Taxogenomic and Comparative Genomic Analysis of the Genus Saccharomonospora Focused on the Identification of Biosynthetic Clusters PKS and NRPS

Abstract

Actinobacteria are prokaryotes with a large biotechnological interest due to their ability to produce secondary metabolites, produced by two main biosynthetic gene clusters (BGCs): polyketide synthase (PKS) and non-ribosomal peptide synthetase (NRPS). Most studies on bioactive products have been carried out on actinobacteria isolated from soil, freshwater or marine habitats, while very few have been focused on halophilic actinobacteria isolated from extreme environments. In this study we have carried out a comparative genomic analysis of the actinobacterial genus Saccharomonospora, which includes species isolated from soils, lake sediments, marine or hypersaline habitats. A total of 19 genome sequences of members of Saccharomonospora were retrieved and analyzed. We compared the 16S rRNA gene-based phylogeny of this genus with evolutionary relationships inferred using a phylogenomic approach obtaining almost identical topologies between both strategies. This method allowed us to unequivocally assign strains into species and to identify some taxonomic relationships that need to be revised. Our study supports a recent speciation event occurring between Saccharomonospora halophila and Saccharomonospora iraqiensis. Concerning the identification of BGCs, a total of 18 different types of BGCs were detected in the analyzed genomes of Saccharomonospora, including PKS, NRPS and hybrid clusters which might be able to synthetize 40 different putative products. In comparison to other genera of the Actinobacteria, members of the genus Saccharomonospora showed a high degree of novelty and diversity of BGCs.

Keywords: Saccharomonospora; actinobacteria; biosynthetic gene clusters; comparative genomic analysis; non-ribosomal peptide synthetase; polyketide synthase; secondary metabolites; taxophylogenomic analysis.

FIGURE 1

Neighbor-joining phylogenetic tree based on the 16S rRNA gene sequence comparison of members of the genus Saccharomonospora and the most closely related genus Prauserella. Bootstrap values ≥ 70% (based on 1,000 pseudo-replicates) are shown above the branches. Strains of Saccharomonospora were colored according to the group they belong to (blue for marine/lake, orange for moderately halophilic terrestrial, green for terrestrial and purple for clinical). Filled circles indicate clusters that were also recovered using maximum-parsimony and maximum-likelihood algorithms. Nocardia brasiliensis DSM 43758^T was used as an outgroup. Bar, 0.01 changes per nucleotide position.

FIGURE 2

Approximately maximum-likelihood phylogenomic tree based on the concatenation of the translated sequence of the 876 single-copy genes shared by the members of the genera Saccharomonospora and Prauserella under study. Bootstrap values ≥ 70% (based on Shimodaira-Hasegawa-like local support) are shown above the branches. Strains of Saccharomonospora were colored according to the group they belong to (blue for marine/lake, orange for moderately halophilic terrestrial, green for terrestrial and purple for clinical). Bar, 0.05 changes per nucleotide position.

FIGURE 3

Heatmap of genome relatedness among members of the genus Saccharomonospora estimated by means of OrthoANI (A) and dDDH (B) values.

FIGURE 4

Synteny plot among highly conserved (A) and partially conserved (B) genomes of Saccharomonospora. Only matches with ≥ 500 bp alignment length and ≥ 90% identity are shown.

FIGURE 5

Comparative pangenome display of the 19 Saccharomonospora strains analyzed in this study. The inner layers represent individual genomes organized regarding their isolation source and their NaCl requirements for growing. In the layers, dark colors indicate the presence of a gene cluster and light color its absence. The core- (978 genes) and the accessory- (11,297 genes) genomes are indicated in purple and read, respectively, in the outmost layer.

FIGURE 6

Pan- and core-genome size evolution within the genus Saccharomonospora. (A) Gene accumulation curves of the pan-genome (blue) and the core-genome (green). The curve is the least squares fit of the power law for the average values. (B) Number of new genes and fit curve (orange) with an increase in the number of Saccharomonospora genomes.

FIGURE 7

Circular diagram of Saccharomonospora BGCs diversity. Genomes of strains with identical BGCs pattern are collapsed in a single band. Each genome is represented by a different colored band (left half of the circle) that can be traced from the organism to the types of BGCs found in that genome (right half of the circle). The width of these bands indicates the number of BGCs of that type. The cluster types are also assigned colors that make up the outer ring next to each genome to easily see what portion of each genome is assigned to a specific BGC. Conversely, the outer ring next to the BGC categories show the proportion of that BGC attributed to each genome represented by the genome color. The RiPP category includes clusters identified as bacteriocin, lanthipeptide or lassopeptide. The Hybrid category includes gene clusters constituted by two or more BGC types. The “Others” category includes uncommon BGCs found in less than five genomes, i.e., betalactone, homoserine lactone, ladderane, linaridin and oligosaccharide. Strains of Saccharomonospora were colored according to the group they belong to (blue for marine/lake, orange for moderately halophilic terrestrial, green for terrestrial and purple for clinical) and the genome size is indicated in parentheses.