryotherapy as eligible and/or referred for further
management.
To improve histologic quality control, a 10% random
sample (via random number table) of negative biopsies and
all CIN2+ positive histologic specimens were independently
reviewed by an external pathologist from Canada who was
blinded to the original diagnosis. Given that the agreement
between the local and external pathologist in Hyderabad, rural
Uttar Pradesh, and Nicaragua was 90.8% (J = 0.71), 93.5%
(J = 0.79), and 75.5% (J = 0.53), respectively, the local pathologist readings were used to assess the final diagnosis.
On the other hand, the agreement for Uganda was 54.5%
(J = 0.22). As a result, all histologic slides from Uganda
were requested for reevaluation by the Canadian pathologist,
whose diagnosis was used as the final diagnosis. We assumed
that women with negative results on all screening tests were
truly negative for any precancer or cancer.
* 2014 IGCS and ESGO
Copyright © 2014 by IGCS and ESGO. Unauthorized reproduction of this article is prohibited.
International Journal of Gynecological Cancer
& Volume 24, Number 3, March 2014
Evaluation of Cervical Cancer Detection
FIGURE 1. 1Age outside 25 to 60 years (n = 20), had hysterectomy (n = 24), pregnant (n = 9), previous CIN/cancer
(n = 5), no sexual experience (Uganda and Nicaragua; n = 0), not married (India only; n = 0), or interfering health risk/
condition (n = 0). 2Vaginal careHPV not tested within protocol timeframe (Nicaragua only; n = 461), missing careHPV
vaginal (0.5 CO; n = 271), missing careHPV cervical (0.5 CO; n = 147), missing VIA (n = 75), or missing Papanicolaou
test (n = 109). 3[+] indicates screen positive and [j] indicates screen negative.
* 2014 IGCS and ESGO
Copyright © 2014 by IGCS and ESGO. Unauthorized reproduction of this article is prohibited.
579
Jeronimo et al
International Journal of Gynecological Cancer
& Volume 24, Number 3, March 2014
and CIN3+, along with the exact binomial 95% confidence
interval (95% CI). Finally, the overall (4 clinical sites combined) clinical performance and 95% CI for detection of
cervical CIN2+ and CIN3+ diagnosis were calculated.
McNemar W2 test was used to assess differences in sensitivity
and specificity for CIN2+ and CIN3+ for paired test results,
and multiple test comparisons were compared using Bonferroniadjusted McNemar W2.
All statistical analyses were conducted in Stata 12.0
(Statacorp, College Station, TX).
RESULTS
FIGURE 2. Percent test positive by screening test and
clinical site.
The positivity of each screening method was calculated
overall by site and age. Also, the percent test positive by all
4 screening tests at each clinical site by severity of histologic diagnosis was determined. In addition, the clinical performance (sensitivity, specificity, positive predictive value
[PPV], negative predictive value [NPV], and test positivity) of
each screening test was determined for detection of CIN2+
From the 4 sites in 3 countries, a total of 20,461 women
were consented and screened (Fig. 1). After excluding those
who did not meet the eligibility criteria (n = 58) or did not
complete all screenings (n = 1063), there were 19,340
(94.5%) eligible, consenting women who completed all
screenings. Of the 19,340 women, 2389 were lost to follow-up
or had missing histologic results, leaving 16,951 (87.6%)
women to be included in this analysis. The breakdown by
country was 4658 (94.6%) of 4925 completely screened
women from rural Uttar Pradesh, India, 4502 (93.8%) of
4799 from Hyderabad, India, 4645 (94.7%) of 4906 from
Nicaragua, and 3146 (66.8%) of 4710 from Uganda.
The mean (SD) age of women was similar in all sites:
rural Uttar Pradesh, 37.9 (7.5) years; Hyderabad, 36.3 (5.9) years;
Nicaragua, 37.9 (6.1) years; and Uganda, 36.8 (8.4) years. Most
FIGURE 3. Percent test positive by age group.
580
* 2014 IGCS and ESGO
Copyright © 2014 by IGCS and ESGO. Unauthorized reproduction of this article is prohibited.
