Fungal Inoculation Research Articles

Boosting species evenness, productivity and weed control in a mixed meadow by promoting arbuscular mycorrhizas.

Lowland meadows represent aboveground and belowground biodiversity reservoirs in intensive agricultural areas, improving water retention and filtration, ensuring forage production, contrasting erosion and contributing to soil fertility and carbon sequestration. Besides such major ecosystem services, the presence of functionally different plant species improves forage quality, nutritional value and productivity, also limiting the establishment of weeds and alien species. Here, we tested the effectiveness of a commercial seed mixture in restoring a lowland mixed meadow in the presence or absence of inoculation with arbuscular mycorrhizal (AM) fungi and biostimulation of symbiosis development with the addition of short chain chito-oligosaccharides (CO). Plant community composition, phenology and productivity were regularly monitored alongside AM colonization in control, inoculated and CO-treated inoculated plots. Our analyses revealed that the CO treatment accelerated symbiosis development significantly increasing root colonization by AM fungi. Moreover, the combination of AM fungal inoculation and CO treatment improved plant species evenness and productivity with more balanced composition in forage species. Altogether, our study presented a successful and scalable strategy for the reintroduction of mixed meadows as valuable sources of forage biomass; demonstrated the positive impact of CO treatment on AM development in an agronomic context, extending previous observations developed under controlled laboratory conditions and leading the way to the application in sustainable agricultural practices.

Fungal symbiont diversity drives growth of Holcus lanatus depending on soil nutrient availability

Abstract Arbuscular mycorrhizal (AM) fungi frequently colonise plant roots and can affect plant morphology and physiology through their contribution to plant nutrition. However, the functional role of AM fungi in the presence of other microbial symbionts, including widespread Mucoromycotina ‘fine root endophytes’ (MFRE) fungi, remains largely unknown. While both AM fungi and MFRE transfer nutrients, including nitrogen, from inorganic and organic sources to host plants, their combined effects on co‐colonised plants have only been investigated in liverworts. Here, we compare the morphology and physiology of the grass Holcus lanatus grown with an AM fungal community versus a more diverse symbiotic fungal community containing both AM fungi and MFRE. Holcus lanatus plants were grown in the presence of either a diverse MFRE+AM fungi soil inoculum or a multi‐species AM fungal inoculum. Plant traits associated with growth were quantified, along with fungal transfer of 15N tracer to plants from a variety of sources (ammonium chloride, alanine, glycine and algal necromass). Holcus lanatus grown with the AM fungal community had greater root and shoot growth during early development and prior to the addition of 15N‐labelled sources, compared with plants grown with the more diverse symbiotic fungal community. When nitrogen sources were made available to the fungal symbionts in the pot microcosms, plants growing with the MFRE+AM fungi soil inoculum had a faster growth rate than plants growing with the AM fungal community. At harvest, H. lanatus grown with the AM fungal community had a larger biomass, and there were no differences in 15N tracer assimilation in plants across the two fungal community treatments. Our results demonstrate that the diversity of fungal inocula in conjunction with soil nutrient availability determine the benefits derived by plants from diverse fungal symbionts. Our research contributes to understanding host plant outcomes in diverse multi‐symbiont scenarios. Read the free Plain Language Summary for this article on the Journal blog.

Comparative Response of Arbuscular Mycorrhizal Fungi versus Endophytic Fungi in Tangor Citrus: Photosynthetic Efficiency and P-Acquisition Traits

Citrus plants are prone to phosphorus (P) deficiency, especially in acidic soil, making them more dependent on root-associated endophytic fungi for growth and development. Beni-Madonna, a hybrid of Citrus nanko × C. amakusa, is a citrus known as “tangor” that is highly popular in China and other parts of the world due to its deep red color and jelly-like flesh. In this study, the inoculation response to two arbuscular mycorrhizal fungi (Diversispora versiformis, Dv; Funneliformis mosseae, Fm) and an endophytic fungus (Serendipita indica, Si) with regard to the growth, leaf gas exchange, light energy efficiency, P levels, acid phosphatase activity, and expression of the purple acid phosphatase (PAP) and phosphate transporter (PT) genes through a potted experiment using Beni-Madonna (tangor) citrus plants grafted on trifoliate orange (Poncirus trifoliata) was studied. Two years following inoculation, the root fungal colonization rates of inoculated plants were significantly increased, accompanied by an increase in plant height and stem diameter, with Si presenting the best effect. Fungal inoculations also significantly increased the leaf chlorophyll index, nitrogen-balance index, photosynthesis rate, stomatal conductance, transpiration rate, photosynthetic efficiency, and quantum yield of PSII, while it reduced photoinhibition and heat dissipation, with Si having the best effect on light energy efficiency. Si significantly increased leaf and root P levels, as well as root CsPAP1‒3, CsPT2, CsPT3, and CsPT7 expression and soil acid phosphatase activity; Fm in mycorrhizal fungi significantly increased leaf and root P levels, as well as root CsPT1 and CsPT7 expression and root/soil acid phosphatase activity. These observations suggested an advantage of Si over the other two AMF in terms of improved plant growth and P acquisition, while Fm displayed prominent effects on increased photosynthetic efficiency.

