Analysis of the photobiont population in lichens using a single-cell manipulator

Lichen thalli are composed of at least two genetically different organisms; a fungus (the mycobiont) and an alga (the photobiont). However, little evi dence exists for genetic homogeneity within the populations of the two bion ts. The aim of our study was to assess the photobiont population of a liche n thallus in this context. To this end it is necessary to analyse a conside rable number of photobiont strains, a task not easily achieved by the tradi tional micropipette method, usually applied for the isolation of photobiont s. Instead a single-cell manipulator was used. This demonstrably facilitate d the establishment of axenic cultures of lichen photobionts. Due to the hi gher precision in selection and a shorter handling time, the yield of algal strains is significantly higher in comparison with the micropipette method . A total of 24 photobiont strains were analysed, obtained from 3 different regions (the apothecial margin, the margin and the center of the thallus) of one and the same thallus of the foliose corticolous lichen Pleurosticta acetabulum (Necker) Elix & Lumbsch. The strains proved to be identical with respect to their morphology, belonging to Trebouxia arboricola Puymaly, an d the absence of an intron at position 1512 in the 18S nrDNA (numbering bas ed on the rDNA gene of Escherichia coli). The ITS-region of one representat ive per location was determined and resulted in identical sequences, again indicating a homogeneous photobiont population in this Lichen thallus. In a second example, we analysed 15 photobiont clones isolated from a specimen of Tremolecia atrata (Ach.) Hertel, a crustose saxicolous lichen species. I n this case the algal morphology, the absence of the 18S nrDNA intron at po sition 1512, and identical ITS nrDNA-sequences of all 15 clones revealed a homogeneous photobiont population. Further applications of the micromanipul ator in the studies of lichens are briefly discussed.