Metabolic engineering of "last-line antibiotic" colistin in Paenibacillus polymyxa.

Colistin, also known as polymyxin E, is a lipopeptide antibiotic used to treat infections caused by multidrug-resistant gram-negative bacteria. It is considered a "last-line antibiotic", but its clinical development is hindered by low titer and impurities resulting from the presence of diverse homologs in microbial fermentation. To ensure consistent pharmaceutical activity and kinetics, it is crucial to have high-purity colistin active pharmaceutical ingredient (API) in the pharmaceutical industry. This study focused on the metabolic engineering of a natural colistin producer strain to produce colistin with a high titer and purity. Guided by genome mining, we identified Paenibacillus polymyxa ATCC 842 as a natural colistin producer capable of generating a high proportion of colistin A. By systematically inactivating seven non-essential biosynthetic gene clusters (BGCs) of peptide metabolites that might compete precursors with colistin or inhibit colistin production, we created an engineered strain, P14, which exhibited an 82% increase in colistin titer and effectively eliminated metabolite impurities such as tridecaptin, paenibacillin, and paenilan. Additionally, we engineered the L-2,4-diaminobutyric acid (L-2,4-DABA) pathway to further enhance colistin production, resulting in the engineered strain P19, which boosted a remarkable colistin titer of 649.3 mg/L - a 269% improvement compared to the original strain. By concurrently feeding L-isoleucine and L-leucine, we successfully produced high-purity colistin A, constituting 88% of the total colistin products. This study highlights the potential of metabolic engineering in improving the titer and purity of lipopeptide antibiotics in the non-model strain, making them more suitable for clinical use. These findings indicate that efficiently producing colistin API in high purity directly from fermentation can now be achieved in a straightforward manner.

Highly stabilized fiber Bragg grating accelerometer based on cross-type diaphragm.

A fiber Bragg grating (FBG) accelerometer based on cross-type diaphragm was proposed and designed, in which the cross-beam acts as a spring element. To balance the sensitivity and stability, the accelerometer structure was optimized. The experimental results show that the designed device has a resonant frequency of 556 Hz with a considerable wide frequency bandwidth of up to 200 Hz, which is consistent with the simulation. The sensitivity of the device is 12.35 pm/g@100 Hz with a linear correlation coefficient of 0.99936. The proposed FBG accelerometer has simple structure and strong anti-interference capability with a maximal cross-error less than 3.26%, which can be used for mechanical structural health monitoring.

Ultrahigh bandwidth measuring approach of an I-FOG based on axial magnetic sensitivity.

The bandwidth is one of the key indicators of the interferometric fiber optic gyroscope (I-FOG) in the application with high frequency jitter. The traditional bandwidth measurement equipment, such as the angular vibration table, can only provide angular vibrations of hundreds of hertz and cannot meet the measurement needs of a high bandwidth gyro. We propose an approach, with which a signal of several thousand hertz can be provided and can measure a high bandwidth of I-FOGs. The bandwidth measurement approach is based on the axial magnetic sensitivity. We present the measurement principle, derive the axial magnetic sensitivity expression of the fiber coil in I-FOGs, and demonstrate the bandwidth measuring system. With this system, the bandwidth of an I-FOG is measured and the experimental result shows that the bandwidth is ∼10 kHz. It is proved that this new, to the best of our knowledge, approach is capable of testing the bandwidth of the I-FOG at ultrahigh frequencies.

Effect of optical fiber end face scratches on laser-induced damage threshold.

The scratches on the fiber end face can enhance the local electrical field, which lowers the damage threshold. The damage mechanism of a high-energy laser is investigated. The effect of scratches on the electric field is simulated by the finite difference time domain (FDTD) solution. The results show that the depth of the scratch has a greater ability to influence the electric field than the width, and multiple scratches have a stronger modulation than a single scratch. In calculation, the damage threshold of the scratch-free end face is 0.456J/c m 2 when the incident light electric field intensity is 50M V/c m, compared to 0.345J/c m 2 in the presence of the scratch on the end face.

