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PubMed1967

Growth of pathogenic Leptospira in chemically defined media.

A protein-free chemically defined medium for cultivation of pathogenic Leptospira was developed. The medium permitted continued serial subculturing of 9 serogroups (52 strains) of the 12 serogroups (61 strains) tested. Growth was initiated from small inocula, and the growth rate and maximal cell yields were similar to those on serum-containing media. The nutritional requirements of serogroups L. canicola, L. pomona, and L. grippotyphosa were studied in a basal medium composed of inorganic salts, a fatty acid, vitamin B(12), and thiamine. All strains tested utilized ammonium chloride as the sole nitrogen source. A fatty acid, vitamin B(12), and ferrous ions were essential. Growth was stimulated by thiamine, potassium, and calcium ions.

Culture Media↗
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↗
PubMed1988

[Characteristics of the interaction of Leptospira with the host organism in the infectious process in golden hamsters].

The comparative evaluation of the interaction of L. icterohaemorrhagiae strain P, L. canicola strain CL and L. hebdomadis strain 650 with golden hamster liver and kidney cells is presented. Three variants of the course of Leptospira infection have been distinguished: (1) the hepato-renal (icteric) variant, caused by the adhesion of leptospires to liver cells with the colonization of their surface and the disaggregation of liver-cell complexes and by the accumulation of leptospires in the kidney interstice; as a consequence, parenchymatous hepatitis and nephroso-nephritis develop, which lead to the death of animals; (2) the renal (anicteric) variant, characterized by the absence of the infective agent and lesions in the liver, by adhesion of leptospires to and their colonization of the nephrothelium of the proximal convoluted tubules of the kidneys; in this case some of the animals die because of renal insufficiency and shock, while in the surviving animals prolonged carrier state develops; (3) the intermediate variant, characterized by the initial process of leptospiral adhesion and colonization in the liver and its subsequent progress in the kidneys.

Animals↗
PubMed1987

Genus-specific antigens in Leptospira revealed by immunoblotting.

Immunoblotting of leptospiral sonicates with heterologous rabbit antisera revealed a distinct cross-reactive pattern which differed with respect to the pathogenic and non-pathogenic leptospiral serovars, and that all serovars tested from Leptospira interrogans, L. biflexa and L. illini contained a common 35 kilodalton (Kd) band. A leptospiral genus-specific antigen preparation produced by ethanol fractionation of L. biflexa serovar patoc reacted by enzyme immunoassay (EIA) with all heterologous serovars tested. Further purification using Sephacryl S-300 gel filtration revealed one major cross-reactive peak and several homologous peaks detectable by EIA. Gel electrophoresis of this peak revealed 3 major protein bands of 35, 34 and 29 Kd by Coomassie blue staining. This peak was further fractionated by high pressure liquid chromatography (HPLC), yielding 7 fractions, one of which cross-reacted. Rabbit antisera to this S-300/HPLC fraction reacted with all serovars tested. Immunoblotting revealed 2 distinct groups of cross-reactive antigens, a 33-35 Kd group that was proteinase K sensitive but not reduced by periodate oxidation, and a 14.4-26.5 Kd group whose activity was reduced by periodate but not proteinase K, indicating the presence of both protein and carbohydrate genus antigens. Immunoblotting L. interrogans serovar pomona flagella with S-300/HPLC antiserum suggested that the 35 Kd band found in all serovars tested was a flagellar component.

Animals↗
PubMed1984

Isolation of an antigenic oligosaccharide fraction from Leptospira interrogans serovar canicola with a monoclonal antibody.

An oligosaccharide fraction containing the antigenic determinant of lipopolysaccharide antigen (TM antigen) from Leptospira interrogans serovar canicola, recognized by a monoclonal antibody (CT3) which agglutinates serovars canicola and broomi, was isolated by formic acid and successive sulphuric acid hydrolyses. Separation of the antigenic compounds was done by Bio-Gel P-2 and Sephadex G-25 gel filtration, and high-performance liquid chromatography with two different columns. The fraction finally obtained was a mixture of two oligosaccharides, both of which migrated as a single spot having a slightly higher mobility than an authentic tetrasaccharide (stachyose) on thin layer chromatography. The fraction contained rhamnose, arabinose and two major and two minor unknown sugars which were shown to be N- or O-acetylated and/or O-methylated sugars by nuclear magnetic resonance. The fraction inhibited the binding of CT3 antibody with TM antigen in enzyme-linked immunosorbent assay and microscopic agglutination of serovar canicola with the antibody. The inhibitory activity was destroyed by periodate oxidation or mild alkaline treatment, but was resistant to sodium borohydride reduction.

