Bacteriophage defends murine gut from Escherichia coli invasion via mucosal adherence.

Bacteriophage are sophisticated cellular parasites that can not only parasitize bacteria but are increasingly recognized for their direct interactions with mammalian hosts. Phage adherence to mucus is known to mediate enhanced antimicrobial effects in vitro. However, little is known about the therapeutic efficacy of mucus-adherent phages in vivo. Here, using a combination of in vitro gastrointestinal cell lines, a gut-on-a-chip microfluidic model, and an in vivo murine gut model, we demonstrated that a E. coli phage, øPNJ-6, provided enhanced gastrointestinal persistence and antimicrobial effects. øPNJ-6 bound fucose residues, of the gut secreted glycoprotein MUC2, through domain 1 of its Hoc protein, which led to increased intestinal mucus production that was suggestive of a positive feedback loop mediated by the mucus-adherent phage. These findings extend the Bacteriophage Adherence to Mucus model into phage therapy, demonstrating that øPNJ-6 displays enhanced persistence within the murine gut, leading to targeted depletion of intestinal pathogenic bacteria.

Development of a CRISPR/Cas12a-based fluorescent detection method of Senecavirus A.

BACKGROUND: Senecavirus A (SVA), identified in 2002, is known to cause porcine idiopathic vesicular disease (PIVD), which presents with symptoms resembling other vesicular diseases. This similarity complicates field diagnosis. Conventional molecular diagnostic techniques are limited by their cost, sensitivity, and requirement for complicated instrumentation. Therefore, developing an effective and accurate diagnostic method is crucial for timely identification and isolation of affected pigs, thereby preventing further disease spread. METHODS: In this study, we developed a highly-specific and ultra-sensitive SVA detection method powered by CRISPR/Cas12a. To enhance the availability in laboratories with varied equipment conditions, microplate reader and ultraviolet light transilluminator were introduced. Moreover, PCR amplification has also been incorporated into this method to improve sensitivity. The specificity and sensitivity of this method were determined following the preparation of the recombinant Cas12a protein and optimization of the CRISPR/Cas12a-based trans-cleavage system. RESULTS: The method demonstrated no cross-reactivity with ten kinds of viruses of swine. The minimum template concentration required to activate substantial trans-cleavage activity was determined to be 106 copies/µL of SVA templates. However, when PCR amplification was incorporated, the method achieved a detection limit of one copy of SVA templates per reaction. It also exhibited 100% accuracy in simulated sample testing. The complete testing process does not exceed three hours. CONCLUSIONS: Importantly, this method utilizes standard laboratory equipment, making it accessible for use in resource-limited settings and facilitating widespread and ultra-sensitive screening during epidemics. Overall, the development of this method not only broadens the array of tools available for detecting SVA but also holds significant promise for controlling the spread of PIVD.

Phosphorus flow characteristics in the waste system of Poyang Lake Watershed over the past 70 years.

With the intensification of human activities, the amount of phosphorus (P)-containing waste has increased. When such waste is not recycled, P is released into the environment, leading to environmental issues such as the eutrophication of water bodies. In this study, based on the material flow analysis method, a P Waste Flow analysis model (P-WFA) was developed to analyze the P flow in the waste system of Poyang Lake, the largest freshwater lake in China. To address the research gap in long-term P flow analysis at the watershed scale, this study quantified the P content in the waste system of the Poyang Lake Watershed from 1950 to 2020. The analysis revealed that from 1950 to 2020, the total P input into the waste system increased from 5.49 × 104 tons in 1950 to 2.28 × 105 tons in 2020. The breeding industry system was identified as the primary source, accounting for 25.19-41.59 % of the total waste system. Over the past 70 years, P loss to surface water from waste systems has been primarily facilitated by manure from the breeding industry, as well as drainage from crop farming systems (77.74 % in 2020). At the same time, the P recycling rate (PRR) of the waste system exhibited an initial increase followed by a decrease, increasing from 44.14 % to 47.75 % before dropping to 44.41 %. Population growth, urbanization, and changes in consumption levels in Jiangxi Province have led to changes in the dietary structure and fertilizer use, consequently affecting the P cycling pattern. This study presents a comprehensive P flow model for waste systems in the Poyang Lake Watershed. This model can be used as a reference to enhance P cycling and manage P loss in other large freshwater lakes.

