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Restriction endonuclease DNA analysis of antigenic variants of leptospires selected by monoclonal antibodies.

The genome of antigenic variant CV (CT3)-1 derived from Leptospira interrogans serovar canicola was compared by cleavage with restriction endonucleases with the parent and serovar bafani, to which the variant was serologically most closely related. No differences were observed between the parent and variant in DNA restriction endonuclease patterns using eight restriction endonucleases. Serovar bafani was different in the patterns from the parent and antigenic variant CV (CT3)-1. The two antigenic variants derived from serovar hebdomadis, HV (H16)-1 and HV (H19)-1 which belonged serologically to serovars jules and hebdomadis, respectively, were compared by restriction endonuclease DNA analysis with the parent and serovar jules. No differences were observed between the parent and variants in DNA restriction endonuclease patterns using the same enzymes. But some differences were observed in DNA restriction endonuclease patterns between HV (H16)-1 and serovar jules. Thus, the antigenic variant selected from the parent by the anti-parent monoclonal antibody and serologically different from the parent, being identified either as a new serovar or as a known one, was found to be similar to the parent by the restriction endonuclease DNA analysis.

Antibodies, Monoclonal↗

Efficacy of macrolides and telithromycin against leptospirosis in a hamster model.

Human studies support the use of beta-lactams and tetracyclines in the treatment of leptospirosis. Additional agents from these and other classes of antimicrobials also have in vitro activity against Leptospira species, though corroborating in vivo data are limited or lacking. We evaluated the therapeutic efficacy of azithromycin, clarithromycin, and telithromycin in a lethal hamster model of leptospirosis using Leptospira interrogans serogroup Canicola serovar Portlandvere. A range of dosages for each antimicrobial was given to the infected animals on days 2 through 7 (5 days) of the 21-day survival model. All untreated control animals survived less than 10 days from infection. Ninety to 100% of doxycycline controls, treated for 5 days with 5 mg/kg of body weight of drug, survived to 21 days. Treatment with azithromycin (daily dose: 6.25, 12.5, 25, 50, 100, or 200 mg/kg) resulted in 100% survival at all evaluated doses. Animals receiving 20 mg/kg or more of clarithromycin (daily dose: 1, 5, 10, 15, 20, 40, 60, or 100 mg/kg) had improved survival. Ninety-eight percent of animals treated with telithromycin (daily dose: 1, 5, 10, 15, 20, or 40 mg/kg) survived. We conclude that all agents tested have demonstrated in vivo efficacy in treating acute leptospirosis. These results provide support for further evaluation of macrolide and ketolide antimicrobial agents in human trials.

Animals↗

Prevalence of canine Lyme disease from an endemic area as determined by serosurvey.

From August 1984 through February 1985, 423 dogs from 43 municipalities in 7 New Jersey counties were evaluated for the presence of antibodies to the Lyme disease spirochete (Borrelia burgdorferi). Of these dogs, 34.7% with no apparent clinical symptoms were serologically reactive (IFA greater than or equal to 1:64); titers in this study ranged from non-reactive to 1:2048. Ninety percent of the dogs surveyed had a current vaccination status to Leptospira interrogans serovars canicola and icterohaemorhagiae. Dogs vaccinated to leptospirosis elicited homologous antibody titers of less than or equal to 1:16 and, therefore, did not interfere with interpretation of antibody levels to B. burgdorferi. Effects of age, degree of outdoor activity, travel history, and location of residence were evaluated. The use of serosurveys of dogs as a tool for Lyme disease surveillance is discussed.

Age Factors↗

Vaccine-associated canine distemper infection in a litter of African hunting dogs (Lycaon pictus).

Four, 57 days old, African hunting dog puppies (Lycaon pictus) from one litter died within three weeks following vaccination with modified-live canine distemper virus (CDV) and killed canine adenovirus type 1, canine parvovirus and Leptospira icterohemorrhagiae and canicola. 18 days post vaccination, the animals developed neurologic disease characterized by episodes of grand mal seizures and circling. Macroscopic, histological and immunohistochemical studies revealed acute systemic CDV infection with acute encephalopathy. Virus isolation attempts using primary dog kidney cells, lung macrophages and Vero cells were negative. Therefore, the question whether the infection was the result of vaccination or natural infection remains open. The benefits and risks regarding the use of modified-live CDV vaccines and killed canine distemper vaccines in exotic carnivores are briefly discussed.

Animals↗

Serological survey for selected diseases in the endangered San Joaquin kit fox (Vulpes macrotis mutica).