Arbuscular mycorrhizal fungi suppress ammonia-oxidizing bacteria but not archaea across agricultural soils

Arbuscular mycorrhizal (AM) fungi are supposedly competing with ammonia-oxidizing microorganisms (AO) for soil nitrogen in form of ammonium. Despite a few studies directly addressing AM fungal and AO interactions, mostly in artificial cultivation substrates, it is not yet clear whether AM fungi can effectively suppress AO in field soils containing complex indigenous microbiomes. To fill this knowledge gap, we conducted compartmentalized pot experiments using four pairs of cropland and grassland soils with varying physicochemical properties. To exclude the interference of roots, a fine nylon mesh was used to separate the rhizosphere and mesh bags, with the latter being filled with unsterile field soils. Inoculation of plants with AM fungus Rhizophagus irregularis LPA9 suppressed AO bacteria (AOB) but not archaea (AOA) in the soils, indicating how soil nitrification could be suppressed by AM fungal presence/activity. In addition, in rhizosphere filled with artificial substrate, AM inoculation did suppress both AOB and AOA, implying more complex interactions between roots, AO, and AM fungi. Besides, we also observed that indigenous AM fungi contained in the field soils eventually did colonize the roots of plants behind the root barrier, and that the extent of such colonization was higher if the soil has previously been taken from cropland than from grassland. Despite this, the effect of experimental AM fungal inoculation on suppression of indigenous AOB in the unsterile field soils did not vanish. It seems that studying processes at a finer temporal scale, using larger buffer zones between rhizosphere and mesh bags, and/or detailed characterization of indigenous AM fungal and AO communities would be needed to uncover further details of the biotic interactions between the AM fungi and indigenous soil AO.

First Report of Lasiodiplodia pseudotheobromae Causing Postharvest Decay of Strawberries in Florida

