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1.
Abstract

One hundred seven Aeromonas spp., 26 Edwardsiella ictaluri, 6 E. tarda, 12 Plesiomonas shigelloides, and 6 Pseudomonas spp. (131 piscine isolates and 26 reference isolates) were studied with 36 biochemical tests from the Minitek system, 20 tests from the API 20E system, and corresponding standard tube tests. Isolates were incubated at 25°C. Arginine dihydrolase, ornithine decarboxylase, mannose, and citrate showed less than 95% agreement between the Minitek system and the tube tests. Arginine dihydrolase, lysine decarboxylase, nitrite reductase, Voges-Proskauer, and citrate showed less than 95% agreement between the API 20E system and the tube tests. The 26 reference isolates were examined with the three systems and were incubated at both 25 and 37°C. There were no major differences between tests run at 25 and 37°C except with nine Aeromonas spp. that did not grow well at 37°C. Both the Minitek and API 20E systems will reproduce standard biochemical tube test results with at least 95% accuracy when used to test warmwater fish pathogens incubated at 25°C. However, the numerical identification databases for both the Minitek and API 20E systems were not usable for identifying fish pathogens.  相似文献   

2.
Coagulase-negative staphylococci (CNS) are the most frequently isolated pathogens from cows with intramammary infection (IMI). Although API STAPH ID 20, a commercially available identification system, and PCR-restriction fragment length polymorphism (PCR-RFLP) of the gap gene (gap PCR-RFLP) have been successfully applied for the identification of CNS isolates from human specimens, their accuracy in the identification of veterinary isolates has not been fully established. In this study, we identified 263 CNS isolates from bovine IMI at species level by partial 16S rRNA gene sequence analysis as the definitive test. Species identification obtained using partial 16S rRNA gene sequence analysis was compared to results from the API STAPH ID 20 and gap PCR-RFLP analysis. Eleven different CNS species were identified by partial 16S rRNA gene sequence analysis. Only 76.0% (200/263) of the species identification results obtained by API STAPH ID 20 matched those obtained by partial 16S rRNA gene sequence analysis, whereas 97.0% (255/263) of the species identification results obtained by the gap PCR-RFLP analysis matched those obtained by partial 16S rRNA gene sequence analysis. The gap PCR-RFLP analysis could be a useful and reliable alternative method for the species identification of CNS isolates from bovine IMI and appears to be a more accurate method of species identification than the API STAPH ID 20 system.  相似文献   

3.
Outer membrane proteins (OMP) of P. multocida (serotype B:2) field isolates (n = 6) and a vaccine strain (P-52) were extracted by a sarkosyl method and characterized using SDS-PAGE and immunoblotting. About 20 polypeptide bands were observed in the profile of the vaccine strain with MW ranging from 16 to 90 kDa and, based on band thickness and intensity of staining, three polypeptides of MW 31, 33 and 37 kDa were considered to be the major OMPs. The profiles of the field isolates showed minor differences when compared with that of the vaccine strain. The OMP of 33 kDa was only expressed by the vaccine strain. Four field isolates expressed an OMP of 39 kDa, which did not appear in the profiles of the remaining two field isolates and the P-52 strain. Similarly, an OMP of 25 kDa was exclusively seen in the profile of a single isolate. By immunoblotting studies, using anti-P. multocida (P-52) whole-cell hyperimmune serum raised in rabbits as well as buffalo immune sera, it became evident that the polypeptide of 37 kDa was the most antigenic OMP in the profiles of all the isolates, including the P-52 strain. Other polypeptides were either weakly antigenic or visible in the profile of only a few of the isolates. The study thus identified the major OMP of P. multocida (B:2) and suggested that this highly antigenic 37 kDa OMP has potential for further protective and immunodiagnostic studies.  相似文献   

4.
One-hundred isolates of Pseudomonas pseudomallei were used to evaluate the API 20E and Microbact 24E rapid identification systems. The API 20E system identified 50% of the isolates using the revised 1979 Manual only, and 63% when referral was made to the computer centre. A higher identification rate (69 and 87%, respectively) was achieved with a longer incubation period of 96 h. The Microbact 24E system identified 84% of the isolates as P. pseudomallei using the revised 1983 Manual, and 100% when referral was made to the computer centre. The Microbact 24E system would appear to be a reliable system for the identification of P. pseudomallei.  相似文献   

