Julio Cesar de Freitas

Isolation and characterization of Streptococcus spp. group B in Nile tilapias (Oreochromis niloticus) reared in hapas nets and earth nurseries in the northern region of Parana State, Brazil

The objective of this study was to isolate and characterize Streptococcus spp. in Nile tilapias (Oreochromis niloticus) reared in net-pens and earth nurseries. Eight intensive tilapia-rearing farms were investigated in north Parana, Brazil from April 1st 2001 to April 30th 2002. The fish were reared in a system of hapas nets on four farms and in earth nurseries on other four farms. A total of 370 samples were analyzed of material collected from 120 fish (brain, liver, kidney, skin scrapes, ascites liquid and eye) that were sown on BHI agar (Brain Heart Infusion) supplemented with 1% yeast extract and sheep blood. Streptococcus spp. was isolated in 36 of the samples (18 brain, eight liver, eight kidney and two ascites liquid) from 25 fish. Streptococci were isolated in both systems, almost in the same proportion. First the streptococci were characterized by the catalase and esculin test, growth in methylene blue and sodium chloride at 6.5%. They were classified in groups by the Slidex Strepto-Kit (BioMerieux, France). The phenotypic characteristics were determined by the Api 20 Strep microtest system (BioMerieux, France). The 36 Streptococcus spp. samples did not present hemolysis and were classified as Lancefield group B. Further 16 samples were identified as Streptococcus agalactiae and 20 were not identified by the Api 20 Strep, but presented the same biochemical profile described for the reference strain of Streptococcus difficile (ND-2-22).

Enterotoxigenicity of Staphylococcus intermedius of canine origin

Seventy-three staphylococcal strains isolated from pyrodermatitis in dogs were classified as Staphylococcus intermedius (52 strains) or Staphylococcus aureus (21 strains) on the basis of acetoin formation, anaerobic mannitol fermentation, aerobic maltose fermentation, pigmentation, coagulation of human plasma, and reaction on crystal violet agar. Enterotoxin was produced by 13 of the 52 S. intermedius strains and 6 of the S. aureus strains. The highest percentage of enterotoxigenic strains produced enterotoxins C (6 strains), D (7 strains), and E (6 strains). Four strains produced the toxic shock syndrome toxin-1. There was little difference in the antibiotic susceptibility between the enterotoxigenic and non-enterotoxigenic strains.

Seroepidemiology and occupational and environmental variables for leptospirosis, brucellosis and toxoplasmosis in slaughterhouse workers in the Paraná State, Brazil

Leptospirosis, brucellosis and toxoplasmosis are widely-distributed zoonosis, being the man an accidental participant of their epidemiological chains. The aim of this paper was to make a seroepidemiological report and identify occupational and environmental variables related to these illnesses in 150 workers in a slaughterhouse in the Northern region of Paraná. For the diagnosis of leptospirosis a microscopical seroagglutination test was applied; for brucellosis, the tamponated acidified antigen test and the 2-mercaptoetanol tests were used, and for toxoplasmosis the indirect immunofluorescence reaction test. For each employee an epidemiological survey was filled, which investigated occupational and environmental variables which could be associated with these infections. Positive results for leptospirosis were found in 4.00% of the samples, for brucellosis in 0.66% of samples and toxoplasmosis in 70.00%. From the three diseases researched, only the results for leptospirosis suggest occupational infection.

Isolation of Leptospira spp from dogs, bovine and swine naturally infected

Leptospira isolation allows definitive diagnosis of the infection. Contamination by microorganisms is one of the inconveniences of the culture. The objective of this study was to describe the isolation of Leptospira from dogs, bovine and swine naturally infected. Urine samples from 14 dogs and three bovines, and kidney, liver, ovary, and uterus body samples from 36 slaughtered sows with unknown health history, were used. The urine and organ samples were cultured in culture medium. Modified Ellinghausen-McCullough-Johnson-Harris medium (EMJH) culture medium was used with addition of 5-fluorouracil, chloramphenicol, vancomycin, nalidixic acid and neomycin. Incubation was performed at 28oC for 24 hours, followed by subculture in modified EMJH without antibiotics. The cultures were assessed weekly for up to eight weeks for the dog and swine samples and for up to 16 weeks for the bovine samples. With this methodology, Leptospira spp could be isolated from 11 dogs, two bovines and liver fragments from two sows.

Evaluation on the Pathogenesis of Streptococcus agalactiae in Nile Tilapia (Oreochromis niloticus)

The pathogenesis of a Streptococcus agalactiae was evaluated in a three-period experiment. Two groups of 40 fishes were intraperitoneally (i.p.) challenged in each experimental period with different infective doses of the pathogen. Doses varied from 1.0 x 106 to 1.5 x 108 CFU/fish. One group of 40 tilapia i.p. injected with tryptic soy broth (TSB) was used as a control group in each period. Mortalities varied from 67.5% in group 8 (infective dose 1.0 x 106 CFU/fish) to 90.0% in group 1 (infective dose 1.5 x 108 CFU/fish). Significant differences in mortalities were found only between group 8 and each of the other groups, except group 5 (infective dose 6.0 x 106 CFU/fish; mortality 75.0%). The highest mortality coefficients were observed in days 1-2 after inoculation (accumulated mortality 44.4%), and a second peak of mortality occurred at days 6-7. Challenged fishes from all the groups showed alterations in behaviour and similar clinical signs. These were anorexia, lethargy, erratic swimming, exophthalmia and ascites. Macroscopically, skin hemorrhage, splenomegaly, hepatomegaly with organ paleness and visceral adherences were observed. S. agalactiae was re-isolated from all the fishes from the experimental groups submitted to bacteriological examination. The illness observed in tilapia naturally infected with S. agalactiae was experimentally reproduced in this study, and the clinical signs produced were similar to those reported from the natural infections.

