Further-attenuated measles vaccine: Virus passages affect viral surface protein expression,immunogenicity and histopathology pattern in vivo

Measles vaccine viruses Leningrad-16 (L-16) and Moscow-5 (M-5, an L-16-derived clonal variant), at passage levels used for vaccination and after ten further low-multiplicity passages on quail embryo (QE) cells, were compared for (1) immunogenicity, (2) histopathological lesions induced in vivo and (3) surface protein expression within infected cells and on the virion surface. At the 10th passage, viruses evoked a poorer neutralizing antibody response in guinea pigs, induced an earlier appearance of more pronounced pathological lesions and replicated faster in Vero cells that the original viruses. H protein expression increased 1.8–2.3-fold after 10 passages of the L-16 variant, but remained virtually unaltered for the M-5 variant. F protein expression of both 10th-passage variants was 0.5–0.8 that of the original virus variants. A similar two-fold decrease in F protein expression was noted after a single virus passage in guinea pigs. The data implicate the loss of F protein as a cause of reduced immunogenicity of further attenuated measles vaccines.     

The Brucella abortus vaccine strain B19 carries a deletion in the erythritol catabolic genes

Abstract Brucella abortus B19, an avirulent strain obtained by spontaneous mutation, is used worldwide as a vaccine for the control of bovine brucellosis. B19 differs from other B. abortus strains in its sensitivity to erythritol. We took advantage of a previously obtained erythritol sensitive Tn 5 insertion mutant of B. abortus 2308 to clone the chromosomal region containing erythritol catabolic genes from this representative pathogenic strain and from the vaccine strain B19. Physical mapping with restriction endonucleases and nucleotide sequence determination revealed the existence of a 702 bp long deletion, occurring between two short direct repeats, in the chromosome of B19. This deletion rendered the B19 strain sensitive to erythritol. Two oligonucleotides whose sequences flank this deletion provided an easy method to differentiate B19 from all other B. abortus isolates.     

狂犬病毒CTN—1株在Vero细胞上的适应传代研究

本文报导了用我国狂犬病毒固定毒人二倍体细胞适应株（ＣＴＮ－１）进行Ｖｅｒｏ细胞适应传代研究。通过连续传代培养,滴度可达８．０１ｏｇＬＤ５０／ｍｌ,达到了ＷＨＯ规定的不需浓缩的标准。病毒用０．０１ＭＯＩ感染细胞其产量与１Ｍｏｌ感染量相仿。病毒增殖高峰在４－５天,维持达１５天无明显下降,且可连续收获４－５次。因此,该毒种符合ＷＨＯ提出的疫苗生产毒种要求,可用于狂犬病疫苗生产。     

Protective activity of a cell-free Klebsiella vaccine in infection in mice caused by Streptococcus pneumoniae serotypes

The capacity of dried Klebsiella cell-free vaccine, obtained from strain No. 204 by the disintegration of microbial mass with hydroxylamine, for protecting mice from pneumococcal infection caused by S. pneumoniae, serotypes 3, 4 and 9N, has been studied. Klebsiella vaccine has been found to possess immunostimulating potency with respect to the S. pneumoniae serotypes under study. On day 5 this potency is manifested to a greater extent than 24 hours after immunization. The combination of Klebsiella vaccine with Proteus vulgaris, Staphylococcus aureus and Escherichia coli K-100 antigens enhances the stimulation of nonspecific resistance.     

Effect of immunization with an enterobacterial vaccine on the rate of plating out of Shigella from the intestinal mucosa of mice orally infected

The oral and subcutaneous immunization of mice with the vaccine prepared from S. minnesota strain R595, chemotype Re, known as enterobacterial vaccine, was found to significantly decrease the number of shigellae in cultures obtained by the inoculation of homogenized mucosal samples taken from the large intestine of mice, the vaccine prepared from S. minnesota isogeneous strain SF 1111 with the intact structure of lipopolysaccharide had no such activity. Antisera, obtained by the immunization of rabbits with enterobacterial vaccine, contained high titers of antibodies to Re-glycolipid and were capable of decreasing the isolation rate of shigellae from homogenized mucosal samples taken from the large intestine of mice; at the same time S. flexneri were found capable of binding with antibodies to glycolipid of Re-chemotype.     