Blood from endangered San Joaquin kit foxes (Vulpes macrotis mutica) inhabiting the Elk Hills Naval Petroleum Reserve, Kern County, and the Elkhorn Plain, San Luis Obispo County, California, was collected in 1981, 1982 and 1984 and sera were tested for antibodies against 10 selected pathogens. Proportions of kit fox sera containing antibodies against pathogens were: canine parvovirus, 100% in 1981-1982 and 67% in 1984; infectious canine hepatitis virus, 6% in 1981-1982 and 21% in 1984; canine distemper virus, none in 1981-1982 and 14% in 1984; Francisella tularensis, 8% in 1981-1982 and 31% in 1984; Brucella abortus, 8% in 1981-1982 and 3% in 1984; Brucella canis, 14% in 1981-1982 and none in 1984; Toxoplasma gondii, 6% in 1981-1982; Coccidioides immitis, 3% in 1981-1982; and Yersinia pestis and Leptospira interrogans serotypes canicola, grippotyphosa, hardjo, icterohaemorrhagiae, and pomona, none in 1981-1982. Although antibodies against selected pathogens were present, no clinical indications of disease were observed in these fox populations.

Animals↗

Serologic survey for selected microbial pathogens in bison from Kansas.

A serologic survey was conducted on an American bison (Bison bison) herd in Kansas for antibodies against Brucella spp., Leptospira interrogans serovar canicola, pomona, grippotyphosa, icterohaemorrhagiae, and hardjo, Anaplasma spp., bluetongue virus, infectious bovine rhinotracheitis virus and bovine viral diarrhea virus. There was an increase in prevalence of bluetongue antibodies from 38% in 1987 to 100% in 1989 in animals greater than or equal to 24-mo-old. Prevalences of antibodies against the other livestock pathogens were either negative or at levels associated with previous vaccination.

Anaplasmosis↗

Serological survey for diseases in free-ranging coyotes (Canis latrans) in Yellowstone National Park, Wyoming.

From October 1989 to June 1993, we captured and sampled 110 coyotes (Canis latrans) for various diseases in Yellowstone National Park, Wyoming (USA). Prevalence of antibodies against canine parvovirus (CPV) was 100% for adults (> 24 months old), 100% for yearlings (12 to 24 months old), and 100% for old pups (4 to 12 months old); 0% of the young pups (< 3 months old) had antibodies against CPV. Presence of antibodies against canine distemper virus (CDV) was associated with the age of the coyote, with 88%, 54%, 23%, and 0% prevalence among adults, yearlings, old pups, and young pups, respectively. Prevalence of CDV antibodies declined over time from 100% in 1989 to 33% in 1992. The prevalence of canine infectious hepatitis (ICH) virus antibodies was 97%, 82%, 54%, and 33%, for adults, yearlings, old pups, and young pups, respectively. The percentage of coyotes with ICH virus antibodies also declined over time from a high of 100% in 1989 to 31% in 1992, and 42% in 1993. Prevalence of antibodies against Yersinia pestis was 86%, 33%, 80%, and 7%, for adults, yearlings, old pups, and young pups, respectively, and changed over time from 57% in 1991 to 0% in 1993. The prevalence of antibodies against Francisella tularensis was 21%, 17%, 10%, and 20%, for adults, yearlings, old pups, and young pups, respectively. No coyotes had serologic evidence of exposure to brucellosis, either Brucella abortus or Brucella canis. No coyotes were seropositive to Leptospira interrogans (serovars canicola, hardjo, and icterohemorrhagiae). Prevalence of antibodies against L. interrogans serovar pomona was 7%, 0%, 0%, and 9%, for adults, yearlings, old pups, and young pups, respectively. Antibodies against L. interrogans serovar grippotyphosa were present in 17% of adults and 0% of yearlings, old pups, and young pups. Many infectious canine pathogens (CPV, CDV, ICH virus) are prevalent in coyotes in Yellowstone National Park, with CPV influencing coyote pup survival during the first 3 months of life; eight of 21 transmitted pups died of CPV infection in 1992. The potential impact of these canine pathogens on wolves (C. lupus) reintroduced to Yellowstone National Park remains to be documented.

Adenoviruses, Canine↗

C-reactive protein in dogs.

