Differential transcriptome analysis of glandular and filamentous trichomes in Artemisia annua

Background: The medicinal plant Artemisia annua is covered with filamentous trichomes and glandular, artemisinin producing trichomes. A high artemisinin supply is needed at a reduced cost for treating malaria. Artemisinin production in bioreactors can be facilitated if a better insight is obtained in the biosynthesis of artemisinin and other metabolites. Therefore, metabolic activities of glandular and filamentous trichomes were investigated at the transcriptome level. Results: By laser pressure catapulting, glandular and filamentous trichomes as well as apical and sub-apical cells from glandular trichomes were collected and their transcriptome was sequenced using Illumina RNA-Seq. A de novo transcriptome was assembled (Trinity) and studied with a differential expression analysis (edgeR). A comparison of the transcriptome from glandular and filamentous trichomes shows that MEP, MVA, most terpene and lipid biosynthesis pathways are significantly upregulated in glandular trichomes. Conversely, some transcripts coding for specific sesquiterpenoid and triterpenoid enzymes such as 8-epi-cedrol synthase and an uncharacterized oxidosqualene cyclase were significantly upregulated in filamentous trichomes. All known artemisinin biosynthesis genes are upregulated in glandular trichomes and were detected in both the apical and sub-apical cells of the glandular trichomes. No significant differential expression could be observed between the apical and sub-apical cells. Conclusions: Our results underscore the vast metabolic capacities of A. annua glandular trichomes but nonetheless point to the existence of specific terpene metabolic pathways in the filamentous trichomes. Candidate genes that might be involved in artemisinin biosynthesis are proposed based on their putative function and their differential expression level

Reef communities associated with ‘dead’ cold-water coral framework drive resource retention and recycling in the deep sea

Cold-water coral (CWC) reefs create hotspots of metabolic activity in the deep sea, in spite of the limited supply of fresh organic matter from the ocean surface (i.e. phytodetritus). We propose that ‘dead’ coral framework, which harbours diverse faunal and microbial communities, boosts the metabolic activity of the reefs, through enhanced resource retention and recycling. Analysis of a video transect across a 700-540 m-deep CWC mound (Rockall Bank, North-East Atlantic) revealed a high benthic cover of dead framework (64%). Box-cored fragments of dead framework were incubated on-board and showed oxygen consumption rates of 0.078–0.182 μmol O2 (mmol organic carbon, i.e. OC)-1 h-1, indicating a substantial contribution to the total metabolic activity of the CWC reef. During the incubations, it was shown that the framework degradation stage influences nitrogen (re)cycling, corresponding to differences in community composition. New (less-degraded) framework released ammonium (0.005 ± 0.001 μmol NH4+ (mmol OC) 1 h 1), probably due to the activity of ammonotelic macrofauna. In contrast, old (more-degraded) framework released nitrate (0.015 ± 0.008 μmol NO3- (mmol OC)- 1 h- 1), indicating that nitrifying microorganisms recycled fauna-excreted ammonium to nitrate. Furthermore, the framework community removed natural dissolved organic matter (DOM) from the incubation water (0.005–0.122 μmol C (mmol OC)- 1 h- 1). Additional feeding experiments showed that all functional groups and macrofauna taxa of the framework community incorporated 13C-enriched (‘labelled’) DOM, indicating widespread DOM uptake and recycling. Finally, the framework effectively retained 13C-enriched phytodetritus, (a) by physical retention on the biofilm-covered surface and (b) by biological filtration through suspension-feeding fauna. We therefore suggest that the dead framework acts as a ‘filtration-recycling factory’ that enhances the metabolic activity of CWC reefs. The exposed framework, however, is particularly vulnerable to ocean acidification, jeopardizing this important aspect of CWC reef functioning

Tiger reefs: Self‐organized regular patterns in deep‐sea cold‐water coral reefs

Complexity theory predicts that self-organized, regularly patterned ecosystems store more biomass and are more resilient than spatially uniform systems. Self-organized ecosystems are well-known from the terrestrial realm, with “tiger bushes” being the archetypical example and mussel beds and tropical coral reefs the marine examples. We here identify regular spatial patterns in cold-water coral reefs (nicknamed “tiger reefs”) from video transects and argue that these are likely the result of self-organization. We used variograms and Lomb–Scargle analysis of seven annotated video transects to analyze spatial patterns in live coral and dead coral (i.e., skeletal remains) cover at the Logachev coral mound province (NE Atlantic Ocean) and found regular spatial patterns with length scales between 62 and 523 m in live and dead coral distribution along these transects that point to self-organization of cold-water coral reefs. Self-organization theory shows that self-organized ecosystems can withstand large environmental changes by adjusting their spatial configuration. We found indications that cold-water corals can similarly adjust their spatial configuration, possibly providing resilience in the face of climate change. Dead coral framework remains in the environment for extended periods of time, providing a template for spatial patterns that facilitates live coral recovery. The notion of regular spatial patterns in cold-water coral reefs is interesting for cold-water coral restoration, as transplantation will be more successful when it follows the patterns that are naturally present. This finding also underlines that anthropogenic effects such as ocean acidification and bottom trawling that destroy the dead coral template undermine cold-water coral resilience. Differences in the pattern periodicities of live and dead coral cover further present an interesting new angle to investigate past and present environmental conditions in cold-water coral reefs

