One-step triplex TaqMan quantitative reverse transcription polymerase chain reaction for the detection of feline coronavirus, feline panleukopenia virus, and feline leukemia virus.

Background and Aim: Feline coronavirus (FCoV), feline panleukopenia virus (FPV), and feline leukemia virus (FeLV) are prevalent throughout China and significantly threaten cat health. These viruses cause similar manifestations and pathological damage. Rapid and accurate diagnosis depends on detection in the laboratory. This study aimed to establish a reliable and rapid method for accurate detection of FCoV, FPV, and FeLV so that a definite diagnosis can be made and effective measures can be taken to prevent and control viral infection. Materials and Methods: We designed three pairs of specific primers and probes for the detection of FCoV 5' untranslated region, FPV viral protein 2, and FeLV pol genes. Recombinant plasmid constructs were generated for use as standard plasmid constructs. Optimal reaction conditions, including primer and probe concentrations, reaction cycles, and annealing temperatures, were obtained on the basis of optimization tests. One-step triplex real-time reverse transcription-quantitative polymerase chain reaction (RT-qPCR) was successfully established to simultaneously detect FCoV, FPV, and FeLV. The specificity, sensitivity, and repeatability of the assay were analyzed, and its applicability was validated by testing 1175 clinical samples. Results: One-step triplex RT-qPCR had a high degree of specificity only for the detection of FCoV, FPV, and FeLV; it had high sensitivity with limits of detection of 139.904, 143.099, and 152.079 copies/reaction for p-FCoV, p-FPV, and p-FeLV standard plasmid constructs, respectively, and it had reliable repeatability with 0.06%-0.87% intra-assay coefficients of variations. A total of 1175 clinical samples were examined for FCoV, FPV, and FeLV using triplex RT-qPCR, and the FCoV, FPV, and FeLV positivity rates were 18.47%, 19.91%, and 47.57%, respectively. The clinical sensitivity and specificity of one-step triplex RT-qPCR were 93.07% and 97.99%, respectively. Conclusion: We developed a rapid and reliable one-step triplex RT-qPCR method for the detection of FCoV, FPV, and FeLV, which could be used as a diagnostic tool for clinical monitoring and diagnosis.

Design and evaluation of mRNA encoding recombinant neutralizing antibodies for botulinum neurotoxin type B intoxication prophylaxis.

Among all natural and synthetic toxins, botulinum neurotoxins (BoNTs), produced by Clostridium botulinum in an anaerobic environment, are the most toxic polymer proteins. Currently, the most effective modalities for botulism prevention and treatment are vaccination and antitoxin use, respectively. However, these modalities are associated with long response time for active immunization, side effects, and donor limitations. As such, the development of more promising botulism prevention and treatment modalities is warranted. Here, we designed an mRNA encoding B9-hFc - a heavy-chain antibody fused to VHH and human Fc that can neutralize BoNT serotype B (BoNT/B) effectively - and assessed its expression in vitro and in vivo. The results confirmed that our mRNA demonstrates good expression in vitro and in vivo. Moreover, a single mRNA lipid nanoparticle injection effectively prevents BoNT/B intoxication in vivo, with effects comparable to those of protein antibodies. In conclusion, we explored and clarified whether mRNA drugs encoding neutralizing antibodies prevent BoNT/B intoxication. Our results provide an efficient strategy for further research on the prevention and treatment of intoxication by botulinum toxin.

A multifunctional evanescent wave biosensor for the universal assay of SARS-CoV-2 variants and affinity analysis of coronavirus spike protein-hACE2 interactions.

