Streptomyces isolated from bird feathers as a potential source for novel antibiotics.

Streptomyces, the main source of antibiotics essential for human health, are widely distributed in nature among terrestrial, oceanic and atmospheric environments. New trends in antibiotic discovery are focused in the search for novel bioactive strains in unexplored habitats. We provide here evidence of the presence of diverse Streptomyces populations in wild bird feathers, such as the seagull, Larus michahellis, collected at Northern Spain; the sparrow, Passer domesticus, and the hoopoe, Upupa epops, both collected in Southern Spain. Taxonomic identification of fourteen bioactive strains, by sequencing their 16S rRNA gene and phylogenetic analyses, revealed that all of them are homologous to a total of 10 different Streptomyces. Strains from seagull samples are homologous to other antibiotic producers previously isolated from atmospheric, marine and terrestrial environments in the Cantabrian Sea region, Northern Spain. Isolates form Southern feather samples, from a house sparrow and a Eurasian hoopoe, are homologues to Streptomyces strains previously isolated mainly from soils along the Mediterranean region. The most relevant feature is that they are producers of diverse antibiotics with activity against Gram-positive, Gram-negative bacteria and fungi. We report here the successful activation of silent antibiotic biosynthetic pathways in response to changes in environmental conditions, such as incubation temperature and salinity of the culture medium, in agreement with the OSMAC approach, One Strain Many Compounds. The finding of bioactive Streptomyces in bird's plumage might be of relevance, not only in the ecology of Streptomyces-birds associations, but also in medicine and biotechnology since they can be regarded as a potential source for novel antibiotics.

NT-PROBNP as a screening tool for low-risk patent ductus arteriousus: a follow-up validation study.

The purpose of the study is to test whether NT-proBNP serves as a screening tool for low-risk patent ductus arteriosus and safely avoids routine early echocardiography. This is a prospective observational study in preterm infants ≤32 weeks of gestational age. Infants with ≥5100 pg/ml (positive screening) at 48-72 hours of life received comprehensive echocardiography and were treated according to shunt severity. Infants with NT-proBNP below 5100 pg/ml (negative screening) were managed expectantly. The main outcome was need for ductus treatment within the first 7 days of life. One hundred twenty-five infants were included; 82 had a negative NT-proBNP screening and 43 had a positive NT-proBNP screening. No infant (0%) with a negative screening was treated for ductus while 26 (60.4%) with a positive screening were treated (p < 0.001). NT-proBNP avoided a 65.6% of routine echocardiograms. NT-proBNP had an excellent performance to predict PDA treatment (AUC = 0.967).Conclusion: NT-proBNP at 48-72 hours of life has an excellent performance to detect low risk and avoids unnecessary echocardiograms. This may contribute to optimize PDA management in terms of resource utilization.

Desertomycin G, a New Antibiotic with Activity against Mycobacterium tuberculosis and Human Breast Tumor Cell Lines Produced by Streptomyces althioticus MSM3, Isolated from the Cantabrian Sea Intertidal Macroalgae Ulva sp.

The isolation and structural elucidation of a structurally new desertomycin, designated as desertomycin G (1), with strong antibiotic activity against several clinically relevant antibiotic resistant pathogens are described herein. This new natural product was obtained from cultures of the marine actinomycete Streptomyces althioticus MSM3, isolated from samples of the intertidal seaweed Ulva sp. collected in the Cantabrian Sea (Northeast Atlantic Ocean). Particularly interesting is its strong antibiotic activity against Mycobacterium tuberculosis clinical isolates, resistant to antibiotics in clinical use. To the best of our knowledge, this is the first report on a member of the desertomycin family displaying such activity. Additionally, desertomycin G shows strong antibiotic activities against other relevant Gram-positive clinical pathogens such as Corynebacterium urealyticum, Staphylococcus aureus, Streptococcus pneumoniae, Streptococcus pyogenes, Enterococcus faecium, Enterococcus faecalis, and Clostridium perfringens. Desertomycin G also displays moderate antibiotic activity against relevant Gram-negative clinical pathogens such as Bacteroides fragilis, Haemophilus influenzae and Neisseria meningitidis. In addition, the compound affects viability of tumor cell lines, such as human breast adenocarcinoma (MCF-7) and colon carcinoma (DLD-1), but not normal mammary fibroblasts.

