Use of a monoclonal antibody to ovine IgE for fly strike studies in sheep.

A new monoclonal raised against sheep IgE was used to examine sera and wound exudates from sheep which had been struck by Lucilia cuprina in the field. The antibody was also used to detect the presence of IgE in sera and skin sections from sheep which had been artificially infected with fly larvae 3 times. Neither total, nor L. cuprina specific circulating IgE could be detected in serum or wound exudates from struck sheep. Cell bound IgE was, however, identified by the monoclonal in skin sections from struck sheep and from a control sheep which had not been struck. No difference in the number of IgE positive cells was observed between the control and 2 of the 3 artificially infected sheep, and none of the latter showed an increase in IgE positive cells even after 3 infections. One sheep showed twice as many IgE positive cells as the other treated sheep and the third larval infection was difficult to establish and limited in size and severity. This suggests a relationship between innate resistance to strike and the number of IgE positive cells present in skin.

Characterization of Pseudomonas maltophilia Isolates from Fleece Rot.

A total of 286 isolates of Pseudomonas maltophilia was collected from sheep exhibiting brown or yellowish fleece rot and from fly-strike lesions. Enzyme activities for 10 of the isolates were examined by plate tests and with the API ZYM system and compared with the enzymatic profile of a human type strain of P. maltophilia. The fact that ovine isolates of P. maltophilia are biochemically similar to pathogenic human strains suggests there may be an association between this organism and the brown to yellow type of fleece rot.

Immunisation of sheep against experimental Pseudomonas aeruginosa dermatitis and fleece-rot associated body strike.

Sheep immunised with an experimental Pseudomonas aeruginosa vaccine and unvaccinated control sheep were challenged by induction of experimental dermatitis with the homologous strain. All of 6 control sheep developed ulcerative dermatitis, and 2 of the 6 challenge sites were struck by larvae of Lucilia cuprina. Neither severe dermatitis nor strike occurred in 6 vaccinated sheep. These results were confirmed in an experimental challenge using 3 different serotypes of P. aeruginosa on each of 3 vaccinated and control sheep, although fly-strikes did not occur. In a field trial of the same vaccine, none of 26 vaccinated sheep developed severe exudative, fleece-rot lesions nor were any fly-struck, whereas 61 of 115 control sheep developed severe, exudative, fleece-rot lesions, 21 of these becoming struck by L. cuprina. The isolates of P. aeruginosa recovered from the field challenge experiment were a different serotype to that used in the vaccine.

Characterisation of isolates of Pseudomonas aeruginosa from sheep.

Fleece rot isolates of Pseudomonas aeruginosa were characterised in terms of biochemical reactions, serological typing and production of keratinase, lipase and the potentially dermal necrotic enzymes exotoxin A, protease, elastase phospholipase and lecithinase. The similarities generally obtained between these characteristics of isolates from sheep and man suggests a role for P. aeruginosa and mechanisms of pathogenesis in the development of the dermatitis associated with fleece rot.

Experimental production of dermatitis in sheep with Pseudomonas aeruginosa.

The attachment, to sheep skin, for 4 days, of control wool pads saturated with sterile culture medium which contained a bacteriostat, induced only a mild dermatitis, whereas wool pads saturated with Pseudomonas aeruginosa culture induced a subacute dermatitis characterised by scaling, microabscess formation and seropurulent exudate. Changes similar to the latter were observed in skin affected by natural fleece-rot which developed spontaneously after 7 days of artificial wetting and in which P. aeruginosa was the predominant species of bacteria. An exacerbatory, if not causal, role for this organism is suggested in the development of the dermatitis associated with fleece-rot and in the exudation of seropurulent material, a step essential in the development of body strike.

Caseous lymphadenitis in goats.

Two dairy goat stud herds were surveyed for Corynebacterium ovis infection by clinical examination and serum tests using both the haemolysis inhibition and double immunodiffusion techniques for detection of C. ovis antitoxin. In one herd, of 53 goats 11 had abscesses from which C. ovis was recorded; 33 and 29 goats were positive to the haemolysis inhibition and double immunodiffusion tests respectively. Serums from 10 and 9 of 23 kids gave reactions positive to the respective tests with 19 of 23 kids having been born to does with demonstrable antibody. In a second herd of 57 goats 16 had superficial abscesses containing C. ovis; serums from 34 and 45 goats gave positive reactions to the haemolysis inhibition and double immunodiffusion tests respectively. The prevalence of caseous lymphadenitis in these stud herds was a cause of economic loss to the owners through euthanasia of severely affected goats, reduced milk production and emaciation, the cost of labour and drugs for treatment, jeopardisation of income from boarding or mating with outside stock and the appearance of goats at stud shows, and sale of valuable breeding stock to local and export markets.

