Influence of enrofloxacin and chloramphenicol on the level of IgY in serum and egg yolk after immunostimulation of hens with Salmonella enteritidis antigens

In the chicken, maternal antibodies are transferred into the egg and subsequently transported into the developing embryo. IgG (called IgY) is the primary immunoglobulin isotype of the egg yolk. Their level in serum depends on the correct function of immunological system in laying hens. Many factors have a direct or indirect influence on antibody level in fowl. One of them is a commonly used antibiotic, but its influence on avian immune system is still unknown. The objective of the study was to determine the effect of enrofloxacin and chloramphenicol on the level of IgY antibody in serum and egg yolk after immunostimulation of hens with living cells of Salmonella enterica subsp. enterica serovar Enteritidis and lipopolisaccharide. Forty adult egg-laying Arbor Acres and Isa 215 hens (32 and 50 weeks old) from the reproductive flocks and 1640 of their eggs were used for the investigation. No clinical symptoms of any diseases were observed in birds during the entire breeding period. Additionally the birds were checked as free from Salmonella spp. in the beginning of the experiment. The birds were divided into 6 experimental and 2 control groups (5 birds in one group). The hens in the experimental groups were immunized with S. Enteritidis antigens: living bacteria and lipopolisaccharide and treated with enrofloxacin or chloramphenicol. Antibiotics were administered in drinking water for 10 days (from 3rd to 13th day of experiment). To indicate anti-S. Enteritidis, antibodies in sera and egg yolk were used indirectly on ELISA based on lipopolisaccharide from S. Enteritidis. As conjugate these were applied anti-chicken IgY with horseradish peroxidase and ABTS with H2O2 as obtained. Additionally, to detect antibody in serum, a rapid slide test was used with Pullognost and Enterognost standard antigens made in the laboratory. The study revealed that both antibiotics tested decreased the level of specific IgY in laying hens immunized with living bacteria and lipopolisaccharide. It seems that antibiotics have a suppressive effect on the immunological system. The strongest immunosuppressive effect was exerted by chloramphenicol.     

抗奶牛乳腺炎多价卵黄抗体的制备及含量测定

本研究的目的是制备抗奶牛乳腺炎主要致病菌的多价卵黄抗体并检测特异性抗体的含量。采用金黄色葡萄球菌、无乳链球菌和大肠杆菌混合抗原免疫产蛋母鸡。通过水稀释法分离卵黄抗体。采用ELISA法检测抗体效价和特异性抗体含量。多价抗体与大肠杆菌的结合效价最高可达25600,与金黄色葡萄球菌和无乳链球菌的结合效价为12800。每毫升卵黄液中大肠杆菌、金黄色葡萄球菌和无乳链球菌特异性卵黄抗体（egg yolk immunoglobulin,IgY）的最高含量分别为2.13、2.01和1.92 mg。免疫后特异性抗体含量随时间变化趋势与抗体效价变化趋势一致。     

Resistance of broiler chickens to Escherichia coli respiratory tract infection induced by passively transferred egg-yolk antibodies

Egg-yolk antibodies induced by immunizing hens with selected Escherichia coli antigens were evaluated for their ability to protect broiler chickens against respiratory/septicemic disease caused by avian pathogenic E. coli (APEC). Seven groups of broiler breeder hens were vaccinated three times, 1 week apart with live E. coli, killed E. coli, E. coli antigens [lipopolysaccharide (LPS), type 1 pilus adhesin (FimH), P pilus adhesin (PapG), aerobactin outer membrane receptor (IutA)] or phosphate buffered saline (PBS). An O78 APEC strain was used for preparation of all the antigens. Egg yolk immunoglobulins (IgY) were purified from eggs of each group and antibody activity in serum and purified IgY was determined by enzyme-linked immunosorbent assay (ELISA). IgY (100mg) was injected intramuscularly into 11-day-old broiler chickens, which were challenged 3 days later with homologous (O78) or heterologous (O1 or O2) E. coli by the intra-air sac route. Mortality was recorded and surviving chickens were euthanized 1 week after the challenge and examined for macroscopic lesions. Passive antibodies against all antigens except FimH were protective (90-100%) against the homologous challenge, but only anti-PapG and anti-IutA were effective against heterologous challenge. Anti-PapG IgY provided the greatest protection against the three serogroups of E. coli used for challenge. Hence vaccination of broiler breeders to induce anti-PapG and anti-IutA antibodies may provide passive protection of progeny chicks against respiratory/septicemic disease caused by APEC.     

