Bacteria and archaea are under-characterized in palaeoecological studies, despite their ubiquity, high diversity, and tight integration with the abiotic, biotic, and human-influenced environments. The complexity of their assemblages and difficulties in separating living- from paleo-prokaryotes render research challenging. Here, we present an ancient metagenomic time series of prokaryotes from a sediment core of Lake Constance, spanning the last 13 500 years of natural and anthropogenic impact. We mapped DNA to reference genomes and characterized the DNA damage of taxa as collectively increasing with time. By constructing co-abundance networks, we recognize major assemblage groups, containing both dead and living microbes, that show specific dynamics: short-term and often low-abundance assemblages are linked to the Pleistocene-Holocene transition, floods, and human activities. Noticeably, certain lineages harbouring microbes common in human-impacted environments expanded during the Middle Ages and Modern time. Some abundant taxa associated with various freshwater and soil environments persisted through millennia. By extricating different sources and trajectories of change, we demonstrate the power of prokaryotic sedimentary DNA in revealing nature- and human-caused long-term eco-evolutionary consequences.

Ecological connectivity shapes ecosystem responses to climate change and is thought to underpin stability, yet its millennial-scale dynamics remain poorly resolved. We asked how spatial and temporal connectivity of plant metacommunities changed over the last 40 ka and which processes drove it. We analysed and compiled plant sedimentary DNA from 20 lake cores across Beringia (Siberia, Alaska) to investigate community dynamics and, for a high-resolution subset, applied beta- and zeta-diversity to track connectivity. Vegetation changed coherently across the glacial–Holocene transition, with trait shifts mirroring functional composition. Connectivity peaked during the late MIS3 and Last Glacial Maximum—likely aided by the Bering Land Bridge, mass effects and facilitation—collapsed during the Deglacial with rapid turnover, and rebounded in the Holocene as shrub and boreal communities expanded. Temporal zeta within sites exceeded spatial zeta, indicating strong local persistence and resilience. Tundra sites uninvaded by forest maintained continuous species pools. Overall, these patterns underscore the value of a metacommunity perspective for assessing millennial-scale connectivity changes. ### Competing Interest Statement The authors have declared no competing interest. European Research Council, https://ror.org/0472cxd90 Deutsche Forschungsgemeinschaft, https://ror.org/018mejw64 U.S. National Science Foundation, https://ror.org/021nxhr62 Bundesministerium für Bildung und Forschung

Bacteria and archaea are currently under-characterised in palaeoecological studies, despite their ubiquity, high diversity and tight integration with the biotic and abiotic environment and human activity. The complexity of their assemblages, and the difficulties in separating living- from paleo-prokaryotes renders analyses challenging. Here we present an ancient prokaryote metagenomic time-series from a sediment core of Lake Constance, a large and deep perialpine lake from temperate Europe, spanning the last 13,500 years of natural and anthropogenic impact. We mapped DNA to reference genomes and estimated the DNA damage of taxa, which displayed a monotonic relationship with time. By constructing co-abundance networks we recognize major microbial assemblages, containing both ancient and living microbes, that show specific dynamics. Short-term and often low-abundance assemblages are linked to the Pleistocene-Holocene transition, floods and human activities. Noticeably, certain lineages harbouring microbes common in human-impacted environments expanded during the Middle Ages and Modern time. Some abundant taxa that were linked to various freshwater and soil environments persisted through millennia. By extricating various sources and trajectories of change, we demonstrate the power of prokaryotic sedimentary DNA in revealing long-term eco-evolutionary outcomes caused by both nature- and humans. ### Competing Interest Statement The authors have declared no competing interest.

