Immunomodulation by dietary vitamin C in healthy and aflatoxin B1-induced immunocompromised rohu (Labeo rohita)

The aim of this study was to examine the immunomodulatory effect of high levels of dietary vitamin C in healthy and immunocompromised rohu (Labeo rohita) treated with aflatoxin B1 (AFB1). Four groups of rohu were fed experimental diets containing either no vitamin C or supplemented with vitamin C at 500 ppm for 60 days. On the first day of feeding, one group fed the high vitamin C diet and one fed the vitamin C deficient diet, were injected intraperitoneally with a single doses of AFB1 at 1.25 mg kg(-1) body weight. The effect of AFB1 and high dietary vitamin C on specific and non-specific immunity, and disease resistance against Aeromonas hydrophila were examined in the rohu. The ability of vitamin C to counteract immunosuppression induced by AFB1 was also examined. Specific immunity indicated by haemagglutination and haemolysin titres against sheep red blood cells (SRBC), and bacterial agglutination appeared to be unaffected by either the AFB1 treatment or the vitamin C enriched diet. A significant reduction was observed in the non-specific immunity of AFB1-treated fish, however, indicated by lowered bactericidal and lysozyme activities. High dietary vitamin C, on the other hand, enhanced the non-specific immunity of fish, including an enhanced phagocytic ratio and increased serum lysozyme activity. Feeding a high level of dietary vitamin C to AFB1-treated fish increased these parameters to levels similar to those found in control fish. High dietary vitamin C significantly (p < 0.05) enhanced protection against Aeromonas hydrophila infection in both healthy and immunocompromised fish. Results from this study help to establish the beneficial effect of dietary vitamin C on AFB1-induced immunosuppression, as well as confirming the immunostimulatory effect of vitamin C in rohu.     

Evaluation of emamectin benzoate for the control of experimentally induced infestations of Argulus sp. in goldfish and koi carp

The effect of 0.2% emamectin benzoate (SLICE; Intervet/ Schering-Plough Animal Health, Roseland, New Jersey) administered in top-dressed, pelleted commercial fish feed was evaluated for control of freshwater Argulus sp. in goldfish Carassius auratus and koi carp, a variant of common carp Cyprinus carpio, in freshwater aquaria at 24-25 degrees C. Sixteen individually housed goldfish were each exposed to 37 Argulus. The number of fish lice attached to each fish at the start of the experiment was not determined; however, the total number of motile fish lice in each aquarium (on fish and in the water) was determined at the start and end of each experiment. Eight goldfish were fed the control diet (0 microg x kg fish biomass(-1) x d(-1)) and eight were fed the medicated diet (50 microg x kg fish biomass(-1) x d(-1)) for seven consecutive days. After treatment, fish louse infestation in controls was 20.5 +/- 1.5 (mean +/- SE) lice per fish. No Argulus were found on fish in the treated group. In a separate experiment, 10 individually housed koi were each exposed to 128 Argulus. Five koi were fed the control diet and five were fed a low-dose medicated diet (5 microg x kg fish biomass(-1) x d(-1)) for 7 d. After treatment, fish louse infestation among the controls was 14.6 +/- 3.8 lice per koi. No Argulus were found on koi in the treated group. Hence, a 7-d regimen of oral emamectin benzoate controlled experimental infestation of Argulus when administered to goldfish at 50 microg x kg fish biomass(-1) x d(-1) and to koi at 5 microg x kg fish biomass(-1) x d(-1).     

