Expression profiles and characterization of microRNAs responding to chitin in Arthrobotrys oligospora.

Extracellular proteases, such as chitinases secreted by Arthrobotrys oligospora (A. oligospora), play a crucial role in the process of nematode infection. However, post-transcriptional regulation of gene expression involving microRNAs (miRNAs) in A. oligospora remains scarcely described. Hereto, transcriptome sequencing was carried out to analyze the expression profiles of chitin-responsive miRNAs in A. oligospora. Based on the RNA-seq data, the differential expression of miRNAs (DEmiRNAs) in response to chitin was screened, identified and characterized in A. oligospora. Meanwhile, the potential target genes were predicted by the online tools miRanda and Targetscan, respectively. Furthermore, the interaction of DEmiRNA with it's target gene was validated by a dual-luciferase reporter assay system. Among 85 novel miRNAs identified, 25 miRNAs displayed significant differences in expression in A. oligospora in response to chitin. Gene Ontology (GO) analysis showed that the potential genes targeted by DEmiRNAs were enriched in the biological processes such as bio-degradation, extracellular components and cell cycle. KEGG analysis revealed that the target genes were mainly involved in Hippo, carbon and riboflavin metabolic pathway. Outstandingly, chitinase AOL_s00004g379, which is involved in the hydrolysis metabolic pathway of chitin, was confirmed to be a target gene of differential miR_70. These findings suggest that chitin-responsive miRNAs are involved in the regulation of cell proliferation, predator hyphae growth and chitinase expression through the mechanisms of post-transcriptional regulation, which provides a new perspective to the molecular mechanisms underlying miRNAs-mediated control of gene expression in A. oligospora.

Molecular characterization and phylogenetic analysis of porcine epidemic diarrhea virus in Xinjiang, China, from 2020 to 2022.

In recent years, the pig industry in Xinjiang, China, has been severely impacted by outbreaks of porcine epidemic diarrhea (PED), despite vaccination efforts. In this study, we investigated the genetic characteristics of currently prevalent porcine epidemic diarrhea virus (PEDV) strains in the region. We collected 548 samples from animals with suspected PED on large-scale pig farms in Xinjiang. Of these, 258 tested positive for PEDV by RT-PCR, yielding an overall positivity rate of 47.08%. S1 gene sequencing and phylogenetic analysis were conducted on 23 randomly selected RT-PCR-positive samples. Three endemic strains of PEDV (PEDV/CH/XU/2020, PEDV/CH/XK/2020, and PEDV/CH/XA/2020) were isolated, and their complete genome sequences were analyzed for evidence of genetic recombination. Sequence comparison of the S gene indicated significant variations in the S1 gene of the Xinjiang strains compared to the vaccine strains CV777, AJ1102, and LWL, with 90.2%-98.5% nucleotide sequence identity. Notably, both the N-terminal and C-terminal domains of the S protein showed significant variation. Genetic evolutionary analysis identified the GIIa subtype as the dominant genotype among the epidemic strains in Xinjiang. Recombination analysis revealed inter-subtype recombination events in the PEDV/CH/XK/2020 and XJ1904-34 strains. These findings highlight the extensive genetic variation in the predominant GIIa genotype of PEDV in Xinjiang, which does not match the genotype of the currently used vaccine strains. These data may guide further efforts toward the development of effective vaccines for the control of PED.

STM1863, a Member of the DUFs Protein Family, Is Involved in Environmental Adaptation, Biofilm Formation, and Virulence in Salmonella Typhimurium.

Salmonella Typhimurium (STM) is an important zoonotic Gram-negative pathogen that can cause infection in a variety of livestock and poultry. Meanwhile, as an important foodborne pathogen, the bacterium can survive in various stressful environments and transmits through the fecal-oral route, posing a serious threat to global food safety. To investigate the roles of STM1863, a member of the DUFs protein family, involved in STM environmental adaptation, biofilm formation, and virulence. We analyzed the molecular characteristics of the protein encoded by STM1863 gene and examined intra- and extracellular expression levels of STM1863 gene in mouse macrophages. Furthermore, we constructed STM1863 gene deletion and complementation strains and determined its environmental adaptation under stressful conditions such as acid, alkali, high salt, bile salt, and oxidation. And the capacity of biofilm formation and pathogenicity of those strains were analyzed and compared. In addition, the interaction between the promoter of STM1863 gene and RcsB protein was analyzed using DNA gel electrophoresis migration assay (electrophoretic mobility shift assay [EMSA]). The experiments revealed that acid adaptability and biofilm formation ability of STM1863 gene deletion strain were significantly weakened compared with the parental and complementary strains. Moreover, the adhesion and invasion ability of STM1863 deletion strain to mouse macrophages was significantly decreased, while the median lethal dose (LD50) increased by 2.148-fold compared with the parental strain. In addition, EMSA confirmed that RcsB protein could bind to the promoter sequence of STM1863 gene, suggesting that the expression of STM1863 gene might be modulated by RcsB. The present study demonstrated for the first time that STM1863, a member of the DUFs protein family, is involved in the modulation of environmental adaptation, biofilm formation, and virulence.

