Friday, February 23, 2018

Weekend reads

And another load of DNA barcoding must-reads right on time for the weekend.

BACKGROUND:
Understanding feedback between above- and below-ground processes of biological communities is a key to the effective management of natural and agricultural ecosystems. However, as above- and below-ground food webs are often studied separately, our knowledge of material flow and community dynamics in terrestrial ecosystems remains limited.
RESULTS:
We developed a high-throughput sequencing method for examining how spiders link above- and below-ground food webs as generalist predators. To overcome problems related to DNA-barcoding-based analyses of arthropod-arthropod interactions, we designed spider-specific blocking primers and Hexapoda-specific primers for the selective PCR amplification of Hexapoda prey sequences from spider samples. By applying the new DNA metabarcoding framework to spider samples collected in a temperate secondary forest in Japan, we explored the structure of a food web involving 15 spider species and various taxonomic groups of Hexapoda prey. These results support the hypothesis that multiple spider species in a community can prey on both above- and below-ground prey species, potentially coupling above- and below-ground food-web dynamics.
CONCLUSIONS:
The PCR primers and metabarcoding pipeline described in this study are expected to accelerate nuclear marker-based analyses of food webs, illuminating poorly understood trophic interactions in ecosystems.

Different second-generation sequencing technologies may have taxon-specific biases when DNA metabarcoding prey in predator feces. Our major objective was to examine differences in prey recovery from bat guano across two different sequencing workflows using the same fecal DNA extracts. We compared results between the Ion Torrent PGM and the Illumina MiSeq with similar library preparations and the same analysis pipeline. We focus on repeatability and provide an R Notebook in an effort toward transparency for future methodological improvements. Full documentation of each step enhances the accessibility of our analysis pipeline. We tagged DNA from insectivorous bat fecal samples, targeted the arthropod cytochrome c oxidase I (COI) minibarcode region, and sequenced the product on both second-generation sequencing platforms. We developed an analysis pipeline with a high operational taxonomic unit (OTU) clustering threshold (i.e., ≥ 98.5%) followed by copy number filtering to avoid merging rare but genetically similar prey into the same OTUs. With this workflow, we detected 297 unique prey taxa, of which 74% were identified at the species-level. Of these, 104 (35%) prey OTUs were detected by both platforms, 176 (59%) OTUs were detected by the Illumina MiSeq system only, and 17 (6%) OTUs were detected using the Ion Torrent system only. Costs were similar between platforms but the Illumina MiSeq recovered six times more reads and four additional insect orders than did Ion Torrent. The considerations we outline are particularly important for long-term ecological monitoring; a more standardized approach will facilitate comparisons between studies and allow faster recognition of changes within ecological communities.

The recent recolonization of Central Europe by the European gray wolf (Canis lupus) provides an opportunity to study the dynamics of parasite transmission for cases when a definitive host returns after a phase of local extinction. We investigated whether a newly established wolf population increased the prevalence of those parasites in ungulate intermediate hosts representing wolf prey, whether some parasite species are particularly well adapted to wolves, and the potential basis for such adaptations. We recorded Sarcocystis species richness in wolves and Sarcocystis prevalence in ungulates harvested in study sites with and without permanent wolf presence in Germany using microscopy and DNA metabarcoding. Sarcocystis prevalence in red deer (Cervus elaphus) was significantly higher in wolf areas (79.7%) than in control areas (26.3%) but not in roe deer (Capreolus capreolus) (97.2% vs. 90.4%) or wild boar (Sus scrofa) (82.8% vs. 64.9%). Of 11 Sarcocystis species, Sarcocystis taeniata and Sarcocystis grueneri occurred more often in wolves than expected from the Sarcocystis infection patterns of ungulate prey. Both Sarcocystis species showed a higher increase in prevalence in ungulates in wolf areas than other Sarcocystis species, suggesting that they are particularly well adapted to wolves, and are examples of "wolf specialists". Sarcocystis species richness in wolves was significantly higher in pups than in adults. "Wolf specialists" persisted during wolf maturation. The results of this study demonstrate that (1) predator-prey interactions influence parasite prevalence, if both predator and prey are part of the parasite life cycle, (2) mesopredators do not necessarily replace the apex predator in parasite transmission dynamics for particular parasites of which the apex predator is the definitive host, even if meso- and apex predators were from the same taxonomic family (here: Canidae, e.g., red foxes Vulpes vulpes), and (3) age-dependent immune maturation contributes to the control of protozoan infection in wolves.

