PCR检测方法在陕西省布鲁氏菌种型鉴定中的应用

目的 将PCR检测技术应用于陕西省布鲁氏菌的种型鉴定中,建立适用于该省的标准化分子分型技术,为布病防治提供科学依据。方法 用BCSP31 - PCR 及AMOS - PCR对收集的3株标准菌株羊16M、牛544A、猪1330S以及89株陕西省1957年以来保存的部分地方株进行种型鉴定。结果 菌14025、14045经BCSP31 - PCR、AMOS - PCR均未条带,排除是布氏菌,与生化鉴定结果一致;其余菌株BCSP31 - PCR均出现223 bp大小的条带;菌62018、74011、74012经AMOS - PCR出现285 bp左右条带,证实为猪1型菌株,菌92008未出现条带,该方法不能用于犬种菌的鉴定,其余菌株均出现731 bp 左右的条带,为羊种菌。结论 PCR方法特异、快速、准确, 在实际工作中,可用BCSP31 - PCR方法鉴定到属,AMOS - PCR检测方法鉴定到种,可作为布鲁氏菌种属快速鉴定方法之一。     

江西省羊种布鲁氏菌分子流行病学特征分析

目的 掌握江西省羊种布鲁氏菌分子特征,了解分离菌株流行病学间相关性。方法 采用多位点可变数目串联重复序列分析方法（MLVA）对江西省17个县（区） 25株人分离羊种布鲁氏菌进行分析。结果 25株羊种布鲁氏菌聚类可分为24个独立基因型,相似度为67.00%～100.00%,辛普森指数在0.000～0.773之间,MLVA8型有3个基因型,60.00%（15/25）为42型,32.00%（8/25）为43型,8.00%（2/25）为63型。MLVA11型有7个基因型,116、125型为主要基因型,两型菌株占56.00%（14/25）。结论 25株人分离羊种布鲁氏菌基因具有高度遗传多样性,菌株基因型繁多,各菌株间无明显流行病学相关性,表明江西省羊种布鲁氏菌传染来源复杂。     

肠炎沙门菌可变数目串联重复序列位点用于多位点可变数目串联重复序列分型分析的评价

目的 评价不同肠炎沙门菌可变数目串联重复序列(VNTR)位点用于多位点可变数目串联重复序列(MLVA)分型的可行性.方法 选取已报道使用的肠炎沙门菌11个VNTR位点,对中国不同时间和地区分离的16株菌进行初步评价,选取具有单一扩增条带的位点进行104株肠炎沙门茵的MLVA分型分析.对这些菌株同时进行脉冲场凝胶电泳(PFGE)分析,比较MLVA分型方法与PFGE分型方法对菌株分型能力的强弱.结果 初筛得到7个VNTR位点用于扩大菌株的分析,这些位点将104株菌分为16个MLVA型别,D值为0.7222,这些菌株同时被分为22个PFGE型别,D值为0.7974.对两种方法各自所分的最大组包含的菌株进行比较,发现PFGE具有更强的分辨能力;从频数分布看,PFGE方法分型结果比较分散,MLVA分型较为集中.结论 用于国际肠炎沙门菌分型具有扩增多态性的VNTR位点在国内分离株中并不都具有多态性结果,在MLVA方法学建立中应选择更多的VNTR位点进行广泛的筛选才有利于国际实验室间的方法的统一.     

布鲁氏菌疫苗株与标准株的比较

布鲁氏菌鉴定需要标准株作为对照,标准株具有高致病性。本研究对牛、羊、猪种生物Ⅰ型的标准株（544A、16M、1330S）与疫苗株（S19、M5、S2）进行比较,探索疫苗株作为对照菌株用于鉴定的可能性。     

肠炎沙门菌可变数目串联重复序列位点用于多位点可变数目串联重复序列分型分析的评价

目的 评价不同肠炎沙门菌可变数目串联重复序列(VNTR)位点用于多位点可变数目串联重复序列(MLVA)分型的可行性.方法 选取已报道使用的肠炎沙门菌11个VNTR位点,对中国不同时间和地区分离的16株菌进行初步评价,选取具有单一扩增条带的位点进行104株肠炎沙门茵的MLVA分型分析.对这些菌株同时进行脉冲场凝胶电泳(PFGE)分析,比较MLVA分型方法与PFGE分型方法对菌株分型能力的强弱.结果 初筛得到7个VNTR位点用于扩大菌株的分析,这些位点将104株菌分为16个MLVA型别,D值为0.7222,这些菌株同时被分为22个PFGE型别,D值为0.7974.对两种方法各自所分的最大组包含的菌株进行比较,发现PFGE具有更强的分辨能力;从频数分布看,PFGE方法分型结果比较分散,MLVA分型较为集中.结论 用于国际肠炎沙门菌分型具有扩增多态性的VNTR位点在国内分离株中并不都具有多态性结果,在MLVA方法学建立中应选择更多的VNTR位点进行广泛的筛选才有利于国际实验室间的方法的统一.     

