IN-SITU ANALYSIS OF THE BACTERIAL COMMUNITY IN THE GUT OF THE EARTHWORM LUMBRICUS-TERRESTRIS L BY WHOLE-CELL HYBRIDIZATION

The bacterial community in the gut of the earthworm Lumbricus terrestr is was analyzed by whole-cell hybridization with 16S rRNA targeted oli gonucleotide probes. Whole-cell hybridization protocols using fluoresc ence-, peroxidase-, or digoxigenin-labeled oligonucleotide probes faci litated detection of significant fractions of bacterial cells stained with 4',6-diamidino-2-phenylindole (DAPI) in the fore-, mid-, and hind -gut and cast of the earthworm. The application of peroxidase- and dig oxigenin-labeled probes, however, was hampered by several methodologic al drawbacks: the requirement of enzymatic permeabilization, the diffu se images of stained cells, and the incompatibility with DAPI staining used as control. Quantitative analysis of the bacterial community was also influenced by its considerable variability in different individu al earthworms. Though the number of bacteria detected by DAPI staining as well as by whole-cell hybridization with the fluorescent eubacteri al probe Eub338 generally showed a significant increase in the number of bacteria towards the end of the gut, a decrease in bacterial number s could be found in some earthworms. In situ analysis of the bacterial community in the fore-, mid-, and hind-gut of one individual earthwor m by whole-cell hybridization with the fluorescent eubacterial probe E ub338 recorded 15, 30, and 25% of DAPI-stained bacteria, respectively. In the cast 37% of the bacteria were detected. Similar to counts obta ined by DAPI and by whole-cell hybridization with probe Eub338, the nu mber of bacteria belonging to the alpha- beta-, and gamma-subgroups of proteobacteria increased significantly towards the end of the gut and remained high m the cast. While the most significant difference in th e counts of bacteria belonging to the ol-subgroup was obtained between the hind-gut and cast, bacterial populations of the beta- and gamma- subgroups of proteobacteria increased most prominently between the for e- and hind-gut.