Cephalopods are primarily active predators throughout life. Flying squids (family Ommastrephidae) represents the most widely distributed and ecologically important family of cephalopods. While the diets of adult flying squids have been extensively studied, the first feeding diet of early paralarvae remains a mystery. The morphology of this ontogenetic stage notably differs from other cephalopod paralarvae, suggesting a different feeding strategy. Here, a combination of Laser Capture Microdissection (LCM) and DNA metabarcoding of wild-collected paralarvae gut contents for eukaryotic 18S v9 and prokaryotic 16S rRNA was applied, covering almost every life domain. The gut contents were mainly composed by fungus, plants, algae and animals of marine and terrestrial origin, as well as eukaryotic and prokaryotic microorganisms commonly found in fecal pellets and particulate organic matter. This assemblage of gut contents is consistent with a diet based on detritus. The ontogenetic shift of diet from detritivore suspension feeding to active predation represents a unique life strategy among cephalopods and allows ommastrephid squids to take advantage of an almost ubiquitous and accessible food resource during their early stages. LCM was successfully applied for the first time to tiny, wild-collected marine organisms, proving its utility in combination with DNA metabarcoding for dietary studies.

Wednesday, February 21, 2018

New shark species described

An Atlantic sixgill pup found off the coast of Belize. Credit: Ivy Baremore/MarAlliance 
Colleagues around the world are finding new species every day showing us how much we still have to learn about our planet's biodiversity. That being said, it is fairly rare that a new species of shark is described or resurrected based on additional evidence. 

Scientists of the Florida Institute of Technology confirmed after decades of uncertainty that sixgill sharks residing in the Atlantic Ocean are a different species than their counterparts in the Indian and Pacific oceans. With ancestors dating back over 250 million years, sixgill sharks are among the oldest creatures on Earth. Yet the fact that they reside at extreme ocean depths, sometimes hundreds of meters below the surface, has made them especially challenging to study.

Only by using two mitochondrial genes, COI and ND2, could the group confirm that bigeye sixgill sharks from the Atlantic Ocean (Belize, Gulf of Mexico, and Bahamas) diverged from those in the Pacific and Indian Oceans (Japan, La Reunion, and Madagascar). It also turns out that with up to 2m in length, Atlantic sixgill sharks are far smaller than their Indo-Pacific relatives, which can grow to 5m or longer. 

With their new classification, Atlantic sixgill sharks will now have a better chance at long-term survival, because we now know there are two unique species, we have a sense of the overall variation in populations of sixgills. We understand that if we overfish one of them, they will not replenish from elsewhere in the world.

Tuesday, February 20, 2018

Extinction cascades

Current species extinction rates are at unprecedentedly high levels. While human activities can be the direct cause of some extinctions, it is becoming increasingly clear that species extinctions themselves can be the cause of further extinctions, since species affect each other through the network of ecological interactions among them. There is concern that the simplification of ecosystems, due to the loss of species and ecological interactions, increases their vulnerability to such secondary extinctions. It is predicted that more complex food webs will be less vulnerable to secondary extinctions due to greater trophic redundancy that can buffer against the effects of species loss.

In other words more complex food webs are likely less vulnerable to extinction cascades because there is a greater chance that other species can step in and buffer against the effects of species loss. Researchers from the University of Exeter used communities of plants and insects to experimentally test this prediction. The removal of a parasitoid wasp species led to secondary extinctions of other, indirectly linked, species at the same trophic level. This effect was much stronger in simple communities than for the same species within a more complex food web. 

The study results demonstrate that biodiversity loss can increase the vulnerability of ecosystems to secondary extinctions which, when they occur, can then lead to further simplification causing run-away extinction cascades. 

In case you want to read and learn more about the interesting discussions relating to extinction cascades I recommend a blog post that I found particularly helpful. Another good source of information is - in case you have access to TREE - a review article (the figure to this post I took from this article).



Friday, February 16, 2018

Weekend reads

In time for the weekend this time - here in Canada a long one. Lots of interesting and diverse topics.

