Strong impact of anthropogenic contamination on the co-occurrence patterns of a riverine
microbial community
Running title:
Co-occurrence network of riverine microbiome
Anyi Hu
1
, Feng Ju
2
, Liyuan Hou
1,3
, Jiangwei Li
1
, Xiaoyong Yang
1
, Hongjie Wang
1,3
,
Sikandar I. Mulla
1
, Qian Sun
1
, Helmut Bürgmann
2*
, Chang-Ping Yu
1,4*
1. CAS Key Laboratory of Urban Pollutant Conversion, Institute of Urban Environment, Chinese
Academy of Sciences, Xiamen 361021, China
2. Eawag, Swiss Federal Institute of Aquatic Science and Technology, Department of Surface
Waters-Research and Management, 6047 Kastanienbaum, Switzerland
3. University of Chinese Academy of Sciences, Beijing 100049, China
4. Graduate Institute of Environmental Engineering, National Taiwan University, Taipei 106, Taiwan
*Correspondence to:
Dr. Chang-Ping Yu, cpyu@iue.ac.cn
CAS Key Laboratory of Urban Pollutant Conversion, Institute of Urban Environment, Chinese Academy
of Sciences, Xiamen 361021, China, Telephone ＆ Fax: +86-592-6190768.
Dr. Helmut Bürgmann, Helmut.Buergmann@eawag.ch
Eawag, Swiss Federal Institute of Aquatic Science and Technology, Department of Surface
Waters-Research and Management, 6047 Kastanienbaum, Switzerland, Telephone:+41 58 765 2165 Fax:
+41 58 765 2168.
This article has been accepted for publication and undergone full peer review but has not been
through the copyediting, typesetting, pagination and proofreading process which may lead to
differences between this version and the Version of Record. Please cite this article as an
‘Accepted Article’, doi: 10.1111/1462-2920.
13942
This article is protected by copyright. All rights reserved.
This is the peer reviewed version of the following article: Hu, A., Ju, F., Hou, L., Li, J.,
Yang, X., Wang, H., … Yu, C. -P. (2017). Strong impact of anthropogenic contamination on the
co-occurrence patterns of a riverine microbial community: Co-occurrence network of riverine
microbiome. Environmental Microbiology, in press, which has been published in final form at
http://doi.org/10.1111/1462-2920.13942. This article may be used for non-commercial purposes
in accordance with Wiley Terms and Conditions for Self-Archiving.
Originality-Significance Statement
Emerging organic contaminants such as pharmaceuticals and personal care products (PPCPs)
represent a great threat to the aquatic biome, yet in comparison to toxic effects on macrobiota, the impact
of PPCPs on riverine microbiota are very poorly understood. Here we demonstrated that α-diversity of
riverine microbial communities can decrease under the stress caused by excessive nutrients and PPCPs.
Moreover, PPCPs are shown to play a more important role in shaping the microbial co-occurrence
ecological network in the lotic ecosystems studied than regular physico-chemical factors. Our work
provides new insights into our understanding of how organic micropollutants affect the assembly of
aquatic microbial communities.
Summary
Although the health of rivers is threatened by multiple anthropogenic stressors with increasing
frequency, it remains an open question how riverine microbial communities respond to emerging
micropollutants. Here, by using 16S rDNA amplicon sequencing of 60 water samples collected during
different hydrological seasons, we investigated the spatio-temporal variation and the co-occurrence
patterns of microbial communities in the anthropogenically impacted Jiulong River in China. The results
indicated that the riverine microbial co-occurrence network had a non-random, modular structure, which
was mainly shaped by the taxonomic relatedness of co-occurring species. Fecal indicator bacteria may
survive for prolonged periods of time in river water, but they formed an independent module which had
fewer interactions with typical freshwater bacteria. Multivariate analysis demonstrated that nutrients and
micropollutants (i.e. pharmaceuticals and personal care products, PPCPs) exerted combined effects in
shaping α- and β-diversity of riverine microbial communities. Remarkably, we showed that a hitherto
unrecognized disruptive effect of PPCPs on the abundance variations of central species and module
communities was stronger than the influence of physico-chemical factors, suggesting the key role played
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by micropollutants for the microbial co-occurrence relationships in lotic ecosystems. Overall, our findings
provide novel insights into community assembly in aquatic environments experiencing anthropogenic
stresses.
