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. 2019 Dec 10;19(1):1046.
doi: 10.1186/s12879-019-4681-0.

Association of CMV genomic mutations with symptomatic infection and hearing loss in congenital CMV infection

G Clement Dobbins  1 , Amit Patki  2 , Dongquan Chen  3   4 , Hemant K Tiwari  2 , Curtis Hendrickson  5 , William J Britt  6   3 , Karen Fowler  6 , Jake Y Chen  3 , Suresh B Boppana  6   5 , Shannon A Ross  7   8
Affiliations

Association of CMV genomic mutations with symptomatic infection and hearing loss in congenital CMV infection

G Clement Dobbins et al. BMC Infect Dis. .

Erratum in

Abstract

Background: Congenital cytomegalovirus (cCMV) infection is the most common congenital infection and a leading cause of long-term neurological and sensory sequelae, the most common being sensorineural hearing loss (SNHL). Despite extensive research, clinical or laboratory markers to identify CMV infected children with increased risk for disease have not been identified. This study utilizes viral whole-genome next generation-sequencing (NGS) of specimens from congenitally infected infants to explore viral diversity and specific viral variants that may be associated with symptomatic infection and SNHL.

Methods: CMV DNA from urine specimens of 30 infants (17 asymptomatic, 13 symptomatic) was target enriched and next generation sequenced resulting in 93% coverage of the CMV genome allowing analysis of viral diversity.

Results: Variant frequency distribution was compared between children with symptomatic and asymptomatic cCMV and those with (n = 13) and without (n = 17) hearing loss. The CMV genes UL48A, UL88, US19 and US22 were found to have an increase in nucleotide diversity in symptomatic children; while UL57, UL20, UL104, US14, UL115, and UL35 had an increase in diversity in children with hearing loss. An analysis of single variant differences between symptomatic and asymptomatic children found UL55 to have the highest number, while the most variants associated with SNHL were in the RL11 gene family. In asymptomatic infants with SNHL, mutations were observed more frequently in UL33 and UL20.

Conclusion: CMV genomes from infected newborns can be mapped to 93% of the genome at a depth allowing accurate and reproducible analysis of polymorphisms for variant and gene discovery that may be linked to symptomatic and hearing loss outcomes.

Keywords: Cytomegalovirus (CMV); Next generation sequencing (NGS); Sensory neural hearing loss (SNHL); Viral diversity.

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Conflict of interest statement

The authors declare that they have no competing interests.

Figures

Fig. 1
Fig. 1
Phylogenetic analysis of CMV envelope glycoproteins gB and gN consensus sequences. a A maximum phylogenetic tree was constructed from consensus sequences generated for UL55 from each patient. b Similar analysis was carried out for UL73 (asymptomatic newborns with normal hearing in black; asymptomatic newborns with hearing loss in blue; symptomatic newborns with normal hearing in orange; and symptomatic newborns with hearing loss in red). Abbreviations: NB, new born; Sym, symptomatic; Asym, asymptomatic; NOHL, normal hearing; HL, hearing loss
Fig. 2
Fig. 2
Principal component analysis for patterns among clusters of variants and symptoms. For novel gene discovery, the SNP density was determined 2 k bps ahead of transcription start sites. The Principle Component Analysis (PCA) was used to show similarity based on variances of all values (SNP density) of all samples. Each principle component (PC1, 2, 3…) with PC1 and PC2 indicating the most variance were included to show similarity/dissimilarity of samples from highest to lowest. A Fisher exact test was used to correlate clinical outcomes and generate a significant gene list clustered with hearing loss (HL) (a) or symptomatic infection (b). c A heatmap displaying hierarchical clustering performed on samples (horizontal axes) and genes (vertical) with high level SNP densities associated with outcomes. The color indicates SNP density values above (red) average and below (green) average (no color) after z-normalization (average = 0 and SD = 1)
Fig. 3
Fig. 3
CMV variants in symptomatic and asymptomatic children. a Variant density of CMV genomes isolated from newborns (17 asymptomatic, 13 symptomatic) calculated in 1000 bp windows with Merlin as the reference strain. b Variants more frequent in symptomatic infection (Fisher’s exact test p < 0.05) are plotted with the genome position. The coding regions with the highest number of such variants are listed in their relative genomic position. c UL55 NSVs in relation to known antigenic domains (AD). Top panel shows the amino acid sequence of CMV strain Merlin. Bottom panel shows where NSVs (indicated by arrows below the reference strain) are more likely seen in infants with symptomatic congenital CMV in relation to antigenic domains
Fig. 4
Fig. 4
CMV variation from children with normal hearing and SNHL. Variant density of CMV genomes isolated from 17 children with normal hearing and 13 children with SNHL calculated in a 1000 bp sliding window with Merlin as the reference strain. Variants more frequent in viruses from children with SNHL (Fisher’s exact test p < 0.05) are plotted with the genome position. The coding regions with the highest number of variants are listed with the genomic position
Fig. 5
Fig. 5
CMV variants from asymptomatic children with normal hearing and SNHL. Variant density of CMV genomes isolated from 11 asymptomatic newborns with normal hearing and 6 asymptomatic newborns that developed SNHL calculated in a 1000 bp sliding window with Merlin as the reference strain. Regions with variants more frequent in viruses from children with asymptomatic infection and SNHL (Fisher’s exact test p < 0.05) are plotted with the genome position
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