Overexpression of ZmDHN15 Enhances Cold Tolerance in Yeast and Arabidopsis.

Maize (Zea mays L.) originates from the subtropical region and is a warm-loving crop affected by low-temperature stress. Dehydrin (DHN) protein, a member of the Group 2 LEA (late embryogenesis abundant proteins) family, plays an important role in plant abiotic stress. In this study, five maize DHN genes were screened based on the previous transcriptome sequencing data in our laboratory, and we performed sequence analysis and promoter analysis on these five DHN genes. The results showed that the promoter region has many cis-acting elements related to cold stress. The significantly upregulated ZmDHN15 gene has been further screened by expression pattern analysis. The subcellular localization results show that ZmDHN15 fusion protein is localized in the cytoplasm. To verify the role of ZmDHN15 in cold stress, we overexpressed ZmDHN15 in yeast and Arabidopsis. We found that the expression of ZmDHN15 can significantly improve the cold resistance of yeast. Under cold stress, ZmDHN15-overexpressing Arabidopsis showed lower MDA content, lower relative electrolyte leakage, and less ROS (reactive oxygen species) when compared to wild-type plants, as well as higher seed germination rate, seedling survival rate, and chlorophyll content. Furthermore, analysis of the expression patterns of ROS-associated marker genes and cold-response-related genes indicated that ZmDHN15 genes play an important role in the expression of these genes. In conclusion, the overexpression of the ZmDHN15 gene can effectively improve the tolerance to cold stress in yeast and Arabidopsis. This study is important for maize germplasm innovation and the genetic improvement of crops.

A Novel Senescence-Specific Gene (ZmSAG39) Negatively Regulates Darkness and Drought Responses in Maize.

The papain-like cysteine proteases (PLCPs) is a subfamily of cysteine proteases that plays an important role in leaf senescence, and some of its members are involved in the regulation of plant growth and development under stress. In this study, we cloned a new gene, ZmSAG39, from maize. Expression profile analysis showed that ZmSAG39 was induced by darkness and drought treatments. In addition, the ZmSAG39 overexpression in maize accelerated the senescence of maize leaves under darkness and drought treatments. However, the knockout of ZmSAG39 in maize enhanced the resistance of maize to darkness and drought stresses and reduced the degree of senescence of maize leaves. Under drought stress, compared with WT plants, the knockout lines had a higher seed germination rate, seedling survival rate and chlorophyll content, and lower reactive oxygen species (ROS) level and malondialdehyde (MDA) content. In addition, quantitative real-time PCR (qRT-PCR) analysis showed that ZmSAG39 negatively regulated some stress-related genes but positively regulated senescence-related genes under darkness and drought stress conditions. To summarize, these results indicate that ZmSAG39 is a senescence-related gene and plays a negative role in response to darkness and drought stresses. This study laid a theoretical foundation for the innovation of maize germplasm resources with high quality, high yield and strong stress resistance.

[Detect 15 pesticides fungicide in edible fungi by multiplug filtration clean-up pretreatment technology with ultra-high performance liquid chromatography-tandem mass spectrometry].

OBJECTIVE: To establish a method for determination of 15 pesticides residue in edible fungi by multiplug filtration clean-up(m-PFC) pretreatment technique with ultra-high performance liquid chromatography-tandem triple quadruple mass spectrometry(UPLC-MS/MS). METHODS: The interferences of edible fungus samples were removed by extracting with acetonitrile and filtration type extraction column, which were fat, carbohydrates, water-soluble vitamins. Samples were separated with column of Waters ACQUITY UPLC®HSS T3(2.1 mm×100 mm, 1.8 µm), and were scanned by multiple reaction monitoring mode(MRM). Samples were quantified with matrix matching standard curve external standard method. RESULTS: The recoveries of 15 target compounds at the spiked levels of 10, 20, 50 µg/kg were 82.5%-118.5%, and the relative standard deviations were between 6.1% and 23.1%(n=6). The detection limit of 15 target compounds was 1-3 µg/kg, and the limit of quantification was 3-10 µg/kg. CONCLUSION: This method improves the efficiency of pretreatment, has good stability and high sensitivity, and could be used for the detection of 15 pesticides in edible fungi.