A Novel Spectrophotometric Method for Determination of Percarbonate by Using N, N-Diethyl-P-Phenylenediamine as an Indicator and Its Application in Activated Percarbonate Degradation of Ibuprofen.

Sodium percarbonate (SPC) concentration can be determined spectrophotometrically by using N, N-diethyl-p-phenylenediamine (DPD) as an indicator for the first time. The ultraviolet-visible spectrophotometry absorbance of DPD•+ measured at 551 nm was used to indicate SPC concentration. The method had good linearity (R2 = 0.9995) under the optimized experimental conditions (pH value = 3.50, DPD = 4 mM, Fe2+ = 0.5 mM, and t = 4 min) when the concentration of SPC was in the range of 0-50 µM. The blank spiked recovery of SPC was 95-105%. The detection limit and quantitative limit were 0.7-1.0 µM and 2.5-3.3 µM, respectively. The absorbance values of DPD•+ remained stable within 4-20 min. The method was tolerant to natural water matrix and low concentration of hydroxylamine (<0.8 mM). The reaction stoichiometric efficiency of SPC-based advanced oxidation processes in the degradation of ibuprofen was assessed by the utilization rate of SPC. The DPD and the wastewater from the reaction were non-toxic to Escherichia coli. Therefore, the novel Fe2+/SPC-DPD spectrophotometry proposed in this work can be used for accurate and safe measurement of SPC in water.

Transport and interaction of cadmium and active sludge extracellular polymeric substances in saturated sand influenced by ionic strength and composition.

Artificial groundwater recharge with reclaimed water (secondary effluent from wastewater treatment plants) has become an important approach to solving water scarcity worldwide. Microorganisms in activated sludge can secrete many extracellular polymeric substances (EPS). However, information on the impact of EPS on the movement of heavy metals in porous media and their environmental effects on underground networks is limited. To assess this risk, we extracted EPS from the aerobic section of a wastewater treatment plant using hot sodium hydroxide and conducted experiments using one-dimensional sand columns to investigate how ion composition and strength affect the movement and interaction of cadmium (Cd) and EPS in porous media. The results showed that EPS facilitated Cd migration in porous media. Sodium (Na) and calcium (Ca) ions promoted the migration of Cd in porous media and EPS-Cd complexation. The effect of Ca was more significant than that of Na. As the Na adsorption ratio increased, the migration ability of Cd in porous media and the complexation ability of EPS with Cd decreased. Therefore, when estimating the migration of EPS and Cd in subsurface environments, careful consideration should be given to prevent the risk of groundwater pollution.

The small RNA STnc1480 contributes to the regulation of biofilm formation and pathogenicity in Salmonella typhimurium.

Salmonella Typhimurium (STM) is one of the most important food-borne bacteria that seriously harms livestock and human beings, which is capable of regulating the expression of its own genes in a variety of ways to adapt to a wide variety of adverse environmental stresses. To understand the regulatory roles of sRNA STnc1480 on the capability of STM, the STnc1480 gene-deficient strain △STnc1480 and its complement strain △STnc1480/STnc1480 were generated, and the impacts of STnc1480 gene deficiency on the capability of responding to different environmental stresses, biofilm(BF)formation and pathogenicity were analyzed, respectively. Then the target genes that were regulated by STnc1480 were also analyzed and explored. Compared with parent and complement strains, the deficiency of the STnc1480 gene significantly reduced the BF formation. Moreover, the capacities of adhesion and invasiveness of the △STnc1480 strain to macrophages were also significantly reduced, while the LD50 in mice was significantly increased. The bacterial loads in liver and spleen were significantly reduced, and the pathological damage was alleviated. It was confirmed that the STnc1480 could be complementary to the 5'-UTR (-52 to -71 bases) region of lpfA mRNA. The bacterial dual-plasmid reporting system confirmed that STnc1480 was capable of interacting with the mRNA of the lpfA gene, suggesting that STnc1480 can regulate the 5'-UTR of the lpfA mRNA at post-transcription level to reduce the expression of the bacterial fimbria, thus reducing the BF formation and pathogenicity of STM.

