重组高产H1N1亚型猪流感疫苗株的构建与鉴定

为制备一株猪流感灭活疫苗的候选毒株,本研究利用反向遗传学技术,将A/swine/Guangdong/1/11(H1N1)毒株的HA、NA基因和A/PR/8/34毒株的PB2、PB1、PA、NP、M、NS基因进行重组,成功拯救出鸡胚高度适应性毒株rH1N1。抗原性分析显示rH1N1保留了亲本毒株良好的抗原性。rH1N1接种10日龄鸡胚48 h后,血凝价达到210,表明该毒株能在鸡胚中高水平复制。该毒株为H1N1亚型猪流感灭活疫苗的研制奠定了坚实的物质基础。     

番鸭细小病毒VP3基因的原核表达及多克隆抗体的制备

为获得针对番鸭细小病毒VP3蛋白的多克隆抗体,根据已发表的该病毒基因序列设计一对引物,利用PCR扩增出VP3基因,将其克隆到原核表达载体p ET-32a,转化感受态细胞BL21(DE3)。经IPTG诱导后,SDS-PAGE检测重组蛋白。将重组蛋白切胶免疫BALB/c小鼠,制备抗番鸭细小病毒部分结构蛋白的多克隆抗体。结果显示成功获得VP3基因,构建的原核表达重组质粒成功表达预期的重组蛋白,免疫小鼠获得的多克隆抗体能与番鸭细小病毒及VP3蛋白反应。表明制备的多克隆抗体可用于MDPV的检测,并为进一步研究MDPV奠定了基础。     

Effects of Compound Plant Nutrients on Rumen Fermentation Parameters,Rumen Microbiota and Fatty Acid Composition of Longissimus dorsi Muscle in Finishing Small Tail Han Sheep北大核心CSCD

This study was conducted to investigate the effects of compound p1ant nutrients (CPN) on rumen fermentation parameters, rumen microbiota and fatty acid composition of longissimus dorsi musc1e in finishing sma11 tai1 Han sheep. Sixteen 4-month-o1d finishing sma11 tai1 Han sheep with an initia1 body weight (BW) of (24.18±0.31) kg were random1y divided into two groups, name1y, with 8 rep1icates per group and 1 sheep per rep1icate. The sheep in the contro1 group were fed a basa1 diet, whereas the sheep of the contro1 group (CPN group) was fed the basa1 diet supp1ementation with 3‰ CPN. The experiment 1asted for 97 days after 7 days adaption. The resu1ts showed as fo11ows: compared with the contro1 group, 1) adding CPN decreased the concentration of ammonia nitrogen (NH3-N) (P<0.05); 2) CPN supp1ementation affected beta diversity of rumen microbiota; 3) the re1ative abundance of Acidobacteriota, Erysipe1otrichaceae_UCG-002, Lactobacillus, and Megasphaera were enhanced by adding CPN (P<0.05), whereas, the re1ative abundance of Bacteroidetes and Prevotella was 1ower (P<0.05); 4) the supp1ementation of CPN had no significant effect on fatty acid composition of longissimus dorsi musc1e of fattening sma11-tai1ed Han sheep (P>0.05); 5) the contents of tota1 MUFA, C18:1n9c, C14:1, C16:1, C18:2n6c, C18:3n3 and n-3PUFA in the longissimus dorsi musc1e were corre1ated with the re1ative abundance of Megasphaera, Erysipe1otrichaceae_UCG-002, Succiniclasticum and Ruminococcus (P<0.05). In conc1usion, CPN can regu1ate the rumen microbiota structure and reduce the rumen NH3-N concentration of fattening sma11-tai1ed Han sheep. In production practice, CPN can be used as a rumen eco1ogica1 regu1ator. © 2023 Authors. All rights reserved.     

The effects of 17β-estradiol on cardiomyocyte hypertrophy induced by endothelin

AIM:To investigate the effect of 17β-estradiol (E₂) on myocardial hypertrophy induced by endothelin-1(ET-1) and the related mechanism. METHODS:Myocardial cells from neonate rats were cultured in vitro and myocardial hypertrophy model was established with ET-1.The effects of 17β-estradiol on myocardial hypertrophy were observed. The role of ERK1/2 in the effects of 17β-estradiol was also detected. RESULTS:Compared with control group,ET-1 increased cell protein content,cell surface area and ［³H］^-Leucine ［³H］^-Leu) incorporation. Pretreatment with E₂ for 24 h could inhibit the increase in cell protein content,cell surface area and ［³H］^-Leu incorporation induced by ET-1.ET-1 significantly stimulated ERK1/2 activity,which was prevented by pretreatment with E₂.Tamoxifen,estradiol receptor antagonist,partially inhibited the effect of E₂. The ability of ET-1 to stimulate ［³H］^-Leu incorporation was significantly blocked by PD98059,which could enhance the inhibitory effect of E₂ on the increase of ［³H］^-Leu incorporation in cardiomyocytes induced by ET-1.CONCLUSION:E₂ can inhibit cardiomyocyte hypertrophy induced by ET-1. This effect is mediated by estrogen receptor. ERK1/2 signal pathway is closely correlated with the inhibitory effect of E₂ on cardiomyocyte hypertrophy induced by ET-1.     

