Soil Bacteria Isolated From Tunisian Arid Areas Show Promising Antimicrobial Activities Against Gram-Negatives

Arid regions show relatively fewer species in comparison to better-watered biomes, but the competition for the few nutrients is very distinct. Here, in total 373 bacterial strains were isolated from rhizospheric soils obtained from three different sampling sites in Tunisia. Their potential for the production of antimicrobial compounds was evaluated. Bacterial strains, showing antibacterial activity against pathogenic bacteria, were isolated from all three sites, one strain from the Bou-Hedma national park, 15 strains from Chott-Djerid, and 13 strains from Matmata, respectively. The dominant genus was Bacillus, with 27 out of 29 strains. Most interestingly, 93% of the isolates showed activity against Gram-positive and Gram-negative test bacteria. Strain Bacillus sp. M21, harboring high inhibitory potential, even against clinical isolates of Gram-negative bacteria, was analyzed in detail to enable purification and identification of the bioactive compound responsible for its bioactivity. Subsequent HPLC-MS and NMR analyses resulted in the identification of 1-acetyl-β-carboline as active component. Furthermore, fungicides of the bacillomycin and fengycin group, which in addition show antibiotic effects, were identified. This work highlights the high potential of the arid-adapted strains for the biosynthesis of specialized metabolites and suggest further investigation of extreme environments, since they constitute a promising bioresource of biologically active compounds

Draft Genome Sequences of the Obligatory Marine Myxobacterial Strains Enhygromyxa salina SWB005 and SWB007

The two marine myxobacterial strains Enhygromyxa salina SWB005 and SWB007 were isolated from coastal soil samples using Escherichia coli as bait for these predatory strains. These strains produce unique specialized metabolites. Genomes were assembled into 312 contigs for E. salina SWB005 (9.0 Mbp) and 192 contigs for E. salina SWB007 (10.6 Mbp)

Antimicrobial Potential of Bacteria Associated with Marine Sea Slugs from North Sulawesi, Indonesia

Nudibranchia, marine soft-bodied organisms, developed, due to the absence of a protective shell, different strategies to protect themselves against putative predators and fouling organisms. One strategy is to use chemical weapons to distract predators, as well as pathogenic microorganisms. Hence, these gastropods take advantage of the incorporation of chemical molecules. Thereby the original source of these natural products varies; it might be the food source, de novo synthesis from the sea slug, or biosynthesis by associated bacteria. These bioactive molecules applied by the slugs can become important drug leads for future medicinal drugs. To test the potential of the associated bacteria, the latter were isolated from their hosts, brought into culture and extracts were prepared and tested for antimicrobial activities. From 49 isolated bacterial strains 35 showed antibiotic activity. The most promising extracts were chosen for further testing against relevant pathogens. In that way three strains showing activity against methicillin resistant Staphylococcus aureus and one strain with activity against enterohemorrhagic Escherichia coli, respectively, were identified. The obtained results indicate that the sea slug associated microbiome is a promising source for bacterial strains, which hold the potential for the biotechnological production of antibiotics

Identification, cloning, expression and functional interrogation of the biosynthetic pathway of the polychlorinated triphenyls ambigol A-C fromFischerella ambigua108b

The terrestrial cyanobacteriumFischerella ambigua108b produces the three polychlorinated triphenyls ambigol A-C that exhibit interesting antimicrobial, antiviral and cytotoxic activities. They are structurally related to polybrominated diphenylethers synthesized by diverse marine bacteria that are known to be highly toxic and are bioaccumulating in natural food webs. All ambigols display unusual connectivities: Ambigols A and B exhibit chlorination and ambigol C biaryl-ether bonds in the relativemetaposition at the central phenol unit, which is flanked by two 2,4-dichlorophenol units in all three compounds. Here we report on the identification of the biosynthetic gene cluster (BGC) reponsible for ambigol production inF. ambigua. After bioinformatic discovery of a putative ambigol BGC (ab) containing 10 genes, we cloned and heterologously expressed this cluster inSynechococcus elongatusPCC 7942 using Direct Pathway Cloning (DiPaC).In vivoandin vitrocharacterization of the two cytochrome P450 enzymes present in theabBGC revealed complementary selectivity for either biaryl-ether bond (Ab2) or biaryl formation (Ab3) and provided a biosynthetic route to the ambigols

Mutasynthetic Production and Antimicrobial Characterization of Darobactin Analogs

