189 research outputs found
FcpB Is a Surface Filament Protein of the Endoflagellum Required for the Motility of the Spirochete Leptospira
International audienceThe spirochete endoflagellum is a unique motility apparatus among bacteria. Despite its critical importance for pathogenesis, the full composition of the flagellum remains to be determined. We have recently reported that FcpA is a novel flagellar protein and a major component of the sheath of the filament of the spirochete Leptospira. By screening a library of random transposon mutants in the spirochete Leptospira biflexa, we found a motility-deficient mutant harboring a disruption in a hypothetical gene of unknown function. Here, we show that this gene encodes a surface component of the endoflagellar filament and is required for typical hook- and spiral-shaped ends of the cell body, coiled structure of the endoflagella, and high velocity phenotype. We therefore named the gene fcpB for flagellar-coiling protein B. fcpB is conserved in all members of the Leptospira genus, but not present in other organisms including other spirochetes. Complementation of the fcpBâ mutant restored the wild-type morphology and motility phenotypes. Immunoblotting with anti-FcpA and anti-FcpB antisera and cryo-electron microscopy of the filament indicated that FcpB assembled onto the surface of the sheath of the filament and mostly located on the outer (convex) side of the coiled filament. We provide evidence that FcpB, together with FcpA, are Leptospira-specific novel components of the sheath of the filament, key determinants of the coiled and asymmetric structure of the endoflagella and are essential for high velocity. Defining the components of the endoflagella and their functions in these atypical bacteria should greatly enhance our understanding of the mechanisms by which these bacteria produce motility
Quantification of Leptospira interrogans Survival in Soil and Water Microcosms
Leptospira interrogans is the etiological agent of leptospirosis, a globally distributed zoonotic disease. Human infection usually occurs through skin exposure with water and soil contaminated with the urine of chronically infected animals. In this study, we aimed to quantitatively characterize the survival of Leptospira interrogans serovar Copenhageni in environmental matrices. We constructed laboratory microcosms to simulate natural conditions and determined the persistence of DNA markers in soil, mud, spring water and sewage using a quantitative PCR (qPCR) and a propidium monoazide (PMA)-qPCR assay. We found that L. interrogans does not survive at high concentrations in the tested matrices. No net growth was detected in any of the experimental conditions and in all cases the concentration of the DNA markers targeted decreased from the beginning of the experiment following an exponential decay with a decreasing decay rate over time. After 12 and 21 days of incubation the spiked concentration of 106L. interrogans cells/ml or g decreased to approximately 100 cells/ml or g in soil and spring water microcosms, respectively. Furthermore, culturable L. interrogans persisted at concentrations under the limit of detection by PMA-qPCR or qPCR for at least 16 days in soil and 28 days in spring water. Altogether, our findings suggest that the environment is not a multiplication reservoir but a temporary carrier of L. interrogans Copenhageni, although the observed prolonged persistence at low concentrations may still enable the transmission of the disease.IMPORTANCE Leptospirosis is a zoonotic disease caused by spirochetes of the genus Leptospira that primarily affects impoverished populations worldwide. Although leptospirosis is transmitted by contact with water and soil, little is known about the ability of the pathogen to survive in the environment. In this study, we quantitatively characterized the survival of L. interrogans in environmental microcosms and found that although it cannot multiply in water, soil or sewage, it survives for extended time periods (days to weeks depending on the matrix). The survival parameters obtained here may help to better understand the distribution of pathogenic Leptospira in the environment and improve the predictions of human infection risks in areas where such infections are endemic
Linking rattiness, geography and environmental degradation to spillover Leptospira infections in marginalised urban settings: An eco-epidemiological community-based cohort study in Brazil
