Elsio Wunder | University of Connecticut at Storrs (original) (raw)

Papers by Elsio Wunder

Research paper thumbnail of Cathelicidin Insufficiency in Patients with Fatal Leptospirosis

Leptospirosis causes significant morbidity and mortality worldwide; however, the role of the host... more Leptospirosis causes significant morbidity and mortality worldwide; however, the role of the host immune response in disease progression and high case fatality (>10–50%) is poorly understood. We conducted a multi-parameter investigation of patients with acute leptospi-rosis to identify mechanisms associated with case fatality. Whole blood transcriptional profiling of 16 hospitalized Brazilian patients with acute leptospirosis (13 survivors, 3 deceased) revealed fatal cases had lower expression of the antimicrobial peptide, cathelici-din, and chemokines, but more abundant pro-inflammatory cytokine receptors. In contrast, survivors generated strong adaptive immune signatures, including transcripts relevant to antigen presentation and immunoglobulin production. In an independent cohort (23 survivors , 22 deceased), fatal cases had higher bacterial loads (P = 0.0004) and lower anti-Lep-tospira antibody titers (P = 0.02) at the time of hospitalization, independent of the duration of illness. Low serum cathelicidin and RANTES levels during acute illness were independent risk factors for higher bacterial loads (P = 0.005) and death (P = 0.04), respectively. To investigate the mechanism of cathelicidin in patients surviving acute disease, we administered LL-37, the active peptide of cathelicidin, in a hamster model of lethal leptospirosis and

Research paper thumbnail of Mastite bovina : avaliação microbiológica do leite, com ênfase nas leveduras isoladas de casos de mastite clínica e subclínica, na região do planalto médio-RS, em 2005 e 2006

Research paper thumbnail of Patogênese da leptospirose: estudo sobre os fatores envolvidos na virulência e disseminação do agente durante a infecção no modelo animal de hamster

Research paper thumbnail of Diversity of yeasts from bovine mastitis in Southern Brazil

Revista Iberoamericana De Micologia, Oct 1, 2008

Mastitis is one of the most serious problems in the dairy cattle farms. The great majority of the... more Mastitis is one of the most serious problems in the dairy cattle farms. The great majority of the cases are caused by bacteria, but lately there have been an increasing number of reports about cases of mycotic etiology. The objective of this work was to characterize the yeasts and yeast-like fungi associated with milk of cows with mastitis. Milk samples (n = 248) from a dairy belt situated around the region of Passo Fundo, hinterland of the state of Rio Grande do Sul, Southern Brazil, were analyzed. Aliquots of 0.1 ml of milk were inoculated on yeast extract-malta agar with chloramphenicol. After a period of incubation of 3-5 days at 22-25 °C, the counting of the morphologically distinct colonies was performed, as well as the isolation and identification through phenotypical and physiological criteria. It was possible to isolate 68 yeast species from 43 (17.3%) of the samples. The most frequent genera were Candida (37.9%), Pichia (19.1%), Cryptococcus (10.3%) and Rhodotorula (10.3%).

Research paper thumbnail of Leveduras associadas à mastite clínica e subclínica

Research paper thumbnail of Doença de Johne: isolamento do Mycobacterium avium subsp. paratuberculosis (Map) em um rebanho leiteiro infectado na região sul do Brasil

Doença de Johne: isolamento do Mycobacterium avium subsp. paratuberculosis (Map) em um rebanho le... more Doença de Johne: isolamento do Mycobacterium avium subsp. paratuberculosis (Map) em um rebanho leiteiro infectado na região sul do Brasil. ...

Research paper thumbnail of Doença de Johne : isolamento do Mycobacterium avium subsp. paratuberculosis e, sua infecção em rebanho leiteiro, no RS

Doença de Johne: isolamento do Mycobacterium avium subsp. paratuberculosis (Map) em um rebanho le... more Doença de Johne: isolamento do Mycobacterium avium subsp. paratuberculosis (Map) em um rebanho leiteiro infectado na região sul do Brasil. ...

Research paper thumbnail of Mastite bovina : agentes identificados no Rio Grande do Sul