Basal-Level Effects of (p)ppGpp in the Absence of Branched-Chain Amino Acids in Actinobacillus pleuropneumoniae.

The (p)ppGpp-mediated stringent response (SR) is a highly conserved regulatory mechanism in bacterial pathogens, enabling adaptation to adverse environments, and is linked to pathogenesis. Actinobacillus pleuropneumoniae can cause damage to the lungs of pigs, its only known natural host. Pig lungs are known to have a low concentration of free branched-chain amino acids (BCAAs) compared to the level in plasma. We had investigated the role for (p)ppGpp in viability and biofilm formation of A. pleuropneumoniae Now, we sought to determine whether (p)ppGpp was a trigger signal for the SR in A. pleuropneumoniae in the absence of BCAAs. Combining transcriptome and phenotypic analyses of the wild type (WT) and an relA spoT double mutant [which does not produce (p)ppGpp], we found that (p)ppGpp could repress de novo purine biosynthesis and activate antioxidant pathways. There was a positive correlation between GTP and endogenous hydrogen peroxide content. Furthermore, the growth, viability, morphology, and virulence were altered by the inability to produce (p)ppGpp. Genes involved in the biosynthesis of BCAAs were constitutively upregulated, regardless of the existence of BCAAs, without accumulation of (p)ppGpp beyond a basal level. Collectively, our study shows that the absence of BCAAs was not a sufficient signal to trigger the SR in A. pleuropneumoniae (p)ppGpp-mediated regulation in A. pleuropneumoniae is different from that described for the model organism Escherichia coli Further work will establish whether the (p)ppGpp-dependent SR mechanism in A. pleuropneumoniae is conserved among other veterinary pathogens, especially those in the Pasteurellaceae family.IMPORTANCE (p)ppGpp is a key player in reprogramming transcriptomes to respond to nutritional challenges. Here, we present transcriptional and phenotypic differences of A. pleuropneumoniae grown in different chemically defined media in the absence of (p)ppGpp. We show that the deprivation of branched-chain amino acids (BCAAs) does not elicit a change in the basal-level (p)ppGpp, but this level is sufficient to regulate the expression of BCAA biosynthesis. The mechanism found in A. pleuropneumoniae is different from that of the model organism Escherichia coli but similar to that found in some Gram-positive bacteria. This study not only broadens the research scope of (p)ppGpp but also further validates the complexity and multiplicity of (p)ppGpp regulation in microorganisms that occupy different biological niches.

Self-Illuminating Photodynamic Therapy with Enhanced Therapeutic Effect by Optimization of the Chemiluminescence Resonance Energy Transfer Step to the Photosensitizer.

The major obstacles to the wider application of photodynamic therapy (PDT) are drawbacks of the current photosensitizers and the tissue penetration limit of the common outer light source. In the present study, the chemiluminescence (CL) from the luminol-H2O2-horseradish peroxidase reaction was explored as a potential inner light source for the intracellular activation of carbon dots (CDs)-based PDT system. To fully use the light and enhance the overall PDT yield, the nanocarrier of CDs, the light of CL, and the PDT agent chlorin e6 (Ce6) were carefully selected and designed to form an efficient and united system. Bright-yellow-emissive CDs (y-CDs) were synthesized through purposeful regulation of the absorption and emission spectra to enhance the overlapping areas in the chemiluminescence resonance energy transfer (CRET) and fluorescence resonance energy transfer (FRET) processes. Our results reflected CL-induced y-CDs-Ce6 system (10 µM) successfully generated reactive oxygen species (ROS, 35.93%), killed ∼90% SMMC-7721 cells in vitro, and significantly delayed tumor growth in vivo. On the basis of immunohistochemical observations of proliferating cell nuclear antigen (PCNA) and platelet/endothelial cell adhesion molecule-1 (PECAM-1 or CD31) results, we concluded that the CL-induced y-CDs-Ce6 system had excellent performance in cancer therapy. The enhanced therapeutic effect was ascribed to two pathways: a direct CRET process and another process of CRET with subsequent y-CD-mediated FRET (CRET-to-FRET).

