Development of a quantitative PCR assay for detecting Egeria densa in environmental DNA samples
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TECHNICAL NOTE
Development of a quantitative PCR assay for detecting Egeria densa in environmental DNA samples Dorothy M. Chase1 · Lauren M. Kuehne2 · Julian D. Olden2 · Carl O. Ostberg1 Received: 26 September 2019 / Accepted: 14 May 2020 © This is a U.S. Government work and not under copyright protection in the US; foreign copyright protection may apply 2020
Abstract Brazilian elodea (Egeria densa) is an invasive freshwater plant that demonstrates widespread ecological impacts in freshwater ecosystems and causes substantial economic damage. Here, we developed an environmental DNA assay for detection of E. densa to provide resource managers with a tool for early detection, identification, and monitoring of invasive populations. Keywords Egeria densa · eDNA · Environmental DNA · Invasive aquatic plants · Quantitative PCR · qPCR Brazilian elodea (Egeria densa) is a macrophyte native to South America. Their introduction to numerous regions of the world has led to infestations that negatively impacted aquatic ecosystems (de Winton and Clayton 1996; Roberts et al. 1999; Lund et al. 2007; Santos et al. 2011) and disrupted ecosystem function and services, causing economic impacts (Hussner et al. 2017; Yarrow et al. 2009). Early detection of invasive species and understanding of their distribution is essential to preventing further spread (Vander Zanden and Olden 2008; Hussner et al. 2017). Traditional methods for surveying invasive aquatic plants can be time consuming and costly. Surveys of aquatic environmental DNA (eDNA) are an alternative that can provide improved detectability relative to traditional field survey methods (Ficetola et al. 2008; Jerde et al. 2011; Larson et al. 2020) while requiring less effort (Biggs et al.; 2015; Sigsgaard et al. 2015; Evans et al. 2017). Here, we developed and validated an eDNA assay as a tool for early detection of E. densa in aquatic environments.
Electronic supplementary material The online version of this article (https://doi.org/10.1007/s12686-020-01152-w) contains supplementary material, which is available to authorized users. * Dorothy M. Chase [email protected] 1
U.S. Geological Survey, Western Fisheries Research Center, Seattle, WA 98115, USA
School of Aquatic and Fishery Sciences, University of Washington, Seattle, WA 98195, USA
2
To develop the E. densa assay, we retrieved DNA sequences for several genes from E. densa and Elodea native to North America (Elodea canadensis, Elodea nuttallii, and Elodea bifoliate) from GenBank (Supplemental File). Sequences were aligned using MEGA 7.0.21 (Kumar et al. 2016) and surveyed for sequence variation. The ITS1-5.8sITS2 region was selected for assay development because it contained many candidates for comparison and many interspecific, single nucleotide polymorphisms (SNPs). The E. densa assay (EdITS1) was developed using the PrimerQuest Tool (Integrated DNA Technologies). The assay amplifies 79 bases of the internal transcribed spacer-1 (ITS1) using primers (F: 5′-GGTCAATGGCAATTCCTTCTTG-3′; R: 5′-GCGCACCACC
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