Esmond et al. (2026) A multi-tracer approach to constraining water sources of culturally and ecologically significant natural springs: Combining environmental isotopes and environmental DNA

Identification

• Journal: Journal of Hydrology
• Year: 2026
• Date: 2026-03-06
• Authors: Monica Esmond, Diοni I. Cendón, Harald Hofmann, Mark de Bruyn, Matthew Currell
• DOI: 10.1016/j.jhydrol.2026.135245

Research Groups

• Australian Rivers Institute, Griffith University, Nathan, QLD, Australia
• Australian Nuclear Science and Technology Organisation, Lucas Heights, NSW, Australia
• School of Biological, Earth and Environmental Sciences, UNSW Sydney, NSW, Australia
• Environment, Commonwealth Scientific and Industrial Research Organisation (CSIRO), Dutton Park, QLD, Australia
• Australian Research Centre for Human Evolution, Griffith University, Nathan, QLD, Australia
• School of Environment and Science, Griffith University, Nathan, QLD, Australia

Short Summary

This study developed an eco-hydrogeological approach, integrating geochemical tracers (hydrochemistry, stable and radio-isotopes) with environmental DNA (eDNA), to create a robust conceptual model of groundwater flow paths and water sources for Great Artesian Basin springs in Carnarvon Gorge, Australia. The findings revealed that vertical inter-aquifer flow and multiple recharge zones control spring dynamics, with eDNA proving more sensitive than isotopes in distinguishing hydraulically separated flow paths and recharge areas.

Objective

• To employ an ecohydrogeological-geochemical approach to develop a robust conceptual model for the source aquifer(s) of highly ecologically and culturally significant springs in the Great Artesian Basin (GAB), Australia.
• To determine what groundwater sources and flow paths contribute to discharge at different spring vents within a geologically complex region near the GAB recharge zone.
• To assess how the integration of ecological signatures derived from eDNA metabarcoding with environmental isotope and hydrochemical tracers can improve hydrogeological conceptual model development.

Study Configuration

• Spatial Scale: Carnarvon Gorge, central Queensland, Australia, located at the eastern margin of the Great Artesian Basin. The study included 16 spring sites and 7 surface water sites within the gorge system.
• Temporal Scale: Geochemical samples were collected over 9 consecutive days in August 2024, with one spring sampled in July 2023. eDNA samples were collected across all sample areas (n = 20) during the August 2024 period. Radioisotope tracers (tritium, radiocarbon, chlorine-36) provide insights into groundwater residence times ranging from modern to hundreds of thousands of years.

Methodology and Data

• Models used:
  • PHREEQC (charge balance calculations)
  • R software (for statistical analyses including Shapiro-Wilk test, Spearman's rank correlation, Principal Component Analysis (PCA), Non-metric Multidimensional Scaling (nMDS), Permutational Multivariate Analysis of Variance (PERMANOVA), Permutational Analysis of Multivariate Dispersions (PERMDISP), Generalised Additive Models (GAMs))
  • GGally package (for correlation matrices)
  • ampvis2 package (for nMDS visualization)
  • vegan package (for PERMANOVA, PERMDISP, GAMs)
  • mgcv package (for GAM diagnostics)
  • SINTAX classification algorithms (for eDNA taxonomic assignment)
• Data sources:
  • Field measurements: pH, temperature (degrees Celsius), specific conductance (microsiemens per centimetre), alkalinity (milligrams per litre as HCO3).
  • Geochemical tracers:
    • Strontium isotope ratios (87Sr/86Sr)
    • Major ions (cations by ICP-AES, anions by IC)
    • Stable water isotopes (δ2H and δ18O, reported in permil (‰) deviations from VSMOW)
    • Radioisotopes: Tritium (3H, reported in Tritium units (TU)), Radiocarbon (14C, reported as percent modern carbon (pMC)), Chlorine-36 (36Cl/Cl ratios, reported as × 10−15).
  • Environmental DNA (eDNA): Metabarcoding analysis of prokaryotic and eukaryotic taxa from water samples, filtered through 1.2 micrometre cellulose acetate filters.
  • External data: Global Network of Isotopes in Precipitation (GNIP) data (Charleville, Mt Isa), published Surat Basin groundwater strontium isotope data, Shuttle Radar Topographic Mission (SRTM) 1 second Digital Elevation Model (DEM), Bureau of Meteorology rainfall and discharge data.

