Synthesis of Novel 4-Acyloxy-2'-bromo-6'-chloropodophyllotoxin Derivatives Displaying Significant Insecticidal Activity against Mythimna separata.

In order to explore novel natural product-based insecticidal agent, two important intermediates (2 and 3) and 4-acyloxy-2'-bromo-6'-chloropodophyllotoxin derivatives (4a-f and 5a-f) were designed and prepared, and their structures were confirmed by 1H NMR, 13C NMR, HRMS, ESI-MS, optical rotation and melting point (mp). The stereochemical configuration of compound 4b was unambiguously confirmed by single-crystal X-ray diffraction. Moreover, we evaluated the insecticidal activity of target compounds 4a-f and 5a-f against a serious agricultural pest of Mythimna separata by using the leaf-dipping method. Among all tested compounds, compounds 4d, 5d and 5f exhibited stronger insecticidal activity with a final mortality rate exceeding 60%. Especially compound 5d exhibited the best insecticidal activity, with a final mortality rate of 74.1%. It has been proven that introducing bromine or chlorine atoms at the C-2', C-2' and C-6' positions of the E ring of podophyllotoxin can produce more potent compounds. In addition, the configuration of the C-4 position is important for insecticidal activity, and 4ß-configuration is optimal. This will pave the way for further design, structural modification, and development of derivatives of podophyllotoxin as insecticidal agents.

Lactiplantibacillus plantarum postbiotic protects against Salmonella infection in broilers via modulating NLRP3 inflammasome and gut microbiota.

Salmonella infection is a major concern in poultry production which poses potential risks to food safety. Our previous study confirmed that Lactiplantibacillus plantarum (LP) postbiotic exhibited a strong antibacterial capacity on Salmonella in vitro. This study aimed to investigate the beneficial effects and underlying mechanism of LP postbiotic on Salmonella-challenged broilers. A total of 240 one-day-old male yellow-feathered broilers were pretreated with 0.8% deMan Rogosa Sharpe (MRS) medium or 0.8% LP postbiotic (LP cell-free culture supernatant, LPC) in drinking water for 28 d, and then challenged with 1×109 CFU Salmonella enterica serovar Enteritidis (SE). Birds were sacrificed 3 d postinfection. Results showed that LPC maintained the growth performance by increasing body weight (BW), average daily gain (ADG), and average daily feed intake (ADFI) in broilers under SE challenge. LPC significantly attenuated SE-induced intestinal mucosal damage. Specifically, it decreased the intestinal injury score, increased villus length and villus/crypt, regulated the expression of intestinal injury-related genes (Villin, matrix metallopeptidase 3 [MMP3], intestinal fatty acid-binding protein [I-FABP]), and enhanced tight junctions (zona occludens-1 [ZO-1] and Claudin-1). SE infection caused a dramatic inflammatory response, as indicated by the up-regulated concentrations of interleukin (IL)-1ß, IL-6, TNF-α, and the downregulation of IL-10, while LPC pretreatment markedly reversed this trend. We then found that LPC inhibited the activation of NOD-like receptor thermal protein domain associated protein 3 (NLRP3) inflammasome by decreasing the gene expression of Caspase-1, IL-lß, and IL-18. Furthermore, LPC suppressed NLRP3 inflammasome activation by inhibiting nuclear factor-kappa B (NF-κB) signaling pathway (the reduced levels of toll-like receptor 4 [TLR4], myeloid differentiation factor 88 [MyD88], and NF-κB). Finally, our results showed that LPC regulated gut microbiota by enhancing the percentage of Ligilactobacillus and decreasing Alistipes and Barnesiella. In summary, we found that LP postbiotic was effective to protect broilers against Salmonella infection, possibly through suppressing NLRP3 inflammasome and optimizing gut microbiota. Our study provides the potential of postbiotics on prevention of Salmonella infection in poultry.

Lactiplantibacillus plantarum Postbiotics Suppress Salmonella Infection via Modulating Bacterial Pathogenicity, Autophagy and Inflammasome in Mice.