There is great need for therapeutics against multidrug-resistant, Gram-negative bacterial pathogens. Recently, darobactin A, a novel bicyclic heptapeptide that selectively kills Gram-negative bacteria by targeting the outer membrane protein BamA, was discovered. Its efficacy was proven in animal infection models of Escherichia coli, Klebsiella pneumoniae, and Pseudomonas aeruginosa, thus promoting darobactin A as a promising lead compound. Originally discovered from members of the nematode-symbiotic genus; Photorhabdus; , the biosynthetic gene cluster (BGC) encoding the synthesis of darobactin A can also be found in other members of the class; Gammaproteobacteria; . Therein, the precursor peptides DarB to -F, which differ in their core sequence from darobactin A, were identified; in silico; . Even though production of these analogs was not observed in the putative producer strains, we were able to generate them by mutasynthetic derivatization of a heterologous expression system. The analogs generated were isolated and tested for their bioactivity. The most potent compound, darobactin B, was used for cocrystallization with the target BamA, revealing a binding site identical to that of darobactin A. Despite its potency, darobactin B did not exhibit cytotoxicity, and it was slightly more active against Acinetobacter baumannii isolates than darobactin A. Furthermore, we evaluated the plasma protein binding of darobactin A and B, indicating their different pharmacokinetic properties. This is the first report on new members of this new antibiotic class, which is likely to expand to several promising therapeutic candidates.; IMPORTANCE; Therapeutic options to combat Gram-negative bacterial pathogens are dwindling with increasing antibiotic resistance. This study presents a proof of concept for the heterologous-expression approach to expand on the novel antibiotic class of darobactins and to generate analogs with different activities and pharmacokinetic properties. In combination with the structural data of the target BamA, this approach may contribute to structure-activity relationship (SAR) data to optimize inhibitors of this essential outer membrane protein of Gram-negative pathogens

Marine Myxobacteria as a Source of Antibiotics—Comparison of Physiology, Polyketide-Type Genes and Antibiotic Production of Three New Isolates of Enhygromyxa salina

Three myxobacterial strains, designated SWB004, SWB005 and SWB006, were obtained from beach sand samples from the Pacific Ocean and the North Sea. The strains were cultivated in salt water containing media and subjected to studies to determine their taxonomic status, the presence of genes for the biosynthesis of polyketides and antibiotic production. 16S rDNA sequence analysis revealed the type strain Enhygromyxa salina SHK-1T as their closest homolog, displaying between 98% (SWB005) and 99% (SWB004 and SWB006) sequence similarity. All isolates were rod-shaped cells showing gliding motility and fruiting body formation as is known for myxobacteria. They required NaCl for growth, with an optimum concentration of around 2% [w/v]. The G + C-content of genomic DNA ranged from 63.0 to 67.3 mol%. Further, the strains were analyzed for their potential to produce polyketide-type structures. PCR amplified ketosynthase-like gene fragments from all three isolates enhances the assumption that these bacteria produce polyketides. SWB005 was shown to produce metabolites with prominent antibacterial activity, including activity towards methicillin resistant Staphylococcus aureus (MRSA) and Staphylococcus epidermidis (MRSE)

Orientia tsutsugamushi is highly susceptible to the RNA polymerase switch region inhibitor corallopyronin a In Vitro and In Vivo

Scrub typhus is a potentially lethal infection caused by the obligate intracellular bacterium; Orientia tsutsugamushi; Reports on the emergence of doxycycline-resistant strains highlight the urgent need to develop novel antiinfectives against scrub typhus. Corallopyronin A (CorA) is a novel α-pyrone compound synthesized by the myxobacterium; Corallococcus coralloides; that was characterized as a noncompetitive inhibitor of the switch region of the bacterial RNA polymerase (RNAP). We investigated the antimicrobial action of CorA against the human-pathogenic Karp strain of; O. tsutsugamushi; in vitro; and; in vivo; The MIC of CorA against; O. tsutsugamushi; was remarkably low (0.0078 μg/ml), 16-fold lower than that against; Rickettsia typhi; In the lethal intraperitoneal; O. tsutsugamushi; mouse infection model, a minimum daily dose of 100 μg CorA protected 100% of infected mice. Two days of treatment were sufficient to confer protection. In contrast to BALB/c mice, SCID mice succumbed to the infection despite treatment with CorA or tetracycline, suggesting that antimicrobial treatment required synergistic action of the adaptive immune response. Similar to tetracycline, CorA did not prevent latent infection of; O. tsutsugamushi; in vivo; However, latency was not caused by acquisition of antimicrobial resistance, since; O. tsutsugamushi; reisolated from latently infected BALB/c mice remained fully susceptible to CorA. No mutations were found in the CorA-binding regions of the β and β' RNAP subunit genes; rpoB; and; rpoC; Inhibition of the RNAP switch region of; O. tsutsugamushi; by CorA is therefore a novel and highly potent target for antimicrobial therapy for scrub typhus