卵黄抗体提取方法及其在畜禽细菌性肠道疾病防治中的应用

细菌性肠道感染是畜禽养殖过程中的常见疾病，易引起畜禽采食量下降、生产性能降低等问题，严重者甚至造成死亡。在养殖过程中多采用抗生素对其进行治疗，但由于抗生素长期大量使用，特别是不合理使用，使药物残留和细菌耐药性问题日益凸显。因此，发展安全可靠的抗生素替代品已然成为当今畜牧行业的研究热点。卵黄抗体（egg yolk immunoglobulins，IgY）是通过免疫接种产蛋鸡后，在蛋黄中表达的一种免疫球蛋白，可用于相应疾病的预防和治疗。IgY具有产量大、性质稳定、安全高效等诸多优点，目前IgY作为抗生素替代品已被广泛用于畜禽养殖过程中，并取得了显著的效果。文章介绍了IgY的分子结构、理化性质和作用机理，总结并比较了5种IgY提取方法的去脂率、抗体效价、总蛋白含量等指标，最后详细介绍了IgY在防治畜禽常见细菌性感染中的研究现状和应用前景，以期为IgY的高效生产及疾病防治提供研究依据和技术支持。     

In vitro reactivity and growth inhibition of EPEC serotype O111 and STEC serotypes O111 and O157 by homologous and heterologous chicken egg yolk antibody

IgY is a chicken egg yolk antibody which has been used for treatment and prophylaxis of gastrointestinal infections. Our aim was to verify if IgY obtained from chickens immunized with EPEC O111, STEC O111 and STEC O157 is able to show in vitro reactivity and biological activity towards the three bacteria. IgY was obtained from eggs laid before and after immunization with each bacterium. The preparations of IgY anti-EPEC O111 and anti-STEC O111 shared high reactivity detected by ELISA and growth inhibition ability towards both bacteria EPEC O111 and STEC O111. Nevertheless, the preparation of IgY anti-STEC O157 showed high reactivity and growth inhibitory effect only towards the homologous strain. Our results showing in vitro biological activity of IgY reinforce its use as an alternative for the treatment or prophylaxis of E. coli infections and encourage the development of in vivo studies for a possible future human therapeutic use.     

免疫卵黄特异性阻断家蚕浓核病感染的研究

用家蚕浓核病病原(Bombyxmoridensonucleosisvirus,BmDNV)免疫蛋鸡收集的卵黄,通过体外中和与体内中和生物试验测定鸡免疫卵黄对DNV的防治效果。结果表明,免疫的鸡卵黄能特异性阻断DNV感染家蚕,达到预防的目的,而非免疫卵黄未表现有阻断作用。结果提示:鸡免疫卵黄中含有抗DNV的有效抗体。     

Characterization of specific egg yolk immunoglobulin (IgY) against mastitis-causing Escherichia coli

The objective of this study was to estimate the in vitro activity of egg yolk immunoglobulin (IgY) against mastitis-causing Escherichia coli. Specific IgY was produced by hens immunized with formaldehyde killed E. coli O111 in long-standing immunization response (titer > or =6400 for 100 days) and was isolated from yolks with a purity of 86% by water dilution, salt precipitations and ultrafiltration. Enzyme-linked immunosorbent assay (ELISA) indicated the produced IgY specifically targeted E. coli O111 and five other E. coli strains which were isolated from mastitic cows. The growth inhibition activity of the specific IgY to bacteria was dose-dependent with an effective concentration of 20mg purified IgY per milliliter. The phagocytic activity of E. coli either by milk macrophages (MPhi) or by polymorphonuclear neutrophil leukocytes (PMN) in the presence of specific IgY was significantly higher than that with nonspecific IgY or without IgY (p<0.05), suggesting that it enhanced phagocytic activity. The current work suggests that this specific IgY has potential as a therapeutic treatment for mastitis in dairy cows.     

IgY stability in eggs stored at room temperature or at +4°C

1. The aim of this study was to investigate the stability of immunoglobulin-Y (IgY) in stored eggs from immunised hens. 2. Eggs from individual hens were randomised and stored for up to one month at room temperature, or for up to 6 months at +4°C. IgY was extracted from the egg yolks and the antibody activities were tested by ELISA. 3. There was no significant reduction in antibody titres with egg storage under these conditions. 4. Egg yolks of immunised chickens provide an inexpensive source of large amounts of polyclonal antibodies for use in immunotherapy and immunoassays. By collecting eggs from different immunised hens and pooling their yolks, it should be possible to reduce batch-to-batch variation.     

