In Part 1 of this 2-part presentation, Matt Binnicker, Ph.D., focused on cytomegalovirus infection. In Part 2, he presents a case study of an Epstein-Barr virus (EBV) infection in a transplant patient. EBV infection is associated with the development of posttransplant lymphoproliferative disorder, and accurate assessment of EBV viral load is essential to manage infected transplant recipients.
Presenters and Credentials:
Matt Binnicker, Ph.D., Director of the Clinical Virology Laboratory in the Division of Clinical Microbiology at Mayo Clinic in Rochester, Minnesota
Our speaker for this program is Dr. Matt Binnicker, Associate Professor of Laboratory Medicine and Pathology and Director of the Clinical Virology Laboratory in the Division of Clinical Microbiology at Mayo Clinic in Rochester, Minnesota.Welcome to Mayo Medical Laboratories Hot Topics. These presentations provide short discussions of current topics and may be helpful to you in your practice. Cytomegalovirus and Epstein-Barr virus are viral pathogens that may cause significant morbidity and mortality following transplantation. This 2-part presentation discusses the clinical presentations of infection, reviews the available testing for these viruses, and provides guidance on selecting the appropriate tests for your patients. Part 1 focused on CMV in the transplant population. In Part 2, the potential clinical impact of EBV in transplant patients will be discussed, and a case study will help illustrate how lab testing can be used to diagnose and monitor patients with EBV infection.
Dr. Binnicker, thank you for presenting today.
Thanks for the introduction, and thanks for joining for me for this update on viral load testing in the transplant population.
Before I begin, I should mention that I don’t have any corporate or financial conflicts of interest to disclose.
Also, I’d encourage you to consider several points in regards to test utilization as you view this presentation. First, how is the testing that we’ll discuss going to be used in your practice? Second, when should the tests be ordered and, finally, how will the results impact patient management?
In part 2 of this series, I’ll highlight the role of Epstein Barr virus, or EBV, in transplant recipients. Like cytomegalovirus or CMV, which we talked about in part 1 of this series, EBV is a member of the herpes virus family that most individuals are exposed to by early adulthood. Because of this, it’s an important cause of both acute and reactivated disease in immunosuppressed patients.
In this presentation, we’re going discuss the range of clinical presentations resulting from EBV infection in the transplant population, and review the clinical utility of quantitative molecular assays for the diagnosis and monitoring of this virus. And finally, I’ll highlight the limitations of current test methods and the impact these limitations may have on result interpretation. So let’s get started. To help illustrate the type of disease that EBV can cause in the transplant population and the role that laboratory testing can play in diagnosis and management.
I’d like to review a patient case. This is a case of a 74-year-old man with a history of primary sclerosing cholangitis who received a liver transplant. Unfortunately, this patient had significant postoperative complications including ischemic cholangitis requiring a second liver transplant just 3 weeks later. Two months after the second transplant, the patient presents to his care provider with elevated liver enzymes, and a subsequent liver biopsy shows acute cholangitis. He was treated with a course of antibiotics and initially showed signs of improvement.
Two months later, the patient presents for a routine follow-up evaluation and is once again found to have elevated liver enzymes. He doesn’t report any fever or chills, but does indicate that he has experienced some back pain recently. The patient’s current medications include acyclovir, cellcept, prednisone, and prograf. Due to his elevated liver enzymes and complaints of back pain, an MRI and PET scan are ordered, which reveal the following.
The MRI, showed numerous intrahepatic focal rim enhancing lesions, which were felt to most likely represent metastasis.
And the PET scan showed innumerable hypermetabolic focal hepatic lesions scattered throughout the liver parenchyma.
So these were obviously very concerning findings, and additional laboratory testing was performed. A CMV viral load was reported as negative or undetectable, but the patient’s EBV viral load was positive at 19.4 million copies/mL. Interestingly, a review of the patient’s record showed an EBV viral load of 1,100 copies just 3 months earlier. The EBV serostatus prior to the second transplant showed a seromismatch, with the donor being positive and the recipient being seronegative.
Liver Biopsy (40X magnification)
A liver biopsy was performed, and the results are shown on the next 2 slides. Here, you can see a significant area of blue-staining cells that are consistent with malignancy.
EBV ISH (20X magnification)
And, on the subsequent slide, we can see that an in situ hybridization for EBV shows a high degree of reactivity, especially in the area labeled as "tumor."
Case Study- Outcome
Based on these findings, the patient was diagnosed with EBV-associated PTLD and treated with weekly rituximab over the period of 1 month. His EBV viral load was monitored over this time, and showed a dramatic reduction while on therapy, dropping from 19.4 million copies/mL to negative just 1 month later.