Background: Zoonotic spillover from animal reservoirs is responsible for a significant global public health burden, but the processes that promote spillover events are poorly understood in complex urban settings. Endemic transmission of Leptospira, the agent of leptospirosis, in marginalised urban communities occurs through human exposure to an environment contaminated by bacteria shed in the urine of the rat reservoir. However, it is unclear to what extent transmission is driven by variation in the distribution of rats or by the dispersal of bacteria in rainwater runoff and overflow from open sewer systems. Methods: We conducted an eco-epidemiological study in a high-risk community in Salvador, Brazil, by prospectively following a cohort of 1401 residents to ascertain serological evidence for leptospiral infections. A concurrent rat ecology study was used to collect information on the fine-scale spatial distribution of 'rattiness', our proxy for rat abundance and exposure of interest. We developed and applied a novel geostatistical framework for joint spatial modelling of multiple indices of disease reservoir abundance and human infection risk. Results: The estimated infection rate was 51.4 (95%CI 40.4, 64.2) infections per 1000 follow-up events. Infection risk increased with age until 30 years of age and was associated with male gender. Rattiness was positively associated with infection risk for residents across the entire study area, but this effect was stronger in higher elevation areas (OR 3.27 95% CI 1.68, 19.07) than in lower elevation areas (OR 1.14 95% CI 1.05, 1.53). Conclusions: These findings suggest that, while frequent flooding events may disperse bacteria in regions of low elevation, environmental risk in higher elevation areas is more localised and directly driven by the distribution of local rat populations. The modelling framework developed may have broad applications in delineating complex animal-environment-human interactions during zoonotic spillover and identifying opportunities for public health intervention
Spatial and temporal dynamics of pathogenic Leptospira in surface waters from the urban slum environment
Leptospirosis has emerged as an important urban health problem as slum settlements have expanded worldwide. Yet the dynamics of the environmentally transmitted Leptospira pathogen has not been well characterized in these settings. We used a stratified dense sampling scheme to study the dynamics of Leptospira abundance in surface waters from a Brazilian urban slum community. We collected surface water samples during the dry, intermediate and rainy seasons within a seven-month period and quantified pathogenic Leptospira by quantitative PCR (qPCR). We used logistic and linear mixed models to identify factors that explained variation for the presence and concentration of Leptospira DNA. Among 335 sewage and 250 standing water samples, Leptospira DNA were detected in 36% and 34%, respectively. Among the 236 samples with positive results geometric mean Leptospira concentrations were 152 GEq/mL. The probability of finding Leptospira DNA was higher in sewage samples collected during the rainy season when increased leptospirosis incidence occurred, than during the dry season (47.2% vs 12.5%, respectively, p = 0.0002). There was a marked spatial and temporal heterogeneity in Leptospira DNA distribution, for which type of water, elevation, and time of day that samples were collected, in addition to season, were significant predictors. Together, these findings indicate that Leptospira are ubiquitous in the slum environment and that the water-related risk to which inhabitants are exposed is low. Seasonal increases in Leptospira presence may explain the timing of leptospirosis outbreaks. Effective prevention will need to consider the spatial and temporal dynamics of pathogenic Leptospira in surface waters to reduce the burden of the disease
A live attenuated-vaccine model confers cross-protective immunity against different species of the Leptospira genus.
Mastite bovina : avaliação microbiolĂłgica do leite, com ĂȘnfase nas leveduras isoladas de casos de mastite clĂnica e subclĂnica, na regiĂŁo do planalto mĂ©dio-RS, em 2005 e 2006.
A mastite bovina Ă© comprovadamente um dos maiores problemas atuais da pecuĂĄria leiteira no Brasil e no mundo, acarretando ĂŽnus aos produtores, Ă indĂșstria e ao consumidor final. Foram analisadas 240 amostras de leite proveniente de animais com mastite clĂnica e de animais com mastite subclĂnica, criados em propriedades leiteiras de alta produção, da regiĂŁo do Planalto MĂ©dio, Rio Grande do Sul. Houve isolamento de 250 agentes bacterianos em 218 amostras (90,8%) e 65 agentes leveduriformes em 39 amostras (16,25%). Foram identificados 10 gĂȘneros e 12 espĂ©cies de bactĂ©rias, sendo os gĂȘneros Staphylococcus, Corynebacterium e Nocardia responsĂĄveis por 88% dos isolados. Foram identificados 08 gĂȘneros (29 espĂ©cies) de leveduras alĂ©m de 02 gĂȘneros de fungos semelhantes a leveduras (04 espĂ©cies), sendo os gĂȘneros Candida, Pichia, Cryptococcus e Rhodotorula responsĂĄveis por 80% das espĂ©cies isoladas.Apenas 03 amostras apresentaram crescimento puro de leveduras, o restante estando associado com isolamentos bacterianos, o que demonstra uma importante relação entre fungos e bactĂ©rias na etiologia das mastites. Mais de 90% dos agentes leveduriformes isolados tiveram crescimento em temperaturas acima de 37ÂșC, demonstrando seu potencial patogĂȘnico. Verificou-se que a prevalĂȘncia de mastite fĂșngica na regiĂŁo Ă© significante e fortemente relacionada com a presença de agentes bacterianos, o tratamento inadequado com produtos antimicrobianos e a saĂșde do Ășbere.The bovine mastitis is one of the largest problems of dairy herds in Brazil and all over the world, resulting in losses to producers, to the industry and to the final consumer. High production dairy herds were analyzed in Planalto MĂ©dio, Rio Grande do Sul, were 240 samples with clinical and subclĂnica mastitis were