Marie-Mathilde Perrineau

Effects of Medicago truncatula Genetic Diversity, Rhizobial Competition, and Strain Effectiveness on the Diversity of a Natural Sinorhizobium Species Community

ABSTRACT We investigated the genetic diversity and symbiotic efficiency of 223 Sinorhizobium sp. isolates sampled from a single Mediterranean soil and trapped with four Medicago truncatula lines. DNA molecular polymorphism was estimated by capillary electrophoresis-single-stranded conformation polymorphism and restriction fragment length polymorphism on five loci (IGSNOD, typA, virB11, avhB11, and the 16S rRNA gene). More than 90% of the rhizobia isolated belonged to the Sinorhizobium medicae species (others belonged to Sinorhizobium meliloti), with different proportions of the two species among the four M. truncatula lines. The S. meliloti population was more diverse than that of S. medicae, and significant genetic differentiation among bacterial populations was detected. Single inoculations performed in tubes with each bacterial genotype and each plant line showed significant bacterium-plant line interactions for nodulation and N2 fixation levels. Competition experiments within each species highlighted either strong or weak competition among genotypes within S. medicae and S. meliloti, respectively. Interspecies competition experiments showed S. meliloti to be more competitive than S. medicae for nodulation. Although not highly divergent at a nucleotide level, isolates collected from this single soil sample displayed wide polymorphism for both nodulation and N2 fixation. Each M. truncatula line might influence Sinorhizobium soil population diversity differently via its symbiotic preferences. Our data suggested that the two species did not evolve similarly, with S. meliloti showing polymorphism and variable selective pressures and S. medicae showing traces of a recent demographic expansion. Strain effectiveness might have played a role in the species and genotype proportions, but in conjunction with strain adaptation to environmental factors.

Active dispersal by wild Triatoma infestans in the Bolivian Andes

Triatoma infestans is the main vector of Chagas disease and target of control programmes in the Southern Cone countries. So far Bolivia is the only country where true T. infestans wild foci are documented. The dispersal ability for wild T. infestans was studied at microgeographical scale in Bolivian Andes, to assess the possibility for wild populations to actively recolonize insecticide‐treated villages. Nine microsatellite loci were used to detect the extent of gene flow between neighbouring collecting sites. The detection of restricted gene flow between close but distinct sylvatic sites supports the hypothesis that wild T. infestans does not disperse by flying at high altitude (2 750 m asl). It gradually disperses over small distances by walking within a ‘patch’ of continuous land cover. The genetic differentiation detected between sylvatic and domestic populations suggests a limited short‐term role of wild insects in the process of recolonization of insecticide‐treated houses in the Andes.

Genetic diversity of symbiotic #Bradyrhizobium elkanii# populations recovered from inoculated and non-inoculated #Acacia mangium# field trials in Brazil

Acacia mangium is a legume tree native to Australasia. Since the eighties, it has been introduced into many tropical countries, especially in a context of industrial plantations. Many field trials have been set up to test the effects of controlled inoculation with selected symbiotic bacteria versus natural colonization with indigenous strains. In the introduction areas, A. mangium trees spontaneously nodulate with local and often ineffective bacteria. When inoculated, the persistence of inoculants and possible genetic recombination with local strains remain to be explored. The aim of this study was to describe the genetic diversity of bacteria spontaneously nodulating A. mangium in Brazil and to evaluate the persistence of selected strains used as inoculants. Three different sites, several hundred kilometers apart, were studied, with inoculated and non-inoculated plots in two of them. Seventy-nine strains were isolated from nodules and sequenced on three housekeeping genes (glnII, dnaK and recA) and one symbiotic gene (nodA). All but one of the strains belonged to the Bradyrhizobium elkanii species. A single case of housekeeping gene transfer was detected among the 79 strains, suggesting an extremely low rate of recombination within B. elkanii, whereas the nodulation gene nodA was found to be frequently transferred. The fate of the inoculant strains varied depending on the site, with a complete disappearance in one case, and persistence in another. We compared our results with the sister species Bradyrhizobium japonicum, both in terms of population genetics and inoculant strain destiny.

Bradyrhizobia Nodulating the Acacia mangium × A. auriculiformis Interspecific Hybrid Are Specific and Differ from Those Associated with Both Parental Species

ABSTRACT In the context of an increasing utilization of the interspecific hybrid Acacia mangium × A. auriculiformis as a plantation tree in the tropical humid zone, its symbiotic characterization was carried out in comparison with that of its two parental species. Rhizobium strains of diverse geographical origins were isolated from root nodules of the hybrid and its parents. Almost all Acacia hybrid isolates were fast growing on yeast extract-mannitol medium, in contrast to those isolated from both parental species, which were mostly slow growing. The rhizobium strains were characterized through partial sequencing of the rRNA operon. In the phylogenetic tree, almost all strains isolated from the hybrid were grouped together in a clade close to Bradyrhizobium japonicum, while all strains isolated from both parental species were close to Bradyrhizobium elkanii. Inoculation experiments performed under in vitro or greenhouse conditions showed that all strains were infective with their original hosts but exhibited very variable degrees of effectivity according to the host plant tested. Thus, homologous strain-host associations were more effective than heterologous ones. This shows that there is still a high potential for isolating and testing new strains from hybrids to be used as inoculants in the context of large-scale afforestation programs.

Monoxenic nodulation process of Acacia mangium (Mimosoideae, Phyllodineae) by Bradyrhizobium sp

Acacia mangium Willd., a native tree in Australia and Papua New Guinea, has been introduced to countries in Asia and South America where plantations have been established that cover several hundred thousand ha. The present study investigated the early stages of the nodulation process in A. mangium using an homologous Australian Bradyrhizobium strain. After optimizing the axenic nodulation, histological and cytological studies were conducted using light and electron microscopy. These documented the proliferation of Bradyrhizobium, the lysis of mucilage at the root surface, root hair deformation and initiation, as well as the development and growth of multiple infection threads. A belt of tannin-filled cells was shown to surround the central nodular fixation zone. The nodules were of the indeterminate type and the bacteroids had a rod shape, without size modification and with few polyhydroxybutyrate (PHB) granules. Several bacteroids can share the same symbiosome. A. mangium exhibits both classical and novel features in its nodulation.

Effective arbuscular mycorrhizal fungi in the roots of Tetraclinis articulata and Lavandula multifida in Moroccan Tetraclinis woodlands

The diversity of arbuscular mycorrhizal fungi (AMF) in roots of two plant species (arar, Tetraclinis articulata and lavender, Lavandula multifida) that exist alone or co-occur in Moroccan Tetraclinis woodlands was characterized using molecular techniques. The interaction between the two plant species on their root AMF communities was also studied. We analysed the large subunit (LSU) ribosomal RNA gene amplified by nested PCR from a root DNA extract using AM fungal-specific primers. A total of 200 cloned fragments from five root bulk samples of the two plant species were analysed by restriction fragment length polymorphism (RFLP) and 30 of them were sequenced. Phylogenetic analyses demonstrated that these RFLP types correspond to eight phylotypes: five belonged to the Gigasporaceae and three to the Glomeraceae. The highest diversity was found in the roots of T. articulata alone, followed by the association T. articulata/L. multifida and, finally, L. multifida alone. Many AMF were shared between the two species, although a few of them had a preference for one or the other. In addition, PCA analysis showed clearly that the AMF community colonizing T. articulata differed significantly from that colonizing L. multifida. Our results reveal an impact of plant interactions on the AMF community.

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.