Abstract:

Consumption of inadequately pasteurized dairy products is the most common means of transmission of brucellosis. This report describes two foodborne outbreaks that occurred in families infected after consumption of fresh home-made cheese bought in different Argentine provinces. High resolution variable number of tandem repeats (VNTR)-based analysis revealed two well-defined groups comprising essentially identical profiles and corresponding to the two different outbreaks. Similar clinical findings in members of the same family could indicate that the differential virulence of different bacterial clones, as indicated by VNTR data, could have influenced the course of the disease. We observed the importance of adequate treatment in early stages of the disease; combination therapy and extended treatment for 6 weeks or longer yielded significantly better results. The risk of the foodborne transmission of this zoonotic disease and disease prevention should be considered.

Author affiliation: Lucero, Nidia E. ANLIS Dr.C.G.Malbrán. Instituto Nacional de Enfermedades Infecciosas. Laboratorio de Brucelosis; Argentina.

Author affiliation: Jacob, Néstor R. Hospital Dr. Cosme Argerich. Departamento de Infectologia; Argentina.

Author affiliation: Tenenbaum, Marina. Hospital de Pediatria Dr. Ricardo Gutierrez. Departamento de Infectologia; Argentina.

Author affiliation: Escobar, Gabriela I. ANLIS Dr.C.G.Malbrán. Instituto Nacional de Enfermedades Infecciosas. Laboratorio de Brucelosis; Argentina.

Author affiliation: Groussaud, Pauline. Veterinary Laboratories Agency. Department of Statutory and Exotic Bacterial Diseases. Brucellosis Reference Unit; Reino Unido.

Abstract:

Canine brucellosis, caused by Brucella canis, provokes epidydimitis, testicular atrophy and sterility in male dogs, while in females the major symptom is miscarriage. Transmission to humans may be through contact with semen, urine and/or aborted fetuses of infected animals. Our study, observational and cross-sectional, focused on dogs in lower class neighborhoods and slums with a high rate of unmet basic needs (UBN) in 8 areas of the city of Buenos Aires. We studied 219 dogs: 184 females and 35 males, that tested negative to the buffered plate antigen test (BPAT), which ruled out infection with smooth species of Brucella. We detected anti-B. canis antibodies in 16 dogs (7.3%): 9 females and 7 males, relevant data since the urine of males is considered one of the vectors for the spread of the infection. Although we could run blood cultures on only 175 animals, we isolated B. canis in 3 (2 females and 1 male). Only 3 of the owners of dogs that tested positive consented to a serological diagnosis and two of them were positive. We highlight that the agar gel immunodiffusion test (IGID) proved to have low sensitivity, having detected only one of the 16 positive cases and none of the three confirmed by isolation. We conclude that in the areas studied, the detection of serologically positive dogs and the isolation of B. canis in 3 cases are indicators of the health hazard for the population exposed to it

Author affiliation: Boeri, Eduardo. Instituto de Zoonosis Luis Pasteur; Argentina.

Author affiliation: Escobar, Gabriela I. ANLIS Dr.C.G.Malbrán. Instituto Nacional de Enfermedades Infecciosas. Servicio de Brucelosis; Argentina .

Author affiliation: Ayala, Sandra M. ANLIS Dr.C.G.Malbrán. Instituto Nacional de Enfermedades Infecciosas. Servicio de Brucelosis; Argentina.

Author affiliation: Sosa-Estani, Sergio. ANLIS Dr.C.G.Malbrán. Centro Nacional de Diagnóstico e Investigación en Endemo-Epidemias; Argentina.

Author affiliation: Lucero, Nidia E. ANLIS Dr.C.G.Malbrán. Instituto Nacional de Enfermedades Infecciosas. Servicio de Brucelosis; Argentina.

Abstract:

The transmission of Brucella canis to man commonly occurs through contact with infected dogs or their secretions, or through direct laboratory exposure. The disease is underdiagnosed due to a general lack of serological testing facilities and misconceptions concerning its prevalence. This report shows the potential use of an indirect ELISA (IELISA) for the diagnosis of human brucellosis caused by B. canis in a population of patients negative by smooth-Brucella antigen tests but positive by rapid slide agglutination test (RSAT). One hundred and ten sera from asymptomatic people found negative by tests using smooth Brucella abortus antigen and by RSAT showed an IELISA specificity of 100 % when a cut-off value of 27 % positivity (%P) was selected. For 17 sera from patients with positive B. canis culture or in close contact with culture-positive dogs, the IELISA sensitivity was 100 % with the same cut-off value. The positive patients presented clinical symptoms similar to brucellosis caused by other species of Brucella and some of them received antibiotic treatment and made good progress. Using this cut-off value, we studied 35 patients with negative blood cultures but positive RSATs, and IELISA detected 18 as positive; of the 17 IELISA-negative, two were RSAT-positive at dilution 1 : 2 and 15 were weakly positive with pure serum. These samples were probably from patients at an early stage of infection or indicate false-positive results. No cross-reaction was observed among the sera from nine cases with a diagnosis other than brucellosis, but cross-reactivity was evident in sera from patients infected with smooth-Brucella species. Since routine brucellosis diagnosis does not include B. canis investigation, infection with this species may be more widespread than is currently suspected. The RSAT could be a suitable screening test for the diagnosis of B. canis human brucellosis, and a supplementary technique, such as IELISA, performed on all positive RSAT samples that were negative by B. abortus antigen could ensure diagnostic specificity and confirm the diagnosis.

