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1.
The use of an enzyme-linked immunosorbent assay (ELISA) as a means of detecting dermonecrotoxin-producing strains of Pasteurella multocida was investigated. The assay was evaluated as a means to identify toxigenic P. multocida isolates recovered from nasal secretions of swine with atrophic rhinitis. The sensitivity and specificity of the ELISA for detecting dermonecrotoxin-producing P. multocida strains were compared to those of mouse-inoculation and cytotoxicity assays. The ELISA was highly sensitive and more specific than animal inoculation or tissue culture assay and is thus a more effective method for screening swine herds for the presence of toxigenic strains of P. multocida. The ELISA is a rapid, effective, economical way to identify toxigenic P. multocida isolates.  相似文献   

2.
Heat-labile toxin from a cell sonicate of a virulent type-D strain of Pasteurella multocida was purified by ammonium sulfate precipitation followed by ion exchange chromatography, gel filtration chromatography, and polyacrylamide gel electrophoresis. Toxic activity was assayed during toxin purification by cytopathic effect in Vero or bovine embryonic lung cell cultures. Toxicity for cells correlated with dermonecrosis in guinea pig skin. Toxicity was accounted for by a single protein with a molecular weight of 149,000, as determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Rabbits were inoculated intranasally with purified toxin to determine whether toxin had a role in the induction of pneumonia in rabbits infected with P multocida. Pneumonia, pleuritis, acute hepatic necrosis, and splenic lymphoid atrophy were found in 4 of 5 rabbits. One of 5 rabbits had bilateral turbinate atrophy. Western blotting with monoclonal antibodies to toxin from a P multocida isolate causing atrophic rhinitis in pigs revealed the toxin that induces pleuritis and pneumonia in rabbits to be the same or a closely related toxin.  相似文献   

3.
Toxigenic strains of Pasteurella multocida were readily differentiated from non-toxigenic strains by an agarose overlay method using bovine turbinate cells or bovine lung cells. Cells which were young and densely confluent were best suited to this assay. The incubation period required to distinguish toxigenic strains was dependent on the confluence of the monolayers, which was affected by the seeding rate, cell passage level and growth time prior to overlay. The agarose overlay method correctly identified 11 of 11 reference strains of Pasteurella multocida, and visible cytotoxic changes were present in the monolayers after 48 to 65 h. Outbreaks of the enzootic form of atrophic rhinitis in 2 New South Wales piggeries were associated with the isolation of toxigenic type D strains of P. multocida.  相似文献   

4.
Chromosomal DNA from 13 different selected Pasteurella multocida spp. multocida strains of serotypes A and D were isolated and compared. All 10 toxigenic strains were recognized by a DNA probe which included the toxA gene coding for the Pasteurella multocida toxin (PMT). None of the three nontoxigenic strains reacted with the DNA probe. Toxin from the 10 toxigenic strains were isolated and compared. All were found to possess the biological characteristics previously described for the PMT isolated from P. multocida ssp. multocida NCTC 12178, including molecular mass of approx. 143 kDa and reactivity with a series of monoclonal antibodies. Toxin prepared from different toxigenic strains could not be differentiated immunologically by tandem crossed immunoelectrophoresis, Toxin, which was affinity purified from four of the strains and subsequently inactivated by formaldehyde, was cross-protective when used for vaccination of mice before challenge with PMT. It is concluded that the toxin from toxigenic strains of P. multocida ssp. multocida must be very similar, if not identical.  相似文献   

5.
Study of the toxin-producing ability of Pasteurella multocida in mice   总被引：1，自引：0，他引：1  
Cell-free sonicated extracts and broth cultures of Pasteurella multocida strains of pig origin were examined for their lienotoxicity in mice. P. multocida strains represented capsular types A and D with or without dermonecrotoxic (DNT) activity in the guinea pig skin test. Mouse lienotoxicity test was suitable for determining the toxigenicity of P. multocida strains only when bacterium-free extracts were tested. In that case both toxigenic type A and D strains were lethal to intravenously inoculated mice and caused a remarkable reduction in spleen mass when sublethal doses were used. The extracts of atoxic strains were not lethal and induced splenic hyperplasia. By testing viable cells no correlation was demonstrable between toxin production and virulence of P. multocida to mice. In one experiment the concentrated sterile culture fluids of a toxigenic type D P. multocida and a toxigenic B. bronchiseptica strain were compared. The former caused deaths and splenic atrophy among mice, while the latter was nontoxic and induced slight hyperplasia of the spleen. This fact indicates that P. multocida secretes its toxin into the culture fluid.  相似文献   

