Ecological Restoration Mechanism Of Salt-tolerant Submerged Macrophyte Myriophyllum Spicatum In Low Salinity Sea Area

Eutrophication of freshwater and coastal marine ecosystems has become a global problem,which poses a great threat to the aquatic ecosystem and human health.Reducing point source pollution from sewage treatment plants and non-point source pollution from agricultural production has been proved an effective measure to prevent the deterioration of eutrophication.While,in-situ remediation is an important way to improve the quality of polluted water.At present,the restoration and construction of submerged plant communities were used for freshwater ecosystem restoration.Large-scale algae cultivation and harvesting were used in marine system.However,few effective ecological restoration measures for low salinity estuary water with relatively serious eutrophication were reported.In this paper,the mechanism of ecological restoration by cultivation salt-tolerant submerged plant Myriophyllum spicatum in a low salinity sea area was studied.It was observed that M.spicatum cultivated by the suspension method showed fast vertical growth during the initial periods.A great number of new shoots and some adventitious roots were formed at this stage.On the other hand,M.spicatum samples that cultivated at the bottom of the water body were found to grow slowly during the initial phases due to the low light intensity.As water clarity gradually increased and exceeded over 1 m?in September?,the bottom-cultivated M.spicatum showed fast growth and were found to sprawl around with the growth rate similar to that of the suspension-cultivated M.spicatum.After about one year of salt adaptation,M.spicatum germinated and grew naturally in 2017,and gradually formed the dominant population.The biomass of M.spicatum increased significantly from August 2017?p<0.05?,reaching the maximum biomass of 4029.70 g·m^-22 in October.The specific growth rate reached a maximum of 4.6%in September.Through two years'monitoring,it was found that M.spicatum could successfully complete its life cycle in the study area of Hangzhou Bay with water temperature 3.9-33.4?and salinity4.9-12.7.The reproductive mode of M.spicatum was sexual reproduction and vegetative reproduction,mainly by vegetative reproduction.After ecological restoration by cultivation M.spicatum,water transparency increased rapidly from 0.5-0.7m to 2.0-2.6m?average 2.3m?,which was significantly higher than that of non-restoration area?p<0.05,n=15?.The mean value of dissolved oxygen in the restored area was 9.3 mg/L,which was significantly higher than that in the non-restored area?p<0.05,n=15?,and the mean value of pH was 9.2,which was significantly higher than that in the non-restored area?p<0.05,n=13?.After ecological restoration,the average removal efficiencies of two years ecological restoration were 15.2%,31.0%,25.1%and 29.5%,respectively.The removal efficiencies of total nitrogen and total phosphorus were 13.0%and 45.2%respectively.The removal rates of NH₄⁺-N,NO₃^--N and NO₂^--N in the sediment were 51.9%,32.3%and 53.6%respectively,and the increase rate of active phosphorus was 13.2%.The annual average contents of C,N and P in M.spicatum were 38.73%,3.19%and0.34%respectively.The annual total biomass increment of M.spicatum was 427.6g·m^-2,and the annual absorption of C,N and P were 170.4 g·m^-2,14.7 g·m^-22 and 1.49g·m^-2,respectively.A total of 64 species of phytoplankton were observed in the study area in 2016,including 51 species in the restoration area and 48 species in the non-restoration area.Bacillariophyta was the main phyla,followed by chlorophyta,and 54.7%of the phytoplankton species was overlapped between the two areas.A total of 103 species of phytoplankton were detected in the study area in 2017.There were 54phytoplankton species in the restoration area,and diatomaceae was the dominant group.79 species were found in non-restoration area,and diatomaceae,chlorophyta,gymnophyta and Cyanophyta were the main groups.The phytoplankton species overlapped between the two areas accounted for 29.1%of the total species.The dominant species in the restoration area were Melosira,Chlorella,Cyclotella,Cocconeis,Navicula and Trachelomonas.The dominant species in the non-restoration area are Microcystis,Chlorella,Cyclotella,and Trachelomonas.A single dominant species of Microcystis appeared in the non-remediated areas in August and September2016,with dominance of 0.99 and 0.98,respectively.The average cell density of phytoplankton in the restored area in 2016 and 2017 was 252.17×10⁴ cells L^-11 and1.3×10⁴ cells L^-1,respectively,while that in the non-restored area was 1280.05×10⁴cells L^-11 and 12.3×10⁴ cells L^-1,respectively.Shannon diversity index and Pielou evenness index in the restoration area were higher than those in the non-restoration area.In 2016,NH₄⁺-N and silicate were the main environmental factors affecting