Antibodies to the second Epstein-Barr virus nuclear antigen in non-Hodgkin lymphomas

Human antibodies against the first and second Epstein-Barr virus encoded nuclear antigens (EBNA-1 and EBNA-2) were analysed by modified ELISA reaction on immunoblots: 5 out of 20 EBNA-1 positive, healthy donor sera reacted with the EBNA-2. Healthy EBV seronegatives contained no antibodies against EBNA-2. The antibodies against this EBNA-2 developed several months after acute EBV infections. Four out of 8 infectious mononucleosis sera failed to react EBNA-2 in the late stage. In the group of EBV seropositive patients with non-Hodgkin lymphoma, 14 out of 18 sera contained EBNA-2 antibodies.     

Detection of antibodies to the Epstein-Barr virus nuclear antigens in the sera from patients with systemic lupus erythematosus

The sera from 65 patients with systemic lupus erythematosus (SLE) were examined by the immunoblotting method to detect antibodies to Epstein-Barr virus (EBV)-associated antigens, especially EBV nuclear antigens (EBNA), and compared with the sera from 66 healthy subjects roughly age- and sex-matched to the patients. Most sera from patients with SLE defined three major EBV-associated antigens with molecular weights (MW) of 70,000 (70K), 90K and 140K in Raji cells, which must correspond to the EBNA-1, 2, and 3, respectively. Approximately 70% of the sera from SLE patients demonstrated the antibodies to the 90K and 140K antigens, whereas the positive rates of these two antibodies were less than 10% in the sera from healthy subjects. The differences of these positive rates of the antibodies between SLE patients and healthy subjects were statistically highly significant. Antibody to EBNA-1 was conspicuously detected in the sera from both SLE patients and healthy subjects, although the difference between the two groups was still significant. The possible role of EBV infection was discussed on the basis of the pathogenesis of SLE.     

Use of bacterially expressed GST/EBNA-1 fusion proteins for detection of antibodies in sera from patients with nasopharyngeal carcinoma and healthy donors

Epstein-Barr virus nuclear antigen-1 (EBNA-1) is a protein expressed consistently in EBV infected cells and in EBV related malignant tissues. Antibodies against EBNA-1 may therefore possibly be used as a marker for disease screening. Western blot analysis of serum antibodies was performed using GST (glutathione-S-transferase) fusion proteins containing different regions of EBNA-1 as antigens. Serum samples were collected from 38 patients with nasopharyngeal carcinoma (NPC) and 38 healthy individuals in Taiwan. All samples were found IgG positive for EBNA-1 when a truncated protein GST/E1 (70-102, 325-641) was used as the antigen. Thirty-three out of 38 NPC sera (86.8%) were positive for IgA antibody against EBNA-1. The positive rate was higher in comparison with IgA antibody against VCA (65.7%) or antibody against DNase (60.5%). Only 2.6% of sera from normal individuals were positive for an IgA response against EBNA-1. The major antigenic determinants for NPC serum IgA response were between amino acid(aa) 390 to aa 459 when different portions of EBNA-1 were used as antigens. The results suggest that IgA response against EBNA-1 could be used in combination with other EBV serology markers for NPC screening.     

Anti-Epstein-Barr virus-nuclear antigen-1, -2A and -2B antibodies in rheumatoid arthritis patients and their relatives.

We have examined serum antibodies to Epstein-Barr virus Nuclear Antigen (EBNA)-1, -2A and -2B, in addition to antibodies to viral capsid antigen and early antigen in 100 rheumatoid arthritis patients and 50 of their relatives. Using indirect immunofluorescence on transfected cells and Western-blot technique, we have found increased frequency and titres of antibodies to EBNA-2B in patients and, to a lesser degree, in their family members, whereas other anti-Epstein-Barr virus antibodies appeared to be similar to controls. Cross-inhibition experiments were carried out and show that antibodies to EBNA-2A are distinct from those to -2B, and vice versa.     

Cross-reactivity between the EBNA-1 p107 peptide, collagen, and keratin: implications for the pathogenesis of rheumatoid arthritis.