International Journal of Gynecological Cancer
& Volume 24, Number 3, March 2014
Evaluation of Cervical Cancer Detection
TABLE 1. Percent test positive by each of the 4 tests at each clinical site by severity of histologic diagnosis
Vaginal careHPV
Rural Uttar Pradesh (n = 4658)
Negative (n = 4547)
CIN1 (n = 79)
CIN2 (n = 19)
CIN3 (n = 9)
Cancer (n = 4)
GCIN2 (n = 4626)
CIN2+ (n = 32)
GCIN3 (n = 4645)
CIN3+ (n = 13)
Hyderabad (n = 4502)
Negative (n = 4385)
CIN1 (n = 41)
CIN2 (n = 27)
CIN3 (n = 31)
Cancer (n = 18)
GCIN2 (n = 4426)
CIN2+ (n = 76)
GCIN3 (n = 4453)
CIN3+ (n = 49)
Nicaragua (n = 4645)
Negative (n = 4340)
CIN1 (n = 214)
CIN2 (n = 28)
CIN3 (n = 61)
Cancer (n = 2)
GCIN2 (n = 4554)
CIN2+ (n = 91)
GCIN3 (n = 4582)
CIN3+ (n = 63)
Uganda (n = 3146)
Negative (n = 2676)
CIN1 (n = 383)
CIN2 (n = 62)
CIN3 (n = 19)
Cancer (n = 6)
GCIN2 (n = 3059)
CIN2+ (n = 87)
GCIN3 (n = 3121)
CIN3+ (n = 25)
Cervical careHPV
VIA
Papanicolaou Test
89
9
6
5
2
98
13
104
7
(2.0)
(11.4)
(31.6)
(55.6)
(50.0)
(2.1)
(40.6)
(2.2)
(53.9)
104 (2.3)
15 (19.0)
6 (31.6)
8 (89.0)
3 (75.0)
119 (2.6)
17 (53.1)
125 (2.7)
11 (84.6)
228 (5.0)
22 (27.9)
6 (31.6)
1 (11.1)
0 (0)
230 (5.4)
7 (21.9)
256 (5.5)
1 (7.7)
90 (2.0)
28 (35.4)
6 (31.6)
7 (77.8)
1 (25.0)
118 (2.6)
14 (43.8)
124 (2.7)
8 (61.5)
189
18
19
25
14
207
58
226
39
(4.3)
(43.9)
(70.4)
(80.7)
(77.8)
(4.7)
(76.3)
(5.1)
(79.6)
194
23
24
27
17
217
68
241
44
(4.4)
(56.1)
(89.0)
(87.1)
(94.4)
(4.9)
(89.5)
(5.4)
(89.8)
361 (8.2)
17 (41.5)
9 (33.3)
16 (51.6)
17 (94.4)
378 (8.5)
42 (55.3)
387 (8.7)
33 (67.4)
92
11
15
24
17
103
56
118
41
(2.1)
(26.8)
(55.6)
(77.4)
(94.4)
(2.3)
(73.7)
(2.7)
(83.7)
553
99
18
43
0
652
61
670
43
(12.7)
(46.3)
(64.3)
(70.5)
(0)
(14.3)
(67.0)
(14.6)
(68.3)
424
89
19
51
1
513
71
582
52
(9.8)
(41.6)
(67.9)
(83.6)
(50.0)
(11.3)
(78.0)
(11.6)
(82.5)
856 (19.7)
129 (60.3)
18 (64.3)
38 (62.3)
2 (100.0)
985 (21.6)
58 (63.7)
1003 (21.9)
40 (63.5)
219
39
10
25
2
258
37
268
27
(5.1)
(18.2)
(35.7)
(41.0)
(100.0)
(40.7)
(12.5)
(5.9)
(42.9)
419
133
49
15
3
552
67
601
18
(15.7)
(34.7)
(79.0)
(79.0)
(50.0)
(18.1)
(77.0)
(19.3)
(72.0)
408
150
56
17
4
558
77
614
21
(15.3)
(39.2)
(90.3)
(89.5)
(66.7)
(18.2)
(88.5)
(19.7)
(84.0)
799 (29.9)
222 (58.0)
44 (71.0)
14 (73.7)
6 (100.0)
1021 (33.4)
64 (73.6)
1065 (34.1)
20 (80.0)
1272
300
42
15
3
1572
60
1614
18
(47.5)
(78.3)
(67.7)
(79.0)
(50.0)
(51.4)
(69.0)
(51.7)
(72.0)
Values are presented as n (percent positive).