6.
Gnotobiotic pig antisera to purified toxoid from a capsule type A or D strain of Pasteurella multocida contained large quantities of antitoxin but comparatively little antibody to a crude lysate of P. multocida. These sera given intraperitoneally to further pigs were almost completely protective against turbinate atrophy after intranasal inoculation of dilute acetic acid and infection with type D toxigenic P. multocida. In contrast, antisera to a crude lysate or bacterin of toxigenic P. multocida which contained large titres of antibody to P. multocida lysate, but no detectable antitoxin, were not protective. Colonisation by toxigenic P. multocida was significantly reduced in protected pigs and was similar to colonisation by nontoxigenic P. multocida in pigs untreated or treated with dilute acetic acid. These results indicated (1) that antitoxin was protective and cross protective between toxins from different capsule types; and (2) that the toxin was the main colonisation factor produced by toxigenic bacteria in the acetic acid model of infection and that immunity to it did not eliminate infection.  相似文献   

7.
Blood sera of 1171 pigs, not vaccinated against PAR, were tested for antibodies against P. multocida toxin in a neutralization test with EBL cell cultures. 277 sera were from 18 herds with clinical PAR; 104 of them (37.5%) had neutralizing antibody titres from 1:2 to 1:1024. No antitoxin was detected in the sera of 866 pigs in 25 PAR nonsuspicious herds. In one nonsuspicious herd, however, 11 of 28 sera were neutralizing in the 1:2 or 1:4 dilution. 30 sows had been vaccinated with inactivated toxigenic P. multocida and/or with the P. multocida toxoid. 21 of these sows had neutralizing sera with titres between 1:8 and 1:4096. The neutralization test presented can be applied for herd diagnosis of PAR and for demonstration of vaccination effects.  相似文献   

8.
This paper is the first report of the production of a dermonecrotic toxin by pasteurella strains that do not belong to the species Pasteurella multocida subspecies multocida. Four strains, isolated from cattle with atrophic rhinitis, were characterised phenotypically. The strains were related to pasteurellaceae, but their taxonomic position remained unclear. The strains produced a toxin that caused a haemorrhagic dermonecrosis in guinea pigs and was lethal to mice. Both effects were neutralised by an antiserum against the purified dermonecrotic toxin of P multocida subspecies multocida. Western blot analysis of culture filtrates of the bovine strains revealed a protein, with the same molecular weight as dermonecrotic toxin, which reacted with both polyclonal and monoclonal antibodies against the toxin. In an immunodiffusion test, anti-dermonecrotic toxin serum did not discriminate between the toxin of the bovine strains and the toxin of P multocida subspecies multocida. It is concluded that these atypical pasteurella strains produce a toxin that is closely related to the dermonecrotic toxin of P multocida subspecies multocida.  相似文献   

9.
Three doses (75 micrograms, 25 micrograms, and 25 micrograms) of purified toxin isolated from a toxigenic strain of type D Pasteurella multocida were given (by atomizer) into the right nasal cavities of each of 10 gnotobiotic pigs on the 21st, 24th, and 27th days of age, respectively. Inoculated pigs (usually 2) and 1 noninoculated control pig each were necropsied on 3, 6, 9, 12, and 15 days after inoculations were given. Severe bilateral atrophy of turbinates occurred in all toxin challenge-exposed pigs. Atrophy was more severe in the inoculated nasal cavity than that in the noninoculated side in 2 of the 10 pigs. Microscopic changes in turbinates of toxin challenge-exposed pigs were more severe in pigs killed at later dates. Dominant changes included degeneration and necrosis of osteoblasts, markedly accelerated osteoclastic osteolysis, replacement of the osseous core by a highly cellular mesenchymal stroma, and multifocal atrophy of submucosal glands. Seemingly, a protein toxin isolated from toxigenic type D strains of P multocida produced rapid atrophy of turbinates and may be a contributing factor in development of clinical progressive atrophic rhinitis in swine.  相似文献   