鸡抗鹿链球菌卵黄抗体的研制与初步治疗实验

用链球菌灭活菌苗免疫产蛋母鸡，监测卵黄抗体效价达到试验要求收集鸡蛋，提取卵黄中的卵黄免疫球蛋白（IgY）。临床发病鹿群用药物和IgY配合治疗，观察IgY的作用和疗效。     

抗猪流行性腹泻病毒卵黄抗体的制备及应用

本试验利用猪流行性腹泻病毒(porcine epidemic diarrhea virus,PEDV)SC株免疫产蛋鸡,收集高免卵黄,采用水稀释法和水稀释-盐析法获得鸡源抗PEDV卵黄抗体,采用已建立的ELISA方法对卵黄抗体效价进行测定。以3日龄无母源抗体的易感仔猪为试验动物,对抗PEDV卵黄抗体的治疗效果及安全性进行测定,并进一步利用自然发病猪场的治疗效果进行验证。结果表明,经PEDV SC株细胞毒免疫的鸡可在2次加强免疫后15 d产生高水平的特异性抗体。在实验室治疗试验中,攻毒治疗组仔猪的存活率达60%,攻毒对照组存活率为20%;用于自然发病猪场时,饲喂抗PEDV卵黄抗体饲料的仔猪存活率亦为60%,而自然发病对照组的存活率为10%。以上结果表明抗PEDV卵黄抗体对感染PEDV的仔猪有一定治疗作用。     

抗牛多杀性巴氏杆菌高免卵黄抗体IgY的制备及间接血凝检测方法的建立

本研究通过自制牛源荚膜血清A型多杀性巴氏杆菌灭活菌苗免疫产蛋鸡,采用卡拉胶结合硫酸铵沉淀法提取卵黄抗体IgY,并采用间接血凝方法检测抗体效价。结果表明,抗原致敏红细胞的最佳浓度是800μg/mL,免疫后第7周抗体效价达到高峰,效价为1：1024,高效价持续5周开始下降,测定提取的IgY浓度为8.258mg/mL,无菌检测及动物安全性实验表明制备的卵黄抗体安全可靠。本研究制备了抗牛多杀性巴氏杆菌卵黄抗体,为防治由荚膜血清A型多杀性巴氏杆菌所致的犊牛肺炎提供了新的手段。     

Supplemental defatted microalgae affects egg and tissue fatty acid composition differently in laying hens fed diets containing corn and flaxseed oil

Effects of defatted microalgae (DFA) on fatty acid composition of eggs and tissues of hens have not been well studied. This experiment was to determine how low concentrations of supplemental DFA in the presence or absence of flaxseed oil (FO) altered fatty acid profiles of egg yolk, liver, adipose tissue, and plasma of layers. A total of 60 Shaver Leghorn layer hens (individually caged, 20-wk old) were allotted into 6 groups (n = 10) and fed a corn/soybean meal basal diet containing 0, 3, or 5% DFA and 0 (1.5% corn oil, CO) or 1.5% FO for 6 wk. BW, ADFI, egg production rate, and egg component weights were not affected by any of the dietary treatments at various time points, but egg yolk color was changed (P < 0.05) with increasing concentrations of DFA. The concentration of n-3 fatty acids in egg yolk and plasma were elevated by FO up to 9-fold (P < 0.05) starting at wk 1 or 2, and 39 to 83% (P < 0.05) by DFA. Supplemental DFA improved (P < 0.05) n-6/n-3 fatty acid ratios in egg yolk and plasma from 13 to 23 and 7 to 13, respectively, only in hens fed diets without FO. Fatty acid profiles of liver and adipose tissue (at wk 6) displayed responses to supplemental DFA and FO similar to those of egg yolk or plasma. In conclusion, supplemental 3 or 5% DFA caused moderate enrichments of n-3 fatty acids and decreased the n6/n3 fatty acid ratios of egg yolk and tissues in hens fed diets without FO.     

Protection of neonatal calves against fatal enteric colibacillosis by administration of egg yolk powder from hens immunized with K99-piliated enterotoxigenic Escherichia coli.

The protective effects of egg yolk powder prepared from hens vaccinated with heat-extracted antigens from K99-piliated enterotoxigenic Escherichia coli (ETEC) strain 431 were evaluated in a colostrum-fed calf model of ETEC-induced diarrhea caused by a heterologous strain (B44). The antibody powder was obtained by spray-drying the water-soluble protein fraction of egg yolks after removing the lipid and fatty components by precipitation with hydroxypropylmethylcellulose phthalate. A total of 16 colostrum-fed calves were studied to determine whether the orally administered antibody powder would prevent fatal bovine colibacillosis caused by a virulent ETEC strain. Clinical response of individual calves was monitored and evaluated in the context of these variables: fecal consistency score, intestinal colonization, weight loss, and mortality. Control calves that were treated with vehicle (milk with egg yolk powder from nonimmunized hens) had severe diarrhea and dehydration and died within 72 hours after infection was manifested. In contrast, calves fed milk containing egg yolk powder with antipili agglutinin titers of 1:800 and 1:1,600 had transient diarrhea, 100% survival, and good body weight gain during the course of the study. Results indicate that the orally administered egg yolk powder protected against ETEC-induced diarrhea in neonatal calves and that the protective components may have been the antibodies raised by vaccination of chickens against ETEC.     

