Acylhomoserine lactone (AHLs)-mediated quorum-sensing (QS) processes seem to be common in the marine environment and among marine pathogenic bacteria, but no data are available on the prevalence of bacteria capable of interfering with QS in the sea, a process that has been generally termed 'quorum quenching' (QQ). One hundred and sixty-six strains isolated from different marine dense microbial communities were screened for their ability to interfere with AHL activity. Twenty-four strains (14.4%) were able to eliminate or significantly reduce N-hexanoyl-l-homoserine lactone activity as detected by the biosensor strain Chromobacterium violaceum CV026, a much higher percentage than that reported for soil isolates, which reinforces the ecological role of QS and QQ in the marine environment. Among these, 15 strains were also able to inhibit N-decanoyl-l-homoserine lactone activity and all of them were confirmed to enzymatically inactivate the AHL signals by HPLC-MS. Active isolates belonged to nine different genera of prevalently or exclusively marine origin, including members of the Alpha- and Gammaproteobacteria (8), Actinobacteria (2), Firmicutes (4) and Bacteroidetes (1). Whether the high frequency and diversity of cultivable bacteria with QQ activity found in near-shore marine isolates reflects their prevalence among pelagic marine bacterial communities deserves further investigation in order to understand the ecological importance of AHL-mediated QS and QQ processes in the marine environment.

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Many bacteria use quorum sensing (QS) to coordinate responses to environmental changes. In Gram-negative bacteria, the most extensively studied QS systems rely on the use of N-acylhomoserine lactones (AHLs) signal molecules. Some bacteria produce enzymes that are able to inactivate AHL signals produced by other bacteria and hence interfere with QS-mediated processes via a phenomenon known as quorum quenching. Acylase-type AHL degradation activity has been found in the biomass of the filamentous nitrogen-fixing cyanobacterium Anabaena (Nostoc) sp. PCC 7120, being absent from the culture media. The gene all3924 has been identified and cloned whose product exhibits homology to the acylase QuiP of Pseudomonas aeruginosa PAO1, demonstrating that it is at least partially responsible for the AHL-acylase activity. The recombinant enzyme, which was named auto-inducer inhibitor from Cyanobacteria (AiiC), shows broad acyl-chain length specificity. Because the presence of AHLs in the biomass of nitrogen-fixing cultures of Anabaena sp. PCC 7120 has been described recently, AiiC could represent a self-modulatory system to control the response to its own QS signals but could also be involved in the interference of signalling within complex microbial communities in which Cyanobacteria are present.

Acinetobacter baumannii presents a typical luxI/luxR quorum sensing (QS) system (abaI/abaR) but the acyl-homoserine lactone (AHL) signal profile and factors controlling the production of QS signals in this species have not been determined yet. A very complex AHL profile was identified for A. baumannii ATCC17978 as well as for A. nosocomialis M2, but only when cultivated under static conditions, suggesting that surface or cell-to-cell contact is involved in the activation of the QS genes. The analysis of A. baumanni clinical isolates revealed a strain-specific AHL profile that was also affected by nutrient availability. The concentration of OHC12-HSL, the major AHL found in A. baumannii ATCC17978, peaked upon stationary-phase establishment and decreases steeply afterwards. Quorum quenching (QQ) activity was found in the cell extracts of A. baumannii ATCC17978, correlating with the disappearance of the AHLs from the culture media, indicating that AHL concentration may be self-regulated in this pathogen. Since QQ activity was observed in strains in which AidA, a novel α/β-hydrolase recently identified in A. baumannii, is not present, we have searched for additional QQ enzymes in A. baumannii ATCC17978. Seven putative AHL-lactonase sequences could be identified in the genome and the QQ activity of 3 of them could be confirmed. At least six of these lactonase sequences are also present in all clinical isolates as well as in A. nosocomialis M2. Surface-associated motility and biofilm formation could be blocked by the exogenous addition of the wide spectrum QQ enzyme Aii20J. The differential regulation of the QQ enzymes in A. baumannii ATCC17978 and the full dependence of important virulence factors on the QS system provides a strong evidence of the importance of the AHL-mediated QS/QQ network in this species.

