Cervical cancer screening using cytology has significantly reduced cervical cancer mortality by detecting precancerous lesions. Screening is most effective in women aged 25-29 years old, as this is when high-grade lesions are most commonly detected. The Bethesda system provides standardized terminology for cervical cytology reports and categorizes results as negative, atypical, low-grade or high-grade abnormalities. Co-testing with HPV testing and cytology is superior to cytology alone and has been adopted in updated cervical cancer screening guidelines.

1. ROLE OF CERVICAL CYTOLOGY IN
SCREENING
Dr. Nirupama Kothari
Consultant Pathologist &Lab head,
Srl Jodhpur

2. Cytologic screening for cervical cancer
• Cervical cancer screening has decreased morbidity and
mortality
– Deaths from cervical cancer decreased from 26,000 to
less than 5,000 between 1941 and 1997

3. Principles of screening

4. • CIN3 has most frequently been found and treated in
women 25-29 years of age . This is also an age group in
which reversible HPV infection is commonly found
making the distinction between LSIL, a predominantly
reversible condition, and HSIL, which carries a risk of
progression to invasive cancer, the main focus for
cytology.
• While symptomatic clinically evident cervical cancer is
rare in women under 30 years of age, occult and
microinvasive cancers may be found in women aged
25-29 years in well-screened populations (Castanon et
al. 2013).

5. • HSIL and LSIL in screened populations
• In the some well-established screening
programmes , CIN II-III is most frequently found in
women aged 25-29 years
• Occult screen-detected cancer may also be found
in women aged 25-29 years
• LSIL and HPV infection are commonest in women
below 30 years of age
• Cytological distinction between HSIL and LSIL is
the key to accurate screening

7. Screening frequency
• Yearly until three consecutive normal pap smears, then
may decrease frequency to every three years
– Annual screening for high-risk women is highly
recommend.

8. When to screen
• Start within 3 years of onset of sexual activity or by age
of 21, whichever is first.
• Risk factors for cervical dysplasia
– Early onset of sexual activity
– Multiple sexual partners
– Tobacco
– Oral contraceptives

9. When to stop routine screening
• Age 65 and “adequate recent screening”
– Three consecutive normal pap smears
– No abnormal pap smears in last 10 years
– No history of cervical or uterine cancer
• Hysterectomy for benign disease
• Hysterectomy for invasive cervical cancer

10. Role of cytopathology
• Early detection of unsuspected diseases (malignant
or pre-malignant lesions).
• Confirmation of suspected diseases without surgical
trauma.
• Diagnosis of hormonal imbalance.
• Useful in follow up the course of disease or
monitoring therapy.

11. Advantage of Cytopathology
• Rapid diagnosis - Inexpensive - Simple
• It is better in evaluating the infectious diseases.
• Supplement or replace frozen section or biopsy
• No injury to tissue allowing repeated sampling
• It is better for hormonal assay
• Cytopathological smear cover a wider surface than that
involved in surgical biopsy.

12. Screening Methods
• Conventional Cytology
• Liquid-based cytology
• Cytology with or without HPV DNA test for oncogenic
types
• Automated screening.
• In 1998, the FDA approved the AutoPap System (now
called FocalPoint™)
• (TriPath Imaging, Burlington, NC) as a primary
• screener for cervicovaginal smears.
• FocalPoint

13. Two Types of Screening
• Conventional Pap Smear
• Cervical cell sample manually “smeared” onto
slide for screening.
• Liquid-Based :Cervical cell sample put into
liquid medium for suspension before
automated thin layer/monolayer slide
preparation .
• ThinPrep System ,SurePath

14. ACCURACY AND REPRODUCIBILITY

15. 15
History of the Conventional
Pap Smear
• Developed by Dr. George N.
Papanicolaou in 1940’s
• Most common cancer screening
test
• Critical aspect of annual
gynecologic examination
Ferris et al. Modern Colposcopy. 2004: 2-4, 49.
Photo accessed from http://www.cytology-iac.org/Cytopaths/1998/cytoFall98.htm

16. What is Pap Smear?
Here scrappings of cells in cervix are taken

18. Liquid base cytology

21. Liquid Based Cytology – lab processing

25. Importance of quality control in
controlling cervical cancer incidence
when risk of disease was increasing
• Quality control of all aspects of the screening
programme is required, from taking the
samples, screening and reporting the cytology,
following up women with cytological
abnormalities, and carrying out colposcopy,
treatment of high-grade abnormalities and
follow up after treatment (Arbyn et al. 2010).

