Diversity, origin and characterisation of fungal communities associated to fungus-growing termite (Isoptera, Termitidae) combs

La diversité fongique des meules de plusieurs espèces de Macrotermitinae a été analysée au niveau taxinomique, fonctionnel et génétique à l’aide d’une approche polyphasique associant plusieurs techniques complémentaires. L’objectif étant d’évaluer la spécificité des taxons fongiques associés aux meules ainsi que les relations qu’ils entretiennent avec les termites champignonnistes. Une grande variété de phylotypes cultivables appartenant majoritairement au phylum des Ascomycètes a été obtenue par isolement et séquençage des ITS fongiques, et peu de séquences se sont révélées être spécifiques à un genre de Macrotermitinae particulier. Les profils physiologiques obtenus ont mis en évidence la nature saprophytique de la majorité des phylotypes et confirmé l’absence de taxons spécifiques. Par PCR-DGGE de l’ADN total de meules, 100% des phylotypes ITS et 28S fongiques identifiés étaient affiliés au genre Termitomyces. La technique Suicide Polymerase Endonuclease Restriction (SuPER) a été adaptée à la mycoflore des meules pour limiter l’impact de l’ADN majoritaire du Termitomyces symbiotique. Celle-ci a permis la détection de plusieurs autres populations fongiques. Les analyses phylogénétiques ont montré d’une part la spécificité des Xylaria associés aux meules de Macrotermitinae bien qu’aucune co-évolution n’ait été observée avec les termites hôtes et d’autre part leur affiliation dans un sous-genre spécifique. Une analyse préliminaire des facteurs d’inhibition a également révélé l’implication des termites dans la régulation des communautés fongiques des meules. Dans leur ensemble, nos résultats illustrent clairement l’influence des Macrotermitinae sur les communautés fongiques telluriques pendant leurs différentes activités.Fungal diversity of several Macrotermitinae fungus combs was analyzed at taxonomic, functional and genetic levels using a polyphasic approach. The aim of this thesis was to evaluate the specificity of fungal strains from combs and to elucidate their relationship with fungus-growing termites. A large variety of culturable phylotypes mainly belonging to Ascomycota phylum was retrieved using conventional isolation techniques followed by sequencing of ITS1-5.8S-ITS2 region. Based on the obtained results, there is evidence for any speciesspecificity between these taxa and a given genus of Macrotermitinae. This finding was supported by the physiological profile of some representative phylotypes which revealed the saprophytic nature of most of the isolates. By PCR-DGGE analysis of fungal ITS and LSU, all of the sequences were belonged to Termitomyces genus. The Suicide Polymerase Endonuclease Restriction method was adapted to the analysis of comb mycoflora for restricting the impact of the dominant Termitomyces DNA. As expected, this latter technique revealed non-Termitomyces fungal populations. Phylogenetic analysis also showed the specificity of termiteassociated Xylaria although they do not evolved with termite hosts, and also their affiliation to a new genus or at least a specific sub-genus. Preliminary investigation revealed the implication of termite workers in fungal regulation in fungus combs. All in one, our results clearly underline the great impact of fungus-growing termite species on soil fungal community during their activities

Diversité, origine et caractérisation de la mycoflore des meules de Macrotermitinae (Isoptera, Termitidae)

