A population genomics approach shows widespread geographical distribution of cryptic genomic forms of the symbiotic fungus Rhizophagus irregularis.

Arbuscular mycorrhizal fungi (AMF; phylum Gomeromycota) associate with plants forming one of the most successful microbe-plant associations. The fungi promote plant diversity and have a potentially important role in global agriculture. Plant growth depends on both inter- and intra-specific variation in AMF. It was recently reported that an unusually large number of AMF taxa have an intercontinental distribution, suggesting long-distance gene flow for many AMF species, facilitated by either long-distance natural dispersal mechanisms or human-assisted dispersal. However, the intercontinental distribution of AMF species has been questioned because the use of very low-resolution markers may be unsuitable to detect genetic differences among geographically separated AMF, as seen with some other fungi. This has been untestable because of the lack of population genomic data, with high resolution, for any AMF taxa. Here we use phylogenetics and population genomics to test for intra-specific variation in Rhizophagus irregularis, an AMF species for which genome sequence information already exists. We used ddRAD sequencing to obtain thousands of markers distributed across the genomes of 81 R. irregularis isolates and related species. Based on 6 888 variable positions, we observed significant genetic divergence into four main genetic groups within R. irregularis, highlighting that previous studies have not captured underlying genetic variation. Despite considerable genetic divergence, surprisingly, the variation could not be explained by geographical origin, thus also supporting the hypothesis for at least one AMF species of widely dispersed AMF genotypes at an intercontinental scale. Such information is crucial for understanding AMF ecology, and how these fungi can be used in an environmentally safe way in distant locations

Quantitative genetic variation in symbiotic fungi and its effect on fungal gene expression

Arbuscular mycorrhizal fungi (AMF) are wide-spread soil microorganisms that form a symbiotic relationship with most terrestrial plant species. AMF colonize plants roots, where they exchange inorganic soil nutrients for plant carbohydrates and lipids. This mycorrhizal association dates to more than 400 Myr ago and it has been hypothesized that it may have enabled the transition of plants from aquatic to terrestrial ecosystems. AMF have been repeatedly shown to significantly influence plants growth, plant community structure, plants resistance to various biotic (pathogens) and abiotic stresses (drought, pollutants, salinity, etc.). Therefore, AMF potentially represent a major tool to improve current agricultural practices and increase crop yields. However, the variability observed in these beneficial effects is very large and, to this date, unpredictable. Because different AMF isolates have differential effects on plants, it has become paramount to understand the molecular differences among AMF isolates. Several genetic studies have shown that extensive variation exists within the model AMF species Rhizophagus irregularis. R. irregularis isolates can be homokaryon or dikaryon, harbouring either one or two major populations of nucleus genotypes. In this thesis, I investigated some of the genetic and epigenetic differences between homokaryon and dikaryon R. irregularis isolates. In chapter 2, I studied the presence of the DNA methylation mark N6-methyldeoxyadenine (6mA) in the genome of R. irregularis and I found that it is most likely a functional epigenetic mark, and that genetically indistinguishable homokaryon isolates differed in their epigenome. In chapter 3, I investigated the generation of quantitative genetic variation during the clonal reproduction of a dikaryon AMF; and its potential impact on gene expression. I found that the two nucleus genotypes could be found at different ratios in a set of clonally produced offspring, and that the nuclear ratios directly impacted the transcriptome. Lastly, in chapter 4, I continued to investigate the contribution to trait variation of epigenetics in homokaryons, and variable nucleus genotype ratios in dikaryons. I studied the potential variation during the critical pre-symbiotic phase of spore germination, and I found that dikaryons generated greater plasticity and variation in molecular responses to changes in the environment than homokaryons. This thesis showed that both factors, epigenetics and variable nuclear ratios in dikaryons, are most likely contributing to the still unexplained trait variation in AMF and variable effects on plants. -- Les champignons mycorhiziens arbusculaires (AMF) sont des micro-organismes du sol très répandus qui entretiennent une relation symbiotique avec la plupart des espèces végétales terrestres. Les AMF colonisent les racines des plantes, où ils échangent les nutriments inorganiques du sol contre des glucides et des lipides. Cette association mycorhizienne existe depuis plus de 400 millions d'années et l'hypothèse a été émise qu'elle aurait permis la transition des plantes des écosystèmes aquatiques aux écosystèmes terrestres. Il a été démontré à plusieurs reprises que les AMF influencent de manière significative la croissance des plantes, la structure des communautés végétales, et la résistance des plantes à divers stress biotiques (ex., pathogènes) et/ou abiotiques (ex., sécheresse, polluants, salinité). Par conséquent, les AMF représentent potentiellement un outil essentiel pour améliorer nos pratiques agricoles courantes pour augmenter les rendements des cultures. Cependant, la variabilité observée dans les effets bénéfiques résultant de l'application d'AMF sur les plantes est très importante et, à ce jour, imprévisible. Étant donné que différents isolats de la même espèce d'AMF ont des effets différentiels sur les plantes, il est devenu primordial de comprendre les différences moléculaires entre ces isolats. Plusieurs études génétiques ont montré qu'il existe une grande variation dans les isolats de l'espèce modèle de AMF, Rhizophagus irregularis. Les isolats de R. irregularis peuvent être homocaryon ou dikaryon, abritant soit une ou deux populations majeures de génotypes de nucleus, respectivement. Dans cette thèse, j'ai étudié certaines des différences génétiques et épigénétiques entre les isolats homokaryon et dikaryon de R. irregularis. Dans le chapitre 2, j'ai étudié la présence de la marque de méthylation de l'ADN N6-méthyldeoxyadénine (6mA) dans le génome de R. irregularis. J'ai trouvé qu'il s'agit très probablement d'une marque épigénétique fonctionnelle, et que les isolats homokaryons génétiquement indiscernables différaient dans leur épigénome. Dans le chapitre 3, j'ai étudié la génération d'une variation génétique quantitative pendant la reproduction clonale d'un AMF dikaryon et son impact potentiel sur l'expression génétique. J'ai découvert que les deux génotypes de noyaux pouvaient se trouver à des ratios différents dans un ensemble de descendants produits par clonage, et que les ratios de noyaux avaient un impact direct sur le transcriptome. Dans le chapitre 4, j'ai continué à étudier la contribution de l'épigénétique à la variation des caractères chez les homocaryons et les rapports variables des génotypes de noyau chez les dikaryons. J'ai étudié la variation potentielle pendant la phase critique pré-symbiotique de la germination des spores, et j'ai découvert que les dikaryons généraient une plus grande plasticité, et une plus grande variation, que les homokaryons, dans les réponses moléculaires aux changements de l'environnement. Cette thèse a montré que deux facteurs, l'épigénétique et les rapports nucléaires variables chez les dikaryons, contribuent très probablement à la variation des traits encore inexpliquée chez les AMF qui peuvent être à l'origine de leurs effets variables sur les plantes