collected. There was the isolation of 250 bacterial agents in 218 samples (90,8%) and 65 yeast organisms in 39 samples (16,25%). There was identification of 10 genera and 12 species of bacteria, and 88% of the isolates were from genera Staphylococcus, Corynebacterium and Nocardia. There was the identification of 08 genera (29 species) of yeasts and 02 genera of yeast-like organisms (04 species), and 80% of the isolates were from genera Candida, Pichia, Cryptococcus and Rhodotorula. Only 03 samples presented pure growth of yeasts, the remaining was associated with bacterial isolations, which demonstrates an important relationship between yeasts and bacteria in the etiology of the mastitis. More than 90% of the yeast isolates had growth in temperatures above 37ÂșC, which demonstrates their pathogenic potential. It was verified that in this particularly region, the occurrence of mastitis caused by yeasts has a significant importance and are closely related with the bacterial agents' presence, the inadequate treatment with antibiotic products and udderâs health
PatogĂȘnese da leptospirose: estudo sobre os fatores envolvidos na virulĂȘncia e disseminação do agente durante a infecção no modelo animal de hamster
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Previous issue date: 2010Fundação Oswaldo Cruz. Centro de Pesquisas Gonçalo Moniz. Salvador, Bahia, BrasilA leptospirose é uma zoonose de importùncia global e um importante problema de
saĂșde pĂșblica principalmente em paĂses em desenvolvimento. Ă causada por bactĂ©rias do
gĂȘnero Leptospira, uma espiroqueta mĂłvel e de alta morbidade capaz de se disseminar nos
tecidos e causar doença crÎnica em animais hospedeiros. Uma barreira importante para o
controle e prevenção da doença tem sido o pouco conhecimento da patogĂȘnese do agente, em
parte pela falta de ferramentas disponĂveis e eficazes de manipulação genĂ©tica. Um dos
objetivos desse estudo foi caracterizar duas cepas mutantes de Leptospira interrogans. A
interrupção do gene lipl32, que codifica para a proteĂna LipL32, a mais abundante proteĂna no
gĂȘnero Leptospira e expressa somente na superfĂcie das leptospiras patogĂȘnicas, foi realizada
através da inserção do transposon Himar1 no sorovar Manilae. A cepa mutante não
apresentou nenhuma diferença de crescimento ou de aderĂȘncia em componentes da matriz
celular, comparada com a cepa parental. O mutante foi capaz de produzir doença aguda no
modelo animal de hamster e causar colonização crÎnica no modelo animal de rato, mostrando
que LipL32 não possui um papel nestes modelos de infecção. A interrupção do gene ligB foi
realizada com a utilização, pela primeira vez em leptospiras patogĂȘnicas, da tĂ©cnica de
recombinação homĂłloga por mutagĂȘnese dirigida, onde o gene que codifica para a proteĂna
LigB teve uma parte substituĂda por um cassete de resistĂȘncia de espectinomicina (Spcr). Essa
proteĂna, identificada como um possĂvel fator de virulĂȘncia, reconhecida pelo soro de
pacientes infectados e importante para a aderĂȘncia em componentes da matriz celular,
mostrou não ser importante para a infecção aguda ou crÎnica, quando testada frente aos
modelos animais, alĂ©m de nĂŁo ser necessĂĄria para a aderĂȘncia em cultura de cĂ©lulas. Outro
objetivo desse trabalho foi estudar a cinética de disseminação da Leptospira interrogans no
modelo animal de hamster, utilizando uma dose alta (108 leptospiras) e baixa (250 leptospiras)
de inóculo, além de diferentes rotas de infecção. Nossos resultados demonstraram que
leptospiras se disseminam rapidamente em todos os tecidos 01 hora após a infecção com uma
alta dose de inĂłculo e que possivelmente a carga do agente nos tecidos Ă© mais importante para
a patogĂȘnese do que a sua habilidade para a disseminação. TambĂ©m demonstramos que a
motilidade não é essencial para disseminação, mas pode ser essencial para a carga nos tecidos
e letalidade.Leptospirosis is a worldwide zoonosis and a major public health problem especially
important in developing countries. Caused by bacteria of Leptospira genus, a motile lifethreatening
spirochete which is able to disseminate to tissues and causes chronic carriage in
animal hosts. A significant barrier to the control and prevention of leptospirosis has been the
limited understanding of its pathogenesis, due in part to the lack of tools available for the
genetic manipulation of this pathogen. One of the purposes of this study was to characterize
two mutant strains of Leptospira interrogans. Interruption of lipL32 gene, encoding LipL32,
the most abundant protein of pathogenic leptospires and its major outermembrane lipoprotein,
was achieved in serovar Manilae using transposon mutagenesis with Himar1. The mutant had
normal morphology and growth rate compared to the wild type and was equally adherent to
extracellular matrix. The mutant was able to cause acute severe disease manifestations in the
hamster model and chronic colonization in the rat model, showing that LipL32 doesnât play a
role in neither of those models of infection. Interruption of ligB gene was achieved using the
homologous recombination by target mutagenesis for the first time in pathogenic leptospires
and a spectinomycin resistance (Spcr) gene replaced a portion of the ligB coding sequence.
