SdiA, an N-Acylhomoserine Lactone Receptor, Becomes Active during the Transit of Salmonella enterica through the Gastrointestinal Tract of Turtles

encode a LuxR-type AHL receptor, SdiA, they cannot synthesize AHLs. In vitro, it is known that SdiA can detect AHLs produced by other bacterial species..We conclude that the normal gastrointestinal microbiota of most animal species do not produce AHLs of the correct type, in an appropriate location, or in sufficient quantities to activate SdiA. However, the results obtained with turtles represent the first demonstration of SdiA activity in animals

Essential Domains of Anaplasma phagocytophilum Invasins Utilized to Infect Mammalian Host Cells

Anaplasma phagocytophilum causes granulocytic anaplasmosis, an emerging disease of humans and domestic animals. The obligate intracellular bacterium uses its invasins OmpA, Asp14, and AipA to infect myeloid and non-phagocytic cells. Identifying the domains of these proteins that mediate binding and entry, and determining the molecular basis of their interactions with host cell receptors would significantly advance understanding of A. phagocytophilum infection. Here, we identified the OmpA binding domain as residues 59 to 74. Polyclonal antibody generated against a peptide spanning OmpA residues 59 to 74 inhibited A. phagocytophilum infection of host cells and binding to its receptor, sialyl Lewis x (sLex-capped P-selectin glycoprotein ligand 1. Molecular docking analyses predicted that OmpA residues G61 and K64 interact with the two sLex sugars that are important for infection, α2,3-sialic acid and α1,3-fucose. Amino acid substitution analyses demonstrated that K64 was necessary, and G61 was contributory, for recombinant OmpA to bind to host cells and competitively inhibit A. phagocytophilum infection. Adherence of OmpA to RF/6A endothelial cells, which express little to no sLex but express the structurally similar glycan, 6-sulfo-sLex, required α2,3-sialic acid and α1,3-fucose and was antagonized by 6-sulfo-sLex antibody. Binding and uptake of OmpA-coated latex beads by myeloid cells was sensitive to sialidase, fucosidase, and sLex antibody. The Asp14 binding domain was also defined, as antibody specific for residues 113 to 124 inhibited infection. Because OmpA, Asp14, and AipA each contribute to the infection process, it was rationalized that the most effective blocking approach would target all three. An antibody cocktail targeting the OmpA, Asp14, and AipA binding domains neutralized A. phagocytophilumbinding and infection of host cells. This study dissects OmpA-receptor interactions and demonstrates the effectiveness of binding domain-specific antibodies for blocking A. phagocytophilum infection

Putative adhesins of Anaplasma marginale: major surface polypeptides 1a and 1b

Genes for the MSP1a and MSP1b subunits of the Anaplasma marginale surface antigen complex MSP1 were previously cloned and expressed in Escherichia coli. We report here the localization of MSP1a and MSP1b polypeptides on the surface of recombinant E. coli by using a live cell indirect immunofluorescent antibody assay. Recombinant E. coli cells expressing the msp1 alpha gene or the msp1 beta gene encoding the MSP1a and MSP1b polypeptide subunits, respectively, were shown by a culture recovery adhesion assay and by direct microscopic examination to specifically adhere to bovine erythrocytes. This adhesion was more than additive when both genes were coexpressed in a single recombinant construct. Similarly, these recombinants hemagglutinated bovine erythrocytes in a microtiter hemagglutination assay. Inhibition of recombinant E. coli adhesion to bovine erythrocytes and hemagglutination inhibition were observed in the presence of homologous monospecific polyclonal antiserum raised against purified MSP1a or MSP1b polypeptide. These data suggest that the MSP1a and MSP1b polypeptides have functions as adhesins on A. marginale initial bodies, probably during erythrocyte invasion

Diagnostic Delay in HPV-Related Oropharyngeal Squamous Cell Carcinoma

Introduction Human papillomavirus-related (HPV + ) oropharyngeal squamous cell carcinoma (OPSCC) is increasing in incidence and presents diagnostic challenges given its unique clinical presentation

Induced immune response of Escherichia coli BL21 expressing recombinant MSP1a and MSP1b proteins of Anaplasma marginale

This work aims to evaluate the potential of immunization with E. coli BL21 expressing the recombinant rMSP1a and rMSP1b proteins of Anaplasma marginale. E. coli BL21 was transformed with recombinant plasmids pET102/msp1&#945; and pET101/msp1&#946;, and rMSP1a and rMSP1b were expressed after induction by IPTG. BALB/c mice were vaccinated with formolized BL21/rMSP1a and BL21/rMSP1b, and the production in mice sera of whole IgG was determined by ELISA. The mice immunized with BL21/rMSP1a showed a better humoral response for whole IgG when compared to the mice immunized with BL21/rMSP1b; these mice exhibited a small response after the second vaccination. Sera of mice immunized with BL21/rMSP1a reacted via western blot with BL21 and rMSP1a, with molecular masses varying from 70 to 105 kDa. Sera of mice immunized with BL21/rMSP1b reacted with BL21 and rMSP1b with a molecular mass of 100 kDa. These results demonstrate that BL21 containing rMSP1a and rMSP1b in the outer membrane were able to produce an immune response in mice, reinforcing its use in vaccine models against bovine anaplasmosis.<br>Esse trabalho avaliou o potencial de imunização de Escherichia coli BL21 expressando as proteínas recombinantes rMSP1a e rMSP1b de Anaplasma marginale. A E. coli BL21 foi transformada com os plasmídios recombinantes pET102/msp1&#945; e pET101/msp1&#946; e as proteínas rMSP1a e rMSP1b foram expressas após indução com IPTG. Camundongos BALB/c foram vacinados com BL21/rMSP1a e BL21/rMSP1b formolisadas, e a produção de IgG total foi determinada pelo teste de ELISA nos soros dos camundongos imunizados. Os camundongos imunizados com a BL21/rMSP1a mostraram uma melhor resposta humoral para IgG total, comparada à resposta apresentada pelos camundongos imunizados com BL21/rMSP1b; estes camundongos exibiram uma menor resposta após a segunda vacinação. Soros de camundongos imunizados BL21/rMSP1a reagiram pelo western blot com BL21 e rMSP1a, com massa molecular variando de 70 a 105 kDa. Soro de camundongos imunizados com BL21/rMSP1b reagiram com BL21 e rMSP1b com massa molecular de 100 kDa. Esses resultados demonstram que BL21 contendo rMSP1a e rMSP1b na membrana externa foram capazes de produzir resposta imune em camundongos, reforçando o seu uso em modelos de vacina contra a anaplasmose bovina