Symposium on Laboratory Diagnosis

Diagnostic Virology

Anne A. Gershon, M.D. *

It is written in the Talmud: "According to the effort is the reward."

More appropriate to the pediatric virologist is the question: "Is the re-
ward worth the effort?" In contrast to other new diagnostic procedures,
virologic techniques are slow, expensive, and time-consuming. Often,
the patient's infection has subsided before the laboratory diagnosis is
made. Still, many situations do exist in which the virologist may be of
aid to the pediatrician. For example, appropriate public health measures
may be initiated when it is known whether a severely ill young Asian
visitor has smallpox or chickenpox. Documentation of a specific viral
infection may be of prognostic value as well. The diagnosis of infectious
mononucleosis rather than leukemia may be confirmed in a teenager
with fatigue, lymphadenopathy, and splenomegaly. Therapeutic regi-
mens, moreover, may be instituted on the basis of a virologic diagnosis,
an example being cesarian section of a pregnant woman with herpes
progenitalis. Often, the virologist can test for susceptibility to a given
disease, such as mumps, which may be of importance in specific in-
stances. Finally, documented virologic diagnosis is a necessity in vaccine
surveillance. Although of little immediate importance to the practitioner,
the occurrence of infection in a child previously vaccinated against that
disease - mumps, for example - is of vital importance to the investigator.
Ultimately, it may become important to the clinician as well.

METHODS EMPLOYED IN DIAGNOSTIC VIROLOGY

Viruses, being intracellular parasites, multiply only within a living

host. Depending upon individual growth requirements, viruses may be
propagated by inoculation of infected material into tissue cultures,
animals, or embryonate eggs. Tissue cultures are derived from animal
cells which are known to support viral growth; the cells are either sus-
pended in or grown under a nutrient medium in a cell sheet or mono-
layer on a glass or plastic surface. After inoculation, cultures must be
observed for as long as several weeks. Viral growth is noted indirectly by
cell death or by specific changes in cellular morphology termed the
"'Instructor, Department of Pediatrics, New York University School of Medicine

Pediatric Clinics of North America- Vol. 18, No.1, February, 1971 73

74 ANNE A. GERSHON

"cytopathic effect." Changes in the characteristics of the tissue culture

cells, such as the inability to support the growth of other viruses
(interference) or the ability to adsorb guinea pig red blood cells to the cell
surface (hemadsorption), may also be used to indicate viral growth.
Certain agents such as Coxsackie A virus and many arboviruses can best
be grown by inoculation into newborn mice. Embryonate eggs are useful
for growth of vaccinia and variola viruses as well as herpes simplex and
mumps viruses.
After a virus has been successfully propagated from a specimen,
two further problems must be resolved. First the virus must be identified.
Second, a causal relationship between the virus and disease must be
established to rule out the possibility that the virus was merely a "silent
passenger." Certain characteristics, such as the time necessary for
growth and the type of tissue culture which supports growth, provide
hints as to the identity of the virus. Viral isolates may be further identi-
fied specifically by serologic tests. Serologic techniques may similarly
be useful in solving the second problem. Acute and convalescent serum
specimens which show at least a four-fold rise of antibody titer against
a given virus indicate a recent infection with that virus (Table 1).
Measurement of various types of antibody is an important function
of the diagnostic virology laboratory. One important type of antibody is
characterized by its ability to fix complement. Complement fixing anti-
bodies against a given virus are usually transient, and therefore elevation
of their titer suggests recent infection. There is some degree of cross
reaction between closely related viruses such as herpes simplex and
varicella-zoster. Thus increased titers of cross-reacting antibodies are
nonspecific. Because complement fixing antibodies may be transient,
their absence does not necessarily indicate susceptibility to disease. The
complement fixation test requires 24 hours to perform and is one of the
most rapid techniques employed by the virologist. Therefore, it is useful
for screening against several suspected agents. The complement fixation
test is also useful, as mentioned earlier, to detect virus in an isolate. In
this situation an aliquot of tissue culture in which the virus is thought
to have grown is used as the antigen against serum known to contain
antibodies to that virus.