Attenuated Actinobacillus pleuropneumoniae double-deletion mutant S-8∆clpP/apxIIC confers protection against homologous or heterologous strain challenge.

BACKGROUND: Actinobacillus pleuropneumoniae is the etiological agent of porcine pleuropneumonia, which leads to large economic losses to the swine industry worldwide. In this study, S-8△clpP△apxIIC, a double-deletion mutant of A. pleuropneumoniae was constructed, and its safety and protective efficacy were evaluated in pigs. RESULTS: The S-8△clpP△apxIIC mutant exhibited attenuated virulence in a murine (BALB/c) model, and caused no detrimental effects on pigs even at a dose of up to 1.0 × 109 CFU. Furthermore, the S-8△clpP△apxIIC mutant was able to induce a strong immune response in pigs, which included high levels of IgG1 and IgG2, stimulated gamma interferon (IFN-γ), interleukin 12 (IL-12), and interleukin 4 (IL-4) production, and conferred effective protection against the lethal challenge with A. pleuropneumoniae serovars 7 or 5a. The pigs in the S-8△clpP△apxIIC immunized groups have no lesions and reduced bacterial loads in the lung tissue after challenge. CONCLUSIONS: The data obtained in this study suggest that the S-8△clpP△apxIIC mutant can serve as a highly immunogenic and potential live attenuated vaccine candidate against A. pleuropneumoniae infection.

The Lon protease homologue LonA, not LonC, contributes to the stress tolerance and biofilm formation of Actinobacillus pleuropneumoniae.

Lon proteases are a family of ATP-dependent proteases that are involved in the degradation of abnormal proteins in bacteria exposed to adverse environmental stress. An analysis of the genome sequence of Actinobacillus pleuropneumoniae revealed the unusual presence of two putative ATP-dependent Lon homologues, LonA and LonC. Sequence comparisons indicated that LonA has the classical domain organization of the LonA subfamily, which includes the N-terminal domain, central ATPase (AAA) domain, and C-terminal proteolytic (P) domain. LonC belongs to the recently classified LonC subfamily, which includes Lon proteases that contain neither the N-terminal domain of LonA nor the transmembrane region that is present only in LonB subfamily members. To investigate the roles of LonA and LonC in A. pleuropneumoniae, mutants with deletions in the lonA and lonC genes were constructed. The impaired growth of the △lonA mutant exposed to low and high temperatures and osmotic and oxidative stress conditions indicates that the LonA protease is required for the stress tolerance of A. pleuropneumoniae. Furthermore, the △lonA mutant exhibited significantly reduced biofilm formation compared to the wild-type strain. However, no significant differences in stress responses or biofilm formation were observed between the △lonC mutant and the wild-type strain. The △lonA mutant exhibited reduced colonization ability and attenuated virulence of A. pleuropneumoniae in the BALB/c mouse model compared to the wild-type strain. Disruption of lonC gene did not significantly influence the colonization and virulence of A. pleuropneumoniae. The data presented in this study illustrate that the LonA protease, but not the LonC protease, is required for the stress tolerance, biofilm formation and pathogenicity of A. pleuropneumoniae.

Decrease of colonization in the chicks' cecum and internal organs of Salmonella enterica serovar Pullorum by deletion of cpdB by Red system.

