Epstein-Barr virus latent gene EBNA-1 genetic diversity among transplant patients compared with patients with infectious mononucleosis.

INTRODUCTION: As a step toward evaluating the association between Epstein-Barr virus genetic diversity and post-transplant lymphoproliferative disorder (PTLD), we conducted a preliminary study to compare the genetic diversity of the EBNA-1 gene among transplant patients and patients with infectious mononucleosis (IM). METHODS: We sequenced the EBNA-1 gene in blood samples from study subjects using Sanger methodology. The sequences were aligned with a reference strain and compared with publicly available sequences. RESULTS: We analyzed 33 study samples and 25 publicly available sequences along with the reference strain B95-8. The evaluable samples were from sixteen patients with IM (median age 14.0 years, range 2-24) and 17 transplant patients. There were six children without PTLD (median age 1.93 years, range 0.79-7.46) and 11 who developed PTLD (median age 5.67 years, range 0.96-17.45). A predominant EBNA-1 variant (P-thr) was identified across the study groups. Differences were observed between the samples from the IM patients compared with the transplant samples. CONCLUSION: The predominant EBNA-1 strain is in contrast to reports of the predominant strain in North America. The results suggest differences between the EBNA-1 strains among the study groups. Further studies will examine the relationship between EBNA-1 strains and PTLD occurrence and outcomes.

The genetic diversity of Epstein-Barr virus in the setting of transplantation relative to non-transplant settings: A feasibility study.

This study examines EBV strains from transplant patients and patients with IM by sequencing major EBV genes. We also used NGS to detect EBV DNA within total genomic DNA, and to evaluate its genetic variation. Sanger sequencing of major EBV genes was used to compare SNVs from samples taken from transplant patients vs. patients with IM. We sequenced EBV DNA from a healthy EBV-seropositive individual on a HiSeq 2000 instrument. Data were mapped to the EBV reference genomes (AG876 and B95-8). The number of EBNA2 SNVs was higher than for EBNA1 and the other genes sequenced within comparable reference coordinates. For EBNA2, there was a median of 15 SNV among transplant samples compared with 10 among IM samples (p = 0.036). EBNA1 showed little variation between samples. For NGS, we identified 640 and 892 variants at an unadjusted p value of 5 × 10(-8) for AG876 and B95-8 genomes, respectively. We used complementary sequence strategies to examine EBV genetic diversity and its application to transplantation. The results provide the framework for further characterization of EBV strains and related outcomes after organ transplantation.

Correlates of illness severity in infectious mononucleosis.

INTRODUCTION: Understanding the spectrum and frequencies of Epstein-Barr virus (EBV) complications and markers of illness severity in immunocompetent patients with primary EBV infection will inform management of patients with EBV-related illnesses. OBJECTIVES: To determine the clinical and laboratory correlates of illness severity among infants, children and youth with infectious mononucleosis (IM). METHODS: Study subjects with confirmed IM were prospectively enrolled. Illness severity was assessed at baseline and at six weeks using a scoring tool. Peripheral blood viral loads served as a measure of viral burden. RESULTS: Among 32 children and young adults with IM, the median age was 16 years (range two to 24 years). The predominant clinical findings were lymphadenopathy (23 of 32 [72%]), pharyngitis (16 of 32 [50%]), fever (nine of 32 [28%]) and splenomegaly (six of 32 [19%]). With respect to symptoms or signs that persisted to at least six weeks after illness onset, the predominant complaint was lymphadenopathy in 35% of subjects available for reassessment. Deranged liver function tests were present at presentation in up to 44% of subjects. Patients with the highest viral loads at presentation had significantly higher illness severity scores associated with fatigue (P=0.02). Other than the scores associated with fatigue, viral load values were not significantly correlated with the illness severity scores at baseline and at six weeks. CONCLUSION: In IM, viral loads are not necessarily correlated with illness severity, with the exception of fatigue. EBV-related hepatitis is common in IM, confirming the status of this virus as a relatively common cause of transient hepatitis in children and youth. This entity is not necessarily a marker of disease severity.

INTRODUCTION: Il faut comprendre le spectre et la fréquence des complications et des marqueurs de gravité du virus d'Epstein-Barr (VEB) chez les patients immunocompétents atteints d'une infection primaire par le VEB pour étayer la prise en charge des patients ayant une maladie liée à ce virus. OBJECTIFS: Déterminer les corrélats cliniques et de laboratoire de gravité de la maladie chez des nourrissons, des enfants et des adolescents atteints de mononucléose infectieuse (MI). MÉTHODOLOGIE: Les sujets atteints d'une MI confirmée ont été enrôlés prospectivement dans l'étude. Les chercheurs ont évalué la gravité de la maladie en début d'étude et au bout de six semaines au moyen d'un outil d'évaluation. Les charges virales du sang périphérique ont été utilisées pour mesurer le fardeau viral. RÉSULTATS: Les 32 enfants et jeunes adultes atteints de MI avaient un âge médian de 16 ans (plage de deux à 24 ans). Les principales observations cliniques étaient une lymphadénopathie (23 cas sur 32 [72 %]), une pharyngite (16 sur 32 [50 %]), de la fièvre (neuf sur 32 [28 %]) et une splénomégalie (six sur 32 [16 %]). Parmi les signes ou symptômes qui ont persisté au moins six semaines après l'apparition de la maladie, la lymphadénopathie était le principal problème chez 35 % des sujets disponibles pour être réévalués. Jusqu'à 44 % des sujets présentaient des anomalies dans les tests de fonction hépatique. Les patients dont la charge virale était la plus élevée à la présentation obtenaient des indices de gravité de la maladie associée à la fatigue considérablement plus marqués (P=0,02). À part les indices associés à la fatigue, les valeurs de la charge virale n'avaient pas de corrélation significative avec les indices de gravité de la maladie en début d'étude et au bout de six semaines. CONCLUSION: En cas de MI, les charges virales ne sont pas nécessairement corrélées avec la gravité de la maladie, à l'exception de la fatigue. L'hépatite liée au VEB est courante en présence de MI. Ainsi, ce virus est une cause relativement fréquente d'hépatite transitoire chez les enfants et les adolescents. Cette entité n'est pas nécessairement un marqueur de gravité de la maladie.

Gene expression using microarrays in transplant recipients at risk of EBV lymphoproliferation after organ transplantation: preliminary proof-of-concept.

We hypothesized that aspects of the virus-host interaction could be measured to help predict progression to EBV-PTLD. We examined the spectrum of host genes differentially expressed and any relevant clustering in children at risk of EBV lymphoproliferation after organ transplantation. We compared the genes expressed among patients with different levels of viral loads. Gene expression was measured by microarray analysis of RNA from CD19+ B lymphocytes using the Human Genome U133 Plus 2.0 GeneChip. Among 27 samples from 26 transplant recipients, the viral load categories were: low or undetectable loads (LU), n = 14; high or intermediate loads (HI), n = 13. There were seven healthy EBV-seropositive (P) and -seronegative controls (N). Median time of post-transplantation was 0.5 yr (range 0.1-3.8). We identified 24-54 differentially expressed genes in each of four comparisons of HI vs. P, LU vs. P, HI vs. LU, and P vs. N. We identified patterns of 563 gene expressions, creating five clusters aligned with levels of viral load. PTLD occurred in four of five clusters. In summary, we demonstrated varying degrees of alignment between levels of VL and gene clusters. Analyses for differential expression of genes showed genes that could be implicated in the pathogenesis of EBV-PTLD.