Effects of Ergosan on the expression of cytokine genes in the liver of juvenile rainbow trout (Oncorhynchus mykiss) exposed to enteric red mouth vaccine

The effects of administration of the immunomodulator Ergosan, an algal extract containing alginic acid, in juvenile rainbow trout (Oncorhynchus mykiss) exposed to AquaVac vaccination, were tested. Juveniles treated with Ergosan, 95 days after the beginning of first solid feeding and control fish fed solely on commercial diet, were vaccinated by immersion in AquaVac solution. The time-course of the effects of vaccination on liver immunorelated gene modulation and on the tolerance to stress manipulation connected with the vaccination was investigated. Liver and plasma sampling was performed at the following times: T=pre-vaccination, T0=5min, T1=2h, T2=8h, T3=24h, T4=48h and T5=72h post-vaccination. Interleukin-1beta (IL-1beta), interleukin-8 (IL-8), tumor necrosis factor alpha 2 (TNF alpha 2) and heat shock protein 70 (Hsp70) gene expression in trout liver was monitored by real-time PCR using Acidic Ribosomal Phosphoprotein P0 (ARP) as internal standard. The evaluation of the plasma cortisol levels was performed by EIA. In AquaVac-vaccinated fish, both the gene expression of Hsp70 and the plasma cortisol levels during the time-course were significantly (P<0.05) lower in Ergosan-treated fish with respect to control, indicating the positive role of Ergosan on handling stress tolerance. This study also demonstrated the stimulatory properties of Ergosan on cytokine genes expression involved in innate immune response: liver IL-1beta, IL-8 and TNF alpha 2 gene expression was significantly (P<0.05) higher in trout fed on Ergosan compared to control, indicating a positive role of this feed additive in improving the immune responsiveness to AquaVac vaccine.     

氨氮胁迫下饥饿与复投喂对黄颡鱼肝脏中脂质代谢相关酶活性及相关基因表达的影响

为了研究氨氮胁迫下饥饿与复投喂对黄颡鱼脂质代谢的影响,将360尾黄颡鱼幼鱼[初始体重为(14.95±0.03)g]随机分配到12个养殖桶中,每桶30尾.对照组人工饱食投喂42 d,试验组饥饿14 d后恢复投喂28 d.对照组和试验组分别被暴露到总氨氮浓度为0(低氨氮处理,正常养殖环境)和5.70 mg/L(高氨氮处理,...     

RNAi suppression of zebrafish peptidoglycan recognition protein 6 (zfPGRP6) mediated differentially expressed genes involved in Toll-like receptor signaling pathway and caused increased susceptibility to Flavobacterium columnare

In Drosophila, Toll signaling cascade, which resembles the mammalian Toll-like receptor (TLR)/IL-1R signaling pathways and regulates the expression of anti-microbial peptide genes, mainly relies on peptidoglycan recognition proteins (PGRPs) for the detection of bacterial pathogens. To explore the effect of zebrafish peptidoglycan recognition protein 6 (zfPGRP6) on Toll-like receptor signaling pathway, RNA interference (siRNA) and real time quantitative PCR (RQ-PCR) methods were used to identify differentially expressed genes regulated by zfPGRP6. The target genes included TLR2, TLR3, TLR5, TLR7, TLR8, IL1R, Sterile-alpha and Armadillo motif containing protein (SARM), myeloid differentiation factor 88 (MyD88) and nuclear factor (NF)-kappa B2 (p100/p52). The results of RQ-PCR showed that RNAi-mediated suppression of zfPGRP6 significantly down-regulated the expression of TLR2, TLR5, IL1R, SARM, MyD88 and p100/p52. The expression of beta-defensin-1 was also down-regulated in those embryos silenced by zfPGRP6. In challenge experiments to determine the anti-bacterial response to Gram-negative bacteria, RNAi knock-down of zfPGRP6 markedly increased susceptibility to Flavobacterium columnare.     

小肽对大黄鱼仔鱼生长和小肠发育的影响及其在低温胁迫下的抗氧化应激效应

本试验分为生长试验和低温胁迫试验2部分.先分别投喂大黄鱼(Larimichthys cro-cea)仔鱼经不同浓度(0、0.5、1.0、2.0和3.0 mL/m3)小肽营养强化后的轮虫和卤虫12 d,以探讨小肽对大黄鱼仔鱼生长和小肠发育的影响;再将大黄鱼仔鱼暴露在温度为12℃的水体中24 h,以探讨小肽对低温胁迫下大黄...     

