Daniel Mousain

The ectomycorrhizal symbiosis between Lactarius deliciosus and Pinus sylvestris in forest soil samples: symbiotic efficiency and development on roots of a rDNA internal transcribed spacer-selected isolate of L. deliciosus.

Abstract. The effect on plant growth of pre-inoculation of Pinus sylvestris with the ectomycorrhizal (ECM) edible basidiomycete Lactarius deliciosus (isolate D45) under controlled conditions, and the development on roots of this basidiomycete, were investigated in γ-irradiated and unsterilized containers containing different forest soil cores or a perlite-vermiculite mixture. Five months after planting, L. deliciosus mycorrhizal plants exhibited greater growth than the non-mycorrhizal ones in all soil types, i.e. up to a 325% increase in shoot height in the sterilized soils. The experiment demonstrated the dependency of P. sylvestris seedlings upon ECM symbiosis for their survival in γ-irradiated, microbiologically disturbed soil samples. Furthermore, in two soils, the growth of L. deliciosus-inoculated seedlings was greater in the sterilized soil samples than in the non-sterilized ones, i.e. 46% and 132% increase in shoot height under sterilized soil conditions. In containers randomly sampled from each soil type, the degree of root colonization by the inoculated isolate, calculated as the number of mycorrhizal root tips divided by the total number of root tips ×100, ranged from 80% to 35%. Within the short term, the inoculated isolate developed rapidly on roots, dominated, and hampered ectomycorrhiza formation by various unidentified (but not Lactarius) resident ECM fungi in unsterilized soil types. Results indicate that the ECM species L. deliciosus is worth investigating to ascertain if other isolates benefit pine growth like the isolate D45, and are therefore also attractive candidates for forestry applications in the Mediterranean area.

Bacteria of the genus Rhodopseudomonas (Bradyrhizobiaceae) : obligate symbionts in mycelial cultures of the black truffles Tuber melanosporum and Tuber brumale

BackgroundThis work aimed at characterizing 12 isolates of the genus Tuber including Tuber melanosporum (11 isolates) and Tuber brumale (one isolate). This was done using internal transcribed spacer (ITS) sequences, confirming their origin.ResultsAnalysis of their mating type revealed that both MAT1-1 and MAT1-2 exist within these isolates (with 3 and 8 of each, respectively). We observed that each of these cultures was consistently associated with one bacterium that was intimately linked to fungal growth. These bacterial associates failed to grow in the absence of fungus. We extracted DNA from bacterial colonies in the margin of mycelium and sequenced a nearly complete 16S rDNA gene and a partial ITS fragment. We found they all belonged to the genus Rhodopseudomonas, fitting within different phylogenetic clusters. No relationships were evidenced between bacterial and fungal strains or mating types. Rhodopseudomonas being a sister genus to Bradyrhizobium, we tested the nodulation ability of these bacteria on a promiscuously nodulating legume (Acacia mangium), without success. We failed to identify any nifH genes among these isolates, using two different sets of primers.ConclusionsWhile the mechanisms of interaction between Tuber and Rhodopseudomonas remain to be elucidated, their interdependency for in vitro growth seems a novel feature of this fungus.

Intraspecific variability of Pisolithus spp. as a response to changes in soil characteristics in a Moroccan cork oak plantation

Soil characteristics may be an important factor in structuring fungal communities. We relate the above- and below-ground distribution of the ectomycorrhizal fungi (EMF) Pisolithus spp. to changes in soil chemistry in a Quercus suber plantation located in the Maâmora forest (Morocco). Intrageneric variability of 115 fruit bodies was studied, using morphological characterization, restriction fragment length polymorphism (RFLP) analyses and internal transcribed spacer (ITS) sequencing. Two Pisolithus spp. genotypes were identified: 97% of the fruit bodies represent Pisolithus arhizus; the remaining 3% correspond to Pisolithus species 4, as previously described by Martin et al. [New Phytologist 153 (2002) 345-357]. Multivariate analysis (PCA) showed that species structure was strongly correlated with soil characteristics. P. arhizus fruit bodies were frequently found in the Eastern part of the plot at low molasic clay, N and P contents, while Pisolithus species 4 were collected only in the Western part at the high molasic clay, N and P contents. To determine whether this change in fruit body structure was expressed at the belowground level, we used morphological and molecular techniques to characterize the Pisolithus ectomycorrhizas. Only ectomycorrhizas of P. arhizus were identified at the low P–N–molasic clay site and disappeared completely at the high P–N–red clay site, where no Pisolithus species 4 mycorrhizas were found. However, autocorrelation among soil parameters makes it difficult to isolate the effects of individual parameters. These results show the local-scale impact of natural spatial heterogeneity on an ectomycorrhizal fungal genus.

Molecular phylogeny of Pisolithus species from Moroccan forest woodlands

The phylogenetic relationships among 200 Pisolithus basidiomata collected from pine, oak, and eucalypt forests and rockrose scrubs in Morocco were investigated. Using PCR-RFLP analysis of the internal transcribed spacer (ITS) of ribosomal DNA, this collection could be divided into 5 groups, by using PCR-RFLP analysis of the internal transcribed spacer (ITS) of ribosomal DNA. The ITS of a representative basidioma of each group was sequenced, and a phylogenetic analysis was performed. The dendrogram suggests that at least five Pisolithus species are present in Morocco. Pisolithus basidiomata collected in the Pinus and Quercus forests correspond to Pisolithus arrhizus and P. species 4 as previously described by Martin and colleagues in 2002. Those collected from the eucalyptus forests, under E. gomphocephala and E. camaldulensis, correspond to P. albus and P. microcarpus. Basidiomata collected from the rockrose scrubs, under Cistus crispus, C. monspeliensis or C. salviifolius, are all identified as Pisolithus species 3. Phylogenetic analyses showed that our different Pisolithus grouped well with Pisolithus specimens from other geographical origins.