Background:No investigation on daptomycin production at the transcriptional regulatory level has been reported. Results: The autoregulator AtrA directly regulates daptomycin gene cluster expression, and atrA is the transcriptional target of AdpA. Conclusion:The AtrA-mediated transcriptional signaling pathway directly regulates daptomycin production. Significance: We reveal for the first time the transcriptional regulatory mechanism of daptomycin production for its potential rational genetic engineering.

Duchenne and Becker muscular dystrophies (DMD/BMD) are the most commonly inherited neuromuscular disease. However, accurate and convenient molecular diagnosis cannot be achieved easily because of the enormous size of the dystrophin gene and complex causative mutation spectrum. Such traditional methods as multiplex ligation-dependent probe amplification plus Sanger sequencing require multiple steps to fulfill the diagnosis of DMD/BMD. Here, we introduce a new single-step method for the genetic analysis of DMD patients and female carriers in real clinical settings and demonstrate the validation of its accuracy. A total of 89 patients, 18 female carriers and 245 non-DMD patients were evaluated using our targeted NGS approaches. Compared with traditional methods, our new method yielded 99.99% specificity and 98.96% sensitivity for copy number variations detection and 100% accuracy for the identification of single-nucleotide variation mutations. Additionally, this method is able to detect partial deletions/duplications, thus offering precise personal DMD gene information for gene therapy. We detected novel partial deletions of exons in nine samples for which the breakpoints were located within exonic regions. The results proved that our new method is suitable for routine clinical practice, with shorter turnaround time, higher accuracy, and better insight into comprehensive genetic information (detailed breakpoints) for ensuing gene therapy.

Klebsiella pneumoniae MGH78578, a clinical isolate, showed high level of resistance to many antimicrobial agents. We cloned genes responsible for drug resistance from chromosomal DNA of K. pneumoniae MGH78578 by shotgun method using Escherichia coli KAM32, a drug hypersensitive strain, as host. We obtained 43 hybrid plasmids that made host cells resistant to several antimicrobial agents. We classified them into 17 groups based on growth properties in the presence of each one of 9 antimicrobial agents and on restriction patterns of each hybrid plasmid. Analysis of the cloned genes must be very useful for investigation of major parts of multidrug resistance systems including multidrug efflux pumps in K. pneumoniae MGH78578 in which genome sequence is available.

In this work, the bioinspired reversible switch between underwater superoleophobicity/superaerophobicity and oleophilicity/ aerophilicity and improved antireflective property were successfully demonstrated on the nanosecond laser-structured titanium surfaces. Titanium materials were first transformed to be superhydrophobic after nanosecond laser ablation and low-temperature annealing treatments, showing oleophilicity/aerophilicity in water. If the surfaces were prewetted with absolute ethanol and then immersed into water, the surfaces showed superoleophobicity/superaerophobicity. More importantly, the underwater oleophilicity/aerophilicity of the surfaces could be easily recovered by natural drying, and the switch between the underwater superoleophobicity/superaerophobicity and oleophilicity/aerophilicity could be repeated many cycles. Moreover, based on the original antireflective performance of the surface of the laser-ablated micro/ nanoscale structures, we demonstrated that the inspired improved antireflective property could be skillfully realized by the prewetting treatment. The developed bioinspired multifunctional materials provide a versatile platform for the potential applications, such as controlling oil droplets, bubbles, and optical behavior.

