Mechanistic dose-response modelling of animal challenge data shows that intact skin is a crucial barrier to leptospiral infection

Leptospirosis is a widespread and potentially life-threatening zoonotic disease caused by spirochaetes of the genus Leptospira. Humans become infected primarily via contact with environmental reservoirs contaminated by the urine of shedding mammalian hosts. Populations in high transmission settings, such as urban slums and subsistence farming communities, are exposed to low doses of Leptospira on a daily basis. Under these conditions, numerous factors determine whether infection occurs, including the route of exposure and inoculum dose. Skin wounds and abrasions are risk factors for leptospirosis, but it is not known whether broken skin is necessary for spillover, or if low-dose exposures to intact skin and mucous membranes can also cause infection. To establish a quantitative relationship between dose, route and probability of infection, we performed challenge experiments in hamsters and rats, developed mechanistic dose-response models representing the spatial dynamics of within-host infection and persistence, and fitted models to experimental data. Results show intact skin is a strong barrier against infection, and that broken skin is the predominant route by which low-dose environmental exposures cause infection. These results identify skin integrity as a bottleneck to spillover of Leptospira and underscore the importance of barrier interventions in the prevention of leptospirosis. This article is part of the theme issue 'Dynamic and integrative approaches to understanding pathogen spillover'

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

Submitted by Ana Maria Fiscina Sampaio ([email protected]) on 2015-07-08T17:32:08Z No. of bitstreams: 1 Costa F Patterns....pdf: 334080 bytes, checksum: 20a586e75f720e668fc7fe9bf6aa27c6 (MD5)Approved for entry into archive by Ana Maria Fiscina Sampaio ([email protected]) on 2015-07-08T17:56:52Z (GMT) No. of bitstreams: 1 Costa F Patterns....pdf: 334080 bytes, checksum: 20a586e75f720e668fc7fe9bf6aa27c6 (MD5)Made available in DSpace on 2015-07-08T17:56:52Z (GMT). No. of bitstreams: 1 Costa F Patterns....pdf: 334080 bytes, checksum: 20a586e75f720e668fc7fe9bf6aa27c6 (MD5) Previous issue date: 2015Fundação Oswaldo Cruz. Centro de Pesquisas Gonçalo Moniz. Salvador, BA, Brasil / Universidade Federal da Bahia. Instituto de Saúde Coletiva. Salvador, BA, Brasil / Yale School of Public Health. Department of Epidemiology of Microbial Diseases. New Haven, Connecticut, USA / University of Liverpool. Institute of Integrative Biology. United KingdomYale School of Public Health. Department of Epidemiology of Microbial Diseases. New Haven, Connecticut, USAFundação Oswaldo Cruz. Centro de Pesquisas Gonçalo Moniz. Salvador, BA, BrasilYale School of Public Health. Department of Epidemiology of Microbial Diseases. New Haven, Connecticut, USAMinistério da Saúde. Secretaria Municipal de Saúde. Centro de Controle do Zoonoses. Salvador, BA, BrasilFundação Oswaldo Cruz. Centro de Pesquisas Gonçalo Moniz. Salvador, BA, Brasil / Yale School of Public Health. Department of Epidemiology of Microbial Diseases. New Haven, Connecticut, USAFundação Oswaldo Cruz. Centro de Pesquisas Gonçalo Moniz. Salvador, BA, Brasil / Yale School of Public Health. Department of Epidemiology of Microbial Diseases. New Haven, Connecticut, USAUniversity of Liverpool. Institute of Integrative Biology. United KingdomYale School of Public Health. Department of Epidemiology of Microbial Diseases. New Haven, Connecticut, USABACKGROUND: 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. METHODOLOGY/PRINCIPAL FINDINGS: Our study, conducted in Salvador, Brazil, established the average load of leptospires in positive kidneys to be 5.9 x 106 per mL (range 3.1-8.2 x106) genome equivalents (GEq), similar to the 6.1 x 106 per ml (range 2.2-9.4 x106) average obtained from paired urines, with a significant positive correlation (R2=0.78) between the two. Based on bivariate and multivariate modeling, we found with both kidney and urine samples that leptospiral loads increased with the age of rats (based on the index of body length to mass), MAT titer and the presence of wounding/scars, and varied with site of capture. Some associations were modified by sex but trends were apparent. Combining with data on the demographic properties and prevalence of leptospiral carriage in rat populations in Salvador, we estimated that daily leptospiral loads shed in the urine of a population of 82 individuals exceeded 9.1 x 1010 leptospires. CONCLUSIONS/SIGNIFICANCE: These factors directly influence the risk of leptospiral acquisition among humans and provide essential epidemiological information linking properties of rat populations with risk of human infection

Correlation between the GEq load of leptospires in the paired kidney and urine samples of <i>Rattus norvegicus</i> from Brazil, 2010.

<p>Correlation between the GEq load of leptospires in the paired kidney and urine samples of <i>Rattus norvegicus</i> from Brazil, 2010.</p

Bivariate analyses of <i>Leptospira</i> load in kidney and urine of wild Norway rats.

<p>¹Standard Deviation</p><p>²Bold items reflect significant differences (P<0.05 ANOVA adjusted by Bonferroni correction method).</p><p>Values given are log10 genome equivalents per mm³ of kidney or per ml of urine.</p><p>Bivariate analyses of <i>Leptospira</i> load in kidney and urine of wild Norway rats.</p

Estimates of Leptospira shedding loads in urine from 82 Norway rats based on properties of the demographic structure of the rat population from Salvador, Brazil.

