Search

ABSTRACT Porcine deltacoronavirus (PDCoV) is a newly emerged enteric pathogen that causes diarrhea, dehydration, vomiting, and in suckling piglets, a high rate of death. Here, we used endocytosis inhibitors, dominant-negative mutants, and small interfering RNA (siRNA) interference to investigate PDCoV entry into swine testicle (ST) cells. Results showed that chlorpromazine, dynasore, and methyl-β-cyclodextrin (MβCD) inhibited PDCoV internalization; immunofluorescence assay (IFA) of dominant-negative (DN) mutants and siRNA interference further demonstrated these results, indicating that PDCoV utilizes clathrin-mediated endocytosis and requires dynamin and cholesterol to enter ST cells. The role of endosomes in PDCoV infection was also investigated and we found that PDCoV does not require Rab5, Rab7, or Rab11 for productive infection, indicating that the transport of PDCoV in ST cells does not require the participation of the endosomal system. Our findings will provide additional understanding of the entry mechanisms of PDCoV and possible antiviral targets. IMPORTANCE Porcine deltacoronavirus (PDCoV) is a newly emerged enteric virus threatening pig industries worldwide. Our previous work showed that PDCoV enters porcine kidney (PK-15) cells through a caveolae-dependent pathway, but the entry mechanism for PDCoV into swine testicle (ST) cells remains unclear. Mechanisms of virus entry can be different with different virus isolates and cell types. Here, we determined that PDCoV enters ST cells via clathrin-mediated endocytosis. Additionally, we found that PDCoV entry does not require Rab5, Rab7, or Rab11. These findings provide additional understanding of the entry mechanisms of PDCoV and possible antiviral targets.

Enterovirus G belongs to the family Picornaviridae and are associated with a variety of animal diseases. We isolated and characterized a novel EV-G2 strain, CHN-SCMY2021, the first genotype 2 strain isolated in China. CHN-SCMY2021 is about 25 nm diameter with morphology typical of picornaviruses and its genome is 7341 nucleotides. Sequence alignment and phylogenetic analysis based on VP1 indicated that this isolate is a genotype 2 strain. The whole genome similarity between CHN-SCMY2021 and other EV-G genotype 2 strains is 78.3–86.4%, the greatest similarity is to EVG/Porcine/JPN/Iba26–506/2014/G2 (LC316792.1). Recombination analysis indicated that CHN-SCMY2021 resulted from recombination between 714,171/CaoLanh_VN (KT265894.2) and LP 54 (AF363455.1). Except for ST cells, CHN-SCMY2021 has a broad spectrum of cellular adaptations, which are susceptible to BHK-21, PK-15, IPEC-J2, LLC-PK and Vero cells. In piglets, CHN-SCMY2021 causes mild diarrhea and thinning of the intestinal wall. The virus was mainly distributed to intestinal tissue but was also found in heart, liver, spleen, lung, kidney, brain, and spinal cord. CHN-SCMY2021 is the first systematically characterized EV-G genotype 2 strain from China, our results enrich the information on the epidemiology, molecular evolution and pathogenicity associated with EV-G.

The craft beer was continuously sterilized by the non-thermal plasma reactor, and its sterilization and quality were evaluated. The total count of aerobic plate, yeast and Escherichia coli were used as microbial indexes, the effects of plasma sterilization and pasteurization on the bactericidal effect of craft beer were studied. Meanwhile, the beer color, foam hold and pH were evaluated to obtain the optimum sterilization condition with the response surface. The results showed that, compared with pasteurization, the sterilization rate after plasma treatment was almost the same, but the chroma, brightness and bubble retention were better, which was closer to the original beer. The total number of colonies and Escherichia coli were almost completely killed, and the number of yeast was reduced by 97.5% with the 40 kV plasma treatment voltage, 10 s and 60 mL/min gas flow rate. According to the analysis of Design Expert software, the sterilization effect and brightness were the best when the voltage, plasma treatment times and gas flow were 40 kV, 10 s and 60 mL/min, respectively. The predicted values were consistent with the measured values. The influence of each factors were voltage > time > gas flow rate.

Residues of fipronil and its metabolites in paddy fish soil were detected using high performance liquid chromatography tandem mass spectrometry (HPLC-MS/MS). Samples were extracted with oscillation and acetonitrile in constant temperature, and subsequently purified by solid phase extraction using ENVI 18 column and 10 mg GCB. The target compounds were separated on an Agilent ZORBAX Extend C18 column (2.1 mm× 100 mm×1.8 μm), and determined using electrospray ionization (ESI) source at negative mode with multiple reation monitoring (MRM). The analytes were quantified with external standard using the matrix matched standard calibration curve method. The results indicated that the average spiked recoveries of fipronil, fipronil desulfinyl, fipronilsulfone and fipronilsulfide in paddyfish soil were 83.5%~110.3% with the relative standard deviations (RSDs) of 1.9%~11.5%. Good linearities were obtained in the range of 0.5~100 μg/L, and the correlation coefficients were greater than 0.998, the limits of detection (LODs) were 1.0 μg/kg, and the limits of quantification (LOQs) were 3.0 μg/kg. This method is simple, accurate and reliable, which can meet the detection requirements of fipronil and its metabolites in paddy fish soil.