Transcriptomic and metabolomic dissection of skeletal muscle of crossbred Chongming white goats with different meat production performance.

BACKGROUND: The transcriptome and metabolome dissection of the skeletal muscle of high- and low- growing individuals from a crossbred population of the indigenous Chongming white goat and the Boer goat were performed to discover the potential functional differentially expressed genes (DEGs) and differential expression metabolites (DEMs). RESULTS: A total of 2812 DEGs were detected in 6 groups at three time stages (3,6,12 Month) in skeletal muscle using the RNA-seq method. A DEGs set containing seven muscle function related genes (TNNT1, TNNC1, TNNI1, MYBPC2, MYL2, MHY7, and CSRP3) was discovered, and their expression tended to increase as goat muscle development progressed. Seven DEGs (TNNT1, FABP3, TPM3, DES, PPP1R27, RCAN1, LMOD2) in the skeletal muscle of goats in the fast-growing and slow-growing groups was verified their expression difference by reverse transcription-quantitative polymerase chain reaction. Further, through the Liquid chromatography-mass spectrometry (LC-MS) approach, a total of 183 DEMs in various groups of the muscle samples and these DEMs such as Queuine and Keto-PGF1α, which demonstrated different abundance between the goat fast-growing group and slow-growing group. Through weighted correlation network analysis (WGCNA), the study correlated the DEGs with the DEMs and identified 4 DEGs modules associated with 18 metabolites. CONCLUSION: This study benefits to dissection candidate genes and regulatory networks related to goat meat production performance, and the joint analysis of transcriptomic and metabolomic data provided insights into the study of goat muscle development.

Detection and Analysis of Antidiarrheal Genes and Immune Factors in Various Shanghai Pig Breeds.