Standardization of the seeding material in producing live anthrax vaccine]

Technology of obtaining dry concentrated seeding material of the anthrax bacillus STI-1 vaccine strain was worked out. The use of dry seeding material for making dry anthrax vaccine rendered the preparations obtained more standard, reduced the time required for their production, led to increase of AKM-SH productivity, and to greater profitability of the vaccine production. The vaccine preparations obtained with the use of dry seeding material did not differ from control by immunogenicity.     

The efficacy of killed Trypanosoma evansi vaccines in mice.

Four strains of Trypanosoma evarsi (D3, D4, D5 and D6) isolated from German shepherd dogs were inoculated into mice, and infected blood was used to prepare 9 separate killed vaccines. White mice inoculated with 1:100 diluted PBS vaccine, 0.5% carbol vaccine, or 100% Lugol vaccine showed survival rates of more than 60%. Among these 3 vaccines PBS vaccine and 0.5% carbol vaccine showed higher survival rates at 1:500 and 1:1000 dilutions, respectively. When young mice (15-20 g) were immunized with PBS vaccine, they resisted challenge with homologous strains, D3 strain in single injection, D6 strain in double injections and all strains in 5 injections. Protection however was not observed in old mice (25-30 g) give the same vaccine preparation. When mice were vaccinated with a single injection of D3 vaccine and challenged with heterologous strains, only those challenged with D4 strain at 10-5 dilution showed a survival rate of more than 60%. There was no difference in protective ability among PBS vaccine, agar adjuvant and kaolin adjuvant vaccines. Agglutinating antibody was demonstrated in mice receiving 5 injections of PBS vaccine.     

The possibility of creating a vaccinal strain of Brucella abortus 19-BA with multiple antibiotic resistance]

The possibility of preparing B. abortus vaccine strain 19-BA with multiresistance to antibiotics was shown. The strain was obtained by the spontaneous induction of resistance to rifampicin with the subsequent transformation of nonconjugative hybrid plasmid pOVI which, in addition to rifampicin resistance, governed the resistance of brucellae to tetracycline, doxycycline, ampicillin, and streptomycin. Experiments on guinea pigs demonstrated the immunization with both multiresistant vaccine strain GSA1 and B. abortus initial vaccine strain 19-BA.     

Beagle狗肾细胞及用于制备乙脑活疫苗的研究

Ｂｅａｇｌｅ乳狗（５－１０日龄）肾块用热消化法制成细胞批放液氮保存,检定合格后做下列试验：（１）复苏培养长满单层后更换维持液,其细胞能维持４天以上,克服了常规消化培养细胞维持时间短（３６小时）的缺陷。（２）传代１４－２株病毒时,１代病毒的滴度即达６．０４－６．２４ＬｏｇＰＦＵ／ｍｌ,１代后的各代（至１１代）病毒滴度无明显提高。（３）该细胞接种１４－２株病毒时不产生明显的破坏性病变（ＣＰＥ）,在收毒前采取冻溶１次比冻溶前的病毒滴度提高０．２７－０．５ＬｏｇＰＦＵ／ｍｌ。以ＢＤＫ细胞传１代的１４－２株病毒作生产毒种,收毒前冻溶１次等工艺法制备五批疫苗,全面检定结果均符合《乙脑活疫苗规程》的质量标准。其中病毒滴度为６．０９－６．２４ＬｏｇＰＦＵ／ｍｌ;免疫效价（ＩＤ５０＝ｍｌ）为１．９－４．０×１０－５,与相同滴度的原毒株１４－２ＰＨＫ苗对照无明显差异（ｔ＝０．９６８Ｐ＞０．０５）,表明其免疫原性与原毒株相似。     

The effect of aerosolization on the changes in major biological properties of the vaccine strain of Francisella tularensis

The exposure of F. tularensis vaccine strain 15/10 in the form of aerosol for more than 10 minutes results in the decrease of its virulence, immunogenicity and the content of species-specific antigen and in the increase of the dissociation level. Proceeding from this fact, during aerosol immunization with this vaccine strain exposure must not exceed 5 minutes for unstabilized aerosol and 10 minutes for aerosol stabilized with 5% glycerin. Under these conditions the properties of F. tularensis strain 15/10 are retained practically on the initial level.     

The new vaccine strains (or variants) of Francisella tularensis

Abstract A colony (N83) of the vaccine strain of Francisella tularensis (15/10) and a strain (N268) isolated from a water sample in nature were revealed for susceptibility to cultivation at 42°C. Both strains had low virulence for white mice and were avirulent for guinea pigs but possessed high immunogenicity in these animals. The spontaneous mutant of vaccine strain 15/10 showed resistance to doxicycline and rifmanpicine (15/10 Dox ^r40 Rif ^r40). The obtained mutant had biological characteristics similar to the parent vaccine strain. It provided immunity in experimental animals when vaccination and antimicrobial agents were used in combination.     