PDCoV is an emerging enteropathogenic coronavirus that mainly causes acute diarrhea in piglets, seriously affecting pig breeding industries worldwide. To date, the molecular mechanisms of PDCoV-induced immune and inflammatory responses or host responses in LLC-PK cells in vitro are not well understood. HSP90 plays important roles in various viral infections. In this study, HSP90AB1 knockout cells (HSP90AB1KO) were constructed and a comparative transcriptomic analysis between PDCoV-infected HSP90AB1WT and HSP90AB1KO cells was conducted using RNA sequencing to explore the effect of HSP90AB1 on PDCoV infection. A total of 1295 and 3746 differentially expressed genes (DEGs) were identified in PDCoV-infected HSP90AB1WT and HSP90AB1KO cells, respectively. Moreover, most of the significantly enriched pathways were related to immune and inflammatory response-associated pathways upon PDCoV infection. The DEGs enriched in NF-κB pathways were specifically detected in HSP90AB1WT cells, and NF-κB inhibitors JSH-23, SC75741 and QNZ treatment reduced PDCoV infection. Further research revealed most cytokines associated with immune and inflammatory responses were upregulated during PDCoV infection. Knockout of HSP90AB1 altered the upregulated levels of some cytokines. Taken together, our findings provide new insights into the host response to PDCoV infection from the transcriptome perspective, which will contribute to illustrating the molecular basis of the interaction between PDCoV and HSP90AB1.

(1) Background: Porcine deltacoronavirus (PDCoV) is a newly emerged enteric virus affecting pig breeding industries worldwide, and its pathogenic mechanism remains unclear. (2) Methods: In this study, we preliminarily identified the endocytic pathway of PDCoV in PK-15 cells, using six chemical inhibitors (targeting clathrin-mediated endocytosis, caveolae-mediated endocytosis, macropinocytosis pathway and endosomal acidification), overexpression of dominant-negative (DN) mutants to treat PK-15 cells and proteins knockdown. (3) Results: The results revealed that PDCoV entry was not affected after treatment with chlorpromazine (CPZ), 5-(N-ethyl-N-isopropyl) amiloride (EIPA)or ammonium chloride (NH4Cl), indicating that the entry of PDCoV into PK-15 cells were clathrin-, micropinocytosis-, PH-independent endocytosis. Conversely, PDCoV infection was sensitive to nystatin, dynasore and methyl-β-cyclodextrin (MβCD) with reduced PDCoV internalization, indicating that entry of PDCoV into PK-15 cells was caveolae-mediated endocytosis that required dynamin and cholesterol; indirect immunofluorescence and shRNA interference further validated these results. (4) Conclusions: In conclusion, PDCoV entry into PK-15 cells depends on caveolae-mediated endocytosis, which requires cholesterol and dynamin. Our finding is the first initial identification of the endocytic pathway of PDCoV in PK-15 cells, providing a theoretical basis for an in-depth understanding of the pathogenic mechanism of PDCoV and the design of new antiviral targets.

How to scientifically reflect the soil environmental quality of a large area through limited soil point monitoring is a major direction of current research. The Huaihe River Basin in Anhui Province was selected as the study area and soil environmental quality monitoring for five major soil pollution types during five consecutive years was conducted. The monitoring project includes Cd, Hg, As, Pb, Cr, Cu, Zn and Ni in the soil. The single pollution index and Nemerow pollution index methods were used to evaluate 426 monitoring sites. The Kriging interpolation method was used to plot the distribution of heavy metal elements in regional soil. In general, the soil environmental quality of the Anhui section of the Huaihe River Basin is relatively safe according to the evaluation results. 96.48% of the surveyed sites belong to clean and relatively clean levels, but some sites were severe. According to the distribution of heavy metal elements in the evaluation area, the overall content is low and the local area is slightly higher. The method of special type monitoring was applied and the characteristics of regional distribution through soil point monitoring results are discussed. The proposed method has great application potential. This method is suitable for preliminary investigation and research of large-scale environmental quality of soil.

Isobaric ions interfere with two isotopes of Ag during determination by Inductively Coupled Plasma-Mass Spectrometry (ICP-MS), i.e., 90Zr16O1H and 91Zr16O on 107Ag, and 92Zr16O1H and 93Nb16O on 109Ag. Although an interference correction coefficient method was used for the calibration of the analytical results, the analytical errors remain serious. In the proposed method, samples were closed decomposed by nitric acid, hydrofluoric acid, and perchloric acid. Ammonia was used to precipitate Zr and the purified Ag was determined by ICP-MS using 103Rh as the internal standard. The interference caused by the residual Zr was calibrated by the interference coefficient method. The method has a detection limit (3σ) of 4.1 ng/g and relative standard deviation (n=12) ranging from 2.5% to 7.8%. Determination of Ag in 53 primary certified materials of geochemical exploration indicates no loss of Ag. Using ammonium as the reagent, separation of Zr and Ag was successfully achieved due to the formation of the stable Ag-ammonium complex and Zr-hydroxide precipitate. The separation of Ag from Zr has reduced the interferences of Zr on Ag. Compared with the high-pressure sealed decomposition method and the P507 extraction resin separation method, the closed sample decomposition method is much simpler with less acid used and a lower blank.