phytoplankton abundance in the restored and non-restored areas,respectively.In 2017,silicate and pH were the main environmental factors affecting phytoplankton abundance in the restored and non-restored areas,respectively.A total of 42 phyla,700 genera and 2,080 OTUs of bacterioplankton were identified in the study area by high throughput sequencing.Among them,1,162 and1,523 OTU were identified in the restoration area and non-restoration area,respectively.29.1%of the bacterioplankton species was overlapped between the two areas.The main groups of bacterioplankton in the restored area were Proteobacteria,Bacteroides,Cyanobacteria and Actinomycetes,with abundances of 30.2%,24.7%,23.5%and 9.6%,respectively.Proteobacteria,Cyanobacteria,Actinomycetes,and Bacteroides were the main groups in the non-restored area,with abundances of 34.9%,32.6%,15.8%and 8.6%,respectively.Bacterioplankton showed significant seasonal differences in both regions.Among the bacteria with fifteen most abundant genera,the abundances of CL500-29,hgcI,SAR11 and OM43 were significantly lower in the restored area than in the non-restored area,and Bacteroidetes-norank,NS3a,Fluviicola,Rhodobacteraceae-unclassified,MWH-UniP1and Candidatus Aquiluna were significantly higher in the restored area than in the non-restored area.NO₃^--N and PO₄^3--P were the main environmental factors affecting the seasonal variation of bacterioplankton abundance in the restored and non-restored areas,respectively.A total of 7923 OTUs belonging to 996 genera and 59 phyla of bacteria in sediment were identified in the study area.Among them,5717 OTUs were found in the restoration area and 6744 OTUs were found in the non-restoration area.57.3%of the OTUs was overlapped between the two areas.The bacterial richness and diversity of sediments in the restored area were significantly lower than those in the non-restored area?p<0.05?.The main bacterial groups in the restored sediments were Proteobacteria?33.7%?,Bacteroides?17.4%?,Planctomycetes?11.4%?and Cyanobacteria?10.9%?,and Proteobacteria?40.0%?,Bacteroides?9.8%?,Chloroflexi?9.4%?and Planctomycetes?8.8%?.Among the bacteria with fifteen most abundant genera,13 genera were significant differences between the restored and non-restored areas.NO₂^--N,NH₄^--N and TN were the main environmental factors affecting the seasonal variation of bacterial abundance in the sediments of the restoration area.NH₄^--N,TP and PO₄^3--P were the main environmental factors affecting the seasonal variation of bacterial abundance in the sediments of the non-restoration area.A total of 436 OTUs belonging to 783 genera and 47 phyla of epiphytic bacteria were identified in the restoration area in four seasons.Proteobacteria was the dominant group in the four seasons,followed by bacteroides.Alphaproteobacteria was dominant in summer,autumn and winter,and Betaproteobacteria was dominant in spring.The diversity of epiphytic bacteria was the lowest in spring.Using metagenomic technology,a total of 14714 species of attached microorganisms were identified,including four domains:Bacteria,Viruses,Eukaryota and Archaea,mainly bacteria,accounting for 93.77%-97.45%,with an average of95.79%.In October 2017,January 2018 and April 2018,the total COG function of microorganisms was 40017,and the total COG function of microorganisms overlapped among the three months was 27653?69.10%?.The COG function of microorganisms in adjacent seasons was closer.The functional annotations of COG in adherent microorganisms mainly involve E-amino acid transport and metabolism?7.65%?,L-replication,recombination and repair?6.57%?,M-cell membrane biogenesis?5.94%?,C-energy production and conversion?5.82%?,P-inorganic ion transport and metabolism?5.70%?,G-carbohydrate transport and metabolism?4.88%?and T-signal transduction pathway?4.87%?.There are more than 100 microorganisms involved in the nitrogen cycle,including bacteria,archaea and eukaryotes.The number and abundance of COG genes related to nitrogen fixation,denitrification and ammoniation were higher.In conclusion,salt-tolerant submerged macrophyte M.spicatum can adapt to the low salinity water body of Hangzhou Bay and form dominant populations.Through ecological restoration of M.spicatum,1.It can significantly improve water transparency,dissolved oxygen and pH,and reduce nutrients in water and sediment,and keep nutrient concentration in water at a low level for a long time,so as to keep water quality stable;2.It can significantly inhibit the growth of phytoplankton,especially microcystis,and change phytoplankton community structure,and improve the diversity and evenness of phytoplankton;3.It can change the community structure of bacterioplankton and sediment bacteria and reduce bacterial diversity.4.It can provide additional niche for attached microorganisms.These attached microorganisms are abundant,diverse and functional,and play a potential role in the biogeochemical cycle.