Arthropod communities in the tropics are increasingly impacted by rapid changes in land use. Because species showing distinct seasonal patterns of activity are thought to be at higher risk of climate-related extirpation, global warming is generally considered a lower threat to arthropod biodiversity in the tropics than in temperate regions. To examine changes associated with land use and weather variables in tropical arthropod communities, we deployed Malaise traps at three major anthropogenic forests (secondary reserve forest, oil palm forest, and urban ornamental forest (UOF)) in Peninsular Malaysia and collected arthropods continuously for 12 months. We used metabarcoding protocols to characterize the diversity within weekly samples. We found that changes in the composition of arthropod communities were significantly associated with maximum temperature in all the three forests, but shifts were reversed in the UOF compared with the other forests. This suggests arthropods in forests in Peninsular Malaysia face a double threat: community shifts and biodiversity loss due to exploitation and disturbance of forests which consequently put species at further risk related to global warming. We highlight the positive feedback mechanism of land use and temperature, which pose threats to the arthropod communities and further implicates ecosystem functioning and human well-being. Consequently, conservation and mitigation plans are urgently needed.

Currently, freshwater zooplankton sampling and identification methodologies have remained virtually unchanged since they were first established in the beginning of the XX century. One major contributing factor to this slow progress is the limited success of modern genetic methodologies, such as DNA barcoding, in several of the main groups. This study demonstrates improved protocols which enable the rapid assessment of most animal taxa inhabiting any freshwater system by combining the use of light traps, careful fixation at low temperatures using ethanol, and zooplankton-specific primers. We DNA-barcoded 2,136 specimens from a diverse array of taxonomic assemblages (rotifers, mollusks, mites, crustaceans, insects, and fishes) from several Canadian and Mexican lakes with an average sequence success rate of 85.3%. In total, 325 Barcode Index Numbers (BINs) were detected with only three BINs (two cladocerans and one copepod) shared between Canada and Mexico, suggesting a much narrower distribution range of freshwater zooplankton than previously thought. This study is the first to broadly explore the metazoan biodiversity of freshwater systems with DNA barcodes to construct a reference library that represents the first step for future programs which aim to monitor ecosystem health, track invasive species, or improve knowledge of the ecology and distribution of freshwater zooplankton.

During the past 50 years, the molecular clock has become one of the main tools for providing a time scale for the history of life. In the era of robust molecular evolutionary analysis, clock calibration is still one of the most basic steps needing attention. When fossil records are limited, well-dated geological events are the main resource for calibration. However, biogeographic calibrations have often been used in a simplistic manner, for example assuming simultaneous vicariant divergence of multiple sister lineages. Here, we propose a novel iterative calibration approach to define the most appropriate calibration date by seeking congruence between the dates assigned to multiple allopatric divergences and the geological history. Exploring patterns of molecular divergence in 16 trans-Bering sister clades of echinoderms, we demonstrate that the iterative calibration is predominantly advantageous when using complex geological or climatological events-such as the opening/reclosure of the Bering Strait-providing a powerful tool for clock dating that can be applied to other biogeographic calibration systems and further taxa. Using Bayesian analysis, we observed that evolutionary rate variability in the COI-5P gene is generally distributed in a clock-like fashion for Northern echinoderms. The results reveal a large range of genetic divergences, consistent with multiple pulses of trans-Bering migrations. A resulting rate of 2.8% pairwise Kimura-2-parameter sequence divergence per million years is suggested for the COI-5P gene in Northern echinoderms. Given that molecular rates may vary across latitudes and taxa, this study provides a new context for dating the evolutionary history of Arctic marine life.

Properly designed (randomized and/or balanced) experiments are standard in ecological research. Molecular methods are increasingly used in ecology, but studies generally do not report the detailed design of sample processing in the laboratory. This may strongly influence the interpretability of results if the laboratory procedures do not account for the confounding effects of unexpected laboratory events. We demonstrate this with a simple experiment where unexpected differences in laboratory processing of samples would have biased results if randomization in DNA extraction and PCR steps do not provide safeguards. We emphasize the need for proper experimental design and reporting of the laboratory phase of molecular ecology research to ensure the reliability and interpretability of results.

