Epstein-Barr Virus Type Research Articles

The Epstein-Barr virus EBNA-2 gene in oral hairy leukoplakia: strain variation, genetic recombination, and transcriptional expression.

Oral hairy leukoplakia (HLP) lesions frequently contain defective Epstein-Barr virus (EBV) genomes with deletions in the EBNA-2 gene that abundantly replicate and persist within the lesion. To characterize these viral strains and recombinant variants, the EBNA-2 gene in EBV DNA from several different HLP biopsy specimens was analyzed. Amplification of EBNA-2 coding sequences by PCR demonstrated the presence in HLP of intact EBNA-2 genes as well as a variety of internally deleted variants of both EBNA-2A and EBNA-2B. Some of the deletion variants evolved within the HLP lesion from intact EBNA-2 genes, while other variants appeared to be transmissible strains that directly infected the lesion. Intrastrain recombination within the HLP lesion also generated variation within the EBNA-2 polyproline region. Cloning and sequencing of HLP cDNA demonstrated transcription from the internally deleted EBNA-2 open reading frame, indicating that these variant genes are expressed in HLP. Comparative analysis of the HLP EBNA-2 sequences confirmed previous findings of EBV coinfection with multiple types and strains. Sequence variation of these wild-type genes demonstrated that EBNA-2A sequences distinguish at least two separate strains and a variety of substrains of EBV type 1. Two of the HLP EBNA-2A sequences contained amino acid changes in a cytotoxic T-cell epitope within an otherwise highly conserved region of the gene. These data indicate that EBV coinfection, strain variation, and recombination within the EBNA-2 gene are common features of HLP and suggest that the expression of internally deleted EBNA-2 variants could contribute to EBV pathogenesis in permissive infection.

Genetic and Biochemical Evidence That EBNA 2 Interaction with a 63-kDa Cellular GTG-Binding Protein Is Essential for B Lymphocyte Growth Transformation by EBV

Epstein-Barr virus (EBV) nuclear protein 2 (EBNA 2) is an acidic transcriptional transactivator of virus and cell gene expression and is essential for growth transformation of primary B lymphocytes. EBNA 2 transactivation of response elements (E2REs) can be mediated by interaction with a GTGGGAA-specific DNA-binding factor(s). We now purify the factor by S-Sepharose and EBNA 2 affinity chromatography and identify it as a single 63-kDa protein. The protein is shown to specifically coimmunoprecipitate with EBNA 2 from lymphoblasts transfected with an EBNA 2 FLAG expression vector. Mutation of GTG to TCT in a GTGGGAA motif common to the Cp, LMP2, and LMP1 promoters results in loss of recognition by p63. EBNA 2 amino acids 310-336 are sufficient for p63 binding. The only motif in this 27 amino acid sequence which is common to the EBNA 2 genes of EBV types 1 and 2 is GPPWWPP (I/V) (C/R) DP, which is therefore likely to mediate p63 interaction. Mutation of WW to SS or FF ablates interaction with p63, indicating that both the hydrophobic and aromatic characteristics of WW are essential for its "key" interaction with p63. EBNA 2 with a WW mutated to SS is also unable to marker rescue primary B lymphocyte transforming virus from cells infected with an EBNA 2-deleted virus, white otherwise isogeneic wild-type EBNA 2 readily marker rescues transforming virus in parallel experiments. EBNA 2 transactivation through the Cp E2RE is completely abolished by the WW to SS mutation while transactivation of -234 to +40 LMP1 E2RE is only partially affected. These genetic and biochemical experiments support the hypothesis that EBNA 2 WW interaction with a p63 GTGGGAA-binding protein is essential for EBV-mediated cell growth transformation because it specifically associates EBNA 2 with its response elements. This enables the EBNA 2 acidic domain to transcriptionally transactivate specific genes.

DNA-binding studies of the Epstein-Barr virus nuclear antigen 2 (EBNA-2): evidence for complex formation by latent membrane protein gene promoter-binding proteins in EBNA-2-positive cell lines.