An unusually heavy load of Epstein-Barr virus (EBV) infection and autoimmunity to collagen are believed to be contributing factors to the pathogenesis of rheumatoid arthritis (RA). The present report presents data showing that p107, the major epitope of the EBV-encoded EBNA-1 antigen, cross-reacts with denatured collagen (DC) and keratin (K), suggesting a new likely link among RA, EBV-1, and these autoantigens. A radioimmunoassay using antigen-coated microtiter plates was used to demonstrate antibodies in sera of patients with RA and sera of healthy donors against p107, DC, and K. Specificity of the antibodies was ascertained by inhibition tests with the homologous antigens. Cross-reactivity among anti-p107, anti-DC, and anti-K antibodies was assayed by the ability of a given antigen to block the binding of nonpurified or affinity-purified antibodies to plates coated with another antigen. Most of the sera contained antibodies to all three antigens, but only anti-DC antibodies were present in higher titers in RA sera. Preincubation of sera with p107 appreciably reduced their binding to plates coated with DC or K. On the other hand, preincubation with DC (in solution or bound to Sepharose) did not result in consistent reduction of anti-p107 titers. Tests with affinity-purified antibodies revealed the existence of two antibodies populations, one of which reacted preferentially with p107, the other with DC. The cross-reactivity of the anti-p107 antibodies with DC and K suggests that such antibodies, produced by RA patients following persistent stimulation with EBV, might react in vivo with collagen (and keratin) exposed in previously damaged areas and thus reinforce the disease process.     

Rat monoclonal antibodies differentiating between the Epstein-Barr virus nuclear antigens 2A (EBNA2A) and 2B (EBNA2B)

Rat monoclonal antibodies were produced against the C-terminus of Epstein-Barr virus nuclear antigens 2A (EBNA2A) and 2B (EBNA2B) expressed as bacterial trpE fusion proteins. The initial screening was performed using a soluble bacterial extract containing the fusion proteins. Positive hybridomas were confirmed by immunofluorescence on SF158 (Spodoptera frugiperda) insect cells infected with recombinant baculovirus (Autographa californica nuclear polyhedrosis virus) and expressing the complete EBNA2A or EBNA2B genes. We selected a panel of antibodies which reacted either with both antigens or specifically with EBNA2A or with EBNA2B. The antibodies were extensively characterized using immunoprecipitation, Western blotting, epitope mapping on synthesized peptide segments of EBNA2A, immunocytology, and immunohistology on both cryostat sections and paraffin sections of AIDS-associated primary central nervous system lymphomas.     

Synthesis of fusion proteins of Epstein-Barr virus nuclear antigens in E. coli and their antigenicity.

Expression and yield in E. coli of a panel of fusion proteins containing various domains of Epstein-Barr virus nuclear antigens, EBNA-1, EBNA-2, EBNA-3, EBNA-4 and EBNA-6, were scrutinized. The antigenicity of the EBNA fusion proteins against human sera was examined. Monospecific antisera to the different EBNA domains were produced by immunizing guinea pigs and rabbits. An EBNA-6 fusion polypeptide was useful for separating anti-EBNA-6 antibody from human sera by immunoaffinity purification. The applications of the fusion proteins to clinical diagnosis are discussed.     

Characterization of an Epstein-Barr virus nuclear antigen 2 variant (EBNA 2B) by specific sera

The Jijoye Epstein-Barr virus (EBV) strain is characterized by a substitution of 1.8 kb in the C-terminal part of EBNA 2 gene compared to B95-8 or M-ABA virus. Protein immunoblot analysis using human sera against EBNA 2 indicated that an immunological variant to the EBNA 2 of B95-8 (type A) is encoded by the Jijoye virus (type B). In order to generate a specific EBNA 2B antiserum the NaeI/NsiI DNA fragment of the Jijoye virus containing 237 bp of the C-terminus from the EBNA 2B gene was cloned in an E. coli expression vector (pME3). The resulting fusion protein contained 79 C-terminal amino acids of the viral protein and a 37,000 Da part of the bacterial anthranilate synthase. Rabbit antisera generated against this fusion protein reacted specifically with two proteins of 73,000 and 77,000 Da from Jijoye cells and three other cell lines carrying type B virus, while no proteins could be identified in the type B cell line BL 29. In addition, using these sera directed against the pME3 fusion protein, no reaction could be observed with the EBNA 2A protein from the B95-8 and several other cell lines containing type A virus.     