贵州省2010-2012年6株羊种布鲁氏菌分子流行病学特征研究

目的：了解贵州省近年羊种布鲁氏菌分离株的分子流行病学特征。方法应用布鲁氏菌属外膜蛋白31基因PCR（BCSP31-PCR）和基于插入序列IS711的牛、羊、绵羊附睾和猪种布鲁氏菌特异的PCR（AMOS-PCR）对贵州省2010-2012年间分离的6株布鲁氏菌进行鉴定,采用16个可变数量串联重复序列（VNTR）的多位点可变数目重复序列分析（MLVA-16）方法对上述菌株进行分型,并做聚类分析。结果6株菌经BCSP31-PCR鉴定为布鲁氏菌属细菌、AMOS-PCR鉴定为羊种布鲁氏菌。MLVA-16分析显示,6株菌在部分VNTR位点上存在重复数目差异,菌株ZY和ZA为MT63型,菌株LL3、LL4和LL11为MT67型,菌株SQ为MT72型。聚类分析显示,6株菌株中的ZY、ZA、LL3、LL4和LL9株与来自云南、福建和广东省羊3型布鲁氏菌聚类最近,菌株SQ与其他菌株聚类相对较远。结论2010-2012年贵州省布鲁氏菌分离株均为羊种生物3型,菌株间存在重复序列多态性,与来自云南、福建和广东省羊3型布鲁氏菌聚类最近。     

碳青霉烯类耐药鲍氏不动杆菌3种分子分型方法的对比研究

目的通过对碳青霉烯类耐药鲍氏不动杆菌进行3种分子流行病学调查方法的对比研究,了解脉冲场凝胶电泳(PFGE)、多位点序列分析(MLST)和多位点可变数目串联重复序列分析(MLVA)3种方法在对碳青霉烯类耐药鲍氏不动杆菌复合体的流行病学调查中的优缺点和分辨率。方法收集某三级医院2011-2012年分离自痰标本的碳青霉烯类耐药鲍氏不动杆菌复合体212株,采用PFGE、MLST和MLVA方法对其进行分子流行病学分析,运用Simpson相关系数(Simpson's index of diversity)和Wallace coefficient,对3种方法的分辨率和同质性进行比较。结果 212株碳青霉烯类耐药鲍氏不动杆菌复合体,其中149株获得PFGE可分析结果,200株获得MLST可分析结果,205株获得MLVA可分析性结果;PFGE结果显示,B型菌株为优势菌株(102株,占68.46%)。MLST分型结果显示,ST195占优势(60株,占30%),其次为ST208(43株,占21.5%)和ST643(33株,占16.5%)。MLVA分型结果显示,D型菌株占优势(151株,占73.66%),其次为N型菌株(8株,占3.9%),K、M、T、W型菌株并列第三(均为4株,各占1.95%)。优势菌株主要分布在ICU。PFGE、MLST、MLVA的Simpson相关系数(D值)分别为0.46、0.76、0.94。MLVA与MLST的Wallace coefficient值略高为0.76。结论 PFGE是区域内分析克隆时间和空间分布的重要工具,本研究中院内流行的碳青霉烯类耐药鲍氏不动杆菌主要来自于ICU;MLST可全球范围内开展抗菌药物抵抗菌株,以及新变异菌株的流行病学研究;MLVA数据可用于不同实验室之间的相互交换、整合与分析。MLVA的分辨率较高,MLVA与MLST的同质性较高。     