5.
An outbreak of pasteurellosis with high mortality was recorded in indigenous pigs in India. The presence of Pasturella multocida in samples collected from dead pigs was detected by smear examination and isolation, and later by P. multocida specific polymerase chain reaction (PM-PCR). P. multocida was detected in all the samples collected from dead pigs, with nine strains ultimately isolated. All the isolates were positive by PM-PCR. Six isolates showed CAPA and three were of CAPD capsular types. All the isolates were negative for toxigenic gene (toxA). The isolates were sensitive to oxytetracycline, doxycycline, gentamycin, erythromycin, ampicillin, amoxycillin, chloramphenicol and enrofloxacin and resistant to sulphadiazine and cloxacillin. The PCR assays used in this study have been shown to be useful diagnostic tools for P. multocida detection and characterization.  相似文献   

6.
In a 2-year period 54 feral cats were captured in Grenada, West Indies, and a total of 383 samples consisting of swabs from rectum, vagina, ears, eyes, mouth, nose and wounds/abscesses, were cultured for aerobic bacteria and campylobacters. A total of 251 bacterial isolates were obtained, of which 205 were identified to species level and 46 to genus level. A commercial bacterial identification system (API/Biomerieux), was used for this purpose. The most common species was Escherichia coli (N = 60), followed by Staphylococcus felis/simulans (40), S. hominis (16), S. haemolyticus (12), Streptococcus canis (9), Proteus mirabilis (8), Pasteurella multocida (7), Streptococcus mitis (7), Staphylococcus xylosus (7), S. capitis (6), S. chromogenes (4), S. sciuri (3), S. auricularis (2), S. lentus (2), S. hyicus (2), Streptococcus suis (2) and Pseudomonas argentinensis (2). Sixteen other isolates were identified to species level. A molecular method using 16S rRNA sequencing was used to confirm/identify 22 isolates. Salmonella or campylobacters were not isolated from rectal swabs. E. coli and S. felis/simulans together constituted 50% of isolates from vagina. S. felis/simulans was the most common species from culture positive ear and eye samples. P. multocida was isolated from 15% of mouth samples. Coagulase-negative staphylococci were the most common isolates from nose and wound swabs. Staphylococcus aureus, or S. intemedius/S. pseudintermedius were not isolated from any sample. Antimicrobial drug resistance was minimal, most isolates being susceptible to all drugs tested against, including tetracycline.  相似文献   

7.
The ability of two commercial rapid identification systems, API 20E and BBL Crystal E/NF, to reliably identify bacterial isolates from the internal organs of reared sea bass were compared. The tests gave different results: API 20E identified bacteria as Pseudomonas spp. with 37% accuracy, while BBL Crystal E/NF identified them as Flavobacterium odoratum with 99% accuracy. Although F. odoratum is not a marine fish pathogen, conventional tests conducted with the same isolates were more indicative of them being Flavobacterium spp. than Pseudomonas spp., suggesting that BBL Crystal E/NF was more reliable in this identification. Both systems were found to be applicable for diagnostics of marine fish pathogens, but should be used with caution because of possible misinterpretation.  相似文献   

8.
Four hundred and twenty-two isolates of Enterobacteriaceae from veterinary sources were used to evaluate the API20E system. The accuracy of the individual tests, compared to the conventional equivalent was established at a minimum of 88%. The identification level of the system was determined to be 86%, a level considerably lower than that recorded for human clinical and food based studies. If the API20E system is adopted for use in veterinary laboratories, then the limitations of the system must be clearly understood.  相似文献   

9.
AIMS: To examine pigs at slaughter in New Zealand for the presence of Pasteurella multocida, and to determine for isolates, their biochemical profiles, somatic and capsular types, and the presence or absence of the HSB and toxA genes, associated with haemorrhagic septicaemia (HS) and progressive atrophic rhinitis (PAR), respectively.