Postharvest decay of strawberry (Fragaria × ananassa Duch.) is a major factor causing fruit losses. Strawberries were obtained from various harvests at cooling facilities located in Dover and Plant City, FL during the 2018-19 and 2019-20 seasons. After the fruits were incubated at 22ºC for up to 5 days (d) to promote disease development, Lasiodiplodia decay was observed at up to 3% from some harvests, exhibiting gray mycelia on small lesions that gradually covered the whole fruit. The fungus was isolated onto potato dextrose agar (PDA). Five isolates (SBD18-14, SBD18-277, SBD18-279, SBD19-02 and SBD19-57) were characterized. Fungal mycelia were initially grayish white and then gradually changed to gray to dark gray on PDA at 25oC, and later produced black pigments (Fig. S1). Pycnidia were observed from inoculated strawberries at 14 d. Isolates shared similar conidia morphology: aseptate, hyaline, ellipsoid to ovoid, measuring L × W: 24.0-34.0 (28.3) × 13.0-16.0 (14.3) μm (n =100). Mature conidia were brown, one septate, measuring L × W: 25.0-33.0 (28.8) × 13.0-16.0 (14.5) μm (n =100). The isolates were identified as Lasiodiplodia spp. morphologically (Alves et al. 2008). DNA was extracted from fungal mycelia using an OmniPrep DNA extraction kit, and PCR amplification of ITS and EF1-α genes was performed following the conditions described by White et al. (1990) with some modifications using primers ITS1F-F/ITS4-R (Gardes and Bruns, 1993; White et al., 1990) and EF1-668-F/EF1-1251-R (Alves et al., 2008), respectively. The BLASTn in GenBank showed that the sequences obtained had 99.61 to 100% homology with those of ITS (EF622077) and EF1-α (EF622057) from L. pseudotheobromae CBS116459 (an ex-type strain) (Alves et al., 2008). Sequences of the isolates have been deposited in GenBank with accessions OP326017 to OP326021 for ITS, and OP356202 to OP356206 for EF1-α. Phylogenetic analysis showed that these isolates clustered in the same clade (bootstrap value at 64) with L. pseudotheobromae (Fig. S2). Two fungal inoculum types (mycelia and conidia), two fruit inoculation methods (injury and non-injury) and five fungal isolates were used for pathogenicity tests. Fungal mycelia (2-day-old) on PDA plug (5 mm) or 10 µL of conidial suspension (106 spores/mL) was placed onto each injury (1 x 1 mm in size) or a non-injury area on the surfaces of five strawberry fruits (cv. Florida Brilliance). PDA plug alone or water drops placed on injury or non-injury areas on fruits served as respective controls. Inoculated and control fruits were incubated in a covered plastic container with 100% RH at 22ºC. The experiment was repeated twice. Decay initially appeared as soft and lightly discolored tissue at inoculation areas 2 d post-inoculation (dpi) that extended quickly thereafter. Brown to dark lesions on both injury- and non-injury fruits inoculated with conidia or mycelia were observed at 3 dpi. Decay and gray mycelia gradually developed over the whole fruit at 6 dpi, and pycnidia were observed after 14 dpi (Fig. S1). Disease incidence of 100% was observed on all tests. Control fruits did not develop decay. The results indicate that these isolates are pathogenic to strawberries and infect fruit via both non-injured and injured fruit surfaces. The inoculated fungal isolates were re-isolated, thus, fulfilling Koch’s postulates. L. theobromae, Neofusicoccum parvum/N. ribis species complex causing strawberry fruit rot in Florida fields was reported (Oliveira et al., 2019), but not L. pseudotheobromae. To our knowledge, this is the first report of postharvest decay caused by L. pseudotheobromae A.J.L. Phillips, A. Alves & Crous on strawberries in Florida and in the USA, and it should be considered in strawberry disease management.

Attenuation of Zucchini mosaic virus disease in cucumber plants by mycorrhizal symbiosis

Key messageArbuscular mycorrhizal fungi generated systemic acquired resistance in cucumber to Zucchini yellow mosaic virus, indicating their prospective application in the soil as a sustainable, environmentally friendly approach to inhibit the spread of pathogens.The wide spread of plant pathogens affects the whole world, causing several plant diseases and threatening national food security as it disrupts the quantity and quality of economically important crops. Recently, environmentally acceptable mitigating practices have been required for sustainable agriculture, restricting the use of chemical fertilizers in agricultural areas. Herein, the biological control of Zucchini yellow mosaic virus (ZYMV) in cucumber (Cucumis sativus L.) plants using arbuscular mycorrhizal (AM) fungi was investigated. Compared to control plants, ZYMV-infected plants displayed high disease incidence (DI) and severity (DS) with various symptoms, including severe yellow mosaic, mottling and green blisters of leaves. However, AM fungal inoculation exhibited 50% inhibition for these symptoms and limited DS to 26% as compared to non-colonized ones. The detection of ZYMV by the Enzyme-Linked Immunosorbent Assay technique exhibited a significant reduction in AM-inoculated plants (5.23-fold) compared with non-colonized ones. Besides, mycorrhizal root colonization (F%) was slightly reduced by ZYMV infection. ZYMV infection decreased all growth parameters and pigment fractions and increased the malondialdehyde (MDA) content, however, these parameters were significantly enhanced and the MDA content was decreased by AM fungal colonization. Also, the protein, proline and antioxidant enzymes (POX and CAT) were increased with ZYMV infection with more enhancements due to AM root colonization. Remarkably, defence pathogenesis-related (PR) genes such as PR-a, PR-b, and PR-10 were quickly expressed in response to AM treatment. Our findings demonstrated the beneficial function of AM fungi in triggering the plant defence against ZYMV as they caused systemic acquired resistance in cucumber plants and supported their potential use in the soil as an environment-friendly method of hindering the spread of pathogenic microorganisms sustainably.

Impact of bacterial and fungal inoculants on the resident rhizosphere microbiome and the volatilome of tomato plants under leaf herbivory stress.

