Download Patients with Suspected Herpes Simplex Encephalitis: Rethinking

BRIEF REPORT
Patients with Suspected Herpes
Simplex Encephalitis: Rethinking
an Initial Negative Polymerase
Chain Reaction Result
Ana A. Weil, Carol A. Glaser, Zahwa Amad, and Bagher Forghani
Viral and Rickettsial Disease Laboratory, Division of Communicable Disease
Control, California Department of Health Services, Richmond
A statewide encephalitis diagnostic project of the California
State Department of Health Services found that herpes simplex virus 1 DNA may not be detectable by molecular methods early in the clinical course of herpes simplex encephalitis.
We describe 3 patients with herpes simplex encephalitis (HSE)
whose initial PCR results were negative for herpes simplex virus
1 (HSV-1).
Patient 1. A 10-year-old previously healthy white girl with
chief complaints of increasing lethargy and fever was seen in
the emergency department. At the time of admission, she was
aphasic, inattentive, and drooling. She had swollen lips and
experienced difficulty opening her mouth. Her vital signs were
as follows: temperature, 39.1C; pulse rate, 126 beats/min; and
respiratory rate, 24 breaths/min. She responded to her name
with a nonverbal sign of acknowledgment and visually tracked
her parents, but she could not follow commands. At admission,
her complete blood count was within the normal range, and
findings of chest and neck imaging were also normal. Three
days before admission, the patient had experienced 2 episodes
of vomiting, somnolence, and symptoms of an upper respiratory infection. The patient had briefly experienced disorientation and had “difficulty finding words” 1 day before admission; at that time, she was febrile and lethargic.
Evaluation of the patient’s medical history disclosed no abnormal findings. Findings of a review of systems were unReceived 10 September 2001; revised 19 November 2001; electronically published 21 March
2002.
Financial support: Centers for Disease Control and Prevention Emerging Infections Program
(U50/CCU915546-03).
Reprints or correspondence: Dr. Bagher Forghani, Viral and Rickettsial Disease Laboratory,
California Dept. of Health Services, 850 Marina Bay Pkwy., Richmond, CA 94804
([email protected]).
Clinical Infectious Diseases 2002; 34:1154–7
2002 by the Infectious Diseases Society of America. All rights reserved.
1058-4838/2002/3408-0019$03.00
1154 • CID 2002:34 (15 April) • BRIEF REPORTS
remarkable. The patient had experienced no recent episodes of
rash, dysphasia, dysarthria, weight loss, headache, neck stiffness,
or changes in vision. She was not taking any medications on
a regular basis, and no significant animal exposure or foreign
travel was identified.
Analysis of a CSF sample obtained from lumbar puncture (LP)
performed at the time of admission showed a WBC count of 90
cells/mL (75% lymphocytes, 14% monocytes, and 11% neutrophils), a protein level of 26 g/dL, and a glucose level of 73 mg/
dL (table 1). A CT scan of the head showed a small area of
decreased attenuation in the left frontal lobe and associated mass
effect in the left lateral lobe. Treatment with doxycycline and
acyclovir was initiated. CSF samples obtained during the initial
LP were found to be negative for HSV-1 by PCR performed at
3 laboratories: the hospital laboratory (targeted HSV-1 screening), a private laboratory (targeted HSV-1 screening), and the
state laboratory (consensus sequence PCR testing for herpesviruses, including HSV-1) (table 2).
On day 2 of hospitalization, the patient began to develop oral
apraxia, and MRI showed multifocal, bilateral T2 prolongation
with patchy enhancement, which was most extensive in the left
temporal lobe. She had a complex partial seizure on day 3 of
hospitalization.
Because of temporal lobe involvement and the lack of another
etiology, HSE was still suspected. On day 4 of hospitalization, a
second LP was performed; analysis of the CSF sample obtained
revealed a WBC count of 113 cells/mL (83% lymphocytes), a
protein level of 107 g/dL, and a glucose level of 59 mg/dL (table
1). HSV-1 was detected by PCR performed at both the state
laboratory and the hospital laboratory.
