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Klebsiella oxytoca M5al is an excellent 1,3-propanediol (1,3-PD) producer, but too much lactic acid yielded greatly lessened the fermentation efficiency for 1,3-PD. To counteract the disadvantage, four lactate deficient mutants were obtained by knocking out the ldhA gene of lactate dehydrogenase (LDH) of K. oxytoca M5al. The LDH activities of the four mutants were from 3.85 to 6.92% of the parental strain. The fed-batch fermentation of 1,3-PD by mutant LDH3, whose LDH activity is the lowest, was studied. The results showed that higher 1,3-PD concentration, productivity, and molar conversion rate from glycerol to 1,3-PD can be gained than those of the wild type strain and no lactic acid is produced under both anaerobic and microaerobic conditions. Sucrose fed during the fermentation increased the conversion and sucrose added at the beginning increased the productivity. In fed-batch fermentation with sucrose as cosubstrate under microaerobic conditions, the 1,3-PD concentration, conversion, and productivity were improved significantly to 83.56 g l-1, 0.62 mol mol-1, and 1.61 g l(-1) h-1, respectively. Furthermore, 60.11 g l(-1) 2,3-butanediol was also formed as major byproduct in the broth.

Nanoparticles (NPs) decorated with functional ligands are promising candidates for cancer diagnosis and treatment. However, numerous studies have shown that chemically coupled targeting moieties on NPs lose their targeting capability in the biological milieu because they are shielded or covered by a \"protein corona\". Herein, we construct a functional magnetosome that recognizes and targets cancer cells even in the presence of protein corona. Magnetosomes (BMPs) were extracted from magnetotactic bacteria, (MSR-1), and decorated with trastuzumab (TZ) via affibody (RA) and glutaraldehyde (GA). The engineered BMPs are referred to as BMP-RA-TZ and BMP-GA-TZ. Their capacities to combine HER2 were detected by ELISA, the quantity of plasma corona proteins was analyzed using LC-MS. The efficiencies of targeting SK-BR-3 were demonstrated by confocal laser scanning microscopy and flow cytometry. Both engineered BMPs contain up to ~0.2 mg TZ per mg of BMP, while the quantity of HER2 binding to BMP-RA-TZ is three times higher than that binding to BMP-GA-TZ. After incubation with normal human plasma or IgG-supplemented plasma, GA-TZ-containing BMPs have larger hydrated radii and more surface proteins in comparison with RA-TZ-containing BMPs. The TZ-containing BMPs all can be targeted to and internalized in the HER2-overexpressing breast cancer cell line SK-BR-3; however, their targeting efficiencies vary considerably: 50-75% for RA-TZ-containing BMPs and 9-19% for GA-TZ-containing BMPs. BMPs were incubated with plasma (100%) and cancer cells to simulate human in vivo environment. In this milieu, BMP-RA-TZ uptake efficiency of SK-BR-3 reaches nearly 80% (slightly lower than for direct interaction with BMP-RA-TZ), whereas the BMP-GA-TZ uptake efficiency is <17%. Application of the RA scaffold promotes and orients the arrangement of targeting ligands and reduces the shielding effect of corona proteins. This strategy improves the targeting capability and drug delivery of NP in a simulated in vivo milieu.