Polaronic defects in monolayer CeO2: Quantum confinement effect and strain engineering.

We uncover the structure, stability, and electronic properties of polaronic defects in monolayer (ML) CeO2 by means of first-principles calculations, with special attention paid to the quantum confinement effect induced by dimensionality reduction. Results show that the polaron can be more stabilized in ML CeO2 than in the bulk, while formation of oxygen vacancy (Vo2+) and polaron-vacancy complexes [(Vo2+-1polaron)1+, (Vo2+-2polaron)0] tends to be more difficult. The polaronic defect states sit deeper in energy within the bandgap of ML CeO2 compared to the bulk case. We further demonstrate that the epitaxial strain in ceria film, as normally exists when grown on metal substrate, plays a crucial role in regulating the defect energetics and electronic structures. In particular, the formation energies of polarons, Vo2+, (Vo2+-1polaron)1+, and (Vo2+-2polaron)0, generally decrease with tensile strain, leading to controllable defect concentration with strain and temperature. This study not only provides physical insights into the polaronic defects in ultrathin oxide films, but also sheds light on their potential technological applications in nanoelectronics, fuel cells, and catalysts.

Influence of coracoglenoid space on scapular neck fracture stability: biomechanical study.

BACKGROUND: The anatomical variation of the coracoglenoid space has the potential to influence the stability of scapular neck fractures. This paper aimed to investigate the mechanical mechanism underlying the influence of different coracoglenoid space types on scapular neck fractures by morphometric analysis and biomechanical experiments. METHODS: The morphology of 68 dried scapulae (left: 36; right: 32) was studied. Two variables, the length of the coracoglenoid distance (CGD) and the coracoglenoid notch (CGN), were measured. The distribution of CGN/CGD × 100% was used to identify the morphology of the coracoglenoid space. Each specimen was tested for failure under static axial compression loading. The average failure load, stiffness, and energy were calculated. RESULTS: Two coracoglenoid space types were identified. The incidence of Type I (''hook'' shape) was 53%, and that of Type II (''square bracket'' shape) was 47%. The CGD and CGN were significantly higher for type I than type II (13.81 ± 0.74 mm vs. 11.50 ± 1.03 mm, P < 0.05; 4.74 ± 0.45 mm vs. 2.61 ± 0.45 mm, P < 0.05). The average maximum failure load of the two types was 1270.82 ± 318.85 N and 1529.18 ± 467.29 N, respectively (P = 0.011). The stiffness and energy were significantly higher for type II than type I (896.75 ± 281.14 N/mm vs. 692.91 ± 217.95 N/mm, P = 0.001; 2100.38 ± 649.54 N × mm vs. 1712.71 ± 626.02 N × mm, P = 0.015). CONCLUSIONS: There was great interindividual variation in the anatomical morphology of the coracoglenoid space. Type I (hook-like) spaces bore lower forces, were less stiff, and bore less energy, which may constitute an anatomical predisposition to scapular neck fractures.

Biological Characteristics of Recombinant Arthrobotrys oligospora Chitinase AO-801.

Chitinase AO-801 is a hydrolase secreted by Arthrobotrys oligospora during nematode feeding, while its role remained elusive. This study analyzed the molecular characteristics of recombinant chitinase of Arthrobotrys oligospora (reAO-801). AO-801 belongs to the typical glycoside hydrolase 18 family with conserved chitinase sequence and tertiary structure of (α/ß)8 triose-phosphate isomerase (TIM) barrel. The molecular weight of reAO-801 was 42 kDa. reAO-801 effectively degraded colloidal and powdered chitin, egg lysate, and stage I larval lysate of Caenorhabditis elegans. The activity of reAO-801 reached its peak at 40˚C and pH values between 4-7. Enzyme activity was inhibited by Zn2+, Ca2+, and Fe3+, whereas Mg2+ and K+ potentiated its activity. In addition, urea, sodium dodecyl sulfate, and 2-mercaptoethanol significantly inhibited enzyme activity. reAO-801 showed complete nematicidal activity against C. elegans stage I larvae. reAO-801 broke down the C. elegans egg shells, causing them to die or die prematurely by hatching the eggs. It also invoked degradation of Haemonchus contortus eggs, resulting in apparent changes in the morphological structure. This study demonstrated the cytotoxic effect of reAO-801, which laid the foundation for further dissecting the mechanism of nematode infestation by A. oligospora.

