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Production and characterization of monoclonal antibodies to lipopolysaccharide antigen of Leptospira interrogans serovar kremastos and canicola.

Monoclonal antibodies to Leptospira interrogans were provided by cell fusions between a myeloma cell line, P3/X63-Ag8.653, and spleen cells of BALB/c mice immunized with two different leptospiral lipopolysaccharide antigens. One antigen was prepared from Leptospira interrogans serovar kremastos strain Kyoto and the other from serovar canicola strain Hond Utrecht IV. Eighteen hybridoma cell lines secreting monoclonal antibodies to the former organisms and five hybridoma cell lines secreting monoclonal antibodies to the latter organisms were established during 6 month cultivation. On the basis of their microscopic agglutination reactivities, 18 anti-kremastos Kyoto monoclonal antibodies were classified into 10 distinct groups, and 5 anti-canicola monoclonal antibodies into 3 distinct groups.

Agglutination Tests↗

Evaluation of a recombinant LipL41 antigen of Leptospira interrogans serovar canicola in ELISA for serodiagnosis of bovine leptospirosis.

The efficacy of a recombinant leptospiral lipoprotein LipL41 as an antigen for conducting enzyme-linked immunosorbent assay (ELISA) for diagnosis of bovine leptospirosis was evaluated. Using known positive and known negative cattle sera the recombinant antigen was found to be highly reactive in the concentration of 100 ng/well. Using a total of 321 field cattle sera the sensitivity of ELISA as compared to microscopic agglutination test (MAT) was calculated to be 100% whereas the specificity was 85.3%. The seropositivity of leptospirosis among bovine population was found to be 21.18% having the predominance of serovars Sejroe and Pomona. It was concluded that rLipL41 protein could be a putative diagnostic candidate for serodiagnosis of bovine leptospirosis.

Agglutination Tests↗

Immunity to leptospirosis: antiserums in dogs and hamsters.

Resistance to clinical and renal leptospirosis in dogs injected with antiserum to Leptospira interrogans serotype canicola and Leptospira interrogans serotype icterohaemorrhagiae was demonstrated. The relationship between resistance induced in serum-injected hamsters was also demonstrated, using criteria of leptospiremia, renal infection, leptospiruria, and clinical signs of disease in dogs and death in hamsters.

Animals↗

Immunity to leptospirosis: bacterins in dogs and hamsters.

Resistance to clinical and renal leptospirosis in dogs vaccinated with a bacterin containing Leptospira interrogans serotype canicola and Leptospira interrogans serotype icterohaemorrhagiae was demonstrated. The relationship between resistance induced in vaccinated dogs and that induced in vaccinated hamsters was also demonstrated, using criteria of leptospiremia, renal infection, leptospiruria, and clinical signs of disease in dogs and death in hamsters.

Animals↗

Leptospira interrogans serovar canicola: a causal agent of sow abortions in Arequipa, Peru.

An outbreak of abortions, stillbirths, mummified piglets and neonatal deaths in a pig herd in Arequipa, Peru is described. A total of 31 of 240 sows aborted between May and September 1988. When sera were examined 12 of 14 had very high titres of antibody to canicola PC125 and canicola Hond Utrecht, but there were also high titres of antibody to other leptospiral serovars. A detailed investigation was made and serovar canicola PC125 was isolated from the urine of four sows which had aborted and the kidney of one slaughter pig. Antibodies to various serovars of Leptospira were demonstrated in 11 of 17 sows which had aborted, two of six sows which had normal litters, nine of 18 boars, four of 39 slaughter pigs and four of 14 workers on the farm. The outbreak was brought under control by treatment and vaccination coupled with a thorough cleaning of the farm and control of the wild animal population. It is suggested that the infection was brought onto the farm by wild animals and that the disease is more common in Arequipa than was previously supposed.

Abortion, Veterinary↗

Selection of antigenic variants from Leptospira interrogans serovar canicola by means of anti-canicola monoclonal antibody.

