奶牛乳和血中大肠杆菌和轮状病毒抗体检测比较

用轮状病毒和大肠杆菌给妊娠后期奶牛免疫,使之产生抗这两种病原的抗体,定期采血和乳,分别用试管凝集反应和反向间接血凝抑制试验检测大肠杆菌和轮状病毒抗体,并比较血和乳中抗体相互关系,试验结果说明：奶牛在未免疫情况下,只在初乳中才能检出抗体;在人工免疫情况下,在初乳和近２个月的常乳中均可检测到抗体,但乳中抗体比血中抗体低２个滴度左右。     

奶牛乳腺炎疫苗免疫试验报告

对奶牛乳腺炎疫苗进行免疫试验，结果显示：试验所用疫苗对奶牛临床乳腺炎的免疫效果较明显，并使奶牛乳中体细胞数大幅下降，避免了乳中体细胞数升高对奶牛产量的不良影响。在不考虑奶牛重复感染临床乳腺炎情况下，疫苗对奶牛临床乳腺炎的保护率可达到84．75％。     

免疫乳中特异性抗体对犊牛腹泻的被动保护作用

以9株人肠道病原菌(包括埃希氏菌属3株、杀门菌属3株、志贺菌属3株)作为免疫原制成混合疫苗,对怀孕奶牛进行系统免疫,以获得对人致病肠道杆菌具有特异性的牛乳抗体。犊牛饲喂试验表明,免疫乳对犊牛的肠道疾病的发生起到了预防和辅助治疗作用,从而促进犊牛的生长发育,这是由于免疫乳中特异性抗体IgG所决定的。     

免疫乳与血中抗轮状病毒和大肠埃希氏菌抗体的比较研究

以引起婴儿腹泻的轮状病毒和大肠埃希氏菌为免疫原给妊娠后期乳牛免疫,定期采乳和血,分别用试管凝集反应和反向间接血凝抑制试验检测抗大肠埃希氏菌和轮状病毒抗体,比较血和乳中抗体相互关系。结果表明,乳牛在免疫前只能在初乳中检出抗体;免疫后在初乳和近２个月常乳中均能检测到抗体,两者呈平行关系,仅乳中抗体效价比血中抗体效价低２个滴度。     

奶牛乳房炎无乳链球菌Fc-Sip和金黄色葡萄球菌Fc-FnBPB-ClfA亚单位疫苗的免疫试验研究

为探究由IgG FcRn介导奶牛乳房炎无乳链球菌(S. agalactiae,Sgc)和金黄色葡萄球菌(Staphylococcus aureus,Sau)保护性抗原亚单位疫苗的免疫效果,在本实验室前期研究基础上,本研究按体积11的比例将Sgc的Fc-Sip和Sau的Fc-FnBPB-ClfA两种蛋白分别与1%的盐酸左旋咪唑(1%LH)充分混合,制备两种亚单位疫苗。将即将干奶(妊娠220 d)、CMT试验为"++"以上的奶牛随机分为4组,分别将Fc-Sip+Fc-FnBPB-ClfA二联亚单位疫苗(A组)、Fc-Sip亚单位疫苗(B组)、Fc-FnBPB-ClfA亚单位疫苗(C组)通过首次肌肉免疫和二次乳头管灌注免疫方式进行免疫试验。通过免疫前后对乳样细菌分离、体细胞数测定及免疫血清抗体滴度测定,评价上述亚单位疫苗的免疫效果。结果显示,免疫前免疫组(A、B、C)奶牛的乳样细菌分离率分别为80%、90%、80%,其中Sgc检出率分别为20%、30%、10%,Sau检出率分别为30%、20%、10%;首免90 d后3个免疫组奶牛乳样细菌分离率分别降至20%、20%、40%,其中Sgc的检出率分别降至0、0、10%,Sau的检出率分别降至10%、10%、0,对照组免疫前后以上各项检测无明显变化。使用利拉伐牛奶体细胞检测仪检测免疫前后试验奶牛乳样中的体细胞数,免疫前免疫组(A、B、C)奶牛体细胞数均值分别为6.19×105个/mL、5.16×105个/mL、5.49×105个/mL,首免90 d后3个免疫组奶牛乳样中体细胞数均值分别为2.05×105个/mL、2.04×105个/mL、2. 52×105个/mL,而对照组奶牛乳样体细胞数与免疫前无明显变化。检测首免后0、7 d、14 d、28 d、60 d、90 d的血清抗体滴度,结果显示,首免后7 d,免疫组(A、B、C)80%的奶牛血清抗体滴度为11 024;第14 d后,免疫组(A、B、C)90%奶牛血清抗体滴度达到12 048;第28 d后,免疫组(A、B、C)80%奶牛血清抗体滴度在14 096～18 192,比对照组平均上升3～4个抗体滴度,对照组奶牛血清抗体滴度一直保持在1256～1512。综合分析以上试验结果显示免疫A组(Fc-Sip+Fc-FnBPB-ClfA)综合免疫效果在各免疫组中最佳。本研究结果表明Fc-Sip+Fc-FnBPB-ClfA二联亚单位疫苗对无乳链球菌和金黄色葡萄球菌性奶牛乳房炎具有较好的治疗和预防作用,为奶牛隐形乳房炎的防治提供了实验依据。     