Women with a negative diagnosis either did not have a histologic diagnosis because all of their screening tests were negative or they had at
least 1 positive screening test but no biopsy was taken or the worst diagnosis on biopsy was negative. The positive cut point for vaginal and
cervical careHPV was 1.0 rlu/pc. A positive VIA was an observed acetowhite lesion. A positive Papanicolaou test was atypical squamous cells
of undetermined significance or worse.
* 2014 IGCS and ESGO
Copyright © 2014 by IGCS and ESGO. Unauthorized reproduction of this article is prohibited.
581
International Journal of Gynecological Cancer
Jeronimo et al
& Volume 24, Number 3, March 2014
TABLE 2. Clinical performance of the 4 screening tests for detection of CIN2+ and CIN3+
Rural Uttar Pradesh
Hyderabad
Nicaragua
Uganda
4658
4502
End point: CIN2+
76
4645
3146
91
87
No. women
Number with CIN2+
Sensitivity, % (95% CI)
Vaginal careHPV
Cervical careHPV
VIA
Papanicolaou test
Specificity, % (95% CI)
Vaginal careHPV
Cervical careHPV
VIA
Papanicolaou test
PPV, % (95% CI)
Vaginal careHPV
Cervical careHPV
VIA
Papanicolaou test
NPV, % (95% CI)
Vaginal careHPV
Cervical careHPV
VIA
Papanicolaou test
Number with CIN3+
Sensitivity, % (95% CI)
Vaginal careHPV
Cervical careHPV
VIA
Papanicolaou test
Specificity, % (95% CI)
Vaginal careHPV
Cervical careHPV
VIA
Papanicolaou test
PPV, % (95% CI)
Vaginal careHPV
Cervical careHPV
VIA
Papanicolaou test
NPV, % (95% CI)
Vaginal careHPV
Cervical careHPV
VIA
Papanicolaou test
582
32
40.6
53.1
21.9
43.8
(23.7, 59.4)
(34.7, 70.9)
(9.3, 40.0)
(26.4, 62.3)
76.3
89.5
55.3
73.7
(65.2,
(80.3,
(43.4,
(62.3,
85.3)
95.3)
66.7)
83.1)
67.0
78.0
63.7
40.7
(56.4,
(68.1,
(53.0,
(30.5,
76.5)
86.0)
73.6)
51.5)
77.0
88.5
73.6
69.0
(66.8,
(79.9,
(63.0,
(58.1,
85.4)
94.3)
82.4)
78.5)
97.9
97.4
94.6
97.4
(97.4,
(96.9,
(93.9,
(97.0,
95.3
95.1
91.5
97.7
(94.7,
(94.4,
(90.6,
(97.2,
95.9)
95.7)
92.3)
98.1)
85.7
88.7
78.4
94.3
(84.6,
(87.8,
(77.1,
(93.6,
86.7)
89.6)
79.6)
95.0)
82.0
81.8
66.6
48.6
(80.5,
(80.3,
(64.9,
(46.8,
83.3)
83.1)
68.3)
50.4)
11.7
12.5
2.7
10.6
(6.4,
(7.5,
(1.1,
(5.9,
21.9
23.9
10.0
35.2
(17.1, 27.4)
(19.0, 29.2)
(7.3, 13.3)
(27.8, 43.2)
8.7
12.2
5.6
12.5
(6.6,
(9.6,
(4.3,
(9.0,
10.8
12.1
5.9
3.7
(8.5,
(9.7,
(4.6,
(2.8,
99.6
99.7
99.4
99.6
(99.4,
(99.5,
(99.2,
(99.4,
99.6 (99.3, 99.8)
99.8 (99.6, 99.9)
99.2 (98.8, 99.4)
99.5 (99.3, 99.7)
End point: CIN3+
49
99.2
99.5
99.1
98.8
(98.9,
(99.2,
(98.7,
(98.4,
99.3
99.6
99.0
99.6
(98.9, 99.6)
(99.3, 99.8)
(98.4, 99.4)
(99.1,99.9)
98.3)
97.9)
95.2)
97.9)
19.2)
19.3)
5.5)
17.2)
99.8)
99.8)
99.6)
99.8)
13
10.9)
15.1)
7.1)
16.9)
99.5)
99.7)
99.4)
99.9)
63