Bias control approach based on VMD and LIA demodulation of a lithium niobate polarization controller.

A bias control approach is an automatic lock working point algorithm based on variational mode decomposition (VMD) and lock-in amplification (LIA) demodulation for a lithium niobate polarization controller (LNPC). Commonly, the dither voltage applied to the LNPC is much smaller than the bias voltage to avoid the influence of the dither signal on the output light, which reduces the polarization control accuracy of the LNPC. In this paper, we use VMD and LIA, with which the polarization control accuracy of LNPC can be improved, to extract and amplify the dither signal to compensate the drift half-wave of LNPC. The light intensity fluctuations of the output polarized light in vertical or horizontal directions are less than 0.017%.

Enhancing the nonlinearity of optomechanical system via multiple mechanical modes.

We theoretically investigate the nonlinear dynamics of an optomechanical system, where the system consists of N identical mechanical oscillators individually coupled to a common cavity field. We find that the optomechanical nonlinearity can be enhanced N times through theoretical analysis and numerical simulation in such a system. This leads to the power thresholds to observe the nonlinear behaviors (bistable, period-doubling, and chaotic dynamics) being reduced to 1/N. In addition, we find that changing the sign (positive or negative) of the coupling strength partly does not affect the threshold of driving power for generating corresponding nonlinear phenomena. Our work may provide a way to engineer optomechanical devices with a lower threshold, which has potential applications in implementing secret information processing and optical sensing.

Cysteine protected cells from H2O2-induced damage and promoted long-chain fatty acids synthesis in vivo to improve γ-aminobutyric acid production in Levilactobacillus brevis.

Levilactobacillus brevis NPS-QW-145 isolated from kimchi is deficient in glutamate dehydrogenase-encoding gene (gdhA) to form glutamate, hence it required exogenous supplementation of glutamate/monosodium glutamate (MSG) for decarboxylation reaction to produce γ-aminobutyric acid (GABA). However, GABA conversion rate from MSG was relatively low. The individual effect of 20 amino acids on regulating GABA biosynthesis was investigated. Cysteine was selected to significantly improve GABA production from MSG. It was found that Lb. brevis was capable of producing H2O2, cysteine protected Lb. brevis against H2O2-induced oxidative damage to increase cell viability for the enhancement of GABA production. Moreover, cysteine promoted glucose consumption to produce acetyl-CoA for synthesizing long-chain fatty acids to significantly up-regulate GABA biosynthesis. These findings deciphered antioxidative capability of cysteine in Lb. brevis 145 and provided a theoretical basis for fatty acids synthesis-mediated GABA synthesis in Lb. brevis 145, and possibly in other lactic acid bacteria.

Highly sensitive Mach-Zehnder interferometric micromagnetic field sensor based on 3D printing technology.

A two-photon 3D printed polymer magnetic sensing device based on a Mach-Zehnder interferometer (MZI) is proposed. One arm of the MZI contains a hollow cavity and two connecting open channels that can be filled with magnetic fluids (MFs) and sealed by the UV curable adhesive, forming a magneto-optical component of the interferometer. As the magnetic field changes, the refractive index (RI) of the MF changes, and the effective RI of the guiding mode of the waveguide changes accordingly, which results in a change in the phase of the MZI. The interferometric spectra can be used to evaluate the sensing sensitivity. The MZI structure with a hollow length of 40 µm is fabricated, and the microstructure is encapsulated with MF, demonstrating a highly sensitive magnetic field device. The experimental results show that the magnetic field sensitivity of the fabricated magnetic field device is -1.675nm/Oe. For a spectrometer with a resolution of 1 pm, the minimal detectable magnetic field resolution of the sensor is up to 59.7 nT with good stability.

A Novel Approach to Double the Sensitivity of Polarization Maintaining Interferometric Fiber Optic Gyroscope.