<p>¹ Number of rats in each mass/age class (NR.)</p><p>²<i>Leptospira</i> prevalence in kidney (PREV: <a href="http://www.plosntds.org/article/info:doi/10.1371/journal.pntd.0003819#pntd.0003819.t001" target="_blank">Table 1</a>).</p><p>³ Volume (ml) of urine shed per 24 hours (VOL) as described by Donaldson [<a href="http://www.plosntds.org/article/info:doi/10.1371/journal.pntd.0003819#pntd.0003819.ref043" target="_blank">43</a>].</p><p>⁴ Genomic equivalents of <i>Leptospira</i> per ml (LOAD: <a href="http://www.plosntds.org/article/info:doi/10.1371/journal.pntd.0003819#pntd.0003819.t001" target="_blank">Table 1</a>).</p><p>⁵ Based on density of rats (DENS) captured around households.</p><p>Estimates of Leptospira shedding loads in urine from 82 Norway rats based on properties of the demographic structure of the rat population from Salvador, Brazil.</p

Multivariate linear regression models of the relation between Leptospira load in kidneys and urine.

<p>¹AIC excluding MAT = 93.5</p><p>²AIC excluding MAT = 150.7</p><p>³AIC excluding MAT = 92.4</p><p>⁴AIC excluding MAT = 86.4</p><p>⁵Bold items reflect significant differences (P<0.05).</p><p>Multivariate linear regression models of the relation between Leptospira load in kidneys and urine.</p

Lvr, a Signaling System That Controls Global Gene Regulation and Virulence in Pathogenic Leptospira

Leptospirosis is an emerging zoonotic disease with more than 1 million cases annually. Currently there is lack of evidence for signaling pathways involved during the infection process of Leptospira. In our comprehensive genomic analysis of 20 Leptospira spp. we identified seven pathogen-specific Two-Component System (TCS) proteins. Disruption of two these TCS genes in pathogenic Leptospira strain resulted in loss-of-virulence in a hamster model of leptospirosis. Corresponding genes lvrA and lvrB (leptospira virulence regulator) are juxtaposed in an operon and are predicted to encode a hybrid histidine kinase and a hybrid response regulator, respectively. Transcriptome analysis of lvr mutant strains with disruption of one (lvrB) or both genes (lvrA/B) revealed global transcriptional regulation of 850 differentially expressed genes. Phosphotransfer assays demonstrated that LvrA phosphorylates LvrB and predicted further signaling downstream to one or more DNA-binding response regulators, suggesting that it is a branched pathway. Phylogenetic analyses indicated that lvrA and lvrB evolved independently within different ecological lineages in Leptospira via gene duplication. This study uncovers a novel-signaling pathway that regulates virulence in pathogenic Leptospira (Lvr), providing a framework to understand the molecular bases of regulation in this life-threatening bacterium

Table7.XLS

<p>Leptospirosis is an emerging zoonotic disease with more than 1 million cases annually. Currently there is lack of evidence for signaling pathways involved during the infection process of Leptospira. In our comprehensive genomic analysis of 20 Leptospira spp. we identified seven pathogen-specific Two-Component System (TCS) proteins. Disruption of two these TCS genes in pathogenic Leptospira strain resulted in loss-of-virulence in a hamster model of leptospirosis. Corresponding genes lvrA and lvrB (leptospira virulence regulator) are juxtaposed in an operon and are predicted to encode a hybrid histidine kinase and a hybrid response regulator, respectively. Transcriptome analysis of lvr mutant strains with disruption of one (lvrB) or both genes (lvrA/B) revealed global transcriptional regulation of 850 differentially expressed genes. Phosphotransfer assays demonstrated that LvrA phosphorylates LvrB and predicted further signaling downstream to one or more DNA-binding response regulators, suggesting that it is a branched pathway. Phylogenetic analyses indicated that lvrA and lvrB evolved independently within different ecological lineages in Leptospira via gene duplication. This study uncovers a novel-signaling pathway that regulates virulence in pathogenic Leptospira (Lvr), providing a framework to understand the molecular bases of regulation in this life-threatening bacterium.</p

Table8.DOC

<p>Leptospirosis is an emerging zoonotic disease with more than 1 million cases annually. Currently there is lack of evidence for signaling pathways involved during the infection process of Leptospira. In our comprehensive genomic analysis of 20 Leptospira spp. we identified seven pathogen-specific Two-Component System (TCS) proteins. Disruption of two these TCS genes in pathogenic Leptospira strain resulted in loss-of-virulence in a hamster model of leptospirosis. Corresponding genes lvrA and lvrB (leptospira virulence regulator) are juxtaposed in an operon and are predicted to encode a hybrid histidine kinase and a hybrid response regulator, respectively. Transcriptome analysis of lvr mutant strains with disruption of one (lvrB) or both genes (lvrA/B) revealed global transcriptional regulation of 850 differentially expressed genes. Phosphotransfer assays demonstrated that LvrA phosphorylates LvrB and predicted further signaling downstream to one or more DNA-binding response regulators, suggesting that it is a branched pathway. Phylogenetic analyses indicated that lvrA and lvrB evolved independently within different ecological lineages in Leptospira via gene duplication. This study uncovers a novel-signaling pathway that regulates virulence in pathogenic Leptospira (Lvr), providing a framework to understand the molecular bases of regulation in this life-threatening bacterium.</p