13.5)
14.9)
7.5)
4.7)
25
53.8
84.6
7.69
61.5
(25.1, 80.8)
(54.6, 98.1)
(0.2, 36.0)
(31.6, 86.1)
79.6
89.8
67.3
83.7
(65.7,
(77.8,
(52.5,
(70.3,
89.8)
96.6)
80.1)
92.7)
68.3
82.5
63.5
42.9
(55.3,
(70.9,
(50.4,
(30.5,
79.4)
90.9)
75.3)
56.0)
72.0
84.0
80.0
72.0
(52.0,
(63.9,
(59.3,
(50.6,
87.9)
95.5)
93.2)
87.9)
97.8
97.3
94.5
97.3
(97.3,
(96.8,
(93.8,
(96.8,
98.2)
97.8)
95.1)
97.8)
94.9
94.6
91.3
97.4
(94.2,
(93.9,
(90.4,
(96.8,
95.6)
95.2)
92.1)
97.8)
85.4
88.4
78.1
94.2
(24.3,
(87.4,
(76.9,
(93.4,
86.4)
89.3)
79.3)
94.8)
80.7
80.3
65.9
48.3
(79.3,
(78.9,
(64.2,
(46.5,
82.1)
81.7)
67.5)
50.1)
6.3
8.1
0.39
6.0
(2.6, 12.6)
(4.1, 14.0)
(0.01, 2.15)
(2.7, 11.6)
14.7
15.4
7.9
25.8
(10.7, 19.6)
(11.4, 20.2)
(5.5, 10.9)
(19.2, 33.3)
6.0
8.9
3.8
9.2
(4.4,
(6.7,
(2.8,
(6.1,
2.9
3.3
1.8
1.1
(1.7,
(2.1,
(1.1,
(0.7,
99.9
99.9
99.7
99.9
(99.7,
(99.8,
(99.5,
(99.7,
99.8
99.9
99.6
99.8
(99.6,
(99.7,
(99.4,
(99.6,
99.5
99.7
99.4
99.2
(99.2,
(99.5,
(99.0,
(98.9,
99.7
99.8
99.8
99.5
(99.4,
(99.6,
(99.4,
(99.1,
100.0)
100.0)
99.9)
100.0)
99.9)
100.0)
99.9)
99.9)
8.0)
11.5)
5.2)
13.0)
99.7)
99.9)
99.6)
99.42
4.6)
5.0)
2.8)
1.7)
99.9)
100.0)
99.9)
99.8)
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International Journal of Gynecological Cancer
& Volume 24, Number 3, March 2014
TABLE 3. Overall (4 clinical sites combined) clinical
performance (sensitivity, specificity, PPV, NPV, and test
positivity) for detection of CIN2+ and CIN3+
No. cases
Positivity, %
Vaginal careHPV
Cervical careHPV
VIA
Papanicolaou test
Sensitivity, % (95% CI)
Vaginal careHPV 69.6
Cervical careHPV 81.5
VIA
59.8
Papanicolaou test
58.4
Specificity, % (95% CI)
Vaginal careHPV 90.9
Cervical careHPV 91.6
VIA
84.2
Papanicolaou test
87.7
PPV, % (95% CI)
Vaginal careHPV 11.7
Cervical careHPV 14.2
VIA
6.1
Papanicolaou test
7.5
NPV, % (95% CI)
Vaginal careHPV 99.4
Cervical careHPV 99.7
VIA
99.2
Papanicolaou test
99.2
CIN2+
CIN3+
286
150
10.08
9.67
16.55
13.08
(63.9,
(76.5,
(53.9,
(52.4,
74.9)
85.8)
65.5)
64.2)
71.3
85.3
62.7
62.7
(63.4,
(78.6,
(54.4,
(54.4,
78.4)
90.6)
70.4)
70.4)
(90.5,
(91.1,
(83.6,
(87.2,
91.4)
92.0)
84.7)
88.2)
90.5
91.0
83.9
87.4
(90.0,
(90.6,
(83.3,
(86.8,
90.9)
91.4)
84.4)
87.9)
(10.2, 13.3)
(12.6, 16.0)
(5.2, 7.1)
(6.5, 8.7)
6.3
7.8
3.4
4.2
(5.2,
(6.6,
(2.7,
(3.4,
99.7
99.9
99.6
99.6
(99.6,
(99.9,
(99.5,
(99.5,
(99.3,
(99.6,
(99.0,
(99.0,
99.5)
99.7)
99.3)
99.3)
7.5)
9.2)
4.1)
5.2)
99.8)
99.9)
99.7)
99.7)
of the women in rural Uttar Pradesh (97.2%) and Hyderabad
(94.1%) were married (proxy measure for sexual experience),
and all (100%) women in Nicaragua and Uganda reported
having had sexual experience or intercourse.