The Epstein-Barr virus (EBV) nuclear antigen 2 (EBNA-2) protein is essential for the immortalization of human primary B cells by EBV. EBNA-2 trans-activates cellular and viral genes like CD23, c-fgr, latent membrane protein 1 (LMP1) and terminal protein 1 (TP1). Trans-activation of the TP1 promoter and of the BamHI C promoter has already been investigated in detail and appears to be mediated via protein-protein interactions and not by direct binding of EBNA-2 type A (of EBV type 1) to the DNA. EBNA-2 is able to trans-activate the expression of the LMP gene in several cell lines. Various reports have delineated the cis-acting elements of the LMP promoter through which EBNA-2 mediates trans-activation. To determine whether EBNA-2 also trans-activates the LMP promoter by protein-protein interactions, we performed a series of gel retardation assays and competition experiments with LMP promoter fragments of different sizes. We determined that the protein-binding region on the LMP promoter was within a 42 bp fragment encompassing nucleotides -135 to -176 relative to the LMP transcriptional start site. None of the DNA fragments investigated indicated interaction of EBNA-2 with the DNA via protein-protein interactions. No significant differences between EBNA-2-positive and EBNA-2-negative nuclear extracts could be seen in the gel retardation assay under conditions that clearly showed binding of EBNA-2A to the TP1 promoter. However, analysis of sucrose gradient fractions in the gel retardation assay provided evidence that the LMP promoter-binding proteins form a complex of higher M(r) in EBNA-2-positive cell extracts. These complexes were destroyed by detergent. We deduce from these results that EBNA-2-positive cells might indeed contain specific complexes bound to the LMP promoter which are, however, too labile to be detected in a standard gel retardation assay.

Sequence variation in the Epstein-Barr virus latent membrane protein 1.

The sequence of the latent membrane protein 1 (LMP-1) gene was analysed in Epstein-Barr virus (EBV) isolates from specific regions representing both type 1 and type 2 EBV. A predominant strain marked by an XhoI restriction enzyme polymorphism (REP) within the LMP-1 gene has been identified in type 1 EBV in nasopharyngeal carcinoma (NPC) from Southern China. This polymorphism was also present in type 2 EBV in NPC from Alaska. In this study, the sequence of the LMP-1 gene was determined in these samples representing type 1 and type 2 EBV and was compared with the prototype lymphoid strains. Consistent nucleotide variation in the amino terminus of LMP-1 was identified in strains marked by the XhoI REP. These changes were present in both EBV type 1 and type 2 strains. Three types of sequence variation were detected in the carboxy terminus of LMP-1. The LMP-1 sequences differed in the number of an 11 amino acid repeat element. In the prototype EBV type 1 (B95-8) sequence and in the type 1 Raji and type 2 HR-1 strains, the third repeat element contained an insertion of 5 amino acids that were also the first five unique amino acids after the last partial repeat element. The third variation was a deletion of amino acids 343 to 352 of the B95-8 LMP-1. This deletion was detected in the type 1 Chinese EBV strains, but was not detected in the type 2 Alaskan strains although the Chinese and Alaskan strains have nearly identical amino acid changes at the amino terminus. Numerous other amino acid changes were detected in the carboxy terminus which did not cosegregate with either EBV type, amino acid changes in the amino terminus, or specific geographic regions. These data indicate that EBV strains can be distinguished by sequence differences within LMP-1 and that unlike the divergence between type 1 and type 2 EBV in Epstein-Barr nuclear antigen sequences, different EBV types are nearly identical in LMP-1 sequence.

Characterization of a human herpes virus-6 (HHV-6) and epstein-barr virus (EBV) associated leukemic cell line, J6-1