Serum concentrations of C-reactive protein (CRP) in dogs with various diseases or undergoing various procedures were measured by specific immunoassay. In 20 healthy dogs from various sources, values were all less than 5 mg/L, but in 22 healthy dogs from a single source, values ranged from less than 5 mg/L in 14 dogs and from 8 to 67 mg/L in 8 dogs. Increased concentrations of serum CRP were attained 24 hours after injection of casein (n = 9; median 188 mg/L), ovariohysterectomy (n = 11; median, 144 mg/L), or elective, nonacute orthopedic surgery (n = 10; median, 83 mg/L). After inoculation of Leptospira interrogans serovar canicola (n = 5), the behavior of serum CRP as an acute-phase reactant provided a sensitive and precise objective reflection of in vivo response. The CRP concentration in random single-serum samples from 73 dogs with other inflammatory and noninflammatory disorders ranged from normal (less than 5 mg/L) to 246 mg/L and generally correlated with the extent and activity of disease.

Animals↗

Development of a modified live, canine origin parvovirus vaccine.

A modified live, canine origin parvovirus vaccine was tested for safety, efficacy, and clinical performance. The vaccine protected dogs from challenge of immunity with canine parvovirus (CPV) that caused clinical illness in all nonvaccinated dogs. Vaccinates all developed CPV serum neutralization antibody titers, with a mean value of 1,664. Challenge virus was not isolated from vaccinates, but feces from nonvaccinated dogs were CPV-positive for up to 4 days following challenge. In a pathogenicity test, dogs inoculated orally with 10 times the label dose remained clinically normal. In a reversion-to-virulence test, the vaccine strain remained nonpathogenic through 6 passages in seronegative test dogs. An immunologic interference test demonstrated that test dogs developed antibodies for all antigens in a combined canine distemper virus-adenovirus 2-parainfluenza virus-parvovirus vaccine and a Leptospira interrogans serovars canicola and icterohaemorrhagiae bacterin. A total of 1,796 doses of the multivalent preparation was administered in a field study, with veterinary practitioners reporting 36 local and 5 generalized reactions.

Animals↗

Comparative study between microscopic agglutination and complement fixation tests in experimental canine leptospirosis.

In order to evaluate the antibody response to pathologic leptospira from both serovars icterohaemorrhagiae and canicola, dogs were experimentally inoculated with these leptospiras. Each of the serovar were inoculated in 15 dogs and the antibody response in the acute phase of infected was followed by microscopic agglutination and complement fixation tests. Complement fixing antibodies were detected in both groups at the 4th day post infection and agglutinins appeared in circulation at the 7th day post infection when homologous antigens were used. On the 63rd day post infection with serovar canicola and on 56th day post infection with serovar icterohaemorrhagiae, complement fixing antibodies could no longer be detected but, in both experimental groups, specific agglutinins were presented in significant titers.

Agglutination Tests↗

Lipid metabolic changes in experimentally induced leptospiral infection with serovars australis, canicola and icterohaemorrhagiae.

Changes in lipid fractions were evaluated in young guinea pigs when infected with 1 ml of 7 day old live cultures of leptospira interrogans serovars australis, canicola and icterohaemorrhagiae. Statistically significant elevation in triglycerides, very low density lipoprotein and phospholipid and a significant reduction in high density lipoprotein (HDL) in all the groups was observed. Cholesterol and low density lipoprotein showed ascending trend in icterohaemorrhagiae group, whereas they were normal in other groups. The results suggest that increase in triglycerides, phospholipid and decrease in HDL in a suspected case of leptospirosis may be considered as markers.

Animals↗

A serosurvey of leptospirosis in Connecticut peridomestic wildlife.

Recently, leptospirosis has gained attention as a re-emerging infection in domestic dogs in the northeastern United States. In order to gain insight into the epizootiology of leptospirosis in this region, 109 small wild mammals (31 raccoons (Procyon lotor), 30 skunks (Mephitis mephitis), 28 opossums (Didelphis virginiana), and 20 gray squirrels (Sciurus carolinensis)) collected between February 27 and September 17, 2001 were tested for serologic evidence of exposure to five common Leptospira serovars (serovars pomona, icterohemorrhagiae, canicola, hardjo, grippotyphosa). Evidence of exposure to leptospirosis was detected in 36% of raccoons tested; icterohemorrhagiae was the predominant reactive serovar in these animals. Sera from 13% of skunks showed evidence of exposure to serovar grippotyphosa. One squirrel exhibited high antibody titers to serovars grippotyphosa and canicola. All 28 opossums examined tested negative to the five Leptospira serovars. Results from this serosurvey suggest that common peridomestic wildlife species should be considered as potential sources of leptospirosis to dogs and humans in Connecticut. Additional investigation is warranted to clarify their role in the epidemiology of this zoonotic disease in the northeastern United States.

