Topsoil Bacterial Community Changes and Nutrient Dynamics Under Cereal Based Climate-Smart Agri-Food Systems.

Soil microorganisms play a critical role in soil biogeochemical processes, nutrient cycling, and resilience of agri-food systems and are immensely influenced by agronomic management practices. Understanding soil bacterial community and nutrient dynamics under contrasting management practices is of utmost importance for building climate-smart agri-food systems. Soil samples were collected at 0-15 cm soil depth from six management scenarios in long-term conservation agriculture (CA) and climate-smart agriculture (CSA) practices. These scenarios (Sc) involved; ScI-conventional tillage based rice-wheat rotation, ScII- partial CA based rice-wheat-mungbean, ScIII- partial CSA based rice-wheat-mungbean, ScIV is partial CSA based maize-wheat-mungbean, ScV and ScVI are CSA based scenarios, were similar to ScIII and ScIV respectively, layered with precision water & nutrient management. The sequencing of soil DNA results revealed that across the six scenarios, a total of forty bacterial phyla were observed, with Proteobacteria as dominant in all scenarios, followed by Acidobacteria and Actinobacteria. The relative abundance of Proteobacteria was 29% higher in rice-based CSA scenarios (ScIII and ScV) and 16% higher in maize-based CSA scenarios (ScIV and ScVI) compared to conventional-till practice (ScI). The relative abundance of Acidobacteria and Actinobacteria was respectively 29% and 91% higher in CT than CSA based rice and 27% and 110% higher than maize-based scenarios. Some taxa are present relatively in very low abundance or exclusively in some scenarios, but these might play important roles there. Three phyla are exclusively present in ScI and ScII i.e., Spirochaetes, Thermi, and Euryarchaeota. Shannon diversity index was 11% higher in CT compared to CSA scenarios. Maize based CSA scenarios recorded higher diversity indices than rice-based CSA scenarios. Similar to changes in soil bacterial community, the nutrient dynamics among the different scenarios also varied significantly. After nine years of continuous cropping, the soil organic carbon was improved by 111% and 31% in CSA and CA scenarios over the CT scenario. Similarly, the available nitrogen, phosphorus, and potassium were improved by, respectively, 38, 70, and 59% in CSA scenarios compared to the CT scenario. These results indicate that CSA based management has a positive influence on soil resilience in terms of relative abundances of bacterial groups, soil organic carbon & available plant nutrients and hence may play a critical role in the sustainability of the intensive cereal based agri-food systems.

Transcriptome Profiling of Haloxylon persicum (Bunge ex Boiss and Buhse) an Endangered Plant Species under PEG-Induced Drought Stress.

Haloxylon persicum is an endangered western Asiatic desert plant species, which survives under extreme environmental conditions. In this study, we focused on transcriptome analysis of H. persicum to understand the molecular mechanisms associated with drought tolerance. Two different periods of polyethylene glycol (PEG)-induced drought stress (48 h and 72 h) were imposed on H. persicum under in vitro conditions, which resulted in 18 million reads, subsequently assembled by de novo method with more than 8000 transcripts in each treatment. The N50 values were 1437, 1467, and 1524 for the control sample, 48 h samples, and 72 h samples, respectively. The gene ontology (GO) and Kyoto encyclopedia of genes and genomes (KEGG) pathway analysis resulted in enrichment of mitogen-activated protein kinase (MAPK) and plant hormone signal transduction pathways under PEG-induced drought conditions. The differential gene expression analysis (DGEs) revealed significant changes in the expression pattern between the control and the treated samples. The KEGG analysis resulted in mapping transcripts with 138 different pathways reported in plants. The differential expression of drought-responsive transcription factors depicts the possible signaling cascades involved in drought tolerance. The present study provides greater insight into the fundamental transcriptome reprogramming of desert plants under drought.

Comprehensive History of CSP Genes: Evolution, Phylogenetic Distribution and Functions.

In this review we present the developmental, histological, evolutionary and functional properties of insect chemosensory proteins (CSPs) in insect species. CSPs are small globular proteins folded like a prism and notoriously known for their complex and arguably obscure function(s), particularly in pheromone olfaction. Here, we focus on direct functional consequences on protein function depending on duplication, expression and RNA editing. The result of our analysis is important for understanding the significance of RNA-editing on functionality of CSP genes, particularly in the brain tissue.

