Synonym

• HSV
• HSV-1
• HSV-2

Tubes

The specimens collected are:

• Blood sample
• Red or tiger top tube (for serology)
• Lavender or pink top tube (for PCR and culture)
• 5-7 mL of venous blood
• Cerebrospinal fluid (for PCR, culture, serology
• Spinal fluid collection tube
• 0.5-1 mL of CSF
• Sample is collected from the affected site with a sterile swab and placed in a suitable transport container (provided by lab for viral culture, PCR)
• Vesicular skin lesions
• Genital area lesions
• Nasopharynx
• Oropharynx
• Bronchoalveolar lavage (BAL)
• Conjunctiva
• Tissue
• Bone marrow
• Lavender or pink top tube (for culture)
• 3-5 mL of bone marrow
• Tzanck smear
• Microscope slides and container with fixative (95% ethyl alcohol/provided by lab)
• Vesicle or fresh blister scrapings

Note on obtaining specimen from vesicle:

The fluid within the vesicle is needed for testing. Unroof several vesicles with an 18-25 gauge needle and use the swab to soak up the vesicular fluid.

Note on obtaining other specimens:

• For body fluids or other respiratory specimens (washings, lavage), use sterile screw-capped jar/tube (provided by lab)
• Tzanck preparation: A fresh blister is opened (as described above) then the base of the vesicle is gently scraped around the rim and is quickly spread evenly on a microscope slide with the fixative. Immediately re-immerse the slide in fixative. After collection, leave the slides in the fixative (95% alcohol) for 10 minutes and then air dry. Repeat the process for additional areas for better diagnostic yield. Label the slides with the patient’s name, specimen source etc.
• If anticipated time between collection and inoculation of cell culture is >3 hrs, store and transport specimen at 4° C. Do not freeze or allow the specimen to dry up
• If delayed, separate the serum and store at 4° C for 1–2 days or at –20° C for up to 1 year
• Reject samples
• If collected in Calcium alginate transport media for culture
• Hemolyzed, lipemic blood, or icteric samples
• Heparinized samples
• Handle blood samples gently to prevent hemolysis
• Send samples to lab immediately

Info

• Herpes simplex virus testing is a group of tests to detect the presence of herpes simplex virus (type 1 and 2), its DNA, or its antigen and antibodies
• Herpes simplex virus is a DNA virus of the herpesviridae family that exists as two main strains: Type–1 (HSV-1) causes most oral mucocutaneous infections and type–2 virus (HSV-2) causes most genital and neonatal herpes. Either type can cause invasive infection
• The tests to detect herpes simplex infection include:
• Tzanck test (microscopic examination of tissue scrapings)
• Herpes culture
• Herpes antibody testing
• Direct antigen testing
• Herpes DNA testing

Clinical

• The clinical utility of herpes simplex virus testing include:
• As an aid in the diagnosis of persons presenting with signs and symptoms of HSV infection (PCR or culture)
• As a screening test for HSV in persons who are at risk of developing HSV infection
• As a screening test in bone marrow recipients and donors (antibody testing)
• During pregnancy as a part of the TORCH panel
• Testing of infants born to mothers infected with HSV during delivery
• Evaluation of recent or past HSV infection (antibody testing)
• Universal screening for HSV antibodies is not yet recommended
• HSV causes a wide spectrum of disease conditions such as:
• Cold sores / Oral herpes / gingivostomatitis
• Neonatal HSV (Encephalitis)
• Genital herpes (primary and recurrent)
• Keratoconjunctivitis / herpes keratitis
• Aseptic meningitis / Encephalitis
• Herpetic whitlow
• Herpes dermatitis
• Visceral herpes (esophagitis, pneumonitis, hepatitis)
• The manifestations and clinical course of HSV infection depend on site of infection, age, gender, and immune status of host
• HSV-1 usually affects children between the age of 6 months and 3 years
• HSV-2 most commonly infects those between 18 and 25 years of age
• Recurrences of HSV-2 are 20% more common in males than in females
• Orofacial herpes
• This condition is most commonly caused by HSV-1 and is usually acquired during childhood (1-2 years), but can be acquired at any age.
• HSV-1 is primarily transmitted through respiratory droplets or direct contact with infected saliva and contaminated utensils
• Primary infection in children is usually mild, minimally symptomatic, or may manifest as painful sores on lips, gums, tongue, roof of the mouth, and inside the cheeks, with fever and myalgia
• Adults with primary infection are often more symptomatic
• Infection remains latent in the sensory (trigeminal) ganglia and may recur (reactive) due to certain stimuli such as fever, physical or emotional stress, menstruation, ultraviolet light exposure, local skin trauma, and axonal injury
• Genital herpes
• Over 50 million people are infected in the U.S. and 1 million new infections occur each year with genital herpes
• This infection is both widespread and contagious
• It is caused mainly by HSV-2 and in some cases by HSV-1. It is transmitted through sexual contact with an infected partner
• The incubation period ranges from 1 to 26 days (average 7 days) after exposure, with 90% of those infected having mild or no symptoms
• In males, the lesion commonly involves the glans penis, skin, mucosal surfaces of the prepuce, and frenal areas, and in females, it involves mucosal surfaces of the labia minora, clitoral hood, skin of the labia majora, buttocks, and thighs in severe cases. It is associated with fever, myalgia, headaches, discharges, painful urination, and inguinal lymphadenopathy
• Cross-infection of type 1 and 2 viruses may occur from oro-genital contact
• Neonatal herpes
• The incidence of neonatal HSV infection is estimated to be 1 in 3,000-20,000 live births per year, primarily caused by HSV-2 (80%) and HSV-1 (15-30%) through vertical transmission (maternal to fetal transmission).
• Approximately 70% of cases occur when the mother is asymptomatically shedding virus near the time of delivery
• Infected newborns may be premature and/or have low birth weight
• This infection can result in microcephaly, hydrocephalus, chorioretinitis, or vesicular skin lesions
• Postnatally acquired infection may present with three patterns such as:
• Disease localized to the skin, eye, or mouth
• Encephalitis, with or without skin, eye or mouth involvement
• Disseminated infection that involves multiple sites, including the central nervous system, lung, liver, adrenals, skin, eye, or mouth
• Complications associated with neonatal HSV infection include:
• Seizures
• Psychomotor retardation
• Spasticity
• Blindness
• Learning disabilities
• Death
• Persons who are immunodeficient, such as HIV/AIDS patients, those on chemotherapy, or those undergoing organ transplant have more frequent and severe outbreaks of HSV infection, which can be fatal
• There is currently no cure (but suppressive therapy is available) for HSV infections and once a primary infection of a HSV establishes latency, it will persist in the host's cells for life
• HSV along with human papilloma virus (HPV) infection is associated with higher risk of developing cervical cancer

