Soil Chemistry and Soil History Significantly Structure Oomycete Communities in Brassicaceae Crop Rotations.

Oomycetes are critically important in soil microbial communities, especially for agriculture, where they are responsible for major declines in yields. Unfortunately, oomycetes are vastly understudied compared to bacteria and fungi. As such, our understanding of how oomycete biodiversity and community structure vary through time in the soil remains poor. Soil history established by previous crops is one factor known to structure other soil microbes, but this has not been investigated for its influence on oomycetes. In this study, we established three different soil histories in field trials; the following year, these plots were planted with five different Brassicaceae crops. We hypothesized that the previously established soil histories would structure different oomycete communities, regardless of their current Brassicaceae crop host, in both the roots and rhizosphere. We used a nested internal transcribed spacer amplicon strategy incorporated with MiSeq metabarcoding, where the sequencing data was used to infer amplicon sequence variants of the oomycetes present in each sample. This allowed us to determine the impact of different soil histories on the structure and biodiversity of the oomycete root and rhizosphere communities from the five different Brassicaceae crops. We found that each soil history structured distinct oomycete rhizosphere communities, regardless of different Brassicaceae crop hosts, while soil chemistry structured the oomycete communities more during a dry year. Interestingly, soil history appeared specific to oomycetes but was less influential for bacterial communities previously identified from the same samples. These results advance our understanding of how different agricultural practices and inputs can alter edaphic factors to impact future oomycete communities. Examining how different soil histories endure and impact oomycete biodiversity will help clarify how these important communities may be assembled in agricultural soils. IMPORTANCE Oomycetes cause global plant diseases that result in substantial losses, yet they are highly understudied compared to other microbes, like fungi and bacteria. We wanted to investigate how past soil events, like changing crops in rotation, would impact subsequent oomycete communities. We planted different oilseed crops in three different soil histories and found that each soil history structured a distinct oomycete community regardless of which new oilseed crop was planted, e.g., oomycete communities from last year's lentil plots were still detected the following year regardless of which new oilseed crops we planted. This study demonstrated how different agricultural practices can impact future microbial communities differently. Our results also highlight the need for continued monitoring of oomycete biodiversity and quantification.

A Comprehensive Insight of Current and Future Challenges in Large-Scale Soil Microbiome Analyses.

In the last decade, various large-scale projects describing soil microbial diversity across large geographical gradients have been undertaken. However, many questions remain unanswered about the best ways to conduct these studies. In this review, we present an overview of the experience gathered during these projects, and of the challenges that future projects will face, such as standardization of protocols and results, considering the temporal variation of microbiomes, and the legal constraints limiting such studies. We also present the arguments for and against the exhaustive description of soil microbiomes. Finally, we look at future developments of soil microbiome studies, notably emphasizing the important role of cultivation techniques.

Disentangling arbuscular mycorrhizal fungi and bacteria at the soil-root interface.

Arbuscular mycorrhizal fungi (AMF) are essential components of the plant root mycobiome and are found in approximately 80% of land plants. As obligate plant symbionts, AMF harbor their own microbiota, both inside and outside the plant root system. AMF-associated bacteria (AAB) possess various functional traits, including nitrogen fixation, organic and inorganic phosphate mobilization, growth hormone production, biofilm production, enzymatic capabilities, and biocontrol against pathogen attacks, which not only contribute to the health of the arbuscular mycorrhizal symbiosis but also promote plant growth. Because of this, there is increasing interest in the diversity, functioning, and mechanisms that underlie the complex interactions between AMF, AAB, and plant hosts. This review critically examines AMF-associated bacteria, focusing on AAB diversity, the factors driving richness and community composition of these bacteria across various ecosystems, along with the physical, chemical, and biological connections that enable AMF to select and recruit beneficial bacterial symbionts on and within their structures and hyphospheres. Additionally, potential applications of these bacteria in agriculture are discussed, emphasizing the potential importance of AMF fungal highways in engineering plant rhizosphere and endophyte bacteria communities, and the importance of a functional core of AAB taxa as a promising tool to improve plant and soil productivity. Thus, AMF and their highly diverse bacterial taxa represent important tools that could be efficiently explored in sustainable agriculture, carbon sequestration, and reduction of greenhouse gas emissions related to nitrogen fertilizer applications. Nevertheless, future studies adopting integrated multidisciplinary approaches are crucial to better understand AAB functional diversity and the mechanisms that govern these tripartite relationships.