Metabarcoding of lake sediments may reveal current and past biodiversity, but little is known about the degree to which taxa growing in the vegetation are represented in environmental DNA (eDNA) records. We analysed composition of lake and catchment vegetation and vascular plant eDNA at 11 lakes in northern Norway. Out of 489 records of taxa growing within 2 m from the lake shore, 17-49% (mean 31%) of the identifiable taxa recorded were detected with eDNA. Of the 217 eDNA records, 73% and 12% matched taxa recorded in vegetation surveys within 2 m and up to about 50 m away from the lakeshore, respectively, whereas 16% were not recorded in the vegetation surveys of the same lake. The latter include taxa likely overlooked in the vegetation surveys or growing outside the survey area. The percentages detected were 61, 47, 25, and 15 for dominant, common, scattered, and rare taxa, respectively. Similar numbers for aquatic plants were 88, 88, 33 and 62%, respectively. Detection rate and taxonomic resolution varied among plant families and functional groups with good detection of e.g. Ericaceae, Roseaceae, deciduous trees, ferns, club mosses and aquatics. The representation of terrestrial taxa in eDNA depends on both their distance from the sampling site and their abundance and is sufficient for recording vegetation types. For aquatic vegetation, eDNA may be comparable with, or even superior to, inlake vegetation surveys and therefore be used as an tool for biomonitoring. For reconstruction of terrestrial vegetation, technical improvements and more intensive sampling is needed to detect a higher proportion of rare taxa although DNA of some taxa may never reach the lake sediments due to taphonomical constrains. Nevertheless, eDNA performs similar to conventional methods of pollen and macrofossil analyses and may therefore be an important tool for reconstruction of past vegetation.

Marine Arctic Fishes


Today a post about some of the work I have been doing over the past years with colleagues from Norway, Russia, Denmark, and the US. Incredible tedious work in particular by the lead author Cathrine Mecklenburg. My part was everything barcoding and interpretation of studies utilizing molecular genetics in the widest sense. Two volumes totalling some 740 pages.



Marine Fishes of the Arctic Region is intended for all who do research in and monitoring of marine eco­systems in the Arctic. It presents accounts for 205 species with maps of global distribution and descriptions of morphology and habitat, as well as a photographic identification guide. Information on 24 other species present only in the fringes of the Arctic Region or taxonomically problematic is given in the introductions to the fish families. As the Arctic continues to warm, more cold-temperate species are expected to enter the region and the distribution of true Arctic species will likely retract as the area of ice-covered cold water shrinks. The maps in this atlas can be used to compare future changes in distributions. The identification guide will be particu­larly helpful for identifying cold-water species, since fewer identification tools are available for this group of fishes.

We're pretty proud of this body of work and hope that it will be helpful and inspire future research. Since the Arctic Ocean is new in an evolutionary time scale, studying species from this area can give insight into the ongoing processes govern­ing zoogeographical patterns like migration and local adaptations. Understanding such processes is of particu­lar relevance in light of global climate change.


Tuesday, February 13, 2018

International wildlife trade

Increasing our understanding of the drivers of international wildlife trade is critical as unsustainable harvesting of wildlife can lead to population decline and the extinction of species. While there is currently a database of legal trade in restricted species, it relies on the submission of annual reports which can be undermined by weak domestic legislation and governance hence we are not getting a complete picture of the industry.

Researchers from the National University of Singapore conducted an in-depth study on international wildlife trade data. They used a new gravity-underreporting model to carry out a comprehensive analysis and comparison of over 370,000 records of wildlife trade between 2004 and 2013 across three groups -- mammals, avian and reptiles.  The key findings established from the analysis show market forces the drive the movement of wildlife products around the globe, and demonstrate that our understanding of illegal and legal wildlife trade is biased towards certain species and regions of the globe. The findings also implied that wildlife trade networks may be more complicated than previously thought, undermining enforcement and conservation efforts. Other findings of the study:

  • Illegal products entering the USA come predominantly from Canada, Mexico and China
  • Illegal products entering the USA were less likely to be intercepted if they were coming from Africa, central Asia, Eastern Europe and Pacific Island states suggesting the existence of complex trade networks and the potential for the laundering of illegal products through legal markets
  • Different drivers and markets exist for mammalian, avian and reptilian trade, suggesting a nuanced approach to regulation and monitoring, which accounts for these differences, is required for effective conservation.
  • The Convention on the International Trade in Endangered Species of Wild Fauna and Flora (CITES) success depends on products considered, and trade in less well studied groups such as orchids, timber or corals are likely to be less well regulated by CITES.

Regulatory authorities, such as CITES, should leverage this information to improve existing conservation efforts and policies and extend efforts into areas that are currently underrepresented.

