Rotavirus RT-PCR and sequencing (reference centre activity)

Test description

RT-PCR and sequencing of VP7 and VP4 

Purpose of the test

• PCRs of VP7 and VP4: to amplify the VP7 gene and the VP4 gene of rotavirus.
• Sequence analysis of VP7 and VP4: to determine the G genotype and P genotype of infectious rotavirus, and to distinguish between the vaccine and wild type rotavirus strains.

Criteria for performing this test in the context of reference activities

The rotavirus G and P genotype can be determined by sequence analysis of rotavirus strains from samples previously diagnosed as rotavirus positive by the sender. It is mainly recommended to send samples from rotavirus positive children with gastroenteritis, despite complete inoculation with RotarixTM or RotaTeqTM , to determine the G and P genotypes.

Instructions for samples 

• Sample collection: faeces samples should be collected in properly labelled faeces containers. Preferably, 1 g (≈ 1000µl) or more should be sent. The minimum sample quantity is 0.1 g (≈ 100µl).
• Storage conditions: Although rotavirus can survive a long time in faeces at room temperature, it is advisable to store samples at 2-8°C, (or -20°C if possible) until shipment. The faeces sample can be sent by post or chilled to the NRC in a hermetically sealed container.  

Instructions for transport 

• Transport conditions: faecal samples can be sent by regular mail to the NRC or refrigerated in a hermetically sealed container. Avoid sending samples at the end of the week to avoid samples staying at room temperature for several days over the weekend.
• Specifications of transport medium: no specific transport medium is required. 

Unacceptable requests 

• Insufficient sample
• Application form not completed in full. This is absolutely essential. Every effort should be made to obtain minimum clinical data, including vaccination status of the patient. 

Turn around time (and frequency of analysis) 

RT-PCR and sequence analysis: a report is sent to all requesting clinicians once a year.

Reporting of test results 

Results will be sent via letter, email and/or fax according to the requesting laboratory or physician’s wishes.