The conventional detection model of passive adaptation to pathogen mutations, i.e., developing assays using corresponding antibodies or nucleic acid probes, is difficult to address frequent outbreaks of emerging infectious diseases. In particular, adaptive mutations observed in coronaviruses, which increase the affinity of the spike protein with the human cellular receptor hACE2, play pivotal roles in the transmission and immune evasion of coronaviruses. Herein, we developed a multifunctional optical fiber evanescent wave biosensor for the universal assay of coronavirus and affinity analysis of the spike protein interacting with hACE2, namely, My-SPACE. By competitively binding with Cy5.5-hACE2 between coronavirus spike proteins in mobile buffer and that modified on optical fibers from the SARS-CoV-2 wild type, My-SPACE could automatically detect SARS-CoV-2 and its variants within 10 min. My-SPACE demonstrated greater sensitivity and faster results than ELISA for SARS-CoV-2 variants, achieving 100% specificity and 94.10% sensitivity in detecting the Omicron variant in 18 clinical samples. Moreover, the interaction between hACE2 and the coronavirus spike protein was accurately characterized across SARS-CoV-2 mutants, SARS-CoV and hCoV-NL63. The accuracy of the affinity determined by My-SPACE was verified by SPR. This approach enables preliminary assessment of the transmissibility and hazards of emerging coronaviruses. The sensor fibers of My-SPACE can be reused more than 40 times, and the device is compact and easy to use; moreover, it is available as a rapid and cost-effective on-site detection tool adapted to coronavirus variability and as an effective assessment platform for early warning of coronavirus transmission risk.

Fish ubiquitin-specific protease 8 (USP8) inhibits IFN production through autophagy-lysosomal dependent degradation of IRF7.

Interferon regulatory factor 7 (IRF7) is considered the master regulator of virus-induced interferon (IFN) production. However, to avoid an autoimmune response, the expression of IRF7 must be tightly controlled. In this study, we report that zebrafish ubiquitin-specific protease 8 (USP8) promotes IRF7 degradation through an autophagy-lysosome-dependent pathway to inhibit IFN production. First, zebrafish usp8 is induced upon spring viremia of carp virus (SVCV) infection and polyinosinic/polycytidylic acid (poly I:C) stimulation. Second, overexpression of USP8 suppresses SVCV or poly I:C-mediated IFN expression. Mechanistically, USP8 interacts with IRF7 and promotes its degradation via an autophagy-lysosome-dependent pathway. Finally, USP8 significantly suppresses cellular antiviral responses and enhances SVCV proliferation. In summary, our discoveries offer a perspective on the role of zebrafish USP8 and provide additional understanding of the regulation of IRF7 in host antiviral immune response.

Rapid and sensitive parallel on-site detection of antibiotics and resistance genes in aquatic environments using evanescent wave dual-color fluorescence fiber-embedded optofluidic nanochip.

Environmental antibiotics and antibiotic resistance genes (ARGs) pose considerable threat to humans and animals; thus, the rapid and sensitive parallel detection of these pollutants from a single sample is urgently required. However, traditional multiplexed analytic technologies detect only one type of target (e.g., small molecules or nucleic acids) per assay. To address this issue, Evanescent wave Dual-color fluorescence Fiber-embedded Optofluidic Nanochip (EDFON) was fabricated by integrating a fiber-embedded optofluidic nanochip with evanescent wave dual-color fluorescence technology. The EDFON was used for the parallel quantitative detection of sulfamerazine (SMR) and MCR-1 with high sensitivity and specificity by combining a heterogeneous immunoassay with a homogenous hybridization chain reaction based on time-resolved effects. LODs of 0.032 µg/L and 35 pM was obtained for SMR and MCR-1, respectively, within 20 min. To our best knowledge, the EDFON is the first device for the simultaneous detection of two type of targets in each test, which is highly valuable to prevent the global threats of antibiotics and ARGs. Comparison with liquid chromatography-mass spectrometry showed a strong linear relationship (R2 = 0.998) for SMR pollution in the Qinghe River, with spiked SMR and MCR-1 negative surface and wastewater samples showing recovery rates of 91.8-113.4%. These results demonstrate the excellent accuracy and reliability of the EDFON, with features such as multi-analyte detection, field-deployment, and minimal-equipment, rendering it revolutionary for environmental monitoring, food safety, and medical diagnostics.

Associations between life's essential 8 and sarcopenia in US adults: a cross-sectional analysis.