Tiger reefs: Self-organized regular patterns in deep-sea cold-water coral reefs

Abstract Complexity theory predicts that self-organized, regularly patterned ecosystems store more biomass and are more resilient than spatially uniform systems. Self-organized ecosystems are well-known from the terrestrial realm, with “tiger bushes” being the archetypical example and mussel beds and tropical coral reefs the marine examples. We here identify regular spatial patterns in cold-water coral reefs (nicknamed “tiger reefs”) from video transects and argue that these are likely the result of self-organization. We used variograms and Lomb–Scargle analysis of seven annotated video transects to analyze spatial patterns in live coral and dead coral (i.e., skeletal remains) cover at the Logachev coral mound province (NE Atlantic Ocean) and found regular spatial patterns with length scales between 62 and 523 m in live and dead coral distribution along these transects that point to self-organization of cold-water coral reefs. Self-organization theory shows that self-organized ecosystems can withstand large environmental changes by adjusting their spatial configuration. We found indications that cold-water corals can similarly adjust their spatial configuration, possibly providing resilience in the face of climate change. Dead coral framework remains in the environment for extended periods of time, providing a template for spatial patterns that facilitates live coral recovery. The notion of regular spatial patterns in cold-water coral reefs is interesting for cold-water coral restoration, as transplantation will be more successful when it follows the patterns that are naturally present. This finding also underlines that anthropogenic effects such as ocean acidification and bottom trawling that destroy the dead coral template undermine cold-water coral resilience. Differences in the pattern periodicities of live and dead coral cover further present an interesting new angle to investigate past and present environmental conditions in cold-water coral reefs

Quantitative Bias in Illumina TruSeq and a Novel Post Amplification Barcoding Strategy for Multiplexed DNA and Small RNA Deep Sequencing

Here we demonstrate a method for unbiased multiplexed deep sequencing of RNA and DNA libraries using a novel, efficient and adaptable barcoding strategy called Post Amplification Ligation-Mediated (PALM). PALM barcoding is performed as the very last step of library preparation, eliminating a potential barcode-induced bias and allowing the flexibility to synthesize as many barcodes as needed. We sequenced PALM barcoded micro RNA (miRNA) and DNA reference samples and evaluated the quantitative barcode-induced bias in comparison to the same reference samples prepared using the Illumina TruSeq barcoding strategy. The Illumina TruSeq small RNA strategy introduces the barcode during the PCR step using differentially barcoded primers, while the TruSeq DNA strategy introduces the barcode before the PCR step by ligation of differentially barcoded adaptors. Results show virtually no bias between the differentially barcoded miRNA and DNA samples, both for the PALM and the TruSeq sample preparation methods. We also multiplexed miRNA reference samples using a pre-PCR barcode ligation. This barcoding strategy results in significant bias

Tiger reefs: Self‐organized regular patterns in deep‐sea cold‐water coral reefs

Complexity theory predicts that self‐organized, regularly patterned ecosystems store more biomass and are more resilient than spatially uniform systems. Self‐organized ecosystems are well‐known from the terrestrial realm, with “tiger bushes” being the archetypical example and mussel beds and tropical coral reefs the marine examples. We here identify regular spatial patterns in cold‐water coral reefs (nicknamed “tiger reefs”) from video transects and argue that these are likely the result of self‐organization. We used variograms and Lomb–Scargle analysis of seven annotated video transects to analyze spatial patterns in live coral and dead coral (i.e., skeletal remains) cover at the Logachev coral mound province (NE Atlantic Ocean) and found regular spatial patterns with length scales between 62 and 523 m in live and dead coral distribution along these transects that point to self‐organization of cold‐water coral reefs. Self‐organization theory shows that self‐organized ecosystems can withstand large environmental changes by adjusting their spatial configuration. We found indications that cold‐water corals can similarly adjust their spatial configuration, possibly providing resilience in the face of climate change. Dead coral framework remains in the environment for extended periods of time, providing a template for spatial patterns that facilitates live coral recovery. The notion of regular spatial patterns in cold‐water coral reefs is interesting for cold‐water coral restoration, as transplantation will be more successful when it follows the patterns that are naturally present. This finding also underlines that anthropogenic effects such as ocean acidification and bottom trawling that destroy the dead coral template undermine cold‐water coral resilience. Differences in the pattern periodicities of live and dead coral cover further present an interesting new angle to investigate past and present environmental conditions in cold‐water coral reefs