Periodic Reporting for period 1 - EcoScan (guano-based monitoring of ecosystems – a novel approach to capture ecological processes underlying ecosystem health)
Berichtszeitraum: 2020-06-01 bis 2022-05-31
Biodiversity loss, exacerbated by climate change is a great challenge facing all life on earth. Rapid and informative assessments of biodiversity are needed to detect and combat declines in biodiversity. Although many monitoring programs exist using various technologies and approaches (e.g. remote sensing, acoustics, visual observations, citizen science), such approaches tend to have limited taxonomic scope and tell little about the underlying processes that precede these end states. Many researchers are working on developing novel methods for the ‘Next Generation of Biodiversity Monitoring’ to complement existing monitoring approaches. Such new methods should standardisable across a large spatial scales and be amenable to automation. Environmental DNA (eDNA) based monitoring is acknowledged as a key element in future protocols for terrestrial and aquatic biomes.
In our project, we aimed to test whether bats could contribute as landscape samplers to help deliver an improved eDNA-based monitoring for terrestrial ecosystems.
Why bats?
• Many monitoring schemes are site based, whereas bats actively sample at the landscape-scale, in a relatively fixed radius of a few kilometres around their roost.
• eDNA is typically characterised by low amounts of degraded DNA and the associations between taxa detected can be difficult to infer, as it not certain that they were present in the same place at the same time. The short gut retention time of insectivorous bats ensures that freshly collected droppings contain a relatively well defined temporal sampling (12-24 hrs) of taxa who were present in the same sampling area, with good quality DNA.
• Bats eat not only a wide diversity of prey items but huge quantities too – up to their own body weight per night during energetically demanding periods – ensuring diverse and plentiful samples.
• Depending on species, temperate bat species either exclusively or regularly use the same sites as day roosts, ensuring a reliable and predictable collection point for fresh droppings.
Using bats as samplers, it is possible to retrieve the DNA or RNA of many different groups due to trophic aggregation in the diet. In our project, we focused on bats, insects, plants, fungi and viruses. Plants form the base of ecosystem functioning and accumulate increased viral- and fungal load in response to stressors like climatic events. The fungi and viruses which grow on plants are ingested along with plant matter by insect herbivores who similarly have microorganisms (viruses and fungi) and may be eaten by a secondary consumer such as a bat. By collecting and sequencing bat droppings, we can get a rich temporal snap-shot of multiple levels of diversity in the environment. However, we can go far beyond simple presence-absence of the species in these groups. We can also reconstruct the interactions between them to understand ecological relationships, how these change over time, and in relation to climate and land use.
In our project, we aimed to test whether bats could contribute as landscape samplers to help deliver an improved eDNA-based monitoring for terrestrial ecosystems.
Why bats?
• Many monitoring schemes are site based, whereas bats actively sample at the landscape-scale, in a relatively fixed radius of a few kilometres around their roost.
• eDNA is typically characterised by low amounts of degraded DNA and the associations between taxa detected can be difficult to infer, as it not certain that they were present in the same place at the same time. The short gut retention time of insectivorous bats ensures that freshly collected droppings contain a relatively well defined temporal sampling (12-24 hrs) of taxa who were present in the same sampling area, with good quality DNA.
• Bats eat not only a wide diversity of prey items but huge quantities too – up to their own body weight per night during energetically demanding periods – ensuring diverse and plentiful samples.
• Depending on species, temperate bat species either exclusively or regularly use the same sites as day roosts, ensuring a reliable and predictable collection point for fresh droppings.
Using bats as samplers, it is possible to retrieve the DNA or RNA of many different groups due to trophic aggregation in the diet. In our project, we focused on bats, insects, plants, fungi and viruses. Plants form the base of ecosystem functioning and accumulate increased viral- and fungal load in response to stressors like climatic events. The fungi and viruses which grow on plants are ingested along with plant matter by insect herbivores who similarly have microorganisms (viruses and fungi) and may be eaten by a secondary consumer such as a bat. By collecting and sequencing bat droppings, we can get a rich temporal snap-shot of multiple levels of diversity in the environment. However, we can go far beyond simple presence-absence of the species in these groups. We can also reconstruct the interactions between them to understand ecological relationships, how these change over time, and in relation to climate and land use.
We worked with regional bat associations to find suitable sampling sites and volunteers. We were lucky to have a large team of very dedicated individuals across France, who collected bat droppings for the project at the same time each month in summer, during two years. Their incredible effort has laid a solid foundation for the project.
In the lab, we first had to develop the protocol to recover the focal taxonomic groups as we needed to start from large volumes (many droppings pooled to represent the colony) and to extract viruses, many of which have RNA genomes which are fragile and degrade quickly, in addition to the DNA of the bats, insects, plants and fungi – all from the same starting material. Although protocols exist for each of these aspects, a single optimised protocol addressing all aspects wasn’t available. Within the first year of the project, we successfully optimised a combined viral enrichment metagenomics protocol with a multi-taxon metabarcoding approach.
The majority of the diversity recovered was insect taxa (5,547 OTUs), which was expected as the two bats used are insectivorous. Significant regional variation in the diet was revealed (e.g. Brittany versus the Mediterranean), and orders of soft-bodied insects were found to predominate at some sites, though these orders were not previously reported as being an important part of the diet. Likewise, substantial temporal variation was also evident. A high diversity of fungal and plant taxa were recovered (4,660 and ~1000 OTUs, respectively). More than half of the viruses recovered were those of insects, with the next largest proportion being ‘unknown’ viruses. This is a common occurrence in viral metagenomics due to the massive diversity of this group. Nevertheless, plant and fungal viruses were also detected. The project aims and progress were presented on a dedicated website, and at three international conferences in 2020 and 2021.