Cardiovascular disease (CVD) is closely associated with sarcopenia. We aimed to examine the relationship between Life's Essential 8 (LE8) and the incidence of sarcopenia among adults in the United States. In this study, a cross-sectional analysis was conducted using data from the National Health and Nutrition Examination Survey from 2013 to 2018 and included 5999 adult participants. LE8 score was categorized into low (< 49), moderate (49-79), and high CVH (≥ 79) groups and consisted of health behavior score and health factor score based on American Heart Association definitions. Sarcopenia was defined according to The Foundation for the National Institutes of Health Sarcopenia Project. Multivariate logistic regressions, restricted cubic spline regressions, and subgroup analyses were used to assess the association between LE8 and sarcopenia. LE8 and its subscales score were negatively associated with the incidence of sarcopenia in US adults.

A triplex crystal digital PCR for the detection of genotypes I and II African swine fever virus.

African swine fever (ASF) is a highly contagious and lethal viral disease that causes severe hemorrhagic fever in pigs. It keeps spreading around the world, posing a severe socioeconomic risk and endangering biodiversity and domestic food security. ASF first outbroke in China in 2018, and has spread to most provinces nationwide. Genotypes I and II ASF virus (ASFV) as the etiological pathogens have been found in China. In this study, three pairs of specific primers and probes targeting the ASFV B646L gene, F1055L gene, and E183L gene were designed to detect universal, genotype I, and genotype II strains, respectively. A triplex crystal digital PCR (cdPCR) was established on the basis of optimizing various reaction conditions. The assay demonstrated remarkably sensitive with low limits of detection (LODs) of 5.120, 4.218, 4.588 copies/reaction for B646L, F1055L, and E183L gene, respectively; excellent repeatability with 1.24-2.01% intra-assay coefficients of variation (CVs) and 1.32-2.53% inter-assay CVs; good specificity for only detection of genotypes I and II ASFV, without cross-reactivity with PCV2, PRV, SIV, PRRSV, PEDV, FMDV, and CSFV. The triplex cdPCR was used to test 1,275 clinical samples from Guangxi province of China, and the positivity rates were 5.05, 3.22, and 1.02% for genotype I, genotype II, and co-infection of genotypes I and II, respectively. These 1,275 clinical samples were also detected using a reported reference triplex real-time quantitative PCR (qPCR), and the agreements of detection results between these two methods were more than 98.98%. In conclusion, the developed triplex cdPCR could be used as a rapid, sensitive, and accurate method to detect and differentiate genotypes I and II strains of ASFV.

Structural insights into the Clp protein degradation machinery.

The Clp protease system is important for maintaining proteostasis in bacteria. It consists of ClpP serine proteases and an AAA+ Clp-ATPase such as ClpC1. The hexameric ATPase ClpC1 utilizes the energy of ATP binding and hydrolysis to engage, unfold, and translocate substrates into the proteolytic chamber of homo- or hetero-tetradecameric ClpP for degradation. The assembly between the hetero-tetradecameric ClpP1P2 chamber and the Clp-ATPases containing tandem ATPase domains from the same species has not been studied in depth. Here, we present cryo-EM structures of the substrate-bound ClpC1:shClpP1P2 from Streptomyces hawaiiensis, and shClpP1P2 in complex with ADEP1, a natural compound produced by S. hawaiiensis and known to cause over-activation and dysregulation of the ClpP proteolytic core chamber. Our structures provide detailed information on the shClpP1-shClpP2, shClpP2-ClpC1, and ADEP1-shClpP1/P2 interactions, reveal conformational transition of ClpC1 during the substrate translocation, and capture a rotational ATP hydrolysis mechanism likely dominated by the D1 ATPase activity of chaperones.IMPORTANCEThe Clp-dependent proteolysis plays an important role in bacterial homeostasis and pathogenesis. The ClpP protease system is an effective drug target for antibacterial therapy. Streptomyces hawaiiensis can produce a class of potent acyldepsipeptide antibiotics such as ADEP1, which could affect the ClpP protease activity. Although S. hawaiiensis hosts one of the most intricate ClpP systems in nature, very little was known about its Clp protease mechanism and the impact of ADEP molecules on ClpP. The significance of our research is in dissecting the functional mechanism of the assembled Clp degradation machinery, as well as the interaction between ADEP1 and the ClpP proteolytic chamber, by solving high-resolution structures of the substrate-bound Clp system in S. hawaiiensis. The findings shed light on our understanding of the Clp-dependent proteolysis in bacteria, which will enhance the development of antimicrobial drugs targeting the Clp protease system, and help fighting against bacterial multidrug resistance.

