To analyze the association of Epstein-Barr virus (EBV) with gastrointestinal non-Hodgkin's lymphomas arising in immunocompetent patients, 56 consecutive cases of gastrointestinal lymphomas (B-cell: 52-cases, T-cell: 3 cases, T/NK-cell: 1 case) occurring in the stomach (33 cases), intestine (22 cases) and esophagus (1 case) were investigated for the presence of EBV using polymerase chain reaction analysis as a screening method followed by EBER-1 RNA and DNA in situ hybridization (ISH) and immunohistochemistry for the expression of latent membrane protein 1 (LMP-1). Forty-seven cases demonstrated extractable DNA and EBV DNA was detected only in 4 cases. Among the, RNA (EBER-1) and DNA ISH analysis confirmed the presence of the EBV genome in tumor cells in 3 cases (T/NK-cell lymphoma of ileum, gastric high-grade B-cell lymphoma of mucosa-associated lymphoid tissue, gastric diffuse large B-cell lymphoma). Only the T/NK cell lymphoma showed diffuse positivity of tumor cells while 2 gastric B-cell lymphomas demonstrated a scattered positive reaction and no cases expressed LMP-1. Nine cases without extractable DNA by the PCR method showed no nuclear signal by EBER-1 ISH. These findings suggest that most sporadic primary gastrointestinal lymphomas in Korea are not associated with EBV.
Gastrointestinal Neoplasms/virology* ; Gastrointestinal Neoplasms/pathology ; Genome, Viral ; Herpesvirus 4, Human/isolation & purification* ; Herpesvirus 4, Human/genetics ; Human ; In Situ Hybridization ; Korea ; Lymphoma/virology* ; Lymphoma/pathology ; Polymerase Chain Reaction

The development of a multiplex polymerase chain reaction (PCR) method for rapid and accurate detection and typing of herpes simplex virus type 1 (HSV-1), and type-2 (HSV-2), cytomegalovirus (CMV) and Epstein-Barr virus (EBV) is very important for clinical diagnosis to allow the deliver of therapy as early as possible. Large scale amplifications by multiplex PCR of viral DNA can lower the cost and time for viral diagnosis. In this study, therefore sensitive quadruplex PCR was achieved by optimizing parameters such as primers, and 1.5 mM magnesium and 200 uM dNTPs concentrations. The concentrations of HSV-1, HSV-2, CMV and EBV primers were 0.5, 0.3, 0.25 and 0.25 pmoles, respectively. Optimal annealing temperature was 54 degrees C. Employing these conditions, we could detect 10 copies of reconstructed template plasmid DNA, which were cloned to vectors containing target sequences of viral DNA. PCR products of 271 bp for HSV-1, 231 bp for HSV-2, 368 bp for CMV, and 326 bp for EBV were separated on 5.0% polyacrylamide gel electrophoresis and confirmed by direct sequencing. The present study showed that the quadruplex PCR assay described herein has potential application in clinical diagnosis, when rapid, accurate detection and typing of viruses HSV-1, HSV-2, CMV or EBV are necessary.
Cytomegalovirus/classification/*isolation & purification ; Herpesvirus 1, Human/classification/*isolation & purification ; Herpesvirus 2, Human/classification/*isolation & purification ; Herpesvirus 4, Human/classification/*isolation & purification ; Human ; *Polymerase Chain Reaction/methods ; Sensitivity and Specificity ; Support, Non-U.S. Gov't

BACKGROUNDThe dilemma of pathogens identification in patients with unidentified clinical symptoms such as fever of unknown origin exists, which not only poses a challenge to both the diagnostic and therapeutic process by itself, but also to expert physicians.

METHODSIn this report, we have attempted to increase the awareness of unidentified pathogens by developing a method to investigate hitherto unidentified infectious pathogens based on unbiased high-throughput sequencing.