A mastite é uma doença de alta freqüência em rebanhos leiteiros e a que mais onera a produção lei... more A mastite é uma doença de alta freqüência em rebanhos leiteiros e a que mais onera a produção leiteira. As perdas econômicas são causadas tanto a nível de fazenda, pela diminuição na produção de leite, custo com mão-de-obra, honorários profissionais, medicamentos, morte ou descarte precoce de animais, como a nível de laticínios, pela queda na qualidade do produto final e diminuição no rendimento industrial para a fabricação dos seus derivados e pelas alterações na composição do leite afetado. Estas perdas acabam sendo indiretamente transmitidas também aos consumidores.O objetivo principal deste trabalho foi o isolamento e a identificação dos agentes bacterianos associados à mastite bovina, principalmente agentes fastidiosos relacionados à esta síndrome e de importância na saúde pública, tais como Prototheca spp., Nocardia spp., Mycoplasma spp.Foram analisadas 223 amostras de leite, previamente congeladas, cultivadas à 37ºC em placas de ágar sangue e McConkey em aerobiose, e também no meio de PPLO em microaerofilia.Foram isolados os seguintes microorganismos: Streptococcus spp. (18, 85%), Corynebacterium bovis (18, 40%), Staphylococcus spp. (14, 75%), Nocardia spp. (6, 75%), Prototheca spp. (1, 35%) e enterobacteriáceas (1, 80%), sendo 5, 80% de amostras contaminas e 32, 30% sem apresentarem crescimento.Das 223 amostras totais, 21 foram de mastite clínica (9, 42%). Destas amostras, foi isolado Nocardia spp. de 28, 58%, sendo que 47, 60% foram de amostras sem crescimento. Foram isoladas também Corynebacterium spp. e Streptococcus spp.Os resultados aqui demonstrados comprovam que a grande maioria das mastites são causadas por bactérias comumente isoladas na rotina laboratorial. Porém, existe um grande número de infecções que ficam sem elucidação, principalmente nos casos de mastite clínica. A continuidade deste trabalho se baseia em uma maior estudo nestas amostras, para determinarmos a causa específica da infecção. (PIBIC/CNPq-UFRGS).

Research paper thumbnail of Aborto eqüino por Leptospira sp

Research paper thumbnail of Mastite com lesões sistêmicas por Staphylococus aureus subesp. aureus em coelhos

Research paper thumbnail of Diagnostic survey of bovine abortion with special reference to Neospora caninum infection: Importance, repeated abortion and concurrent infection in aborted fetuses in Southern Brazil

The Veterinary Journal, 2006

The protozoa Neospora caninum is an important cause of bovine abortion world-wide. The objective ... more The protozoa Neospora caninum is an important cause of bovine abortion world-wide. The objective of this survey was to determine the distribution pattern of infectious abortion in Southern Brazil with special reference to N. caninum infection. A total of 161 bovine aborted fetuses from 149 farms were analysed during a 1.5 year period. The cause of abortion was identified in 51.5% of cases. Overall, 23% (37/161) of the fetuses were considered to be infected with N. caninum. Bacterial infection accounted for 17.4% (28/161) of cases, fungal infection for 3.1% (5/161) of cases and viral aetiology for 1.8% (3/161). Six fetuses had concurrent infection with N. caninum and Leptospira spp. Data from 111 fetuses and the respective aborted cows were analysed to investigate the association between previous abortion and current N. caninum infection. The prevalence of N. caninum-infected fetuses from cows with and without a history of previous abortion was 44% (11/25) and 24.4% (21/86), respectively. Cows aborting a N. caninum-infected fetuses were 2.4 times more likely to have aborted previously than cows aborting for other reasons (95% CI of odds ratio = 0.9-6.8, P = 0.06).

Research paper thumbnail of Cathelicidin Insufficiency in Patients with Fatal Leptospirosis

PLOS Pathogens, 2016

Leptospirosis causes significant morbidity and mortality worldwide; however, the role of the host... more Leptospirosis causes significant morbidity and mortality worldwide; however, the role of the host immune response in disease progression and high case fatality (>10–50%) is poorly understood. We conducted a multi-parameter investigation of patients with acute leptospi-rosis to identify mechanisms associated with case fatality. Whole blood transcriptional profiling of 16 hospitalized Brazilian patients with acute leptospirosis (13 survivors, 3 deceased) revealed fatal cases had lower expression of the antimicrobial peptide, cathelici-din, and chemokines, but more abundant pro-inflammatory cytokine receptors. In contrast, survivors generated strong adaptive immune signatures, including transcripts relevant to antigen presentation and immunoglobulin production. In an independent cohort (23 survivors , 22 deceased), fatal cases had higher bacterial loads (P = 0.0004) and lower anti-Lep-tospira antibody titers (P = 0.02) at the time of hospitalization, independent of the duration of illness. Low serum cathelicidin and RANTES levels during acute illness were independent risk factors for higher bacterial loads (P = 0.005) and death (P = 0.04), respectively. To investigate the mechanism of cathelicidin in patients surviving acute disease, we administered LL-37, the active peptide of cathelicidin, in a hamster model of lethal leptospirosis and

Research paper thumbnail of Real-time PCR reveals rapid dissemination of Leptospira interrogans after intraperitoneal and conjunctival inoculation of hamsters

Infection and immunity, Jan 2, 2016

The pathogen Leptospira interrogans is a highly motile spirochete that causes acute and fulminant... more The pathogen Leptospira interrogans is a highly motile spirochete that causes acute and fulminant infections in humans and other accidental hosts. Hematogenous dissemination is important for infection by this pathogen, but remains poorly understood because few animal model studies have used sensitive tools to quantify bacteria. We evaluated the kinetics of leptospiral infection in Golden Syrian hamsters by a sensitive quantitative real time PCR (TaqMan) with lipl32 as the target gene. The dissemination and bacterial burden was measured after intraperitoneal infection with a high dose (10(8) leptospires) or low dose (2.5 x 10(2) leptospires). We also examined the conjunctival challenge route to mimic the natural history of infection. Quantification of leptospires in perfused animals revealed that pathogens were detected in all organs of intraperitoneally infected hamsters including the eye and brain within 1 hour after inoculation of 10(8) virulent L. interrogans Peaks of 10(5)-10(8)...