New 3-Hydroxyquinaldic Acid Derivatives from Cultures of the Marine Derived Actinomycete Streptomyces cyaneofuscatus M-157.

Fractionation of the bioactive extract of a culture of the marine derived actinomycete Streptomyces cyaneofuscatus M-157 led to the isolation of the known 3-hydroxyquinaldic acid (4), its amide (5) and three new derivatives (1⁻3) containing different amino acid residues. The structures of the new molecules (1⁻3), including their absolute configuration, were determined by the analysis of their ESI-TOF MS and one-dimensional (1D) and two-dimensional (2D) NMR spectra and advanced Marfey's analysis of their hydrolyzation products. Compound 3 spontaneously dimerized in solution to give the disulfide derivative 6. Unfortunately, none of the new compounds isolated confirmed the antimicrobial activity found in the bacterial extract, perhaps indicating that such antibacterial activity might be due to presence in the extract at the trace level of larger bioactive 3-hydroxyquinaldic acid derivatives from which compounds 1⁻3 are biosynthetic precursors. Cytotoxicity tests confirmed the moderate and weak IC50 values of 15.6 and 51.5 µM for compounds 5 and 1, respectively.

Anthracimycin B, a Potent Antibiotic against Gram-Positive Bacteria Isolated from Cultures of the Deep-Sea Actinomycete Streptomyces cyaneofuscatus M-169.

The potent antimicrobial extract of a culture of the marine derived actinomycete Streptomyces cyaneofuscatus M-169 was fractionated by reversed phase flash chromatography and preparative HPLC to yield the new Gram-positive antibiotic, anthracimycin B (1), together with its congener, anthracimycin (2). The structure of the new compound was established by analysis of its ESI-TOF MS and 1D and 2D NMR spectra, and comparison with data published for anthracimycin and anthracimycin BII-2619 (3). Notably, anthracimycin seemed to be the major and almost unique component of the extract detected by HPLC-UV-MS, making our S. cyanofuscatus strain an excellent candidate for further biosynthetic studies of this potent antibiotic.

Pharmacological Potential of Phylogenetically Diverse Actinobacteria Isolated from Deep-Sea Coral Ecosystems of the Submarine Avilés Canyon in the Cantabrian Sea.

Marine Actinobacteria are emerging as an unexplored source for natural product discovery. Eighty-seven deep-sea coral reef invertebrates were collected during an oceanographic expedition at the submarine Avilés Canyon (Asturias, Spain) in a range of 1500 to 4700 m depth. From these, 18 cultivable bioactive Actinobacteria were isolated, mainly from corals, phylum Cnidaria, and some specimens of phyla Echinodermata, Porifera, Annelida, Arthropoda, Mollusca and Sipuncula. As determined by 16S rRNA sequencing and phylogenetic analyses, all isolates belong to the phylum Actinobacteria, mainly to the Streptomyces genus and also to Micromonospora, Pseudonocardia and Myceligenerans. Production of bioactive compounds of pharmacological interest was investigated by high-performance liquid chromatography (HPLC) and gas chromatography-mass spectrometry (GC-MS) techniques and subsequent database comparison. Results reveal that deep-sea isolated Actinobacteria display a wide repertoire of secondary metabolite production with a high chemical diversity. Most identified products (both diffusible and volatiles) are known by their contrasted antibiotic or antitumor activities. Bioassays with ethyl acetate extracts from isolates displayed strong antibiotic activities against a panel of important resistant clinical pathogens, including Gram-positive and Gram-negative bacteria, as well as fungi, all of them isolated at two main hospitals (HUCA and Cabueñes) from the same geographical region. The identity of the active extracts components of these producing Actinobacteria is currently being investigated, given its potential for the discovery of pharmaceuticals and other products of biotechnological interest.