Various studies have addressed the impact of microbial inoculants on the composition of the resident microbiome. How microbial inoculants impact plant metabolism and interact with the resident rhizobiota under herbivory stress remains elusive. Here, we investigated the impact of two bacterial and two fungal inoculants, inoculated as single species and as a synthetic community, on the rhizosphere microbiome and volatilome of tomato plants (Solanum lycopersicum) comparing nonstress conditions to exposed to leaf herbivory by Spodoptera exigua. Based on amplicon sequencing analysis, rhizobacterial community composition was significantly affected by all four inoculants and the magnitude of this effect was dependent on herbivory stress. Fungal community composition was altered by the microbial inoculants but independent of herbivory stress. The rhizosphere volatilome was impacted by the microbial inoculation and differences between treatments were evened under herbivory stress. Each microbial inoculant caused unique changes in the volatilome of stressed plants but also shared similar responses, in particular the enhanced production of dimethyl disulfide and benzothiazole. In conclusion, the introduction of microbial inoculants in the tomato rhizosphere caused unique as well as common changes in the rhizosphere microbiome and volatilome, but these changes were minor compared to the microbiome changes induced by herbivory stress.

Divergent responses between lineages of arbuscular mycorrhizal fungi to soil phosphorus and nitrogen availability

Arbuscular mycorrhizal (AM) associations are multifunctional. Two important functions they perform are facilitating nutrient uptake in host plants and protecting plants from biotic stress, among other functions. AM fungal taxa vary in how capably they perform these functions and can also respond differently to environmental selection. Therefore, there is a need to better understand how particular environmental variables might alter the response of AM fungal communities. Here, we analysed data from a DNA-based survey of fungal communities in soils collected throughout Australia to observe relationships among soil nitrogen and phosphorus availability and the abundance of two AM fungal taxa that reportedly vary in function – the Gigasporaceae (putatively more important for nutrient uptake) and Glomeraceae (putatively more important for biotic stress). Relationships were assessed in three vegetation types – grasslands, forests and woodlands – to assess whether associations with soil nutrition varied depending on carbon availability for AM fungi. Fungi from the Gigasporaceae decreased in frequency as available phosphorus increased, while those from the Glomeraceae increased or were unresponsive as available phosphorus increased. Similar patterns were observed for nitrate availability, although only in woodlands. These patterns are consistent with expectations that AM fungi from the Gigasporaceae, in general, are better suited to alleviate nutrient limitation in hosts as soil chemical fertility decreases. This knowledge may aid in implementing optimal strategies involving AM fungal inoculum best suited to the local conditions of future land management and agricultural projects.

Volatile organic compounds as potential biomarkers of Cadophora luteo‐olivacea presence on kiwifruits

AbstractCadophora luteo‐olivacea is the causal agent of the skin‐pitting disease of kiwifruit, a syndrome that appears after 4–5 months of cold storage. However, it is assumed that the infection takes place in the field during fruit development. The present work takes into consideration the production of volatile organic compounds (VOCs) of Actinidia deliciosa ‘Hayward’ at different phenological phases as potential C. luteo‐olivacea infection biomarkers. In vitro assays were conducted to gain knowledge on the effect of kiwifruit VOCs on pathogen conidial germination and mycelial growth. VOCs produced by kiwifruit either inoculated or not with C. luteo‐olivacea were analysed at different phenological phases by SPME/GC–MS analysis. In particular, ethanol, o‐xylene, d‐limonene and acetic acid showed a significant increase in the presence of fungal inoculation. Ethanol and d‐limonene were also detected as volatile metabolites of the pathogen. The effect of each compound (ethanol, o‐xylene, d‐limonene and acetic acid) was tested on the fungal conidial germination at different concentrations, showing a growth stimulation at lower amounts. These results show how the production of some VOCs can contribute to the knowledge of fruit–pathogen interaction in the field with the aim of developing future tools for early disease detection and consequent effective control.

