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PubMed1964

CULTIVATION OF LEPTOSPIRAE. II. GROWTH AND LYSIS IN SYNTHETIC MEDIUM.

Stalheim, O. H. V. (University of Wisconsin, Madison), and J. B. Wilson. Cultivation of leptospirae. II. Growth and lysis in synthetic medium. J. Bacteriol. 88:55-59. 1964.-Differences were found in the ability of leptospirae to grow in a synthetic medium; 43 strains, consisting of 16 serotypes, were tested and designated as either type I or type II. Type I leptospirae did not grow; type II grew and could be subcultured. The lytic effect of several lipids was measured with Leptospira pomona and L. canicola as representatives of type I and II leptospirae, respectively. L. pomona organisms were rapidly lysed by the monoolein of the synthetic medium and by other lipids as well; L. canicola cells were consistently more resistant. Although both organisms incorporated similar amounts of label when incubated in the presence of oleic-1-C(14) acid, only L. canicola grew in a modified, nonlytic synthetic medium. No differences were found in susceptibility to lysis between virulent and avirulent L. canicola organisms. Mutant type I leptospirae grown in synthetic medium had increased resistance to lysis by surface-active agents; they were poorly agglutinated by antiserum. The role of protein in the growth and antigenicity of type I leptospirae is discussed.

Bacteriolysis↗
PubMed1983

Analysis of Leptospira antigens by crossed immunoelectrophoresis.

Serovars icterohaemorrhagiae, canicola, and arboreae of pathogenic Leptospira interrogans and serovar patoc of saprophytic L. biflexa were examined by crossed immunoelectrophoresis. A close antigenic relationship was found between the interrogans serovars, particularly between icterohaemorrhagiae and canicola. To a much lesser extent cross-reactions were found between interrogans serovars and patoc. Comparison of three different antigenic preparations of icterohaemorrhagiae by tandem crossed immunoelectrophoresis and by absorption experiments with the patoc reference system showed the presence of several common broadly reactive antigens. Antigen 1 of the reference system, a genus-specific heat-stable protein antigen, was found in all antigenic preparations. It is a diagnostically important antigen.

Animals↗
PubMed1984

Serodiagnosis of canine leptospirosis by solid-phase enzyme-linked immunosorbent assay.

An enzyme-linked immunosorbent assay (ELISA) to detect antibodies to Leptospira interrogans serotype canicola in dogs was developed and evaluated. Comparison of the ELISA with the microscopic agglutination test (MAT) showed that, during the first two weeks after an experimental infection with serotype canicola, the ELISA detected antibody at higher dilutions than the MAT. After the second week post-infection both tests detected antibody at almost equal titres (r = 0.89). The outer envelope (OE) antigen of serotypes icterohaemorrhagiae, copenhageni and canicola was fairly serotype-specific, whereas the pellet (P) antigen showed more cross-reactivity. Both OE and P antigen of Leptospira biflexa strain Patoc I could be used as cross-reacting antigen in the ELISA. Compared to the MAT, the ELISA has some technical advantages. It is suggested that the ELISA would be useful as a screening test.

Agglutination Tests↗
PubMed1984

Leptospiral attachment to cultured cells.

Each virulent strain of copenhageni, canicola and pomona of Leptospira interrogans attached effectively to MDCK cells and primary dog kidney cells, while the avirulent or less virulent line of the same strain and avirulent strains belonging to the same serovars and the avirulent reference strains of other serovars did not. Inhibition of the attachment of the virulent copenhageni to MDCK cells was found in the presence of the homologous immunoglobulin G Fab fragment. Strains of L. biflexa attached to the animal cells, but they differed from those of virulent L. interrogans in their capability to attach to glass.

Adhesiveness↗
PubMed1976

Some leptospira agglutinins detected in domestic animals in British Columbia.

During a period of six years 7,555 bovine sera, 421 canine sera, 251 porcine sera and 135 equine sera were tested for agglutinins to Leptospira interrogans serotypes canicola, grippotyphosa, hardjo, icterohemorrhagiae, pomona and sejroe. The bovine sera reacted predominantly with hardjo and/or sejroe at a rate of 15% compared to 3.5% with pomona. Breeding or abortion problems were associated with pomona but not with sejroe/hardjo agglutinins. The canine sera reacted to canicola (9.9%y and icterohemorrhagiae (5.4%), tcted predominantly with canicola (8.9%) and icterohemorrhagiae (8.1%).

Agglutinins↗
PubMed1972

Isolation of the outer sheath of Leptospira and its immunogenic properties in hamsters.

