Study On Mangrove Sediment Microbiome And Its Functioning Of Response And Removal Of BaP Contamination

Mangrove ecosystem is one of the most productive ecosystems in the world,playing a key role in biogeochemical cycles.Located in the boundary between the sea and the land,mangrove ecosystems exhibit unique environmental characteristics,leading to massive amounts of distinctive microorganisms.These microorganisms are essential to drive the element flows and filter the pollutants.Polycyclic aromatic hydrocarbons(PAHs)are typical organic pollutants consisted of more than one benzene ring.With the rapid development of economy,PAHs contamination and its remediation has drawn worldwide attention.Microbial degradation is one of the most important ways to remove PAHs in natural environments,and mangrove microbes display its promise for PAH remediation strategies.However,due to the physicochemical characteristics of high-molecular-weight PAHs(HMW-PAHs),their biodegradation is still a great challenge.In natural environments,PAHs are always degraded by the cooperation among different populations,and the cooperation will be stronger along with the increase of benzene rings the PAHs possess.Thus,revealing the PAHs microbial degradation at community level and the cooperation pattern among members would be an efficient way to break through the HMWPAHs bottleneck.Considering the advantages of mangrove microorganisms,the present study focused on the HMW-PAHs degradation issue,regarding the Yunxiao mangrove microbiome as study object.Appling 16S rRNA gene high-throughput sequencing and shotgun metagenomics to the microcosms exhibited based on Yunxiao mangrove sediments with Benzo[a]pyrene(BaP)as contaminant,trying to provide a clue to HMW-PAHs bioremediation from the dynamics of community diversity,structural succession,microbe-to-microbe interactions and function.The following shows the main results and conclusions of this study:1.The present study revealed the structure and function characteristics of mangrove microbiome.At the first place,the present study investigated the Yunxiao mangrove microbiome.Physicochemical assays characterized the Yunxiao mangrove sediments as carbon(C)-rich(the concentration of organic carbon was 21.79±1.35 g/kg),sulfur(S)-rich(the concentration of total sulfur was 4.27±1.09 g/kg),and nitrogen(N)-limited(the concentration of nitrite and ammonium were 15.24±5.60 μg/kg and 7.61±0.76 mg/kg)environment.Then phylogenetic analysis profiling a distinctive microbiome with an unexpected high frequency of Chloroflexi and Nitrospirae appeared to be an adaptive characteristic of microbial structure in S-rich habitat.Metagenome sequencing analysis revealed the metabolic potential of microbiota.At KEGG pathway level,the metabolic pathways of N and S cycling at the community-level were routed through ammonification and dissimilatory nitrate reduction to ammonium for N conservation in this Nlimited habitat,and dissimilatory sulfate reduction.These reducing routed metabolic pathways could significantly contributed to the high productivity of mangrove ecosystems via overcoming the nutrient limitation.At KEGG level 2,the genes related to hydrocarbon metabolism were most dominant(4.32%).Within,pyruvate metabolism,TCA cycle and glycolysis were among most abundant categories.The genes related to xenobiotics biodegradation and metabolism also showed high frequencies(0.96%).The dominance of these two function categories indicated the great ability of decomposing organic carbon.Thus,metagenomics revealed the ability of organic carbon decomposition of mangrove microbiome,indicating its potential in organic carbon biodegradation and purification.2.The present study revealed the dynamics of microbial diversity and structure in response to BaP contamination and(or)nitrate stimulation.According to the environmental characteristics of Yunxiao mangrove sediments,the present study exhibited BaP-contaminated and nitrate-stimulated microcosms(BN)and nitratestimulated microcosms(N)besides BaP-contaminated(B)and non-contaminated microcosms(CK),in order to explore the microbial response to HMW-PAHs without nitrogen limitation.At the first place,mangrove microbial community showed great BaP degradation and nitrate utilization ability,with 58.90%and 63.25%BaP degradation ratio in BN and B,and 91.42%and 91.74%nitrate degradation ratio in BN and N,but the nitrate stimulation did not show effect on its BaP degradation efficiency.Based on 130 16S rRNA amplicons datasets obtained using highthroughput sequencing,the study traced the diversity and microbial succession in response to BaP contamination and nitrate stimulation.Bioinformatic analysis indicated both BaP and nitrate addition significantly altered the microbial community structures,along with the structural succession at temporal scale.But BaP and nitrate exhibited different effects on the mangrove microbial community:when nitrate significantly decreased the microbial diversity and the community tended to form a simpler cluster with phylogenetically closed members with similar niches,BaP