Identification of a novel protein in the genome sequences of Leptospira interrogans with the ability to interact with host's components.

BACKGROUND: Leptospirosis is an infectious disease that affects humans and animals worldwide. The etiological agents of this disease are the pathogenic species of the genus Leptospira. The mechanisms involved in the leptospiral pathogenesis are not full understood. The elucidation of novel mediators of host-pathogen interaction is important in the detection of virulence factors involved in the pathogenesis of leptospirosis. OBJECTIVE: This work focused on identification and characterization of a hypothetical protein of Leptospira encoded by the gene LIC10920. METHODS: The protein of unknown function was predicted to be surface exposed. Therefore, the LIC10920 gene was cloned and the protein expressed in Escherichia coli BL21 (DE3) Star pLysS strain. The recombinant protein was purified by metal affinity chromatography and evaluated with leptospirosis human serum samples. The interaction with host components was also performed. RESULTS: The recombinant protein was recognized by antibodies present in leptopsirosis human serum, suggesting its expression during infection. Immunofluorescence and intact bacteria assays indicated that the bacterial protein is surface-exposed. The recombinant protein interacted with human laminin, in a dose-dependent and saturable manner and was named Lsa24.9, for Leptospiral surface adhesin, followed by its molecular mass. Lsa24.9 also binds plasminogen (PLG) in a dose-dependent and saturable fashion, fulfilling receptor ligand interaction. Moreover, Lsa24.9 has the ability to acquire PLG from normal human serum, exhibiting similar profile as observed with the human purified component. PLG bound Lsa24.9 was able of generating plasmin, which could increase the proteolytic power of the bacteria. CONCLUSIONS: This novel leptospiral protein may function as an adhesin at the colonization steps and may help the invasion process by plasmin generation at the bacterial cell surface.

Identification of a novel protein in the genome sequences of Leptospira interrogans with the ability to interact with host's components

Background Leptospirosis is an infectious disease that affects humans and animals worldwide. The etiological agents of this disease are the pathogenic species of the genus Leptospira. The mechanisms involved in the leptospiral pathogenesis are not full understood. The elucidation of novel mediators of host-pathogen interaction is important in the detection of virulence factors involved in the pathogenesis of leptospirosis. Objective This work focused on identification and characterization of a hypothetical protein of Leptospira encoded by the gene LIC10920. Methods The protein of unknown function was predicted to be surface exposed. Therefore, the LIC10920 gene was cloned and the protein expressed in Escherichia coli BL21 (DE3) Star pLysS strain. The recombinant protein was purified by metal affinity chromatography and evaluated with leptospirosis human serum samples. The interaction with host components was also performed. Results The recombinant protein was recognized by antibodies present in leptopsirosis human serum, suggesting its expression during infection. Immunofluorescence and intact bacteria assays indicated that the bacterial protein is surface-exposed. The recombinant protein interacted with human laminin, in a dose-dependent and saturable manner and was named Lsa24.9, for Leptospiral surface adhesin, followed by its molecular mass. Lsa24.9 also binds plasminogen (PLG) in a dose-dependent and saturable fashion, fulfilling receptor ligand interaction. Moreover, Lsa24.9 has the ability to acquire PLG from normal human serum, exhibiting similar profile as observed with the human purified component. PLG bound Lsa24.9 was able of generating plasmin, which could increase the proteolytic power of the bacteria. Conclusions This novel leptospiral protein may function as an adhesin at the colonization steps and may help the invasion process by plasmin generation at the bacterial cell surface.

Identification of a novel protein in the genome sequences of Leptospira interrogans with the ability to interact with host's components

Background Leptospirosis is an infectious disease that affects humans and animals worldwide. The etiological agents of this disease are the pathogenic species of the genus Leptospira. The mechanisms involved in the leptospiral pathogenesis are not full understood. The elucidation of novel mediators of host-pathogen interaction is important in the detection of virulence factors involved in the pathogenesis of leptospirosis. Objective This work focused on identification and characterization of a hypothetical protein of Leptospira encoded by the gene LIC10920. Methods The protein of unknown function was predicted to be surface exposed. Therefore, the LIC10920 gene was cloned and the protein expressed in Escherichia coli BL21 (DE3) Star pLysS strain. The recombinant protein was purified by metal affinity chromatography and evaluated with leptospirosis human serum samples. The interaction with host components was also performed. Results The recombinant protein was recognized by antibodies present in leptopsirosis human serum, suggesting its expression during infection. Immunofluorescence and intact bacteria assays indicated that the bacterial protein is surface-exposed. The recombinant protein interacted with human laminin, in a dose-dependent and saturable manner and was named Lsa24.9, for Leptospiral surface adhesin, followed by its molecular mass. Lsa24.9 also binds plasminogen (PLG) in a dose-dependent and saturable fashion, fulfilling receptor ligand interaction. Moreover, Lsa24.9 has the ability to acquire PLG from normal human serum, exhibiting similar profile as observed with the human purified component. PLG bound Lsa24.9 was able of generating plasmin, which could increase the proteolytic power of the bacteria. Conclusions This novel leptospiral protein may function as an adhesin at the colonization steps and may help the invasion process by plasmin generation at the bacterial cell surface.

