AIM To assess whether Epstein-Barr virus (EBV) reactivation is triggered by persistent apical periodontitis-related microbes using in vitro and ex vivo methodologies. METHODOLOGY Surgically removed human periapical granulomas (n = 50) and healthy gingival tissues (n = 10) were analysed to determine the presence of EBV and seven persistent apical periodontitis-related microbes. In addition, real-time polymerase chain reaction was used to detect the mRNA expression of BZLF-1, an immediate-early gene of EBV. Expression of latent membrane protein (LMP)-1 and ZEBRA, an early lytic protein of EBV encoded by BZLF-1, was also examined using triple-colour immunofluorescence staining. n-Butyric acid produced by the microbes was quantified, and luciferase assays were performed in association with bacterial lysates. In addition, Daudi cells were cultured with bacterial lysates, and the expression levels of BZLF-1 mRNA and ZEBRA protein were determined. RESULTS EBV DNA and BZLF-1 mRNA were detected in 47 out of 50 periapical granulomas, but not in healthy gingival tissues. The EBV DNA copy number and the number of Fusobacterium nucleatum were significantly positively correlated with BZLF-1 expression in periapical granulomas. The number of Prevotella intermedia was slightly correlated with BZLF-1 expression; however, the other microbes were not. CD79a-positive B cells in periapical granulomas, but not those in healthy gingival tissues, expressed both LMP-1 and ZEBRA. n-Butyric acid production was the highest in F. nucleatum and the lowest in P. intermedia. Enterococcus faecalis, Candida albicans and the other tested microbes did not produce n-butyric acid. An F. nucleatum lysate exhibited significantly increased BZLF-1-luciferase activity in the same manner of commercial butyric acid, whereas P. intermedia did not. F. nucleatum also induced the expression of BZLF-1 mRNA and ZEBRA protein by Daudi cells, indicating that EBV reactivation was induced. CONCLUSION Among the persistent apical periodontitis-related bacteria that were tested, F. nucleatum most strongly reactivated latent EBV, whereas E. faecalis and C. albicans as well as the other microbes did not.

中文翻译：

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持久性根尖周牙病原体激活爱泼斯坦-巴尔病毒。

目的使用体外和离体方法评估持久性根尖性牙周炎相关微生物是否触发了爱泼斯坦-巴尔病毒（EBV）的再激活。方法学对手术切除的人根尖周肉芽肿（n = 50）和健康的牙龈组织（n = 10）进行分析，以确定是否存在EBV和7种持久性根尖周炎相关微生物。此外，实时聚合酶链反应用于检测EBV的早期基因BZLF-1的mRNA表达。还使用三色免疫荧光染色检查了潜在膜蛋白（LMP）-1和ZEBRA（由BZLF-1编码的EBV的早期裂解蛋白）的表达。定量由微生物产生的正丁酸，并与细菌裂解物一起进行荧光素酶测定。此外，用细菌裂解液培养Daudi细胞，并测定BZLF-1 mRNA和ZEBRA蛋白的表达水平。结果在50例根尖肉芽肿中，有47例检测到EBV DNA和BZLF-1 mRNA，但在健康的牙龈组织中未检出。根尖肉芽肿中EBV DNA的拷贝数和核梭状芽胞杆菌的数目与BZLF-1表达显着正相关。中间型普氏杆菌的数量与BZLF-1的表达略有相关。但是，其他微生物则没有。根尖肉芽肿中的CD79a阳性B细胞表达LMP-1和ZEBRA，而健康牙龈组织中不表达。正丁酸产量在核镰刀菌中最高，而在中间镰刀菌中最低。粪肠球菌，白色念珠菌和其他经过测试的微生物未产生正丁酸。F. 核酸裂解物以与商业丁酸相同的方式显示出BZLF-1-荧光素酶活性的显着增加，而中间假单胞菌则没有。F. nucleatum还通过Daudi细胞诱导BZLF-1 mRNA和ZEBRA蛋白的表达，表明诱导了EBV激活。结论在所测试的持续性根尖周炎相关细菌中，核仁镰刀菌能最强地激活潜在的EBV，而粪肠球菌和白色念珠菌以及其他微生物则没有。