Single And Combined Effects Of Amino Polystyrene And Perfluorooctane Sulfonate On Hydrogen-producing Thermophilic Bacteria And The Interaction Mechanisms

The widespread use of plastic products and the ecological risks caused by their improper handling have attracted more and more attention.Moreover,these plastics are decomposed into micro(nano)-plastics(?<5mm)by the action of light,microbiological or mechanical forces after entering the environment,which will induce serious ecological risks and health hazards.A large amount of sewage is mixed with microplastics into the sewage treatment plant.After sewage treatment process,more than 98.3%of the microplastics can be removed,resulting in the sludge of the sewage treatment plant becoming the pool and risk source with the highest microplastic content in the terrestrial environment.The concentration of microplastics in the sludge of sewage plants in China is 1.0-56.4 particles/g(dry weight of sludge).The anaerobic digestion of sludge to produce hydrogen is one of the important ways for sludge resource utilization.However,research on the impact of the presence of microplastics.on sludge resource utilization and energy utilization is still scarce.Exploring the toxic effect and mechanism of micro(nano)plastics on microorganisms in the anaerobic hydrogen production system has positive significance for sludge recycling.On the other hand,because of the small volume and large specific surface area,micro(nano)plastics inevitably adsorb hydrophobic organic micro-pollutants(POPs).PFOS is a typical persistent POPs and generally present in wastewater.Due to PFOS monomers have greater toxicity and strong adsorption affinity with microplastics(300-1800ng/g).They will have a joint toxic effect on digestive hydrogen-producing microorganisms.And PFOS is an additive for plastic products,which is detected in micro(nano)plastics in different water bodies.Therefore,in this study,the thermophilic hydrogen-producing bacteria in sludge will be used as the exposure object and glucose will be used as the substrate.Nano-plastic polystyrene with amino groups(NPS:70 nm,0.2 mg/L)and perfluorooctanesulfonate(PFOS:0.1,1 and 5 mg/L)was used as the target pollutant.To analyze the metabolic toxicity and action mechanism of NPS and PFOS alone and combined exposure on thermophilic hydrogen-producing bacteria.The specific research results are as follows:(1)The thermophilic hydrogen-producing bacteria were exposed to different concentrations of NPS and PFOS at T=55 ? and pH 5.7 for 7 days.The cumulative amount of hydrogen and the amount of VFAs were used as indicators of its metabolic toxicity to thermophilic bacteria.The results showed that a single NPS had a significant interference with the metabolism of the thermophilic bacteria.The hydrogen production decreased by 53.9%compared with the blank group,and the highest hydrogen production caused by a single PFOS decreased by 19.1%compared with the blank group.However,the compound of NPS+PFOS produced an antagonistic effect,resulting in a decrease in hydrogen production by 27.4%-31.6%compared with the blank group.Further analysis by 16S rRNA showed that the number of OTUs in the sludge particles was 37 OTUs less than the blank(94 OTUs)when NPS alone exposed.However-,when the NPS and PFOS added at the same time the quantity of OTU was 17 OTUs less than the blank.Which indicates that the toxicity of single NPS is stronger than that of a mixed pollutant of NPS+PFOS.In other words,the combined toxicity type of NPS and PFOS against thermophilic hydrogen-producing bacteria is antagonistic.Nevertheless,neither the single action nor the combined action had same end products and they didn't change the type of hydrogen production.(2)The extracellular polymers contain various enzymes,nucleic acids and lipids,and are an important barrier for microorganisms to resist external interference and toxicity(e.g.nanoparticles).The changes in composition and structure have an important influence on the physiological functions of microorganisms.Therefore,the composition,surface characteristics and key hydrolase enzyme activities of three extracellular polymers were analyzed.According to the data,we can know that NPS alone and NPS+PFOS complex not only changed the composition of extracellular polymers(EPSs)and changed the ?-bond structure of proteins,but also reduced the surface charge and hydrolase activity in EPSs.Scanning electron microscopy also showed that the structure of the sludge is more loose than the blank.There are a large number of nanoparticles are attached to the surface of the sludge.The adsorption of NPS on the surface of the granular sludge will hinder the exchange of materials and energy inside and outside the cell,thereby reducing the metabolism of bacteria.(3)In order to clarify the toxic mechanism of NPS and PFOS on thermophilic hydrogen-producing bacteria,this study uses the fluorescent probe 4',6-diamidino-2-phenylindole(DAPI)and dichlorodihydrofluorescein diacetate(H2DCF-DA)respectively to detect the effects of NPS alone and complex NPS+PFOS exposure on the thermophilic cell permeability and intracellular reactive oxygen species levels.The results show that compared with NPS exposure alone,membrane permeable dyes are less permeable to thermophilic bacteria exposed to NPS+PFOS and thermophilic bacteria produce less reactive oxygen species,with a maximum of 4.32%less.Moreover,the active oxygen produced by thermophilic bacteria was 0.77-1.71 times less than NPS alone exposure,but it was still significantly higher than the control group,which was 1.45-1.86 times of the blank group.That is,the addition of PFOS reduced the toxic effects of NPS.In summary,the mechanism of toxicity of NPS alone and composite NPS+PFOS to thermophilic hydrogen-producing bacteria is mainly to induce increased permeability of cell membranes and oxidative stress.