Dilution-to-Extinction Platform for the Isolation of Marine Bacteria-Producing Antitumor Compounds.

Gram-negative marine bacteria are an underexplored source of new chemical entities for a wide range of applications. Even though, some have shown a high antitumor activity. This chapter describes an isolation and screening protocol based on the Dilution-to-Extinction approach coupled with an antiproliferative test oriented to the discovery of new cytotoxic compounds synthesized by marine bacteria. In addition to the discovery of new bioactive secondary metabolites, this protocol provides a high-throughput isolation and screening platform for discarding no bioactive strains during the first steps of the drug discovery process.

Detection of a pederin-like compound using a dilution-to-extinction-based platform for the isolation of marine bacteria in drug discovery strategies.

The continued development of culturing technologies for the discovery of new molecules from marine microbes is of paramount importance for drug discovery. Coupled with this, the use of the high-throughput approach shows promise for increasing the number of Gram-negative and non-filamentous bacteria cultures that can be surveyed, since they show a lower potential of bioactivity. In this work, we propose a new strategy of high-throughput cultivation of bacteria inspired by a dilution-to-extinction (DTE) methodology for the isolation of, and screening for, new cytotoxic compound producing marine bacteria. A marine sponge tissue was directly used as inoculum and the results were compared with the data obtained through the direct plating isolation method. Enterobacterial repetitive intergenic consensus polymerase chain reaction (ERIC-PCR) genomic fingerprinting indicated the isolation of four bioactive strains, three of them producers of a pederin-like compound, and the fourth one able to synthesize a different compound, still unidentified, rendered by the DTE approach, in comparison with one bioactive strain identified through the plating method. Analyses based on the 16S rRNA gene data showed the existence of two different species belonging to the genus Labrenzia. The efficiency and diversity ratio in the number of isolates and compounds are discussed. In view of the results, the proposed DTE approach proved to be efficient for the isolation of new cytotoxic compounds of marine origin and pave the way for future potential applications.