Attenuation of Chikungunya virus vaccine strain 181/clone 25 is determined by two amino acid substitutions in the E2 envelope glycoprotein

Chikungunya virus (CHIKV) is the mosquito-borne alphavirus that is the etiologic agent of massive outbreaks of arthralgic febrile illness that recently affected millions of people in Africa and Asia. The only CHIKV vaccine that has been tested in humans, strain 181/clone 25, is a live-attenuated derivative of Southeast Asian human isolate strain AF15561. The vaccine was immunogenic in phase I and II clinical trials; however, it induced transient arthralgia in 8% of the vaccinees. There are five amino acid differences between the vaccine and its parent, as well as five synonymous mutations, none of which involves cis-acting genome regions known to be responsible for replication or packaging. To identify the determinants of attenuation, we therefore tested the five nonsynonymous mutations by cloning them individually or in different combinations into infectious clones derived from two wild-type (WT) CHIKV strains, La Reunion and AF15561. Levels of virulence were compared with those of the WT strains and the vaccine strain in two different murine models: infant CD1 and adult A129 mice. An attenuated phenotype indistinguishable from that of the 181/clone 25 vaccine strain was obtained by the simultaneous expression of two E2 glycoprotein substitutions, with intermediate levels of attenuation obtained with the single E2 mutations. The other three amino acid mutations, in nsP1, 6K, and E1, did not have a detectable effect on CHIKV virulence. These results indicate that the attenuation of strain 181/clone 25 is mediated by two point mutations, explaining the phenotypic instability observed in human vaccinees and also in our studies.     

轮状病毒基因重配LH9株(G4型)传代稳定性研究

对兰州生物制品研究所自行构建的轮状病毒基因重配株G4型LH9株在Vero细胞中传代的稳定性进行研究。将基因重配LH9毒株接种于Vero细胞上从7代传至17代,经电镜检查、病毒滴度测定、病毒基因组RNA电泳图谱分析、基因序列测定分析、轮状病毒型别鉴定及其它相关检定,结果均符合药典要求。证实该毒株为A群G4P〔12〕型轮状病毒,在Vero细胞传代中具有良好的稳定性,为疫苗研制提供安全有效的依据。     

LIVE TULAREMIA VACCINE I. : Host-Parasite Relationship in Monkeys Vaccinated Intracutaneously or Aerogenically.

Eigelsbach, H. T. (Fort Detrick, Frederick, Md.), J. J. Tulis, M. H. McGavran and J. D. White. Live tularemia vaccine. I. Host-parasite relationship in monkeys vaccinated intracutaneously or aerogenically. J. Bacteriol. 84:1020-1027. 1962.-Bacteriological, histological, immunohistochemical, and serological studies were made on monkeys administered live tularemia vaccine strain LVS by either of two routes. Comparative data are presented on nonvaccinated monkeys exposed via the respiratory route to a highly virulent strain of Pasteurella tularensis. Tissue changes resulting from either aerogenic or intracutaneous vaccination were mild, and consisted primarily of the proliferation of histiocytes without the formation of granulomas. The vaccine strain was isolated from the site of vaccination of animals inoculated dermally, from the lungs of animals vaccinated aerogenically, and from the regional lymph nodes, liver, and spleen of both groups; it was not isolated from the blood or bone marrow. Proliferation of the vaccine strain at the site of dermal inoculation and in the lungs of animals exposed aerogenically was observed within 24 hr; in both groups, the maximal viable population was reached within 3 days and maintained through the 10th day. A reduction in the number of viable vaccine organisms had begun by the 14th day; isolations were obtained only from the regional lymph nodes on the 28th day, and the vaccine strain was not isolated from any of the tissues cultured on the 90th day. Because the monkey is less resistant to tularemia than is man, the benign response of this animal to live tularemia vaccine indicates that the vaccine might also be safe for man when administered by either the dermal or respiratory route.     

Nucleotide sequence of a neurovirulent variant of the type 2 oral poliovirus vaccine.