Branimycins B and C, Antibiotics Produced by the Abyssal Actinobacterium Pseudonocardia carboxydivorans M-227.

Two new antibiotics, branimycins B (2) and C (3), were produced by fermentation of the abyssal actinobacterium Pseudonocardia carboxydivorans M-227, isolated from deep seawater of the Avilés submarine Canyon. Their structures were elucidated by HRMS and NMR analyses. These compounds exhibit antibacterial activities against a panel of Gram-positive bacteria, including Corynebacterium urealyticum, Clostridium perfringens, and Micrococcus luteus, and against the Gram-negative bacterium Neisseria meningitidis. Additionally, branimycin B displayed moderate antibacterial activity against other Gram-negative bacteria such as Bacteroides fragilis, Haemophilus influenzae, and Escherichia coli, and branimycin C against the Gram-positive Enterococcus faecalis and methicillin-sensitive and methicillin-resistant Staphylococcus aureus.

Paulomycin G, a New Natural Product with Cytotoxic Activity against Tumor Cell Lines Produced by Deep-Sea Sediment Derived Micromonospora matsumotoense M-412 from the Avilés Canyon in the Cantabrian Sea.

The present article describes a structurally novel natural product of the paulomycin family, designated as paulomycin G (1), obtained from the marine strain Micromonospora matsumotoense M-412, isolated from Cantabrian Sea sediments collected at 2000 m depth during an oceanographic expedition to the submarine Avilés Canyon. Paulomycin G is structurally unique since-to our knowledge-it is the first member of the paulomycin family of antibiotics lacking the paulomycose moiety. It is also the smallest bioactive paulomycin reported. Its structure was determined using HRMS and 1D and 2D NMR spectroscopy. This novel natural product displays strong cytotoxic activities against different human tumour cell lines, such as pancreatic adenocarcinoma (MiaPaca_2), breast adenocarcinoma (MCF-7), and hepatocellular carcinoma (HepG2). The compound did not show any significant bioactivity when tested against a panel of bacterial and fungal pathogens.

Lobophorin K, a New Natural Product with Cytotoxic Activity Produced by Streptomyces sp. M-207 Associated with the Deep-Sea Coral Lophelia pertusa.

The present article describes the isolation of a new natural product of the lobophorin family, designated as lobophorin K (1), from cultures of the marine actinobacteria Streptomyces sp. M-207, previously isolated from the cold-water coral Lophelia pertusa collected at 1800 m depth during an expedition to the submarine Avilés Canyon. Its structure was determined using a combination of spectroscopic techniques, mainly ESI-TOF MS and 1D and 2D NMR. This new natural product displayed cytotoxic activity against two human tumor cell lines, such as pancreatic carcinoma (MiaPaca-2) and breast adenocarcinoma (MCF-7). Lobophorin K also displayed moderate and selective antibiotic activity against pathogenic Gram-positive bacteria such as Staphylococcus aureus.

Atmospheric Dispersal of Bioactive Streptomyces albidoflavus Strains Among Terrestrial and Marine Environments.

Members of the Streptomyces albidoflavus clade, identified by 16S rRNA sequencing and phylogenetic analyses, are widespread among predominant terrestrial lichens (Flavoparmelia caperata and Xanthoria parietina) and diverse intertidal and subtidal marine macroalgae, brown red and green (Phylum Heterokontophyta, Rhodophyta, and Chlorophyta) from the Cantabrian Cornice. In addition to these terrestrial and coastal temperate habitats, similar strains were also found to colonize deep-sea ecosystems and were isolated mainly from gorgonian and solitary corals and other invertebrates (Phylum Cnidaria, Annelida, Echinodermata, Arthropoda, and Porifera) living up to 4700-m depth and at a temperature of 2-4 °C in the submarine Avilés Canyon. Similar strains have been also repeatedly isolated from atmospheric precipitations (rain drops, snow, and hailstone) collected in the same area throughout a year observation time. These ubiquitous strains were found to be halotolerant, psychrotolerant, and barotolerant. Bioactive compounds with diverse antibiotic and cytotoxic activities produced by these strains were identified by high-performance liquid chromatography (HPLC) and database comparison. These include antibacterials (paulomycins A and B), antifungals (maltophilins), antifungals displaying also cytotoxic activities (antimycins and 6-epialteramides), and the antitumor compound fredericamycin. A hypothetical dispersion model is here proposed to explain the biogeographical distribution of S. albidoflavus strains in terrestrial, marine, and atmospheric environments.

