Ammonium transporters in grasses : molecular and functional characterization with special reference to the arbuscular mycorrhizal symbiosis

Most herbaceous plants live in symbiosis with arbuscular mycorrhizal (AM) fungi. AM fungi colonize the roots of their host plant symbionts and provide them with mineral nutrients, especially phosphorus (P) and nitrogen (N) and receive, in exchange, photosynthetically fixed carbon. In this work, we focused on the role of N in the AM symbiosis formed between Glomus mosseae or Rhizophagus irregularis and different plants belonging to the Poaceae: sorghum (Sorghum bicolor), maize (Zea mays), rice (Oryza sativa), foxtail millet (Setaria italic) and purple false brome (Brachypodium distachyon). It had been shown that AM fungi can take up N in form of nitrate, ammonium and amino acid and transfer it to the plant in form of ammonium. Thus, we hypothesized that some plant ammonium transporters (AMT) might be up-regulated at the interface between plant and fungus in the AM symbiosis. As described in chapter 2, we established mycorrhized and non-mycorrhized sorghum plants and gave them different N treatments: no nitrogen, nitrate or ammonium. We found out that two AMTs, AMT3;1 and AMT4 were induced in mycorrhized plants (AM-inducible AMTs) independently of their N status. In sorghum, the pattern of expression of AMT3;1 and AMT4 was assessed with a split-root experiment combined with laser microdissection technology. Expression of both AMTs was not systemic in the roots of the plant. However, at a small scale, systemic expression around cells containing arbuscules could be observed. We conclude that expression of AMT3;1 and AMT4 could be part of the prepenetration response of the plant, preparing the cells to receive a new arbuscule. In addition, using immunolocalization, we localized the protein of AMT3;1 at the level of mature arbuscules. As described in chapter 3, the up-regulation of AMT3;1 and AMT4 was conserved in all four Poaceae species studies. As the core Poaceae divided in two groups about -55 million years ago separating sorghum, foxtail millet and maize from rice and purple false brome, we assume that AMT3;1 and AMT4 were already induced by AM fungi in a common ancestor of all these plants. In chapter 4, we looked at the fungal side and at the effect of the different N treatments on the expression of fungal transporters and enzymes of the N cycle. Our results show that the source of N has an impact on the transcriptional regulation of enzymes from the fungal N cycle. Expression of the corresponding genes was modified in the fungal extraradical mycelium as well as in the intraradical mycelium. In chapter 5, we studied the time needed by the AM fungus Glomus mosseae to transfer N from a 15N-labeled source to sorghum plants. Labeled N was present in the plant leaves already after 48 hours revealing a very rapid transfer. This finding highlights the underestimated role of AM symbiosis in N-acquisition by the plant

Inorganic Nitrogen Uptake and Transport in Beneficial Plant Root-Microbe Interactions

Arbuscular mycorrhiza (AM), ectomycorrhiza (ECM) and nitrogen (N) fixation through rhizobia symbioses (RS) play a critical role for plant nutrient use efficiency in natural ecosystems, usually characterized by nutrient limitation, especially regarding nitrogen and phosphate. Substantial evidence has accumulated about how the rational use of microsymbionts' properties should significantly contribute to decreasing fertilizer and pesticide use in agriculture and forestry. Understanding the mechanisms underlying high N use efficiency by mycorrhizal/rhizobial plants and carbon allocation in a context of mutualistic biotrophic interactions is critical for managing both croplands and forests while taking care of the ecosystem services rendered by microbial symbionts. Availability, uptake and exchange of nutrients in biotrophic interactions drive plant growth and modulate biomass allocation, and these parameters are central to plant yield, a major outcome in the context of high biomass production. To unravel the symbiotic N "transportome" blueprint from various host plant combinations, it is critical to facilitate the first steps favoring the manipulation of crops toward greater nitrogen use efficiency and mycorrhizal or rhizobial ability. The present review addresses current knowledge on inorganic N transport in mycorrhizal/rhizobial symbiosis

Evaluation of antibacterial properties of fluoride-containing mouth rinses differing in their acidic compound using a Streptococcus mutans biofilm

