Toksikologija / Toxicology


Testing for pesticides in Croatian birds of prey

The research project being set up at the Sokolarski centre in Šibenik, Croatia is a project aimed at monitoring effects of organophosphate and carbamate pesticides on birds of prey. This is done by monitoring the levels of the enzyme Acetylcholinesterase which is found in birds blood plasma and is effected by organophosphate and caramate pesticides.

Firstly, what is Acetylcholinesterase?

Acetylcholinesterase (AChE) is an enzyme that breaks down cholinesters in the body. The most well known function of this is the breakdown of A-acetylcholine, the ester that transmits signals across nerve synapses. However, forms of acetylcholinesterase are found in most organs, red blood cells and blood plasma. The studies into acetylcholinesterase began in 1914 and as such it is one of the more understood enzymes even though it has several different forms. The inhibition of AChE by organophosphate or carbamate pesticides leads to a build up of A-acetylcholine in the nerve synapse which can cause twitching, paralysis, breathing problems and in extreme cases is fatal.i

Why focus on Acetylcholinesterase in birds of prey?

Acetylcholinesterase is an enzyme found in all animals and insects in one form or another. For this reason it is the enzyme targeted by many pesticides. The most common of which are organophosphate and carbamate pesticides which inhibit the enzyme at its active site. Many chlorinated hydrocarbons have now been banned after it was shown that their long life time in ecosystems was having a super bad effect on entire ecosystems; Organophosphates have risen because they have a comparatively very short lifetime (weeks or months instead of years), they are now one of the most widely used pesticides in commercial farming. The idea is that because insects are so small a much smaller amount of organophosphate is needed to paralyse them and kill them then for larger organisms. However, the usage of these pesticides still needs to be monitored as they are fairly toxic even in small doses. Currently Croatia has no regulations on pesticide use and with the upcoming move into the EU parameters for monitoring and regulating pesticide use will need to be introduced. Looking at Acetylcholinesterase in birds of prey allows us to see the effect of these pesticides on top predators and provides a clear pre-indicator of excessive pesticide use that may be harmful to humans as well as having detrimental effects on surrounding ecosystems.

What are we hoping to achieve?

We will accomplish two main objectives with this project. The first will be a cheap and efficient way of diagnosing pesticide poisoning in birds of prey brought into the centre, allowing them to be treated. The second is an ongoing source of quantative data to support new pesticide regulations in Croatia that will ensure the minimum risk to humans and the environment.

How will the project work?

This project will take place in two stages. Due to the high variation in AChE between species the first stage will be to build up a source of reference values for the normal levels of plasma AChE in individual species. This will be done by taking blood samples of species brought in to the Sokolarski centre twice a day for up to two weeks. This will give the body time to metabolise any residue pesticides while the birds are on a regulated diet. It will also give multiple sample comparisons to reduce margins for errors in those birds that aren’t available in large sample numbers. In each case plasma will be separated from whole blood using a centrifuge and then tested for concentrations of the enzyme using a variation of the Ellman methodii.

The next stage of the project will be to establish a pesticide screening into all birds brought into the centre and to compare their levels of AChE against the established base values. This variation will then be recorded against the species and the geographical area where it was found, and monitored so that any trends in enzyme variation can be monitored and in the case of significant variations appropriate actions can be taken.