The aim of this study was to identify effective genetic markers for the Antigen Processing Associated Transporter 1 (TAP1), α (1,2) Fucosyltransferase 1 (FUT1), Natural Resistance Associated Macrophage Protein 1 (NRAMP1), Mucin 4 (MUC4) and Mucin 13 (MUC13) diarrhea-resistance genes in the local pig breeds, namely Shanghai white pigs, Fengjing pigs, Shawutou pigs, Meishan pigs and Pudong white pigs, to provide a reference for the characterization of local pig breed resources in Shanghai. Polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLR) and sequence sequencing were applied to analyze the polymorphisms of the above genes and to explore the effects on the immunity of Shanghai local pig breeds in conjunction with some immunity factors. The results showed that both TAP1 and MUC4 genes had antidiarrheal genotype GG in the five pig breeds, AG and GG genotypes of the FUT1 gene were detected in Pudong white pigs, AA antidiarrheal genes of the NRAMP1 gene were detected in Meishan pigs, the AB type of the NRAMP1 gene was detected in Pudong white pigs, and antidiarrheal genotype GG of the MUC13 gene was only detected in Shanghai white pigs. The MUC13 antidiarrhea genotype GG was only detected in Shanghai white pigs. The TAP1 gene was moderately polymorphic in Shanghai white pigs, Fengjing pigs, Shawutou pigs, Meishan pigs and Pudong white pigs, among which TAP1 in Shanghai white pigs and Shawutou pigs did not satisfy the Hardy-Weinberg equilibrium. The FUT1 gene of Pudong white pigs was in a state of low polymorphism. NRAMP1 of Meishan pigs and Pudong white pigs was in a state of moderate polymorphism, which did not satisfy the Hardy-Weinberg equilibrium. The MUC4 genes of Shanghai white pigs and Pudong white pigs were in a state of low polymorphism, and the MUC4 genes of Fengjing pigs and Shawutou pigs were in a state of moderate polymorphism, and the MUC4 genes of Fengjing pigs and Pudong white pigs did not satisfy the Hardy-Weinberg equilibrium. The MUC13 gene of Shanghai white pigs and Pudong white pigs was in a state of moderate polymorphism. Meishan pigs had higher levels of IL-2, IL-10, IgG and TNF-α, and Pudong white pigs had higher levels of IL-12 than the other pigs. The level of interleukin 12 (IL-12) was significantly higher in the AA genotype of the MUC13 gene of Shanghai white pigs than in the AG genotype. The indicator of tumor necrosis factor alpha (TNF-α) in the AA genotype of the TAP1 gene of Fengjing pigs was significantly higher than that of the GG and AG genotypes. The indicator of IL-12 in the AG genotype of the Shawutou pig TAP1 gene was significantly higher than that of the GG genotype. The level of TNF-α in the AA genotype of the NRAMP1 gene of Meishan pigs was markedly higher than that of the AB genotype. The IL-2 level of the AG type of the FUT1 gene was obviously higher than that of the GG type of Pudong white pigs, the IL-2 level of the AA type of the MUC4 gene was dramatically higher than that of the AG type, and the IgG level of the GG type of the MUC13 gene was apparently higher than that of the AG type. The results of this study are of great significance in guiding the antidiarrhea breeding and molecular selection of Shanghai white pigs, Fengjing pigs, Shawutou pigs, Meishan pigs and Pudong white pigs and laying the foundation for future antidiarrhea breeding of various local pig breeds in Shanghai.

Evaluation of Dry Ice for Short-Term Storage and Transportation of Frozen Boar Semen.

To address the safety problems posed by the transportation of boar semen using LN, this study was conducted on the short-term storage of frozen boar semen in dry ice (-79 °C). Boar semen frozen in LN was transferred to dry ice, kept for 1 day, 3 days, 5 days, 7 days, or 8 days, and then moved back to LN. The quality of frozen semen stored in LN or dry ice was determined to evaluate the feasibility of short-distance transportation with dry ice. The results showed that 60 °C for 8 s was the best condition for thawing frozen semen stored in dry ice. No significant differences in spermatozoa motility, plasma membrane integrity, or acrosome integrity were observed in semen after short-term storage in dry ice compared to LN (p > 0.05). There were no significant changes in antioxidant properties between storage groups either (p > 0.05). In conclusion, dry ice could be used as a cold source for the short-term transportation of frozen boar semen for at least 7 days, without affecting sperm motility, morphological integrity, or antioxidant indices.

Virological evaluation of natural and modified attapulgite against porcine epidemic diarrhoea virus.

BACKGROUND: The Porcine Epidemic Diarrhea Virus (PEDV) has caused significant economic losses in the global swine industry. As a potential drug for treating diarrhea, the antiviral properties of attapulgite deserve further study. METHODS: In this study, various methods such as RT-qPCR, Western blot, viral titer assay, Cytopathic Effect, immunofluorescence analysis and transmission electron microscopy were used to detect the antiviral activity of attapulgite and to assess its inhibitory effect on PEDV. RESULTS: When exposed to the same amount of virus, there was a significant decrease in the expression of the S protein, resulting in a viral titer reduction from 10-5.613 TCID50/mL to 10-2.90 TCID50/mL, which represents a decrease of approximately 102.6 folds. Results of cytopathic effect and indirect immunofluorescence also indicate a notable decrease in viral infectivity after attapulgite treatment. Additionally, it was observed that modified materials after acidification had weaker antiviral efficacy compared to powdered samples that underwent ultrasonic disintegration, which showed the strongest antiviral effects. CONCLUSION: As a result, Attapulgite powders can trap and adsorb viruses to inhibit PEDV in vitro, leading to loss of viral infectivity. This study provides new materials for the development of novel disinfectants and antiviral additives.

