Haemolytic and cytotoxic activities of the Tween 80-extracted putative haemolysin of Pasteurella multocida B:2.

The objective of this study was to investigate the haemolytic and cytotoxic activity of Pasteurella multocida B:2 strains, originally from cases of haemorrhagic septicaemia in cattle. All six P. multocida B:2 strains were non-haemolytic on sheep blood agar (SBA) and horse blood agar (HBA) when grown aerobically and on SBA anaerobically but they were haemolytic on HBA when grown anaerobically. No haemolytic activity against horse red blood cells was detected in culture supernates from aerobically or anaerobically grown cultures and only very weak haemolytic activity was obtained in supernates or pellet fractions from sonicated cells. However, after repeated extraction of sonicated cells with Tween 80, haemolytic activity was found in various cell fractions, both Tween-soluble and -insoluble. The Tween-extracted putative haemolysin and other bacterial fractions were also cytotoxic for mouse macrophage-like J774.2 cells. Further characterisation of the putative haemolysin revealed it to be a heat-labile, non-pore-forming protein of molecular weight >10 kDa whose activity was completely destroyed by trypsin and greatly reduced with protease and proteinase K treatment. Congo red also reduced the haemolytic activity. Non-denaturing gel-electrophoresis and RBC agar overlay revealed clear haemolytic zones but suggested that Tween was bound to some component of the P. multocida B:2 fractions and was responsible, to some extent, for the haemolytic activity observed. However, the effect of heat and other reagents on the Tween-extracted fractions and the lack of haemolytic activity in different Tween-extracted cell fractions of organisms other than P. multocida suggested that some proteinaceous component of the organism could indeed act as a haemolysin. This putative haemolysin may be one of the virulence attributes of P. multocida, but its characterisation and role in pathogenesis require further study.

Prevalence of aerobic and anaerobic uterine bacteria during peripartum period in normal and dystocia-affected buffaloes.

Parturition complications predispose establishment of uterine infections, which in turn affect subsequent fertility. The aim of present study was to characterize and compare the type of bacterial flora prevalent within the uterine lumen of dystocia-affected buffaloes and compare them with the normally calving buffaloes. The study was conducted on 40 buffaloes; of which 10 calved normally (Group I) and 30 were treated for dystocia (Group II). Bacteriological examination was performed using uterine swabs, which were collected before delivery, immediately after delivery and day's 24-60 postpartum. A total of 30 uterine swabs from Group I and 79 swabs from Group II were collected, of which 19 (63.3%) and 71 (89.9%) yielded significant bacterial growth, respectively. A total of 205 isolates belonging to 10 different genera of bacteria were identified, 8 facultative anaerobes and 2 obligate anaerobes. In Group II, 91.6% of the bacteria positive swabs (n = 71) yielded mixed cultures, whereas the remainder being pure cultures. In contrast, 89.5% of the bacteria positive swabs of Group I (n = 19) yielded pure cultures. Mixed infections comprised mostly Arcanobacter (Actinomyces) pyogenes together with obligate anaerobes, Fusobacterium spp. and Bacteroides spp. In Group II, the frequency of incidental and coliform group bacteria was highest at the time of parturition, i.e., before and immediately after delivery, and decreased to nil during the 24-60-day postpartum period. However, in Group I, the incidental and coliform group of bacteria present at the time of parturition apparently persisted beyond the period when uterine involution is complete. The frequency of obligate anaerobes and A. pyogenes at the time of parturition was nil in the Group I while they predominated in dystocia-affected buffaloes (Group II). During the postpartum period of 24-60 days, the frequency of both obligate anaerobes and A. pyogenes increased significantly in Group II, whereas in Group I, only occasional isolates were obtained. To conclude, at the time of calving the prevalence of obligate anaerobes and A. pyogenes occurring in combination was highest in dystocia-affected buffaloes, and further increased in the postpartum period suggesting that these infections act synergistically.

Sequential pathological studies in Asian water buffaloes infected intratracheally with Absidia corymbifera.

Zygomycosis was produced experimentally in Asian water buffalo calves (Bubalus bubalis) by intratracheal inoculation of sporangiospores of Absidia corymbifera. Infected animals exhibited dullness, depression, partial anorexia, initial pyrexia, mucopurulent nasal and ocular discharge and coughing during the first week. There was no mortality at any stage of the experiment, which continued for 30 days. The gross and microscopic lesions were restricted to the lungs and there was no dissemination of the fungus to other organs. Gross and microscopic changes in the lungs were observed up to the 20th day post-infection. Gross lesions consisted of pneumonic consolidation of the antero-ventral lobes of the lungs. Microscopic changes consisted of granulomatous reactions with well developed pulmonary granulomas. Distorted hyphae of A. corymbifera were demonstrated in tissue sections up to 15 days post inoculation. Re-isolation of the fungus was achieved consistently for up to 15 days. It is concluded that intratracheal inoculation of A. corymbifera in buffalo calves leads to significant pathological changes in the lungs, but the disease appears to be self limiting 20 days following inoculation.

