Planctomycetes as novel source of bioactive molecules

Marine environments are a fruitful source of bioactive compounds some of which are the newest leading drugs in medicinal therapeutics. Of particular importance are organisms like sponges and macroalgae and their associated microbiome. Planctomycetes, abundant in macroalgae biofilms, are promising producers of bioactive compounds since they share characteristics, like large genomes and complex life cycles, with the most bioactive bacteria, the Actinobacteria. Furthermore, genome mining revealed the presence of secondary metabolite pathway genes or clusters in 13 analyzed Planctomycetes genomes. In order to assess the antimicrobial production of a large and diverse collection of Planctomycetes isolated from macroalgae from the Portuguese coast, molecular, and bioactivity assays were performed in 40 bacteria from several taxa. Two genes commonly associated with the production of bioactive compounds, nonribosomal peptide synthetases (NRPS), and polyketide synthases (PKS) genes were screened. Molecular analysis revealed that 95% of the planctomycetes potentially have one or both secondary bioactive genes; 85% amplified with PKS-I primers and 55% with NRPS primers. Some of the amplified genes were confirmed to be involved in secondary metabolite pathways. Using bioinformatic tools their biosynthetic pathways were predicted. The secondary metabolite genomic potential of strains LF1, UC8, and FC18 was assessed using in silico analysis of their genomes. Aqueous and organic extracts of the Planctomycetes were evaluated for their antimicrobial activity against an environmental Escherichia coli, E. coli ATCC 25922, Pseudomonas aeruginosa ATCC 27853, Staphylococcus aureus ATCC 25923, Bacillus subtilis ATCC 6633, and a clinical isolate of Candida albicans. The screening assays showed a high number of planctomycetes with bioactive extracts revealing antifungal (43%) and antibacterial (54%) activity against C. albicans and B. subtilis, respectively. Bioactivity was observed in strains from Rhodopirellula lusitana, R. rubra, R. baltica, Roseimaritima ulvae, and Planctomyces brasiliensis. This study confirms the bioactive capacity of Planctomycetes to produce antimicrobial compounds and encourages further studies envisaging molecule isolation and characterization for the possible discovery of new drugs. © 2016 Graça, Calisto and Lage.This research was partially supported by the Strategic Funding UID/Multi/04423/2013 through national funds provided by FCT—Foundation for Science and Technology and European Regional Development Fund (ERDF), in the framework of the programme PT2020 and by the Structured Program of R&D&I INNOVMAR – Innovation and Sustainability in the Management and Exploitation of Marine Resources (reference NORTE-01-0145-FEDER-000035, Research Line NOVELMAR), funded by the Northern Regional Operational Programme (NORTE2020) through the European Regional Development Fund (ERDF)

Planctomycetes as a vital constituent of the microbial communities inhabiting different layers of the meromictic lake sælenvannet (Norway)

Meromictic lakes are permanently stratified lakes that display steep gradients in salinity, oxygen and sulphur compounds tightly linked to bacterial community structure and diversity. Lake Sælenvannet is a meromictic lake located south of Bergen, Norway. The 26 m deep lake is connected to the open sea and permanently stratified into two layers separated by a chemocline. The upper water layer is brackish with major input from water runoff from the surroundings. The bottom layer consists of old saline water with low or no oxygen concentrations. Bacteria from phylum Planctomycetes are reported to be ubiquitous in lake environments. They are involved in the degradation of complex carbon sources in aquatic environments and are also linked to anaerobic processes such as fermentation and sulphur reduction. To study Planctomycete distribution along a chemical gradient, we sampled the water column throughout Lake Sælenvannet in 2012 and profiled the microbial community using 16S rRNA amplicon sequencing (metabarcoding) with 454 pyrosequencing. Planctomycetes related 16S rRNA gene sequences were found to be present both in the oxic and anoxic parts of the lake and showed an uneven distribution throughout the water column, with the highest relative abundance of 10% found in the saline anoxic layer at 15 m depth. In a follow-up study in 2014, samples from eight different depths were collected for enrichment and isolation of novel Planctomycetes. This study resulted in successful isolation in pure culture of 10 isolates affiliated to four different genera from the family Planctomycetaceae. One strain closely related to Blastopirellula cremea was isolated from 9 m depth, and two novel strains affiliated to the genera Stieleria and Gimesia were isolated at 7 and 9 m depths, respectively. Furthermore, seven isolates with identical 16S rRNA gene sequences were retrieved from seven different depths which varied greatly in salinity and chemical composition. These isolates likely represent a new species affiliated to Rubinisphaera. The adaptation of this novel Planctomycete to water depths spanning the entire chemical gradient could indicate a high phenotypic plasticity and/or a very efficient survival strategy. Overall, our results show the presence of a diverse group of Planctomycetes in Lake Sælenvannet, with a strong potential for novel adaptations to chemical stress factors.This study was supported through the NRC funded CaveIce project number 531891 and by the Department of Biological Sciences, University of Bergen, Norway. The sequencing was done at the Norwegian Sequencing Centre in Oslo, Norway