不同干燥工艺加工的苜蓿粗多糖对蛋鸡生产性能、蛋品质和血清、鸡蛋中抗体含量的影响

本试验旨在研究不同干燥工艺加工的苜蓿粗多糖对蛋鸡生产性能、蛋品质、器官指数和血清、鸡蛋中抗体含量的影响。选取54只38周龄的京红1号产蛋鸡,随机分为3组,每组18个重复,每个重复1只鸡,单笼饲养。对照组饲喂玉米-豆粕型基础饲粮,试验组分别饲喂在基础饲粮中添加1 000 mg/kg微波真空干燥所得苜蓿粗多糖1(ACPS1)和减压烘干干燥所得苜蓿粗多糖2(ACPS2)。试验期65 d。结果表明:与对照组相比,1)在试验第28~36天,ACPS1组蛋鸡的平均日采食量显著增加(P0.05);饲粮中添加不同干燥工艺加工的苜蓿粗多糖对蛋鸡各阶段的产蛋率和平均日增重均无显著影响(P0.05)。2)试验第63天,ACPS1组蛋鸡的蛋黄颜色显著增加(P0.05);试验第30天,饲粮中添加不同干燥工艺加工的苜蓿粗多糖对蛋鸡的蛋壳颜色、蛋黄颜色、蛋白高度和哈氏单位均无显著影响(P0.05)。3)ACPS1组蛋鸡的肝脏指数显著增加(P0.05),ACPS2组的卵巢指数显著降低(P0.05);且ACPS1组的肝脏指数和卵巢指数均显著高于ACPS2组(P0.05)。饲粮中添加不同干燥工艺加工的苜蓿粗多糖对蛋鸡的脾脏指数和输卵管指数均无显著影响(P0.05)。4)ACPS1组蛋鸡血清中免疫球蛋白Y(Ig Y)含量显著增加(P0.05),ACPS1和ACPS2组血清中新城疫抗体(NDV-Ab)、蛋黄中NDV-Ab、蛋清中免疫球蛋白A(Ig A)和免疫球蛋白M(Ig M)含量均显著增加(P0.05);且ACPS1组蛋黄中NDV-Ab、蛋清中Ig A和Ig M含量均显著高于ACPS2组(P0.05)。以上结果表明,饲粮中添加苜蓿粗多糖能加深蛋鸡的蛋黄颜色,增加血清和鸡蛋中的抗体含量,且ACPS1的效果更佳。     

Correlation analysis of the total IgY level in hen serum,egg yolk and offspring serum

The correlation between IgY levels of the serum and the yolk has been well documented in wild and domestic birds. The levels of total yolk IgY can be an index of the general health status of birds and may contribute to breeding programs when fitness of the offspring is a concern. We measured the levels of total serum IgY and yolk IgY in three different breeds (White Leghorn, Silkie and Dongxiang blue-shell) using indirect ELISA, and found that there was a significantly positive correlation between the levels of total serum IgY and total yolk IgY in all three breeds (White Leghorn: r = 0.404, P < 0.001, n = 100; Silkie: r = 0.561, P < 0.001, n = 70; Dongxiang blue-shell: r = 0.619, P < 0.001, n = 30). We also measured the total serum IgY levels in the 3-day-old offspring hatched from the Silkie hens and results were significantly correlated for serum IgY levels (r = 0.535, P < 0.001, n = 70) and the yolk IgY levels (r = 0.481, P < 0.001, n = 70). The regression analysis showed simple linear regression between IgY levels in hen serum, yolk and offspring serum. Our results suggest that total IgY level could be used as an index for chicken fitness.     