Soil bacterial diversity under conservation agriculture-based cereal systems in Indo-Gangetic Plains.

In Indo-Gangetic plains (IGP) of India, natural resources (soil, water, and environment) are degrading under the conventional-till (CT)-based management practices in rice-wheat cropping system. A long-term field experiment was conducted to understand the soil bacterial diversity and abundance under different sets of management scenarios (Sc). The study comprised of four scenarios, namely, -Sc.I CT-based rice-wheat system (farmers' practice); Sc.II, partial conservation agriculture (CA) based in which rice is under CT-wheat and mungbean under zero-tillage (ZT); Sc.III, full CA-based in which rice-wheat-mungbean are under ZT and Sc.IV, where maize-wheat-mungbean are under ZT. These scenarios varied in cropping system, tillage, and crop residue management practices. Using Illumina MiSeq sequencing technology, the variable regions V3-V4 of 16S rRNA were sequenced and the obtained reads were analyzed to study the diversity patterns in the scenarios. Results showed the presence of 53 bacterial phyla across scenarios. The predominant phyla in all scenarios were Proteobacteria, Acidobacteria, Actinobacteria, and Bacteroidetes which accounted for more than 70% of the identified phyla. However, the rice-based systems (Sc.I, Sc.II, and Sc.III) were dominated by phylum Proteobacteria; however, maize-based system (Sc.IV) was dominated by Acidobacteria. The class DA052 and Acidobacteriia of Acidobacteria and Bacteroidetes of Bacteroidia were exceptionally higher in Sc.IV. Shannon diversity index was 8.8% higher in Sc.I, 7.5% in Sc.II, and 2.7% in Sc.III compared to Sc.IV. The findings revealed that soil bacterial diversity and abundance are influenced by agricultural management practices as bacterial diversity under full CA-based management systems (Sc.III and Sc.IV) was lower when compared to farmer's practice (Sc.I) and partial CA (Sc.II) scenarios.

Draft genome of a high value tropical timber tree, Teak (Tectona grandis L. f): insights into SSR diversity, phylogeny and conservation.

Teak (Tectona grandis L. f.) is one of the precious bench mark tropical hardwood having qualities of durability, strength and visual pleasantries. Natural teak populations harbour a variety of characteristics that determine their economic, ecological and environmental importance. Sequencing of whole nuclear genome of teak provides a platform for functional analyses and development of genomic tools in applied tree improvement. A draft genome of 317 Mb was assembled at 151× coverage and annotated 36, 172 protein-coding genes. Approximately about 11.18% of the genome was repetitive. Microsatellites or simple sequence repeats (SSRs) are undoubtedly the most informative markers in genotyping, genetics and applied breeding applications. We generated 182,712 SSRs at the whole genome level, of which, 170,574 perfect SSRs were found; 16,252 perfect SSRs showed in silico polymorphisms across six genotypes suggesting their promising use in genetic conservation and tree improvement programmes. Genomic SSR markers developed in this study have high potential in advancing conservation and management of teak genetic resources. Phylogenetic studies confirmed the taxonomic position of the genus Tectona within the family Lamiaceae. Interestingly, estimation of divergence time inferred that the Miocene origin of the Tectona genus to be around 21.4508 million years ago.

TGF-ß2-induced EMT is dampened by inhibition of autophagy and TNF-α treatment.

Hepatocellular carcinoma (HCC) typically develops in a chronic inflammatory setting causal to release of a plethora of growth factors and cytokines. However, the molecular effect of these cytokines on HCC progression is poorly understood. In this study, we exposed HCC cells to TGF-ß2 (Transforming Growth Factor-ß2), which resulted in a significant elevation of EMT (Epithelial to Mesenchymal Transition) like features. Molecular analysis of EMT markers showed an increase at both RNA and protein levels upon TGF-ß2 administration along with up-regulation of TGF-ß-induced Smad signaling. Induction of EMT was associated with a simultaneous increase in reactive oxygen species (ROS) and cytostasis of TGF-ß2-treated cells. Importantly, quenching of ROS resulted in a significant promotion of TGF-ß2-induced EMT. Furthermore, cells treated with TGF-ß2 also showed an enhanced autophagic flux. Interestingly, inhibition of autophagy by chloroquine-di-phosphate (CQDP) or siRNA-mediated ablation of ATG5 drastically inhibited TGF-ß2-induced EMT. Autophagy inhibition significantly increased ROS levels promoting apoptosis. It was further observed that pro-inflammatory cytokine like, TNF-α (Tumor Necrosis Factor-α) can antagonize TGF-ß2-induced response by down-regulating autophagy, increasing ROS levels and thus inhibiting EMT in HCC cells. This inhibitory effect of TNF-α is serum-independent. Transcriptomic analysis through RNA sequencing was further performed which validated that TGF-ß2-induced autophagic genes are inhibited by TNF-α treatment suppressing EMT. Our study suggests that autophagy plays a pro-metastatic role facilitating EMT by regulating ROS levels in HCC cells and TNF-α can suppress EMT by inhibiting autophagy. We provide unique mechanistic insights into the role of TGF-ß2 in HCC cells, along with appropriate cues to effectively control the disease.

