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1.
Pasteurella multocida is a capsulated, gram-negative cocco-bacillus that can cause serious disease in a wide range of mammals and birds. P. multocida strains are classified into 16 serovars based on lipopolysaccharide (LPS) antigens. LPS is an essential virulence factor of P. multocida; mutants expressing severely truncated LPS are completely attenuated in chickens. LPS is also a major immunogen of P. multocida and protection against infections caused by P. multocida is generally considered to be serovar specific. In this review we summarize current knowledge of the structure and genetics of LPS assembly of P. multocida strains belonging to five different serovars. These include strains belonging to serovars 1 and 3, the most common serovars found in the poultry industry, and strains belonging serovars 2 and 5, the serovars associated with bovine haemorrhagic septicaemia outbreaks. A number of the serovars are genetically related; serovars 1 and 14 share the same LPS outer core biosynthesis locus, but due to a mutation within the phosphocholine biosynthesis gene, pcgA, the serovar 14 strain produces a truncated LPS structure. Similarly serovars 2 and 5 share an identical LPS outer core locus and express near-identical LPS structures. However, due to a single point mutation in the phosphoethanolamine (PEtn) transferase gene, lpt_3, the serovar 2 strain does not elaborate a PEtn residue on heptose II. Knowledge of the genetic basis for the LPS structures expressed by P. multocida will facilitate the development of rapid molecular methods for typing and diagnosis and will be essential for a rational approach to vaccine formulation.  相似文献   

2.
The susceptibility of two highly virulent (VP161 and VP138) and two less virulent (VP17 and VP21) strains of Pasteurella multocida to phagocytic uptake and killing by chicken macrophages was compared using in vitro phagocytosis and bactericidal assays. When compared with VP17 and VP21, particularly after they were preopsonised with specific immune serum, VP161 and VP138 were more resistant to phagocytosis by chicken macrophages. The uptake of these bacteria increased following the removal of the bacterial capsules with hyaluronidase. All strains preopsonised with specific immune serum were killed to some extent by chicken macrophages. However, the percentages of killing for VP17 and VP21 were higher than those of VP161 and VP138. When the capsules of VP161 and VP138 were removed, the susceptibility of the bacteria to bactericidal activity of chicken macrophages increased. It can be concluded that the virulent strains of P. multocida were more resistant to phagocytosis and phagocytic killing by chicken macrophages compared with the less virulent strains. The hyaluronic acid capsule was considered to be important in the resistance, but might not be the only factor contributing to the resistance since the less virulent strains of P. multocida also possess capsules.  相似文献   

3.
The role of a 39kDa protein of avian Pasteurella multocida in pathogenesis of fowl cholera was investigated using monoclonal antibodies (Mabs). Mabs were prepared by immunization of BALB/c mice with a crude capsular extract (CCE) of P. multocida strain P-1059 (serovar A:3). Totally eight hybridomas producing Mab were obtained. Immunoblot analysis of the hybridomas revealed that all the Mabs recognized a 39kDa protein of CCE. Treatment of CCE antigen with proteinase K or periodic acid indicated that the epitope recognized was proteinaceous. The Mabs reacted with a major 39kDa protein of CCE from encapsulated strains but not with any protein of non-capsulated strains indicating that a direct correlation between encapsulation and the 39kDa protein. Immunoelectron microscopy on strain P-1059 and the non-capsulated derivative P-1059B (serovar -:3) reacting with the Mabs and gold-labeled anti-mouse IgG indicated that the protein is associated with the capsule. The Mabs significantly inhibited the adherence of encapsulated P. multocida strains to chicken embryo fibroblast cells, but only slightly that of non-capsulated strains. Mice passively immunized with the Mabs were protected from lethal challenge with virulent strains P-1059 and X-73 (serovar A:1). Thus the capsular 39kDa protein was determined to be an adherence factor and a cross-protective antigen of avian P. multocida type A strains.  相似文献   

4.
A 39 kDa protein of avian Pasteurella multocida strain P-1059 (serovar A:3) was purified from a crude capsular extract by immunoaffinity chromatography by using a ligand of purified mouse monoclonal antibody to the 39 kDa capsular protein of the strain. Protective activity of the purified 39 kDa protein antigen was determined by inoculation of ddY mice twice with 25 or 125 mug of the protein and challenge-exposure with 10 or 50 LD(50) of strains P-1059 or X-73 (serovar A:1). The results showed that the antigen gave high protection (60 to 100%). These results indicated that the 39 kDa protein of avian P. multocida is a cross-protective antigen over serovars A:1 and A:3.  相似文献   