Key words: Microbial community, spatio-temporal variation, co-occurrence network, nutrient,
pharmaceuticals and personal care products, Jiulong River
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Introduction
The essential roles of microorganisms in mediating global biogeochemical cycles and maintaining
ecosystem services are well known. The recent application of high-throughput sequencing provides an
increasingly clear view of the specific and unique microbiomes that characterize different natural or
artificial environments, such as freshwater, ocean, soil, sediment, hot springs, animal guts and bioreactors
(Tamames et al., 2010). In lake ecosystems, a number of microbial taxa have been termed 'typical
freshwater bacteria' (TFB), which is based on phylogenetic analysis of 16S rRNA gene pools from lake
bacterioplankton, mainly including α-, β- and γ-Proteobacteria, Actinobacteria, Bacteroidetes,
Cyanobcateria and Verrucomicrobia, (Glöckner et al., 2000; Zwart et al., 2002; Newton et al., 2011).
Many prior investigations have demonstrated that TFB are widely distributed not only in lake water, but
also in the surface waters of rivers and streams (Hu et al., 2014; Savio et al., 2015; Meziti et al., 2016).
However, the relative abundance of TFB was found to generally decrease in those parts of the lotic
ecosystems that suffer strong disturbances from human activities; Specific allochthonous signals, such as
inflows of sewage and the associated human and animal gut bacteria, could also be detected when
studying the composition and dynamics of riverine microbial communities (Hu et al., 2014; Meziti et al.,
2016). These findings highlight that rivers are among the most vulnerable ecosystems in the context of a
growing human population along the riverside and increasing anthropogenic pressure.
Meanwhile, rivers are critical sources of water for drinking, agriculture and industrial production as
well as valuable reservoirs of biodiversity (Shiklomanov, 1998), not to mention essential channels for
connecting terrestrial and marine ecosystems (Vörösmarty et al., 2010). However, the ecological
mechanisms controlling microbial community assembly and interspecies interactions in
anthropogenically disturbed rivers have not been resolved with respect to complex abiotic and biotic
environmental factors.
So far, evidence has accumulated that both deterministic (species sorting) and stochastic (mass effect)
processes govern the assembly of aquatic microbial communities (Savio et al., 2015; Nino-Garcia et al.,
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2016). In lotic ecosystems, local physico-chemical factors such as temperature, pH, dissolved oxygen
(DO), nitrogen and phosphorus nutrients as well as organic matter usually control species sorting to a
considerable extent (Hu et al., 2014; Staley et al., 2015), whereas the hydrological structure of lotic
ecosystems (flow direction, fluvial network, water residence time and riparian construction) may have an
influence on balancing deterministic and stochastic processes (Widder et al., 2014; Savio et al., 2015;
Nino-Garcia et al., 2016). Moreover, chemical micropollutants discharged from riparian human activities,
such as pharmaceuticals and personal care products (PPCPs), may act as potential selective forces on
riverine microbial communities considering their intensive/widespread use, resistance to wastewater
treatment and biodegradation, environmental persistence, and known ecotoxicological effects on aquatic
life (Schwarzenbach et al., 2006; Rosi-Marshall and Royer, 2012; Liu and Wong, 2013).
The potential influences of PPCPs on the diversity and function of freshwater microbial
communities were also assessed by means of several laboratory-scale microcosm or mesocosm
experiments (Yergeau et al., 2012; Lawrence et al., 2015; Sabater et al., 2016). It has been suggested that
the pseudo-persistent PPCPs may have contrasting ecological effects on the aquatic community: PPCPs
may become carbon and energy sources to certain members of the natural microbial community via
biodegradation (subsidy effect), while being toxic to others (stress effect) - even though their
environmental concentrations are generally low (ng/L ~ μg/L range) (Luo et al., 2014). Both subsidy and
stress effects of PPCPs would, however, be expected to lead to changes in the diversity and composition
of freshwater microbial communities and relevant biogeochemical processes (Yergeau et al., 2012).
Apart from abiotic environmental factors, the composition and dynamics of microbial communities
are also shaped by ecological factors such as top-down control (i.e. grazing and viral lysis) and
microbe-microbe interactions (Chaffron et al., 2010; Faust and Raes, 2012; Lima-Mendez et al., 2015).
Microbe-microbe interactions specifically have received little attention until recently (Faust and Raes,
2012). Due to advances in high-throughput sequencing technologies, the increased availability of
sequence data across large spatio-temporal scales has provided an opportunity to address this gap in
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