A newly constructed Agrobacterium-mediated transformation system based on the hisB auxotrophic marker for genetic manipulation in Aspergillus niger.

The filamentous fungus Aspergillus niger is widely exploited as an industrial workhorse for producing enzymes and organic acids. So far, different genetic tools, including CRISPR/Cas9 genome editing strategies, have been developed for the engineering of A. niger. However, these tools usually require a suitable method for gene transfer into the fungal genome, like protoplast-mediated transformation (PMT) or Agrobacterium tumefaciens-mediated transformation (ATMT). Compared to PMT, ATMT is considered more advantageous because fungal spores can be used directly for genetic transformation instead of protoplasts. Although ATMT has been applied in many filamentous fungi, it remains less effective in A. niger. In the present study, we deleted the hisB gene and established an ATMT system for A. niger based on the histidine auxotrophic mechanism. Our results revealed that the ATMT system could achieve 300 transformants per 107 fungal spores under optimal transformation conditions. The ATMT efficiency in this work is 5 - 60 times higher than those of the previous ATMT studies in A. niger. The ATMT system was successfully applied to express the DsRed fluorescent protein-encoding gene from the Discosoma coral in A. niger. Furthermore, we showed that the ATMT system was efficient for gene targeting in A. niger. The deletion efficiency of the laeA regulatory gene using hisB as a selectable marker could reach 68 - 85% in A. niger strains. The ATMT system constructed in our work represents a promising genetic tool for heterologous expression and gene targeting in the industrially important fungus A. niger.

CYP2C19 genetic polymorphism in the Vietnamese population.

Background: Genetic polymorphism of CYP2C19 has been shown to affect enzyme activity and thereby contribute to inter-individual variability in drug metabolism and response. The complete genetic variation of CYP2C19 in Vietnam still remains obscure even though data of common alleles in Vietnamese Kinh have been reported.Aim: To establish the extent of CYP2C19 polymorphism in Vietnamese.Subjects and methods: The promoter and all nine exons of CYP2C19 in 100 healthy unrelated Vietnamese Kinh subjects were sequenced. Additionally, the CYP2C19 variants, *2, *3 and *17 were analysed by RFLP-PCR in 275 subjects of four minor ethnic groups in Vietnam (Tay, Muong, H'Mong and Nung).Results: In 100 Kinh subjects, the percentages of CYP2C19*1, CYP2C19*2, CYP2C19*3 and CYP2C19*17 alleles were 76%, 20.5%, 2.5% and 1%, respectively. Three novel variants in introns 2, 5 and 8 had no impact on mRNA splicing according to the Human Splicing Finder. The prevalence of CYP2C19*17 in Vietnamese Kinh was significantly lower compared with figures found in Western Asia and Europe, while CYP2C19*2 frequency was statistically higher than that in Western Asia and several countries in Europe. The frequency of CYP2C19*2 in Kinh was significantly lower than in the other four ethnic minorities.Conclusion: These results provide information on CYP2C19 polymorphism in the Vietnamese population, which could be useful for optimising drug therapies and precision medicine studies.

Correction: Nguyen et al. Green Silver Nanoparticles Formed by Phyllanthus urinaria, Pouzolzia zeylanica, and Scoparia dulcis Leaf Extracts and the Antifungal Activity. Nanomaterials 2020, 10, 542.

In the original publication [...].

MFSD7c functions as a transporter of choline at the blood-brain barrier.

Mutations in the orphan transporter MFSD7c (also known as Flvcr2), are linked to Fowler syndrome. Here, we used Mfsd7c knockout (Mfsd7c-/-) mice and cell-based assays to reveal that MFSD7c is a choline transporter at the blood-brain barrier (BBB). We performed comprehensive metabolomics analysis and detected differential changes of metabolites in the brains and livers of Mfsd7c-/-embryos. Particularly, we found that choline-related metabolites were altered in the brains but not in the livers of Mfsd7c-/- embryos. Thus, we hypothesized that MFSD7c regulates the level of choline in the brain. Indeed, expression of human MFSD7c in cells significantly increased choline uptake. Interestingly, we showed that choline uptake by MFSD7c is greatly increased by choline-metabolizing enzymes, leading us to demonstrate that MFSD7c is a facilitative transporter of choline. Furthermore, single-cell patch clamp analysis showed that the import of choline by MFSD7c is electrogenic. Choline transport function of MFSD7c was shown to be conserved in vertebrates, but not in yeasts. We demonstrated that human MFSD7c is a functional ortholog of HNM1, the yeast choline importer. We also showed that several missense mutations identified in patients exhibiting Fowler syndrome had abolished or reduced choline transport activity. Mice lacking Mfsd7c in endothelial cells of the central nervous system suppressed the import of exogenous choline from blood but unexpectedly had increased choline levels in the brain. Stable-isotope tracing study revealed that MFSD7c was required for exporting choline derived from lysophosphatidylcholine in the brain. Collectively, our work identifies MFSD7c as a choline exporter at the BBB and provides a foundation for future work to reveal the disease mechanisms of Fowler syndrome.

Green Silver Nanoparticles Formed by Phyllanthus urinaria, Pouzolzia zeylanica, and Scoparia dulcis Leaf Extracts and the Antifungal Activity.

