Utility of a field deployable qPCR instrument for analyzing freshwater quality

Abstract

A significant limitation of quantitative real-time polymerase chain reaction (qPCR) technology for environmental research has been the lack of portability of the amplification and detection equipment needed to perform qPCR assays in the field. The ability to perform qPCR assays in the field would significantly improve the utility of the technology, enabling quicker risk management decisions in acute environmental emergencies, and speeding the pace of environmental research. We tested a portable qPCR instrument, Liberty16, to evaluate if this instrument would be useful for qPCR analysis in our fieldwork. Parallel testing of the Liberty16 with our existing LightCycler 480 Instrument II revealed similar repeatability and sensitivity for the detection of targets in known and blind freshwater samples by qPCR. Both instruments were able to detect all target sequences at 20 copies per reaction and up to 66% targets in a 10-fold dilution. In seven of eight blind freshwater samples, results from both instruments were consistent for detected or nondetected target. In the other sample, the target was only detected by the Liberty16. Portable instruments of this type may open up new avenues for researchers to perform qPCR analyses in the field.

Artificial DNA in hydrology

“…The suggest investing in reducing DNA tracer preparation time, automating sample processing and qPCR, and refining tracer detectability in different hydrological matrices. Furthermore, they mentioned the potential for rapid detection of DNA tracers in the field as portable PCR and qPCR devices become increasingly available (Billington et al, 2021). This advancement would enable more efficient and real‐time monitoring of DNA tracers in hydrological systems.…”

“…whereby n k is the number concentration of aggregate k, α ij sticking efficiency of smaller particles i and j, η ij collision efficiency of smaller particles i and j, n i number concentration of particle i, n j number concentration of particle j, α ik sticking efficiency of particle i and aggregate k, η ik collision efficiency of particle i and aggregate k. In words, Equation (1) describes a particle number mass balance whereby the rate of change of number concentration of k aggregates is equal to the rate of increase of k particles due to aggregation of smaller particles i and j plus the rate of decrease in concentration of k particles due to aggregation of k aggregate with any other particle. Essentially, Equation (1) is the Smoluchowski equation (Smoluchowski, 1917) which, until today, remains the fundamental theory of flocculation (Benjamin, 2013). The collision efficiency for colloids in surface water, η, can be determined from (Praetorius et al, 2020):…”

“…According to Afrooz et al (2014), Gallego-Urrea et al ( 2011), Labille et al (2015), Petosa et al (2010), Praetorius et al (2014), aggregate rates and sticking efficiency values are typically calculated by tracking particle size over time using Dynamic Light Scattering (DLS), laser diffraction (LD), or nanoparticle tracking analysis, or by following settling of aggregates via batch methods (Barton et al, 2014;Geitner et al, 2017;Tang et al, 2023). In case of hetero-aggregation including sedimentation of hetero-aggregates to the stream bed, then Equations ( 1) and ( 2) [e.g., (Benjamin, 2013)] need to be extended, which is beyond the scope of this review.…”