The fine structure of Fusiformis nodosus with special reference to the location of antigens associated with immunogenicity.
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Agglutination Tests↗
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A gene region in Dichelobacter nodosus encoding a lipopolysaccharide epitope.
Dichelobacter nodosus is a Gram-negative anaerobic bacterium that is the causative organism of footrot in sheep. A D. nodosus locus responsible for a modification of the host lipopolysaccharide (LPS) in Escherichia coli was cloned and sequenced. Genetic studies showed that the modification occurred within the inner-core region of the host LPS, most likely to one or more of the heptose molecules. Antibodies eluted from the modified LPS reacted preferentially with the lipid-A-core region of D. nodosus LPS, suggesting that the cloned epitope was present in this region of the D. nodosus LPS. The gene responsible for the modification, IpsA, potentially encoded a polypeptide of approximately 37 kDa which was highly basic, a characteristic of enzymes which interact with the acidic inner LPS core. The IpsA gene appeared to be arranged in a complex operon with a downstream gene, prfC, which encoded a protein with similarity to E. coli peptide-chain release factor 3.
Amino Acid Sequence↗
Delineation of the virulence-related locus (vrl) of Dichelobacter nodosus.
Dichelobacter nodosus is the primary pathogen implicated in ovine footrot. In this paper we have delineated a 27 kb locus, termed the virulence-related locus (vrl), that was essentially specific for virulent D. nodosus isolates. The precise ends of this locus were mapped and the sequences of the junction regions from the virulent strain A198 were compared to corresponding sequences from the benign isolate C305. The left end of the vrl locus was located in a sequence similar to that of the small stable 10Sa RNA molecule of Escherichia coli, next to a phage-attachment-site-like sequence, which indicated that the vrl locus might have arisen by the integration of a phage. However, no attachment-like sequence could be found at the right end of the vrl locus. In the chromosome of the benign strain the sequences bordering vrl were not contiguous but were separated by about 3 kb. It was concluded that the divergence of the benign and virulent strains at this locus was a multi-step process. Several potential ORFs were identified at the junction regions but only one ORF, encoding a 126 kDa protein, was expressed in a T7 expression system in E. coli.
Amino Acid Sequence↗
An electron microscope study of Bacteroides nodusus: ultrastructure of organisms from primary isolates and different colony types.
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Agglutination Tests↗
Characterization of a spirochaete isolated from a case of bovine digital dermatitis.
AIMS: The aim of the study was to characterize a spirochaete isolated from the lesions of a cow with digital dermatitis (DD). METHODS AND RESULTS: The characterization was on the basis of its light and electron microscopic appearance, enzymic profile and DNA sequence analysis of its flagellin and 16S rRNA genes. The spirochaete was 6-8-microm long and 0.2-0.3 microm in diameter, and possessed seven to eight periplasmic flagella, with three to five helical turns. The enzymic profile of the bacterium resembles, but is not identical to that of Treponema brennaborense. Its flagellin gene sequence was identical to that of Treponema phagedenis but distinct from that of an ovine spirochaete. Analysis of a 1477-bp region of the 16S rRNA genes indicated that this is a Treponema species and that it is indistinguishable from some isolates made from cases of bovine DD in the United States. Finally, electron microscopy revealed the presence of myovirus-like bacteriophage particles in all cultures of the treponeme examined. CONCLUSIONS: The spirochaete isolate was identified as a Treponema species closely related to some isolates from the United States (by 16S rDNA) and to T. phagedenis (by flagellin gene sequence) and is associated with bacteriophage particles. SIGNIFICANCE AND IMPACT OF THE STUDY: The fact that the isolates with the same or very similar 16S rDNA sequences have been obtained from cases of bovine DD in cattle in different countries at different times, lends further support to the hypothesis that treponemes play a role in the pathogenesis of this disease.
Animals↗
The influence of chronic pain on the analgesic effects of the alpha 2-adrenoceptor agonist, xylazine, in sheep.
A comparison of the analgesic potency of the alpha 2-adrenoceptor agonist, xylazine, in control healthy sheep and sheep suffering chronic pain from footrot, indicated that the analgesic effectiveness of xylazine was significantly reduced in the animals experiencing chronic pain. This was measured by recording the threshold to a mechanically applied pressure stimulus. Furthermore, when the condition was apparently resolved, by conventional treatment over a period of 2 to 3 weeks, the decreased analgesic effectiveness of the alpha 2-agonist was still apparent although the animals were clinically cured of the footrot.
