Matthew A. Barnes

Environmental Conditions Influence eDNA Persistence in Aquatic Systems

Environmental DNA (eDNA) surveillance holds great promise for improving species conservation and management. However, few studies have investigated eDNA dynamics under natural conditions, and interpretations of eDNA surveillance results are clouded by uncertainties about eDNA degradation. We conducted a literature review to assess current understanding of eDNA degradation in aquatic systems and an experiment exploring how environmental conditions can influence eDNA degradation. Previous studies have reported macrobial eDNA persistence ranging from less than 1 day to over 2 weeks, with no attempts to quantify factors affecting degradation. Using a SYBR Green quantitative PCR assay to observe Common Carp ( Cyprinus carpio ) eDNA degradation in laboratory mesocosms, our rate of Common Carp eDNA detection decreased over time. Common Carp eDNA concentration followed a pattern of exponential decay, and observed decay rates exceeded previously published values for aquatic macrobial eDNA. Contrary to our expectations, eDNA degradation rate declined as biochemical oxygen demand, chlorophyll, and total eDNA (i.e., from any organism) concentration increased. Our results help explain the widely divergent, previously published estimates for eDNA degradation. Measurements of local environmental conditions, consideration of environmental influence on eDNA detection, and quantification of local eDNA degradation rates will help interpret future eDNA surveillance results.

Conservation in a cup of water: estimating biodiversity and population abundance from environmental DNA

Three mantras often guide species and ecosystem management: (i) for preventing invasions by harmful species, ‘early detection and rapid response’; (ii) for conserving imperilled native species, ‘protection of biodiversity hotspots’; and (iii) for assessing biosecurity risk, ‘an ounce of prevention equals a pound of cure.’ However, these and other management goals are elusive when traditional sampling tools (e.g. netting, traps, electrofishing, visual surveys) have poor detection limits, are too slow or are not feasible. One visionary solution is to use an organism’s DNA in the environment (eDNA), rather than the organism itself, as the target of detection. In this issue of Molecular Ecology, Thomsen et al. (2012) provide new evidence demonstrating the feasibility of this approach, showing that eDNA is an accurate indicator of the presence of an impressively diverse set of six aquatic or amphibious taxa including invertebrates, amphibians, a fish and a mammal in a wide range of freshwater habitats. They are also the first to demonstrate that the abundance of eDNA, as measured by qPCR, correlates positively with population abundance estimated with traditional tools. Finally, Thomsen et al. (2012) demonstrate that next‐generation sequencing of eDNA can quantify species richness. Overall, Thomsen et al. (2012) provide a revolutionary roadmap for using eDNA for detection of species, estimates of relative abundance and quantification of biodiversity.

The ecology of environmental DNA and implications for conservation genetics

Environmental DNA (eDNA) refers to the genetic material that can be extracted from bulk environmental samples such as soil, water, and even air. The rapidly expanding study of eDNA has generated unprecedented ability to detect species and conduct genetic analyses for conservation, management, and research, particularly in scenarios where collection of whole organisms is impractical or impossible. While the number of studies demonstrating successful eDNA detection has increased rapidly in recent years, less research has explored the “ecology” of eDNA—myriad interactions between extraorganismal genetic material and its environment—and its influence on eDNA detection, quantification, analysis, and application to conservation and research. Here, we outline a framework for understanding the ecology of eDNA, including the origin, state, transport, and fate of extraorganismal genetic material. Using this framework, we review and synthesize the findings of eDNA studies from diverse environments, taxa, and fields of study to highlight important concepts and knowledge gaps in eDNA study and application. Additionally, we identify frontiers of conservation-focused eDNA application where we see the most potential for growth, including the use of eDNA for estimating population size, population genetic and genomic analyses via eDNA, inclusion of other indicator biomolecules such as environmental RNA or proteins, automated sample collection and analysis, and consideration of an expanded array of creative environmental samples. We discuss how a more complete understanding of the ecology of eDNA is integral to advancing these frontiers and maximizing the potential of future eDNA applications in conservation and research.

