fundamentals of eDNA

I was really jazzed about this paper by Diaz (2020) after reading it. It brought up a lot of questions for me, as well as brought to light some interesting parallels to my own research. I think that SPM has great potential in the field of eDNA, but it clearly has some limitations that this paper glossed over a bit. Particularly when talking about time frames for eDNA and what you’re looking for. If you’re not careful in planning and sampling method, then its possible that you’d get a signal from species that were not in the system at the time but their DNA was present in surface sediments. Additionally, if you pool samples like the study did, you won’t be able to use this kind of research to look at variations caused by seasonality or fish migrations, which is something most researchers strive toward. And, what are the differences in these sediment trap methods? Is it possible that contamination from bottom sediments can be ruled out with the use of that rotating system? And how long does it really take to get sufficient DNA in SPM? Is it attaching itself to it or is it picked up from the bottom? I think the possibilities and the yield of DNA produced by SPM is exciting for the field of eDNA, but I also think that it needs way more development to be seen as a real possibility.

On a note about the applicability to my research, I was intrigued by their discussion of bacterial signals in their PCR amplification using the MiFish primer set, as well as some resulting contamination introduced from their lab environment by another fish (Danios). I’ve also been using the MiFish primer set and been recieving this bacterial signal, and after much trial and error we have reduced it but not completely gotten rid of it. What does this say about the efficiency of this primer set in detecting fish? How fine tuned and accurate and sensitive is it if it’s being overwhelmed by bacterial amplfication? And are there adjustments we can make to the set to avoid this other than touchdown PCRs and ampure bead cutting? And as for contamination during metabarcoding, what are the most common lab practices during your library preparation that cause this contamination? What practices can you use to better improve upon sterility and cautious preparation of samples?

I really liked the discussion and thinking this paper generated!! Its super useful to deeply discuss methodology and how it can be improved upon in a study when you’re a grad student doing your own research :)