This report characterizes the J6-1 cell line derived from a Chinese acute myelomonocytic leukemia patient and previoulsy reported to be associated with EBV. These studies showed that J6-1 cells were also infected with HHV-6 as demonstrate at the DNA level by PCR and Southern blot hybridization and by expression of HHV-6 early membrane antigen on the J6-1 cell surface. Further characterization showed J6-1 was co-infected with EBV type 2. Generally, cells infected with EBV type 2 do not grow wellin vitro, However, J6-1, although difficult to maintainin vitro, has been growth for 15 years. Possibly, co-infection with HHV-6 confers this property. In this regard, J6-1 cells exhibited density dependent growth which could be inhibited with an anti-HHV-6-MA monoclonal antibody (MAb). In contrast, anti-HHV-6-VCA MAb stimulate the J6-1 cell proliferation. Electron microscopic analysis showed that, morphologically, there were two types of J6-1 cell, one with lymphoblastoid features and one with a monocytoid appearance. Accordingly, the flow profile of the J6-1 cell line showed heterogeneity with two populations comprised of CD15-, CD19+cells with low light scatter (small cells) and a population with greater light scatter (larger cells) which was CD15+, CD19+, The population was negative for progenitor cell markers (CD33, 34), and T cell markers. Southern analysis showed no T cell receptor rearrangement, however there was a clonal JH and kappa light chain expressing population. Glycocytochemical analysis showed several endogenous lectin receptors on the J6-1 cell surface: BSA-Xylose, BSA-Rhamnose, BSA-Gal, BSA-Lac. This cell line shares many characteristics with other monocytic/lymphoblastoid cell lines isolated elsewhere and provides circumstantial evidence linking Herpes viruses, as least as co-factors, to leukemia cell growth.

Biologic aspects of AIDS-related lymphoma.

Lymphoma is one of the defining manifestations of AIDS. Most of these lymphoproliferations are high-grade B-cell non-Hodgkin's lymphoma. Unlike lymphoproliferations that arise in other settings of immunodeficiency, HIV-related lymphomas have a variable association with Epstein-Barr virus (EBV) and also contain alterations in c-myc and p53. EBV infection appears to precede clonal expansion, and its latent expression pattern (Epstein-Barr nuclear antigen1+/Epstein Barr nuclear antigen 2-/latent membrane protein+) is unique among non-Hodgkin's lymphomas. Both EBV types A and B are present in HIV-related lymphomas. Mutations in c-myc include translocations and point mutations. Other altered loci include ras and bcl-6. Although all of these somatic alterations can be detected in lymphomas arising in the general population, their accumulation in a relatively short period (6 to 8 years) after HIV infection suggests an acceleration of underlying mechanisms.

Epidemiology, pathogenesis, animal models, and treatment of Sjögrenʼs syndrome

Sjögren's syndrome is characterized by dry eyes (xerophthalmia) and dry mouth (xerostomia). Although general agreement exists about the ocular features of Sjögren's syndrome, significant controversy surrounds the classification criteria for defining the oral component. This has resulted in confusion in both the clinical and the research literature. The recent litigation involving silicone breast implants has forced clinicians to evaluate critically whether the condition of a particular patient fulfills specific diagnostic criteria for Sjögren's syndrome. Research studies have focused on potential genetic and environmental factors in the pathogenesis of Sjögren's syndrome. Studies of various ethnic populations have demonstrated an association with particular HLA-DR and -DQ alleles. However, the associated alleles are different for each ethnic group. No single environmental agent has been identified as a critical cofactor. Studies have concentrated on herpesviruses (Epstein-Barr virus and human herpesvirus type 6), hepatitis C virus, and retroviruses. Epstein-Barr virus isolated from patients has altered the ability to transform and lytically infect particular types of lymphocytes. Hepatitis C can lead to sicca symptoms, even in patients with relatively normal salivary gland biopsy findings. One report of Japanese patients indicated the presence of human T cell lymphotropic virus type I-like tax genes in the salivary biopsy specimens of a subset of patients and no gag, pol, or env sequences; this finding suggested a potential infection by a defective retrovirus. Studies on the pathogenesis have indicated that cytokines produced in the salivary gland are similar to T helper type 1 lymphocytes (interferon gamma).(ABSTRACT TRUNCATED AT 250 WORDS)

Detection of A-Type and B-Type Epstein-Bart Virus in Throat Washings and Lymphocytes

Peripheral blood lymphocytes, obtained from 94 individuals, were assayed for the presence and type of Epstein-Barr virus (EBV) using primers specific for A-type and B-type EBV in the polymerase chain reaction (PCR). Samples from 16 individuals (17%) were negative for EBV sequences. Of the remaining individuals A-type EBV was detected in 35%, B-type in 21%, and both A- and B-type EBV in 27%. Samples of throat washings were also collected from 33 healthy donors and the presence and type of EBV was determined using PCR. EBV was not detected in 12 donors. However, of those who were excreting EBV, A-type EBV was present in 11 donors (52%), B-type in 7 donors (33%), and both A- and B-types in the remaining 3 donors (14%). These results suggest that infection with B-type EBV and coinfections with both A- and B-type EBV are more common than previously thought.