Feasibility of planctomycetes as a nutritional or supplementary food source for Daphnia spp

Daphnia magna is widely used as a standard organism in ecotoxicology assays. It plays a key role in energy transfer in freshwater food webs as a primary consumer, grazing on microalgae, yeast and bacteria. Daphnids are commonly reared in the laboratory using microalgae cultures but alternative or complementary sources are important to reduce the dependency on a single food source. The role played in nature by planctomycetes as a food source for other higher trophic levels is still unknown. In this study, we aimed to evaluate the potential of Rhodopirellula rubra strain LF2 as a nutritional or a supplementary food source for D. magna and Daphnia longispina. Life-history assays were conducted with daphnids fed with R. rubra in exponential and stationary growth phases, in three concentrations. Additionally, its adequacy as a supplement to the microalga Raphidocelis subcapitata was tested. In general, both daphnids showed impairment in all the parameters evaluated, especially when fed with R. rubra. However, when daphnids were fed with the two food sources, no changes were recorded for the rate of population increase. At the tested concentrations, R. rubra was not a good alternative food source in the daphnid diet. © EDP Sciences, 2016.This research was partially supported by the Strategic Funding UID/Multi/04423/2013 through national funds provided by FCT (Foundation for Science and Technology) and the European Regional Development Fund (ERDF), in the framework of the programme PT2020

Current screening methodologies in drug discovery for selected human diseases

The increase of many deadly diseases like infections by multidrug-resistant bacteria implies re-inventing the wheel on drug discovery. A better comprehension of the metabolisms and regulation of diseases, the increase in knowledge based on the study of disease-born microorganisms’ genomes, the development of more representative disease models and improvement of techniques, technologies, and computation applied to biology are advances that will foster drug discovery in upcoming years. In this paper, several aspects of current methodologies for drug discovery of antibacterial and antifungals, anti-tropical diseases, antibiofilm and antiquorum sensing, anticancer and neuroprotectors are considered. For drug discovery, two different complementary approaches can be applied: classical pharmacology, also known as phenotypic drug discovery, which is the historical basis of drug discovery, and reverse pharmacology, also designated target-based drug discovery. Screening methods based on phenotypic drug discovery have been used to discover new natural products mainly from terrestrial origin. Examples of the discovery of marine natural products are provided. A section on future trends provides a comprehensive overview on recent advances that will foster the pharmaceutical industry. © 2018 by the authors.Acknowledgments: This work was funded by the H2020-TWINN-2015 project BLUEandGREEN (grant 692419). We acknowledge the FCT Project UID/Multi/04423/2013 and the Interreg Atlantic Area project BLUEHUMAN (EAPA_151/2016) funded by the European Regional Development Fund

Pink‐ and orange‐pigmented Planctomycetes produce saproxanthin‐type carotenoids including a rare C45 carotenoid