Salmonella enterica serovar Pullorum (S. Pullorum) is a worldwide poultry pathogen of considerable economic importance, particularly in those countries with a developing poultry industry. A variety of genes that affect S. Pullorum colonization in chickens had been identified. 2',3'-cyclic phosphodiesterase (cpdB) is the bifunctional enzyme which possess 2',3'-cyclic phosphodiesterase as well as 3'-nucleotidase activity. To assess the role of cpdB of S. Pullorum in colonization of cecum and internal organs in poultry, seven-day-old chicks were infected with 10(9) CFU/ml of a cpdB mutant and wild type strain. High number of cpdB mutant and wild type strain colonized the internal organs shortly after infection, but no colonization of cpdB mutant were observed from internal organs at day 10 post-infection, meanwhile, wild type bacteria in internal organs were observed at day 16 post-infection. Furthermore, the colonization of cpdB mutant in the cecum was seriously decreased from 6 days post-infection simultaneously wild type strain was increased and seriously decreased at day 8 post-infection. At day 12 post-infection, no cpdB mutant was observed from cecum, however high numbers of wild type strain were isolated at day 16 post-infection. It is concluded that cpdB is involved in long-term colonization of S. Pullorum in the chicks' cecum and internal organs. In addition, deletion of cpdB from S. Pullorum was not affect the morphology and growth of bacteria.

A Sub-1 ppm/°C Reference Voltage Source with a Wide Input Range.

With the continuous advancement of electronic technology, the application of high-voltage integrated circuits is becoming increasingly prevalent in fields such as power systems, medical devices, and industrial automation. The reference circuit within high-voltage integrated circuits must not only exhibit insensitivity to temperature variations but also maintain stability across a broad voltage supply. This paper presents a bandgap reference (BGR) source capable of operating over a wide input range. This BGR employs a high-order curvature compensation method to eliminate nonlinear voltage terms, resulting in minimal temperature drift. The circuit achieves an impressive temperature coefficient (TC) of 0.88 ppm/°C over a temperature range from -40 °C to 130 °C. To ensure stable operation within a 4-40 V range, the design incorporates a pre-regulation circuit that stabilizes the supply voltage of the BGR core at a fixed value, thereby enhancing the ability to withstand variations in power supply voltage.

Zmhdz9, an HD-Zip transcription factor, promotes drought stress resistance in maize by modulating ABA and lignin accumulation.

Maize is the largest crop in the world in terms of both planting area and total yield, and it plays a crucial role in ensuring global food and feed security. However, in recent years, with climate deterioration, environmental changes, and the scarcity of freshwater resources, drought has become a serious limiting factor for maize yield and quality. Drought stress-induced signals undergo a series of transmission processes to regulate the expression of specific genes, thereby affecting the drought tolerance of plants at the tissue, cellular, physiological and biochemical levels. Therefore, in this study we investigated the HD-Zip transcription factor gene Zmhdz9, and yeast activation experiments demonstrated that Zmhdz9 exhibited transcriptional activation activity. Under drought stress, high abscisic acid (ABA) and lignin levels significantly improved drought resistance in maize. Yeast two-hybrid, bimolecular fluorescence complementation (BIFC) and pull-down experiments showed that Zmhdz9 interacted with ZmWRKY120 and ZmTCP9, respectively. Overexpression of Zmhdz9 and gene editing of ZmWRKY120 or ZmTCP9 improved maize drought resistance, indicating their importance in the drought stress response. Furthermore, Zmhdz9 promoted the direct transcription of ZmWRKY120 in the W-box, activating elements of the ZmNCED1 promoter, which encodes a key enzyme in ABA biosynthesis. Additionally, Zmhdz9 promoted direct transcription of ZmTCP9 in the GGTCA motif, activating elements of the ZmKNOX8 promoter, which encodes a key enzyme in lignin synthesis. This study showed that the regulation of ABA and lignin by Zmhdz9 is essential for drought stress resistance in maize.

AE Characteristic and Mechanical Behaviors of Red Sandstone with Two Prefabricated Close-Collinear-Equal Length Cracks under Compression.

Preexisting cracks are of great significance for the stability of rocks, they can influence the growth and propagation pattern of cracks. For revealing the failure characteristics of rocks with prefabricated cracks, the mechanical behaviors of red sandstone with two close-collinear-equal length cracks were studied by acoustic emission (AE) monitoring experiment. The results indicated that the stress of red sandstone specimens showed an obvious stepwise growth phenomenon, and AE ring counts can better characterize the growth of microcracks of rocks. Moreover, AE events were mainly distributed near the prefabricated cracks of rocks. Then, a novel modified Kachanov method for analyzing collinear-close cracks was proposed. The influences of crack length and crack spacing on the interaction force of cracks were discussed. Then, it can be found that this modified method had a good applicability for two close-collinear-equal length cracks of rocks under compression.