Research paper thumbnail of A novel flagellar sheath protein, FcpA, determines filament coiling, translational motility and virulence for theLeptospiraspirochete

Molecular Microbiology, 2016

Leptospira are unique among bacteria based on their helical cell morphology with hook-shaped ends... more Leptospira are unique among bacteria based on their helical cell morphology with hook-shaped ends and the presence of periplasmic flagella (PF) with pronounced spontaneous supercoiling. The factors that provoke such supercoiling, as well as the role that PF coiling plays in generating the characteristic hook-end cell morphology and motility, have not been elucidated. We have now identified an abundant protein from the pathogen L. interrogans, exposed on the PF surface, and named it Flagellar-coiling protein A (FcpA). The gene encoding FcpA is highly conserved among Leptospira and was not found in other bacteria. fcpA(-) mutants, obtained from clinical isolates or by allelic exchange, had relatively straight, smaller-diameter PF, and were not able to produce translational motility. These mutants lost their ability to cause disease in the standard hamster model of leptospirosis. Complementation of fcpA restored the wild-type morphology, motility and virulence phenotypes. In summary, we identified a novel Leptospira 36-kDa protein, the main component of the spirochete's PF sheath, and a key determinant of the flagella's coiled structure. FcpA is essential for bacterial translational motility and to enable the spirochete to penetrate the host, traverse tissue barriers, disseminate to cause systemic infection, and reach target organs. This article is protected by copyright. All rights reserved.

Research paper thumbnail of Acquisition of negative complement regulators by the saprophyte Leptospira biflexa expressing LigA or LigB confers enhanced survival in human serum

Research paper thumbnail of What Makes a Bacterial Species Pathogenic?:Comparative Genomic Analysis of the Genus Leptospira

PLOS Neglected Tropical Diseases, 2016

Leptospirosis, caused by spirochetes of the genus Leptospira, is a globally widespread, neglected... more Leptospirosis, caused by spirochetes of the genus Leptospira, is a globally widespread, neglected and emerging zoonotic disease. While whole genome analysis of individual pathogenic, intermediately pathogenic and saprophytic Leptospira species has been reported, comprehensive cross-species genomic comparison of all known species of infectious and non-infectious Leptospira, with the goal of identifying genes related to pathogenesis and mammalian host adaptation, remains a key gap in the field. Infectious Leptospira, comprised of pathogenic and intermediately pathogenic Leptospira, evolutionarily diverged from non-infectious, saprophytic Leptospira, as demonstrated by the following computational biology analyses: 1) the definitive taxonomy and evolutionary relatedness among all known Leptospira species; 2) genomically-predicted metabolic reconstructions that indicate novel adaptation of infectious Leptospira to mammals, including sialic acid biosynthesis, pathogen-specific porphyrin metabolism and the first-time demonstration of cobalamin (B12) autotrophy as a bacterial virulence factor; 3) CRISPR/Cas systems demonstrated only to be present in pathogenic Leptospira, suggesting a potential mechanism for this clade's refractoriness to gene targeting; 4) finding Leptospira pathogen-specific specialized protein secretion systems; 5) novel virulence-related genes/gene families such as the Virulence Modifying (VM) (PF07598 paralogs) proteins and pathogen-specific adhesins; 6) discovery of novel, pathogen-specific protein modification and secretion mechanisms including unique lipoprotein signal peptide motifs, Sec-independent twin arginine protein secretion motifs, and the absence of certain canonical signal recognition particle proteins from all Leptospira; and 7) and demonstration of infectious Leptospira-specific signal-responsive gene expression, motility and chemotaxis systems. By identifying large scale changes in infectious (pathogenic and intermediately pathogenic) vs. non-infectious Leptospira, this work provides new insights into the evolution of a genus of bacterial pathogens. This work will be a comprehensive roadmap for understanding leptospirosis pathogenesis. More generally, it provides new insights into mechanisms by which bacterial pathogens adapt to mammalian hosts.

