Role of coaggregation among periodontopathic bacteria in formation of periodontopathic biofilm

目的：由革兰氏阴性杆状体和螺旋体引起的牙周炎。牙龈卟啉单胞菌是慢性牙周病的主要病原体，该微生物具有多种毒力因子，包括菌毛、脂多糖和蛋白酶，常与单宁菌和密螺旋体共分离。这三种微生物的组合被称为“红色复合物”，并且被认为在慢性牙周炎的进展中起重要作用。在这个项目中，我们研究了这些牙周病细菌之间的共聚集和协同作用，以阐明牙周病生物膜形成的潜在机制。材料与方法：采用共聚集缓冲液中660 nm吸光度的方法，对牙龈p.s ingivalis ATCC 33277与齿牙p.s denticola ATCC 35405的共聚集进行评价。为了明确牙龈卟啉卟啉配体参与共聚集，构建了缺乏牙龈蛋白酶和菌毛的敲除突变体。采用含0.4 m过滤单元的双室共培养系统，对生物膜的形成进行了评价。结果与讨论：在Mfa1缺陷突变体、kgp、RgpA和RgpB缺陷突变体和kgp、RgpA和HagA缺陷突变体中，牙龈假单胞菌和齿齿假单胞菌之间的共聚集反应减弱。这些结果表明牙龈痛和Mfa1参与了共聚集反应。Kgp、RgpA和RgpB缺陷菌株和Kgp、RgpA和HagA缺陷菌株的活性显著降低。此外，在Mfa1 FimA突变体中观察到共聚集活性。这表明牙龈痛和HagA的血凝素/黏附结构域在共聚集反应中起关键作用。使用我们的双室系统，与单独培养相比，与牙龈假单胞杆菌共同培养的核梭杆菌的生物膜形成增加了大约四倍。综上所述，这些结果表明牙龈假单胞菌与其他牙周病病原体的共聚集反应不仅参与龈沟生物膜的定植，还参与牙周病生物膜的形成。少

Objects : Periodontitis is caused by Gram-negative rods and spirochetes. Porphyromonas gingivalis is a major pathogen of chronic periodontal disease, This microorganism has a number of virulence factors, including fimbriae, lipopolysaccharides and proteases, and is often co-isolated with Tannerella forsythensis and Treponema denticola. The combination of these three microorganisms is known as the "red complex", and is thought to play an important role in the progression of chronic periodontitis. In this project, we investigated coaggregation and synergistic effects among these periodontopathic bacteria to clarify the underlying mechanisms of periodontopathic biofilm formation.Materials and Methods : Coaggregation between P.gingivalis ATCC 33277 and T.denticola ATCC 35405 was evaluated by absorbance at 660 nm in coaggregation buffer. To clarify which P.gingivalis ligands were involved in coaggregation, knockout mutants lacking gingipains and fimbriae were constructed. Synergistic effect … More s on biofilm formation were evaluated by using a two-compartment co-culture system incorporating a 0.4 m filter unit.Results and Discussion : Coaggregation reaction between P.gingivalis and T.denticola was decreased in the Mfa1 deficient mutant, kgp, RgpA and RgpB deficient mutant and Kgp, RgpA and HagA deficient mutant. These results indicated that gingipains and Mfa1 were involved in the coaggregation reaction. In particular, activity decreased significantly in the Kgp, RgpA and RgpB deficient strain and Kgp, RgpA and HagA deficient strain. In addition, coaggregation activity was observed in the Mfa1 FimA mutant. This suggests that the hemagglutinin/ahesion domains of gingipains and HagA play a key role in the coaggregation reaction. Using our two-compartment system, Fusobacterium nucleatum showed an approximately four-fold increase in biofilm formation in co-culture with P.gingivalis compared with that obtained with single culture alone. Taken together, these results suggest that the coaggregation reaction of P.gingivalis with other periodontopathogens is involved not only in the colonization of gingival crevicular biofilms, but also in the formation of periodontopathic biofilms. Less