ISOLATION OF LEPTOSPIRA SEROVARS CANICOLA AND COPENHAGENI FROM CATTLE URINE IN THE STATE OF PARANÁ, BRAZIL

In 2001, 698 urine samples were randomly collected from cattle at a slaughterhouse in the State of Parana, Brazil. Direct examination using dark field microscopy was carried out immediately after collection. Five putative positive samples were cultured in modified EMJH medium, yielding two positive cultures (LO-14 and LO-10). Typing with monoclonal antibodies revealed that the two isolates were similar to Canicola (LO-14) and Copenhageni (LO-10). Microscopic agglutination test results show that Hardjo is the most common serovar in cattle in Brazil. Rats and dogs are the common maintenance hosts of serovars Copenhageni and Canicola. The excretion of highly pathogenic serovars such as Copenhageni and Canicola by cattle can represent an increasing risk for severe leptospirosis is large populations, mainly living in rural areas.

Leptospirosis in slaughtered sows: serological and histopathological investigation

Swine leptospirosis is recognized world wide as an important cause of reproductive failure. Serum and histo-pathological examinations have been constantly used to diagnose this disease. This study was carried out on slaughtered sows from Northern Parana State to compare serum findings with kidney, liver, ovary and uterus histo-pathological examinations. The microscopic serum-agglutination carried out on 36 animals detected 24 with titles > 100 and 12 negative ( 100 serological titles and five from the negative group. The results showed that all the 16 animals had indicative lesions in the liver or in one of the kidneys. There were no lesions in the ovaries or uterus. Microscopic serum-agglutination title readings between 20 and 80 were detected in animals of the group considered negative with title < 100.

Molecular characterization, serotyping, and antibiotic susceptibility profile of Leptospira interrogans serovar Copenhageni isolates from Brazil

Leptospira interrogans serogroup Icterohaemorrhagiae is the major serogroup infecting humans worldwide, and rodents and dogs are the most significant transmission sources in urban environments. Knowledge of the prevalent serovars and their maintenance hosts is essential to understand the epidemiology of leptospirosis. In this study, 20 Leptospira isolates were evaluated by pulsed-field gel electrophoresis (PFGE), variable number tandem-repeat analysis (VNTR), serotyping, and determination of antimicrobial resistance profile. Isolates, originated from bovine, canine, human, and rodent sources, were characterized by microscopic agglutination test with polyclonal and monoclonal antibodies and were identified as L. interrogans serogroup Icterohaemorrhagiae serovar Copenhageni. MICs of antimicrobials often used in veterinary medicine were determined by broth microdilution test. Most of tested antibiotics were effective against isolates, including penicillin, ampicillin, and ceftiofur. Higher MIC variability was observed for fluoroquinolones and neomycin; all isolates were resistant to trimethoprim/sulfamethoxazole and sulphadimethoxine. Isolates were genotyped by PFGE and VNTR; both techniques were unable to discriminate between serovars Copenhageni and Icterohaemorrhagiae, as expected. PFGE clustered all isolates in 1 pulsotype, indicating that these serovars can be transmitted between species and that bovine, rodent, and dogs can maintain them in the environment endangering the human population.

Occurrence of Ehrlichia canis and Anaplasma platys in household dogs from northern Parana

Canine monocytic ehrlichiosis caused primarily by Ehrlichia canis and canine thrombocytic anaplasmosis induced by Anaplasma platys are important emerging zoonotic tick-borne diseases of dogs. There is evidence that these pathogens can also affect humans. This study evaluated the presence of E. canis and A. platys in blood samples collected from 256 domiciled dogs in the municipality of Jataizinho, located in north region of the State of Parana, Brazil, by PCR assay. The occurrence of E. canis and A. platys was 16.4% (42/256) and 19.4% (49/256), respectively; while 5.47% (14/256) of the dogs evaluated were co-infected by these two organisms. The presence of E. canis and A. platys was not significantly associated with the variables evaluated (sex, age, outdoor access, and presence of ticks during blood collection). Infection of dogs by E. canis was associated with anemia and thrombocytopenia, while infection induced by A. platys was related only to thrombocytopenia. Canine monocytic ehrlichiosis and canine thrombocytic anaplasmosis should be included in the differential diagnoses when these hematological alterations are observed during routine laboratory evaluation of dogs.

Fatores de risco associados a soropositividade para leptospirose em matrizes suínas

Leptospirosis is a widespread zoonosis of great social and economic importance for swine production sistem. Two hundred and ninety-eight sows from farms in the north of Parana state were studied to individualize management practices or environmental condition that are related to risk of leptospirosis infection. All the animals were submitted to the microscopic seroaglutination test and an epidemiological questionnaire was filled out for each one to investigate variables that could be associated with this infection. The results were submitted to the c2 test, followed by multivariate analysis. Antibody titers of >100 were detected in 132 sows (44.3%) most frequently against icterohaemorrhagiae serovar (98.5%). The final model of multivariate analysis showed the risk factors to be flooded areas near the installation (OR=1.73; 1.04<IC95%<2.80), canal type water source (OR=1.58; 0.98<IC95%<2.55) and absence of cleaning of the water reservoirs (OR=2.25; 0.79<IC95%<6.43).