Development and deployment of high-throughput retrotransposon-based markers reveal genetic diversity and population structure of Asian bamboo

Bamboo, a non-timber grass species, known for exceptionally fast growth, is a commercially viable crop. Long terminal repeat (LTR) retrotransposons, the main class I mobile genetic elements in plant genomes, are highly abundant (46%) in bamboo contributing to genome diversity. They play significant roles in the regulation of gene expression, chromosome size and structure as well as in genome integrity. Inter-retrotransposon amplified polymorphism (IRAP) is a high-throughput method to study the genetic diversity of plant species. Till date, there are no markers based on Transposable Elements (TEs) for the bamboo genome and no reports on bamboo genetic diversity using the IRAP method. Phyllostachys is an Asian bamboo, the largest group in the bamboo subfamily, Bambusoideae, and it is of great economic value due to its fast growth. The structure of LTR-retrotransposon superfamilies, Ty3-gypsy and Ty1-copia, were analysed in the bamboo genome using LTRharvest and LTRdigest software. A total of 98,850 LTR retrotransposons with both ends of intact LTR sequences were identified, grouped into 64,281 clusters/scaffolds, using CD-HIT software. Among the total of 64,281 clusters, 13 clusters had more than 30 copy numbers of LTR sequences and at least one copy had all intact protein domains such as gag protein and polyprotein. Based on the high copy numbers of conserved LTR sequences, a total of 16 IRAP primers were developed. All these IRAP primers were used to study the genetic diversity and population structure of the Asian bamboo. AMOVA analysis was done for 58 Asian bamboo species collected from nine different provinces of China, from Italy and America. In the bamboo species, these IRAP primers produced a total of 3340 amplicons with an average of 98% polymorphism. The 58 Asian bamboo species were grouped into two major clusters and four sub-clusters, based on UPGMA analysis. UPGMA cluster analysis was corroborated by statistical analyses of genetic similarity coefficients. Structure analysis showed that the bamboo species could be divided into four subpopulations (K = 4): SP1, SP2, SP3 and SP4. All SPs had an admixture of alleles. AMOVA analysis showed that higher genetic variations occurred within populations (75%) rather than among populations (25%). The study highlights the usability of IRAP in Asian bamboo to determine inter-species variability using retrotransposon markers.Peer reviewe

Long terminal repeats (LTR) and transcription factors regulate PHRE1 and PHRE2 activity in Moso bamboo under heat stress

Background LTR retrotransposons play a significant role in plant growth, genome evolution, and environmental stress response, but their regulatory response to heat stress remains unclear. We have investigated the activities of two LTR retrotransposons, PHRE1 and PHRE2, of moso bamboo (Phyllostachys edulis) in response to heat stress. Results The differential overexpression of PHRE1 and PHRE2 with or without CaMV35s promoter showed enhanced expression under heat stress in transgenic plants. The transcriptional activity studies showed an increase in transposition activity and copy number among moso bamboo wild type and Arabidopsis transgenic plants under heat stress. Comparison of promoter activity in transgenic plants indicated that 5'LTR promoter activity was higher than CaMV35s promoter. Additionally, yeast one-hybrid (Y1H) system and in planta biomolecular fluorescence complementation (BiFC) assay revealed interactions of heat-dependent transcription factors (TFs) with 5'LTR sequence and direct interactions of TFs with pol and gag. Conclusions Our results conclude that the 5'LTR acts as a promoter and could regulate the LTR retrotransposons in moso bamboo under heat stress.Peer reviewe

Metagenomic Analysis of the Bioremediation of Diesel-Contaminated Canadian High Arctic Soils

As human activity in the Arctic increases, so does the risk of hydrocarbon pollution events. On site bioremediation of contaminated soil is the only feasible clean up solution in these remote areas, but degradation rates vary widely between bioremediation treatments. Most previous studies have focused on the feasibility of on site clean-up and very little attention has been given to the microbial and functional communities involved and their ecology. Here, we ask the question: which microorganisms and functional genes are abundant and active during hydrocarbon degradation at cold temperature? To answer this question, we sequenced the soil metagenome of an ongoing bioremediation project in Alert, Canada through a time course. We also used reverse-transcriptase real-time PCR (RT-qPCR) to quantify the expression of several hydrocarbon-degrading genes. Pseudomonas species appeared as the most abundant organisms in Alert soils right after contamination with diesel and excavation (t = 0) and one month after the start of the bioremediation treatment (t = 1m), when degradation rates were at their highest, but decreased after one year (t = 1y), when residual soil hydrocarbons were almost depleted. This trend was also reflected in hydrocarbon degrading genes, which were mainly affiliated with Gammaproteobacteria at t = 0 and t = 1m and with Alphaproteobacteria and Actinobacteria at t = 1y. RT-qPCR assays confirmed that Pseudomonas and Rhodococcus species actively expressed hydrocarbon degradation genes in Arctic biopile soils. Taken together, these results indicated that biopile treatment leads to major shifts in soil microbial communities, favoring aerobic bacteria that can degrade hydrocarbons

Spatial patterns of microbial diversity and activity in an aged creosote-contaminated site

Restoration of polluted sites via in situ bioremediation relies heavily on the indigenous microbes and their activities. Spatial heterogeneity of microbial populations, contaminants and soil chemical parameters on such sites is a major hurdle in optimizing and implementing an appropriate bioremediation regime. We performed a grid-based sampling of an aged creosote-contaminated site followed by geostatistical modelling to illustrate the spatial patterns of microbial diversity and activity and to relate these patterns to the distribution of pollutants. Spatial distribution of bacterial groups unveiled patterns of niche differentiation regulated by patchy distribution of pollutants and an east-to-west pH gradient at the studied site. Proteobacteria clearly dominated in the hot spots of creosote pollution, whereas the abundance of Actinobacteria, TM7 and Planctomycetes was considerably reduced from the hot spots. The pH preferences of proteobacterial groups dominating in pollution could be recognized by examining the order and family-level responses. Acidobacterial classes came across as generalists in hydrocarbon pollution whose spatial distribution seemed to be regulated solely by the pH gradient. Although the community evenness decreased in the heavily polluted zones, basal respiration and fluorescein diacetate hydrolysis rates were higher, indicating the adaptation of specific indigenous microbial populations to hydrocarbon pollution. Combining the information from the kriged maps of microbial and soil chemistry data provided a comprehensive understanding of the long-term impacts of creosote pollution on the subsurface microbial communities. This study also highlighted the prospect of interpreting taxa-specific spatial patterns and applying them as indicators or proxies for monitoring polluted sites