INTRODUCTION
Zooplankton form some of the most abundant and diverse biological
communities in aquatic environments and constitute an integral component
of both marine and freshwater ecosystems
(Bucklin,
Lindeque, Rodriguez-Ezpeleta, Albaina, & Lehtiniemi, 2016; Djurhuus et
al., 2018; Xiong et al., 2019). Estimating the richness of zooplankton
species remains a priority in several fields of research, from ecology
to conservation biology
(Carugati,
Corinaldesi, Dell’Anno, & Danovaro, 2015; Gaston, 2009). Such studies
are particularly important in marine ecosystems where ecological
processes are often maintained by both complex biotic interactions and
environmental drivers
(Palumbi et al.,
2009; Steinberg & Landry,
2017).
For example, zooplankton communities play a central role in regulating
biogeochemical cycles by transferring carbon from primary producers to
higher trophic levels
(Bucklin et al.,
2019; Everaert, Deschutter, De Troch, Janssen, & De Schamphelaere,
2018; Lindeque, Parry, Harmer, Somerfield, & Atkinson, 2013; Steinberg
& Landry,
2017).
Moreover, due to their small size, limited capacity for self-dispersal,
and high sensitivity to environmental change, zooplankton are also
considered to be useful bio-indicators that may be used to evaluate the
health of marine ecosystems
(Buttay, Miranda,
Casas, González-Quirós, & Nogueira, 2015; Johnston, Mayer-Pinto, &
Crowe, 2015; Parmar, Rawtani, & Agrawal, 2016; Yang & Zhang,
2019).
Long-term studies have reported that both environmental change and
ecological processes shape the spatial and temporal abundance of
zooplankton and the taxonomic composition of their communities
(Buttay et al.,
2015).
As such, environmental change may lead to the loss of zooplankton
biodiversity
(Gazonato Neto,
Silva, Saggio, & Rocha, 2014; Parmar et al.,
2016),
which in turn may affect ecosystem services and result in economic
consequences
(Beaugrand,
Edwards, & Legendre, 2010; Bucklin et al., 2016; Everaert et al., 2018;
Johnston et al.,
2015).
The Gulf of Mexico (GoM) is an example of a system that is subject to
changing environmental conditions. For example, the large scale
near-surface circulation in the GoM is largely dominated by the
energetic Loop Current (LC; Damien et al., 2018). The northward
penetration of the LC is often accompanied by the formation and release
of anticyclonic LC eddies (LCEs), which travel towards the western
boundary of the GoM
(Damien
et al., 2018; Sheinbaum, Athié, Candela, Ochoa, & Romero-Arteaga,
2016). Together with other smaller cyclonic and anticyclonic eddies that
are formed within the gulf, LCEs are considered to constitute the
principal source of mesoscale variability within this ecosystem
(Damien
et al., 2018; Hamilton, 2007; Jouanno et al., 2016). Moreover, LCEs and
smaller eddies have been found to constrain the distributions of
nutrients, zooplankton, and other planktonic species in the gulf
(Biggs &
Ressler, 2001; Linacre et al., 2015). Based on the distribution of
chlorophyll
(Damien et al.,
2018; Muller-Karger et al., 2015; Salmerón-García, Zavala-Hidalgo,
Mateos-Jasso, & Romero-Centeno, 2011), the GoM may be conceptually
divided into two main areas: (1) a central oligotrophic area and (2) the
eutrophic inshore waters that semi-surround it (Damien et al., 2018;
Uribe-Martínez, Aguirre-Gómez, Zavala-Hidalgo, Ressl, & Cuevas, 2019).
Nevertheless, few studies have evaluated plankton distributions in the
eutrophic inshore waters of the GoM
(Biggs &
Ressler, 2001; Damien et al., 2018) nor the distribution patterns of
zooplankton and other planktonic species over the entire gulf. This
constitutes the principle limitation in determining if the zooplankton
distribution within the GoM also follows the pattern that has been
established for chlorophyll.