Spherical forms of Leptospira interrogans serotype canicola Hond Utrecht IV were induced with 1 m NaCl. Electron micrographs of these salt-altered cells (SAC) revealed that the outer envelope or sheath had pulled away from the protoplasmic cylinder. Treatment of SAC with 0.02% sodium lauryl sulfate solubilized the sheath and released the protoplasmic cylinder. Further processing of the solubilized sheath yielded a pellet which displayed a membrane structure in electron micrographs. The released protoplasmic cylinder showed loss of intracellular organization and the outer envelope present in normal cells. Immunization of hamsters with whole formalized cells, SAC, or sheath in doses as low as 10 mug/animal protected them from death upon challenge with virulent canicola 27.

Animals↗
PubMed1984

[Incidence of antibodies against leptospira in the blood of breeding sows].

Within a year, 4438 blood sera of sows coming from 25 farms of the Strakonice district were subjected to serological examination by the reaction of microagglutination lysis (RMAL) with 12 strains of Leptospira. The reaction was positive in 3.22% of all the blood sera. Separate significant titres were demonstrated in the reaction with the Leptospira organisms of four serological groups, including Grippotyphosa, Icterohaemorrhagiae, Hebdomadis and Australis. The reactions with the Leptospira of the Canicola serological group were observed only individually as coagglutination with a simultaneous occurrence of antibodies to Leptospira of the Grippotyphosa serological group. The highest number of reactions 119 (75.8%) was demonstrated in titre 400.

Animals↗
PubMed1976

Leptospirosis in Trinidad and Grenada, with special reference to the mongoose.

In Trinidad, six Leptospira isolates were made from 957 febrile patients between 1968 amd 1972. In addition, CF antibodies were detected in 6-6% of febrile patients and human survey sera collected during this period. In 1972 alone, 10-4% (38/363) of sera examined had CF titres consistent with positive exposure to the disease. Grenada does not report leptospirosis, but the disease is common in mongooses in both Trinidad and Grenada. Serogroups Icterohaemorrhagiae, Pomona and Canicola are present in Grenada with 35-2% of mongooses examined being seropositive. Five Canicola strains of Leptospira were isolated from mongooses in Trinidad and serological studies showed that this was the most common serogroup from mongooses on the island. A total of 31 strains recorded from at least seven different serogroups and eight named serotypes have been isolated from humans, rats and mongooses in Trinidad. Human leptospirosis is probably more common in the Caribbean than the medical records indicate.

Animals↗
PubMed1982

Immunogenicity and humoral and cell-mediated immune responses to leptospiral whole cell, outer envelope, and protoplasmic cylinder vaccines in hamsters and dogs.

The immunogenicity of 2 leptospiral cell structural components, the outer envelope (OE) and protoplasmic cylinder (PC), as well as the leptospire whole cell (WC), was compared in hamsters and dogs. The 50% protective dose for hamsters against death (PD50D) and kidney infection (PD50k) was evaluated for Leptospira interrogans serovars canicola, icterohaemorrhagiae, pomona, and grippotyphosa. All 3 immunogens had similar PD50D values. However, the PD50K values for OE and WC vaccines ranged from 0.05 to 0.80 micrograms (dry weight), whereas the PC vaccines ranged from 7.0 to 14.0 micrograms. Cellular and humoral responses of dogs to serovar canicola WC, OE, and PC vaccines were monitored for 21 weeks. Little difference was observed among the canine humoral responses to the different preparations. Protoplasmic cylinder vaccines sensitized the greater population of lymphoid cells followed by OE and WC. Cross-reactivity was greater in the blastogenic response of lymphoid cell populations than in the humoral response.

Animals↗
PubMedDate not supplied

Isolation and characterization of partially purified leptospiral antigens.

The methanol extract of Leptospira interrogans serovar canicola was purified by precipitation with acetone or acetone and chloroform. The antigenicity of the antigen was not altered by heating or treatment with pepsin and pronase. However the antigenicity was lost when the antigen was treated with periodic acid. Chemical analysis revealed the presence of 40% carbohydrate (22% methylpentose, 28% hexoses), 4% protein, 20% lipid and 2.7% phosphate. The complement fixation test with sera from patients with leptospirosis agreed with the microscopic agglutination reaction.

Agglutination Tests↗
PubMed1975

[Attempts at obtaining a pentavalent vaccine against leptospirosis].

Obtained was a five-valent vaccine (Leptospira pomona, L. canicola, L. grippotyphosa, L. mitis, and L. icterohaemorrhagiae). It was found that storage at 4degreesC does not impair the immunogenicity of the vaccine in the course of one year. Three groups of rabbits were used to test its immunogenicity--group I, injected once, subcutaneously, with 1 cu. cm of the vaccine; group II, injected subcutaneously, twice, at an interval of 7 days, with 1 cu. cm of the vaccine each time; and group III, injected once, muscularly, with 1 cu. cm. The animals were kept under observation for twelve months. It was established that in the case of intramuscular treatment there were highest antibody titers in the blood serum persisting up to the twelfth month.

