| Aim: Epstein-Barr virus(Epstein-Barr virus, EBV) is closely related to the occurrence of human lymphoma, the distribution has certain regional and population orientation, in addition to the environmental factors and genetic factors, whether EBV gene variants associated with EBV positive lymphoma has been closely watched. Polymerase chain reaction(PCR) combined with DNA sequencing technology was used to detect EBV nuclear antigen 1(EBNA1) gene sequences in the lymphoma tissues in Qingdao region, it was aimed to explore whether EBNA1 coding gene sequence variation has statistical significance in the EBV positive lymphoma tissues and to clarify whether the variation is regional or tumor specific.Methods: In situ hybridization(ISH) was used to detect EBV-encoded small RNA 1(EBER1) in six hundred and twenty-two lymphoma tissues from Qingdao city in Shandong province. EBER1 was used to select EBV positive tumor cases. Nested-PCR and DNA sequencing were applied to determine the gene polymorphisms of EBNA1 in EBV positive specimens. DNAStar software was used to contrast and analyze EBNA1 coding gene sequence detection results, the sequence variation types of EBNA1 gene were classified according to the signature changes at amino acid(AA) 487, definition of EBV type 1/type 2 in EBV positive lymphom was performed by using standard PCR assays according to the deletion mutantion of EBNA3 C gene sequence.Results: ①one hundred and ten of 622 cases of lymphomas was EBV positive lymphomas, positive rate was 17.68%. The positive rate of Hodgkin’s lymphoma(HL) was 33.33%(35/105), the positive rate of NK/T cell lymphoma was 78.67%(59/75), the positive rate of diffuse large B cell lymphoma(DLBCL) was 1.34%(5/372) and the positive rate of T cell lymphoma was 15.71%(11/70). ②Based on the signature changes at amino acid residue 487 and other common mutations EBNA1 can be divided into 5 kinds of subtypes, in this study four types were detected, including Variant(V)-val, Prototype(P)-thr, V-leu and P-ala, and the detection rate was 67.3%(74/110), 15.5%(17/110), 3.6%(4/110) and 10.9%(12/110) respectively.③The distribution of the four EBNA1 subtypes in the lymphoma tissues had no significant difference(P=0.383). Compared with gastric carcinoma(GC), nasopharyngeal carcinoma(NPC) and throat washings(TW) from healthy people, there is also no significant difference for lymphomas(P>0.05). There was no significant difference for the distribution of type 1, 2 and type 1+2 in four groups of lymphomas(P=0.089). Compared the lymphomas with NPC, EBVa GC and TWs respectively, there were also statistically significant difference(P=0.002, P=0.007, P=0.001). The distribution of EBNA1 subtypes(V-val, P-thr, V-leu, P-ala) in the three groups of lymphomas which were classified by EBV types(type1, type2, type1+2) has no significant difference(P=0.553).Conclusion: ①V-val, P-thr, V-Leu and P-ala four EBNA1 subtypes were detected in EBV positive lymphoma samples, and V-val is the most common subtype of lymphoma in the Qingdao area, because the V-val subtype is also the most common subtype in Asia, and the result is different from the relevant report in other parts, therefore it showed that V-val subtype can represent regional correlation. ② The distribution of EBNA1 subtypes in EBV positive lymphoma, NPC, EBVa GC tissues and TWs in healthy people, and among four kinds of EBV positive lymphoma in Qingdao area there was no significant difference, it showed the distribution of EBNA1 subtypes had no tumor specificity, and didn’t have the selection bias in development of lymphoma, which further support its regional correlation characteristics. ③ EBV positive lymphoma tissues were detected more EBV co-infections of Type1 and Type2, especially was evident in T cell lymphoma, which was different from NPCs, EBVa GC and TWs. It was speculated that Type2 EBV may be more susceptible to primary T cells, EBV co-infection of Type 1 and Type 2 may have synergistic effect. ④There is no correlation between EBV 1/2 type and EBNA1 subtypes in lymphoma tissues. |
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