Kurt P. Snipes

Molecular fingerprinting of Pasteurella multocida associated with progressive atrophic rhinitis in swine herds.

Ninety-six nasal isolates of Pasteurella multocida from swine herds with progressive atrophic rhinitis were characterized by restriction endonuclease analysis (REA) of whole-cell DNA, ribotyping, and plasmid analysis. For REA, bacterial DNA was digested with SmaI and electrophoresed in 0.7% agarose, and fragments were visualized with UV light. For ribotyping, EcoRI-digested and electrophoresed restriction fragments of whole-cell DNA were transferred to nitrocellulose membranes, hybridized with γ-32P-labeled Escherichia coli ribosomal RNA, and visualized by autoradiography. Phenotypes of isolates were toxigenic capsular type D (n = 51), nontoxigenic type D (n = 28), nontoxigenic type A (n = 16), and toxigenic type A (n = 1). Plasmids of various sizes were evident in 92.2% and 17.9% of toxigenic and nontoxigenic D strains, respectively, but were absent from all type A strains. Among the 4 phenotypes, there were 17 REA profiles and 6 ribotypes. For 3 of 17 REA patterns, multiple ribotypes were evident, and several REA types were evident in 5 of 6 ribotypes. Thirty-seven isolates of toxigenic capsular type D from Australian herds were either SmaI type B or C and ribotype 2, whereas 14 toxigenic D isolates from the USA and other countries were more heterogeneous (7 REA types and 6 ribotypes). The fingerprinting results provided evidence in support of the hypothesis of a single source infection in Australia associated with the introduction of breeding pigs from overseas.

Pasteurella multocida in Wild Mammals and Birds in California: Prevalence and Virulence for Turkeys

Samples collected from the oropharynx of wild mammals and birds trapped on 36 turkey farms in California were evaluated for the presence of Pasteurella multocida. A total of 966 animals were collected from 18 premises that had experienced an outbreak of fowl cholera within the past 2-8 months; samples were collected from 16 of these 18 premises within 2-8 weeks of outbreak notification and while the infected flock was still present. A total of 939 animals were trapped from an additional 18 premises that had not reported any outbreaks of fowl cholera within at least 4 months, if ever. Forty-eight isolates of P. multocida, of a variety of somatic serotypes, were recovered from 6 species of mammals and 3 species of birds. On only 2 of 7 premises was the somatic serotype of the isolates obtained from wildlife the same as the isolate obtained from tissues of turkeys that had died of fowl cholera on the same premises. Tests for virulence to turkeys were conducted with 31 of the isolates. Seventeen of these isolates caused mortality in turkeys. Wide ranges in mortality rates and median times to death were observed.

Association of Complement Sensitivity with Virulence of Pasteurella multocida Isolated from Turkeys

Two strains of Pasteurella multocida, both derivatives of strain P1059, were compared for virulence for 14-week-old turkeys and sensitivity to turkey plasma. Strain P1059-1, a nalidixic-acid-resistant mutant of P1059 with an LD50 of approximately 10(3) colony-forming units (CFU), was more resistant to the bactericidal effects of fresh turkey plasma at 37 C than avirulent strain P1059-1A. P1059-1A, with an LD50 of approximately 10(8) CFU, is an acapsular variant of P1059-1 that spontaneously arose after prolonged passage on artificial medium. The bactericidal effect on P1059-1A was removed when turkey plasma was treated with heat or with zymosan, maneuvers that removed hemolytic complement activity from turkey plasma.

Homogeneity of characteristics of Pasteurella multocida isolated from turkeys and wildlife in California, 1985-88.

Five hundred twenty isolates of Pasteurella multocida, collected in California from September 1985 to November 1988, were characterized in the laboratory. Characteristics examined included serotype, capsular type, biotype (subspecies), and possession of plasmid DNA. Three hundred thirty-three isolates recovered from turkeys dying from fowl cholera, 88 isolates from liver turkeys in flocks with fowl cholera outbreaks in the recent past, and 99 isolates from wildlife captured on fowl cholera-outbreak and non-outbreak turkey premises were studied in this manner. Characteristics were fairly homogeneous among isolates, especially those obtained from turkeys. The majority of isolates were serotype 3,4, capsular type A, subspecies multocida, and lacked plasmid DNA. Common serotypes of isolates from turkeys and wildlife sampled on the same premises were noted in eight of 13 cases examined.

Differentiation of field isolates of Pasteurella multocida serotype 3,4 from live vaccine strain by genotypic characterization.