Antibodies to a new viral citrullinated peptide, VCP2: fine specificity and correlation with anti-cyclic citrullinated peptide (CCP) and anti-VCP1 antibodies

Anti-citrullinated protein/peptide antibodies (ACPA) are a hallmark of rheumatoid arthritis (RA) and can be measured using different citrullinated substrates. In this paper we describe a new viral citrullinated peptide - VCP2 - derived from the Epstein-Barr virus-encoded protein EBNA-2 and analyse its potential as substrate for ACPA detection. Analysing sera from 100 RA patients and 306 controls, anti-VCP2 immunoglobulin (Ig)G were found in 66% of RA sera, IgM in 46% and IgA in 39%, compared with less than 3% of control sera. Anti-VCP2 IgG was associated with erosive arthritis, the presence of rheumatoid factor and anti-VCP1 and anti-cyclic citrullinated peptide (CCP) antibodies. Anti-VCP2 antibodies were detected in 1% and anti-VCP1 antibodies in 4% of CCP-negative RA sera; conversely, 3% of the VCP-negative sera were CCP-positive. Taken together, these data suggest that VCP2 could offer a valuable tool for ACPA detection. Inhibition assays showed that two non-overlapping epitopes - a citrulline-glycine stretch shared between VCP1 and VCP2 and the N-terminal portion of the VCP2 sequence - were targeted by anti-VCP2 antibodies. Moreover, in some RA sera that tested positive in CCP and VCP2 assays, preincubation with VCP2 inhibited binding to CCP, whereas in other sera the binding was unaffected. Thus, the reactivity with more than one ACPA substrate might be due in some RA patients to antibody populations with different specificities, and in others to cross-reactive antibody populations. Finally, affinity-purified anti-VCP2 antibodies immunoprecipitated deiminated Epstein-Barr virus nuclear antigen (EBNA-2) from an EBNA-2-transfected cell line, suggesting that viral sequences may be involved in the generation of the ACPA response.     

Infectious mononucleosis patients temporarily recognize a unique,cross-reactive epitope of Epstein-Barr virus nuclear antigen-1

The spectrum of antibodies against Epstein-Barr virus nuclear antigen-1 (EBNA-1) in patients with a recent history of infectious mononucleosis and nonaffected EBV-positive individuals has been characterized by epitope mapping. Sera were evaluated for antibodies to all unique maximally overlapping octapeptides of EBNA-1. Both normal controls and patients with infectious mononucleosis produce IgG antibodies that recognize the glycine-alanine-rich portion of EBNA-1, as previously described. All EBNA-1 IgG-positive infectious mononucleosis patients tested, however, consistently produce IgG specific for an additional epitope (aa 398-412 PPPGRRPFFHPVGEA) near the middle of the EBNA-1 protein. This region was not found to be antigenic in healthy EBV-seropositive individuals. This region does, however, cross-react with the sequence PPPGMRPP from the common lupus spliceosomal autoantigen Sm B' in several infectious mononucleosis patient sera. Patients with recent clinical infectious mononucleosis temporarily recognize a unique cross-reactive epitope of EBNA-1 not bound by antibodies from non-infectious mononucleosis EBV-positive sera or those with a distant history of IM.     

