Snail / schistosome compatibility Réunion Scientifique mardi 3 Avril 2007 Snail / schistosome compatibility from populations to mechanisms André THERON UMR 5244 BIOLOGIE ET ECOLOGIE TROPICALE ET MEDITERRANEENNE University of Perpignan - 66860 Perpignan - France

Mollusc / Trematode models life cycles Schistosoma mansoni Biomphalaria glabrata

Mollusc / Trematode models Schistosome life cycle Schistosoma mansoni Adults Egg Miracidium Cercariae Biomphalaria glabrata

Snail infection in Field populations Most of the field surveys on natural populations of Biomphalaria snails report : The typically low Infection rates variations of population size high mortality rates genetic bottleneck (extinction / recolonisation, founder effect) dry season (1-5%) rainy season rainy season

? 1 2 Ecologie moléculaire de la transmission : 100% 1% recrutement et diversité génétique Génotypage intra-hôte : - parasites adultes HD (microsatellites) - Parasite larvaires HI (microsatellites) 1 2 100% Limited contact ? RESISTANT/ SUSCEPTIBLE 1% Théron et al., 2004. Parasitology.

Resistance / Susceptibility 2 Polymorphism of host susceptibility/resistance SUSCEPTIBLE HOST RESISTANT HOST Laboratory strains Experimental infections 50 - 60 % 0 % 50 % 100 % 50 40 30 20 10 1 2 nMi Selected «RESISTANT» strain selected «SUSCEPTIBLE» strain

Selected “Resistant“strains: 2 Selected “Resistant“strains: EVOLUTIONARY IMPLICATIONS >> selected «RESISTANT» strain … Evolutionary inferences (Co-evolution, Life history and Red Queen theories, …) Routinely used to investigate : host defence mechanisms Genetics of resistance Genes of resistance Resistance is heritable and dominant (Richards et al., 1992 ). Molecular markers associated with resistance (Spada et al., 2002 ). Mating behaviour associated with resistance (Webster et al., 2003 ). Maintenance of resistance/susceptibility polymorphism Fitness Cost associated with resistance (Webster & Woolhouse, 2001 ). trade-offs between the FITNESS COST associated with RESISTANCE (reduced fertility) and the fitness costs resulting from parasitism (increased mortality). (Webster & Davis, Parasitology , 123, 2001)

Are Biomphalaria snails RESISTANT to Schistosoma mansoni ? Laboratory selected “resistant“strains: use and limitation. However, despite extensive efforts and use of various complementary biomolecular approaches, no linkage group, no genetic or functional marker has been assigned to any locus controlling resistance. Théron & Coustau (J. Helminthol., 2005) raised the question : Are Biomphalaria snails RESISTANT to Schistosoma mansoni ? based on some arguments …

B. glabrata resistance : a relative concept SUCCESSFUL INFECTION Developed sporocyst SUSCEPTIBLE HOST RESISTANT HOST Encapsulated sporocysts FAILURE of INFECTION

We conclude that … Compatibility is tested independently for each entering miracidium. The phenotype (resistant vs susceptible) of the host is expressed as a function of the parasite genotype it harbors. The phenotype (un-infective vs infective) of the parasite is expressed as a function of the host genotype it enters. This means that the CONCEPT of RESISTANCE (and its evolutionary implication) appears un-adapted to the B. glabrata / S. mansoni system

2 Apparent Resistance Warning Selected “Resistant“strains: EVOLUTIONARY IMPLICATIONS >> “Resistant“ selected Apparent Resistance Resistance is heritable and dominant (Richards et al., 1992 ). Molecular markers associated with resistance (Spada et al., 2002 ). Mating behaviour associated with resistance (Webster et al., 2003 ). Fitness Cost associated with resistance (Webster & Woolhouse, 2001 ). Warning … Evolutionary inferences (co-evolution, life history and Red Queen theories, …)

Polymorphism of compatibility “Resistance“ / Infectivity in Field populations 2 RESISTANT/ SUSCEPTIBLE Going back to the field ……, 3 Genotype-by-genotype Interaction Polymorphism of compatibility Compatible Un-Compatible GENETIC DIVERSITY

- + + - + 3 Compatibility polymorphism and B. glabrata « resistance » in the field ? Genotype-by-Genotype interaction Matching genotype Snail genotypic diversity - + + - Parasite genotypic diversity 100 % +

Populational validation exposed snails directly collected in the site S. mansoni eggs obtained from a substantial number of naturally infected vertebrate hosts 100 % compatibility - increasing miracidial dose exposures 0 % 50 % 100 % 50 40 30 20 10 1 2 nMi Dans Fond Dubelloy Belle Plaine

