Microbial phyllosphere populations are more complex than previously realized

Phyllosphere microbial communities were evaluated on leaves of field-grown plant species by culture-dependent and -independent methods. Denaturing gra dient gel electrophoresis (DGGE) with 165 rDNA primers generally indicated that microbial community structures were similar on different individuals o f the same plant species, but unique on different plant species. Phyllosphe re bacteria were identified from Citrus sinesis (cv. Valencia) by using DGG E analysis followed by cloning and sequencing of the dominant rDNA bands. O f the 17 unique sequences obtained, database queries showed only four strai ns that had been described previously as phyllosphere bacteria. Five of the 17 sequences had 165 similarities lower than 90% to database entries, sugg esting that they represent previously undescribed species. In addition, thr ee fungal species were also identified. Very different 165 rDNA DGGE bandin g profiles were obtained when replicate cv. Valencia leaf samples were cult ured in BIOLOG EcoPlates for 4.5 days. All of these rDNA sequences had 97-1 00% similarity to those of known phyllosphere bacteria, but only two of the m matched those identified by the culture independent DGGE analysis. Like o ther studied ecosystems, microbial phyllosphere communities therefore are m ore complex than previously thought, based on conventional culture-based me thods.