The percent test positive varied greatly by screening test
and clinical site, but there was a strong correlation within
clinical site (ie, the percent test positive for all tests tended to
cluster by clinical site; Fig. 2). In rural Uttar Pradesh, the
percent test positive for vaginal HPV, cervical HPV, VIA, and
Papanicolaou testing were 2.4%, 2.9%, 5.5%, and 2.8%, respectively. In Hyderabad, the percent test positive for vaginal
HPV, cervical HPV, VIA, and Papanicolaou testing were
5.9%, 6.3%, 9.3%, and 3.5%, respectively. In Nicaragua, the
percent test positive for vaginal HPV, cervical HPV, VIA, and
Papanicolaou testing were 15.4%, 12.6%, 22.5%, and 6.4%,
respectively. In Uganda, the percent test positive for vaginal
HPV, cervical HPV, VIA, and Papanicolaou testing were
19.6%, 20.2%, 34.4%, and 51.8%, respectively. Notably,
there was also a correlation between percent test positive and
the prevalence of CIN2+.
Evaluation of Cervical Cancer Detection
The age-group specific patterns of percent test positive are shown in Figure 3. Generally, the percent test positive by VIA was much higher than that for other tests in
the youngest age groups. The percent test positive by VIA
declined sharply with increasing age (ptrend G 0.001 for rural
Uttar Pradesh, Nicaragua, and Uganda; ptrend = 0.04 for
Hyderabad) so it converged with the percent test positive for
the other tests. The percent test positive for vaginal and
cervical careHPV decreased with increasing age in Nicaragua
(ptrend = 0.12 for both) and Uganda (ptrend G 0.001 for both)
but did not change significantly with age for the two India
sites. The percentage of positive Paps did not change with
age except for those read in Uganda, in which the percentage of positive Paps was very high and surprisingly increased
with increasing age (ptrend G 0.001).
Table 1 shows the percentage of women who resulted
positive for each test by geographic site and by severity of the
histologic diagnosis. As a rule, the percentage of women who
resulted positive for all tests increased with increasing severity
of histologically confirmed disease, with the only exception
being Papanicolaou testing in Uganda, where the percentage of
positive tests was greatest among those with CIN1 and then
declined with increasing severity of diagnosis. Among women
with negative histologic finding who had a positive screening
test, VIA was the most common false-positive in rural Uttar
Pradesh, Hyderabad, and Nicaragua, and Papanicolaou test had
the highest false-positive rate in Uganda. Women with CIN3+
were most likely to result positive by cervical careHPV at all
sites (range, 82.5% to 89.8%).
The clinical performance characteristics of the 4 tests
are reported in Table 2 for CIN2+ and CIN3+ by geographic
site. Cervical careHPV was the most sensitive and had the
best NPV for CIN2+ and CIN3+ at all clinical sites. Vaginal
careHPV typically had the second or third best sensitivity and
NPV. Papanicolaou test performance was quite variable;
Papanicolaou testing was very accurate with high sensitivity
and specificity in Hyderabad, resulting in a very high PPV,
whereas it was very insensitive in Nicaragua and nonspecific in Uganda. Visual inspection with acetic acid had the
lowest specificity and PPV at all sites and had very poor
sensitivity in rural Uttar Pradesh (21.9% for CIN2+ and
7.69% for CIN3+).