Protective Immunity and Reduced Renal Colonization Induced by Vaccines Containing Recombinant Leptospira interrogans Outer Membrane Proteins and Flagellin Adjuvant.

Leptospirosis is a global zoonotic disease caused by different Leptospira species, such as Leptospira interrogans, that colonize the renal tubules of wild and domestic animals. Thus far, attempts to develop effective leptospirosis vaccines, both for humans and animals, have failed to induce immune responses capable of conferring protection and simultaneously preventing renal colonization. In this study, we evaluated the protective immunity induced by subunit vaccines containing seven different recombinant Leptospira interrogans outer membrane proteins, including the carboxy-terminal portion of the immunoglobulinlike protein A (LigA(C)) and six novel antigens, combined with aluminum hydroxide (alum) or Salmonella flagellin (FliC) as adjuvants. Hamsters vaccinated with the different formulations elicited high antigen-specific antibody titers. Immunization with LigA(C), either with alum or flagellin, conferred protective immunity but did not prevent renal colonization. Similarly, animals immunized with LigA(C) or LigA(C) coadministered with six leptospiral proteins with alum adjuvant conferred protection but did not reduce renal colonization. In contrast, immunizing animals with the pool of seven antigens in combination with flagellin conferred protection and significantly reduced renal colonization by the pathogen. The present study emphasizes the relevance of antigen composition and added adjuvant in the efficacy of antileptospirosis subunit vaccines and shows the complex relationship between immune responses and renal colonization by the pathogen.

Antibacterial activity and chemical compounds of leaves and branches of Protium hebetatum / Atividade anti bacteriana e compostos químicos das folhas e ramos de Protium hebetatum

ABSTRACT The extracts and fractions of leaves and branches of Protium hebetatum D. C. Daly (Burseraceae) were investigated for their antibacterial activity and chemical composition. The methanol extract of branches (EMG) was considered active against the Escherichia coli and the Proteus vulgaris, showing an inhibition zone of 13 mm, and was selected for bioassay-guided phytochemical fractionation. From the technique of broth microdilution, the extract was considered a moderate inhibitor against Staphylococcus aureus, Pseudomonas aeruginosa and Enterococcus faecalis, with a minimum inhibitory concentration (MIC) of 1 mg/mL. The dichloromethane fraction was considered a moderate inhibitor against S. aureus (MIC of 1 mg/mL) and a potent inhibitor against E. faecalis (MIC of 0.5 mg/mL). F1, F2, F5 and F6 from chromatographic column of dichloromethane fraction were considered moderate inhibitors against S. aureus (MIC of 1 mg/mL). Through analysis by a gas chromatography mass spectrometry, eighteen compounds were identified, from which thirteen (isoeugenol, p-vinylguaiacol, metoxyeugenol, coumarin, 5-hydroxy-scopoletin, 4,7-dihydroxy-6-metoxicromam-2-one, 4[(1E]-3-hydroxy-1-propenyl)-2-methoxyphenol, piperonal, scoparon, o-guaiacol, spathulenol, seringol and antiarol) are unprecedented in these species. We also identified the triterpenes α-amyrin and β-amyrin, the steroids stigmasterol and sitosterol and the coumarin scopoletin, which was closely linked to the antibacterial activity of the samples.

RESUMO Atividade antibacteriana e compostos químicos de folhas e galhos de Protium hebetatum. Extratos e frações de folhas e galhos de Protium hebetatum D. C. Daly (Burseraceae) foram investigados quanto sua atividade antibacteriana e composição química. O extrato metanólico dos galhos (EMG) foi considerado ativo contra Escherichia coli e Proteus vulgaris, apresentando um halo de inibição de 13 mm, sendo selecionado para um fracionamento fitoquímico biomonitorado. A partir da técnica de microdiluição em caldo o EMG foi considerado um inibidor moderado contra Staphylococcus aureus, Pseudomonas aeruginosa e Enterococcus faecalis, apresentando uma concentração inibitória mínima (CIM) de 1mg/mL. A fração diclorometânica foi considerada inibidora moderada contra S. aureus (CIM de 1 mg/mL) e inibidora potente contra E. faecalis (CIM de 0,5 mg/mL). F1, F2, F5 e F6 provenientes da fração diclorometânica foram consideradas inibidoras moderadas contra S. aureus (CIM de 1 mg/mL). Através da análise por cromatografia gasosa acoplada a espectrometria de massa, foram identificados dezoitos compostos, dos quais treze (isoeugenol, p-vinilguaiacol, metoxieugenol, cumarina, 5-hidroxi-escopoletina, 4,7-dihidroxi-6-metoxicromam-2-ona, 4[(1E]-3-hidroxi-1-propenil)-2-methoxifenol, piperonal, escoparona, o-guaiacol, espatulenol, seringol e antiarol) foram identificados pela primeira vez nesta espécie. Foram também identificados os triterpenos α-amirina e β-amirina, os esteroides estigmasterol e sitosterol e a cumarina escopoletina, que estão intimamente ligados à atividade antibacteriana da espécie.

