Bioprospecting Portuguese Atlantic coast cyanobacteria for bioactive secondary metabolites reveals untapped chemodiversity

Abstract Continental Portugal has an extensive Atlantic Ocean coastline; however little is known about the diversity of its marine cyanobacteria, with only a few reports published. Cyanobacteria are a prolific source of bioactive compounds with promising therapeutic applications. Previously, several cyanobacterial strains (Subsections I–IV) were isolated from the Portuguese Atlantic coast and characterized using a polyphasic approach. Preliminary screening indicated that these cyanobacteria lacked the genes for proteins involved in the production of conventional cyanotoxins. However, it was previously shown that extracts of marine Synechocystis and Synechococcus were toxic to invertebrates, with crude extracts causing stronger effects than partially purified ones. To further evaluate the potential of our temperate region Portuguese isolates to produce bioactive compounds, a PCR screening for the presence of genes encoding non-ribosomal peptide synthetases (NRPSs) and polyketide synthases (PKSs), targeting the adenylation (A) and ketosynthase (KS) domains respectively, was performed. DNA fragments were obtained for more than 80% of the strains tested, and the results revealed that PKS genes are more ubiquitous than NRPS genes. The sequences obtained were used in an in silico prediction of the PKS and NRPS systems. RT-PCR analyses revealed that these genes are transcribed under routine laboratory conditions in several selected strains. Furthermore, LC–MS analysis coupled with molecular networking, a mass spectrometric tool that clusters metabolites with similar MS/MS fragmentation patterns, was used to search for novel or otherwise interesting metabolites revealing an untapped chemodiversity.