Phosphorus forms in the sediment of seagrass meadows affected mainly by fungi rather than bacteria: a preliminary study based on ³¹P-NMR and high-throughput sequencing

Microorganisms play an important role in the circulation of phosphorus (P) in the sediment of coastal wetland ecosystems. In this study, solution ³¹P nuclear magnetic resonance (NMR) was used to determine different forms of P in the sediments of four different seagrass meadows and a bare tidal flat, while high-throughput 16S and ITS rRNA gene sequencing was used to determine the microbial community composition. The solution ³¹P-NMR spectra revealed six forms of the P compounds detected by the NaOH-EDTA extraction of sediments, where Ortho-P was the most dominant P compound, followed by Mono-P. The Po compounds were more varied in the seagrass meadow sediments and more abundant compared to the bare tidal flat. Bacterial communities in the sediments collected from E. acoroides and fungal communities in the bare tidal flat were relatively different from those at the other sites. The relative abundance of P-cycling-related fungi belonging to the phylum Ascomycota was 26.20% and was much higher than that of bacteria (only 0.29%) belonging to the class Bacilli. Mono-P was the major factor determining the distribution of P-cycling-related fungi and negatively correlated with the relative abundance of Aspergillus and Trichoderma. We believe that fungi can affect P forms in the sediment of seagrass meadows more than bacteria.

Acknowledgements
This work was supported by the Natural Science Foundation of Hainan Province (No. 418MS074), Young Talents’ Science and Technology Innovation Project of Hainan Association for Science and Technology (No. QCXM201811), the Key Project of Natural Science Foundation of China (No. 41730529) and the Regional Project of Natural Science Foundation of China (No. 41766004).

References

Baldwin, D.S. (2013). Organic phosphorus in the aquatic environment. Environ. Chem. 10(6): 439–454. DOI: 10.1071/EN13151.
Benitez-Nelson, C. R., O'Neill, L., Kolowith, L.C., Pellechia, P. & Thunell, R. (2004). Phosphonates and particulate organic phosphorus cycling in an anoxic marine basin. Limnol. Oceanogr. 49(5): 1593–1604. DOI: 10.4319/lo.2004.49.5.1593.
Bhattacharyya, P.N. & Jha, D.K. (2012). Plant growth-promoting rhizobacteria (PGPR): emergence in agriculture. World J. Microbiol. Biotechnol. 28(4): 1327–1350. DOI: 10.1007/s11274-011-0979-9.
Bramha, S.N., Mohanty, A.K., Padhi, R.K., Panigrahi, S.N. & Satpathy, K.K. (2014). Phosphorus speciation in the marine sediment of Kalpakkam coast, southeast coast of India. Environ. Monit. Assess. 186(10): 6003–6015. DOI: 10.1007/s10661-014-3836-0.
Brodersen, K.E., Koren, K., Mosshammer, M., Ralph, P.J., Kuhl, M. et al. (2017). Seagrass-mediated phosphorus and iron solubilization in tropical sediments. Environ. Sci. Technol. 51(24): 14155–14163. DOI: 10.1021/acs.est.7b03878.
Brodersen, K.E., Siboni, N., Nielsen, D.A., Pernice, M., Ralph, P.J. et al. (2018). Seagrass rhizosphere microenvironment alters plant-associated microbial community composition. Environ. Microbiol. 20(8): 2854–2864. DOI: 10.1111/1462-2920.14245.
Cade-Menun, B.J. (2005). Characterizing phosphorus in environmental and agricultural samples by P-31 nuclear magnetic resonance spectroscopy. Talanta. 66(2): 359–371. DOI: 10.1016/j.talanta.2004.12.024.
Cade-Menun, B.J. (2015). Improved peak identification in 31P-NMR spectra of environmental samples with a standardized method and peak library. Geoderma. 257–258: 102–114. DOI: 10.1016/j.geoderma.2014.12.016.
Cade-Menun, B.J. & Liu, C.W. (2014). Solution phosphorus-31 nuclear magnetic resonance spectroscopy of soils from 2005 to 2013: A review of sample preparation and experimental parameters. Soil Sci. Soc. Am. J. 78(1): 19–37. DOI: 10.2136/sssaj2013.05.0187dgs.
Cucio, C., Engelen, A.H., Costa, R. & Muyzer, G. (2016). Rhizosphere microbiomes of european seagrasses are selected by the plant, but are not species specific. Front. Microbiol. 7: 440. DOI: 10.3389/fmicb.2016.00440.