CRISPR/Cas12a powered air-displacement enhanced evanescent wave fluorescence fiber-embedded microfluidic biochip for nucleic acid amplification-free detection of Escherichia coli O157:H7.

Simple yet ultrasensitive and contamination-free quantification of environmental pathogenic bacteria is in high demand. In this study, we present a portable clustered regularly interspaced short palindromic repeats-associated protein 12a (CRISPR/Cas12a) powered Air-displacement enhanced Evanescent wave fluorescence Fiber-embedded microfluidic Biochip (AEFB) for the high-frequency and nucleic acid amplification-free ultrasensitive detection of Escherichia coli O157:H7. The performance of AEFB was dramatically enhanced upon employing a simple air-solution displacement process. Theoretical assays demonstrated that air-solution displacement significantly enhances evanescent wave field intensity on the fiber biosensor surface and increases the V-number in tapered fiber biosensors. Consequently, light-matter interaction is strengthened, and fluorescence coupling and collection efficiency are improved, considerably enhancing sensitivity. By integrating the CRISPR biosensing mechanism, AEFB facilitated rapid, accurate, nucleic acid amplification-free detection of E.coli O157:H7 with polymerase chain reaction (PCR)-level sensitivity (176 cfu/mL). To validate its practicality, AEFB was used to detect E.coli O157:H7 in surface water and wastewater. Comparison with RT-PCR showed a strong linear relationship (R2 = 0.9871), indicating the excellent accuracy and reliability of this technology in real applications. AEFB is highly versatile and can be easily extended to detect other pathogenic bacteria, which will significantly promote the high-frequency assessment and early-warning of bacterial contamination in aquatic environments.

Bifunctional MOF-Encapsulated Cobalt-Doped Carbon Dots Nanozyme-Powered Chemiluminescence/Fluorescence Dual-Mode Detection of Aflatoxin B1.

A novel bifunctional MOF-encapsulated cobalt-doped carbon dots nanozyme (Co-CD/PMOF) with excellent peroxidase-mimic catalytic activity and fluorescence property was synthesized and employed to fabricate a chemiluminescence/fluorescence (CL/FL) dual-mode immunosensor for AFB1 detection. Co-CD/PMOF could catalyze the luminol/H2O2 system to generate robust and long-lasting CL signals due to the slow diffusion effect and continuous generation of •OH, O2•-, and 1O2 species. Differing from traditional flash-type CL emissions, this glow-type CL emission is helpful to fabricate a sensitive and accurate CL sensing platform. Then the CL/FL dual-mode detection of AFB1 was developed using antibody-functionalized Co-CD/PMOF as the signal-amplifying nanoprobe. The CL mode assay based on indirect competitive immune principle was carried out on a chemiluminescence optical fiber platform, where the AFB1-OVA-functionalized optical fiber probe was employed for biorecognition, separation, and signal conducting. The AFB1 detection range and LOD were 0.63-69.36 ng/mL and 0.217 ng/mL, respectively. Using AFB1 antibody-functionalized immunomagnetic beads for capturing and separation, the FL mode detection of AFB1 was established based on the sandwich immune principle. A linear range of 0.54-51.91 ng/mL and a LOD of 0.027 ng/mL were obtained. This work designed a sensitive, rapid, and reliable nanozyme-powered dual-mode assay strategy and provided technical support in the field of environmental monitoring and food safety.