Author affiliation: Lucero, Nidia E. ANLIS Dr.C.G.Malbrán. Laboratorio de Brucelosis; Argentina.

Author affiliation: Escobar, Gabriela I. ANLIS Dr.C.G.Malbrán. Laboratorio de Brucelosis; Argentina.

Author affiliation: Ayala, Sandra M. ANLIS Dr.C.G.Malbrán. Laboratorio de Brucelosis; Argentina.

Author affiliation: Jacob, Néstor R. Hospital Cosme Argerich. Infectologia; Argentina.

Abstract:

Fluorescence polarization immunoassay (FPA) uses molecular rotational properties to measure antibody binding toantigendirectly. Thepotential useof thismethodwas assessed incomparison to a competitive enzyme immunoassay (CELISA) and conventional serological tests for the diagnosis of brucellosis on a total of 587 human sera. Based on 340 sera from asymptomatic blood donors with no evidence of brucellosis, the specificity of the FPAwas97·9% using a cut-off value of 72mP. Sera fromBrucella-infected patients (11Brucellamelitensis,32Brucella abortus,32Brucella suis and oneBrucellasp.) yielded a sensitivity estimate of 96·1%. In tests on 84 sera from suspected brucellosis patients, the FPA detected 80 cases. Of 87 sera from patients with probable infection, 15 were detected by both CELISA and FPA, three by CELISA only and four by FPA only. The discrepancies inbothgroups involvedserawith low, declining titres. TheFPAuses a sampleof 40 l serum, takesabout 5 min tocompleteandhasbeendemonstrated tobeaccurate for thedetectionof antibodies toB. abortus, B. melitensisandB. suisand for identifying patients suffering relapses. Because of the ease of the procedure, it could be readily adopted for use in clinical laboratories and blood banks.

Author affiliation: Lucero, Nidia E. ANLIS Dr.C.G.Malbrán. Laboratorio de Brucelosis; Argentina.

Author affiliation: Escobar, Gabriela I. ANLIS Dr.C.G.Malbrán. Laboratorio de Brucelosis; Argentina.

Author affiliation: Ayala, Sandra M. ANLIS Dr.C.G.Malbrán. Laboratorio de Brucelosis; Argentina.

Author affiliation: Silva Paulo, Patricia, Comision Nacional de Energia Atomica. Centro Atomico de Ezeiza; Argentina.

Author affiliation: Nielsen, Klaus. Canadian Food Inspection Agency. Animal Diseases Research Institute; Canada.

Abstract:

The diagnosis of B. canis infection in dogs is based on bacteriological examination and serological methods including agglutination and gel diffusion tests. Bacteriological studies are the only methods that have been considered specific but, as intermittent periods of abacteraemia may occur, a negative blood culture cannot be used as a criterion for excluding canine brucellosis. Close contact between people and infected dogs increases the risk of transmission; however, its impact on public health is probably underestimated due to lack of reporting and inadequate diagnostic services. This paper describes an indirect enzyme-linked immunoassay (IELISA) procedure for the diagnosis of brucellosis caused byB. canis in a population of normal and infected dogs previously screened by the buffered plate antigen test (BPAT) and rapid slide agglutination test (RSAT). The serological survey was performed with 446 field sera. The 270 sera from the asymptomatic group found negative by BPA, RSAT and blood culture showed IELISA specificities of 96.7% and 100%, respectively, when cut-off values of OD 0.237 and 0.281 were selected. For 52 sera from culture-positive dogs, IELISA sensitivity was 100% with cut-off values of OD414 0.237 and 0.281. OD414 0.281 was selected because this value provided the highest accuracy with minimal false-negative and false-positive results. This cut-off value was used to study 124 blood culture-negative but RSAT positive sera. IELISA produced 107 positive results; the 17 sera that were negative by IELISA presented a wide range of reactivities by RSAT (2 were RSAT positive at 1 in 2 dilution and 15 were weakly positive with pure serum). These samples were probably from animals at an early stage of the infection or were false-positive results. The IELISA described here detects IgG and IgA antibodies that are useful for evaluating the clinical status of dogs. Although RSAT is a practical screening test, a supplementary technique such as IELISA should be used on all positive RSAT samples to ensure diagnostic specificity. Furthermore, people in contact with infected dogs could be investigated for possible transmission. The procedure described in this study was relatively simple and could have widespread applications.

