Herpes Simplex Workup

Updated: May 24, 2021
  • Author: Folusakin O Ayoade, MD; Chief Editor: Michael Stuart Bronze, MD  more...
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Workup

Laboratory Studies

Direct Method 

This method is used to demonstrate the presence of HSV in a suspicious lesion or in genital secretions. The vesicle should be unroofed with a sterile needle or scalpel, and a sterile Dacron or rayon swab with a plastic shaft should be rotated firmly in the base of the lesion to allow epithelial cells to be collected onto the swab. Ideally, more than one lesion should be sampled. Similarly for ulcerative lesions, a swab should be firmly rotated in the base of one or more lesions. The specimen should be held at 4°C and transported to the laboratory for further processing within 48 h. The test sensitivity declines if the vesicular lesion has been present for >24h and in patients with recurrent lesions than in those with first episodes. [21, 22]  

Culture

Herpes simplex virus (HSV) infection is best confirmed by isolation of the virus in tissue culture (the criterion standard for diagnosis). Tissue culture success is operator-dependent, but this modality can yield positive results within 48 hours of inoculation.

Characteristic cytopathic effect with ballooning of cells and cell death are observed, and death of the entire monolayer of cells may be rapid.

Immunofluorescent staining of the tissue culture cells can be used to quickly identify HSV and can distinguish between types 1 and 2. 

Tzank smear

This test is rarely used for diagnosis. The characteristic cytologic changes induced by HSV can be demonstrated in Tzank smears (see Procedures); however, this procedure does not distinguish between HSV-1 and HSV-2.

Rapid diagnosis (usually within an hour) is possible based on the histological appearance of the lesion.

Multinucleated giant cells and epithelial cells containing eosinophilic intranuclear inclusion bodies distinguish the lesions of herpesviruses.

Punch biopsy provides more reliable material for histological examination, particularly when lesions are infected with bacteria and fungi.

Polymerase chain reaction

Detection of HSV DNA in clinical specimens is possible with polymerase chain reaction (PCR) techniques. PCR is more sensitive than culture, allowing rapid laboratory diagnosis and increasing overall HSV detection rate by 24%. [23]  PCR detection of viral DNA is the gold standard for diagnosis of CNS infections. [23]  

In HSV encephalitis, PCR using CSF provides a rapid, noninvasive diagnostic technique that is as sensitive as brain biopsy. [24]

PCR has been used to detect HSV-2 as the cause of recurrent meningitis (Mollaret) and has shown a strong association between HSV-1 and Bell's palsy.

PCR can be used to detect asymptomatic viral shedding.

Direct fluorescent antigen (Immunofluorescence)

Cells scraped from ulcer bases can be stained with a direct fluorescent antibody, used to distinguish HSV-1 from HSV-2. Additionally, tissue culture cells can also be stained (see above). This procedure can usually be performed within 2-3 hours. [25]

Antibody testing

Antibody testing can demonstrate a primary seroconversion, particularly with HSV-1 in childhood. [1] If serology results are negative while viral culture of specimen is positive, one can assume primary infection.

Because of sero–cross-reactivity, HSV-1 and HSV-2 are not generally distinguishable unless a glycoprotein G antibody assay is available. Although there is a very close serological relationship between HSV-1 and HSV-2, they each encode a serologically distinct glycoprotein G ,gG-1 and gG-2. [21]  Testing for HSV-specific immunoglobulin M (IgM) antibodies is not available.

Antibody titer increases generally do not occur during recurrences of HSV infection. Therefore, the test is generally not used for the diagnosis of mucocutaneous HSV relapse.

Serum HSV antibody measurements are not of utility in the diagnosis of HSV encephalitis in adults. [26]

Antibody testing has been the mainstay of large-scale epidemiologic studies.

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Imaging Studies

Brain imaging studies in HSV encephalitis generally demonstrate focal localization in the temporal area that is associated with edema and contrast enhancement.

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Procedures

Tzanck preparation

Tzanck preparation is a time-honored procedure for assisting in the diagnosis of cutaneous herpesvirus infections. However, it does not easily distinguish HSV-1, HSV-2, and varicella-zoster virus.

Typically, an intact vesicle is used from which the vesicular fluid is aspirated by puncture with a sterile tuberculin syringe. This fluid can be used for viral culture or PCR.

Aspiration should facilitate complete collapse of the vesicle because it is not multiloculated as cutaneous poxvirus infections can be.

After aspiration, the vesicle should be unroofed aseptically.

Using a sterile instrument, the floor of the newly produced ulcer can then be scraped. The obtained material can be spread on a glass microscope slide and then dried and fixed for staining.

Staining can be performed with a Papanicolaou smear stain or, alternatively, whatever is available will suffice (eg, Gram, Giemsa, or Wright stain).

A positive result is the finding of multinucleate giant cells.

Direct fluorescent antigen

Using appropriate immunofluorescent antibody reagents, the smear can be used to distinguish different herpesviruses and nonherpesviruses that may be present (eg, vaccinia, smallpox). Viral inclusion bodies appear in UV microscope as bright green intranuclear particles. 

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