Agglutination Tests↗

Evaluation of a rapid slide agglutination test for the diagnosis of acute canine leptospirosis.

The standard method for the diagnostic of leptospirosis is the microscopic agglutination test (MAT). Nevertheless, it is time-consuming and presents risk of infection for laboratory personal. Several attempts to simplify and reduce the time, skill and expense required for serological testing were made. Rapid slide agglutination tests (RSAT) have been used nowadays as a reliable screening test for the detection of acute and recent infections by many laboratories. The purpose of that study was to evaluate a new antigen preparation for the rapid diagnoses of acute canine leptospirosis and to compare it with MAT results in clinically suspect serum samples. Two hundred and thirteen serum samples from dogs of both sexes and different ages were tested and 141 (66.2%) animals were considered as positive by MAT. The most frequent serovar was icterohaemorrhagiae, followed by canicola. RSAT results were very similar to those observed on MAT, with 139 (65.3%) reactive animals. Correlation between RSAT and MAT was positive (0.82) and significant (p < 0.01). Concordance of results was of 93.4% (199/ 213) of the animals correctly diagnosed by RSAT. Considering MAT as the standard test, sensitivity of RSAT was calculated on 94.3% (133/141) and specificity on 91.7% (66/72).

Agglutination Tests↗

[Outbreak of human leptospirosis after a flood in Reconquista, Santa Fe, 1998].

Since April of 1998 a high number of leptospirosis cases were detected, coming from the area of Reconquista Central Hospital in Santa Fe province. Since January of that year a notable increase in rainfall and river levels was observed causing inundation. As screening test, macroscopic agglutination (MAT) using 10 serotypes of L. interrogans. Among the 122 patients studied 71 were TR positive and 52 were also ELISA positive, leptospirosis diagnosis being confirmed in 40 of them. Five infecting serogroups were identified: Icterohaemorrhagiae (7/40), Ballum (5/40), Sejroe (3/40), Pomona (3/40) and Canicola (2/40). In the remaining cases (20/40), co-agglutinins were found at the same titer against two or more serotypes of leptospires. Infection prevalence was higher in men and productive age (21 to 40 years). The clinical symptoms more frequently observed were headache, fever and myalgias. All cases occurred after the rains and in the period when the area was flooded. Their clinical presentation, time distribution, geographical localization and high frequency of contact with the risk factor inundation could indicate that, independently of search activities, there was an outbreak.

Adolescent↗

Serological survey of pre-weaned New Zealand fur seals (Arctocephalus forsteri) for brucellosis and leptospirosis.

AIM: To conduct a longitudinal serological survey for evidence of Brucella spp and Leptospira spp infection of pre-weaned New Zealand fur seals in a colony on the Otago Peninsula. METHODS: Seal pups were repeatedly captured on a monthly basis from February through to July 2001. Pups were tagged at first capture and a blood sample was taken at each capture event. A total of 163 sera were collected from 118 seal pups. Where sufficient volume was collected, the sera were tested for leptospirosis using the microscopic agglutination test (MAT), and for brucellosis using the competitive enzyme-linked immunosorbent assay (ELISA) for Brucella abortus. RESULTS: None of 128 sera from 101 seals tested positive to the ELISA for B. abortus. All tests for Leptospira interrogans serovars Grippotyphosa, Copenhageni, Bratislava and Leptospira borgpetersenii serovar Ballum were negative at a cut-off of <1/100 dilution. Positive or suspicious titres were found to L. interrogans serovars Canicola and Pomona and L. borgpetersenii serovar Hardjo. The highest titres (12,800) were found to serovar Pomona. The titre to serovar Pomona in one seal rose from <1/50 in March to 12,800 in April and was <1/50 when re-sampled in July. The titre to serovar Pomona in another seal dropped from 12,800 in May to <1/50 in June. These seals also had titres to serovar Hardjo, which rose or fell in the same manner. All suspicious or positive titres occurred in late April and early May, when the pups were approximately 4-5 months old. In June and July, all seals tested were negative. CONCLUSIONS: There was no serological evidence of Brucella infection in the pre-weaned fur seals at the colony. Positive titres to serovars Pomona, Hardjo, or Canicola suggest that a Leptospira species was present at the colony, however isolation or visualisation of the organism is required to confirm this. Care should be exercised when handling New Zealand fur seals to prevent human infection or inadvertent transfer of leptospirosis to another marine mammal species.

Age Factors↗