Brassicaceae host plants mask the feedback from the previous year's soil history on bacterial communities, except when they experience drought.

Soil history operates through time to influence the structure and biodiversity of soil bacterial communities. Examining how different soil histories endure will help clarify the rules of bacterial community assembly. In this study, we established three different soil histories in field trials; the following year these plots were planted with five different Brassicaceae species. We hypothesized that the previously established soil histories would continue to structure the subsequent Brassicaceae bacterial root and rhizosphere communities. We used a MiSeq 16S rRNA metabarcoding strategy to determine the impact of different soil histories on the structure and biodiversity of the bacterial root and rhizosphere communities from the five different Brassicaceae host plants. We found that the Brassicaceae hosts were consistently significant factors in structuring the bacterial communities. Four host plants (Sinapis alba, Brassica napus, B. juncea, B. carinata) formed similar bacterial communities, regardless of different soil histories. Camelina sativa host plants structured phylogenetically distinct bacterial communities compared to the other hosts, particularly in their roots. Soil history established the previous year was only a significant factor for bacterial community structure when the feedback of the Brassicaceae host plants was weakened, potentially due to limited soil moisture during a dry year. Understanding how soil history is involved in the structure and biodiversity of bacterial communities through time is a limitation in microbial ecology and is required for employing microbiome technologies in improving agricultural systems.

Inter-Kingdom Networks of Canola Microbiome Reveal Bradyrhizobium as Keystone Species and Underline the Importance of Bulk Soil in Microbial Studies to Enhance Canola Production.

The subterranean microbiota of plants is of great importance for plant growth and health, as root-associated microbes can perform crucial ecological functions. As the microbial environment of roots is extremely diverse, identifying keystone microorganisms in plant roots, rhizosphere, and bulk soil is a necessary step towards understanding the network of influence within the microbial community associated with roots and enhancing its beneficial elements. To target these hot spots of microbial interaction, we used inter-kingdom network analysis on the canola growth phase of a long-term cropping system diversification experiment conducted at four locations in the Canadian Prairies. Our aims were to verify whether bacterial and fungal communities of canola roots, rhizosphere, and bulk soil are related and influenced by diversification of the crop rotation system; to determine whether there are common or specific core fungi and bacteria in the roots, rhizosphere, and bulk soil under canola grown in different environments and with different levels of cropping system diversification; and to identify hub taxa at the inter-kingdom level that could play an important ecological role in the microbiota of canola. Our results showed that fungi were influenced by crop diversification, which was not the case on bacteria. We found no core microbiota in canola roots but identified three core fungi in the rhizosphere, one core mycobiota in the bulk soil, and one core bacterium shared by the rhizosphere and bulk soil. We identified two bacterial and one fungal hub taxa in the inter-kingdom networks of the canola rhizosphere, and one bacterial and two fungal hub taxa in the bulk soil. Among these inter-kingdom hub taxa, Bradyrhizobium sp. and Mortierella sp. are particularly influential on the microbial community and the plant. To our knowledge, this is the first inter-kingdom network analysis utilized to identify hot spots of interaction in canola microbial communities.

Into the wild blueberry (Vaccinium angustifolium) rhizosphere microbiota.

The ability of wild blueberries to adapt to their harsh environment is believed to be closely related to their symbiosis with ericoid mycorrhizal fungi, which produce enzymes capable of organic matter mineralization. Although some of these fungi have been identified and characterized, we still know little about the microbial ecology of wild blueberry. Our study aims to characterize the fungal and bacterial rhizosphere communities of Vaccinium angustifolium (the main species encountered in wild blueberry fields). Our results clearly show that the fungal order Helotiales was the most abundant taxon associated with V. angustifolium. Helotiales contains most of the known ericoid mycorrhizal fungi which are expected to dominate in such a biotope. Furthermore, we found the dominant bacterial order was the nitrogen-fixing Rhizobiales. The Bradyrhizobium genus, whose members are known to form nodules with legumes, was among the 10 most abundant genera in the bacterial communities. In addition, Bradyrhizobium and Roseiarcus sequences significantly correlated with higher leaf-nitrogen content. Overall, our data documented fungal and bacterial community structure differences in three wild blueberry production fields.

Expression of N-cycling genes of root microbiomes provides insights for sustaining oilseed crop production.

