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ABSTRACT Isolation 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 situations. IMPORTANCE Bottom-up and top-down controls greatly influence marine microbial community composition and dynamics, which in turn have effects on their culturability. We isolated a high amount of heterotrophic bacterial strains from experiments where seawater environmental conditions had been manipulated and found that decreasing grazing and viral pressure as well as rising nutrient availability are key factors increasing the success in culturing marine bacteria. Our data hint at factors influencing culturability and underpin bacterial cultures as a powerful way to discover new taxa.

Ramon Margalef was a pioneering scientist who introduced an interdisciplinary approach to ecological studies. His studies were among the first to incorporate various concepts in the literature of aquatic ecology, covering topics such as organisms, ecosystem interactions and evolution. To bring Margalef’s approach into current scientific studies, in this review we explore his vision of aquatic ecology within four interrelated fields of study: ecological theory, microbial diversity, biogeochemical cycles and global environmental changes. Taking inspiration from his studies, we analyse current scientific challenges and propose an integrated approach, considering the unifying concept of Margalef’s Mandala with the aim of improving future studies on aquatic microbial ecology.

Abstract Background Isolation of marine microorganisms is fundamental to gather information about their physiology, ecology and genomic content. To date, most of the bacterial isolation efforts have focused on the photic ocean leaving the deep ocean less explored. We have created a marine culture collection of heterotrophic bacteria (MARINHET) using a standard marine medium comprising a total of 1561 bacterial strains, and covering a variety of oceanographic regions from different seasons and years, from 2009 to 2015. Specifically, our marine collection contains isolates from both photic (817) and aphotic layers (744), including the mesopelagic (362) and the bathypelagic (382), from the North Western Mediterranean Sea, the North and South Atlantic Ocean, the Indian, the Pacific, and the Arctic Oceans. We described the taxonomy, the phylogenetic diversity and the biogeography of a fraction of the marine culturable microorganisms to enhance our knowledge about which heterotrophic marine isolates are recurrently retrieved across oceans and along different depths. Results The partial sequencing of the 16S rRNA gene of all isolates revealed that they mainly affiliate with the classes Alphaproteobacteria (35.9%), Gammaproteobacteria (38.6%), and phylum Bacteroidetes (16.5%). In addition, Alteromonas and Erythrobacter genera were found the most common heterotrophic bacteria in the ocean growing in solid agar medium. When comparing all photic, mesopelagic, and bathypelagic isolates sequences retrieved from different stations, 37% of them were 100% identical. This percentage increased up to 59% when mesopelagic and bathypelagic strains were grouped as the aphotic dataset and compared to the photic dataset of isolates, indicating the ubiquity of some bacterial isolates along different ocean depths. Finally, we isolated three strains that represent a new species, and the genome comparison and phenotypic characterization of two of these strains (ISS653 and ISS1889) concluded that they belong to a new species within the genus Mesonia. Conclusions Overall, this study highlights the relevance of culture-dependent studies, with focus on marine isolated bacteria from different oceanographic regions and depths, to provide a more comprehensive view of the culturable marine bacteria as part of the total marine microbial diversity.