In this paper, a novel optical approach to double the sensitivity to angular rate of interferometric fiber optic gyroscope (IFOG) is proposed. Two fiber polarization combiner/splitters (FPCSs), as the key components, are added in the traditional IFOG light path. The FPCSs are able to either combine two orthogonal polarizations transmitting at two different polarization-maintaining fibers (PMFs) into the two orthogonal axes of one PMF, respectively, or split two polarizations transmitting at the two orthogonal axes of one PMF into two polarizations to transmit at two different PMFs, respectively. Through the specific placement and coupling of these two FPCSs, the incident light can transmit twice along the polarization-maintaining fiber coil (PMFC). The novel approach is verified experimentally and the experimental results show consistency with the theoretical analysis. The proposed approach is able to double the sensitivity of IFOGs and can increase the signal-to-noise ratio (SNR) without increasing the length of PMFC, which is very susceptible to environmental influences and is of great significance in the technical improvement of IFOGs, as well as the miniaturization of IFOGs.

A CRISPR/Cas9 Toolbox for Multiplexed Plant Genome Editing and Transcriptional Regulation.

The relative ease, speed, and biological scope of clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated Protein9 (Cas9)-based reagents for genomic manipulations are revolutionizing virtually all areas of molecular biosciences, including functional genomics, genetics, applied biomedical research, and agricultural biotechnology. In plant systems, however, a number of hurdles currently exist that limit this technology from reaching its full potential. For example, significant plant molecular biology expertise and effort is still required to generate functional expression constructs that allow simultaneous editing, and especially transcriptional regulation, of multiple different genomic loci or multiplexing, which is a significant advantage of CRISPR/Cas9 versus other genome-editing systems. To streamline and facilitate rapid and wide-scale use of CRISPR/Cas9-based technologies for plant research, we developed and implemented a comprehensive molecular toolbox for multifaceted CRISPR/Cas9 applications in plants. This toolbox provides researchers with a protocol and reagents to quickly and efficiently assemble functional CRISPR/Cas9 transfer DNA constructs for monocots and dicots using Golden Gate and Gateway cloning methods. It comes with a full suite of capabilities, including multiplexed gene editing and transcriptional activation or repression of plant endogenous genes. We report the functionality and effectiveness of this toolbox in model plants such as tobacco (Nicotiana benthamiana), Arabidopsis (Arabidopsis thaliana), and rice (Oryza sativa), demonstrating its utility for basic and applied plant research.

Effective screen of CRISPR/Cas9-induced mutants in rice by single-strand conformation polymorphism.

KEY MESSAGE: A method based on DNA single-strand conformation polymorphism is demonstrated for effective genotyping of CRISPR/Cas9-induced mutants in rice. Clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated 9 (Cas9) has been widely adopted for genome editing in many organisms. A large proportion of mutations generated by CRISPR/Cas9 are very small insertions and deletions (indels), presumably because Cas9 generates blunt-ended double-strand breaks which are subsequently repaired without extensive end-processing. CRISPR/Cas9 is highly effective for targeted mutagenesis in the important crop, rice. For example, homozygous mutant seedlings are commonly recovered from CRISPR/Cas9-treated calli. However, many current mutation detection methods are not very suitable for screening homozygous mutants that typically carry small indels. In this study, we tested a mutation detection method based on single-strand conformational polymorphism (SSCP). We found it can effectively detect small indels in pilot experiments. By applying the SSCP method for CRISRP-Cas9-mediated targeted mutagenesis in rice, we successfully identified multiple mutants of OsROC5 and OsDEP1. In conclusion, the SSCP analysis will be a useful genotyping method for rapid identification of CRISPR/Cas9-induced mutants, including the most desirable homozygous mutants. The method also has high potential for similar applications in other plant species.

Nonreciprocal phase shift caused by magnetic-thermal coupling of a polarization maintaining fiber optic gyroscope.

A theory for nonreciprocal phase shift caused by cross coupling generated in a polarization maintaining (PM) fiber optic gyroscope (FOG) under the combined action of magnetic and temperature fields is proposed. The magnetic-thermal coupling in the FOG originates from the interaction of the magnetic field, fiber twist, birefringence caused by thermal stress, and the intrinsic and bending birefringence of the fiber. The cross coupling changes with temperature. When the PM fiber has a diameter of 250 µm, beat length of 3 mm, length of 500 m, twist rate of 1 rad/m, and optical source wavelength of 1310 nm, the maximum degree of magnetic-thermal coupling generated by a 1 mT radial magnetic field within the temperature range of -20°C to 60°C is -5.47%.