To study the beneficial effects of ivabradine in dilated cardiomyopathy mice, which evolved from coxsackievirus B3-induced chronic viral myocarditis. Four-to-five-week-old male balb/c mice were inoculated intraperitoneally with coxsackievirus B3 (Strain Nancy) on day 1, day 14 and day 28. The day of the first virus inoculation was defined as day 1. Thirty-five days later, the surviving chronic viral myocarditis mice were divided randomly into two groups, a treatment group and an untreated group. Ivabradine was administered by gavage for 30 consecutive days in the treatment group, and the untreated group was administered normal saline. Masson’s trichrome stain was used to evaluate the fibrosis degree in myocardial tissue. The expression levels of tumor necrosis factor-α (TNF-α), interleukin-1β (IL-1β), interleukin-6 (IL-6), Collagen I, Collagen III and p38-MAPK signaling pathway proteins were detected by western blot. Electrocardiogram was used to investigate the heart rate and rhythm. The thickness of the ventricular septum and left ventricular posterior wall, left ventricular end diastolic dimension, left ventricular end systolic dimension, left ventricular ejection fractions and fractional shortening were studied by echocardiography. Compared with the untreated chronic viral myocarditis mice, ivabradine significantly increased the survival rate, attenuated the myocardial lesions and fibrosis, improved the impairment of the left ventricular function, diminished the heart dimension, decreased the production of collagen I and collagen III, reduced the expression of the proinflammatory cytokines TNF-α, IL-1β, and IL-6, and lowered the production of phospho-p38MAPK. The findings indicate the therapeutic effect of ivabradine in preventing the progression from viral myocarditis to dilated cardiomyopathy in mice with chronic viral myocarditis induced by coxsackievirus B3, is associated with inhibition of the p38MAPK pathway, downregulated inflammatory responses and decreased collagen expression. Ivabradine appears a promising approach for the treatment of patients with viral myocarditis.

This study investigated the compensating mediating mechanism of resilience in the relationship between loneliness and life satisfaction in Thai and Chinese college students during a risk period of coronavirus disease 2019 transmission. A total of 262 Thai and 247 Chinese college students were enrolled in the study between December 2020 and January 2021. A measurement model was constructed with the items of loneliness, resilience, and life satisfaction by using structural equation models. Subsequently, structural mediating models were constructed according to the valid items and a multigroup comparison was performed. The relationship between loneliness and life satisfaction differed significantly between Thai and Chinese college students as a result of the mediating effect of resilience. Resilience in Thai college students partly compensated for the negative influence of loneliness on life satisfaction, and resilience in Chinese college students compensated completely for the negative influence of loneliness on life satisfaction.

The stable slurry was prepared by ball-milling with a certain proportion of silica fume as silicon source,glucose as carbon source and metal niobium (Nb) as additives mixed with alcohol water. After the slurry was dried and pulverized, put the powder into a tubular furnace heated to 650℃ for 2h under nitrogen atmosphere to prepare precursor, the heating rate was 5℃/min. Treated the precursor under vacuum by carbothermal reduction method to prepare silicon carbide (SiC) powder. The effect of temperature and additive content on the morphology of synthesised silicon carbide and the impurity removal order on product purity were explored. After firing at 1300℃,1400℃,1500℃ for 2 h, the 3C-SiC powders are detected, and as the temperature increased, the crystallinity of the product become better. When the content of the additive is 1% of the quality of the silica fume, the particle size of the silicon carbide synthesized at the temperature of 1500℃ is even and the dispersion is better. As for the impurity removal order, removed SiO2 first, then removed C can effectively remove the impurities in the product.

Parnassia wightiana Wall. ex Wight & Arn., a perennial herb, is an alpine plant distributed across three biodiversity hotspots in China. This species offers an excellent study system to analyze the distribution pattern and genetic structure of high mountainous plant populations. Three chloroplast DNA (cpDNA) regions (rpl32-trnL, trnL-F, and trnS-G) from 442 individuals of 39 populations and internal transcribed spacers (ITS) from 418 individuals of 38 populations were sequenced and analyzed. We identified a total of 26 haplotypes based on the concatenated sequences of cpDNA and a total of 36 based on ITS sequences. A hierarchical analysis of molecular variance indicated significant phylogeographic structure among populations. Mismatch analysis of multipeak and Tajima's D neutral test demonstrated Parnassia wightiana populations within China had not experienced abrupt expansion recently. The allopatric fragmentation resulted in the geographic isolation due to environmental heterogeneity, producing a high proportion of private haplotypes in the distribution of Parnassia wightiana within China. Key words: Parnassia wightiana Wall. ex Wight & Arn., cpDNA, ITS, genetic diversity, phylogeography, Introduction Climatic oscillations during the Cenozoic period, especially the Quaternary glacial and interglacial cycle, shaped the distribution of species (Hewitt 2004). Phylogeography is a discipline that deals with the spatial [...]