This Lig protein, previously identified as a putative virulence factor, recognized by the sera of
infected patients and important for the adherence in extracellular matrix components, showed
not to be important for acute or chronic infection when tested with animal models and itâs not
required to mediate bacterial adherence to cultured cells. Another purpose of this study was to
determine and analyze the kinetics of dissemination of Leptospira interrogans in the hamster
model, using a high (108 leptospires) and low (250 leptospires) dose of inoculum and different
routes of infection. Our results demonstrated that leptospires can rapidly disseminate through
all tissues after 1 hour post-challenge with a high inoculum dose and that perhaps the load of
the agent in target tissues is more important for pathogenesis than its ability for dissemination.
We also demonstrate that motility is not essential for dissemination but can be essential for
tissue load and lethality
Mastite bovina : avaliação microbiolĂłgica do leite, com ĂȘnfase nas leveduras isoladas de casos de mastite clĂnica e subclĂnica, na regiĂŁo do planalto mĂ©dio-RS, em 2005 e 2006.
A mastite bovina Ă© comprovadamente um dos maiores problemas atuais da pecuĂĄria leiteira no Brasil e no mundo, acarretando ĂŽnus aos produtores, Ă indĂșstria e ao consumidor final. Foram analisadas 240 amostras de leite proveniente de animais com mastite clĂnica e de animais com mastite subclĂnica, criados em propriedades leiteiras de alta produção, da regiĂŁo do Planalto MĂ©dio, Rio Grande do Sul. Houve isolamento de 250 agentes bacterianos em 218 amostras (90,8%) e 65 agentes leveduriformes em 39 amostras (16,25%). Foram identificados 10 gĂȘneros e 12 espĂ©cies de bactĂ©rias, sendo os gĂȘneros Staphylococcus, Corynebacterium e Nocardia responsĂĄveis por 88% dos isolados. Foram identificados 08 gĂȘneros (29 espĂ©cies) de leveduras alĂ©m de 02 gĂȘneros de fungos semelhantes a leveduras (04 espĂ©cies), sendo os gĂȘneros Candida, Pichia, Cryptococcus e Rhodotorula responsĂĄveis por 80% das espĂ©cies isoladas.Apenas 03 amostras apresentaram crescimento puro de leveduras, o restante estando associado com isolamentos bacterianos, o que demonstra uma importante relação entre fungos e bactĂ©rias na etiologia das mastites. Mais de 90% dos agentes leveduriformes isolados tiveram crescimento em temperaturas acima de 37ÂșC, demonstrando seu potencial patogĂȘnico. Verificou-se que a prevalĂȘncia de mastite fĂșngica na regiĂŁo Ă© significante e fortemente relacionada com a presença de agentes bacterianos, o tratamento inadequado com produtos antimicrobianos e a saĂșde do Ășbere.The bovine mastitis is one of the largest problems of dairy herds in Brazil and all over the world, resulting in losses to producers, to the industry and to the final consumer. High production dairy herds were analyzed in Planalto MĂ©dio, Rio Grande do Sul, were 240 samples with clinical and subclĂnica mastitis were collected. There was the isolation of 250 bacterial agents in 218 samples (90,8%) and 65 yeast organisms in 39 samples (16,25%). There was identification of 10 genera and 12 species of bacteria, and 88% of the isolates were from genera Staphylococcus, Corynebacterium and Nocardia. There was the identification of 08 genera (29 species) of yeasts and 02 genera of yeast-like organisms (04 species), and 80% of the isolates were from genera Candida, Pichia, Cryptococcus and Rhodotorula. Only 03 samples presented pure growth of yeasts, the remaining was associated with bacterial isolations, which demonstrates an important relationship between yeasts and bacteria in the etiology of the mastitis. More than 90% of the yeast isolates had growth in temperatures above 37ÂșC, which demonstrates their pathogenic potential. It was verified that in this particularly region, the occurrence of mastitis caused by yeasts has a significant importance and are closely related with the bacterial agents' presence, the inadequate treatment with antibiotic products and udderâs health
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