Effects of short-term PM2.5 exposure on blood lipids among 197,957 people in eastern China.

Globally, air pollution is amongst the most significant causes of premature death. Nevertheless, studies on the relationship between fine particulate matter (PM2.5) exposure and blood lipids have typically not been population-based. In a large, community-based sample of residents in Yixing city, we assessed the relationship between short-term outdoor PM2.5 exposure and blood lipid concentrations. Participants who attended the physical examination were enrolled from Yixing People's hospital from 2015 to 2020. We collected general characteristics of participants, including gender and age, as well as test results of indicators of blood lipids. Data on daily meteorological factors were collected from the National Meteorological Data Sharing Center ( http://data.cma.cn/ ) and air pollutant concentrations were collected from the China Air Quality Online Monitoring and Analysis Platform ( https://www.aqistudy.cn/ ) during this period. We applied generalized additive models to estimate short-term effects of ambient PM2.5 exposure on each measured blood lipid-related indicators and converted these indicators into dichotomous variables (non- hyperlipidemia and hyperlipidemia) to calculate risks of hyperlipidemia associated with PM2.5 exposure. A total of 197,957 participants were included in the analysis with mean age 47.90 years (± SD, 14.28). The increase in PM2.5 was significantly associated with hyperlipidemia (odds ratio (OR) 1.003, 95% CI 1.001-1.004), and it was still significant in subgroups of males and age < 60 years. For every 10 µg/m3 increase in PM2.5, triglyceride levels decreased by 0.5447% (95% CI - 0.7873, - 0.3015), the low-density lipoprotein cholesterol concentration increased by 0.0127 mmol/L (95% CI 0.0099, 0.0156), the total cholesterol concentration increased by 0.0095 mmol/L (95% CI 0.0053, 0.0136), and no significant association was observed between PM2.5 and the high-density lipoprotein cholesterol concentration. After excluding people with abnormal blood lipid concentrations, the associations remained significant except for the high-density lipoprotein cholesterol concentration. PM2.5 was positively correlated with low-density lipoprotein cholesterol and total cholesterol, and negatively correlated with triglyceride, indicating PM2.5 can potentially affect health through blood lipid levels.

Draft genome sequence of Actinobacillus pleuropneumoniae serotype 7 strain S-8.

The Gram-negative bacterium Actinobacillus pleuropneumoniae is the etiological agent of porcine pleuropneumonia, a respiratory disease that leads to severe economic losses in the swine industry. For years, scientists working with it have lacked a reliable genome sequence for comparison with other Actinobacillus species. Here, we report the draft genome sequence of A. pleuropneumoniae serotype 7 (strain S-8), isolated from swine lung in China in 1992.

Experimental Study on the Spatial-Temporal Failure Characteristics of Red Sandstone with a Cemented Structural Surface under Compression.

A geological structural surface frequently appears in underground engineering and it significantly affects the stability of engineering structures. To elucidate the spatial-temporal failure characteristics of discontinuities between rocks, red sandstone with a firmly cemented discontinuity was investigated under uniaxial compression. The results revealed that adding a strong binder to rock discontinuities can improve the shear strength and stability of rock mass. The stress curve indicated that red sandstone specimens had elastic-brittle failure characteristics without obvious plastic deformation. The acoustic emission ring counts did not have large fluctuations, and its values were smaller (less than 2.5 × 104/s) at elastic deformation stage, which was termed as the "quiet period". The acoustic emission ring counts initially exhibited an obvious jump increase phenomenon at the forthcoming brittle failure stage before stress curve produced an obvious turning point, and then, it entered the "relatively quiet period". Based on the acoustic emission index (rise angle and average frequency), at the initial destruction stage of rocks, the types of internal fractures were simple and clearly (tensile cracks or shear cracks), and the fracture location was also relatively scattered inside specimens. Subsequently, the internal fracture developed into mixed tensile and shear cracks, and then, the fracture location exhibited a progressive evolution trend in the vertical space of specimens.