Heterologous Expression of Pseudouridimycin and Description of the Corresponding Minimal Biosynthetic Gene Cluster

Pseudouridimycin (PUM) was recently discovered from Streptomyces sp. DSM26212 as a novel bacterial nucleoside analog that competes with UTP for access to the RNA polymerase (RNAP) active site, thereby inhibiting bacterial RNAP by blocking transcription. This represents a novel antibacterial mode of action and it is known that PUM inhibits bacterial RNAP in vitro, inhibits bacterial growth in vitro, and was active in vivo in a mouse infection model of Streptococcus pyogenes peritonitis. The biosynthetic gene cluster (BGC) was previously identified and characterized by knockout experiments. However, the minimal set of genes necessary for PUM production was not proposed. To identify the minimal BGC and to create a plug-and-play production platform for PUM and its biosynthetic precursors, several versions of a redesigned PUM BGC were generated and expressed in the heterologous host Streptomyces coelicolor M1146 under control of strong promotors. Heterologous expression allowed identification of the putative serine/threonine kinase PumF as an enzyme essential for heterologous PUM production and thus corroboration of the PUM minimal BGC

Seven New Alkaloids Isolated from Marine Flavobacterium <i>Tenacibaculum discolor</i> sv11

Marine flavobacterium Tenacibaculum discolor sv11 has been proven to be a promising producer of bioactive nitrogen-containing heterocycles. A chemical investigation of T. discolor sv11 revealed seven new heterocycles, including the six new imidazolium-containing alkaloids discolins C-H (1–6) and one pyridinium-containing alkaloid dispyridine A (7). The molecular structure of each compound was elucidated by analysis of NMR and HR-ESI-MS data. Furthermore, enzymatic decarboxylation of tryptophan and tyrosine to tryptamine and tyramine catalyzed by the decarboxylase DisA was investigated using in vivo and in vitro experiments. The antimicrobial activity of the isolated compounds (1–7) was evaluated. Discolin C and E (1 and 3) exhibited moderate activity against Gram-positive Bacillus subtilis DSM10, Mycobacterium smegmatis ATCC607, Listeria monocytogenes DSM20600 and Staphylococcus aureus ATCC25923, with MIC values ranging from 4 μg/mL to 32 μg/mL

Identification of sequestered chloroplasts in photosynthetic and non-photosynthetic sacoglossan sea slugs (Mollusca, Gastropoda)

BACKGROUND: Sacoglossan sea slugs are well known for their unique ability among metazoans to incorporate functional chloroplasts (kleptoplasty) in digestive glandular cells, enabling the slugs to use these as energy source when starved for weeks and months. However, members assigned to the shelled Oxynoacea and Limapontioidea (often with dorsal processes) are in general not able to keep the incorporated chloroplasts functional. Since obviously no algal genes are present within three (out of six known) species with chloroplast retention of several months, other factors enabling functional kleptoplasty have to be considered. Certainly, the origin of the chloroplasts is important, however, food source of most of the about 300 described species is not known so far. Therefore, a deduction of specific algal food source as a factor to perform functional kleptoplasty was still missing. RESULTS: We investigated the food sources of 26 sacoglossan species, freshly collected from the field, by applying the chloroplast marker genes tufA and rbcL and compared our results with literature data of species known for their retention capability. For the majority of the investigated species, especially for the genus Thuridilla, we were able to identify food sources for the first time. Furthermore, published data based on feeding observations were confirmed and enlarged by the molecular methods. We also found that certain chloroplasts are most likely essential for establishing functional kleptoplasty. CONCLUSIONS: Applying DNA-Barcoding appeared to be very efficient and allowed a detailed insight into sacoglossan food sources. We favor rbcL for future analyses, but tufA might be used additionally in ambiguous cases. We narrowed down the algal species that seem to be essential for long-term-functional photosynthesis: Halimeda, Caulerpa, Penicillus, Avrainvillea, Acetabularia and Vaucheria. None of these were found in Thuridilla, the only plakobranchoidean genus without long-term retention forms. The chloroplast type, however, does not solely determine functional kleptoplasty; members of no-retention genera, such as Cylindrobulla or Volvatella, feed on the same algae as e.g., the long-term-retention forms Plakobranchus ocellatus or Elysia crispata, respectively. Evolutionary benefits of functional kleptoplasty are still questionable, since a polyphagous life style would render slugs more independent of specific food sources and their abundance