We noted an important trade off in sensitivity and false
positivity (1 specificity) of VIA for CIN2+ and CIN3+
by clinical site (Figure 1, Supplemental Digital Content 2,
http://links.lww.com/IGC/A200). The more sensitive VIA
was read, the less specific and the greater percentage of falsepositives. Interestingly, there was a correlation with the
population risk of CIN2+; the higher the risk of CIN2+, the
greater was the sensitivity, the lower the specificity, and
the greater the false positivity.
Table 3 provides a summary of the combined, multisite
clinical performance of the 4 testing modalities for detection
of CIN2+ and CIN3+. Cervical careHPV had the highest
sensitivity (81.5% for CIN2+ and 85.3% for CIN3+) and
specificity (91.6% for CIN2+ and 91.0% for CIN3+) in
this study for both end points (P G 0.01, unadjusted and
Bonferroni-adjusted McNemar W2 for all comparisons) and
therefore had the best PPV and NPV as well. Vaginal careHPV
* 2014 IGCS and ESGO
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583
International Journal of Gynecological Cancer
Jeronimo et al
had the next highest sensitivity, specificity, PPV, and NPV for
CIN2+ specificity (P G 0.05, unadjusted and Bonferroni-adjusted
McNemar W2 vs VIA and Papanicolaou test) and consequently
the next highest PPV and NPV.
DISCUSSION
We reported on a multisite study to compare different
cervical cancer screening options in low- and middle-income
countries. We found that the objective tests, HPV DNA testing using clinician-collected or self-collected specimens, had
better clinical performance than the subjective tests, VIA and
Papanicolaou test. It is increasingly clear that there is a ‘‘menu of
options’’ for choice of screening tests, with trade offs in clinical
performance versus costs and infrastructure requirements. Although additional costing evaluations are under development to
explore the cost per CIN2+ detected and treated, it is important
to emphasize that ‘‘one size does not fit all,’’ and in countries
with limited screening, the goal should be to explore new approaches to expand current screening. The choice of test has
important downstream implications for patient management and
care. Although one of the criticisms of HPV DNA testing is its
low specificity, in our study, VIAwas less specific and had lower
PPV than careHPV. As a consequence, a VIA-based program
will refer 50% and above more women to either colposcopy in
more traditional screening programs in which diagnostic verification is required or to unnecessary treatment in a screen-andtreat program, such as the trial conducted in South Africa.25
We noted that the performance of all of the tests was site
dependent, even for nonsubjective tests such as careHPV. We
also observed that the test positivity (and therefore specificity) for all 4 tests tended to cluster by site rather than method.
This was presumably due to differences by site in exposure
to the underlying cause of cervical cancer, carcinogenic HPV
infections, and seemed to be correlated with the prevalence of
precancerous lesions as markers of cancer risk. A previous
report found that the prevalence of carcinogenic HPV correlated with cervical cancer incidence.26
We also found that the performance of VIA differed
greatly by geographic site, with a strong inverse relationship of
specificity with sensitivity for VIA (ie, the more specific VIA
performed, the less sensitive it was). Our results contradict a
previous meta-analysis which reported that VIA accuracy was
not influenced by study region or capacity of screener.12 We
can only speculate on possible explanations. First, one of the
challenges in assessing the true performance of VIA is that
disease ascertainment is based on colposcopy and directed
biopsies, which rely on the same acetowhitening phenomenon,
leading to inflated sensitivity due to autocorrelation. The performance of all tests was different in 1 of the study sites (Uttar
Pradesh). Because all screening tests were affected in this site,
we hypothesized that there was a local factor influencing it. The
study allocated additional funding and effort to review all the
steps and processes in this site, but we could not find a factor
for these results.
Our results also highlight the importance of having adequate local conditions at the time of implementing any cervical
cancer screening program. A good example was the performance of Papanicolaou test in our Uganda site; although the
584
& Volume 24, Number 3, March 2014
project was careful to select a senior pathologist for the local
reading of the Papanicolaou tests, the performance of that local
expert was inadequate and the results were clearly unreliable.
