Sônia Regina Pinheiro

European 1: a globally important clonal complex of Mycobacterium bovis.

We have identified a globally important clonal complex of Mycobacterium bovis by deletion analysis of over one thousand strains from over 30 countries. We initially show that over 99% of the strains of M. bovis, the cause of bovine tuberculosis, isolated from cattle in the Republic of Ireland and the UK are closely related and are members of a single clonal complex marked by the deletion of chromosomal region RDEu1 and we named this clonal complex European 1 (Eu1). Eu1 strains were present at less than 14% of French, Portuguese and Spanish isolates of M. bovis but are rare in other mainland European countries and Iran. However, strains of the Eu1 clonal complex were found at high frequency in former trading partners of the UK (USA, South Africa, New Zealand, Australia and Canada). The Americas, with the exception of Brazil, are dominated by the Eu1 clonal complex which was at high frequency in Argentina, Chile, Ecuador and Mexico as well as North America. Eu1 was rare or absent in the African countries surveyed except South Africa. A small sample of strains from Taiwan were non-Eu1 but, surprisingly, isolates from Korea and Kazakhstan were members of the Eu1 clonal complex. The simplest explanation for much of the current distribution of the Eu1 clonal complex is that it was spread in infected cattle, such as Herefords, from the UK to former trading partners, although there is evidence of secondary dispersion since. This is the first identification of a globally dispersed clonal complex M. bovis and indicates that much of the current global distribution of this important veterinary pathogen has resulted from relatively recent International trade in cattle.

Sorovares de leptospiras predominantes em exames sorológicos de bubalinos, ovinos, caprinos, eqüinos, suínos e cães de diversos estados brasileiros

From 1984 to 1997 15.558 sorologic tests to leptospirosis were performed (MAT with 24 leptospires serovars) in 284 sheeps, 879 boffaloes, 983 dogs, 1.941 goats, 2.903 horses and 8.568 swines. The distribuition of animals examined by species and state were: sheeps - SP (100%); buffaloes - SP (100%); dogs - SP (80.7%), RS (0.10%), SC (0.10%) and PI (19.0%); goats - SP (33.1%), PB (63.7%) and CE (3.2%); horses - SP (79.3%), RS (9.98%), SC (0.62%), PR (2.5%), RJ (0.17%), MG (1.96%), MT (3.99%), PB (1.3%) and PI (0.03%); swines - SP (61.91%), RS (0.3%), SC (5.95%), PR (3.67%), RJ (0.88%), MG (24.38%), GO (1.12%), SE (0.2%), PE (0.90%), CE (0.34%) and MA (0.1%). From the sheeps tested, 54.5% were examined between 1996 and 97. 33.3% between 1989 and 1990 and 12.2% in the other years; buffaloes - 21.7% from 1984 to 95 and 78.83% between 1996 and 97; dogs - 16.91% from 1984 to 92 and 83.09% from 1993 to 97; goats - 6.97% from 1984 to 91 and 93.09% from 1992 to 97, althought 49% were concerning to 1992; horses - 18.1% from 84 to 90 and 81.9% from 1991 to 97; swines - 61.16% concerning 1990, 91, 95 and 96. The proportion of reactors to at least one leptospira serovar presented the following average and most frequent serovars: sheeps - 0.70% and icterohaemorrhagiae like most frequent serovar; buffaloes - 43.7% and serovars hardjo and pomona; dogs - 17.7% and serovar icterohaemorrhagiae in SP and pyrogenes in PI; goats - 4.17% and icterohaemorrhagiae and grippotyphosa in CE, icterohaemorrhagiae in PB and pyrogenes in SP; horses - 29% and serovar icterohaemorrhagiae in PR, SC, SP, RJ e MG, grippotyphosa in MT, pyrogenes in PB and patoc in RS; swines -24.46% and serovar grippotyphosa and icterohaemorrhagiae in MG, pomona in RS, pomona and icterohaemorrhagiae in PE and RJ, autumnalis in CE and icterohaemorrhagiae in GO, PR, SC e SP.