Expression of Toll-like receptors and downstream genes in lipopolysaccharide-induced porcine alveolar macrophages

The aim of the present study was to determine the age-related kinetic changes of Toll-like receptors (TLRs) and downstream genes expression, and secretion of cytokine in lipopolysaccharide (LPS) stimulated porcine alveolar macrophages (AM). For this purpose, AMs were isolated from 5-day-old newborn piglets and 120-day-old young pigs. mRNA expression and cytokine measurement was determined by quantitative real-time PCR and ELISA, respectively. First, AMs were incubated for 24 h in the absence or presence of increasing concentrations of LPS. Results showed the up-regulation of TLRs 2, 4, 5 and 9 mRNA from all concentrations of LPS used, as compared to non-stimulated cells, and TLR4 was the highest expression in both ages (P<0.05). Furthermore, quantitative analysis demonstrated increased expression of mRNAs encoding TLRs 2, 4, 5 and 9, LBP, CD14, MD2, MyD88, IRAK4 and TRAF6 in both ages in a time-dependant manner (P<0.05). Overall, LPS inducible mRNA for TLR4, LBP, CD14 and MyD88 had higher expression in newborn piglets compared with those of young pigs (P<0.05). The level of cytokine protein IL6 and TNFα in supernatant fluid significantly varied with time of incubation and age of animals. Their concentration increased immediately at 1 h after LPS stimulation and remained significantly higher up to 48 h in both ages. Production of pro-inflammatory cytokine protein IL6 and TNFα in supernatant was significantly higher in young pigs than those of piglets. This study suggests that differential age-related changes in the expression of TLRs and downstream genes, and pro-inflammatory cytokine could contribute to a different age-related innate immune response during pulmonary infection. Further investigation is warranted to determine the precise effects of LPS on porcine AMs by means of a functional study across a wider age range.     

军曹鱼幼鱼维生素C需要量的研究

选用初体均重为25.85 g的军曹鱼幼鱼225尾随机分为5组,每组设3个重复,每个重复15尾.分别投喂添加5种不同剂量维生素C(添加量分别为0、37.5、75、150和300 mg/kg)的试验饲料,试验期为56 d,观察维生素c对军曹鱼幼鱼生长、组织维生素C积累量及非特异性免疫的影响.结果表明,随着饲料中维生素C添加量增加,军曹鱼幼鱼增重率和特定生长率呈现出上升后下降趋势,在75 mg/kg时达到最高值,显著高于0和300 mg/kg组(P<0.05).全鱼粗蛋白质含量随着饲料中维生素C添加量的升高亦呈现先上升后下降趋势,在75 mg/kg时达到最高值,全鱼干物质、粗蛋白质、粗脂肪、灰分含量在各组之间均无显著差异(P>0.05).血清维生素C积累量随着饲料中维生素C添加量的增加呈现先上升后下降趋势,在150 mg/kg时达到最高值且显著高于0 mg/kg组(P<0.05);脑和肝脏中维生素C含量在300 mg/kg时达到最高值.300 mg/kg组脑中维生素C含量显著高于0,37.5和75 mg/kg组(P<0.05),肝脏中维生素C含量显著高于0,37.5和150 mg/kg组(P<0.05).随着饲料中维生素C添加量的增加,溶菌酶活性呈现先上升后下降趋势,在75 mg/kg达到最高,显著高于0,37.5和300 mg/kg组(P<0.05);血清总抗氧化能力呈现上升趋势,75、150和300 mg/kg组显著高于0 mg/kg组(P<0.05);肾抗氧离子自由基在150 mg/kg组达到最高,但各组之间无显著差异(P>0.05).以鱼体增重率和血清溶菌酶含量为指标,经折线模型回归分析求得军曹鱼幼鱼饲料中维生素C的适宜添加量分别为70和80.72 mg/kg.     