Phytoconstituents presenting in herbal plant broths are the biocompatible, regenerative, and cost-effective sources that can be utilized for green synthesis of silver nanoparticles. Different plant extracts can form nanoparticles with specific sizes, shapes, and properties. In the study, we prepared silver nanoparticles (P.uri.AgNPs, P.zey.AgNPs, and S.dul.AgNPs) based on three kinds of leaf extracts (Phyllanthus urinaria, Pouzolzia zeylanica, and Scoparia dulcis, respectively) and demonstrated the antifungal capacity. The silver nanoparticles were simply formed by adding silver nitrate to leaf extracts without using any reducing agents or stabilizers. Formation and physicochemical properties of these silver nanoparticles were characterized by UV-vis, Fourier transforms infrared spectroscopy, scanning electron microscope, transmission electron microscope, and energy dispersive X-ray spectroscopy. P.uri.AgNPs were 28.3 nm and spherical. P.zey.AgNPs were 26.7 nm with hexagon or triangle morphologies. Spherical S.dul.AgNPs were formed and they were relatively smaller than others. P.uri.AgNPs, P.zey.AgNPs and S.dul.AgNPs exhibited the antifungal ability effective against Aspergillus niger, Aspergillus flavus, and Fusarium oxysporum, demonstrating their potentials as fungicides in the biomedical and agricultural applications.

Bacterial community structures and biodegradation kinetic of Tiamulin antibiotic degrading enriched consortia from swine wastewater.

The antibiotic tiamulin (TIA) is common and widely used medication for dysentery eradication in swine productions. Tiamulin persists in livestock manure, and its residues have been found in various environment. This work obtained four tiamulin-degrading enriched bacterial consortia from a covered anaerobic lagoon system and a stabilized pond system of swine farms. Tiamulin was efficiently removed by the enriched cultures at the concentrations between 2.5 and 200 mg/L, with a removal of 60.1-99.9% during 16 h and a degradation half-life of 4.5-15.7 h. The stabilized pond system cultured with taimulin solely could eliminate tiamulin at the highest rates. The logistic substrate degradation model fit most of the experimental data. Next-generation amplicon sequencing was conducted, and it was found that the bacterial community was significantly impacted by the inoculum source, nutrient addition, and high tiamulin concentrations. Principal coordinate analysis (PCoA) indicated the similarity of bacterial communities in the original enriched samples and the 2.5 mg/L tiamulin-removed cultures. The 200 mg/L consortia were rather different and became similar to the other 200 mg/L consortia from different sources and cultures without nutrient supplementation. Shannon and Simpson indices suggested a reduction in bacterial diversity at high concentrations. The microbes that had high growth in the most efficient enriched culture, or which were abundant in all samples, or which increased with higher tiamulin concentrations were likely to be the major tiamulin-degrading bacteria. This is the first report suggested the possible roles of Achromobacter, Delftia, Flavobacterium, Pseudomonas, and Stenotrophomonas in tiamulin degradation.

Therapeutic Development of Interrelated Metabolic and Neurodegenerative Disorders.

BACKGROUND: Metabolic syndromes such as insulin resistance, type 2 diabetes and obesity share common pathogenic pathways with some age-related neurodegenerative disorders. Impaired insulin signaling, inflammation, mitochondrial dysfunction and ER stress can be both causatives and consequences in both groups of the diseases. Patients with chronic metabolic disorders therefore have potential risks to develop neurological diseases in late-age phase and vice versa those who with neurodegenerative diseases also have impairments in metabolic signaling. METHOD: In this review, we summarize about the interrelation between pathogenic pathways, common drug targets as well as known and developing therapeutics for these "modern" diseases. RESULTS: There are conventional medicines for insulin resistance associated metabolic disorders such as insulin analogues, insulin sensitizers and ER stress releasers which have been suggested in the treatments of some neurodegenerative diseases. Some used or tested therapeutics such as bromocriptine, memantine and α-2A adrenergic antagonists for Parkinson's and Alzheimer's diseases, vice versa, were promisingly shown as alternative or complementary drugs for metabolic syndromes. CONCLUSION: Therefore, it is important and possible to consider contemporary control and intervention for both diseases.

Suggestion of suitable animal models for in vivo studies of protein tyrosine phosphatase 1b (PTP1B) inhibitors using computational approaches.

PTP1B is a prototypic enzyme of the superfamily protein tyrosine phosphatases (PTPs) which are critical regulators of tyrosine phosphorylation-dependent signaling events. It is a highly plausible candidate for designing therapeutic inhibitors of obesity and type 2 diabetes (T2D). In this study, a detailed comparative analysis to reveal the evolutionary relationship of human PTP1B among related vertebrates has been addressed. The phylogenetic trees were constructed with maximum likelihood algorithm by PhyML package on the basis of multiple sequence alignment (MSA) by ClustalΩ and T-coffee. Mutational variability of the sequences corresponding to the 3D structure (pdb: 2vev) was analyzed with Consurf software. The comparative analysis by inhibitor docking to different models was made to confirm the suitability of models. As a result, the PTP1B or PTP non-receptor type 1 homologies show high conservativity where about 70% positions on primary structures are conserved. Within PTP domain (3-277), the most variable positions are 12, 13, 19 and 24 which is a part of the second aryl binding site. Moreover, there are important evolutional mutations that can change the conformation of the proteins, for instance, hydrophilic N139 changed to hydrophobic Gly (mPTP1B); E132 to proline in the hydrophobic core structure or Y46 to cystein in pTyr recognition loop. These variations/differences should be taken into account for rational inhibitor design and in choosing suitable animal models for drug testing and evaluation. Moreover, our study suggests critically potential models which are Heterocephalus glaber, Tupaia chinensis, Sus scrofa, and Rattus norvegicus in addition to the best one Macaca fascicularis. Among these models, the H.glaber and R.norvegicus are preferable over M.musculus thanks to their similarity in binding affinity and binding modes to investigated PTP1B inhibitors.