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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.

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.

Background Epstein–Barr virus (EBV) is associated with several diseases, including infectious mononucleosis (IM) and malignant disorders, including post-transplant lymphoproliferative disorder (PTLD). The relationship between strains of the virus and disease manifestations or illness severity is of interest. Such strains have been defined by genetic variations in the major viral genes. Data involving the patterns of genetic diversity of the virus in different populations are required. We examined the genetic diversity of the BZLF1 gene, which is a major lytic gene of the virus. Methods We sequenced the BZLF1 gene of EBV following amplification from DNA that was extracted from blood obtained from pediatric bone marrow transplant (BMT) patients and children and young adults with IM. Sequencing was done by Sanger methodology (dideoxy DNA sequencing) and the sequences were aligned with a reference strain of EBV using Geneious software. The variant burden and types of single nucleotide variants (SNV) were compared across the 3 exons of the BZLF1 gene. Results We sequenced the BZLF1 gene using 21 patients with IM (median age 14, age range 2–19 years) and 11 who underwent bone marrow transplantation (median age 6, range 3–13 years). Three of 11 BMT patients developed post-transplant lymphoproliferative disorder (PTLD). Among the 3 exons, exon 1 had the greatest diversity across both study groups. There was a tendency for less diversity among PTLD samples, with no sample containing >1 single nucleotide variant (SNV) in contrast to the other samples. In samples that contained SNVs, there was a non-statistically significant trend for more SNVs to occur among the IM samples compared with PTLD samples (median 4.5 and 0, respectively; P > 0.05). Additionally, 2/11 (18.2%) BMT sequences contained more than 1 SNV compared with 7/21 (33.3%) IM sequences (P > 0.05). Conclusion There was a tendency for more genetic diversity among samples from patients with IM compared with bone marrow transplant patients, notably those with PTLD. Further studies will determine if this tendency is due to selective pressures in the transplant setting, including but not limited to the use of antiviral agents directed at the lytic phase of EBV. Disclosures All authors: No reported disclosures.

Infectious mononucleosis is caused by Epstein-Barr virus and can lead to complications, including hepatitis and hematological abnormalities, in a subset of patients. The authors of this article assessed measures of illness severity as well as viral load at presentation and six weeks later among a cohort of individuals, 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.

Background The Epstein–Barr virus (EBV) is associated with several diseases, including infectious mononucleosis as well as malignant disorders. The relationship between strains of the virus and disease manifestation or illness severity is of interest. Such strains have been defined by genetic variations in the major viral genes. As a first step toward a better understanding of the relationship between strains and clinical outcomes, data are required on the patterns of genetic diversity of the virus in different populations. In this study, we examined the genetic diversity of the BZLF-1 gene, which is a major lytic gene of the virus. Methods We sequenced the BZLF-1 gene of EBV following amplification from DNA that was extracted from blood and saliva from previously healthy Canadian children and young adults with infectious mononucleosis. Sequencing was done by Sanger methodology (dideoxy DNA sequencing) and the sequences were aligned with a reference strain of EBV using Geneious software. The variant burden and types of single nucleotide variants were compared in blood and saliva samples. Results Twenty-six samples were obtained from 24 patients less than 24 years of age (16 saliva and 10 blood samples). Two subjects provided paired blood and saliva samples at the same visit. Among 36 single nucleotide variations (SNVs), 22% were common to both blood and saliva samples. There was a nonstatistically significant trend for more SNVs among blood compared with saliva samples (median 6 and 1, ranges 0–8 and 0–9, respectively). Of the 3 exons of BZLF-1, exon 1 had the greatest frequency of SNVs compared with exons 2 and 3. Among the paired samples of blood and saliva, there were different genetic variants of the BZLF-1 gene in the blood compared with the saliva samples obtained from patients with infectious mononucleosis. Conclusion Among patients with infectious mononucleosis, different genetic variants of EBV may be present in blood compared with saliva. Blood samples revealed viral strains with a tendency for more genetic diversity compared with saliva. The potential compartmentalization of strains is of relevance in sample selection for the evaluation of the potential clinical impact of the genetic diversity of EBV. In addition, the potential impact on disease pathogenesis is of interest. Disclosures All authors: No reported disclosures.