Protozoa enhance foraging efficiency of arbuscular mycorrhizal fungi for mineral nitrogen from organic matter in soil to the benefit of host plants.

Dead organic matter (OM) is a major source of nitrogen (N) for plants. The majority of plants support N uptake by symbiosis with arbuscular mycorrhizal (AM) fungi. Mineralization of N is regulated by microfauna, in particular, protozoa grazing on bacteria. We hypothesized that AM fungi and protozoa interactively facilitate plant N nutrition from OM. In soil systems consisting of an OM patch and a root compartment, plant N uptake and consequences for plant carbon (C) allocation were investigated using stable isotopes. Protozoa mobilized N by consuming bacteria, and the mobilized N was translocated via AM fungi to the host plant. The presence of protozoa in both the OM and root compartment stimulated photosynthesis and the translocation of C from the host plant via AM fungi into the OM patch. This stimulated microbial activity in the OM patch, plant N uptake from OM and doubled plant growth. The results indicate that protozoa increase plant growth by both mobilization of N from OM and by protozoa-root interactions, resulting in increased C allocation to roots and into the rhizosphere, thereby increasing plant nutrient exploitation. Hence, mycorrhizal plants need to interact with protozoa to fully exploit N resources from OM.

The Phosphate Inhibition Paradigm: Host and Fungal Genotypes Determine Arbuscular Mycorrhizal Fungal Colonization and Responsiveness to Inoculation in Cassava With Increasing Phosphorus Supply.

A vast majority of terrestrial plants are dependent on arbuscular mycorrhizal fungi (AMF) for their nutrient acquisition. AMF act as an extension of the root system helping phosphate uptake. In agriculture, harnessing the symbiosis can potentially increase plant growth. Application of the AMF Rhizophagus irregularis has been demonstrated to increase the yields of various crops. However, there is a paradigm that AMF colonization of roots, as well as the plant benefits afforded by inoculation with AMF, decreases with increasing phosphorus (P) supply in the soil. The paradigm suggests that when fertilized with sufficient P, inoculation of crops would not be beneficial. However, the majority of experiments demonstrating the paradigm were conducted in sterile conditions without a background AMF or soil microbial community. Interestingly, intraspecific variation in R. irregularis can greatly alter the yield of cassava even at a full application of the recommended P dose. Cassava is a globally important crop, feeding 800 million people worldwide, and a crop that is highly dependent on AMF for P uptake. In this study, field trials were conducted at three locations in Kenya and Tanzania using different AMF and cassava varieties under different P fertilization levels to test if the paradigm occurs in tropical field conditions. We found that AMF colonization and inoculation responsiveness of cassava does not always decrease with an increased P supply as expected by the paradigm. The obtained results demonstrate that maximizing the inoculation responsiveness of cassava is not necessarily only in conditions of low P availability, but that this is dependent on cassava and fungal genotypes. Thus, the modeling of plant symbiosis with AMF under different P levels in nature should be considered with caution.

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.

Aligning molecular studies of mycorrhizal fungal diversity with ecologically important levels of diversity in ecosystems.

Arbuscular mycorrhizal fungi (AMF) occur in the roots of most plants and are an ecologically important component of the soil microbiome. Richness of AMF taxa is a strong driver of plant diversity and productivity, thus providing a rationale for characterizing AMF diversity in natural ecosystems. Consequently, a large number of molecular studies on AMF community composition are currently underway. Most published studies, at best, only address species or genera-level resolution. However, several experimental studies indicate that variation in plant performance is large among plants colonised by different individuals of one AMF species. Thus, there is a potential disparity between how molecular community ecologists are currently describing AMF diversity and the level of AMF diversity that may actually be ecologically relevant. We propose a strategy to find many polymorphic loci that can define within-species genetic variability within AMF, or at any level of resolution desired within the Glomermycota. We propose that allele diversity at the intraspecific level could then be measured for target AMF groups, or at other levels of resolution, in environmental DNA samples. Combining the use of such markers with experimental studies on AMF diversity would help to elucidate the most important level(s) of AMF diversity in plant communities. Our goal is to encourage ecologists who are trying to explain how mycorrhizal fungal communities are structured to take an approach that could also yield meaningful information that is relevant to the diversity, functioning and productivity of ecosystems.

Bacteria with Phosphate Solubilizing Capacity Alter Mycorrhizal Fungal Growth Both Inside and Outside the Root and in the Presence of Native Microbial Communities.