Together, the EcoScan team is working on analysing the temporal variation in diversity, and associations between taxa. We will also examine how climate and land-use impacts on diversity and ecological interactions. Although the number of sites used in EcoScan is low, our proof-of-concept study has nevertheless demonstrated the great potential for non-invasive biodiversity monitoring with bats as landscape samplers.
In the lab, we first had to develop the protocol to recover the focal taxonomic groups as we needed to start from large volumes (many droppings pooled to represent the colony) and to extract viruses, many of which have RNA genomes which are fragile and degrade quickly, in addition to the DNA of the bats, insects, plants and fungi – all from the same starting material. Although protocols exist for each of these aspects, a single optimised protocol addressing all aspects wasn’t available. Within the first year of the project, we successfully optimised a combined viral enrichment metagenomics protocol with a multi-taxon metabarcoding approach.
The majority of the diversity recovered was insect taxa (5,547 OTUs), which was expected as the two bats used are insectivorous. Significant regional variation in the diet was revealed (e.g. Brittany versus the Mediterranean), and orders of soft-bodied insects were found to predominate at some sites, though these orders were not previously reported as being an important part of the diet. Likewise, substantial temporal variation was also evident. A high diversity of fungal and plant taxa were recovered (4,660 and ~1000 OTUs, respectively). More than half of the viruses recovered were those of insects, with the next largest proportion being ‘unknown’ viruses. This is a common occurrence in viral metagenomics due to the massive diversity of this group. Nevertheless, plant and fungal viruses were also detected. The project aims and progress were presented on a dedicated website, and at three international conferences in 2020 and 2021.
Together, the EcoScan team is working on analysing the temporal variation in diversity, and associations between taxa. We will also examine how climate and land-use impacts on diversity and ecological interactions. Although the number of sites used in EcoScan is low, our proof-of-concept study has nevertheless demonstrated the great potential for non-invasive biodiversity monitoring with bats as landscape samplers.
-The protocol optimisation for a combined metagenomic and metabarcoding approach from pooled samples, with carefully planned temporal and spatial sampling, has extended the limits of genetic surveys which typically focus on one taxonomic group, sampled once.
- The metabarcoding of insects resulted in the most detailed overview to date of the diet for two bat species (Rhinolophus hipposideros, Eptesicus serotinus): 9 sites per species in three biogeographic regions, sampled each month May-September during two years.
- The metabarcoding approach detected invasive species and could be used as a surveillance system for these taxa. For example, we detected the tiger mosquito, Aedes albopictus, which is also a vector of disease, and high numbers of the box tree moth Cydalima perspectalis which causes severe defoliation to box (Buxus) plants.
- Microbial diversity is rarely included in terrestrial monitoring, though it plays a key role in ecosystem health. In general, an undue focus is placed on pathogenic fungi and viruses, which are the minority of taxa in these highly diverse groups. In our project, the total viral and and fungal diversity was surveyed, revealing the enormous taxonomic richness in these understudied groups, which also include many highly beneficial taxa.
- Although they comprised only a small number, we also detected important pathogens, including viruses impacting pollinators or vineyards, and several species of fungal plant pathogens – therefore our monitoring approach can also be used for pathogen surveillance.
- Novel methods to deliver the Next Generation of Biodiversity monitoring are badly needed. In this study, we tested the feasibility of combining the known advantages of non-invasive eDNA methods with landscape sampling outsourced to bats. The emerging results confirm that this is an exciting avenue for biodiversity monitoring which can deliver rapid multi-taxonomic surveys and shed light on the ecological processes underlying shifts in diversity or interactions. Methods like these will be critical for enabling fast interventions when ecosystems are under stress.
- The metabarcoding of insects resulted in the most detailed overview to date of the diet for two bat species (Rhinolophus hipposideros, Eptesicus serotinus): 9 sites per species in three biogeographic regions, sampled each month May-September during two years.
- The metabarcoding approach detected invasive species and could be used as a surveillance system for these taxa. For example, we detected the tiger mosquito, Aedes albopictus, which is also a vector of disease, and high numbers of the box tree moth Cydalima perspectalis which causes severe defoliation to box (Buxus) plants.
- Microbial diversity is rarely included in terrestrial monitoring, though it plays a key role in ecosystem health. In general, an undue focus is placed on pathogenic fungi and viruses, which are the minority of taxa in these highly diverse groups. In our project, the total viral and and fungal diversity was surveyed, revealing the enormous taxonomic richness in these understudied groups, which also include many highly beneficial taxa.
- Although they comprised only a small number, we also detected important pathogens, including viruses impacting pollinators or vineyards, and several species of fungal plant pathogens – therefore our monitoring approach can also be used for pathogen surveillance.
- Novel methods to deliver the Next Generation of Biodiversity monitoring are badly needed. In this study, we tested the feasibility of combining the known advantages of non-invasive eDNA methods with landscape sampling outsourced to bats. The emerging results confirm that this is an exciting avenue for biodiversity monitoring which can deliver rapid multi-taxonomic surveys and shed light on the ecological processes underlying shifts in diversity or interactions. Methods like these will be critical for enabling fast interventions when ecosystems are under stress.