Diversity and antagonistic potential of bacteria associated with bryophytes from nutrient-poor habitats of the Baltic Sea Coast

Abstract

Very little is known about the interaction of bryophytes with bacteria. Therefore, we analyzed bacteria associated with three bryophyte species, Tortula ruralis, Aulacomnium palustre, and Sphagnum rubellum, which represent typical moss species of three nutrient-poor plant communities at the southern Baltic Sea coast in Germany. By use of two cultivation-independent techniques, denaturing gradient gel electrophoresis and single-strand conformation polymorphism analysis of the 16S ribosomal DNA, a high degree of moss specificity was found for associated bacterial communities. This specificity could be further evidenced by a cultivation-dependent approach for the following parameters: (i) plate counts of bacteria on R2A medium, (ii) proportion of antagonistic isolates, (iii) antagonistic activity as well as spectrum against pathogens, and (iv) diversity and richness of antagonistic isolates. The proportion of isolates with antagonistic activity against the pathogenic model fungus Verticillium dahliae was highest for S. rubellum (31%), followed by A. palustre (17%) and T. ruralis (5%). A high percentage (99%) of moss-associated antagonistic bacteria produced antifungal compounds. The high recovery of antagonistic isolates strongly suggests that bryophytes represent an ecological niche which harbors a diverse and hitherto largely uncharacterized microbial population with yet unknown and untapped potential biotechnological applications, e.g., for biological control of plant pathogens.

FIG.1.

Scanning electron micrographs of the surfaces of gametophytes of T. ruralis (top),  A. palustre (center), and S. rubellum (bottom). Bars, 8 μm; arrowheads indicate bacterial cells.

FIG. 2.

SSCP profiles showing the bacterial communities of the bryophytes A. palustre (lanes 1 to 4), S. rubellum (lanes 5 to 8), and T. ruralis (lanes 9 to 12). The fingerprints were generated by separation of 16S rDNA fragments amplified with universal primers. As a marker (lanes M), a 1-kb ladder was used. Bands indicated by arrows were purified and sequenced (for results, see Table 1).

FIG. 3.

Dendrogram based on amplified 16S rDNA fragments of the moss-associated communities from T. ruralis from Rostock-Warnemünde (Twm), T. ruralis from the island of Vilm (Tiv), and T. ruralis (T), S. rubellum (S), and A. palustre (A) from Ribnitzer Großes Moor, obtained by using universal primers and separated by SSCP. The patterns obtained were grouped by UPGMA.

FIG. 4.

Phylogenetic dendrogram calculated with all 16S rDNA sequences that were recovered from bacteria associated with the mosses T. ruralis (T), A. palustre (A), and S. rubellum (S), grouped by the neighbor-joining method. Additionally, type strain sequences of Burkholderia cepacia, Collimonas fungivorans, E. coli, P. putida, and Serratia plymuthica were added. Most of the isolates belong to three different cluster groups. Bar indicates 10% sequence divergence.

FIG. 5.

Dendrogram showing the relationships of 38 Burkholderia isolates (a) and 16 Pseudomonas isolates (b) from Tortula, Aulacomnium, and Sphagnum based on BOX-PCR fingerprints grouped by UPGMA. Double-headed vertical arrows indicate the similarity for the groupings.