Direct genotyping of cytomegalovirus envelope glycoproteins from toddler's saliva samples

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• 作者：J. Grosjean ; S. Hantz ; S. Cotin ; M.C. Baclet ; C. Mengelle ; L. Trapes ; B. Virey ; F. Undreiner ; P. Brosset ; C. Pasquier ; F. Denis ; S. Alain
• 关键词：Genotype ; Cytomegalovirus ; Saliva ; PCR-RFLP ; Toddler ; Glycoproteins
• 刊名：Journal of Clinical Virology
• 出版年：2009
• 出版时间：December 2009
• 年：2009
• 卷：46
• 期：Supplement 4
• 页码：S43-S48
• 全文大小：628 K

文摘

lass=""h4"">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.

lass=""h4"">Objectives

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)).

lass=""h4"">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.

lass=""h4"">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.

lass=""h4"">Conclusion

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.