Dell'Anno, A. & Danovaro, R. (2005). Extracellular DNA plays a key role in deep-sea ecosystem functioning. Science 309(5744): 2179–2179. DOI: 10.1126/science.1117475.
Ding, S., Bai, X., Fan, C. & Zhang, L. (2010). Caution needed in pretreatment of sediments for refining phosphorus-31 nuclear magnetic resonance analysis: Results from a comprehensive assessment of pretreatment with ethylenediaminetetraacetic acid. J. Environ. Qual. 39(5): 1668–1678. DOI: 10.2134/jeq2009.0396.
Dipta, B., Bhardwaj, S., Kaushal, M., Kirti, S. & Sharma, R. (2019). Obliteration of phosphorus deficiency in plants by microbial interceded approach. Symbiosi. 78(2): 163–176. DOI: 10.1007/s13199-019-00600-y.
Duarte, C.M. & Chiscano, C.L. (1999). Seagrass biomass and production: A reassessment. Aquat. Bot. 65(1–4): 159–174. DOI: 10.1016/S0304-3770(99)00038-8.
Ettinger, C.L., Voerman, S.E., Lang, J.M., Stachowicz, J.J. & Eisen, J.A. (2017). Microbial communities in sediment from Zostera marina patches, but not the Z. marina leaf or root microbiomes, vary in relation to distance from patch edge. Peerj. 5: e3246. DOI: 10.7717/peerj.3246.
Fraser, M.W., Gleeson, D.B., Grierson, P.F., Laverock, B. & Kendrick, G.A. (2018). Metagenomic evidence of microbial community responsiveness to phosphorus and salinity gradients in seagrass sediments. Front. Microbiol. 9: 1703. DOI: 10.3389/fmicb.2018.01703.
Garcia-Martinez, M., Lopez-Lopez, A., Calleja, M.L., Marba, N. & Duarte, C.M. (2009). Bacterial community dynamics in a seagrass (Posidonia oceanica) meadow sediment. Estuaries Coasts 32(2): 276–286. DOI: 10.1007/s12237-008-9115-y.
Huang, X., Huang, L., Li, Y., Xu, Z., Fong, C.W. et al. (2006). Main seagrass beds and threats to their habitats in the coastal sea of South China. Chin. Sci. Bull. 51: 136–142. DOI: 10.1007/s11434-006-9136-5.
Hurtado-McCormick, V., Kahlke, T., Petrou, K., Jeffries, T., Ralph, P.J. et al. (2019). Regional and microenvironmental scale characterization of the Zostera muelleri seagrass microbiome. Front. Microbiol. 10: 1011. DOI: 10.3389/fmicb.2019.01011.
Jensen, H.S., McGlathery, K.J., Marino, R. & Howarth, R.W. (1998). Forms and availability of sediment phosphorus in carbonate sand of Bermuda seagrass beds. Limnol. Oceanogr. 43(5): 799–810. DOI: 10.4319/lo.1998.43.5.0799.
Jiang, Y.F., Ling, J., Wang, Y.S., Chen, B., Zhang, Y.Y. et al. (2015). Cultivation-dependent analysis of the microbial diversity associated with the seagrass meadows in Xincun Bay, South China Sea. Ecotoxicology. 24(7–8): 1540–1547. DOI: 10.1007/s10646-015-1519-4.
Khan, M.S., Zaidi, A. & Wani, P.A. (2007). Role of phosphate-solubilizing microorganisms in sustainable agriculture – A review. Agron. Sustainable Dev. 27(1): 29–43. DOI: 10.1051/agro:2006011.
Komatsu, T., Umezawa, Y., Nakakoka, M., Supanwanid, C. & Kanamoto, Z. (2004). Water flow and sediment in Enhalus acoroides and other seagrass beds in the Andaman Sea, off Khao Bae Na, Thailand. Coast. Mar. Sci. 29(1): 63–68. DOI: 10.15083/00040824.
Koukol, O., Novak, F. & Hrabal, R. (2008). Composition of the organic phosphorus fraction in basidiocarps of saprotrophic and mycorrhizal fungi. Soil Biol. Biochem. 40(9): 2464–2467. DOI: 10.1016/j.soilbio.2008.04.021.
Li, W., Joshi, S.R., Hou, G., Burdige, D.J., Sparks, D.L. et al. (2015). Characterizing phosphorus speciation of Chesapeake Bay sediments using chemical extraction, 31P-NMR, and X-ray absorption fine structure spectroscopy. Environ. Sci. Technol. 49(1): 203–211. DOI: 10.1021/es504648d.
