ACCEPTABLE SPECIMEN:
1. Dermal lesion/vesicle, mucosal lesion/vesicle, eye swab, vaginal swab, rectal swab, or skin biopsy.
2. Spinal fluid collected by lumbar puncture, ventricular or suboccipital tap, or through shunt.

Note: Swab specimens collected from multiple sites (eye, skin, mouth, rectal) on neonatal patients for purposes of HSV screening may be combined into one VTM tube for PCR testing.

Causes for Rejection:
1. Specimen not labeled with patient's name, MRN, date/time of collection, collector's initials.
2. Specimens previously used by other laboratories - "shared" specimen not acceptable.
3. Calcium Alginate swab or any swab with a wooden shaft, or swab transported in gel.

Collect specimen as follows:
LESION:
1. Aseptically unroof lesion.
2. Firmly sample the base of lesion with a plastic shaft polyester swab moistened with sterile saline to collect infected epithelial cells.
3. Place swab into the tube of Viral Transport Media (VTM) breaking off shaft of swab, if necessary. Cap tube tightly.
4. Double-bag the labeled VTM tube and submit to the laboratory.

VESICLE:
1. Cleanse vesicle with sterile saline.
2. Carefully aspirate vesicle fluid with syringe. Alternately, open the vesicle with a needle or a scalpel blade and use a plastic shaft polyester swab to collect fluid and cellular material by vigorously sampling the base of the lesion.
3. Place vesicle fluid into VTM tube. Alternately, break off swab into VTM tube. Cap tube tightly.
4. Double-bag the labeled VTM tube and submit to the laboratory.

EYE:
1. Collect material from the lower conjunctiva with a flexible, fine-shafted polyester swab moistened with sterile saline.
2. Place the swab into the VTM tube and cap tightly.
3. Double-bag the labeled VTM tube and submit to the laboratory.

RECTAL:
1. Carefully insert a plastic shaft polyester swab just beyond rectal sphincter.
2. Rotate swab gently to sample anal crypts.
3. Place the swab into the VTM tube breaking off shaft as necessary and cap tightly.
4. Double-bag the labeled VTM tube and submit to the laboratory.

VAGINAL:
1. Use a plastic shaft polyester swab to vigorously swab the lesion if present.
2. If lesion is not present, remove mucus from the cervix with a swab and discard swab.
3. Firmly sample the endocervix with a fresh plastic shaft polyester swab by rotating the swab for 5 seconds.
4. Carry out a vulvar sweep with a second swab.
5. Place the swabs in the VTM tube breaking off shaft as necessary and cap tightly.
6. Double-bag the labeled VTM tube and submit to the laboratory.

SPINAL FLUID:
1. Decontaminate skin with povidone iodine.
2. Perform sterile lumbar puncture; ventricular or suboccipital tap.
3. Send 1-5 mL of spinal fluid in a screw-capped, sterile collection tube.
4. Submit immediately to the laboratory.

Cause for Rejection:
3. Insufficient volume of CSF.
4. Patient over 2 y.o. with CSF protein <= 50 mg/dL and leukocyte count <= 5. Specimens from infants <= 2 y.o. will be tested regardless.

SHUNT FLUID - CSF:
1. Decontaminate skin and catheter with povidone iodine.
2. Aseptically aspirate fluid through shunt.
3. Send 1-5 mL of shunt fluid in a screw-capped, sterile collection tube.
4. Submit immediately to the laboratory.


Note: Submit CSF in sterile specimen container (not VTM).

NOTE for BLOOD:
Collect 4-5 mL blood in EDTA (LAV) tube. Order GEN CODE B; with comment "blood for HSV PCR to Viracor". Send specimen to the External Referral Laboratory.

DELIVER IMMEDIATELY TO MICROBIOLOGY IN A TIGHTLY SEALED CONTAINER WITH NO EXTERNAL SPILLAGE. Refrigerate specimen if transport is delayed.

Causes for Rejection:
1. Container container leaking or contaminated.
2. Specimen delayed in transit.

Test is indicated for the evaluation of patients with signs or symptoms of aseptic meningitis and/or encephalitis.  It is also indicated as an aid to diagnose HSV infection in patients with skin, mucosal, genital, or ocular lesions.  This assay should be used with other criteria including symptoms and clinical presentation.

Herpes simplex virus (HSV-1 and HSV-2) can cause a variety of diseases including: oral or perioral ulcers, genital and non-genital skin lesions, ocular infections, encephalitis, meningitis, and disseminated disease in the neonate.  Molecular methods (i.e., PCR) are now the diagnostic gold standard owing to superior sensitivity and more rapid turn around time as compared to culture.

CSF:
It is difficult to recover HSV from CSF by culture. PCR is more sensitive and is now the standard of care. This leads to more focused therapy for infected patients and prevents prolonged hospitalization of patients that do not have HSV in the CNS.

LESIONS, OCULAR INFECTIONS, DISSEMINATED DISEASE:
While viral culture has remained the standard diagnostic method for isolating HSV from anatomic locations other than the central nervous system, real-time HSV PCR assays have emerged as a more sensitive method to confirm HSV infection in clinical specimens obtained from genital ulcers, and mucocutaneous sites.  PCR is particularly useful for the detection of asymptomatic HSV shedding.  Earlier diagnosis by real-time HSV PCR assays may reduce the transmission of infection during reactivation syndromes that are manifested only by asymptomatic viral shedding.

Positive results are reported to indicate the specific HSV virus Type I or II detected. Occasionally, results will not be available due to inhibitory substances in the specimen (see limitations).

NOTE: Consult with the Clinical Microbiology Laboratory if susceptibility testing is warranted.

A negative result does not preclude active disease.  Specimen collection technique in addition to the age of the lesion is likely to influence test performance.

Specimens grossly contaminated with blood, oral washes, or other substances may inhibit the PCR and produce false negative results. The high sensitivity of PCR amplification requires that the specimens be processed in an environment where contamination of the specimens by HSV DNA is not likely to occur.

Real-Time Polymerase Chain Reaction (PCR)

The performance of this PCR test was verified in the Clinical Microbiology Laboratory at Duke University Medical Center. This test is run in accordance with the Diasorin MDX package insert and as approved by the U.S. Food and Drug Administration.