Antibodies, Monoclonal↗
PubMed1990

Linkage of ribosomal RNA genes in Leptospira.

We determined the linkage of 16S, 23S, and 5S rRNA genes in several strains of Leptospira and Leptonema by DNA-DNA hybridization. Almost all the hybridizations in all leptospires used in these experiments gave two radioactive bands and the results strongly suggest that the number of the 16S and the 23S rRNA genes in those strains is two, respectively. In contrast with the larger rRNAs, the number of 5S rRNA gene was different. In the strains of leptospires, L. biflexa, which were non-parasitic, there are two genes for 5S rRNA, whereas only one gene for 5S rRNA is carried in L. interrogans, which were originally isolated as parasitic. Southern hybridization experiments suggest that those rRNA genes are interspersed on the leptospiral chromosome.

DNA, Bacterial↗
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↗
PubMed1979

Axial filament involvement in the motility of Leptospira interrogans.

Motility mutants of Leptospira interrogans serovar illini were isolated and analyzed by dark-field and electron microscopy. Mutants were obtained by screening for small colonies after nitrosoguanidine treatment. One class of mutants did not have hook- or spiral-shaped ends. In addition, the axial filaments from these mutants were not coiled. An analysis of revertants of two of the mutants in this class indicated that the mutations were pleiotropic with respect to motility, hook- and spiral-shaped ends, and axial filament coiling. We conclude that the axial filaments and the hook- and spiral-shaped ends are involved in L. interrogans motility.

Cell Wall↗
PubMed1979

Radioimmunoassay system using a serovar-specific lipopolysaccharide antigen of Leptospira.

The serovar-specific main (TM) antigen of Leptospira interrogans serovar kremastos strain Kyoto was labeled with sodium boro[3H]hydride after 1 h of oxidation with periodate. When 50 ng of the labeled compound was employed, 50% of the compound was bound to the antibodies contained in 7.5 x 10(-2) nl of anti-kremastos Kyoto serum. Under this condition, only 9 ng of the homologous TM antigen was required for 50% inhibition of serovar kremastos Kyoto 3H-TM antigen-anti-kremastos Kyoto serum binding, whereas 5,000 times as much TM antigen of serovar hebdomadis, which belongs to the same Hebdomadis serogroup as serovar kremastos, was required for the same inhibition. The TM antigens from serovars pomona, icterohaemorrhagiae, and copenhageni, which belong to different serogroups, showed no inhibition in an amount of up to 3 x 10(3), 2 x 10(5), and 2 x 10(5) ng, respectively. In the inhibition study using the serovar hebdomadis TM antigen, inhibition values slightly fluctuated due to the antigen's insolubility. When the TM antigens were solubilized with 0.1% sodium taurodeoxycholate, the fluctuation in inhibition values was minimized, and the cross-reactivity of the serovar hebdomadis TM antigen with anti-kremastos Kyoto serum was diminished, while the inhibitory activity of the serovar kremastos Kyoto TM antigen was enhanced.

Antibodies, Bacterial↗
PubMed1980

Isolation of Leptospira biflexa from commercially prepared deionized water labeled "sterile for tissue culture".

Leptospira biflexa were isolated from urine cultures of two patients with clinical and laboratory findings compatible with leptospirosis. Neither patient had detectable leptospiral agglutinins. The source of the L. biflexa was water labeled " "sterile for tissue culture" purchased from M. A. Bioproducts, formerly Microbological Associates. M. A. Bioproducts water is sterilized by filtration through a 0.22-mum-pore size membrane filter. Leptospiral contamination can occur in products sterilized by filtration. Heat sterilization of water eliminates this possibility.

Adult↗
PubMed1981

Surface colonies of Leptospira interrogans.

Observations of colonial growth of Leptospira interrogans serovar pomona, BJ, on soft-agar (1% agar) plates revealed the presence of both surface and subsurface colonies. The organisms from both types of colonies appeared to be similar in motility, morphology, antigenic composition, and pathogenicity. Passage in vivo tended to produce a higher incidence of surface colonies, whereas passage in vitro tended to produce a higher incidence of subsurface colonies, suggesting the importance of environmental factors in determining the type of colony produced.

Antigens, Bacterial↗
PubMed1975

Failure of transformation in leptospira with two genetic markers of streptomycin resistance and the ability to grow in a synthetic medium.

