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1.
Western blot analysis was performed from the culture supernatant of 59 rabbit Staphylococcus aureus strains, classified as high and low virulence strains according to their epidemiological behaviour in commercial rabbitries, bio-, phage- and RAPD-type. Fourteen extracellular antigen bands (A-N) were recognised using sera of rabbits immunised with washed, viable high virulence S. aureus bacteria. Eleven of these bands were found in high virulence as well as in low virulence strains. The band A, approximately 78 kDa, was not seen in any of the 27 high virulence strains, except for one strain which was also typical in other aspects, was detected in all, but one of the low virulence strains. The M and N bands with molecular masses of approximately 29 and 27 kDa, respectively, were recognised in all high virulence strains except for the atypical strain, but in none of the low virulence strains. This indicates that the latter two antigens may be virulence-associated markers for S. aureus strains from rabbits.  相似文献   

2.
In the present study, an in vivo rabbit skin infection model was developed to reproduce the lesions caused by high and low virulence Staphylococcus aureus strains from rabbits. "O"-shaped dermal skin lesions were induced on the shaved flanks of anaesthetised rabbits using a tattoo pin and pincers. The induced lesions on the flanks of four groups of 10 rabbits were then inoculated by topical application of 0.1 ml of 10(8)cfu S. aureus bacteria. One group was inoculated with a typical high virulence (HV) S. aureus strain from rabbits, one group received an atypical HV strain and two groups were inoculated with low virulence (LV) strains. Five animals were kept as negative controls. The development, appearance and size of abscesses were scored daily for a period of 2 weeks. The infection model showed reproducible results for the different S. aureus inoculation groups. Inoculation of the skin with the typical HV strain resulted in significantly larger abscesses than those caused by the LV strains. The atypical HV strain caused abscesses of a size intermediate to that obtained with the HV and LV strains. In rabbits infected with LV strains, most of the lesions had healed by day 14 post-inoculation. The devised infection model is able to reliably reproduce the virulence properties of HV and LV S. aureus strains.  相似文献   

3.
Staphylococcus aureus is an important cause of pododermatitis, subcutaneous abscesses and mastitis in rabbits. On rabbitry level, two types of S. aureus infections can be distinguished. In the first type, caused by low virulence strains, the infection affects only a small number of animals. The second type of infection is caused by high virulence strains and spreads throughout the rabbit flock. The pathogenic capacity of a particular S. aureus strain is attributed to a combination of invasive properties and extracellular factors such as toxin production. Therefore, 22 high virulence and 37 low virulence S. aureus isolates were compared regarding the prevalence of genes encoding exfoliative toxins, leucotoxins and superantigens. This study revealed a clearly significant difference between HV and LV rabbit S. aureus strains. All typical HV isolates were positive for the egc cluster, containing the enterotoxin(like) genes seg, sei, selm, seln, selo and selu, whereas these genes were not detected in any of the LV isolates. Further research is necessary to clarify the importance of the egc cluster in the pathogenesis of infections with high virulence S. aureus strains in rabbits.  相似文献   

4.
At rabbit flock level, two types of Staphylococcus aureus infections can be distinguished. In the first type, caused by low virulence strains, the infection remains limited to a small number of animals. The second type of infection is caused by the high virulence strains, which spread throughout the rabbitry. The pathogenetic capacity of a particular S. aureus strain is attributed to a combination of extracellular factors and invasive properties such as adherence and biofilm formation. Twenty eight high virulence and 34 low virulence S. aureus isolates recovered from rabbits between 1998 and 2003 were used to study slime production on Congo red Agar (CRA) and prevalence of bap, icaA and icaD associated with biofilm formation. Furthermore these strains were screened for the presence of bbp, clfA, clfB, cna, ebpS, eno, fnbA, fnbB and fib encoding microbial surface components recognizing adhesive matrix molecules (MSCRAMMs). The presence of icaA and icaD was not correlated with slime production on CRA. Bap was absent in all strains. All rabbit S. aureus strains harboured clfA and clfB. The prevalences of ebpS, eno, fnbA and fib did not reveal striking differences between high and low virulence strains. FnbB prevalence in high virulence isolates was lower than in low virulence isolates and cna was absent in high virulence strains. It was remarkable that only high virulence strains were positive for bbp. Further research is necessary to elucidate the significance of bbp in the pathogenesis of high virulence strains.  相似文献   