Development of a human herpesvirus 6 species-specific immunoblotting assay

In order to assess the full spectrum of human herpesvirus 6A (HHV-6A)- and HHV-6B-associated diseases, we sought to develop an HHV-6 species-specific serological assay based on immunoblot analysis. The immunodominant proteins encoded by open reading frame U11, p100 for HHV-6A (strain U1102) and 101K for HHV-6B (strain Z29), were selected to generate virus species-specific antigens. Recombinant p100 and 101K were produced in a prokaryotic expression system. The expression of these proteins was confirmed by using anti-His tag and 101K-specific monoclonal antibodies. HHV-6 species-specific antibodies were detected by immunoblotting in patient sera. Eighty-seven serum samples obtained from various subjects were utilized to determine the reliability of the method for clinical use. Ten of twelve exanthem subitum convalescent-phase sera reacted exclusively with 101K, whereas none of twelve acute-phase sera reacted with either protein. Two of three sera collected from HHV-6A-infected patients reacted with p100 and 101K. Although all five acute and convalescent-phase sera obtained from transplant recipients reacted exclusively with 101K, two of six convalescent-phase sera obtained from patients with drug-induced hypersensitivity syndrome reacted with both p100 and 101K. Of 38 sera obtained from healthy adults, 31 were positive for 101K antibody, while 4 reacted with both proteins. However, PCR analysis of peripheral blood mononuclear cells and saliva from these subjects did not detect HHV-6A DNA. In conclusion, this novel serological assay based on immunoblot analysis using recombinant HHV-6A p100 and HHV-6B 101K allowed us to discriminate between HHV-6A- and HHV-6B-specific antibodies.     

Expression of Epstein-Barr virus nuclear antigens 3, 4, and 6 are altered in cell lines containing B-type virus

A high proportion of HIV-positive sera were found to react with 130- and 180-kDa antigens which were present in the Jijoye cell line. The majority of the HIV-positive sera which detected these antigens also contained antibodies to Epstein-Barr virus (EBV) nuclear antigen 2B (EBNA2B) suggesting a relationship between B-type EBV strains and the expression of the 130K/180K antigens. Cell lines were established by infection of B lymphocytes with different A- and B-type strains of EBV. Incubation of these lines with sera from individuals infected with either A-type or B-type EBV strains demonstrated that the 130K and 180K antigens were only expressed by cell lines containing B-type virus. Sera from individuals infected with A-type EBV did not react with the 180K antigen in any cell lines but could detect EBNAs 3, 4, and 6 antigens in the A-type cell lines. Restriction enzyme analysis of the BamHI E region of the EBV genome revealed marked differences between the A and B types of the virus. These results demonstrate that expression of antigens encoded from the BamHI E region of EBV (EBNAs 3, 4, and 6) are altered in cell lines transformed by B-type strains of EBV.     

Antibodies against membrane antigens of cytomegalovirus infected cells in sera of patients with a cytomegalovirus infection

The appearance of a new virus specific antigen was demonstrated by an indirect immunofluorescence technique on cell surfaces of CMV infected human fibroblasts, 48–72 hr after inoculation. The development of antibodies to these membrane antigens was followed in thirty-nine serial sera from twelve patients with a virologically and/or serologically confirmed CMV infection. In all patients except one, membrane antibodies could be detected. Sera from four patients collected before infection were negative, as were sera taken 1–13 days after onset of symptoms. From the end of the second week of illness CMV-membrane antibodies as well as CMV macroglobulin antibodies to intracellular antigens were detectable. The membrane antibodies persisted until 335 days after onset of illness. They were mainly of the IgM class. Most positive sera also reacted with non-infected fibroblasts but to a lesser degree. This reaction became negative after prior absorption of the sera with non-infected fibroblasts, whereas the reaction with CMV-infected cells remained positive.Controls consisted of forty-five sera from healthy persons and nine sera from patients suffering from other herpes virus infections with antibodies to herpes simplex, zoster/varicella and Epstein-Barr virus. All but two sera were negative in the membrane immunofluorescence test.     

