Testing the sequence of successional processes in miniature ecosystems.

Dispersal, environmental filtering, and biotic interactions define the species inventory of local communities. Along successional gradients, these assembly processes are predicted to sequentially vary in their relative importance with dispersal as the dominating process early in succession, followed by environmental filtering and biotic interactions at later stages. While observational data from field studies supported this prediction, controlled experiments confirming a sequence of successional processes are still lacking. We designed miniature ecosystems to explicitly test these assumptions under controlled laboratory conditions. Our "Ecosystems on a Plate" (EsoaP) are 3D-printed customized microplates with 24 connected wells allowing us to track dispersal, niche filtering, and biotic interactions among bacteria and plants in time and space. Within EsoaPs, we created heterogeneous habitat landscapes by well-specific nutrient levels or by providing plant seedlings as mutualistic partners in a checkerboard pattern. Bacteria of a single strain were released in one well and subsequently distributed themselves within the plates. We measured the spatial distribution of bacterial abundances at two time points as a function of abiotic or biotic heterogeneity. Bacterial abundance distribution confirmed a shift from initial dispersal-dominated processes to later niche filtering and biotic interactions as more important processes. Our approach follows the principles of open science as the affordable availability of 3D printers as well as shared STL files makes EsoaPs disseminatable and accessible to all levels of society, facilitating future experimental research. IMPORTANCE: Hypotheses regarding the underlying processes of ecological successions have primarily emerged from and have been tested in observational studies, lacking substantial support through controlled experiments. The design of such experiments should focus on testing contemporary ecological theories at the intersection of community assembly and successional research. To achieve this, we developed and employed 3D-printed "Ecosystems on a Plate" (EsoaP) within controlled laboratory settings. EsoaPs surmount several limitations of nanoscale instruments that had hindered their application in ecologically meaningful research. By sharing 3D printing designs, experimental protocols, and data openly, we facilitate reproducibility of our experiments by researchers across diverse ecological disciplines. Moreover, our approach facilitates cost-effective replication of experiments, democratizing access to tools for ecological research, and thus holds the potential to serve as a model for future studies and educational purposes.

Adding experimental precision to the realism of field observations: Plant communities structure bacterial communities in a glacier forefield.

Ecological studies are aligned along a realism-precision continuum ranging from field observations to controlled lab experiments that each have their own strengths and limitations. Ecological insight may be most robust when combining approaches. In field observations along a successional gradient, we found correlations between plant species composition and soil bacterial communities, while bacterial Shannon diversity was unrelated to vegetation characteristics. To add a causal understanding of the processes of bacterial community assembly, we designed lab experiments to specifically test the influence of plant composition on bacterial communities. Using soil and seeds from our field site, we added different combinations of surface-sterilised seeds to homogenised soil samples in microcosms and analysed bacterial communities 4 months later. Our results confirmed the field observations suggesting that experimental plant community composition shaped bacterial community composition, while Shannon diversity was unaffected. These results reflect intimate plant-bacteria interactions that are important drivers of plant health and community assembly. While this study provided insights into the role of plants underlying the assembly of bacterial communities, we did not experimentally manipulate other drivers of community assembly such as abiotic factors. Therefore, we recommend multi-factorial laboratory experiments to quantify the relative importance of different factors contributing to microbial composition.

Exploring the Frequency and Distribution of Ecological Non-monotonicity in Associations among Ecosystem Constituents.

Complex links between biotic and abiotic constituents are fundamental for the functioning of ecosystems. Although non-monotonic interactions and associations are known to increase the stability, diversity, and productivity of ecosystems, they are frequently ignored by community-level standard statistical approaches. Using the copula-based dependence measure qad, capable of quantifying the directed and asymmetric dependence between variables for all forms of (functional) relationships, we determined the proportion of non-monotonic associations between different constituents of an ecosystem (plants, bacteria, fungi, and environmental parameters). Here, we show that up to 59% of all statistically significant associations are non-monotonic. Further, we show that pairwise associations between plants, bacteria, fungi, and environmental parameters are specifically characterized by their strength and degree of monotonicity, for example, microbe-microbe associations are on average stronger than and differ in degree of non-monotonicity from plant-microbe associations. Considering directed and non-monotonic associations, we extended the concept of ecosystem coupling providing more complete insights into the internal order of ecosystems. Our results emphasize the importance of ecological non-monotonicity in characterizing and understanding ecosystem patterns and processes. Supplementary Information: The online version contains supplementary material available at 10.1007/s10021-023-00867-9.

