Diagnosis Of Epstein-Barr Virus Infection Research Articles

Epstein-Barr virus infection: from awareness of medical staff to the organization of epidemic control measures

The actuality of the infection caused by the Epstein — Barr virus (EBV) is due to its ubiquitous spread, the increase in morbidity, the chronic course of the infectious process, the high risk of the pathogen spread in organized groups of children and adults. A leading role in the timely identification and isolation of patients from medical organizations plays the awareness of medical workers on the issues of epidemiology, clinic and diagnosis of EBV infection. In order to assess the awareness of medical workers on these issues, a survey was conducted of 1,023 employees of various medical organizations in Moscow, including 514 doctors of therapeutic departments of hospitals, 342 otorhinolaryngologists, 167 dentists and assistants dentists. The conducted study revealed low awareness of medical personnel on all questions of the questionnaire, which creates a risk of nosocomial spread of the pathogen and requires an in-depth study of the epidemic process of EBV infection in the conditions of medical care for the subsequent development of a set of preventive and epidemic-control measures.

Evaluation of Clinical Application of Chemiluminescence and Real-time, Fluorescence-based Quantitative PCR in Diagnosis of Epstein-Barr Virus Infection

Objective: To compare the effects of clinical application of chemiluminescence and real-time, fluorescence-based quantitative PCR in the detection Epstein-Barr virus (EBV). Methods: The data of chemiluminescence and real-time fluorescent quantitative PCR from 97 patients who were suspected of being infected with EBV from January 2016 to January 2019 in our hospital were analyzed. The specific stage of EBV infection was analyzed, and the differences in results of the two detection methods were compared. Results: Chemiluminescence method was used to detect EBV infection during the active phase. The sensitivity of the chemiluminescence method was 76.7% (56/73) and the real-time quantitative PCR method was 90.4% (66/73). There was a statistical difference between the two detection methods (P<0.05). Conclusion: There was no statistical difference in positive predictive values between the chemiluminescence method and the real-time, fluorescence-based quantitative PCR method in the detection of EBV infection, but the sensitivity of chemiluminescence method is slightly lower than the real-time quantitative PCR method. It is noteworthy that chemiluminescence method is convenient and fast while the real-time, fluorescence-based quantitative PCR method is more accurate, which can provide a more accurate reference for clinical treatment.

Assessment of the epidemic situation in the diagnosis of Epstein-Barr virus infection in persons 15 years and older with a description of the clinical case

In order to assess the epidemic situation in the diagnosis of infection caused by the Epstein-Barr virus, a retrospective epidemiological analysis of the incidence of infectious mononucleosis for the period from 1990 to 2018 was conducted in the Moscow region using generally accepted methods of statistical data processing. The description of the clinical case was based on the results of dynamic observation of the patient reflected in the outpatient chart. It is shown that against the background of increasing morbidity rates, there is an epidemiological problem with infectious mononucleosis. Despite the fact that children 0–14 years old are traditionally at risk for infectious mononucleosis, the epidemic process involves the adult population, whose incidence rates also tend to increase. The growth rate of in dicators in the age groups 0–14 and 15 years and older is comparable. The active form of EBV infection in adults can occur in the form of reactivation, which is illustrated by the clinical case described in the article. This case of reactivation of EB infection in an adult clearly illustrates the need to assess the current epidemic situation in differential diagnosis and inform clinicians of different specialties on the epidemiology of EB infection.

Evaluation of two algorithms for diagnosis of Epstein-Barr virus infection

Evaluation of two algorithms for diagnosis of Epstein-Barr virus infection

Epistein-Barr Virus Infection with Concurrent Pancreatitis and Hepatitis: A Rare Disease Entity

Concurrent acute hepatitis and acute pancreatitis is a rare disease entity. Most commonly it can be attributed to infectious etiologies such as hepatitis A, Hepatitis B and Hepatitis E. Although not as common, Epstein Barr Virus (EBV) should be considered in the differential diagnosis in patients with combined hepatitis and pancreatitis. We report a case of a 25 year-old male admitted with 2-day history of abdominal pain, nausea and dark stools. Laboratory findings were significant for mixed hepatocellular and cholestatic liver injury. Abdominal imaging showed evidence of acute pancreatitis, without biliary obstruction. Serologic tests and radiologic imaging excluded common infectious, autoimmune and/or structural etiologies. The diagnosis of EBV infection was made by a positive IgM antibody to the viral capsid antigen and Epstein-Barr nuclear antigen. The patient had a prolonged hospitalization for persistent abdominal pain and lag in improvement in his cholestatic liver injury. This disease process has been described in 6 children and no cases in adults have been reported to our knowledge.