Integration of mRNA and microRNA analysis reveals the molecular mechanisms underlying drought stress tolerance in maize (Zea mays L.).

Drought is among the most serious environmental issue globally, and seriously affects the development, growth, and yield of crops. Maize (Zea mays L.), an important crop and industrial raw material, is planted on a large scale worldwide and drought can lead to large-scale reductions in maize corn production; however, few studies have focused on the maize root system mechanisms underlying drought resistance. In this study, miRNA-mRNA analysis was performed to deeply analyze the molecular mechanisms involved in drought response in the maize root system under drought stress. Furthermore, preliminary investigation of the biological function of miR408a in the maize root system was also conducted. The morphological, physiological, and transcriptomic changes in the maize variety "M8186" at the seedling stage under 12% PEG 6000 drought treatment (0, 7, and 24 h) were analyzed. With prolonged drought stress, seedlings gradually withered, the root system grew significantly, and abscisic acid, brassinolide, lignin, glutathione, and trehalose content in the root system gradually increased. Furthermore, peroxidase activity increased, while gibberellic acid and jasmonic acid gradually decreased. Moreover, 32 differentially expressed miRNAs (DEMIRs), namely, 25 known miRNAs and 7 new miRNAs, and 3,765 differentially expressed mRNAs (DEMRs), were identified in maize root under drought stress by miRNA-seq and mRNA-seq analysis, respectively. Through combined miRNA-mRNA analysis, 16 miRNA-target gene pairs, comprising 9 DEMIRs and 15 DEMRs, were obtained. In addition, four metabolic pathways, namely, "plant hormone signal transduction", "phenylpropane biosynthesis", "glutathione metabolism", and "starch and sucrose metabolism", were predicted to have important roles in the response of the maize root system to drought. MiRNA and mRNA expression results were verified by real-time quantitative PCR. Finally, miR408a was selected for functional analysis and demonstrated to be a negative regulator of drought response, mainly through regulation of reactive oxygen species accumulation in the maize root system. This study helps to elaborate the regulatory response mechanisms of the maize root system under drought stress and predicts the biological functions of candidate miRNAs and mRNAs, providing strategies for subsequent mining for, and biological breeding to select for, drought-responsive genes in the maize root system.

Improvement of cold tolerance in maize (Zea mays L.) using Agrobacterium-mediated transformation of ZmSAMDC gene.

Maize (Zea mays L.) is a food crop sensitive to low temperatures. As one of the abiotic stress hazards, low temperatures seriously affect the yield of maize. However, the genetic basis of low-temperature adaptation in maize is still poorly understood. In this study, maize S-adenosylmethionine decarboxylase (SAMDC) was localized to the nucleus. We used Agrobacterium-mediated transformation technology to introduce the SAMDC gene into an excellent maize inbred line variety GSH9901 and produced a cold-tolerant transgenic maize line. After three years of single-field experiments, the contents of polyamines (PAs), proline (Pro), malondialdehyde (MDA), antioxidant enzymes and ascorbate peroxidases (APXs) in the leaves of the transgenic maize plants overexpressing the SAMDC gene significantly increased, and the expression of elevated CBF and cold-responsive genes effectively increased. The agronomic traits of the maize overexpressing the SAMDC gene changed, and the yield traits significantly improved. However, no significant changes were found in plant height, ear length, and shaft thickness. Therefore, SAMDC enzymes can effectively improve the cold tolerance of maize.

Quaternized chitosan/cellulose composites as enhanced hemostatic and antibacterial sponges for wound healing.