MiRNomics Reveals Breast Cancer Cells Cultured on 3D Scaffolds Better Mimic Tumors in Vivo than Conventional 2D Culture.

Tissue-engineering-based three-dimensional (3D) models offer several advantages over conventional two-dimensional (2D) cultures and can mimic tissues in vivo. Although studies have analyzed the changes in the expression of genes and proteins that might mediate in vivo-like signaling, the changes in the post-transcriptional control of gene expression that are critical in fine-tuning of signaling events has never been studied. In this study, we used next-generation sequencing (NGS) to analyze the changes in the post-transcriptional regulation in MDA-MB-231 breast cancer cells cultured on 3D scaffolds. The changes in the expression of several known microRNAs were similar to the changes reported in highly invasive cancers and their profiles highly correlated with xenotumors and human breast tumors. To elucidate the role of miRNAs in modulating metastatic potential, we integrated the miRNA and the mRNA microarray data and developed networks for major pathways implicated in metastasis. From these networks, we identified several key miRNA-mRNA interactions that might contribute to the invasive behavior and aid in developing a miRNA signature for highly invasive breast cancers. This report on the differential regulation of miRNAs in breast cancer cells cultured on scaffolds demonstrates that 3D culture better mimics the tissue in vivo with novel insights into the roles of miRNAs in modulating metastatic progression.

Mitochondrial genome of the North African Sahara Honeybee, Apis mellifera sahariensis (Hymenoptera: Apidae).

We present the complete mitochondrial genome of honey bee subspecies, Apis mellifera sahariensis (Apidae) belonging to the African lineage. The assembled circular genome has a length of 16,569 bp which comprises 13 protein coding genes, 22 transfer RNA genes, two ribosomal RNA genes, and AT rich region.

Biotype Characterization, Developmental Profiling, Insecticide Response and Binding Property of Bemisia tabaci Chemosensory Proteins: Role of CSP in Insect Defense.

Chemosensory proteins (CSPs) are believed to play a key role in the chemosensory process in insects. Sequencing genomic DNA and RNA encoding CSP1, CSP2 and CSP3 in the sweet potato whitefly Bemisia tabaci showed strong variation between B and Q biotypes. Analyzing CSP-RNA levels showed not only biotype, but also age and developmental stage-specific expression. Interestingly, applying neonicotinoid thiamethoxam insecticide using twenty-five different dose/time treatments in B and Q young adults showed that Bemisia CSP1, CSP2 and CSP3 were also differentially regulated over insecticide exposure. In our study one of the adult-specific gene (CSP1) was shown to be significantly up-regulated by the insecticide in Q, the most highly resistant form of B. tabaci. Correlatively, competitive binding assays using tryptophan fluorescence spectroscopy and molecular docking demonstrated that CSP1 protein preferentially bound to linoleic acid, while CSP2 and CSP3 proteins rather associated to another completely different type of chemical, i.e. α-pentyl-cinnamaldehyde (jasminaldehyde). This might indicate that some CSPs in whiteflies are crucial to facilitate the transport of fatty acids thus regulating some metabolic pathways of the insect immune response, while some others are tuned to much more volatile chemicals known not only for their pleasant odor scent, but also for their potent toxic insecticide activity.

The influence of menstrual cycle and endometriosis on endometrial methylome.

