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1 Department of Clinical and Experimental Medicine, Division Microbiology – St Orsola General Hospital, University of Bologna, Via Massarenti 9, 40138 Bologna, Italy
2 Center of Information Biology, National Institute of Genetics, Mishima, Japan
3 Department of Microbiology, SEALS, Prince of Wales Hospital, Randwick, NSW, Australia
4 Medical and Research Services, VA Medical Center and Division of Infectious Diseases, Oregon Health Sciences University, Portland, OR, USA
5 Department of Pediatrics and Microbiology, University of Alabama, Birmingham, AL, USA
6 Institute of Clinical and Molecular Virology, University of Erlangen-Nurnberg, Germany
Correspondence
Paola Dal Monte
dalmonte{at}med.unibo.it
Human cytomegalvirus (HCMV) ORF UL73 is a polymorphic locus, encoding the viral glycoprotein gpUL73-gN, a component of the gC-II envelope complex. The previously identified gN genomic variants, denoted gN-1, gN-2, gN-3 and gN-4, were further investigated in this work by analysing a large panel of HCMV clinical isolates collected from all over the world (223 samples). Sequencing and phylogenetic analysis confirmed the existence of the four gN genotypes, but also allowed the identification of a novel subgroup belonging to the gN-3 genotype, which was designated gN-3b. The number of non-synonymous (dN) and synonymous (dS) nucleotide substitutions and their ratio (dN/dS) were estimated among the gN genotypes to evaluate the possibility of positive selection. Results showed that the four variants evolved by neutral (random) selection, but that the gN-3 and gN-4 genotypes are maintained by positive selective pressure. The 223 HCMV clinical isolates were subdivided according to their geographical origin, and four main regions of gN prevalence were identified: Europe, China, Australia and Northern America. The gN variants were found to be widespread and represented within the regions analysed without any significant difference, and no new genotype was detected. Finally, for clinical and epidemiological purposes, a rapid and low-cost method for genetic grouping of the HCMV clinical isolates was developed based on the RFLP revealed by SacI, ScaI and SalI digestion of the PCR-amplified UL73 sequence. This technique enabled us to distinguish all four gN genomic variants and also their subtypes.
The UL73 sequences representative of the clinical strains tested in this present work have been assigned the following GenBank accession numbers: AF390748–AF390796, AF395118–AF395122, AF390835–AF390847, AF309969–AF309975, AF309980–AF309987, AF309989–AF310006, AF396722–AF396745, AF309976–AF309979, AF309988, AF396713–AF396721, AF390797–AF390834, AF390848–AF390857.
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