- Background: Lake Baikal is one of the oldest freshwater lakes and has constituted a stable environment for millions of years, in stark contrast to small, transient bodies of water in its immediate vicinity. - 80% of the more than 2500 so far described animal species are endemics [2]. - Full list of author information is available at the end of the article. - independent evolution of the Baikal endemics [3, 4] that may have highly specialized adaptations to the specific environmental conditions of Lake Baikal, as it is ultra- oligotrophic with low levels of ions and dissolved organic carbon, has high oxygen levels and the low average tem- perature of 6 °C in the littoral zone [5–7]. - Ponds in the direct vicinity of Baikal and isolated waters at the shores of the lake are inhabited by species of the Euro-Siberian (Palearctic) fauna, which is completely distinct from the fauna of the open Lake Baikal [1].. - 2019 Open Access This article is distributed under the terms of the Creative Commons Attribution 4.0 International License (http://creativecommons.org/licenses/by/4.0. - The environmental conditions in the ponds in the vicin- ity of Baikal and the isolated bays are in many regards highly different to those of the open littoral zone of Lake Baikal, including higher seasonal and daily fluctuations of the water temperature, higher levels of organic substances in those shallow waters, as well as different hydrodynamic regimes [1, 9, 10]. - lacus- tris and the most common in the upper littoral of Baikal endemic gammarids Eulimnogammarus verrucosus (Gerstfeldt, 1858) and E. - with heavy metals, can change the environmen- tal condition of the Baikal littoral zone. - lacustris over the Eulimnogam- marus species in the changing environment. - We explored the transcriptomes of individuals of the three species upon acclimation to pristine Lake Baikal conditions, i.e., Baikal water at 6 °C and from short term (3–24 h) species-specific treatments with cadmium and elevated temperature.. - In order to obtain references for the downstream dif- ferential expression analyses, we performed de novo transcriptome assembly for each of the species. - Technical characteristics of the assemblies are shown in Additional file 1: Table S1.. - This method reliably identified and taxonomically assigned 25% of the transcripts. - of the different transcript sequences and about 25% of the overall expression, the closest matches were meta- zoan proteins. - As we could not interpret the unannotated transcripts, we did not include them into this reduced set, even though they may rep- resent sequences highly specific for this group of the organisms.. - Intra- and inter-species variability of transcript abundance For an estimate to which degree transcript levels differ in all three species and to control for samples that might have been mislabeled (Additional file 1), the raw transcriptome reads obtained from each species were mapped to each transcriptome assembly generated for each of the three species. - 1 Overview of the assembly composition and completeness. - To account for the difference in mapping rates and reduce the level of noise, we performed each of the three pos- sible interspecies comparisons twice using each of the two assemblies as references. - unless stated otherwise, the same criteria for differential expres- sion are applied throughout the paper) and selected only those terms that were overrepresented in each of the three species-pairs comparisons regardless of the assembly that was used as a reference and with a significance level with a p-value <. - The dif- ferential expression patterns of the transcripts associated with the GO terms translation and translational elonga- tion in the three species indicate a species-specific protein synthesis activity in the order E. - However, the annotations of all these transcripts are rather vague as they are based on matches of the sequences with sequences from a phylogenetically rather distant taxon, the leech Helobdella robusta Shankland, Bissen &. - verru- cosus, and no genes were differentially expressed in the other two species. - Lethal temperatures: Temperature sensitivities of the studied species were estimated by determining the species- specific lethal temperatures. - Individuals of the three. - The lethal temperatures for 50% of the individuals (LT50) differed considerably between the species ranging from 25.2 to 27.2 °C (Fig. - 3a) and mirroring the differences in temperature preferences of the species determined previously [15, 16]. - verrucosus the range of the temperatures causing no and 100% lethality was with 24–26 °C comparatively narrow, whereas this range was wider for E. - Thus, although LTx values of the different species were in the lower temperature range similar, the higher LTx values varied more across the species due to the different slopes of the regression curves (Fig. - The species-specific LT10 values were chosen for the temperature stress treatments in the studies of the transcriptomic responses of the amphipod species to thermal stress.. - The more temperature- sensitive species, i.e., the species with lower LT50, showed higher numbers of responsive genes (Fig. - 3 Temperature sensitivities of the different amphipod species and their transcriptional responses to elevated temperature. - mortalities of the studied species at different temperatures. - c, GO terms enriched in the transcripts up-regulated under acute heat shock. - cyaneus in the 3-h LT10 treat- ment, as well as down-regulated (approx. - in the 24-h LT10 treatment in E. - Thus, here we attempted to determine expression changes of the respective genes.. - No other cox-related tran- scripts was differentially expressed in any of the studied species, even though other cox contigs were present in all assemblies.. - Some of the groups were characteristic only for one combination of species and exposure time. - The top MF and CC GO terms enriched in genes down- regulated in the Eulimnogammarus species in response to heat shock were also connected to ribosomes and translation, such as structural constituent of ribosome (GO:0003735), rRNA binding (GO:0019843), translation elongation factor activity (GO:0003746), GTPase activity (GO:0003924), ribosome (GO:0005840) and cytosolic large ribosomal subunit (GO:0022625).. - in the Baikal Eulimnogammarus species, but not in the Holarctic G. - The toxic sensitivities of the examined amphipod species to cadmium were determined earlier. - The only common GO term enriched in the genes up-regulated under