Arbuscular mycorrhizal fungi (AMF) and phosphate solubilizing Pseudomonas bacteria (PSB) could potentially interact synergistically because PSB solubilize phosphate into a form that AMF can absorb and transport to the plant. However, very little is known about the interactions between these two groups of microorganisms and how they influence the growth of each other. We tested whether different strains of bacteria, that have the capacity to solubilize phosphate, are able to grow along AMF hyphae and differentially influence the growth of AMF both outside the roots of carrot in in vitro conditions and inside the roots of potato in the presence of a microbial community. We found strong effects of AMF on the growth of the different bacterial strains. Different bacterial strains also had very strong effects on the growth of AMF extraradical hyphae outside the roots of carrot and on colonization of potato roots by AMF. The differential effects on colonization occurred in the presence of a microbial community. Our results show that these two important groups of rhizosphere microorganisms indeed interact with each other. Such interactions could potentially lead to synergistic effects between the two groups but this could depend on whether the bacteria truly solubilize phosphate in the rhizosphere in the presence of microbial communities.

The role of community and population ecology in applying mycorrhizal fungi for improved food security.

The global human population is expected to reach ∼9 billion by 2050. Feeding this many people represents a major challenge requiring global crop yield increases of up to 100%. Microbial symbionts of plants such as arbuscular mycorrhizal fungi (AMF) represent a huge, but unrealized resource for improving yields of globally important crops, especially in the tropics. We argue that the application of AMF in agriculture is too simplistic and ignores basic ecological principals. To achieve this challenge, a community and population ecology approach can contribute greatly. First, ecologists could significantly improve our understanding of the determinants of the survival of introduced AMF, the role of adaptability and intraspecific diversity of AMF and whether inoculation has a direct or indirect effect on plant production. Second, we call for extensive metagenomics as well as population genomics studies that are crucial to assess the environmental impact that introduction of non-local AMF may have on native AMF communities and populations. Finally, we plead for an ecologically sound use of AMF in efforts to increase food security at a global scale in a sustainable manner.

ALTA PRESENCIA DE CADMIO RESULTA EN BAJA DIVERSIDAD DE HONGOS FORMADORES DE MICORRIZAS ARBUSCULARES ASOCIADOS A CACAO (Theobroma cacao L.) / High cadmium concentration resulted in low arbuscular mycorrhizal fungi community diversity associated to cocoa (Theobroma cacao L.)

RESUMEN Los hongos formadores de micorrizas arbusculares (HFMA) son simbiontes obligados presentes en la rizósfera de plantas de cacao y la diversidad de sus comunidades se modifica, dependiendo de diversos factores como la presencia de cadmio (Cd) en el suelo. La persistencia de HFMA en suelos enriquecidos naturalmente con Cd podría ser un indicador de su capacidad para tolerar esta condición. Esta investigación caracterizó la estructura de la comunidad de HFMA locales presentes en la rizósfera de plantas de cacao en dos suelos con baja (B-Cd: 0,1 mg kg-1) y alta (A-Cd: 20,9 mg kg-1) concentración de Cd. Esporas de HFMA se identificaron mediante claves taxonómicas y su abundancia, riqueza y diversidad se determinó en muestras de suelo originales y después de su multiplicación mediante cultivos trampa. Las comunidades de HFMA se compararon usando análisis de componentes principales (ACP) e índices de diversidad alfa y beta. Los resultados indican que A-Cd presentó valores significativamente menores de abundancia (21 %), riqueza (20 %) y diversidad (11 %) de morfoespecies de HFMA con respecto a B-Cd. Las dos comunidades de HFMA presentaron cinco de siete géneros en común, pero solo cuatro de las 23 morfoespecies descritas se encontraron en ambas comunidades. El análisis de diversidad beta y el ACP determinaron baja similaridad y tasa de recambio entre las comunidades de HFMA. La dominancia de Diversispora spurca, Rhizoglomus sp. y Claroideoglomus etunicatum en A-Cd sugiere que estas morfoespecies son estrés-tolerantes y candidatos potenciales para el desarrollo de estrategias de mitigación en suelos con Cd.

ABSTRACT Arbuscular mycorrhizae fungi (AMF) are obligate symbionts present in rhizosphere of cocoa plants and their community diversity is modified depending on several factors, such as cadmium (Cd) presence in soil. AMF persistence on Cd natural enriched soils might be an indicator of their tolerance and their potential in biotechnological applications. In this research we characterized local AMF community structure present in cocoa rhizosphere soils with low (B-Cd: 0.1 mg kg-1) and high (A-Cd: 20.9 mg kg-1) natural Cd concentrations. AMF spore identification was carried out using taxonomic keys and their abundance, richness and diversity were determined in original samples and after multiplication process using onion trap cultures. AMF communities were compared using alpha and beta diversity indexes and principal component analysis (PCA). The results indicated that A-Cd presented significative lower values of abundance (21 %), richness (20 %) and diversity (11 %) of AMF morphospecies in comparison with B-Cd. Both AMF communities presented five of seven genera in common, but only four of 23 morphospecies described were found in two communities. Low similarity and turnover were found among AMF communities throughout beta diversity analysis and PCA. Dominance of Diversispora spurca, Rhizoglomus sp. and Claroideoglomus etunicatum in A-Cd suggests that these morphospecies are stress-tolerant and they are potential candidates for the development of mitigation strategies in cocoa plants under Cd stress.