The trends we have established in this study highlight the need for regulatory bodies to look beyond the existing databases and take into account the uncertainty surrounding our current understanding of wildlife trade in their conservation efforts. For example, capacity building to improve our ability to regulate and monitor trade in less well studied species and in countries with higher levels of corruption are essential if we want to prevent trade driven extinctions globally

Monday, February 12, 2018

Weekend reads (after the weekend)

A little late this time but still some very interesting papers for you. You can hold on to them for the coming weekend or read them right away. Your choice.

Soil is often collected from a suspect's tire, vehicle, or shoes during a criminal investigation and subsequently submitted to a forensic laboratory for analysis. Plant and insect material recovered in such samples is rarely analyzed, as morphological identification is difficult. In this study, DNA barcoding was used for taxonomic identifications by targeting the gene regions known to permit discrimination in plants [maturase K (matK) and ribulose 1,5-biphosphate carboxylase (rbcL)] and insects [cytochrome oxidase subunit I (COI)]. A DNA barcode protocol suitable for processing forensic-type biological fragments was developed and its utility broadly tested with forensic-type fragments (e.g., seeds, leaves, bark, head, legs; n, 213) isolated from soils collected within Virginia, USA (n, 11). Difficulties with PCR inhibitors in plant extracts and obtaining clean Sanger sequence data from insect amplicons were encountered during protocol development; however, the final protocol produced sequences specific to the expected locus and taxa. The overall quantity and quality of DNA extracted from the 213 forensic-type biological fragments was low (< 15 ng/μL). For plant fragments, only the rbcL sequence data was deemed reliable; thus, taxonomic identifications were limited to the family level. The majority of insect sequences matched COI in both GenBank and Barcode of Life DataSystems; however, they were identified as an undescribed environmental contaminant. Although limited taxonomic information was gleaned from the forensic-type fragments processed in this study, the new protocol shows promise for obtaining reliable and specific identifications through DNA barcoding, which could ultimately enhance the information gleaned from soil examinations.

Gelatinous zooplankton are a large component of the animal biomass in all marine environments, but are considered to be uncommon in the diet of most marine top predators. However, the diets of key predator groups like seabirds have conventionally been assessed from stomach content analyses, which cannot detect most gelatinous prey. As marine top predators are used to identify changes in the overall species composition of marine ecosystems, such biases in dietary assessment may impact our detection of important ecosystem regime shifts. We investigated albatross diet using DNA metabarcoding of scats to assess the prevalence of gelatinous zooplankton consumption by two albatross species, one of which is used as an indicator species for ecosystem monitoring. Black-browed and Campbell albatross scats were collected from eight breeding colonies covering the circumpolar range of these birds over two consecutive breeding seasons. Fish was the main dietary item at most sites; however, cnidarian DNA, primarily from scyphozoan jellyfish, was present in 42% of samples overall and up to 80% of samples at some sites. Jellyfish was detected during all breeding stages and consumed by adults and chicks. Trawl fishery catches of jellyfish near the Falkland Islands indicate a similar frequency of jellyfish occurrence in albatross diets in years of high and low jellyfish availability, suggesting jellyfish consumption may be selective rather than opportunistic. Warmer oceans and overfishing of finfish are predicted to favour jellyfish population increases, and we demonstrate here that dietary DNA metabarcoding enables measurements of the contribution of gelatinous zooplankton to the diet of marine predators.

Fungi play a key role in soil-plant interactions, nutrient cycling and carbon flow and are essential for the functioning of arctic terrestrial ecosystems. Some studies have shown that the composition of fungal communities is highly sensitive to variations in environmental conditions, but little is known about how the conditions control the role of fungal communities (i.e., their ecosystem function). We used DNA metabarcoding to compare taxonomic and functional composition of fungal communities along a gradient of environmental severity in Northeast Greenland. We analysed soil samples from fell fields, heaths and snowbeds, three habitats with very contrasting abiotic conditions. We also assessed within-habitat differences by comparing three widespread microhabitats (patches with high cover of Dryas, Salix, or bare soil). The data suggest that, along the sampled mesotopographic gradient, the greatest differences in both fungal richness and community composition are observed amongst habitats, while the effect of microhabitat is weaker, although still significant. Furthermore, we found that richness and community composition of fungi are shaped primarily by abiotic factors and to a lesser, though still significant extent, by floristic composition. Along this mesotopographic gradient, environmental severity is strongly correlated with richness in all fungal functional groups: positively in saprotrophic, pathogenic and lichenised fungi, and negatively in ectomycorrhizal and root endophytic fungi. Our results suggest complex interactions amongst functional groups, possibly due to nutrient limitation or competitive exclusion, with potential implications on soil carbon stocks. These findings are important in the light of the environmental changes predicted for the Arctic.