Molecular Characteristics and Potent Immunomodulatory Activity of Fasciola hepatica Cystatin.

Cystatin, a cysteine protease inhibitor found in many parasites, plays important roles in immune evasion. This study analyzed the molecular characteristics of a cystatin from Fasciola hepatica (FhCystatin) and expressed recombinant FhCystatin (rFhcystatin) to investigate the immune modulatory effects on lipopolysaccharide-induced proliferation, migration, cytokine secretion, nitric oxide (NO) production, and apoptosis in mouse macrophages. The FhCystatin gene encoded 116 amino acids and contained a conserved cystatin-like domain. rFhCystatin significantly inhibited the activity of cathepsin B. rFhCystatin bound to the surface of mouse RAW264.7 cells, significantly inhibited cell proliferation and promoted apoptosis. Moreover, rFhCystatin inhibited the expression of cellular nitric oxide, interleukin-6, and tumor necrosis factor-α, and promoted the expression of transforming growth factor-ß and interleukin-10. These results showed that FhCystatin played an important role in regulating the activity of mouse macrophages. Our findings provide new insights into mechanisms underlying the immune evasion and contribute to the exploration of potential targets for the development of new drug to control F. hepatica infection.

First-principles screening of single transition metal atoms anchored on two-dimensional C9N4 for the nitrogen reduction reaction.

Compared to the Haber-Bosch process, the electrochemical nitrogen reduction reaction (NRR) can convert N2 into NH3 under ambient conditions, and thus has attracted considerable attention in recent years. However, it remains a challenge to fabricate NRR catalysts with high faradaic efficiency and yield rate. In this work, by systematic first-principles calculations, we investigate the structure, stability and catalytic performance of single metal atoms anchored on porous monolayer C9N4 (M@C9N4) for the electrochemical NRR. A total of 25 transition metals (Sc-Zn, Zr-Mo, Ru-Ag, Hf-Au) were explored, and we screened out four promising systems, i.e., Nb, Ta, Re and W@C9N4, which not only exhibit high catalytic activity with low limiting potentials of -0.3, -0.42, -0.49 and -0.25 V, respectively, but also have superior selectivity that suppresses the competitive hydrogen evolution reaction. The physical origin lies in the coupling between the d orbitals of the transition metals and the 2π* orbital of N2, which activates the N2 molecule and facilitates the reduction process. Our proposed systems are kinetically and thermodynamically stable, which may shed light on future design and fabrication of high-efficiency single atom catalysts for various technologically important chemical reactions.

Development and evaluation of multi-epitope protein p72 (MeP72) for the serodiagnosis of African swine fever.

African swine fever (ASF) is an acute and severe infectious disease that seriously endangers the global porcine industry. In order to develop ASF serodiagnostic reagents with high specificity and sensitivity, in the present study, the antigenic epitopes of P72 protein of African swine fever virus (ASFV) were analyzed, and the ASFV multi-epitope fusion gene MeP72 in tandem with the dominant linear epitopes was constructed. The recombinant multi-epitope fusion MeP72 (reMeP72) was prepared in Escherichia coli. A colloidal gold-based immunochromatographic assay (CGIA) based on reMeP72 was developed for the detection of antibodies against ASFV. A total of 139 pig clinical serum samples were used for assessment of the potential diagnostic value of reMeP72. The results showed that CGIA did not cross-react with positive sera of viruses, such as classical swine fever virus (CSFV), porcine circovirus type 2 (PCV2), pseudorabies virus (PRV), porcine reproductive and respiratory syndrome virus (PRRSV) and swine influenza virus (SIV), showing high specificity. Sensitivity analysis showed that CGIA could detect ASFV-positive serum at a dilution of 1:64. Compared with commercial ASFV kits, the sensitivity and specificity of ASFV CGIA based on reMeP72 protein were 85.7% and 97.6%, respectively. The agreement rate of the two methods was 96.4%, showing a good detection performance. The results indicated that the reMeP72 was of potential value for the serodiagnosis of ASF. Keywords: African swine fever virus; P72 gene; antigenic protein; colloidal gold-based immunochromatographic assay.