It is essential to enhance our antibacterial arsenal in the first-aid hemostatic treatment due to the healing delay and even death from the bacteria-contaminated wounds. Herein, serial quaternized chitosan with varying degrees of substitution (QCSX) was prepared by glycidyl trimethyl ammonium chloride modification. Then the obtained QCSX was conjugated with dialdehyde cellulose (DAC) through Schiff base reaction to obtain the corresponding composite sponges (2QCSX-DAC). The surface morphology, chemical structures, and physical characters of mechanical measurement, water uptake behavior, porosity, and degradation tests were determined. Furthermore, in vitro and in vivo biological assays were performed. The obtained 2QCSX-DAC sponges exhibit abundant porous structures, moderate mechanical properties, excellent water uptake performance, and effective bactericidal rates against Staphylococcus aureus and Escherichia coli. Moreover, these porous composite sponges have superior blood coagulation abilities with the blood coagulation time reduced by 76.6% and 59.8% compared with blank control and Celox™ as well as low hemolysis rates (<5%). Meanwhile, 2QCS3-DAC had benign cytotoxicity of L929 cells in vitro and could accelerate the infected wound healing of rats at the early stage in vivo. Overall, this composite sponge appears to be a viable wound dressing for daily wound care in civilian hospitals and emergency hemostasis on battlefields.

A CRISPR/Cas12a-assisted rapid detection platform by biosensing the apxIVA of Actinobacillus pleuropneumoniae.

Actinobacillus pleuropneumoniae is an important respiratory pig pathogen that causes substantial losses in the worldwide swine industry. Chronic or subclinical infection with no apparent clinical symptoms poses a challenge for preventing transmission between herds. Rapid diagnostics is important for the control of epidemic diseases. In this study, we formulated an A. pleuropneumoniae species-specific apxIVA-based CRISPR/Cas12a-assisted rapid detection platform (Card) that combines recombinase polymerase amplification (RPA) of target DNA and subsequent Cas12a ssDNase activation. Card has a detection limit of 10 CFUs of A. pleuropneumoniae, and there is no cross-reactivity with other common swine pathogens. The detection process can be completed in 1 h, and there was 100% agreement between the conventional apxIVA-based PCR and Card in detecting A. pleuropneumoniae in lung samples. Microplate fluorescence readout enables high-throughput use in diagnostic laboratories, and naked eye and lateral flow test readouts enable use at the point of care. We conclude that Card is a versatile, rapid, accurate molecular diagnostic platform suitable for use in both laboratory and low-resource settings.

Enhancement of bacteriolysis of shuffled phage PhiX174 gene E.

Bacterial ghosts that are generated using the regulated PhiX174 lysis gene E offer a new avenue for the study of inactivated vaccines. Here, we constructed a library of mutant gene E using a gene-shuffling technique. After screening and recombination with the prokaryotic non-fusion expression vector pBV220, two lysis plasmids were selected. Among which, a novel mutant E gene (named mE), consisting of a 74-bp non-encoding sequence at 5'-end and a 201-bp gene ΔE, significantly increased the lysis effect on prokaryotic Escherichia coli and Salmonella enteritidis. Moreover, lysis efficiency, as measured by the OD600 value, reached 1.0 (109 CFU), avoiding the bottleneck problem observed with other bacterial lysis procedures, which results in a low concentration of bacteria in suspension, and consequent low production of bacterial ghosts. Our results may provide a promising avenue for the development of bacterial ghost vaccines.

Polysaccharides-modified chitosan as improved and rapid hemostasis foam sponges.

Serial hemostatic sponges consisting of polysaccharides-modified chitosan foam sponges were prepared by Schiff base crosslinking reaction between the deacetylated chitosan and oxidized dialdehyde cellulose. Such composite foam sponges were characterized by scanning electron microscopy and Fourier-transform infrared spectroscopy to confirm their morphology and compositions. Then the coagulation process was evaluated in vitro by thrombus elasticity meters. Furthermore, the hemostasis experiments on mouse tail vein and rabbit femoral artery were also performed in vivo. The results strongly indicated that such synergistic cellulose-modified chitosan foam sponges showed comprehensively excellent water-absorbing quality, improved mechanical performance, low hemolysis rates, benign cytotoxicity, good resilience ability after repeated compression, and superior hemostasis capability both in vitro and in vivo. Furthermore, the hemostatic mechanism is via adhering/activating the red blood cell/platelet to form robust blood clots through the endogenous coagulation pathway, which serves as a good candidate for emergency trauma treatment in daily civilian and military hemostasis.