The effects of diverse microbial community structures, driven by arbuscular mycorrhizal fungi inoculation, on carbon release from a paddy field

The effects of diverse microbial community structures, driven by arbuscular mycorrhizal fungi inoculation, on carbon release from a paddy field

Microbial Consortia Versus Single-Strain Inoculants as Drought Stress Protectants in Potato Affected by the Form of N Supply

This study investigated the drought protection effects of six fungal and bacterial inoculants and ten consortia thereof on vegetative growth, nutritional status, and tuberization of potato under controlled and field conditions. It was hypothesized that microbial consortia offer improved drought protection as compared with single strains, due to complementary or synergistic effects, with differential impacts also of N fertilization management. Under NO3− fertilization, a 70% reduction in water supply over six weeks reduced shoot and tuber biomass of non-inoculated plants by 30% and 50%, respectively, and induced phosphate (P) limitation compared to the well-watered control. The P nutritional status was significantly increased above the deficiency threshold by three single-strain inoculants and eight consortia. This was associated with the presence of the arbuscular mycorrhizal fungus (AMF) inoculant Rhizophagus irregularis MUCL41833 (five cases) and stimulation of root growth (five cases). Additionally, Bacillus amyloliquefaciens FZB42 and AMF + Pseudomonas brassicacearum 3Re2-7 significantly reduced irreversible drought-induced leaf damage after recovery to well-watered conditions. However, the microbial inoculants did not mitigate drought-induced reductions in tuber biomass, neither in greenhouse nor in field experiments. By contrast, NH4+-dominated fertilization significantly increased tuber biomass under drought stress (534%), which was further increased by additional AMF inoculation (951%). This coincided with (i) improved enzymatic detoxification of drought-induced reactive oxygen species (ROS), (ii) improved osmotic adjustment in the shoot tissue (glycine betaine accumulation), (iii) increased shoot concentrations of ABA, jasmonic acid, and indole acetic acid, involved in drought stress signaling and tuberization, and (iv) reduced irreversible drought-induced leaf damage. Additional application of bacterial inoculants further improved ROS detoxification by increasing the production of antioxidants but stimulated biomass allocation towards shoot growth at the expense of tuber development. The results demonstrated that microbial consortia could increase the probability of drought protection effects influenced by the form of N supply. However, protective effects on vegetative growth do not necessarily translate into yield benefits, which can be achieved by adequate combination of inoculants and fertilizers.

Effect of sub-optimal moisture levels on the quality of groundnut (Arachis hypogaea L.) during storage in triple-layer hermetic storage bags

Storage is an important aspect of groundnut, as the in-shell and shelled kernels are prone to infestation by insects, pests, and fungi. Among several storage pests, the groundnut bruchid, Caryedon serratus, causes serious losses. Farmers often resort to different management practices, including hermetic storage, to control it. The moisture content of the commodity plays an important role in insect infestation during storage. Drying to safe moisture levels before storage is known to prevent the activity of various living organisms, such as storage pests. However, drying to low levels of moisture may not be economical for farmers, as they may not have access to devices to accurately check product moisture. In this regard, we wanted to demonstrate the efficacy of triple-layer hermetic storage bags in preventing the damage caused by C. serratus when the groundnuts are stored at intermediate (10%) and high (14%) levels of moisture compared to traditionally used bags such as polypropylene bags and jute bags. Groundnut pods at 10% moisture content and 14% moisture content were separately inoculated with adult bruchids and a toxigenic strain of Aspergillus flavus fungal inoculum before storing them for 6 months. Results from groundnut samples taken at two-month intervals indicated that groundnut pods stored in triple-layer hermetic bags were completely free from infestation by C. serratus by recording a zero number of eggs laid, number of pupae, adult emergence, percentage of loss, and percentage of damage up to 6 months of storage, by creating low oxygen (hypoxia) and high carbon dioxide (hypercarbia) conditions. Results also indicate no loss of pod weight stored in triple-layer bags, but a slight reduction in germination percentage was recorded due to a slight increase in fungal activity, but the reduction was significantly less in triple-layer plastic bags compared to other bag types. Similarly, biochemical constituents such as oil and protein content were slightly reduced in triple-layer plastic bags when pods were stored at a 10% moisture level, but a higher reduction was observed at a 14% moisture level. However, the reduction was very high and significant in other bag types at both 10 and 14% moisture levels.

Overexpression of cucumber CYP82D47 enhances resistance to powdery mildew and Fusarium oxysporum f. sp. cucumerinum.