Table 1. Methods of Diagnostic Virology

LABORATORY METHOD PURPOSE

Culture To propagate virus from infected material

(e.g., pharyngeal swabs, urine, cerebrospinal
fluid)

Serologic tests 1. To identify a virus isolated from a patient

Complement fixation
Hemagglutination inhibition
Neutralization
Fluorescent antibody
Gel diffusion
Hemagglutination
2. To document a specific antibody rise against
a virus following an acute disease
3. To identify an antigen in material collected
from a patient (e.g., vesicle fluid or cerebrospinal
fluid)
4. To test for susceptibility to a specific disease
DIAGNOSTIC VIROLOGY 75
Another type of antibody that is measured is termed neutralizing
antibody. Such antibodies are detected by their ability to inhibit viral
growth. In contrast to complement fixing antibodies, neutralizing anti-
bodies are more persistent and usually more specific. Absence of this
type of antibody to a specific virus usually indicates susceptibility to
the disease caused by that virus. A disadvantage of this test is that it
requires that the agent be propagated in the laboratory. For this reason,
the test may take several weeks to complete. In addition, neutralization
tests are not available for all viruses.
The hemagglutination inhibition test is another serologic technique
used by the virologist. As the name suggests, the basis of this test is that
antibodies against specific viruses inhibit the natural ability of that
virus to agglutinate red blood cells. Myxoviruses (mumps, measles, in-
fluenza), adenoviruses, arboviruses, certain enteroviruses, poxviruses,
and rubella virus all have the ability to cause hemagglutination. Like
neutralizing antibodies, those that prevent hemagglutination are usually
present long after recovery from the infection. The hemagglutination
inhibition test is useful, therefore, to determine susceptibility to infection
as well as to document a recent infection, if paired sera are examined. It
may also be used to identify a viral isolate. In contrast to neutralization
tests, hemagglutination inhibition tests are both easy and rapid to
perform.
In summary, the following procedures are employed in diagnostic
virology: (1) inoculation of animals and tissue culture, (2) serological
tests to detect complement fixing, hemagglutination inhibiting, and
:i i
neutralizing antibodies. These techniques are used to document infec- I

tion on the basis of a significant rise in antibody titer, to identify a cul-

tured virus, or to determine susceptibility to a given disease.
Several other immunologic tests are also frequently utilized in diag-
nostic virology. Quantitative measurements of IgM antibodies by gel
diffusion or the fluorescent antibody technique may be helpful in de-
termining whether intrauterine infection has taken place. Gel diffusion
has also been used to determine the type of antigen present in skin
lesions such as crusts or vesicles. 6 • 26 The fluorescent antibody technique
has also been especially useful recently in the diagnosis of infectious
mononucleosis.
The pediatrician himself may often utilize another immunologic
tool, the skin test. While this type of test is commonly used in the de-
termination of susceptibility to mumps, recent evidence suggests that
it is of questionable value because of both false positive and false nega-
tive results. 4 However the skin test is of use in the diagnosis of cat
scratch disease.

SUBMISSION OF MATERIAL TO LABORATORIES

Most large medical centers employ a clinical virologist who is

trained and equipped to carry out viral diagnostic procedures. If this
facility is not available, a state health department can usually do the
necessary studies. It is advisable to consult the laboratory to determine
76 ANNE A. GERSHON

what type of specimens they require and how these specimens are to
be collected and transported.
In general, body fluids which are to be cultured should be submitted
in a sterile tube. Since most viruses are destroyed by drying, swabs
should be dipped in a sterile salt solution (for example, Hanks balanced
salt solution). Tissues may be sent in any sterile container containing
Hanks solution to prevent drying. Many viruses are unstable at refrigera-
tor or ordinary freezer temperatures. Therefore, specimens for culture
should be packed in dry ice. Ideally, specimens should be inoculated
immediately upon collection. Material for culture should be obtained
as early in the course of the disease as possible, to provide the best chance
of recovering the virus.
It must be emphasized that to document a viral infection on the
basis of an antibody rise it is necessary to obtain an acute serum, taken
as early in the disease as possible, and a convalescent serum, taken 2 to 4
weeks later. Only then is there a possibility of documenting a four-fold
or greater rise in antibody titer. The antibody titer of only one serum
specimen from a patient is usually meaningless, because the antibodies
may have been present for years. It is worthwhile to obtain an acute
serum from any severely ill patient in whom no diagnosis has been
established. This serum can be stored, and if needed later it may be used
for viral studies. When testing for susceptibility, one serum specimen
will suffice. Serum specimens may be stored frozen, or they may be kept
in the refrigerator for a limited period. They may be mailed to the
laboratory without being under refrigeration (Table 2).
Table 2. Specimens That Should be Sent to the Laboratory
for Viral Diagnosis