Research paper thumbnail of Spatiotemporal Determinants of Urban Leptospirosis Transmission: Four-Year Prospective Cohort Study of Slum Residents in Brazil

PLOS Neglected Tropical Diseases, 2016

Rat-borne leptospirosis is an emerging zoonotic disease in urban slum settlements for which there... more Rat-borne leptospirosis is an emerging zoonotic disease in urban slum settlements for which there are no adequate control measures. The challenge in elucidating risk factors and informing approaches for prevention is the complex and heterogeneous environment within slums, which vary at fine spatial scales and influence transmission of the bacterial agent. We performed a prospective study of 2,003 slum residents in the city of Salvador, Brazil during a four-year period (2003-2007) and used a spatiotemporal modelling approach to delineate the dynamics of leptospiral transmission. Household interviews and Geographical Information System surveys were performed annually to evaluate risk exposures and environmental transmission sources. We completed annual serosurveys to ascertain leptospiral infection based on serological evidence. Among the 1,730 (86%) individuals who completed at least one year of follow-up, the infection rate was 35.4 (95% CI, 30.7-40.6) per 1,000 annual follow-up events. Male gender, illiteracy, and age were independently associated with infection risk. Environmental risk factors included rat infestation (OR 1.46, 95% CI, 1.00-2.16), contact with mud (OR 1.57, 95% CI 1.17-2.17) and lower household elevation (OR 0.92 per 10m increase in elevation, 95% CI 0.82-1.04). The spatial distribution of infection risk was highly heterogeneous and varied across small scales. Fixed effects in the spatiotemporal model accounted for the majority of the spatial variation in risk, but there was a significant residual component that was best explained by the spatial random effect. Although infection risk varied between years, the spatial distribution of risk associated with fixed and random effects did not vary temporally. Specific "hot-spots" consistently had higher transmission risk during study years. The risk for leptospiral infection in urban slums is determined in large part by structural features, both social and environmental. Our findings indicate that topographic factors such as household elevation and inadequate drainage increase risk by promoting contact with mud and suggest that the soil-water interface serves as the environmental reservoir for spillover transmission. The use of a spatiotemporal approach allowed the identification of geographic outliers with unexplained risk patterns. This approach, in addition to guiding targeted community-based interventions and identifying new hypotheses, may have general applicability towards addressing environmentally-transmitted diseases that have emerged in complex urban slum settings.

Research paper thumbnail of Efficient Detection of Pathogenic Leptospires Using 16S Ribosomal RNA

PLOS ONE, 2015

Pathogenic Leptospira species cause a prevalent yet neglected zoonotic disease with mild to life-... more Pathogenic Leptospira species cause a prevalent yet neglected zoonotic disease with mild to life-threatening complications in a variety of susceptible animals and humans. Diagnosis of leptospirosis, which primarily relies on antiquated serotyping methods, is particularly challenging due to presentation of non-specific symptoms shared by other febrile illnesses, often leading to misdiagnosis. Initiation of antimicrobial therapy during early infection to prevent more serious complications of disseminated infection is often not performed because of a lack of efficient diagnostic tests. Here we report that specific regions of leptospiral 16S ribosomal RNA molecules constitute a novel and efficient diagnostic target for PCR-based detection of pathogenic Leptospira serovars. Our diagnostic test using spiked human blood was at least 100-fold more sensitive than corresponding leptospiral DNA-based quantitative PCR assays, targeting the same 16S nucleotide sequence in the RNA and DNA molecules. The sensitivity and specificity of our RNA assay against laboratory-confirmed human leptospirosis clinical samples were 64% and 100%, respectively, which was superior then an established parallel DNA detection assay. Remarkably, we discovered that 16S transcripts remain appreciably stable ex vivo, including untreated and stored human blood samples, further highlighting their use for clinical detection of L. interrogans. Together, these studies underscore a novel utility of RNA targets, specifically 16S rRNA, for development of PCRbased modalities for diagnosis of human leptospirosis, and also may serve as paradigm for detection of additional bacterial pathogens for which early diagnosis is warranted.

Research paper thumbnail of Patterns in Leptospira Shedding in Norway Rats (Rattus norvegicus) from Brazilian Slum Communities at High Risk of Disease Transmission

PLOS Neglected Tropical Diseases, 2015

We address some critical but unknown parameters of individuals and populations of Norway rats (Ra... more We address some critical but unknown parameters of individuals and populations of Norway rats (Rattus norvegicus) that influence leptospiral infection, maintenance and spirochetal loads shed in urine, which contaminates the environment ultimately leading to human infection.

Research paper thumbnail of Proteomic Features Predict Seroreactivity against Leptospiral Antigens in Leptospirosis Patients

Journal of proteome research, Jan 11, 2014

With increasing efficiency, accuracy, and speed we can access complete genome sequences from thou... more With increasing efficiency, accuracy, and speed we can access complete genome sequences from thousands of infectious microorganisms; however, the ability to predict antigenic targets of the immune system based on amino acid sequence alone is still needed. Here we use a Leptospira interrogans microarray expressing 91% (3359) of all leptospiral predicted ORFs (3667) and make an empirical accounting of all antibody reactive antigens recognized in sera from naturally infected humans; 191 antigens elicited an IgM or IgG response, representing 5% of the whole proteome. We classified the reactive antigens into 26 annotated COGs (clusters of orthologous groups), 26 JCVI Mainrole annotations, and 11 computationally predicted proteomic features. Altogether, 14 significantly enriched categories were identified, which are associated with immune recognition including mass spectrometry evidence of in vitro expression and in vivo mRNA up-regulation. Together, this group of 14 enriched categories a...