Agricultural production is dependent on inputs of nitrogen (N) whose cycle relies on soil and crop microbiomes. Crop diversification has increased productivity; however, its impact on the expression of microbial genes involved in N-cycling pathways remains unknown. Here, we assessed N-cycling gene expression patterns in the root and rhizosphere microbiomes of five oilseed crops as influenced by three 2-year crop rotations. The first phase consisted of fallow, lentil or wheat, and the second phase consisted of one of five oilseed crops. Expression of bacterial amoA, nirK and nirS genes showed that the microbiome of Ethiopian mustard had the lowest and that of camelina the highest potential for N loss. A preceding rotation phase of lentil significantly increased the expression of nifH gene by 23% compared with wheat and improved nxrA gene expression by 51% with chemical fallow in the following oilseed crops respectively. Lentil substantially increased biological N2 fixation and reduced denitrification in the following oilseed crops. Our results also revealed that most N-cycling gene transcripts are more abundant in the microbiomes associated with roots than with the rhizosphere. The outcome of our investigation brings a new level of understanding on how crop diversification and rotation sequences are related to N-cycling in annual cropping systems.

Holobiont chronobiology: mycorrhiza may be a key to linking aboveground and underground rhythms.

Circadian clocks are nearly ubiquitous timing mechanisms that can orchestrate rhythmic behavior and gene expression in a wide range of organisms. Clock mechanisms are becoming well understood in fungal, animal, and plant model systems, yet many of these organisms are surrounded by a complex and diverse microbiota which should be taken into account when examining their biology. Of particular interest are the symbiotic relationships between organisms that have coevolved over time, forming a unit called a holobiont. Several studies have now shown linkages between the circadian rhythms of symbiotic partners. Interrelated regulation of holobiont circadian rhythms seems thus important to coordinate shifts in activity over the day for all the partners. Therefore, we suggest that the classical view of "chronobiological individuals" should include "a holobiont" rather than an organism. Unfortunately, mechanisms that may regulate interspecies temporal acclimation and the evolution of the circadian clock in holobionts are far from being understood. For the plant holobiont, our understanding is particularly limited. In this case, the holobiont encompasses two different ecosystems, one above and the other below the ground, with the two potentially receiving timing information from different synchronizing signals (Zeitgebers). The arbuscular mycorrhizal (AM) symbiosis, formed by plant roots and fungi, is one of the oldest and most widespread associations between organisms. By mediating the nutritional flux between the plant and the many microbes in the soil, AM symbiosis constitutes the backbone of the plant holobiont. Even though the importance of the AM symbiosis has been well recognized in agricultural and environmental sciences, its circadian chronobiology remains almost completely unknown. We have begun to study the circadian clock of arbuscular mycorrhizal fungi, and we compile and here discuss the available information on the subject. We propose that analyzing the interrelated temporal organization of the AM symbiosis and determining its underlying mechanisms will advance our understanding of the role and coordination of circadian clocks in holobionts in general.

Expression of putative circadian clock components in the arbuscular mycorrhizal fungus Rhizoglomus irregulare.

Arbuscular mycorrhizal fungi (AMF) are obligatory plant symbionts that live underground, so few studies have examined their response to light. Responses to blue light by other fungi can be mediated by White Collar-1 (WC-1) and WC-2 proteins. These wc genes, together with the frequency gene (frq), also form part of the endogenous circadian clock. The clock mechanism has never been studied in AMF, although circadian growth of their hyphae in the field has been reported. Using both genomic and transcriptomic data, we have found homologs of wc-1, wc-2, and frq and related circadian clock genes in the arbuscular mycorrhizal fungus Rhizoglomus irregulare (synonym Rhizophagus irregularis). Gene expression of wc-1, wc-2, and frq was analyzed using RT-qPCR on RNA extracted from germinating spores and from fungal material cultivated in vitro with transformed carrot roots. We found that all three core clock genes were expressed in both pre- and post-mycorrhizal stages of R. irregulare growth. Similar to the model fungus Neurospora crassa, the core circadian oscillator gene frq was induced by brief light stimulation. The presence of circadian clock and output genes in R. irregulare opens the door to the study of circadian clocks in the fungal partner of plant-AMF symbiosis. Our finding also provides new insight into the evolution of the circadian frq gene in fungi.

Petroleum hydrocarbon contamination, plant identity and arbuscular mycorrhizal fungal (AMF) community determine assemblages of the AMF spore-associated microbes.

