High throughput shotgun sequencing of eRNA reveals taxonomic and derived functional shifts across a benthic productivity gradient

Abstract

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The Promise and Pitfalls of Environmental DNA and RNA Approaches for the Monitoring of Human and Animal Pathogens from Aquatic Sources

“…eRNA analysis can also be used simultaneously to complement eDNA-based approaches (Zaiko et al 2018 , Merou et al 2020 ). In recent studies of marine fanworms ( Sabella spallanzanii ) and bioindicator microeukaryote groups such as nematodes, the combination of eDNA analysis with high-throughput shotgun sequencing of eRNA has helped to discriminate between “contemporary intracellular (living) and extracellularly persistent (legacy) genetic fragments” (von Ammon et al 2019 , p. 1, Broman et al 2020 ). The analysis of eRNA provides discriminating data between viable and inviable cells, and more accurate data concerning the presence and spread of pathogens in aquatic environments (Goldberg et al 2016 , Broman et al 2020 , Merou et al 2020 ).…”

“…In recent studies of marine fanworms ( Sabella spallanzanii ) and bioindicator microeukaryote groups such as nematodes, the combination of eDNA analysis with high-throughput shotgun sequencing of eRNA has helped to discriminate between “contemporary intracellular (living) and extracellularly persistent (legacy) genetic fragments” (von Ammon et al 2019 , p. 1, Broman et al 2020 ). The analysis of eRNA provides discriminating data between viable and inviable cells, and more accurate data concerning the presence and spread of pathogens in aquatic environments (Goldberg et al 2016 , Broman et al 2020 , Merou et al 2020 ). Single-species qPCR-based eRNA approaches have already been successfully used in numerous wildlife health studies, including the detection and quantification of a protozoan parasite ( Bonamia ostreae ) in seawater (Merou et al 2020 ).…”

“…For each particular study, extensive evaluation of study design should be considered to ensure the cost effectiveness of an eDNA- or eRNA-based approach as opposed to traditional approaches, because, as the current costs stand, on the basis of study design, eDNA-based methods can be more expensive than traditional methods and eRNA-based approaches are significantly more expensive than eDNA-based approaches (Smart et al 2016 , Broman et al 2020 , Winkworth et al 2020 ). However, the continued rapid fall in high-throughput sequencing costs, increased multiplexing potential of qPCR or ddPCR (which partitions the sample before amplification, allowing for rare or low-abundance detection), and the development of more cost-effective and resource-conscious methodologies such as LAMP-based assays is bridging these economic gaps (Smart et al 2016 , Broman et al 2020 , Winkworth et al 2020 ). As an example, it has been estimated that when detecting New Zealand mudsnails, an invasive species in the United States, it is four to eight times less expensive to use qPCR-based eDNA approaches—at US$35–$80 per sample—than traditional approaches (collecting, sorting, and identifying samples) that cost US$300 per sample (Goldberg et al 2013 ).…”

“…Current practices rely on single species from a designated habitat to assign scores to infer a community-based assessment—as opposed to using biodiversity to track changes in populations (Seymour et al 2020 ). It will be important for future effective biomonitoring to incorporate traditional community biomonitoring scores with molecular-informed population-based assessments, such as eDNA metabarcoding and eRNA shotgun sequencing, in order to incorporate a wide range of taxa that is traditionally overlooked (Broman et al 2020 , Seymour et al 2020 ). Therefore, methodologies originating from the study of microbial diversity—and, subsequently, megafauna presence and diversity—are now being refocused on disease-causing microbes of animals and humans.…”

“…An alternative environmental genetic detection tool is the analysis of eRNA, a method that uses similar detection approaches to the analysis of eDNA but that, instead, is targeted against RNA molecules (transcripts). Environmental RNA-based approaches have helped to discriminate between “contemporary intracellular (living or recently shed) and extracellularly persistent (legacy) genetic fragments” (von Ammon et al 2019 , Broman et al 2020 ). Environmental RNA analysis has proven to be a reliable predictor for both living species and viral pathogens in aquatic habitats and is the accurate and efficient methodology applied in our second case study, the COVID-19 pandemic-related early viral detection from human wastewater (Kumar et al 2020 , Randazzo et al 2020 , Wu et al 2020 ).…”