METHODS: Swabs from 173 lungs, 158 palatine tonsils and 82 nasal passages of pigs at two abattoirs in New Zealand were cultured for P. multocida using conventional techniques, and isolated colonies were subjected to biochemical tests for identification of biovars. Somatic serotyping was conducted using an agar gel immunodiffusion (AGID) test. Polymerase chain reaction (PCR) assays were used to confirm phenotypic identification of colonies using species-specific primers, capsule type using serogroup-specific primers and multiplex PCR, and to test for the presence of HSB and toxA genes.

RESULTS: Pasteurella multocida was isolated from 11/173 (6.4%) lung, 32/158 (20.2%) palatine tonsil and 5/82 (6.1 %) nasal swab samples, a total of 48 isolates from 413 samples (11.6%). Isolation rates per farm ranged from 1–53% of tissue samples collected from pigs 5–6 months of age. On phenotypic characterisation, isolates were allocated to seven main biovars, viz 1, 2, 3, 5, 9, 12, and a dulcitol-negative variant of Biovar 8, the majority (30/48) being Biovar 3. Of the 42 isolates for which somatic serotyping was conducted, 10% were Serovar 1, 79% were Serovar 3, 2% were Serovar 6,1, 2% were Serovar 12, and 7% could not be typed. All 48 isolates were confirned as P. multocida using a species-specific PCR. In the capsular multiplex PCR, 92% of isolates were Capsular (Cap) type A, 2% were Cap D, and 6% could not be typed. None of the samples were positive for the HSB or toxA genes.

CONCLUSION: Serovars or capsular types of P. multocida associated with HS or PAR in pigs were not detected. Establishment of species-specific, capsular and toxin PCR assays allowed the rapid screening of isolates of P. multocida, while serotyping provided an additional tool for epidemiological and tracing purposes.  相似文献   

10.
The gram-negative bacterium Pasteurella multocida constitutes a heterogeneous species associated with wide range of disease in many animals. Isolates are classified into five groups based on capsular antigen (capA, B, D, E and F). Recently, a new valuable PCR-based method was introduced to determine the epidemiological correlation between P. multocida infection and existence of virulence genes including tbpA, pfhA, toxA and hgbB. However, this method is tedious and laborious. Thus, in the current study, we designed a reliable multiplex PCR method for rapid detection of virulence genes in P. multocida. Eighty seven strains of P. multocida isolated from various clinically healthy and infected hosts were examined by uniplex PCR method for each virulence associated genes. Based on our improved and simplified multiplex PCR method, rapid detection of four virulence genes was accomplished. It is proposed that its implementation may benefit the epidemiological investigations.  相似文献   

11.
The API 20E System and the Encise Enterotube were evaluated for the identification of the Enterobacteriaceae isolated from clinical specimens of animal origin at a veterinary diagnostic laboratory. Compared to conventional tubed media, the API 20E System identified 235 of 240 isolates (97.9%) correctly. The Encise Enterotube correctly identified 229 of the 240 isolates (95.4%). Thus, both these identification systems could be used to replace conventional methods for identifying members of this family isolated from animal origin.  相似文献   

12.
《Veterinary microbiology》1997,54(2):167-183
The outer membrane proteins (OMPs) of P. multocida serotypes A3 (7 isolates), A4 (2 isolates), A3,4 and A2 (one isolate each) obtained from pneumonic cattle (10 isolates) and from one pig isolate were investigated to identify potential immunogens. SDS-PAGE of P. multocida OM isolated by SDG centrifugation of spheroplasts revealed eight major OMPs. Outer membranes isolated by sarcosyl extraction or SDG had similar protein composition on Coomassie blue-stained SDS-PA gel and on immunoblots. Two major OMPs (Mrs of 35 and 46 kDa at 100°C) demonstrated heat modifiability with apparent Mrs of 30 and 34 kDa at 37°C, respectively. The N-terminal aa sequences of these heat modifiable proteins revealed homology with E. coli OmpA and Hib P1 proteins, respectively. Protease treatment of whole cells followed by western immunoblots using bovine convalescent sera identified several immunogenic, surface-exposed and conserved OMPs among the eleven P. multocida isolates examined. The whole organism SDS-PAGE profiles of the eleven P. multocida isolates differed such that six patterns were seen. These patterns could potentially be used as a typing system for P. multocida bovine isolates based on the molecular weights of whole cell proteins. The above observations have potentially important implications relative to the immunity to infection.  相似文献   