Biodiversity is essential for the resilience and stability of life, yet it is highly dynamic and has continuously evolved throughout Earth’s history. The biodiversity concept encompasses three hierarchical levels of equal importance to fundamental ecological processes: diversity at the ecosystem, species and genetic levels. The current biodiversity crisis calls for an urgent need to understand the causes and consequences of widespread diversity losses at all three levels. Breakthroughs in palaeogenomics have increased the ecological and temporal scales on which we can use genomic information to study past biodiversity, reaching as far back as the Early Pleistocene. In this Review, we explore the possibilities and limitations of using palaeogenomics for studying all aspects of biodiversity. We explore how incorporating palaeogenomics into biodiversity research can provide clues about ecosystem composition, trophic interactions, species distributions, adaptation, evolution and extinction through time, in response to natural processes and as a consequence of human impact. We report how palaeogenomics can be applied to address a wide range of topics across all three hierarchical levels of biodiversity, and we show how advances within the field are making palaeogenomics an invaluable tool for understanding past and present declines in biodiversity, and in helping to predict future losses. This Review explores the utility of palaeogenomics for exploring past biodiversity trends and ecosystem responses to a changing world, including natural climate cycles and increasing human activity.

Little is known about the genetic diversity and stability of natural populations over millennial time scales, although the current biodiversity crisis calls for heightened understanding. Marine phytoplankton, the primary producers forming the basis of food webs in the oceans, play a pivotal role in maintaining marine ecosystems health and serve as indicators of environmental change. This study examines the genetic diversity and shifts in allelic composition in the diatom species Skeletonema marinoi over ~8000 years in the Baltic Sea by analyzing chloroplast and mitochondrial genomes. Sedimentary ancient DNA (sedaDNA) demonstrates the stability and resilience of genetic composition and diversity of this species across millennia in the context of major climate events. Accelerated change in allelic composition is observed from historical periods onwards, coinciding with times of intensifying human activity, like the Roman Empire, the Viking Age, and the Hanseatic Age, suggesting that anthropogenic stressors have profoundly impacted this species for the last two millennia. The data indicate a very high natural stability and resilience of the genomic composition of the species and underscore the importance of uncovering genomic disruptions caused by human impact on organisms, even those not directly exploited, to better predict and manage future biodiversity.

Global warming is a major threat to marine biodiversity and ecosystem functioning, with consequences that are yet largely unknown. To frame these consequences, we need to understand how marine ecosystems respond to warming and related environmental changes. Ecosystem models have proven to be a valuable tool in this respect, but their projections vary considerably. A major limitation in current ecosystem models may be that they largely ignore evolutionary processes, which nonetheless can be relevant on the simulated timescales. In addition, ecosystem models are usually fit to contemporary data and used predictively afterwards, without further validation that they are equally applicable to past (and, by inference, future) scenarios. A promising approach to validate evolutionary ecosystem models is the use of biological archives such as natural sediments, which record long-term ecosystem changes. Since the ecosystem changes present in sediment records are affected by evolution, evolution needs to be represented in ecosystem models not only to realistically simulate the future, but also the sediment record itself. The sediment record, in turn, can provide the required constraints on long-term evolutionary changes, along with information on past environmental conditions, biodiversity, and relative abundances of taxa. Here, we present a framework to make use of such information to validate evolutionary ecosystem models and improve model projections of future ecosystem changes. Using the example of phytoplankton, key players in marine systems, we review the existing literature and discuss (I) which data can be derived from ancient sedimentary archives, (II) how we can integrate these data into evolutionary ecosystem models to improve their projections of climate-driven ecosystem changes, and (III) future perspectives and aspects that remain challenging.