Characterization and Comparative Analysis of Whole-Transcriptome Sequencing in High- and Low-Fecundity Chongming White Goat Ovaries during the Estrus Phase.

Reproductive performance is one of the most important economic traits in the goat industry. Increasing the number of goats is an effective measure to improve production efficiency and reduce production costs. Ovaries are important reproductive organs in female mammals that directly affect the estrous cycle and reproductive abilities. Understanding the complex transcription network of non-coding RNAs (lncRNAs, circRNAs, and miRNAs) and messenger RNA (mRNA) could lead to significant insights into the ovarian regulation of the reproductive processes of animals. However, the whole-transcriptome analysis of the non-coding RNAs and mRNA of the ovaries in Chongming white goats between high-fecundity (HP) and low-fecundity (LP) groups is limited. In this study, a whole-transcriptome sequencing approach was used to identify lncRNA, circRNA, miRNA, and mRNA expression in the ovaries of Chongming white goats during the estrus phase using RNA-Seq technology. More than 20,000 messenger RNAs (mRNAs), 10,000 long non-coding RNAs (lncRNAs), 3500 circular RNAs (circRNAs), and 1000 micro RNAs (miRNAs) were identified. A total of 1024 differential transcripts (724 mRNAs, 112 lncRNAs, 178 circRNAs, and 10 miRNAs) existing between the HP and the LP groups were revealed through a bioinformatics analysis. They were enriched in the prolactin signaling pathway, the Jak-STAT signaling pathway, and the GnRH signaling pathway, as well as various metabolic pathways. Differentially expressed mRNAs (such as LYPD6, VEGFA, NOS3, TNXB, and EPHA2) and miRNAs (such as miR-10a-5p) play key roles in the regulation of goat ovaries during the estrus phase. The enrichment of pathways related to reproduction, such as the Hippo, Hedgehog, PI3K-AKT, and MAPK signaling pathways, suggests that they might be involved in the prolificacy of goat ovaries. Overall, we identified several gene modules associated with goat fecundity and provided a basis for a molecular mechanism in the ovaries of Chongming white goats.

PAK1-Dependent Regulation of Microtubule Organization and Spindle Migration Is Essential for the Metaphase I-Metaphase II Transition in Porcine Oocytes.

P21-activated kinase 1 (PAK1) is a critical downstream target that mediates the effect of small Rho GTPase on the regulation of cytoskeletal kinetics, cell proliferation, and cell migration. PAK1 has been identified as a crucial regulator of spindle assembly during the first meiotic division; however, its roles during the metaphase I (MI) to metaphase II (MII) transition in oocytes remain unclear. In the present study, the potential function of PAK1 in regulating microtubule organization and spindle positioning during the MI-MII transition was addressed in porcine oocytes. The results showed that activated PAK1 was co-localized with α-tubulin, and its expression was increased from the MI to MII stage (p < 0.001). However, inhibiting PAK1 activity with an inhibitor targeting PAK1 activation-3 (IPA-3) at the MI stage decreased the first polar body (PB1) extrusion rate (p < 0.05), with most oocytes arrested at the anaphase-telophase (ATI) stage. IPA-3-treated oocytes displayed a decrease in actin distribution in the plasma membrane (p < 0.001) and an increase in the rate of defects in MII spindle reassembly with abnormal spindle positioning (p < 0.001). Nevertheless, these adverse effects of IPA-3 on oocytes were reversed when the disulfide bond between PAK1 and IPA-3 was reduced by dithiothreitol (DTT). Co-immunoprecipitation revealed that PAK1 could recruit activated Aurora A and transform acidic coiled-coil 3 (TACC3) to regulate spindle assembly and interact with LIM kinase 1 (LIMK1) to facilitate actin filament-mediated spindle migration. Together, PAK1 is essential for microtubule organization and spindle migration during the MI-MII transition in porcine oocytes, which is associated with the activity of p-Aurora A, p-TACC3 and p-LIMK1.