cadmium treatment is the response to stress (GO:0006950), which includes mostly the transcripts annotated as putative 70 kDa heat shock protein family members (Additional file 4). - In addi- tion, in the case of E. - b, GO terms enriched in the transcripts up-regulated under cadmium treatment. - c, GO terms enriched in the transcripts down-regulated under these conditions. - The p-values shown on top of the boxplots come from the DESeq2 analysis, except for the value in parentheses calculated with the Mann-Whitney test. - However, none of the transcripts was differentially expressed according to our standards, even though the majority of transcripts in Baikal species had higher median abundance values under cadmium treat- ment (Additional file 1. - Moreover, most of the transcripts had low expression: only six transcripts in E. - Expression levels of the two most abundant transcripts are shown in Fig. - Thus, the questions of specificity of the putative products of these transcripts and regulation of their expression in response to heavy metals remain open.. - The GO terms overrepresented among down-regulated genes were mostly the same as featured in the response to elevated temperature (Fig. - To test these hypotheses, we built scatterplots for all genes that were differentially expressed in response to at least one of the treatments after 24-h exposure (Fig. - 5), especially in the case of E. - This result was corrob- orated by comparison of the numbers of DE genes (see above).. - lacustris occurs in the Holarctic and can be regarded a potential invader species to Baikal. - Animals were either kept in the conditions simulating those of the Baikal littoral (control), subjected to the temperature stress (LT10) or chemical stress (LC10 CdCl 2. - To compare transcriptional profiles between species, we analyzed interspecies differences in the control samples.. - Each point corresponds to a gene which was differentially expressed in at least one of the conditions indicated along the axes. - Exposure of the species to two different stressors clearly reveals similar response in the two Baikal species, which. - each of the transcriptomes contained transcripts with best hits annotated as Hsf1 in other arthropod species, but none of them were differentially expressed. - However, changes in the expression of some transcripts were stressor-specific. - Alter- natively, it is possible that the observed characteristics of the transcription profile are inherent for these species. - Taken together with the results of the interspecies comparison, these results nev- ertheless allow us to suggest that inhibition of ribosomal protein gene expression is associated with stress response and that gene expression in G. - Thus, it is possible that the adaptation potential of the former species is not much wider than for Baikal endemics: even though G. - individuals were collected in the Baikal littoral near the Bolshie Koty village (south- west coast of Baikal N E). - lacus- tris individuals were collected in Lake 14 in the vicinity of Bolshie Koty N E). - In addition, groups of animals were kept at constantly rising temperature (0.8 °C/day) from 6 to 24.4 °C and frozen at 12.4, 18, and 24.4 °C with par- allel control groups kept at 6 °C under the whole time of the experiment. - The average concentration of Cd measured at the end of the exposure was never below 75% of the nomi- nal concentration in the beginning of the exposure (2.2 vs.. - verrucosus, a pool of four to five individuals in the case of E. - Moreover, up to approximately 27% of the forward reads and approximately 20% of the reverse reads were assigned to overrepresented sequences. - Finally, de novo transcriptome assembly was per- formed with Trinity [41] v2.4.0 with the -SS_lib_type FR parameter enabled to utilize the strand specificity of the libraries.. - Technical characteristics of the assemblies are shown in Additional file 1: Table S1. - These assemblies will be referred to with the first letter of the genus, two let- ters of the species name, conditions used and all (e.g., EveBCdTP1_all).. - Quality control of the assemblies was performed with TransRate [42] v1.0.3. - Each transcript was annotated with the blastx ana- log of the accelerated blastp-like program diamond [43]. - 1a) and served as references for mapping reads in the differential expression analysis.. - Additional figures and tables in the pdf format.. - Table S1: Characteristics of the assembly.. - Terms that are robust regardless of the assembly used as a reference are shown in bold. - For explanations of the column names please see the caption for Additional file 2. - LCx: Lethal concentration for x% of the population. - LTx: Lethal temperature for x% of the population. - PD and LRD lead the analysis of the data. - FJ, CO, and SH contributed to the primary analysis of the high-throughput sequencing data. - JH and SS acknowledge the support in part by the Initiative and Networking Fund of the Helmholtz Association (VH-NG738). - PD acknowledges the support of DAAD / Ministry of Science and Higher Education of the Russian Federation (Mikhail Lomonosov Programme).. - DB acknowledges the grant of the President of the Russian Federation (MK . - The funding bodies had no role in the design of the study and collection, analysis, and interpretation of data and in writing the manuscript.. - lacustris), and the differential expression data are available in the GEO repository under the accession number GSE129069.. - The study was approved by the Animal Subjects Research Committee of the Institute of Biology at Irkutsk State University.. - Late Cretaceous-Cenozoic history of the Lake Baikal depression and formation of its unique biodiversity.. - Checklist of the Amphipoda (Crustacea) from continental waters of Russia, with data on alien species.. - Macrozoobenthic communities of underwater landscapes in the shallow-water zone of southern Lake Baikal. - On the ciliate fauna associated with the marine species of the amphipod genus Gammarus J.G. - Metallothionein diversity and distribution in the tree of life: a multifunctional protein. - Large-scale transcriptome analysis of retroelements in the migratory locust, Locusta migratoria.. - Genomic expression programs in the response of yeast cells to environmental changes. - Comparative analysis of the transcriptional responses to low and high temperatures in three rice planthopper species. - Genes of the pig, Sus scrofa, reconstructed with EvidentialGene
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