Research paper thumbnail of Cathelicidin Insufficiency in Patients with Fatal Leptospirosis

Leptospirosis causes significant morbidity and mortality worldwide; however, the role of the host... more Leptospirosis causes significant morbidity and mortality worldwide; however, the role of the host immune response in disease progression and high case fatality (>10–50%) is poorly understood. We conducted a multi-parameter investigation of patients with acute leptospi-rosis to identify mechanisms associated with case fatality. Whole blood transcriptional profiling of 16 hospitalized Brazilian patients with acute leptospirosis (13 survivors, 3 deceased) revealed fatal cases had lower expression of the antimicrobial peptide, cathelici-din, and chemokines, but more abundant pro-inflammatory cytokine receptors. In contrast, survivors generated strong adaptive immune signatures, including transcripts relevant to antigen presentation and immunoglobulin production. In an independent cohort (23 survivors , 22 deceased), fatal cases had higher bacterial loads (P = 0.0004) and lower anti-Lep-tospira antibody titers (P = 0.02) at the time of hospitalization, independent of the duration of illness. Low serum cathelicidin and RANTES levels during acute illness were independent risk factors for higher bacterial loads (P = 0.005) and death (P = 0.04), respectively. To investigate the mechanism of cathelicidin in patients surviving acute disease, we administered LL-37, the active peptide of cathelicidin, in a hamster model of lethal leptospirosis and

Research paper thumbnail of Mastite bovina : avaliação microbiológica do leite, com ênfase nas leveduras isoladas de casos de mastite clínica e subclínica, na região do planalto médio-RS, em 2005 e 2006

Research paper thumbnail of Patogênese da leptospirose: estudo sobre os fatores envolvidos na virulência e disseminação do agente durante a infecção no modelo animal de hamster

Research paper thumbnail of Diversity of yeasts from bovine mastitis in Southern Brazil

Revista Iberoamericana De Micologia, Oct 1, 2008

Mastitis is one of the most serious problems in the dairy cattle farms. The great majority of the... more Mastitis is one of the most serious problems in the dairy cattle farms. The great majority of the cases are caused by bacteria, but lately there have been an increasing number of reports about cases of mycotic etiology. The objective of this work was to characterize the yeasts and yeast-like fungi associated with milk of cows with mastitis. Milk samples (n = 248) from a dairy belt situated around the region of Passo Fundo, hinterland of the state of Rio Grande do Sul, Southern Brazil, were analyzed. Aliquots of 0.1 ml of milk were inoculated on yeast extract-malta agar with chloramphenicol. After a period of incubation of 3-5 days at 22-25 °C, the counting of the morphologically distinct colonies was performed, as well as the isolation and identification through phenotypical and physiological criteria. It was possible to isolate 68 yeast species from 43 (17.3%) of the samples. The most frequent genera were Candida (37.9%), Pichia (19.1%), Cryptococcus (10.3%) and Rhodotorula (10.3%).

Research paper thumbnail of Leveduras associadas à mastite clínica e subclínica

Research paper thumbnail of Doença de Johne: isolamento do Mycobacterium avium subsp. paratuberculosis (Map) em um rebanho leiteiro infectado na região sul do Brasil

Doença de Johne: isolamento do Mycobacterium avium subsp. paratuberculosis (Map) em um rebanho le... more Doença de Johne: isolamento do Mycobacterium avium subsp. paratuberculosis (Map) em um rebanho leiteiro infectado na região sul do Brasil. ...

Research paper thumbnail of Doença de Johne : isolamento do Mycobacterium avium subsp. paratuberculosis e, sua infecção em rebanho leiteiro, no RS

Doença de Johne: isolamento do Mycobacterium avium subsp. paratuberculosis (Map) em um rebanho le... more Doença de Johne: isolamento do Mycobacterium avium subsp. paratuberculosis (Map) em um rebanho leiteiro infectado na região sul do Brasil. ...

Research paper thumbnail of Mastite bovina : agentes identificados no Rio Grande do Sul