Epstein-barr virus in patients with polymorphic reticulosis (lethal midline granuloma) from china and japan

Polymorphic reticulosis is one of several diseases constituting lethal midline granuloma (LMG). Previous immunohistochemical studies suggested a T-cell nature of proliferating cells; the term nasal T-cell lymphoma (NTL-LMG) has since been used widely. The authors' previous study in Asian countries showed the clustering of Mongolian patients with NTL-LMG, but the frequency varied with geographic area; it was much higher in Korea and southwest Japan (Okinawa) than in Shanghai and Honshu, Japan. Recently an etiologic role of Epstein-Barr virus (EBV) for the development of NTL-LMG has been postulated. In this study, the presence of EBV and human T-cell lymphocytic leukemia virus type 1 (HTLV-1) genomes were examined in NTL-LMG patients from Southwest Japan (Okinawa, 10 patients), another Japanese district (Honshu, 21 patients), and Shanghai, China (5 patients). All of the tissues from different geographic sites were analyzed at one central location. Immunohistochemistry showed that proliferating large cells were positive for CD43 and/or CD45RO, identical with reported NTL-LMG cases. Polymerase chain reaction (PCR) revealed the presence of EBV genome in the NTL-LMG lesions, but the frequency varied according to the geographic area: 67% in Okinawa, 33% in Honshu, and 100% in Shanghai. In situ hybridization provided positive signals in the nuclei of proliferating cells. Expression of latent membrane protein in the proliferating cells of cases positive for EBV by PCR and in situ hybridization was confirmed. The results suggest that the EBV may play a role in the development of NTL-LMG. However, the variation of frequency of EBV genome in different geographic locations suggests that EBV infection may not be an indispensable condition for the disease.

Molecular characterization of CD30+ anaplastic large-cell lymphoma: high frequency of c-myc proto-oncogene activation

Anaplastic large-cell lymphoma (ALCL) represents a morphologically distinct type of non-Hodgkin's lymphoma (NHL) characterized phenotypically by the expression of the CD30 antigen, a new member of the nerve growth factor gene family. The lymphoid origin of ALCL has been documented using immunohistochemical and molecular genetic analyses. However, very little is known so far regarding the precise pathogenetic mechanisms involved in its development and progression. Therefore, we investigated bcl-2, p53, and retinoblastoma gene (Rb) expression immunohistochemically; the occurrence of bcl-2, c-myc, and Rb gene rearrangements using Southern blotting; and the presence of ras and p53 gene somatic mutations by single-strand conformation polymorphism assay in a panel of 18 well-characterized ALCLs. In addition, the presence of Epstein-Barr (EBV) and human T-cell lymphotropic virus type I (HTLV-I) genomes were investigated using polymerase chain reaction. We identified abnormal c-myc gene products in 6 of 18 cases (33%) of ALCL. On the other hand, the bcl-2 and Rb genes were not rearranged and K-, N-, and H-ras gene somatic mutations were not found. Significant levels of p53 protein expression were found in more than 60% of ALCLs, but only a single ALCL carried a p53 gene mutation (exon 5). Only 3 ALCL cases, all occurring in human immunodeficiency virus-infected patients, were positive for EBV genomes. On the other hand, contrary to previous findings, no HTLV-I products could be identified. Despite the fact that the c-myc proto- oncogene appears to be frequently altered in ALCL, no pathognomonic abnormality could be identified and therefore additional studies and new strategies should be designed to identify the pathogenetic mechanisms involved in the development of ALCL.