5.
High virulence rabbit Staphylococcus aureus strains cause chronic and spreading problems of mastitis, pododermatitis and subcutaneous abscesses on rabbit flock level, whereas infections with low virulence strains are limited to individual rabbits. In the present report, 13 high virulence rabbit S. aureus strains, selected out of a large collection of strains isolated in five European countries between 1983 and 2004, were genotyped using pulsed-field gel electrophoresis, spa typing, multilocus sequence typing (MLST) and accessory gene regulator (agr) group typing. Two low virulence rabbit S. aureus strains were also included in the study. The results indicate the clonal origin of high virulence rabbit S. aureus strains present in Europe. Furthermore, the results of MLST and spa typing form a basis for international epidemiology of rabbit S. aureus strains, as these DNA sequence-based typing techniques can easily be used for intercentre comparisons.  相似文献   

6.
High virulence rabbit Staphylococcus aureus strains, which are clonal in origin, are responsible for the spread of chronic staphylococcosis at the rabbit flock level. The aim of the present study was to develop a multiplex PCR assay that can be used for the identification of these high virulence strains. Two targets of the assay were the bbp and the selm genes, which have recently been shown to occur specifically in high virulence isolates. A third target was a sequence designated "flank", which was derived from a previously generated high virulence specific RAPD pattern. Furthermore, the femA gene, which is specific for S. aureus, was incorporated in order to avoid false negative results due to insufficient DNA preparation. The multiplex PCR was successful at differentiating the 26 typical high virulence and 50 low virulence rabbit S. aureus strains incorporated in the present study. Therefore it is useful for the initial screening of newly acquired breeding stock, in order to prevent the intake of high virulence strains in rabbitries.  相似文献   

7.
Four groups of 12 rabbits each were inoculated in the nose with strains with suspected differences in virulence. In the two groups infected with strains from severe outbreaks, belonging to a rabbit-pathogenic biotype-phage type combination, 6-12 rabbits were found positive at the successive bacteriological samplings over a period of 28 days. In the two other groups, infected with strains obtained from rabbitries without a history of staphylococcosis, the number of Staphylococcus aureus positive animals quickly became negative but increased again after 1 week to 1-5 positive animals until the end of the experiment. Two rabbits in each group inoculated with a high virulence strain developed purulent skin lesions, while in the groups inoculated with low virulence strains, all animals remained clinically healthy. Results indicate that colonisation capacity is an important virulence determinant in rabbit staphylococcosis.  相似文献   

8.
Bacterial superantigens are one of the major virulence factors produced by Streptococcus pyogenes and Staphylococcus aureus. The two novel superantigen encoding genes seeM and seeL were described for S. equi subsp. equi which is known as the causative agent of strangles in equids. In the present study previously characterized S. equi subsp. equi strains and strains of various other animal pathogenic streptococcal species and subspecies were investigated for the presence of the superantigen encoding genes seeM and seeL by polymerase chain reaction. According to these studies seeL and seeM appeared to be a constant characteristic of all investigated S. equi subsp. equi strains. Surprisingly, one S. equi subsp. zooepidemicus strain (S.z. 122) was also positive for both genes. The species identity of this S. equi subsp. zooepidemicus strain could additionally be confirmed by sequencing the 16S rRNA gene and the 16S-23S rDNA intergenic spacer region. The superantigen encoding genes could not be found among additionally investigated S. equi subsp. zooepidemicus strains or among strains of seven other streptococcal species. The seeL and seeM genes of the S. equi subsp. equi strain S.e. CF32 and the genes szeL and szeM of the S. equi subsp. zooepidemicus strain S.z. 122 were cloned and sequenced. A sequence comparison revealed a high degree of sequence homology between seeL, szeL, speL and seeM, szeM and speM, respectively. The superantigenic toxins L and M seemed to be widely distributed virulence factors of S. equi subsp. equi, rare among S. equi subsp. zooepidemicus but did not occur among a number of other animal pathogenic streptococcal species.  相似文献   

9.
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10.
Sodium-dodecyl-sulphate polyacrylamide gel electrophoresis (SDS-PAGE) was used to study the protein variability of Mycoplasma hyopneumoniae isolates. Fifty-six M. hyopneumoniae isolates from 6 different countries and 37 different herds were used. From eight herds, more than one isolate was available. All SDS-PAGE patterns of isolates originating from different herds were clearly divergent. Intra-species protein variability was quantified using the reference strain J and seven field strains all obtained from different herds and classified according to virulence. Between the field strains, a variability of 25% was found, while the culture-adapted strain J was clearly divergent and showed 30% variability with the field strains. No clustering according to virulence was obtained, but a protein band of about 181 kDa was present in the two highly virulent isolates whereas this protein band was absent in the moderately and low virulent isolates. Protein patterns of isolates derived from different animals from the same herd, were identical or differed in only a few protein bands. This study clearly indicates that, in agreement with previous studies on genomic diversity of M. hyopneumoniae isolates, proteomic variability within the species is high. Our study did not find clear evidence that more than one M. hyopneumoniae isolate circulates within a herd at a specific time point. The minor differences found between M. hyopneumoniae isolates from the same herd might reflect the organism's ability to alter its proteomic expression profile under field conditions.  相似文献   