Our study aimed to explore the effects of postbiotics on protecting against Salmonella infection in mice and clarify the underlying mechanisms. Eighty 5-week-old C57BL/6 mice were gavaged daily with Lactiplantibacillus plantarum (LP)-derived postbiotics (heat-killed bacteria, LPBinactive; culture supernatant, LPC) or the active bacteria (LPBactive), and gavaged with Salmonella enterica Typhimurium (ST). The Turbidimetry test and agar diffusion assay indicated that LPC directly inhibited Salmonella growth. Real-time PCR and biofilm inhibition assay showed that LPC had a strong ability in suppressing Salmonella pathogenicity by reducing virulence genes (SopE, SopB, InvA, InvF, SipB, HilA, SipA and SopD2), pili genes (FilF, SefA, LpfA, FimF), flagellum genes (FlhD, FliC, FliD) and biofilm formation. LP postbiotics were more effective than LP on attenuating ST-induced intestinal damage in mice, as indicated by increasing villus/crypt ratio and increasing the expression levels of tight junction proteins (Occludin and Claudin-1). Elisa assay showed that LP postbiotics significantly reduced ST-induced inflammation by regulating the levels of inflammatory cytokines (the increased IL-4 and IL-10 and the decreased TNF-α) in serum and ileum (p < 0.05). Furthermore, LP postbiotics inhibited the activation of NOD-like receptor thermal protein domain-associated protein 3 (NLRP3) inflammasome by decreasing the protein expression of NLRP3 and Caspase-1, and the gene expression of Caspase-1, IL-1ß and IL-18. Meanwhile, both LPC and LPB observably activated autophagy under ST infection, as indicated by the up-regulated expression of LC3 and Beclin1 and the downregulated p62 level (p < 0.05). Finally, we found that LP postbiotics could trigger an AMP-activated protein kinase (AMPK) signaling pathway to induce autophagy. In summary, Lactiplantibacillus plantarum-derived postbiotics alleviated Salmonella infection via modulating bacterial pathogenicity, autophagy and NLRP3 inflammasome in mice. Our results confirmed the effectiveness of postbiotics agents in the control of Salmonella infection.

Synthesis and Characterization of High Glycolic Acid Content Poly(glycolic acid-co-butylene adipate-co-butylene terephthalate) and Poly(glycolic acid-co-butylene succinate) Copolymers with Improved Elasticity.

Poly(glycolic acid) (PGA) is a biodegradable polymer with high gas barrier properties, mechanical strength, and heat deflection temperature. However, PGA's brittleness severely limits its application in packaging, creating a need to develop PGA-based copolymers with improved elasticity that maintain its barrier properties and hydrolytic degradability. In this work, a series of PGBAT (poly(glycolic acid-co-butylene) adipate-co-butylene terephthalate) copolymers containing 21-92% glycolic acid (nGA) with Mw values of 46,700-50,600 g mol-1 were synthesized via melt polycondensation, and the effects of altering the nGA on PGBAT's thermomechanical properties and hydrolysis rate were investigated. Poly(glycolic acid-co-butylene succinate) (PGBS) and poly(glycolic acid-co-butylene terephthalate) (PGBT) copolymers with high nGA were synthesized for comparison. DSC analysis revealed that PGBAT21 (nGA = 21%) and PGBAT92 were semicrystalline, melting between 102.8 and 163.3 °C, while PGBAT44, PGBAT86-89, PGBT80, and PGBS90 were amorphous, with Tg values from -19.0 to 23.7 °C. These high nGA copolymers showed similar rates of hydrolysis to PGA, whereas those containing <50% GA showed almost no mass loss over the testing period. Their mechanical properties were highly dependent upon their crystallinity and improved significantly after annealing. Of the high nGA copolymers, annealed PGBS90 (Mw 97,000 g mol-1) possessed excellent mechanical properties with a modulus of 588 MPa, tensile strength of 30.0 MPa, and elongation at break of 171%, a significant improvement on PGA's elongation at break of 3%. This work demonstrates the potential of enhancing PGA's flexibility by introducing minor amounts of low-cost diols and diacids into its synthesis.

Study on Management of Blood Transfusion Therapy in Patients with Hereditary Spherocytosis.

Hereditary spherocytosis (HS) is a chronic hemolytic disorder caused by inherited defects in the red blood cell membrane. This study discusses the treatment strategy for the decline in hemoglobin level in three HS probands with moderately severe or severe hemolysis and summarizes the appropriate laboratory tests that help improve clinical management of blood transfusion in HS patients. Three probands who were diagnosed with HS in our hospital and their family members were included in this study. Clinical data of the three families were reviewed to summarize their hematopoietic characteristics. DNA from all family members of the 3 HS probands was amplified by polymerase chain reaction (PCR) and sequenced by the Sanger method to assess genetic relation for HS. Based on the sequencing results, the type of mutated membrane protein in each proband was analyzed using the eosin-5'-maleimide (EMA) binding test and sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE). The hemoglobin level was reduced in all 3 probands after different levels of infection. The fluorescence of EMA-labeled red blood cell (RBC) was decreased. DNA sequencing showed that His54Pro, Leu1858Val, and 6531-12C>T compound heterozygous mutations were present in the SPTA1 gene of patient I-1, Arg344Gln and c.609+86G>A heterozygous mutations were present in the SLC4A1 gene of patient II-1, and Leu2032Pro homozygous mutation was present in the SPTB gene of patient III-1. SDS-PAGE results demonstrated that the concentration of band 3 was reduced in II-1, whereas the levels of the corresponding mutant proteins in the other probands were unchanged. The family members of the respective patients presented mutations in major genes causing HS. The Leu2032Pro mutation identified in patient III-1 is a new missense mutation of the SPTB gene in the Chinese population that has never been reported in literature previously. The presence or absence of acute or chronic infections is a critical deciding factor for the treatment and clinical management of HS patient via blood transfusion. For patients with infections, hemoglobin concentration can be restored once the infection is controlled, thus obviating the need for proper infection control before blood transfusion.