Main Results

• Geochemical tracers indicate that spring discharge is derived from multiple recharge zones and groundwater flow paths, primarily controlled by vertical hydraulic gradients and inter-aquifer connectivity.
• A clear elevation gradient was observed, with upper tableland (UT) springs exhibiting elevated radioisotopic activities (e.g., up to 1.77 TU for 3H, 103.6 pMC for 14C), suggesting shorter flow paths and modern recharge.
• Lower gorge (LG) springs showed the lowest radioisotopic activities (e.g., <0.07 TU for 3H, 59.4 pMC for 14C, 84.05 × 10−15 for 36Cl), indicative of longer, intermediate flow paths and older recharge, influenced by larger summer monsoonal rainfall events.
• Stable water isotopes corroborated these findings, with enriched ratios for shorter flow paths and depleted ratios for longer, deeper flow paths.
• Strontium isotope ratios (87Sr/86Sr) helped constrain dominant water sources, identifying influences from the Precipice Sandstone, Evergreen Formation, and Hutton Sandstone, and confirming multiple recharge zones.
• Surface waters in Carnarvon Creek exhibited high geochemical homogeneity, representing a composite of various spring inputs.
• eDNA metabarcoding (549 prokaryotic, 579 eukaryotic taxa) demonstrated greater sensitivity than isotopic tracers in distinguishing groundwater from the same aquifer(s) but emerging at different springs hydraulically separated by gorge topography.
• eDNA analysis confirmed the distinct outlier status of spring BB 1 (Bowen Basin) due to its unique microbial community composition, despite similar abiotic characteristics to nearby Surat Basin springs.
• Tritium (3H) showed a statistically significant non-linear relationship with eDNA beta diversity, visualized effectively using Generalised Additive Models (GAMs), supporting the hypothesized elevation-driven vertical hydraulic gradient.

Contributions

• This study introduces an innovative eco-hydrogeological approach by integrating environmental DNA (eDNA) with a comprehensive suite of geochemical tracers (hydrochemistry, stable and radio-isotopes) to develop a more robust conceptual model of groundwater flow paths and water sources for natural springs.
• It is among the first studies to demonstrate the enhanced value of combining ecological (eDNA) with isotopic tracers, particularly highlighting eDNA's superior sensitivity in distinguishing hydraulically separated groundwater flow paths and recharge areas that isotopic tracers alone could not resolve.
• Provides the first detailed synthesis of hydrogeological tracer data for the culturally and ecologically significant springs of Carnarvon Gorge, a previously poorly characterized region within the Great Artesian Basin.
• Emphasizes the critical need for multi-tracer approaches to inform evidence-based management and protection strategies for springs and their connected ecosystems, especially in the face of climate change and resource extraction pressures.
• Advocates for the integration of local-scale field data and culturally inclusive practices in water management initiatives to safeguard unique environmental and cultural values.

Funding

• AINSE Ltd. Postgraduate Research Award (PGRA)
• Australian Contaminated Land Consultants Association (ACLCA)

Citation

@article{Esmond2026multitracer,
  author = {Esmond, Monica and Cendón, Diοni I. and Hofmann, Harald and Bruyn, Mark de and Currell, Matthew},
  title = {A multi-tracer approach to constraining water sources of culturally and ecologically significant natural springs: Combining environmental isotopes and environmental DNA},
  journal = {Journal of Hydrology},
  year = {2026},
  doi = {10.1016/j.jhydrol.2026.135245},
  url = {https://doi.org/10.1016/j.jhydrol.2026.135245}
}