肠炎沙门菌可变数目串联重复序列位点用于多位点可变数目串联重复序列分型分析的评价

目的 评价不同肠炎沙门菌可变数目串联重复序列(VNTR)位点用于多位点可变数目串联重复序列(MLVA)分型的可行性.方法 选取已报道使用的肠炎沙门菌11个VNTR位点,对中国不同时间和地区分离的16株菌进行初步评价,选取具有单一扩增条带的位点进行104株肠炎沙门茵的MLVA分型分析.对这些菌株同时进行脉冲场凝胶电泳(PFGE)分析,比较MLVA分型方法与PFGE分型方法对菌株分型能力的强弱.结果 初筛得到7个VNTR位点用于扩大菌株的分析,这些位点将104株菌分为16个MLVA型别,D值为0.7222,这些菌株同时被分为22个PFGE型别,D值为0.7974.对两种方法各自所分的最大组包含的菌株进行比较,发现PFGE具有更强的分辨能力;从频数分布看,PFGE方法分型结果比较分散,MLVA分型较为集中.结论 用于国际肠炎沙门菌分型具有扩增多态性的VNTR位点在国内分离株中并不都具有多态性结果,在MLVA方法学建立中应选择更多的VNTR位点进行广泛的筛选才有利于国际实验室间的方法的统一.

Abstract：

Objective To evaluate the feasibility of the application of variable-number tandem repeat(VNTR)loci of Salmonella Enteritidis(S. enteritidis)in subtyping mutiple-locus variable-number tandem repeat analysis(MLVA).Methods A total of 16 isolates of S.enteritidis from different place and time in China were preliminarily assessed by choosing 11 reported VNTR loci.the loci with single amplified bands were picked to subtype all 104 S.enteritidis isolates.The isolates were also analyzed by pulse field gel electrophoresis(PFGE)to compare the superiority or inferiority of MLVA method and PFGE method.Results Seven VNTR loci were selected from the preliminary screening to expand the analysis,and the 7 VNTR loci had grouped 104 of S.enteritidis isolates into either 16 MLVA subtypes or 22 PFGE subtypes.with the D value at 0.7222 and 0.7974,respectively.Comparing with the isolates in MLVA subtypes.the isolates in PFGE showed a stronger resolving power.Meanwhile the results in PFGE showed a more disperse frequency distribution than those in MLVA.Conclusion These results indicate that some VNTR locus which have shown a good polymorphism intemationaUy,may fail to show polymorphism in China.thereby.more VNTR loci should be included in MLVA and the wide screening may benefit the unity of global laboratorial methods.     

贵州省-起人间布鲁氏菌病疫情的菌株鉴定及遗传特征分析

目的鉴定贵州省-起人间布鲁氏菌病(布病)疫情来源菌株(GZZA)并分析其遗传特征。方法应用传统方法和聚合酶链反应(PCR)鉴定菌株, 采用多位点可变数目串联重复序列(MLVA)-16分析其遗传特征。结果菌株(GZZA)采用传统方法和PCR鉴定为羊种生物3型布鲁氏菌, MLVA-16分析显示该菌株与布鲁氏菌各型代表菌株中的羊种生物3型布鲁氏菌聚类最近, 但在重复序列(VNTR)位点bruce42、bruce04、brace09和brucel6与羊种生物3型布鲁氏菌存在重复数目差异。结论该起疫情来源菌株(GZZA)鉴定为羊种生物3型布鲁氏菌, 其遗传特征与羊种生物3型菌株最接近, 但部分VNTR位点存在重复数目差异。     

MLVA基因分型方法研究世界多地区布鲁氏菌的流行病学特征

目的 通过多位点串联重复序列分析（MLVA）研究世界多国家和地区分离的布鲁氏菌分子流行病学特征。方法 选择11个可变数目串联重复序列位点,使用BioNumerics软件,采用非加权配对算术平均法,对1953－2013年全球48个国家和地区分离的布鲁氏菌VNTR资料进行聚类分析,绘制系统发育树和最小生成树,分析菌株的流行分布特征。结果 布鲁氏菌系统发育树的进化关系与经典的生物分型方法基本上吻合,但猪种生物5型菌株与其他猪种生物1、2、3和4型菌株关系较远;鲸种布鲁氏菌也分为2个部分,并且关系较远。2005－2008年出现了全球布鲁氏菌病（布病）流行,中国是布病多发区,主要流行株为羊种布鲁氏菌,其次是牛种布鲁氏菌,猪种布鲁氏菌主要出现在中国南部省份,犬种布鲁氏菌只在犬中出现,人类没有发现病例。结论 布鲁氏菌具有种的系统发育树特征,并且具有分离的时间、地域和宿主的特异性,这些特征对于布病的防控具有重要意义。     

Brucella suis S2, brucella melitensis Rev. 1 and Brucella abortus S19 living vaccines: residual virulence and immunity induced against three Brucella species challenge strains in mice