The role of lncRNA HCG18 in human diseases.

A substantial number of long noncoding RNAs (lncRNAs) have been identified as potent regulators of human disease. Human leukocyte antigen complex group 18 (HCG18) is a new type of lncRNA that has recently been proven to play an important role in the occurrence and development of various diseases. Studies have found that abnormal expression of HCG18 is closely related to the clinicopathological characteristics of many diseases. More importantly, HCG18 was also found to promote disease progression by affecting a series of cell biological processes. This article mainly discusses the expression characteristics, clinical characteristics, biological effects and related regulatory mechanisms of HCG18 in different human diseases, providing a scientific theoretical basis for its early clinical application.

Genetic and Evolutionary Analysis of Porcine Deltacoronavirus in Guangxi Province, Southern China, from 2020 to 2023.

Porcine deltacoronavirus (PDCoV) has shown large-scale global spread since its discovery in Hong Kong in 2012. In this study, a total of 4897 diarrheal fecal samples were collected from the Guangxi province of China from 2020 to 2023 and tested using RT-qPCR. In total, 362 (362/4897, 7.39%) of samples were positive for PDCoV. The S, M, and N gene sequences were obtained from 34 positive samples after amplification and sequencing. These PDCoV gene sequences, together with other PDCoV S gene reference sequences from China and other countries, were analyzed. Phylogenetic analysis revealed that the Chinese PDCoV strains have diverged in recent years. Bayesian analysis revealed that the new China 1.3 lineage began to diverge in 2012. Comparing the amino acids of the China 1.3 lineage with those of other lineages, the China 1.3 lineage showed variations of mutations, deletions, and insertions, and some variations demonstrated the same as or similar to those of the China 1.2 lineage. In addition, recombination analysis revealed interlineage recombination in CHGX-MT505459-2019 and CHGX-MT505449-2017 strains from Guangxi province. In summary, the results provide new information on the prevalence and evolution of PDCoV in Guangxi province in southern China, which will facilitate better comprehension and prevention of PDCoV.

Substrate binding and catalytic mechanism of the Se-glycosyltransferase SenB in the biosynthesis of selenoneine.

Selenium is an essential multifunctional trace element in diverse organisms. The only Se-glycosyltransferase identified that catalyzes the incorporation of selenium in selenoneine biosynthesis is SenB from Variovorax paradoxus. Although the biochemical function of SenB has been investigated, its substrate specificity, structure, and catalytic mechanism have not been elucidated. Here, we reveal that SenB exhibits sugar donor promiscuity and can utilize six UDP-sugars to generate selenosugars. We report crystal structures of SenB complexed with different UDP-sugars. The key elements N20/T23/E231 contribute to the sugar donor selectivity of SenB. A proposed catalytic mechanism is tested by structure-guided mutagenesis, revealing that SenB yields selenosugars by forming C-Se glycosidic bonds via spontaneous deprotonation and disrupting Se-P bonds by nucleophilic water attack, which is initiated by the critical residue K158. Furthermore, we functionally and structurally characterize two other Se-glycosyltransferases, CbSenB from Comamonadaceae bacterium and RsSenB from Ramlibacter sp., which also exhibit sugar donor promiscuity.

Structural basis of antiphage immunity generated by a prokaryotic Argonaute-associated SPARSA system.

Argonaute (Ago) proteins are ubiquitous across all kingdoms of life. Eukaryotic Agos (eAgos) use small RNAs to recognize transcripts for RNA silencing in eukaryotes. In contrast, the functions of prokaryotic counterparts (pAgo) are less well known. Recently, short pAgos in conjunction with the associated TIR or Sir2 (SPARTA or SPARSA) were found to serve as antiviral systems to combat phage infections. Herein, we present the cryo-EM structures of nicotinamide adenine dinucleotide (NAD+)-bound SPARSA with and without nucleic acids at resolutions of 3.1 Å and 3.6 Å, respectively. Our results reveal that the APAZ (Analogue of PAZ) domain and the short pAgo form a featured architecture similar to the long pAgo to accommodate nucleic acids. We further identified the key residues for NAD+ binding and elucidated the structural basis for guide RNA and target DNA recognition. Using structural comparisons, molecular dynamics simulations, and biochemical experiments, we proposed a putative mechanism for NAD+ hydrolysis in which an H186 loop mediates nucleophilic attack by catalytic water molecules. Overall, our study provides mechanistic insight into the antiphage role of the SPARSA system.

