Diagnostic Cytology
respiratory tract or ocular symptoms. Morphologically, HSV 1
and HSV 2 cellular changes appear identical.
Although congenital or neonatal transmission may occur,
HSV 2 generally occurs after puberty and the onset of sexual
activity. Cutaneous lesions, commonly vesicles, tend to occur
in the same area repeatedly; the interval between successive
eruptions varies considerably even in the same individual.
Stress, menses, and other unrelated ailments may precipitate an
eruption in an otherwise healthy person. Following the initial
infection, the virus remains dormant in the sacral (S2 through
S4) dorsal root ganglia in the spinal cord. McDougall56 has
documented its presence in the spinal cord.
Recently, there has been an increase in the occurrence of HSV
2 cases; it is generally attributed to changed sexual and social
habits. Using seroepidemiologic data, an association of HSV 2
and cervical cancer has been reported by Kessler and others.55,57-63
The precise role of HSV 2 in the development of human cervical
cancer is far from resolved; evidence, however, accepts HPV as
the most important causative infection.
Herpes Simplex Genitalis Virus, Type 2
For nearly 2500 years,
people have used the word herpes in medical literature. Corey
has briefly discussed the history of genital herpes.64 John Astruc
first described genital herpes in 1736 in the French literature.
More than 100 cases of "herpes progenitalis" were reported in
the late nineteenth century. Lipschutz established experimental
transmission of herpes in human beings in 1921.65 He concluded
that there were differences between oral (HSV 1) and genital
herpes (HSV 2) infections.
HSV 2 infection is one of the most common sexually transmit-
ted genital infections; more than 300,000 new cases are recorded
in the United States annually. The prevalence of infection varies
depending on the group studied. Although in general popula-
tions the incidence of infection is not well established, genital
HSV infection was diagnosed among 4.2% of those attending
the Sexually Transmitted Disease Clinic in Seattle, Washington,
in 1980. Women presenting at student health services have been
found to have HSV 2 infection about seven to ten times more
commonly than gonorrhea. Data from the Centers for Disease
Control and Prevention (CDC) suggest that the prevalence of
HSV 2 is increasing and that the infection is occurring in social
groups that previously did not have the disease.
Primary infection may be asymptomatic or accompanied by
severe constitutional symptoms. Commonly, fever, headache,
and myalgia occur before the appearance of mucocutaneous
lesions. Visible lesions appear between 2 and 7 days following
exposure to the virus. Local pain and itching, dysuria, vaginal
discharge, and inguinal lymphadenopathy may be present. The
lesions are painful and often multiple. Large ulcerations that
start as papules or vesicles spread rapidly. They form pustules
that coalesce and break down. Unless complicated by second-
ary infection, these ulcers heal in 5 to 10 days with reepitheli-
alization. Residual scarring is uncommon. Systemic symptoms
and inguinal lymphadenopathy occur mainly in primary HSV 2
The cytologic diagnosis of HSV infection is important and
must be made on well-preserved cells that have typical diagnos-
tic features and have not been altered by air-drying, fixation, or
inflammation. Such a diagnosis may determine proper man-
agement of patients, especially pregnant women with geni-
tal ulcerations. An HSV diagnosis, with its social and medical
implications, should be rendered only when unequivocal
evidence is present.
In addition to diagnosis in the standard Pap test, direct sam-
pling of visible lesions can be performed. Such smears should be
prepared from the edge and bed of the ulceration, not from the
contents of the vesiculae. The latter generally contain serosan-
guineous material with acute inflammatory cells, eosinophils,
and some macrophages. Although use of air-dried smears and
their examination after Romanowsky stain (Tzanck prepara-
tion)66 have been advocated, we do not recommend this for
genital lesion diagnoses. Heavy inflammation, cellular obscur-
ing, and degeneration often make interpretation difficult and
may severely compromise the diagnostic value of air-dried
smears. Cellular samples obtained from the cleared ulcer beds
should be immediately fixed in 95% ethanol and examined after
Papanicolaou staining.
The virus may infect the immature squamous, metaplastic,
and endocervical columnar cells. Initially, the changes are pro-
plastic and somewhat nonspecific. The infected cells can occur
singly, in groups, and in tissue fragments. There is cytomegaly
and karyomegaly, and the nucleocytoplasmic ratio is not much
altered. These cells demonstrate a combination of reactive (pro-
plastic) and degenerative (retroplastic) changes. The nuclei of
the infected cells show changes in the chromatin structure con-
sisting of hydropic or ballooning degeneration. The chromatin
material becomes extremely finely divided and is uniformly
dispersed in the nuclear sap. The chromatin-parachromatin
interphase is obliterated, and nuclei assume a faintly hematoxy-
linophilic, homogenized appearance. Some chromatin material
may be matted against the inner leaf of the nuclear envelope,
which may appear uniformly thick and conspicuous. The altered
nuclear morphology is commonly referred to as ground glass,
bland, gelatinous, glassy, or opaque. In some cases the redistri-
bution of chromatin may result in a beaded appearance of the
nuclear margins. Nucleoli may be present and conspicuous, may
have associated chromatin, and may not appear typically bright
acidophilic. Although the nucleoli generally remain round or
oval, sometimes irregular shapes may be observed.
In the later stages of HSV infection, the cells undergo the
effects of viral replication and DNA integration. The cells may
assume multinucleation, which is observed in nearly 80% of the
smears from cases of genital HSV infection. The infected nuclei
may have the same homogeneous chromatin pattern described
previously. The nuclei appear tightly packed within the cells
and reveal distinct internuclear molding (Fig. 7.42). At times
they may be overlapping and not molding. Large and single
intranuclear inclusions appear within these nuclei. The nuclear
inclusions are generally round or oval. They can be angulated
and sharp (Fig. 7.43). They lack structure and are densely eosi-
nophilic. Depending on the staining procedure employed, they
may appear cherry red. A clear zone, or halo, which separates it
from the nuclear membrane, surrounds the intranuclear inclu-
sion. Most often the halo is as clear as the background of the
slide. Sometimes it may retain delicate, homogeneous, diffuse
hematoxylinophilia. Small, inconspicuous chromatin granules
can occur in the peri-inclusion halo. Inclusions may occur in
infected single cells observed in nearly one-third of the cases of
HSV 2 infection. Intranuclear inclusions may not be present in
all of the nuclei within the multinucleated giant cells.
The cytoplasm in the infected cells at this early stage of HSV
infection is dense. It may lose its transparent appearance and
become opaque. Often it stains bright cyanophilic.
HSV-infected cells can become atypical; the enlarged cells may
assume bizarre shapes (Fig. 7.44). They may be hyperchromatic
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