Cytochrome P450s are a large family of protein-encoding genes in plant genomes, many of which have not yet been comprehensively characterized. Here, a novel P450 gene, CYP82D47, was isolated and functionally characterized from cucumber (Cucumis sativus L.). Quantitative real-time reverse-transcription polymerase chain reaction analysis revealed that CYP82D47 expression was triggered by salicylic acid (SA) and ethephon (ETH). Expression analysis revealed a correlation between CYP82D47 transcript levels and plant defense responses against powdery mildew (PM) and Fusarium oxysporum f. sp. cucumerinum (Foc). Although no significant differences were observed in disease resistance between CYP82D47-RNAi and wild-type cucumber, overexpression (OE) of CYP82D47 enhanced PM and Foc resistance in cucumber. Furthermore, the expression levels of SA-related genes (PR1, PR2, PR4, and PR5) increased in CYP82D47-overexpressing plants 7days post fungal inoculation. The levels of ETH-related genes (EIN3 and EBF2) were similarly upregulated. The observed enhanced resistance was associated with the upregulation of SA/ETH-signaling-dependent defense genes. These findings indicate the crucial role of CYP82D47 in pathogen defense in cucumber. CYP82D47-overexpressing cucumber plants exhibited heightened susceptibility to both diseases. The study results offer important insights that could aid in the development of disease-resistant cucumber cultivars and elucidate the molecular mechanisms associated with the functions of CYP82D47.

Variation Among Japanese Miso Breweries in Indoor Microbiomes is Mainly Ascribed to Variation in Type of Indoor Surface

Miso is a microbially-fermented soybean food. The miso brewery indoor microbiome contributes to miso fermentation. Japanese breweries are not climate-controlled, so indoor spaces are strongly affected by the prevailing climate. Because climate influences microorganism distribution, our first hypothesis is that latitude, as a proxy for climate, is a major determinant of brewery indoor microbiome structure. Breweries vary in interior surface materials and in the way operations (steaming, processing, fermenting) are apportioned among rooms. Therefore, our second hypothesis is that more variability in indoor microbiomes exists among breweries than can be ascribed to a latitudinal gradient. Most miso produced today is inoculated with commercial microbial strains to standardize fermentation. If commercial strains outcompete indigenous microbes for membership in the indoor microbiome, this practice may homogenize indoor microbiomes among regions or breweries. Therefore, our third hypothesis is that inoculant fungal species dominate indoor fungal communities and make it impossible to distinguish communities among breweries or across their latitudinal gradient. We tested these hypotheses by sampling indoor surfaces in several breweries across a latitudinal gradient in Japan. We found that latitude had a significant but relatively small impact on indoor fungal and bacterial communities, that the effect of brewery was large relative to latitude, and that inoculant fungi made such small contributions to the indoor microbiome that distinctions among breweries and along the latitudinal gradient remained apparent. Recently, the Japanese Ministry of Agriculture, Forestry and Fisheries specified fungal inoculants to standardize miso production. However, this may not be possible so long as the indoor microbiome remains uncontrolled.

Does arbuscular mycorrhizal fungi inoculation influence soil carbon sequestration?

Whether arbuscular mycorrhizal fungi (AMF) inoculation promotes soil C sequestration is largely unknown. Here, meta-analysis and logistic regression were applied to study the ecological effects of AMF inoculation on soil organic C (SOC) turnover and plant growth under different inoculation manipulations, plant traits, and soil conditions. Results showed that AMF inoculation generally increased SOC stock and plant biomass accumulation. Soil sterilization, unsterilized inoculum wash (a filtrate of mycorrhizal inoculum excluding AMF) addition in non-mycorrhizal treatments, experimental type, and inoculated AMF species influenced soil microbial biomass C (MBC) but had no impact on SOC turnover. Plant root system, initial SOC content, and soil pH were the key factors that influenced the AMF-mediated SOC turnover. AMF inoculation in fertile or acidic soils might deplete SOC. The symbiosis between tap-rooted plants and AMF was more likely to sequestrate C into the soil compared to fibrous-rooted plants. Moreover, plant total dry biomass largely relied on its own photosynthetic pathway although AMF was introduced. Collectively, our results suggest that AMF inoculation is a promising approach for soil C sequestration.