The root-associated microbiome is a key determinant of pollutant degradation, soil nutrient availability and plant biomass productivity, but could not be examined in depth prior to recent advances in high-throughput sequencing. Arbuscular mycorrhizal fungi (AMF) form symbioses with the majority of vascular plants. They are known to enhance mineral uptake and promote plant growth and are postulated to influence the processes involved in phytoremediation. Amplicon sequencing approaches have previously shown that petroleum hydrocarbon pollutant (PHP) concentration strongly influences AMF community structure in in situ phytoremediation experiments. We examined how AMF communities and their spore-associated microbiomes were structured within the rhizosphere of three plant species growing spontaneously in three distinct waste decantation basins of a former petrochemical plant. Our results show that the AMF community was only affected by PHP concentrations, while the AMF-associated fungal and bacterial communities were significantly affected by both PHP concentrations and plant species identity. We also found that some AMF taxa were either positively or negatively correlated with some fungal and bacterial groups. Our results suggest that in addition to PHP concentrations and plant species identity, AMF community composition may also shape the community structure of bacteria and fungi associated with AMF spores.

A Diverse Soil Microbiome Degrades More Crude Oil than Specialized Bacterial Assemblages Obtained in Culture.

UNLABELLED: Soil microbiome modification may alter system function, which may enhance processes like bioremediation. In this study, we filled microcosms with gamma-irradiated soil that was reinoculated with the initial soil or cultivated bacterial subsets obtained on regular media (REG-M) or media containing crude oil (CO-M). We allowed 8 weeks for microbiome stabilization, added crude oil and monoammonium phosphate, incubated the microcosms for another 6 weeks, and then measured the biodegradation of crude oil components, bacterial taxonomy, and functional gene composition. We hypothesized that the biodegradation of targeted crude oil components would be enhanced by limiting the microbial taxa competing for resources and by specifically selecting bacteria involved in crude oil biodegradation (i.e., CO-M). Postincubation, large differences in taxonomy and functional gene composition between the three microbiome types remained, indicating that purposeful soil microbiome structuring is feasible. Although phylum-level bacterial taxonomy was constrained, operational taxonomic unit composition varied between microbiome types. Contrary to our hypothesis, the biodegradation of C10 to C50 hydrocarbons was highest when the original microbiome was reinoculated, despite a higher relative abundance of alkane hydroxylase genes in the CO-M microbiomes and of carbon-processing genes in the REG-M microbiomes. Despite increases in the relative abundances of genes potentially linked to hydrocarbon processing in cultivated subsets of the microbiome, reinoculation of the initial microbiome led to maximum biodegradation. IMPORTANCE: In this study, we show that it is possible to sustainably modify microbial assemblages in soil. This has implications for biotechnology, as modification of gut microbial assemblages has led to improved treatments for diseases like Clostridium difficile infection. Although the soil environment determined which major phylogenetic groups of bacteria would dominate the assemblage, we saw differences at lower levels of taxonomy and in functional gene composition (e.g., genes related to hydrocarbon degradation). Further studies are needed to determine the success of such an approach in nonsterile environments. Although the biodegradation of certain crude oil fractions was still the highest when we inoculated with the diverse initial microbiome, the possibility of discovering and establishing microbiomes that are more efficient in crude oil degradation is not precluded.

Independent mitochondrial and nuclear exchanges arising in Rhizophagus irregularis crossed-isolates support the presence of a mitochondrial segregation mechanism.