Additional information

• Laboratory diagnosis of HSV infection includes:
• Tzanck test
• Cytopathologic examination of scrapings from herpes lesions to detect inclusion bodies (which are indicative of HSV infection)
• Multinucleated giant cells
• Cowdry type A intranuclear inclusion bodies
• The test is quick, but the diagnostic sensitivity is only 50-70%, and it cannot distinguish between HSV type 1 and 2 strains
• Cannot be used to differentiate between HSV and varicella zoster virus
• Herpes culture
• Virus isolation by culture is performed on the fluid sample taken from the lesion as early as possible (first three days of appearance), which gives close to 100% accuracy
• This technique has markedly less yield from older ulcerated sores, recurrent lesions, or latency
• Once the virus is grown in culture, determination of strain can be made (HSV-1 vs. HSV-2)
• The characteristic cytopathic effects (CPE) of HSV can be detected within 24-48 hrs of culture for 50% of HSV strains, and 5-7 days are required to detect remaining HSV strains
• The culture is generally not productive beyond more than five days after the patient becomes symptomatic
• Herpes DNA testing
• Polymerase chain reaction (PCR) is the method of choice for diagnosis, with a sensitivity of 95% and specificity of 99-100%
• PCR can identify specific strains of HSV and also asymptomatic viral shedding
• PCR is recommended in the following conditions:
• Specimen likely to have a low viral load such as in CSF (in herpes encephalitis, as the test is more rapid than the viral culture)
• CSF in unusual CNS syndromes or recurrent meningitis
• Viral cultures are negative
• Immunosuppressed or AIDS patients
• Infant has suspected neonatal herpes or keratitis
• Lesions are several days old and culture is likely to be negative
• Herpes antibody testing
• This test is useful in detection of recent or previous HSV infections, especially in persons presenting with lesions already healed or absent, where culture or PCR will not be useful
• It is estimated that 50-80% of the general population have antibodies to HSV-1 and 20-30% to HSV-2
• In primary HSV infection, significant IgG or IgM titer elevations do not occur for 10-14 days in mucocutaneous disease and for 3-4 weeks in herpes encephalitis
• Serial rising titers of IgM and IgG (four-fold increase) is suggestive of active HSV infection
• Various antibody testing methods are available to detect and distinguish between HSV type 1 and 2 infections such as:
• Western blot test, which is the traditional gold standard for researchers with an accuracy rate of 99%, but it is expensive, time consuming, and not widely available
• HerpeSelect, which is a combination of ELISA and immunoblot with a high sensitivity, but takes 1-2 weeks for results
• POCkit, which is a rapid, cost-effective test on finger prick blood sample with a good sensitivity, but detects only HSV-2 strain
• The limitations of antibody testing is that the positive result implies previous exposure to one or both of these viruses, but not whether the person is infectious, or if the particular genital syndrome is due to HSV.
• Repeat testing may be indicated in 10-14 days if negative or equivocal test results are seen
• Direct antigen testing
• This test is performed on the samples collected from vesicle fluid, CSF, or the material collected from eye
• It is a rapid, type specific, and sensitive method for detecting HSV by direct immunofluorescence using fluorescein labelled monoclonal antibodies specific for HSV antigens
• The diagnostic sensitivity is 80% in acute vesicular lesions and 60-75% in resolving lesions or asymptomatic shedding
• Factors interfering with the test results include:
• Hemolytic, lipemic, or icteric blood samples
• Contaminated or heat-inactivated specimens
• Administration of antiviral drugs before specimen collection
• Related laboratory tests include:
• Complete blood count
• CSF testing
• Sexually transmitted disease testing
• Chlamydia
• Human immunodeficiency virus (HIV)
• Human papilloma virus and genital warts
• Syphilis
• Trichomonas
• TORCH panel