Epstein–Barr virus serology and PCR: conflicting results in an immunocompetent host. A case report and review of literature

We present the case of a 27-year-old immunocompetent man who progressively developed a generalized lymphadenopathy and B symptoms. Results of Epstein–Barr virus (EBV) serology were suggestive for a past infection, but the EBV viral load in whole blood was high. Also, core needle biopsy of the largest lymph node showed an image which could fit an EBV-driven reactive lymphoproliferation. Despite the absence of an immune disorder, all medical evidence points to an EBV-driven lymphoproliferative proces. In immunocompetent patients, it seems extremely uncommon to detect a high EBV viral load in the absence of serological evidence of an acute EBV infection or reactivation. We reviewed literature on this topic and on the selection of the appropriate sample type for EBV PCR, as this is known to be a critical point. Serological testing for the diagnosis of EBV infection is the gold standard in immunocompetent patients. Measuring EBV viral load is only recommended when dealing with immunocompromised patients. Although extremely rare, this case report shows that there is a place for EBV PCR in certain situations in immunocompetent patients. Besides, there is still no consensus regarding the specimen of choice for the determination of the EBV viral load. The preferred specimen type seems to depend on the patient’s underlying condition.

Diagnosis of Epstein-Barr Virus Infection with EBV IgM/IgG and Monospot Test Evaluation

Diagnosis of Epstein-Barr Virus Infection with EBV IgM/IgG and Monospot Test Evaluation

P669 Inflammatory bowel disease and Epstein-Barr virus: analysis of our experience

Background: Infection by the Epstein Barr virus (EBV) has been related to inflammatory bowel disease (IBD), but its implication in pathogenesis or exacerbation of the disease is uncertain. Its role in development of lymphoproliferative syndromes associated to immunosuppression is however well known, mainly in transplant patients and in IBD since introduction of thiopurines. Our objective was to report the association of immunosuppression and EBV as risk factors for the lymphoproliferative syndrome, including the lymphoplasmacytic infiltrate that may precede development of lymphoma. Methods: A retrospective study of patients with IBD in which histology showed a polytypic lymphoplasmacytic infiltrate (lambda and kappa chains) or lymphoma associated to EBV between December 2010 and October 2013. General patient and IBD characteristics, prior treatments, histological findings at the time of diagnosis of EBV infection, and subsequent course were recorded. Presence of EBV was tested using EBER probe hybridization with the respective messenger RNA. Results: 13 patients with IBD with a mean disease duration since diagnosis of 125.46±84.94 months were enrolled. 7 patients (53.8%) were male, and mean age was 40.46±9.98 years. As regards type of IBD, 7/13 (53.8%) had Crohn’s disease (CD), 5/13 (38.5%) ulcerative colitis (UC), and 1/13 (7.7%) non-classifiable IBD. In 11 patients, histology revealed polytypic lymphoplasmacytic infiltrate associated to EBV with no evidence of malignancy, and lymphoma associated to EBV was found in two patients. At diagnosis, patients were being treated with corticosteroids (7/13, 53.8%), azathioprine (5/13, 38.5%), and biologics (7/13, 53.8%). Another two patients had taken azathioprine before diagnosis. Mean time of treatment with azathioprine was 75.15±34.9 months. Mean duration of treatment with azathioprine was 75.14±34.85 months. All patients were in the exacerbation phase of IBD. None of the 11 patients developed lymphoma from the polytypic lymphoplasmacytic infiltrate associated to EBV after a mean follow-up (months) of 18±7.5 despite use of immunosuppressants and/or biologics in 8/13 (72.73%) of them. Conclusions: A lymphoplasmacytic infiltrate associated to EBV is a histological finding of uncertain significance made in a subgroup of patients with long-standing IBD, severe inflammatory activity, and chronic use of immunosuppressant and/or biologic drugs. Its implication in sequential development of lymphoproliferative syndromes and follow-up measures to be taken are not clear. Because of the increased incidence of lymphoma and its association to EBV since the introduction of thiopurines, we think that, based on these findings, longer follow-up with colonoscopy should be performed to assess changes in the lesion over time. P670 Efficacy of vaccination against hepatitis A and B in inflammatory bowel disease patients K. Karmiris1 *, E. Voudoukis1, M. Choustoulaki2, A. Sfiridaki2, G.A. Paspatis1. 1Venizeleio General Hospital, Gastroenterology, Heraklion, Crete, Greece, 2Venizeleio General Hospital, Blood Bank Center, Heraklion, Crete, Greece

