Evolution of the streptomycin and viomycin biosynthetic clusters and resistance genes

The distribution of the streptomycin (strA) and viomycin (vph) resistance genes was examined in Streptomyces isolates. It was hypothesised that non-antibiotic producers that are niche competitors with producers will need to possess resistance to the antibiotic and will thus have acquired resistance genes. A detailed phylogenetic study, utilizing a novel multilocus sequence typing (MLST) scheme, was made of a collection of isolates and types strains with a Streptomyces griseus phenotype in addition to type strains from known producers of streptomycin and related compounds. strA and vph were found either within a biosynthetic gene cluster or independently. S. griseus strains possessing the streptomycin cluster formed part of a clonal complex and have been readily isolated from soil originating in every continent except Antarctica. Few copies of strA were detected in soil total community DNA, none of which were identical to the gene from the streptomycin cluster. All S. griseus strains possessing solely strA belonged to two clades and were closely related to streptomycin producers. The strA in the resistance-only strains is likely to have originated from the self-resistance gene of another aminoglycoside cluster and arrived in those S. griseus strains via horizontal gene transfer. S. griseus strains with only vph also formed two clades and were more distantly related to the producers than to one another. The high sequence divergence of the viomycin resistance genes also suggests that the vph homologue arrived in these two groups from another peptide antibiotic cluster via horizontal gene transfer. The expression of the strA gene was constitutive in resistance-only strains from both subgroups whereas streptomycin producers showed peak strA expression in late log phase which correlates with the switch on of streptomycin biosynthesis. One example of horizontal gene transfer of the streptomycin cluster was discovered, to a Streptomyces platensis strain, which contained a cluster with 84% sequence identity and almost identical gene structure and arrangement to that of the S. griseus cluster. Its expression pattern was also highly similar to that of S. griseus producers, but at a much lower level. Whilst there is evidence that antibiotics have diverse roles in nature, this work clearly supports the co-evolution of resistance in the presence of antibiotic biosynthetic capability within closely related soil dwelling bacteria. This reinforces the view that, for some antibiotics at least, the primary role is one of antibiosis during competition in soil for resources

Evolution of the streptomycin and viomycin biosynthetic clusters and resistance genes

The distribution of the streptomycin (strA) and viomycin (vph) resistance genes was examined in Streptomyces isolates. It was hypothesised that non-antibiotic producers that are niche competitors with producers will need to possess resistance to the antibiotic and will thus have acquired resistance genes. A detailed phylogenetic study, utilizing a novel multilocus sequence typing (MLST) scheme, was made of a collection of isolates and types strains with a Streptomyces griseus phenotype in addition to type strains from known producers of streptomycin and related compounds. strA and vph were found either within a biosynthetic gene cluster or independently. S. griseus strains possessing the streptomycin cluster formed part of a clonal complex and have been readily isolated from soil originating in every continent except Antarctica. Few copies of strA were detected in soil total community DNA, none of which were identical to the gene from the streptomycin cluster. All S. griseus strains possessing solely strA belonged to two clades and were closely related to streptomycin producers. The strA in the resistance-only strains is likely to have originated from the self-resistance gene of another aminoglycoside cluster and arrived in those S. griseus strains via horizontal gene transfer. S. griseus strains with only vph also formed two clades and were more distantly related to the producers than to one another. The high sequence divergence of the viomycin resistance genes also suggests that the vph homologue arrived in these two groups from another peptide antibiotic cluster via horizontal gene transfer. The expression of the strA gene was constitutive in resistance-only strains from both subgroups whereas streptomycin producers showed peak strA expression in late log phase which correlates with the switch on of streptomycin biosynthesis. One example of horizontal gene transfer of the streptomycin cluster was discovered, to a Streptomyces platensis strain, which contained a cluster with 84% sequence identity and almost identical gene structure and arrangement to that of the S. griseus cluster. Its expression pattern was also highly similar to that of S. griseus producers, but at a much lower level. Whilst there is evidence that antibiotics have diverse roles in nature, this work clearly supports the co-evolution of resistance in the presence of antibiotic biosynthetic capability within closely related soil dwelling bacteria. This reinforces the view that, for some antibiotics at least, the primary role is one of antibiosis during competition in soil for resources.EThOS - Electronic Theses Online ServiceGBUnited Kingdo

Designing and implementing an assay for the detection of rare and divergent NRPS and PKS clones in European, Antarctic and Cuban soils

The ever increasing microbial resistome means there is an urgent need for new antibiotics. Metagenomics is an underexploited tool in the field of drug discovery. In this study we aimed to produce a new updated assay for the discovery of biosynthetic gene clusters encoding bioactive secondary metabolites. PCR assays targeting the polyketide synthases (PKS) and non-ribosomal peptide synthetases (NRPS) were developed. A range of European soils were tested for their biosynthetic potential using clone libraries developed from metagenomic DNA. Results revealed a surprising number of NRPS and PKS clones with similarity to rare Actinomycetes. Many of the clones tested were phylogenetically divergent suggesting they were fragments from novel NRPS and PKS gene clusters. Soils did not appear to cluster by location but did represent NRPS and PKS clones of diverse taxonomic origin. Fosmid libraries were constructed from Cuban and Antarctic soil samples; 17 fosmids were positive for NRPS domains suggesting a hit rate of less than 1 in 10 genomes. NRPS hits had low similarities to both rare Actinobacteria and Proteobacteria; they also clustered with known antibiotic producers suggesting they may encode for pathways producing novel bioactive compounds. In conclusion we designed an assay capable of detecting divergent NRPS and PKS gene clusters from the rare biosphere; when tested on soil samples results suggest the majority of NRPS and PKS pathways and hence bioactive metabolites are yet to be discovere