11.
In the present study 20 staphylococci isolated from lymph node abscesses of 19 goats of two herds in Western Poland could be identified as Staphylococcus aureus subsp. anaerobius. All 20 strains grew under microaerobic conditions, were negative in the catalase test, showed the typical phenotypic properties of 5. aureus and could genotypically be identified by a positive sa442, 235 rDNA, nuc, coa and spa PCR reaction. The variable regions of the coa and spa gene of the 20 strains appeared with uniform amplicon sizes, respectively. All 20 strains were negative for 12 additionally investigated enterotoxin encoding genes, tst and ssl7 and positive for the gene cap8. Identical properties could be observed for S. aureus subsp. anaerobius DSM 20714. Amplification and sequencing of kat gene of a single Staphylococcus aureus subsp. anaerobius strain of the present study and S. aureus subsp. anaerobius DSM 20714 revealed a complete identity of the kat sequences of both strains and a katB sequence obtained from GenBank (AJ000471). The bacteria were additionally investigated for relatedness by macrorestriction analysis of chromosomal DNA with subsequent pulsed-field gel electrophoresis (PFGE), yielding, corresponding to the above mentioned PCR results, identical PFGE patterns for all 20 Staphylococcus aureus subsp. anaerobius strains isolated in Western Poland and the S. aureus subsp. anaerobius reference strain DSM 20714.This indicates the clonal identity of the strains isolated in Western Poland and the S. aureus subsp. anaerobius reference strain. The route of infection of the two herds in Western Poland with a bacterial clone originally isolated in Spain remains unclear.  相似文献   

12.
根据GenBank上公布的金黄色葡萄球菌肠毒素A（staphylococcus enterotoxin A,SEA）基因的全序列,设计一对特异性引物扩增内蒙古分离株的SEA基因序列。经基因克隆和序列测定,表明扩增的基因片段长度为582 bp,与标准菌株ATCC13565的SEA基因片段序列相似性为100%,与GenBank上公布的金黄色葡萄球菌菌株（EF520720.1）SEA基因相似性达到99.14%。本研究结果为进一步研究建立牛乳中SEA分子检测技术奠定了实验基础。  相似文献   

13.
Recently whole genome sequencing of Staphylococcus aureus has revealed the genes encoding cysteine proteases such as staphopain and SspB. In this study, we cloned and sequenced the structural gene (ScpA) encoding a cysteine (thiol) protease of S. aureus strain CH-91 from a chicken with dermatitis using polymerase chain reaction (PCR) and inverse PCR methods. The sequence information revealed a coding sequence (CDS) of 1200 nucleotides encoding the ScpA preproenzyme of 399 amino acids with a molecular mass of 45,071 Da. The deduced amino acid sequence of the ScpA differed at many positions from those of staphopain and SspB with identities of 64 and 42%, respectively. In the Southern blot analysis with a total DNA of S. aureus strain CH-91, the ScpA probe hybridized with a single 7.7 kb XbaI fragment or 2.8 and 0.8 kb EcoRI fragments, whereas the staphopain and SspB probes did not hybridize with these DNA fragments. These results suggest that this ScpA gene is a single-copy gene and is a novel gene, which is not found in the published whole genome sequences of S. aureus. In immunoblot, PCR, and Southern blot assays, the ScpA or its gene was detected in high protease-producing strains from chickens, but was not recognized in bovine and porcine strains or low protease-producing avian strains. These results indicate that the ScpA of CH-91 type may be specific to the high protease-producing strains of S. aureus from chickens, namely, there is a strain specificity of the ScpA.  相似文献   

14.
The virulence towards mice of Staphylococcus aureus strains from bovine mastitis was enhanced upon growth in milk whey compared to homologous organisms grown in tryptic soy broth (TSB). In the mouse mastitis model, S. aureus grown in milk whey caused more severe lesions than homologous strains grown in TSB. Staphylococcus aureus strain F1440 grown in milk whey induced 75% mortality and local necrotic reaction in subcutaneously inoculated mice, whereas the homologous strain grown in TSB caused only 5% mortality and slight skin reaction. Extracellular capsule on milk whey-grown, S. aureus could not be demonstrated. However, diffuse type colony morphology could be correlated with an increased virulence of S. aureus towards mice.  相似文献   

