Ballast Water Exchange and Invasion Risk Posed by Intracoastal Vessel Traffic: An Evaluation Using High Throughput Sequencing.

Ballast water remains a potent vector of non-native aquatic species introductions, despite increased global efforts to reduce risk of ballast water mediated invasions. This is particularly true of intracoastal vessel traffic, whose characteristics may limit the feasibility and efficacy of management through ballast water exchange (BWE). Here we utilize high throughput sequencing (HTS) to assess biological communities associated with ballast water being delivered to Valdez, Alaska from multiple source ports along the Pacific Coast of the United States. Our analyses indicate that BWE has a significant but modest effect on ballast water assemblages. Although overall richness was not reduced with exchange, we detected losses of some common benthic coastal taxa (e.g., decapods, mollusks, bryozoans, cnidaria) and gains of open ocean taxa (e.g., certain copepods, diatoms, and dinoflagellates), including some potentially toxic species. HTS-based metabarcoding identified significantly differentiated biodiversity signatures from individual source ports; this signal persisted, though weakened, in vessels undergoing BWE, indicating incomplete faunal turnover associated with management. Our analysis also enabled identification of taxa that may be of particular concern if established in Alaskan waters. While these results reveal a clear effect of BWE on diversity in intracoastal transit, they also indicate continued introduction risk of non-native and harmful taxa.

Comparison of Larval Fish Detections Using Morphology-Based Taxonomy versus High-Throughput Sequencing for Invasive Species Early Detection.

When first introduced, invasive species typically evade detection; DNA barcoding coupled with high-throughput sequencing (HTS) may be more sensitive and accurate than morphology-based taxonomy, and thereby improve invasive (or rare) species detection. We quantified the relative error of species detection between morphology-based and HTS-based taxonomic identification of ichthyoplankton collections from the Port of Duluth, Minnesota, an aquatic non-native species introduction 'hot-spot' in the Laurentian Great Lakes. We found HTS-based taxonomy identified 28 species and morphology-based taxonomy 30 species, of which 27 were common to both. Among samples, 76% of family-level taxonomic assignments agreed; however, only 42% of species assignments agreed. Most errors were attributed to morphology-based taxonomy, whereas HTS-based taxonomy error was low. For this study system, for most non-native fishes, the detection probability by randomized survey for larvae was similar to that by a survey that is optimized for non-native species early detection of juveniles and adults. We conclude that classifying taxonomic errors by comparing HTS results against morphology-based taxonomy is an important step toward incorporating HTS-based taxonomy into biodiversity surveys.

Early detection monitoring for non-indigenous fishes; comparison of survey approaches during two species introductions in a Great Lakes port.

Assessing relative performance of different sampling methods used for early detection monitoring (EDM) is a critical step in understanding the likelihood of detecting new non-indigenous species (NIS) in an environment of interest. EDM performance metrics are typically based on the probability of detecting established NIS or rare indigenous species; however, detection probability estimates for these proxies may not accurately reflect survey effectiveness for newly introduced NIS. We used data from three different EDM survey approaches that varied by targeted life-stage (adult-juvenile versus ichthyoplankton), media (physical fish versus environmental DNA), and taxonomic method (morphology-based versus DNA-based taxonomy) to explore relative detection sensitivity for recently introduced white bass (Morone chrysops) and gizzard shad (Dorosoma cepedianum) in the Port of Duluth-Superior, a NIS introduction hot spot within the Laurentian Great Lakes. Detection efficiency, measured by the effort (number of samples) required to achieve 95% probability of detection, differed by EDM approach and species. Also, the relative sensitivity (detection rate) of each survey approach differed by species. For both species, detection in surveys using DNA-based taxonomy was generally as good or better than the adult-juvenile survey using morphology-based taxonomy. While both species appear to have been detected at early stages of invasion, white bass were likely present up to 5 years prior to initial detection, whereas gizzard shad may have been detected in the first year of introduction. We conclude that using complimentary sampling methods can help to balance the strengths and weaknesses of each approach and provide more reliable early detection of new invaders.

Sensitivity and accuracy of high-throughput metabarcoding methods for early detection of invasive fish species.

High-throughput DNA metabarcoding has gained recognition as a potentially powerful tool for biomonitoring, including early detection of aquatic invasive species (AIS). DNA based techniques are advancing, but our understanding of the limits to detection for metabarcoding complex samples is inadequate. For detecting AIS at an early stage of invasion when the species is rare, accuracy at low detection limits is key. To evaluate the utility of metabarcoding in future fish community monitoring programs, we conducted several experiments to determine the sensitivity and accuracy of routine metabarcoding methods. Experimental mixes used larval fish tissue from multiple "common" species spiked with varying proportions of tissue from an additional "rare" species. Pyrosequencing of genetic marker, COI (cytochrome c oxidase subunit I) and subsequent sequence data analysis provided experimental evidence of low-level detection of the target "rare" species at biomass percentages as low as 0.02% of total sample biomass. Limits to detection varied interspecifically and were susceptible to amplification bias. Moreover, results showed some data processing methods can skew sequence-based biodiversity measurements from corresponding relative biomass abundances and increase false absences. We suggest caution in interpreting presence/absence and relative abundance in larval fish assemblages until metabarcoding methods are optimized for accuracy and precision.

An examination of fitness costs of glyphosate resistance in the common morning glory, Ipomoea purpurea.

Fitness costs are frequently invoked to explain the presence of genetic variation underlying plant defense across many types of damaging agents. Despite the expectation that costs of resistance are prevalent, however, they have been difficult to detect in nature. To examine the potential that resistance confers a fitness cost, we examined the survival and fitness of genetic lines of the common morning glory, Ipomoea purpurea, that diverged in the level of resistance to the herbicide glyphosate. We planted a large field experiment and assessed survival following herbicide application as well as fitness of the divergent selection lines in the absence of the herbicide. We found that genetic lines selected for increased resistance exhibited lower death compared to control and susceptible lines in the presence of the herbicide, but no evidence that resistant lines produced fewer seeds in the absence of herbicide. However, susceptible lines produced more viable seeds than resistant or control lines, providing some evidence of a fitness cost in terms of seed germination, and thus potential empirical support for the expectation of trait trade-offs as a consequence of adaptation to novel environments.