Roxana Colombo

The PAMPA datasets: a metagenomic survey of microbial communities in Argentinean pampean soils

BackgroundSoil is among the most diverse and complex environments in the world. Soil microorganisms play an essential role in biogeochemical cycles and affect plant growth and crop production. However, our knowledge of the relationship between species-assemblies and soil ecosystem processes is still very limited. The aim of this study was to generate a comprehensive metagenomic survey to evaluate the effect of high-input agricultural practices on soil microbial communities.ResultsWe collected soil samples from three different areas in the Argentinean Pampean region under three different types of land uses and two soil sources (bulk and rhizospheric). We extracted total DNA from all samples and also synthetized cDNA from rhizospheric samples. Using 454-FLX technology, we generated 112 16S ribosomal DNA and 14 16S ribosomal RNA amplicon libraries totaling 1.3 M reads and 36 shotgun metagenome libraries totaling 17.8 million reads (7.7 GB). Our preliminary results suggested that water availability could be the primary driver that defined microbial assemblages over land use and soil source. However, when water was not a limiting resource (annual precipitation >800 mm) land use was a primary driver.ConclusionThis was the first metagenomic study of soil conducted in Argentina and our datasets are among the few large soil datasets publicly available. The detailed analysis of these data will provide a step forward in our understanding of how soil microbiomes respond to high-input agricultural systems, and they will serve as a useful comparison with other soil metagenomic studies worldwide.

Continuous and long-term monoxenic culture of the arbuscular mycorrhizal fungus Gigaspora decipiens in root organ culture.

Establishment of arbuscular mycorrhizal (AM) germplasm collections is complex because of the obligate biotrophic nature of AM fungi. Only a few AM species are routinely maintained in monoxenic culture with Ri T-DNA transformed roots as host. Incorporation of new AM species into this culture system is important for molecular, physiological, and taxonomical studies. Here we report for the first time the successful monoxenic culture of Gigaspora decipiens (JA2 strain) with transformed carrot (Daucus carota) roots. In vitro cultures were established from field-collected spores; sub-culture of newly in vitro formed spores was established over five successive generations for a period of 6 y. Although initial culture of field-collected spores was difficult successive sub-cultures appeared to be adapted to the in vitro growing conditions. The JA2 strain of G. decipiens completed its life cycle while maintaining its morphological characteristics, stability, and propagule viability under the monoxenic conditions over several generations. This stable and homogeneous monoxenic material obtained for G. decipiens is part of the Banco de Glomeromycota In Vitro (BGIV, http://www.bgiv.com.ar), and could facilitate morphological, physiological, and molecular analysis of this AM species.

Evaluation of Arbuscular Mycorrhizal Fungi Capacity to Alleviate Abiotic Stress of Olive (Olea europaea L.) Plants at Different Transplant Conditions

The capacity of roots to sense soil physicochemical parameters plays an essential role in maintaining plant nutritional and developmental functions under abiotic stress. These conditions generate reactive oxygen species (ROS) in plant tissues causing oxidation of proteins and lipids among others. Some plants have developed adaptive mechanisms to counteract such adverse conditions such as symbiotic association with arbuscular mycorrhizal fungi (AMF). AMF enhance plant growth and improve transplant survival by protecting host plants against environmental stresses. The aim of this study was to evaluate the alleviation of transplanting stress by two strains of Rhizophagus irregularis (GC2 and GA5) in olive. Our results show that olive plants have an additional energetic expense in growth due to an adaptative response to the growing stage and to the mycorrhizal colonization at the first transplant. However, at the second transplant the coinoculation improves olive plant growth and protects against oxidative stress followed by the GA5-inoculation. In conclusion, a combination of two AMF strains at the beginning of olive propagation produces vigorous plants successfully protected in field cultivation even with an additional cost at the beginning of growth.

Differential efficiency of two strains of the arbuscular mycorrhizal fungus Rhizophagus irregularis on olive ( Olea europaea ) plants under two water regimes

The water regime affects a wide variety of physiological and biochemical processes in plants including an increased production of reactive oxygen species (ROS) capable of causing oxidative damage to proteins, DNA and lipids. Arbuscular mycorrhizal fungi (AMF) colonize a wide range of plant species though the ability of different AMF species to promote host growth or contribute to plant water deficit resistance varies. The first phase of olive tree cultivation takes place in a nursery where plants usually suffer stress by drying. Currently, olive production systems do not use of AMF to counteract this problem. To study the colonization strategies of two AMF strains and their efficiency with respect to growth and their effect on enzymatic activities, we inoculated them individually and co-inoculated then on olive plants under nursery growing conditions. The results showed the benefits generated by these fungi in terms of growth and survival rate. Co-inoculation, particularly, improved growth and reduced the damage due to water stress, partly as a result of the activation of the antioxidant defenses in the olive plant host.

Combined effects of arbuscular mycorrhizal fungi and exogenous cytokinins on pomegranate (Punica granatum) under two contrasting water availability conditions

To our knowledge, there are no studies on the interactive effects of inoculation with arbuscular mycorrhizal fungi and cytokinin addition to plants under drought stress. We investigated the potential protective effect of arbuscular mycorrhizae on pomegranate plants, combined with exogenous cytokinin addition, under two contrasting soil water availability regimes. Our results showed that exogenous cytokinin addition enhances plant biomass, shoot to root ratio and water content, as well as increasing the anthocyanin content. However, a combination of AM fungal inoculation and cytokinin addition did not result in a synergistic protective effect against water stress. Plants were equally well protected against this stress by cytokinin spraying alone. The improvement of pomegranate growth was due mainly to exogenous cytokinin addition. Photosynthesis was promoted both by mycorrhizal inoculation alone and by exogenous cytokinin addition. The main protection against oxidative stress caused by drought was via enhanced accumulation of anthocyanins when the plants were sprayed with cytokinins. When cytokinins were used, the photosynthesis apparatus was also protected.