BACKGROUND: Alterations in endometrial DNA methylation profile have been proposed as one potential mechanism initiating the development of endometriosis. However, the normal endometrial methylome is influenced by the cyclic hormonal changes, and the menstrual cycle phase-dependent epigenetic signature should be considered when studying endometrial disorders. So far, no studies have been performed to evaluate the menstrual cycle influences and endometriosis-specific endometrial methylation pattern at the same time. RESULTS: Infinium HumanMethylation 450K BeadChip arrays were used to explore DNA methylation profiles of endometrial tissues from various menstrual cycle phases from 31 patients with endometriosis and 24 healthy women. The DNA methylation profile of patients and controls was highly similar and only 28 differentially methylated regions (DMRs) between patients and controls were found. However, the overall magnitude of the methylation differences between patients and controls was rather small (Δß ranging from -0.01 to -0.16 and from 0.01 to 0.08, respectively, for hypo- and hypermethylated CpGs). Unsupervised hierarchical clustering of the methylation data divided endometrial samples based on the menstrual cycle phase rather than diseased/non-diseased status. Further analysis revealed a number of menstrual cycle phase-specific epigenetic changes with largest changes occurring during the late-secretory and menstrual phases when substantial rearrangements of endometrial tissue take place. Comparison of cycle phase- and endometriosis-specific methylation profile changes revealed that 13 out of 28 endometriosis-specific DMRs were present in both datasets. CONCLUSIONS: The results of our study accentuate the importance of considering normal cyclic epigenetic changes in studies investigating endometrium-related disease-specific methylation patterns.

Using RNA sequencing for identifying gene imprinting and random monoallelic expression in human placenta.

Given the possible critical importance of placental gene imprinting and random monoallelic expression on fetal and infant health, most of those genes must be identified, in order to understand the risks that the baby might meet during pregnancy and after birth. Therefore, the aim of the current study was to introduce a workflow and tools for analyzing imprinted and random monoallelic gene expression in human placenta, by applying whole-transcriptome (WT) RNA sequencing of placental tissue and genotyping of coding DNA variants in family trios. Ten family trios, each with a healthy spontaneous single-term pregnancy, were recruited. Total RNA was extracted for WT analysis, providing the full sequence information for the placental transcriptome. Parental and child blood DNA genotypes were analyzed by exome SNP genotyping microarrays, mapping the inheritance and estimating the abundance of parental expressed alleles. Imprinted genes showed consistent expression from either parental allele, as demonstrated by the SNP content of sequenced transcripts, while monoallelically expressed genes had random activity of parental alleles. We revealed 4 novel possible imprinted genes (LGALS8, LGALS14, PAPPA2 and SPTLC3) and confirmed the imprinting of 4 genes (AIM1, PEG10, RHOBTB3 and ZFAT-AS1) in human placenta. The major finding was the identification of 4 genes (ABP1, BCLAF1, IFI30 and ZFAT) with random allelic bias, expressing one of the parental alleles preferentially. The main functions of the imprinted and monoallelically expressed genes included: i) mediating cellular apoptosis and tissue development; ii) regulating inflammation and immune system; iii) facilitating metabolic processes; and iv) regulating cell cycle.

DNA methylome profiling of human tissues identifies global and tissue-specific methylation patterns.

BACKGROUND: DNA epigenetic modifications, such as methylation, are important regulators of tissue differentiation, contributing to processes of both development and cancer. Profiling the tissue-specific DNA methylome patterns will provide novel insights into normal and pathogenic mechanisms, as well as help in future epigenetic therapies. In this study, 17 somatic tissues from four autopsied humans were subjected to functional genome analysis using the Illumina Infinium HumanMethylation450 BeadChip, covering 486 428 CpG sites. RESULTS: Only 2% of the CpGs analyzed are hypermethylated in all 17 tissue specimens; these permanently methylated CpG sites are located predominantly in gene-body regions. In contrast, 15% of the CpGs are hypomethylated in all specimens and are primarily located in regions proximal to transcription start sites. A vast number of tissue-specific differentially methylated regions are identified and considered likely mediators of tissue-specific gene regulatory mechanisms since the hypomethylated regions are closely related to known functions of the corresponding tissue. Finally, a clear inverse correlation is observed between promoter methylation within CpG islands and gene expression data obtained from publicly available databases. CONCLUSIONS: This genome-wide methylation profiling study identified tissue-specific differentially methylated regions in 17 human somatic tissues. Many of the genes corresponding to these differentially methylated regions contribute to tissue-specific functions. Future studies may use these data as a reference to identify markers of perturbed differentiation and disease-related pathogenic mechanisms.

Molecular evidence of RNA editing in Bombyx chemosensory protein family.