Analgesia↗
Comparative in vitro susceptibility of Bacteroides and Fusobacterium isolated from footrot in sheep to 28 antimicrobial agents.
The agar dilution method was used to determine the bacteriostatic activity of 28 antimicrobial agents against 141 strains to the genus Bacteroides and 29 strains from the genus Fusobacterium. All organisms were isolated from clinical cases of ovine footrot. The strains were isolated from 125 Merino sheep, over a period of 2 years, from January 1987 to December 1988. The three ureidopenicillins studied (azlocillin, mezlocillin and piperacillin) proved to be the most effective antimicrobial agents. Chloramphenicol, metronidazole and tinidazole effectively inhibited the growth of Bacteroides spp., while phosphomycin was active against Fusobacterium spp.
Animals↗
The effects of opioid and alpha 2 adrenergic blockade on non-steroidal anti-inflammatory drug analgesia in sheep.
The analgesic effects of the non-steroidal anti-inflammatory drugs (NSAIDs) flunixin and dipyrone were assessed in healthy sheep with no pre-existing inflammation, and in sheep with a chronic inflammatory lesion, using a mechanical noxious stimulus. Saline and dexamethasone were given as controls. Blood taken from healthy sheep after NSAID administration was assayed for thromboxane B2 (TxB2) to compare the ability of these drugs to inhibit cyclo-oxygenase. Both flunixin and dipyrone produced a small but statistically significant rise in pain thresholds (18% and 21% of maximum possible effect respectively) in the healthy sheep which peaked at 30 min and had returned to pre-drug values by 2-3 h. In the lame sheep a similar effect occurred but the response was smaller, much more variable and tended to be prolonged. Saline and dexamethasone had no effect on thresholds over 6 h in either group of sheep. The rise in thresholds was prevented by pre-treatment with naloxone (an opioid antagonist) or atipamezole (an alpha 2-adrenergic antagonist) in the healthy sheep. Naloxone and atipamezole had no effect on thresholds when given alone to healthy sheep. Both NSAIDs inhibited the production of TxB2 to a similar extent. These results indicate that central mechanisms may be involved in NSAID analgesia.
Adrenergic alpha-Antagonists↗
Minimum inhibitory concentrations for 25 selected antimicrobial agents against Dichelobacter nodosus and Fusobacterium strains isolated from footrot in sheep of Portugal and Spain.
The agar dilution method was used to determine the inhibitory activity of 25 antimicrobial agents against 69 strains of Dichelobacter nodosus and 108 strains of the genus Fusobacterium, all of which were isolated from 90 clinical cases of ovine footrot between October 1998 and November 2000. In the case of the micro-organisms belonging to the genus Fusobacterium, the six beta-lactams studied (benzyl penicillin, ampicillin, cloxacillin, cefadroxil, cefuroxime and cephalexine) proved to be, in general, the most effective antimicrobial agents. Chloramphenicol, clindamycin and doxycycline were also quite active against Fusobacterium spp. With regard to the 69 strains of D. nodosus tested, the levels of resistance remain low.
Animals↗
Efficacy of the E-test in evaluating the antimicrobial susceptibility of anaerobic bacteria of veterinary interest.
A study was made of the sensitivity of 39 clinical isolates of anaerobic bacteria to 10 antimicrobial agents. Minimal inhibitory concentrations (MICs) were calculated using a new method--the E-test (AB Biodisk, Solna, Sweden)--and compared with those obtained using the conventional agar dilution method. Agreement between the MICs obtained by the two methods with a variation of +/- 2 dilutions was 78.7%. The E-test, though less sensitive than the conventional agar dilution method, may be of value in clinical veterinary practice when rapid selection of treatment for a given infectious process is required.
Animals↗
Virulence regions and virulence factors of the ovine footrot pathogen, Dichelobacter nodosus.
