Assessment of Passive Traps Combined with High-Throughput Sequencing To Study Airborne Fungal Communities J Aguayo, C Fourrier-Jeandel, C Husson, R Ioos. Applied and Environmental Microbiology 84 (11), e02637-17

ABSTRACT

Techniques based on high-throughput sequencing (HTS) of environmental DNA have provided a new way of studying fungal diversity. However, these techniques suffer from a number of methodological biases which may appear at any of the steps involved in a metabarcoding study. Air is one of the most important environments where fungi can be found, because it is the primary medium of dispersal for many species. Looking ahead to future developments, it was decided to test 20 protocols, including different passive spore traps, spore recovery procedures, DNA extraction kits, and barcode loci. HTS was performed with the Illumina MiSeq platform targeting two subloci of the fungal internal transcribed spacer. Multivariate analysis and generalized linear models showed that the type of passive spore trap, the spore recovery procedure, and the barcode all impact the description of fungal communities in terms of richness and diversity when assessed by HTS metabarcoding. In contrast, DNA extraction kits did not significantly impact these results. Although passive traps may be used to describe airborne fungal communities, a study using specific real-time PCR and a mock community showed that these kinds of traps are affected by environmental conditions that may induce losses of biological material, impacting diversity and community composition results.

IMPORTANCE The advent of high-throughput sequencing (HTS) methods, such as those offered by next-generation sequencing (NGS) techniques, has opened a new era in the study of fungal diversity in different environmental substrates. In this study, we show that an assessment of the diversity of airborne fungal communities can reliably be achieved by the use of simple and robust passive spore traps. However, a comparison of sample processing protocols showed that several methodological biases may impact the results of fungal diversity when assessed by metabarcoding. Our data suggest that identifying these biases is of paramount importance to enable a correct identification and relative quantification of community members.

The Hydrophobin-Like OmSSP1 May Be an Effector in the Ericoid Mycorrhizal Symbiosis. S Casarrubia, S Daghino, A Kohler, E Morin, HR Khouja, C Venault-Fourrey,… Front. Plant Sci., 01 May 2018 |

Abstract

Mutualistic and pathogenic plant-colonizing fungi use effector molecules to manipulate the host cell metabolism to allow plant tissue invasion. Some small secreted proteins (SSPs) have been identified as fungal effectors in both ectomycorrhizal and arbuscular mycorrhizal fungi, but it is currently unknown whether SSPs also play a role as effectors in other mycorrhizal associations. Ericoid mycorrhiza is a specific endomycorrhizal type that involves symbiotic fungi mostly belonging to the Leotiomycetes (Ascomycetes) and plants in the family Ericaceae. Genomic and RNASeq data from the ericoid mycorrhizal fungus Oidiodendron maius led to the identification of several symbiosis-upregulated genes encoding putative SSPs. OmSSP1, the most highly symbiosis up-regulated SSP, was found to share some features with fungal hydrophobins, even though it lacks the Pfam hydrophobin domain. Sequence alignment with other hydrophobins and hydrophobin-like fungal proteins placed OmSSP1 within Class I hydrophobins. However, the predicted features of OmSSP1 may suggest a distinct type of hydrophobin-like proteins. The presence of a predicted signal peptide and a yeast-based signal sequence trap assay demonstrate that OmSSP1 is secreted. OmSSP1 null-mutants showed a reduced capacity to form ericoid mycorrhiza with Vaccinium myrtillus roots, suggesting a role as effectors in the ericoid mycorrhizal interaction.

First Report of Phytophthora ramorum causing Japanese Larch dieback in France N Schenck, C Saurat, C Guinet, C Fourrier-Jeandel, L Roche, A Bouvet, … Plant Disease