SN2 Reaction at the Amide Nitrogen Center Enables Hydrazide Synthesis.

Nucleophilic substitutions are fundamentally important transformations in synthetic organic chemistry. Despite the substantial advances in bimolecular nucleophilic substitutions (SN2) at saturated carbon centers, analogous SN2 reaction at the amide nitrogen atom remains extremely limited. Here we report an SN2 substitution method at the amide nitrogen atom with amine nucleophiles for nitrogen-nitrogen (N-N) bond formation that leads to a novel strategy toward biologically and medicinally important hydrazide derivatives. We found the use of sulfonate-leaving groups at the amide nitrogen atom played a pivotal role in the reaction. This new N-N coupling reaction allows the use of O-tosyl hydroxamates as electrophiles and readily available amines, including acyclic aliphatic amines and saturated N-heterocycles as nucleophiles. The reaction features mild conditions, broad substrate scope (>80 examples), excellent functional group tolerability, and scalability. The method is applicable to late-stage modification of various approved drug molecules, thus enabling complex hydrazide scaffold synthesis.

Bimetallic Nanozyme: A Credible Tag for In Situ-Catalyzed Reporter Deposition in the Lateral Flow Immunoassay for Ultrasensitive Cancer Diagnosis.

The lateral flow immunoassay (LFIA) is a sought-after point-of-care testing platform, yet the insufficient sensitivity of the LFIA limits its application in the detection of tumor biomarkers. Here, a colorimetric signal amplification method, bimetallic nanozyme-mediated in situ-catalyzed reporter deposition (BN-ISCRD), was designed for ultrasensitive cancer diagnosis. The bimetallic nanozyme used, palladium@iridium core-shell nanoparticles (Pd@Ir NPs), had ultrahigh enzyme-like activity, which was further explained by the electron transfer of Pd@Ir NPs and the change in the Gibbs free energy during catalysis through density functional theory calculations. With gastric cancer biomarkers pepsinogen I and pepsinogen II as model targets, this assay could achieve a cutoff value of 10 pg/mL, which was 200-fold lower than that without signal enhancement. The assay was applied to correctly identify 8 positive and 28 negative clinical samples. Overall, this BN-ISCRD-based LFIA showed great merits and potential in the application of ultrasensitive disease diagnosis.

Tetrandrine (TET) inhibits African swine fever virus entry into cells by blocking the PI3K/Akt pathway.

African Swine Fever Virus (ASFV) infection causes an acute and highly contagious disease in swine, resulting in significant economic losses and societal harm worldwide. Currently, there are no effective vaccines or antiviral drugs available for ASFV. Tetrandrine (TET) is extracted from the traditional Chinese herb Stephania tetrandrae, possesses diverse biological functions such as anti-inflammatory, anti-tumor, and antiviral activities. The study comprehensively evaluated the anti-ASFV effect of TET and validated it through biological assays. The dose-dependent inhibition of TET against ASFV was confirmed and a novel mechanism of TET's anti-ASFV activity was elucidated. TET effectively inhibits ASFV during internalization by blocking macropinocytosis through the inhibition of the PI3K/Akt pathway. The specific inhibitor LY294002, targeting the PI3K/Akt pathway, exhibits similar antiviral activity against ASFV as TET. Furthermore, the inhibitory effect of TET against other viruses such as Lumpy Skin Disease Virus (LSDV) and Porcine Epidemic Diarrhea Virus (PEDV) was also identified. Our findings suggest that TET effectively inhibits ASFV and reveal the potential for broad-spectrum antiviral drugs targeting the PI3K/Akt pathway.