Novel Phosphorus-Nitrogen-Containing Ionic Liquid Modified Metal-Organic Framework as an Effective Flame Retardant for Epoxy Resin.

Metal-organic frameworks (MOFs) have shown great potential in flame retardant applications; however, strategies for fully exploiting the advantages of MOFs in order to further enhance the flame retardant performance are still in high demand. Herein, a novel MOF composite was designed through the generated cooperative role of MOF (NH2-MIL-101(Al)) and a phosphorus-nitrogen-containing ionic liquid ([DPP-NC3bim][PMO]). The ionic liquid (IL) was composed of imidazole cation modified with diphenylphosphinic group (DPP) and phosphomolybdic acid (PMoA) anions, which can trap the degrading polymer radicals and reduce the smoke emission. The MOF acts as a porous host and can avoid the agglomeration of ionic liquid. Meanwhile, the -NH2 groups of NH2-MIL-101(Al) can increase the compatibility with epoxy resin (EP). The framework is expected to act as an efficient insulating barrier to suppress the flame spread. It was demonstrated that the MOF composite (IL@NH2-MIL-101(Al)) is able to effectively improve the fire safety of EP at low additions (3 wt. %). The LOI value of EP/IL@NH2-MIL-101(Al) increased to 29.8%. The cone calorimeter results showed a decreased heat release rate (51.2%), smoke production rate (37.8%), and CO release rate (44.8%) of EP/IL@NH2-MIL-101(Al) with respect to those of neat EP. This strategy can be extended to design other advanced materials for flame retardant.

Hierarchically Assembled Graphene Oxide Composite Membrane with Self-Healing and High-Efficiency Water Purification Performance.

Graphene oxide (GO) with a two-dimensional lamellar structure and single-atom thickness has exhibited advantages in water purification by stacking to a continuous membrane. However, a proper method to further increase the separation property of the GO membrane is still urgently needed. Besides, damage to the membrane during the full-scale application processes and the resulted consequential loss are prevalent problems need to be solved. Here, a hierarchically assembled GO composite membrane was developed that can achieve high-efficiency water purification performance and self-healing property via the synergistic effect of the metal-organic framework (MOF) and the coated hydrophilic layer of chitosan. The intercalated MOF effectively expanded the channel space of GO and endowed the channels with molecular-sieving property. Meanwhile, the coated chitosan layer can selectively adsorb water and achieve self-healing through the cross-linking reaction. The prepared GO composite membrane shows largely improved water flux (14.62 L m-2 h-1 bar-1), increased 344% than the water flux of the GO membrane, high rejection ratio (>99% for dyes), and good antifouling performance. In addition, the damaged GO composite membrane can recover its water flux (95%) and rejection ratio (96%) through a facile self-healing process.

[Application of High Resolution Melting Curve Analysis in Detection of SLC4A1 Gene Mutation in Patients with Hereditary Spherocytosis].

OBJECTIVE: To investigate the feasibility and clinical significance of high resolution melting(HRM) curve analysis to detect SLC4A1 gene D38A and K56E mutations in the patients with hereditary spherocytosis(HS). METHODS: Peripheral blood was collected from 23 cases of HS for routine tests and their genomic DNA was extracted by routine technique. Specific primers of mutation sites D38A and K56E of SLC4A1 gene were designed. The HRM method was used to analyze all the samples, and then the results of HRM were verified with DNA sequencing technology. RESULTS: Among 23 specimens of HS patients, 6 cases of heterozygous mutant gene were detected by HRM technology, including 3 cases of D38A mutation and 3 cases of K56E mutation, which were confirmed by DNA sequencing. CONCLUSION: The HRM technology can correctly detect 2 common mutation sites including D38A and K56E in SLC4A1 gene in an efficient, fast, and reliable way, which not only can be used for clinical diagnosis, but also expected to be a new method for clinical researchers to define gene mutation spectrum in HS patients.

Study of the micro-climate and bacterial distribution in the deadspace of N95 filtering face respirators.

It is common for people to use N95 filtering facepiece respirators (FFRs) in daily life, especially in locations where particulate matter (PM2.5) concentration is rising. Wearing N95 FFRs is helpful to reduce inhalation of PM2.5. Although N95 FFRs block at least 95% of particles from the atmosphere, the deadspace of N95 FFRs could be a warm, wet environment that may be a perfect breeding ground for bacterial growth. This work studies the micro-climate features including the temperature distribution and water vapor condensation in the deadspace of an N95 FFR using the computational fluid dynamics (CFD) method. Then, the temperature and relative humidity inside the same type of N95 FFR are experimentally measured. There is a good agreement between the simulation and experimental results. Moreover, an experiment is conducted to study the distribution of bacteria sampled from the inner surface of an N95 FFR after donning.