26. • Cervical cytology provides, without damage to
the underlying tissue, a cellular sample of the
entire transformation zone (TZ) of the cervix
where most precancerous lesions develop
. When samples are taken for liquid-based
cytology it provides a medium on which HPV
testing and other molecular tests can be carried
out. HPV testing is frequently used for triage of
atypical or borderline cytology and as a test of
cure after treatment of high-grade precancerous
lesions

27. • The WHO guidelines for cervical cancer
screening only recommend cytological
screening “in countries where an
appropriate/high-quality screening strategy
with cytology followed by colposcopy already
exists”, where “either an HPV test or cytology
followed by colposcopy could be used.”

28. Guidelines for taking cellular samples
• Cytology samples should be taken at mid-cycle
• Menstrual endometrial cells may lead to
misinterpretation of the cytology
• Accurate personal information including name,
date of birth, screening history and symptoms (if
any) should be recorded on the request form
• Any or all of this information could be relevant to
the cytology result and recommendation for
clinical management

29. Devices for collecting cervical cellular
samples

33. Pap Test - Key points
• Effective, despite low sensitivity
• Most effective when tested at regular,
repeated intervals
• Most effective when laboratory practices
are optimized
• Role of pap test in cervical cancer
prevention is evolving

34. Samples obscured by blood,
inflammatory exudate or air-drying
• TBS regards a conventional smear as inadequate
if more than 75% is obscured by blood, exudate
or air-drying artefact but allows “quality indicator
comments” to be made if less than 75% is
obscured.
• TBS 2004 eliminated the previous ‘satisfactory
but limited by’ category, and liquid-based
cytology virtually eliminated most causes of this
troublesome category leaving scant cellularity as
the sole remaining cause of unsatisfactory
cytology (Siebers et al. 2012).

35. Number of cells per x40 high-power
field and equivalent number of total
cells on a LBC slide
• 3.8 cells per high-power field = 5000 cells with
ThinPrep
• 9.0 cells per high-power field = 5000 cells with
SurePath
• 10 cells per high-power field = 13,000 cells
with ThinPrep

37. DIAGNOSTIC TERMINOLOGY AND
REPORTING SYSTEMS

46. Bethesda system:why?
• 1.Need for standard system of nomenclature
• So that results are comparable
• 2.A clear statement of adequacy of specimen
• 3.General categorization.
• 4.Appropriateness of making
recommendations for further evaluation if
clinically indicated.

47. Why? Bethesda system
• It is important to distinguish cervical
intraepithelial neoplasia from benign &
reactive atypias.
• More recent information pointing to an
apparant biologic dichotomy between mere
“infection and genuine “neoplasia” has
resulted in latest entry “bethesda sysytem

48. Why? Bethesda system
• The Bethesda system terminology suggests
the disease is not a continuum but rather a
discontinous two disease system emphasized
by terminology of low(6&11) and high
grade(16-18) squamous intraepithelial lesion.

49. The Bethesda system criteria for
adequacy as a minimum requirement
• Conventional smear cellularity at least 8,000 to
12,000 cells
• Liquid-based cellularity at least 5,000 cells (‘grey
area’ of 5,000 to 20,000 cells may include a
comment on the report)
• Conventional smear inadequate if >75% is
obscured by blood, exudate or air-drying artefact
• TZ sampling reported but not a criterion for
adequacy

50. Guidelines for reporting negative
cytology
• Negative for intraepithelial lesion or
malignancy

51. • Benign changes that need not be reported in a negative
report
• Hormonal patterns (post-partum or atrophic)
• Repair changes
• Microglandular hyperplasia
• Tubal metaplasia
• Sampling of the lower uterine segment
• Irradiation changes
• Alterations due to inflammation or the presence of an
intrauterine device
• Benign changes occasionally seen after hysterectomy
• Endometrial cells in women under 40 years of age

52. • Organisms that may be mentioned in a negative
report
• Trichomomas vaginalis
• Candida
• Actinomyces-like organisms
• Herpesvirus multinucleated cells.
• These organisms may be mentioned because they
may have clinical relevance but are not related to
intraepithelial lesions.

