Simple and efficient preservation of fish environmental RNA in filtered water samples via RNAlater

Jo, Toshiaki; Matsuda, Nao; Hirohara, Takaya; Yamanaka, Hiroki

doi:10.21203/rs.3.rs-2170577/v1

Cited by 2 publications

(2 citation statements)

References 53 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…The general assumption has been that the eRNA released by macroorganisms into the environment is labile and degrades too rapidly to be reliably detected or quantified (but see Cristescu, 2019). However, the RNA shed by macroorganisms into their surrounding environment can be successfully extracted and quantified (T. Jo et al, 2022; Kagzi et al, 2022; Littlefair et al, 2022; Marshall et al, 2021; Miyata et al, 2021; Tsuri et al, 2021; von Ammon et al, 2019; Wood et al, 2020). Despite this evidence supporting the extractability of eRNA, environmental transcriptomics, defined here as eRNA based gene expression profiling, remained untested.…”

Section: Introductionmentioning

confidence: 99%

Environmental transcriptomics under heat stress: Can environmental RNA reveal changes in gene expression of aquatic organisms?

Hechler

Yates

Chain

et al. 2022

Preprint

View full text Add to dashboard Cite

To safeguard biodiversity in a changing climate, we require taxonomic information about species turnover and insights into the health of organisms. Environmental DNA approaches are increasingly used for species identification, but cannot provide functional insights. Transcriptomic methods reveal the physiological states of macroorganisms, but are currently species specific and require tissue sampling or animal sacrifice, making community-wide assessments challenging. Here, we test if broad functional information (activity level of the transcribed genes) can be harnessed from environmental RNA (eRNA), which includes extra-organismal RNA from macroorganisms along with whole microorganisms. We exposed Daphnia pulex as well as phytoplankton prey and microorganism colonizers to control (20 °C) and heat stress (28 °C) conditions for seven days. We sequenced eRNA from tank water (after complete removal of Daphnia) as well as RNA from Daphnia tissue, enabling comparisons of extra-organismal and organismal RNA based gene expression profiles. Both RNA types detected similar gene expression responses. Using eRNA, we identified 42 Daphnia genes to be differentially expressed following heat stress. Of these, 62% were also differentially expressed and exhibited similar levels of relative expression in organismal RNA. Both RNA types recovered similar Daphnia heat stress responses via gene ontology terms. In addition to the extra-organismal Daphnia response, eRNA detected community-wide heat stress responses consisting of 199 differentially expressed genes across 9 taxa, and distinct functional profiles via KEGG orthologs. Our study demonstrates that environmental transcriptomics based on eRNA can non-invasively reveal gene expression responses of macroorganisms following environmental changes, with broad potential implications for the biomonitoring of ecological health across the trophic chain.

show abstract

Section: Introductionmentioning

confidence: 99%

Environmental transcriptomics under heat stress: Can environmental RNA reveal changes in gene expression of aquatic organisms?

Hechler

Yates

Chain

et al. 2022

Preprint

View full text Add to dashboard Cite

show abstract

“…However, the current filter methods to detect environmental RNAs in water can mainly measure RNAs remaining in the filters, neglecting dissolved RNAs in the filtrates. Filters used for filtering water samples to measure environmental RNAs have different pore sizes, such as 0.2~0.22 µm (Wu and Liu, 2018;Wood et al, 2020;Hempel et al, 2022), 0.45 µm (von Ammon et al, 2019Tsuri et al, 2020;Miyata et al, 2021;Zaiko et al, 2022;Jo et al, 2022b;Miyata et al, 2022), 0.7 µm (Kagzi et al, 2022;Hechler et al, 2022;Jo et al, 2022a;Littlefair et al, 2022), 1.2 µm (Marshall et al, 2021;Zaiko et al, 2022), 1.6 µm (Pochon et al, 2017), and 5 µm (Zaiko et al, 2022). A study indicated that filters with different pore sizes could collect naked RNAs (Zaiko et al, 2022), but the RNA amounts in the filtrates were unknown in that study.…”

Section: Introductionmentioning

confidence: 99%

Degradation and release of dissolved environmental RNAs from zebrafish cells

2023

Preprint

View full text Add to dashboard Cite

Environmental RNAs in water are gradually being applied in aquatic ecological surveys, water pollution monitoring, etc., but the current methods to detect environmental RNAs in water can mainly measure the RNAs in the filters that are used for filtering water samples, neglecting dissolved environmental RNAs in water. The sources and degradation profiles of dissolved environmental RNAs in water remain unknown. The present study was conducted to measure the permeability of extracted RNAs from zebrafish cells through filters, the degradation of extracted RNAs from zebrafish cells in tubes, and the release rate and degradation of dissolved environmental RNAs from living zebrafish cells and dying zebrafish cells, aiming to provide dynamic information from dissolved environmental RNAs in water. The results showed that there were no significant differences between the levels of extracted RNAs from zebrafish cells before filtration with 0.45 um filters and those in the filtrates. The extracted RNAs from zebrafish cells degraded in water in the tubes, and after 2 months, more than 15% of RNAs in the groups of RNAs in water were still detected. The half-life of all the RNAs in the tubes was approximately 20~43 days. During the 6-day experiment of the release and degradation of dissolved RNAs from living cells, an average of 0.00041~0.0017 pg dissolved RNAs (760000~3200000 RNA bases) were secreted per cell per day into the liquid environment. During the 6-day experiment of the release and degradation of dissolved RNAs from dying cells, approximately 4.2 pg of dissolved RNAs released by a dying zebrafish cell in water could be detected. The dissolved environmental RNAs in water from zebrafish cells degraded faster in the presence of zebrafish cells: under the conditions without zebrafish cells, the average survival rate of the dissolved environmental RNAs in water per day was 98.4%/day; under the conditions with living zebrafish cells, the average survival rate per day was 49.7%/day; and under the conditions with dying zebrafish cells, the average survival rate per day was 34.9%/day. The estimated levels of dissolved environmental RNAs in water in fish tanks were too low to be detected by the current techniques. Although the methods in the present study need to be improved, this study may provide information to develop new ways to measure the dynamics of dissolved environmental RNAs in water and quantitatively analyze RNAs released into liquid environments of living and dying cells.

show abstract

Simple and efficient preservation of fish environmental RNA in filtered water samples via RNAlater

Cited by 2 publications

References 53 publications

Environmental transcriptomics under heat stress: Can environmental RNA reveal changes in gene expression of aquatic organisms?

Environmental transcriptomics under heat stress: Can environmental RNA reveal changes in gene expression of aquatic organisms?

Degradation and release of dissolved environmental RNAs from zebrafish cells

Contact Info

Product

Resources

About