[Analysis of SPTA1 gene mutations in a patient with hereditary elliptocytosis].

OBJECTIVE: To detect disease-causing mutations in a patient with hereditary elliptocytosis. METHODS: Sodium dodecyl sulfate polyacrylamide gel electropheresis (SDS-PAGE) was used to identify the type of erythrocyte membrane protein defect. Potential mutations of the exons and adjacent introns of relevant genes were analyzed by Sanger sequencing. RESULTS: SDS-PAGE has failed to detect any difference between the patient and healthy controls. However, Sanger sequencing has detected three mutations in the SPTA1 gene in the patient, which included c.5077A>C (p.Lys1693Gln) missense mutation in exon 36, c.5572C>G (p.Leu1858Val) missense mutation in exon 40, and a IVS45nt-12C>T in intron 45. The father and grandmother of the patient were both heterozygous for c.5077A>C mutation, while her mother was heterozygous for c.5572C>G and IVS45nt-12C>T mutations. CONCLUSION: The hereditary elliptocytosis in the patient may be attributed to the synergistic action of c.5077A>C, c.5572C>G and IVS45nt-12C>T mutations of the SPTA1 gene.

Analysis of the causes of the misdiagnosis of hereditary spherocytosis.

Hereditary spherocytosis (HS) is an inherited hemolytic disease with clinical diversities. The aim of the present study was to examine the reasons for prolonged misdiagnosis and mistherapy of HS in a Chinese patient, and to summarize the laboratory screening and treatment methods for this disease in increasing the knowledge towards HS. Clinical data of the proband was reviewed. The proband was first screened by detection of eosin-5'-maleimide (EMA)-labeled red blood cells (RBCs) using flow cytometry. The type of protein defect in the extracted RBC membrane proteins was confirmed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). Mutant fragments were verified using direct DNA sequencing and matrix-assisted laser desorption/ionization time of flight (MALDI-TOF) mass spectroscopy. The proband showed a significant hemolytic tendency and significant reduction in the number of EMA-labeled RBCs. DNA sequencing indicated three site mutations in the SPTA1 gene, including His54Pro, Leu1858Val and 6531-12C>T. Additional DNA analysis of the three mutations in the parents of the proband showed that both the Leu1858Val and 6531­12C>T mutations were carried by the father and the His54Pro mutation was carried by the mother. Moreover, the mutated peptides were identified by MALDI-TOF mass spectroscopy. HS has diverse clinical manifestations and is easily missed, misdiagnosed and mistreated. Therefore, a comprehensive analysis involving a routine blood test, blood smear, EMA labeling (flow cytometry) and SDS-PAGE can effectively distinguish HS from thalassemia, glucose-6-phosphate deficiency, iron-deficiency anemia and autoimmune hemolytic anemia.

Novel compound heterozygous SPTA1 mutations in a patient with hereditary elliptocytosis.

Hereditaryelliptocytosis (HE) is a hereditary hemolytic disease, characterized by the presence of many elliptical erythrocytes in the peripheral blood that is caused by abnormal cytoskeletal proteins in the erythrocyte membrane. In the present study, a novel, causal HE mutation was reported. Routine blood examinations were performed on the proband and their family, and the fluorescence intensity of eosin­5­maleimide (EMA)­labeled erythrocytes was determined via flow cytometry. Subsequently, DNA was extracted from the peripheral blood of the proband and their family members, and amplified by quantitative polymerase chain reaction. The Sanger sequencing approach was used to determine and identify gene mutations, which were verified by matrix­assisted laser desorption­ionization time of flight (MALDI­TOF) mass spectrometry. To exclude genetic polymorphisms, newly identified mutations were subjected to large­scale gene screening using high­resolution melt analysis. Protein expression levels in the erythrocyte membrane of the proband were determined via SDS­PAGE, which demonstrated that, compared with healthy controls, the proband exhibited a reduction in EMA­labeled erythrocytes. In addition, DNA analysis demonstrated that the proband carried three mutations in the spectrin α chain erythrocytic 1 (SPTA1) gene: c.161A>C, c.5572C>G and 6531­12C>T. The corresponding mutant polypeptides were also analyzed by MALDI­TOF mass spectroscopy. SDS­PAGE analysis indicated that the proband exhibited normal levels of erythrocyte membrane proteins. In the present study, a novel HE case with a His54Pro mutation in the SPTA1 gene was reported. The results suggested that the His54Pro mutation influenced the role of erythrocyte membrane proteins without reducing its level of expression.