A Study On The Response Of Pseudomonas Putida KT2440 And Shewanella Oneidensis MR-1 Biofilm To Heavy Metals

Microorganisms often coexist in multiple species and are coated by extracellular polymers secreted by themselves in the natural environment.This form of survival is defined as biofilm.Interspecific interactions in biofilms are the key to community response to external stress,and are critical to community stability and biofilm spatial structure.Previous studies on biofilms have focused on water or medical fields,but research on soil environments has received little attention.The biggest difference between soil and aquatic environment is that the soil is water unsaturated in most cases.Soil moisture not only affects the migration and transformation of substances,but also directly changes the physiological characteristics of microorganisms.Therefore,it is necessary to consider the factor of moisture when studying the response of microorganisms to external pollutants such as heavy metals in the soil environment.In this study,two common strains-Pseudomonas putida KT2440 and Shewanella oneidensis MR-1 were selected as experimental strains to culture biofilm under water saturated and unsaturated conditions.Fluorescent protein labeling technology,real-time quantitative PCR(qPCR),atomic force microscopy(AFM),optical coherence tomography(OCT)and other in-situ observation techniques were used to characterize biofilm properties and explore the survival advantages of co-culture biofilm include microbes response to heavy metals at different concentrations.The main results are as follows:1)In blank or low heavy metal conditions,the biomass of P.putida and S.oneidensis co-culture community is only 56.4%-72.3% of the sum of the individual cultures under water saturated or unsaturated conditions.However,under the treatment of high concentration of Cu and Pb,the advantage of co-culture began to manifest.Co-culture biomass exceeded the sum of the individual cultures.In this community structure,P.putida was the dominant species,and its population ratio reached 97%.At high concentrations of heavy metals,P.putida cause a strong exploiting effect on S.oneidensis,so the presence of S.oneidensis promotes the growth of P.putida and increases the biomass of the entire community.In addition,it was found via fluorescence microscopy that P.putida mainly formed biofilm at the air-liquid interface,while S.oneidensis mainly formed biofilm at the solid-liquid interface in the co-culture system.2)Under water unsaturated conditions,the surface cell volume and surface area of the co-cultured unsaturated biofilm are smaller as 75% of that in the culture alone.Since S.oneidensis is located at the bottom of the unsaturated biofilm,most P.putida was located at the top so that the co-cultured biofilm roughness did not change significantly compared to the cultured P.putida alone biofilm.Under water saturated conditions,bacteria cannot grow under high concentrations of Cu and Pb when cultured alone,but co-culture can form a air-liquid interface biofilm with a thickness of 80-100 ?m,even cell assemble occur(130-150 mM).Co-culture not only allows bacteria to form biofilms under high concentrations of heavy metals but also secretes more extracellular extracellular polymers as TOC and TN is 1.1-2.68 times that of cultures alone.