Placed in a carbonate-rich catchment at the northern foot of the Alps, Lake Constance is one of the largest lakes in Central Europe, and the hard water lake chemistry makes it a natural laboratory to investigate in detail the carbonate formation, preservation, and the interactions with the detrital carbonate fraction, the latter with a major dolomite component. The highly reactive lacustrine carbonates are great paleoenvironment archives - due to the climate sensitivity in their formation either as erosional sources of suspended matter or through lake internal processes; still, they are prone to diagenetic overprints that shall be considered in a major context. For that, the (post-)glacial sediment deposits were investigated up to 24 m long cores recovered from the northernmost deep portions of Lake Constance, at approximately 200 mwd. The benthic carbon cycle and mineral sources and (trans)formations, were investigated through porewaters and sedimentary solid phases analyses for stable isotope signatures (CHOS), and major- and trace element compositions. The water isotope signatures display a trend towards lighter data with depth, indicating the development in the lake water composition with time, superimposed by diffusion processes in the sediments. The dissolved inorganic carbon (DIC) concentrations increase below the sediment-water interface and are generally higher in the postglacial sediments, indicating that mineralization rates followed the enhancement of lake productivity under milder climate conditions. The δ13C-DIC shows the lightest values in the glacial sediments and displays an enrichment in Holocene sediments, together with pore water hydrochemistry, indicative of organic matter (OM) oxidation, carbonate dissolution, and potential involvement of methane. The sulfur isotope record indicates that minor dissimilatory sulfate reduction is involved in OM degradation. Bulk carbonate C and O isotope signatures show strong variations partly induced by non-authigenic dolomite. Vertical variations in the composition of porewaters, bulk, and acid-soluble phases, in combination with SEM investigations, allow insights about the (trans)formation of authigenic mineral phases and the destruction of allochthonous carbonates, as possible modifiers of the sedimentary record.

Dormancy is a widespread key life history trait observed across the tree of life. Many plankton species form dormant cell stages that accumulate in aquatic sediments and, under anoxic conditions, form chronological records of past species and population dynamics under changing environmental conditions. Here we report on the germination of a microscopic alga, the abundant marine diatom Skeletonema marinoi Sarno et Zigone, that had remained dormant for up to 6871 +/- 140 years in anoxic sediments of the Baltic Sea and resumed growth when exposed to oxygen and light. Resurrected diatom strains, representing cohorts from six different time points of the past 6871 +/- 140 years, are genetically differentiated, and fundamental physiological functions such as growth and photosynthesis have remained stable through time despite distinct environmental dynamics. Showing that resurrection and full functional recovery, in comparison to 3 +/- 2 years of dormancy, is possible after millennial resting, we emphasize the relevance of dormancy and living sediment archives. For the future, sediment archives, together with the resurrection approach, would offer a powerful tool to trace adaptive traits over millennia under distinct climatic conditions and elucidate the underlying mechanisms.

Ancient lakes contain unique and very vulnerable fauna. Determining and understanding the origin of such biodiversity is a key factor in promoting conservation and management actions in some of the most singular ecosystems on the planet. Lake Ohrid in the Balkans is known as a natural laboratory for speciation, containing a high number of endemic species. However, the identity and origin of the planktonic cladoceran Diaphanosoma is uncertain. Representatives of the genus were long considered to have invaded the lake, but recent morphological studies have suggested that they belonged to the endemic taxon in the Balkans, D. macedonicum. Here, phylogenetic methods based on two mitochondrial gene fragments (COI and 16S) were used to identify Diaphanosoma specimens from the ancient Lake Ohrid and Lake Prespa in the Balkans and compare them with other species in Europe, including those living in nearby water bodies. Molecular evidence showed that D. macedonicum was constrained to the ancient lakes Ohrid, Prespa, and Mikri Prespa, which suggests reproductive isolation within the lakes. Phylogenetic analyses supported previous morphological assessments and situated D. macedonicum within the D. mongolianum species group, which contains three sibling species (D. mongolianum, D. lacustris, and D. macedonicum). Nuclear markers are needed to study intraspecific gene flow in these organisms and discard a potential formation of hybrids.