Exploring the roles of ribosomal peptides in prokaryote-phage interactions through deep learning-enabled metagenome mining.

BACKGROUND: Microbial secondary metabolites play a crucial role in the intricate interactions within the natural environment. Among these metabolites, ribosomally synthesized and post-translationally modified peptides (RiPPs) are becoming a promising source of therapeutic agents due to their structural diversity and functional versatility. However, their biosynthetic capacity and ecological functions remain largely underexplored. RESULTS: Here, we aim to explore the biosynthetic profile of RiPPs and their potential roles in the interactions between microbes and viruses in the ocean, which encompasses a vast diversity of unique biomes that are rich in interactions and remains chemically underexplored. We first developed TrRiPP to identify RiPPs from ocean metagenomes, a deep learning method that detects RiPP precursors in a hallmark gene-independent manner to overcome the limitations of classic methods in processing highly fragmented metagenomic data. Applying this method to metagenomes from the global ocean microbiome, we uncover a diverse array of previously uncharacterized putative RiPP families with great novelty and diversity. Through correlation analysis based on metatranscriptomic data, we observed a high prevalence of antiphage defense-related and phage-related protein families that were co-expressed with RiPP families. Based on this putative association between RiPPs and phage infection, we constructed an Ocean Virus Database (OVD) and established a RiPP-involving host-phage interaction network through host prediction and co-expression analysis, revealing complex connectivities linking RiPP-encoding prokaryotes, RiPP families, viral protein families, and phages. These findings highlight the potential of RiPP families involved in prokaryote-phage interactions and coevolution, providing insights into their ecological functions in the ocean microbiome. CONCLUSIONS: This study provides a systematic investigation of the biosynthetic potential of RiPPs from the ocean microbiome at a global scale, shedding light on the essential insights into the ecological functions of RiPPs in prokaryote-phage interactions through the integration of deep learning approaches, metatranscriptomic data, and host-phage connectivity. This study serves as a valuable example of exploring the ecological functions of bacterial secondary metabolites, particularly their associations with unexplored microbial interactions. Video Abstract.

Identification of intratumor bacteria-associated prognostic risk score in adrenocortical carcinoma.

A landmark study by Poore et al. showed intratumor bacteria (ITBs) playing a critical role in most cancers by reproduction of The Cancer Genome Atlas (TCGA) transcriptome data. A recent study by Salzberg et al. argued that ITBs, being overstated as a methodology by Poore et al., were problematic. We previously reported that ITBs were prognostic in adrenocortical carcinoma (ACC), a highly aggressive rare disease using data by Poore et al., and here, we aimed to answer whether ITBs truly existed and were prognostic in ACC. ACC samples from our institutes underwent 16S rRNA sequencing [adrenocortical carcinoma blocks from Huashan Hospital and China Medical University (HS) cohort]. The ITB profile was compared to TCGA data processed by Poore et al. (TCGA-P) and TCGA data processed by Salzberg et al. (TCGA-S), respectively. The primary outcome was overall survival (OS). A total of 26 ACC cases (HS cohort) and 10 paraffin controls were sequenced. The TCGA cohort encompassed 77 cases. Two and four amid the top 10 abundant genera in HS cohort were not detected in TCGA-P and TCGA-S, respectively. Neither was alpha or beta diversity associated with survival nor could ACC be subtyped by ITB signature in the HS cohort. Notably, a five-genera ITB risk score (Corynebacterium, Mycoplasma, Achromobacter, Anaerococcus, and Streptococcus) for OS trained in the HS cohort was validated in both TCGA-P and TCGA-S cohorts and was independently prognostic. Whereas ITB signature on the whole may not be associated with ACC subtypes, certain ITB features are associated with prognosis, and a risk score could be generated and validated externally. IMPORTANCE: In this report, we looked at the role of ITBs in ACC in patients with different race and sequencing platforms. We found a five-genera ITB risk score consistently predicted overall survival in all cohorts. We conclude that certain ITB features are universally pathogenic to ACC.