Chemosensory proteins (CSPs) are small scavenger proteins that are mainly known as transporters of pheromone/odor molecules at the periphery of sensory neurons in the insect antennae and in the producing cells from the moth female pheromone gland. Sequencing cDNAs of RNA encoding CSPs in the antennae, legs, head, pheromone gland and wings from five single individual adult females of the silkworm moth Bombyx mori showed that they differed from genomic sequences by subtle nucleotide replacement (RDD). Both intronless and intronic CSP genes expressed RDDs, although in different rates. Most interestingly, in our study the degree of RDDs in CSP genes were found to be tissue-specific. The proportion of CSP-RDDs was found to be significantly much higher in the pheromone gland. In addition, Western blot analysis of proteins in different tissues showed existence of multiple CSP protein variant chains particularly found in the pheromone gland. Peptide sequencing demonstrated the occurrence of a pleiad of protein variants for most of all BmorCSPs from the pheromone gland. Our findings show that RNA editing is an important feature in the expression of CSPs and that a high variety of RDDs is found to expand drastically thus altering the repertoire of CSP proteins in a tissue-specific manner.

Biotype expression and insecticide response of Bemisia tabaci chemosensory protein-1.

Chemosensory proteins (CSPs) are a group of small soluble proteins found so far exclusively in arthropod species. These proteins act in chemical communication and perception. In this study, a gene encoding the Type 1 CSP (BtabCSP1) from the agricultural pest Bemisia tabaci (whitefly) was analyzed to understand sequence variation and expression specificity in different biotypes. Sequence analysis of BtabCSP1 showed significant differences between the two genetically characterized biotypes, B and Q. The B-biotype had a larger number of BtabCSP1 mutations than the Q-biotype. Similar to most other CSPs, BtabCSP1 was more expressed in the head than in the rest of the body. One-step RT-PCR and qPCR analysis on total messenger RNA showed that biotype-Q had higher BtabCSP1 expression levels than biotype-B. Females from a mixed field-population had high levels of BtabCSP1 expression. The interaction of BtabCSP1 with the insecticide thiamethoxam was investigated by analyzing the BtabCSP1 expression levels following exposure to the neonicotinoid, thiamethoxam, in a time/dose-response study. Insecticide exposure increased BtabCSP1 expression (up to tenfold) at 4 and 24 h following 50 or 100 g/ml treatments.

Autoimmune regulator is acetylated by transcription coactivator CBP/p300.

The Autoimmune Regulator (AIRE) is a regulator of transcription in the thymic medulla, where it controls the expression of a large set of peripheral-tissue specific genes. AIRE interacts with the transcriptional coactivator and acetyltransferase CBP and synergistically cooperates with it in transcriptional activation. Here, we aimed to study a possible role of AIRE acetylation in the modulation of its activity. We found that AIRE is acetylated in tissue culture cells and this acetylation is enhanced by overexpression of CBP and the CBP paralog p300. The acetylated lysines were located within nuclear localization signal and SAND domain. AIRE with mutations that mimicked acetylated K243 and K253 in the SAND domain had reduced transactivation activity and accumulated into fewer and larger nuclear bodies, whereas mutations that mimicked the unacetylated lysines were functionally similar to wild-type AIRE. Analogously to CBP, p300 localized to AIRE-containing nuclear bodies, however, the overexpression of p300 did not enhance the transcriptional activation of AIRE-regulated genes. Further studies showed that overexpression of p300 stabilized the AIRE protein. Interestingly, gene expression profiling revealed that AIRE, with mutations mimicking K243/K253 acetylation in SAND, was able to activate gene expression, although the affected genes were different and the activation level was lower from those regulated by wild-type AIRE. Our results suggest that the AIRE acetylation can influence the selection of AIRE activated genes.

Expansion of signal pathways in the ectomycorrhizal fungus Laccaria bicolor- evolution of nucleotide sequences and expression patterns in families of protein kinases and RAS small GTPases.

