Human cytomegalovirus strain diversity and dynamics reveal the donor lung as a major contributor after transplantation

Mixed human cytomegalovirus (HCMV) strain infections are frequent in lung transplant recipients (LTRs). To date, the influence of the donor (D) and recipient (R) HCMV serostatus on intra-host HCMV strain composition and viral population dynamics after transplantation is only poorly understood. Here, we investigated ten pre-transplant lungs from HCMV-seropositive donors and 163 sequential HCMV-DNA-positive plasma and bronchoalveolar lavage samples from fifty LTRs with multiviremic episodes post-transplantation. The study cohort included D+R+ (38 per cent), D+R− (36 per cent), and D−R+ (26 per cent) patients. All samples were subjected to quantitative genotyping by short amplicon deep sequencing, and twenty-four of them were additionally PacBio long-read sequenced for genotype linkages. We find that D+R+ patients show a significantly elevated intra-host strain diversity compared to D+R− and D−R+ patients (P = 0.0089). Both D+ patient groups display significantly higher viral population dynamics than D− patients (P = 0.0061). Five out of ten pre-transplant donor lungs were HCMV DNA positive, whereof three multiple HCMV strains were detected, indicating that multi-strain transmission via lung transplantation is likely. Using long reads, we show that intra-host haplotypes can share distinctly linked genotypes, which limits overall intra-host diversity in mixed infections. Together, our findings demonstrate donor-derived strains as the main source of increased HCMV strain diversity and dynamics post-transplantation. These results foster strategies to mitigate the potential transmission of the donor strain reservoir to the allograft, such as ex vivo delivery of HCMV-selective immunotoxins prior to transplantation to reduce latent HCMV.