Increased algicidal activity of Aeromonas veronii in response to Microcystis aeruginosa: interspecies crosstalk and secondary metabolites synergism.

Toxic Microcystis spp. blooms constitute a serious threat to water quality worldwide. Aeromonas veronii was isolated from Microcystis sp. colonies collected in Lake Kinneret. Spent Aeromonas media inhibits the growth of Microcystis aeruginosa MGK isolated from Lake Kinneret. The inhibition was much stronger when Aeromonas growth medium contained spent media from MGK suggesting that Aeromonas recognized its presence and produced secondary metabolites that inhibit Microcystis growth. Fractionations of the crude extract and analyses of the active fractions identified several secondary metabolites including lumichrome in Aeromonas media. Application of lumichrome at concentrations as low as 4 nM severely inhibited Microcystis growth. Inactivation of aviH in the lumichrome biosynthetic pathway altered the lumichrome level in Aeromonas and the extent of MGK growth inhibition. Conversely, the initial lag in Aeromonas growth was significantly longer when provided with Microcystis spent media but Aeromonas was able to resume normal growth. The longer was pre-exposure to Microcystis spent media the shorter was the lag phase in Aeromonas growth indicating the presence of, and acclimation to, secondary MGK metabolite(s) the nature of which was not revealed. Our study may help to control toxic Microcystis blooms taking advantage of chemical languages used in the interspecies communication.

Identification and characterization of haemofungin, a novel antifungal compound that inhibits the final step of haem biosynthesis.

OBJECTIVES: During recent decades, the number of invasive fungal infections among immunosuppressed patients has increased significantly, whereas the number of effective systemic antifungal drugs remains low and unsatisfactory. The aim of this study was to characterize a novel antifungal compound, CW-8/haemofungin, which we previously identified in a screen for compounds affecting fungal cell wall integrity. METHODS: The in vitro characteristics of haemofungin were investigated by MIC evaluation against a panel of pathogenic and non-pathogenic fungi, bacteria and mammalian cells in culture. Haemofungin mode-of-action studies were performed by screening an Aspergillus nidulans overexpression genomic library for resistance-conferring plasmids and biochemical validation of the target. In vivo efficacy was tested in the Galleria mellonella and Drosophila melanogaster insect models of infection. RESULTS: We demonstrate that haemofungin causes swelling and lysis of growing fungal cells. It inhibits the growth of pathogenic Aspergillus, Candida, Fusarium and Rhizopus isolates at micromolar concentrations, while only weakly affecting the growth of mammalian cell lines. Genetic and biochemical analyses in A. nidulans and Aspergillus fumigatus indicate that haemofungin primarily inhibits ferrochelatase (HemH), the last enzyme in the haem biosynthetic pathway. Haemofungin was non-toxic and significantly reduced mortality rates of G. mellonella and D. melanogaster infected with A. fumigatus and Rhizopus oryzae, respectively. CONCLUSIONS: Further development and in vivo validation of haemofungin is warranted.

Secondary Metabolites of Aeromonas veronii Strain A134 Isolated from a Microcystis aeruginosa Bloom.

Aeromonas veronii strain A134 was isolated from Microcystis aeruginosa colonies collected from Lake Kinneret (Sea of Galilee), Israel. The Aeromonas culture media inhibited the growth of M. aeruginosa (strain MGK). The crude extract of a large-scale culture of A. veronii A134 was separated in a few chromatographic steps to yield three new secondary metabolites, 9-chlorolumichrome (1), veronimide (2) and veronipyrazine (3), along with a known lumichrome and several known diketopiperazines. The structures of the new compounds were established by analyses of the data from 1D and 2D NMR experiments and HRMS data of the compounds, as well as a single-crystal x-ray analysis of synthetic 1. The structure elucidation and proposed biogenesis of the new compounds are described below.

Compounds from the marine sponge Cribrochalina vasculum offer a way to target IGF-1R mediated signaling in tumor cells.

In this work two acetylene alcohols, compound 1 and compound 2, which were isolated and identified from the sponge Cribrochalina vasculum, and which showed anti-tumor effects were further studied with respect to targets and action mechanisms. Gene expression analyses suggested insulin like growth factor receptor (IGF-1R) signaling to be instrumental in controlling anti-tumor efficacy of these compounds in non-small cell lung cancer (NSCLC). Indeed compounds 1 and 2 inhibited phosphorylation of IGF-1Rß as well as reduced its target signaling molecules IRS-1 and PDK1 allowing inhibition of pro-survival signaling. In silico docking indicated that compound 1 binds to the kinase domain of IGF-1R at the same binding site as the well known tyrosine kinase inhibitor AG1024. Indeed, cellular thermal shift assay (CETSA) confirmed that C. vasculum compound 1 binds to IGF-1R but not to the membrane localized tyrosine kinase receptor EGFR. Importantly, we demonstrate that compound 1 causes IGF-1Rß but not Insulin Receptor degradation specifically in tumor cells with no effects seen in normal diploid fibroblasts. Thus, these compounds hold potential as novel therapeutic agents targeting IGF-1R signaling for anti-tumor treatment.

Marine sponge Cribrochalina vasculum compounds activate intrinsic apoptotic signaling and inhibit growth factor signaling cascades in non-small cell lung carcinoma.

Marine-derived compounds have been explored and considered as possible antitumor agents. In this study, we analyzed extracts of the sponge Cribrochalina vasculum for their ability to inhibit tumor cell proliferation. Screening identified two acetylenic compounds of similar structure that showed strong tumor-specific toxicity in non-small cell lung carcinoma (NSCLC) cells and small-cell lung carcinoma cells, and less prominent toxicity in ovarian carcinoma, while having no effect on normal cells. These acetylenic compounds were found to cause a time-dependent increase in activation of apoptotic signaling involving cleavage of caspase-9, caspase-3, and PARP, as well as apoptotic cell morphology in NSCLC cells, but not in normal fibroblasts. Further analysis demonstrated that these compounds caused conformational change in Bak and Bax, and resulted in loss of mitochondrial potential and cytochrome c release in NSCLC cells. Moreover, a decreased phosphorylation of the growth factor signaling kinases Akt, mTOR, and ERK was evident and an increased phosphorylation of JNK was observed. Thus, these acetylenic compounds hold potential as novel therapeutic agents that should be further explored for NSCLC and other tumor malignancies.