Probiotic preparation reduces the faecal water genotoxicity in chickens fed with aflatoxin B1 contaminated fodder

The aim of the study was to evaluate the influence of a probiotic preparation on the genotoxicity of faecal water of broiler chickens fed with a fodder contaminated with aflatoxin B₁ (AFB₁) at 1 or 5 mg per kg. Human blood lymphocytes were exposed to chicken’s faecal water samples and DNA damage was measured using the comet assay. Genotoxicity of faecal water did not depend on the AFB₁ concentration in the fodder. The mean DNA damage, measured as the percentage of DNA in the tail of the comets, for chickens fed with fodder with AFB₁ at 1 mg/kg was 16.80 ± 0.66, at 5 mg/kg – 16.73 ± 1.51 and in the controls – 12.79 ± 0.66. The supplementation of fodder with the probiotic preparation decreased the extent of DNA damage to 10.02 ± 0.39 for 1 mg/kg AFB₁ and to 11.89 ± 0.72 for 5 mg/kg.     

The impact of weight loss on circulating cytokines in Beagle dogs

Chronic low-grade inflammation in obesity is characterized by an increased production of pro-inflammatory and chemotactic cytokines that are contributing to insulin resistance and related co-morbidities. Cytokines act in networks and exhibit pleiotropic effects so we investigated the circulating levels of a wide array of cytokines (pro and anti-inflammatory, chemotactic and growth factors) in a canine model of weight loss. The dogs served as their own control in order to study the impact of weight loss independent of potential confounding factors, such as history of excess weight or gender. While low-grade inflammation had been previously investigated in obese dogs by measuring changes in adipokines, acute phase proteins and key pro-inflammatory cytokines, to the best of our knowledge this is the first study to evaluate how weight loss impacts a wide array of circulating cytokines.Eighteen overweight Beagle dogs were recruited (six spayed females and 12 neutered males), and none of them were grossly obese according to the body condition score (BCS). All the dogs reached an ideal weight by the end of the program. Parameters were assessed before (baseline), at mid-point (month 3) and at end-point (month 6). Plasma GM-CSF, IL-2, Il-4, IL-6, IL-7, IL-8, IL-10, IL-15, IL-18, IFNγ, IP-10, TNFα, monocyte chemotactic protein 1 (MCP-1), keratinocyte chemokine (KC) were measured with canine multiplex immunoassays. Fat mass was assessed by dual energy X-ray absorption (DEXA).Several cytokines decreased throughout the weight loss program (p < 0.01) and were correlated with the percentage of fat measured by DEXA (p < 0.05): chemotactic (MCP-1), growth factors (GM-CSF, IL-7 and IL-2), and pro-inflammatory (KC and IL-18). We could not show trends for several cytokines, possibly because their level may be lower than the assay sensitivity: anti-inflammatory (IL-4 and IL-10), and pro-inflammatory (IL-6 and TNFα).In conclusion, while our findings for several pro-inflammatory and chemotactic cytokines are in accordance with human and rodent studies, we may have identified additional cytokines, such as growth factors, related to obesity-induced low-grade inflammation. Considering the weight loss was enabled by an adjusted diet, the role of this association of cytokines in insulin resistance and related co-morbidities needs to be clarified. Our results could help better understand the cytokine biology in dogs, and as such are relevant for further elucidating the relationship between immune function and metabolism/nutrition.     