Fecundity of the exotic applesnail, Pomacea insularum

Abstract International trade frequently moves mollusks around the globe, thereby increasing their opportunity to cause ecological and economic damage. Recent genetic studies have confirmed the identity of South American applesnails (Pomacea insularum) in the southeastern US, but limited literature exists on this species. Understanding fecundity provides direct insight into the invasive potential of mollusks. Our study documents P. insularum fecundity in Texas and offers comparisons with the closely related global invader P. canaliculata. We quantified P. insularum clutch and hatchling physical characteristics and examined field and laboratory hatching success. Clutches contained thousands of eggs (mean = 2064 eggs), and clutch size tended to increase over the reproductive season. Clutches exhibited average field and laboratory hatching efficiencies (number of hatchlings/total number of eggs) near 70 and 30%, respectively. Notably, several clutches hatched at 80% or higher in the field. Exotic P. insularum deposit more eggs/clutch than the related invader P. canaliculata, but we do not yet know how seasonal hatching efficiencies compare. However, even with a conservative estimate of 1 to 10% survival to adulthood, the average P. insularum clutch yields 14 to 144 new applesnails. The high fecundity of P. insularum translates into considerable ecological impact because adult females can contribute >1 clutch/wk over an extended growing season in the southeastern US. The need for research has increased with the emergence of P. insularum populations in the fragile Florida Everglades. We advocate life-history studies like ours to help understand the invasion potential of applesnails and other invasive mollusks.

Molecular Detection of Invasive Species in Heterogeneous Mixtures Using a Microfluidic Carbon Nanotube Platform

Screening methods to prevent introductions of invasive species are critical for the protection of environmental and economic benefits provided by native species and uninvaded ecosystems. Coastal ecosystems worldwide remain vulnerable to damage from aquatic species introductions, particularly via ballast water discharge from ships. Because current ballast management practices are not completely effective, rapid and sensitive screening methods are needed for on-site testing of ships in transit. Here, we describe a detection technology based on a microfluidic chip containing DNA oligonucleotide functionalized carbon nanotubes. We demonstrate the efficacy of the chip using three ballast-transported species either established (Dreissena bugensis) or of potential threat (Eriocheir sinensis and Limnoperna fortuneii) to the Laurentian Great Lakes. With further refinement for on-board application, the technology could lead to real-time ballast water screening to improve ship-specific management and control decisions.

Viability of Aquatic Plant Fragments following Desiccation

Abstract Desiccation following prolonged air exposure challenges survival of aquatic plants during droughts, water drawdowns, and overland dispersal. To improve predictions of plant response to air exposure, we observed the viability of vegetative fragments of 10 aquatic plant species (Cabomba caroliniana, Ceratophyllum demersum, Elodea canadensis, Egeria densa, Myriophyllum aquaticum, Myriophyllum heterophyllum, Myriophyllum spicatum, Potamogeton crispus, Potamogeton richardsonii, and Hydrilla verticillata) following desiccation. We recorded mass loss, desiccation rate, and plant fragment survival across a range of air exposures. Mass loss accurately predicted viability of aquatic plant fragments upon reintroduction to water. However, similar periods of air exposure differentially affected viability between species. Understanding viability following desiccation can contribute to predicting dispersal, improving eradication protocols, and disposing of aquatic plants following removal from invaded lakes or contaminated equipment. Nomenclature: Brazilian egeria, Egeria densa Planch., common elodea, Elodea canadensis Michx., coontail, Ceratophyllum demersum L., curlyleaf pondweed, Potamogeton crispus L., Eurasian watermilfoil, Myriophyllum spicatum L., fanwort, Cabomba caroliniana Gray, hydrilla, Hydrilla verticillata (L. f.) Royle, parrotfeather, Myriophyllum aquaticum (Vell.) Verdc., Richardsons pondweed, Potamogeton richardsonii (A. Bennett) Rydb., variable-leaf watermilfoil, Myriophyllum heterophyllum Michx Management Implications: A framework for assessing the vulnerability of ecosystems to invasion by aquatic weeds must consider many aspects of species invasions: which species will arrive, how will they get there, and will they establish and generate impacts following introduction. Knowledge about physiological responses to stressors provides critical input to such a framework. In our study, we compared the viability of vegetative fragments of 10 aquatic plant species following variable periods of desiccation. We found that while desiccation expectedly decreased plant viability, desiccation rates and tolerances differed significantly between plant species. The species-specific nature of desiccation warrants species-specific management actions. Our results suggest that boat launch inspection programs should be extra vigilant in their search for species that are relatively desiccation-resistant or –tolerant, such as emergent Myriophyllum aquaticum. On the other hand, our finding that some common invasive plants (Ceratophyllum demersum, Hydrilla verticillata) are comparatively intolerant to air exposure suggests that for these species, greater attention should be given to specific situations that promote insulation of vegetative material (e.g., entrainment in nets or anchor wells, burial in sediments), plant propagules that are more tolerant or resistant to desiccation (e.g., seeds or tubers), and alternative invasion pathways (e.g., water garden contaminants).

