Cloning, expression, and characterization of the MSP1a and MSP1b recombinant proteins from PR1 Anaplasma marginale strain, Brazil.

The Anaplasma marginale is a bacterium that has obligate intraerythrocytic multiplication in cattle causing important economic loss. The A. marginale major surface protein 1 (MSP1) complex, heterodimer composed of MSP1a and MSP1b, has been identified as adhesins for bovine erythrocytes. The objectives of this study were to sequences the msp1beta gene and produce and characterize recombinant MSP1a and MSP1b from a Brazilian strain of A. marginale, PR1. The msp1alpha and msp1beta genes from the PR1 strain were cloned and expressed in E. coli BL21 Star using the vectors pET102 and pET101/D-TOPO. Antibodies were produced against the recombinant proteins and were shown to react with rMSP1a and rMSP1b demonstrating a molecular mass of 70kDa to 105kDa and 100kDa, respectively for these proteins. Bovine erythrocytes were agglutinated by BL21/rMSP1a and BL21/rMSP1b and, this agglutination was inhibited by the presence of the IgY anti-rMSP1a, confirming the adhesion function of these proteins. Additionally, using the IgY anti-rMSP1a and rMSP1b in a IFI, the presence of rMSP1a and rMSP1b was confirmed on the outer membrane of the recombinant E. coli BL21. Our results show that the msp1beta gene from the PR1 strain has both the conserved region and contain the defined polymorphism regions previously described for other strains of A. marginale. The results from this study confirm adhesive functions for rMSP1a and rMSP1b from PR1 strain in bovine erythrocytes invasion.

Rickettsial infection in capybaras (Hydrochoerus hydrochaeris) from São Paulo, Brazil: serological evidence for infection by Rickettsia bellii and Rickettsia parkeri / Infección por rickettsia en capibaras (Hydrochoerus hydrochaeris) de São Paulo, Brasil: evidencia serológica de infección por Rickettsia bellii y Rickettsia parkeri

Introduction. In Brazil, capybaras (Hydrochoerus hydrochaeris) are important hosts for Amblyomma ticks, which in turn can transmit rickettsiae to humans and animals. Therefore, capybaras are potential sentinels for rickettsial infection. Objective. The present study evaluated rickettsial infection in capybaras in different areas of the state of São Paulo, where rickettsiosis has never been reported. Materials and methods. Blood sera from 73 capybaras from six localities in São Paulo were tested by indirect immunofluorescence assay using Rickettsia rickettsii, Rickettsia parkeri, and Rickettsia bellii antigens. Capybara spleens were tested by PCR, targeting a fragment of the rickettsial gltA gene. Ticks were collected from each capybara sample and taxonomically identified to species. Results. A total of 94 positively reacting capybara samples, 19 (26.0 percent), 25 (34.2 percent), and 50 (68.5 percent) capybara sera reacted to R. rickettsii, R. parkeri, and R. bellii, respectively. Twenty-five capybara sera showed titers to R. bellii at least four-fold higher than to any of the other two antigens. These sera were considered homologous to R. bellii. Using the same criteria, 3 capybara sera were considered homologous to R. parkeri were be considered homologous to R. rickettsii. No rickettsial DNA was detected in capybara spleen samples. Ticks collected on capybaras were Amblyomma dubitatum and Amblyomma cajennense. Conclusions. The first evidence is reported of R. bellii natural infection in vertebrate hosts, and the first evidence of R. parkeri infection in capybaras. While R. parkeri is known to infect and cause disease in humans, no similar evidence for human infection has been indicated by R. bellii.