53. Guidelines for reporting abnormal
cytology
The Bethesda terminology

98. Atypical and borderline changes
• The words ‘borderline’ and ‘atypical’ are used
when there is genuine doubt as to whether
the changes are neoplastic or reactive
• While ASC-US and borderline changes border
on LSIL, a minority, defined as ASC-H, cannot
exclude HSIL or cancer

99. ASC-US / borderline changes in
squamous cells, NOS
•
• There are two situations in which the ASC-US
category applies:
• Cytoplasmic changes suggesting HPV effect with
minimal nuclear change.
• Cytological preparations in which it is genuinely
difficult to distinguish benign, reactive or
degenerative nuclear changes from LSIL
•

100. • ASC-H / borderline changes, high-grade
dyskaryosis not excluded.
• AGC / borderline changes in endocervical
cells

101. Normal cytology and benign reactive
changes
• A benign cervical smear or LBC slide will contain:
• Squamous epithelial cells from the ectocervix
• Superficial squamous cells
• Intermediate squamous cells
• Parabasal squamous cells
• Navicular cells
• Columnar epithelium from the endocervical canal
• Endocervical cells
• Metaplastic squamous cells from the transformation zone
• Commensal vagina flora
• Lactobacilli, Gardnerella vaginalis, Leptothrix vaginalis
• Cervical mucus
•

102. A benign cervical smear or LBC slide may
contain
Exfoliated endometrial cells (dependant of
stage of menstrual cycle)
Epithelial cells
Superficial stromal cells
Deep stromal cells
Histiocytes, leucocytes, and red blood cells
Contaminants
e.g. spermatozoa, talc granules

103. Basic cytological pattern
• The type of epithelial cells seen in a cervical smear is
determined by:
• The degree of maturation of the cervical epithelium
• The location of the squamocolumnar junction
• The presence of metaplastic change in the
transformation zone
• The stage in the menstrual cycle when the smear is
taken
• The effect of hormones in pregnancy or ingested oral
contraceptives or hormone replacement therapy

104. TRANSFORMATION ZONE

105. NORMAL SQUAMOUS CELL CYTOLOGY

106. TRANSFORMATION ZONE SAMPLING
ENDOCERVICAL CELLS
IMMATURE METAPLASTIC
SQUAMOUS CELLS

107. ENDOMETRIAL CELLS
1/3
(a) Classical ‘top hat’ appearance of endometrial cells
surrounding stroma

108. POSTMENOPAUSAL SMEARS
High power view,40x
Sheets of atrophic cells with a uniform pattern and no
inflammation

109. Organisms seen in cervical cytology
Candida albicans

110. Actinomycosis Trichomonas vaginalis

111. Herpes virus infection of the cervix
(a) Margination of chromatin and
ground glass appearance of nuclei
Margination of chromatin,
multinucleation, cytomegaly and
eosinophilic inclusions.

112. Non-Epithelial Cells
sperms
Lymphocytes Polymorphs

113. Typical atrophic cell pattern
SCREENIN POWER VIEW 4X
Atrophic pattern with mild inflammation: note polymorph
infiltration, enlarged nuclei, dense hyperchromasia but relatively
low NC ratio for immature cell

114. • Potentially reversible low-grade and
borderline lesions
• Borderline, ASC-US and LSIL are more often
reported than HSIL
• HPV-positive rates in women with normal
cytology are even higher than ASC-US in
young women

115. • Excision of high-grade CIN carries a risk of premature rupture of
membranes in pregnancy: the risk is related to the depth of biopsy.
• Rigorous quality control is essential to avoid over-diagnosis as well
as under-diagnosis.
• Measurement of accuracy combines sensitivity, specificity, positive
predictive value and negative predictive value.
• There is a trade off between sensitivity (reflecting false negatives)
and specificity (reflecting false positives).
• In a population-based screening test of a rare condition NPV and
specificity may be less relevant than sensitivity and PPV as
measures of accuracy.
• Declining prevalence of disease may compromise PPV and
sensitivity.

116. • Pitfalls in cervical cytology
• Classic cytological appearances of high-grade
intraepithelial lesions are frequently not
encountered in routine practice and there are
many mimics of benign and neoplastic changes
that may lead to errors in diagnosis. Pitfalls in
cervical cytology can conveniently be divided into
three categories:
• Potential false negatives
• Potential false positives
• Unnecessary atypical/borderline diagnoses

117. Co-testing is superior to cytology alone in
cervical cancer
 Clinical evidence favors co-testing with
HPVdetection+Cytology tests
 The negative predictive value of co-testing is
higher than cytology alone
 Co-testing has been incorporated in cervical
screening guidelines in 2012
http://www.acog.org/~/media/Districts/District%20II/PDFs/USPSTF_Cervical_Ca_Screening_Guidelines.pdf?dmc=1

118. Is HPV+Cytology (co-testing) better
than cytology alone ??