10.
产毒素多杀性巴氏杆菌菌落双重PCR检测方法的建立   总被引：1，自引：0，他引：1  
为建立快速特异的PCR方法以及同时检测并区分产毒素与非产毒素多杀性巴氏杆菌,本研究根据GenBank登录的多杀性巴氏杆菌KMT1基因和toxA毒素基因序列,设计合成了2对特异引物。特异性试验表明产毒素多杀性巴氏杆菌C51-6扩增出了460bp和1854bp的2条目的片段,而不产毒素多杀性巴氏杆菌、大肠埃希菌、胸膜肺炎放线杆菌、猪链球菌、支气管败血波氏杆菌、副猪嗜血杆菌和鸡白痢沙门菌的扩增均为阴性;敏感性试验表明该PCR方法能从含450CFU的菌液中扩增出相应的目的片段。同时用豚鼠皮肤坏死试验和小鼠致死试验对该PCR方法进行了验证。  相似文献   

11.
The cell-free antigen (CFA) obtained from the culture supernatant of Pasteurella multocida (P. multocida) and the toxin (PMT) purified from CFA were inactivated and mixed with oil adjuvant to prepare a trial vaccine. Both of the mice immunized with CFA and PMT toxoid vaccine were noticeably protected against intratracheal challenge with toxigenic strains of P. multocida. Nevertheless, the protective indices of the mice immunized with CFA vaccine indicate that it is more protective and clears away the bacteria more promptly than in the mice immunized with PMT vaccine. The results suggested that CFA would possibly be good as an effective antigen to toxigenic strains of P. multocida infection.  相似文献   

12.
Four herds of pigs were selected which had different degrees of clinical atrophic rhinitis and used different specific counter-measures. In two of them, the clinical signs occurred spasmodically and were slight. The sows, suckling pigs and growing pigs in all the herds were sampled for toxigenic Pasteurella multocida. In one of the slightly affected herds (herd D), the weaners were moved to a second farm for finishing. No toxigenic P multocida were found at the breeding farm, but 50 per cent of the large growing pigs were positive. It seemed that the organism had entered only at the finishing farm and that the mild clinical signs were due to the infection starting in older pigs than usual. In the second mildly affected herd, 47 per cent of the sows and 42 per cent of the growers were infected. Three toxigenic isolates from this herd produced as severe turbinate damage experimentally in specific-pathogen-free pigs as a stock pathogenic strain. Except in herd D, toxigenic P multocida were found in all the age groups of pigs sampled. However, the pattern of distribution of the organism within the herds was not obviously correlated with the severity of the disease. In a fifth herd there were obvious cases of clinical atrophic rhinitis, with marked turbinate atrophy, from which toxi-genic P multocida were recovered in abundance. Subsequently, the clinical disease disappeared and, despite extensive and repeated sampling, the organism was not found again.  相似文献   

13.
Between March and July 1987, a study was undertaken to determine the prevalence of and factors associated with toxigenic type D Pasteurella multocida infection in New South Wales pig herds. Toxigenic type D P. multocida was isolated from the nasal cavities of pigs in one (2%) of 50 randomly selected herds. Toxigenic isolates were also recovered from 2 (8%) of a separate group of 25 herds that had purchased pigs from a known infected piggery in South Australia (herd SA). Snout abnormalities were present in 9.4%, 3.2% and 1.8% of grower pigs in the 3 affected herds. Isolation of toxigenic P. multocida was significantly associated (p less than 0.0001) with the occurrence of clinically affected pigs in the herd. Purchase of at least 5 pigs from herd SA was associated with an elevated risk (p less than 0.05) of isolation of toxigenic P. multocida.  相似文献   