RESULTSOur observations show that this method supplements current diagnostic technology that predominantly relies on information derived five cases from the intensive care unit. This methodological approach detects viruses and corrects the incidence of false positive detection rates of pathogens in a much shorter period. Through our method is followed by polymerase chain reaction validation, we could identify infection with Epstein-Barr virus, and in another case, we could identify infection with Streptococcus viridians based on the culture, which was false positive.

CONCLUSIONSThis technology is a promising approach to revolutionize rapid diagnosis of infectious pathogens and to guide therapy that might result in the improvement of personalized medicine.

Female ; Herpesvirus 4, Human ; genetics ; isolation & purification ; High-Throughput Nucleotide Sequencing ; methods ; Humans ; Male ; Viridans Streptococci ; genetics ; isolation & purification

Herpesvirus 4, Human ; genetics ; isolation & purification ; Humans ; Infectious Mononucleosis ; drug therapy ; pathology ; virology ; RNA, Viral ; isolation & purification ; Sensitivity and Specificity

OBJECTIVETo compare the detection rates of Epstein-Barr virus (EBV) DNA in the serum/plasma between apparently healthy adults (AHAs) and nasopharyngeal carcinoma (NPC) patients in attempt to evaluate the efficiency of EBV DNA assay for serodiagnosis of NPC.

METHODSThe plasma and serum were obtained from 58 AHAs and 66 untreated NPC patients. EBV DNA W-fragment was detected using nested ploymerase chain reaction (PCR). Immunoenzymatic assay for titration of IgA-VCA was also adopted.

RESULTSEBV DNA detection rate (84.85%) in the plasma/serum of 66 NPC patients was significantly higher than that (10.34%) in 58 AHAs. The sensitivity of plasma/serum EBV DNA assay (0.8485) was higher than that (0.8030) of titrating IgA-VCA (positive criterion >/= 1:40) though the specificities of these two tests were the same (0.8966). The correct rate, predictive value of a positive test, and Odds ratio of dual positivity (0.8387, 0.9792 and 141.0, respectively) were higher than those of single positivity either to plasma/serum EBV assay (0.5242, 0.7333 and 1.1423, respectively) or to IgA-VCA >/= 1:40 test (0.4839, 0.5385 and 1.0480, respectively).

CONCLUSIONThe EBV DNA detection in the plasma/serum using nested PCR may be a useful indicator for serodiagnosis of NPC.

Antigens, Viral ; blood ; DNA, Viral ; blood ; Herpesvirus 4, Human ; isolation & purification ; Humans ; Immunoglobulin A ; blood ; Nasopharyngeal Neoplasms ; diagnosis ; virology ; Serologic Tests

Child ; Epstein-Barr Virus Infections ; complications ; Female ; Giardia lamblia ; isolation & purification ; Giardiasis ; complications ; Herpesvirus 4, Human ; isolation & purification ; Humans ; Lymphohistiocytosis, Hemophagocytic ; drug therapy ; etiology

The precise etiology of hemolytic uremic syndrome (HUS) is unknown. However, it has been associated with bacterial (Shigella, Salmonella, E. coli, S. pneumoniae), Bartonella, and viral (coxsackie, ECHO, influenza, varicella. Epstein-Barr) infections and with endotoxemia. Recently, we experienced a case of HUS in a 16-year-old boy who was in the acute phase of an Epstein-Barr virus (EBV) infection. He had typical manifestations of HUS and EBV infection. He also transiently presented disseminated intravascular coagulation. His renal dysfunction recovered by supportive care, including hemodialysis, plasmapheresis, antihypertensive medication and aspirin. We present this case with a review of the literature as the second report of HUS associated with EBV infection.
Adolescence ; Follow-Up Studies ; Hemolytic-Uremic Syndrome/virology* ; Hemolytic-Uremic Syndrome/therapy ; Herpesviridae Infections/diagnosis* ; Herpesvirus 4, Human/isolation & purification* ; Human ; Male ; Renal Dialysis ; Tumor Virus Infections/diagnosis*