BackgroundStreptomyces chattanoogensis L10 is the industrial producer of natamycin and has been proved a highly efficient host for diverse natural products. It has an enormous potential to be developed as a versatile cell factory for production of heterologous secondary metabolites. Here we developed a genome-reduced industrial Streptomyces chassis by rational ‘design-build-test’ pipeline.ResultsTo identify candidate large non-essential genomic regions accurately and design large deletion rationally, we performed genome analyses of S. chattanoogensis L10 by multiple computational approaches, optimized Cre/loxP recombination system for high-efficient large deletion and constructed a series of universal suicide plasmids for rapid loxP or loxP mutant sites inserting into genome. Subsequently, two genome-streamlined mutants, designated S. chattanoogensis L320 and L321, were rationally constructed by depletion of 1.3 Mb and 0.7 Mb non-essential genomic regions, respectively. Furthermore, several biological performances like growth cycle, secondary metabolite profile, hyphae morphological engineering, intracellular energy (ATP) and reducing power (NADPH/NADP+) levels, transformation efficiency, genetic stability, productivity of heterologous proteins and secondary metabolite were systematically evaluated. Finally, our results revealed that L321 could serve as an efficient chassis for the production of polyketides.ConclusionsHere we developed the combined strategy of multiple computational approaches and site-specific recombination system to rationally construct genome-reduced Streptomyces hosts with high efficiency. Moreover, a genome-reduced industrial Streptomyces chassis S. chattanoogensis L321 was rationally constructed by the strategy, and the chassis exhibited several emergent and excellent performances for heterologous expression of secondary metabolite. The strategy could be widely applied in other Streptomyces to generate miscellaneous and versatile chassis with minimized genome. These chassis can not only serve as cell factories for high-efficient production of valuable polyketides, but also will provide great support for the upgrade of microbial pharmaceutical industry and drug discovery.Electronic supplementary materialThe online version of this article (10.1186/s12934-019-1055-7) contains supplementary material, which is available to authorized users.

A simple and low-crosstalk 1 × 4 silicon mode (de)multiplexer based on multimode grating-assisted-couplers is proposed. Mode transitions can be flexibly controlled by designing the grating period at the phase-matching condition. Due to the contra-directional coupling, precise control of the coupling strength and the coupling length are not needed in the system. Calculation results show that the insertion loss and the 3 dB bandwidths of the device are 0.2 dB and 3.7 nm, 0.34 dB and 7.6 nm, and 0.21 dB and 11.8 nm for the channels which (de)multiplex to the 1st, 2nd, and 3rd modes of the bus waveguide, respectively.

Gamma-butyrolactones (GBLs) produced by several Streptomyces species have been shown to serve as quorum-sensing signaling molecules for activating antibiotic production. The GBL system of Streptomyces chattanoogensis L10, a producer of antifungal agent natamycin, consists of three genes: scgA, scgX, and scgR. Both scgA and scgX contribute to GBL production, while scgR encodes a GBL receptor. ⌬scgA and ⌬scgX mutants of S. chattanoogensis behaved identically: they had a growth defect in submerged cultures and delayed or abolished the morphological differentiation and secondary metabolites production on solid medium. ScgR could bind to the promoter region of scgA and repress its transcription. Moreover, scgA seems also to be controlled by a GBL-mediated negative-feedback system. Hence, it is apparent that GBL biosynthesis is tightly controlled to ensure the correct timing for metabolic switch. An additional direct ScgR-target gene gbdA was identified by genomic SELEX and transcriptional analysis. Comparative proteomic analysis between L10 and its ⌬scgA mutant revealed that the GBL system affects the expression of more than 50 proteins, including enzymes involved in carbon uptake system, primary metabolism, and stress response, we thus conclude that scgR-scgA-scgX constitute a novel GBL regulatory system involved in nutrient utilization, triggering adaptive responses, and finally dictating the switch from primary to secondary metabolism. Quorum sensing (QS) is a cell-cell communication processin which bacteria use the production and detection of extracellular chemicals called autoinducers to monitor cell population density and synchronize community behavior through regulation of their gene expression in response to changes in cell density. Acyl homoserine lactones (AHLs) are a major class of autoinducers used by Gram-negative proteobacteria for intraspecies quorum sensing (23) and has been studied intensively over the past decade. Streptomyces, probably as well as its related genera, use ␥-butyrolactones (2,3-di-substituted-␥-butyrolactones) as autoinducers, the chemical structure of which is similar to that of AHLs except for the carbon side chain (31). In addition, other signal molecules, such as PI factor [2,3-diamino-2,3-bis(hydroxymethyl)-1,4-butanediol] and AHFCAs (2-alkyl-4-hydroxymethylfuran-3-carboxylic acids), have been described to play a similar role to that of GBLs in Streptomyces (6, 26).Streptomycetes are Gram-positive soil bacteria that undergo a developmental program that leads to sporulating aerial hyphae. They are also characterized by a complex secondary metabolism, which makes them the largest antibiotic-producing genus, producing over two-thirds of the clinically used antibiotics of natural origin. The ␥-butyrolactone (GBL) system of Streptomyces, typically consisting of a GBL synthase and a cognate receptor, is drawing more attention because of its close association with production of several antibiotics (31). Currently, there are four such systems with a known GBL and receptor, including the ...

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