Planctomycetes, are ubiquitous and environmentally important Gram-negative aquatic bacteria with key roles in global carbon and nitrogen cycles. Many planctomycetal species have a pink or orange colour and have been suggested to produce carotenoids. Potential applications as food colorants or anti-oxidants have been proposed. Hitherto, the planctomycetal metabolism is largely unexplored and the strain pigmentation has not been identified. For a holistic view on the complex planctomycetal physiology we analyzed carotenoid profiles of the pink-pigmented strain Rhodopirellula rubra LF2T and of the orange strain Rubinisphaera brasiliensis Gr7. During LC-MS/MS analysis of culture extracts we were able to identify three saproxanthin-type carotenoids including a rare C45 carotenoid. These compounds, saproxanthin, dehydroflexixanthin and 2’-isopentenyldehydrosaproxanthin, derive from the common carotenoid precursor lycopene and are characterized by related end groups, namely a 3-hydroxylated β-carotene-like cyclohexene ring as one end group and simple hydration on the other end of the molecule. Based on the observed molecule structure we present putative pathways for their biosynthesis. Results support Planctomycetes as a promising, yet mostly untapped source of carotenoids

Bioactivities and extract dereplication of actinomycetales isolated from marine sponges

In the beginning of the twenty-first century, humanity faces great challenges regarding diseases and health-related quality of life. A drastic rise in bacterial antibiotic resistance, in the number of cancer patients, in the obesity epidemics and in chronic diseases due to life expectation extension are some of these challenges. The discovery of novel therapeutics is fundamental and it may come from underexplored environments, like marine habitats, and microbial origin. Actinobacteria are well-known as treasure chests for the discovery of novel natural compounds. In this study, eighteen Actinomycetales isolated from marine sponges of three Erylus genera collected in Portuguese waters were tested for bioactivities with the main goal of isolating and characterizing the responsible bioactive metabolites. The screening comprehended antimicrobial, anti-fungal, anti-parasitic, anti-cancer and anti-obesity properties. Fermentations of the selected strains were prepared using ten different culturing media. Several bioactivities against the fungus Aspergillus fumigatus, the bacteria Staphylococcus aureus methicillin-resistant (MRSA) and the human liver cancer cell line HepG2 were obtained in small volume cultures. Screening in higher volumes showed consistent anti-fungal activity by strain Dermacoccus sp. #91-17 and Micrococcus luteus Berg02-26. Gordonia sp. Berg02-22.2 showed anti-parasitic (Trypanosoma cruzi) and anti-cancer activity against several cell lines (melanoma A2058, liver HepG2, colon HT29, breast MCF7 and pancreatic MiaPaca). For the anti-obesity assay, Microbacterium foliorum #91-29 and #91-40 induced lipid reduction on the larvae of zebrafish (Danio rerio). Dereplication of the extracts from several bacteria showed the existence of a variety of secondary metabolites, with some undiscovered molecules. This work showed that Actinomycetales are indeed good candidates for drug discovery.This research was partially supported by the Strategic Funding UID/Multi/04423/2013 through national funds provided by FCT – Foundation for Science and Technology and European Regional Development Fund (ERDF), in the framework of the programme PT2020, the EU H2020-TWINN-2015, BLUEandGREEN – Boosting scientific excellence and innovation capacity in biorefineries based on marine resources (Project No. 692419) and the European ERA-NET Marine Biotechnology project CYANOBESITY (ERA-MBT/0001/2015), financed by national funds through FCT (Foundation for Science and Technology, Portugal). Ralph Urbatzka was supported by a FCT postdoc grant (SFRH/BPD/112287/2015). The MEDINA authors disclosed the receipt of financial support from Fundación MEDINA, a public-private partnership of Merck Sharp & Dohme de España S.A./Universidad de Granada/Junta de Andalucía. Moreover, some of the equipment used in this work was supported by the Ministerio de Ciencia e Innovación and the European Union (Grant INP-2011-0016-PCT-010000-ACT6)