New oligonucleotide primers for P-typing of rotavirus strains: Strategies for typing previously untypeable strains.
BACKGROUND
The use of molecular methods for rotavirus characterisation provides increased sensitivity for typing, and allows the identification of putative reassortant strains. However, due to the constant accumulation of point mutations through genetic drift; and to the emergence of novel genotypes; and possibly zoonotic transmission and subsequent reassortment, the reagents and methods used for genotyping require close monitoring and updating.


OBJECTIVE
To design and evaluate a new VP4 consensus oligonucleotide primer pair that provides increased sensitivity and allows typing of strains that were untypeable using available methods.


METHODS
A total of 489 rotavirus-positive faecal specimens from studies conducted between 1996 and 2006 were used for the evaluation of the new VP4 primers which was performed in the WHO Rotavirus Collaborating and Reference centres in the US, Australia, South Africa and the UK.


RESULTS
The new primer pair allowed P-typing of rotavirus strains and provided increased sensitivity, allowing typing of a significant number of strains that previously could not be P-typed.


CONCLUSIONS
This study highlights the importance of a constant reconsideration of primer sequences employed for the molecular typing of rotaviruses.
Simmonds
Mirjam Kühne
MK
Enteric Virus Unit, Virus Reference Department, Centre for Infections, Health Protection Agency, London NW9 5EQ, UK.
Armah
George
G
Asmah
Richard
R
Banerjee
Indrani
I
Damanka
Susan
S
Esona
Mathew
M
Gentsch
Jon R
JR
Gray
Jim J
JJ
Kirkwood
Carl
C
Page
Nicola
N
Iturriza-Gómara
Miren
M
D001315:Australia; D036022:Capsid Proteins / Q000235:genetics; D017931:DNA Primers / Q000235:genetics*; D005243:Feces / Q000821:virology; D006801:Humans; D012367:RNA, Viral / Q000235:genetics; D012401:Rotavirus / Q000145:classification* / Q000235:genetics* / Q000302:isolation & purification; D012400:Rotavirus Infections / Q000821:virology; D012680:Sensitivity and Specificity; D013019:South Africa; D006113:United Kingdom; D014481:United States
D036022:Capsid Proteins; D017931:DNA Primers; D012367:RNA, Viral; C060339:VP4 protein, Rotavirus