Patient 2. A 37-year-old man presented with fever, memory
loss, photophobia, and disorientation. MRI revealed lateral temporal lobe enhancement. Examination of a CSF sample obtained
from an initial LP performed at the time of admission revealed
pleocytosis and a slightly increased protein level (table 1). This
sample was found to be negative for HSV-1 on the basis of PCR
performed at the state laboratory and at an outside laboratory.
Seven days later, a CSF sample obtained during a second LP was
found to be positive for HSV-1 by PCR done at the state laboratory and at a different outside laboratory (table 2).
Patient 3. A 78-year-old woman had onset of HSE that
was characterized by fever and altered mental status that rapidly
progressed to seizures and coma. Because clinical deterioration
and significant temporal lobe edema were seen on MRI, she
underwent a temporal lobectomy. Her first CSF sample, which
was obtained at the time of admission, showed pleocytosis and
Table 1.
Clinical and laboratory characteristics of patients with herpes simplex encephalitis who had an initial negative PCR result.
Age in
years,
Patient sex
Race
WBCs/mL (% differential)
Clinical findings
Imaging method and finding(s)
At LP 1
At LP 2
MRI: edema of left temporal lobe
90 (75% L, 11% N)
113 (83% L, 13% M, 4% N)
1
10, M
W
Fever, lethargy, disorientation
aphasia, seizure
2
37, M
W
Fever, headache, disorientation CT: normal; MRI: lateral temporal
memory loss, confusion
lobe enhancement
330 (100% L)
3
78, F
W
Fever, lethargy disorientation,
seizures, coma
720 (50% L, 49% M)
NOTE.
CT: hemorrhage, edema of right
temporal lobe; MRI: edema
1356 (100% L)
624 (84% L, 9% M, 7% N)
Protein level,
mg/dL
Glucose level,
mg/dL
Serum IgG index
Acute ConvalesAt LP 1 At LP 2 At LP 1 At LP 2 phase cent phase
26
107
73
59
1.39
16.9
85
131
53
53
6.57
20.7
132
253
69
49
5.05
17.0
L, lymphocytes; LP 1, first lumbar puncture; LP 2, second lumbar puncture; M, monocytes; N, neutrophils; W, white. For each patient, the LP 1 was performed at the time of admission.
Table 2. Results of PCR analysis of CSF samples obtained from patients who
had an initial negative PCR result.
Patient
No. of days
between onset
of CNS symptoms
and LP 1
HSV PCR result (laboratory
a
where PCR was performed )
for sample from
LP 1
LP 2
No. of days
between LP 1
and LP 2
1
3
Negative (A, B, D)
Positive (A, D)
4
2
2
Negative (A, C)
Positive (A, B)
7
3
1
Negative (A)
Positive (A, B)
4
NOTE.
HSV, herpes simplex virus; LP 1, first lumbar puncture; LP 2, second lumbar puncture.
a
A, laboratory A (i.e., the California State Health Department Viral and Rickettsial Disease Laboratory; uses consensus-sequence PCR for herpesviruses and reports a detection rate of ⭐100
copies/mL); B, laboratory B (i.e., a private laboratory other than the hospital laboratory; uses targeted
HSV-1 PCR and reports a detection rate of ⭐10 HSV-1 copies/mL); C, laboratory C (i.e., a private
laboratory other than the hospital laboratory; uses targeted HSV-1 PCR and reports a detection rate
of ⭐100 HSV-1 copies/mL); D, laboratory D (i.e., the hospital laboratory; uses targeted HSV-1 PCR
and reports a detection rate of ⭐14 HSV-1 particles /mL).
an increased protein level; this sample was found to be negative
for HSV-1 by PCR performed at the state laboratory (table 1).
A second CSF sample, which was obtained during LP performed 4 days later, tested positive for HSV-1 by PCR performed at the state laboratory and at an outside laboratory
(table 2).
Discussion. HSV has been established as a common cause
of acute, necrotizing encephalitis that may present at any age.
With an estimated frequency of 1 case in 250,000–500,000 people per year, HSE is the most common cause of sporadic encephalitis in the United States [1]. Without receiving prompt
antiviral therapy, up to 70% of patients with HSE can die [2].