5.
The outer membrane protein (OMP) profiles of two strains of capsular type A Pasteurella multocida isolated from the lungs of pigs with enzootic pneumonia were studied. Sarkosyl extracted OMPs from P. multocida grown under iron-restricted and iron-replete conditions were subjected to sodium dodecyl sulfate-polyacrylamide gel electrophoresis and Western blot analysis. Results showed that the iron-regulated outer membrane proteins (IROMPs) with molecular masses of 74 kDa, 94 kDa, 99 kDa and 109 kDa were expressed by strain A52, while 74 kDa, 82 kDa, 94 kDa and 99 kDa IROMPs were expressed by strain B80. Swine immune sera, obtained from pigs which were first immunized with a polyvalent P. multocida type A and type D bacterin and subsequently challenged with type A strain of P. multocida, contained antibodies against the IROMPs. These antibodies cross-reacted with the IROMPs expressed by avian strain P1059 of P. multocida. Convalescent-phase serum obtained from turkeys which survived fowl cholera, also cross-reacted with the IROMPs from porcine strains of P. multocida. These results suggested that IROMPs from porcine and avian strains of P. multocida may share common epitopes that were recognized by swine immune serum as well as turkey convalescent-phase serum.  相似文献   

6.
为了解禽多杀性巴氏杆菌(Pasteurella multocida,Pm)分离菌株的荚膜血清型、菌体血清型与外膜蛋白型之间的相关性,首先对自行分离的10个菌株采用间接血凝试验和琼脂扩散试验进行鉴定(均为A:1);然后采用超声波破碎、高速离心和十二烷基肌氨酸钠提取外膜蛋白,通过SDS-PAGE电泳的方法对上述10个菌株与C48-1(A:1)、X73(A:1)、P1059(A:3)、CU(A:3,4)等一起进行外膜蛋白(Outer membrane proteins,OMP)分型研究。结果表明:14个禽多杀性巴氏杆菌菌株以2个主要蛋白OmpH和OmpA的差异为依据分为3种主要OMP型;依据次要蛋白的差异,OMP-1,3菌株进一步分为OMP型1.1,1.2和3.1,3.2,其中OMP型1.1有6个菌株,OMP型1.2有5个菌株;OMP型2有1株(YZ7031);P1059为OMP型3.1;CU为OMP型3.2;另外,血清型为A:1的12个菌株中有11个菌株外膜蛋白型均为OMP-1型,血清型为A:3、A:3,4的菌株外膜蛋白型属于3型。说明禽多杀性巴氏杆菌血清型与特定的外膜蛋白型具有很强的相关性。  相似文献   

7.
为确定禽多杀性巴氏杆菌(P.multocida)的保护性抗原外膜蛋白(OMPs)和脂多糖(LPS)在抵抗感染中的作用,本研究提取了禽P.multocida CVCC 474的OMPs和LPS成分,将该两种成分分别与弗氏佐剂混合制备免疫原,进行动物免疫。小鼠分为4组:OMPs组、LPS组、禽P.multocida弱毒活疫苗组和PBS对照组,每组16只。各组均免疫3次,每次间隔两周。间接ELISA检测免疫后小鼠血清特异性抗体水平,MTT法检测小鼠脾淋巴细胞增殖情况。以禽P.multocida强毒株CVCC 474进行攻毒,计算小鼠死亡数及保护率。实验结果表明,动物免疫后,OMPs组与LPS组免疫小鼠特异性血清抗体水平持续升高,与弱毒活疫苗组水平相近,与PBS组相比差异极显著(p<0.01)。脾淋巴细胞增殖试验表明,OMPs组、LPS组和弱毒活疫苗组的SI值极显著高于PBS对照组(p<0.01),但3个免疫组之间则无明显差异(p>0.05)。攻毒保护试验结果显示,OMPs免疫组的保护率与弱毒活疫苗相当,为8/10,高于LPS免疫组的保护率(7/10),表明OMPs作为研制禽P.multocida亚单位疫苗具有良好的...  相似文献   