Effect of miR-181b on cell proliferation and expression of Bcl-2 and SP1 in HepG2 cells

AIM: To investigate the role of miR-181b in the expression of Bcl-2 and SP1 at mRNA and protein levels in the human hepatoma G2 cells (HepG2), and to explore the effect of miR-181b on the regulation of HepG2 cell proliferation. METHODS: The synthetic double-strand complementary DNA based on the sequence of miR-181b was inserted into the vector of miRNASelect^TM pEGP-miR. The microRNA high-expression plasmid was cloned, and the sequences were identified. The miR-181b plasmid was transfected into HepG2 cells with liposomes. The stable cell line was screened by puromycin. The mRNA and protein levels of Bcl-2 and SP1 were measured by RT-PCR and Western blotting, respectively. Methyl thiazolyl tetrazolium (MTT) method was used to analyze the proliferation of HepG2 cells. RESULTS: The Western blotting results showed that miR-181b inhibited the protein expression of Bcl-2 and SP1. The result of RT-PCR also indicated that the mRNA expression of Bcl-2 and SP1 was suppressed. Compared with the control, the growth rate of HepG2 with high expression of miR-181b was significantly decreased.CONCLUSION: miR-181b inhibits the proliferation of HepG2, which may be related to the down-regulation of Bcl-2 and SP1.     

猪抗酶基因1(AZ1)5’调控区单核甘酸多态性与屠宰性状的相关性分析

通过PCR-RFLP方法检测了长白与蓝塘猪F2代群体抗酶基因1(AZ1)5’调控区-792～-433单核苷酸多态性,共获得3个SNP:-731C＞T,-584A＞C和-476G＞A.-713位点处T为优势等位基因,频率为0.777,优势基因型TT频率为0.598;-584位点处A为优势等位基因,频率为0.769,优势基...     

Negligible contribution from roots to soil-borne phospholipid fatty acid fungal biomarkers 18:2ω6,9 and 18:1ω9

The phospholipid fatty acid biomarkers 18:1ω9, 18:2ω6,9 and 18:3ω3,6,9 are commonly used as fungal biomarkers in soils. They have, however, also been found to occur in plant tissues, such as roots. Thus, the use of these PLFAs as fungal biomarkers in sieved soil, which may still contain small remains of roots, has been questioned. We used data from a recent beech tree girdling experiment to calculate the contribution of roots to these biomarkers and were able to demonstrate that not more than 0.61% of 18:1ω9 and 18:2ω6,9 in sieved soil samples originated from roots (but 4% of 18:3ω3,6,9). Additionally, the abundance of the biomarker 18:2ω6,9 in the soil was found to be highly correlated to ectomycorrhizal root colonization, which further corroborates its fungal origin. PLFA biomarkers were substantially reduced in vital roots from girdled trees compared to roots of control trees (by up to 76%), indicating that the major part of PLFAs measured in roots may actually originate from ectomycorrhizal fungi growing inside the roots. We calculated, that even a near to 50% reduction in fine root biomass - as observed in the girdling treatment - accounted for only 0.8% of the measured decrease of 18:2ω6,9. Our results demonstrate that both 18:1ω9 and 18:2ω6,9 are suitable biomarkers for detecting fungal dynamics in soils and that especially 18:2ω6,9 is a reliable biomarker to study mycorrhizal dynamics in beech forests.     

香蕉枯萎病菌myosin-1基因的功能分析及外施dsRNA介导的抗性评估

 香蕉枯萎病是由尖孢镰刀菌古巴专化型(Fusarium oxysporum f. sp. cubense,Foc)引起的香蕉毁灭性土传病害,其中 4号生理小种(Foc4)能感染几乎所有的香蕉品系,危害最严重。SMART在线软件分析myosin-1基因具有肌球蛋白马达蛋白(myosin motor domain, MMD),肌动蛋白尾结构TH1(myosin tail)和 Src家族同源结构域SH3(src homology domain 3),与禾谷镰刀菌中氰烯菌酯靶标基因myosin-5具高度的蛋白同源性,相似性高达83%。利用Split-marker基因重组技术获得Foc4的myosin-1基因敲除突变体,Δmyosin-1突变株丧失了对氰烯菌酯的敏感性,菌丝生长缓慢,产孢量减少且孢子畸形,对香蕉致病力严重下降,证实myosin-1是氰烯菌酯在Foc4中的作用靶标基因。外施靶向myosin-1体外转录的dsRNA,能抑制菌丝的生长,降低菌丝活性;菌丝膨胀扭曲分枝增多,出现典型的球状结构,与氰烯菌酯处理后的表型一致。在盆栽活体人工接种实验中,体外施用dsRNA可以明显抑制枯萎病外部症状的发展,推迟发病时间,赋予寄主抗性,结果说明体外施用dsRNA可以作为新型杀菌剂防治香蕉枯萎病。综上,myosin-1基因作为氰烯菌酯在Foc4中的靶标基因具有高度的序列保守,在调控菌丝生长发育,产孢以及致病力等方面发挥重要作用,而外施dsRNA具有防治香蕉枯萎菌的巨大潜力。