For this reason, rapid and accurate diagnostic procedures are
essential for management of the disease.
PCR analysis of CSF samples is recommended as the standard
method for diagnosing HSE [1]. Compared with brain biopsy,
which was the previous diagnostic standard, PCR is preferred
because of its high sensitivity and specificity as well as its noninvasive nature. The sensitivity and specificity of PCR for the
detection of HSV-1 have been documented as 91% and 92%,
respectively, in patients with biopsy-proven disease [1].
PCR has been documented as a useful method for diagnosing
HSE early in the course of infection [3]. However, there have
been reports of individuals with HSE for whom a CSF sample
obtained early in the course of disease was negative for HSV1 by PCR but for whom CSF samples obtained a few days later
were found to be positive by PCR [4–6]. In a prospective study
of 27 adult patients with acute viral encephalitis, 10 patients
had HSE diagnosed by PCR analysis of CSF samples. For 2 of
these patients, the CSF samples obtained at admission were
found to be negative for HSV-1 DNA, but samples obtained 4
and 7 days later had positive test results [4]. In a retrospective
study of 22 patients with HSV-1 encephalitis, 1 patient had a
negative PCR result for a CSF sample obtained on the third
day after the onset of disease but had a positive PCR result for
1156 • CID 2002:34 (15 April) • BRIEF REPORTS
a CSF sample obtained from an LP performed 5 days later [5].
There are 2 additional studies in which patients with HSE whose
initial PCR results were negative for HSV-1 early in the course
of infection were found to be positive for HSV-1 when a CSF
sample obtained subsequently was tested [6, 7].
The California Encephalitis Project (CEP) performs diagnostic tests on samples obtained from immunocompetent, previously healthy patients with encephalitis who are 16 months
of age. Although the state laboratory tests CSF samples for HSV1 DNA by use of a consensus-sequence PCR test, the CEP
strongly encourages that all patient samples also be analyzed
by PCR for targeted HSV-1 at another laboratory. Therefore,
the majority of patients enrolled in the CEP have a minimum
of 2 HSV-1 PCR analyses performed on CSF samples obtained
from an initial LP. As of 15 December 2000, the CSF samples
of 291 patients with encephalitis who were enrolled in the CEP
had undergone PCR for the detection of HSV-1. A commercial
PCR kit (Argene-Biosoft) was used for testing for herpesviruses.
This kit uses consensus primers (stair primers), that are capable
of amplifying HSV-1, HSV-2, cytomegalovirus, Epstein-Barr
virus, varicella-zoster virus, and human herpesvirus 6 in a single
reaction [8]. For serologic testing, an antibody assay for the
detection of HSV IgG is performed on paired serum samples
by use of an EIA, and an index value is determined for each
sample [9].
A total of 11 (3.8%) of 291 patients in the CEP had PCR
results that were positive for HSV-1. Three (27%) of these 11
patients had negative results of initial PCR testing for HSV-1
in CSF samples obtained at the time of admission; these 3
patients later had positive PCR results for CSF samples obtained
at 4, 7, and 4 days after the first LP. These patients had serologic
test results that supported the diagnoses of HSE and clinical
courses consistent with HSE (table 1). For all 3 patients, the
first CSF sample was obtained at the time of treatment initiation
(table 2). It has been speculated that an increase in the HSV-
1 DNA level occurs during the first few days of administration
of antiviral therapy, which may be responsible for this finding
[10]. HSE typically evolves over several days and is associated
with progressive deterioration of consciousness. The 3 patients
that we describe had CNS symptoms for only 1–3 days before
admission. This would suggest that these 3 patients presented
relatively early during the clinical course of HSE, and they may
have had a very low number of copies of the target HSV-1
DNA present in their initial CSF sample.
Conventional PCR methods for detection of HSV-1 were
used by the different laboratories involved in evaluation of these
patients; the methods had comparable sensitivities (table 2).