Evaluation of 4 methods for the serological diagnosis of Epstein-Barr virus infection using an immunofluorescence assay as the reference method.

Tests specific for VCA IgM, VCA IgG, and EBNA IgG are used to diagnose Epstein-Barr virus (EBV) infections and interpret disease status. The immunofluorescence assay (IFA) is accepted as the "gold standard" test. The purpose of this study was to evaluate the performance of 4 methods in comparison with IFA. In total, 101 serum samples were obtained from clinically suspected cases of EBV infection between May 2010 and May 2012 and evaluated by IFA. All serum samples were analyzed by an immunoblot assay, enzyme-linked fluorescent assay (ELFA), enzyme immunoassay (EIA), and immunochromatographic assay (ICA). ELFA and ICA results were in good agreement with IFA for the detection ofVCA IgM, VCA IgG, and EBNA IgG. The results of the immunoblot assay agreed less well with IFA for EBNA IgG, while EIA results were not in agreement with IFA for EBNA IgG or VCA IgM. Among the tests studied, ELFA and ICA appear to be suitable methods for the diagnosis and staging of EBV when considering cost-effectiveness, turnaround times, need for a specialist, and IFA concordance.

Acute EBV infection masquerading as "In-situ Follicular Lymphoma": a pitfall in the differential diagnosis of this entity

We present the case of a 30 year-old man who was referred for evaluation of diffuse lymphadenopathy. Six weeks prior, he noticed darkening of his urine associated with pale stools, nausea and an eventual 30 lb weight loss within a month. The initial laboratory findings showed elevation of the liver enzymes. A CT scan showed mesenteric and periaortic lymphadenopathy with the largest lymph node measuring 2.8 cm. Other laboratory results were otherwise unremarkable (including a normal LDH) with the exception of positive serum antibodies against Epstein-Barr virus (EBV) associated antigens (IgM+ and IgG+). An excisional biopsy of 4 of the small neck lymph nodes showed a normal architecture with prominent follicles and an intact capsule. But, by immunohistochemistry two of the follicles showed aberrant coexpression of BCL-2, in addition to CD10 and BCL-6. In-situ hybridization for early Epstein-Barr virus mRNA (EBER) and immunohistochemistry for latent membrane protein-1 (LMP-1) stained both scattered positive cells, as well as BCL-2 positive B-cells. Although an original diagnosis of in-situ follicular lymphoma was favored at an outside facility, additional interphase fluorescence in situ hybridization (FISH) studies for t(14;18);(IGH-BCL2) rearrangement (performed on the BCL-2 + follicles microdissected from the tissue block; Abott probe dual colour fusion) and molecular studies (IGH gene rearrangement by PCR, also performed on the microdissected follicles) were negative. Serologic studies (positive EBV antibodies) and immunostains in conjunction with the molecular studies confirmed the reactive nature of the changes. Our case also shows direct immunopathogenic evidence of BCL-2 expression among the EBV-infected cells, which has to our knowledge not been previously documented in vivo. A diagnosis of EBV infection should, therefore, be considered when confronted with BCL-2 expression in germinal centers, particularly in younger individuals, as the diagnosis of FLIS may lead to extensive and invasive haematologic work-ups.Virtual slidesThe virtual slide(s) for this article can be found here: http://www.diagnosticpathology.diagnomx.eu/vs/1323656318940068