The association between collagen gene polymorphisms and intracranial aneurysms: a meta-analysis.

The association between single nucleotide polymorphisms (SNPs) of the collagen gene and intracranial aneurysm (IA) pathogenesis remains controversial. Thus, in this study, a meta-analysis was performed to evaluate the association between collagen gene SNPs and the incidence of IA. A systematic search of major online databases up to March 2017 was performed. Five genetic models (allelic, dominant, recessive, heterozygous, and homozygous models) were used to analyze the associations. A total of 14 trials with 13,709 patients were included. Four collagen genes, COL1A2 (21 SNPs), COL3A1 (7 SNPs), COL4A1 (6 SNPs), and COL4A2 (1 SNP), were analyzed. We observed that rs42524 in the COL1A2 gene was associated with a significant increase in the risk of IA in Japanese patients (allelic model: OR, 1.94; 95% CI, 1.03-3.64; p = 0.04); the rs1800255 polymorphism in the COL3A1 gene was significantly correlated with Chinese IA patients (allelic model: OR, 1.50; 95% CI, 1.30-1.73; p < 0.001); and rs2621215 was significantly correlated with IA for the heterozygous model (OR, 1.58; 95% CI, 1.15-2.17; p = 0.005) and the dominant model (OR, 1.49; 95% CI, 1.09-2.02; p = 0.012). Furthermore, in the COL4A1 gene, there was a significant relationship between the rs3783107 polymorphism and a Dutch IA population (allelic model: OR, 1.23; 95% CI, 1.06-1.42; p = 0.006), and the prevalence ratio of mutation carriers in the Dutch population was significantly higher than that in the Japanese population (ROR, 1.31; 95% CI, 1.07-1.63; p = 0.008). The rs1800255 polymorphism in COL3A1 is robustly correlated with IA in the Chinese population. Three COL1A2 SNPs-rs42524, rs1800238, and rs2621215-should be studied further.

The Regulatory Roles of ncRNA Rli60 in Adaptability of Listeria monocytogenes to Environmental Stress and Biofilm Formation.

Listeria monocytogenes is a facultative anaerobic Gram-positive bacterium. It is well adapted to external environments and able to infect both humans and animals. To understand the impacts of ncRNA Rli60 on the adaptability of L. monocytogenes to environmental stresses and biofilm formation, a rli60 deletion strain of L. monocytogenes (LM-Δrli60) was constructed using splicing by overlap extension PCR (SOE-PCR) and homologous recombination and then compared it with wild-type strain L. monocytogenes EGD-e in the aspects of adaptability to environmental stresses by measuring their growth under stresses of different temperatures, and acidic, alkaline, hypertonic and alcoholic conditions, and capability of biofilm formation by using crystal violet staining, as well as the transcriptional levels of genes (gltB and gltC) related to the biofilm formation by real-time quantitative PCR (qRT-PCR). The results showed that (1) the growth of LM-Δrli60 strain was significantly slower under environmental stresses of low temperature (30 °C), high temperature (42 °C), as well as alkaline and alcoholic conditions, (2) the amount of biofilm formed by LM-Δrli60 was attenuated, and (3) the transcriptional levels of gltB and gltC genes at 24 h and 48 h in LM-Δrli60 revealed a significant reduction. Overall, the results confirmed that ncRNA Rli60 plays important roles in regulating the adaptability of L. monocytogenes to environmental stresses and biofilm formation possibly through impacting the expression of gltB and gltC genes.

Knock-down of stress inducible OsSRFP1 encoding an E3 ubiquitin ligase with transcriptional activation activity confers abiotic stress tolerance through enhancing antioxidant protection in rice.