14.
A total of 163 pigs from nine farrow-to-finish herds representing various levels of atrophic rhinitis (AR) were selected for postslaughter examination of AR and pneumonia. Nasal swabs and lungs were cultured for detection of Bordetella bronchiseptica and Pasteurella multocida. Seventy-three pigs were examined at eight weeks of age and 90 contemporaries at six months of age. Mean AR scores were 1.21 and 1.11 for the eight week and six month old pigs, respectively (0 = normal, 3 = severe). In individual pigs increasing AR score was related to increasing pneumonia score in eight week old pigs but not in six month old hogs. In eight week old pigs, B. bronchiseptica and P. multocida were isolated more frequently from pigs with higher AR scores. From nasal swabs of six month old hogs, Bordetella was almost never recovered while Pasteurella was frequently isolated score. Toxigenic type DP. multocida was isolated from nasal cultures of only seven (4%) pigs and from lung cultures of only one pig. Pasteurella was never isolated from lungs of the eight week old pigs and Bordetella never from the six month old hogs. The isolation rate of P. multocida, predominantly type A, from lungs of six month old pigs increased from 11% in grossly normal lungs to 86% in lungs with severe pneumonia. Pigs from one herd free from lesions of AR and pneumonia were also examined; type AP. multocida was isolated from nasal cultures of one of six eight week old pigs. Somatic antigens of P. multocida were determined for 94 nasal and 20 lung isolates. Somatic serovar 3 was found in 93% of the nasal isolates and in all lung isolates.(ABSTRACT TRUNCATED AT 250 WORDS)  相似文献   

15.
A luminol-dependent chemiluminescence (LDCL) assay was used to evaluate the response of bovine polymorphonuclear leukocytes; (neutrophils [PMN]) to living and heat-killed Escherichia coli, Pasteurella multocida (type A, serotype 3), and P haemolytica (biotype A, serotype 1), and to heat-killed P haemolytica and sterile culture supernatant from living P haemolytica. Control cultures containing PMN that had not been phagocytically stimulated with bacteria had a modest increase in LDCL during the initial 10 minutes of incubation, followed by a gradual decline throughout the 120-minute incubation period. Bovine PMN emitted LDCL more efficiently when the cells were exposed to living E coli or P multocida than when they were exposed to the same bacteria killed by heat. The mean LDCL values for reaction mixtures containing living E coli or P multocida peaked at 30 minutes of incubation and remained above values for mixtures containing the same heat-killed bacteria. Kinetics of the LDCL response of bovine PMN to heat-killed P haemolytica were similar (although reduced in amplitude) to that observed with killed E coli or P multocida. The LDCL response of bovine PMN to living P haemolytica was not like that for E coli or P multocida, and was characterized by the development of a peak response at 10 minutes followed by a precipitous decrease in responsiveness and a subsequent complete cessation of LDCL. Addition of sterile culture supernatant from living P haemolytica to test samples containing heat-killed P haemolytica induced a response similar to that obtained with the living microorganism.(ABSTRACT TRUNCATED AT 250 WORDS)  相似文献   

16.
A total of 855 pig lungs were collected at slaughter and evaluated macroscopically. Bacteriological examinations were carried out on tissue samples from chronic pleuropneumonic lesions (n = 196) and from chronic bronchopneumonic lesions with suppuration (n = 14). Samples from normal lung tissue (n = 22) were also included. Pasteurella multocida was isolated from 54%, Actinobacillus (Haemophilus) pleuropneumoniae from 11%, and Streptococcus spp. from 14% of the pneumonic lesions, respectively. From normal lung tissue P. multocida was isolated from 3 (14%) of the samples, A. pleuropneumoniae was not recovered and streptococci were isolated from only 1 (5%) of these samples. The above mentioned bacterial species were recovered either in pure cultures or mixed with various other microbes. A total of 109 P. multocida strains were further characterized by capsular serotyping and testing for production of dermonecrotic toxin. Ninety-nine (91%) of the strains were capsular type A 10 (9%) were type D. Out of the type A and the type D strains 94% and 90% were toxigenic, respectively. Most of the A. pleuropneumoniae strains were serotype 2. Strains of serotypes 1 and 7 were also identified. The majority of the streptococci were identified as either Streptococcus suis or Streptococcus dysgalactiae. Actinomyces pyogenes was isolated from 14% of the lesions and anaerobic bacteria from 18%, respectively. The significance of the various bacterial species in relation to the development of chronic pneumonic lesions is discussed. Special attention is paid to P. multocida, and it is concluded that this bacterial species is probably of importance for the development of both types of chronic pneumonias.  相似文献   