The ectomycorrhizal fungus Laccaria bicolor has the largest genome of all fungi yet sequenced. The large genome size is partly a result of an expansion of gene family sizes. Among the largest gene families are protein kinases and RAS small guanosine triphosphatases (GTPases), which are key components of signal transduction pathways. Comparative genomics and phylogenetic analyses were used to examine the evolution of the two largest families of protein kinases and RAS small GTPases in L. bicolor. Expression levels in various tissues and growth conditions were inferred from microarray data. The two families possessed a large number of young duplicates (paralogs) that had arisen in the Laccaria lineage following the separation from the saprophyte Coprinopsis cinerea. The protein kinase paralogs were dispersed in many small clades and the majority were pseudogenes. By contrast, the RAS paralogs were found in three large groups of RAS1-, RAS2- and RHO1-like GTPases with few pseudogenes. Duplicates of protein kinases and RAS small GTPase have either retained, gained or lost motifs found in the coding regions of their ancestors. Frequent outcomes during evolution were the formation of pseudogenes (nonfunctionalization) or proteins with novel structures and expression patterns (neofunctionalization).

Paralysis of nematodes: shifts in the transcriptome of the nematode-trapping fungus Monacrosporium haptotylum during infection of Caenorhabditis elegans.

The transcriptional response in the parasitic fungus Monacrosporium haptotylum and its nematode host Caenorhabditis elegans were analysed during infection using cDNA microarrays. The array contained 2684 fungal and 372 worm gene reporters. Dramatic shifts occurred in the transcriptome of M. haptotylum during the different stages of the infection. An initial transcriptional response was recorded after 1 h of infection when the traps adhered to the cuticle, but before immobilization of the captured nematodes. Among the differentially expressed genes were two serine protease genes (spr1 and spr2), and several homologues to genes known to be regulated in other pathogenic fungi. After 4 h, when approximately 40% of the nematodes were paralysed, we identified an upregulated cluster of 372 fungal genes which were not regulated during the other phases of the infection. This cohort contained a large proportion (79%) of genes that appear to be specific for M. haptotylum and closely related species. These genes were of two different classes: those translating into presumably functional peptides and those with no apparent protein coding potential (non-coding RNAs). Among the infection-induced C. elegans genes were those encoding antimicrobial peptides, protease inhibitors and lectins.

Evolution of nucleotide sequences and expression patterns of hydrophobin genes in the ectomycorrhizal fungus Paxillus involutus.

Hydrophobins are small, secreted proteins that play important roles in the development of pathogenic and symbiotic fungi. Evolutionary mechanisms generating sequence and expression divergence among members in hydrophobin gene families are largely unknown. Seven hydrophobin (hyd) genes and one hyd pseudogene were isolated from strains of the ectomycorrhizal fungus Paxillus involutus. Sequences were analysed using phylogenetic methods. Expression profiles were inferred from microarray experiments. The hyd genes included both young (recently diverged) and old duplicates. Some young hyd genes exhibited an initial phase of enhanced sequence evolution owing to relaxed or positive selection. There was no significant association between sequence divergence and variation in expression levels. However, three hyd genes displayed a shift in the expression levels or an altered tissue specificity following duplication. The Paxillus hyd genes evolve according to the so-called birth-and-death model in which some duplicates are maintained for a long time, whereas others are inactivated through mutations. The role of subfunctionalization and/or neofunctionalization for preserving the hyd duplicates in the genome is discussed.

Screening for rapidly evolving genes in the ectomycorrhizal fungus Paxillus involutus using cDNA microarrays.

We have examined the variations in gene content and sequence divergence that could be associated with symbiotic adaptations in the ectomycorrhizal fungus Paxillus involutus and the closely related species Paxillus filamentosus. Strains with various abilities to form mycorrhizae were analysed by comparative genomic hybridizations using a cDNA microarray containing 1076 putative unique genes of P. involutus. To screen for genes diverging at an enhanced and presumably non-neutral rate, we implemented a simple rate test using information from both the variations in hybridizations signal and data on sequence divergence of the arrayed genes relative to the genome of Coprinus cinereus. C. cinereus is a free-living saprophyte and is the closest evolutionary relative to P. involutus that has been fully sequenced. Approximately 17% of the genes investigated were detected as rapidly diverging within Paxillus. Furthermore, 6% of the genes varied in copy numbers between the analysed strains. Genome rearrangements associated with this variation including duplications and deletions may also play a role in adaptive evolution. The cohort of divergent and duplicated genes showed an over-representation of either orphans, genes whose products are located at membranes, or genes encoding for components of stress/defence reactions. Some of the identified genomic changes may be associated with the variation in host specificity of ectomycorrhizal fungi. The proposed procedure could be generally applicable to screen for rapidly evolving genes in closely related strains or species where at least one has been sequenced or characterized by expressed sequence tag analysis.