(Methylthio)phenol semiochemicals are exploited by deceptive orchids as sexual attractants for Campylothynnus thynnine wasps

Until recently, (methylthio)phenols as natural products had only been reported from bacteria. Now, four representatives of this class of sulfurous aromatic compounds have been discovered as semiochemicals in the orchid Caladenia crebra, which secures pollination by sexual deception. In this case, field bioassays confirmed that a 10:1 blend of 2-(methylthio)benzene-1,4-diol (1) and 4-hydroxy-3-(methylthio)benzaldehyde (2) sexually attracts the male thynnine wasp Campylothynnus flavopictus (Tiphiidae:Thynnineae), the exclusive pollinator of C. crebra. Here we show with field bioassays that another undescribed species of Campylothynnus (sp. A) is strongly sexually attracted to a 1:1 blend of compounds 1 and 2, which elicits very high attempted copulation rates (88%). We also confirm that this Campylothynnus species is a pollinator of Caladenia attingens subsp. attingens. Chemical analysis of the flowers of this orchid revealed two (methylthio)phenols, compound 2 and 2-(methylthio)phenol (3), as candidate semiochemicals involved in pollinator attraction. Thus, (methylthio)phenols are likely to be more widely used than presently known. The confirmation of this Campylothynnus as a pollinator of C. attingens subsp. attingens at our study sites was unexpected, since elsewhere this orchid is pollinated by a different thynnine wasp (Thynnoides sp). In general, sexually deceptive Caladenia only use a single species of pollinator, and as such, this unusual case may offer a tractable study system for understanding the chemical basis of pollinator switching in sexually deceptive orchids.BB and RDP: Australian Research Council (ARC) Discovery Early Career Researcher Awards (DE 160101313 and DE150101720), RDP: the Australian Orchid Foundation and the Holsworth Wildlife Research Endowment, RP and GRF: ARC grant (LP130100162) and RP: ARC grant (DP150102762)

Three Chemically Distinct Floral Ecotypes in Drakaea livida, an Orchid Pollinated by Sexual Deception of Thynnine Wasps

Sexually deceptive orchids are unusual among plants in that closely related species typically attract different pollinator species using contrasting blends of floral volatiles. Therefore, intraspecific variation in pollinator attraction may also be underpinned by differences in floral volatiles. Here, we tested for the presence of floral ecotypes in the sexually deceptive orchid Drakaea livida and investigated if the geographic range of floral ecotypes corresponded to variation in pollinator availability. Pollinator choice trials revealed the presence of three floral ecotypes within D. livida that each attracts a different species of thynnine wasp as a pollinator. Surveys of pollinator distribution revealed that the distribution of one of the ecotypes was strongly correlated with that of its pollinator, while another pollinator species was present throughout the range of all three ecotypes, demonstrating that pollinator availability does not always correlate with ecotype distribution. Floral ecotypes differed in chemical volatile composition, with a high degree of separation evident in principal coordinate analysis. Some compounds that differed between ecotypes, including pyrazines and (methylthio)phenols, are known to be electrophysiologically active in thynnine wasp antennae. Based on differences in pollinator response and floral volatile profile, the ecotypes represent distinct entities and should be treated as such in conservation management

The volatile chemistry of orchid pollination

Covering: up to September 2022 Orchids are renowned not only for their diversity of floral forms, but also for their many and often highly specialised pollination strategies. Volatile semiochemicals play a crucial role in the attraction of a wide variety of insect pollinators of orchids. The compounds produced by orchid flowers are as diverse as the pollinators they attract, and here we summarise some of the chemical diversity found across orchid taxa and pollination strategies. We focus on compounds that have been experimentally demonstrated to underpin pollinator attraction. We also highlight the structural elucidation and synthesis of a select subset of important orchid pollinator attractants, and discuss the ecological significance of the discoveries, the gaps in our current knowledge of orchid pollination chemistry, and some opportunities for future research in this field

Three Chemically Distinct Floral Ecotypes in Drakaea livida, an Orchid Pollinated by Sexual Deception of Thynnine Wasps

Sexually deceptive orchids are unusual among plants in that closely related species typically attract different pollinator species using contrasting blends of floral volatiles. Therefore, intraspecific variation in pollinator attraction may also be underpinned by differences in floral volatiles. Here, we tested for the presence of floral ecotypes in the sexually deceptive orchid Drakaea livida and investigated if the geographic range of floral ecotypes corresponded to variation in pollinator availability. Pollinator choice trials revealed the presence of three floral ecotypes within D. livida that each attracts a different species of thynnine wasp as a pollinator. Surveys of pollinator distribution revealed that the distribution of one of the ecotypes was strongly correlated with that of its pollinator, while another pollinator species was present throughout the range of all three ecotypes, demonstrating that pollinator availability does not always correlate with ecotype distribution. Floral ecotypes differed in chemical volatile composition, with a high degree of separation evident in principal coordinate analysis. Some compounds that differed between ecotypes, including pyrazines and (methylthio)phenols, are known to be electrophysiologically active in thynnine wasp antennae. Based on differences in pollinator response and floral volatile profile, the ecotypes represent distinct entities and should be treated as such in conservation management.The Holsworth Wildlife Research Endowment and the Australian Systematic Botany Society are thanked for their provision of research funding. AMW was supported by an Australian Government Research Training Program (RTP), and BB and RD were supported by Australian Research Council (ARC) Discovery Early Career Researcher Awards (DE 160101313 and DE150101720)