AbstractSedimentary ancient DNA (sedaDNA) offers an important opportunity for investigating long‐term community dynamics. Nevertheless, sedaDNA is challenging since DNA is degraded and fragmented over time. Of particular interest for such sedaDNA studies are phytoplankton communities, which are sensitive environmental indicators and important producers in aquatic systems. So far, only a few suitable metabarcoding primers for sedaDNA targeting phytoplankton exist. In this study, we introduce new metabarcoding primers targeting cyanobacteria and dinoflagellates. They amplify short, ~200‐bp ribosomal 16S and 18S DNA fragments. We compared these primers against published ones, uncovering distinct communities captured by different primer sets. The newly designed dinoflagellate and cyanobacterial primers revealed unique sets of amplicon sequence variants (ASVs) compared to published primers, highlighting the impact of primer choice on describing community composition. We also explored the effect of amplicon length on metabarcoding success over a sample age. Observed trends suggest that amplification success decreases with longer amplicons, probably as a result of DNA degradation in older sediment samples. Lastly, strong DNA preservation challenges emerged in sediment samples older than 7000 BP, corresponding with oxic phases of the Baltic Sea bottom water. This emphasizes the importance of age, sediment type, and preservation conditions when interpreting sedaDNA results. Despite limitations in temporal resolution, the study shows that sedaDNA‐based fluctuations in the phytoplankton community are consistent with well‐known environmental stages. More research is necessary to understand (1) DNA preservation and its impact on reconstructed communities and (2) impact of abiotic conditions on phytoplankton communities.

Ancient environmental DNA (aeDNA) from lake sediments has yielded remarkable insights for the reconstruction of past ecosystems, including suggestions of late survival of extinct species. However, translocation and lateral inflow of DNA in sediments can potentially distort the stratigraphic signal of the DNA. Using three different approaches on two short lake sediment cores of the Yamal peninsula, West Siberia, with ages spanning only the past hundreds of years, we detect DNA and identified mitochondrial genomes of multiple mammoth and woolly rhinoceros individuals—both species that have been extinct for thousands of years on the mainland. The occurrence of clearly identifiable aeDNA of extinct Pleistocene megafauna (e.g. >400 K reads in one core) throughout these two short subsurface cores, along with specificities of sedimentology and dating, confirm that processes acting on regional scales, such as extensive permafrost thawing, can influence the aeDNA record and should be accounted for in aeDNA paleoecology.

Marginal sea ecosystems, such as the Baltic Sea, are severely affected by anthropogenic pressures, such as climate warming, pollution, and eutrophication, which increased in the course of the past century. Biodiversity monitoring data and assessment of environmental status in such systems have typically been carried out only for the past few decades, if at all, and knowledge on pre-impact stability and good ecological status is limited. An extension of monitoring time series can potentially be achieved through analyses of paleoecological records, e.g. for phytoplankton, which form the base of the food web and are highly susceptible to environmental changes. Within the phytoplankton community, dinoflagellates and diatoms play a significant role as primary producers, and their relative dominance in the spring bloom, calculated as Dia/Dino index, is used as an indicator for the environmental status of the Baltic Sea. To extend time series on the dominance patterns and include non-fossilized dinoflagellates, we here establish a simple droplet digital PCR (ddPCR) reaction on ancient DNA from sediment cores that decodes phytoplankton dynamics. We focus on two common spring bloom species, the diatom Skeletonema marinoi and the dinoflagellate Apocalathium malmogiense, for which we evaluate a DNA based dominance index. It performs very well in comparison to DNA metabarcoding and modern monitoring and can elucidate past species dominance across the past century and across millennia in different basins of the Baltic. For the past century, we see a dominance shift already starting before the mid-20th century in two of the Baltic Sea basins, thus substantially predating current monitoring programs. Shifts are only partly coeval among the cores and the index shows different degrees of stability. This pattern is confirmed across millennia, where a long-term stable relationship between the diatom and the dinoflagellate is observed in the Eastern Gotland Basin, while data from the Gulf of Finland bear testimony to a much more unstable relationship. This confirms that good ecological status based on the dominance pattern of diatoms and dinoflagellates must be established locally and exemplifies how sediment core DNA can be employed to extend monitoring data.

