APPLICATION OF THE SCHOLANDER-PRESSURE-BOMB TO STUDIES ON ENDOPHYTIC BACTERIA OF PLANTS

The Scholander pressure bomb system, which expresses vascular plant sa p, was compared with the trituration method, in which roots are surfac e disinfested and triturated, for recovery of endophytic bacteria. The two methods were compared for recovery of indigenous and introduced e ndophytes from roots of several plant genera. The pressure bomb method was acceptable for routine recovery of endophytes from cotton (Gossyp ium hirsutum), soybean (Glycine max), and bean (Phaseolus vulgaris), b ut owing to tissue collapse under pressure, the method did not work re liably for cucumber (Cucumis sativa) or tomato (Lycopersicon esculentu m) seedlings. High bacterial densities on the root surface, experiment ally obtained by dipping cotton roots into a suspension of Enterobacte r asburiae JM22 immediately prior to processing, did not affect the po pulation densities of recovered indigenous endophytic bacteria by the pressure bomb technique but resulted in increased bacterial densities for the trituration method. Internal populations of JM22 following app lication as a seed treatment were statistically equivalent with the tr ituration and pressure bomb techniques. Analysis of taxonomic diversit y of a group of indigenous endophytes recovered with the trituration a nd pressure bomb techniques indicated some differences between the two groups. The total number of bacterial genera and species recovered wa s greater using the pressure bomb method. Gram-positive species, such as Bacillus spp., were more frequently isolated with the trituration m ethod than with the pressure bomb method. Agrobacterium radiobacter an d less common species were more often isolated using the pressure bomb technique. Pseudomonas spp. and Phyllobacterium spp. were recovered w ith equal frequencies using both techniques. These results suggest tha t the two techniques sample two different internal habitats available for colonization by endophytic bacteria, i.e., the trituration method recovering mainly endophytes residing in the root cortex and the press ure bomb method detecting vascular colonists. A combination of both me thods is recommended for understanding the full pattern of internal pl ant colonization by endophytic bacteria.