seemed to trigger the increase of microbial diversity and niche differentiation,and stimulate more phylogenetically distant members with different functions,leading to a more complex and multi-orient succession.We also figured out the potential responding bacteria based on their changes of relative abundance,including OP3,WS6,Lentisphaerae,Microgenomates,Parcubacteria和 Verrucomicrobia.3.The present study demonstrated the dynamics of microbe-to-microbe interactions in response to BaP contamination and(or)nitrate stimulation,and identified the keystone taxa and keystone guilds.Keystone taxa are drivers of microbiome structure and functioning.Thus,identifying the keystone taxa under certain environmental conditions based on microbe-to-microbe interactions,and describing their functions and the interactional changes they cause,is a promising way to understand microbial response to BaP at community level.Applying 130 16S rRNA gene datasets to molecular ecological networks pipeline,4 time-series co-occurrence networks were obtained,and the microbe-to-microbe interactions were revealed.The results showed that,the microbial community turned into a centralized interaction pattern with better information transfer efficiency responding to nitrate stimulation.The members and keystone taxa involved in nitrate-stimulated network showed monotonous pattern,with Sulfurimonas as dominant keystone taxa.The comparative genomics described the great ability in nitrate acquisition without the potential in aromatic pollutants degradation,not only answering the question why nitrate did not enhance the BaP degradation,but also providing a clue to the assembly pattern of microbial community when it was out of nitrogen limitation.Responding to BaP addition,the community resisted against BaP contamination through intensifying the mutualistic interactions among a group of Proteobacteria,Acidobacteria and Bacteroidetes populations,in accordance with our enrichment systems.They clustered into a highly-related module,in which the group comprising of Novosphingobium,Robiginitalea and Marinicella was identified as a keystone guild which we highly and positively connected with neighbors,and we proposed this group as BaP keystone-accessory guild.Predicted metagenomic results further suggested their cooperation pattern and highlighted the potential significance of the keystone-accessory guild in BaP degrading processes,mainly through biofilm formation,aromatic degradation and compounds transport.4.The present study reconstructed the processes of BaP microbial responses and degradation at community level.The present study further explored the functional response of mangrove microbial community to BaP contamination and its BaP degrading potential based on shotgun metagenomics.5 metagenomic datasets were obtained(one from in-situ sample,4 from 4 treated microcosms on the 60th day).The dominant population analysis showed the enrichment of genes related to carbon metabolism,PAHs degradation and oxidation resistance in BaP-contaminated samples,describing the potential resistant and degrading processes of microbial community to BaP:along with the resistance to oxidative stress by peroxidases and cytochrome P450,the BaP degradation converted complex BaP into simpler hydrocarbon compounds and flowed into TCA cycle and fatty acid metabolisms,when the products could be utilized by microorganisms.Aiming at evaluating the degrading potential of microbial community to BaP,this study investigated the diversity and metabolic features of ring-hydroxylation dioxygenases(RHOs),and the results detected 8 types of RHOs,and one of them showed ability of BaP degradation.In combination of binning approach,we further located these RHOs in SG8-30 from Gammaproteobacteria,UBA8639 from Nitropsirota and FEB-10 from Acidobacteria,indicating their key roles in BaP degradation.We also located the coding genes of other key PAH degrading enzymes including pht,ligA/B,pcaG/H and nahG in MAGs,and reconstructed the BaP degrading pathway.On the other hand,we focused on Sulfurovum as a representative functioned as auxiliary member in BaP degradation based on network analysis.The present study binned their genomes and applied them to comparative genomics with other representative genomes from public database,uncovering their metabolic advantages under BaP contamination and their contribution to community-level BaP degradation through multiple strategies such as oxidation resistance and amino acid supply.Thus,we located the geens related to quorum sensing,biofilm formation,thiol compounds exporters and amino acid exporters in MAGs to identify the auxiliary populations.Ultimately,collecting all these clues together,the present study reconstructed the processes of BaP microbial responses and degradation at community level in Yunxiao mangrove sediments.Overall,the present study displayed a comprehensive view of mangrove microbiome structure and functioning,and its potential in HMW-PAHs degradation,providing theoretical basis for the exhibition of cost-efficient bioremediation strategies.