Experimental candidal mastitis in goats: clinical, haematological, biochemical and sequential pathological studies.

The present study, first of its kind, was conducted with the objectives to understand hitherto little known aspects of candidal mastitis, like its sequential pathology, pathogenesis and clinico-biochemical changes. For this purpose, unilateral intramammary inoculation of 10 goats with Candida albicans (1.2 x 10(7) yeast cells) resulted in the development of mastitis, with gross and microscopic lesions being restricted to the infected udder halves only and without dissemination of infection to the opposite uninfected udder halves as well as other organs of the body. The experiment was continued for 40 days and after infection, there was sharp fall in milk yield and Candida albicans was directly demonstrated in the milk and re-isolated from the milk and udder tissues up to 30th day after inoculation. An increase in total immunoglobulins in the milk and plasma along with increase in total plasma proteins were also observed. Haematology revealed leukocytosis and neutrophilia. Microscopically, there was acute purulent mastitis, which later became chronic, nonpurulent and interstitial with formation of granulomas. It was concluded that Candida albicans was highly pathogenic to the lactating goat mammary gland even without immunosuppression or antibiotic treatment, resulting in severe irreversible tissue damage and nearly complete agalactia.

Epidemiology of inclusion body hepatitis in poultry in northern India from 1990 to 1994.

The epidemiology of inclusion body hepatitis (IBH) was studied in poultry in northern India, from April 1990 to March 1994, to evaluate the various factors responsible for causing and determining the severity of the disease. Broiler chicks and Japanese quail (Coturnix coturnix japonica) were the species examined. The factor observed to be most commonly associated with IBH was the presence of aflatoxins in the feed at higher than permissible levels, i.e. 20 parts per billion. Avian adenovirus-1 was isolated from the livers of affected birds. In the final year of the study, a number of outbreaks of IBH caused heavy mortalities among three to five-week-old broiler chicks, which displayed typical IBH lesions in addition to hydropericardium.

Clinico-pathological studies on experimental cryptococcal mastitis in goats.

Unilateral intramammary inoculation of 10 goats with Cryptococcus neoformans (2 x 10(6) yeast cells) resulted in the development of mastitis, with gross and microscopic lesions being restricted to the infected udder halves only and there was no dissemination of infection to the opposite uninfected udder halves as well as to other organs of the body. The experiment was continued for 40 days, with 2 animals each from the infected and control groups being killed on 5th, 10th, 20th, 30th and 40th day post-inoculation (DPI). Initial enlargement of the infected udder halves was followed by marked decrease in size leading to very small, firm and nodular udder halves. After infection, there was also sharp fall in the milk yield. Cryptococcal organisms were demonstrated in the mastitic milk and udder impression smears with special stains. C. neoformans was reisolated from the milk of the only infected udder halves up to 25th DPI. Microscopically, there was initially acute diffuse purulent mastitis which later on became chronic, characterised by marked infiltration of lymphocytes, macrophages, extensive fibrosis and development of multiple granulomas. The cryptococcal organisms could be demonstrated in the udder sections only up to 30th DPI. It is concluded that intramammary inoculation of Cryptococcus neoformans in goats leads to severe mastitis with sharp fall in milk yield.

Outbreak of mycotic tracheitis in domestic fowl.

An outbreak of mycotic tracheitis was observed in 8000 2-month-old, female White Leghorn birds. The birds were showing difficult respiration and there was mortality of 7-8 birds daily in the flock. On post-mortem examination of the affected birds, the trachea was found to be occluded with a white caseous mass. Microscopically hyphae were found invading the tracheal epithelium, cartilage and serosal layer along with infiltration of macrophages and lymphocytes. Aspergillus fumigatus was isolated in pure culture from the trachea. The birds responded to oral copper sulphate treatment. The ubiquitous occurrence of the organism and the conditions of the harvesting season have been found to be responsible for the outbreak of the disease.

Mucormycosis of the bovine udder.

Nine cross-bred cows suffering from mucormycosis of the udder were observed. Histopathological and cultural examination of the lesions revealed the presence of Mucor spec.

Dermatomycosis in dogs.

During the routine examination of dogs for cutaneous lesions, 205 dogs were screened for fungi other than dermatophytes. Twenty-two dogs (10.8%) revealed the presence of non-dermatophytic fungi suspicious for representing the etiologic agents of the skin lesions. The fungi isolated were Alternaria sp. (2.9%), Penicillium sp. (2.4%), Aspergillus fumigatus (2.0%), Mucor sp. (1.5%), Cladosporium sp. (1.5%) and Fusarium sp. (0.5%). No dermatophyte was isolated in association with these fungi. The incidence of these infections was found to be greater in warm and humid climate.