Viral Diagnosis: Mucocutaneous, Enteric, and Systemic

HSV NAAT is more sensitive than culture. If lesions are absent, type-specific glycoprotein G serology can support selected diagnoses, but it cannot identify the site or time of infection. Repeat serology 12 weeks after a suspected recent acquisition when the initial result is negative. Low-positive HSV-2 enzyme immunoassay results require confirmation with a second method because false reactivity is common. HSV IgM is not type specific and is not recommended.^{11 Centers for Disease Control and Prevention. Sexually Transmitted Infections Treatment Guidelines, 2021. 2021.}

PCR of lesion material is the preferred confirmation method for both primary varicella and herpes zoster. Saliva, nasopharyngeal secretions, urine, and blood are less sensitive than lesion material for an uncomplicated rash.^{22 Centers for Disease Control and Prevention. Laboratory testing for varicella-zoster virus. 2024.} For suspected mpox, notify the health department as required by the jurisdiction and arrange testing without using a sharp instrument on the lesion.^{33 Centers for Disease Control and Prevention. Collecting and handling specimens for monkeypox PCR testing. 2026.}

High-risk HPV assays detect viral DNA or E6/E7 mRNA and are used for cervical screening and the management of abnormal cervical results according to the assay's labeling. A validated assay may report HPV-16 and HPV-18 separately and pool other high-risk types. The result is interpretable only for an approved or validated specimen and clinical indication. Select an assay by its clinical performance and intended use. The viral gene targeted by an assay is not a stand-alone measure of oncogenic risk.^{11 Centers for Disease Control and Prevention. Sexually Transmitted Infections Treatment Guidelines, 2021. 2021.}

Hand-foot-mouth disease and herpangina are usually clinical diagnoses. Severe, neurologic, cardiac, or outbreak-associated disease may warrant enterovirus reverse-transcription polymerase chain reaction (RT-PCR) and typing. Vesicle material is useful for rash disease, cerebrospinal fluid (CSF) for meningitis, and respiratory or stool specimens for selected syndromes. Because fecal shedding is prolonged, a positive stool result does not by itself establish the cause of a neurologic illness.

Measles, rubella, and parvovirus B19

Suspected measles and rubella require prompt public-health notification because clinical findings alone are not specific enough for outbreak decisions.

For measles, collect serum for IgM and a nasopharyngeal or throat swab for rRT-PCR at first contact. Urine may be collected in addition to a respiratory specimen. PCR has its greatest yield from the first day of rash through the next 3 days and can remain positive for 10 to 14 days. IgM may be absent during the first 72 hours of rash. If an early IgM result is negative and PCR is negative or was not done, collect another serum specimen 3 to 10 days after symptom onset. Recent measles-mumps-rubella (MMR) vaccination can produce IgM and vaccine-virus RNA, so public-health genotyping may be needed.^{44 Centers for Disease Control and Prevention. Laboratory testing for measles. 2024.} The incubation period from exposure to prodrome averages 11 to 12 days. Rash appears about 14 days after exposure, with a range of 7 to 21 days.^{55 Gastanaduy P, et al. Measles. 2021.} Measles IgM provides presumptive evidence and can also be falsely reactive with parvovirus B19, rubella, roseola, rheumatoid factor, or nonspecific assay reactivity. An isolated positive IgM result without compatible epidemiology requires public-health interpretation and molecular or supplemental confirmation.^{66 Centers for Disease Control and Prevention. Measles serology testing.}

For suspected rubella, collect serum for IgM and a respiratory specimen for RT-PCR as early as possible. Rubella IgM may be negative on the first rash day and is most sensitive near day 5. It can also be reactive after recent vaccination or because of another rash illness, rheumatoid factor, or nonspecific assay reactivity. Never use IgM to screen an asymptomatic pregnant patient. Rubella IgG assesses immunity, and IgG avidity can help distinguish a recent primary response from remote infection or vaccination in selected cases.^{77 Centers for Disease Control and Prevention. Serology testing for rubella. 2024.}

IgG avidity is the overall strength of antibody binding and increases as the response matures. Collection timing therefore limits interpretation. High avidity in the third trimester cannot exclude infection during the first trimester. It also cannot identify the cause of an earlier rash or establish the infant's infection status. Communicate this timing limitation with the result, and review any earlier serology with the clinical and public-health services.^{77 Centers for Disease Control and Prevention. Serology testing for rubella. 2024.}

Parvovirus B19 serology can establish susceptibility, prior immunity, or a recent immune response in an immunocompetent patient. Maternal IgG and IgM do not diagnose fetal hydrops. B19 DNA testing is especially useful in an immunocompromised patient, persistent anemia or aplastic crisis, and selected fetal investigations because antibody may be absent.^{88 Centers for Disease Control and Prevention. Parvovirus B19.} Human herpesvirus 6 (HHV-6) serology has little value for routine roseola diagnosis. A high, persistent HHV-6 DNA concentration in whole blood can reflect inherited chromosomally integrated HHV-6 without active infection.

