Expanding success in the isolation of abundant marine bacteria after reduction in grazing and viral pressure and increase in nutrient availability

21 pages, 6 figures, 1 table.-- Data Availability: All supplemental figures and tables can be found on GitHub (https://github.com/x-rv/Manuscript-2023). The 16S rRNA gene sequences of the isolates obtained in this study were deposited in GenBank under accession numbers OP342842 to OP344484. Amplicon sequencing data of the V4-V5 region of the 16S rRNA gene used in this study are publicly available in the European Nucleotide Archive under BioProject PRJEB60085Isolation of microorganisms is a useful approach to gathering knowledge about their genomic properties, physiology, and ecology, in addition to allowing the characterization of novel taxa. We performed an extensive isolation effort on samples from seawater manipulation experiments that were carried out during the four astronomical seasons in a coastal site of the northwest Mediterranean to evaluate the impact of grazing, viral mortality, resource competition reduction, and light presence/absence on bacterioplankton growth. Isolates were retrieved using two growth media, and their full 16S rRNA gene was sequenced to assess their identity and calculate their culturability across seasons and experimental conditions. A total of 1,643 isolates were obtained, mainly affiliated to the classes Gammaproteobacteria (44%), Alphaproteobacteria (26%), and Bacteroidia (17%). Isolates pertaining to class Gammaproteobacteria were the most abundant in all experiments, while Bacteroidia were preferentially enriched in the treatments with reduced grazing. Sixty-one isolates had a similarity below 97% to cultured taxa and are thus putatively novel. Comparison of isolate sequences with 16S rRNA gene amplicon sequences from the same samples showed that the percentage of reads corresponding to isolates was 21.4% within the whole data set, with dramatic increases in the summer virus-reduced (71%) and diluted (47%) treatments. In fact, we were able to isolate the top 10 abundant taxa in several experiments and from the whole data set. We also show that top-down and bottom-up controls differentially affect taxa in terms of culturability. Our results indicate that culturing marine bacteria using agar plates can be successful in certain ecological situationsThis research was supported by grant RTI2018-101974-B-C22 funded by the Spanish Ministry of Economy and Competitiveness and grants TM2015-70340-R, RTI2018-101025-B-100 and PID2021-125469NB-C32 funded by the Spanish Ministry of Science and Innovation. This work acknowledges the Severo Ochoa Centre of Excellence accreditation (CEX2019-000928-S). Xavier Rey-Velasco was supported by Spanish FI-SDUR and FPU grantsPeer reviewe