Quantitative and Rapid DNA Detection by Laser Transmission Spectroscopy

Laser transmission spectroscopy (LTS) is a quantitative and rapid in vitro technique for measuring the size, shape, and number of nanoparticles in suspension. Here we report on the application of LTS as a novel detection method for species-specific DNA where the presence of one invasive species was differentiated from a closely related invasive sister species. The method employs carboxylated polystyrene nanoparticles functionalized with short DNA fragments that are complimentary to a specific target DNA sequence. In solution, the DNA strands containing targets bind to the tags resulting in a sizable increase in the nanoparticle diameter, which is rapidly and quantitatively measured using LTS. DNA strands that do not contain the target sequence do not bind and produce no size change of the carboxylated beads. The results show that LTS has the potential to become a quantitative and rapid DNA detection method suitable for many real-world applications.

Geographic selection bias of occurrence data influences transferability of invasive Hydrilla verticillata distribution models

Due to socioeconomic differences, the accuracy and extent of reporting on the occurrence of native species differs among countries, which can impact the performance of species distribution models. We assessed the importance of geographical biases in occurrence data on model performance using Hydrilla verticillata as a case study. We used Maxent to predict potential North American distribution of the aquatic invasive macrophyte based upon training data from its native range. We produced a model using all available native range occurrence data, then explored the change in model performance produced by omitting subsets of training data based on political boundaries. We also compared those results with models trained on data from which a random sample of occurrence data was omitted from across the native range. Although most models accurately predicted the occurrence of H. verticillata in North America (AUC > 0.7600), data omissions influenced model predictions. Omitting data based on political boundaries resulted in larger shifts in model accuracy than omitting randomly selected occurrence data. For well-documented species like H. verticillata, missing records from single countries or ecoregions may minimally influence model predictions, but for species with fewer documented occurrences or poorly understood ranges, geographic biases could misguide predictions. Regardless of focal species, we recommend that future species distribution modeling efforts begin with a reflection on potential spatial biases of available occurrence data. Improved biodiversity surveillance and reporting will provide benefit not only in invaded ranges but also within under-reported and unexplored native ranges.

Confronting species distribution model predictions with species functional traits

Abstract Species distribution models are valuable tools in studies of biogeography, ecology, and climate change and have been used to inform conservation and ecosystem management. However, species distribution models typically incorporate only climatic variables and species presence data. Model development or validation rarely considers functional components of species traits or other types of biological data. We implemented a species distribution model (Maxent) to predict global climate habitat suitability for Grass Carp (Ctenopharyngodon idella). We then tested the relationship between the degree of climate habitat suitability predicted by Maxent and the individual growth rates of both wild (N = 17) and stocked (N = 51) Grass Carp populations using correlation analysis. The Grass Carp Maxent model accurately reflected the global occurrence data (AUC = 0.904). Observations of Grass Carp growth rate covered six continents and ranged from 0.19 to 20.1 g day−1. Species distribution model predictions were correlated (r = 0.5, 95% CI (0.03, 0.79)) with observed growth rates for wild Grass Carp populations but were not correlated (r = −0.26, 95% CI (−0.5, 0.012)) with stocked populations. Further, a review of the literature indicates that the few studies for other species that have previously assessed the relationship between the degree of predicted climate habitat suitability and species functional traits have also discovered significant relationships. Thus, species distribution models may provide inferences beyond just where a species may occur, providing a useful tool to understand the linkage between species distributions and underlying biological mechanisms.

DNA-based species detection capabilities using laser transmission spectroscopy.

Early detection of invasive species is critical for effective biocontrol to mitigate potential ecological and economic damage. Laser transmission spectroscopy (LTS) is a powerful solution offering real-time, DNA-based species detection in the field. LTS can measure the size, shape and number of nanoparticles in a solution and was used here to detect size shifts resulting from hybridization of the polymerase chain reaction product to nanoparticles functionalized with species-specific oligonucleotide probes or with the species-specific oligonucleotide probes alone. We carried out a series of DNA detection experiments using the invasive freshwater quagga mussel (Dreissena bugensis) to evaluate the capability of the LTS platform for invasive species detection. Specifically, we tested LTS sensitivity to (i) DNA concentrations of a single target species, (ii) the presence of a target species within a mixed sample of other closely related species, (iii) species-specific functionalized nanoparticles versus species-specific oligonucleotide probes alone, and (iv) amplified DNA fragments versus unamplified genomic DNA. We demonstrate that LTS is a highly sensitive technique for rapid target species detection, with detection limits in the picomolar range, capable of successful identification in multispecies samples containing target and non-target species DNA. These results indicate that the LTS DNA detection platform will be useful for field application of target species. Additionally, we find that LTS detection is effective with species-specific oligonucleotide tags alone or when they are attached to polystyrene nanobeads and with both amplified and unamplified DNA, indicating that the technique may also have versatility for broader applications.