Hi Andrea;

The answer is yes. I know in particular studies made with Phytophthora infestans, the potato late blight pathogen, where DNA is collected directly from blighted leaves and used for isolate fingerprinting with SSR markers. There is a report about this type of study within the Proceedings of the 14th Euroblight workshop (Limassol, 2013), available online from the Euroblight website (http://euroblight.net/workshop-proceedings-1996-2013/). Ref is on page 75 of this volume.

There are probably others, but I have this one on hand. It is interesting in the fact that it shows that the sampling on FTA cards allows to obtain DNA in sufficient amount and quality not only to detect the pathogen, but even to type it genetically. This might be only the case when symptoms are large and soft enough, but this is a widepread situation for foliar pathogens of annual plants at least. I guess it would also work with bacteria, by the way.

Hi Andrea and Didier

Next week we will publish a news about results from the 2013 Monitoring project - where we sampled P. infestans with FTA cards. Read about it here: http://euroblight.net/pathogen-characteristics-and-host-resistance/p.-infestans-monitoring-europe/.

Lessons from the 2013 project:

• This was a low blight pressure year but we received well over 650 samples.

• Sampling indeed was the most critical process, the sampling process needs extra attention. Samples that failed in the SSR analysis were from all regions of Europe

• When SSR characterization failed this was mostly for one of two reasons:

o No (or not enough) DNA present – probably due to dry lesions or confusion with early blight

o A mixture of (at least) two genotypes was detected.

• Consistent labelling of the cards is also of critical importance to facilitate smooth processing of the cards.

For future samplings it is therefore extra important to instruct the people sampling to:

• ONLY sample leaflets with single lesions

• ONLY sample lesions that are clearly sporulating (P. infestans) (or incubate drier samples in a plastic bag with some damp tissue until they sporulate)

The DNA detection system is very sensitive so please minimise the chance of cross-contamination of cards, before, during and after sampling. Avoid touching the sampling areas with anything other than the blight sample and place the ‘used’ cards in separate bags for all subsequent handling and shipping.

More a comment and another question rather than answering the original question.

This is really a great idea....I am interested in powdery mildews, and was wondering if this (Immobilisation of nucleic acids on the FTA card) could be pushed for qRT PCRs of fungal genes......Would you get better results even for a rather small biomass in early time points of infection?

Hi Laurence

I'm not at all an expert in qRT PCR, so the following impression might be quite wrong.

However, my feeling is that qRT PCR requires some form of standardization to be interpreted correctly - because what you're after then is a quantitative assessment of a metabolic activity, and also because they are many steps that can go wrong with a RT-PCR, not least the RNA quality/purity. I would therefore be quite careful in using FTA cards for this purpose, although they might possibly be useful if you work with very standardised samples.

The only way to be sure is probably to have a go at it and compare with more traditional means of nucleic acid sampling and preparation.

Good luck!

Thanks a lot for your answers. Im looking for a rapid method for sampling pine needles. For the detection of Phytophtora pinifolia by a conventional PCR. I've read that it's a bit complicated to fix DNA from this tissues but with some grinding of the needles after the application on the card i may overcome this problem...do you think its possible??