Invasive Dreissena Mussel Coastal Transport From an Already Invaded Estuary to a Nearby Archipelago Detected in DNA and Zooplankton Surveys.

Coastal waters of Lake Superior are generally inhospitable to the establishment of invasive Dreissena spp. mussels (both Dreissena polymorpha and Dreissena bugensis). Dreissena have inhabited the Saint Louis River estuary (SLRE; largest commercial port in the Laurentian Great Lakes) for over three decades, but only in the last few years have small colonies been found in the Apostle Islands National Lakeshore (APIS, an archipelago situated 85 km to the east of SLRE) A 2017 survey determined a low abundance Dreissena spatial distribution in APIS, with the largest colonies on the north and west islands which suggested potential veliger transport from the SLRE via longshore currents. Our objective in this study was to determine if Dreissena veligers are transported by currents at low densities along the south shore of Lake Superior from the SLRE to APIS. To do so, we used both eDNA (water and passive substrate samples) and zooplankton collection methods at eight sites evenly spaced between the SLRE and APIS with three sampling times over five weeks. Dreissena veligers were consistently detected along the south shore, although at low abundances (veligers per m3 range = 0-690, median = 8), and for every 1 km increase in distance from the SLRE, both veliger counts and water eDNA copy numbers decreased on average by 5 and 7%, respectively. D. polymorpha (suited to estuary habitats) was detected two times more than D. bugensis (better suited to deep-lake habitats). There was not a trend in the veliger size distribution along the south shore, and temperature and calcium concentrations fluctuated around the threshold for Dreissena veliger and adult development, averaging 11.0°C and 14.8 ppm, respectively. Three zooplankton taxa representative of the estuary community-Daphnia retrocurva, Diaphanosoma birgei, and Mesocyclops copepodites-decreased as the distance from the SLRE increased mirroring Dreissena veliger abundance patterns. Findings represent multiple sources of evidence of a propagule "conveyor belt" for Dreissena along the south shore of Lake Superior. We conclude that veligers are functioning as a propagule, using coastal currents to spread from the point of invasion, thereby traversing coastal habitat previously reported as inhospitable to distant habitats suitable for colonization.

Experimental paradigm for in-laboratory proxy aquatic studies under conditions of static, non-flow-through chemical exposures.

Endocrine-disrupting chemicals (EDCs) such as 17α-ethynylestradiol, 17ß-estradiol, estrone, and para-nonylphenol have been measured in wastewater-treatment plant effluents, surface waters, sediments, and sludge and have been shown to induce liver-specific vitellogenin (vtg) messenger RNA in male fathead minnows (Pimephales promelas). The purpose of the present study was to establish minimal concentrations of select EDCs necessary to induce transcription of vtg in 48-h static renewal exposures, as measured by quantitative real-time thermal cycle amplification. Adult males were exposed to 17α-ethynylestradiol, 17ß-estradiol, estrone, and para-nonylphenol. Dose-dependent increases in vtg expression were significant with all chemicals tested. The lowest concentrations of these chemicals to induce measurable vtg expression, with significant difference from respective controls, were 17α-ethynylestradiol, 2.2 ng L(-1); para-nonylphenol, 13.9 µg L(-1); 17ß-estradiol, 42.7 ng L(-1); and estrone, 46.7 ng L(-1), measured as 48-h average concentrations. The present experiments were designed to frame a commonly acceptable approach for investigators who conduct static, in-laboratory proxy environmental aquatic exposures. The present study highlights the need for investigators to report in peer-reviewed submissions the observed concentration values for minimal induction levels when measuring molecular responses to chemical exposures by means of real-time polymerase chain reaction, quantitative polymerase chain reaction, or other "omic" technologies.

17alpha-ethynylestradiol-induced vitellogenin gene transcription quantified in livers of adult males, larvae, and gills of fathead minnows (Pimephales promelas).

We have applied a method for quantifying relative levels of messenger RNA (mRNA) transcription to assess chemically induced gene expression in fathead minnows (Pimephales promelas). Synthetic oligonucleotides designed for the fathead minnow vitellogenin gene transcription product were used in a reverse transcription polymerase chain reaction (RT-PCR) protocol. This sensitive and rapid strategy detected vitellogenin gene transcription in livers of male fathead minnows exposed to concentrations as low as 2 ng/L of the endocrine-disrupting compound 17alpha-ethynylestradiol for 24 h. Surprisingly, vitellogenin transcription products also were detected in gill tissue and in 48-h-old posthatch fathead minnow larvae. Relative levels of vitellogenin gene induction among individuals were quantified in a single-step reaction (PCR multiplex) with 18S rRNA universal primers and Competimers concurrently with fathead minnow vitellogenin oligonucleotides. This quantitative approach will markedly enhance detection of the first cellular event of estrogenic exposure to aquatic ecosystems in both field and laboratory systems. Use of the model provides sensitivity of detection at a concentration below those that cause mortality or visible signs of stress in fish or other aquatic organisms. The model may also provide an in vivo screening method for estrogenlike endocrine-disrupting compounds.