不同品种、不同周龄鸡蛋营养成分比较研究

本试验选用北京油鸡、白来航、海兰褐3个品种,每个品种随机挑选17或19周龄(北京油鸡)、42周龄、70周龄母鸡收集鸡蛋进行蛋黄营养成分的测定。结果表明:不同品种3个产蛋时期的蛋黄比例、蛋白比例、蛋黄维生素E含量都有极显著差异(P<0.01)。其中白来航、海兰褐在不同产蛋时期的蛋黄、蛋白比例呈现相同的变化趋势,即蛋黄比例为70周龄蛋最高,蛋白比例为初产蛋最高。3个品种不同产蛋时期维生素E含量变化亦势趋相同,初产蛋最高。3个品种42周龄蛋中的必需氨基酸比例显著高于其他2个时间点(P<0.01)。商业品种与中国地方鸡种所产蛋在蛋黄、蛋白比例、维生素B2变化趋势迥异,无统一规律可循。     

Effects of IgY antibody on the development of Marek's disease.

The effects of passive immunization with immunoglobulin Y (IgY) on the pathogenesis of Marek's disease (MD) were examined in an experimental line of White Leghorn chickens highly susceptible to MD. Purified IgY with anti-MDV antibody activity, when injected into chicks, delayed the development of MDV viremia and lesions until 9 days postinoculation (PI) with Marek's disease virus (MDV). The blastogenic response of spleen cells to concanavallin-A was depressed at 6 days PI in the birds without passive immunization, whereas it was not totally depressed until 17 days in birds passively immunized with IgY anti-MDV antibody.     

抗CP型、NCP型牛病毒性腹泻病毒高免卵黄抗体的制备

本研究采用致细胞病变（cytopathic,CP）型牛病毒性腹泻病毒（bovine viral diarrhea virus,BVDV）标准毒和非致细胞病变（noncytopathic,NCP）型新疆优势毒株免疫产蛋鸡,用改良PEG法提取卵黄抗体(IgY),并对提取的IgY采用SDS-PAGE检测纯度,间接ELISA检测免疫后每隔7 d的抗体效价,并测定所得抗体对NCP型BVDV的中和效价。结果表明,用该法提取的IgY纯度较高;间接ELISA结果证明,经过4次免疫后,抗CP型BVDV的效价达到1∶32000,抗NCP型BVDV的效价达到1∶40000,3个月后再次检测,卵黄抗体效价未见明显下降。最后一次免疫14 d的抗体对NCP型BVDV的中和效价达到1×10_^-3。     

Effects of supplemental copper and chromium on the serum and egg traits of laying hens

1. This study investigated the effects of supplemental copper and chromium on the serum and egg traits of laying hens using 144 forty-five-week-old White Leghorn layers. The hens were randomly assigned into 9 groups to conduct this 3 copper (0, 125 and 250 mg/kg as copper sulphate) x 3 chromium (0, 800 and 1600 microg/kg as chromium picolinate) factorial trial. 2. Egg yolk cholesterol was significantly reduced by supplementary copper (Cu) and chromium (Cr) and there was an interaction between Cu and Cr supplementation. Egg production, egg weight, eggshell strength and eggshell thickness were not influenced by Cu or Cr supplementation. 3. Serum cholesterol concentration was decreased by Cu supplementation and very low-density lipoprotein (VLDL) was reduced markedly by both Cu and Cr. On the other hand, high-density lipoprotein (HDL) was significantly increased by Cu and Cr. Interaction occurred between Cu and Cr in the VLDL and HDL parameters. Supplmentary Cu also reduced VLDL-cholesterol (VLDL-C) and enhanced HDL-cholesterol (HDL-C), while supplemental Cr had a contrary effect on these two parameters. 4. If minimum egg cholesterol content is the criterion, this study suggests that 125 mg/kg of Cu together with 800 to 1600 microg/kg of Cr provide adequate supplementary concentrations for laying hens.     

Development and application of an ELISA for the detection of duck antibodies against Riemerella anatipestifer antigens in egg yolk of vaccinees and in serum of their offspring

A direct and an indirect antibody enzyme-linked immunosorbent assay for duck yolk IgY and duck serum IgY was developed and tested on egg yolk and serum of ducks vaccinated with Riemerella anatipestifer (Ra). Tests were performed either with primary antibodies labelled with horseradish peroxidase or with alkaline phosphatase-labelled secondary antibodies reacting with specifically bound rabbit anti-duck IgY antibodies, respectively. Ra-specific IgYs in egg yolk from three ducks increased rapidly at day 8 after the first of two vaccinations. In two ducks, the IgY titre persisted on a high plateau for 3 months. The concentration of Ra-specific IgYs in the serum of the progeny of vaccinees decreased between day 3 and day 10 after hatching. The fraction of total IgYs decreased less but also significantly. It was shown that antibodies were vertically transmitted and therefore protect offspring against Ra infection at least during the first week after hatching. The test design with anti-IgY rabbit antibodies is further suitable to detect other specific antibodies if respective antigens were fixed on solid phases.