Large‐scale multiplex polymerase chain reaction assay for diagnosis of viral reactivations after allogeneic hematopoietic stem cell transplantation

Abstract

Viral reactivations following hematopoietic stem cell transplantation are thought to result from the breakdown of both cell‐mediated and humoral immunity. As a result, many viruses could be reactivated individually or simultaneously. Using a multiplex polymerase chain reaction (PCR), we prospectively examined many kinds of viral DNAs at a time in 105 patients who underwent allogeneic hematopoietic stem cell transplantation. In total, 591 whole blood samples were collected weekly from pre‐ to 42 days post‐transplantation and the following 13 viruses were tested; herpes simplex virus 1 (HSV‐1), HSV‐2, varicella‐zoster virus (VZV), Epstein–Barr virus (EBV), cytomegalovirus (CMV), human herpes virus 6 (HHV‐6), HHV‐7, HHV‐8, adenovirus, BK virus (BKV), JC virus (JCV), parvovirus B19, and hepatitis B virus (HBV). Several viral DNAs were detected in 12 patients before hematopoietic stem cell transplantation. The detection rate gradually increased after transplantation and peaked at 21 days. The most frequently detected virus was HHV‐6 (n = 63; 60.0%), followed by EBV (n = 11; 10.5%), CMV (n = 11; 10.5%), and HHV‐7 (n = 9; 8.6%). Adenovirus and HBV were each detected in one patient (1.0%). Detection of HHV‐6 DNA was significantly more common among patients undergoing cord blood transplantation or with steroid treatment. EBV DNA tended to be more common in patients treated with anti‐thymocyte globulin. Multiplex PCR was useful for detecting many viral reactivations after hematopoietic stem cell transplantation, simultaneously. Cord blood transplantation, steroid treatment, or anti‐thymocyte globulin use was confirmed to be risk factors after transplantation. J. Med. Virol. 87:1427–1435, 2015. © 2015 Wiley Periodicals, Inc.

HHV‐6 encephalitis may complicate the early phase after allogeneic hematopoietic stem cell transplantation: Detection by qualitative multiplex PCR and subsequent quantitative real‐time PCR

“…The plasmid DNAs were synthesized by Nihon Techno Service Company, Ibaragi, Japan. Using the serially diluted plasmid DNAs as standards, we confirmed that a minimum of 50 copies of each virus could be detected by this system [Inazawa et al, ].…”

“…Median age was 34 years (ranging from 6 months to 69 years) (Table ). Detailed patient characteristics were previously described [Inazawa et al, ].…”

“…The multiplex PCR assay was conducted as previously described [Ito et al, ; Inazawa et al, ]. The assay was designed to detect genomic DNA of the 13 viruses mentioned above.…”

Herpesviruses dUTPases: A New Family of Pathogen-Associated Molecular Pattern (PAMP) Proteins with Implications for Human Disease

“…Reactivation of multiple herpesviruses has been reported in transplant recipients [157,158,159], patients with sepsis [160] and drug-induced hypersensitivity syndrome (DISS), also known as drug rash, or those with eosinophilia and systemic symptomology (DRESS) [161,162,163,164,165,166]. The temporal order in which these viruses are reactivated and whether they interact to contribute to symptomology remains unknown.…”

Multiplex polymerase chain reaction for pathogen detection in donor/recipient corneal transplant tissue and donor storage solution

“…Primers and probes for HHV types 1–8 and the PCR conditions have been described previously 16, 17 , as have the primers and probes for the other viruses 18 and the primers and probes for Candida 18S and Aspergillus 18S 10, 19–21 . Amplification of the human β-globulin gene served as an internal positive extraction and amplification control.…”