Molecular Characterization of CD30+ Anaplastic Large-Cell Lymphoma: High Frequency of c-myc Proto-Oncogene Activation

Anaplastic large-cell lymphoma (ALCL) represents a morphologically distinct type of non-Hodgkin's lymphoma (NHL) characterized phenotypically by the expression of the CD30 antigen, a new member of the nerve growth factor gene family. The lymphoid origin of ALCL has been documented using immunohistochemical and molecular genetic analyses. However, very little is known so far regarding the precise pathogenetic mechanisms involved in its development and progression. Therefore, we investigated bcl-2, p53, and retinoblastoma gene (Rb) expression immunohistochemically; the occurrence of bcl-2, c-myc, and Rb gene rearrangements using Southern blotting; and the presence of ras and p53 gene somatic mutations by single-strand conformation polymorphism assay in a panel of 18 well-characterized ALCLs. In addition, the presence of Epstein-Barr (EBV) and human T-cell lymphotropic virus type I (HTLV-I) genomes were investigated using polymerase chain reaction. We identified abnormal c-myc gene products in 6 of 18 cases (33%) of ALCL. On the other hand, the bcl-2 and Rb genes were not rearranged and K-, N-, and H-ras gene somatic mutations were not found. Significant levels of p53 protein expression were found in more than 60% of ALCLs, but only a single ALCL carried a p53 gene mutation (exon 5). Only 3 ALCL cases, all occurring in human immunodeficiency virus-infected patients, were positive for EBV genomes. On the other hand, contrary to previous findings, no HTLV-I products could be identified. Despite the fact that the c-myc proto-oncogene appears to be frequently altered in ALCL, no pathognomonic abnormality could be identified and therefore additional studies and new strategies should be designed to identify the pathogenetic mechanisms involved in the development of ALCL.

Clonality of Benign Lymphoid Hyperplasia in Orbit and Conjunctiva

In order to thoroughly characterize the clonal population of lymphoid hyperplasia of the orbit and conjunctiva, we investigated six cases which were histologically proven to be benign lymphoid hyperplasia. We analyzed the clonal rearrangements of the antigen receptors and bcl-2 gene, Epstein-Barr virus (EBV), and human T-cell leukemia virus type 1 (HTLV-I) by Southern blot and/or polymerase chain reaction (PCR), and performed in situ hybridization for mRNA of kappa and lambda immunoglobulin. Five cases showed rearrangements of immunoglobulin heavy chain gene (JH) and/or light chain gene (J kappa), and the monoclonal V-J recombination of JH in PCR. However, the rearranged bands were much more faint than was the germ-line band. We considered the monoclonal population of B cells small. Two of the five cases recurred locally after four and nine years respectively. Because benign lymphoid hyperplasias frequently contain an occult monoclonal B-cell population, a follow-up should be conducted. The remaining case in our investigation showed a rearrangement of the T-cell-receptor gene and proviral DNA of HTLV-I, and it showed rapid progress to adult T-cell leukemia after the biopsy. EBV and bcl-2 gene rearrangements were not observed in any of the six cases we studied.

Changes in the dominant Epstein--Barr virus type during human immunodeficiency virus infection

Two types of Epstein-Barr virus (EBV), EBV-1 and EBV-2, were identified on the basis of DNA sequence divergence in the genes for nuclear proteins EBNA 2, 3a, 3b and 3c. In the present study, we conducted an immunological and genomic analysis in a human immunodeficiency virus (HIV)-infected population to determine the prevalence of the two types, and whether the identified type was stable over years. The EBNA-2 serotyping and genotyping showed that HIV-infected patients were highly infected by EBV-2, and that the dominant strain was mostly retained. However, during a follow-up study, a change in the dominant viral strain was observed in two patients. A first HIV-positive patient (patient A), although having a stable level of anti-EBNA-2A and -2B antibodies, showed a change in the genotype and antigen produced in spontaneously established lymphoblastoid cell lines (LCL). The sequence analysis of LCLs confirmed the emergence of the EBV-2 type population. A strain from a second HIV-positive patient (patient B) was clearly identified as EBV-2: the genotype from a saliva sample and from sequential LCLs belonged to EBV-2, as well as the antigen produced from LCLs, and serum antibodies. After a 5-year continuous EBV-2 infection, a reactivation of the EBV-1 strain was observed. In both cases, sequence analysis of the EBNA-2 gene showed, only with EBV-1, the presence of EBV variants related to the B95-8 prototype. Two mutations (at nucleotides 49212 and 49304) were found in both patients A and B, whereas an additional mutation (at nucleotide 49237) was characteristic of the patient A. No mutation relative to the prototype B95-8 strain was observed in a subsequent analysis of this EBNA-2 region from LCLs obtained from two HIV-negative patients predominantly infected by EBV-1. Therefore, we speculate that these mutations may be EBV-1 mutations specifically occurring during HIV infection.