Author affiliation: Lucero, Nidia E. ANLIS Dr.C.G.Malbrán. Laboratorio de Brucelosis; Argentina.

Author affiliation: Escobar, Gabriela I. ANLIS Dr.C.G.Malbrán. Laboratorio de Brucelosis; Argentina.

Author affiliation: Ayala, Sandra M. ANLIS Dr.C.G.Malbrán. Laboratorio de Brucelosis; Argentina.

Author affiliation: Lopez, G. Municipalidad Lomas de Zamora. Centro de Zoonosis; Argentina.

Abstract:

Clinical diagnosis of canine brucellosis is not sensitive enough and a negative blood culture cannot rule out the disease. Indirect methods of serological testing such as agar gel immunodiffusion (AGID), rapid slide agglutination test (RSAT) and indirect enzyme linked immunoassay (IELISA) are preferred for routine diagnosis. Since Brucella canis shares antigenic components with the Brucella ovis and Brucella abortus RB51 strain, it would seem that either strain could be used as antigen. We present data on AGID and IELISA tests using the B. ovis antigen, RSAT and IELISA using the B. canis antigen and IELISA using the B. abortus RB51 antigen. The cut-off values were adjusted by the ROC analysis; the IELISA-B. ovis cut-off value was 23 (%P) and the IELISA-B. abortus RB51, 24 (%P), with 100% sensitivity and 98.8% specificity. RSAT detected 100% of positive cases, while AGID was less sensitive. The sera from dogs treated with antibiotic showed that %P correlated well with the clinical course. Sera from dogs presumptively infected with B. suis were negative in all tests performed with the rough Brucella strains. RSAT is a very sensitive screening test and IELISA-B. canis, B. ovis and B. abortus RB51 could be used as confirmatory tests, since they show good specificity and sensitivity.

Las técnicas más usadas en el diagnóstico de brucelosis canina son la inmunodifusión en gel de agar (AGID), la microaglutinación en portaobjetos (RSAT) y el ELISA indirecto ya que el diagnóstico clínico es poco sensible y el bacteriológico no excluye la enfermedad. Como Brucella canis comparte componentes antigénicos con Brucella ovis y Brucella abortus RB51, estas cepas podrían ser usadas indistintamente como antígenos. En este trabajo presentamos datos sobre las pruebas de AGID e IELISA con antígeno B. ovis, RSAT e IELISA con antígeno B. canis e IELISA con antígeno B. abortus RB51. Los puntos de corte ajustados con la curva ROC fueron (%P) 23 para IELISA-B. ovis y (%P) 24 para IELISA-RB51 con 100% de sensibilidad y 98,8% de especificidad. RSAT detectó el 100% de los casos positivos, pero AGID fue menos sensible. Los sueros de los perros tratados con antibióticos tuvieron un %P que correlacionó con la evolución clínica. Los perros presumiblemente infectados con B. suis fueron negativos a todos los ensayos realizados con antígenos de cepas de Brucella rugosas. RSAT es muy sensible como prueba tamiz y los IELISA con antígenos de B. canis, B. ovis y B. abortus RB51 podrían ser usados como confirmatorios ya que han demostrado alta sensibilidad y especificidad.

Author affiliation: Escobar, Gabriela I. ANLIS Dr.C.G.Malbrán; Argentina.

Author affiliation: Boeri, Eduardo. Hospital Luis Pasteur; Argentina.

Author affiliation: Ayala, Sandra M. ANLIS Dr.C.G.Malbrán; Argentina.

Author affiliation: Lucero, Nidia E. ANLIS Dr.C.G.Malbrán; Argentina.

Abstract:

Brucella canis is considered a rare cause of human brucellosis. The clinical importance of this infection may have been underestimated so far because of difficulties with presumptive diagnosis. The case described here presented symptoms compatible with brucellosis but the routine tests using Brucella abortus antigen were negative. The infection would have remained undiagnosed if culture had not been positive. This case illustrates the potential for a favourable outcome in Brucella canis diagnosis and supports recommendations for the use of B. canis serology. The infection should be suspected in patients with compatible symptoms and negative serology for B. abortus antigen.

Author affiliation: Lucero, Nidia E. ANLIS Dr.C.G.Malbrán. Laboratorio de Brucelosis; Argentina.

Author affiliation: Jacob, Nestor O. Hospital Cosme Argerich. Unidad de Transplante Renal; Argentina.

Author affiliation: Ayala, Sandra M. ANLIS Dr.C.G.Malbrán. Laboratorio de Brucelosis; Argentina.

Author affiliation: Escobar, Gabriela I. ANLIS Dr.C.G.Malbrán. Laboratorio de Brucelosis; Argentina.

Author affiliation: Tuccillo, Patricia. Hospital Naval Pedro Mallo; Argentina.

Author affiliation: Jacques, Isabelle. Institut National de la Recherche Agronomique. Laboratoire de Pathologie Infectieuse et Immunologie; Francia.