[Investigation of Brucella canis seropositivity by in-house slide agglutination test antigen in healthy blood donors].
Canine brucellosis which is due to Brucella canis, is transmitted to man by infected dogs or their secretions. The symptoms of canine brucellosis are similar to the symptoms of brucellosis caused by other Brucella species and endocarditis or meningitis may develop in untreated cases. There is limited data regarding B.canis infections in man and the current status of the disease is insufficiently evaluated in our country.
Serological diagnosis of brucellosis is classically based on standard slide and tube agglutination tests. However, the antigens used in these tests detect antibodies that develop against species (B.melitensis, B.abortus, B.suis) with “smooth” lipopolysaccharides in their cell wall. B.canis has “rough” lipopolysaccharide in its cell wall and thus these classical tests can not detect antibodies against B.canis.
Besides there is no commercial slide agglutination test which uses B.canis antigens. The aim of this study was to investigate the B.canis seropositivity by slide agglutination test (SAT), using homemade B.canis antigen, in healthy subjects and to determine the prevalence of B.canis infection in our population. A total of 1930 blood donors (age range: 18-55 years) who were admitted to the blood donation centers of different hospitals in Kocaeli province (located at Northwestern part of Turkey) between January-December 2010, have been included in the study.
All of the subjects were negative in terms of Rose-Bengal plate test (B.abortus antigen test). Undiluted serum samples were initially screened by SAT, and those which were found positive were retested by SAT in the dilutions of 1/25 – 1/200. Confirmation of the positive results was performed by using 2-mercaptoethanol (2-ME) SAT. The test antigen (Alton antigen) was prepared from the less mucoid M(-) variant of B.canis, and 1/1048 titered dog antiserum was used as positive control.
Of the 1930 blood donors sera, 40 (2.1%) were found positive with SAT, whereas 16 of them yielded equivocal positive (12 were 1/50, 4 were 1/100 titers) and 15 yielded positive (≥ 1/200 titer) results with 2-ME SAT. As a result, B.canis seropositivity rate in the healthy subjects in this study was estimated as 1.6% (31/1930). The integration of B.canis SAT to the routine serological tests applied for brucellosis diagnosis might aid to the data related to brucellosis epidemiology.
B.canis seroprevalence determined as 1.6% in this study supplied a basic data about the infection in our country. However, larger scale, multicenter studies with different patient and risk groups should be conducted to further evaluate the epidemiology of B.canis infections in Turkey.
Immunochemical characterization of antigens of Brucella canis and their use in seroprevalence study of canine brucellosis.
To explore immunochemical characterization of antigens of Brucella canis (B. canis), and the use in seroprevalence study of canine brucellosis.
External hot phosphate buffer saline extract (HPBSE) and internal sonicated (SA) antigens were prepared from B. canis strain MEX 51 and immunochemically characterized. These antigens were used to test 527 serum samples of dogs by 2-mercaptoethanol-tube agglutination test (2 ME-TAT), agar gel immunodiffusion test (AGID), dot-ELISA and indirect enzyme-linked immunosorbent assay (I-ELISA) to assess the seroprevalence of canine brucellosis.
The protein content of HPBSE and SA antigens was 0.387 mg/mL and 0.195 mg/mL, respectively, whereas carbohydrate content was 0.174 mg/mL and 0.150 mg/mL, respectively. The sodium dodecyl sulfate-polyacrylamide gel electrophoresis (12.5%) of HPBSE and SA, revealed 6 and 8 visible peptide bands ranging from 18-80 kDa and 12-45 kDa, respectively. Western blot analysis showed immunodominant bands of MW 12, 28, 39 and 45 kDa for HPBSE and 20-24 kDa for SA. The AGID revealed HPBSE as more specific antigen than SA but both I-ELISA and dot-ELISA indicated SA antigen to be more specific and reliable than HPBSE. The seroprevalence of canine brucellosis was 2.27% by 2ME-TAT, 1.5% by AGID, 3.03% by dot-ELISA and 16.12% by I-ELISA.
On the basis of the results of present study, we concluded that HPBSE is suitable antigen for AGID, which is more specific; whereas SA antigen is suitable for I-ELISA, which is highly sensitive. Therefore, initial screening of serum samples should be carried out by I-ELISA followed by confirmation with AGID.
Doppler ultrasonographic assessment of maternal and fetal blood flow in abnormal canine pregnancy.
The aim of this study was to describe the changes of uterine artery, umbilical artery and fetal abdominal aorta, renal and internal carotid arteries blood flow in abnormal canine pregnancy. Twenty-two, Brucella-negative pregnant bitches were retrospectively classified into abnormal (which had either interrupted their pregnancy between days 52 and 60 or had perinatal death>>60% of the litter; n=11).