Animals↗
PubMed1981

Isolation of antigenically active components from leptospiral serovar-specific lipopolysaccharide antigen by alkaline treatment.

The serovar-specific main antigen (TM antigen) of Leptospira interrogans serovar canicola, which as lipopolysaccharide properties, was treated with 0.1 N sodium hydroxide. This treatment degraded the antigen into two major antigenic components, one of high and one of low molecular weight. The component with the lower molecular weight (approximately 4,000 daltons) consisted mainly of carbohydrates, having lost almost all of the fatty acid and protein components of the original antigen. Although the substance lacked immunoprecipitable activity, it continued to show serovar-specific inhibitory potency in a radioimmunoassay system as well as in a microscopic immunoagglutination reaction of the organisms. The antigenic activity of the compound was also reduced by periodate oxidation as was that of the TM antigen. A component with the same chemical and physicochemical properties was also produced by alkaline treatment from a different serotype TM antigen (serovar kremastos Kyoto), but it showed no antigenic activity.

Bacterial Proteins↗
PubMed1984

Determination of specific anti-leptospiral immunoglobulins M and G in sera of experimentally infected dogs by solid-phase enzyme-linked immunosorbent assay.

The development and evaluation of an enzyme-linked immunosorbent assay (ELISA) to detect specific anti-leptospiral IgM and IgG in sera of dogs experimentally infected with Leptospira interrogans serotype canicola are reported. In all dogs specific anti-leptospiral IgM was detected from the second half of the first week after infection, the maximum being attained during the second week. Subsequently the IgM titre gradually decreased. Specific anti-leptospiral IgG was detected later and increased gradually to reach almost the same level as the IgM titre after two to three months. During the initial stage of the infection, when the microscopic agglutination titre was still negative or very low, a high IgM titre was accompanied by a negative or very low IgG titre in every case. After the initial stage a substantial IgG titre was also detectable. It is suggested that the test is suitable for serodiagnostic purposes, particularly for the diagnosis of a current infection in an individual.

Animals↗
PubMed1985

Characterization of monoclonal antibodies to Treponema pallidum.

Thirteen hybrid cell lines which produce mouse monoclonal antibodies to Treponema pallidum, the causative agent of syphilis, have been established. All of the monoclonal antibodies react with T. pallidum, Nichols strain, in ELISA and in immunofluorescence assays, but do not react with normal rabbit testicular tissue in the ELISA. Two of these antibodies were demonstrated to react with the nonpathogenic treponemes T. phagedenis, biotype Reiter, T. refringens (Noguchi strain), T. vincentii, and T. denticola (strains 11 and W), as well as with Borrelia recurrentis, Leptospira interrogans, serogroup Canicola, and the swine pathogen T. hyodysenteriae. The remaining 11 antibodies react with four recently isolated strains of T. pallidum, but with none of the related nonpathogens nor with Borrelia or Leptospira. Thus, our results to date indicate that these monoclonal antibodies may identify antigenic determinants that are specific either for T. pallidum alone or for those treponemes which are pathogenic for humans. The molecular specificities of six of the 13 antibodies were determined by Western blotting. We anticipate potential usefulness of these antibodies in the investigation of the antigenic structure of T. pallidum, the taxonomic study of the pathogenic and nonpathogenic treponemes, and in the diagnosis of syphilis.

Animals↗
PubMed1992

[Detection of leptospiral DNA in the serum of 175 patients with early leptospirosis by polymerase chain reaction].

We have developed a sensitive assay for leptospira, using the polymerase chain reaction (PCR). On the basis of the published nucleotides sequence of 23S rRNA gene from Leptospira interrogans serovar canicola strain Moulton, primers were chosen to produce an amplified fragment of 123 bp. Primer A: 5'GAT CTA ATT CGC TGT AGC AGG3' and primer B: 5'ACT TTC ACC CTC TAT GGT CGG3' Eight different svs. of Leptospira interrogans could all be detected by PCR, but the DNAs from L. biflexa. Leptonema bacteria, virus and human could not produce the specific amplified fragment. The assay detected approximately 10 fg of purified leptospiral DNA and 1 microliter serum of experimental animal. Positive results were obtained from simulated positive samples containing a single organism leptospiral DNA. The diagnostic test (proved by "gold standards": Clinical diagnosis; blood culture and MAT) showed that the sensitivity was 92.00%; the specificity 94.35%; the accuracy 92.54%; the positive predictive value 98.17%; the negative predictive value 78.13%; the positive likelihood ratio 16.25; and the negative likelihood ratio 0.0848. The diagnosis of early leptospirosis by using PCR may become a significant addition to diagnostic means.

Animals↗