Rotavirus and Norovirus Antigen

Rotavirus and norovirus are leading causes of acute viral gastroenteritis. Rotavirus group A (a reovirus) causes severe dehydrating diarrhoea mainly in children under 5 years, and norovirus (a calicivirus) causes epidemic vomiting and diarrhoea at all ages.

Diagnosis is by detection of viral antigen in stool by enzyme immunoassay or immunochromatographic rapid test (rotavirus; sensitivity 90 to 99% for rotavirus, specificity over 95%), or detection of viral RNA by RT-PCR, including multiplex gastrointestinal panels that detect rotavirus, norovirus GI and GII, adenovirus 40/41, astrovirus and sapovirus.

Antigen tests for norovirus have lower sensitivity (about 50 to 70%) and are used for outbreaks, whereas RT-PCR is the reference method for norovirus.

Rotavirus causes about 120 000 to 200 000 deaths each year in children under 5 years (mostly in sub-Saharan Africa and South Asia) and about 25 to 40% of hospital admissions for gastroenteritis before vaccine introduction. Rotavirus vaccines have reduced hospitalisation by 50 to 90% in countries with high coverage.

Norovirus causes about 685 million illnesses each year, around 200 million in children under 5 years, and is the commonest cause of gastroenteritis outbreaks in care homes, hospitals, cruise ships, schools and restaurants. Incubation is 1 to 3 days for rotavirus and 12 to 48 hours for norovirus, and illness lasts 3 to 8 days (rotavirus) and 1 to 3 days (norovirus).

Indications. Acute watery diarrhoea with vomiting in infants and young children requiring admission. Outbreaks of gastroenteritis in hospitals, nursing homes, schools and cruise ships (to identify the pathogen and control spread). Immunocompromised patients with persistent diarrhoea (chronic norovirus infection after transplantation). Hospital acquired diarrhoea.

Surveillance of vaccine impact. Differential diagnosis of bloody or prolonged diarrhoea (bacterial and parasitic causes). Not required for routine uncomplicated gastroenteritis in healthy adults.

Specimen. Fresh liquid stool 2 to 5 mL (or 1 to 2 g) in a clean, leak proof container, collected within 48 to 72 hours of onset when viral shedding is highest. Stored at 2 to 8 °C for up to 72 hours, and frozen for longer storage. Rectal swabs are acceptable for some PCR assays. Vomitus is rarely used.

Reference values. Rotavirus antigen: not detected. Norovirus antigen or RNA: not detected. Results are reported as positive or negative, and PCR panels may report cycle threshold values (low cycle threshold means high viral load).

Interpretation.

• Positive rotavirus antigen in a child with acute watery diarrhoea: confirms rotavirus gastroenteritis. A positive result in a recently vaccinated infant (within 1 to 2 weeks of the live oral vaccine) can reflect vaccine virus shedding, and interpretation needs history and genotyping.
• Positive norovirus PCR with compatible illness: confirms norovirus. Low level detection (high cycle threshold) in an asymptomatic or recovering person can reflect prolonged shedding (shedding continues for 1 to 3 weeks, and months in immunocompromised people).
• Negative antigen test: does not exclude infection, because antigen tests for norovirus are insensitive. PCR confirms.
• Multiplex panels detect co-infections, which are common (10 to 30%) and often of unclear significance. Pathogens at low load may not explain symptoms.
• In outbreaks, norovirus is likely when all four outbreak criteria are met (vomiting in over half of cases, mean incubation 24 to 48 hours, mean illness duration 12 to 60 hours, and no bacterial pathogen found), and testing of at least 2 to 5 stool samples confirms the cause.
• Genotyping of norovirus (GII.4 variants) and rotavirus (G and P types) is used for surveillance and vaccine monitoring.
• Nosocomial rotavirus and norovirus are common, and a positive test triggers infection control.

Critical values and findings needing immediate action.

• Infant or child with signs of severe dehydration (sunken eyes, lethargy, poor skin turgor, absent urine, shock): immediate rehydration (oral rehydration solution, or intravenous isotonic fluid for shock), zinc supplementation, and monitoring of electrolytes. Antigen testing never delays rehydration.
• Hospital or care home outbreak: contact isolation (soap and water handwashing, as alcohol gel is less effective against norovirus), cohorting, closure of wards, exclusion of staff for 48 hours after symptoms resolve, and notification of public health.
• Immunocompromised patient with persistent diarrhoea and positive norovirus or rotavirus: review of immunosuppression, nitazoxanide in selected cases, and nutritional support.
• Neonates with rotavirus: risk of necrotising enterocolitis, close monitoring.
• Intussusception in an infant after rotavirus vaccination (rare, 1 to 6 in 100 000): abdominal pain, vomiting, red currant stool, urgent imaging.

Interferences and pitfalls.

• Antigen tests miss many norovirus infections, and a negative test does not exclude norovirus. PCR on stool from several patients is used in outbreaks.
• Rotavirus antigen can be detected for several days after live vaccine administration, giving positive results in vaccinated infants.
• Asymptomatic carriage and prolonged shedding make a positive test hard to interpret in older children and adults with another cause of diarrhoea.
• Multiplex panels can overdetect low level pathogens, and results should be correlated with clinical findings.
• Stool samples contaminated with urine, barium, mineral oil or some preservatives can interfere with antigen tests.
• Inadequate collection time (late sample, after shedding falls) lowers sensitivity.
• Viral gastroenteritis does not need antibiotics, and testing is for infection control and diagnosis, not for guiding antimicrobials.
• Bacterial (Salmonella, Shigella, Campylobacter, Escherichia coli), parasitic (Giardia, Cryptosporidium, Entamoeba) and toxin (Clostridioides difficile) causes need testing if the illness is prolonged, bloody, severe or follows antibiotics or travel.
• Rotavirus vaccination is a primary prevention measure, and efficacy is lower in low income settings (about 50 to 60%) than in high income ones (85 to 98%).

Special groups.

• Infants and children: highest burden, with dehydration and hospital admission. Zinc supplementation reduces duration and severity.
• Pregnancy: norovirus and rotavirus cause mild disease, with rehydration the main treatment.
• Older adults and care home residents: norovirus causes severe dehydration, acute kidney injury and death, with outbreaks in institutions.
• Immunocompromised patients: chronic infection with prolonged shedding, malnutrition and graft dysfunction.
• Regions: rotavirus remains a leading killer of children in sub-Saharan Africa and South Asia, where vaccine coverage, access to oral rehydration and zinc, and nutrition are the main determinants of mortality.

Follow up tests. Multiplex gastrointestinal PCR panel, stool culture for bacterial pathogens, Clostridioides difficile testing, stool microscopy for parasites, Giardia and Cryptosporidium antigen, serum electrolytes, urea and creatinine, glucose, blood gas in severe dehydration, full blood count, stool calprotectin when chronic diarrhoea persists, abdominal ultrasound for intussusception, genotyping of rotavirus and norovirus in surveillance, and review of vaccination status.