Serological diagnosis of Epstein-Barr virus infection: Problems and solutions

Serological tests for antibodies specific for Epstein-Barr virus (EBV) antigens are frequently used to define infection status and for the differential diagnosis of other pathogens responsible for mononucleosis syndrome. Using only three parameters [viral capsid antigen (VCA) IgG, VCA IgM and EBV nuclear antigen (EBNA)-1 IgG],it is normally possible to distinguish acute from past infection: the presence of VCA IgM and VCA IgG without EBNA-1 IgG indicates acute infection, whereas the presence of VCA IgG and EBNA-1 IgG without VCA IgM is typical of past infection. However, serological findings may sometimes be difficult to interpret as VCA IgG can be present without VCA IgM or EBNA-1 IgG in cases of acute or past infection, or all the three parameters may be detected simultaneously in the case of recent infection or during the course of reactivation. A profile of isolated EBNA-1 IgG may also create some doubts. In order to interpret these patterns correctly, it is necessary to determine IgG avidity, identify anti-EBV IgG and IgM antibodies by immunoblotting, and look for heterophile antibodies, anti-EA (D) antibodies or viral genome using molecular biology methods. These tests make it possible to define the status of the infection and solve any problems that may arise in routine laboratory practice.

The role of IgG avidity determination in diagnosis of Epstein-Barr virus infection in immunocompetent and immunocompromised patients

There is a high degree of variability in the serologic response to Epstein-Barr virus (EBV) infection, especially in viral capsid antigen (VCA)-IgM antibodies. Therefore, additional tests are needed to confirm primary infection. We evaluated the value of IgG avidity determination in diagnosis of EBV infection in immunocompetent and immunocompromised patients. A total of 236 serum samples from immunocompetent patients with symptoms suggestive of EBV infection were tested for the presence of VCA-IgM/IgG antibodies and IgG avidity. Using IgG avidity, acute primary infection was confirmed in 56.7% of the immunocompetent patients with positive and in 1.8% of patients with negative VCA-IgM. Recent primary infection was documented in 8.9% of the IgM positive and 3.5% of the IgM negative patients. In patients with indeterminate serology (equivocal IgM), 6.7% were classified by avidity index (AI) as acute primary infection, 10.0% as post-acute and 83.3% as past infection cases. Concerning the 32 immunocompromised patients, recent primary infection was documented in 3 of the 14 IgM positive patients. High AI was detected in 11 of these patients, indicating an IgM response due to reactivation. Determination of IgG avidity in combination with classical serologic markers seems to be a reliable method to confirm primary infection both in immunocompetent and immunocompromised patients. It may be especially useful to differentiate cases of primary infection in patients with undetectable VCA-IgM antibodies or indeterminate routine EBV serology.

PX-6 Comparative evaluation of indirect immunofluorescence assay and ELISA methods for the diagnosis of Epstein-Barr virus infection

A commercial line blot using recombinant antigens was compared with a commercial ELISA and ‘in-house’ IFA (reference test). Two panels were evaluated: Panel A was selected to distinguish between primary infections (89), past infections (20) and seronegatives (8) in immunocompetent individuals. In panel B, patients with a high number of reactivations were included: immunosuppressed patients (37), lymphoma (19), nasopharyngeal carcinoma (10), chronic fatigue syndrome (14). Blood donors (43) and cross-reactive sera (29) were added as controls. Line blot and IFA were concordant in 94% of primary infections, 100% of seronegatives and 100% of past infections, similar to ELISA. Results differed significantly with regard to reactivations. When compared with IFA, the incidence of reactivations was overestimated by the blot, 24 and 58% in blood donors and cross-reactive sera, respectively. ELISA showed a similar problems with 21 and 34% indeterminate results, respectively. The line blot is easy to carry out, has a good concordance with the reference IFA for primary infections, and is, therefore, a sufficient choice for distinguishing primary infection from seronegative and past infection. EBV reactivation assessment will require other methods such as EBV viral load.

