Skip to main page content
U.S. flag

An official website of the United States government

Dot gov

The .gov means it’s official.
Federal government websites often end in .gov or .mil. Before sharing sensitive information, make sure you’re on a federal government site.

Https

The site is secure.
The https:// ensures that you are connecting to the official website and that any information you provide is encrypted and transmitted securely.

Access keys NCBI Homepage MyNCBI Homepage Main Content Main Navigation
. 2022 Mar 4;14(1):33.
doi: 10.1186/s13148-022-01255-1.

Methyl-qPCR: a new method to investigate Epstein-Barr virus infection in post-transplant lymphoproliferative diseases

Chloé Borde #  1 , Frédérique Quignon #  2 , Corinne Amiel #  3 , Joël Gozlan #  4 , Vincent Marechal #  1 , Eolia Brissot #  5   6
Affiliations

Methyl-qPCR: a new method to investigate Epstein-Barr virus infection in post-transplant lymphoproliferative diseases

Chloé Borde et al. Clin Epigenetics. .

Abstract

Epstein-Barr virus DNA viral load is used as a surrogate marker to start Rituximab in transplant recipients at risk of developing PTLD. However, an elevated EBV DNAemia does not discriminate lymphoproliferation and replication. We designed a new molecular assay (methyl-qPCR) to distinguish methylated versus unmethylated viral genomes. In blood, viral genomes were highly methylated in EBV primary infections, PTLD and 4/5 transplant recipients with high viral load. The only patient with under-methylated EBV genomes did not respond to rituximab. Methyl-qPCR is a convenient method to discriminate between latent and lytic EBV genomes and could be useful in treatment decisions.

Keywords: DNA methylation; Epigenetics; Epstein–Barr virus; Post-transplant lymphoproliferative disease.

PubMed Disclaimer

Conflict of interest statement

The authors declare no conflict of interest.

Figures

Fig. 1
Fig. 1
Evaluation of latent versus lytic EBV DNA by methyl-qPCR. Methylation Index was measured on various EBV containing samples. EBV + Akata cell line during latency (A) and reactivation (B). EBV-positive saliva from healthy patients (n = 10) (C). Peripheral blood from patients with primary EBV infection (n = 9) (D); Peripheral blood from confirmed PTLD (post-transplant lymphoproliferative diseases) (n = 8) (E); Peripheral blood from HSCT recipients with no proof of PTLD (n = 5) (F). All samples were submitted to methyl-sensitive qPCR on four distinct regions of the viral genome. Each dot represents one individual sample. Whole extracted DNA were digested with either MspI or HpaII, two isoschizomers with different sensitivities to CpG methylation. HpaII is methylation sensitive, whereas MspI is methylation insensitive. For each region specified above, results are expressed in comparison with non-digested DNA. Data are expressed as mean ± SD
Fig. 2
Fig. 2
Representation of mean methylation index in Akata cells during latency and reactivation; saliva (n = 10) or blood of patients with primary infection (n = 9); confirmed PTLD (post-transplant lymphoproliferative diseases) (n = 8); or no proven PTLD (n = 5). Each dot represents one individual sample. Data are expressed as mean ± SD. *p < 0.05; ****p < 0.0001
See this image and copyright information in PMC

References

    1. Swerdlow S, Campo E, Harris N, et al. WHO classification of tumours of haematopoietic and lymphoid tissues. https://publications.iarc.fr/Book-And-Report-Series/Who-Classification-O.... Accessed 21 Dec 2020.
    1. Dierickx D, Habermann TM. Post-transplantation lymphoproliferative disorders in adults. N Engl J Med. 2018;378(6):549–562. doi: 10.1056/NEJMra1702693. - DOI - PubMed
    1. Landgren O, Gilbert ES, Rizzo JD, et al. Risk factors for lymphoproliferative disorders after allogeneic hematopoietic cell transplantation. Blood. 2009;113(20):4992–5001. doi: 10.1182/blood-2008-09-178046. - DOI - PMC - PubMed
    1. Hoshino Y, Kimura H, Tanaka N, et al. Prospective monitoring of the Epstein-Barr virus DNA by a real-time quantitative polymerase chain reaction after allogenic stem cell transplantation. Br J Haematol. 2001;115(1):105–111. doi: 10.1046/j.1365-2141.2001.03087.x. - DOI - PubMed
    1. Tomblyn M, Chiller T, Einsele H, et al. Guidelines for preventing infectious complications among hematopoietic cell transplantation recipients: a global perspective. Biol Blood Marrow Transplant. 2009;15(10):1143–1238. doi: 10.1016/j.bbmt.2009.06.019. - DOI - PMC - PubMed

Publication types

MeSH terms