E3 ubiquitin ligases are involved in a variety of physiological processes. This study demonstrated the function of a previously unknown rice RING finger E3 ligase, Oryza sativa Stress-related RING Finger Protein 1 (OsSRFP1) in stress responses in rice. OsSRFP1 was ubiquitously expressed in various rice organs, with the higher expression levels in roots, panicles and culm nodes. The transcript of OsSRFP1 was induced by cold, dehydration, salt, H2O2 and abscisic acid treatments. Interestingly, the OsSRFP1-overexpressing plants were less tolerant to salt, cold and oxidative stresses than wild type plants; while the RNA interference silencing of OsSRFP1 plants were more tolerant than wild type without yield penalty. Compared with the wild type, amounts of free proline and activities of antioxidant enzymes were increased in the RNAi plants but decreased in the overexpression plants under cold stress, which were inversely correlated with the malondialdehyde and hydrogen peroxide (H2O2) levels in the tested lines. Microarray analysis showed that expression of numerous genes involving in ROS homeostasis was altered in the OsSRFP1-overexpressing plants under normal and cold conditions. In vitro ubiquitination assays showed that OsSRFP1 possessed E3 ubiquitin ligase activity and the intact RING domain was essential for the activity. Moreover, OsSRFP1 might function in transcriptional regulation with nuclear localization. Taken together, our results demonstrate that OsSRFP1 may have dual functions in post-translational and transcriptional regulations in modulating abiotic stress responses in rice, at least in part, by negatively regulating antioxidant enzymes-mediated reactive oxygen species removal.

Expression of serine proteinase P186 of Arthrobotrys oligospora and analysis of its nematode-degrading activity.

The nematode-trapping fungi possess a unique capability of predating and invading nematodes. As a representative nematode-trapping fungus, Arthrobotrys oligospora has been widely used to study the interactions between nematode-trapping fungi and their hosts. Serine proteinase is one of the important virulence factors during process of invasion of the nematode-trapping fungi into nematodes. In this study, using reverse transcription polymerase chain reaction, we amplified the gene sequence of serine proteinase 186 from A. oligospora, cloned it into pPIC9K vector and expressed it in the yeast Pichia pastoris. The expressed recombinant serine proteinase186 (reP186) was purified via Ni-affinity chromatography. The in vitro nematode-degrading activity of reP186 was analyzed. Sodium dodecyl sulfate polyacrylamide gel electrophoresis and Western blot analysis revealed that reP186 with molecular weight of 33 kDa was successfully obtained. ReP186 was capable of degrading a series of protein substrates including casein, gelatin, bovine serum albumin, denatured collagen and nematode cortical layer. The reP186 exhibited the maximal activity at pH 8.0 and 55 °C and was highly sensitive to the inhibitor, phenylmethanesulfonylfluoride. Treatment of Caenorhabditis elegans and Haemonchus contortus with reP186 for 12, 24 and 36 h, respectively, resulted in 62, 88 and 100 % of killing rates for C. elegans, and 52, 65 and 84 % of killing rates for H. contortus, respectively, indicating a relatively strong nematode-degrading bioactivity of reP186.

The efficacies of 5 insecticides against hard ticks Hyalomma asiaticum, Haemaphysalis longicornis and Rhipicephalus sanguineus.

At present, chemical-based tick control strategies are still the most efficient and widely used methods in control of ticks and tick-borne diseases. In this study, the efficacies of lambda-cyhalothrin, beta-cypermethrin, emamectin benzoate, spirotetramat and hexaflumuron in vitro were evaluated against Hyalomma asiaticum, Haemaphysalis longicornis and Rhipicephalus sanguineus that are widespread and able to transmit a variety of human and animal diseases in China. The results showed that the LC (lethal concentration) 50 of lambda-cyhalothrin, beta-cypermethrin, emamectin benzoate, spirotetramat and hexaflumuron were 22.05, 107.35, 287.62, 432.25 and over 6250 mg/L to Hy. asiaticum engorged nymphs, respectively. The LC50 of lambda-cyhalothrin and beta-cypermethrin were each to 100.69 mg/L and 340.05 mg/L against Hy. asiaticum unfed adults. In addition, 50 mg/L of lambda-cyhalothrin could completely inhibit engorged females of the 3 tick species to lay eggs. These results indicate that lambda-cyhalothrin has the highest efficacy and broadest spectrum for against the 3 tick species. The present study provides some information for selecting chemical acaricides in control ticks and tick-borne-diseases, as well for preparing acaricide mixtures to improve killing efficacy, and retard the advent of tick-resistance of acaricides in China.

Detection of genetic variations in Orf virus isolates epidemic in Xinjiang China.

