Fvwc1 and Fvwc2, orthologues of the wc-1 and wc-2 genes encoding for proteins of the white collar complex (WCC) in Neurospora crassa were cloned from Fusarium verticillioides and lack-of-function wc mutants were obtained by targeted gene disruption. Photo-conidiation was found to be absent in F. verticillioides, on the contrary, the wild type strain produced less conidia under continuous illumination than in the dark. Inactivation of any of the wc genes led to total female sterility, without affecting male fertility or asexual conidiation. No loss in colonization capability/invasive growth of the wc mutants was observed, when assessed on tomato fruits. Both Fvwc1 and Fvwc2 showed constitutive expression in the wild type cultures incubated in the dark and exposure to light caused only negligible increases in their transcription. Both Fvwc1 and Fvwc2 were down-regulated in a ΔFvmat1-2-1 gene disruption mutant, lacking a functional mating type (mat1-2-1) gene, suggesting that the MAT1-2-1 product has a positive regulatory effect on the white collar genes.

Conidium production was significantly reduced in three independent ΔFvMAT1-2-1 gene disruption mutants of F. verticillioides as compared to the wild type parental strain, when fungi were incubated on carrot agar either under diurnal illumination conditions or in total darkness. The conidiation specific con10 gene was expressed constitutively at low levels in the wild type strain, whereas levels of con10 transcripts were drastically reduced in the the ΔFvMAT1-2-1/M15 mutant. These findings demonstrate that mutants, lacking the functional mat1-2-1 mating type gene have lost not only their sexual reproduction capability, but became also retarded in asexual sporulation indicating that mating type genes have positive functions during the asexual phase of the fungal life cycle.

BACKGROUND:The luxS gene in Shewanella oneidensis was shown to encode an autoinducer-2 (AI-2)-like molecule, the postulated universal bacterial signal, but the impaired biofilm growth of a luxS deficient mutant could not be restored by AI-2, indicating it might not have a signalling role in this organism.FINDINGS:Here, we provide further evidence regarding the metabolic role of a luxS mutation in S. oneidensis. We constructed a luxS mutant and compared its phenotype to a wild type control with respect to its ability to remove AI-2 from the medium, expression of secreted proteins and biofilm formation. We show that S. oneidensis has a cell-dependent mechanism by which AI-2 is depleted from the medium by uptake or degradation at the end of the exponential growth phase. As AI-2 depletion is equally active in the luxS mutant and thus does not require AI-2 as an inducer, it appears to be an unspecific mechanism suggesting that AI-2 for S. oneidensis is a metabolite which is imported under nutrient limitation. Secreted proteins were studied by iTraq labelling and liquid chromatography mass spectrometry (LC-MS) detection. Differences between wild type and mutant were small. Proteins related to flagellar and twitching motility were slightly up-regulated in the luxS mutant, in accordance with its loose biofilm structure. An enzyme related to cysteine metabolism was also up-regulated, probably compensating for the lack of the LuxS enzyme. The luxS mutant developed an undifferentiated, loosely-connected biofilm which covered the glass surface more homogenously than the wild type control, which formed compact aggregates with large voids in between.CONCLUSIONS:The data confirm the role of the LuxS enzyme for biofilm growth in S. oneidensis and make it unlikely that AI-2 has a signalling role in this organism.

Background The autoinducer-2 (AI-2) group of signalling molecules are produced by both Gram positive and Gram negative bacteria as the by-product of a metabolic transformation carried out by the LuxS enzyme. They are the only non species-specific quorum sensing compounds presently known in bacteria. The luxS gene coding for the AI-2 synthase enzyme was found in many important pathogens. Here, we surveyed its occurrence in a collection of 165 marine isolates belonging to abundant marine phyla using conserved degenerated PCR primers and sequencing of selected positive bands to determine if the presence of the luxS gene is phylogenetically conserved or dependent on the habitat. Results The luxS gene was not present in any of the Alphaproteobacteria (n = 71) and Bacteroidetes strains (n = 29) tested; by contrast, these bacteria harboured the sahH gene, coding for an alternative enzyme for the detoxification of S -adenosylhomocysteine (SAH) in the activated methyl cycle. Within the Gammaproteobacteria (n = 76), luxS was found in all Shewanella, Vibrio and Alteromonas isolates and some Pseudoalteromonas and Halomonas species, while sahH was detected in Psychrobacter strains. A number of Gammaproteobacteria (n = 27) appeared to have neither the luxS nor the sahH gene. We then studied the production of AI-2 in the genus Shewanella using the Vibrio harveyi bioassay. All ten species of Shewanella tested produced a pronounced peak of AI-2 towards the end of the exponential growth phase in several media investigated. The maximum of AI-2 activity was different in each Shewanella species, ranging from 4% to 46% of the positive control. Conclusion The data are consistent with those of fully sequenced bacterial genomes and show that the potential for lu xS related signalling is dependent on phylogenetic affiliation rather than ecological niche and is largest in certain groups of Gammaproteobacteria in the marine environment. This is the first report on AI-2 production in Shewanella species; its signalling role in these organisms remains to be elucidated.

More than 100 bacterial isolates from various marine habitats were screened for AHL production by using gfp reporter constructs based on the lasR system of Pseudomonas aeruginosa and the luxR system of Vibrio fischeri. Of the 67 Alphaproteobacteria tested, most of which belonged into the so-called Roseobacter clade, 39 induced fluorescence in either one or both sensor strains up to 103-fold compared to controls. Acylated homoserine lactones were identified by GC-MS analysis and shown to have chain lengths of C8, C10, C13-C16, and C18. One or two double bonds were often present, while a keto or hydroxyl group occurred only rarely in the side chain. Most strains produced several different AHLs. C18-en-HSL and C18-dien-HSL were produced by Dinoroseobacter shibae, an aerobic anoxygenic phototrophic bacterium isolated from dinoflagellates, and are among the longest AHLs found to date. Z7-C14-en-HSL, which has previously been detected in Rhodobacter sphaeroides, was produced by Roseovarius tolerans and Jannaschia helgolandensis. This signal molecule was synthesised and shown to induce a similar response to the culture supernatant in the respective sensor strain. The widespread occurrence of quorum-sensing compounds in marine Alphaproteobacteria, both free-living strains and those associated to eukaryotic algae, points to a great importance of this signalling mechanism for the adaptation of the organisms to their widely different ecological niches.

Conidium production was significantly reduced in three independent ΔFvMAT1-2-1 gene disruption mutants of F. verticillioides as compared to the wild type parental strain, when fungi were incubated on carrot agar either under diurnal illumination conditions or in total darkness. The conidiation specific con10 gene was expressed constitutively at low levels in the wild type strain, whereas levels of con10 transcripts were drastically reduced in the the ΔFvMAT1-2-1/M15 mutant. These findings demonstrate that mutants, lacking the functional mat1-2-1 mating type gene have lost not only their sexual reproduction capability, but became also retarded in asexual sporulation indicating that mating type genes have positive functions during the asexual phase of the fungal life cycle.