A simplified double immunodiffusion technique for detection of Corynebacterium ovis antitoxin.

A method is described for obtaining yields of Corynebacterium ovis exotoxin sufficiently high to allow use of unconcentrated supernatant and serum as the reactants in a double immunodiffusion test. C ovis supernatant with a haemolytic titre or rabbit dermal necrosis titre of 1:16,394 or greater produced readily detectable precipitation lines with specific antitoxin in the range of concentrations commonly found in sera from cases of natural C ovis infection. When used as a diagnostic screening test, sera from 32 confirmed cases of caseous lymphadenitis in sheep and goats were positive whereas sera from 16 sheep free of caseous lymphadenitis were negative. Sera from 10 out of 16 kids which had been suckled by mothers from an affected flock of goats were positive.

A haemolysis inhibition test for detection of antibody to Corynebacterium ovis exotoxin.

A haemolysis inhibition test for detection of antibody to Corynebacterium ovis exotoxin is described. The basis of the test is te inhibition, by immune serum, of the haemolytic activity of C ovis exotoxin for sheep red cells. When the test was applied to sera from sheep which had been vaccinated with C ovis cells and exotoxin, the geometric mean titre increased from < 2 to 137 whereas in sheep vaccinated with C ovis cells only, the mean titre remained < 2. Experimental infection with a toxigenic C ovis strain produced titres ranging from 2 to 32 in 10 out of 12 sheep but a nontoxigenic C ovis strain did not produce a response in any of six sheep. The test was successful in diagnosing 24 out of 30 sheep with caseous lymphadenitis evident at autopsy. Antitoxin was also detected in the serum of six out of seven lambs suckled by ewes which had positive titres.

In vitro direct haemagglutination by Corynebacterium ovis exotoxin.

Haemagglutinating activity of toxigenic Corynebacterium ovis culture supernatants was estimated by titration in phosphate buffered saline diluent against sheep erythrocytes. The haemagglutination titre was closely related to the haemolysis and rabbit dermonecrosis titres of culture supernatants and to the toxigenicity of different C ovis strains. The substance responsible for haemagglutination was diffusible and its concentration was unrelated to cell yield in cultures. Corresponding inactivation of all three activities by antitoxin, toxoid formation with formalin, heat and Seitz filtration suggested that haemagglutination is another activity of C ovis exotoxin.

Conditions for in vitro haemolytic activity by Corynebacterium ovis exotoxin.

A substance, concluded to be Corynebacterium ovis exotoxin on the basis of properties shared, was found to have haemolytic activity below pH 6 and red cell adhesion activity at neutral or slightly alkaline pH. An acidified, solid blood agar medium was used to demonstrate the extent of haemolysis that could be obtained at optimal pH and its inhibition by immune serum. Culture in a liquid medium giving rise to a suitably acidic pH allowed titration for haemolytic activity by the exotoxin and use of supernatant in a haemolysis-inhibition test to detect antitoxin.

Non-specific agglutination of Corynebacterium ovis by precolostral and young lamb sera.

An agglutination reaction, relatively coarse compared to that obtained with immune sera, was observed when Corynebacterium ovis cells were incubated with lamb sera collected before and after ingesting colostrum. The reaction did not appear to involve immunoglobulin since it was not affected by treatment of precolostral lamb serum with rivanol, mercaptoethanol or rabbit antisheep serum. IgM was detected in only three of 10 precolostral samples with agglutinating activity and immunofluorescence studies showed that it was not specific for C ovis. In view of the non-specific clumping factor, tests which detect antitoxin would be preferable for immune surveys in young lambs.

Experimental induction of caseous lymphadenitis in sheep by intralymphatic inoculation of Corynebacterium ovis.

Lesions histologically similar to those of natural caseous lymphadenitis developed in sheep popliteal lymph nodes following injection of viable Corynebacterium ovis cells into afferent lymphatic ducts. Histopathological examination of affected lymph nodes suggested that exotoxin might be involved in the establishment, progressive development and persistence of caseous lymphadenitis lesions through localised cytotoxicity.