Role of an arbuscular mycorrhizal fungus in vegetation restoration as indicated by bacterial diversity and microbial metabolic limitation in soil underlying moss biocrusts

Arbuscular mycorrhizal (AM) fungi contribute to the revegetation of degraded ecosystems such as coal mining areas and may indirectly affect the development of biological soil crusts (biocrusts). However, bacterial diversity and microbial nutrient cycling in the soil underlying biocrusts are seldom considered when assessing ecological restoration effects. Bacterial community composition was quantified and compared using 16S rRNA gene sequencing, metabolic limitation of microbes via extracellular enzymatic stoichiometry and their association with carbon use efficiency (CUE) in soil underlying moss biocrusts in vegetated areas inoculated with an AM fungus or uninoculated and in uninoculated unvegetated areas. The AM fungal inoculum significantly influenced bacterial community composition and diversity, extracellular enzyme activities and CUE in soil underlying moss biocrusts mainly by affecting soil nutrients. Correlation analysis revealed that Actinobacteria, Armatimonadetes, Bacteroidetes, Chlorobi, Cyanobacteria, Saccharibacteria, and Verrucomicrobia were positively correlated with soil nutrients carbon (C), nitrogen (N) and phosphorus (P). A vector analysis of extracellular enzyme activity indicates that soil microbial communities underlying moss biocrusts were limited by P but microbial communities in areas inoculated with the AM fungus had the lowest relative P limitation. The alleviation of microbial P limitation increased the microbial CUE and bacterial phylum community and alpha diversity, leading to an increase in subsoil C content underlying moss biocrusts. This indicates that microbial communities in soil underlying moss biocrusts under in restoration areas containing the AM fungus were more stable under environmental stress, and the inclusion of AM fungal inoculation may be recommended as the preferred option for ecosystem restoration in arid and semi-arid coal mining areas.

Limited effects of plant-beneficial fungi on plant volatile composition and host-choice behavior of Nesidiocoris tenuis.

Biological control using plant-beneficial fungi has gained considerable interest as a sustainable method for pest management, by priming the plant for enhanced defense against pathogens and insect herbivores. However, despite promising outcomes, little is known about how different fungal strains mediate these beneficial effects. In this study, we evaluated whether inoculation of tomato seeds with the plant-beneficial fungi Beauveria bassiana ARSEF 3097, Metarhizium brunneum ARSEF 1095 and Trichoderma harzianum T22 affected the plant's volatile organic compound (VOC) profile and the host-choice behavior of Nesidiocoris tenuis, an emerging pest species in NW-European tomato cultivation, and the related zoophytophagous biocontrol agent Macrolophus pygmaeus. Results indicated that fungal inoculation did not significantly alter the VOC composition of tomato plants. However, in a two-choice cage assay where female insects were given the option to select between control plants and fungus-inoculated plants, N. tenuis preferred control plants over M. brunneum-inoculated plants. Nearly 72% of all N. tenuis individuals tested chose the control treatment. In all other combinations tested, no significant differences were found for none of the insects. We conclude that inoculation of tomato with plant-beneficial fungi had limited effects on plant volatile composition and host-choice behavior of insects. However, the observation that N. tenuis was deterred from the crop when inoculated with M. brunneum and attracted to non-inoculated plants may provide new opportunities for future biocontrol based on a push-pull strategy.

Insecticidal and antioxidant potential of volatile compounds and nanoparticles from Tulbaghia violacea Harv. inoculated with endophytic fungus Beauveria bassiana (Bals.-Criv.) Vuill

This study aimed to investigate the effect of the entomopathogenic fungus Beauveria bassiana on the growth and ability of Tulbaghia violacea to synthesise nanoparticles and volatile compounds. Tulbaghia violacea was inoculated by drenching with different concentrations of B. bassiana conidial concentrations (0 mL−1, 1 × 106 mL−1, 1 × 107, and 1 × 108 conidia mL−1) and the colonization of plant tissue in the different fungal treatments was evaluated. A UV–VIS spectrometer (SPECTROstar nano) was used to determine the presence of nanoparticles while volatile constituents in the plants were analysed with chromatography-mass spectrometry. Polyphenols, alkaloids, flavonoids and antioxidant contents were determined using referenced methods. At least 57 % of plants were successfully colonized at the highest concentration (1 × 108 conidia mL−1). The results showed that treating B. bassiana with T. violacea had no significant effect (P>0.05) on the growth parameters (height, number of leaves, aerial part fresh weight, aerial part dry weight, root fresh weight, and root dry weight) of T. violacea after four weeks post-treatment. The aqueous extracts of T. violacea from both control and B. bassiana-treated plants successfully synthesized gold nanoparticles. Furthermore, the aqueous extracts of plants and gold nanoparticles from both fungus and control treatments were equally toxic to mealybug and induced high mortality. The Gas chromatography–mass spectrometry (GC–MS) results revealed that the number of compounds detected in plants subjected to the different concentrations of fungal inoculum and control did not vary significantly (P>0.05) in leaves and roots. The polyphenol contents in the leaves were not significantly different (P>0.05) among treatments. Similarly, the antioxidant activities were not significantly influenced (P>0.05) by fungal treatment on both leaves and roots. However, T. violacea extracts reduced aqueous gold to gold nanoparticles in both control and fungus-treated plants.