A mastite é uma doença de alta freqüência em rebanhos leiteiros e a que mais onera a produção lei... more A mastite é uma doença de alta freqüência em rebanhos leiteiros e a que mais onera a produção leiteira. As perdas econômicas são causadas tanto a nível de fazenda, pela diminuição na produção de leite, custo com mão-de-obra, honorários profissionais, medicamentos, morte ou descarte precoce de animais, como a nível de laticínios, pela queda na qualidade do produto final e diminuição no rendimento industrial para a fabricação dos seus derivados e pelas alterações na composição do leite afetado. Estas perdas acabam sendo indiretamente transmitidas também aos consumidores.O objetivo principal deste trabalho foi o isolamento e a identificação dos agentes bacterianos associados à mastite bovina, principalmente agentes fastidiosos relacionados à esta síndrome e de importância na saúde pública, tais como Prototheca spp., Nocardia spp., Mycoplasma spp.Foram analisadas 223 amostras de leite, previamente congeladas, cultivadas à 37ºC em placas de ágar sangue e McConkey em aerobiose, e também no meio de PPLO em microaerofilia.Foram isolados os seguintes microorganismos: Streptococcus spp. (18, 85%), Corynebacterium bovis (18, 40%), Staphylococcus spp. (14, 75%), Nocardia spp. (6, 75%), Prototheca spp. (1, 35%) e enterobacteriáceas (1, 80%), sendo 5, 80% de amostras contaminas e 32, 30% sem apresentarem crescimento.Das 223 amostras totais, 21 foram de mastite clínica (9, 42%). Destas amostras, foi isolado Nocardia spp. de 28, 58%, sendo que 47, 60% foram de amostras sem crescimento. Foram isoladas também Corynebacterium spp. e Streptococcus spp.Os resultados aqui demonstrados comprovam que a grande maioria das mastites são causadas por bactérias comumente isoladas na rotina laboratorial. Porém, existe um grande número de infecções que ficam sem elucidação, principalmente nos casos de mastite clínica. A continuidade deste trabalho se baseia em uma maior estudo nestas amostras, para determinarmos a causa específica da infecção. (PIBIC/CNPq-UFRGS).

Research paper thumbnail of Aborto eqüino por Leptospira sp

Research paper thumbnail of Mastite com lesões sistêmicas por Staphylococus aureus subesp. aureus em coelhos

Research paper thumbnail of Diagnostic survey of bovine abortion with special reference to Neospora caninum infection: Importance, repeated abortion and concurrent infection in aborted fetuses in Southern Brazil

The Veterinary Journal, 2006

The protozoa Neospora caninum is an important cause of bovine abortion world-wide. The objective ... more The protozoa Neospora caninum is an important cause of bovine abortion world-wide. The objective of this survey was to determine the distribution pattern of infectious abortion in Southern Brazil with special reference to N. caninum infection. A total of 161 bovine aborted fetuses from 149 farms were analysed during a 1.5 year period. The cause of abortion was identified in 51.5% of cases. Overall, 23% (37/161) of the fetuses were considered to be infected with N. caninum. Bacterial infection accounted for 17.4% (28/161) of cases, fungal infection for 3.1% (5/161) of cases and viral aetiology for 1.8% (3/161). Six fetuses had concurrent infection with N. caninum and Leptospira spp. Data from 111 fetuses and the respective aborted cows were analysed to investigate the association between previous abortion and current N. caninum infection. The prevalence of N. caninum-infected fetuses from cows with and without a history of previous abortion was 44% (11/25) and 24.4% (21/86), respectively. Cows aborting a N. caninum-infected fetuses were 2.4 times more likely to have aborted previously than cows aborting for other reasons (95% CI of odds ratio = 0.9-6.8, P = 0.06).

Research paper thumbnail of Cathelicidin Insufficiency in Patients with Fatal Leptospirosis

PLOS Pathogens, 2016

Leptospirosis causes significant morbidity and mortality worldwide; however, the role of the host... more Leptospirosis causes significant morbidity and mortality worldwide; however, the role of the host immune response in disease progression and high case fatality (>10–50%) is poorly understood. We conducted a multi-parameter investigation of patients with acute leptospi-rosis to identify mechanisms associated with case fatality. Whole blood transcriptional profiling of 16 hospitalized Brazilian patients with acute leptospirosis (13 survivors, 3 deceased) revealed fatal cases had lower expression of the antimicrobial peptide, cathelici-din, and chemokines, but more abundant pro-inflammatory cytokine receptors. In contrast, survivors generated strong adaptive immune signatures, including transcripts relevant to antigen presentation and immunoglobulin production. In an independent cohort (23 survivors , 22 deceased), fatal cases had higher bacterial loads (P = 0.0004) and lower anti-Lep-tospira antibody titers (P = 0.02) at the time of hospitalization, independent of the duration of illness. Low serum cathelicidin and RANTES levels during acute illness were independent risk factors for higher bacterial loads (P = 0.005) and death (P = 0.04), respectively. To investigate the mechanism of cathelicidin in patients surviving acute disease, we administered LL-37, the active peptide of cathelicidin, in a hamster model of lethal leptospirosis and

Research paper thumbnail of Real-time PCR reveals rapid dissemination of Leptospira interrogans after intraperitoneal and conjunctival inoculation of hamsters

Infection and immunity, Jan 2, 2016

The pathogen Leptospira interrogans is a highly motile spirochete that causes acute and fulminant... more The pathogen Leptospira interrogans is a highly motile spirochete that causes acute and fulminant infections in humans and other accidental hosts. Hematogenous dissemination is important for infection by this pathogen, but remains poorly understood because few animal model studies have used sensitive tools to quantify bacteria. We evaluated the kinetics of leptospiral infection in Golden Syrian hamsters by a sensitive quantitative real time PCR (TaqMan) with lipl32 as the target gene. The dissemination and bacterial burden was measured after intraperitoneal infection with a high dose (10(8) leptospires) or low dose (2.5 x 10(2) leptospires). We also examined the conjunctival challenge route to mimic the natural history of infection. Quantification of leptospires in perfused animals revealed that pathogens were detected in all organs of intraperitoneally infected hamsters including the eye and brain within 1 hour after inoculation of 10(8) virulent L. interrogans Peaks of 10(5)-10(8)...