Elevated immunoglobulin G antibodies to the proline-rich amino-terminal region of Epstein-Barr virus nuclear antigen-2 in sera from patients with systemic connective tissue diseases and from a subgroup of Sjögren's syndrome patients with pulmonary involvements

Associations of Epstein-Barr virus (EBV) and autoimmune diseases have been hypothesized. We have analysed IgG antibodies to EBV nuclear antigen (EBNA)-2 in sera from Japanese patients with autoimmune systemic connective tissue diseases (CTD), exemplified by systemic lupus erythematosus (SLE), primary Sjogren's syndrome (SS), rheumatoid arthritis (RA), systemic sclerosis (SSc) and secondary SS (classical CTDs complicated with SS). An enzyme-linked immunosorbent assay (ELISA) which uses glutathione-S-transferase polypeptides fused to EBV nuclear antigen (EBNA)-2 and EBNA-1 was developed. Ratios of IgG antibody reactivity to whole IgG concentrations of sera were calculated to normalize EBNA-2 and EBNA-1 antibody levels to the hypergammaglobulinaemia that occurs in CTD. The ELISA optical density OD(450) readings of IgG antibodies to both the amino-terminal aa 1-116 of EBNA-2 and carboxyl-terminal aa 451-641 of EBNA-1 were elevated significantly in patients with SLE, primary SS, RA, SSc and secondary SS when compared to EBNA-1. The OD readings were divided by serum IgG concentrations to normalize for the hypergammaglobulinaemia. The specific levels of IgG antibodies to the amino-terminal region of EBNA-2 were elevated in patients with SLE, primary SS or RA, as well as those with secondary SS complicated with SLE or RA. The EBNA-2 amino-terminal region contains a polyproline tract and a proline-rich sequence and has considerable amino acid sequence homology with many cellular proline-rich proteins. High ratios of EBNA-2 aa 1-116 to EBNA-1 aa 451-641 IgG antibody levels which probably suggest reactivation of EBV latent infection were associated significantly with pulmonary involvement in SS patients. These results are consistent with the hypothesis that the sequence similarity between the amino-terminal region of EBNA-2 and proline-rich cellular proteins is associated with pathogenesis in a subpopulation of CTD patients, possibly by the molecular mimicry-epitope shift mechanism.     

Rheumatoid arthritis sera react with a phage-displayed peptide selected by a monoclonal antibody to type II collagen that has homology to EBNA-1.

Antibodies to type II collagen, and to Epstein Barr virus nuclear antigen-1 (EBNA-1) have been associated with rheumatoid arthritis (RA). In studies involving probing of phage-displayed random peptide libraries with an antibody to type II collagen, CII-C1, we observed that among 17 phagotopes selected 5 expressed peptides with homology with the sequence of EBNA-1. The residues in common were RLPFG. Hence we tested sera from 50 patients with RA, of whom 26 had antibodies to native type II collagen, and 43 healthy controls, for reactivity by ELISA with a phagotope selected 4 times, which expressed the peptide RRLPFGSQM. Eight RA sera (16%) but no normal sera reacted with the phagotope (p = 0.025). This reactivity could not be correlated with reactivity of RA sera with EBNA-1 by semi-quantitative western blot, with which reactivity occurred in 78% of RA patients and 81% of controls. Evidence for molecular mimicry was not found insofar as the phagotope did not inhibit reactivity of RA sera with EBNA-1 and CII-C1 was not reactive with EBNA-1. We conclude that the reactivity of the RA sera with the phagotope is most likely due to the phagotope being a mimic of an epitope of type II collagen for a proportion of RA sera.     

Production of herpes B virus recombinant glycoproteins and evaluation of their diagnostic potential

B virus (cercopithecine herpesvirus 1) is the only deadly alphaherpesvirus that is zoonotically transmissible from macaques to humans. The detection of humoral immune responses is the method of choice for the rapid identification of B virus-infected animals. We evaluated the diagnostic potential of recombinant B virus glycoproteins for the detection of immunoglobulin G (IgG) antibodies in monkey and human sera. Glycoproteins B, C, and E and secreted (sgG) and membrane-associated (mgG) segments of glycoprotein G (gG) were expressed in the baculovirus expression system, while gD was expressed in CHO cells. We developed recombinant protein-based IgG enzyme-linked immunosorbent assays (ELISAs) and compared their diagnostic efficacies by using B virus antibody-negative (n = 40) and -positive (n = 75) macaque sera identified by a whole antigen-based ELISA and Western blotting. The diagnostic sensitivities of the gB-, gC-, gD-, and mgG-ELISAs were 100, 97.3, 88.0, and 80.0%, respectively. The specificities of the gB-, gC-, and gD-ELISAs and of the mgG-ELISA were 100 and 97.5%, respectively. In contrast, the sensitivities and specificities of sgG- and gE-ELISAs were low, suggesting that sgG and gE are less effective diagnostic antigens. Sera from nonmacaque monkeys cross-reacted with gB, gC, and gD, and only baboon sera reacted weakly with mgG. Human herpes simplex virus type 1 (HSV-1)- and HSV-2-positive sera pools reacted with gB and gD, whereas sera from B virus-infected individuals reacted with all four antigens. These data indicate that gB, gC, gD, and mgG have a high diagnostic potential for B virus serodiagnosis in macaques, whereas mgG may be a valuable antigen for discrimination between antibodies induced by B virus and those induced by other, closely related alphaherpesviruses, including HSV-1 and -2.     