Drought stress reduces arbuscular mycorrhizal colonization of Poncirus trifoliata (L.) roots and plant growth promotion via lipid metabolism.

Drought stress poses increasingly serious threats to agricultural production in the era of global climate change. Arbuscular mycorrhizal (AM) fungi are well-recognized biostimulants promoting plant tolerance to drought stress. Lipids are indispensable for AM fungal colonization, however, the involvement of lipid metabolism in the drought tolerance conferred by AM fungi is largely unknown. In this study, we inoculated Poncirus trifoliata (L.) with Rhizophagus irregularis DAOM197198 under no drought stress, medium drought stress and severe drought stress, with non-inoculation under respective treatments as control. Results indicated that AM fungal inoculation significantly promoted the drought tolerance of P. trifoliata (L.), with the effect size decreasing along with drought severity. Moreover, the effect size was significantly related to arbuscule abundance. Fatty acid profiling showed that the arbuscule abundance was determined by the AM-specific phospholipids (PLs), whose biosynthesis and delivery were inhibited by drought stress as revealed by qRT-PCR of FatM, RAM1 and STR/STR2. More interestingly, AM fungal inoculation increased the lipid allocation to total PLs and the unsaturation rate of total neutral lipids (NLs), probably indicating the involvement of non-AM-specific lipids in the increased drought tolerance. Taken together, our results demonstrate that lipid metabolism in AM mediates the increased drought tolerance conferred by AM fungal inoculation, with AM-specific and non-AM-specific lipids functioning therein in different ways.

Elucidation of mechanisms underlying active oxygen burst in Citrus sinensis after Diaporthe citri infection using transcriptome analysis.

Reactive oxygen species (ROS) generation is a common disease defense mechanism in plants. However, it is unclear whether Citrus host activates defense response against Diaporthe citri causing citrus melanose disease by producing ROS, and the underlying molecular mechanisms are unknown. DAB staining and RNA-Seq technology were used to compare the active oxygen burst and differential gene expression, respectively, in uninfected and infected Citrus sinensis leaves at different time points during D. citri infection in vivo. The functions of CsRBOH (a significant DEG) were confirmed in N. benthamiana through the Agrobacterium-mediated transient expression system. DAB staining indicated that C. sinensis initiated defense against D. citri infection within 24 h by generating ROS. Illumina sequencing revealed 25,557 expressed genes of C. sinensis. The most upregulated DEGs (n = 1,570) were identified 72 h after fungal inoculation (sample denoted as CD72). In the CD72 vs. Cs (samples at 0 h after fungal inoculation) comparison, the KEGG pathway category with the highest number of genes (n = 62) and most significant enrichment was Protein processing in endoplasmic reticulum, followed by Glutathione metabolism and MAPK signaling pathway-plant. GO analysis revealed that the DEGs of CD72 vs. Cs related to active oxygen burst and chitin recognition were significantly grouped into the regulation of biological processes and molecular functions, with GO terms including response to ROS, response to fungus, and oxidoreductase activity. Remarkably, CsRBOH was significantly enriched in the GO and KEGG analyses, and its expression pattern in qRT-PCR and DAB staining results were consistent. Among the 63 ROS-related DEGs, HSP genes and genes associated with the peroxidase family were highly significant as revealed by protein-protein interaction networks. Furthermore, ROS accumulation, cell death, and upregulation of defense-related genes were observed in N. benthamiana leaves with CsRBOH expressed through the Agrobacterium-mediated transient expression system. Our findings suggested that C. sinensis activates CsRBOH and ROS-related genes, leading to ROS accumulation to resist the invasion by D. citri. This study laid the foundation for future research on molecular mechanisms and breeding of C. sinensis cultivars resistant to citrus melanose.