<i>Streptomyces</i>, Greek Habitats and Novel Pharmaceuticals: A Promising Challenge

Bacteria of the genus Streptomyces produce a very large number of secondary metabolites, many of which are of vital importance to modern medicine. There is great interest in the discovery of novel pharmaceutical compounds derived from strepomycetes, since novel antibiotics, anticancer and compounds for treating other conditions are urgently needed. Greece, as proven by recent research, possesses microbial reservoirs with a high diversity of Streptomyces populations, which provide a rich pool of strains with potential pharmaceutical value. This review examines the compounds of pharmaceutical interest that have been derived from Greek Streptomyces isolates. The compounds reported in the literature include antibiotics, antitumor compounds, biofilm inhibitors, antiparasitics, bacterial toxin production inhibitors and antioxidants. The streptomycete biodiversity of Greek environments remains relatively unexamined and is therefore a very promising resource for potential novel pharmaceuticals

Streptomyces, Greek Habitats and Novel Pharmaceuticals: A Promising Challenge

Bacteria of the genus Streptomyces produce a very large number of secondary metabolites, many of which are of vital importance to modern medicine. There is great interest in the discovery of novel pharmaceutical compounds derived from strepomycetes, since novel antibiotics, anticancer and compounds for treating other conditions are urgently needed. Greece, as proven by recent research, possesses microbial reservoirs with a high diversity of Streptomyces populations, which provide a rich pool of strains with potential pharmaceutical value. This review examines the compounds of pharmaceutical interest that have been derived from Greek Streptomyces isolates. The compounds reported in the literature include antibiotics, antitumor compounds, biofilm inhibitors, antiparasitics, bacterial toxin production inhibitors and antioxidants. The streptomycete biodiversity of Greek environments remains relatively unexamined and is therefore a very promising resource for potential novel pharmaceuticals

Corrigendum to "Coevolution of antibiotic production and counter-resistance in soil bacteria"

P>We present evidence for the coexistence and coevolution of antibiotic resistance and biosynthesis genes in soil bacteria. The distribution of the streptomycin (strA) and viomycin (vph) resistance genes was examined in Streptomyces isolates. strA and vph were found either within a biosynthetic gene cluster or independently. Streptomyces griseus strains possessing the streptomycin cluster formed part of a clonal complex. All S. griseus strains possessing solely strA belonged to two clades; both were closely related to the streptomycin producers. Other more distantly related S. griseus strains did not contain strA. S. griseus strains with only vph also formed two clades, but they were more distantly related to the producers and to one another. The expression of the strA gene was constitutive in a resistance-only strain whereas streptomycin producers showed peak strA expression in late log phase that correlates with the switch on of streptomycin biosynthesis. While there is evidence that antibiotics have diverse roles in nature, our data clearly support the coevolution of resistance in the presence of antibiotic biosynthetic capability within closely related soil dwelling bacteria. This reinforces the view that, for some antibiotics at least, the primary role is one of antibiosis during competition in soil for resources

Soil sites screened for the <i>strA</i> and <i>atpD</i> genes [31], [32].

<p>Soil sites screened for the <i>strA</i> and <i>atpD</i> genes <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0035756#pone.0035756-Tolba1" target="_blank">[31]</a>, <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0035756#pone.0035756-Garcia1" target="_blank">[32]</a>.</p

Phosphotransferase diversity in microcosm soil based on partial sequences derived from <i>strA</i> primers.

<p>Marker sequences (from GenBank) in bold. (SP) = Confirmed streptomycin producer, (AP) = Confirmed producer of other aminoglycoside <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0035756#pone.0035756-DSMZ1" target="_blank">[51]</a>, <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0035756#pone.0035756-Wink1" target="_blank">[52]</a>. Accession numbers in parentheses. The tree was constructed using the neighbour-joining method; the numbers besides the branches indicate the percentage bootstrap value of 10000 replicates. The scale bar indicates 10% nucleotide dissimilarity.</p

Divergence of the (a) Mg²⁺ binding domain and (b) putative nucleotide-binding domain in environmental sequences compared to the <i>S. aureofaciens hur</i> peptide.

<p>Divergence of the (a) Mg²⁺ binding domain and (b) putative nucleotide-binding domain in environmental sequences compared to the <i>S. aureofaciens hur</i> peptide.</p

Level of significance, calculated using the χ² test, of the differences in number of <i>atpD</i> sequences belonging to each group (A = Group A, B = Group B, C = Group C,) and <i>strA</i> sequences belonging to each group (1 = Group 1, 2 = Group 2, 3 = Group 3, O = others) between the main soil sites.

<p>Significant probability values (p<0.05) are followed by an asterisk.</p