Live attenuated Brucella suis S2 vaccine was compared to living vaccines B. abortus S19 and B. melitensis Rev. 1 in mice. Residual virulence was estimated by ability to multiply and persist in spleen and lymph nodes. Immunogenicity was estimated by spleen counts of control and vaccinated mice challenged either with the reference B. abortus 544 strain or with virulent B. melitensis H38 and B. suis 1330 strains. S2 vaccine had lower residual virulence; expressed as 50% recovery time, persistence was 4.3 weeks, compared to 7.1 and 9.0 weeks for S19 and Rev. 1 vaccines. Immunity induced by the three vaccines was similar 45 days after vaccination. At 150 days, immunity by S19 and Rev.1 was still similar against the three challenge strains. In contrast, immunity induced by S2 had declined against the B. melitensis strain. Thus, a recall vaccination may be required for vaccination of sheep to confer a long-lasting immunity.     

Assessment of genetic stability of Brucella melitensis Rev 1 vaccine strain by multiple-locus variable-number tandem repeat analysis

The assessment of the genetic stability is one of the essential elements to guarantee the biological quality of live anti-bacteria vaccines. Live attenuated Brucella melitensis Rev 1 is the most effective vaccine against brucellosis in small ruminants. Thirty-six B. melitensis Rev 1 vaccine strains isolated from human or animal sources from different geographic regions, from different commercial batches or laboratory collections were typed by the multiple-locus variable-number tandem repeat (VNTR) analysis (MLVA) recently described for Brucella spp. Our results demonstrated that B. melitensis Rev 1 group as assayed by MLVA is genetically very homogeneous. We believe that MLVA methodology could be an essential assay to guarantee the quality and stability of live anti-bacterial vaccines being produced worldwide and can be included as in vitro control.     

2型猪链球菌锌转运蛋白A对小鼠免疫保护作用

目的研究猪链球菌(Streptococcus suis,S.suis)锌转运蛋白A(ZnuA)对小鼠接种致死量2型猪链球菌(S.suis 2)菌株的免疫保护作用,为进一步研究S.suis 2亚单位疫苗奠定实验基础。方法采用聚合酶链反应(PCR)检测znuA基因在不同血清型S.suis中的分布情况;蛋白免疫印迹(western blot)检测ZnuA在S.suis 2的表达,利用流式细胞术对ZnuA进行细胞定位,动物实验研究ZnuA蛋白的免疫保护作用。结果除血清型17、21和30型菌株及荷兰分离株T15菌株外,其他30个血清型S.suis菌株、7996菌株以及3株S.suis 2型国内分离菌株基因组中均扩增到目的条带;S.suis 2菌体蛋白与兔抗ZnuA蛋白的血清能发生特异性反应;兔抗ZnuA血清标记S.suis 2的荧光强度明显较高;ZnuA重组蛋白免疫组小鼠死亡率较低。结论 ZnuA是一种具有免疫保护作用的蛋白,可作为S.suis的亚单位疫苗候选分子。     

Brucella suis biovar 2 multi locus sequence type ST16 in wild boars (Sus scrofa) from Abruzzi region,Italy. Introduction from Central-Eastern Europe?

Porcine brucellosis occurs in many countries where pigs are farmed, often representing an underrated problem. B. suis biovar 2 is the most common isolate in Europe, with high prevalence reported in wild boars in which it is generally isolated in the absence of gross lesions. In the last five years, we tested for Brucella spp. 389 lymph nodes of wild boars collected during hunting seasons or during necropsy procedures.In this paper, we describe the first case of isolation of B. suis biovar 2 from a wild boar aborted foetus, and we analyse the genomic relationships with B.suis biovar 2 strains isolated in the past five years in Abruzzi Region, Central Italy. The genetic fingerprint revealed that the isolates under study belong to the MLST ST16 and to the MLVA11 Gt 57, similar to the Central-Eastern European strains. Massive restocking (for hunting purpose) of wild boars from Eastern Europe have been done since 1950 in Italy contributing to the increasing of population size and distribution, as well as to the interbreeding between these foreign breeds and the local population. The contamination of pastures with infected material such as aborted wild boars foetuses can increase the risk of transmission of Brucella among wild and domestic animals. The contact of B. suis with domestic ruminants may also cause serological reactions to brucellosis serological testing, and even unapparent infection, thus hampering the efforts made in the brucellosis eradication campaign.     