Nested PCR is considered more sensitive than conventional
PCR because it includes a second amplification step; theoretically, the procedure is able to detect a single copy of the target
HSV-1 DNA [7, 11]. Because the laboratories involved in diagnostic testing of these patients used conventional PCR, it is
important to be aware of the negative results obtained under
these circumstances. The initial negative PCR results that were
obtained for the cases we report are not likely to have been the
result of inhibitors, because the Elongase DNA polymerase
(Gibco BRL) used by our laboratory is less affected by hemoglobin than is Taq DNA polymerase. In addition, the initial
CSF samples all had !5 RBCs/mL, a finding that suggests that
minimal hemoglobin levels were present. Finally, inhibitors
generally are less problematic for PCR testing of CSF than they
are for PCR testing of other body fluids or tissue extracts.
Observation of initial negative PCR results for a small but
notable number of patients with confirmed HSE infection indicates that a PCR result that is negative for HSV-1 early in
the course of infection does not rule out the presence of HSE.
Diagnosis of HSE should be based on clinical findings in combination with the results of neuroimaging and diagnostic testing. For patients for whom HSV-1 infection is still suspected
(i.e., for those with temporal lobe involvement and no alternative diagnosis), a second LP and subsequent PCR testing for
HSV-1 should be considered.
Acknowledgments
We thank George F. Brooks, Giorgio Cosentino, Cynthia Cossen, David Cottam, Fred Drach, Sabrina Gilliam, Mary Ellen
Guroy, Somayeh Honarmand, David Lennette, Evelyne Lennette, Gordon Shell, James Sejvar, and Erica Steffe for their
valuable assistance.
References
1. Whitley RJ. Herpes simplex viruses. In: Scheld WM, Whitley RJ, Durack DT, eds. Infections of the central nervous system. 2nd ed. Philadelphia: Lippincott-Raven, 1997:73–89.
2. Whitley RJ, Soong S-J, Hirsch MS, et al. Herpes simplex encephalitis:
vidarabine therapy and diagnostic problems. N Engl J Med 1981; 304:
313–8.
3. Cinque P, Cleator T, Weber, Monteyne P, Sindic CJ, van Loon AM.
The role of laboratory investigation in the diagnosis and management
of patients with suspected herpes simplex encephalitis: a consensus
report. J Neurol Neurosurg Psychiatry 1996; 61:339–45.
4. Studahl M, Bergstrom T, Hagsberg L. Acute viral encephalitis in
adults—a prospective study. Scand J Infect Dis 1998; 30:215–20.
5. Guffond T, Dewilde A, Lobert PE, Caparros-Lefebvre D, Hober D,
Wattre P. Significance and clinical relevance of the detection of herpes
simplex virus DNA by the polymerase chain reaction in cerebrospinal
fluid from patients with presumed encephalitis. Clin Infect Dis 1994;18:
744–9.
6. Koskiniemi M, Piiparinen H, Mannonen L, Rantalaiho T, Vaheri A.
Herpes encephalitis is a disease of middle aged and elderly people:
polymerase chain reaction for detection of herpes simplex virus in the
CSF of 516 patients with encephalitis. J Neurol Neurosurg Psychiatry
1996; 60:174–8.
7. Aurelius E, Johansson B, Skoldenberg B, Staland A, Forsgren M. Rapid
diagnosis of herpes simplex encephalitis by nested polymerase chain
reaction assay of cerebrospinal fluid. Lancet 1991; 337:189–92.
8. Minjolle S, Michelet C, Jusselin I, Joannes M, Cartier F, Colimon R.
Amplification of the six major human herpesviruses from cerebrospinal
fluid by a single PCR. J Clin Microbiol 1999; 37:950–3.
9. Forghani B. Enzyme immunoassay systems. In: Lennette EH, ed. Laboratory diagnosis of viral infections. 2nd ed. New York: Marcel Dekker,
1992:105–25.
10. Roos KL. Pearls and pitfalls in the diagnosis and management of central
nervous system infectious diseases. Semin Neurol 1998; 18:185–96.
11. Persing DH. In vitro nucleic acid amplification techniques. In: Persing
DH, Smith TF, Tenover FC, White TJ, eds. Diagnostic molecular microbiology—principle and application. Washington, DC: American Society for Microbiology, 1993:51–87.
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