Multiplex panels are sensitive and fast, but they test only the targets on the panel. Codetections and low-prevalence detections require clinical interpretation, and a positive NAAT does not prove that replication is continuing. Virus culture is not part of the routine enteric workup. During a suspected norovirus outbreak, public-health laboratories use RT-qPCR and may genotype positive specimens to link cases or investigate a source. CDC recommends collecting whole stool during acute illness, ideally within 72 hours, from at least five ill people when specimens are available.^{1010 Centers for Disease Control and Prevention. Norovirus specimen collection and laboratory testing. 2025.} Vomiting can disperse virus-containing droplets onto food and surfaces. Norovirus is transmitted when contaminated material is swallowed. It is not a respiratory airborne infection.

In an immunocompromised patient with suspected cytomegalovirus (CMV) gastrointestinal disease, blood CMV DNA may support the evaluation, but it does not establish CMV colitis or enteritis. Tissue histology with CMV immunohistochemistry is the preferred evidence of tissue-invasive disease.

Order IgM anti-HAV only for patients with a compatible illness, because recent vaccination, cross-reactivity, and nonspecific reactivity can produce misleading results.^{1414 Centers for Disease Control and Prevention. Clinical screening and diagnosis for hepatitis A. 2024.} HBsAg can be transiently reactive after hepatitis B vaccination. HBV DNA can identify current infection when serology is incomplete or discordant.^{1515 Centers for Disease Control and Prevention. Clinical testing and diagnosis for hepatitis B. 2026.} A reactive anti-HCV result shows past or current exposure. Only detected HCV RNA establishes current infection.^{1616 Centers for Disease Control and Prevention. Clinical screening and diagnosis for hepatitis C. 2025.} HEV pregnancy-associated severity is best documented for genotypes 1 and 2 in endemic settings, and that finding does not extend to every genotype.^{1717 World Health Organization. Hepatitis E. 2026.}

HIV diagnosis and viral load

The standard laboratory sequence remains a venous-blood HIV-1/2 antigen/antibody immunoassay, followed by an HIV-1/HIV-2 antibody differentiation immunoassay when reactive, then a diagnostic nucleic acid test (NAT) when the differentiation result is negative or indeterminate. A differentiation result reactive for both types or reported as HIV positive untypable establishes HIV infection but does not prove dual infection. Type-specific HIV-1 and HIV-2 NAT can test for dual infection in that uncommon situation. A persistent HIV-2 indeterminate result with undetectable HIV-1 RNA requires supplemental HIV-2 testing or repeat testing of a new specimen through the algorithm.^{1818 Association of Public Health Laboratories. Suggested Reporting Language for the HIV Laboratory Diagnostic… 2025.}

CDC window-period ranges depend on the method and specimen: NAT usually detects infection 10 to 33 days after exposure, a laboratory antigen/antibody assay on venous blood 18 to 45 days, a rapid finger-stick antigen/antibody assay 18 to 90 days, and antibody-only tests 23 to 90 days. Oral-fluid rapid tests are antibody-only and are less sensitive for recent infection. Preexposure prophylaxis (PrEP) or postexposure prophylaxis (PEP) can suppress early replication and delay RNA, antigen, or antibody detection, so interpret the result with antiretroviral use and exposure timing.^{1919 Centers for Disease Control and Prevention. Getting tested for HIV. 2026.}

A nonreactive antigen/antibody screen does not exclude acute HIV. If recent exposure or an acute retroviral syndrome raises concern, order diagnostic HIV-1 RNA even when the screen is nonreactive. Patients taking or recently exposed to PrEP or PEP often need concurrent laboratory antigen/antibody and HIV-1 RNA testing. Repeat discordant or ambiguous results on a new blood specimen.^{2020 Centers for Disease Control and Prevention. Clinical guidance for PrEP.}

Quantitative HIV-1 RNA assays are primarily monitoring tests. Only assays with a diagnostic claim may be used for diagnosis, and their validated specimen and reporting mode must be followed. Some dual-claim assays accept plasma for quantitative monitoring and accept serum only for qualitative diagnostic testing. Reports distinguish three different results: "target not detected," "detected below the lower limit of quantification," and a quantified value. A change of about 0.5 log₁₀, or threefold, is generally the smallest statistically significant difference between serial measurements. This applies only to results from one platform that are not near an assay limit. National Institutes of Health (NIH) treatment guidelines define virologic failure as the inability to achieve or maintain HIV RNA below 200 copies/mL. A single detectable result followed by renewed suppression is a viral blip and is interpreted as part of the trend.^{2121 Panel on Antiretroviral Guidelines for… Plasma HIV-1 RNA (viral load) and CD4 count monitoring.}

