昆明地区猪链球菌病氢氧化铝胶多价灭活疫苗研制

2006年4月初昆明地区几个集约化猪场的仔猪发生高发病率、高死亡率的疾病,经细菌染色镜检、分离纯化培养、生化试验及血清学鉴定,最后确诊为革兰氏C、D、E、L群链球菌[1]。利用从几个发病猪场的病仔猪体内分离出的链球菌菌株作为疫苗株,研制出猪链球菌病氢氧化铝多价灭活疫苗。试验证明,该苗对小鼠的保护率为96%,优于单价灭活疫苗和链球菌的商品苗,表明该疫苗的安全性好、免疫效力高,完全适于本地区猪链球菌病的免疫防治。     

市场商情

全国首个猪链球菌蜂胶灭活疫苗问世近日，由山东华宏生物工程有限公司自主研发的猪链球菌病蜂胶灭活疫苗获农业部新兽药注册证书（（2012）新兽药正字02号）。该猪链球菌病蜂胶灭活疫苗是马链球菌兽疫亚种与猪链球菌2型的二价苗。利用蜂胶作为免疫佐剂，能大大降低疫苗的免疫应激，目前该产品已批量上市。     

猪链球菌病灭活疫苗安全性与免疫效力试验

我国学者对猪链球菌致病菌的研究证实,国内猪链球菌病的病原主要是C群和D群链球菌,而国外多由R群Ⅱ型链球菌引起猪发生败血型和脑炎型链球菌病.近年来,许多猪场的猪群经猪链球菌疫苗免疫后,猪链球菌病仍时有发生,对分离的致病菌做了分群鉴定后发现,用于免疫疫苗的疫苗株的菌群与当地的流行株不符,结果造成免疫失败.因此在做好该病病原流行株调查基础上,筛选有代表性的、免疫原性好的链球菌作为制苗的疫苗株,研制适合本地区的多价灭活疫苗,已经成为控制该病的有效手段.     

猪链球菌病灭活疫苗(2型,HA9801株)的研制(Ⅴ)——猪链球菌2型灭活疫苗临床使用效果

广东永顺生物制药有限公司研制生产的猪链球菌2型灭活疫苗, 2005-2008年在全国十多个地区使用,证明该疫苗安全性好,副作用小,免疫效果可靠,是防控由猪链球菌2型引起的猪链球菌病的有力武器,具有非常重要的意义.     

猪链球菌3型疫苗候选菌株筛选及3＋9型二价灭活疫苗对小鼠免疫效果评估

【目的】 筛选猪链球菌血清3型疫苗候选菌株,制备猪链球菌3＋9型二价灭活疫苗并评估其在BALB/c小鼠上的免疫保护效果。【方法】 从发病猪病料中分离猪链球菌血清3型菌株,通过蜡螟幼虫和BALB/c小鼠模型筛选出强毒株作为疫苗候选菌株,并对候选菌株进行生物学特性研究。将筛选得到的3型疫苗候选菌株和前期筛选的9型疫苗候选菌株灭活浓缩,使其抗原浓度为2×10¹⁰ CFU/mL,无菌检测后将浓缩抗原液按1：1配比混合,再混合抗原与Summit Poly Solution佐剂按4：1比例混合,制备猪链球菌3＋9型二价灭活疫苗,疫苗中猪链球菌血清3和9型含菌量均为2×10⁹ CFU/mL。将制备的疫苗免疫6周龄BALB/c小鼠,首免后第14天进行二免,二免后第14天进行攻毒。同时设立商品化疫苗免疫组和阴性对照组,评估疫苗的安全性和免疫保护效果。【结果】 PCR鉴定结果显示,23株临床分离株均为血清3型猪链球菌,依次命名为KQ3-1~KQ3-23。分别通过蜡螟和小鼠进行初筛和复筛,筛选到强毒株KQ3-1。毒力基因检测显示,该菌株基因型为gapdh^＋/sly^＋/fbps^-/orf2^-/mrp^-/89K^-/gdh^＋/epf^-;生长曲线显示,该菌株在37 ℃培养8~10 h时生长达到对数生长后期;BALB/c小鼠致病性结果显示,腹腔接种12 h内可引起小鼠精神萎靡、扎堆、毛发耸立、运动迟缓、死亡等临床症状,该菌株的LD₅₀为5.2×10⁷ CFU/只。制备的疫苗免疫小鼠后,小鼠精神状况、采食等均正常,疫苗注射部位无肿胀、硬块等不良反应,无死亡发生,表明该疫苗具有良好的安全性。免疫后进行猪链球菌血清2型菌株ZYS、猪链球菌血清3型菌株KQ3-1和猪链球菌血清9型菌株YT攻毒,对照组死亡率分别为90%、100%和100%,猪链球菌3＋9型二价灭活疫苗免疫组保护率分别为30%、80%和70%,商品化疫苗组的保护效果分别为70%、0和10%。【结论】 本研究研制的疫苗能对猪链球菌血清3和9型强毒株提供良好的免疫保护,该疫苗具备疫苗市场开发的潜在价值。     