We conclude that … Among Biomphalaria and S. mansoni associations, susceptibility of snails in the field is general if we take into account the total amount of genetic diversity present within the whole population of parasites within vertebrate hosts. Snail genetic diversity Parasite genetic diversity Field situation 100 % matching Meeting probability + Matching probability 1-5% infection rates

Apparent resistance in Laboratory strains … Snail genetic diversity Parasite genetic diversity Field situation 100 % matching Laboratory Strains 0 à 60% matching

The functional point of view : Schistosomes use molecular mimicry as infectivity strategy and are confronted to self/non-self recognition processes of the host (general immune defence) Host Host success Matched Molecular Variants failure Un-Matched Molecular Variant Parasite Parasite A genetically determined mechanism theoretically consistent with a matching genotype model

? from populations to mechanisms (André THERON) CONCLUSION … Matched vs Un-matched status of host-parasite combinations : What phenotypic factors interact between host and parasite ? ? What genomic mechanisms generate diversity ? from populations to mechanisms (André THERON) from mechanisms to populations (Guillaume MITTA)

Biomphalaria glabrata Sporocyste I installé chez le mollusque Recherche des gènes acteurs de la compatibilité: Approches moléculaires comparatives entre souches de S. mansoni Biomphalaria glabrata Sporocyste I installé chez le mollusque Sporocyste I en dégénérescence Capsule hémocytaire Compatible Incompatible Schistosoma mansoni

Mother Sporocyst Polymorphic Proteins (MSPPs) Approche protéomique comparative et identification de candidats prometteurs, les MSPPs analogies de structure pour des protéines immuno-dominantes de Plasmodium et Coccidioides : rôle «écran de fumée» Parasites compatibles Parasites incompatibles Mw Mw 250 250 150 150 100 100 75 75 50 50 37 37 25 25 20 20 Mother Sporocyst Polymorphic Proteins (MSPPs) 15 15 pI (NL) pI (NL) 3 10 3 10 répétitions de 9 aa en tandem 234 aa Etc.. O-glycosylations

Les MSPPs et les variants au niveau cDNA REPET B: GGTGACCTCGCATCAGACAAACCCACA B G D L A S D K P T REPET BMOD: GGTGGCCTCGCATCAGACAAACCCACA B G G L A S D K P T REPET G: GATGACTATGCATTGAGCGAACCAACA G D D Y A L S E P T REPET GMOD: GATTACTATGCATTGAGCGAACCAACA G D Y Y A L S E P T REPET M: GGTGACCTCGCATTAGACGAACCAACA M G D L A L D E P T Compatible Incompatible BBBBBBBB BBBBBBBB BBBBBBBBBBB BBBBBBBBBBBBBBB BBBBBB BBBBBBBBBBBBBBBBBBBBBBBBBB GGBBBBBBBBBBB BBBBBBBBBBBBBBBBBBBB GGBBGG BBBBBBBBBBBBBBBBBBBB GGBBGG BBBBBB GGBBG BBBBB GGBBBBG GGGGGGGGGGG GGMGG GGGGGGGGG GG GGGG GGGGGGGGGGG GGG GGGGGGGG GG GGGGG GG

Les MSPPs: ume famille multigénique 3 gènes MSPP1a 2 3 4 5 6 7 8 9 10 11 12 13 14 15 MSPP2a 2 3 4 5 6 7 8 9 10 11 12 13 14 15 MSPP3 2 3 4 5 6 7 8 9 10 11 12 13 14 15 8 pseudogènes exons MSPP1b 2 3 4 5 6 7 8 9 10 MSPP2b 13 * 4 5 6 7 8 9 10 11 12 14 15 MSPP4 2 3 4 5 6 7 8 9 10 MSPP5 2 3 7 8 9 10 MSPP6 2 3 3 4 MSPP7 9 10 11 12 13 14 15 MSPP8 11 12 13 14 15 MSPP9 5 6 11 12 13 14 15 Gènes soumis à des insertions, remaniements fréquents médiés par ces RT Retrotransposons (RT)

Le modèle B. glabrata/S. mansoni: les MSPPs des candidats de choix - Protéines potentiellement impliquées dans le contournement du système de défense de B. glabrata par S. mansoni - Polymorphisme et organisation génomique génératrice de diversité Candidats de choix en tant que facteurs clefs de la compatibilité Candidats pour une 1ère « approche populationnelle »

Polymorphisme de compatibilité et MSPPs en populations naturelles Compatible Compatibles Pop Nat Labo Guadeloupe Brésil Comparaison des MSPPs: Protéines « actrices » de la compatibilité en populations naturelles? 4% Isolats de S. mansoni de Guadeloupe 6%