Transformability of Leptospira was investigated with the two genetic markers, streptomycin (SM) resistance and the ability to grow in SHENBERG's synthetic medium. DNA was extracted from the two mutants of L. icterohaemorrhagiae strain Shibaura, which were SM-resistant and able to grow in SHENBERG's medium. Recipients were 5 strains of L. icterohaemorrhagiae and 1 strain of L. copenhageni. Studies taking careful consideration of the growth phase of the recipient cells, transformation medium, and incubation temperature did not produce the appearance of transformant. Transformation of L. kremastos with SM resistance also showed a negative result.

Culture Media↗
PubMed1964

THE USE OF LEPTOSPIRA BIFLEXA PATOC ANTIGEN IN FIELD INVESTIGATIONS OF LEPTOSPIROSIS.

Hitherto the laboriousness of serological procedures for the laboratory diagnosis of leptospirosis has somewhat limited their usefulness. The authors of this paper report on a simple and sensitive genus-specific serological test for this disease that is within the capabilities of ordinary diagnostic laboratories. They describe the organization and results of a trial carried out in Romania in 1962 of a complement-fixation (CF) test in leptospirosis in which an antigen derived from the Patoc I strain of Leptospira biflexa is used. Human sera examined with this test in nine field laboratories were re-examined at the Cantacuzino Institute in Bucharest with both CF and agglutination tests.Of 152 sera found CF-positive in the field laboratories, 138 were found positive by the agglutination test in Bucharest-representing 90% agreement. There was 88% agreement between the field laboratory and central laboratory results in the CF test.The test makes possible the early detection of human leptospirosis and gives positive reactions with sera from leptospirosis patients irrespective of the causative serotype.

Agglutination Tests↗
PubMed1986

[Effect of the viable cell count on the growth parameters of Leptospira cultures].

The influence of the amount of live cells on the growth characteristics of 41 Leptospira pathogenic strains belonging to 4 serogroups at different stages of growth has been studied. The study has revealed that under the conditions of batch cultivation the maximum concentration of pathogenic leptospires in the inoculum decreases the duration of the lag phase and determines the highest specific growth rate characterizing the individual features of leptospires in the serogroups under study.

Cell Division↗
PubMed1983

Species- and genus-specific antigens in Leptospira, revealed by monoclonal antibodies and enzyme immunoassay.

Mouse antisera to Leptospira interrogans serovar pomona or L. biflexa serovar patoc agglutinated homologous leptospires but not leptospires of serovars from any heterologous serogroup tested. When tested by enzyme immunoassay (EIA), pomona and patoc antisera reacted with all serovars tested from L. interrogans, L. biflexa or L. illini species. A non-agglutinating hybridoma-derived monoclonal antibody reacted by EIA only with serovars of the pathogenic L. interrogans species but not with L. biflexa nor L. illini. The results demonstrated the presence of both genus and species specific non-agglutinating leptospiral antigens which could be detected only by EIA.

Antibodies, Monoclonal↗
PubMed1982

[Nucleotide composition and homology of the DNA of parasitic Leptospira].

The study of DNA in parasitic leptospirae of different serovariants by the method of molecular hybridization has revealed two groups differing in their homology level. The homology of nucleic acids has been found to correlate with serological properties. The serovariants ensuring cross immunity are characterized by the high level of homology of their DNA.

Cross Reactions↗
PubMed1997

[Leptospira interrogans in a canine population of Greater Buenos Aires: variables associated with seropositivity].

We determined the seroprevalence of leptospirosis in a suburban canine population for the purpose of analyzing the association between different individual and environmental variables and seropositivity for leptospirosis. The study, which was cross-sectional, was performed in July 1992 in a neighborhood of Greater Buenos Aires with approximately 9,500 inhabitants and a canine population of around 2,000 animals. We studied a random sample of 223 dogs and obtained a blood sample from each. Each animal's epidemiologic history was obtained by interviewing the housewife. Serologic measurements were performed by the microagglutination technique with the use of 10 different serotypes of Leptospira interrogans. Of the 223 dogs that were tested, 57% showed seropositivity; 82% of the positive sera coagglutinated with two or more serotypes. The most frequently detected serotypes were canicola and pyrogenes. Seroprevalence in females was less common than in males (P < 0.05) and in puppies less than 1 year old it was less common than in older animals (P < 0.01). Street behavior in the dog and the presence of stagnant water in front of the owner's dwelling were the most important of the risk factors examined. The associations between seropositivity on the one hand and contact with trash deposits, hunting behavior and the presence of rodents inside the dwelling on the other were not statistically significant. Different control measures are discussed.

Age Factors↗