Nl Result

• Tzanck test: Not detected
• Herpes culture: HSV-1 and HSV-2 not isolated
• Herpes IgM antibody:
• 0.89 IV (no significant level detected)
• 0.90-1.09 IV (Equivocal, repeat in 10-14 days)
• 1.10 IV (Current/Recent infection)
• Herpes IgG antibody
• 0.89 IV (no significant level detected)
• 0.90-1.09 IV (Equivocal, repeat in 10-14 days)
• 1.10 IV (Current or past infection)
• Direct antigen testing: Negative
• Herpes DNA testing: Negative

High Result

A positive result is typically indicative of infection with HSV.

Low Result

A negative result may indicate no infection with HSV or may indicate a false negative. Repeat testing may be indicated in select cases.

Conditions associated with false negative test results include:

• Inhibitors of PCR (heme) especially in neonates and young infants
• CSF samples collected in the early stage of the disease process, as HSV may involve only a focal area of the brain
• CSF sample collected late, after administration of antiviral therapy
• Antibodies will not be detectable in the early stages of infection
• Infections due to glycoprotein G-deficient virus (5-10%) gives negative antibody test results
• If transport of the sample is delayed, there may not be sufficient virus to detect even though the patient is infected

References

• ARUP's Laboratories®. Herpes Simplex Virus by PCR. [Homepage on the Internet]©2007. Last accessed on February 9, 2007. Available at URL: http://www.aruplab.com/guides/ug/tests/0060041.jsp
• ARUP's Laboratories®. Herpes Simplex Virus Culture with Reflex to HSV Typing. [Homepage on the Internet]©2007. Last accessed on February 9, 2007. Available at URL: http://www.aruplab.com/guides/ug/tests/0065065.jsp
• ARUP's Laboratories®. Herpes Simplex Virus Type 1 and/or 2 Antibodies, IgG & IgM with Reflex to Type 1 & 2 Glycoprotein G-Specific Ab, IgG. [Homepage on the Internet]©2007. Last accessed on February 9, 2007. Available at URL: http://www.aruplab.com/guides/ug/tests/0050916.jsp
• Beauman JG et al. Genital herpes: a review. Am Fam Physician. 2005 Oct 15;72(8):1527-34. Available at URL: http://www.aafp.org/afp/20051015/1527.html
• Bosch FX et al. CHAPTER 2 The epidemiology of human papillomavirus infection and its association with cervical cancer. Int J Gynaecol Obstet. 2006 Nov;94 Suppl 1:S8-S21.
• Drake AL et al. Herpes simplex virus type 2 and risk of intrapartum human immunodeficiency virus transmission. Obstet Gynecol. 2007 Feb;109(2 Pt 1):403-9.
• Elbers JM et al. A 12-year prospective study of childhood herpes simplex encephalitis: is there a broader spectrum of disease? Pediatrics. 2007 Feb;119(2):e399-407.
• eMedicine from WebMD®. Herpes Simplex. [Homepage on the Internet] ©1996-2006. Last updated on June 1, 2006. Last accessed on February 9, 2007. Available at URL: http://www.emedicine.com/EMERG/topic246.htm
• LabTestsOnline®. Herpes. [Homepage on the Internet]©2001-2007. Last reviewed on August 30, 2006. Last accessed on February 9, 2007. Available at URL: http://www.labtestsonline.org/understanding/analytes/herpes/sample.html
• Lingappa JR et al. Clinical and Therapeutic Issues for Herpes Simplex Virus-2 and HIV Co-Infection. Drugs. 2007;67(2):155-74.
• RUDNICK CM et al. Neonatal herpes simplex virus infections. Am Fam Physician. 2002 Mar 15;65(6):1138-42. Available at URL: http://www.aafp.org/afp/20020315/1138.html
• U.S. Preventive Services Task Force. Screening for genital herpes: recommendation statement- Independent Expert Panel. 2005 Mar 18. 11 pages. NGC:004067. Last updated on February 5, 2007. Last accessed on February 9, 2007. Available at URL: http://www.guidelines.gov/summary/summary.aspx?doc_id=6494&nbr=004067&string=hsv
• Whitley RJ. Herpes simplex virus infection. Semin Pediatr.Infect Dis. 2002 Jan;13(1):6-11.