A modified procedure for the propagation of wild type Epstein-Barr virus in cultures of marmoset blood cells

Productive infection of marmoset blood mononuclear cells by the Epstein-Barr virus was generally achieved by the co-cultivation method. By introducing 2 modifications, the success rate of infection was increased from 6% to 79%. The modifications consisted of the selection of human lymphocyte donors to serve as carriers of Epstein-Barr virus in cocultures, and the addition of cyclosporin A to culture media. So far, 10 of 10 wild type oropharyngeal EBV have been propagated successfully in cultures of marmoset blood mononuclear cells by the modified procedure. Fragment length polymorphism study failed to reveal any difference between viral genomes in human lymphocytes and that in marmoset blood mononuclear cells. Antigenic analysis of 8 wild strains showed that all were related to the B95-8 strain of Epstein-Barr virus by the neutralization test.

Association of Epstein-Barr virus types 1 and 2 with acquired immunodeficiency syndrome-related primary central nervous system lymphomas [letter; comment

Association of Epstein-Barr virus types 1 and 2 with acquired immunodeficiency syndrome-related primary central nervous system lymphomas [letter; comment

Characterization of Epstein‐Barr viral strains in parotid gland saliva and peripheral blood of patients with primary Sjögren's syndrome and healthy EBV carriers

Increased Epstein-Barr virus (EBV) replication has been reported in the salivary and lacrimal glands in Sjögren's syndrome (SS). We studied whether or not certain EBV strains would occur preferentially in the peripheral blood and parotid gland saliva of 18 EBV-seropositive patients with primary Sjögren's syndrome (pSS) and 12 EBV-seropositive control persons. Transforming EBV was detected in the blood of 11 of 18 (61%) pSS patients and 9 of 12 controls (75%). Unexpectedly, neither transforming nor Raji-superinfecting EBV strains were detected in SS parotid saliva, whereas these EBV types were detected in control saliva in 7 and 8 cases, respectively (P < 0.001). Transforming EBV strains were further characterized by 'Ebno-typing,' i.e., analysis of the size spectrum of the viral antigens EBNA 1, 2, 3, and 6 in immunoblots of lymphoblastoid cell lines (LCL). Previous work has shown that a single EBV strain (Ebnotype) dominates the blood and oropharynx of healthy carriers and that unrelated individuals carry different EBV strains, reflecting the vast polymorphism of Ebnotypes in the general population. Two unexpected observations were made. First, an identical Ebnotype was detected in 4 unrelated individuals, i.e., in the blood of 1 pSS patient and in the saliva of 3 control persons. Second, carriage of 2 to 4 different Ebnotypes by a single individual was observed in 4 cases, i.e., in the blood of 1 pSS patient, and in the blood and saliva of 3 control persons.(ABSTRACT TRUNCATED AT 250 WORDS)

Nasopharyngeal carcinoma: genetic changes, Epstein-Barr virus infection, or both. A clinical and molecular study of 36 patients.

The pathogenesis of nasopharyngeal carcinoma has been under intense investigation, especially of its peculiar predominance in southern China. The authors previously reported consistent loss of genetic material on the short arm of chromosome 3 in a few nasopharyngeal carcinoma cases. In this study, the authors examined the genetic changes as well as the presence of Epstein-Barr virus (EBV) genome on 36 nasopharyngeal carcinoma primary biopsy specimens of the undifferentiated type and the correlation of the findings to patients' clinical status. The authors examined the DNA from tumor tissue and from matched blood leukocytes of 36 patients who suffered from nasopharyngeal carcinoma by the restriction fragment length polymorphism analysis. The genotyping for EBV was carried out by polymerase chain reaction using primers complementary to both types of EBV and probes specific to EBNA-2A (EBV-A) or EBNA-2B (EBV-B). A consistent deletion at two specific locus of the short arm of chromosome 3 was observed in all informative cases. The authors also found that EBV genome, especially type A, was present in 35 of 36 cases. In the remaining one case, EBV-B was detected. As the same tumor tissue was used for both genetic and viral studies in each case, the results may represent sequential genetic lesions in the pathogenesis and/or summation of genetic events. Moreover, 7 of 32 informative tumors were from patients of early staging (Stages I and II), which suggests the genetic changes may occur in the early development of nasopharyngeal carcinoma. Difference in allele frequency in specific locus was also noted between Asian and white patients for the first time.

