Virus load dynamics of individual CMV‐genotypes in lung transplant recipients with mixed‐genotype infections

Abstract

Human cytomegalovirus (CMV) is a major cause of disease and transplant dysfunction in lung transplant recipients. Simultaneous emergence of more than one CMV-genotype can occur, and appears to be disadvantageous for the patient. In this study, the dynamics of individual CMV-genotypes in blood and lung was assessed within mixed CMV-genotype populations emerging after lung transplantation. In 69 plasma and 76 bronchoalveolar lavage samples of 16 lung transplant recipients with mixed CMV-genotype infections within the first year posttransplantation each of the major glycoprotein B (gB) and glycoprotein H (gH) genotypes was selectively quantified by genotype-specific quantitative TaqMan assays. The data obtained revealed that individually different genotype dynamics occurred for the individual patients and that the relative levels of the genotypes to each other may change over time. The quantitative development was independent of the specific gB-gH-genotype. In 10 of the 16 lung recipients the patient's individual genotype composition was the same in blood and lung. Genotype development during the follow-up was influenced by antiviral treatment. These data show for the first time that the CMV load used as diagnostic tool after transplantation is not always a constant entity but reflects the sum of the individual CMV-genotype dynamics developing over time.

Deep Sequencing Reveals Highly Complex Dynamics of Human Cytomegalovirus Genotypes in Transplant Patients over Time

“…For this test, HCMV DNA was isolated directly from patient samples using a QIAamp RNA isolation kit (Qiagen, Hilden, Germany). Samples with a viral DNA load of at least 5 ϫ 10 4 copies/ml were then tested using gB and gH genotype-specific quantitative PCR assays, which have been described previously (14), to preselect clinical samples with mixed HCMV infections.…”

“…Even cloning of PCR products and subsequent Sanger sequencing do not allow a sensitive assessment of the genotypes present unless an extremely large number of individual clones is sequenced. Improved genotype-specific real-time-PCR-based assays have been established recently for gB and gH genotyping, and these allow simultaneous detection and quantitation of distinct genotypes in mixed infections down to a level of 5% or even less (14,28).In the present study, we determined the relative amounts of different HCMV genotypes in clinical samples from lung transplant patients by using the highly sensitive ultradeep-pyrosequencing (UDPS) technology. UDPS genotyping of three of the most variable genes within the HCMV genome, the gN, gO, and UL139 genes, revealed that there is a substantially higher diversity of HCMV genotype populations in patients with mixed HCMV infection than has been recognized previously and that the HCMV strains show a distinct pattern in…”

“…Even cloning of PCR products and subsequent Sanger sequencing do not allow a sensitive assessment of the genotypes present unless an extremely large number of individual clones is sequenced. Improved genotype-specific real-time-PCR-based assays have been established recently for gB and gH genotyping, and these allow simultaneous detection and quantitation of distinct genotypes in mixed infections down to a level of 5% or even less (14,28).…”

Copy Numbers of Telomeric Repeat Sequences of Human Herpesvirus 6B in Clinical Isolates: Possibility of Mixed Infections

“…Active HHV-6B infection generally occurs only once throughout life (at the time of the primary infection) in immunocompetent individuals, but it may occur several times in transplant recipients (8) or DIHS patients (17). Frequent HHV-6B reactivation, as evidenced by the repeated isolation of the virus during an active viral infection, was observed in hematopoietic stem cell transplant (HSCT) recipients (8, 18).Previous reports detected mixed infections with multiple cytomegalovirus (CMV) strains in the same human herpesvirus subfamily (Betaherpesvirinae subfamily) as HHV-6B in a variety of patient populations, including immunocompetent and immunocompromised patients (19)(20)(21)(22). Furthermore, other studies have suggested that the genotype of the virus may be associated with the severity of the congenital cytomegalovirus infection (23).…”

“…Previous reports detected mixed infections with multiple cytomegalovirus (CMV) strains in the same human herpesvirus subfamily (Betaherpesvirinae subfamily) as HHV-6B in a variety of patient populations, including immunocompetent and immunocompromised patients (19)(20)(21)(22). Furthermore, other studies have suggested that the genotype of the virus may be associated with the severity of the congenital cytomegalovirus infection (23).…”