The Taxonomic Compositions Of Cable Bacteria And Their Functions For Enhancing Pyrene Removal In Typical Polluted River Sediments In The Pearl River Delta

Cable bacteria are a type of multicellular filamentous bacteria which have not pure culture yet,belonging to the family Desulfobulbaceae within the Deltaproteobacteria.They can mediate the long-distance electron transport（LDET）over centimeter-scale distances to generate current to couple sulfide oxidation occurring in suboxic and anoxic sediments with oxygen reduction occurring in oxic sediments.An increasing number of studies have documented that the LDET mediated by cable bacteria play important ecological functions in the biogeochemical cycles in various natural aquatic environments.However,there is little research on cable bacteria in the typical polluted rivers caused by rapid industrialization and urbanization.This study investigated the taxonomic diversity of cable bacteria in the typical polluted river sediments in the Pearl River Delta by building the fluorescent quantitative PCR（q PCR）method targeted dsr B gene for the genus Ca.Electronema and combining with microelectrode analysis,fluorescence in situ hybridization（FISH）and scanning electron microscopy（SEM）,16S r RNA gene full-length cloning library,amplicon and metagenomic sequencing,and other methods.Through the microcosmic simulation experiment by blocking the growth of the cable bacteria with filter membranes,this study also analyzed the functions of cable bacteria for removing organic pollutant pyrene in anaerobic zone of sediments.The main results and conclusions are summarized as follows:（1）This study confirmed the existence of diverse cable bacteria in the typical polluted river sediments for the first time,expanding our knowledge on the habitat and diversity of cable bacteria.The geochemical fingerprint of the LDET mediated by cable bacteria appeared in typical polluted river sediments incubated with saturated oxygenated water for 15days,FISH and SEM also confirmed the filamentous and unique outer membrane structures of the cable bacteria;Subsequently,50 full-length 16S r RNA gene sequences related to cable bacteria were recovered from the clone libraries of the cable bacteria enrichment cultures,and their homology with Ca.Electronema palustris F3（KP728463）ranging from 92.54 to 97.57%.Ten medium and high-quality metagenome-assembled genomes（MAGs）belonging to genus Ca.Electronema were obtained,and they were classified into 5 novel Ca.Electronema species by average nucleotide identity（ANI）analysis;（2）A novel fluorescent quantitative PCR targeted with dsr B gene was developed for qualification and quantification of cable bacteria（Ca.Electronema）to explore the distribution of Ca.Electronema in river sediments in Canton.This method could be sensitive,convenient and rapid to directly detect and quantify the abundance of Ca.Electronema in environmental samples.Based on this method,this study found that Ca.Electronema is widely present in river sediments in Canton and their in-situ abundance was between 42.12-155.29 copys/ng DNA;One of the key factors affecting their in-situ abundance was the concentration of dissolved oxygen in the habitat environment;（3）The Ca.Electronema might have a mixed nutrition lifestyle.The presence or absence of Nha P type Na⁺/H⁺cation-proton antiporter was one of the important reasons for the differences in the habitat of cable bacteria.The reconstructions of metabolic potential for 10 Ca.Electronema MAGs obtained in this study showed that the Ca.Electronema not only had the ability to assimilate organic carbon acetate,but also had the ability to fix inorganic CO₂and CO through the Wood-Ljungdahl（WL）pathway,which indicated that Ca.Electronema might have a mixed nutrition lifestyle.The Ca.Electronema possessed genes related to sulfide oxidation or reduction,nitrate and nitrite reduction,nitrogen fixation and arsenic reduction.In addition,Ca.Electronema might be able to use LDET coupled acetate respiration to obtain carbon sources and energy.Comparison of the core genomes of cable bacterial two candidate genera showed that Ca.Electrothrix had Nha P-type Na⁺/H⁺cation-proton antiporters,while Ca.Electronema did not have genes or proteins with a similar function,which resulted in a low tolerance to salt stress;（4）The enrichment of Ca.Electronema promoted the simultaneous removal of AVS and pyrene in the anaerobic zone of sediments.By experimenting with a microcosmic simulation to block the activity of cable bacteria,this study found that the Ca.Electronema could protect the microorganisms with the functions of organic matter degradation by mediating LDET to remove H₂S,thus improving the bioavailability of pyrene.The synergistic interaction of cable bacteria,sulfate reducing bacteria and pyrene degrading microorganisms promoted the removal of AVS and the degradation of organic pollutant pyrene.In summary,this study confirmed that diverse cable bacteria exist in the typical polluted river sediments and their promotion for the synergistic removal of the AVS and organic pollutant pyrene in the anaerobic zone of sediments through the novel fluorescent quantitative PCR targeted with dsr B and some independent culture technologies,and bio-electrochemistry methods.In addition,this study also has proposed the physiological metabolic potential and environmental adaptability mechanisms of cable bacteria based on metagenomics and comparative genomics,including their mixed nutrition lifestyle and salt/freshwater habitat differences determined by the Nha P-type Na⁺/H⁺antiporter.These findings expanded our scientific understanding of the taxonomy and functions of cable bacteria,and provided a new idea for the development of in situ bioremediation technology in river sediments.