猪链球菌病灭活疫苗（2型，HA9801株）的研制（Ⅲ）——疫苗安全性研究

本试验采用实验室研制的3批猪链球菌病灭活疫苗(2型,HA9801株),批号200501、200502、200503,分别用猪进行单次超剂量免疫、多次单剂量免疫、对不同日龄猪单剂量免疫、怀孕母猪单剂量单次免疫试验,以及用Bal/c小鼠和兔进行安全性试验,证实该疫苗对猪及Bal/c小鼠、兔是安全的。     

科技前沿

<正>国家三类新兽药:猪链球菌病三价灭活疫苗华中农业大学预防兽医学实验室武汉科前动物生物制品有限责任公司,目前采用疫苗进行免疫预防是国内外控制猪链球菌病的根本手段,而市售商品疫苗只有猪链球菌2     

国际首个“猪链球与副猪二联灭活疫苗”获国家新兽药批文

正科前生物投入重兵研发的"猪链球菌病、副猪嗜血杆菌病二联灭活疫苗"喜获国家农业部新兽药批文。该产品是国际首个猪链球菌病、副猪嗜血杆菌病二联疫苗,将为解决困扰我国养猪业的猪链球菌病和副猪嗜血杆菌病的防控难题提供完美解决方案。     

兽药

正农业部批准猪链球菌病、副猪嗜血杆菌病二联灭活疫苗等2种兽药产品为新兽药近日,农业部批准武汉科前生物股份有限公司等8家单位申报的猪链球菌病、副猪嗜血杆菌病二联灭活疫苗(LT株+MD0322株+SH0165株)     

商情新闻

全国首个猪链球菌蜂胶灭活疫苗问世近日,由山东华宏生物工程有限公司自主研发的猪链球菌病蜂胶灭活疫苗获农业部新兽药注册证书((2012)新兽药正字02号)。该猪链球菌病蜂胶灭活疫苗是马链球菌兽疫亚种与猪链球菌2型的二价苗。利用蜂胶作     

Preparation and testing of polyvalent EDTA-Na-extract vaccine from Escherichia coli strains pathogenic to Swine]

Monovalent vaccines were prepared of E. coli strains with pathogenicity to swine by means of a technique described elsewhere. A polyvalent vaccine was obtained by mixing the monovalent vaccines. This polyvalent vaccine was tested by criteria usually applied to vaccine of E. coli strains with pathogenicity to man, and it exhibited the same quality characteristics.     

Immune response to oil-emulsion vaccines with single or mixed antigens of Newcastle disease, avian influenza, and infectious bronchitis

Inactivated Newcastle disease virus (NDV), avian influenza virus (AIV), and infectious bronchitis virus (IBV) antigens were evaluated for immunological efficacy in monovalent and polyvalent vaccines. Vaccinated broilers were bled for hemagglutination-inhibition (HI) tests at 1- or 2-week intervals. Half of the chickens were challenged with the Largo isolate of velogenic viscerotropic (VV) NDV at 8 weeks post-vaccination, and the remainder were challenged with the Massachusetts 41 strain IBV at 9 weeks post-vaccination. Newcastle disease HI titers were reduced significantly (P less than 0.05) from those of monovalent control vaccine groups when IBV antigen was emulsified in mixtures with low (1-3x) concentrated NDV or NDV and AIV antigens. Avian influenza HI titers were significantly (P less than 0.05) lower than those of the control monovalent groups when highly concentrated NDV was part of the polyvalent vaccine. Infectious bronchitis HI titers were higher than those of control monovalent groups in 13 of 15 vaccine groups when IBV antigen was in polyvalent formulations. VV NDV challenge killed all non-NDV vaccinates and induced increased HI titers in NDV vaccinates but no morbidity or mortality. Sixty of 80 IBV vaccinates experienced a fourfold or greater HI titer increase following challenge. All non-IBV vaccinates seroconverted at 1 week post-challenge.     

