Microbial Ecology Molecular Characterization of an Oil-Degrading Cyanobacterial Consortium Olga Sa ´nchez, Elia Diestra, Isabel Esteve and Jordi Mas Departament de Gene `tica i Microbiologia, Facultat de Cie `ncies, Universitat Auto ` noma de Barcelona, 08193, Bellaterra, Spain Received: 8 April 2005 / Accepted: 15 June 2005 / Online publication: 15 December 2005 Abstract Recent studies have shown that the cyanobacterium Microcoleus chthonoplastes forms a consortium with heterotrophic bacteria present within the cyanobacterial sheath. These studies also show that this consortium is able to grow in the presence of crude oil, degrading aliphatic heterocyclic organo-sulfur compounds as well as alkylated monocyclic and polycyclic aromatic hydro- carbons. In this work, we characterize this oil-degrading consortium through the analysis of the 16S rRNA gene sequences. We performed the study in cultures of Microcoleus grown in mineral medium and in cultures of the cyanobacterium grown in mineral medium sup- plemented with crude oil. The results indicate that most of the clones found in the polluted culture correspond to well-known oil-degrading and nitrogen-fixing micro- organisms, and belong to different phylogenetic groups, such as the Alpha, Beta, and Gamma subclasses of Proteobacteria, and the Cytophaga/Flavobacteria/Bacter- oides group. The control is dominated by one predom- inant organism (88% of the clones) closely affiliated to Pseudoxanthomonas mexicana (similarity of 99.8%). The presence of organisms closely related to well-known nitrogen fixers such as Rhizobium and Agrobacterium sug- gests that at least some of the cyanobacteria-associated heterotrophic bacteria are responsible for nitrogen fixation and degradation of hydrocarbon compounds inside the polysaccharidic sheath, whereas Microcoleus provides a habitat and a source of oxygen and organic matter. Introduction Several research groups [5, 17] attribute to cyanobacteria an important role in the biodegradation of organic pollutants. In fact, there is evidence that microbial communities dominated by cyanobacteria can be actively involved in oil degradation [2]. Observations made after oil spills in the Arabian Gulf showed that cyanobacteria grew, forming heavy thick mats on the top of the sedi- ments [4, 39]. Other studies have focused on the capa- city of cyanobacteria isolates to degrade hydrocarbons. Cerniglia et al. [9, 10] observed the degradation of naphthalene, a major component of the water-soluble fraction of crude oil, and biphenyl, by the same strain of Oscillatoria. It has also been reported that phenan- threne can be metabolized by the unicellular marine cyanobacterium Agmenellum quadruplicatum [27]. Also, Oscillatoria salina, Plectonema tenebrans, and Aphano- capsa sp. degraded crude oil when grown in artificial medium and natural seawater [32]. However, it is by no means clear whether oil degradation is carried out by cyanobacteria alone or by heterotrophic bacteria associated to cyanobacteria. Some studies point to heterotrophic bacteria associated to cyanobacteria as coresponsible of hydrocarbon degrada- tion. Al-Hasan et al. [5] demonstrated that nonaxenic cyanobacterial samples containing Microcoleus chthono- plastes and Phormidium corium consumed and oxidized n-alkanes. They found that cyanobacterial growth steadi- ly declined with progressive axenity, and they identified four genera and species of associated heterotrophic bac- teria (Rhodococcus rhodochrous, Arthrobacter nicotianae, Pseudomonas sp., and Bacillus sp.) able to oxidize n- alkanes, although cyanobacteria directly contributed to hydrocarbon uptake and oxidation. On the other hand, Al-Hasan et al. [6] demonstrated that picocyanobacteria from the Arabian Gulf accumulated hydrocarbons from the water body, but did not utilize these compounds; the authors assumed that associated bacteria may be carrying Correspondence to: O. Sa ´nchez; E-mail: olga.sanchez@uab.es. DOI: 10.1007/s00248-005-5061-4 & Volume 50, 580–588 (2005) & * Springer Science+Business Media, Inc. 2005 580