Myceligenerans cantabricum sp. nov., a barotolerant actinobacterium isolated from a deep cold-water coral.

An actinobacterium strain (M-201(T)) was isolated from a deep-sea scleractinian coral (Fam. Caryophillidae) collected at 1500 m depth in the Avilés Canyon in the Cantabrian Sea, Asturias, Spain. Strain M-201(T) grew at pH 6.0-9.0 (optimum pH 7.0), between 4 and 37 °C (optimum 28 °C) and at salinities of 0.5-10.5% (w/v) NaCl (optimum 0.5-3.0%). The peptidoglycan contained the amino acids Lys, Ala, Thr, Glu and one unknown amino acid component, and belonged to type A4α, and the cell-wall sugars are glucose, mannose and galactose. The polar lipids were diphosphatidylglycerol, phosphatidylglycerol, phosphatidylinositol, an unknown phosphoglycolipid and seven unknown glycolipids. The predominant menaquinones were MK-9(H4) and MK-9(H6). Major cellular fatty acids were anteiso-C(15 : 0), iso-C(15 : 0) and anteiso-C(17 : 0). The genomic DNA G+C content was 72.4 mol%. The chemotaxonomic properties supported the affiliation of strain M-201(T) to the genus Myceligenerans . Phylogenetic analysis based on 16S rRNA gene sequences revealed that the organism was most closely related to Myceligenerans crystallogenes CD12E2-27(T) (98.2% 16S rRNA gene sequence similarity). However, it had a relatively low DNA-DNA relatedness value with the above strain (48%). The isolate showed antibiotic activity against Escherichia coli , Micrococcus luteus ATCC 14452 and Saccharomyces cerevisiae var. carlsbergensis. To the best of our knowledge, this is the first report of antibiotic production in the genus Myceligenerans . The differences in phenotypic, metabolic, ecological and phylogenetic characteristics justify the proposal of a novel species of the genus Myceligenerans , Myceligenerans cantabricum sp. nov., with M-201(T) ( = CECT 8512(T) = DSM 28392(T)) as the type strain.

Two Streptomyces species producing antibiotic, antitumor, and anti-inflammatory compounds are widespread among intertidal macroalgae and deep-sea coral reef invertebrates from the central Cantabrian Sea.

Streptomycetes are widely distributed in the marine environment, although only a few studies on their associations to algae and coral ecosystems have been reported. Using a culture-dependent approach, we have isolated antibiotic-active Streptomyces species associated to diverse intertidal marine macroalgae (Phyllum Heterokontophyta, Rhodophyta, and Chlorophyta), from the central Cantabrian Sea. Two strains, with diverse antibiotic and cytotoxic activities, were found to inhabit these coastal environments, being widespread and persistent over a 3-year observation time frame. Based on 16S rRNA sequence analysis, the strains were identified as Streptomyces cyaneofuscatus M-27 and Streptomyces carnosus M-40. Similar isolates to these two strains were also associated to corals and other invertebrates from deep-sea coral reef ecosystem (Phyllum Cnidaria, Echinodermata, Arthropoda, Sipuncula, and Anelida) living up to 4.700-m depth in the submarine Avilés Canyon, thus revealing their barotolerant feature. These two strains were also found to colonize terrestrial lichens and have been repeatedly isolated from precipitations from tropospheric clouds. Compounds with antibiotic and cytotoxic activities produced by these strains were identified by high-performance liquid chromatography (HPLC) and database comparison. Antitumor compounds with antibacterial activities and members of the anthracycline family (daunomycin, cosmomycin B, galtamycin B), antifungals (maltophilins), anti-inflamatory molecules also with antituberculosis properties (lobophorins) were identified in this work. Many other compounds produced by the studied strains still remain unidentified, suggesting that Streptomyces associated to algae and coral ecosystems might represent an underexplored promising source for pharmaceutical drug discovery.