Available DNA barcodes of freshwater zooplankton are scarce in regions like the Iberian Peninsula, which harbours many rare and endemic species and is considered a hotspot of crustacean biodiversity. Recently, a new species of Ceriodaphnia (Cladocera: Daphniidae) was described using morphological analysis of specimens in the Mediterranean region and molecular data on a single locality in southern Spain. In our study, we detected the presence of the newly discovered taxa and here, we provide new DNA sequences on the barcoding region mitochondrial cytochrome c oxidase I together with ecological information of the recently described Ceriodaphnia smirnovi. Additionally, we built a molecular phylogenetic tree and genetically compared these specimens with previously available mitochondrial DNA sequences and with new sequences of the genus recovered in Iberia. Our data suggest that this morphospecies might contain some cryptic taxa and might be more common than previously thought, occupying temporary to semi-permanent ecosystems, with vegetation and highly variable pH and turbidity conditions. Moreover, the existence of a non-identified clade within our phylogenetic tree requires additional morphological research. Our study highlights the need for further research on microcrustacean biota to better constrain its spatial boundaries, phylogenetic relationship and determine species hiding cryptic diversity.

Successful conservation of cryptic species such as representatives of the genus Cobitis presents a challenge for fishery managers. Only Cobitis taenia L. is currently assumed to occur in southwest Germany, where it is classified as critically endangered. Established genetic markers and eDNA analysis identified three Cobitis species in the study area. Cobitis taenia and Cobitis elongatoides were widely distributed in the Rhine River system, primarily forming hybrid species complexes in which polyploid individuals dominate but coexist with diploid parent species. A third non-native species, Cobitis bilineata, has recently become established in the southern part of the Rhine to which it probably migrated naturally via the connected Aare River system. In the Danube River catchment, two populations, including one previously thought to be extinct, comprise both diploid and polyploid representatives of C. elongatoides. Our results provide an important basis for more targeted conservation strategies for Cobitis species currently found in the region.

Sedimentary ancient DNA (sedaDNA) has proven to be a useful tool for palaeoenvironmental studies, but only a handful of studies exist so far for tropical regions. In this study we used sedaDNA to study the temporal succession of Brachionus spp. rotifer mitochondrial DNA haplotypes using two sediment cores from two climatically different alkaline-saline crater lakes from the Kenyan Rift Valley. Data were retrieved from a sediment core (dating back to AD 1800) from Lake Kageinya, located in the remote, hot and hyper-arid Suguta Valley. sedaDNA was used to study the temporal succession of mitochondrial DNA haplotypes of Brachionus spp. rotifers. The results were compared to previously published data from Lake Sonachi, a well-studied lake in the humid and colder mountainous region of Kenya near the town of Naivasha, now supported by a Pb-210 age chronology. Both records extend back before the onset of substantial anthropogenic impact in these regions. The results revealed that climate-rather than anthropogenic impact-was most strongly correlated with haplotype changes in both lakes. During prolonged dry periods (such as from AD 1910 to the late AD 1960s), certain haplotypes persisted. Sudden changes and the emergence of alter native haplotypes were observed when climate became more humid or during episodes of highly variable climate (before AD 1910 and from AD 1960s onwards). Progressive changes in prevailing haplotypes during periods with variable climate could reflect local adaptation and/or be the result of immigration of new haplotypes after the eradication of previous populations during extreme environmental conditions (high temperatures, UV irradiation, pH and salinity). The results indicate that, despite adverse chemical conditions, sedaDNA in tropical lake sediments is preserved for at least a few hundred years. Therefore, its analysis provides a useful complementary palaeoenvironmental proxy for palaeolimnological reconstructions and novel insights on changes in rotifer populations through time.