Antigenic variants were isolated from canicola by a single selection with anti-canicola monoclonal antibody CT-3. The variants were not identical to any serovars of serogroup Canicola and thought to be a new serovar. Variation frequency was calculated at 5.9 X 10(-4). The usefulness of monoclonal antibodies for selection of antigenic variants of leptospiras is discussed.

Agglutination Tests↗

Isolation and biological activities of endotoxin from Leptospira interrogans.

Endotoxins extracted with ethylenediaminetetraacetate (EDTA) from Leptospira interrogans serovars icterohaemorrhagiae and canicola and Leptospira biflexa serovar patoc were tested for various biological activities characteristic of endotoxins. The presence of lipopolysaccharide biological activity was demonstrated by the Limulus amoebocyte lysate test, pyrogenicity in rabbits, complement interaction inhibiting the erythrocyte lysis, and chicken-embryo lethality. The lipopolysaccharides did not induce the local Shwartzman reaction. The lipopolysaccharides of serovars icterohaemorrhagiae and canicola were immunogenic in rabbits and were cytotoxic to chicken-embryo fibroblasts.

Animals↗

USE OF ANTIBIOTICS IN THE PREPARATION OF CANINE KIDNEY TISSUE CULTURE VACCINES TO ELIMINATE LEPTOSPIRAL INFECTION HAZARDS.

The potential leptospiral infection hazard in the use of vaccines prepared from canine kidney monolayer cultures was studied. Cell cultures were prepared from kidneys of dogs experimentally infected with Leptospira serotype canicola. Viable leptospires were found in kidney cell suspensions at the time of seeding, surviving trypsinization either at room temperature for approximately 2 hr or overnight at 4 C, even in the presence of antibiotics. In tissue cultures maintained without antibiotics, leptospires were cultured up to the time of involution of cells at 25 to 34 days of incubation. Cytopathogenic effects of leptospires on cultured kidney cells were not noted; neither was growth of leptospires remarkable. Generally, the leptospire culture titer decreased to 10(-4) or 10(-5) at the 4th hr or 1st day of incubation to 10(-1) or negative by the 30th or 34th day of incubation. The addition of either a combination of penicillin (100 units per ml) plus streptomycin (100 mug/ml) or polymyxin B (50 units per ml) plus dihydrostreptomycin (100 mug/ml) to seeding cell suspensions resulted in the elimination of viable leptospires by the 4th hr of incubation. From cell cultures treated with neomycin (100 mug/ml) or chloramphenicol (100 mug/ml), leptospires were recovered, respectively, after 24 and 48 hr, but not thereafter. It was apparent that antibiotics, particularly the combination of polymyxin B and dihydrostreptomycin, could be effectively used to eliminate leptospires in tissue culture. Other antibiotics with known antileptospiral activities probably would be effective also. If antibiotics are not used in canine kidney tissue culture employed for viral vaccine preparations, rigid testing for the presence of leptospires in donor dogs and tissue-culture vaccine is indicated.

Animals↗

LEPTOSPIRAL COLONIAL MORPHOLOGY.

Stalheim, O. H. V. (University of Wisconsin, Madison) and J. B. Wilson. Leptospiral colonial morphology. J. Bacteriol. 86:482-489. 1963.-A sequence of apparent colonial types was observed with colonies of Leptospira pomona, L. canicola, L. icterohaemorrhagiae, and L. grippotyphosa in agar medium. Although some colonies of these serotypes had a different appearance initially, they eventually developed the mature or final appearance characteristic of the serotype. Colonies of freshly isolated, virulent cultures of L. pomona, L. canicola, and L. icterohaemorrhagiae were similar in appearance to colonies of avirulent strains of the same serotype. Additional studies of three stable and distinct colonial types of a laboratory strain of L. autumnalis revealed no differences in antigenicity, catalase activity, or mouse infectivity; however, differences in susceptibility to lysis by oleic acid were found. Although the colonial variants were stable during several in vitro variations, including growth in the presence of homologous antiserum and mutation to growth in a chemically characterized medium, rapid dissociation in vivo was found.

Animals↗