Resistance genotyping during virologic failure is attempted when RNA remains above 200 copies/mL, although success is less reliable below 500 copies/mL. Baseline genotyping usually includes reverse transcriptase and protease. Integrase is added when transmitted integrase-inhibitor resistance is a concern or after integrase exposure through treatment, PEP, or long-acting cabotegravir PrEP. Proviral-DNA genotyping can help when plasma RNA is too low for sequencing, but it can miss archived mutations. Infants younger than 18 months with perinatal exposure require HIV RNA or DNA NAT on the age- and feeding-specific schedule because maternal antibody can persist. Every positive infant NAT is confirmed promptly with a virologic test on a separate specimen.^{2222 Panel on Treatment of HIV During Pregnancy… Diagnosis of HIV infection in infants and children.}

Yellow fever is most readily detected by NAAT during the first 3 to 4 illness days. IgM and neutralization testing become more useful after that early viremic phase, and the vaccination history is essential because available serology cannot reliably distinguish wild-type infection from a vaccine response.^{3232 Centers for Disease Control and Prevention. Clinical features and diagnosis of yellow fever.}

CDC's acute dengue testing pairs a direct test with IgM in the same illness phase because IgM can begin to appear while RNA and NS1 are declining.^{3333 Centers for Disease Control and Prevention. Clinical testing guidance for dengue. 2025.} Zika testing is narrower because low prevalence increases the proportion of false-reactive IgM results and RNA is detectable only briefly. Testing in pregnancy follows separate exposure-specific algorithms.^{3434 Centers for Disease Control and Prevention. Clinical testing and diagnosis for Zika virus disease. 2025.} Chikungunya IgM also requires confirmation by neutralization when a definitive diagnosis is needed.^{3535 Centers for Disease Control and Prevention. Chikungunya: diagnosis and testing. 2025.}

Interpret flavivirus serology with the illness day, geography, past dengue or other flavivirus infection, and dengue, yellow fever, or Japanese encephalitis vaccination history. Cross-reactivity affects dengue, Zika, West Nile, Japanese encephalitis, and yellow fever assays. PRNT can resolve some nonspecific IgM reactions after a primary exposure, but it may remain inconclusive after several flavivirus infections or vaccinations. A positive IgM result may therefore show presumptive recent infection without identifying the exact virus or date.

West Nile molecular testing has low sensitivity in an immunocompetent patient at neurologic presentation but can be useful after B-cell-depleting therapy or other marked immunosuppression.^{3636 Centers for Disease Control and Prevention. Clinical testing and diagnosis for West Nile virus disease. 2025.} Japanese encephalitis and Oropouche testing are arranged through the health department. Both may need paired or confirmatory serology when an early specimen is negative or cross-reactivity remains.^{37,3837 Centers for Disease Control and Prevention. Clinical features and diagnosis of Japanese encephalitis. 2025.38 Centers for Disease Control and Prevention. Clinical overview of Oropouche virus disease. 2025.}

Primer and probe mismatch can reduce molecular detection as viruses evolve. Follow manufacturer and public-health updates, review unexpected target dropouts or subtype failures, and retain suitable internal controls. Public-health genotyping and sequencing support surveillance and outbreak investigation. They do not replace a validated clinical result. Qualifying results are reported under federal, state, territorial, and local rules, and reference testing is arranged with the receiving public-health laboratory before shipment.

Detection below quantification

A target can be detected at a concentration too low for reliable measurement. Preserve that qualitative finding and the assay's reporting limit. Neither zero nor the lower quantification limit is a measured patient concentration. Quest's Roche cobas HCV guidance distinguishes RNA detected below 15 IU/mL from RNA not detected, and recommends a second specimen to confirm this low-level finding when testing is for initial diagnosis.^{4646 Quest Diagnostics. Hepatitis C virus antibody and RNA testing.}

The laboratory information system should pass the performing laboratory's detection wording, units, and interpretive comments through to the final report. If a numeric field cannot represent "detected below quantification," report a supported text result. Do not convert the result to zero. The referring laboratory never revises a referral result or the information that explains it.^{4747 Test report. 42 CFR §493.1291(a), (i).}

Control failures and result holds

A valid positive amplification control shows that amplification worked for that control. It does not compensate for a failed negative extraction control, which raises concern about contamination. Patient signal strength alone cannot resolve that failure. Hold affected results, investigate under the validated procedure, and obtain acceptable control results before release. Do not relabel patient positives as negatives merely because contamination is possible. The Clinical Laboratory Improvement Amendments (CLIA) require controls to meet the laboratory's and applicable manufacturer's acceptance criteria before patient results are reported.^{4848 Control procedures. 42 CFR §493.1256(d), (f).}

In the Microbiology Postanalytic Reporting lesson, see Automated result release for report holds and Corrected reports and postanalytic quality when a released result is later found invalid.