Infectious cDNAs of the Sabin type 2 poliovirus vaccine virus and a vaccine-derived neurovirulent type 2 strain (P2/117) have been cloned in Escherichia coli. Nucleotide sequence analysis revealed that P2/117 differs from the vaccine strain by just 23 point mutations. Three occur in the 5' noncoding region. The remainder result in a total of 5 coding changes located in VP1, VP4, 2B, and 3D. The likely role of these mutations in the evolution to neurovirulence is discussed.     

A comparison of complete genome sequences of a rabies virus chinese isolate SH06 with the vaccine strains

In this study, we determined the complete nucleotide and deduced amino acid sequence of a primary isolate of rabies virus (SH06) obtained from the brain of a rabid dog. The overall length of the genome was 11 924 nucleotides. Comparison of the genomic sequence showed the homology of SH06 at nucleotide level with full-length genomes of reference vaccine strains ranged from 82.2% with the PV strain to 86.9% with the CTN strain. A full-length genome-based phylogenetic analysis was performed with sequences available from GenBank. Phylogenetic analysis of the complete genome sequences indicated that the SH06 exhibited the highest homology with rabies street virus BD06 and CTN vaccine strain originated from China.     

Subtyping of European foot-and-mouth disease virus strains by nucleotide sequence determination

The VP1-coding regions of foot-and-mouth disease virus strains from 18 recent European outbreaks and of 9 strains isolated more than 20 years ago and used in part as vaccines were determined by direct cDNA sequencing. Comparison of the sequences revealed that most of the isolated outbreak viruses are closely related to the vaccine strains used. Isolates from the Italian epizootic of 1984 to 1985 correspond, for example, to the vaccine strain A5 Parma 62; the outbreak in 1984 in Bernbeuren, Federal Republic of Germany, was induced by A5 Allier 60; outbreaks in 1982 in Funen, Denmark, and in Murchin, German Democratic Republic, were caused by O1 Lausanne 65. Viruses isolated during the 1983 Iberian epizootic show a close relationship to the vaccine strain A5 Allier 60 but were probably derived from another not yet identified vaccine strain from Spain. Only two minor outbreaks in the Federal Republic of Germany, A Aachen in 1976 and O Wuppertal in 1982, did not correspond to the classical European strains but were obviously introduced from outside. We suggest that nucleotide sequence analysis should be used as a standard method of diagnosis, because when compared with other techniques it more clearly reveals the origin and course of epizootics and offers the possibility of preventing further outbreaks.     

马传染性贫血弱毒疫苗株核衣壳蛋白基因的表达与鉴定

从感染驴白细胞的马传染性贫血弱毒疫苗株前病毒DNA中克隆了编码核衣壳蛋白 (pll)的基因 ,在大肠杆菌中得到了表达 ,而表达的蛋白是一种可溶性的融合蛋白 ,其氨基端带有 6个组氨酸的标签 ,因此可以用固定化金属离子亲和层析法在非变性条件下进行纯化。经间接ELISA和免疫印迹试验检测 ,这种表达的融合蛋白可与马传贫阳性血清样品发生反应 ,而与健康马血清无任何反应 ,显示其具有良好的抗原性和特异性 ,可用于马传贫弱毒疫苗株在体内外复制及在接种马体内免疫应答的研究。     

马传贫驴白细胞弱毒疫苗株酸性蛋白p9基因的克隆与表达

从感染驴白细胞的马传贫驴白细胞弱毒疫苗株前病毒DNA中克隆了编码p9蛋白的基因,并在大肠杆菌中进行了表达。所表达的蛋白是一种可溶性的融合蛋白,其氨基端带有6个组氨酸的标签,因此可以用固定化金属离子亲和层析法在非变性条件下进行纯化。在间接酶联免疫吸附试验(ELISA)和免疫印迹试验中,重组的酸性蛋白p9可与马传贫阳性血清样品发生反应,而与健康马血清无任何反应。这表明该重组蛋白具有良好的抗原性和特异性,可用于马传贫弱毒疫苗株在体内外复制及在接种马体内免疫应答的研究 。     

Antigenic activity and the reactogenicity of a concentrated, purified, UV-inactivated cultured rabies vaccine

The results of the study of concentrated, purified, UV-inactivated cell-culture rabies vaccine, obtained from strain Vnukovo-22, passage 33-40, in the primary culture of Syrian hamster kidney cells, demonstrated the pronounced antigenic potency of this vaccine: when introduced intramuscularly in 3-4 injections at certain intervals, it induced the production of virus-neutralizing antibodies in high titers. In tests on volunteers the vaccine proved to be nonreactogenic.