The in vitro mass-produced model mycorrhizal fungus, Rhizophagus irregularis, significantly increases yields of the globally important food security crop cassava.

The arbuscular mycorrhizal symbiosis is formed between arbuscular mycorrhizal fungi (AMF) and plant roots. The fungi provide the plant with inorganic phosphate (P). The symbiosis can result in increased plant growth. Although most global food crops naturally form this symbiosis, very few studies have shown that their practical application can lead to large-scale increases in food production. Application of AMF to crops in the tropics is potentially effective for improving yields. However, a main problem of using AMF on a large-scale is producing cheap inoculum in a clean sterile carrier and sufficiently concentrated to cheaply transport. Recently, mass-produced in vitro inoculum of the model mycorrhizal fungus Rhizophagus irregularis became available, potentially making its use viable in tropical agriculture. One of the most globally important food plants in the tropics is cassava. We evaluated the effect of in vitro mass-produced R. irregularis inoculum on the yield of cassava crops at two locations in Colombia. A significant effect of R. irregularis inoculation on yield occurred at both sites. At one site, yield increases were observed irrespective of P fertilization. At the other site, inoculation with AMF and 50% of the normally applied P gave the highest yield. Despite that AMF inoculation resulted in greater food production, economic analyses revealed that AMF inoculation did not give greater return on investment than with conventional cultivation. However, the amount of AMF inoculum used was double the recommended dose and was calculated with European, not Colombian, inoculum prices. R. irregularis can also be manipulated genetically in vitro, leading to improved plant growth. We conclude that application of in vitro R. irregularis is currently a way of increasing cassava yields, that there is a strong potential for it to be economically profitable and that there is enormous potential to improve this efficiency further in the future.

ANÁLISIS ESPACIAL DE LA INCIDENCIA DE ENFERMEDADES EN DIFERENTES GENOTIPOS DE CACAO (Theobroma cacao L.) EN EL YOPAL (CASANARE), COLOMBIA / Spatial Analysis of Diseases Incidence in Different Cocoa Genotypes (Theobroma cacao L.) in Yopal (Casanare), Colombia

RESUMEN Las enfermedades "pudrición negra del fruto" y "moniliasis" son reportadas como limitantes en plantaciones de cacao en Colombia. Fueron evaluados los patrones espaciales de incidencia para estas dos enfermedades en un cultivo de cacao durante un monitoreo de trece semanas en campo. Semivariogramas e interpolación kriging fueron usados para analizar el comportamiento espacial de "pudrición negra del fruto", los estadísticos Join Count e Indice de Moran se emplearon para estudiar el comportamiento espacial de "moniliasis". La incidencia de "pudrición negra del fruto" presentó estructura de dependencia espacial de acuerdo a los semivariogramas durante el tiempo de evaluación, mientras que la autocorrelación espacial de sitios de infección de "moniliasis" se distribuyó de forma aleatoria entre las plantas de cacao según el Indice de Moran. El rango de dispersión estimado de "pudrición negra del fruto" varió entre 21,74 y 40,23 m y los mapas kriging evidenciaron el progreso espacio-temporal de la enfermedad durante las semanas evaluadas. Las condiciones experimentales dadas y la aproximación de los análisis espaciales, contribuyeron al entendimiento de la dinámica espacial de dos enfermedades limitantes en tres clones de cacao en la Orinoquía Colombiana.

ABSTRACT "Black pod" and "frosty pod" diseases are reported as limiting in cocoa plantations in Colombia. Spatial patterns of incidence for these two diseases were evaluated during a thirteen weeks long period of monitoring in the field. Semivariogram and kriging interpolation were used to analyze the spatial behavior of "black pod" and Join Count and Moran's Index statistics were used to analyze the spatial behavior of "frosty pod". The incidence of "Black pod" has a spatial dependence structure according to semivariogramas during the evaluation time, while spatial autocorrelation of "moniliasis" infection sites was randomly distribuited among cocoa plants according to the Moran's Index. The estimated dispersion range of "Black pod" was from 21.74 to 40.23 m y the kriging maps showed the spatio-temporal progression of the disease during the evaluated weeks. The experimental conditions given and spatial analysis approach, contributed to the understanding of the spatial dynamics of two limiting diseases in three clones of cocoa in the Colombian Orinoquia.