PHARYN· CEREBRO·
DISEASE OR GEAL SPINAL VESICULAR
SYMPTOM SWAB STOOL URINE FLUID FLUID SERA

Neonatal X X X X If lesions are Maternal and infant,

viral disease present repeat in 3 months
Lymphadenopathy X X Acute and
convalescent
Infectious • Acute and
mononucleosis convalescent
Orchitis, parotitis X X If symptoms Acute and
are present convalescent
Rash X X X If lesions are Acute and
present convalescent
Acute respiratory X X Acute and
disease convalescent
Aseptic meningitis, X X X X If lesions are Acute and
encephalitis present convalescent
Myocarditis X X X Acute and
convalescent
Hepatitis X X Acute and
convalescent
Vaccine- X X X If lesions are Acute and
associated disease present convalescent
Susceptibility to One serum
specific disease specimen
DIAGNOSTIC VIROLOGY 77
Finally, the laboratory must be notified that the specimens are being
sent, so that the material can be processed quickly and appropriately.
A brief clinical history should accompany all specimens. There are
numerous viral agents which cause human disease, all with highly
specific but varying growth requirements, and there are numerous dif-
ferent techniques for viral detection as well. The virologist, must, there-
fore, have some notion of what he is looking for, so that he may employ
the proper methods to detect viral infection.

INSTANCES IN WHICH VIRAL STUDIES ARE INDICATED

Having briefly described methods used in diagnostic virology, it is

now possible to discuss specific situations in which viral diagnosis might
profitably be attempted (Table 3).

NEONATAL INFECTIONS

Cytomegalic Inclusion Disease, Rubella, Herpes Simplex Infection

Hepatosplenomegaly, jaundice, chorioretinitis, and thrombocyto-
penic purpura are all symptoms of congenital cytomegalic inclusion dis-
ease, rubella, and herpes simplex infection of the newborn infant.
Despite the similarity of the initial symptoms, the later manifestations
and prognoses of these diseases vary. Thus, it is important to make the
correct diagnosis.
Viral isolation is the most accurate method of diagnosing these dis-
orders, and therefore specimens of urine, cerebrospinal fluid, saliva, and
vesicle fluid, if present, should be obtained for culture. Infants con-
genitally infected may excrete virus for months or years/' 20. 41 and
therefore cultures may be appropriate even in older children. Sera from
both mother and child should be obtained for antibody determinations.
For infants under 12 months of age the indirect fluorescent antibody test
for specific IgM antibodies may be useful. Antibody to cytomegalovirus
and herpes simplex virus has been detected by this method. 14 , 32 Since
IgM antibody is not transferred across the placenta, any IgM present
must have been made by the fetus or infant. Thus if specific IgM is
detected, it indicates actual infection.
The complement fixation test, although less sensitive, may also be
employed, Complement fixing antibodies are transferred across the
placenta. If the titer is positive, therefore, the test must be repeated in a
month or two to determine whether the antibody titer is rising or falling,
In certain infected infants, complement fixing antibody cannot be
demonstrated early in life but becomes detectable after several months,
It may be necessary, therefore, to determine antibody titers of infants
at 2 to 3 months if neonatal sera had no detectable complement fixing
antibody and infection is still suspected, Rubella antibody titers are
measured by the hemagglutination inhibition test and are more fully
discussed in another article (p, 87),
78 ANNE A. GERSHON

Table 3. Syndromes and the Viruses with Which They

Are Commonly Associated

SYNDROME VIRUS

Neonatal jaundice, thrombocytopenia, Cytomegalovirus

hepatosplenomegaly, chorioretinitis Rubella virus
Herpes simplex virus

Lymphadenopathy Epstein Barr virus or herpes-like virus

Cytomegalovirus
Cat scratch disease virus

Parotitis Mumps virus

Parainfluenza virus
Lymphocytic choriomeningitis virus
Enterovirus

Rash Varicella-zoster virus

Herpes-simplex virus
Variola virus
Vaccinia virus
Rubella virus
Coxsackie virus
ECHO virus
Measles virus