8.
Capsules from a range of bacterial species have been shown to be major virulence determinants and capsule has been implicated in virulence in Pasteurella multocida. Moreover, capsular serogroup appears to be related to disease predilection. Haemorrhagic septicaemia strains belong to serogroup B and E, fowl cholera strains to serogroup A and atrophic rhinitis strains to serogroup D. The entire capsule biosynthetic locus of P. multocida A:1 has been cloned and its nucleotide sequence determined (Chung et al., 1998. FEMS Microbiol. Lett. 166, 289-296); however, nothing is known of the P. multocida B:2 capsule locus. In this work we have determined the nucleotide sequence and genetic organisation of the P. multocida M1404 (B:2) capsule locus. By analogy with the cap loci of other bacteria, the nucleotide sequence can be divided into three functional regions. Regions 1 and 3 comprise six genes involved in transport of the polysaccharide capsule to the cell surface. The deduced products of these genes show high similarity to proteins involved in capsule export in other bacteria. Region 2 comprises nine genes which are likely involved in biosynthesis of the polysaccharide capsule. The deduced products of three of these genes (bcbA, bcbB and bcbC) show significant similarity to proteins known to be involved in polysaccharide biosynthesis while the other six show no similarity to known proteins. However, their organisation indicates they are co-transcribed with bcbA, bcbB, bcbC and the Region 1 capsule export genes, suggesting strongly that they are also involved in capsule biosynthesis.  相似文献   

9.
The type 4 fimbrial subunit gene of pasteurella multocida   总被引：1，自引：0，他引：1  
Colonisation of host tissue by Gram- negative bacteria is facilitated by various adhesins, one of which is type 4 fimbriae (pili). These structures have been associated with pathogenesis in several bacterial species, and have been shown to mediate colonisation of epithelial surfaces. Recently, type 4 fimbriae were identified and characterised from P. multocida strains A, B and D. The type 4 fimbrial subunit protein (PtfA) was identified as an 18-kDa protein which was isolated from whole membrane fractions.We report here the isolation and characterisation of the gene (ptfA) encoding the PtfA protein from P. multocida VP161 (serotype A:1). Part of the gene was cloned on a 2-kb genomic DNA fragment. The complete ptfA gene was obtained using inverse PCR. The gene and its flanking regions were characterised, and the deduced PtfA amino acid sequence was compared to type 4 subunit protein sequences from other bacterial species. The ptfA gene was amplified and sequenced from several P. multocida strains. Comparison of these sequences revealed variation within the type 4 subunit gene of P. multocida.  相似文献   

10.
Chickens were protected against fowl cholera by ribosomal vaccines prepared from noncapsulated Pasteurella multocida. Passive hemagglutination (PHA) titers to lipopolysaccharide (LPS) and the degree of protection conferred by ribosomal vaccines were diminished or abolished when ribosomes were chromatographed on an immunoadsorbent column. Addition of subimmunogenic amounts of serotype 1 (homologous) LPS to highly purified ribosomes resulted in vaccines that protected against challenge exposure and produced PHA titers to homologous LPS. Addition of serotype 5 LPS to highly purified ribosomes did not protect chickens against challenge exposure with serotype 1 P multocida, but produced PHA titers to serotype 5 LPS. Combinations of serotype 1 ribosomal RNA and serotype 1 (homologous) LPS did not protect chickens or produce PHA titers to LPS. Purified ribosomes from Brucella abortus, Aspergillus fumigatus, and chicken liver were combined with LPS from P multocida and were evaluated as vaccines. Brucella abortus and A fumigatus ribosomes combined with LPS protected chickens as well as did bacterin made from whole cells of P multocida. Chicken liver ribosomes combined with LPS did not provide protection. To determine whether a protein carrier would substitute for ribosomes, methylated bovine albumin (MBA) was combined with LPS and evaluated as a vaccine. A serologic response to LPS was induced by MBA-LPS vaccine, but the vaccine offered no better protection than when LPS was used alone as vaccine. Ribosome-LPS vaccines produced serologic responses to LPS that were at least 5-fold greater than those produced by MBA-LPS vaccine.  相似文献   