15.
为分析猪链球菌4型(Streptococcus suis type 4,SS4)分离株的病原生物学特性,试验对来自中国不同地区的10株猪链球菌4型进行了多位点序列分型,通过PCR方法对猪链球菌7个保守管家基因aroA、gki、dpr、mutS、recA、thrA、cpn60进行扩增,测序后将结果上传至MLST数据库查找序列型,然后制作聚类分析图来阐明菌株之间的亲缘关系;采用PCR方法对7种主要的毒力基因gdh、mrp、epf、sly、fbps、gapdh与orf2进行鉴定,通过毒力因子谱来分析猪链球菌4型毒力因子的分布;以纯化的10株细菌对BALB/c小鼠进行动物致病性试验,根据小鼠致死数量筛选出最强毒株,并进行新西兰兔致病性试验。结果显示,10株猪链球菌4型经多位点序列分型,6株为ST850型,3株为ST1006型,1株为ST94型;结合菌株分离地区分析发现,广东和江苏地区菌株有较高的同源性,江沪地区菌株出现分化现象,表现为遗传多样性;10株菌株均检测到了gdh、gapdh和orf2毒力基因,7株检测到sly基因,4株检测到fbps基因,根据毒力因子谱发现共有3个毒力基因型,gdh+sly+gapdh+orf2+型有6株,gdh+fbps+gapdh+orf2+型有3株,gdh+sly+fbps+gapdh+orf2+型仅有1株。动物致病性试验表明,10株细菌均能使BALB/c小鼠死亡,其中SH1510的半数致死量低至1×108 CFU,对小鼠的致病性最强,将纯化的SH1510菌液接种新西兰兔,可使新西兰兔出现典型的神经症状并死亡。以上结果为猪链球菌的遗传进化、毒力研究提供了新的数据,丰富了猪链球菌病的研究。  相似文献   

16.
Based on our clinical experience on bovine mastitis, we hypothesized that subtypes of Staphylococcus aureus (S. aureus) exist which differ in their contagious and pathogenic properties. In order to investigate this hypothesis, we analyzed strains of S. aureus isolated from spontaneous intramammary infection (IMI) with their virulence gene patterns and genotypes obtained by PCR amplification of the 16S-23S rRNA intergenic spacer (RS-PCR). The genotypes were then associated with epidemiological and clinical data including 26 herds. The results demonstrated a high association between genotypes and virulence gene patterns as well as between epidemiological and pathogenic properties of S. aureus. In particular, genotype B was related to high contagiosity and increased pathogenicity whereas the other types (C, OG) were found with infection of single cows. Because of the high clinical relevance, our results indicate the need to subtype the IMI-associated strains of S. aureus in the future.  相似文献   

17.
We investigated the ability of a high virulence (STR 357) and a low virulence (STR 598) strain of Streptococcus gallolyticus to attach to the intestinal tract of pigeons. For that purpose, first of all, two groups of six pigeons were anesthetized and ligatures were placed at the beginning of duodenum, jejunum, ileum, and colon. The obtained intestinal loops of the birds of the first and second group were injected with S. gallolyticus strains STR 357 and STR 598, respectively. At 15, 30, and 60 min postinoculation, two pigeons of each group were euthanatized and the various intestinal loops were sampled for histologic, immunohistochemical, and electron microscopic examination. Both the high and low virulence strains were able to adhere to the intestinal mucosa. Indeed, all samples dearly showed numerous coccal-shaped bacteria that stained positively with S. gallolyticus antiserum and were lining up against the intestinal epithelium. Likewise, on electron microscopic examination, cocci were seen in the mucus covering the intestinal epithelium. Second, the association of S. gallyticus strains of differing virulence with the intestinal tissue was determined quantitatively. Experiments were performed as described above. The number of S. gallolyticus bacteria that adhered to the intestinal epithelium was determined by plating out 10-fold serial dilutions of the segments. No significant differences in the number of adhered bacteria were found between the strains of high and low virulence.  相似文献   