Arbuscular Mycorrhizal Fungal Association in Genetically Modified Drought-Tolerant Corn

The commercial use of genetically modified (GM) plants has significantly increased worldwide. The interactions between GM plants and arbuscular mycorrhizal (AM) fungi are of considerable importance given the agricultural and ecological role of AM and the lack of knowledge regarding potential effects of drought-tolerant GM corn ( L.) on AM fungal symbiosis. This work studied AM fungal colonization in five corn lines growing under two different irrigation regimes (30 and 100% of soil field capacity [SFC]). Four of the lines were GM corn, and two of these were drought tolerant. The experiment was conducted for 60 d in a growth chamber under constant irrigation, after which mycorrhization, corn biomass, and days to plant senescence (DTS) were evaluated. Arbuscular mycorrhizal fungal species of the order were predominant in the soil inocula. At the end of the experiment, all plants showed AM colonization. Mycorrhization was higher at 30% SFC than at 100% SFC. Within the same corn line, the AM fungi produced more vesicles in plant roots under drought stress. Among treatments, DTS varied significantly, and drought-tolerant GM corn lines survived longer than the wild-type corn when maintained at 100% SFC. Corn biomass did not vary among treatments, and no correlations were found between DTS or biomass and mycorrhization. We conclude that overexpression of the gene in corn plants under the experimental conditions of this study did not affect AM fungal infectivity and improved the tolerance of the corn to drought stress.

Growth dynamics of geographically different arbuscular mycorrhizal fungal isolates belonging to the 'Rhizophagus clade' under monoxenic conditions

The growth dynamics of extraradical mycelium and spore formation of 14 “Rhizophagus” isolates from different sites in Argentina were evaluated under monoxenic conditions. A modified Gompertz model was used to characterize the development of mycelium and spores for each isolate under the same conditions. The lag time, maximal growth rate and total quantity of both extraradical hyphae and spores were determined. Wide variability among isolates was detected, and all growth parameters were significantly altered by fungal isolate. Discriminant analysis differentiated isolates primarily based on the extent of extraradical hyphae produced, yet such differences did not conclusively correspond to phylogenetic relationships among closely related isolates based on partial SSU sequences. Given that the “Rhizophagus” isolates were grown under controlled conditions for many generations, the expression of phenotypic variability could be attributed to genetic differences that are not completely resolved by phylogenetic analysis employing the small ribosomal gene.

Arbuscular mycorrhizal fungal diversity in high-altitude hypersaline Andean wetlands studied by 454-sequencing and morphological approaches

The Laguna Brava Nature Reserve is a stressful habitat in the Andean Mountains (Argentina) dominated by extreme abiotic factors: high altitude and UV radiance, hypersalinity, alkalinity, and high concentrations of toxic elements in the soil. The sparse native vegetation that inhabits Laguna Brava and Mulas Muertas wetlands is frequently colonized by arbuscular mycorrhizal (AM) fungi. It is, however, unknown which AM species can survive in such a harsh environment and how those environmental conditions influence the AM communities. To answer these questions, 454-amplicon pyrosequencing and morphological (based on spore traits) approaches were used to assess fungal diversity. A total of 23 molecular operational taxonomic units and 14 distinct morphospecies of AM fungi were identified. The morphological characterization of AM fungal communities in Laguna Brava and Mulas Muertas, supported by the molecular data, revealed that Glomeraceae and Claroideoglomeraceae were the dominant families, confirming the predominance of generalist and ruderal AM fungal taxa but with stress-tolerant life history traits. Our results showed that the presence of AM fungi is strongly associated with local environmental variations in Laguna Brava (hypersalinity and high Na+, Sr, As and U contents in soils). The AM fungal communities in Laguna Brava and Mulas Muertas wetlands were similar according to the Simpson diversity index and the ecological distance estimated by Bray Curtis index. These results were also supported by the environmental parameters measured, as they did not vary between the studied sites. This study represents the first characterization of AM fungal community in a high-altitude Andean wetland in Argentina, improving our knowledge about these fungi from extreme environments.

Differential effects of two strains of Rhizophagus intraradices on dry biomass and essential oil yield and composition in Calamintha nepeta.

The aim of this work was to determine the effects of two geographically different strains of Rhizophagus intraradices (M3 and GA5) on the total biomass and essential oil (EO) yield and composition of Calamintha nepeta, with or without phosphorus (P) fertilization, under greenhouse conditions. The plant biomass was not significantly affected by any of the treatments, showing higher values in control plants. Strains had a differential response in their root colonization rates: M3 reduced these parameters while GA5 did not modify them. Both strains affected EO yield in absence of P fertilization: M3 promoted EO yield in C. nepeta plants and GA5 resulted in negative effects. The percentage composition of EO was not significantly modified by either strain or P fertilization. M3 strain could be a potential fungal bioinoculant for production and commercialization of C. nepeta in the aromatic plant market.