The iciHHV-6 Sense: Sensing the Source and Relevance of Human Herpesvirus 6 (HHV-6) DNA in the Transplant Recipient With Inherited Chromosomally Integrated HHV-6

The clinical implications and in vivo viral dynamics of inherited chromosomally integrated human herpesvirus 6 (iciHHV-6) have been an enigma since the condition was first recognized in approximately 1% of the human population in the 1990s [1]. In solid organ transplant (SOT) and hematopoietic cell transplant (HCT) recipients, most HHV-6 infections are due to reactivation of postnatally acquired infection with HHV-6B, with manifestations ranging from asymptomatic DNAemia to fatal end-organ disease [2, 3]. Donor cells are an additional source of HHV-6 in transplantation. Both hematopoietic cell and solid organ grafts may contain latent virus, which can either be episomal or, in the case of an iciHHV-6–positive donor, chromosomally integrated HHV-6 [4, 5]. Clinical quantitative PCR–based testing for HHV-6 DNA cannot readily distinguish transcriptionally silent iciHHV-6 from active, replicating virus and cannot determine whether replicating virus originates from donor or recipient. Dissecting the etiology and relevance of HHV-6 is most challenging after iciHHV-6–positive transplantation (ie, transplant involving an iciHHV-6–positive donor and/or recipient).

In this issue of The Journal of Infectious Diseases, Hannolainen and colleagues describe a unique case of HHV-6B infection of a transplanted liver originating from iciHHV-6B in the recipient [6]. A 2-year-old with biliary atresia underwent a deceased donor, split liver transplant. The transplant was complicated by an early leak at the biliary-enteric anastomosis requiring surgical reconstruction 4 weeks later. Total bilirubin and alanine aminotransferase rose despite the intervention, and repeated liver biopsies over the next 2 months showed areas of micronecrosis and cholangitis with no evidence of graft rejection. Despite a patent hepatic artery, the patient developed nonanastomotic biliary strictures, recurrent bacterial cholangitis, and persistent cholestasis, and ultimately required a second liver transplant. HHV-6 DNAemia >6.0 log₁₀ copies/mL was detected in the recipient's whole blood before the first transplant, and HHV-6 DNAemia persisted with <1 log₁₀ copy/mL fluctuations throughout the initial transplant course. iciHHV-6B was identified in the recipient and 1 parent. HHV-6B DNA, messenger RNA (mRNA), and viral proteins were also detected in several allograft tissue biopsies, raising important questions about HHV-6 transmission between donor and recipient tissues and the potential contributions of recipient-derived HHV-6 in graft failure.

The authors used sophisticated virologic techniques to identify HHV-6B transmission from the recipient's iciHHV-6B to donor cells within the allograft. HHV-6B DNA was detected in the donor liver before implant at low levels (3.1 log₁₀ copies/million cells), then rose and persisted in much higher quantities (>6.0 log₁₀ copies/million cells) in the graft after transplant. Hybrid capture sequencing with targeted enrichment of viral DNA was performed on tissue isolated from the donor liver before implantation and at multiple time points after transplantation. HHV-6B sequences recovered from the pretransplant donor liver were distinct from the recipient's iciHHV-6B sequence (consistent with previous community-acquired infection in the donor), whereas posttransplant sequences matched the recipient's iciHHV-6B sequence. To distinguish HHV-6B infection of the donor liver from graft infiltration with recipient iciHHV-6B–containing cells, the authors determined the relative abundance of donor and recipient mitochondrial DNA (mtDNA) from the same enriched hybrid capture sequencing data. The mtDNA sequences of the graft samples exclusively matched those of the donor, suggesting sequencing of entirely donor tissue, whereases a mixed donor/recipient genetic profile would be expected if the sample contained significant numbers of recipient cells. Viral mRNA transcripts were identified within hepatocytes and biliary cells by multiple methods including reverse-transcription PCR and in situ RNA hybridization. Immunohistochemistry and immunofluorescence detected HHV-6 protein expression in graft biopsy specimens. Taken together, these data support active HHV-6B infection of the liver allograft originating from the recipient's iciHHV-6B, though hybridization capture methods may still have enriched HHV-6 DNA of any origin, including from miniscule amounts of recipient-derived nucleated or cell-free iciHHV-6B–containing DNA [7].

Could HHV-6 infection have contributed to the posttransplant biliary complications? Many elements of the present case can be explained by factors other than HHV-6. Split liver transplant is a recognized risk factor for biliary complications, which occur in 30% of pediatric split liver transplant recipients compared to 17% of pediatric whole liver transplant recipients [8]. Biliary-enteric anastomotic leaks are the most common biliary complication of split liver transplant. Nonanastomotic strictures occur after approximately 5% of liver transplants, either as a complication of hepatic artery thrombosis (which was excluded in this case) or ischemia/immune-mediated injury [9]. Biliary complications likely led to persistent cholestasis. Micronecrosis was present on allograft biopsies throughout the posttransplant course, a nonspecific finding seen with viral hepatitis, reperfusion, or preservation injury [10].