There has been increasing interest in standardized and quantitative Epstein-Barr virus (EBV) DNA testing for the management of EBV disease. We evaluated the performance of the Real-Q EBV Quantification Kit (BioSewoom, Korea) in whole blood (WB). Nucleic acid extraction and real-time PCR were performed by using the MagNA Pure 96 (Roche Diagnostics, Germany) and 7500 Fast real-time PCR system (Applied Biosystems, USA), respectively. Assay sensitivity, linearity, and conversion factor were determined by using the World Health Organization international standard diluted in EBV-negative WB. We used 81 WB clinical specimens to compare performance of the Real-Q EBV Quantification Kit and artus EBV RG PCR Kit (Qiagen, Germany). The limit of detection (LOD) and limit of quantification (LOQ) for the Real-Q kit were 453 and 750 IU/mL, respectively. The conversion factor from EBV genomic copies to IU was 0.62. The linear range of the assay was from 750 to 10⁶ IU/mL. Viral load values measured with the Real-Q assay were on average 0.54 log₁₀ copies/mL higher than those measured with the artus assay. The Real-Q assay offered good analytical performance for EBV DNA quantification in WB.
DNA, Viral/*blood/metabolism ; Epstein-Barr Virus Infections/diagnosis/virology ; Herpesvirus 4, Human/*genetics/isolation & purification ; Humans ; Limit of Detection ; Reagent Kits, Diagnostic ; Real-Time Polymerase Chain Reaction

BACKGROUNDThere were only 3 multiple myeloma (MM) cell lines established in China. In this study, we succeeded in establishing a novel MM cell line and analyzed its biological characteristics.

METHODSMononuclear cells isolated from the peripheral blood (PB) and bone marrow (BM) of a patient with advanced MM (lambda light chain type) were cultured in medium. Cell morphology was analyzed by Wright-Giemsa-staining and cytochemical staining, immunophenotyping by flow cytometry and cytogenetic analysis by chromosome RHG-banding technique. Quantitative fluorescent polymerase chain reaction (PCR) was used to detect Epstein - Barr virus (EBV) DNA.

RESULTSThe established cell line could survive and proliferate in the presence of feeder cells or conditioned medium. The cells secreted lambda light chain and were negative for EBV. The Wright-Giemsa-staining showed typical plasmablast or plasma cell morphology. The cytochemical staining of the cells showed the following reactivity patterns: positive for acid phosphatase, negative for myeloperoxidase. The immunoprofile of the cells was concordant with that of MM cells: positive for CD10, CD28, CD38, CD138, CD56, CD49d, CD44, CD54 and CD58, negative for CD19, CD40, CD95, CD95L, CD34, CD2 and CD5. The cytogenetic analysis showed complex chromosome abnormality of i (1q+), 8q+, 13q+, i (17q), i (18q) and +M. There was no difference in morphology, immunophenotype and cytogenetics between cells from PB and BM.

CONCLUSIONSAn MM cell line secreting lambda light chain named CZ-1 was established. The cells from both PB and BM have the same biological characteristics.

Cell Line, Tumor ; Chromosome Aberrations ; Herpesvirus 4, Human ; isolation & purification ; Humans ; Male ; Middle Aged ; Multiple Myeloma ; genetics ; pathology ; Polymerase Chain Reaction

We report two cases of posttransplant malignant lymphoma(PTML) of B cell origin associated with Epstein-Barr virus(EBV) infection. They were a 52-year-old male and a 37 year-old-female, in whom intermediate-grade diffuse malignant lymphomas of large cell type developed in the submandibular area and jejunum, respectively. DNA and RNA in situ hybridization revealed the presence of EBV-specific DNA and RNA sequences in the tumor cells.
Adult ; Female ; *Herpesvirus 4, Human/isolation & purification ; Humans ; In Situ Hybridization ; Kidney Transplantation/*adverse effects/pathology ; Lymphoma/*complications/pathology/virology ; Male ; Middle Aged ; Transplantation, Homologous ; Tumor Virus Infections/*complications