Lipidomic features of honey bee and colony health during limited supplementary feeding

Honey bee nutritional health depends on nectar and pollen, which provide the main source of carbohydrates, proteins and lipids to individual bees. During malnutrition, insect metabolism accesses fat body reserves. However, this process in bees and its repercussions at the colony level are poorly understood. Using untargeted lipidomics and gene expression analysis, we examined the effects of different feeding treatments (starvation, sugar feeding and sugar + pollen feeding) on bees and correlated them with colony health indicators. We found that nutritional stress led to an increase in unsaturated triacylglycerols and diacylglycerols, as well as a decrease in free fatty acids in the bee fat body. Here, we hypothesise that stored lipids are made available through a process where unsaturations change lipid\u27s structure. Increased gene expression of three lipid desaturases in response to malnutrition supports this hypothesis, as these desaturases may be involved in releasing fatty acyl chains for lipolysis. Although nutritional stress was evident in starving and sugar-fed bees at the colony and physiological level, only starved colonies presented long-term effects in honey production

Identification of the Cat Attractants Isodihydronepetalactone and Isoiridomyrmecin from Acalypha indica

Acalypha indica is a herb that grows throughout the tropical regions of the world. As well as being exploited for medicinal use, the roots of this plant are known to elicit a drug-like effect in cats. Recent research into feral cat control on Christmas Island has investigated whether a preparation of the roots of A. indica might be effective in traps to attract feral cats. However, the volatile nature of the attractants made it unviable for use in traps for more than a few days. In this study, we investigated the volatile components emitted by the plant roots and identified two iridoid compounds, (4R,4aR,7S,7aR)- isodihydronepetalactone and (4R,4aS,7S,7aR)-isoiridomyrmecin, which are known to affect behavioural activity in cats. Synthesis of standards confirmed the stereochemistry of both compounds emitted by the plant. Potential application of these compounds in feral cat control is discussed.The Australian Research Council is also gratefully acknowledged for funding for G.F. (FT110100304) and B.B. (LP130100162)

2-(Tetrahydrofuran-2-yl)acetic Acid and Ester Derivatives as Long-Range Pollinator Attractants in the Sexually Deceptive Orchid Cryptostylis ovata

Sexually deceptive orchids achieve pollination by luring male insects to flowers through chemical and sometimes visual mimicry of females. An extreme example of this deception occurs in Cryptostylis, one of only two genera where sexual deception is known to induce pollinator ejaculation. In the present study, bioassay-guided fractionations of Cryptostylis solvent extracts in combination with field bioassays were implemented to isolate and identify floral volatiles attractive to the pollinator Lissopimpla excelsa. ( S)-2-(Tetrahydrofuran-2-yl)acetic acid [( S)-1] and the ester derivatives methyl ( S)-2-(tetrahydrofuran-2-yl)acetate [( S)-2] and ethyl ( S)-2-(tetrahydrofuran-2-yl)acetate [( S)-3], all previously unknown semiochemicals, were confirmed to attract L. excelsa males in field bioassays. Chiral-phase GC and HPLC showed that the natural product 1 comprised a single enantiomer, its S-configuration being confirmed by synthesis of the two enantiomers from known enantiomers of tetrahydrofuran-2-carboxylic acid.B.B. and R.D.P.: Australian Research Council (ARC) Discovery Early Career Researcher Awards (DE 160101313 and DE150101720), A.M.W.: Australian Government Research Training Program, and Australian Orchid Foundation Grant 319.17, R.P. and G.R.F.: ARC Linkage Program Award (LP130100162). D. Bainbridge is gratefully acknowledged for designing and fabricating the preparative GC collector used in this stud

Complex Sexual Deception in an Orchid Is Achieved by Co-opting Two Independent Biosynthetic Pathways for Pollinator Attraction

