Multiple-Strain Infections of Human Cytomegalovirus With High Genomic Diversity Are Common in Breast Milk From Human Immunodeficiency Virus–Infected Women in Zambia

Abstract

Background In developed countries, human cytomegalovirus (HCMV) is a major pathogen in congenitally infected and immunocompromised individuals, where multiple-strain infection appears linked to disease severity. The situation is less documented in developing countries. In Zambia, breast milk is a key route for transmitting HCMV and carries higher viral loads in human immunodeficiency virus (HIV)–infected women. We investigated HCMV strain diversity. Methods High-throughput sequence datasets were generated from 28 HCMV-positive breast milk samples donated by 22 mothers (15 HIV-infected and 7 HIV-negative) at 4–16 weeks postpartum, then analyzed by genome assembly and novel motif-based genotyping in 12 hypervariable HCMV genes. Results Among the 20 samples from 14 donors (13 HIV-infected and one HIV-negative) who yielded data meeting quality thresholds, 89 of the possible 109 genotypes were detected, and multiple-strain infections involving up to 5 strains per person were apparent in 9 HIV-infected women. Strain diversity was extensive among individuals but conserved compartmentally and longitudinally within them. Genotypic linkage was maintained within hypervariable UL73/UL74 and RL12/RL13/UL1 loci for virus entry and immunomodulation, but not between genes more distant from each other. Conclusions Breast milk from HIV-infected women contains multiple HCMV strains of high genotypic complexity and thus constitutes a major source for transmitting viral diversity.

Mixed cytomegalovirus genotypes in HIV positive mothers show compartmentalization and distinct patterns of transmission to infants

“…Fig. S2 shows that almost all the breast milk samples were highly diverse and therefore likely to contain multiple virus strains, a finding consistent with previous analyses of breast milk from HIV-infected women [24]. In contrast, the cervical and infant samples with the exception of one cervical sample from family 12, showed lower diversity.…”

“…We used next generation sequencing and haplotype reconstruction of individual CMV genomes, obtained from samples of HIV-infected women and their infants, to identify mixed infections, compartmentalization and distinct viral-genotype associations with transmission of CMV from mother-to-infant. Breast milk CMV showed high nucleotide diversity and, as has been previously reported [24], contained a mixture of viral genotypes, some of which were as genetically distant from each other as unrelated GenBank sequences and can therefore be considered distinct viral strains. Cervical samples were of low nucleotide diversity and dominated by a single viral genotype that was, with one exception, present in lower abundance in breast milk.…”

Polymorphisms in Human Cytomegalovirus Glycoprotein O (gO) Exert Epistatic Influences on Cell-Free and Cell-to-Cell Spread and Antibody Neutralization on gH Epitopes

“…These effects could not be explained by changes to the levels of gH/gL complexes in the virion envelope but rather point to changes in the mechanism(s) of gH/gL/gO in the processes of cell-free and cell-to-cell spread. The associated epistasis with the global genetic background highlights a particular challenge for intervention approaches since humans can be superinfected with several combinations of HCMV genotypes and recombination may occur frequently (1)(2)(3)(4)(5)(6)(7)(8). Moreover, these observations could help explain the incomplete protection observed for the natural antibody response against HCMV.…”

“…R ecent application of state-of-the-art genomics approaches have begun to uncover a greater and more complex genetic diversity of human cytomegalovirus (HCMV) than had been appreciated (1)(2)(3)(4)(5)(6)(7)(8). Of the 165 canonical open reading frames (ORFs) in the 235-kbp HCMV genome, 21 show particularly high nucleotide diversity and are distributed throughout the otherwise highly conserved genome.…”

Evolution and Genetic Diversity of Primate Cytomegaloviruses

“…These and other early efforts started to reveal that the genetic diversity of HCMV is high and that mutations frequently arise in cell culture [72]. More recently, the development of high-throughput technologies has allowed the sequencing of a sizable number of HCMV strains, either directly from clinical specimens or after a small number of passages [73][74][75][76][77][78]. To date, more than 300 complete HCMV genomes are available in public databases.…”