Recent developments on the removal of zinc from electric arc furnace dust by using microwave heating.

Electric arc furnace dust (EAFD) is a hazardous by-product of steel production. As global steel output increases, substantial amounts of EAFD are produced, which causes significant environmental issues. EAFD contains quantities of Fe and Zn, which could be reused as raw materials in the steelmaking process. However, zinc oxides can be reduced and vaporized during this process, forming zinc vapor that contaminates equipment surfaces and causes damage. Consequently, various pyrometallurgical methods have been proposed for zinc removal from EAFD. Due to the extensive usage of carbonaceous materials, these methods contribute to significant CO2, raising concerns about greenhouse gas emissions. Microwave heating offers an efficient, energy-saving, and environmentally friendly alternative to pyrometallurgical approaches. EAFD can generate heat under microwave irradiation without carbon addition, which means the CO2 emissions can be reduced by replacing the reductant in the microwave heating process. Furthermore, microwaves enhance zinc removal reactions to a certain extent, resulting in higher efficiency. Thus, employing microwave heating for EAFD processing has significant potential for future development. This paper reviews recent research on using microwave heating for zinc removal from EAFD, focusing on the heating behavior of EAFD in microwaves and the mechanisms of zinc removal. This review will be crucial for researchers working on processing EAFD using microwave heating and could help guide the development of more sustainable and efficient methods.

Life cycle assessment of carbonaceous pellets used in blast furnaces in the context of "double carbon".

As the sole carbonaceous renewable energy source, biomass is distinguished by its abundant yield, widespread distribution, and carbon neutrality. It is integral to the achievement of zero and negative carbon production via conventional carbonaceous pellet technology. This study introduces a cradle-to-gate life cycle assessment methodology for biomass preparation in carbonaceous pellets. We prepare high-quality biochar through a process combining hydrothermal carbonization and pyrolytic carbonization. Biomass high molecular weight extracts are obtained via organic pyrolytic extraction, while biomass high-temperature binders result from the modification and treatment of biochar. Biomass carbonaceous pellets are then formed using hot press technology. The ReCiPe model facilitates a comprehensive life cycle assessment of biomass carbonaceous pellets used in blast furnace production. The study leverages two comprehensive evaluation indicators - renewability, and environmental performance - to enhance the environmental performance of the process system and to maximize energy-saving and emission reduction potential.

Intratumoral microbiota is associated with prognosis in patients with adrenocortical carcinoma.

Adrenocortical carcinoma (ACC) is a rare but aggressive malignancy. Recent studies have discovered a pivotal role of the intratumoral microbiota in various cancers, yet it remains elusive in ACC. Here, we explored the intratumoral microbiome data derived from in silico identification, further validated in an in-house cohort by bacterial 16S rRNA fluorescence in situ hybridization and lipopolysaccharide staining. Unsupervised clustering determined two naturally distinct clusters of the intratumoral microbiome in ACC, which was associated with overall survival. The incorporation of microbial signatures enhanced the prognostic performance of the clinical stage in an immunity-dependent manner. Genetic and transcriptomic association analyses identified significant upregulation of the cell cycle and p53 signaling pathways associated with microbial signatures for worsened prognosis. Our study not only supports the presence of intratumoral bacteria but also implies a prognostic and biological role of intratumoral microbiota in ACC, which can advance a better understanding of the biology of ACC.

CRISPR-detector: fast and accurate detection, visualization, and annotation of genome-wide mutations induced by genome editing events.