Genetically different isolates of the arbuscular mycorrhizal fungus Rhizophagus irregularis induce differential responses to stress in cassava

Water scarcity negatively impacts global crop yields and climate change is expected to greatly increase the severity of future droughts. The use of arbuscular mycorrhizal fungi (AMF) can potentially mitigate the effects of water stress in plants. Cassava is a crop that feeds approximately 800 million people daily. Genetically different isolates of the AMF R. irregularis as well as their clonal progeny have both been shown to greatly alter cassava growth in field conditions. Given that cassava experiences seasonal drought in many of the regions in which it is cultivated, we evaluated whether intraspecific variation in R. irregularis differentially alters physiological responses of cassava to water stress. In a first experiment, conducted in field conditions in Western Kenya, cassava was inoculated with two genetically different R. irregularis isolates and their clonal progeny. All cassava plants exhibited physiological signs of stress during the dry period, but the largest differences occurred among plants inoculated with clonal progeny of each of the two parental fungal isolates. Because drought had not been experimentally manipulated in the field, we conducted a second experiment in the greenhouse where cassava was inoculated with two genetically different R. irregularis isolates and subjected to drought, followed by re-watering, to allow recovery. Physiological stress responses of cassava to drought differed significantly between plants inoculated with the two different fungi. However, plants that experienced higher drought stress also recovered at a faster rate following re-watering. We conclude that intraspecific genetic variability in AMF significantly influences cassava physiological responses during water stress. This highlights the potential of using naturally existing variation in AMF to improve cassava tolerance undergoing water stress. However, the fact that clonal progeny of an AMF isolate can differentially affect how cassava copes with natural drought stress in field conditions, highlights the necessity to understand additional factors, beyond genetic variation, which can account for such large differences in cassava responses to drought