Evaluation and Selection of Multilocus Variable-Number Tandem-Repeat Analysis Primers for Genotyping Brucella abortus Biovar 1 Isolated from Human Patients

ObjectivesBrucellosis is the most common bacterial zoonosis in the world. Multilocus variable-number tandem-repeat (VNTR) analysis (MLVA) is a molecular method for genotyping bacterial species. Brucella abortus biovar I was isolated from most of the brucellosis-suspected patients in Korea. This study was conducted to investigate the ability of various MLVA primers that are used for molecular typing B. abortus isolates and for analyzing their epidemiological data.MethodsA total of 80 human isolates of B. abortus biovar I isolated from human patients and the reference strain were used for MLVA. Genetic diversity was determined by calculating the Simpson's diversity index (DI) of each VNTR locus. The Brucella strains were subcultured 30 times to determine the stability of each locus. The DNA of the strains cultivated in each passage was extracted and subjected to MLVA for further investigation.ResultsThe 15 VNTR loci were selected based on high DI values. The DIs of the 15 VNTR loci showed considerable discrimination power ranging from 59% for Bruce 43 to 87% for Bruce 22. Bruce 09, Bruce 11, Bruce 16, Bruce 42, and Bruce 43 were confirmed to remain stable in vitro among the 15 VNTR loci selected.ConclusionThe results of this study suggest that the five loci subsets may be a useful epidemiological tool for investigating B. abortus biovar 1 outbreak.     

Genomic evidence that the live Chlamydia abortus vaccine strain 1B is not attenuated and has the potential to cause disease

BackgroundThe live, temperature-attenuated vaccine strain 1B of Chlamydia abortus, the aetiological agent of ovine enzootic abortion (OEA), has been implicated in cases of vaccine breakdown. The aim of this study was to understand the nature of this attenuation through sequencing of the vaccine parent strain (AB7) and the derived mutant strains 1B and 1H, as well as to clarify the role of the vaccine strain in causing disease through comparative whole genome analysis.MethodsWhole genome sequencing was performed on: vaccine parent strain AB7; N-methyl-N′-nitro-N-nitrosoguanidine (NTG)-induced temperature attenuated mutant strain 1B grown from the commercial live vaccines Cevac Chlamydia and Enzovax; strain 1H a reverted NTG mutant; and 5 strains isolated from cases of OEA originating from animals from the original vaccine safety trial (2 strains) or from vaccinated ewes or ewes exposed to vaccinated animals (3 strains).ResultsWe confirmed that AB7 is in a different lineage from the reference strain S26/3. The genome of vaccine strain 1B contains ten single nucleotide polymorphisms (SNPs) created by the NTG treatment, which are identical to those found in strain 1H. The strains from OEA cases also cluster phylogenetically very tightly with these vaccine strains.ConclusionsThe results show that C. abortus vaccine strain 1B has an identical genome sequence to the non-attenuated “reverted mutant” strain 1H. Thus, the protection of the 1B vaccine is unlikely to be due to the NTG induced SNPs and is more likely caused by the administration of high doses of C. abortus elementary bodies that stimulate protective immunity. Vaccine-identical strains were also isolated from cases of disease, as well as strains which had acquired 1–3 SNPs, including an animal that had not been vaccinated with either of the commercial live OEA vaccines, indicating that the 1B vaccine strain may be circulating and causing disease.     

Recombinant measles AIK-C strain expressing current wild-type hemagglutinin protein

We constructed a recombinant measles virus cDNA, pIC-MVAIK-H/87-K, in which the hemagglutinin (H) gene of the AIK-C vaccine strain was replaced by the wild-type (MVi/Tokyo.JPN/87-K: genotype D3) H gene and the remaining genes were the same as the AIK-C vaccine strain. To investigate the feasibility of the recombinant vaccine strain expressing wild-type H protein instead of the AIK-C H protein, we constructed two recombinant measles cDNA, having Leu (small plaque-type) and Phe (large plaque-type) at position 278 of the F protein. Infectious chimeric virus strains, MVAIK-H/87-K/S (small plaque-type) and MVAIK-H/87-K/L (large plaque-type), were recovered, which were designed to induce small (S) and large (L) plaques in Vero cells. The MVAIK-H/87-K/S and MVAIK-H/87-K/L did not grow at 39-40 degrees C, similar to the original AIK-C strain, and retained the temperature sensitivity (ts) characteristics. They did not induce cytopathic effect (CPE) in Vero cells but produced CPE in B95a cells, similar to the current wild-type measles MVi/Tokyo.JPN/87-K. From the results of Western blotting, the mobility of the H protein of MVAIK-H/87-K/S and MVAIK-H/87-K/L was similar to that of MVi/Tokyo.JPN/87-K. Hyper-immune sera raised by MVAIK-H/87-K/S neutralized all types of current wild strains. Thus, the chimeric measles virus expressing the current wild H protein demonstrated wild-type H properties with ts characteristics of the vaccine strain, indicating that the construction strategy of recombinant measles virus can cope with the hyper-mutated measles virus.     