Acute respiratory disease RespiratOry syncytial virus

Parainfluenza virus
Adenovirus
Influenza virus
Enterovirus
Rhinovirus

Encephalitis and aseptic meningitis Measles virus

Mumps virus
Polio virus
ECHO virus
Coxsackie virus
Herpes simplex virus

Myocarditis Coxsackie B virus

ECHO virus
Mumps virus
Measles virus

Hepatitis Cytomegalovirus
Hepatitis associated antigen
Epstein-Barr virus

Vaccine-associated disease Measles virus

Polio virus
Mumps virus
Vaccinia virus
Rubella virus
Smallpox virus
--
DIAGNOSTIC VIROLOGY 79
A few more points should be noted in regard to herpes simplex in-
fection. The maternal genital tract has been implicated as the source of
infection of infants. Therefore viral cultures should be obtained on all
pregnant women with symptoms suggestive of herpes progenitalis. A
woman at term with proven herpes progenitalis and intact membranes
should probably have a cesarian section. 31 Although prenatal dissemina-
tion of herpes simplex virus despite intact fetal membranes has been
reported,2B.39 avoidance of contact between the baby and the mother's
genital tract may prevent infection of the infant. Finally, it is important
to diagnose neonatal herpes simplex infection because there have been
recent reports of successful treatment of encephalitis due to this virus
with idoxuridine (IDUR),u
Miscellaneous Infections
It is known that infants with congenital infections may have in-
creased levels of IgM. Screening of infants for silent infections by meas-
urement of IgM, however, has not been widely used. False negative and
false positive results have been found and, in addition, infants with
elevated levels were usually suspected clinically of infection before the
IgM test was completed. This test has not been recommended, there-
fore, as a routine screening test for infected infants.24
Although measurement of hepatitis-associated antigen has recently
been used to diagnose serum hepatitis,25.36 extensive studies for the
detection of neonatal hepatitis have not been reported.
If maternal varicella occurs at the time of delivery, measurement of
varicella antibody in both maternal and cord sera should be performed.
Infants with demonstrable complement fixing antibody are passively
protected from chickenpox, while those without antibody are probably
susceptible. 5
Newborn infants with myocarditis and encephalitis should have
stool, urine, and cerebrospinal fluid cultures for Coxsackie B virus.
Neonatal vaccinia has been reported as a rare complication of
smallpox vaccination during pregnancy.12 Vesicle fluid from a suspected
case should be submitted for culture, as well as serum from mother and
baby for antibody studies.

DISEASES OF CHILDREN-SYMPTOMS INDICATING VIRAL STUDIES

Lymphadenopathy
When the cause for persistent lymphadenopathy cannot be de-
termined by the usual laboratory means, viral studies may provide a clue
to the diagnosis. Among the viral causes of lymphadenopathy, infectious
mononucleosis, acquired cytomegalic inclusion disease, and cat scratch
disease are the most important. Occasionally the salivary gland enlarge-
ment noted in mumps may be confused with lymphadenopathy as well.
Although infectious mononucleosis in childhood is frequently
asymptomatic,IB it may cause fever, pharyngitis, and lymphadenopathy,
especially in adolescents. In adults the diagnosis is usually made by the
 80 ANNE A. GERSHON