Response to and efficacy of vaccination against eastern equine encephalomyelitis virus in emus

OBJECTIVE: To evaluate humoral immune responses of emus vaccinated with commercially available equine polyvalent or experimental monovalent eastern equine encephalomyelitis (EEE) virus and western equine encephalomyelitis (WEE) virus vaccines and to determine whether vaccinated emus were protected against challenge with EEE virus. DESIGN: Cohort study. ANIMALS: 25 emus. PROCEDURE: Birds were randomly assigned to groups (n = 5/group) and vaccinated with 1 of 2 commercially available polyvalent equine vaccines, a monovalent EEE virus vaccine, or a monovalent WEE virus vaccine or were not vaccinated. Neutralizing antibody responses against EEE and WEE viruses were examined at regular intervals for up to 9 months. All emus vaccinated with the equine vaccines and 2 unvaccinated control birds were challenged with EEE virus. An additional unvaccinated bird was housed with the control birds to assess the possibility of contact transmission. RESULTS: All 4 vaccines induced detectable neutralizing antibody titers, and all birds vaccinated with the equine vaccines were fully protected against an otherwise lethal dose of EEE virus. Unvaccinated challenged birds developed viremia (> 10(9) plaque-forming units/ml of blood) and shed virus in feces, oral secretions, and regurgitated material. The unvaccinated pen-mate became infected in the absence of mosquito vectors, presumably as a result of direct virus transmission between birds. CONCLUSIONS AND CLINICAL RELEVANCE: Results indicate that emus infected with EEE virus develop a high-titer viremia and suggest that they may serve as important virus reservoirs. Infected emus shed EEE virus in secretions and excretions, making them a direct hazard to pen-mates and attending humans. Commercially available polyvalent equine vaccines protect emus against EEE virus infection.     

A monovalent attenuated serotype 2 bluetongue virus vaccine confers homologous protection in sheep

An outbreak of bluetongue caused by bluetongue virus serotype 2 virus in certain Mediterranean countries during 1999/2000, presented an opportunity to produce a monovalent type 2 vaccine. Since no data have been published previously on the protection conferred by the current live attenuated bluetongue vaccine strains used in the polyvalent vaccine, a challenge experiment was performed to determine the degree of homologous protection induced by the type 2 vaccine strain. The standard vaccine dose of 5 x 10(4) pfu of vaccine conferred 99.7% protection against clinical disease and no viraemia was detected in the vaccinates.     

Antigenic properties of four serotypes of Pasteurella multocida determined by enzyme-linked immunosorbent assay

Broiler minibreeder hens were used to produce monovalent antisera to bacterins prepared from serotypes 1, 3, 4, and 3 X 4 cross (CU strain) of P. multocida and to a polyvalent fowl cholera bacterin containing serotypes 1, 3, and 4. Antiserum to the CU strain (live vaccine) was also produced. Monovalent enzyme-linked immunosorbent assay (ELISA) plate antigens were prepared by separately sonicating each of the strains. Polyvalent plate antigen (Poly 3) was prepared by combining, in equal amounts after sonication, antigens from serotypes 1, 3, and 4. Each antiserum was assayed against its homologous ELISA plate antigen and against all other heterologous plate antigens, including Poly 3. The strongest reactions, as indicated by the highest absorbance values, were observed in homologous ELISAs. The CU strain may be the best monovalent ELISA plate antigen for detecting antibodies formed in response to a commercial polyvalent bacterin and to vaccinations with the live CU strain. Overall, monovalent serotype 1 (strain X-73) antiserum did not react well with any other heterologous ELISA plate antigen, whereas monovalent antisera of serotypes 4 (strain P-1662) and 3 X 4 (CU strain) reacted equally strongly with monovalent serotype 4 ELISA plate antigen. Background binding of negative serum was significantly lower (P less than 0.05) when using CU plate antigen than when using any of the other plate antigens.     

