A polyphasic approach has been developed to gain knowledge of suitable key indicators for the evaluation of environmental impact of genetically modified Bt 11 and Bt 176 corn lines on soil ecosystems. We assessed the effects of Bt corn (which constitutively expresses the insecticidal toxin from Bacillus thuringiensis, encoded by the truncated Cry1Ab gene) and non-Bt corn plants and their residues on rhizospheric and bulk soil eubacterial communities by means of denaturing gradient gel electrophoresis analyses of 16S rRNA genes, on the nontarget mycorrhizal symbiont Glomus mosseae, and on soil respiration. Microcosm experiments showed differences in rhizospheric eubacterial communities associated with the three corn lines and a significantly lower level of mycorrhizal colonization in Bt 176 corn roots. In greenhouse experiments, differences between Bt and non-Bt corn plants were detected in rhizospheric eubacterial communities (both total and active), in culturable rhizospheric heterotrophic bacteria, and in mycorrhizal colonization. Plant residues of transgenic plants, plowed under at harvest and kept mixed with soil for up to 4 months, affected soil respiration, bacterial communities, and mycorrhizal establishment by indigenous endophytes. The multimodal approach utilized in our work may be applied in long-term field studies aimed at monitoring the real hazard of genetically modified crops and their residues on nontarget soil microbial communities.

Arbuscular mycorrhizal (AM) fungi are essential elements of soil fertility, plant nutrition and productivity, facilitating soil mineral nutrient uptake. Helianthus annuus is a non-model, widely cultivated species. Here we used an RNA-seq approach for evaluating gene expression variation at early and late stages of mycorrhizal establishment in sunflower roots colonized by the arbuscular fungus Rhizoglomus irregulare. mRNA was isolated from roots of plantlets at 4 and 16 days after inoculation with the fungus. cDNA libraries were built and sequenced with Illumina technology. Differential expression analysis was performed between control and inoculated plants. Overall 726 differentially expressed genes (DEGs) between inoculated and control plants were retrieved. The number of up-regulated DEGs greatly exceeded the number of down-regulated DEGs and this difference increased in later stages of colonization. Several DEGs were specifically involved in known mycorrhizal processes, such as membrane transport, cell wall shaping, and other. We also found previously unidentified mycorrhizal-induced transcripts. The most important DEGs were carefully described in order to hypothesize their roles in AM symbiosis. Our data add a valuable contribution for deciphering biological processes related to beneficial fungi and plant symbiosis, adding an Asteraceae, non-model species for future comparative functional genomics studies.

Summary• Inoculation of crop plants by non-native strains of arbuscular mycorrhizal (AM) fungi as bio-enhancers is promoted without clear evidence for symbiotic effectiveness and fungal persistence. To address such gaps, the forage legume Medicago sativa was inoculated in an agronomic field trial with two isolates of Funneliformis mosseae differing in their nuclear rDNA sequences from native strains.• The inoculants were traced by PCR with a novel combination of the universal fungal NS31 and Glomeromycota-specific LSUGlom1 primers which target the nuclear rDNA cistron. The amplicons were classified by restriction fragment length polymorphism and sequencing.• The two applied fungal inoculants were successfully traced and discriminated from native strains in roots sampled from the field up to 2 yr post inoculation. Moreover, field inoculation with inocula of non-native isolates of F. mosseae appeared to have stimulated root colonization and yield of M. sativa.• Proof of inoculation success and sustained positive effects on biomass production and quality of M. sativa crop plants hold promise for the role that AM fungal inoculants could play in agriculture.

We developed an experimental model system to monitor the impact of generically modified (GM) plants on arbuscular mycorrhizal (AM) fungi, a group of non-target soil microorganisms, fundamental for soil fertility and plant nutrition. The system allowed us to study the effects of root exudates of both commercial Bt corn and aubergine plants expressing Dm-AMP1 defensin on different stages of the life cycle of the AM fungal species G. mosseae. Root exudates of Bt 176 corn significantly reduced pre-symbiotic hyphal growth, compared to Bt 11 and non-transgenic plants. No differences were found in mycelial growth in the presence of Dm-AMP1 and control plant root exudates. Differential hyphal morphogenesis occurred irrespective of the plant line, suggesting that both exuded Bt toxin and defensin do not interfere with fungal host recognition mechanisms. Bt 176 affected the regular development of appressoria, 36% of which failed to produce viable infection pegs. Our experimental model system represents an easy assay for testing the impact of GM plants on non-target soil-borne AM fungi

Arbuscular mycorrhizal fungi (AMF) are beneficial soil microorganisms establishing mutualistic symbioses with the roots of the most important food crops and playing key roles in the maintenance of long-term soil fertility and health. The great inter- and intra-specific AMF diversity can be fully exploited by selecting AMF inocula on the basis of their colonization ability and efficiency, which are affected by fungal and plant genotypes and diverse environmental variables. The multiple services provided by AMF are the result of the synergistic activities of the bacterial communities living in the mycorrhizosphere, encompassing nitrogen fixation, P solubilization, and the production of phytohormones, siderophores, and antibiotics. The tripartite association among host plants, mycorrhizal symbionts, and associated bacteria show beneficial emerging properties which could be efficiently exploited in sustainable agriculture. Further in-depth studies, both in microcosms and in the field, performed on different AMF species and isolates, should evaluate their colonization ability, efficiency, and resilience. Transcriptomic studies can reveal the expression levels of nutrient transporter genes in fungal absorbing hyphae in the presence of selected bacterial strains. Eventually, newly designed multifunctional microbial consortia can be utilized as biofertilizers and biostimulants in sustainable and innovative production systems.