11.
The relative virulence for chickens of five strains of Pasteurella multocida was evaluated. Twenty groups, each of ten chickens, were inoculated with a standard dose of 10(5) of each of five strains by the intramuscular (I.m.), intravenous (I.v.), intratracheal (I.tr.) or conjunctival (Co) routes. The highest mortality occurred in the groups dosed I.m. and I.v., followed by I.tr. inoculation. The relative virulence of each strain did not change when inoculated by the different routes. The most virulent strain, VP161, caused 100% mortality by all except the Co route. The least virulent strain, VP17, caused a single mortality by the I.v. route, but gave a high level of protection to birds inoculated by both the I.m. and I.v. routes, when challenged by intramuscular injection with (VP161). There was no protection against I.m. challenge in the birds inoculated by the I.tr. or Co routes. Serum antibody levels measured by ELISA correlated with the level of protection against virulent challenge for groups inoculated I.m. or I.v., but not I.tr. Western blots of pooled sera from each group did not show any specific antigen recognition that might explain the observed differences in protection. Inoculation with strain VP17, (both I.m. and I.tr.) also gave a high level of protection to birds challenged with strain VP161 by intratracheal instillation.  相似文献   

12.
Pasteurella multocida causes various respiratory disease symptoms in pigs, including atrophic rhinitis and pneumonia. In the present study, 69 strains of P. multocida were isolated from 443 pigs with respiratory clinical symptoms at 182 farms located throughout South Korea from 2009 to 2010. A multiplex capsular PCR typing assay revealed that 69 strains of P. multocida isolated in this study had the biosynthetic locus of the capsules of either serogroup A (47 strains, 68.1%) or serogroup D (22 strains, 31.9%). The 22 strains positive for serogroup D-specific primers were divided into four clusters and the 47 strains positive for serogroup A-specific primers were divided into 12 clusters according to the results of Random Amplified Polymorphic DNA (RAPD) analysis. P. multocida strains in the present study were susceptible to most of the antimicrobial agents used. An analysis of antimicrobial resistance and virulence gene pattern combined with RAPD indicated that a certain P. multocida strain appeared to be genetically identical, implying the persistence of the strain within a single farm.  相似文献   

13.
Capsule thickness of avian Pasteurella multocida type A strains was determined by transmission electron microscopy after labeling with polycationic ferritin and compared with their pathogenicity for chickens. The capsule thickness of P. multocida strains Pm-18 and X-73 was 81.4 and 50.1 nm on average, respectively. These strains were highly virulent for chicken, whereas the less virulent strains Pm-1 and Pm-3 had a thin and irregular capsule, 21.0 and 29.8 nm on average, respectively. However, the thickest capsule was observed in strain P-1059, 101.2 nm on average, and the strain revealed moderate virulence. The noncapsulated variant P-1059B, which was derived from strain P-1059, revealed low virulence. The six P. multocida strains were examined with regard to protein content on the capsule of organisms. Amounts of total proteins of crude capsular extract (CCE) from capsulated strains were approximately twice those of the noncapsulated strains. The amount of an antigenic 39 kDa protein in the CCE were found to correlate with the capsule thickness, since heavily capsulated strains exhibited the greatest amount, whereas noncapsulated strains including noncapsulated and low virulent variant P-1059B possessed little 39 kDa protein. The results demonstrated that the capsule thickness and the quantity of a 39 kDa capsular protein of avian P. multocida type A strains correlated with their pathogenicity for chickens.  相似文献   

14.
Lipopolysaccharides (LPS) were extracted from a serotype of each of 2 species of Pasteurella isolated from sheep with respiratory tract infections. Lipopolysaccharides from P haemolytica 82-25 (serotype 1A) or P multocida P-1573 (serotype 12) were mixed with sheep lung surfactant and were incubated for 6 hours at 37 C. After incubation, LPS-surfactant mixtures were centrifuged overnight in sucrose density gradients, and fractions were analyzed. Binding occurred between LPS and surfactant vesicles resulting in a stable complex with densities greater than those with the surfactant alone. The surfactant alone had a density of 1.052 to 1.060 g/ml. Diffuse bands of surfactant had a density of 1.075 to 1.092 when incubated with P haemolytica LPS and a density of 1.069 to 1.105 when incubated with P multocida LPS.  相似文献   

15.
Encapsulated avian strains of Pasteurella multocida possessing an A-type capsule were shown to be resistant to the bactericidal action of turkey serum, whereas unencapsulated variants as well as other unencapsulated strains were not. Removal of the capsule from serum-resistant strain P1059-1 resulted in this strain becoming susceptible to the bactericidal effects of turkey serum. Since complement was consumed when encapsulated or unencapsulated strain P1059-1 was incubated in turkey serum, we conclude that the capsule acts to shield the outer membrane rather than prohibiting the generation of an effective membrane attack complex.  相似文献   