La diversité fongique des meules de plusieurs espèces de Macrotermitinae a été analysée au niveau taxinomique, fonctionnel et génétique à l aide d une approche polyphasique associant plusieurs techniques complémentaires. L objectif étant d évaluer la spécificité des taxons fongiques associés aux meules ainsi que les relations qu ils entretiennent avec les termites champignonnistes. Une grande variété de phylotypes cultivables appartenant majoritairement au phylum des Ascomycètes a été obtenue par isolement et séquençage des ITS fongiques, et peu de séquences se sont révélées être spécifiques à un genre de Macrotermitinae particulier. Les profils physiologiques obtenus ont mis en évidence la nature saprophytique de la majorité des phylotypes et confirmé l absence de taxons spécifiques. Par PCR-DGGE de l ADN total de meules, 100% des phylotypes ITS et 28S fongiques identifiés étaient affiliés au genre Termitomyces. La technique Suicide Polymerase Endonuclease Restriction (SuPER) a été adaptée à la mycoflore des meules pour limiter l impact de l ADN majoritaire du Termitomyces symbiotique. Celle-ci a permis la détection de plusieurs autres populations fongiques. Les analyses phylogénétiques ont montré d une part la spécificité des Xylaria associés aux meules de Macrotermitinae bien qu aucune co-évolution n ait été observée avec les termites hôtes et d autre part leur affiliation dans un sous-genre spécifique. Une analyse préliminaire des facteurs d inhibition a également révélé l implication des termites dans la régulation des communautés fongiques des meules. Dans leur ensemble, nos résultats illustrent clairement l influence des Macrotermitinae sur les communautés fongiques telluriques pendant leurs différentes activités.Fungal diversity of several Macrotermitinae fungus combs was analyzed at taxonomic, functional and genetic levels using a polyphasic approach. The aim of this thesis was to evaluate the specificity of fungal strains from combs and to elucidate their relationship with fungus-growing termites. A large variety of culturable phylotypes mainly belonging to Ascomycota phylum was retrieved using conventional isolation techniques followed by sequencing of ITS1-5.8S-ITS2 region. Based on the obtained results, there is evidence for any speciesspecificity between these taxa and a given genus of Macrotermitinae. This finding was supported by the physiological profile of some representative phylotypes which revealed the saprophytic nature of most of the isolates. By PCR-DGGE analysis of fungal ITS and LSU, all of the sequences were belonged to Termitomyces genus. The Suicide Polymerase Endonuclease Restriction method was adapted to the analysis of comb mycoflora for restricting the impact of the dominant Termitomyces DNA. As expected, this latter technique revealed non-Termitomyces fungal populations. Phylogenetic analysis also showed the specificity of termiteassociated Xylaria although they do not evolved with termite hosts, and also their affiliation to a new genus or at least a specific sub-genus. Preliminary investigation revealed the implication of termite workers in fungal regulation in fungus combs. All in one, our results clearly underline the great impact of fungus-growing termite species on soil fungal community during their activities.PARIS-EST-Université (770839901) / SudocPARIS12-Bib. électronique (940280011) / SudocSudocFranceF

Molecular diversity and host specificity of termite associated Xylaria

International audiencePrevious studies have revealed that some Xylaria species were closely associated with fungus-growing termite nests. However, this relationship had been very rarely investigated and the host-specificity of termite-associated Xylaria was not yet clearly established. 18 Xylaria rDNA-ITS sequences were obtained from fungus combs belonging to 11 Macrotermitinae species from 8 regions. There was low diversity between isolates and 9 different sequences were retrieved. Termite-associated Xylaria were shown to be monophyletic, with three main clades, all including strains from various termite hosts and geographical localities. This new molecular study shows that there is no species specificity with respect to fungus-growing termites which suggests that there may be substrate specialization

Occurrence of fungi in combs of fungus-growing termites (Isoptera: Termitidae, Macrotermitinae)

Fungus-growing termites cultivate their mutualistic basidiomycete Termitomyces species on a substrate called a fungal comb. Here, the Suicide Polymerase Endonuclease Restriction (SuPER) method was adapted for the first time to a fungal study to determine the entire fungal community of fungal combs and to test whether fungi other than the symbiotic cultivar interact with termite hosts. Our molecular analyses show that although active combs are dominated by Termitomyces fungi isolated with direct Polymerase Endonuclease Restriction - Denaturing Gradient Gel Electrophoresis (PCR-DGGE), they can also harbor some filamentous fungi and yeasts only revealed by SuPER PCR-DGGE. This is the first molecular evidence of the presence of non-Termitomyces species in active combs. However, because there is no evidence for a species-specific relationship between these fungi and termites, they are mere transient guests with no specialization in the symbiosis. It is however surprising to notice that termite-associated Xylaria strains were not isolated from active combs even though they are frequently retrieved when nests are abandoned by termites. This finding highlights the implication of fungus-growing termites in the regulation of fungi occurring within the combs and also suggests that they might not have any particular evolutionary-based association with Xylaria species