Isolation of Leptospira santarosai, serovar guaricura from buffaloes (Bubalus bubalis) in Vale do Ribeira, São Paulo, Brazil

In April 1998 urine samples from adult female buffaloes were collected in a farm located in Registro, Vale do Ribeira, Sao Paulo State, Brazil. The urine samples obtained after furosemide injection were immediately transported to the laboratory in liquid modified EMJH medium and seeded, by the serial dilution technique, into Fletchers or modified EMJH-0.2% agar, both of them with 5-fluorouracil 100mg/mL. The intraperitoneoum inoculation of 0.5 mL was also performed with each urine sample in young, adult hamsters (Mesocricetus auratus). All samples seeded directly in culture medium were contaminated. The hamsters did not show any sign of disease and were killed at the 21st post inoculation day. At this time kidney cultures of these animals were performed and from one of them, one leptospira strain (M04-98) was isolated, identified as belonging to serogroup Sejroe by Microscopic Agglutination Test (MAT) with a panel of 36 rabbit sera against serovars representative for the pathogenic serogroups. Subsequently, MAT was carried out with antisera against the 19 reference strains of serogroup Sejroe, revealing a close relationship with serovar guaricura. Afterwards the MAT was done with a panel of 18 monoclonal antibodies representative for serovars of serogroup Sejroe. The histogram closely resembled that of serovar guaricura. So Cross Agglutination Absorption Test (CAAT) was carried out with the buffalo isolate and serovar guaricura, supporting the relationship between the buffalo isolate and serovar guaricura.

Prevalence of Rickettsia felis in the Fleas Ctenocephalides felis felis and Ctenocephalides canis from Two Indian Villages in São Paulo Municipality, Brazil

Abstract:  We evaluated the presence of Rickettsia infection among fleas collected on domestic dogs in two Guarani Indian communities in the suburban area of São Paulo Municipality, Brazil. A total of 114 Ctenocephalides felis felis and 47 Ctenocephalides canis were collected from 40 dogs. A total of 41 C. felis felis (36.0%) and 9 C. canis (19.1%) fleas yielded expected bands by PCR, which were all shown by DNA sequencing to be indentical to the corresponding sequence of a fragment of the Rickettsia felis gltA gene deposited in GenBank. The overall prevalence of R. felis was 31.0% (49/161).

Tuberculosis lymphadenitis in slaughtered swine from S. Paulo State (Brazil): gross lesions, histopathology and demonstration of mycobacteria

