Introduction
It is expected that global warming and the ensuing sea ice melt will
strongly alter the Arctic pelagic environment. This could eventually
result in a modification in the species composition and biomass of
unicellular plankton, changing matter fluxes affecting the entire
pelagic and even benthic systems (Wassmann, 2015). Thus, it is necessary
to generate information on the temporal occurrences of planktonic
species to also better understand their variability and responses
towards different environmental conditions. Acknowledging this, in 1999,
the Alfred-Wegener-Institute Helmholtz Centre for Polar and Marine
Research established the ‘Long-Term Ecological Research Site (LTER)
HAUSGARTEN to carry out regular observations of the ecosystem in the
eastern Fram Strait. This involves an extended sediment trap program
based on annual deployments of moored sediment traps at key stations of
the observation area (Soltwedel et al. 2005; 2016). Despite
uncertainties related to the use of these tools, such as trapping
efficiency, validity of results etc. (Butman 1986; Gust et al. 1994;
Buesseler et al. 2007), they remain useful in gaining insights of
vertical particle flux patterns. Moreover, the deployment of sediment
traps facilitates an understanding of plankton dynamics in the upper
water column all year-round, even in remote areas such as the Arctic
Ocean.
Measurements of bulk parameters like sinking matter and its components
combined with light microscopy can provide an estimate of the pelagic
eukaryotic microbial community in the catchment area above the traps
(Bauerfeind et al., 2009). Unfortunately, these assessments are mainly
focused on larger organisms from the micro-plankton fraction, as surveys
of small eukaryotic microbial species from the nano- and picoplankton
fractions in sediment traps are almost impossible owing to their size
and a simple morphology. For example, besides micro-eukaryotic
stramenopiles (mainly diatoms), pico- and nano-eukaryotic
Mamiellophyceae (mainly Micromonas ) and haptophyceae (mainlyPhaeocystis sp. ) are key Arctic phytoplankton taxa. They
are well known to be major contributors to phytoplankton communities and
biomass (Lovejoy and Potvin, 2011; Metfies, von Appen, Kilias, Nicolaus,
& Nothig, 2016), although their contributions to vertical flux and
particle export are not well understood. Thus, it is particularly
important to elucidate this black box, because changes in the abundance
of these key phytoplankton taxa have been observed in the area of LTER
HAUSGARTEN. These changes have been associated with a warm water anomaly
from 2005 to 2007 (Beszczynska-Moller, Fahrbach, Schauer, & Hansen,
2012; Nöthig et al., 2015), while some of them even remained after the
warm-water event. During this warm period, higher phytoplankton biomass
was observed in the water column, eukaryotic microbial plankton
>3 µm changed in composition, and diatoms that dominated
the summer period before the warm period significantly decreased (Nöthig
et al., 2015). In 2006, Phaeocystis pouchetii started to dominate
the eukaryotic microbial community and remained prominent in the region
since then, while diatom concentrations remained low and small
flagellates increased in abundance (Nöthig et al., 2015).
Over the recent years, 18S rRNA gene meta-barcoding using high
throughput sequencing (HTS) platforms has become an indispensable tool
to generate cultivation-independent and in-depth information on the
biodiversity and community composition of eukaryotic microbes, including
all size fractions, dominant and rare taxa (de Vargas et al., 2015;
Sunagawa et al., 2015). A considerable number of marine surveys have
also taken advantage of ribosomal sequence information to broaden our
understanding of protist diversity and community structure (e.g.,
Gescher et al., 2008; Metfies et al., 2010), and revolutionized the
field of microbial ecology into a semi quantitative method that allowed
testing and modeling of assemblages across time and space (e.g., Vernet
et al., 2017), and even determining links between and among communities
and/or functions (e.g., Lima-Mendez et al., 2015). The 18S rRNA V4
region is the most frequently used marker for this purpose. It is best
suited for HTS-based surveillance of microbial eukaryotes (Dunthorn et
al., 2014), as the combination of V4 with the V5 region provides the
most detailed phylogenetic information on the 18S rRNA gene (Hugerth et
al., 2014). However, different primer-evaluation studies also showed
that most of the known primer sets amplifying the V4 region are limited
in their potential to provide comprehensive biodiversity information as
they discriminate against certain taxa (Bradley, Pinto, & Guest, 2016).
Among these, the primers reported by Stoeck et al. (2010) have been
widely used in assessing microbial eukaryotic biodiversity in different
marine habitats including the Arctic region (e.g., Comeau et al., 2013;
Hardge et al., 2017). Although a truthfully ‘universal’ primer that can
amplify all known representative sequences has not yet been described,
many efforts have been done to at least minimize or lessen potential PCR
biases. Hugerth et al. (2014) for example has systematically designed
primers that greatly performed in silico but were not tested
using mock communities. Their results showed that the majority of primer
sets were able to amplify most but not all taxonomic groups. This is
important since our capacity to describe and predict patterns in the
communities depend on our ability to amplify and detect most members of
the community. Such task has become more challenging since many of the
existing primers were primarily designed based on available sequences of
known species, and accumulating evidence show that many previously
unknown taxa are increasingly detected with more environmental surveys
(Massana et al., 2011). Thus, there is a need to carefully select for
primers to be used depending on the question being asked or the targeted
taxonomic groups, or even use complementary primer sets to retrieve
realistic information on microbial biodiversity in a sample.
This study aimed at elucidating changes in the composition of exported
pelagic phytoplankton communities in response to a warm anomaly in Fram
Strait from 2005-2007. The study is based on using a set of published
and newly developed primer-sets considering their complementarity,
limitations and advantages deduced. We explored major taxonomic
phytoplankton groups including the Chlorophyta, Haptophyta and
Bacillariophyta, and their contribution to the exported eukaryotic
microbial communities during the phases of maximum particulate organic
carbon (POC) flux in spring and autumn from 2000-2011 in the area of
LTER HAUSGARTEN. This study provides new insights on how primer biases
could limit or improve our understanding of ecological dynamics of
microbial communities and their contribution to carbon transport in the
changing oceans, such as the Arctic.