The woolly rhinoceros (Coelodonta antiquitatis) is an iconic species of the Eurasian Pleistocene megafauna, which was abundant in Eurasia in the Pleistocene until its demise beginning approximately 10 000 years ago. Despite the early recovery of several specimens from well-known European archaeological sites, including its type specimen (Blumenbach 1799), no genomes of European populations were available so far, and all available genomic data originated exclusively from Siberian populations. Using coprolites of cave hyenas (Crocuta crocuta spelea) recovered from Middle Palaeolithic layers of two caves in Germany (Bockstein-Loch and Hohlenstein-Stadel), we isolated and enriched predator and prey DNA to assemble the first European woolly rhinoceros mitogenomes, in addition to cave hyena mitogenomes. Both coprolite samples produced copious sequences assigned to C. crocuta (27% and 59% mitogenome coverage, respectively) and woolly rhinoceros (Coelodonta antiquitatis; 27% and 81% coverage, respectively). The sequences suggested considerable DNA degradation, which may limit the conclusions to be drawn; however, the mitogenomes of European woolly rhinoceros are genetically distinct from the Siberian woolly rhinoceros, and analyses of the more complete mitogenome suggest a split of the populations potentially coinciding with the earliest fossil records of woolly rhinoceros in Europe.

Diatoms (Bacillariophyta) are unicellular eukaryotic algae that contribute largely to the primary production of lakes and oceans and play an important role in global carbon, nitrogen and silica cycles. They are abundant and ubiquitous in almost all aquatic environments, they are particularly sensitive to environmental perturbations and are well preserved in the environment and especially in sedimentary archives. For this reason, diatoms are valuable bioindicators applied to appraise environmental status and health of aquatic systems in space and time. The molecular assessment of diatom composition improves the comparability of community assessments because it reduces analytical error through standardisation and automation of sample preparation and taxonomic classification, and provides an alternative to time-intensive and expert-dependent microscopic identification. Moreover, molecular techniques allow for the identification of cryptic diatom diversity and have the potential to characterise adaptational and functional processes in diatoms, allowing a holistic understanding of diatom-environment relationships.

1. Analyses of sedimentary DNA ( sed DNA) have increased exponentially over the last decade and hold great potential to study the effects of anthropogenic stressors on lake biota over time. 2. Herein, we synthesise the literature that has applied a sed DNA approach to track historical changes in lake biodiversity in response to anthropogenic impacts, with an emphasis on the past c. 200 years. 3. We identified the following research themes that are of particular relevance: (1) eutrophication and climate change as key drivers of limnetic communities; (2) increasing homogenisation of limnetic communities across large spatial scales; and (3) the dynamics and effects of invasive species as traced in lake sediment archives. 4. Altogether, this review highlights the potential of sed DNA to draw a more comprehensive picture of the response of lake biota to anthropogenic stressors, opening up new avenues in the field of paleoecology by unrevealing a hidden

How does nature change through time and across space? How might it change in the future? While time travel has long been the domain of science fiction, scientists have approached these questions using techniques that examine the traces of the past remaining in today's environment. The use of ancient environmental DNA (ancient eDNA) is the latest method for paleoenvironmental reconstruction in a line of ancient biomolecular research, with the first research into DNA barcoding appearing near the turn of the last century (Willerslev et al. 2003). This field has principally advanced through a combination of milestone discoveries and a technological revolution that ultimately transformed genetic and genomic analyses into the Big Data Science we recognise today (Fig. 13.1). But what has been the trajectory of this development, and perhaps more importantly, what future potential lies ahead for this research area?

Environmental archives, such as lake sediments, harbour DNA of past and present ecosystems. However, our understanding of the provenance, deposition and distribution of sedimentary DNA in lake systems is largely unknown, limiting the breadth of derived spatiotemporal inferences. By mapping the distribution of aquatic and terrestrial taxa in a large deep lake using metabarcoding, we characterise the spatial heterogeneity of sedimentary DNA and point to its potential driving factors. Taxa composition varies across geographic gradients in the lake, and spatial distribution of DNA is linked to the range and life mode of organisms. Exogenous taxa, such as alpine plants, have the most reliable detection near the mouth of the inflow. Our data reveal that sedimentary DNA is reflecting the mosaic distribution of organisms and organic remains in the environment, and a single location from lakes with watersheds across different elevations, biomes or other diversity boundaries does not capture the full dynamics in the surrounding area.