Developing the metrology for nucleic acid analysis to support surveillance of DNA from the environment
TL;DR
The challenge which may occur when there are trace amounts of DNA (≤ 1 copy) which could be found in an environmental survey sample is focused on and the DNA of an antibiotic resistance gene (ARG) is detected using the high accuracy binary counting method of digital PCR.
Abstract
Monitoring the impact of climate change is necessary for example to ensure that Net Zero targets are met. Bio-measurements in this area are routinely performed to track the effects of climate change by measuring changes in biological diversity (biodiversity) which can threaten our food and clean water supply, trace the spread of antimicrobial resistant (AMR) infections as well as monitor the impact of mitigation strategies put in place to combat climate change and assist in moving towards a low carbon future. These methods encompass molecular methods such as nucleic acid amplification techniques and massively parallel sequencing technology to measure the environmental DNA (eDNA). The application of these methods with confidence is limited by the fact that the measurement infrastructure is in early development in this field meaning that it is not possible to compare the results from different studies and routes to define the technical performance of these methods are lacking. The methods are under development to advance the metrology to support a wide variety of environmental targets relevant to measure our ecosystem health and track mitigation strategies necessary to preserve health. These targets include monitoring air, water and waste quality to monitor antimicrobial resistance associated with human and agricultural use of antibiotics (a ONE Health approach - recognising the interlinked relationship of the health of animals, people and our environment), recording of species migration and extinction and tracking of genetically barcoded engineered organisms used for bioremediation purposes. There are many challenges to measuring eDNA and developing a reference measurement system to support the confidence and traceability in the field and explore these challenges. We have initially focused on the challenge which may occur when there are trace amounts of DNA (≤ 1 copy) which could be found in an environmental survey sample. We were able to detect the DNA of an antibiotic resistance gene (ARG) at a concentration of 0.10 copies/µL using the high accuracy binary counting method of digital PCR. We will further develop reference measurement procedures for the quantification of airborne eDNA, developing controlled release facilities for characterisation of air samplers and preparing prototype reference materials. These approaches, along with a planned inter-laboratory study will support the standardisation of airborne eDNA measurements. In addition to developing these methods, a documentary standard is proposed, which focuses on the barcoding of engineered bio-products, which will help to ensure the comparability of measurements between laboratories. By developing the reference measurement infrastructure in this area will improve the environmental monitoring approaches enabling stakeholders to have confidence in their results which can best inform public health response and shape environmental policy.