Activation and silencing of secondary metabolites in Streptomyces albus and Streptomyces lividans after transformation with cosmids containing the thienamycin gene cluster from Streptomyces cattleya.

Activation and silencing of antibiotic production was achieved in Streptomyces albus J1074 and Streptomyces lividans TK21 after introduction of genes within the thienamycin cluster from S. cattleya. Dramatic phenotypic and metabolic changes, involving activation of multiple silent secondary metabolites and silencing of others normally produced, were found in recombinant strains harbouring the thienamycin cluster in comparison to the parental strains. In S. albus, ultra-performance liquid chromatography purification and NMR structural elucidation revealed the identity of four structurally related activated compounds: the antibiotics paulomycins A, B and the paulomenols A and B. Four volatile compounds whose biosynthesis was switched off were identified by gas chromatography-mass spectrometry analyses and databases comparison as pyrazines; including tetramethylpyrazine, a compound with important clinical applications to our knowledge never reported to be produced by Streptomyces. In addition, this work revealed the potential of S. albus to produce many others secondary metabolites normally obtained from plants, including compounds of medical relevance as dihydro-ß-agarofuran and of interest in perfume industry as ß-patchoulene, suggesting that it might be an alternative model for their industrial production. In S. lividans, actinorhodins production was strongly activated in the recombinant strains whereas undecylprodigiosins were significantly reduced. Activation of cryptic metabolites in Streptomyces species might represent an alternative approach for pharmaceutical drug discovery.

Comparative analysis of a cryptic thienamycin-like gene cluster identified in Streptomyces flavogriseus by genome mining.

In silico database searches allowed the identification in the S. flavogriseus ATCC 33331 genome of a carbapenem gene cluster highly related to the S. cattleya thienamycin one. This is the second cluster found for a complex highly substituted carbapenem. Comparative analysis revealed that both gene clusters display a high degree of synteny in gene organization and in protein conservation. Although the cluster appears to be silent under our laboratory conditions, the putative metabolic product was predicted from bioinformatics analyses using sequence comparison tools. These data, together with previous reports concerning epithienamycins production by S. flavogriseus strains, suggest that the cluster metabolic product might be a thienamycin-like carbapenem, possibly the epimeric epithienamycin. This finding might help in understanding the biosynthetic pathway to thienamycin and other highly substituted carbapenems. It also provides another example of genome mining in Streptomyces sequenced genomes as a powerful approach for novel antibiotic discovery.

Natural products, including a new caboxamycin, from Streptomyces and other Actinobacteria isolated in Spain from storm clouds transported by Northern winds of Arctic origin.