18.
为了解陕西省部分地区猪流行性腹泻病毒(PEDV)的遗传和变异情况,采集陕西省部分地区规模化猪场的5份疑似PEDV感染的猪小肠内容物,进行PEDV S、M和N基因的RT-PCR扩增,并对扩增产物进行序列测定和遗传变异分析。结果表明,5份病料均能扩增出PEDV S、M和N基因,5株病毒分别命名为SXSL、SX-BJ、SX-YL、SX-WN和SX-HZ株。序列分析表明,5株毒株之间的S、M和N基因核苷酸序列的同源性分别为96.7%~99.8%、98.4%~100%和97.2%~99.9%;氨基酸序列的同源性分别为97.4%~99.9%、98.2%~100%和98.2%~100%。该5株病毒与中国疫苗株CV777的S、M和N基因核苷酸序列的同源性分别为93.9%~99.8%、98.1%~100%和95.3%~99.9%,氨基酸序列的同源性为93.6%~99.9%、96.2%~100%和98.2%~100%。遗传进化分析结果显示,5个陕西分离株的S基因与中国疫苗株CV777亲缘关系较远,与近年来中国株、日本株以及韩国株亲缘关系较近。SX-SL株、SX-BJ株和SX-YL株的M和N基因与中国疫苗株CV777亲缘关系较近,且与中国株CHGD-01亲缘关系密切。SX-WN株和SX-HZ株的M和N基因与中国疫苗株CV777亲缘关系较远。该5株病毒的S基因以及SX-WN株和SX-HZ株的M基因和N基因变异程度较大,而SX-SL株、SX-BJ株和SX-YL株三个流行株均与中国株CHGD-01亲缘关系密切,并且与近年在陕西省流行的PEDV也不完全相同。  相似文献   

19.
为了全面了解犬冠状病毒（CCoV）分离毒株JS1706和JS1712基因组3'端主要结构蛋白基因和非结构蛋白基因的分子特征,本研究设计了8组引物进行RT-PCR扩增,产物经测序和拼接后,获得了约8.7 kb基因组片段,该基因组结构及其编码蛋白顺序为5'-S-3abc-E-M-N-7ab-3'。对CCoV JS1706、JS1712株8.7 kb基因组核苷酸序列与α冠状病毒属参考毒株的相同区域核苷酸序列进行比对,结果表明,2个分离株与CCoV Ⅱ型参考毒株相似性最高（83.4%~93.1%）,其次为FCoV Ⅱ型参考毒株（87.1%~87.9%）、TGEV参考毒株（86.1%~86.8%）、CCoV Ⅰ型参考毒株（72.0%~72.1%）和FCoV Ⅰ型参考毒株（67.5%~69.9%）。JS1706、JS1712毒株与同属冠状病毒参考株的结构蛋白S、E、M和N蛋白氨基酸相似性分别为46.4%~95.2%、75.6%~100%、82.8%~99.2%和78.5%~99.7%。说明同属内冠状病毒的S基因变异度大,E、M、N基因相对保守。根据基因组3'端8.7 kb核苷酸序列和S蛋白氨基酸序列相似性比对结果,JS1706和JS1712毒株均与泛嗜型原型株CB/05相似性最高,分别为93.0%~93.1%、94.8%~95.2%,其他结构蛋白包括E、M和N氨基酸序列比对也发现与CB/05株的相似性较高,分别为97.6%~100%、92.4%~93.1%和97.9%。S蛋白氨基酸序列的进一步分析表明,JS1706和JS1712毒株的S蛋白N端有一些特有氨基酸,S蛋白氨基酸序列中没有明显的S1/S2蛋白酶切位点（RRARR）,但在958—963位氨基酸有S2'裂解位点特征基序（KRKYRS）。基于S蛋白氨基酸序列构建的系统发育进化树分析显示,CCoV JS1706和JS1712株与CCoV Ⅱa亚型参考毒株和FCoV Ⅱ型参考毒株聚集形成一个分枝。CCoV JS1706和JS1712株非结构蛋白的编码基因ORF3abc和ORF7,其结构、大小与经典疫苗株INSAVC-1相似,无明显插入、缺失和移码突变。本研究有助于深入了解国内CCoV流行毒株的分子特性,为后续分子流行病学调查、诊断试剂和疫苗研发奠定了基础。  相似文献   

20.
Randomly Amplified Polymorphic DNA (RAPD) typing was performed on 53 rabbit Staphylococcus aureus strains. Twenty-three strains isolated in 13 different rabbitries with chronic problems of staphylococcosis, showed the same RAPD banding pattern. Twenty of these strains belonged to the 'mixed CV-C' biotype and to the phage-type 3A/3C/55/71, previously described to be highly virulent in rabbits, and three strains belonged to other biotypes or phage-types. None of the strains isolated from rabbitries without chronic problems of staphylococcosis showed this specific RAPD pattern. RAPD analysis can be used as a rapid and reliable test method to differentiate between the characteristic genotype corresponding to high virulence and other S. aureus strains from rabbits. This is useful for the diagnosis and prevention of the introduction of these highly virulent strains in industrial rabbitries.  相似文献   

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