Research paper thumbnail of A novel flagellar sheath protein, FcpA, determines filament coiling, translational motility and virulence for theLeptospiraspirochete

Molecular Microbiology, 2016

Leptospira are unique among bacteria based on their helical cell morphology with hook-shaped ends... more Leptospira are unique among bacteria based on their helical cell morphology with hook-shaped ends and the presence of periplasmic flagella (PF) with pronounced spontaneous supercoiling. The factors that provoke such supercoiling, as well as the role that PF coiling plays in generating the characteristic hook-end cell morphology and motility, have not been elucidated. We have now identified an abundant protein from the pathogen L. interrogans, exposed on the PF surface, and named it Flagellar-coiling protein A (FcpA). The gene encoding FcpA is highly conserved among Leptospira and was not found in other bacteria. fcpA(-) mutants, obtained from clinical isolates or by allelic exchange, had relatively straight, smaller-diameter PF, and were not able to produce translational motility. These mutants lost their ability to cause disease in the standard hamster model of leptospirosis. Complementation of fcpA restored the wild-type morphology, motility and virulence phenotypes. In summary, we identified a novel Leptospira 36-kDa protein, the main component of the spirochete's PF sheath, and a key determinant of the flagella's coiled structure. FcpA is essential for bacterial translational motility and to enable the spirochete to penetrate the host, traverse tissue barriers, disseminate to cause systemic infection, and reach target organs. This article is protected by copyright. All rights reserved.

Research paper thumbnail of Acquisition of negative complement regulators by the saprophyte Leptospira biflexa expressing LigA or LigB confers enhanced survival in human serum

Research paper thumbnail of What Makes a Bacterial Species Pathogenic?:Comparative Genomic Analysis of the Genus Leptospira

PLOS Neglected Tropical Diseases, 2016

Leptospirosis, caused by spirochetes of the genus Leptospira, is a globally widespread, neglected... more Leptospirosis, caused by spirochetes of the genus Leptospira, is a globally widespread, neglected and emerging zoonotic disease. While whole genome analysis of individual pathogenic, intermediately pathogenic and saprophytic Leptospira species has been reported, comprehensive cross-species genomic comparison of all known species of infectious and non-infectious Leptospira, with the goal of identifying genes related to pathogenesis and mammalian host adaptation, remains a key gap in the field. Infectious Leptospira, comprised of pathogenic and intermediately pathogenic Leptospira, evolutionarily diverged from non-infectious, saprophytic Leptospira, as demonstrated by the following computational biology analyses: 1) the definitive taxonomy and evolutionary relatedness among all known Leptospira species; 2) genomically-predicted metabolic reconstructions that indicate novel adaptation of infectious Leptospira to mammals, including sialic acid biosynthesis, pathogen-specific porphyrin metabolism and the first-time demonstration of cobalamin (B12) autotrophy as a bacterial virulence factor; 3) CRISPR/Cas systems demonstrated only to be present in pathogenic Leptospira, suggesting a potential mechanism for this clade's refractoriness to gene targeting; 4) finding Leptospira pathogen-specific specialized protein secretion systems; 5) novel virulence-related genes/gene families such as the Virulence Modifying (VM) (PF07598 paralogs) proteins and pathogen-specific adhesins; 6) discovery of novel, pathogen-specific protein modification and secretion mechanisms including unique lipoprotein signal peptide motifs, Sec-independent twin arginine protein secretion motifs, and the absence of certain canonical signal recognition particle proteins from all Leptospira; and 7) and demonstration of infectious Leptospira-specific signal-responsive gene expression, motility and chemotaxis systems. By identifying large scale changes in infectious (pathogenic and intermediately pathogenic) vs. non-infectious Leptospira, this work provides new insights into the evolution of a genus of bacterial pathogens. This work will be a comprehensive roadmap for understanding leptospirosis pathogenesis. More generally, it provides new insights into mechanisms by which bacterial pathogens adapt to mammalian hosts.