BACKGROUND: Arbuscular mycorrhizal fungi (AMF) are members of the phylum Glomeromycota, an early divergent fungal lineage that forms symbiotic associations with the large majority of land plants. These organisms are asexual obligate biotrophs, meaning that they cannot complete their life cycle in the absence of a suitable host. These fungi can exchange genetic information through hyphal fusions (i.e. anastomosis) with genetically compatible isolates belonging to the same species. The occurrence of transient mitochondrial length-heteroplasmy through anastomosis between geographically distant Rhizophagus irregularis isolates was previously demonstrated in single spores resulting from crossing experiments. However, (1) the persistence of this phenomenon in monosporal culture lines from crossed parental isolates, (2) its correlation with nuclear exchanges and (3) the potential mechanisms responsible for mitochondrial inheritance are still unknown. Using the AMF model organism R. irregularis, we tested whether the presence of a heteroplasmic state in progeny spores was linked to the occurrence of nuclear exchanges and whether the previously observed heteroplasmic state persisted in monosporal in vitro crossed-culture lines. We also investigated the presence of a putative mitochondrial segregation apparatus in Glomeromycota by identifying proteins similar to those found in other fungal groups. RESULTS: We observed the occurrence of biparental inheritance both for mitochondrial and nuclear markers tested in single spores obtained from crossed-isolates. However, only one parental mitochondrial DNA and nuclear genotype were recovered in each monosporal crossed-cultures, with an overrepresentation of certain mitochondrial haplotypes. These results strongly support the presence of a nuclear-independent mitochondrial segregation mechanism in R. irregularis. Furthermore, a nearly complete set of genes was identified with putative orthology to those found in other fungi and known to be associated with the mitochondrial segregation in Saccharomyces cerevisiae and filamentous fungi. CONCLUSIONS: Our findings suggest that mitochondrial segregation might take place either during spore formation or colony development and that it might be independent of the nuclear segregation machinery. We present the basic building blocks for a better understanding of the mitochondrial inheritance process and segregation in these important symbiotic fungi. The comprehension of these processes is of great importance since it has been shown that different segregated lines of the same isolate can have variable effects on the host plant.

Mitochondrial comparative genomics and phylogenetic signal assessment of mtDNA among arbuscular mycorrhizal fungi.

Mitochondrial (mt) genes, such as cytochrome C oxidase genes (cox), have been widely used for barcoding in many groups of organisms, although this approach has been less powerful in the fungal kingdom due to the rapid evolution of their mt genomes. The use of mt genes in phylogenetic studies of Dikarya has been met with success, while early diverging fungal lineages remain less studied, particularly the arbuscular mycorrhizal fungi (AMF). Advances in next-generation sequencing have substantially increased the number of publically available mtDNA sequences for the Glomeromycota. As a result, comparison of mtDNA across key AMF taxa can now be applied to assess the phylogenetic signal of individual mt coding genes, as well as concatenated subsets of coding genes. Here we show comparative analyses of publically available mt genomes of Glomeromycota, augmented with two mtDNA genomes that were newly sequenced for this study (Rhizophagus irregularis DAOM240159 and Glomus aggregatum DAOM240163), resulting in 16 complete mtDNA datasets. R. irregularis isolate DAOM240159 and G. aggregatum isolate DAOM240163 showed mt genomes measuring 72,293bp and 69,505bp with G+C contents of 37.1% and 37.3%, respectively. We assessed the phylogenies inferred from single mt genes and complete sets of coding genes, which are referred to as "supergenes" (16 concatenated coding genes), using Shimodaira-Hasegawa tests, in order to identify genes that best described AMF phylogeny. We found that rnl, nad5, cox1, and nad2 genes, as well as concatenated subset of these genes, provided phylogenies that were similar to the supergene set. This mitochondrial genomic analysis was also combined with principal coordinate and partitioning analyses, which helped to unravel certain evolutionary relationships in the Rhizophagus genus and for G. aggregatum within the Glomeromycota. We showed evidence to support the position of G. aggregatum within the R. irregularis 'species complex'.

Genome of an arbuscular mycorrhizal fungus provides insight into the oldest plant symbiosis.

The mutualistic symbiosis involving Glomeromycota, a distinctive phylum of early diverging Fungi, is widely hypothesized to have promoted the evolution of land plants during the middle Paleozoic. These arbuscular mycorrhizal fungi (AMF) perform vital functions in the phosphorus cycle that are fundamental to sustainable crop plant productivity. The unusual biological features of AMF have long fascinated evolutionary biologists. The coenocytic hyphae host a community of hundreds of nuclei and reproduce clonally through large multinucleated spores. It has been suggested that the AMF maintain a stable assemblage of several different genomes during the life cycle, but this genomic organization has been questioned. Here we introduce the 153-Mb haploid genome of Rhizophagus irregularis and its repertoire of 28,232 genes. The observed low level of genome polymorphism (0.43 SNP per kb) is not consistent with the occurrence of multiple, highly diverged genomes. The expansion of mating-related genes suggests the existence of cryptic sex-related processes. A comparison of gene categories confirms that R. irregularis is close to the Mucoromycotina. The AMF obligate biotrophy is not explained by genome erosion or any related loss of metabolic complexity in central metabolism, but is marked by a lack of genes encoding plant cell wall-degrading enzymes and of genes involved in toxin and thiamine synthesis. A battery of mycorrhiza-induced secreted proteins is expressed in symbiotic tissues. The present comprehensive repertoire of R. irregularis genes provides a basis for future research on symbiosis-related mechanisms in Glomeromycota.