presence of a positive heterophile antibody test. It is well known, how-

ever, that this test is often only transiently positive in children.
Recently, infectious mononucleosis has been linked etiologically to
the Epstein-Barr (EB) virus or herpes-like virus in Burkitt's tumor
cells. 9 ,lO,17 Antibodies to this virus are detected by direct or indirect
immunofluorescence, and complement fixation. In contrast to heter-
ophile antibodies, antibodies to Epstein-Barr virus persist for many years.
It has been well documented that the appearance of these antibodies
during the course of a disease indicates infectious mononucleosis. It
has not yet been possible to propagate this agent in tissue culture.
Therefore, cultures are not useful for diagnostic purposes.
Acquired cytomegalic inclusion disease, in which symptoms similar
to infectious mononucleosis have been reported to occur,23 may be
diagnosed by culture of urine, blood, or saliva. Acute and convalescent
sera for complement fixing antibody determinations should also be
submitted to the laboratory. Although complement fixation tests are
performed in 24 hours, the virus grows slowly in tissue culture. Often
it takes 2 to 3 weeks before this diagnosis can be made by propagation
of the virus.
The causative agent of cat scratch disease is unknown. It is thought
to be a member of the lymphogranuloma venereum group,7 although a
virus morphologically similar to herpes simplex virus has also been
implicated. 21 Culture and antibody determinations for diagnostic pur-
poses are unavailable. However, a skin test may be used to assist in the
diagnosis. Pus from a patient with cat scratch disease is used for prepara-
. tion of the antigen. 27
At times mumps may cause swelling of the submaxillary or sub-
lingual glands, and not the parotid glands. On the other hand, parotitis
may be caused by suppuration, neoplasm, or calculus rather than
mumps. In such situations cultures for mumps virus are indicated.
Mumps virus may be grown from the patient's saliva or urine. Serologic
tests also may be helpful in making this diagnosis. Complement fixing
antibody to the "soluble" portion of the virus (believed to be ribonucleo-
protein) rises early in mumps, while antibody to the viral component
(protein coat) appears later. Thus a complement fixation test positive for
antibody against soluble mumps antigen indicates recent mumps in-
fection. 16 A four-fold or greater rise in antibody is also indicative of
mumps.
Rash
The vesicular rash of chickenpox is familiar to all pediatricians.
Viral studies on every patient with varicella would be of little use to doc-
tor or patient. Occasionally, however, in this age of jet travel, it is neces-
sary to distinguish a severe case of varicella from smallpox. This is most
rapidly accomplished by a gel diffusion test which can be used to detect
smallpox26 or varicella-zoster antigen in vesicle fluid. 6 The results of this
laboratory procedure may be obtained within 4 hours. Vesicle fluid from
the patient in question is collected in a capillary pipet. This fluid is
allowed to diffuse against concentrated serum containing known vari-
DIAGNOSTIC VIROLOGY 81
cella antibody on a gel-coated microscope slide. If the patient has
chickenpox, a precipitin line will be visible in the gel within 3 to 5 hours.
Smallpox and chickenpox may also be differentiated by culture of vesicle
fluid, although this procedure takes at least 2 or 3 days. Antibody de-
terminations are of little use for this purpose, since complement-fixing
antibodies may not appear until the second week of either disease.
Herpes simplex infection can produce a zosteriform rash on either
the trunk or the face. 40 When it is necessary to distinguish herpes simplex
from varicella-zoster infection-as, for example, when there is corneal
involvement, or when there is risk of transmission of chickenpox from
a person with zoster- viral studies are indicated. Complement fixation
tests are not adequately specific, since cross-reactions occur between
these two viruses. In addition there is no relationship between presence
of serum complement fixing antibody and recurrences of herpes simplex
infection. Moreover, a rise in antibody titer may not occur during re-
currences. Vesicle fluid cultures are more likely to lead to the correct
diagnosis. Herpes simplex virus will cause a cytopathic effect in human
embryonic lung fibroblasts in as little as 24 hours, whereas varicella-
zoster virus will take at least 2 or 3 days to become apparent. Such iso-
lates may be positively identified by complement fixation. Again, if gel
diffusion tests with vesicle fluid can be performed, these will lead to the
most rapid diagnosis.
If it should be necessary to determine whether measles, rubella,
ECHO, or Coxsackie virus is the cause of a maculopapular rash, acute
and convalescent sera and throat and stool specimens should be obtained.
A mild disease caused by Coxsackie A-16 virus is often seen in young
children. 3s Typically, fever, mouth ulcers, and vesicular lesions on the
palms and soles are present. If there is any question as to the etiology of
these symptoms in a young child, the virus can be isolated from the
mouth and the stool. Sera for acute and convalescent antibody de-
terminations should also be submitted to the laboratory.
Acute Respiratory Disease
Acute infections of the respiratory tract in children are frequent and ·1I'
usually self-limited. Routine virologic studies, which are very costly, 1

