Direct genotyping of cytomegalovirus envelope glycoproteins from toddler's saliva samples.
BACKGROUND
The polymorphism of genes encoding CMV envelope protein is used for strain classification and may influence pathogenesis and/or infectivity. CMV genotyping is usually based on sequencing or acrylamide gel-RFLP, but these methods are not suited to rapid screening of large populations.


OBJECTIVE
We developed a high-throughput method to analyze CMV strains diversity and to detect multiple-strain infection in a large population of toddlers (six daycare centers (DCC) and an emergency unit (EU)).


METHODS
We developed a new PCR-RFLP method coupled with capillary electrophoresis fragment detection for UL55-gB, UL75-gH and UL73-gN genotyping. To detect gB recombinants, gpUL55 typing was applied to two variable zones (NTerminal and central). We applied this method to 212 CMV-positive saliva samples and controlled the results by direct sequencing of PCR products.


RESULTS
We identified 112 strains, that fell into eight groups in UL55-gB, two groups in UL75-gH, and seven groups in UL73-gN. The 79 samples from the emergency unit contained 30 strains, 28 children harboring 2 strains. The samples (n=133) from the six daycare centers contained respectively 4, 1, 6, 1 and 11 strains. Fifteen percent of strains were UL55-gB recombinants.


CONCLUSIONS
Our new method can simultaneously determine gB, gH and gN genotypes and offers more precise classification of CMV strains than previous RFLP-based methods. This could constitute the basis for a new classification, particularly in UL55-gB. Easy direct identification of multiple strains and recombinants in pathological samples could facilitate large epidemiologic studies.
Grosjean
J
J
Université de Limoges, INSERM, France.
Hantz
S
S
Cotin
S
S
Baclet
M C
MC
Mengelle
C
C
Trapes
L
L
Virey
B
B
Undreiner
F
F
Brosset
P
P
Pasquier
C
C
Denis
F
F
Alain
S
S
D002656:Child Day Care Centers; D003587:Cytomegalovirus / Q000145:classification* / Q000235:genetics / Q000302:isolation & purification; D003586:Cytomegalovirus Infections / Q000175:diagnosis / Q000821:virology*; D005838:Genotype; D006801:Humans; D007223:Infant; D016133:Polymerase Chain Reaction / Q000379:methods*; D012150:Polymorphism, Restriction Fragment Length; D020641:Polymorphism, Single Nucleotide; D012463:Saliva / Q000821:virology*; D014759:Viral Envelope Proteins / Q000235:genetics*; D014764:Viral Proteins / Q000235:genetics
C546870:UL55 protein, human cytomegalovirus; D014759:Viral Envelope Proteins; D014764:Viral Proteins