Mecanismos de defensa y respuestas de las plantas en la interacción micorrícica: una revisión / Plant defense mechanisms and responses in the arbuscular mycorrhizal symbiosis: a review

El establecimiento de la simbiosis planta-hongos formadores de micorrizas Arbusculares (HFMA) requiere procesos armónicos a nivel espacio-temporal, que dependen de señales para el reconocimiento, colonización e intercambio bidireccional de nutrientes. Las plantas presentan respuestas de defensa frente a posibles organismos invasores; sin embargo, frente a HFMA estas son débiles, localizadas y no impiden la colonización del hongo. Los beneficios de la simbiosis generalmente se asocian a nutrición vegetal, aunque, también está relacionada con el incremento de la tolerancia-resistencia de plantas a los estreses bióticos. La resistencia inducida HFMA (MIR) es importante en el control de patógenos foliares, comedores de hojas y necrótrofos, encontrándose protección de plantas micorrizadas tanto a nivel local como sistémico, relacionada con los niveles de ácido jasmónico en tejidos. Un mecanismo en la MIR está asociado con el “priming”, que permite una rápida y eficiente respuesta de defensa de plantas micorrizadas. Se han planteado posibles mecanismos de atenuación de las respuestas de defensa, considerando: activación de supresores de defensa; plantas que producen respuestas de defensa frente a HFMA y otras que no las producen, y plantas que suprimen las respuestas de defensa en la simbiosis. Aunque el control de la simbiosisestá regulado básicamente por la planta, aún se desconoce el papel de los HFMA en el debilitamiento de las respuestas de defensa. Recientemente, se ha dado un avance importante en entender los mecanismos mediante los cuales se establece y mantiene la biotrofía del hongo, al describirse la proteína SP7 que interactúa con el factor de transcripción PR, ERF19 en el núcleo de la célula vegetal. Se ha sugerido que SP7 es un efector que actúa oponiéndose al programa de inmunidad de la planta.

Harmonic processes between plant and arbuscular mycorrhyzal fungi (AMF) are required for the symbiosis formation between the two organisms. These processes depend on specific signalling for the plant-fungus recognition, colonisation and bidirectional nutrient exchange. Plants show defence responses against invasive organisms, however they react weakly and localised when challenged by AMF. The benefits derived from the mycorrization are described for the nutritional aspect; however, it is known that mycorrhized plants are more tolerant to biotic stresses. Mycorrhizal induced resistance (MIR) is especially important for the control of foliar pathogens, leaf cutters and necrotrophs. There has also been found that mycorrhizal plants are protected both locally and systemically and their protection is related with jasmonic acid levels at their tissues. One of the most important mechanisms for MIR is the so called “priming” that allows plants to exert a fast and efficient defence response. Possible mechanisms to unravel mycorrhizal plants lower defence systems include: defence suppressor activation, differential plants response towards AMF from inexistent to low, and plant defence response suppression during the AMF symbiosis. The symbiosis control is known to be regulated by the plant, however, no role has been assigned to the AMF for the weakening of the plant defence system. Recently, a big step towards understanding of the fungal role has been made. A protein SP7 that interacts with a PR transcription factor ERF19, in the plant nucleus, has been described. This discovery indicates a possible mechanism to establish and maintain the biotrophic status of the AMF counteracting the immune plant system.

Studies on the diversity of the distinct phylogenetic lineage encompassing Glomus claroideum and Glomus etunicatum.

Morphological and molecular characters were analysed to investigate diversity within isolates of the Glomus claroideum/Glomus etunicatum species group in the genus Glomus. The inter- and intra-isolate sequence diversity of the large subunit (LSU) rRNA gene D2 region of eight isolates of G. claroideum and G. etunicatum was studied using PCR-single strand conformational polymorphism (SSCP)-sequencing. In addition, two isolates recently obtained from Southern China were included in the analysis to allow for a wider geographic screening. Single spore DNA isolation confirmed the magnitude of gene diversity found in multispore DNA extractions. An apparent overlap of spore morphological characters was found between G. claroideum and G. etunicatum in some isolates. Analysis of the sequence frequencies in all G. etunicatum and G. claroideum isolates (ten) showed that four LSU D2 sequences, representing 32.1% of the clones analysed for multispore extraction (564) were found to be common to both species, and those sequences were the most abundant in four of the ten isolates analysed. The frequency of these sequences ranged between 23.2% and 87.5% of the clones analysed in each isolate. The implications for the use of phenotypic characters to define species in arbuscular mycorrhizal fungi are discussed. The current position of G. claroideum/G.etunicatum in the taxonomy of the Glomeromycota is also discussed.