Ovine footrot is a debilitating and highly infectious disease that is primarily caused by the Gram-negative, anaerobic bacterium Dichelobacter nodosus. The major antigens implicated in virulence are the type IV fimbriae and extracellular proteases. The fimbriae show sequence and structural similarity to other type IV fimbriae, this similarity extends to genes that are involved in fimbrial biogenesis. Several acidic and basic extracellular serine proteases are produced by both virulent and benign isolates of D. nodosus. Subtle functional differences in these proteases appear to be important in virulence. In addition, there are two chromosomal regions that have a genotypic association with virulence. The partially duplicated and rearranged vap regions appear to have arisen from the insertion of a plasmid into a tRNA gene via an integrase-mediated site-specific insertion event. The 27 kb vrl region has several genes often found on bacteriophages and has inserted into an ssrA gene that may have a regulatory role in the cell. The determination of the precise role that each of these genes and gene regions has in virulence awaits the development of methods for the genetic analysis and manipulation of D. nodosus.
Animals↗
Electroporation-mediated transformation of the ovine footrot pathogen Dichelobacter nodosus.
Studies on the potential virulence genes of the ovine footrot pathogen Dichelobacter nodosus have been hindered by the lack of a genetic system for this organism. In an attempt to accomplish the transformation of D. nodosus cells, we constructed a plasmid that contained part of a native D. nodosus plasmid and carried a tetracycline resistance gene that was located between the D. nodosus rrnA promoter and terminator. This plasmid was used to transform several D. nodosus strains to tetracycline resistance. Analysis of two independent transformants from each parental strain showed that in nearly all of these derivatives, the plasmid was not replicating independently, but that the tetracycline resistance gene had inserted by homologous recombination into one of the three rrn operons located on the chromosome. In most of the transformants, double reciprocal crossover events had occurred. These results are highly significant for genetic studies in D. nodosus and for footrot pathogenesis studies, since by using reverse genetics it will now be possible to examine the role of putative D. nodosus-encoded virulence genes in the disease process.
Animals↗
Vaccination against ovine footrot using a live bacterial vector to deliver basic protease antigen.
A strain of Corynebacterium pseudotuberculosis, designated Toxminus, that has been rationally attenuated by deletion of the phospholipase D gene, is being developed as a live vaccine vector for the delivery of veterinary vaccine antigens. In the present study a recombinant form of the basic protease gene of the ovine footrot causative bacterium, Dichelobacter nodosus, was introduced into the vector strain using the high copy number plasmid pEP2. This strain secreted the basic protease protein. Vaccination trials in sheep with the recombinant strain demonstrated that, although an IgG immune response was elicited, the animals were not protected from footrot following artificial challenge under pen conditions. Although the sheep were not protected there was evidence that the progression of the disease was slowed.
Animals↗
Transmission of Fusiformis nodosus infection from cattle to sheep.
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Animals↗
Changes in animal disease patterns in the Riverina of New South Wales.
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Animals↗
A degrading proteinase test to distinguish benign and virulent ovine isolates of Bacteroides nodosus.
Bacteroides nodosus was recovered from naturally occurring cases of virulent ovine footrot (VFR) and benign footrot (BFR) using an artificial culture medium which incorporated trypticase, arginine, serine and 5% agar. A degrading proteinase (DP) test was developed which measured the proteinase activity of broth cultures of B. nodosus for a period of time after organism death to assess the stability of the enzyme. The spectrophotometric measurement of the release of dye from a hide powder--azure conjugate by the action of proteinase provided an objective analysis of enzyme activity. The DP test differentiated VFR and BFR isolates and promises to be a useful laboratory method for the diagnosis of benign and virulent footrot in sheep.
Animals↗
Isolation and characterisation of Bacteroides nodosus from foot lesions of cattle in Western Australia.
Thirty one isolates of Bacteroides nodosus were obtained from foot lesions observed on cattle at 3 abattoirs. All isolates were similar to the B. nodosus of ovine benign footrot (BFR) in their response to the degrading proteinase test. At one abattoir, where the interdigital lesions were examined in detail, 9 of 10 isolates were obtained from hyperkeratotic lesions with deep fissures. Traceback to 8 of the farms of origin which carried both sheep and cattle, revealed BFR in sheep on 4 farms. The significance of B. nodosus in interdigital lesions in cattle, and its possible pathogenicity, are discussed.
Animals↗