亚麻籽油对脂多糖刺激仔猪肝脏Toll样受体4和核苷酸结合寡聚化结构域信号通路关键基因表达的影响

本试验旨在研究亚麻籽油对脂多糖(LPS)刺激仔猪肝脏Toll样受体4(TLR4)和核苷酸结合寡聚化结构域(NOD)信号通路关键基因表达的影响。选取24头断奶仔猪,按体重相近原则随机分为4个组,分别为对照组、LPS组、2.5%亚麻籽油组(2.5%亚麻籽油+LPS)、5.0%亚麻籽油组(5.0%亚麻籽油+LPS),每组6个重复,每个重复1头猪,试验期21 d。试验组注射100μg/kg体重的LPS,对照组注射等量的生理盐水。注射LPS或生理盐水4 h后屠宰仔猪,取肝脏,测定TLR4和NOD信号通路关键基因及相关炎性介质的mRNA表达水平。结果表明:1)LPS刺激显著提高了肝脏肿瘤坏死因子-α(TNF-α)、环氧酶2(COX2)、热休克蛋白70(HSP70)的mRNA相对表达量(P0.05),2.5%亚麻籽油可显著降低COX2、TNF-α的mRNA相对表达量(P0.05),5.0%亚麻籽油可显著降低TNF-α的mRNA相对表达量(P0.05)。2)LPS刺激显著提高了肝脏TLR4、髓样分化因子88(My D88)、白细胞介素-1受体相关激酶1(IRAK1)、NOD1、NOD2、受体互作蛋白2(RIPK2)、核因子-κB(NF-κB)的mRNA相对表达量(P0.05);2.5%亚麻籽油可显著降低NOD1、NOD2的mRNA相对表达量(P0.05),有降低RIPK2 mRNA相对表达量的趋势(0.05≤P0.10);5.0%亚麻籽油可显著降低NOD2的mRNA相对表达量(P0.05)。这表明LPS刺激导致仔猪发生炎症反应,亚麻籽油可能通过抑制NOD信号通路进而缓解肝脏炎症反应。     

共轭亚油酸对大黄鱼生长和免疫指标的影响

本试验旨在研究共轭亚油酸(CLA)对大黄鱼(Pseudosciaena crocea,R)生长和免疫指标的影响.选用初体重为(146.25±10.62) g的大黄鱼1 600尾,随机分为4组,每组2个重复,每个重复200尾,分别饲喂在基础日粮中添加了0%、1%、2%和4%共轭亚油酸的试验日粮,试验共进行10周.结果表明,与未添加共轭亚油酸组相比,各共轭亚油酸添加组的饵料系数(FE)、特定生长率(SGR)、肥满度(CF)、肝体比(HSI)和脏体比(VSI)未产生显著影响(P>0.05).添加共轭亚油酸能够显著提高血清中溶菌酶、免疫球蛋白M(IgM)、血清超氧化物歧化酶(SOD)和肝脏SOD活性(P<0.05),并且随共轭亚油酸含量的升高,活性也升高.血清补体(C3、C4)活性2%和4%共轭亚油酸组显著提高(P<0.05),且以2%组活性最高,并显著高于1%和4%组(P<0.05);肌肉SOD 2%和4%共轭亚油酸组显著提高(P<0.05),且以4%组活性最高,但与1%和2%组差异不显著(P>0.05).由此得出,日粮中添加1%～4%CLA不能提高大黄鱼的生长性能,但可显著提高大黄鱼的机体免疫能力.[动物营养学报,2009,21(2):205-211][中文全文见<动物营养学报>网站(www.ChinaJAN.com)中文版2009年21卷2期]     