Since the advent of DNA metabarcoding surveys, the planktonic realm is considered a treasure trove of diversity, inhabited by a small number of abundant taxa, and a hugely diverse and taxonomically uncharacterized consortium of rare species. Here we assess if the apparent underestimation of plankton diversity applies universally. We target planktonic foraminifera, a group of protists whose known morphological diversity is limited, taxonomically resolved and linked to ribosomal DNA barcodes. We generated a pyrosequencing dataset of ~100,000 partial 18S rRNA foraminiferal sequences from 32 size fractioned photic-zone plankton samples collected at 8 stations in the Indian and Atlantic Oceans during the Tara Oceans expedition (2009-2012). We identified 69 genetic types belonging to 41 morphotaxa in our metabarcoding dataset. The diversity saturated at local and regional scale as well as in the three size fractions and the two depths sampled indicating that the diversity of foraminifera is modest and finite. The large majority of the newly discovered lineages occur in the small size fraction, neglected by classical taxonomy. These unknown lineages dominate the bulk [>0.8 µm] size fraction, implying that a considerable part of the planktonic foraminifera community biomass has its origin in unknown lineages.

Niche partitioning through foraging is a mechanism likely involved in facilitating the coexistence of ecologically similar and co-occurring animal species by separating their use of resources. Yet, this mechanism is not well understood in flying insectivorous animals. This is particularly true of bats, where many ecologically similar or cryptic species coexist. The detailed analysis of the foraging niche in sympatric, cryptic sibling species provides an excellent framework to disentangle the role of specific niche factors likely involved in facilitating coexistence. We used DNA metabarcoding to determine the prey species consumed by a population of sympatric sibling Rhinolophus euryale and R. mehelyi whose use of habitat in both sympatric and allopatric ranges has been well established through radio tracking. Although some subtle dietary differences exist in prey species composition, the diet of both bats greatly overlapped (Ojk = 0.83) due to the consumption of the same common and widespread moths. Those dietary differences we did detect might be related to divergences in prey availabilities among foraging habitats, which prior radio tracking on the same population showed are differentially used and selected when both species co-occur. This minor dietary segregation in sympatry may be the result of foraging on the same prey-types and could contribute to reduce potential competitive interactions (e.g. for prey, acoustic space). Our results highlight the need to evaluate the spatial niche dimension in mediating the co-occurrence of similar insectivorous bat species, a niche factor likely involved in processes of bat species coexistence.

The environmental health of aquatic ecosystems is critical to society, yet traditional assessments of water quality have limited utility for some bodies of water such as large rivers. Sequencing of environmental DNA (eDNA) has the potential to complement if not replace traditional sampling of biotic assemblages for the purposes of reconstructing aquatic assemblages and, by proxy, assessing water quality. Despite this potential, there has been little testing of the ability of eDNA to reconstruct assemblages and their absolute and relative utility to infer water quality metrics. Here, we reconstruct phytoplankton communities by amplifying and sequencing DNA from a portion of the 23S rRNA region from filtered water samples along a 2900-km transect in the Mississippi River. Across the entire length, diatoms dominated the assemblage (72.6%) followed by cryptophytes (8.7%) and cyanobacteria (7.0%). There were no general trends in the abundances of these major taxa along the length of the river, but individual taxon abundance peaked in different regions. For example, the abundance of taxa genetically similar to Melosira tropica peaked at approximately 60% of all reads 2750 km upstream from the Gulf of Mexico, while taxa similar to Skeletonema marinoi began to increase below the confluence with the Missouri River until it reached approximately 30% of the reads at the Gulf of Mexico. There were four main clusters of samples based on phytoplankton abundance, two above the confluence with the Missouri and two below. Phytoplankton abundance was a poor predictor of NH4+ concentrations in the water, but predicted 61% and 80% of the variation in observed NO3- and PO43- concentrations, respectively. Phytoplankton richness increased with increasing distance along the river, but was best explained by phosphate concentrations and water clarity. Along the Mississippi transect, there was similar structure to phytoplankton and bacterial assemblages, indicating that the two sets of organisms are responding to similar environmental factors. In all, the research here demonstrates the potential utility of metabarcoding for reconstructing aquatic assemblages, which might aid in conducting water quality assessments.