Possible techniques are the use of proteins such as rabbit immunoglobulin G which you can add to the food and detect later with ELISAs.

Hagler published a lot about this for parasitoids (also a review):

https://www.researchgate.net/publication/233638982_An_Immunomarking_Technique_for_Labeling_Minute_Parasitoids

Another technique is the use of fluorescent dyes which was used e.g. for Tachinidae or here sandflies:

http://www.ncbi.nlm.nih.gov/pubmed/8827598

For the proteins digestion time is surely an issue you have to try out before, and for such dye powders the effect on the insects (and for all techniques surely the recapture rate but that's surely well known).

Article An Immunomarking Technique for Labeling Minute Parasitoids

Possible techniques are the use of proteins such as rabbit immunoglobulin G which you can add to the food and detect later with ELISAs.

Hagler published a lot about this for parasitoids (also a review):

https://www.researchgate.net/publication/233638982_An_Immunomarking_Technique_for_Labeling_Minute_Parasitoids

Another technique is the use of fluorescent dyes which was used e.g. for Tachinidae or here sandflies:

http://www.ncbi.nlm.nih.gov/pubmed/8827598

For the proteins digestion time is surely an issue you have to try out before, and for such dye powders the effect on the insects (and for all techniques surely the recapture rate but that's surely well known).

Article An Immunomarking Technique for Labeling Minute Parasitoids

Another approach may be to use enriched stable isotopes. Some work has been done in aquatic ecosystems using enriched 15N. It has also been used to study seed dispersal in bell peppers.

MACNEALE, K. H., PECKARSKY, B. L. & LIKENS, G. E. (2005). Stable isotopes identify dispersal patterns of stonefly populations living along stream corridors. Freshwater Biology 50, 1117-1130.

Dear Peter and Danswell,

Thank you for your interesting contridutions!!!

Use of the IgG seems a good compromise between sensitivity, field detection (easy and rather cheaper with elisa on several thousand individuals) and long term persistance. Of course, digestion time need to be tested in flies before to run a large assay.

I am less convinced by the use of fluorescent dyes. But may be they could also usefull even in wet tropical climat...

Indeed the use of stable isotope seems perfect in terms of : facility of use, time and stability, low environmental impact. I keep them in mind in case of. The main problem I see using them is in term of detection. A good lab is requested :-)

Thanks again!!!

I Agree with Peter

IgG has been used in many Ant studies, both in the field and lab, It does last for upwards of 1 week in ants. I have also worked with IgG and found it did the job well.

You can check papers by Buczkowski, G. for the Ant systems.

IgG is really easy to mark with as you just mix it with the substance you are feeding to your study system. It also has little environmental impact as it is broken down by the digestive system, it is just a protein, it is also relatively sensitive.

You may try with oil paints, just a little drop on the back resists for at least 2 week after drying

Hi,

I have nothing new to add regarding methods, but for an instance of using isotopes to study dispersal on flies you can check the following reference:

Braack LEO, Retief PF (1986) Dispersal, Density and Habitat Preference of the blow-flies Chrysomyia albiceps (Wd) and Chrysomyia marginalis (Wd) (Diptera: Calliphoridae). Onderstepoort. Journal of Veterinary Research 53: 13-18.

Dear David and Paolo,

Thank you for your contributions!

David did you have an idea about the retention time of the IgG marker in ants (its seems to be +/- 4 days in termites as show in papers of Buczkowski)?

This make me IgG the best and cheapest marker for the moment.

The drop of oil paint is also a good idea with good resistance. But this seems to be less easy to be used on the field on +/- 50 000 diptera before release them.

If this is primarily a mark-release-recapture study, another technique you may want to consider would be marking the flies using fluorescent marking powders. Day-Glo (as well as a few other companies) make many powders commonly used to mark flies for similar studies. However, this process does involve a bit of labor in the hind end regarding the detection of the fluorescent powder on flies and may not hold up incredibly well in tropical high humidity conditions.

Here are a few references to check regarding the fluorescent powder.

Corbett, A., and Rosenheim, J. A. 1996. Quantifying movement of a minute parasitoid, Anagrus epos (Hymenoptera: Mymaridae), using fluorescent dust marking and recaputre. Biol. Control. 6:35-44.

Weldon, C. W. 2006. Marking Queensland fruit fly, Bactrocera tryoni (Froggatt) (Diptera: Tephritidae) with flourescent pigments: pupal emergence, adult mortality, and visibility and persistence of marks. Gen. Appl. Ent. 34: 7-14.