Fifty-five serotype 3,4 isolates of Pasteurella multocida, isolated from turkeys dead from fowl cholera, were characterized (fingerprinted) genotypically for comparison with the serotype 3,4 live fowl cholera vaccine principally used in turkeys in California. Twenty-three isolates were obtained from turkeys vaccinated with the M9 live vaccine, and 32 additional isolates were from turkeys not vaccinated for fowl cholera. Methods of characterization included restriction endonuclease analysis of chromosomal DNA and ribotyping, a technique for highlighting restriction site heterogeneity of highly conserved ribosomal RNA genes and associated sequences using a radiolabeled rRNA probe. Eight different genotypes or ribotypes were detected in these isolates by the above methods. Of 23 isolates from M9-vaccinated turkeys flocks, 19 were the same ribotype as M9. Thirty of 32 isolates recovered from unvaccinated turkeys were different ribotypes from M9. The remaining two isolates resembled M9 and were recovered from two different flocks placed in succession on a turkey farm where a flock placed previously had been vaccinated with M9, suggesting interflock transmission. Ribotyping and restriction endonuclease analysis appear to be useful tools to aid in the determination of the role that the live vaccine plays in fowl cholera epidemiology.

Transmission of Pasteurella multocida on California turkey premises in 1988-89.

Restriction endonuclease analysis (REA) of whole-cell DNA was used to determine possible sources of Pasteurella multocida for each outbreak of fowl cholera occurring in turkey flocks in eight commercial poultry companies in California from October 1988 to September 1989. Over this period, 179 isolates of P. multocida were obtained from dead turkeys in 80 meat and breeder flocks on 43 premises. P. multocida was isolated from wildlife on five premises. Isolates were characterized by subspecies, serotype, presence of plasmid DNA, and REA type. In 52 (65%) flocks, all isolates of P. multocida had the same REA pattern as the M9 live vaccine strain following digestion of DNA with the restriction enzyme SmaI. Field strains of P. multocida were obtained from 27 (34%) flocks, and one flock (1%) yielded both M9 and a field strain of the organism. REA of field strains of P. multocida revealed 17 different SmaI REA types. Based on matching SmaI REA types, potential sources of P. multocida were identified for 15 of the 28 flocks infected with field strains of the organism, and transmission between turkey premises was a possibility in only seven flocks.

Detection of Pasteurella multocida-specific DNA in Turkey Flocks by Use of the Polymerase Chain Reaction

A polymerase chain reaction (PCR)-based assay using primers constructed to amplify the gene (psl) encoding the P6-like protein (Psl) of Pasteurella multocida was developed. After Southern blotting and hybridization with psl, the assay (PCR-H) was found to be specific (it did not detect a variety of other avian bacterial pathogens) and sensitive (detected > or = 10 P. multocida organisms or > or = 24 femtograms of extracted P. multocida DNA). Samples were collected from the oropharynx of randomly selected birds housed on premises that had recently experienced an outbreak of avian cholera (outbreak farms) or from birds housed on premises that had not reported an outbreak of this disease during the preceding 12 mo (control farms). The PCR-H assay detected 11 infected turkeys out of a total of 178 sampled on six outbreak farms as compared with isolation of P. multocida from 23 turkeys by using mouse inoculation. Neither method detected P. multocida in samples collected from 174 turkeys sampled on six control farms. Statistical analysis using the Kappa test demonstrated that the results of the two tests showed poor agreement from five outbreak flocks (K = 0, 0, 0, 0.35, 0.47) and strong agreement from one outbreak flock (K = 0.89). Combined results from the outbreak flocks showed poor agreement (K = 0.49) between the two methods.

Epidemiology and financial impact of fowl cholera in turkeys: a retrospective analysis.

A retrospective study of the epidemiology and financial impact of fowl cholera (FC) in California meat turkeys during 1984 was performed. Data were collected from 64 flocks--23 FC-outbreak flocks and 41 controls (non-outbreak)--raised in the Central Valley of the state. Mean flock age at the time of the FC outbreak was 11.3 weeks. Flocks that reported a colibacillosis outbreak had increased odds (P = 0.11) of also having an FC outbreak. (This association may or may not indicate a cause-effect relationship.) There was no significant difference between FC-outbreak and control flocks in number of diseases reported, age at onset, or duration of diseases or syndromes except age at onset of roundheart disease. The relative mortality rates were 52% higher in FC-outbreak toms and 26% higher in FC-outbreak hens than in their controls. Medication costs were nearly tripled, and the relative condemnation rate was 60% higher in FC-outbreak flocks than in control flocks. The average costs of FC were nearly

Pasteurella multocida Recovered from Live Turkeys: Prevalence and Virulence in Turkeys

0.40 per bird, or

Serum Resistance as an Indicator of Virulence of Pasteurella multocida for Turkeys

18,750 per flock, in an outbreak flock of 50,000 birds, and