Analysis of a large dataset of mycorrhiza inoculation field trials on potato shows highly significant increases in yield.

An increasing human population requires more food production in nutrient-efficient systems in order to simultaneously meet global food needs while reducing the environmental footprint of agriculture. Arbuscular mycorrhizal fungi (AMF) have the potential to enhance crop yield, but their efficiency has yet to be demonstrated in large-scale crop production systems. This study reports an analysis of a dataset consisting of 231 field trials in which the same AMF inoculant (Rhizophagus irregularis DAOM 197198) was applied to potato over a 4-year period in North America and Europe under authentic field conditions. The inoculation was performed using a liquid suspension of AMF spores that was sprayed onto potato seed pieces, yielding a calculated 71 spores per seed piece. Statistical analysis showed a highly significant increase in marketable potato yield (ANOVA, P < 0.0001) for inoculated fields (42.2 tons/ha) compared with non-inoculated controls (38.3 tons/ha), irrespective of trial year. The average yield increase was 3.9 tons/ha, representing 9.5 % of total crop yield. Inoculation was profitable with a 0.67-tons/ha increase in yield, a threshold reached in almost 79 % of all trials. This finding clearly demonstrates the benefits of mycorrhizal-based inoculation on crop yield, using potato as a case study. Further improvements of these beneficial inoculants will help compensate for crop production deficits, both now and in the future.

The large (134.9 kb) mitochondrial genome of the glomeromycete Funneliformis mosseae.

Funneliformis mosseae is among the most ecologically and economically important glomeromycete species and occurs both in natural and disturbed areas in a wide range of habitats and climates. In this study, we report the sequencing of the complete mitochondrial (mt) genome of F. mosseae isolate FL299 using 454 pyrosequencing and Illumina HiSeq technologies. This mt genome is a full-length circular chromosome of 134,925 bp, placing it among the largest mitochondrial DNAs (mtDNAs) in the fungal kingdom. A comparative analysis with publically available arbuscular mycorrhizal fungal mtDNAs revealed that the mtDNA of F. mosseae FL299 contained a very large number of insertions contributing to its expansion. The gene synteny was completely reshuffled compared to previously published glomeromycotan mtDNAs and several genes were oriented in an anti-sense direction. Furthermore, the presence of different types of introns and insertions in rnl (14 introns) made this gene very distinctive in Glomeromycota. The presence of alternative genetic codes in both initiation (GUG) and termination (UGA) codons was another new feature in this mtDNA compared to previously published glomeromycotan mt genomes. The phylogenetic analysis inferred from the analysis of 14 protein mt genes confirmed the position of the Glomeromycota clade as a sister group of Mortierellomycotina. This mt genome is the largest observed so far in Glomeromycota and the first mt genome within the Funneliformis clade, providing new opportunities to better understand their evolution and to develop molecular markers.

Molecular diagnostic toolkit for Rhizophagus irregularis isolate DAOM-197198 using quantitative PCR assay targeting the mitochondrial genome.

Rhizophagus irregularis (previously named Glomus irregulare) is one of the most widespread and common arbuscular mycorrhizal fungal (AMF) species. It has been recovered worldwide in agricultural and natural soils, and the isolate DAOM-197198 has been utilized as a commercial inoculant for two decades. Despite the ecological and economical importance of this taxon, specific markers for quantification of propagules by quantitative real-time PCR (qPCR) are extremely limited and none have been rigorously validated for quality control of manufactured products such as biofertilizers. From the sequencing of 14 complete AMF mitochondrial (mt) genomes, a qPCR assay using a hydrolysis probe designed in the single copy cox3-rnl intergenic region was tested and validated to specifically and accurately quantify the spores of R. irregularis isolate DAOM-197198. Specificity tests were performed using standard PCR and qPCR, and results clearly showed that the primers specifically amplified the isolate DAOM-197198, yielding a PCR product of 106 bp. According to the qPCR analyses on spores produced in vitro, the average copy number of mt genomes per spore was 3172 ± 304 SE (n = 6). Quantification assays were successfully undertaken on known and unknown samples in liquid suspensions and commercial dry formulations to show the accuracy, precision, robustness, and reproducibility of the qPCR assay. This study provides a powerful molecular toolkit specifically designed to quantify spores of the model AMF isolate DAOM-197198. The approach of molecular toolkit used in our study could be applied to other AMF taxa and will be useful to research institutions and governmental and industrial laboratories running routine quality control of AMF-based products.