INTRODUCTION: As the occurrence of tuberculosis lymphadenitis in swine constitutes a public health risk, especially in immunosuppressed individuals, the distribution of tuberculoid lesions and the presence of micobacteria in lymphonodes and hepatic and muscular tissue in swine for slaughter, in the State of S. Paulo, SP (Brazil), in the period from 1993 to 1994, were studied. MATERIAL E METODO: Tuberculous lesions in 60 carcasses of swine, slaughtered at abattoirs during the period 1993-1994, were studied. When macroscopic lesions were observed, a representative sampling of lymphnodes, hepatic and muscular tissues (masseter and diaphragm) were examined using histophatological and micobacterial isolation techniques. RESULTS: The macroscopic lesions were found predominantly in mesenteric lymphnodes. The MAI Complex (M.avium-intracellulare) was isolated mainly in the carcass group that showed lesions and was not found in the control group. Microorganisms of the MAI Complex were not isolated from hepatic and muscular tissues. CONCLUSION: No clear relationship between the type of mycobacteria isolated and the macroscopic lesions observed during the carcass inspection was found.INTRODUCAO: Como a ocorrencia de linfadenites tuberculoides nos suinos oferece riscos a saude publica, sobretudo em individuos imunocomprometidos, foi estudada a distribuicao das lesoes tuberculoides e a presenca de micobacterias em linfonodos, tecido hepatico e muscular de suinos de abate, no Estado de Sao Paulo, SP (Brasil), no periodo de 1993-1994. MATERIAL E METODO: Foram estudadas 60 carcacas de suinos abatidos, sendo que, trinta apresentavam lesoes tuberculoides macroscopicas (grupo A) e trinta estavam livres de tais lesoes (grupo B ou controle). Foram analisadas seis localizacoes: linfonodos (retrofaringeos, jejunais e mediastinicos), tecido hepatico e tecido muscular (masseter e diafragma). Os exames executados foram: histopatologia e o cultivo para micobacterias. RESULTADOS: No grupo A, 14 em 30 carcacas apresentaram granuloma na histopatologia e, em 14, houve o isolamento de representantes do Complexo MAC (Mycobacterium avium-intracellulare). No grupo B nao foram observadas lesoes na histopatologia, e de quatro carcacas houve o isolamento de micobacterias de rapido crescimento. As lesoes macroscopicas foram encontradas predominantemente nos linfonodos mesentericos. O Complexo MAC foi isolado apenas em linfonodos do grupo com lesao, nao sendo isolado em tecido hepatico e muscular. CONCLUSAO: Nao foi possivel o estabelecimento de relacao entre o tipo de micobacteria isolada e as caracteristicas macroscopicas das lesoes.

Inquérito sorológico e fatores de risco para a brucelose por Brucella canis em cães do município de Santana de Parnaíba, Estado de São Paulo

The prevalence of brucellosis due to Brucella canis was investigated in dogs of the Santana de Parnaiba county, State of Sao Paulo, southeastern Brazil, and the risk factors for infection were analyzed. For this purpose, 410 blood samples were collected from dogs during the rabies vaccination campaign, in August 1999. The agar gel immunodiffusion test (AGID), using lipopolysaccharides and protein antigens from Brucella ovis, strain Reo 198, was applied first as a screening test on normal sera, and secondly, for confirmation. The same AGID test was applied to sera treated previously with 2-mercaptoethanol (ME-AGID). The complement fixation test (CFT), using B. ovis antigen, strain 63/290, was applied also as a confirmatory test. For the prevalence analysis, animals presenting positive results in both ME-AGID and CFT were considered positive. The prevalence of brucellosis due to B. canis was 2.2% (95% C.I.=1.01-4.13%). Dogs that were allowed by their owners to stay free outside their home had a higher risk for contracting B. canis infection, with an odds ratio value of 8.73 (95% C.I.=1.48-51.55) and p=0.04.

Fatores de risco para a leptospirose em fêmeas bovinas em idade reprodutiva no Estado da Bahia, Nordeste do Brasil

Risk factors associated with leptospirosis were identified in cows in the state of Bahia, northeastern Brazil. A total of 10,823 cows with > 24 months of age from 1,414 herds were randomly sampled. For the serological diagnosis of Leptospira spp. infection, the microscopic agglutination test (MAT) was carried out using 24 serovars as antigens. A herd was considered positive when presented at least one seropositive animal. Of the 1,414 investigated herds, 1,076 (77.9%; 95% CI = 75.7-80.0%) presented at least one reactant animal at MAT to any serovar. Serovar Hardjo (Hardjoprajitno) was the most prevalent, with 34.49% (95% CI = 31.97-37.14%) of the positive herds. Presence of more than 28 bovine females in reproductive age (OR=2.11; p<0.001), presence of cervids (OR=2.02; p=0.010), animal purchase (OR=1.57; p<0.001), to slaughter animals in the property (OR=1.58; p=0.030) and to share pasture (OR=1.63; p<0.001) were identified as risk factors for leptospirosis due to any serovar. Risk factors for leptospirosis due to serovar Hardjo (Hardjoprajitno) were presence of swine (OR=1.28; p=0.040) and animal purchase (OR=1.48; p<0.001).

