|Biology of Spirochetes|
|HEAD||Dr PICARDEAU Mathieu / firstname.lastname@example.org|
|MEMBERS||Researcher PICARDEAU Mathieu IP Associate Professor, PhD BENAROUDJ Nadia IP Assistant Professor, PhD
Research Engineer BOURHY Pascale IP Engineer, PhD Deputy manager of the NRC for Leptospira
Research fellows LOURDAULT Kristel PhD student WUNDER Elsio PhD student SLAMTI Leyla postdoc student De OLIVERIA Lucas master student
Other staff ZININI Farida IP Technician COUEILLE Solange IP Secretary BREMONT Sylvie IP Technician LANDIER Annie IP Technician ETIEMBLE Sylviane IP Technical staff
Spirochetes are the causative agents of several important animal and human diseases such as syphillis, Lyme disease, and leptospirosis. The unit is composed of a research group on Leptospira and the National Reference Center (NRC) for Leptospirosis which is also a WHO collaborating Center.
In 2010, the main research activities included:
i) Genetics and Genomics in Leptospira
In comparison to other bacterial species, genetics and molecular basis of the pathogenesis of leptospires are in their infancy. We recently provided the first evidence of gene transfer in a pathogenic Leptospira strain. However, the further development and improvement of genetic tools are required. We reported the first demonstration of conjugation between E. coli and Leptospira spp. using RP4 derivative conjugative plasmids. >In 2010, we have transferred fluorescent alleles (gfp and mRFP1) in virulent strains by using the Himar1 transposon (there is no replicative vector available in the pathogens). Generation of transcriptional fusions placing GFP expression under the control of a bacterial promoter can be a valuable tool to examine gene expression patterns during Leptospira infection processes or in in vitro conditions. Imaging technologies should afford a powerful approach to visualizing the infection process, determining the tissue specificity of infection, and the spatial migration of the bacteria.
Electronic microscopy of Leptospira biflexa
The genus Leptospira has both saprophytic and pathogenic members, such as L. biflexa and L. interrogans, respectively. The recent completion of the genome sequences of the pathogen L. interrogans and L. borgpetersenii has provided a basis for understanding these organisms. A major challenge remains the assignment of a biological role to the predicted genes, more than 50 % of which show no similarity to proteins of known function. However, the slow growth of these bacteria, their virulence in humans and a lack of genetic tools make it difficult to work with these pathogens. We therefore sequenced the genome of the saprophyte L. biflexa serovar Patoc strain Patoc1. T>he ease of genetic manipulation in the model bacterium L. biflexa provides us with an ideal opportunity for studies of the biology of Leptospira spp.
ii) Identification of virulence factors
Comparison of the genome sequence of pathogenic and saprophytic Leptospira strains provides an opportunity to identify features that are unique to pathogenic species. The L. interrogans genome contain approximately 3400 predicted coding regions, of which 656 are pathogen-specific genes, i.e. genes not present in the saprophyte L. biflexa. The majority of these genes has no known function, suggesting the presence of pathogenic mechanisms unique to Leptospira. We have obtained random insertion of Himar1 in the pathogen L. interrogans. We found mutants with transposon insertion in genes encoding putative virulence factors. These mutants showed decrease virulence compared to the wild-type strain in the animal model of leptospirosis. This library of random mutants provides an opportunity to investigate genes that contribute to pathogenesis and will provide a better understanding of the biology of L. interrogans. We recently generated a mutant by allelic exchange in L. interrogans, wherein a spectinomycin resistance gene replaced a portion of the coding sequence. Our data provides the first evidence for site-directed homologous recombination in pathogenic Leptospira.
iii) Molecular typing of pathogenic leptospires
The main function of the NRC for leptospirosis, which is also a WHO Collaborating Center, is the surveillance of this zoonosis. The NRC performed more than 4000 diagnosis every year by bacteriological and serological means. It enables the NRC to update knowledge of the epidemiological traits of leptospirosis and the situation of the disease regularly, both in mainland France and in French overseas territories. For example, we investigated an outbreak of leptospirosis after a raid held on the island of Martinique in the Caribbean island in 2009. We also performed epidemiological investigations of leptospirosis in regions of high endemicity such as in Mayotte (Indian Ocean) where we reported the existence of ten potentially new pathogenic Leptospira genotypes. The NRC has also a duty to improve diagnostic techniques. We have developed a rapid and simple typing method based on analysis of the polymorphism of variable-number tandem repeats (VNTR) loci in the pathogen genome
Keywords: Spirochetes, Leptospira, epidemiology, genetics, virulence factors
Ristow, P., P. Bourhy , F.W. McBride, C.P. Figueira, M. Huerre, P. Ave, I.S. Girons, A.I. Ko, and M. Picardeau (2007). The OmpA-like protein Loa22 is essential for leptospiral virulence. PLoS Pathog. 3(7):e97
M. Picardeau, D.M. Bulach, C. Bouchier, R. L. Zuerner, N. Zidane, P. J. Wilson, S. Creno, E. S. Kuczek, S. Bommezzadri, J. C. Davis, A. McGrath, D. Roche, M. J. Johnson, C. Boursaux-Eude, T. Seemann, Z. Rouy, J. I. Rood, A. Lajus, J. K. Davies, C. Midigue, and B. Adler (2008). Genome sequence of the saprophyte Leptospira biflexaprovides insights into the evolution of Leptospiraand the pathogenesis of leptospirosis. PLoS ONE 3(2):e1607.
G. L. Murray, V. Morel, G.M. Cerqueira, J. Croda, A. Srikram, R. Henry, A.I. Ko, O.A. Dellagostin, D.M. Bulach, R. Sermswan, B. Adler, and M. Picardeau (2009). Genome-wide transposon mutagenesis in pathogenic Leptospiraspp.. Infect. Immun. 77:810-816.. A. I. Ko, C. Goarant, and M. Picardeau (2009). Leptospira: The dawn of the molecular genetics era for an emerging zoonotic pathogen. Nature Rev. Microbiol 7:736-47.
P. Bourhy, L. Collet, S. Climent, M. Huerre, P. Ave, C. Giry, F. Pettinelli, and M. Picardeau (2010). Isolation and characterization of new Leptospira genotypes from patients in Mayotte (Indian Ocean).. PLoS Negl Trop Dis. 2010 4:e724.
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Activity Reports 2010 - Institut Pasteur
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