The freshwater microalga Haematococcus pluvialis is one of the best microbial sources of the carotenoid astaxanthin, but this microalga shows low growth rates and low final cell densities when cultured with traditional media. A single-variable optimization strategy was applied to 18 components of the culture media in order to maximize the productivity of vegetative cells of H. pluvialis in semicontinuous culture. The steady-state cell density obtained with the optimized culture medium at a daily volume exchange of 20% was 3.77 x 10(5) cells ml(-1), three times higher than the cell density obtained with Bold basal medium and with the initial formulation. The formulation of the optimal Haematococcus medium (OHM) is (in g l(-1)) KNO3 0.41, Na2HPO4 0.03, MgSO4 x 7H2O 0.246, CaCl2 x 2H2O 0.11, (in mg l(-1)) Fe(III)citrate x H2O 2.62, CoCl2 x 6H2O 0.011, CuSO4 x 5H2O 0.012, Cr2O3 0.075, MnCl2 x 4H2O 0.98, Na2MoO4 x 2H2O 0.12, SeO2 0.005 and (in microg l(-1)]) biotin 25, thiamine 17.5 and B12 15. Vanadium, iodine, boron and zinc were demonstrated to be non-essential for the growth of H. pluvialis. Higher steady-state cell densities were obtained by a three-fold increase of all nutrient concentrations but a high nitrate concentration remained in the culture medium under such conditions. The high cell productivities obtained with the new optimized medium can serve as a basis for the development of a two-stage technology for the production of astaxanthin from H. pluvialis.

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Acinetobacter baumannii is an important pathogen that causes nosocomial infections generally associated with high mortality and morbidity in Intensive Care Units (ICUs). Currently, little is known about the Quorum Sensing (QS)/Quorum Quenching (QQ) systems of this pathogen. We analyzed these mechanisms in seven clinical isolates of A. baumannii. Microarray analysis of one of these clinical isolates, Ab1 (A. baumannii ST-2_clon_2010), previously cultured in the presence of 3-oxo-C12-HSL (a QS signalling molecule) revealed a putative QQ enzyme (α/ß hydrolase gene, AidA). This QQ enzyme was present in all non-motile clinical isolates (67% of which were isolated from the respiratory tract) cultured in nutrient depleted LB medium. Interestingly, this gene was not located in the genome of the only motile clinical strain growing in this medium (A. baumannii strain Ab421_GEIH-2010 [Ab7], isolated from a blood sample). The AidA protein expressed in E. coli showed QQ activity. Finally, we observed downregulation of the AidA protein (QQ system attenuation) in the presence of H2O2 (ROS stress). In conclusion, most of the A. baumannii clinical strains were not surface motile (84%) and were of respiratory origin (67%). Only the pilT gene was involved in surface motility and related to the QS system. Finally, a new QQ enzyme (α/ß hydrolase gene, AidA protein) was detected in these strains.

The abundance of quorum quenching (QQ) activity was evaluated in cultivable bacteria obtained from oceanic and estuarine seawater and compared with the frequency of QQ enzyme sequences in the available marine metagenomic collections. The possible role of the high QQ activity found among marine bacteria is discussed. The low bacterial population encountered in the open sea and the low chemical stability of N-acyl-homoserine lactones (AHLs) in seawater have led to the suggestion that the AHL-mediated quorum sensing (QS) activity may be concentrated in specific microhabitats in the marine environment (3, 11) and that, therefore, the quorum quenching (QQ) processes would be expected to be less frequent in seawater. The presence of AHLs in marine particulate organic carbon has been recently confirmed, with more than 10% of the particle-associated bacteria being identified as AHL producers (10). Even higher QS activity values have been found for isolates from subtidal biofilms (31% [12]) and sponges (20 to 56% [14]), while a similar value was found for marine snow and microalgal isolates (9.3% [9]). Moreover, the addition of exogenous AHLs to incubations containing marine organic particles stimulated the activity of some key hydrolytic enzymes, suggesting a role of QS signals in the regulation of the degradation of particulate organic carbon (10). In a recent work, the importance of QQ processes in marine dense microbial communities was evidenced by the large amount of cultivable marine bacteria active against AHL signals that could be isolated from marine habitats such as sediments, biofilms, and the surface of the alga Fucus vesiculosus (18). These results indicate that QQ could be a usual strategy adopted in the marine environment to achieve competitive advantages at least in surfaces such as biofilms and eukaryotic niches. As an approximation to evaluate the ecological significance of QQ processes in the marine environment and to elucidate if QQ processes are as abundant in marine seawater pelagic microbial communities as previously described for coastal dense communities (18), we studied the presence of QQ activity among isolates from estuarine and open-ocean seawater. This activity was compared with the frequency of sequences homologous to known QQ enzymes in the available long-read marine metagenomic collections.Quorum quenching activity among cultivable bacteria. In order to evaluate if QQ processes are as abundant among cultivable bacteria from marine seawater pelagic microbial communities as previously described for those isolated from marine surfaceassociated communities (18), 464 marine isolates were obtained from three different seawater samples using different culture media and temperatures and screened for detection of AHL-QQ activity using a bioassay based on Chromobacterium violaceum reporter strains as previously described (18). One of the samples was obtained from surface water at 10 m from the shore line in an estuary (42°0.33=0.45.32ЉN, 8°0.53=0.08.23ЉW, Spain), and the other two samples were obta...