Epstein-Barr virus infection of HIV-seropositive individuals is transiently suppressed by high-dose acyclovir treatment

To assess whether oral acyclovir can eliminate persistent Epstein-Barr virus (EBV) infection and thereby prevent EBV-associated lymphoma development in HIV-seropositive homosexual men. Persistent EBV infection was examined in a group of 21 HIV-seropositive homosexual men before, during and after treatment with oral acyclovir at a dose of 800 mg every 6 h (10 individuals) or with a placebo (11 individuals). In 13 individuals, EBV was isolated from the oropharynx before and after treatment (seven cases from the acyclovir-treated group and six from the placebo-treated group). A significant reduction in virus isolation occurred during treatment in the acyclovir-treated group, but not in the placebo-treated group. In seven cases in whom EBV shedding was detected before and after treatment, the EBV strain isolated was identical throughout the study, even when acyclovir had abolished detectable shedding for the duration of the treatment. In two other cases more than one strain was detected. On examination of the EBV type present, 89% of a group of 18 patients consistently shed type A virus, 5.5% type B virus and 5.5% showed evidence of co-infection with both virus types. This compares with figures of 100, 0 and 0%, respectively, in a control group of HIV-seronegative individuals. High-dose acyclovir therapy does not eliminate persistent EBV infection from the oropharynx of healthy HIV-seropositive individuals and therefore would not necessarily prevent lymphoma development. Our results suggest that infection by type B EBV, and co-infections of both A and B type, are more common in HIV-seropositives than HIV-seronegatives.

Subtypes of Epstein-Barr virus in HIV-1-associated and HIV-1-unrelated Hodgkin's disease cases.

Epstein-Barr virus (EBV) type 2 is considered to be a much less potent transformer of lymphocytes than type 1. However, type-2 EBV may be involved in the pathogenesis of non-Hodgkin's lymphomas (NHLs) arising in immunocompromised patients, i.e., subjects with malaria or HIV-1 infection. To determine whether type-2 EBV may also play a role in Hodgkin's disease (HD) developing in immunocompromised patients, we characterized EBV subtypes in EBV-positive HD samples from 10 HIV-1-positive patients as well as from a control population of 24 HIV-1-negative patients. Type-2 EBV was detected in 5/10 HD samples from the HIV-1-positive group (1 case showed concomitant type-1 EBV positivity), but only in 1/20 HD samples from the HIV-1-negative group, indicating that, during HIV-1-induced immunodepression, type-2 EBV may be pathogenetically involved also in HD, as previously reported for HIV-associated NHLs.

Strain Variation in Epstein-Barr Virus Immediate Early Genes

RNase protection analysis of mRNAs for Epstein-Barr virus (EBV) genes activated early in the induction of the productive cycle in Akata cells has confirmed that BZLF1 and BRLF1 behave as immediate early genes. These are likely to be the only virus immediate early genes since analysis of three other transcripts previously suggested to behave as immediate early genes in this system shows that they are, in fact, early genes. Western blotting of BRLF1 and BZLF1 demonstrates expression of these proteins early in induction of the productive cycle and reveals that the BZLF1 protein of Akata virus migrates more slowly on SDS gels than the BZLF1 protein of the prototypic virus strain B95-8. The nucleotide sequence of the BZLF1 gene and the immediate early promoters of Akata EBV have been determined and compared to other EBV strains. RFLP analysis of sequence variation in the BZLF1 gene of EBV strains from many different parts of the world shows a distribution of BZLF1 types which are not simply related to the established A and B EBV types.