Abstract

Phytophthora ramorum Werres, De Cock & Man in’t Veld, an oomycete known in the USA as the causal agent of Sudden Oak Death, has spread across Europe since the early 2000s. It is responsible for damage and death to a wide range of plant species, including mature trees. In 2009 it was identified on Japanese larch (Larix kaempferi) in South-West England (Webber et al., 2010) and since, it has caused severe damages and losses to Larix spp. in the United Kingdom and the Republic of Ireland. There are two lineages of the oomycete EU1 and EU2 found in Europe (King et al., 2015), EU2 being the more aggressive. The symptoms on larch include necrosis and loss of needles, wilting of shoots, dieback of branches and death, often with abundant resin bleeding on trunks and branches. As sporulating hosts, Larix spp. may disperse P. ramorum over long distances. In May 2017, wilting, yellowing/reddening needles and branch mortality was observed on mature Larix kaempferi (about fifty years old) in the forest of Saint-Cadou, Finistère, in the far North western part of France (3° 59’ 49.2’’ W ; 48° 22’ 22.4’’ N). Approximately, 4027% of the trees were affected in May, and 42% later in September 2017. The presence of P. ramorum was suspected, and was first confirmed by testing samples collected from trunks and branches with necrosis and resin bleeds, using the specific conventional PCR method developed by Ioos et al. (2006). The oomycete was also isolated in pure culture, using a Phytophthora selective medium (PARB[H]). The features observed, such as a coralloid mycelium, the presence of numerous, thin-walled chlamydospores (up to 75 µm large) and deciduous, semi-papillate sporangia arranged in clusters, matched those reported for P. ramorum . In June 2017, the presence of P. ramorum was confirmed in another larch stand in Hanvec, Finistère (4° 12’ 45.0” W ; 48° 20’ 10.8” N), using the same identification techniques. In this stand, the prevalence was not precisely estimated, but was deemed much lower than in Saint-Cadou. Based on the analysis of Cox1 partial sequence and the PCR-RFLP pattern described by Van Poucke et al. (2012) on Cox1, the P. ramorum isolates collected in these two forests could be assigned to the EU1 lineage. This is the first report of P. ramorum affecting Japanese larch in France and in mainland Europe. Until now it had only been detected on shrubs in nurseries, green spaces, and in rare circumstances in the natural environment on understory vegetation (rhododendron) in Normandy and Brittany, but not in the vicinity of the infected larch stands. The presence of this pathogen in the natural environment represents a major threat for larch trees, but also for the other potential forest host trees in this region, such as sweet chestnut and might have a severe impact on both forest and ornamental tree species. Research is in progress to learn more about this outbreak, the possible origin of the inoculum, the extension of the disease and its progression.

1. The ash dieback invasion of Europe was founded by two genetically divergent individuals M McMullan, M Rafiqi, G Kaithakottil, BJ Clavijo, L Bilham, E Orton, …Nature Ecology & Evolution, 1

   Abstract

   Accelerating international trade and climate change make pathogen spread an increasing concern. Hymenoscyphus fraxineus, the causal agent of ash dieback, is a fungal pathogen that has been moving across continents and hosts from Asian to European ash. Most European common ash trees (Fraxinus excelsior) are highly susceptible to H. fraxineus, although a minority (~5%) have partial resistance to dieback. Here, we assemble and annotate a H. fraxineus draft genome, which approaches chromosome scale. Pathogen genetic diversity across Europe and in Japan, reveals a strong bottleneck in Europe, though a signal of adaptive diversity remains in key host interaction genes. We find that the European population was founded by two divergent haploid individuals. Divergence between these haplotypes represents the ancestral polymorphism within a large source population. Subsequent introduction from this source would greatly increase adaptive potential of the pathogen. Thus, further introgression of H. fraxineus into Europe represents a potential threat and Europe-wide biological security measures are needed to manage this disease.

N-Acetylglucosaminidase activity, a functional trait of chitin degradation, is regulated differentially within two orders of ectomycorrhizal fungi: Boletales and Agaricales F Maillard, M Didion, L Fauchery, C Bach, M Buée. Mycorrhiza, 1-7

Abstract

Chitin is one of the most abundant nitrogen-containing polymers in forest soil. Ability of ectomycorrhizal (EM) fungi to utilize chitin may play a key role in the EM symbiosis nutrition and soil carbon cycle. In forest, EM fungi exhibit high diversity, which could be based on function partitioning and trait complementarity. Although it has long been recognized that closely related species share functional characteristics, the phylogenetic conservatism of functional traits within microorganisms remains unclear. Because extracellular N-acetylglucosaminidase activity has been proposed as functional trait of chitin degradation, we screened this activity on 35 EM fungi species with or without chitin in the growth medium to (i) describe the functional diversity of EM fungi and (ii) identify potential links between this functional trait and EM fungal phylogeny. We observed large variations of the extracellular N-acetylglucosaminidase activities among the fungal strains. Furthermore, our results revealed two regulation patterns of extracellular N-acetylglucosaminidase activities. Indeed, these chitinolytic activities were stimulated or repressed in the presence of chitin, in comparison to the control treatment. These profiles of extracellular N-acetylglucosaminidase stimulation/repression might be conserved at a high phylogenetic level in the Basidiomycota phylum, as illustrated by the opposite patterns of regulation between Boletales and Agaricales. Finally, the downregulation of this activity by chitin, for some EM fungal groups, might suggest another chitin degradation pathway.