Genomic Dissection through Whole-Genome Resequencing of Five Local Pig Breeds from Shanghai, China.

China has rich genetic resources of local pig breeds. In this study, whole-genome resequencing was performed on five Shanghai local pig breeds, aiming to analyze their population genetic structure and unique genomic characteristics. Tens of millions of single nucleotide variants were obtained through the resequencing of a total of 150 individual pigs from five local pig breeds (Meishan, Fengjing, Shawutou, Pudong White, and Shanghai White) after mapping them with the pig reference genome of Sus scrofa 11.1. The results of admixture structure analysis also clearly demonstrated the genetic differences between the Shanghai local pig breeds and the three commercial pig breeds (Duroc, Landrace, and Yorkshire). The genetic infiltration of Landrace and Yorkshire pig breeds in the SHW breed was detected, which is consistent with the early history of crossbreeding in this breed. Selective sweep analysis between four indigenous Shanghai pig breed populations and three commercial pig breed populations identified 270 and 224 genes with selective signatures in the commercial and indigenous Shanghai pig populations, respectively. Six genes (TGS1, PLAG1, CHCHD7, LCORL, TMEM68, and TMEM8B) were found to be associated with animal growth in the commercial pig population through gene enrichment and protein-protein interaction analysis. In contrast, the MSRB3 gene in the indigenous Shanghai pig population was significantly under selection, which correlated with the long pendulous ear phenotype of the indigenous Shanghai pig population. In conclusion, this study is the first genomic profiling of five representative local pig breeds in Shanghai, which provides molecular genetic data and foundations for better conservation and utilization of local pig breed resources in Shanghai, China.

ProQ binding to small RNA RyfA promotes virulence and biofilm formation in avian pathogenic Escherichia coli.

Avian pathogenic Escherichia coli (APEC) is a notable subpathotype of the nonhuman extraintestinal pathogenic E. coli (ExPEC). Recognized as an extraintestinal foodborne pathogen, the zoonotic potential of APEC/ExPEC allows for cross-host transmission via APEC-contaminated poultry meat and eggs. ProQ, an RNA binding protein, is evolutionarily conserved in E. coli. However, its regulatory roles in the biofilm formation and virulence of APEC/ExPEC have not been explored. In this study, proQ deletion in the APEC strain FY26 significantly compromised its biofilm-forming ability. Furthermore, animal tests and cellular infection experiments showed that ProQ depletion significantly attenuated APEC virulence, thereby diminishing its capacity for bloodstream infection and effective adherence to and persistence within host cells. Transcriptome analysis revealed a decrease in the transcription level of the small RNA (sRNA) RyfA in the mutant FY26ΔproQ, suggesting a direct interaction between the sRNA RyfA and ProQ. This interaction might indicate that sRNA RyfA is a novel ProQ-associated sRNA. Moreover, the direct binding of ProQ to the sRNA RyfA was crucial for APEC biofilm formation, pathogenicity, adhesion, and intracellular survival. In conclusion, our findings provide detailed insight into the interaction between ProQ and sRNA RyfA and deepen our understanding of the regulatory elements that dictate APEC virulence and biofilm development. Such insights are instrumental in developing strategies to counteract APEC colonization within hosts and impede APEC biofilm establishment on food surfaces.

Photo-Triggered, Copper(II) Chloride-Catalyzed Radical Hydroalkylation and Hydrosilylation of Vinylboronic Esters To Access Alkylboronic Esters.

Alkyl boronic acids and their derivatives constitute vital building blocks in organic synthesis and are important motifs identified in medicinal chemistry. Herein, we present a phototriggered, CuCl2-catalyzed radical hydroalkylation and hydrosilylation of vinylboronic esters to alkylboronic esters. This approach exhibits mild reaction conditions, utilization of easily accessible reagents, and scalability up to a gram scale. Further synthetic transformations of the hydrosilylation products and mechanistic studies are also demonstrated.