The main challenges in evaluating the spatial and temporal patterns of
zooplankton communities lie in the taxonomic complexity of their
assemblages, which include a considerable number of morphologically
diverse and cryptic species and a lack of diagnostic characteristics for
immature and larval developmental stages
(Bucklin
et al., 2016). In recent years, molecular methods have generated new and
powerful approaches for assessing zooplankton biodiversity by overcoming
the main limitations associated with traditional taxonomic surveys, such
as the amount of time needed to identify specimens and the need for
advanced taxonomic expertise
(Creer
et al., 2016; Cristescu, 2014; Zhang, Chain, Abbott, & Cristescu,
2018). In particular, the metabarcoding approach
(Creer
et al., 2016) is considered to be one of the most comprehensive means to
holistically evaluate zooplankton assemblages, as it combines DNA
barcoding and high-throughput sequencing to evaluate the taxonomic
composition of a sample with any source of environmental DNA (eDNA;
Stefanni
et al., 2018; Zhang et al., 2018). However, metabarcoding results
strongly depend on having adequate taxonomic coverage and a systematic
resolution of the chosen molecular marker
(Bucklin
et al., 2016; Larke, Beard, Swadling, & Deagle, 2017; Zhang et al.,
2018).
Nuclear-encoded ribosomal RNA fragments, especially hypervariable
regions of the 18S rRNA gene, were prime targets in early studies
because they are able to provide conserved primer binding sites with
broad taxonomic coverage across the eukaryotic domain
(Larke
et al., 2017; Lindeque et al., 2013). Nevertheless, the 18S rRNA gene
often lacks the taxonomic resolution of protein-coding genes, such as
mitochondrial cytochrome oxidase c subunit I (COI;
Larke
et al., 2017; Machida, Leray, Ho, & Knowlton, 2017; Zhang et al.,
2018). Indeed, COI undergoes faster rates of evolution than that of the
18S rRNA gene; hence, its great genetic variability can facilitate the
systematic classification of even closely related taxa
(Zhang
et al., 2018). Nevertheless, the wobble effect (i.e., meaningless
nucleotide change in the position of the third codon) can increase the
occurrence of primer mismatches among taxa and consequentially induce
taxonomic bias during PCR amplification
(Larke
et al., 2017; Piñol, Mir, Gomez-Polo, & Agustí,
2015).
These gene-related limitations have led many researchers to propose the
need to implement a multi-locus approach in metabarcoding surveys, as
the synergistic information of different loci may represent a trade-off
between increasing the resolution needed to identify metazoan taxa
(Carroll
et al.,
2019)
and reducing the occurrence of false negative and/or false positive
outcomes
(Bucklin
et al., 2016; Zhang et al.,
2018).
Accordingly, it has become increasingly accepted that the combined use
of nuclear and mitochondrial genes may facilitate the assessment of
zooplankton communities due to the resulting improved sensitivity for
detecting cryptic as well as intra-species genetic diversity
(Carugati et al.,
2015).
Despite the ecological importance of zooplankton and an improved
understanding of the dynamics of the GoM, few efforts have been made to
describe the composition and distribution of the entire zooplankton
community that is dispersed throughout the deep-water region of the gulf
in the Exclusive Economic Zone (EEZ) of Mexico. Moreover, the extent to
which the physical environment may shape the structure of those
communities remains uncertain. In this study, we hypothesized that the
structure of the zooplankton community would reflect regional and
seasonal environmental features. Accordingly, the main goals of this
study were to (i ) assess the spatial and temporal variability of
the zooplankton community from the deep water region of the GoM within
the Mexican EEZ, (ii ) explore the possibility that structural
changes in the zooplankton community might be related to environmental
factors, and (iii ) describe the regional and temporal diversity
of the community.
In order to address these hypotheses, we characterized the structure and
variability of the zooplankton community using a metabarcoding approach
based on the genetic information of both the hypervariable V9 and
Leray_Folmer 1 regions of the 18S rRNA and COI genes, respectively. We
chose this combination of genes because it currently provides the best
compromise between the detection and resolution of zooplankton taxa.
Specifically, the hypervariable V9 region may be used to amplify across
all known and unknown metazoan phyla due to its highly conserved
flanking regions
(Amaral-Zettler,
McCliment, Ducklow, & Huse,
2009).
In contrast, we posited that COI could provide greater taxonomic
resolution (or α-biodiversity) due to its high variability
(Heimeier,
Lavery, & Sewell,
2010).
For both molecular markers, detected amplicon sequence variants (ASVs)
were used to estimate zooplankton alpha and beta diversity across the
various sampled areas of the GoM. First, we analyzed the genetic
information from three cruises, and we pooled all data from each locus
to evaluate the potential zooplankton patterns in a comprehensive
analysis. Our efforts were directed towards investigating the effects of
temperature, salinity, oxygen, depth, latitude, longitude, water
density, and florescence (as a proxy for chlorophyll abundance) in
shaping the structure of the zooplankton community in order to better
understand the physical-biological interactions controlling spatial,
seasonal, and inter-annual changes of zooplankton assemblages.