Background Magnetic nanoparticles such as magnetosomes modified with antibodies allow a high probability of their interaction with targets of interest. Magnetosomes biomineralized by magnetotactic bacteria are in homogeneous nanoscale size and have crystallographic structure, and high thermal and colloidal stability. Camelidae derived nanobodies (Nbs) are small in size, thermal stable, highly water soluble, easy to produce, and fusible with magnetosomes. We aimed to functionalize Nb-magnetosomes for the analysis of the insecticide fipronil. Results Three recombinant magnetotactic bacteria (CF, CF+ , and CFFF) biomineralizing magnetosomes with different abundance of Nbs displayed on the surface were constructed. Compared to magnetosomes from the wild type Magnetospirillum gryphiswaldense MSR-1, all of the Nb-magnetosomes biosynthesized by strains CF, CF+ , and CFFF showed a detectable level of binding capability to fipronil-horseradish peroxidase (H2-HRP), but none of them recognized free fipronil. The Nb-magnetosomes from CFFF were oxidized with H 2 O 2 or a glutathione mixture consisting of reduced glutathione and oxidized glutathione in vitro and their binding affinity to H2-HRP was decreased, whereas that to free fipronil was enhanced. The magnetosomes treated with the glutathione mixture were employed to develop an enzyme-linked immunosorbent assay for the detection of fipronil in water samples, with average recoveries in a range of 78–101%. Conclusions The economical and environmental-friendly Nb-magnetosomes biomineralized by the bacterial strain MSR-1 can be potentially applied to nanobody-based immunoassays for the detection of fipronil or nanobody-based assays in general.

Background Magnetosomes (BMPs) are organelles of magnetotactic bacteria (MTB) that are responsible for mineralizing iron to form magnetite. In addition, BMP is an ideal biomaterial that is widely used in bio- and nano-technological applications, such as drug delivery, tumor detection and therapy, and immunodetection. The use of BMPs to create multifunctional nanocomposites would further expand the range of their applications. Results In this study, we firstly demonstrate that the extracted BMP can remineralize in vitro when it is exposed to AgNO 3 solution, the silver ions (Ag + ) were transported into the BMP biomembrane (MM) and mineralized into a silver crystal on one crystal plane of Fe 3 O 4 . Resulting in the rapid synthesis of an Ag-Fe 3 O 4 hybrid BMP (BMP-Ag). The synergy between the biomembrane, Fe 3 O 4 crystal , and unmineralized iron enabled the remineralization of BMPs at an Ag + concentration ≥ 1.0 mg mL −1 . The BMP-Ag displayed good biocompatibility and antibacterial activity. At a concentration of 2.0 mg/mL, the BMP-Ag and biomembrane removed Ag-Fe 3 O 4 NPs inhibited the growth of gram-negative and gram-positive bacteria. Thus using BMP-Ag as a wound dressing can effectively enhance the contraction of infected wounds. Conclusions This study represents the first successful attempt to remineralize organelles ex vivo, realizing the biosynthesis of hybrid BMP and providing an important advancement in the synthesis technology of multifunctional biological nanocomposites. Graphical abstract

It is well known that when exposed to human blood plasma, nanoparticles are predominantly coated by a layer of proteins, forming a corona that will mediate the subsequent cell interactions. Magnetosomes are protein-rich membrane nanoparticles which are synthesized by magnetic bacteria; these have gained a lot of attention owing to their unique magnetic and biochemical characteristics. Nevertheless, whether bacterial magnetosomes have a corona after interacting with the plasma, and how such a corona affects nanoparticle-cell interactions is yet to be elucidated. The aim of this study was to characterize corona formation around a bacterial magnetosome and to assess the functional consequences. Magnetosomes were isolated from the magnetotactic bacteria, (MSR-1). Size, morphology, and zeta potential were measured by transmission electron microscopy and dynamic light scattering. A quantitative characterization of plasma corona proteins was performed using LC-MS/MS. Protein absorption was further examined by circular dichroism and the effect of the corona on cellular uptake was investigated by microscopy and spectroscopy. Various serum proteins were found to be selectively adsorbed on the surface of the bacterial magnetosomes following plasma exposure, forming a corona. Compared to the pristine magnetosomes, the acquired corona promoted efficient cellular uptake by human vascular endothelial cells. Using a protein-interaction prediction method, we identified cell surface receptors that could potentially associate with abundant corona components. Of these, one abundant corona protein, ApoE, may be responsible for internalization of the magnetosome-corona complex through LDL receptor-mediated internalization. Our findings provide clues as to the physiological response to magnetosomes and also reveal the corona composition of this membrane-coated nanomaterial after exposure to blood plasma.