13.
This report describes the proliferation and transmission patterns of Pasteurella multocida B:2 among stressful goats, created through dexamethasone injections. Thirty seven clinically healthy adult goats were divided into three groups consisted of 15 goats in group A, 11 goats in group B and the remaining 11 in group C. At the start of the study, all goats of group A were exposed intranasally to 1.97 × 1010 CFU/ml of live P. multocida B:2. Dexamethasone was immediately administered intramuscularly for 3 consecutive days at a dosage rate of 1 mg/kg. The exposed goats were observed for signs of HS for a period of 1 month. At the end of the 1-month period, 11 goats from group B were introduced into and commingled with the surviving goats of group A before all goats from both groups were immediately injected intramuscularly with dexamethasone for 3 consecutive days. The treatment with dexamethasone was then carried out at monthly interval throughout the 3-month study period. Goats of group C were kept separately as negative control. Three surviving goats from each group were killed at 2-week interval for a complete post-mortem examination. Two (13%) goats of group A were killed within 24 hours after intranasal exposure to P. multocida B:2 while another two (13%) goats from the same group were killed on day 40, approximately 10 days after the second dexamethasone injection. All four goats showed signs and lesions typical of haemorrhagic septicaemia. Bacteraemia was detected in 3 goats of group A that were having rectal temperature higher than 41°C. The P. multocida B:2 isolation pattern was closely associated with dexamethasone injections when significantly (p < 0.05) higher rate of isolations from both groups were observed after each dexamethasone injection. Transmission of P. multocida B:2 from goats of group A to group B was successful when P. multocida B:2 was isolated from goats of group B for a period of 28 days. There was a strong correlation between dexamethasone injections, rate of bacterial isolation and serum cortisol level. The IgG level showed an increasing trend 2 weeks after exposure to P. multocida B:2 and remained high throughout the study period.  相似文献   

14.
The aim of the present study was to determine which Pasteurella and Mannheimia species are present in the upper respiratory tract of healthy calves with no history of antimicrobial treatment prior to sampling. The presence of subpopulations of tetracycline-resistant Pasteurellaceae was also investigated. Nasal swabs from 61 loose group-housed, clinically healthy calves, 1 to 4 months old, from 16 dairy herds were inoculated aerobically on a selective medium (Columbia agar with 5% ovine blood and 16 mg/L bacitracin) with or without 4 mg/L oxytetracycline (OTC). A total of 43 strains belonging to the family Pasteurellaceae were isolated from 38 calves (62.3%) out of 13 herds (81.3%). The predominant organisms were Pasteurella multocida subsp. multocida (57.4%), Mannheimia varigena (4.9%) and M. haemolytica (3.2%). Growth of Pasteurellaceae on the OTC-containing medium was seen only with samples from two herds (6 animals; 9.8%), and on only one farm this proved to be an OTC-resistant subpopulation. Minimum inhibitory concentration (MIC) determinations by means of agar dilution confirmed a low prevalence of OTC-resistant Pasteurellaceae, with overall MIC50 and MIC90 values of 0.25 and 32 mg/L, respectively. These data do not support the hypothesis that the relative high frequency of tetracycline-resistant P. multocida isolates from fatal cases of bovine respiratory disease is related to the presence of minor tetracycline-resistance subpopulations within this species.  相似文献   

15.
Pasteurella multocida is the causative agent of pasteurellosis, a major disease in most domestic animals and livestock. In this study, a total of 34 isolates of P. multocida from rabbits and other domestic animals from Mexico with respiratory diseases underwent polymerase chain reaction-based capsular typing. One sheep isolate was found to belong to capsular serogroup D, whereas the rest of the rabbit, sheep, cattle, pig, goat, and duck isolates belonged to capsular serogroup A of P. multocida. This is the first report of capsular type A in P. multocida isolates from rabbits and duck origin in Mexico.  相似文献   

