变异链球菌囊泡密度感应信号分子-2的鉴定及囊泡对生物膜形成的影响OA

Objective To investigate whether membrane vesicles(MVs)of Streptococcus mutans(S.mutans)con-tain autoinducer-2(AI-2)and to preliminarily explore the effects of these MVs on the growth and biofilm formation of S.mutans.Methods MVs were isolated from the S.mutans UA159 strain using differential centrifugation.The isolated MVs were characterized by nanoparticle tracking analysis for particle size and concentration and observed by transmis-sion electron microscopy.The presence of AI-2 was identified using the Vibrio harveyi BB170 bioluminescence assay:the BB170 diluent was supplemented with AB medium(control group),MV extract(MVs group),pre-ultrafiltration su-pernatant(Sup group),or post-ultrafiltration supernatant(Sup-af group).The effects of MVs on growth and biofilm forma-tion were assessed using the S.mutans UA159 strain or a luxS deletion mutant as the control group,compared with ex-perimental groups stimulated with gradient concentrations of MVs(MVs-2.0E+7,MVs-2.0E+8,and MVs-2.0E+9 groups).Growth curves,MTT assay,and colony-forming unit(CFU)counts were used to determine changes in growth ca-pacity.Biofilm formation was evaluated using crystal violet staining,confocal laser scanning microscopy,and the an-throne method for polysaccharide quantification.Results Enriched S.mutans MVs were successfully obtained,with an average particle size of approximately 94.19 nm and a concentration of 1.87E+11 particles/mL.The bioluminescence assay showed that the luminescence intensity of the Sup group was higher than that of the Sup-af group,and the MVs group exhibited higher intensity than the control group.Assessments via growth curves,MTT assay,and CFU counts in-dicated no significant differences in the growth capacity of the various S.mutans strains after treatment with different concentrations of MVs.Crystal violet staining quantification and confocal laser scanning microscopy observations re-vealed that high-concentration MV treatment(2.0E+9 particles/mL group)resulted in lower biofilm mass compared to the control.The anthrone method showed that the production of both water-soluble and water-insoluble polysaccharides was significantly lower in the high-concentration MV group than in the control.Conclusion S.mutans MVs contain the quorum sensing signal molecule AI-2.These MVs do not significantly affect the growth of S.mutans,but they can regulate biofilm formation and exhibit an inhibitory effect at high concentrations.