Contents lists available at ScienceDirectMarine Pollution Bulletinjournal homepage: www.elsevier.com/locate/marpolbulCommunity dynamics and activity of nirS-harboring denitrif i ers insediments of the Indus River EstuaryFozia a , Yanling Zheng a,b,c, ? , Lijun Hou a, ?? , Zongxiao Zhang a , Dengzhou Gao b,c , Guoyu Yin b,c ,Ping Han a,b,c , Hongpo Dong a , Xia Liang a , Yi Yang b,c , Min Liu b,ca State Key Laboratory of Estuarine and Coastal Research, East China Normal University, 500 Dongchuan Road, Minhang District, Shanghai 200241, Chinab School of Geographic Sciences, East China Normal University, 500 Dongchuan Road, Minhang District, Shanghai 200241, Chinac Key Laboratory of Geographic Information Science (Ministry of Education), East China Normal University, 500 Dongchuan Road, Minhang District, Shanghai 200241,ChinaA R T I C L E I N F OKeywords:Denitrif i cationNitrogennirS geneThe Indus River EstuaryA B S T R A C TDenitrif i cation is an important pathway for reactive nitrogen removal from aquatic ecosystems. In this study, thebiodiversity, abundance, and activity of cytochrome cd 1 -type nitrate reductase gene (nirS)-harboring denitrif i ersin the sediments of the Indus River Estuary were examined by molecular and isotope-tracing techniques. Resultsshowed that the nirS-harboring denitrif i er communities showed signif i cant geographical variations along theestuarine salinity gradient. Real-time quantitative PCR showed that the abundance of nirS-harboring denitrif i ersranged from 5.3 × 10 6 to 2.5 × 10 8 copies g −1 , without signif i cant spatiotemporal variation. The potential ratesof denitrif i cation varied from 0.01 to 6.27 μmol N kg −1 h −1 and correlated signif i cantly to TOC and Fe(II)(P < 0.05). On the basis of15 N isotope-tracing experiments, the denitrif i cation process contributed 18.4–99.4%to the total nitrogen loss in the sediments of the Indus River Estuary. This study provides novel insights into themicrobial mechanism of nitrogen removal process in estuarine ecosystems.1. IntroductionNitrogen (N) is an essential constituent of living organisms, butexcess amount of nitrogen can be harmful to aquatic ecosystems(Fowler et al., 2013; Galloway, 2013; Gruber and Galloway, 2008; Luoet al., 2018). In recent decades, reactive nitrogen loading in mostaquatic environments has greatly increased, because of human activ-ities such as the discharge of industrial and municipal wastes, fertilizerapplication, and fossil fuel combustion (Booth and Campbell, 2007;Galloway et al., 2008, 2004; Vitousek and Howarth, 1991). The accu-mulation of reactive nitrogen in aquatic ecosystems has caused severeeutrophication, harmful algal blooms, hypoxia, anoxia, loss of biodi-versity and seawater acidif i cation (Booth and Campbell, 2007; Deeganet al., 2012; Diaz and Rosenberg, 2008; Galloway et al., 2004; Glibertet al., 2005; Paerl, 1997). In this context, a better understanding ofnitrogen removal and associated microbial mechanisms is crucial forcontrolling the pollution of reactive nitrogen and improving waterquality in aquatic ecosystems.Denitrif i cation is a known and sophisticated process eliminatingreactive nitrogen from aquatic ecosystems (Dalsgaard et al., 2012;Zumft, 1997). It is a primary contributor to the removal of reactivenitrogen in natural ecosystems, compared with anaerobic oxidation ofammonium (anammox) (Dalsgaard et al., 2012). Under an anaerobiccondition, denitrif i ers respire nitrate (NO 3 − ) to nitrite (NO 2 − ), nitricoxide (NO), nitrous oxide (N 2 O), and at last to N 2 (Santoro et al., 2011;Zumft, 1997). Denitrif i cation is a biological process for f i xed N removalfrom aquatic and terrestrial ecosystems, which is restricted by dif f erentgroups of microorganisms (Zumft, 1997). These diverse phylogeneticgroups of microorganisms are responsible for the denitrif i cation processby using various types of enzymes, such as nitrate reductase (Nar),nitrite reductase (Nir), nitric oxide reductase (Nor), and nitrous oxidereductase (Nos) (Zumft, 1997). In denitrif i cation, NO 2 − reduction toNO is catalyzed either by cytochrome cd 1 -nirS NO 2 − reductase orcopper containing nirK NO 2 − reductase (Coyne et al., 1989; Hochsteinand Tomlinson, 2003; Sakurai and Kataoka, 2007; Zumft, 1997). Al-though these two genes are similar in physiological and functionalmanner, but possess dif f erent skeleton (Glockner et al., 1993; Zumft,1997). The nirS gene is generally used as a gene marker for thehttps://doi.org/10.1016/j.marpolbul.2020.110971Received 26 November 2019; Received in revised form 4 February 2020; Accepted 10 February 2020? Correspondence to: Y. Zheng, State Key Laboratory of Estuarine and Coastal Research, East China Normal University, 500 Dongchuan Road, Minhang District,Shanghai 200241, China.?? Corresponding author.E-mail addresses: ylzheng@geo.ecnu.edu.cn (Y. Zheng), ljhou@sklec.ecnu.edu.cn (L. Hou).Marine Pollution Bulletin 153 (2020) 110971Available online 15 February 20200025-326X/ © 2020 Elsevier Ltd. All rights reserved.T