The ectomycorrhizal basidiomycete Laccaria bicolor releases a secreted β-1, 4 endoglucanase that plays a key role in symbiosis development. F Zhang, GE Anasontzis, A Labourel, C Champion, M Haon, … The New phytologist

Abstract

In ectomycorrhiza, root ingress and colonization of the apoplast by colonizing hyphae is thought to rely mainly on the mechanical force that results from hyphal tip growth, but this could be enhanced by secretion of cell-wall-degrading enzymes, which have not yet been identified. The sole cellulose-binding module (CBM1) encoded in the genome of the ectomycorrhizal Laccaria bicolor is linked to a glycoside hydrolase family 5 (GH5) endoglucanase, LbGH5-CBM1. Here, we characterize LbGH5-CBM1 gene expression and the biochemical properties of its protein product. We also immunolocalized LbGH5-CBM1 by immunofluorescence confocal microscopy in poplar ectomycorrhiza. We show that LbGH5-CBM1 expression is substantially induced in ectomycorrhiza, and RNAi mutants with a decreased LbGH5-CBM1 expression have a lower ability to form ectomycorrhiza, suggesting a key role in symbiosis. Recombinant LbGH5-CBM1 displays its highest activity towards cellulose and galactomannans, but no activity toward L. bicolor cell walls. In situ localization of LbGH5-CBM1 in ectomycorrhiza reveals that the endoglucanase accumulates at the periphery of hyphae forming the Hartig net and the mantle. Our data suggest that the symbiosis-induced endoglucanase LbGH5-CBM1 is an enzymatic effector involved in cell wall remodeling during formation of the Hartig net and is an important determinant for successful symbiotic colonization.

Intraspecific comparative genomics of isolates of the Norway spruce pathogen (Heterobasidion parviporum) and identification of its potential virulence factors Z Zeng, H Sun, EJ Vainio, T Raffaello, A Kovalchuk, E Morin, S Duplessis, … BMC genomics 19 (1), 220

Abstract

Impact of soil pedogenesis on the diversity and composition of fungal communities across the California soil chronosequence of Mendocino PE Courty, M Buée, JJT Tech, D Brulé, Y Colin, JHJ Leveau, S Uroz Mycorrhiza, 1-1

Abstract

Understanding how soil pedogenesis affects microbial communities and their in situ activities according to ecosystem functioning is a central issue in soil microbial ecology, as soils represent essential nutrient reservoirs and habitats for the biosphere. To address this question, soil chronosequences developed from a single, shared mineralogical parent material and having the same climate conditions are particularly useful, as they isolate the factor of time from other factors controlling the character of soils. In our study, we considered a natural succession of uplifted marine terraces in Mendocino, CA, ranging from highly fertile in the younger terrace (about 100,000 years old) to infertile in the older terraces (about 300,000 years old). Using ITS amplicon pyrosequencing, we analysed and compared the diversity and composition of the soil fungal communities across the first terraces (T1 to T3), with a specific focus in the forested terraces (T2 and T3) on soil samples collected below trees of the same species (Pinus muricata) and of the same age. While diversity and richness indices were highest in the grassland (youngest) terrace (T1), they were higher in the older forested terrace (T3) compared to the younger forested terrace (T2). Interestingly, the most abundant ectomycorrhizal (ECM) taxa that we found within these fungal communities showed high homology with ITS Sanger sequences obtained previously directly from ECM root tips from trees in the same study site, revealing a relative conservation of ECM diversity over time. Altogether, our results provide new information about the diversity and composition of the fungal communities as well as on the dominant ECM species in the soil chronosequence of Mendocino in relation to soil age and ecosystem development.

The origin and evolution of mycorrhizal symbioses: from palaeomycology to phylogenomics. C Strullu-Derrien, MA Selosse, P Kenrick, FM Martin The New phytologist

Abstract