Sexually deceptive orchids lure their specific male pollinators using volatile semiochemicals that mimic female sex pheromones. To date, the semiochemicals known to be involved consist of blends of chemically and biosynthetically related compounds. In contrast, we report that (S)-β-citronellol and 2-hydroxy-6-methylacetophenone, two biosynthetically distinct compounds, are the active semiochemicals in Caladenia plicata, which is pollinated by male Zeleboria sp. thynnine wasps. They are also sex pheromone components of the female Zeleboria. A 1:4 blend elicits a high rate of attempted copulation (∼70%) in bioassays, equivalent to rates observed at orchid flowers. Whereas β-citronellol is well known, 2-hydroxy-6-methylacetophenone appears to be previously unknown as a floral volatile. Production of the two compounds is restricted to glandular sepal tips; thus, differential expression analysis of contrasting floral tissue transcriptomes was employed to illuminate the biosynthesis. As expected, production of (S)-β-citronellol commences with the terpene synthase GES1 catalyzing the irreversible conversion of geranyl diphosphate (GPP) to geraniol. Contrary to prediction, biosynthesis subsequently proceeds in three steps, commencing with the oxidation of geraniol to geranial by alcohol dehydrogenase ADH3, followed by the enantioselective reduction of a double bond in geranial by geranial reductase GER1 to give (S)-β-citronellal. Finally, ADH3-catalyzed reduction of (S)-β-citronellal results in (S)-β-citronellol. In line with previous work on insects showing that 2-hydroxy-6-methylacetophenone is derived from a polyketide pathway, we report a differentially expressed polyketide synthase (PKS) gene candidate. Thus, in this unique example of sexual deception, pollination is achieved by co-opting and regulating two independent biosynthetic pathways of floral volatile compounds

LATERAL BRANCHING OXIDOREDUCTASE acts in the final stages of strigolactone biosynthesis inArabidopsis

Strigolactones are a group of plant compounds of diverse but related chemical structures. They have similar bioactivity across a broad range of plant species, act to optimize plant growth and development, and promote soil microbe interactions. Carlactone, a common precursor to strigolactones, is produced by conserved enzymes found in a number of diverse species. Versions of the MORE AXILLARY GROWTH1 (MAX1) cytochrome P450 from rice and Arabidopsis thaliana make specific subsets of strigolactones from carlactone. However, the diversity of natural strigolactones suggests that additional enzymes are involved and remain to be discovered. Here, we use an innovative method that has revealed a missing enzyme involved in strigolactone metabolism. By using a transcriptomics approach involving a range of treatments that modify strigolactone biosynthesis gene expression coupled with reverse genetics, we identified LATERAL BRANCHING OXIDOREDUCTASE (LBO), a gene encoding an oxidoreductase-like enzyme of the 2-oxoglutarate and Fe(II)-dependent dioxygenase superfamily. Arabidopsis lbo mutants exhibited increased shoot branching, but the lbo mutation did not enhance the max mutant phenotype. Grafting indicated that LBO is required for a graft-transmissible signal that, in turn, requires a product of MAX1. Mutant lbo backgrounds showed reduced responses to carlactone, the substrate of MAX1, and methyl carlactonoate (MeCLA), a product downstream of MAX1. Furthermore, lbo mutants contained increased amounts of these compounds, and the LBO protein specifically converts MeCLA to an unidentified strigolactone-like compound. Thus, LBO function may be important in the later steps of strigolactone biosynthesis to inhibit shoot branching in Arabidopsis and other seed plants

Symbiodinium Genotypic and Environmental Controls on Lipids in Reef Building Corals

BACKGROUND: Lipids in reef building corals can be divided into two classes; non-polar storage lipids, e.g. wax esters and triglycerides, and polar structural lipids, e.g. phospholipids and cholesterol. Differences among algal endosymbiont types are known to have important influences on processes including growth and the photobiology of scleractinian corals yet very little is known about the role of symbiont types on lipid energy reserves. METHODOLOGY/PRINCIPAL FINDINGS: The ratio of storage lipid and structural lipid fractions of Scott Reef corals were determined by thin layer chromatography. The lipid fraction ratio varied with depth and depended on symbiont type harboured by two corals (Seriatopora hystrix and Pachyseris speciosa). S. hystrix colonies associated with Symbiodinium C1 or C1/C# at deep depths (>23 m) had lower lipid fraction ratios (i.e. approximately equal parts of storage and structural lipids) than those with Symbiodinium D1 in shallow depths (<23 m), which had higher lipid fraction ratios (i.e. approximately double amounts of storage relative to structural lipid). Further, there was a non-linear relationship between the lipid fraction ratio and depth for S. hystrix with a modal peak at ∼23 m coinciding with the same depth as the shift from clade D to C types. In contrast, the proportional relationship between the lipid fraction ratio and depth for P. speciosa, which exhibited high specificity for Symbiodinium C3 like across the depth gradient, was indicative of greater amounts of storage lipids contained in the deep colonies. CONCLUSIONS/SIGNIFICANCE: This study has demonstrated that Symbiodinium exert significant controls over the quality of coral energy reserves over a large-scale depth gradient. We conclude that the competitive advantages and metabolic costs that arise from flexible associations with divergent symbiont types are offset by energetic trade-offs for the coral host