鸡源致病性大肠杆菌iss基因原核表达产物的免疫保护作用

本试验以鸡E.coli强致病力菌株HO2的iss基因原核表达产物GST-Iss融合蛋白,免疫SPF鸡获得抗血清。分别检测菌株HO2在抗血清及正常血清中的存活状况,通过比较两者差别,探讨iss基因原核表达产物的免疫保护作用。结果显示,所获鸡大肠杆菌iss基因原核表达产物的抗血清能够减弱菌株的血清抗性,对鸡E.coli强致病力菌株HO2有抑制增殖或杀灭的作用。     

免疫乳及血液中抗伤寒沙门菌抗体消长规律的研究

为了研究免疫乳及血液中抗伤寒沙门菌抗体的消长规律,试验采用伤寒沙门菌作为免疫原对妊娠后期奶牛进行强化免疫,然后定期采集血样和乳样,用微量凝集反应检测样品中伤寒沙门菌的抗体水平,并比较二者的相互关系。结果表明:对照组只有在初乳中检测出较高的抗体效价,并且维持时间较短;试验组中可以观察到高效价抗体且从分娩之日起持续近50d。同时发现在整个试验期间,试验乳中抗体比试验血约低2个效价。     

重组大肠杆菌外膜蛋白OmpT对小鼠免疫保护作用的研究

为评价大肠杆菌(E.coli)外膜蛋白酶T(OmpT)的免疫原性及对小鼠的免疫效率,本研究将原核重组表达纯化的OmpT蛋白(rOmpT)免疫小鼠,并以灭活的E.coli 308-2全菌疫苗株作为对照组,通过检测其抗体效价、体外调理吞噬作用和细胞因子表达水平以及攻毒试验进行保护效率评价。结果表明rOmpT免疫组在二免后14 d其抗体水平可以达到峰值(1∶64 000),并且其抗血清具有显著促进巨噬细胞对E.coli的吞噬作用;同时,重组蛋白与全菌免疫组均能够显著提高小鼠体内IL-4和IFN-γ的表达。而且重组蛋白免疫组对4个进化种系(Phylogenetic group)的E.coli攻毒的免疫保护率均为60%左右,比全菌免疫组高约10%。研究结果显示rOmpT比全菌灭活E.coli能够诱导小鼠产生更有效免疫保护作用,表明rOmpT可以作为E.coli亚单位疫苗候选蛋白。     

牛支原体灭活疫苗免疫程序及母源抗体消长规律研究

为评价不同剂量的自主研发牛支原体灭活氢氧化铝佐剂疫苗对怀孕母牛免疫后抗体消长规律和免疫效果,本试验选择产前30～15d围产母牛70头,随机分为7组,每组10头牛,间隔15d,采用不同剂量免疫,定期采集初乳和犊牛血清,采用已建立的牛支原体间接ELISA检测方法检测初乳乳清和犊牛血清中的抗体滴度,统计分析其抗体消长规律。结果表明,使用自主研发牛支原体灭活氢氧化铝佐剂疫苗免疫怀孕母牛无红肿、发热等症状,是安全可靠的;用不同剂量疫苗免疫怀孕母牛,初乳乳清抗体中牛支原体Ig G抗体水平在奶牛分娩后3h达到一个峰值,随着奶牛产后时间的延长抗体水平呈逐步下降,在奶牛产后96h,抗体值均在0.3以下;A组的抗体最低,B组、C组、D组抗体均低于E组和F组;E组和F组抗体滴度较高,两者无差异。不同剂量疫苗免疫母牛所产犊牛哺食初乳后,犊牛血清中牛支原体Ig G抗体在犊牛出生后24h达到最高,随着犊牛日龄的增加,抗体滴度呈现逐步下降趋势;在A组、B组和C组抗体呈现层次不齐的状况,D组抗体水平高于A组、B组、C组,低于E组和F组;E组和F组抗体水平较高,且维持时间较长。研究结果为牛支原体疫苗的免疫程序制定提供了...     