The leading-edge CRISPR/CRISPR-associated technology is revolutionizing biotechnologies through genome editing. To track on/off-target events with emerging new editing techniques, improved bioinformatic tools are indispensable. Existing tools suffer from limitations in speed and scalability, especially with whole-genome sequencing (WGS) data analysis. To address these limitations, we have developed a comprehensive tool called CRISPR-detector, a web-based and locally deployable pipeline for genome editing sequence analysis. The core analysis module of CRISPR-detector is based on the Sentieon TNscope pipeline, with additional novel annotation and visualization modules designed to fit CRISPR applications. Co-analysis of the treated and control samples is performed to remove existing background variants prior to genome editing. CRISPR-detector offers optimized scalability, enabling WGS data analysis beyond Browser Extensible Data file-defined regions, with improved accuracy due to haplotype-based variant calling to handle sequencing errors. In addition, the tool also provides integrated structural variation calling and includes functional and clinical annotations of editing-induced mutations appreciated by users. These advantages facilitate rapid and efficient detection of mutations induced by genome editing events, especially for datasets generated from WGS. The web-based version of CRISPR-detector is available at https://db.cngb.org/crispr-detector, and the locally deployable version is available at https://github.com/hlcas/CRISPR-detector.

A systematically biosynthetic investigation of lactic acid bacteria reveals diverse antagonistic bacteriocins that potentially shape the human microbiome.

BACKGROUND: Lactic acid bacteria (LAB) produce various bioactive secondary metabolites (SMs), which endow LAB with a protective role for the host. However, the biosynthetic potentials of LAB-derived SMs remain elusive, particularly in their diversity, abundance, and distribution in the human microbiome. Thus, it is still unknown to what extent LAB-derived SMs are involved in microbiome homeostasis. RESULTS: Here, we systematically investigate the biosynthetic potential of LAB from 31,977 LAB genomes, identifying 130,051 secondary metabolite biosynthetic gene clusters (BGCs) of 2,849 gene cluster families (GCFs). Most of these GCFs are species-specific or even strain-specific and uncharacterized yet. Analyzing 748 human-associated metagenomes, we gain an insight into the profile of LAB BGCs, which are highly diverse and niche-specific in the human microbiome. We discover that most LAB BGCs may encode bacteriocins with pervasive antagonistic activities predicted by machine learning models, potentially playing protective roles in the human microbiome. Class II bacteriocins, one of the most abundant and diverse LAB SMs, are particularly enriched and predominant in the vaginal microbiome. We utilized metagenomic and metatranscriptomic analyses to guide our discovery of functional class II bacteriocins. Our findings suggest that these antibacterial bacteriocins have the potential to regulate microbial communities in the vagina, thereby contributing to the maintenance of microbiome homeostasis. CONCLUSIONS: Our study systematically investigates LAB biosynthetic potential and their profiles in the human microbiome, linking them to the antagonistic contributions to microbiome homeostasis via omics analysis. These discoveries of the diverse and prevalent antagonistic SMs are expected to stimulate the mechanism study of LAB's protective roles for the microbiome and host, highlighting the potential of LAB and their bacteriocins as therapeutic alternatives. Video Abstract.

Genomic and metabolic analyses reveal antagonistic lanthipeptides in archaea.

BACKGROUND: Microbes produce diverse secondary metabolites (SMs) such as signaling molecules and antimicrobials that mediate microbe-microbe interaction. Archaea, the third domain of life, are a large and diverse group of microbes that not only exist in extreme environments but are abundantly distributed throughout nature. However, our understanding of archaeal SMs lags far behind our knowledge of those in bacteria and eukarya. RESULTS: Guided by genomic and metabolic analysis of archaeal SMs, we discovered two new lanthipeptides with distinct ring topologies from a halophilic archaeon of class Haloarchaea. Of these two lanthipeptides, archalan α exhibited anti-archaeal activities against halophilic archaea, potentially mediating the archaeal antagonistic interactions in the halophilic niche. To our best knowledge, archalan α represents the first lantibiotic and the first anti-archaeal SM from the archaea domain. CONCLUSIONS: Our study investigates the biosynthetic potential of lanthipeptides in archaea, linking lanthipeptides to antagonistic interaction via genomic and metabolic analyses and bioassay. The discovery of these archaeal lanthipeptides is expected to stimulate the experimental study of poorly characterized archaeal chemical biology and highlight the potential of archaea as a new source of bioactive SMs. Video Abstract.