This in vitro study assessed the antibacterial effect on Streptococcus mutans biofilms of mouth rinses with 700 ppm F- (derived from NaF) that differed only in their acid compounds (malic, citric, tartaric, fumaric, hydrochloric, phosphoric, and lactic acid) used to adjust pH. S. mutans (ATCC™ 25175) was grown for 22 h at 37°C, harvested, resuspended in simulated body fluid and biofilm formation followed for 24 h at 37°C. Thereafter, biofilms were treated with experimental rinses for 30 s and placed in TAM48 isothermal microcalorimeter at 37°C for 72 h. Applying Gompertz growth model the parameters lag time and growth rate were determined from heat flow curves; additionally, reduction of active biofilms was calculated. Moreover, samples were live/dead-stained and analysed by confocal scanning microscopy. All mouth rinses were showing statistically significant lag time and reduction of active biofilm (p < 0.05, A 19.1 ± 2.3 h and 58.5 ± 7.7%, B 15.5 ± 1.1 h and 41.9 ± 5.3%, C 17.6 ± 1.9 h and 53.1 ± 7.5%, D 18.4 ± 2.4 h and 55.8 ± 8.8%, E 20.2 ± 3.3 h and 61.5 ± 10.0%, F 20.2 ± 3.0 h and 61.6 ± 9.3%, and G 18.3 ± 2.5 h and 55.3 ± 8.9%). Interestingly, there were no differences found between the treated groups (p > 0.05, A 0.064 ± 0.004 1/h, B 0.063 ± 0.005 1/h, C 0.065 ± 0.004 1/h, D 0.067 ± 0.004 1/h, E 0.066 ± 0.006 1/h, F 0.067 ± 0.004 1/h, G 0.066 ± 0.006 1/h) for the maximum growth rate. Vitality staining supported these findings. The present investigation demonstrates that the type of acid compounds used to produce the rinses did not show any negative effect on the antimicrobial properties of the tested products as all of them exhibited a similar efficacy against S. mutans biofilms

The largest subunit of RNA polymerase II as a new marker gene to study assemblages of arbuscular mycorrhizal fungi in the field.

Due to the potential of arbuscular mycorrhizal fungi (AMF, Glomeromycota) to improve plant growth and soil quality, the influence of agricultural practice on their diversity continues to be an important research question. Up to now studies of community diversity in AMF have exclusively been based on nuclear ribosomal gene regions, which in AMF show high intra-organism polymorphism, seriously complicating interpretation of these data. We designed specific PCR primers for 454 sequencing of a region of the largest subunit of RNA polymerase II gene, and established a new reference dataset comprising all major AMF lineages. This gene is known to be monomorphic within fungal isolates but shows an excellent barcode gap between species. We designed a primer set to amplify all known lineages of AMF and demonstrated its applicability in combination with high-throughput sequencing in a long-term tillage experiment. The PCR primers showed a specificity of 99.94% for glomeromycotan sequences. We found evidence of significant shifts of the AMF communities caused by soil management and showed that tillage effects on different AMF taxa are clearly more complex than previously thought. The high resolving power of high-throughput sequencing highlights the need for quantitative measurements to efficiently detect these effects

Expression of Sorghum bicolor ammonium transporters upon colonization with arbuscular mycorrhizal fungi

International audienc

Photosynthesis mediates expression of Sorghum bicolor transporters in arbuscular mycorrhizal symbiosis

National audienc

Isolation and identification of desert habituated arbuscular mycorrhizal fungi newly reported from the Arabian Peninsula

Arbuscular mycorrhizal fungi (AMF) are known to facilitate the growth and vigour of many plants, particularly in arid ecosystems. In a survey of AMF in a date palm plantation and two natural sites of a desert in Oman, we generated many single spore-derived cultures of AMF. We identified a number of these isolates based on spore morphotyping and molecular phylogenetic analysis using the sequence of the LSU-rDNA. Here, we presented the characteristics of four species of AMF recovered, namely Claroideoglomus drummondii, Diversispora aurantia, Diversispora spurca and Funneliformis africanum. The four species have been described previously, but for the Arabian Peninsula they are reported here for the first time. Our endeavor of isolation and characterization of some AMF habituated to arid sites of Arabia represents a first step towards application for environmental conservation and sustainable agriculture in this region