16.
Pasteurella multocida and Pasteurella haemolytica produce specific proteins in the outer membrane under iron-depleted conditions. Pasteurella multocida serovar A expresses these proteins of molecular masses of 76 and 96 kDa as determined by electrophoresis. The analogous serovar D produces a further iron-regulated protein of 85 kDa. The Pasteurella haemolytica strains of serovar A1, A6 and T contain iron-regulated outer membrane proteins of molecular masses of 71, 77 and 100 kDa. These proteins possess binding positions for iron ions. Both Pasteurella multocida and Pasteurella haemolytica strains utilize iron from porcine and bovine transferrin, but not from haemin and haemoglobin.  相似文献   

17.
用EDTA—溶菌酶系统处理无荚膜的5:A型禽多杀性巴氏菌,得到无粘肽层的细胞膜.蔗糖密度梯度离心,将细胞膜分成3部分.琥珀酸脱氢酶、还原型辅酶I-2,6 二氯酚靛酚(NADH-DICP)氧化还原酶和NADH氧化酶的活性以及脂多糖含量测定表明,这3部分分别是细胞内膜、细胞外膜和混合膜.用高压液相色谱法从分离的外膜中提纯外膜蛋白,获得2个主要成分P_1和P_2.将P_1和P_2分别免疫小鼠,其保护率分别为20%和90%.SDS-聚丙烯酰胺凝胶电泳表明,P_2是一种主要的外膜蛋白,分子量为52000.  相似文献   

18.
The protein profiles of Pasteurella multocida serotype 1 isolates and the response of chickens to serotype 1 antigens were investigated using SDS-PAGE. Patterns obtained with Coomassie blue staining of soluble protein extracts were similar. The major difference between isolates was the position of one of the major proteins in the 34-38 kDa region. When chickens were experimentally infected with a clinical isolate of P. multocida serotype 1 various proteins were recognised by immunoblotting, including one with a relative molecular weight of 34 kDa; however, no reactions were observed in the region where LPS is known to migrate. When these infection sera were used in an EIA with purified LPS obtained from Heddleston serotype 1 type strain (X-73) they reacted strongly. Serum used for serotyping isolates in the gel diffusion precipitin test recognised many antigens in common with sera from infected birds, but some antigens were specific to typing sera.  相似文献   

19.
Outer membrane protein (OMP)-enriched extracts of avian strains of Pasteurella multocida were examined by use of sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Culture medium did not have a significant effect on the OMP profiles of strains of P multocida examined; however, in vivo propagation had an appreciable effect on the OMP profile composition of the reference strain P-1059. Such bacteria, expressed several additional OMP in the 27-kD, 48-kD, 56-kD, 60-kD, 80-kD, and 94-kD molecular mass regions. These OMP were not detected in the electrophorogram of strain P-1059 grown in vitro. The OMP profiles of reference strains of the 16 serotypes of P multocida did not identify any serotype-specific protein markers. Field strains of serotype A:3 had variation in OMP profiles and did not express OMP that all were identical to that expressed by the reference strain P-1059. The live attenuated CU and M9 bacterial vaccine strains expressed strain-specific OMP markers of 48-kD and 45-kD molecular masses, respectively. These strain-specific OMP markers may be used to differentiate these strains from virulent field strains that are of the same serotype and isolated from turkeys that have succumbed to pasteurellosis as a result of vaccine-related reactions or breakdown in immunity.  相似文献   

20.
In experimental fowl cholera, the intramuscular inoculation of Pasteurella multocida induces tissue damage that implies proteolytic or cytolytic activity of the bacteria. Such activity could not be demonstrated by conventional in vitro tests. The treatment of P. multocida strain VP21 with Tween-80 yielded an extract that lysed washed chicken red cells. Extracts were active to a maximum titre of 64. Haemolytic activity of the extract was neither affected by boiling nor by extremes of pH, indicating the active component was not a simple protein. Treatment with trypsin had no effect, but it was inactivated by Proteinase K. Yields were highest from bacteria grown in dextrose starch- or casein sucrose-yeast broths; were similar if cultured in air or anaerobically, but were reduced if the bacteria were grown in 5% CO(2). Haemolytic activity was eliminated on exposure to serum or serum albumen. The extract from strain VP21 haemolysed red cells from the chicken, rabbit, sheep, horse, bovine and human, with the highest titres observed on chicken cells. Six other avian strains and seven out of 10 strains of P. multocida from other species yielded an extract which haemolysed chicken red cells. The elaboration of this cytotoxic substance in vivo and its role in pathogenesis remains to be determined.  相似文献   

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