Introducción. En Brasil, los capibaras (Hydrochoerus hydrochaeris) son importantes huéspedes para garrapatas del género Amblyomma, las cuales transmiten rickettsiosis a humanos y animales. Por lo tanto, estos roedores pueden ser potenciales centinelas para detectar infección por rickettsia. Objetivos. Este trabajo evaluó la infección por rickettsia en capibaras de diferentes regiones del estado de São Paulo, donde las rickettsiosis nunca han sido reportadas. Materiales y métodos. Se examinarion los sueros de 73 capibaras de seis localidades en São Paulo con la prueba de immunofluorescencia indirecta con antígenos de Rickettsia rickettsii, Rickettsia parkeri y Rickettsia bellii. Los bazos de los capibaras se extrajeron y se analizaron por reacción en cadena de la polimerasa para un fragmento del gene gltA de rickettsia. Las garrapatas se recolectaron de los capibaras y se identificaron hasta especie. Resultados. Diecinueve (26,0%), 25 (34,2%) y 50 (68,5%) sueros de los capibaras reaccionaron con R. rickettsii, R. parkeri y R. bellii, respectivamente. De los 50 sueros que reaccionaron con antígenos de R. bellii, 25 presentaron títulos, por lo menos, cuatro veces mayores que los otros dos antígenos. Estos sueros fueron considerados homólogos de R. bellii. Usando el mismo criterio, tres sueros de los capibaras se consideraron homólogos de R. parkeri. Ningún suero se consideró homólogo de R. rickettsii. No se detectó ADN de rickettsia en bazo. Las garrapatas recolectadas de los capibaras fueron identificadas como Amblyomma dubitatum y Amblyomma cajennense. Conclusiones. Este trabajo reporta la primera evidencia de infección natural por R. bellii en vertebrados y, también, la primera evidencia de infección por R. parkeri en capibaras. Se sabe que R. parkeri infecta y produce enfermedad en humanos; sin embargo, no hay evidencia de infección humana por R. bellii.

Isolation and molecular characterization of a Brazilian strain of Borrelia anserina, the agent of fowl spirochaetosis.

Borrelia anserina the agent of fowl spirochaetosis, has a worldwide distribution, where it is transmitted by Argas spp. ticks. The present study reports the first molecular characterization and in vitro isolation of an avian spirochaete strain from Brazil, presumably identified as B. anserina originated from naturally infected Argas miniatus ticks. DNA fragments of the rrs and flab genes were amplified by PCR and sequenced to determine phylogenetic similarities. The resulting sequences were 99.8% (483 of 484) and 98.7% (754 of 764) similar to GenBank corresponding sequences of B. anserina rrs and flaB genes, respectively. By neighbor-joining phylogenetic analysis, the flaB sequence of the Brazilian strain clustered in a monophyletic group with the sequence of B. anserina under 100% bootstrap support. The isolate was successfully isolated in BSK medium, with seven passages performed. The spirochaete strain isolated in the present study was genetically identified as B. anserina labeled as strain PL.

Rickettsial infection in capybaras (Hydrochoerus hydrochaeris) from São Paulo, Brazil: serological evidence for infection by Rickettsia bellii and Rickettsia parkeri.

INTRODUCTION: In Brazil, capybaras (Hydrochoerus hydrochaeris) are important hosts for Amblyomma ticks, which in turn can transmit rickettsiae to humans and animals. Therefore, capybaras are potential sentinels for rickettsial infection. OBJECTIVE: The present study evaluated rickettsial infection in capybaras in different areas of the state of São Paulo, where rickettsiosis has never been reported. Materials and methods. Blood sera from 73 capybaras from six localities in São Paulo were tested by indirect immunofluorescence assay using Rickettsia rickettsii, Rickettsia parkeri, and Rickettsia bellii antigens. Capybara spleens were tested by PCR, targeting a fragment of the rickettsial gltA gene. Ticks were collected from each capybara sample and taxonomically identified to species. RESULTS: A total of 94 positively reacting capybara samples, 19 (26.0%), 25 (34.2%), and 50 (68.5%) capybara sera reacted to R. rickettsii, R. parkeri, and R. bellii, respectively. Twenty-five capybara sera showed titers to R. bellii at least four-fold higher than to any of the other two antigens. These sera were considered homologous to R. bellii. Using the same criteria, 3 capybara sera were considered homologous to R. parkeri. No sera were be considered homologous to R. rickettsii. No rickettsial DNA was detected in capybara spleen samples. Ticks collected on capybaras were Amblyomma dubitatum and Amblyomma cajennense. CONCLUSIONS: The first evidence is reported of R. bellii natural infection in vertebrate hosts, and the first evidence of R. parkeri infection in capybaras. While R. parkeri is known to infect and cause disease in humans, no similar evidence for human infection has been indicated by R. bellii.