Pathogenicity of Fusobacterium necrophorum strains from man and animals.

Necrobacillosis occurs in man and animals. The typical forms of the disease in animals are caused by Fusobacterium necrophorum biovar A; biovar B strains are much less pathogenic. In this study the pathogenicity for mice of eight human isolates of F. necrophorum was compared with that of animal biovar A and B strains. By subcutaneous inoculation seven of the human strains differed from biovar A but resembled biovar B in (1) producing, at the most, mild local lesions that rapidly healed, and (2) showing no enhancement of infectivity when suspended in sub-lethal doses of Staphylococcus aureus broth culture. The eighth human strain (A2433) resembled biovar A but differed from biovar B in (1) producing severe lesions, and (2) showing greatly enhanced infectivity in the presence of S. aureus. Nonetheless, strain A2433 differed from biovar A, both in the nature of the lesions produced and in its failure to cause severe general signs of illness and rapidly fatal infection. By intravenous inoculation one of two biovar B strains and all except one of the eight human strains produced purulent lesions, often severe, in the liver and elsewhere, but infection was not usually associated with general signs of illness. In contrast, intravenous injection of a biovar A strain gave rise to a rapidly fatal infection, with severe lesions in the liver or elsewhere. The results suggest that the term ''necrobacillosis'' as used in human and veterinary medicine refers to diseases that differ in important respects.     

人源和猪源猪链球菌的同源性研究

目的 进一步鉴定菌种;评价人源株与猪源株猪链球菌的同源性。方法 对分离自猪无菌部位、患者血液和脑脊液的７株猪链球菌Ⅱ型和标准猪链球菌Ⅱ型,用菌体脂肪酸分析和随机引物基因扩增技术进行菌种的表现型和基因型分类。对其结果进行聚类分析和主成分分析。结果 随机引物基因扩增技术分析提示,被检的７株猪链球菌Ⅱ型与标准株一致,均为猪链球菌Ⅱ型;人源株与猪源株同源;分离自病人血液和脑脊液的菌株同源。这些结果在菌体脂     

Evaluation of Brucella abortus S19 vaccines commercialized in Brazil: Immunogenicity,residual virulence and MLVA15 genotyping

Live attenuated Brucella abortus S19 is the most effective vaccine against brucellosis in cattle. The assessment of the immunological parameters is essential to guarantee the biological quality of live anti-bacteria vaccines. The evaluation of genetic stability of live bacterial vaccines is also important in quality control. The aims of the present study were to compare (i) the immunogenicity and residual virulence, and (ii) the genotypic profile (MLVA15) of the eight S19 vaccines commercialized in Brazil to the USDA S19 reference strain. Two batches of each of the eight S19 commercial vaccines used in Brazil (A–H) were tested. They were submitted to the potency and residual virulence in vivo tests recommended by OIE and typed by the multiple-locus variable-number tandem repeat (VNTR) analysis (MLVA) described for Brucella spp. Our results demonstrated that all S19 vaccines commercialized in Brazil would be approved by Brazilian and OIE recommendations for potency and residual virulence. Furthermore, the S19 vaccine is genetically very homogeneous, as all but two batches (from the same manufacturer) tested showed identical MLVA15 profile. The two batches with different profiles presented six repeat units in locus Bruce07, instead of the five found in all other strains, including the USDA S19 reference strain. Although presenting a slightly different profile, this vaccine was also protective, as demonstrated by the immunogenicity and residual virulence assays performed. Therefore, the commercial Brazilian S19 vaccines were in accordance to Brazilian and international standards for immunogenicity and residual virulence tests. Moreover, our results also show that MLVA could be a useful inclusion to the list of in vitro tests required by the official control authorities to be applied to the commercial S19 vaccines, as an efficient assay to guarantee the quality and stability of the vaccine strains.