Li, Y., Zhang, J., Zhang, J., Xu, W. & Mou, Z. (2019). Characteristics of inorganic phosphate-solubilizing bacteria from the sediments of a eutrophic lake. Int. J. Environ. Res. Public Health. 16(12): 2141. DOI: 10.3390/ijerph16122141.
Liu, J., Wang, H., Yang, H., Ma, Y. & Cai, O. (2009). Detection of phosphorus species in sediments of artificial landscape lakes in China by fractionation and phosphorus-31 nuclear magnetic resonance spectroscopy. Environ. Pollut. 157(1): 49–56. DOI: 10.1016/j.envpol.2008.07.031.
Liu H., Pan F., Han X., Song F., Zhang Z. et al. (2019). Response of soil fungal community structure to long-term continuous soybean cropping. Front. Microbiol. 9: 3316. DOI: 10.3389/fmicb.2018.03316.
McRoy, C.P., Nebert, M. & Barsdate, R.J. (1972). Phosphorus cycling in an eelgrass (Zostera marina L.) ecosystem. Limnol. Oceanogr. 17(1): 58–67. DOI: 10.4319/lo.1972.17.1.0058.
Mercl, F., Garcia-Sanchez, M., Kulhanek, M., Kosnar, Z., Szakova, J. et al. (2020). Improved phosphorus fertilization efficiency of wood ash by fungal strains Penicillium sp. PK112 and Trichoderma harzianum OMG08 on acidic soil. Appl. Soil Ecol. 147: 103360. DOI: 10.1016/j.apsoil.2019.09.010.
Nielsen, O.I., Koch, M.S. & Madden, C.J. (2007). Inorganic phosphorus uptake in a carbonate-dominated seagrass ecosystem. Estuaries Coasts. 30(5): 827–839. DOI: 10.1007/bf02841337.
Pagès, A., Welsh, D.T., Robertson, D., Panther, J.G., Schäfer, J. et al. (2012). Diurnal shifts in co-distributions of sulfide and iron(II) and profiles of phosphate and ammonium in the rhizosphere of Zostera capricorni. Estuar. Coast Shelf Sci. 115: 282–290. DOI: 10.1016/j.ecss.2012.09.011.
Paytan, A., Cade-Menun, B.J., McLaughlin, K. & Faul, K.L. (2003). Selective phosphorus regeneration of sinking marine particles: Evidence from 31P-NMR. Mar. Chem. 82(1–2): 55–70. DOI: 10.1016/s0304-4203(03)00052-5.
Prasad, M.B.K. & Ramanathan, A.L. (2010). Characterization of phosphorus fractions in the sediments of a tropical intertidal mangrove ecosystem. Wetlands Ecol. Manage. 18(2): 165–175. DOI: 10.1007/s11273-009-9157-3.
Prüter, J., Leipe, T., Michalik, D., Klysubun, W. & Leinweber, P. (2019). Phosphorus speciation in sediments from the Baltic Sea, evaluated by a multi-method approach. J. Soils Sediments 20(3): 1676–1691. DOI: 10.1007/s11368-019-02518-w.
Reitzel, K., Ahlgren, J., Gogoll, A., Jensen, H.S. & Rydin, E. (2006). Characterization of phosphorus in sequential extracts from lake sediments using 31P-nuclear magnetic resonance spectroscopy. Can. J. Fish. Aquat. Sci. 63(8): 1686–1699. DOI: 10.1139/f06-070.
Richardson, A.E., Lynch, J.P., Ryan, P.R., Delhaize, E., Smith, F.A. et al. (2011). Plant and microbial strategies to improve the phosphorus efficiency of agriculture. Plant Soil. 349(1–2): 121–156. DOI: 10.1007/s11104-011-0950-4.
Sannigrahi, P. & Ingall, E. (2005). Polyphosphates as a source of enhanced P fluxes in marine sediments overlain by anoxic waters: Evidence from 31P-NMR. Geochem. Trans. 6(3): 52–59. DOI: 10.1063/1.1946447.
Schneider, K.D., Cade-Menun, B.J., Lynch, D.H. & Voroney, R.P. (2016). Soil phosphorus forms from organic and conventional forage fields. Soil Sci. Soc. Am. J. 80(2): 328–340. DOI: 10.2136/sssaj2015.09.0340.
Sharma, S.B., Sayyed, R.Z., Trivedi, M.H. & Gobi, T.A. (2013). Phosphate solubilizing microbes: sustainable approach for managing phosphorus deficiency in agricultural soils. Springerplus. 2: 587. DOI: 10.1186/2193-1801-2-587.
