Escherichia coli and its lipopolysaccharide modulate in vitro Candida biofilm formation

Abstract

Demystification of microbial behaviour in mixed biofilms could have a major impact on our understanding of infectious diseases. The objectives of this study were to evaluatein vitrothe interactions of six differentCandidaspecies and a Gram-negative coliform,Escherichia coli, in dual-species biofilms, and to assess the effect ofE. coliLPS onCandidabiofilm formation. A single isolate ofE. coliATCC 25922 and six different species ofCandida,Candida albicansATCC 90028,Candida glabrataATCC 90030,Candida kruseiATCC 6258,Candida tropicalisATCC 13803,Candida parapsilosisATCC 22019 andCandida dubliniensisMYA-646, were studied using a standard biofilm assay. EachCandidaspecies was co-cultured withE. colion a polystyrene surface and biofilm formation was quantified by a c.f.u. assay. The biofilm was then analysed by Live/Dead staining and fluorescence microscopy (confocal laser-scanning microscopy, CLSM), whilst scanning electron microscopy (SEM) was employed to visualize the biofilm architecture. The effect ofE. coliLPS onCandidabiofilm cell activity at defined time intervals was assessed with an XTT reduction assay. A significant quantitative reduction in c.f.u. counts ofC. tropicalis(after 90 min),C. parapsilosis(after 90 min and 24 h),C. krusei(after 24 h) andC. dubliniensis(after 24 and 48 h) was noted on incubation withE. coliin comparison with their monospecies biofilm counterparts (P<0.05). On the other hand, a simultaneous and significant reduction inE. colicell numbers occurred on co-culture withC. albicans(after 90 min), and an elevation ofE. colicell numbers followed co-culture withC. tropicalis(after 24 h) andC. dubliniensis(after 24 h and 48 h) (P<0.05). All quantitative findings were confirmed by SEM and CLSM analyses. By SEM observation, dual-species biofilms demonstrated scanty architecture with reduced visible cell counts at all stages of biofilm development, despite profuse growth and dense colonization in their single-species counterparts. Significantly elevated metabolic activity, as assessed by XTT readings, was observed inE. coliLPS-treatedC. tropicalisandC. parapsilosisbiofilms (after 48 h), whilst this had the opposite effect forC. dubliniensis(after 24 h) (P<0.05). These data indicate thatE. coliandCandidaspecies in a mixed-species environment mutually modulate biofilm development, both quantitatively and qualitatively, and thatE. coliLPS appears to be a key component in mediating these outcomes.