Seroprevalence of toxoplasmosis in a low-income community in the São Paulo municipality, SP, Brazil

Toxoplasmosis is one of the most common zoonoses worldwide. The seroprevalence for T. gondii in human population from Brazil might range from 40 to 80%. The aim of this paper was to study the seroprevalence of T. gondii infection in children from age one to 15 living in a low socioeconomic community, named community of Jardim São Remo in the year of 2002. The community is located in the West area of São Paulo municipality, São Paulo State, Brazil. Antibodies to T. gondii were found in 110 (32.4%, CI 95%: 27.5 - 37.7) of the 339 children tested with indirect immunofluorescent antibody test. The titration of the samples revealed 29 children with serum titer equal to 16, 14 children with 32, 18 children with 64, 21 children with 128, 20 children with 256 and eight children with serum titer > 512. The age dependence of the prevalence of T. gondii infection and the association between seroprevalence for T. gondii and seroprevalence for T. canis suggest that the infection is chiefly postnatal. Seroconversion in infant population of community Jardim São Remo occurs in children as young as two years old, earlier than in the children attended at health centers of São Paulo city. The seroprevalence of T. gondii in children from Jardim São Remo was compared to the prevalence in children from other urban centers of Brazil.

Teste de tuberculinização em caprinos (Capra hircus) experimentalmente sensibilizados

The tuberculin skin test was established with the aim to be applied in the diagnosis of tuberculosis in experimentally sensitized goats. Thirty goats were alocated into three groups with ten animals each. The animals in group A were sensitized with Mycobacterium avium sample D4; group B with Mycobacterium bovis sample AN5; and group C (control) was inoculated with saline solution. The results of the simple cervical test after 72h of bovine tuberculin inoculation was interpreted as follow: positive reaction, when there was an increase in the skin fold thickness greater than 3.9mm; suspicious, when from 1.8 to 3.8mm; and negative when less than 1.7mm. The analysis of the results of the comparative cervical test between M. avium and M. bovis and analysed, 72h after tuberculin, indicated positive reactions, when the increase in skin fold thickness induced by M. bovis was greater than that one induced by the avian tuberculin at least 2.5mm; it was considered suspicious when the difference between the bovine and avian tuberculin reactions was from 1.9 to 2.4mm; and negative when this difference was smaller than 1.8mm. The histological evaluation of the local specific tuberculin response, were performed in skin samples collected from five goats in group A, five in group B, and four in group C, the results were the presence of mononuclear inflammatory infiltrate at 96h after tuberculin inoculation.

The use of MPB70-ELISA for the diagnosis of caprine tuberculosis in Brazil.

After one clinical case that evidenced the outbreak, a complete screening by intradermal tuberculin test was performed in one goat herd in Brazil. The herd was composed by 500 animals and 83 of them (16.6%) showed to be reactive to the comparative double cervical intradermal test. Four months after the test, all the 83 reactive animals were slaughtered and blood samples were collected from 45 of them, for serological assays. From those 45, 32 were randomly chosen for necropsy and histopathological and bacteriological procedures were conducted. Histopathology evidenced at least one characteristic lesion of tuberculosis in each animal, with typical granulommas where acid-fast bacilli (AFB) could be observed. Bacteriology was positive for Mycobacterium bovis in 22 samples (68.7%), therefore confirming the etiology of the outbreak. Sera of 45 animals plus 20 other from a certified free tuberculosis farm were tested in an ELISA using the recombinant M.bovis protein MPB70 as capture antigens. From those, 43 were reactive to the test, with high ODs results, considering a cut-off point established by ROC curve analyzing results (cut-off = 0.8; mean = 0.55; range: 0.157–1.357). These results suggest that MPB70-ELISA can be considered as a reliable tool to diagnose tuberculosis in goat herds, since this assay was capable to correctly detect 95.6% of the animals here examined.