Effect of Medicago sativa L. and compost on organic and inorganic pollutant removal from a mixed contaminated soil and risk assessment using ecotoxicological tests.

Several Gentle Remediation Options (GRO), e.g., plant-based options (phytoremediation), singly and combined with soil amendments, can be simultaneously efficient for degrading organic pollutants and either stabilizing or extracting trace elements (TEs). Here, a 5-month greenhouse trial was performed to test the efficiency of Medicago sativa L., singly and combined with a compost addition (30% w/w), to treat soils contaminated by petroleum hydrocarbons (PHC), Co and Pb collected at an auto scrap yard. After 5 months, total soil Pb significantly decreased in the compost-amended soil planted with M. sativa, but not total soil Co. Compost incorporation into the soil promoted PHC degradation, M. sativa growth and survival, and shoot Pb concentrations [3.8 mg kg(-1) dry weight (DW)]. Residual risk assessment after the phytoremediation trial showed a positive effect of compost amendment on plant growth and earthworm development. The O2 uptake by soil microorganisms was lower in the compost-amended soil, suggesting a decrease in microbial activity. This study underlined the benefits of the phytoremediation option based on M. sativa cultivation and compost amendment for remediating PHC- and Pb-contaminated soils.

Does soil history decline in influencing the structure of bacterial communities of Brassica napus host plants across different growth stages?

Soil history has been shown to condition future rhizosphere microbial communities. However, previous experiments have also illustrated that mature, adult plants can "re-write," or mask, different soil histories through host plant-soil community feedbacks. This leaves a knowledge gap concerning how soil history influences bacterial community structure across different growth stages. Thus, here we tested the hypothesis that previously established soil histories will decrease in influencing the structure of Brassica napus bacterial communities over the growing season. We used an on-going agricultural field experiment to establish three different soil histories, plots of monocrop canola (B. napus), or rotations of wheat-canola, or pea-barley-canola. During the following season, we repeatedly sampled the surrounding bulk soil, rhizosphere, and roots of the B. napus hosts at different growth stages-the initial seeding conditions, seedling, rosette, bolting, and flower-from all three soil history plots. We compared composition and diversity of the B. napus soil bacterial communities, as estimated using 16S rRNA gene metabarcoding, to identify any changes associated with soil history and growth stages. We found that soil history remained significant across each growth stage in structuring the bacterial bulk soil and rhizosphere communities, but not the bacterial root communities. This suggests that the host plant's capacity to "re-write" different soil histories may be quite limited as key components that constitute the soil history's identity remain present, such that the previously established soil history continues to impact the bacterial rhizosphere communities, but not the root communities. For agriculture, this highlights how previously established soil histories persist and may have important long-term consequences on future plant-microbe communities, including bacteria.

The rhizosphere of a drought-tolerant plant species in Morocco: A refuge of high microbial diversity with no taxon preference.

Arid and semi-arid areas are facing increasingly severe water deficits that are being intensified by global climate changes. Microbes associated with plants native to arid regions provide valuable benefits to plants, especially in water-stressed environments. In this study, we used 16S rDNA metabarcoding analysis to examine the bacterial communities in the bulk soil, rhizosphere and root endosphere of the plant Malva sylvestris L. in Morocco, along a gradient of precipitation. We found that the rhizosphere of M. sylvestris did not show significant differences in beta-diversity compared to bulk soil, although, it did display an increased degree of alpha-diversity. The endosphere was largely dominated by the genus Rhizobium and displayed remarkable variation between plants, which could not be attributed to any of the variables observed in this study. Overall, the effects of precipitation level were relatively weak, which may be related to the intense drought in Morocco at the time of sampling. The dominance of Rhizobium in a non-leguminous plant is particularly noteworthy and may permit the utilization of this bacterial taxon to augment drought tolerance; additionally, the absence of any notable selection of the rhizosphere of M. sylvestris suggests that it is not significatively affecting its soil environment.