Casanova, C., Costa, A. I., Natal, D. 2005. Dispersal pattern of the sand fly Lutzomyia neivai (Diptera: Pschyodidae) in cutaneous leishmaniasis endemic rural area in Southeastern Brazil. Mem. Inst. Oswaldo Cruz. 100: 719-724.

http://www.scielo.br/pdf/mioc/v100n7/v100n7a06.pdf

Hi Yves

I do not remember off the top of my head, will go back to my honours thesis and check how long my detection was after application, though I would only be able to tell you tomorrow. As a example of the sensitivity, I marked food and tracked its presence throughout the colony, this included individuals that had not eaten the food but had received it from donors via tropholaxis, I still recorded substantial numbers of positive individuals despite the dilution effect.

you might find this useful:

http://entomology.tfrec.wsu.edu/VPJ_Lab/downloads/immunomarkers2.pdf

Vincent Jones' Lab provides a nice overview on immuno marking with food proteins: http://entomology.tfrec.wsu.edu/VPJ_Lab/immuno.html

Hello - I would be happy to answer questions you might have about the isotope technique. I used it to label stoneflies, for the paper that was listed above. One definite (and obvious) limitation is that it is not useful for multiple recaptures - you have to incinerate the insect to determine if it indeed was isotopically labeled. Another thing to consider, as mentioned above, is the turnover of the isotope you are using. Determining how long the insect will stay "labeled" will depend on several factors, including how enriched it is when it is released and how much and on what it will feed between release and capture. Some advantages of this method over a dust, powder or paint method, is that it does not impact an insects flight ability, color, or perception of it's environment, and you don't have to handle the insects (a big deal for small or delicate taxa). Hope this helps,

Kate Macneale

I have seen a paper where lacewing were fed fluerescent material and then could be found at night with spotlights. This can really help to find back widely dispersed animals, I suppose.

Here is some left field electronic technology that may work (depending on the scale of movement you'd like to detect)- at least it does for honey bees and fruit flies. These tiny chips record insect movement across the landscape, similar to PIT tags and listening stations in larger organisms.

http://csironewsblog.com/2014/01/15/tiny-technology-creates-a-buzz/

http://my.csiro.au/News-and-events/News-Listing-Page/2014/March/24/Queensland-fruit-flies-reveal-their-secrets.aspx

One of my colleagues has used fluorescent powder to trace pollinators -- she dusted flowers and then followed it's spread through her field site by searching for it at night with UV light. If flies were dusted with it, it would often show up where they subsequently land, and it would also transfer to flies they encounter 'intimately'. It would also stay on the flies for life, and the first generation dusted would be very easily identified, even if dead on the ground.

Sorry, I do not know what the technical specs or supplier are, but I gather that several colours were used and should be available.

Dear all,

Thank you for your incredible contributions!!

Like IgGs, the fluorescent marking seems also interesting and promising with all your references. One additional main issue to be tested will be the persistence of this marking under wet tropical field conditions. On the field it could be also easy to manage the detection of several thousand marked flies with fluorescent lights ...

Kate, thank you for your proposition! I keep them in mind. In my idea, it should be only a capture. So incinaration of the insect is not a problem after their morphological identification (if needed) :-)

John, although the electronic technology could be also applied and efficient, I have some difficulties to imagine the cost, the placement and the use of 50 000 chips on flies :-D It should be better and cheaper in this case to use oil paints.

Thank you again!

To add to the fluorescent powders: There are several papers describing how to mark insects automatically when in traps (e.g. Diptera and Lepidoptera). One would be

HALL, M. J. R. (1988), Use of fluorescent pigments for the automatic marking of tsetse flies in traps. Medical and Veterinary Entomology, 2: 171–176. doi: 10.1111/j.1365-2915.1988.tb00067.x

For internal markers Bribosia et al. used Rhodamine B as a fluorescent dye which was provided with the food. They tested this using a very mobile hoverfly species (E. balteatus) and it was even possible to mark the offspring, as the eggs also contained dye: http://www.ncbi.nlm.nih.gov/pubmed/16628909 (I can send a copy of the paper if somebody is interested)

We used aniline dyes to look at black fly dispersal many years ago. After collecting them, we killed them and laid them out on fitler paper. I believe we put a drop of ethanol on each specimen and if it was a marked fly, the filter paper around it became coloured with the specific colour. We were able to mark and release at different sites using different colours ... our recapture rate was pretty low, but I am sure that is because there were so many millions of black flies out!!

Good thing about tracking technology is that you dont loose insects (those that are never recpatured/resighted) and the data is really rich. If only you could tag 20 indivdiuals and follow their every movement, it would be super. I suspect that a small revolution in entomolgy is near due to this technology. See also question about triangulation of butterflies by Safi.

There has been a special issue of marking techniques for insects by the International Journal of Pest Management, 50(3), 2004. You might also find some ideas there.