Orange-derived extracellular vesicles nanodrugs for efficient treatment of ovarian cancer assisted by transcytosis effect.

Extracellular vesicles (EVs) have recently received much attention about the application of drug carriers due to their desirable properties such as nano-size, biocompatibility, and high stability. Herein, we demonstrate orange-derived extracellular vesicles (OEV) nanodrugs (DN@OEV) by modifying cRGD-targeted doxorubicin (DOX) nanoparticles (DN) onto the surface of OEV, enabling significantly enhancing tumor accumulation and penetration, thereby efficiently inhibiting the growth of ovarian cancer. The obtained DN@OEV enabled to inducement of greater transcytosis capability in ovarian cancer cells, which presented the average above 10-fold transcytosis effect compared with individual DN. It was found that DN@OEV could trigger receptor-mediated endocytosis to promote early endosome/recycling endosomes pathway for exocytosis and simultaneously reduce degradation in the early endosomes-late endosomes-lysosome pathway, thereby inducing the enhanced transcytosis. In particular, the zombie mouse model bearing orthotopic ovarian cancer further validated DN@OEV presented high accumulation and penetration in tumor tissue by the transcytosis process. Our study indicated the strategy in enhancing transcytosis has significant implications for improving the therapeutic efficacy of the drug delivery system.

Erratum: Efficacy of Shikonin against Esophageal Cancer Cells and its possible mechanisms in vitro and in vivo: Erratum.

[This corrects the article DOI: 10.7150/jca.21224.].

Development of a triplex real-time quantitative PCR for detection and differentiation of genotypes I and II African swine fever virus.

African swine fever virus (ASFV) was first identified in 1921 and is extensively prevalent around the world nowadays, which has a significant negative impact on the swine industry. In China, genotype II ASFV was first discovered in 2018, and has spread quickly to different provinces in a very short time; genotype I ASFV was first found in 2020, and has been reported in several provinces since then. To establish an accurate method for detection and differentiation of genotypes I and II ASFV, three primers and probes were designed targeting the ASFV B646L gene for different genotypes, the F1055L gene for genotype I, and the E183L gene for genotype II, and a triplex real-time quantitative PCR (qPCR) for differential detection of genotypes I and II ASFV was developed after optimizing the reaction conditions. The assay showed high sensitivity, and the limits of detection (LOD) of the B646L, F1055L, and E183L genes were 399.647 copies/reaction, 374.409 copies/reaction, and 355.083 copies/reaction, respectively; the coefficients of variation (CVs) of the intra-assay and the inter-assay were 0.22-1.88% and 0.16-1.68%, respectively, showing that this method had good repeatability; the assay could detect only ASFV, without cross-reactivity with other swine viruses including PRRSV, PEDV, PDCoV, CSFV, PRV, and PCV2, showing excellent specificity of this method. A total of 3,519 clinical samples from Guangxi province, southern China, were tested by the developed assay, and 8.16% (287/3,519) samples were found to be positive for ASFV, of which 0.17% (6/3,519) samples were positive for genotype I, 7.19% (253/3,519) samples for genotype II, and 0.80% (28/3,519) samples for genotypes I and II. At the same time, these clinical samples were also tested by a previously reported multiplex qPCR, and the agreement between these two methods was more than 99.94%. In summary, the developed triplex qPCR provided a fast, specific and accurate method for detection and differentiation of genotypes I and II ASFV.

A reusable fiber-embedded microfluidic chip for rapid and sensitive on-site detection of kanamycin residues in water environments.