Serological evidence for variation in the incidence of herpesvirus infections in different species of apes.

Sera from captive lowland gorillas, chimpanzees, orangutans, and gibbons were screened by enzyme-linked immunosorbent assay (ELISA) for antibody to herpesviruses serologically related to human herpes simplex virus types 1 and 2 (HSV-1, HSV-2), a baboon virus (SA8), and a macaque herpesvirus (B virus). The incidence of herpesvirus antibodies varied considerably among the different species, gorillas having the highest incidence of seropositivity (65.4%) and orangutans the lowest. The virus specificity of positive sera was further analyzed by examining the kinetics of virus neutralization, competition of reactivity in ELISAs, and immunoblotting against HSV-1, HSV-2, SA8, and B virus antigens. Using these assays, the majority of positive gorilla sera (49 of 53, 92%) were determined to react in a manner identical to human HSV-1 immune sera. The remaining four positive gorilla sera reacted as HSV-2-positive sera. In contrast, the majority of positive chimpanzee sera (5 of 7, 71%) reacted as HSV-2 immune rather than HSV-1 immune. All positive sera from gibbon apes reacted as HSV-1 positive. No orangutan sera were identified which gave positive reactions by ELISAs to any of the four primate herpesviruses tested. Although four orangutan sera gave equivocal results against HSV-1 antigen, further analysis by immunoblotting could not confirm any specific reactivity with any of the primate herpesvirus antigens. Varied reactivity among individual animals with both SA8 and B virus proteins was observed, but none of the seropositive primates detected appeared to be infected with either of these simian viruses. Three gorilla sera had antigen recognition patterns slightly different from those of HSV-2-positive human and chimpanzee sera and another HSV-2-positive gorilla serum, raising the possibility that these animals harbor an indigenous virus related to HSV-2.     

Antibody reactivity to a synthetic peptide (P62) of the Epstein-Barr nuclear antigen in sera of patients with X-linked lymphoproliferative syndrome

An enzyme-linked immunosorbent assay (ELISA) was used to measured IgG antiboody titers againt a synthetic peptide whose sequence was derived from the glycine-alanine repeating region of Epstein-Barr virus nuclear associated antigen 1 (EBNA-1). Antibody titers were determined in sera from 15 normal subjects, sera from 21 normal male siblings of X-linked lymphoproliferative syndrome (XLP) patients, from 20 XLP patients comprising a total of 42 samples, and ten samples before and ten samples after gamma-globulin therapy in ten patients with XLP. Data analysis demonstrated that while there are differences between the ELISA and ACIF, they appear to measure a similar response as demonstrated by their correlation coefficient (0.77) and the GMT to EBNA observed by both methods. No cross-reactivity of cytomegalovirus antibodies to the EBNA-1 peptide was observed by immunobv using adsorption against AD-169 infected MRC-5 cells.. However, non-specific binding was observed if samples were not pre-incubated in a 10% goat serum PBS-Tween 20 solution. This pre-treatment removed the non-specific binding that falsely elevated GMT in approximately 15% of both normal and XLP samples in ELISA. The ELISA system appears to be a sensitive, reproducible and objective test that may be useful for assessing the antibody responses of patients to the EBNA-1 protein.     