Shinohara, R., Imai, A., Kawasaki, N., Komatsu, K., Kohzu, A. et al. (2012). Biogenic phosphorus compounds in sediment and suspended particles in a shallow eutrophic lake: A 31P-nuclear magnetic resonance (31P-NMR) study. Environ. Sci. Technol. 46(19): 10572–10578. DOI: 10.1021/es301887z.
Sun, W., Qian, X., Gu, J., Wang, X. J., Li, Y. et al. (2017). Effects of inoculation with organic-phosphorus-mineralizing bacteria on soybean (Glycine max) growth and indigenous bacterial community diversity. Can. J. Microbiol. 63(5): 392–401. DOI: 10.1139/cjm-2016-0758.
Tapia-Torres, Y., Rodríguez-Torres, M.D., Elser, J.J., Islas, A., Souza, V. et al. (2016). How to live with phosphorus scarcity in soil and sediment: Lessons from bacteria. Appl. Environ. Microbiol. 82(15): 4652–4662. DOI: 10.1128/AEM.00160-16.
Teymouri, M., Akhtari, J., Karkhane, M. & Marzban, A. (2016). Assessment of phosphate solubilization activity of rhizobacteria in mangrove forest. Biocatal. Agric. Biotechnol. 5: 168–172. DOI: 10.1016/j.bcab.2016.01.012.
Turner, B.L., Mahieu, N. & Condron, L.M. (2003). Phosphorus-31 nuclear magnetic resonance spectral assignments of phosphorus compounds in soil NaOH-EDTA extracts. Soil Sci. Soc. Am. J. 67(2): 497–510. DOI: 10.2136/sssaj2003.4970.
Ugarelli, K., Chakrabarti, S., Laas, P. & Stingl, U. (2017). The seagrass holobiont and its microbiome. Microorganisms. 5: 81. DOI: 10.3390/microorganisms5040081.
Vazquez, P., Holguin, G., Puente, M.E., Lopez-Cortes, A. & Bashan, Y. (2000). Phosphate-solubilizing microorganisms associated with the rhizosphere of mangroves in a semiarid coastal lagoon. Biol. Fertil. Soils. 30(5–6): 460–468. DOI: 10.1007/s003740050024.
Wahyudi, A.A.J., Rahmawati, S., Prayudha, B., Iskandar, M.R. & Arfianti, T. (2016). Vertical carbon flux of marine snow in Enhalus acoroides-dominated seagrass meadows. Reg. Stud. Mar. Sci. 5: 27–34. DOI: 10.1016/j.rsma.2016.01.003.
Wainwright, B.J., Zahn, G.L., Zushi, J., Lee, N.L.Y., Ooi, J.L.S. et al. (2019). Seagrass-associated fungal communities show distance decay of similarity that has implications for seagrass management and restoration. Ecol. Evol. 9(19): 11288–11297. DOI: 10.1002/ece3.5631.
Watson, S.J., Cade-Menun, B.J., Needoba, J.A. & Peterson, T.D. (2018). Phosphorus forms in sediments of a river-dominated estuary. Front. Mar. Sci. 5: 302. DOI: 10.3389/fmars.2018.00302.
Xie, F., Li, L., Song, K., Li, G., Wu, F. et al. (2019). Characterization of phosphorus forms in a Eutrophic Lake, China. Sci. Total Environ. 659: 1437–1447. DOI: 10.1016/j.scitotenv.2018.12.466.
Yang, D. & Yang, C. (2009). Detection of seagrass distribution changes from 1991 to 2006 in Xincun Bay, Hainan, with satellite remote sensing. Sensors 9(2): 830–844. DOI: 10.3390/s90200830.
Yuan, H.Z., Pan, W., Ren, L.J., Liu, E.F., Shen, J. et al. (2015). Species and biogeochemical cycles of organic phosphorus in sediments from a river with different aquatic plants located in Huaihe river watershed, China. Int. J. Phytoremediation. 17(1–6): 215–221. DOI: 10.1080/15226514.2013.876969.
Zhao, G., Sheng, Y., Jiang, M., Zhou, H. & Zhang, H. (2019). The biogeochemical characteristics of phosphorus in coastal sediments under high salinity and dredging conditions. Chemosphere. 215: 681–692. DOI: 10.1016/j.chemosphere.2018.10.015.
Zhu, F., Qu, L., Hong, X. & Sun, X. (2011). Isolation and characterization of a phosphate-solubilizing halophilic bacterium Kushneria sp. YCWA18 from Daqiao Saltern on the coast of Yellow Sea of China. Evid. Based Compl. Altern. Med. 2011: 615032. DOI: 10.1155/2011/615032.