The overuse and abuse of antibiotics have led to increased pollution in water environments. Thus, it is crucial to develop a rapid, high-frequency, and cost-effective method for on-site detection of antibiotics. In this regard, a reusable fiber-embedded microfluidic chip was constructed by combining a microfluidic chip with a functionalized fiber bioprobe that served as both a biorecognition element and an optical transducer. The fiber-embedded microfluidic chip enabled the quantitative detection of kanamycin (KANA) by integrating a portable all-fiber evanescent wave fluorescence detection device. Under optimized conditions, quantitative KANA detection was achieved with a detection limit of 0.03 µg L-1 and a linear detection range of 0.21-10.3 µg L-1. The accurate detection of KANA in various water samples can be completed within 25 min without pretreatment. The functionalized fiber-embedded microfluidic chip could be reused more than 200 times without significant performance loss. To demonstrate its suitability for practical applications, the fiber-embedded microfluidic chip was used to investigate KANA residues in surface waters obtained from the Qinghe River in Beijing, China. The results were compared with those of a traditional enzyme-linked immunosorbent assay, which showed a high correlation. Compared to conventional optical microfluidic chips, the proposed fiber-embedded microfluidic chip has several advantages, including its ease of use, miniaturization, cost-effectiveness, reusability, and high flexibility. It is an ideal alternative for rapid, sensitive on-site detection of antibiotics and other trace substances in environmental, food, and medical fields.

Dynamic supramolecular hydrogels mediated by chemical reactions.

Supramolecular self-assembly in a biological system is usually dominated by sophisticated metabolic processes (chemical reactions) such as catalysis of enzymes and consumption of high energy chemicals, leading to groups of biomolecules with unique dynamics and functions in an aqueous environment. In recent years, increasing efforts have been made to couple chemical reactions to molecular self-assembly, with the aim of creating supramolecular materials with lifelike properties and functions. In this feature article, after summarising the work of chemical reaction mediated supramolecular hydrogels, we first focus on a typical example where dynamic self-assembly of molecular hydrogels is activated by in situ formation of a hydrazone bond in water. We discuss how the formation of the hydrazone-based supramolecular hydrogels can be controlled in time and space. After that, we describe transient assembly of supramolecular hydrogels powered by out-of-equilibrium chemical reaction networks regulated by chemical fuels, which show unique properties such as finite lifetime, dynamic structures, and regenerative capabilities. Finally, we provide a perspective on the future investigations that need to be done urgently, which range from fundamental research to real-life applications of dynamic supramolecular hydrogels.

The Septin Gene StSep4 Contributes to the Pathogenicity of Setosphaeria turcica by Regulating the Morphology, Cell Wall Integrity, and Pathogenic Factor Biosynthesis.

Septins are a conserved group of GTP-binding proteins found in all eukaryotes and are the fourth-most abundant cytoskeletal proteins. Septins of some pathogenic fungi are involved in morphological changes related to infection. Our previous studies have identified four core septins (StSep1-4) in Setosphaeria turcica, the causal agent of northern corn leaf blight, while only StSep4 is significantly upregulated during the invasive process. We therefore used forchlorfenuron (FCF), the specific inhibitor of septin, and ΔStSep4 knockout mutants to further clarify the role of septins in S. turcica pathogenicity. FCF treatment caused a dose-dependent reduction in S. turcica colony growth, delayed the formation of infection structures, and reduced the penetration ability. ΔStSep4 knockout mutants displayed abnormal mycelium morphology, slow mycelial growth, conidiation deficiency, delayed appressorium development, and weakened pathogenicity. StSep4 deletion also broke cell wall integrity, altered chitin distribution, decreased the melanin content, and disrupted normal nuclear localization. A transcriptomic comparison revealed that genes differentially expressed between ΔStSep4 and WT were enriched in terms of ribosomes, protein translation, membrane components, and transmembrane transport activities. Our results demonstrate that StSep4 is required for morphology and pathogenicity in S. turcica, making it a promising target for the development of novel fungicides.