Arbuscular mycorrhizal fungi are increasingly used in organic cropping systems to increase yields. Although cover crops are largely used in organic farming, there is little knowledge on the impact of cover crops on native mycorrhizal fungi. Here, we studied the effect of cover crop diversity on mycorrhizal colonization in subsequent organic maize cultivars differing in the level of genetic diversity. Experiments were conducted from 2010 to 2012 in a Mediterranean environment. First Indian mustard (Brassica juncea L. Czern.), hairy vetch (Vicia villosa Roth), a mix of seven cover crop species (Mix), and natural vegetation (Control) were cultivated as winter cover crops. Then, an organically and a conventionally bred maize hybrid and three organically bred composite cross populations were cultivated. Mycorrhizal propagule dynamics were measured. Results at juvenile stage show a higher mycorrhizal colonization in maize plants grown after hairy vetch, of 35.0 %, and Mix cover crops, of 29.4 %, compared to Indian mustard, of 20.9 %, and Control, of 21.3 %. The potential of soil mycorrhization decreased of 56.5 % following Indian mustard, higher than that of other cover crops, of 34.1-47.3 %. This finding could be explained by the release of isothiocyanates in soils. Moreover, maize shoot biomass, nitrogen, and phosphorus content across all maize genotypes at juvenile stage increased with mycorrhizal colonization. These findings provide the first evidence of the greater role played by cover crop identity in the enhancement of early mycorrhizal colonization of the subsequent crop and of soil mycorrhizal activity.

The new paradigm in agriculture, sustainable intensification, is focusing back onto beneficial soil microorganisms, for the role played in reducing the input of chemical fertilizers and pesticides and improving plant nutrition and health. Worldwide, more and more attention is deserved to arbuscular mycorrhizal fungi (AMF), which establish symbioses with the roots of most land plants and facilitate plant nutrient uptake, by means of a large network of extraradical hyphae spreading from colonized roots to the surrounding soil and functioning as a supplementary absorbing system. AMF protect plants from biotic and abiotic stresses and are able to modulate the activity of antioxidant enzymes and the biosynthesis of secondary metabolites (phytochemicals), such as polyphenols, anthocyanins, phytoestrogens and carotenoids, that play a fundamental role in promoting human health. An increasing number of studies focused on the use of AMF symbionts for the production of functional food, with enhanced nutritional and nutraceutical value. Yet, while several plant species were investigated, only few AMF were utilized, thus limiting the full exploitation of their wide physiological and genetic diversity. Here, we will focus on AMF effects on the biosynthesis of plant secondary metabolites with health-promoting activity, and on the criteria for a finely tuned, targeted selection of the best performing symbionts, to be utilized as sustainable biotechnological tools for the production of safe and healthy plant foods.

Arbuscular mycorrhizal fungal (AMF) communities are fundamental in organic cropping systems where they provide essential agro-ecosystem services, improving soil fertility and sustaining crop production. They are affected by agronomic practices, but still, scanty information is available about the role of specific crops, crop rotations and the use of winter cover crops on the AMF community compositions at the field sites. A field experiment was conducted to elucidate the role of diversified cover crops and AMF inoculation on AMF diversity in organic tomato. Tomato, pre-inoculated at nursery with two AMF isolates, was grown following four cover crop treatments: Indian mustard, hairy vetch, a mixture of seven species and a fallow. Tomato root colonization at flowering was more affected by AMF pre-transplant inoculation than by the cover crop treatments. An enormous species richness was found by morphological spore identification: 58 AMF species belonging to 14 genera, with 46 and 53 species retrieved at the end of cover crop cycle and at tomato harvest, respectively. At both sampling times, AMF spore abundance was highest in hairy vetch, but after tomato harvest, AMF species richness and diversity were lower in hairy vetch than in the cover crop mixture and in the mustard treatments. A higher AMF diversity was found at tomato harvest, compared with the end of the cover crop cycle, independent of the cover crop and pre-transplant AMF inoculation. Our findings suggest that seasonal and environmental factors play a major role on AMF abundance and diversity than short-term agronomic practices, including AMF inoculation. The huge AMF diversity is explained by the field history and the Mediterranean environment, where species characteristic of temperate and sub-tropical climates co-occur.