Derived from magnetotactic bacteria (MTB), magnetosomes consist of magnetite crystals enclosed within a lipid bilayer membrane and are known to possess advantages over artificially synthesized nanoparticles because of the narrow size distribution, uniform morphology, high purity and crystallinity, single magnetic domain, good biocompatibility, and easy surface modification. These unique properties have increasingly attracted researchers to apply bacterial magnetosomes (BMs) in the fields of biology and medicine as MRI imaging contrast agents. Due to the concern of biosafety, a long-term follow-up of the distribution and clearance of BMs after entering the body is necessary. In this study, we tracked changes of BMs in major organs of mice up to 135 days after intravenous injection using a combination of several techniques. We not only confirmed the liver as the well-known targeted organs of BMs, but also found that BMs accumulated in the spleen. Besides, two major elimination paths, as well as the approximate length of time for BMs to be cleared from the mice, were revealed. Together, the results not only confirm that BMs have high biocompatibility, but also provide a long-term in-vivo assessment which may further help to forward the clinical applications of BMs as an MRI contrast agent.

Background Magnetosomes (also called bacterial magnetic nanoparticles; BMPs) are biomembrane-coated nanoparticles synthesized by magnetotactic bacteria (MTB). Engineered BMPs fused to protein A (termed ∆F-BMP-FA) bind antibodies (Abs) automatically, and thus provide a series of potential advantages. However, no report so far has systematically evaluated functional applicability of genetically engineered BMPs. Results We evaluated properties of ∆F-BMP-FA, and developed/optimized culture methods for host strain Magnetospirillum gryphiswaldense ΔF-FA, ∆F-BMP-FA extraction conditions, conditions for Ab conjugation to ∆F-BMP-FA surface, and procedures for antigen detection using ∆F-BMP-FA/Ab complexes (termed BMP-A-Ab). Fed-batch culture for 36 h in a 42-L fermentor resulted in yields (dry weight) of 2.26 g/L for strain ΔF-FA and 62 mg/L for ∆F-BMP-FA. Optimal wash cycle number for ∆F-BMP-FA purification was seven, with magnetic separation following each ultrasonication step. Fusion of protein A to BMPs resulted in ordered arrangement of Abs on BMP surface. Linkage rate 962 μg Ab per mg ∆F-BMP-FA was achieved. BMP-A-Ab were tested for detection of pathogen ( Vibrio parahaemolyticus ; Vp) surface antigen and hapten (gentamicin sulfate). Maximal Vp capture rate for BMP-A-Ab was 90% (higher than rate for commercial immunomagnetic beads), and detection sensitivity was 5 CFU/mL. ∆F-BMP-FA also bound Abs from crude mouse ascites to form complex. Lowest gentamicin sulfate detection line for BMP-A-Ab was 0.01 ng/mL, 400-fold lower than that for double Ab sandwich ELISA, and gentamicin sulfate recovery rate for BMP-A-Ab was 93.2%. Conclusion Our findings indicate that engineered BMPs such as ∆F-BMP-FA are inexpensive, eco-friendly alternatives to commercial immunomagnetic beads for detection or diagnostic immunoassays, and have high Ab-conjugation and antigen-adsorption capacity.