Actinobacteria, mostly Streptomyces species, are the main source of natural products essential in medicine. While the majority of producer microorganisms of secondary metabolite are reported from terrestrial or marine environments, there are limited reports of their isolation from atmospheric precipitations. Clouds are considered as atmospheric oases for microorganisms and there is a recent paradigm shift whereby atmospheric-derived Actinobacteria emerge as an alternative source for drug discovery. In this context, we studied a total of 18 bioactive Actinobacteria strains, isolated by sampling nine precipitation events with prevailing Northern winds in the Cantabrian Sea coast, Northern Spain. Backward trajectories meteorological analyses indicate that air masses were originated mostly in the Arctic Ocean, and their trajectory to downwind areas involved the Atlantic Ocean and also terrestrial sources from continental Europe, and in some events from Canada, Greenland, Mauritania and Canary Islands. Taxonomic identification of the isolates, by 16S rRNA gene sequencing and phylogenetic analyses, revealed that they are members of three Actinobacteria genera. Fifteen of the isolates are Streptomyces species, thus increasing the number of bioactive species of this genus in the atmosphere to a 6.8% of the total currently validated species. In addition, two of the strains belong to the genus Micromonospora and one to genus Nocardiopsis. These findings reinforce a previous atmospheric dispersal model, extended herein to the genus Micromonospora. Production of bioactive secondary metabolites was screened in ethyl acetate extracts of the strains by LC-UV-MS and a total of 94 secondary metabolites were detected after LC/MS dereplication. Comparative analyses with natural products databases allowed the identification of 69 structurally diverse natural products with contrasted biological activities, mostly as antibiotics and antitumor agents, but also anti-inflammatory, antiviral, antiparasitic, immunosuppressant and neuroprotective among others. The molecular formulae of the 25 remaining compounds were determined by HRMS. None of these molecules had been previously reported in natural product databases indicating potentially novel metabolites. As a proof of concept, a new metabolite caboxamycin B (1) was isolated from the culture broth of Streptomyces sp. A-177 and its structure was determined by various spectrometric methods. To the best of our knowledge, this is the first novel natural product obtained from an atmospheric Streptomyces, thus pointing out precipitations as an innovative source for discovering new pharmaceutical natural products.

Mutational analysis of the thienamycin biosynthetic gene cluster from Streptomyces cattleya.

The generation of non-thienamycin-producing mutants with mutations in the thnL, thnN, thnO, and thnI genes within the thn gene cluster from Streptomyces cattleya and their involvement in thienamycin biosynthesis and regulation were previously reported. Four additional mutations were independently generated in the thnP, thnG, thnR, and thnT genes by insertional inactivation. Only the first two genes were found to play a role in thienamycin biosynthesis, since these mutations negatively or positively affect antibiotic production. A mutation of thnP results in the absence of thienamycin production, whereas a 2- to 3-fold increase in thienamycin production was observed for the thnG mutant. On the other hand, mutations in thnR and thnT showed that although these genes were previously reported to participate in this pathway, they seem to be nonessential for thienamycin biosynthesis, as thienamycin production was not affected in these mutants. High-performance liquid chromatography (HPLC)-mass spectrometry (MS) analysis of all available mutants revealed some putative intermediates in the thienamycin biosynthetic pathway. A compound with a mass corresponding to carbapenam-3-carboxylic acid was detected in some of the mutants, suggesting that the assembly of the bicyclic nucleus of thienamycin might proceed in a way analogous to that of the simplest natural carbapenem, 1-carbapen-2-em-3-carboxylic acid biosynthesis. The accumulation of a compound with a mass corresponding to 2,3-dihydrothienamycin in the thnG mutant suggests that it might be the last intermediate in the biosynthetic pathway. These data, together with the establishment of cross-feeding relationships by the cosynthesis analysis of the non-thienamycin-producing mutants, lead to a proposal for some enzymatic steps during thienamycin assembly.

Characterization of the gilthead seabream (Sparus aurata L.) transferrin gene: genomic structure, constitutive expression and SNP variation.

Transferrin (Tf) is a multi-function protein with a central role in iron metabolism, and it is this function that is associated with a role in the innate immune system response. The clear link between Tf and immune defense mechanism lead to propose Tf as a candidate gene for disease resistance. In this study, genomic and cDNA sequences of Tf gene in gilthead seabream (Sparus aurata L.) (SaTf gene), were identified and characterized. SaTf gene structure consists of a coding region of 2076 nucleotides divided into 17 exons and a no-coding region that includes 16 introns and spans 5495 nucleotides. The deduced Tf protein for gilthead seabream is composed of 691 amino acids and consists of an initial peptide and two lobes (N- and C-lobes). This gene structure is similar to that of previously described Tf genes in other fish species. RT-PCR analyses carried out in different tissues and two developmental stages showed tissue-and stage-specific Tf expression in gilthead seabream. Finally, by sequencing the transferrin genomic sequences of 20 unrelated seabreams, 31 SNPs were identified. These data allowed the estimation of the frequency of nucleotide substitution in the SaTf gene as 1SNP per 253 bp. SNPs were detected in different regions of the genomic sequence but they were mainly localized in non-coding regions, specifically, SNP frequency in non-coding regions was fifteen-fold higher than within coding regions.