Research paper thumbnail of Spatiotemporal Determinants of Urban Leptospirosis Transmission: Four-Year Prospective Cohort Study of Slum Residents in Brazil

PLOS Neglected Tropical Diseases, 2016

Rat-borne leptospirosis is an emerging zoonotic disease in urban slum settlements for which there... more Rat-borne leptospirosis is an emerging zoonotic disease in urban slum settlements for which there are no adequate control measures. The challenge in elucidating risk factors and informing approaches for prevention is the complex and heterogeneous environment within slums, which vary at fine spatial scales and influence transmission of the bacterial agent. We performed a prospective study of 2,003 slum residents in the city of Salvador, Brazil during a four-year period (2003-2007) and used a spatiotemporal modelling approach to delineate the dynamics of leptospiral transmission. Household interviews and Geographical Information System surveys were performed annually to evaluate risk exposures and environmental transmission sources. We completed annual serosurveys to ascertain leptospiral infection based on serological evidence. Among the 1,730 (86%) individuals who completed at least one year of follow-up, the infection rate was 35.4 (95% CI, 30.7-40.6) per 1,000 annual follow-up events. Male gender, illiteracy, and age were independently associated with infection risk. Environmental risk factors included rat infestation (OR 1.46, 95% CI, 1.00-2.16), contact with mud (OR 1.57, 95% CI 1.17-2.17) and lower household elevation (OR 0.92 per 10m increase in elevation, 95% CI 0.82-1.04). The spatial distribution of infection risk was highly heterogeneous and varied across small scales. Fixed effects in the spatiotemporal model accounted for the majority of the spatial variation in risk, but there was a significant residual component that was best explained by the spatial random effect. Although infection risk varied between years, the spatial distribution of risk associated with fixed and random effects did not vary temporally. Specific "hot-spots" consistently had higher transmission risk during study years. The risk for leptospiral infection in urban slums is determined in large part by structural features, both social and environmental. Our findings indicate that topographic factors such as household elevation and inadequate drainage increase risk by promoting contact with mud and suggest that the soil-water interface serves as the environmental reservoir for spillover transmission. The use of a spatiotemporal approach allowed the identification of geographic outliers with unexplained risk patterns. This approach, in addition to guiding targeted community-based interventions and identifying new hypotheses, may have general applicability towards addressing environmentally-transmitted diseases that have emerged in complex urban slum settings.

Research paper thumbnail of Efficient Detection of Pathogenic Leptospires Using 16S Ribosomal RNA

PLOS ONE, 2015

Pathogenic Leptospira species cause a prevalent yet neglected zoonotic disease with mild to life-... more Pathogenic Leptospira species cause a prevalent yet neglected zoonotic disease with mild to life-threatening complications in a variety of susceptible animals and humans. Diagnosis of leptospirosis, which primarily relies on antiquated serotyping methods, is particularly challenging due to presentation of non-specific symptoms shared by other febrile illnesses, often leading to misdiagnosis. Initiation of antimicrobial therapy during early infection to prevent more serious complications of disseminated infection is often not performed because of a lack of efficient diagnostic tests. Here we report that specific regions of leptospiral 16S ribosomal RNA molecules constitute a novel and efficient diagnostic target for PCR-based detection of pathogenic Leptospira serovars. Our diagnostic test using spiked human blood was at least 100-fold more sensitive than corresponding leptospiral DNA-based quantitative PCR assays, targeting the same 16S nucleotide sequence in the RNA and DNA molecules. The sensitivity and specificity of our RNA assay against laboratory-confirmed human leptospirosis clinical samples were 64% and 100%, respectively, which was superior then an established parallel DNA detection assay. Remarkably, we discovered that 16S transcripts remain appreciably stable ex vivo, including untreated and stored human blood samples, further highlighting their use for clinical detection of L. interrogans. Together, these studies underscore a novel utility of RNA targets, specifically 16S rRNA, for development of PCRbased modalities for diagnosis of human leptospirosis, and also may serve as paradigm for detection of additional bacterial pathogens for which early diagnosis is warranted.

Research paper thumbnail of Patterns in Leptospira Shedding in Norway Rats (Rattus norvegicus) from Brazilian Slum Communities at High Risk of Disease Transmission

PLOS Neglected Tropical Diseases, 2015

We address some critical but unknown parameters of individuals and populations of Norway rats (Ra... more We address some critical but unknown parameters of individuals and populations of Norway rats (Rattus norvegicus) that influence leptospiral infection, maintenance and spirochetal loads shed in urine, which contaminates the environment ultimately leading to human infection.

Research paper thumbnail of Proteomic Features Predict Seroreactivity against Leptospiral Antigens in Leptospirosis Patients

Journal of proteome research, Jan 11, 2014

With increasing efficiency, accuracy, and speed we can access complete genome sequences from thou... more With increasing efficiency, accuracy, and speed we can access complete genome sequences from thousands of infectious microorganisms; however, the ability to predict antigenic targets of the immune system based on amino acid sequence alone is still needed. Here we use a Leptospira interrogans microarray expressing 91% (3359) of all leptospiral predicted ORFs (3667) and make an empirical accounting of all antibody reactive antigens recognized in sera from naturally infected humans; 191 antigens elicited an IgM or IgG response, representing 5% of the whole proteome. We classified the reactive antigens into 26 annotated COGs (clusters of orthologous groups), 26 JCVI Mainrole annotations, and 11 computationally predicted proteomic features. Altogether, 14 significantly enriched categories were identified, which are associated with immune recognition including mass spectrometry evidence of in vitro expression and in vivo mRNA up-regulation. Together, this group of 14 enriched categories a...