Catalytic degradation of crystal violet and methyl orange in heterogeneous Fenton-like processes.

Metals-loaded (Fe3+, Cu2+ and Zn2+) activated carbons (M@AC) with different loading ratios (0.1%, 0.5%, 1%, 5% and 10%) were prepared and employed for catalytic degradation of dye model compounds (crystal violet (CV) and methyl orange (MO)) in wastewater by heterogeneous Fenton-like technique. Compared with Cu@AC and Zn@AC, 0.5% Fe3+ loaded AC (0.5Fe@AC) had better catalytic activity for dyes degradation. The effects of dyes initial concentration, catalyst dosage, pH and hydrogen peroxide (H2O2) volume on the catalytic degradation process were investigated. Cyclic performance, stability of 0.5Fe@AC and iron leaching were explored. Degradation kinetics were well fitted to the pseudo-second-order model (Langmuir-Hinshelwood). Almost complete decolorization (99.7%) of 400 mg L-1 CV was achieved after 30 min reaction under the conditions of CV volume (30 mL), catalyst dosage (0.05 g), H2O2 volume (1 mL) and pH (7.7). Decolorization of MO reached 98.2% under the same conditions. The abilities of pyrolysis char (PC) of dyeing sludge (DS) and metal loaded carbon to remove dye pollutants were compared. The intermediate products were analyzed and the possible degradation pathway was proposed. This study provided an insight into catalytic degradation of triphenylmethane- and aromatic azo-based substances, and utilization of sludge char.

Extracellular vesicles produced by avian pathogenic Escherichia coli (APEC) activate macrophage proinflammatory response and neutrophil extracellular trap (NET) formation through TLR4 signaling.

BACKGROUND: Avian pathogenic Escherichia coli (APEC) is the major pathogen causing important avian diseases in poultry. As an important subtype of extraintestinal pathogenic E. coli, APEC has zoonotic potential and is considered a foodborne pathogen. APEC extracellular vesicles (EVs) may play vital roles in the interaction of the pathogen with its host cells. However, the precise roles played by APEC EVs are still not completely clear, especially in immune cells. RESULTS: In this study, we investigated the relationships between APEC EVs and immune cells. The production and characteristics of the EVs of APEC isolate CT265 were identified. Toll like receptor 4 (TLR4) triggered the cellular immune responses when it interacted with APEC EVs. APEC EVs induced a significant release of proinflammatory cytokines in THP-1 macrophages. APEC EVs induced the macrophage inflammatory response via the TLR4/MYD88/NF-κB signaling pathway, which participated in the activation of the APEC-EV-induced NLRP3 inflammasome. However, the loss of lipopolysaccharide (LPS) from APEC EVs reduced the activation of the NLRP3 inflammasome mediated by TLR4/MYD88/NF-κB signaling. Because APEC EVs activated the macrophage inflammatory response and cytokines release, we speculated that the interaction between APEC EVs and macrophages activated and promoted neutrophil migration during APEC extraintestinal infection. This study is the first to report that APEC EVs induce the formation of neutrophil extracellular traps (NETs) and chicken heterophil extracellular traps. Treatment with APEC EVs induced SAPK/JNK activation in neutrophils. The inhibition of TLR4 signaling suppressed APEC-EV-induced NET formation. However, although APEC EVs activated the immune response of macrophages and initiated NET formation, they also damaged macrophages, causing their apoptosis. The loss of LPS from APEC EVs did not prevent this process. CONCLUSION: APEC-derived EVs induced inflammatory responses in macrophages and NETs in neutrophils, and that TLR4 was involved in the APEC-EV-activated inflammatory response. These findings provided a basis for the further study of APEC pathogenesis.

Phosphine-Promoted Tandem Intermolecular Diels-Alder Reactions with Pentadienyl 4-Nitrobenzoate as a Diene Precursor.