鸡新城疫疫苗对不同鸡群的免疫试验

通过对母源抗体消长规律的测定，从而为鸡新城疫疫苗的首次免疫提供了依据，在此基础上，用不同的鸡新城疫疫苗分别对雏鸡进行免疫，并监测其抗体的消长。根据监测的结果看出，经用ND CO－30和ND（Lasota系）活疫苗滴鼻、点眼的雏鸡，抗体可维持到21d，然后需再次进行免疫；而在7日龄时用ND活疫苗滴鼻、点眼的同时注射ND油乳剂灭活疫苗的雏鸡，抗体可保持到60d以上。经对国产苗和进口苗的比较，两种疫苗的结果基本一致，差异不显著，但在价格方面，国产苗比进口苗占有优势。     

Studies of the pathogenesis of enteric E. coli infections in weaned pigs. The significance of the milk of the dam in preventing the disease.

Milk from sows whose progeny developed post weaning E. coli diarrhoea (PWD milk) and from sows which were immunized by intramuscular vaccination using a homologous strain of E. coli (immune milk) were tested in ligated segments of pig intestine. The results showed that PWD milk neutralized the enterotoxigenic, fluid accumulating capacity of the lysate of the disease-causing E. coli pathogen. A similar effect was seen by using immune milk (Table I). Neither PWD milk nor immune milk contained sufficient antibacterial activity to neutralize the fluid accumulating capacity of live cultures of E. coli O149:K91, while such activity was contained in immune serum. It is concluded that milk from sows whose progeny developed PWD contains antibodies capable of neutralizing the enterotoxigenic effects of the homologous E. coli organisms. It is suggested that the presence in milk from these sows of antibody-mediated activity against enteropathogenic E. coli organisms may be instrumental in preventing the disease in the progeny during the suckling period and consequently, when this protective milk supply stops at weaning, the disease may develop in susceptible animals, mainly because their own production of specific E. coli antibodies is insufficient to prevent PWD.     

Immunity to Escherichia coli in Pigs. The Role of Milk in Protective Immunity to E. coli Enteritis

Milk whey from immunized sows increased the survival time of gnotobiotic piglets infected with Escherichia coli. The survival time of infected piglets fed milk whey from non-vaccinated sow's was the same as that of similar pigs fed condensed cow milk.

The significance of milk antibodies in immune protection against E. coli enteritis is discussed and compared with that of absorbed colostral antibodies. From calculations presented it would appear that seven day old piglets receive approximately 1 g of gamma globulin daily from milk and that this is equal to the piglets total serum gamma globulin content. After seven days of age the gamma globulin content of piglet serum falls, whereas that of milk remains constant; milk is, therefore, potentially a major source of immunoglobulins with protective activity against E. coli associated enteritides.

     

Serological, colostral and milk responses of cows vaccinated with a single dose of a combined vaccine against rotavirus, coronavirus and Escherichia coli F5 (K99)

Twenty-five Ayrshire/Friesian cows were vaccinated once with a new combined vaccine against rotavirus, coronavirus and Escherichia coli F5 (K99) or given a saline placebo 31 days before the first expected calving date. Blood samples were taken from the cows at intervals from vaccination until seven days after calving and from their calves up to 28 days after birth, and colostrum and milk samples were collected from the cows at intervals for 28 days after calving. There was a significant increase in the mean specific antibody titre against all three antigens in the serum of the vaccinated animals (even in the presence of pre-existing antibody) which was accompanied by increased levels of protective antibodies to rotavirus, coronavirus and E coli F5 (K99) in their colostrum and milk for at least 28 days.     