Maintenance of serum immunoglobulin G antibodies to Epstein-Barr virus (EBV) nuclear antigen 2 in healthy individuals from different age groups in a Japanese population with a high childhood incidence of asymptomatic primary EBV infection

Immunoglobulin G (IgG) antibodies to Epstein-Barr virus (EBV) nuclear antigens 2 and 1 (EBNA-2 and EBNA-1, respectively) were studied using sera from healthy individuals of a population with a high incidence of asymptomatic primary EBV infections during infancy or childhood in Japan. Two CHO-K1 cell lines expressing EBNA-2 and EBNA-1 were used for anticomplement and indirect immunofluorescence assays. The positivity rate for EBNA-2 IgG rose in the 1- to 2-year age group, increased and remained at a plateau ( approximately 45%) between 3 and 29 years of age (3- to 4-, 5- to 9-, 10- to 14-, and 15- to 29-year age groups), and then reached 98% by age 40 (>/== 40-year age group). Both seropositivity for EBNA-1 and seropositivity for EBNAs in Raji cells (EBNA/Raji) were detected in the 1- to 2-year age group, remained high, and finally reached 100% by age 40. The geometric mean titer (GMT) of EBNA-2 IgG reached a plateau in the 5- to 9- and 10- to 14-year-old groups and remained elevated in the older age groups (15 to 29 and >/== 40 years). The GMT of EBNA-1 IgGs increased to a plateau in the 1- to 2-year-old group and remained unchanged in the older age groups. The GMT of EBNA/Raji IgGs also reached a plateau in the 1- to 2-year-old group, remained level throughout the 3- to 14-year age groups, and decreased in the 15- to 29-year-olds. EBNA-2 IgGs emerged earlier than EBNA-1 IgGs in 8 of 10 patients with infectious mononucleosis, who were between 1 and 27 years old, and declined with time in three of eight cases. These results suggest that EBNA-2 IgG antibodies evoked in young children by asymptomatic primary EBV infections remain elevated throughout life, probably because of reactivation of latent and/or exogenous EBV superinfection.     

Maintenance of Serum Immunoglobulin G Antibodies to Epstein-Barr Virus (EBV) Nuclear Antigen 2 in Healthy Individuals from Different Age Groups in a Japanese Population with a High Childhood Incidence of Asymptomatic Primary EBV Infection

Immunoglobulin G (IgG) antibodies to Epstein-Barr virus (EBV) nuclear antigens 2 and 1 (EBNA-2 and EBNA-1, respectively) were studied using sera from healthy individuals of a population with a high incidence of asymptomatic primary EBV infections during infancy or childhood in Japan. Two CHO-K1 cell lines expressing EBNA-2 and EBNA-1 were used for anticomplement and indirect immunofluorescence assays. The positivity rate for EBNA-2 IgG rose in the 1- to 2-year age group, increased and remained at a plateau (~45%) between 3 and 29 years of age (3- to 4-, 5- to 9-, 10- to 14-, and 15- to 29-year age groups), and then reached 98% by age 40 (≥40-year age group). Both seropositivity for EBNA-1 and seropositivity for EBNAs in Raji cells (EBNA/Raji) were detected in the 1- to 2-year age group, remained high, and finally reached 100% by age 40. The geometric mean titer (GMT) of EBNA-2 IgG reached a plateau in the 5- to 9- and 10- to 14-year-old groups and remained elevated in the older age groups (15 to 29 and ≥40 years). The GMT of EBNA-1 IgGs increased to a plateau in the 1- to 2-year-old group and remained unchanged in the older age groups. The GMT of EBNA/Raji IgGs also reached a plateau in the 1- to 2-year-old group, remained level throughout the 3- to 14-year age groups, and decreased in the 15- to 29-year-olds. EBNA-2 IgGs emerged earlier than EBNA-1 IgGs in 8 of 10 patients with infectious mononucleosis, who were between 1 and 27 years old, and declined with time in three of eight cases. These results suggest that EBNA-2 IgG antibodies evoked in young children by asymptomatic primary EBV infections remain elevated throughout life, probably because of reactivation of latent and/or exogenous EBV superinfection.