17.
Colonisation of type D Pasteurella multocida was studied in five groups of seven SPF piglets each. Piglets of Group 1 were kept together with seven 5-week-old piglets obtained from a large herd infected with toxigenic P. multocida for 16 weeks (contact infection). These piglets were made free from toxigenic Bordetella bronchiseptica by local immunisation. Piglets of Group 2 were inoculated with 5 x 10(7) colony-forming units (cfu) of P. multocida washed from the nasal mucosa of piglets free from toxigenic B. bronchiseptica with fetal calf serum. Piglets of Group 3 were inoculated intranasally with 5 x 10(7) cfu of P. multocida washed from yeast-extract proteose-peptone cystine (YPC)-blood agar with fetal calf serum. Piglets of Group 4 were inoculated with 5 x 10(7) cfu of P. multocida grown in a YPC-based broth without blood. Piglets of Group 5 served as controls. The piglets of Group 1 did not contract P. multocida infection from infected contact piglets. After a single inoculation one of four, while after three inoculations two of three piglets of Group 2 became infected by P. multocida. After a single inoculation none of four, while after three inoculations one of three piglets of Group 3 were colonised by P. multocida. Both single and repeated inoculation failed in piglets of Group 4.  相似文献   

18.
The objective of the experimental study was to examine whether aerial ammonia alone could predispose the respiratory system of pigs to infection with toxigenic Pasteurella multocida type A. Two groups of 5 pigs each were continuously exposed to 50 ppm ammonia and less than 5 ppm ammonia, respectively, for a 59-day period (from 37 kg to 90 kg bodyweight) followed by necropsy. In an aerosol chamber all pigs were exposed to an aerosol of toxigenic P. multocida type A (mean bacterial concentration in the aerosol-exposure chamber: 10(5) colony forming units/m3; exposure period: 25 min) at day 10, 21, 35 and 49 after the onset of ammonia exposure. During the experiment none of the pigs showed clinical signs of pneumonia nor did they develop visible distortion of the snout. None of the pigs had gross lesions in the lungs at necropsy and toxigenic P. multocida was not detected by culture from the lungs from any of the pigs. The chance of recovering toxigenic P. multocida from nasal swabs (collected during experiment) was 2-4 times greater in the test group compared to the control group. The average daily weight gain was lower for the ammonia exposed pigs compared to the control group. In conclusion the results from this study suggest that ammonia in concentrations of 50 ppm is unlikely to predispose growing pigs to pulmonary infection with toxigenic P. multocida.  相似文献   

19.
猪多杀性巴氏杆菌的分离鉴定及生物学特性研究   总被引：17，自引：2，他引：17  
用PCR方法配合生化鉴定，从有肺炎症状猪的肺脏及进行性萎缩性鼻炎(Progressive atrophic rhinitis，PAR)症状猪的鼻拭子中分离出66株多杀性巴氏杆菌(Pasteurella multocida，Pm)。然后做了药敏试验，并用PCR方法对这66株Pm进行分型及毒素基因的检测，用豚鼠皮肤坏死试验及小鼠致死试验对产毒素多杀性巴氏杆菌(Toxigenie Pasteurella multocida，T^ Pm)进一步鉴定。结果显示PCR鉴定与生化鉴定Pm结果完全一致；PCR分型表明有46株为D型Pm，18株为A型：Pm，1株为B型Pm，1株无法定型；有8株用PcR检测为T^ Pm；豚鼠皮肤坏死试验及小鼠致死试验对这8株T^ Pm的进一步鉴定也表明均为产毒素菌株。所鉴定的8株T^ Pm都为D型，都分离于有严重PAR症状的猪。  相似文献   

20.
Thirty-eight clinical isolates of Pasteurella multocida, recovered from a continuous flow, farrow-to-finish swine herd, were characterized by capsular serotyping and restriction endonuclease analysis (REA) in order to study the epidemiology of P. multocida pneumonia. Twenty-three of the 38 isolates obtained in the study belonged to serotype A. They displayed three REA patterns after digestion with HpaII, of which one designated A-3 represented 70% of the samples. The remaining 15 isolates were serotype D. Four different REA patterns were observed in the type D isolates. The REA type D-1 was most prevalent and accounted for 47% of the serotype D isolates. All serotype A isolates were nontoxigenic, whereas five (33%) of the serotype D isolates were toxigenic. Vertical transmission of P. multocida could not be demonstrated, and was probably not a major route of infection. The results of this study suggest that strains of P. multocida virulent for pigs exist and cause swine pneumonic pasteurellosis in continuous flow herds by horizontal transmission.  相似文献   

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