Foot-and-mouth disease virus typing by complement fixation and enzyme-linked immunosorbent assay using monovalent and polyvalent antisera.

An indirect "sandwich" enzyme-linked immunosorbent assay (ELISA) using polyvalent and monovalent antisera was compared with the 50% complement fixation (CF50) test for the detection of foot-and-mouth disease (FMD) O, A, and C virus types. ELISA was more sensitive than CF50 tests when polyvalent antisera were used for detecting the 3 types of virus in epithelial samples, whereas ELISA using monovalent antisera was the least sensitive technique. The ELISA performed with polyvalent antisera was 9 times more sensitive for detecting FMD virus than that with monovalent antisera. However, viral isolation in cell culture was the most sensitive detection system. The combined use of ELISA with polyvalent antisera and cell culture inoculations was the most effective procedure for identifying FMD virus in epithelial samples from the field.     

New serotype 2 and attenuated serotype 1 Marek's disease vaccine viruses: comparative efficacy

Attempts were made, through selection of optimum viral strains, to develop improved vaccines against Marek's disease (MD). Seven attenuated serotype 1 strains and 22 avirulent serotype 2 strains, both alone and in combination with the FC126 strain of serotype 3, were screened for protective efficacy against challenge with virulent and very virulent MD viral strains. The three viruses selected as most promising were evaluated alone and in various combinations and compared with commercially available vaccines, including FC126, bivalent (FC126 + SB-1), and CV1988/C, in 12 separate assays. Two of these new viruses--301B/1 (serotype 2) and Md11/75C/R2 (serotype 1)--were exceptionally protective compared with prototype vaccine strains. Four new monovalent and polyvalent vaccines based on these two isolates protected chickens better than FC126 alone or CV1988/C alone. Three of these new vaccines provided better protection than the bivalent (FC126 + SB-1) vaccine. Protective synergism was noted commonly between viruses of serotypes 2 and 3 but only sporadically between serotypes 1 and 2 or between serotypes 1 and 3. Strain CVI988/C was protective but was no better than FC126 alone, and it was less effective than bivalent (FC126 + SB-1) vaccine, even when used as a bivalent vaccine with FC126 or SB-1.     

Development and evaluation of an enzyme-linked immunosorbent assay for detection, typing, and subtyping of vesicular stomatitis virus.

An indirect sandwich enzyme-linked immunosorbent assay (ELISA) has been used for vesicular stomatitis virus (VSV) typing using sets of monovalent and polyvalent rabbit/guinea pig antisera for identification of VSV types New Jersey (VNJ) and Indiana (VIND). The VIND polyvalent antiserum (VIND-P) detects any strain of the 3 subtypes of the VIND type (VIND-1, VIND-2, and VIND-3) with the same strong reactivity. It is also possible to subtype the VIND strains using VIND-P rabbit antiserum as capture antibody and monovalent VIND-1, VIND-2, or VIND-3 guinea pig antisera as detector. The ELISA proposed has about 10 times more sensitivity and provides 10% more positive results than does the complement fixation 50% (CF50) test when epithelial samples are tested.     

Immunization of chickens and turkeys against avian influenza with monovalent and polyvalent oil emulsion vaccines.

Chickens and turkeys vaccinated with inactivated virus oil-emulsion vaccines containing different concentrations of either 1 (monovalent) or 4 (polyvalent) strains of avian influenza virus (AIV) were challenged-exposed with virulent AIV A/chicken/Scotland/59 or A/turkey/Ontario/7732/66. Four of 6 vaccines protected completely against postexposure mortality. Vaccine valency did not alter the serologic and challenge-exposure responses of chickens vaccinated with AIV A/turkey/Wisconsin/68, which was the virus component common to both monovalent and polyvalent vaccines. The magnitude of the serologic responses and protection against challenge-exposure were dependent on the concentration of virus in the vaccines. These data indicate that control of virulent AIV in chickens and turkeys by vaccination with inactivated vaccines may be feasible.