Thank Maren and Freerk. I will have a look to that.

In our lab we looked at the flourescent approach for a marked recapture experiment. Labelling was robust enough to detect insects after a week or so but was time consuming due to the relatively low levels of fluorecence we achieved (and a huge by-catch). A second method we examined was to feed adult flies a sugar solution containing a red food dye. The insects could be squashed for several days later and still yield a red solution although whether this would work in the field is anyone's guess. The frass of the insect (Musca domestica) also remains, somewhat surprisingly, coloured for several days afterwards and so placing adults on filter paper overnight gives readily identiable red spots.

Hi Howard,

Nice to read your answer and experience.

Could you suggest some dyes that could be ingested? Please could you send me your protocole with the red food dye and may be some photos of the result afetr squashing :-)

Best

Hi Yves,

I'll dig the stuff out this evening and get it to you tomorrow.

Cheers,

Howard

:-) Thank you!!

Hi Yves,

As promised, a report I did a few years ago - the bits you are interested in are on Pages 20-21 and 39-43. Apologies that I have not cut it out from the rest of the report (which was very hit-and-miss). Hope this is of some use. Please note that I did not get the chance to validate the procedure in the real world as the funding got pulled.

All the best,

Howard

Depends how big your flies are. Commercially available “queen bee tags” might work. They are round, slightly domed paper tags printed with numbers from 1-99 and are available in 5 colors. We used them to mark snails with a shell length of 6 mm or greater. We attached them with super glue. They generally remained attached for 5 to 6 months with a few lasting a full year. Check: http://www.beeworks.com/catalog/index.php?main_page=product_info&products_id=125

Thank you Howard! Very interesting!!

Nice idea to glue a tag. May be it could work on flies (they have size of thorax around 5 mm, I think). But probably the super glue will not allow the flies to keep their health :-) Neverthelless one other glue could be tested. Thank you :-)

The bee tags come with glue and applicator and since they are attached ot honey bees I expect the glue is safe for flies. The tags are about 2 mm diameter.

That seems cool :-) Thank you again!

See this paper, page 59

Thank you Yves!

We used spray for shoes to mark ants (in collaboration with Rainer Rosengren, Finland). Three colors are long lasting, silver, blue and white.

Dear Daniel,

Interesting idea! Did you know what is the composition of the shoes' spray?

I have to check to the lab, but at least no greenhousesgaz and the ants survived very well, but you should just press slwoly to send little drops of paint and not spraying. Little dots on the back of ants stayed for nearly a year!

Very interesting!!

Thank you!!

I have thought about playing around with some non toxic flourescent powder that floresces under UV light, if applied liberally to bats I have found it sticking on for a couple of weeks. Have some interest in testing matrix permeability with insect releases, thinking that coating them thoroughly in powder, releasing, then searching at night with a UV flashlight might work -- but would need to test it to see longevity.

It is stuff like this

http://www.amazon.com/Ultraviolet-Theft-Detection-Powder-Flashlight/dp/B0038AM1QU/ref=sr_1_3?ie=UTF8&qid=1398825409&sr=8-3&keywords=flourescent+powder

Here are a few references that I've seen on this. If you need a pdf of these send me a message.

-Hagler J.R., C.G. Jackson. 2001 Methods for marking insects: Current techniques and future prospects. Annu. Rev. Entomol. 46:511-543. doi: 10.1146/annurev.ento.46.1.511.

-Schellhorn N.A., G. Siekmann, C. Paull, G. Furness, G. Baker. 2004 The use of dyes to mark populations of beneficial insects in the field. Int. J. Pest Manage. 50:153-159.

-Silberbauer L., M. Yee, A. Del Socorro, S. Wratten, P. Gregg, M. Bowie. 2004 Pollen grains as markers to track the movements of generalist predatory insects in agroecosystems. Int. J. Pest Manage. 50:165-171. doi: 10.1080/09670870410001731880.

Dear Kimberly, Eric,

Thank for your contributions!!

Nice to imagine fluorescent bats flying in the dark sky :-D

Dear Yves - sorry for late response

Rebecca Hood-Nowotny has demonstrated the use of enriched 13C as a marker - as suggested earlier, but her papers dealt with terrestrial flies. Attached fyi

Further, I'd recommend 13C over 15N as a marker 'cos C more likely to be incorporated into the "innate tissues" (only a relative term in insects, I have found)

And

Hi Peter!

Good idea too and sugestion!!

Thank you!

For mosquitoes, the most common method is to use fluorescent dusts. Lots of examples. See the book Mosquito Ecology by John Silver It lasts well enough for mosquitoes, but I guess the rate of loss of marks will depend on how smooth or hairy your flies are... But it's easy!