16.
Two Carter type B Pasteurella multocida isolates, Izatnagar 52 and 25, isolated from cases of haemorrhagic septicaemia ( ), were used in a modified subtractive hybridisation technique with the specific aim of cloning unique DNA sequences related to the pathogenesis of HS. Biochemical and protein analyses have shown these isolates to be similar, but reports indicate that they have differences in pathogenicity. The subtracted inserts were screened against genomic DNA from a wide range of P multocida isolates, with two distinct fragments demonstrating specific hybridisation with Carter type B isolates that cause . No identity was observed with either Carter type E isolates or non-HS type B strains. The clones were sequenced and a search of the GenBank database revealed significant identity of the clone A3b (296 nt) to P haemolytica lipoprotein, whereas there was no significant identity with 6b (956 nt). Both these fragments had a high level of identity (72·8 to 76·9 per cent) to the H infuenzae Rd genome.  相似文献   

17.
The objective of this work was to describe trends in multiple-class antimicrobial resistance present in clinical isolates of Escherichia coli F4, Pasteurella multocida and Streptococcus suis from Ontario swine 1998–2010. Temporal changes in multiple-class resistance varied by the pathogens examined; significant yearly changes were apparent for the E. coli and P. multocida data. Although not present in the E. coli data, significant increases in multiple-class resistance within P. multocida isolates occurred from 2003 to 2005, coinciding with the expected increase in antimicrobials used to treat clinical signs of Porcine Circovirus Associated Disease (PCVAD) before it was confirmed. Prospective temporal scan statistics for multiple-class resistance suggest that significant clusters of increased resistance may have been found in the spring of 2004; months before the identification of the PCVAD outbreak in the fall of 2004.  相似文献   

18.
The prevalence of capsular and somatic serotypes were studied among 123 Pasteurella multocida strains isolated from chickens (n = 94), ducks (22), quails (4), turkeys (2) and geese (1) from different geographical regions of India. All strains exhibited similar cultural and morphological characteristics. Ninety-two of the isolates belonged to serotype A:1, the most prevalent serotype, with serotypes A:3, A:1,3, D:3 and F:3 having two isolates each. Only one isolate was positive for serotypes A:4 and D:1. Twenty isolates were untyped. A multiplex capsular PCR assay generated amplicons of sizes 460, 1044, 657 and 854 bp in 106 isolates identified as capsular serotype-A, 15 in serotype D and two in serotype F. Capsular types B and E were not detected in any of the avian isolates studied. The present findings suggest that a multiplex capsular PCR assay may be suitable for the rapid initial identification serotypes P. multocida during epidemiological studies of fowl cholera.  相似文献   

19.
Using the Analytab (API 20E) Enterobacteriaceae system of biochemical identification, a total of 506 Escherichia coli isolates from different animal species were coded numerically or biotyped. Fifty-four different biotypes were identified, 11 accounting for 83.1% of the isolates examined. Three of these profiles accounted for 65.3% of the isolates and were found in almost all animal species. Some of the biotypes were found in only one animal species: six in cattle, five in horses, 15 in pigs, two in sheep, two in birds, one in dogs and one in a porpoise. Biotypes, as determined here, could not be related to a particular pathology and more work is needed to assess the extent and significance of this relative biotype specificity among animal species. The use of other, more sophisticated, typing systems, i.e. plasmid "fingerprinting", or restriction endonuclease analysis of chromosomal DNA, would have to be investigated.  相似文献   

20.
Repetitive extragenic palindromic sequence-based PCR (REP-PCR) was used to characterize 67 field isolates of Pasteurella multocida originating from different animal species and geographical regions of India. REP-PCR was found to be rapid and reproducible (three repeats were done). These isolates yielded different 23 profiles which were clustered into eight groups. The discrimination index was moderate (D value 0.83). Somatic and antigenic typing of the isolates did not reveal any correlation with REP-PCR profiles. There was no host-specific, type-specific, region-specific or pathenogenicity-specific pattern. The REP profiles of isolates obtained from wild animals were similar to those obtained from domestic animals. Two common bands were present in all the isolates irrespective of somatic or antigenic types. The results were not comparable with earlier findings, which had shown high discrimination index and correlation with disease presentation. Saxena, M.K., Singh, V.P., Kumar, A.A., Chaudhuri, P., Singh, V.P., Shivachandra, S.B., Biswas, A. and Sharma, B., 2006. REP–PCR analysis of Pasteurella multocida isolates from wild and domestic animals in India. Veterinary Research Communications, 30(8), 851–861  相似文献   

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