A phosphine-promoted tandem Diels-Alder reaction using pentadienyl 4-nitrobenzoate (α-vinyl MBH adduct) as a diene precursor with 3-olefinic oxindoles or CF3-activated ketones as dienophiles has been developed. The reaction proceeds through the formation of a pentadienyl phosphonium intermediate via SN2'' addition, which acts as both a D-A diene and a precursor for the exomethylene moiety. This method offers a metal-free and step-efficient approach for synthesizing exomethylene-bearing spirooxindoles and dihydropyrans, which are privileged structures found in natural products.

Epidemiological characteristics of SHV, cmlv, and FosA6-producing carbapenem-resistant Klebsiella pneumoniae based on whole genome sequences in Jiangsu, China.

Carbapenem-resistant Klebsiella pneumoniae (CRKP), particularly those with high virulence, cause invasive disease in clinical settings. An epidemiological investigation was conducted on the evolution, virulence, and antimicrobial resistance of CRKP isolates in two tertiary teaching hospitals in Jiangsu, China from November 2020 to December 2021. There were 31 different CRKP strains discovered. We performed whole genome sequencing (WGS) on 13 SHV, cmlv, and FosA6-producing CRKP to reveal molecular characteristics. Five ST15/ST11 isolates had CRISPR-Cas systems. By conjugation tests, KPC-2 can be transmitted horizontally to E. coil. A conjugative pHN7A8-related multi-resistance plasmid (KPC-2, blaCTX-M-65, blaTEM-1, fosA3, catII, and rmtB) was first discovered in CRKP clinical isolates. Using bacteriological testing, a serum killing assay, and an infection model with Galleria mellonella, three ST11-K64 KPC-2 generating carbapenem-resistant hypervirulent Klebsiella pneumoniae (CR-hvKP) were identified. These strains harbored a virulent plasmid and an IncFII-family pKPC/pHN7A8 conjugative plasmid, which led to hypervirulence and resistance. One of these CR-hvKPs, which co-harbored KPC-2, NDM-6, SHV-182, SHV-64, and blaCTX-M-122 genes, was first discovered. Importantly, this CR-hvKP strain also produced biofilm and had non-inferior fitness. The widespread use of ceftazidime/avibactam might provide this CR-hvKP with a selective advantage; hence, immediate action is required to stop its dissemination. Another important finding is the novel ST6136 in K. pneumoniae. Finally, the sterilization efficiency rates of Fe2C nanoparticles in CRKP were more than 98%. Furthermore, our novel antibacterial Fe2C nanoparticles may also provide a therapeutic strategy for infections.

A Novel Methacryloyl Chitosan Hydrogel Microneedles Patch with Sustainable Drug Release Property for Effective Treatment of Psoriasis.

Psoriasis is a chronic and recurrent skin disease that often requires long-term treatment, and topical transdermal drug delivery can reduce systemic side effects. However, it is still a challenge in efficient transdermal drug delivery for psoriasis treatment due to low penetration efficiency of most drugs and the abnormal skin conditions of psoriasis patients. Here, a safe and effective methacryloyl chitosan hydrogel microneedles (CSMA hMNs) patch is developed and served as a sustained drug release platform for the treatment of psoriasis. By systematically optimizing the CSMA preparation, CSMA hMNs with excellent morphological characteristics and strong mechanical properties (0.7 N needle-1 ) are prepared with a concentration of only 3% (w/v) CSMA. As a proof-of-concept, methotrexate (MTX) and nicotinamide (NIC) are loaded into CSMA hMNs patch, which can produce a sustained drug release of 80% within 24 h in vitro. In vivo experiments demonstrated that the CSMA hMNs patch can effectively inhibit the skin thickening and spleen enlargement of psoriatic mice and has a good biosafety profile at sufficient therapeutic doses. This study provides a new idea for the preparation of hMN systems using modified CS or other biocompatible materials and offers an effective therapeutic option for psoriasis treatment.