are not indicated. The most common viral pathogens of the upper and
lower respiratory tract include respiratory syncytial viruses, adeno-
viruses, enteroviruses, myxoviruses and paramyxoviruses. Occasion-
ally, it is necessary to distinguish between pertussis and adenoviral in-
fection, especially when a child who is inadequately immunized against
pertussis presents with a severe cough. t. 33 Adenovirus may be cultured
from nasopharyngeal swabs and stool. Antibody studies are useful, since
adenoviruses share a common complement fixing antigen. Specific
diagnosis is made by hemagglutination inhibition tests.
Encephalitis and Aseptic Meningitis
There are many viruses which cause encephalitis and aseptic
meningitis. When the clinician encounters a case of aseptic meningitis,
acute and convalescent sera for antibody studies and stool for culture
82 ANNE A. GERSHON

should be obtained. The most frequent viral causes of aseptic meningitis

are mumps and the enteroviruses - the ECHO and Coxsackie viruses.
The most common viruses which cause encephalitis are the arbo-
viruses, measles, mumps, and herpes simplex viruses. Acute and con-
valescent sera should be obtained, as well as blood, cerebrospinal fluid,
stool, and mouth swabs for culture. These tests will suffice to confirm
the diagnosis of an arbovirus infection such as Venezuelan, eastern,
or western equine encephalitis. The diagnosis of mumps meningoen-
cephalitis is easily made clinically unless the encephalitis occurs, as it
occasionally does, without parotid swelling. Again, in this case, a com-
plement fixation test for antibodies against the soluble and viral compo-
nents of mumps virus or acute and convalescent sera to demonstrate a
rise in mumps antibody titer will be of use.
The incidence of measles has declined in the United States because
of wide-spread use of measles vaccine. Consequently, measles en-
cephalitis has become rare. This diagnosis is usually made clinically
when a patient convalescing from measles develops fever and central
nervous system abnormalities. The disease is thought to represent an
allergic phenomenon.
The etiologic agent of Dawson's encephalitis or subacute sclerosing
panencephalitis is also probably measles virus. Affected children develop
dementia and seizures many years after measles infection. Electron
microscopy of brain tissue from these patients has revealed structures
resembling measles virus. 22 A high antibody titer to measles virus is an
associated finding. Recently, measles virus has been propagated from
brain tissue from patients with subacute sclerosing panencephalitis. 34
If one suspects that a patient has this disease, the diagnosis may be
made by the clinical findings as well as high serum antibody titers to
measles virus.
Herpes simplex causes a severe encephalitis in infants, children,
and adults. Serum antibody titers may not be elevated, and it may not
be possible to isolate the virus from cerebrospinal fluid. In such a case,
if one has good reason to suspect herpes simplex encephalitis, a biopsy
of the brain may be indicated, to obtain a positive diagnosis.t 5 Recently,
as already mentioned, this disease has been treated successfully with
IDUR.
Myocarditis
Known causes of myocarditis include ECHO virus, Coxsackie B
virus (especially in newborns) and rarely, mumps and measles virus.
Acute and convalescent sera, and throat, urine, and stool specimens
should be submitted to the laboratory for culture if these diagnoses
are suspected.
Unexplained Fever
Fever of unknown origin without other symptoms is not usually an
indication for viral studies. In certain instances, infectious mononucleo-
sis or cytomegalic inclusion disease may be suspected. Viral diagnosis of
these diseases has already been discussed.
DIAGNOSTIC VIROLOGY 83
Hepatitis
Recently it has been noted that in long-incubation type or "serum"
hepatitis a new antigen, termed hepatitis associated antigen (HAA)
(also termed "Australia antigen") is present in the serum. 2 • 3. 25. :3(l This
agent is currently under investigation as the possible cause of long-
incubation type hepatitis. Hepatitis associated antigen is detectable by
gel diffusion and by complement fixation. 36 • 37 It is useful for differentiat-
ing between the two types of hepatitis, and for screening blood donors.
This antigen appears early in the course of hepatitis, before jaundice
occurs. In some patients, this antigen persists for months or years, while
in others it is more transient. A comparable agent has not been identified
for short incubation or "infectious" hepatitis.
Acquired cytomegalic inclusion disease and infectious mononucle-
osis may also produce abnormal liver function tests and jaundiceY To
differentiate between these diseases, samples of urine and saliva for
culture, and paired sera for antibody tests against cytomegalovirus and
Epstein-Barr virus should be submitted to the laboratory.
Diseases Following Vaccination
Illnesses which follow vaccination against viral diseases may be
divided into two categories: (1) immediate complications related to the
vaccine, and (2) late occurrence of the specific disease due to vaccine
failure and subsequent exposure.
Included in the first category are complications following polio,
measles, and smallpox vaccination. There are no contraindications to i
the use of live oral polio vaccine (OPV) in children. Although OPV is I
thought to be one of the safest vaccines in use today, there have been I
rare reports of vaccine-associated cases of poliomyelitis. H} In such
situations, examination of stool specimens and of sera may indicate
whether the infection is vaccine-associated. It can be determined by
serologic testing of the viral isolate whether wild or vaccine virus is re-
sponsible for the illness. In addition, if the disease is due to a virus other
than polio, this will be elucidated by culture and serology.
Encephalitis has been reported as a rare complication of measles
vaccination. A recent report of all cases in the United States from 1965
to 1967, during which time approximately 15 million doses of vaccine
were administered, described 23 cases of neurologic disorders following
vaccinationYo In no instance was proof available that the illness was
vaccine-induced. Several children showed evidence of herpes simplex
infection of the nervous system. Thus it is apparent that if neurologic
symptoms occur following administration of measles vaccine, diag-
nostic procedures are of great importance. Cerebrospinal fluid, stool,
and swabs of the throat should be collected and tested to identify the
responsible vi.rus. Acute and convalescent sera should also be obtained.
Complications following smallpox vaccination are usually fairly
obviously related to the vaccine, and cultures may not be necessary.
Occasionally, one may have to differentiate between varicella, or im-
petigo, and disseminated vaccinia or herpes simplex in a patient with
eczema. Swabs of the infected area or vesicle fluid may be submitted to
84 ANNE A. GERSHON