Peptide mimics selected from immune sera using phage display technology can replace native antigens in the diagnosis of Epstein-Barr virus infection

There is an expanding area of small molecule discovery, especially in the area of peptide mimetics. Peptide sequences can be used to substitute for the entire native antigen for use in diagnostic assays. Our approach is to select peptides that mimic epitopes of the natural immune response to Epstein–Barr virus (EBV) that may be recognised by antibodies typically produced after infection with EBV. We screened a random peptide library on sera from rabbits immunised with a crude preparation of EBV and serum antibodies from a patient with a high titer of EBV antibodies. We selected four peptides (Eb1–4) with the highest relative binding affinity with immune rabbit sera and a single peptide with high affinity to human serum antibodies. The peptides were coupled to the carrier molecule BSA and the recognition of the peptides by IgM antibodies in clinical samples after infection with EBV was measured. The sensitivities were Eb1 94%, Eb2, 3, 4 88%, H1 81% and all had 100% specificity. This study illustrates that the phage display approach to select epitope mimics can be applied to polyclonal antibodies and peptides that represent several diagnostically important epitopes can be selected simultaneously. This panel of EBV peptides representing a wide coverage of immunodominant epitopes could replace crude antigen preparations currently used for capture in commercial diagnostic tests for EBV.

Comparative evaluation of ELISA and immunoblot assay for diagnosis of Epstein-Barr virus infection

Comparative evaluation of ELISA and immunoblot assay for diagnosis of Epstein-Barr virus infection

Peptide Mimotopes Selected from a Random Peptide Library for Diagnosis of Epstein-Barr Virus Infection

Epstein-Barr virus (EBV) is a ubiquitous, worldwide infectious agent that causes infectious mononucleosis, affecting >90% of the world's population. Currently, enzyme-linked immunosorbent assay, mostly with purified preparations of EBV cell extracts to capture immunoglobulin M (IgM) antibodies in patients' serum, is used for primary diagnosis. Our objective was to determine whether a small set of peptides could contain sufficient immunogenic information to replace solid-phase antigens in EBV diagnostics. Using monoclonal antibodies, we selected four peptides that mimic different epitopes of EBV from a phage-displayed random peptide library. To assess their diagnostic value, we screened a panel of 62 individual EBV IgM sera for their reactivities with the peptides alone. For all peptides, there was a clear distinction between the EBV-positive and the EBV-negative samples, resulting in 100% specificity. The sensitivities were 88%, 85%, 71%, and 54% for peptides F1, A3, gp125, and A2, respectively. Any combination of peptides increased the sensitivity, indicating that individual peptides react with different subsets of antibodies. Furthermore, when the F1 and the gp125 peptides were coupled to bovine serum albumin and screened against 216 serum samples, there were dramatic improvements in sensitivities (95% and 92%, respectively) and little cross-reactivity with the other peptides encountered during acute viral infections, including rheumatoid factor. This study shows the potential for the use of peptide mimotopes as alternatives to the complex antigens used in current serodiagnostics for EBV infection.

Gianotti-Crosti syndrome and allergic background.

The aim of the study was to verify whether there is a relationship between Gianotti-Crosti syndrome and an allergic background in children. Twenty-nine children affected by Gianotti-Crosti syndrome were first screened for a large panel of microbiological examinations, including serological and cultural tests for viruses and bacteria. A causative agent was identified in only 10 cases (34.4%). In five cases a diagnosis of Epstein-Barr virus infection was made on the basis of significant titres of anti-Epstein-Barr virus antibodies (IgM) associated with constitutional symptoms (fever, pharyngitis-tonsillitis). Our data concur with several clinical studies demonstrating that Epstein-Barr virus is now the most common viral agent associated with Gianotti-Crosti syndrome. For allergic evaluation, a group of 59 age- and sex-matched children investigated for recurrent infections were used as controls. The presence of atopic dermatitis (24.1%) in those with Gianotti-Crosti syndrome was significantly higher (p < 0.005) than in the control group (6.8%). In addition, a more common family history for atopy was 51.7% vs. 31% (p < 0.027) and the percentage of patients with total IgE greater than +2 SD for age higher than in controls (27.6% vs. 13.7%), as was the percentage of specific IgE present (31% vs. 17.2%). These results indicate that atopy is significantly associated with Gianotti-Crosti syndrome.

