CEN/TC 264/WG 29 - Ambient air - Monitoring of genetically modified organisms (GMO)

This Technical Specification describes a procedure through which pollen – in particular pollen of genetically modified organisms (GMO) – can be sampled by means of bee colonies.
Bee colonies, especially the foraging bees, actively roam an area and are therefore area related samplers. Pollen sampling depends on the collection activity of the bees and the availability of pollen sources within the spatial zone according to the bees' preferences (supply of melliferous plants). A colony of bees normally forages over an area of up to 5 km radius (median 1,6 km, mean 2,2 km), in rare cases some bees may also forage in greater distances up to 10 km and more [26].
Foragers fix the gathered pollen on the outside of their hind legs (pollen loads, also known as pollen pellets). Inside the hive they place these pollen loads into comb cells close to the brood nest (bee bread). Furthermore, foragers gather nectar and honeydew. Nectar contains pollen which fell from the anthers of the blossom into the nectar drop, or pollen which was dispersed by the wind and sticks in the nectar of other blossoms or adheres to the sticky honeydew of plants. Nectar and honeydew are converted to honey and stored by the bees in the beehive.
Honey, pollen load and bee-bread may be used as sample matrices for the subsequent analysis of pollen as it is possible to concentrate sufficient amounts of pollen for microscopic and molecular biological diagnostics.
Microscopic analysis is used to identify the various pollen types and to quantify the exposure to the target pollen types in question. GMO exposure is analyzed by molecular-biological methods: For analysis of pollen DNA quantitative PCR methods are used and described here in this Technical Specification. The analysis of GMO specific proteins and toxins in pollen is possible, too, using ELISA, but to this date the method has not been evaluated enough in pollen matrices for standardization in this Technical Specification.

This Technical Specification describes a procedure for the use of the passive samplers Sigma-2 and PMF to sample airborne pollen. Both are designed to sample coarse aerosol particles. Collected samples are used to analyze pollen input with regard to pollen type and amount, and input of transgenic pollen. The Sigma-2 passive sampler here provides a standardized sampling method for direct microscopic pollen analysis and quantifying the aerial pollen input at the site. The PMF yields sufficient amounts of pollen to additionally carry out molecular-biological diagnostics for detection of GMO.
Essential background information on performing GMO monitoring is given in Guideline VDI 4330 Part 1 [4], which is based on an integrated assessment of temporal and spatial variation of GMO cultivation (sources of GMO), the exposure in the environment and biological/ecological effects. Ideally, the pollen sampling using technical samplers for GMO monitoring should be undertaken in combination with the biological collection of pollen by bees (FprCEN/TS 16817-2).
The application of technical passive samplers and the use of honey bee colonies as active biological collectors complement each other in a manifold way when monitoring the exposure to GMO pollen. Technical samplers provide results regarding the pollen input at the sampling site in a representative way, whereas with biological sampling by honey bee colonies, pollen from flowering plants in the area is collected according to the bees' collection activity. Thus, this method represents GMO exposure to roaming insects. By combining the two sampling methods these two main principles of exposure are represented. Furthermore, a broad range of pollen species is covered.
The sample design depends on the intended measuring objective. Some examples are given in 6.2.