Bioactive Natural Products in Actinobacteria Isolated in Rainwater From Storm Clouds Transported by Western Winds in Spain.

Actinobacteria are the main producers of bioactive natural products essential for human health. Although their diversity in the atmosphere remains largely unexplored, using a multidisciplinary approach, we studied here 27 antibiotic producing Actinobacteria strains, isolated from 13 different precipitation events at three locations in Northern and Southern Spain. Rain samples were collected throughout 2013-2016, from events with prevailing Western winds. NOAA HYSPLIT meteorological analyses were used to estimate the sources and trajectories of the air-mass that caused the rainfall events. Five-day backward air masses trajectories of the diverse events reveals a main oceanic source from the North Atlantic Ocean, and in some events long range transport from the Pacific and the Arctic Oceans; terrestrial sources from continental North America and Western Europe were also estimated. Different strains were isolated depending on the precipitation event and the latitude of the sampling site. Taxonomic identification by 16S rRNA sequencing and phylogenetic analysis revealed these strains to belong to two Actinobacteria genera. Most of the isolates belong to the genus Streptomyces, thus increasing the number of species of this genus isolated from the atmosphere. Furthermore, five strains belonging to the rare Actinobacterial genus Nocardiopsis were isolated in some events. These results reinforce our previous Streptomyces atmospheric dispersion model, which we extend herein to the genus Nocardiopsis. Production of bioactive secondary metabolites was analyzed by LC-UV-MS. Comparative analyses of Streptomyces and Nocardiopsis metabolites with natural product databases led to the identification of multiple, chemically diverse, compounds. Among bioactive natural products identified 55% are antibiotics, both antibacterial and antifungal, and 23% have antitumor or cytotoxic properties; also compounds with antiparasitic, anti-inflammatory, immunosuppressive, antiviral, insecticidal, neuroprotective, anti-arthritic activities were found. Our findings suggest that over time, through samples collected from different precipitation events, and space, in different sampling places, we can have access to a great diversity of Actinobacteria producing an extraordinary reservoir of bioactive natural products, from remote and very distant origins, thus highlighting the atmosphere as a contrasted source for the discovery of novel compounds of relevance in medicine and biotechnology.

Identification of transcriptional activators for thienamycin and cephamycin C biosynthetic genes within the thienamycin gene cluster from Streptomyces cattleya.

Two regulatory genes, thnI and thnU, were identified in the thienamycin (thn) gene cluster from Streptomyces cattleya. ThnI resembles LysR-type transcriptional activators and ThnU belongs to the SARP family of transcriptional activators. Their functional role was established after independent inactivation by gene replacement together with transcriptional analysis involving reverse transcription polymerase chain reaction (RT-PCR). Deletion of thnI abolished thienamycin production showing its involvement in thienamycin biosynthesis. Gene expression analysis applied to the thn gene cluster demonstrated that ThnI is a transcriptional activator essential for thienamycin biosynthesis that regulates the expression of nine genes involved in thienamycin assembly and export (thnH, thnJ, thnK, thnL, thnM, thnN, thnO, thnP and thnQ). Unexpectedly, the thnU disrupted mutant was not affected in thienamycin production but turned out to be essential for cephamycin C biosynthesis. Transcript analysis applied to early and late structural genes for cephamycin C biosynthesis (pcbAB and cmcI), revealed that ThnU is the transcriptional activator of these cephamycin C genes although they are not physically linked to the thn cluster. In addition, it was shown that deletion of thnI has an upregulatory effect on pcbAB and cmcI transcription consistent with a significant increase in cephamycin C biosynthesis in this mutant.