Multiple Streptomyces species with distinct secondary metabolomes have identical 16S rRNA gene sequences

Abstract

Microbial diversity studies using small subunit (SSU) rRNA gene sequences continue to advance our understanding of biological and ecological systems. Although a good predictor of overall diversity, using this gene to infer the presence of a species in a sample is more controversial. Here, we present a detailed polyphasic analysis of 10 bacterial strains isolated from three coastal lichens Lichina confinis, Lichina pygmaea and Roccella fuciformis with SSU rRNA gene sequences identical to the type strain of Streptomyces cyaneofuscatus. This analysis included phenotypic, microscopic, genetic and genomic comparisons and showed that despite their identical SSU rRNA sequences the strains had markedly different properties, and could be distinguished as 5 different species. Significantly, secondary metabolites profiles from these strains were also found to be different. It is thus clear that SSU rRNA based operational taxonomy units, even at the most stringent cut-off can represent multiple bacterial species, and that at least for the case of Streptomyces, strain de-replication based on SSU gene sequences prior to screening for bioactive molecules can miss potentially interesting novel molecules produced by this group that is notorious for the production of drug-leads.

Figure 1

Colony morphology on oatmeal agar (ISP 3) plates of each of the color group recorded from the 10 test strains. (A) Edge of colony showing aerial spore mass & diffusible pigments, and (B) Colony reverse.

Figure 2

Tree showing diversity between the strains with identical SSU rRNA gene sequences. Branch colors represent the lichen sources as follows: blue R. fuciformis, green L. confinis, red L. pygmaea. Phylogenetic tree inferred on concatenated sequences of the 5 protein coding sequences (atpD, gyrB, recA, rpoB and trpD) using neighbor-joining analyses. Confidence in tree topology was calculated by bootstrap test amongst 1000 replicates. There were a total of 3844 positions in the final dataset.

Figure 3

Upper triangular matrix of mean ΔTm values. ΔTm is the difference in melting temperature of the homologous genomic DNA and hybrid DNA preparation corresponding to their 50% decrease in fluorescence in melting curves. The values in the brackets are standard error of the mean between 3 replicates. Hybrid pairs showing less than 5 degree difference are considered as same genomic species.