Epstein-Barr virus reactivation in kidney transplant recipients: relevance of serologic study

IT HAS been known for years that Epstein-Barr virus (EBV) and the development of posttransplant lymphoproliferative disorders (PTLD) are closely related. However, there are few data regarding the evolution of EBVspecific serologic markers as well as their actual clinical relevance in the transplant population. In the immunocompetent patient, serologic diagnosis of EBV infection relies on detecting antibodies against several EBV-specific antigens: viral capsid antigen (VCA), early antigen (EA), and nuclear antigens (EBNA), which appear at different times after infection. Nevertheless, in the immunosuppressed host interpretation of the former serologic antibodies is hampered by the inability of these patients to mount suitable serologic responses. The aims of our study were to (1) prospectively follow-up several EBV serologic markers in kidney transplant (KT) recipients, (2) determine the EBV reactivation rate in this group of patients, and (3) establish their clinical value and correlation with different KT variables.

Routine use of real-time quantitative PCR for laboratory diagnosis of Epstein-Barr virus infections.

Real-time quantitative polymerase chain reaction (PCR) on the LightCycler instrument (LC-PCR) was developed to measure the Epstein-Barr virus (EBV) load in clinical samples. LC-PCR detected two copies of the EBV genome per 500 ng of DNA. Its specificity was confirmed by assays in EBV-negative cell lines, other human herpesviruses and EBV-seronegative individuals. Excellent inter-assay reproducibility of LC-PCR was obtained in 43 samples (r = 0.983). LC-PCR results were compared with a routinely used ELISA-PCR of 150 samples and a good correlation was found (r = 0.956). A total of 88 individuals were studied, including healthy EBV-seropositive adults (n = 32), patients with EBV-associated disease (n = 34), and HIV-infected patients (n = 22); 37.5% of PBMC samples from healthy individuals contained EBV DNA, while no serum sample was positive. The viral load was significantly higher in PBMCS and saliva specimens in patients recently infected with HIV (19 and 39,400 copies/microg DNA, respectively), as well as in AIDS patients (122 and 331,130 copies/microg DNA) than in the control population (0 and 35 copies/microg DNA). This study confirmed that EBV load measurement with LC-PCR is helpful in the management of EBV-related post-transplantation lymphoproliferative disorders and probably of EBV-associated primary central nervous system B-cell lymphoma.

CHARACTERISTICS OF EPSTEIN-BARR VIRUS PRIMARY INFECTION IN PEDIATRIC LIVER TRANSPLANT RECIPIENTS

41 Background: Pediatric liver transplant recipients are at high risk of Epstein-Barr virus (EBV) infection following surgery. However the incidence of clinical symptoms and the graft function at the time of acute infection remains poorly documented. We report here the characteristics of primary EBV infection when occurring in pediatric liver transplant recipients. Patients and methods: Clinical and biological patterns associated with EBV infection were systematically searched in a cohort of 38 liver transplanted children (median age at OLT 20 months, range 5-158) between 1996 and 1998. Diagnosis of EBV infection was made by detection of EBV genome in peripheral blood lymphocytes using polymerase chain reaction (PCR) and/or by detection of anti-VCA IgM antibodies. Echography was used to detect abdominal lymphoid tissue. Results: Only 5 children (13%) had pretransplant immunity for EBV. Twenty-six (68.5%) developed primary EBV infection within a median of 30 days (range 15-90) following OLT. Ten had positive PCR with negative IgM. Sixteen patients had positive IgM among which PCR was positive whenever tested (n=5). Seven (18.5%) remained EBV negative on follow-up. Four (15%) had clinical symptoms at the time of infection, including necrotic tonsilitis (n=3) and impaired general condition (n=2). Two of them had concomitant PTLD liver nodules (n=2). A third PTLD occurred in one additional patient (gastric involvement) 300 days after primary asymptomatic infection. Overall incidence of PTLD was 10.5%. All three over-PTLD had full recovery following arrest of immunosuppression, one of them treated with anti-CD 20 antibodies (Mabthera®, Roche, Switzerland). No single patient had abnormal liver function tests (AST, ALT, GammaGT) related to EBV infection. Three of four PTLD had hypergammaglobulinemia, but only one of 34 non PTLD patient (cystic fibrosis). Conclusion: EBV primary infection occurs in 80% of seronegative patients within 3 months after OLT. PCR is more reliable than IgM for early diagnosis. Infection is not associated to changes of LFT's. A minority (15%) have clinical symptoms, being at high risk of full PTLD. Hypergammaglobulinemia is closely associated with PTLD.