Spatial distribution of Rhodopseudomonas palustris ecotypes on a local scale

Abstract

The number, spatial distribution, and significance of genetically distinguishable ecotypes of prokaryotes in the environment are poorly understood. Oda et al. (Y. Oda, B. Star, L. A. Huisman, J. C. Gottschal, and L. J. Forney, Appl. Environ. Microbiol. 69:xxx-xxx, 2003) have shown that Rhodopseudomonas palustris ecotypes were lognormally distributed along a 10-m transect and that multiple strains of the species could coexist in 0.5-g sediment samples. To extend these observations, we investigated the clonal diversity of R. palustris in 0.5-g samples taken from the corners and center of a 1-m square. A total of 35 or 36 clones were recovered by direct plating from each sample and were characterized by BOX A1R repetitive element-PCR genomic DNA fingerprinting. Isolates with fingerprint images that were >/=80% similar to each other were defined as the same genotype. Among the 178 isolates studied, 32 genotypes were identified, and each genotype contained between 1 and 40 isolates. These clusters were consistent with minor variations found in 16S rRNA gene sequences. The Shannon indices of the genotypic diversity within each location ranged from 1.08 (5 genotypes) to 2.18 (13 genotypes). Comparison of the rank abundance of genotypes found in pairs of locations showed that strains from three locations were similar to each other, with Morisita-Horn similarity coefficients ranging from 0.59 to 0.71. All comparisons involving the remaining two locations resulted in coefficients between 0 and 0.12. From these results we inferred that the patterns of ecotype diversity at the sampling site are patchy at a 1-m scale and postulated that factors such as mixing, competitive interactions, and microhabitat variability are likely to be responsible for the maintenance of the similarities between some locations and the differences between others.

FIG. 1.

Sampling scheme used in this study. The superscript numbers indicate which cluster of BOX A1R rep-PCR genotypes was predominant in each sample. Cluster 1 contained genotypes 1, 3, 4, 7, and 9; cluster 2 contained genotypes 2 and 6; and cluster 3 contained genotypes 5 and 8.

FIG. 2.

ERIC, REP, and BOX-PCR fingerprints of a representative strain from each of the nine major genotypes found in this study. The genotypes are indicated by G followed by the number of the genotype. All dendrograms were produced by using Pearson's product-moment coefficient and a UPGMA clustering algorithm.

FIG. 3.

Dendrogram of 178 BOX-PCR fingerprints of R. palustris strains produced by using Pearson's product-moment coefficient and a UPGMA clustering algorithm. The genotypes are indicated by G followed by the number of the genotype. Major clusters of strains associated with one site or group of sites are indicated by the gray vertical bars. The control data show the level (∼85%) at which the fingerprint of a single strain (Azorhizobium caulinodans ORS-571) can vary across PCRs and gels. The vertical line indicates 80% similarity among fingerprints, which was used to define groups of related genotypes. Numbers in parentheses indicate the proportion of isolates in the indicated genotypes (G1 to G9) that came from the indicated site(s) (A to E).

FIG. 4.

Comparison of BOX-PCR and 16S rRNA gene sequences for a subset of strains (66 isolates). Both dendrograms were generated as described in the legend to Fig. 2. Numbers in parentheses indicate the proportion of strains from each similarity grouping that originated in the indicated site(s).