Methods for thermal inactivation of pathogens in mozzarella: a comparison between stretching and pasteurization / Métodos para inativação térmica de patógenos em mozarela: comparação entre filagem e pasteurização

This study aimed to evaluate the efficiency of stretching in the reduction of pathogens when compared to milk pasteurization, the official method to ensure safe cheese production. Whole buffalo milk was contaminated with Mycobacterium fortuitum, Listeria monocytogenes, Salmonella typhimurium, and Staphylococcus aureus. Part of the milk was used in mozzarella production and the other part was submitted to holder pasteurization. Pathogens were quantified before and after thermal processing (mozzarella stretching and milk pasteurization). Pasteurization and stretching led to the following reductions in log cycles, respectively: 4.0 and 6.3 for Mycobacterium sp.; 6.0 and 8.4 for Listeria sp.; >6.8 and 4.5 for Staphylococcus sp.; and >8.2 and 7.5 for Salmonella sp.

Este estudo teve como objetivo avaliar a eficácia da filagem na redução de patógenos,em comparação coma pasteurizaçãodo leite, que é o método oficialpara garantir aprodução de queijos seguros. Leite de búfala integral foi contaminado com Mycobacterium fortuitum, Listeria monocytogenes, Salmonella typhimurium e Staphylococcus aureus. Parte desse leite foi empregada na fabricação da mozarela e outra parte foi submetida à pasteurização lenta. Os patógenosforam quantificadosantes e após os processos térmicos (filagem da mozarela e pasteurização do leite). As reduções, em ciclos logarítmicos, causadas pela pasteurização e pela filagem, respectivamente, foram: 4,0 e 6,3 de Mycobacterium sp., 6,0 e 8,4 de Listeria sp., >6,8 e 4,5 de Staphylococcus sp. e >8,2 e 7,5 de Salmonella sp.

Anticorpos anti-Leptospira spp. em ovinos do município de Monte Negro, estado de Rondônia / Anti-Leptospira spp. antibodies in ovines from Monte Negro municipality, Rondonia state, Brazil

O presente estudo determinou a prevalência de anticorpos anti-Leptospira spp. em ovinos do Município de Monte Negro, RO. Foram examinados soros de 141 ovinos de raça, idade e sexo variados provenientes de 15 fazendas, pela técnica de Soroaglutinação Microscópica. Doze (80,0%) propriedades apresentaram pelo menos um animal reagente. Títulos de anticorpos iguais ou superiores a 100 foram detectados em 47 (33,3%) animais, e os sorovares mais frequentes foram Patoc (29,7%), Autumnalis (14,8%), Pyrogenes (10,6%), Australis (4,2%), Bratislava (4,2%), Hardjo (4,2%), Icterohaemorrhagiae (4,2%), Castellonis (2,1%) e Hebdomadis (2,1%). Em 11 (23,4%) soros não foi possível a determinação do provável sorovar envolvido na reação. Alerta-se também para a possibilidade de infecção no homem, tendo em vista as características regionais de fronteira agrícola amazônica.

The present study determined the prevalence of anti-Leptospira spp.antibodies in 141 ovines from 15 farms of the Monte Negro Municipality, Rondonia State, Brazil, by the microscopic agglutination test. Twelve (80.0%) farms presented at least 1 reactive animal. Antibodies titers of ? 100 were detected in 47 (33.3%) animals, the most frequent serovars being Patoc (29.7%), Autumnalis (14.8%), Pyrogenes (10.6%), Australis (4.2%), Bratislava (4.2%), Hardjo (4.2%), Icterohaemorrhagiae (4.2%), Castellonis (2.1%) and Hebdomadis (2.1%). In 11 (23.4%) sera it was not possible to determine the most frequent serovar involved. The results raise a warning as to the possibility of infection in the human being by Leptospira in light of the regional characteristics of the Amazon agricultural frontier.

Aneurisma da arteria esplenica. / Aneurysm of splenic artery

Apresentam os autores a experiencia de dois casos de aneurismas de arteria esplenica agudos no servico de cirurgia do HECC. Fazem revisao da literatura enfatizando o exame fisico e radiologico simples do abdomen dados fundamentais para o diagnostico em que o sinal do "anel de sinete" devera ser sempre pesquisado. Referem ainda os magnificos resultados obtidos com a aneurismectomia e esplenectomia