HHV-6 DNAemia occurs in 40%–80% of liver transplant recipients within the first 3 months after transplant, but severe HHV-6 disease after liver transplantation is rare [2, 11]. Critical illness itself is a risk factor for HHV-6 DNAemia after liver transplant, confounding interpretation of HHV-6 DNAemia when identified during evaluation for unexplained organ dysfunction [11]. Potential “indirect” effects of HHV-6 have been postulated to influence transplant outcomes in the absence of HHV-6 end-organ disease. HHV-6 DNAemia is associated with acute graft-versus-host disease (aGVHD) and all-cause mortality after HCT and acute graft rejection after liver transplant [3, 11]. In the present case, upregulation of host genes associated with viral infections persisted throughout the clinical course. Host gene signatures have been used to implicate HHV-6B as a pulmonary pathogen after HCT but are challenging to interpret in a single patient without profiles from aviremic controls [12].

Does iciHHV-6 have clinical implications in transplantation? iciHHV-6 is a persistent source of HHV-6 DNA, and therefore donor or recipient iciHHV-6 is a plausible risk factor for HHV-6–associated conditions after transplant. In addition to the present case of HHV-6B reactivation from iciHHV-6B, reactivation of HHV-6A from iciHHV-6A has been described in case reports of immunocompromised patients with severe illnesses, including recipients of organs (kidney, liver) from a donor with iciHHV-6A and a child with iciHHV-6A, X-linked severe combined immunodeficiency, and hemophagocytic syndrome [5, 13, 14]. In HCT, iciHHV-6–positive transplant is associated with an increased risk of aGVHD but not HHV-6 encephalitis or other neurologic conditions [15, 16]. Systemic evaluation of outcomes after SOT involving iciHHV-6 in the donor and/or recipient is lacking.

Ascertaining the extent to which HHV-6 (or iciHHV-6) directly contributes to any clinical condition remains challenging. Perhaps it is more meaningful to consider whether identifying and treating HHV-6 improves outcomes. In the case presented by Hannolainen et al, antivirals (ganciclovir and later valganciclovir) were initiated within days to weeks after transplant, yet the patient's condition continued to deteriorate [6]. (Val)ganciclovir and foscarnet are frequently used to treat HHV-6 infections based on their in vitro antiviral activity, but data from prospective, randomized, placebo-controlled studies assessing relationships between these antivirals and HHV-6–associated clinical outcomes are limited [3, 17, 18]. HHV-6 encephalitis after HCT is associated with significant morbidity and mortality despite treatment; fewer than half of treated patients make a full neurologic recovery, but comparative outcomes absent of therapy are unknown [19]. Lack of clinical response to antivirals does not necessarily vindicate HHV-6 as a pathogen. Poor response may be due to inadequate antiviral concentrations or virally triggered inflammatory processes that propagate independent of ongoing viral replication, although autopsy studies have not shown evidence of virally triggered neuroinflammation in HHV-6 encephalitis [17]. Unfortunately, neither antiviral prophylaxis nor preemptive therapy has been shown to reduce the risk of HHV-6 encephalitis after HCT [3]. Preemptive approaches targeting HHV-6 and HHV-7 after liver transplantation failed to improve graft outcomes in a randomized clinical trial [18]. Myelosuppression and nephrotoxicity are common adverse effects of (val)ganciclovir and foscarnet, respectively, and judicious use of antivirals is particularly important in liver transplant recipients who are prone to bone marrow insults and kidney injury. Routine screening, surveillance, or preemptive therapy for HHV-6 after SOT is not recommended [20].

Changes in HHV-6 DNA loads following introduction of antivirals should be interpreted cautiously, especially after iciHHV-6–positive transplant. Antivirals do not affect HHV-6 DNAemia due to iciHHV-6 but may treat superimposed HHV-6 reactivation. In the absence of iciHHV-6, HHV-6 DNA load measurements depend on sample type (ie, plasma vs whole blood) and are subject to variations in sample processing and laboratory techniques. Immune reconstitution or reduction in immunosuppression intensity may facilitate spontaneous clearance of HHV-6 DNAemia in transplant recipients [3, 18]. In individuals with iciHHV-6 in hematopoietic cells, whole blood HHV-6 DNA load varies by total white blood cell count [3]. Tissue injury and cell turnover with release of human DNA from iciHHV-6–containing cells can increase HHV-6 DNA concentrations, complicating interpretation of viral loads after iciHHV-6–positive transplant [3, 16]. Routine screening for iciHHV-6 in transplant donors or recipients is unlikely to be cost-effective, but evaluation for donor or recipient iciHHV-6 in cases of unexplained, persistent, high-level HHV-6 DNAemia can help inform antiviral management and assuage clinician and patient concerns.

The case presented by Hannolainen et al is a remarkable scientific achievement in virology and transplantation. While the full clinical-pathologic spectrum of HHV-6 reactivation after iciHHV–6 positive liver transplant is still elusive, the presented methods are a giant step forward in sensing the source of HHV-6.

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Author notes