Community, Creeks & Critters

"Community, Creeks & Critters - using eDNA technology to connect people, place, and science" was a project conducted throughout 2023 and 2024. It aimed to inspire local community members to become volunteer citizen scientists and was funded by the NSW Department of Premier and Cabinet as part of the NSW Social Cohesion Grants Round 2 – Unsung Heroes – Innovation in Volunteering program.

The project built upon a successful 2022 pilot initiative (Using Hornsby Platypus eDNA as a Catalyst for Healthy Waterways) funded by the Sydney Water Community Grants program, which used eDNA technology to detect platypus DNA in rural creeklines across Hornsby Shire. The “Creeks & Critters” project expanded this approach to employ broader eDNA metabarcoding techniques that could detect a wider range of species beyond just platypus. Additionally, it broadened the scope of eDNA sampling to include more creeks across Hornsby Shire, with sampling events planned to capture seasonal variation in Spring 2023 and Autumn 2024.

Environmental DNA (eDNA) is a cutting-edge, non-invasive technology used for wildlife monitoring and detection. All animals release DNA into their surroundings, such as through mucus, faeces, urine, gametes, and skin cells.

Scientists extract and analyse this DNA from water, air, or soil samples to trace the presence of various species. In this project, eDNA samples from water captured genetic evidence of species inhabiting local creeklines.

How does eDNA work?

There are two methods for identifying sources of eDNA – Barcoding & Metabarcoding:

• eDNA Barcoding: This method targets a specific gene to detect a single species. It was utilised in the Hornsby platypus eDNA pilot project.
• eDNA Metabarcoding: This method allows for the simultaneous identification of many taxa within the same sample. The main difference between barcoding and metabarcoding is that metabarcoding does not focus on one specific organism, but instead aims to determine species composition within a sample.

Limitations of eDNA monitoring

Environmental DNA (eDNA) analysis has become a powerful tool in ecology and conservation biology for detecting the presence of species in aquatic environments. However, it does come with limitations:

1. Sampling Bias: eDNA analysis of the local waterways relied on filtering a sample of water (generally 400mL to 2000mL during both Creeks & Critters seasonal sampling events) in order to capture fragments of DNA (e.g. mucus, faeces, urine, gametes, and skin cells). This meant that capture of the DNA was subject to the limitations of sampling, including where and when samples were collected. The sampling method will only detect DNA that happens to be in the volume of water sampled. Therefore, it's possible to miss species that are present in the area but not in the sampled water.
2. Temporal and Spatial Variability: Environmental conditions, such as water flow, temperature, and the presence of inhibitors, can affect the distribution and degradation of DNA in the environment. Consequently, eDNA detection can vary temporally and spatially, leading to inconsistent results.
3. Sensitivity to Detection: The sensitivity of eDNA detection can vary depending on the target species, their abundance, and their behaviour. Rare or elusive species may not shed enough DNA into the environment for detection, leading to false negatives. Reptiles are renown for not shedding much DNA.
4. Community Composition: The diversity and abundance of species in a given area can affect the likelihood of detecting their DNA in a water sample. Species with higher biomass or shedding more DNA are more likely to be detected compared to rare or less abundant species.
5. DNA Persistence: DNA can degrade over time due to environmental factors such as sunlight, microbial activity, and water chemistry. This degradation can reduce the detectability of DNA, especially in certain environments.
6. Cross-contamination: There is a risk of cross-contamination between samples, which can lead to false positives if proper precautions are not taken during sample collection, processing, and analysis.
7. Reference Databases: The accuracy of species identification in eDNA analysis depends on the availability and quality of reference databases. Incomplete or inaccurate databases can lead to misidentification of species or failure to detect species not represented in the database. In Creeks & Critters, no Eastern Water Dragons or Red-bellied Black Snakes were detected due to these species not being represented in the projects eDNA analytical laboratory database.

An important goal of the "Creeks & critters" project was to encourage the local community to get involved and learn more about creek critters while picking up citizen science-based eDNA sampling and survey skills. Through local print and social media promotion, and by volunteers sharing this knowledge within local networks, the project aimed to deepen understanding of aquatic biodiversity, foster community connections, and contribute to the promotion of healthy waterways.

Citizen scientists participated in field work by collecting water samples from creeks using special, easy to use eDNA sampling kits. Samples were then sent off for laboratory analysis where a DNA reference library detected genetic traces (such as skin particles, mucus, faeces, and urine) left by different species in local creeks.

The collection of this valuable eDNA data not only facilitated a baseline assessment of creek biodiversity across the Shire, but also offered insights into local aquatic wildlife. This knowledge could prove crucial in the event of a natural disaster.