High milk neutrophil chemiluminescence limits the severity of bovine coliform mastitis

Polymorphonuclear neutrophil (PMN) function changes during mastitis. To investigate the contribution of milk PMN to the severity of Escherichia coli (E. coli) mastitis, chemiluminescence (CL) of blood and milk PMN and their efficiency to destroy coliform bacteria in the mammary gland were examined following the induction of E. coli mastitis in early lactating cows. To better assess and define the degree of mastitis severity, cows were classified as moderate and severe responders according to milk production loss in the non-infected quarters at post-infection hour (PIH) 48. There was an inverse relationship between pre-infection milk PMN CL and colony-forming units at PIH 6. In moderate cows, the pre-infection blood and milk PMN CL was approximately 2-fold higher than that of severe cows. The probability of severe response increased with decreasing pre-infection PMN CL. At the beginning of the infection blood and milk PMN CL was consistently higher, and milk PMN CL increased faster after infection in moderate cows. At PIH > 48 milk PMN CL in severe cows exceeded that of moderate cows. The somatic cell count (SCC) in moderate cows increased faster than colony-forming units, whereas in severe cows the results were reversed. The kinetics of CL activity for blood and milk PMN before and during the early phase of infection confirmed an impairment in PMN CL activity for severe responding cows. High pre-infection blood and milk PMN CL and the immediate increase of milk PMN CL and SCC after infection limited bacterial growth thereby facilitating the recovery of E. coli mastitis in moderate cows. Our study strengthens the idea that pre-existing milk PMN (a static part of the udder's immune defense) functions as a "cellular antibiotic" before and during infection, and low milk PMN CL is a risk factor for bovine coliform mastitis.     

Effect of selenium and vitamin E on antibody production by dairy cows vaccinated against Escherichia coli.

Sixty clinically healthy Holstein cows were randomly assigned to one of four groups according to their age and parity and vaccinated in late pregnancy (day 190) with a multivalent vaccine against Escherichia coli. The 15 cows in the first group (SeE) were injected intramuscularly with a solution of sodium selenite (0.1 mg Se/kg bodyweight) and vitamin E (alpha-tocopherol acetate, 8 U/kg bodyweight), the cows in the second group (Se) received only selenium and the cows in the third group (E) received only vitamin E at the same doses and by the same route of administration; the cows in the fourth group were used as controls. The vaccination and the injections of selenium and vitamin E were repeated 42 days later. The concentration of selenium in whole blood and of vitamin E in serum was determined by fluorometric methods. Specific antibody titres against E coli were determined in serum samples by ELISA. The results showed that the injection of selenium either alone or in combination with vitamin E significantly improved the production of specific antibodies against E coli, and that the production of specific antibodies was greater after the administration of selenium alone.     

Use of enzyme-linked immunosorbent assay to measure bovine milk and serum antibodies to alpha toxin, beta toxin, and capsular antigens of Staphylococcus aureus

An enzyme-linked immunosorbent assay (ELISA) procedure was used to quantitate milk and serum antibodies (IgG) to Staphylococcus aureus alpha and beta toxins, and S. aureus 2-8 and Smith diffuse strain capsular antigens. Milk samples were collected on two occasions. A comparison was made between levels of milk antibodies specific for the two toxins and capsular antigens for 41 cows that were infected with S. aureus on both sampling dates, and 18 cows not S. aureus-infected on either date. Staphylococcus aureus-infected cows were grouped according to somatic cell counts. All groups of infected cows, regardless of somatic cell counts, had significantly higher milk antibody levels to alpha and beta toxins than did the non-infected cows (P less than .002). Serum samples taken for 13 infected and 4 non-infected cows also indicated that significant elevations in anti-alpha toxin and anti-beta toxin IgG were present in S. aureus-infected cows, compared to non-infected cows. A similar immune response was not seen to capsular antigens, however. No significant differences were present between the two groups of cows for either milk or serum antibodies to Smith diffuse strain capsular antigens. Milk antibodies to 2-8 capsule were significantly elevated only in infected cows with somatic cell counts greater than 10(6)/ml, compared to non-infected cows; no differences were present for serum antibodies to 2-8 capsule between infected and non-infected cows. These results indicate that significant increases in milk (and possibly serum) antibodies to alpha and beta toxins are present in cows with chronic staphylococcal mastitis, apparently resulting from a systemic immune response to these toxins. There does not appear to be a similar immune response to capsular antigens.     