Succession comprises a sequence of threshold-induced community assembly processes towards multidiversity.

Research on successions and community assembly both address the same processes such as dispersal, species sorting, and biotic interactions but lack unifying concepts. Recent theoretical advances integrated both research lines proposing a sequence of stochastic and deterministic processes along successional gradients. Shifts in ecosystem states along successional gradients are predicted to occur abruptly once abiotic and biotic factors dominate over dispersal as main driver. Considering the multidiversity composed of five organismal groups including plants, animals, and microbes, our results imply that stochastic, likely dispersal-dominated, processes are replaced by rather deterministic processes such as environmental filtering and biotic interactions after around 60 years of succession in a glacier forefield. The niche-based character of later successional processes is further supported by a decline in multi-beta-diversity. Our results may update concepts of community assembly by considering multiple taxa, help to bridge the gap between research on successions and community assembly, and provide insights into the emergence of multidiverse and complex ecosystems.

Accuracy of mutual predictions of plant and microbial communities vary along a successional gradient in an alpine glacier forefield.

Receding glaciers create virtually uninhabited substrates waiting for initial colonization of bacteria, fungi and plants. These glacier forefields serve as an ideal ecosystem for studying transformations in community composition and diversity over time and the interactions between taxonomic groups in a dynamic landscape. In this study, we investigated the relationships between the composition and diversity of bacteria, fungi, and plant communities as well as environmental factors along a successional gradient. We used random forest analysis assessing how well the composition and diversity of taxonomic groups and environmental factors mutually predict each other. We did not identify a single best indicator for all taxonomic and environmental properties, but found specific predictors to be most accurate for each taxon and environmental factor. The accuracy of prediction varied considerably along the successional gradient, highlighting the dynamic environmental conditions along the successional gradient that may also affect biotic interactions across taxa. This was also reflected by the high accuracy of predictions of plot age by all taxa. Next to plot age, our results indicate a strong importance of pH and temperature in structuring microbial and plant community composition. In addition, taxonomic groups predicted the community composition of each other more accurately than environmental factors, which may either suggest that these groups similarly respond to other not measured environmental factors or that direct interactions between taxa shape the composition of their communities. In contrast, diversity of taxa was not well predicted, suggesting that community composition of one taxonomic group is not a strong driver of the diversity of another group. Our study provides insights into the successional development of multidiverse communities shaped by complex interactions between taxonomic groups and the environment.

Divergent assembly processes? A comparison of the plant and soil microbiome with plant communities in a glacier forefield.

Community assembly is a result of dispersal, abiotic and biotic characteristics of the habitat as well as stochasticity. A direct comparison between the assembly of microbial and 'macrobial' organisms is hampered by the sampling of these communities in different studies, at different sites or on different scales. In a glacier forefield in the Austrian Alps, we recorded the soil and plant microbiome (bacteria and fungi) and plants that occurred in the same landscape and in close proximity in the same plots. We tested five predictions deduced from assembly processes and revealed deviating patterns of assembly in these community types. In short, microbes appeared to be less dispersal limited than plants and soil microbes, and plants strongly responded to abiotic factors whereas the leaf microbiome was plant species specific and well buffered from environmental conditions. The observed differences in community assembly processes may be attributed to the organisms' dispersal abilities, the exposure of the habitats to airborne propagules and habitat characteristics. The finding that assembly is conditional to the characteristics of the organisms, the habitat and the spatial scale under consideration is thus central for our understanding about the establishment and the maintenance of biodiversity.