TLRs介导的信号通路在马链球菌感染RAW264.7细胞中的作用

为探究Toll样受体(TLRs)介导的信号通路在马链球菌马亚种(S.equi)感染小鼠巨噬细胞RAW264.7中的作用,收集S.equi感染后不同时间点的RAW264.7细胞,提取总RNA并反转录成cDNA,利用实时荧光定量PCR技术检测细胞Toll样受体1、2、6(TLR1、TLR2、TLR6)、接头蛋白骨髓分化蛋白88(MyD88)及细胞因子IL-1、IL-6、IL-10、IL-12、TNF-αmRNA的表达情况。结果显示,S.equi感染RAW264.7细胞后6h时,TLR1、TLR2、TLR6与MyD88mRNA水平均较对照组没有显著差异(P>0.05);感染后12h时,TLR1、TLR2和TLR6mRNA表达量未出现明显上升(P>0.05),而MyD88mRNA水平极显著升高(P<0.01);感染后24h时,TLR1、TLR2和TLR6mRNA表达水平出现极显著升高(P<0.01),MyD88mRNA表达没有显著变化(P>0.05),且IL-10和IL-12mRNA水平与对照组相比极显著升高(P<0.01),IL-1、IL-6和TNF-αmRNA水平均极显著下降(P<0.01)。结果表明,TLRs介导的信号通路参与S.equi感染RAW264.7细胞的免疫应答反应。     

虹鳟酪氨酸酶基因的生物信息学分析及其在不同发育阶段和组织中的表达分析

酪氨酸酶（tyrosinase,Tyr）基因在调控动物色素合成的过程中发挥关键作用。为了解tyr基因与虹鳟（Oncorhynchus mykiss）体色变异的关系,本研究对虹鳟tyr-1和tyr-2基因进行了蛋白序列分析,并利用实时荧光定量PCR检测两种基因在野生型虹鳟（虹鳟）和黄色突变型虹鳟（金鳟）体色发生的19个不同时期和成鱼13种不同组织中的表达差异。蛋白序列分析结果表明,Tyr-1和Tyr-2都是亲水性蛋白,且蛋白结构主要是无规则卷曲和α-螺旋;实时荧光定量PCR结果显示,tyr-1基因在胚胎各时期均有表达,tyr-1基因表达量在虹鳟受精期最高,桑椹期次之,且显著高于其他时期（P<0.05）;tyr-1基因表达量在金鳟16-细胞期最高,且显著高于其他时期（P<0.05）;tyr-2基因分别从虹鳟囊胚期和金鳟原肠期开始表达,表达量均在心跳期达到最高且显著高于其他时期（P<0.05）。出膜后,tyr-1、tyr-2基因分别在虹鳟5日龄和金鳟3月龄表达量最高。在胚胎各时期中,tyr-1、tyr-2基因在虹鳟中表达量普遍高于金鳟,出膜后普遍低于金鳟。在各组织中,tyr-1基因在皮肤、眼睛、肾脏、肝脏等组织中有较高表达,tyr-2基因主要在皮肤、眼睛中有较高表达,在其他组织中表达量较低。以上结果表明,tyr-1和tyr-2基因与虹鳟体色变异具有一定的相关性,为深入阐明虹鳟体色变异机制和体色遗传改良奠定了理论基础。     

麝鼠TNFRSF13 C和TNFRSF4基因表达分析

麝鼠(Ondatra zibethicus)是一种珍贵的毛皮动物和香料动物,在繁殖季节能分泌麝鼠香,而其香腺泌香机制尚不清楚。TNFRSF13 C(BAFF受体)和TNFRSF4(OX40受体)基因是TNF受体超家族的成员,参与免疫调节、炎症、凋亡,还与自身免疫病和多种器官的形成有关。本研究通过RT-qPCR技术检测繁殖期与非繁殖期麝鼠的心、肝、肺、肾、睾丸、肌肉及香腺中TNFRSF13 C和TNFRSF4基因的转录水平,结果显示两者在繁殖期和非繁殖期7个组织中均有表达,在繁殖期麝鼠的心、肝、肺、肾、睾丸、香腺中表达水平普遍高于非繁殖期,并且差异显著(P≤0.05),而在肌肉中无明显表达差异(P>0.5)。由此推测TNFRSF13 C和TNFRSF4基因参与了麝鼠心、肝、肺、肾、睾丸和香腺发育。     