DNA data of this kind had never been gathered before within the Bushland Shire, making this the most significant citizen science and creek biodiversity research project in Hornsby’s history.

Project statistics

• 94 citizen scientists participated in the Spring eDNA sampling
• 54 citizen scientists participated in the Autumn eDNA sampling (participation dropped off due to the Sampling Saturdays having to be cancelled and rescheduled 4 times due to ongoing rainfall events and saturated catchments)
• 91 proposed creek sampling site locations were nominated by the community via an online mapping tool
• 85 citizen scientists attended one of three eDNA sampling training workshops
• 41 local waterway sites across Hornsby Shire were sampled for eDNA
• 164 individual eDNA filters were sent off for laboratory analysis (i.e. 2 replicate samples @ 41 sampling sites x 2 sampling events)
• Spring 2023 sampling: 97 vertebrate taxa (i.e. 6 frog, 25 fish, 6 reptile, 36 bird, and 24 mammal taxa), including 1 threatened species; Grey-headed flying fox (Pteropus poliocephalus) and 5 decapod taxa were detected
• Autumn 2024 sampling: 79 vertebrate taxa (i.e. 3 frog, 19 fish, 4 reptile, 34 bird, and 19 mammal taxa), including 3 threatened species; Powerful owl (Ninox strenua), Grey-headed flying-fox (Pteropus poliocephalus) and Pilotbird (Pycnoptilus floccosus) and 5 decapod taxa were detected.
• Longfin eel (Anguilla reinhardtii) and Sydney spiny crayfish (Euastacus australasiensis) were respectively the most commonly detected vertebrate and decapod taxon.
• An innovative Data Dashboard was developed to display the eDNA species detection data
• A baseline biodiversity assessment of Hornsby creeklines was created.
• 55 citizen scientists attended the final project presentation.

The project identified a diverse array of native vertebrate species, including fish, frogs, birds, reptiles, eels, turtles, rakali, flying foxes and microbats. Additionally, a number of feral species were detected, highlighting the impact of non-native fauna on local ecosystems. The eDNA analysis also revealed the presence of decapod species such as freshwater shrimp, spiny crayfish and common yabby, providing a richer understanding of the aquatic biodiversity within the Shire.

Some interesting findings:

• Longfin Eels were the most common detected species at 35 of the 41 sampling sites. Eels are considered one of the most tolerant fish species and can survive environmental hazards like high water temperatures or low dissolved oxygen concentrations. That means they can generally live in habitats where other species cannot survive.
• Cox’s Gudgeon was the second most common species detected at 30 of the 41 sites.
• Leaf Green Tree Frog and the Common Froglet were the third and fourth most common species detected at 25 and 22 of the 41 sites.
• Grey-Headed Flying Fox was detected and is considered a threatened species in NSW.
• About the size of a platypus, the white-tailed native otter-like Rakali was detected at only 2 of the 41 sites
• A significant number of new species were recorded for the very first time within Hornsby Shire.
• 12 introduced species were detected.
• The Saw-shelled turtle, considered an introduced species, was detected at Joe Crafts Creek. There are concerns that its presence may pose a potential threat to the local Eastern Longnecked turtle population.

July/August 2023 – Project promotion

A call-out for citizen scientists was made through local print and social media

August/September 2023 – Registration and Site Nomination

A Hornsby Council YourSay project webpage was set up to facilitate community engagement. Participants registered their interest in becoming citizen scientists and nominated preferred eDNA sampling sites by marking locations on an online map, with some sharing valuable local knowledge. Most nominations were for a participant’s local creek or a favoured bushwalking spot.

September/October 2023 – Finalising Sampling Sites

October 2023 – SPRING eDNA Training and Sampling

Citizen scientists attended workshops to learn eDNA sampling techniques and collected water samples during designated "Sampling Saturdays". Workshops featured presentations from a local platypus expert and included a safety briefing and eDNA kit distribution.

• Workshops were held at Hornsby Library and Galston Community Centre.
• Sampling Saturdays operated from base camps at Hornsby Council Chambers and Galston Community Centre.

April-May 2024 – AUTUMN eDNA Training and Sampling

A training workshop went ahead as scheduled, but base camp "Sampling Saturdays" had to be postponed on four occasions due to extreme wet weather events saturating catchments and creating unsuitable and unsafe sampling conditions. Sampling resumed when conditions improved, with the final sampling session completed on the 1st of June.

August 2024 – Final Project Presentation

A project presentation was held on 22 August 2024 showcasing findings and celebrating the contributions of the project’s valued citizen scientists.