Clinical Manifestations in the Transplant Population
This is a classic case of EBV-associated posttransplant lymphoproliferative disorder, or PTLD, which can be caused by primary infection with the virus, but more commonly, by reactivation of the virus following severe immunosuppression. PTLD is characterized by nonspecific constitutional symptoms, similar to infectious mononucleosis. Patients may present with fever, weight loss, and fatigue. Importantly, greater than 50% of PTLD patients present with extranodal masses of the GI tract, lung, liver, or the transplanted allograft.
EBV–PTLD Risk Factors
There are a number of factors that place an individual at greater risk for developing PTLD, and those are shown on this slide. For the solid organ transplant population, these risk factors include an EBV seromismatch, especially a donor positive, recipient negative categorization. In addition, the risk is higher in children than in adults, and certain types of solid organ transplants, such as small bowel, lung, or heart/lung transplants are at greater risk for PTLD. Finally, any patient with severe immunosuppression is at high risk. For hematopoietic stem cell transplant patients, those individuals with T-cell depletion, an HLA-mismatch, or with severe immunosuppression are at greatest risk for developing PTLD.
PTLD–Always due to EBV?
One question you might have is whether PTLD is always due to EBV infection, or whether other infections or factors can lead to its occurrence. Fortunately, the overall incidence of PTLD is relatively low, ranging from 1% to 10% in stem cell transplants, liver, renal, or heart/lung transplants. The incidence is higher, at 11% to 33% in intestinal or multiorgan transplants. Most of these cases occur during the first year posttransplant. Current data suggest that the vast majority, or somewhere between 70% and 90% of PTLD cases, are associated with EBV, which is either established by real-time PCR testing or histopathology. For those cases where EBV cannot be directly linked to PTLD, the etiology is still not well understood and work is underway to better define other potential causes. How about the association of the EBV viral load and the risk of PTLD? Can we make some predictions about an individual’s risk of developing the disease based on the amount of virus in the blood?
EBV Viral Load and Risk of PTLD
Well, a few studies have addressed this issue, and one in particular from 2012 performed a retrospective review of over 1,100 patients, which consisted of 700 solid organ transplant recipients and about 430 stem cell transplants. In this study, the EBV viral load was monitored on whole blood, and EBV DNAemia was defined as at least 1,000 copies/mL, while negative results were defined as those patients that never had a positive result, or had at least 3 negative tests. This study found that a higher proportion of solid organ transplant recipients had EBV viremia following the transplant compared to stem cell recipients.
Among their cohort of over 1,100 patients, 66 were diagnosed with PTLD. Interestingly, not all patients with PTLD had EBV detected in their blood. Furthermore, there were patients with a high EBV viral load that did not have PTLD and some patients with a relatively low viral load developed the disease. One conclusion that the authors were able to come to was that a higher peak EBV viral load was associated with a greater risk of progressing to PTLD.
PTLD–Screening and Diagnostic Recommendations
So what type of strategy should we employ for screening and diagnosing PTLD in transplant recipients? In the uncomplicated stem cell transplant recipient, the current strategy is to perform an initial viral load for EBV on the day of transplant, followed by weekly monitoring for the first 3 months. After this period, viral load testing can be performed monthly for the next 9 months. In the uncomplicated kidney transplant recipient, the recommendation is to get a viral load during the first week after the transplant, and then monitor monthly for the first 3 to 6 months postop. After this, the patient can be screened every 3 months until 1 year posttransplant. So although there remains quite a bit of uncertainty regarding how to best diagnose and monitor for PTLD, it’s relatively clear that quantitative real-time PCR testing plays an important role.
PTLD–Role of qPCR in Diagnosis
EBV viral loads are generally elevated in PTLD patients. Importantly, EBV quantitative PCR may have a higher negative predictive value than positive predictive value. In other words, a negative result by PCR provides pretty good assurance that a patient doesn’t have PTLD, while a positive result has overall poor specificity for diagnosing a patient with the disease. A true challenge that remains is the lack of test standardization in clinical laboratories performing EBV real-time PCR. This makes it difficult to compare results among laboratories and define a clinically significant threshold for disease. This lack of standardization means that in the future, clinical labs will need to consider implementing an international standard by which results can be compared among testing sites. And even more importantly, laboratories and providers should encourage their partners in industry to develop FDA-approved viral load assays for EBV.
In conclusion, the diagnosis of PTLD should be made through a combination of multiple laboratory and clinical findings, including radiology, EBV quantitative real-time PCR results, and, ultimately, tissue biopsy of the mass with review by a pathologist.
I’d like to thank you for taking the time to join me for this update on viral load testing in the transplant population.
Please feel free to contact Mayo Medical Laboratories if you have any questions about the testing that we discussed in this series.