中华鲟幼鲟维生素C营养需要的研究

本研究旨在确定中华鲟幼鲟对维生素C(VC)的营养需要量。配制含VC分别为0.5、59.8、119.5、239.1、478.1和956.2 mg/kg饲料的6组日粮(分别编号为C0.5、C59.8、C119.5、C239.1、C478.1和C956.2),连续投喂中华鲟幼鲟8周后,以生长速度、组织VC含量和部分相关酶活性来确定中华鲟对VC的需求量。结果表明,C0.5和C59.8组鲟鱼的增重率、饲料效率、蛋白质效率都低于C119.5组,进一步提高饲料VC含量,也没有显著变化(P>0.05);C478.1组的肝脏和肌肉组织VC含量达最高水平;C239.1组血清溶菌酶活性最高;C239.1组SOD活性最低。根据平均增重率和饲料效率做折线法分析表明,维持中华鲟最佳生长的饲料VC含量为108.5 mg/kg;肝脏中VC含量达最大时饲料VC添加量为309.4 mg/kg;肌肉中VC含量达最大时饲料VC添加量为273.3 mg/kg。     

mRNA profile of Nellore calves after primary infection with Haemonchus placei

Haemonchus parasites are responsible for many losses in animal production. However, few studies are available, especially of zebu cattle. In this study, we investigated mRNA differences of immune response genes in na?ve Nellore calves infected with Haemonchus placei, relating these differences to patterns of cellular infiltrate. Calves were infected with 15,000 H. placei L3 larvae and after 7 days lymph node and abomasum tissues were collected. IL-2, IL-4, IL-8, IL-12, IL-13, IFN-γ, MCP-1, lysozyme, pepsinogen and TNF-α genes were evaluated by qPCR. Mast cells, eosinophils and globular leukocytes were counted by abomasum histology. In the infected group, IL-4, IL-13 and TNF-α were up-regulated in the abomasal lymph node. In the abomasum, IL-13 increased and TNF-α was down-regulated (p<0.05). No differences were detected for mast cells and eosinophil counts in abomasal tissue (p>0.05). We conclude that for this infection time, there was Th2 polarization but that cellular infiltrate in abomasal tissue takes longer to develop.     

Relationships between the first ovulation postpartum and polymorphism in genes relating to function of immunity, metabolism and reproduction in high-producing dairy cows

The decrease in fertility and conception rates of high-producing dairy cows is one of the major negative impacts for today's producers. The recovery of ovarian activity postpartum is affected by the status of immunity, metabolism and reproduction and plays a critical role in subsequent fertility after parturition in the cow. In the present study we investigated the relationships between polymorphisms in genes relating to the above functions and the first postpartum ovulation as a marker of the recovery of ovarian function in the cow. In immune function related-factors, the occurrence of first postpartum ovulation within 3 weeks in the C/C genotypes of tumor necrosis factor α (TNFα) exon (55.4%) and the A/G genotypes of TNFα promoter (55.4%) was significantly higher than that in T/T genotypes of TNFα exon (14.3%) and A/A genotypes of TNFα promoter (14.3%). Moreover, anovulatory cows with the T/T genotype of TNFα exon and the A/A genotype of TNFα promoter tended to have a prolonged days open compared with those of the other genotypes of TNFα polymorphisms. In metabolic function-related factors, ovulatory and anovulatory cows had a different distribution for alleles of the growth hormone receptor, but there were no significant differences in genotype and allele frequency of insulin-like growth factor-I polymorphism. No significant relationships were found between ovarian function after parturition and polymorphisms for reproduction-related genes. In conclusion, polymorphisms of TNFα gene both in exon and promoter regions have a strong association with the early first ovulation within 3 weeks after parturition in the high-producing dairy cow. Taken together, polymorphisms of TNFα gene could be strongly related to early first ovulation after parturition, thus being an effective tool of selection for improving reproductive performance in the high-producing dairy cow.