Lactoferrin Concentration in Milk of Bovine Clinical Mastitis

The lactoferrin (LF) concentration in the milk from dairy cows with clinical mastitis was determined to evaluate the relationship between the LF concentration (LFC) in milk and the non-specific defensive capability of the udder. The mean LFC in 368 milk samples from 319 cows with clinical mastitis was significantly higher (p<0.01) than that of normal cows. The mean LFC in milk from quarters infected with Mycoplasma bovis or Staphylococcus aureus was significantly higher (p<0.05) than that of quarters infected with coagulase-negative staphylococci (CNS). In Escherichia coli mastitis, the level of LFC in milk from cows with peracute mastitis was significantly lower (p<0.01) than that from cows with acute mastitis. In cases of mastitis due to E. coli, the mean LFC in milk from cows that needed more than 10 days to recover from the mastitis or were not cured was significantly lower (p<0.05) than that for cows which took less than 10 days to be cured. The mean LFC in milk from cows with peracute E. coli mastitis was significantly lower (p<0.05) than that for cows with mastitis associated with environmental streptococci or CNS, although these low LF levels were somewhat increased after 46 h from the occurrence of mastitis. These results suggest that the decreased levels of LF in peracute E. coli mastitis may be associated with the progress of inflammation in the early phase of mastitis.     

The influence of colostral leukocytes on the immune system of the neonatal calf. II. Effects on passive and active immunization

The influence of colostral leukocytes on the concentration of immunoglobulins and antibodies against an enterotoxigenic strain of E. coli in the sera of newborn calves was investigated for four weeks using four experimental groups. The calves received either complete colostrum (COL-, n = 16), cell-supplemented milk substitute (MS+, n = 7) or pure milk substitute (MS-, n = 6) during the first three days of life. The cows were not specifically immunized. The sera of the COL+ calves had significantly higher concentrations of antibodies against E. coli mainly of IgG1 specificity on the second day of life as compared to those of the COL-. The sera of the COL+ calves contained significantly more IgM on days 2 and 5 and slightly more IgA during the first week. Both COL groups had equal concentrations of serum IgG. It appears that colostral leukocytes which are an integral part of the colostrum enhance the passive immunity of the neonatal calf, especially in regard of antibodies and immunoglobulin classes which are essential for intestinal immunity. The concentration of IgM in the sera of the MS+ calves was reduced, that of IgG did not rise to appreciable amounts; the IgA synthesis started one week later as compared to the MS- group. The administration of isolated colostral cells led to an impairment of the natural active immunization.     

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.     

Kinetics of local and systemic isoforms of serum amyloid A in bovine mastitic milk

The aim of the present study was to characterise the serum amyloid A (SAA) response to intramammary inoculation of Escherichia coli and to examine the distribution of hepatically and extrahepatically produced SAA isoforms in plasma and milk from cows with mastitis. Milk and plasma SAA concentrations were determined before and after experimental induction of E. coli mastitis in six dairy cows. The milk SAA response was characterised by low or undetectable levels before inoculation, very rapid and large increases in concentration after inoculation, and rapid decline towards baseline levels after resolution of disease. In plasma from cows with experimentally induced E. coli mastitis, four hepatically derived SAA isoforms with apparent isoelectric point (pI) values of 5.8, 6.2, 6.8 and 7.4 were demonstrated by denaturing isoelectric focusing. In milk three highly alkaline isoforms with apparent pI values above 9.3 appeared 12 h post-inoculation. These isoforms were not present in any of the plasma samples, and it therefore seems likely that they were locally produced, tissue-specific isoforms. At 24-36 h post-inoculation one or more acidic isoforms corresponding to those found in plasma appeared in the milk samples. The isoforms demonstrated in plasma from cows with E. coli mastitis were also present in serum obtained from three cows with clinical Streptococcus uberis mastitis. In conclusion, experimentally induced E. coli mastitis is accompanied by a prominent SAA response. The results of the present study indicate that SAA accumulation in mastitic milk is the result of both local synthesis of SAA and of hepatically derived SAA gaining access to the milk due to increased permeability of the blood-milk barrier.