We found that Papanicolaou testing at the Hyderabad site
performed better than at the other sites. We conducted a post hoc
analysis in which we had a second pathologist in Hyderabad
reread a subset of the Papanicolaou tests (n = 994) without
knowledge of the ongoing study. Restricted to the subset read by
both pathologists, the second read was less sensitive (20.8% vs
81.8%) and slightly more specific (98.6% vs 97.2%) for CIN2+
than the original study reading (Table 2, Supplemental Digital
Content 3, http://links.lww.com/IGC/A201). The change in performancebetween the pathologists may be the variability because of the subjective nature of the test and/or the impact of
a priori knowledge of the study purpose versus routine practice.
In some studies, very high sensitivity for cytology has been reported, and we wonder if it is a study artifact. However, we
cannot differentiate between the 2 possible explanations, and
they should be the subject of further investigation.
We acknowledge that our study was limited with respect
to verification bias; women who resulted negative and had no
visible lesions were not referred to colposcopy and were not
biopsy examined. As a result, our results may overestimate the
clinical sensitivity of the tests evaluated and the results.
However, among the women who resulted positive, we highly
encouraged colposcopists to take a biopsy even if only a slight
abnormality was observed during colposcopy. Furthermore,
both vaginal and cervical careHPV were performed on all
women in conjunction with VIA and Papanicolaou test; it is
very likely that most women with CIN2+ were detected.
The careHPV test is more sensitive for detecting CIN2+
and CIN3+ than VIA or Papanicolaou test. Hence, we conclude that it is possible to perform high-quality tests for HPV
DNA in urban and rural areas in low-resource settings.
careHPV adds another screening option for low-resource
settings with the advantage of having a very high sensitivity
for detecting precancer and cancer and the feasibility of
performing cervical cancer screening without the need for a
pelvic evaluation. It is important to consider that vaginal
careHPV testing has lower sensitivity than cervical careHPV,
as reported by previous studies,21 but the loss on sensitivity
has to be evaluated in the context of potential for expansion
of screening coverage if vaginal samples self-collected by
women are used for primary screening.
Finally, it is important to emphasize that the performance
of the screening test is important, but the screening test is only
1 component of the population-based program required to
affect cervical cancer incidence and mortality.
ACKNOWLEDGMENTS
The Screening Technologies to Advance Rapid
TestingYUtility and Program Planning (START-UP) Study
Group includes Pushpa Sodhani (Institute of Cytology and
Preventive Oncology, Uttar Pradesh, India); Sanjay Gupta
(Institute of Cytology and Preventive Oncology, Uttar
Pradesh, India); Suresh Bhambhani (Institute of Cytology
and Preventive Oncology, Uttar Pradesh, India); Haawa
Nakawagi (Department of Obstetrics and Gynecology, Mulago
* 2014 IGCS and ESGO
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International Journal of Gynecological Cancer
& Volume 24, Number 3, March 2014
Hospital, Kampala, Uganda); Carol Nakisige (Department
of Obstetrics and Gynecology, Mulago Hospital, Kampala,
Uganda); Swarnalata Gowrishankar, MD (Department of
Pathology, Apollo Hospitals, Hyderabad, India); Meenakshi
Swain (Department of Pathology, Apollo Hospitals, Hyderabad,
India); Emmanuel Mugisha, PhD (PATH, Reproductive Health
Program, Kampala, Uganda); Edward Kumakech, MPH (PATH,
Reproductive Health Program, Kampala, Uganda); Magda
Sequeira (PATH, Reproductive Health Program, Nicaragua);
Geovanni Alvarado (PATH, Reproductive Health Program,
Nicaragua); Irfan Khan (PATH, Reproductive Health Program,
New Delhi, India); Philip E. Castle, PhD (Global Cancer Initiative, Chestertown, MD); and Alice Lytwyn, MD (Department
of Pathology and Molecular Medicine, McMaster University,
Hamilton, Ontario, Canada).
11.
12.
13.
14.
15.
16.
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585