Orf is an exanthemous viral disease seriously threatening the goat and sheep industry and widely epidemic in the goat and sheep populations in Xinjiang, China. In order to investigate the genetic variability of the orf virus (ORFV), three virus isolates (SHZ1, SHZ2, and SHZ3) were isolated by PCR and Vero cell culture using the clinical samples from the lips of the lambs suspected of ORFV infection. The isolates were further verified by electron microscopy and animal infection experiments. The protective antigen genes B2L, F1L, and virulence genes VIR, GIF, and VEGF in the isolates were cloned, sequenced and analyzed for genetic evolution. The results showed that B2L and F1L were relatively conservative with homology 86.7-97.9%, while VIR, GIF, particularly VEGF were considerably variable with homology 71.5-97.9% at amino acid sequence level, respectively. Phylogenetic tree analysis based on B2L and VIR showed that the isolates SHZ1 and SHZ2 were closely related with the Taiwan isolates. This is the first report to confirm that there have been genetic variations in the Xinjiang ORFV isolates. The findings provide molecular evidence about the genetic variability of the major antigenic and virulence genes in the virus isolates epidemic in Xinjiang.

STnc1280, a trans-coding sRNA is involved in virulence modulation via targeting gldA mRNA in Salmonella Typhimurium.

Introduction. Salmonella Typhimurium (STM) is a food-borne Gram-negative bacterium, which can infect humans and a wide range of livestock and poultry, causing a variety of diseases such as septicaemia, enteritis and abortion.Hypothesis/Gap Statement. We will decipher the impacts of sRNA STnc1280 on STM virulence and provide a theoretical basis to reveal the regulatory role and molecular mechanism of STnc1280.Aim. The main objective of this study was to clarify whether sRNA STnc1280 exerts regulatory roles on STM pathogenicity.Methodology. The STnc1280 gene was amplified and its molecular characteristics were analysed in this study. Then, STnc1280 gene deletion strain (STM-ΔSTnc1280) and the complementary strain (ΔSTnc1280/STnc1280) were constructed by λ-Red homologous recombination method, respectively, to analyse of adhesion and invasive ability and pathogenicity of different strains. Subsequently, the potential target gene regulated by STnc1280 was predicted using target RNA2 software, followed by the verification of the interaction between STnc1280 and target mRNA using the dual plasmid reporter system (DPRS). Furthermore, the mRNA and protein level of target gene was determined using qRT-PCR and Western blot, respectively.Results. The results revealed that the cell adhesion and invasive ability and pathogenicity of STM-ΔSTnc1280 were significantly reduced compared to STM-SL1344 strain, indicating that the deficiency of STnc1280 gene significantly influenced STM pathogenicity. The DPRS results showed that STnc1280 can interact with the mRNA of target gene gldA, thus suppressing the expression of lacZ gene. Furthermore, the level of gldA mRNA was not influenced in STM-ΔSTnc1280, but the expression of GldA protein decreased significantly.Conclusion. Combining the bioinformatic analysis, these findings suggested that STnc1280 may bind to the SD sequence of gldA mRNA, hindering the binding of ribosomes to gldA mRNA, thereby inhibiting the expression of GldA protein to modulate the virulence of STM.

Construction and immunological evaluation of truncated hepatitis B core particles carrying HBsAg amino acids 119-152 in the major immunodominant region (MIR).

Hepatitis B capsid protein expressed in Escherichia coli can reassemble into icosahedral particles, which could strongly enhance the immunogenicity of foreign epitopes, especially those inserted into its major immunodominant region. Herein, we inserted the entire 'α' antigenic determinant amino acids (aa) 119-152 of HBsAg into the truncated HBc (aa 1-144), between Asp(78) and Pro(79). Prokaryotic expression showed that the mosaic HBc was mainly in the form of inclusion bodies. After denaturation with urea, it was dialyzed progressively for protein renaturation. We observed that before and after renaturation, mosaic HBc was antigenic as determined by HBsAg ELISA and a lot of viruslike particles were observed after renaturation. Thus, we further purified the mosaic viruslike particles by (NH4)2SO4 precipitation, DEAE chromatography, and Sepharose 4FF chromatography. Negative staining electron microscopy demonstrated the morphology of the viruslike particles. Immunization of Balb/c mice with mosaic particles induced the production of anti-HBs antibody and Th1 cell immune response supported by ELISPOT and CD4/CD8 proportions assay. In conclusion, we constructed mosaic hepatitis core particles displaying the entire 'α' antigenic determinant on the surface and laid a foundation for researching therapeutic hepatits B vaccines.