the laboratory for culture. When encephalitis occurs following smallpox

vaccine, samples of urine, stool and cerebrospinal fluid should be
cultured, since it is important to determine, if possible, whether the
encephalitis is related to the vaccine or is merely coincidental.
Illnesses which appear to be mumps or measles may occur months
or years after vaccine administration. Whether or not these diseases are
due to vaccine failure is of great importance in vaccine surveillance, and
ultimately, of course, to clinicians. Several cases of measles have been
reported many years after vaccination. 29 This has been reported in
children who were vaccinated under 1 year of age and who received gam-
ma globulin with the vaccine. When measles is suspected in a previously
vaccinated child, urine and throat swabs should be submitted for culture.
Antibody determinations of acute and convalescent sera may provide
confirmatory evidence of recent infection.
Some children who received mumps vaccine have developed
parotitis months or years later. However, not all parotitis is due to
mumps. Bacterial and other viral infections, as well as parotid duct cal-
culi, may produce similar symptoms.:15 • 42 Mouth swabs and urine speci-
mens should be submitted for culture in post-vaccination parotitis. In
addition, acute and convalescent sera for antibody determinations
against mumps virus should be collected.

SUSCEPTIBILITY

At present there are neutralization or hemagglutination inhibition

tests, which measure antibody of long duration, for many viral diseases.
Tests for immunity or susceptibility may be performed for measles,
mumps, rubella, and polio. Unfortunately, there is as yet no reliable
routine test for immunity to chickenpox.

SUMMARY

A review of the indications for viral diagnostic procedures has been

presented. Specific diagnosis is unnecessary and impractical in every
patient with a suspected viral illness. In certain instances, however, the
documentation of a viral disease by culture or serology may be of great
value to the patient. The pediatrician must therefore use his judgment
as to when to study the patient, and should resist the temptation simply
to tell an anxious parent, "Don't worry; it's just a virus."

REFERENCES

1. Blattner, R. J.: The whooping cough syndrome. J. Pediat., 65:150,1964.

2. Blumberg, B. S., Alter, H. J., and Visnick, S.: A "new" antigen in leukemia sera. J.A.M.A.,
191 :541, 1965.
3. Blumberg, B. S., Gerstley, J. S., Hungerford, D. A., London, W. T., and Sutnick, A. I.: A
serum antigen (Australia antigen) in Down's syndrome, leukemia, and hepatitis. Ann.
Intern. Med., 66:924,1967.
-
DIAGNOSTIC VIROLOGY 85
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