Therapeutic outcomes of colorectal cancer (CRC) are influenced by intestinal microbiota and metabolites. To leverage the tumor-tropism and microbiota-regulating properties of bacteria, we developed oncolytic magnetotactic bacteria (MSR-CPT/APPs) loaded with camptothecin (CPT) and anti-PD-L1 peptide (APP) for targeted chemo-magnetothermal immunotherapy of CRC. To further achieve oral delivery, MSR-CPT/APPs were coated with mulberry leaf lipids and Pluronic F127 (LPs). When exposed to an alternating magnetic field, MSR-CPT/APP@LPs penetrated colonic mucus and reached deep-seated tumors. They elevated proinflammatory cytokine secretion, prolonged T cell recruitment, and reduced immunosuppressive cell proportions by activating the cGAS-STING pathway, inducing immunogenic cell death, and facilitating macrophage polarization to M1 phenotype. Oral MSR-CPT/APP@LPs increased the relative abundances of crucial commensal microorganisms (e.g., Lachnospiraceae family and Alistipes) and influenced their metabolites, like elevating beneficial metabolites (short-chain fatty acids and citrulline) levels and decreasing harmful metabolites (l-glutamine and kynurenic acid) amounts, thereby remodeling the tumor immune microenvironment. Overall, MSR-CPT/APP@LPs foster a supportive intestinal environment and mitigate immunosuppression, achieving the eradication of both primary and distant colorectal tumors. TOC 1. After oral administration, MSR-CPT/APP@LP magnetic motors achieve stable gastrointestinal transit, accumulate in the tumor tissues, and modulate the metabolic program of intestinal microflora. [Display omitted] •Oncolytic magnetotactic bacteria for targeted CRC therapy was constructed.•Protective shells and AMF exposure allow oral bacteria to safely reach deep tumors.•MSR-CPT/APP@LPs plus AMF exposure elicit innate and adaptive immune immunity.•MSR-CPT/APP@LPs plus AMF exposure greatly reprogram gut microbiota and metabolites.•MSR-CPT/APP@LPs plus AMF exposure foster an immune-supportive microenvironment.

Magnetosomes (MAGs) extracted from magnetotactic bacteria are well-defined membrane-enveloped single-domain magnetic nanoparticles. Due to their superior magnetic and structural properties, MAGs constitute potential materials that can be manipulated via genetic and chemical engineering for use in biomedical and biotechnological applications. However, the long-term effects exerted by MAGs on cells are of concern in the context of in vivo applications. Meanwhile, it remains relatively unclear which mechanisms are employed by cells to process and degrade MAGs. Hence, a better understanding of MAGs’ degradation and fundamental signal modulations occurring throughout this process is essential. In the current study, we investigated the potential actions of MAGs on endothelial cells over a 10-day period. MAGs were retained in cells and found to gradually gather in the lysosome-like vesicles. Meanwhile, iron-ion release was observed. Proteomics further revealed a potential cellular mechanism underlying MAGs degradation, in which a group of proteins associated with vesicle biogenesis, and lysosomal enzymes, which participate in protein hydrolysis and lipid degradation, were rapidly upregulated. Moreover, the released iron triggered the regulation of the iron metabolic profiles. However, given that the levels of cell oxidative damage were relatively stable, the released iron ions were handled by iron metabolic profiles and incorporated into normal metabolic routes. These results provide insights into the cell response to MAGs degradation that may improve their in vivo applications.

Magnetosome formation by Magnetospirillum gryphiswaldense MSR-1 is dependent on iron and oxygen levels. We used transcriptome to evaluate transcriptional profiles of magnetic and non-magnetic MSR-1 cells cultured under high-iron and low-iron conditions. A total of 80 differentially expressed genes (DEGs) were identified, including 53 upregulated and 27 downregulated under high-iron condition. These DEGs belonged to the functional categories of biological regulation, oxidation-reduction process, and ion binding and transport, and were involved in sulfur metabolism and cysteine/methionine metabolism. Comparison with our previous results from transcriptome data under oxygen-controlled conditions indicated that transcription of mam or mms was not regulated by oxygen or iron signals. 17 common DEGs in iron- and oxygen-transcriptomes were involved in energy production, iron transport, and iron metabolism. Some unknown-function DEGs participate in iron transport and metabolism, and some are potential biomarkers for identification of Magnetospirillum strains. IrrA and IrrB regulate iron transport in response to low-oxygen and high-iron signals, respectively. Six transcription factors were predicted to regulate DEGs. Fur and Crp particularly co-regulate DEGs in response to changes in iron or oxygen levels, in a proposed joint regulatory network of DEGs. Our findings provide new insights into biomineralization processes under high- vs. low-iron conditions in magnetotactic bacteria.