Honeybees Learn Odour Mixtures via a Selection of Key Odorants

BACKGROUND The honeybee has to detect, process and learn numerous complex odours from her natural environment on a daily basis. Most of these odours are floral scents, which are mixtures of dozens of different odorants. To date, it is still unclear how the bee brain unravels the complex information contained in scent mixtures. METHODOLOGY/PRINCIPAL FINDINGS This study investigates learning of complex odour mixtures in honeybees using a simple olfactory conditioning procedure, the Proboscis-Extension-Reflex (PER) paradigm. Restrained honeybees were trained to three scent mixtures composed of 14 floral odorants each, and then tested with the individual odorants of each mixture. Bees did not respond to all odorants of a mixture equally: They responded well to a selection of key odorants, which were unique for each of the three scent mixtures. Bees showed less or very little response to the other odorants of the mixtures. The bees' response to mixtures composed of only the key odorants was as good as to the original mixtures of 14 odorants. A mixture composed of the other, non-key-odorants elicited a significantly lower response. Neither an odorant's volatility or molecular structure, nor learning efficiencies for individual odorants affected whether an odorant became a key odorant for a particular mixture. Odorant concentration had a positive effect, with odorants at high concentration likely to become key odorants. CONCLUSIONS/SIGNIFICANCE Our study suggests that the brain processes complex scent mixtures by predominantly learning information from selected key odorants. Our observations on key odorant learning lend significant support to previous work on olfactory learning and mixture processing in honeybees.This work was supported by a grant from the Commonwealth Scientific and Industrial Research Organisation Food Futures Flagship Collaborative Research Fund (CBR3_45865_9 W2003, http://www.csiro.au/org/FoodFuturesFlagship.html). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript

Hop Harvest Timing

In the Northeast, hop harvest generally begins in mid-August and continues through mid-September. Harvest date is primarily dependent on the hop variety but weather can delay or hasten maturation and impact when harvest will occur. In addition to weather, various pests, such as spider mites and downy mildew, can similarly impact harvest timing. The time at which you harvest hops can affect the various qualities of your finished product. Alpha and beta acid content peaks before many essential oils have fully developed. Delaying harvest can provide time for these oils to develop but increases the amount of time the hops are left vulnerable to disease and fall rains which can result in degradation of resins

Organosulfate Formation in Biogenic Secondary Organic Aerosol

Organosulfates of isoprene, α-pinene, and β-pinene have recently been identified in both laboratory-generated and ambient secondary organic aerosol (SOA). In this study, the mechanism and ubiquity of organosulfate formation in biogenic SOA is investigated by a comprehensive series of laboratory photooxidation (i.e., OH-initiated oxidation) and nighttime oxidation (i.e., NO3-initiated oxidation under dark conditions) experiments using nine monoterpenes (α-pinene, β-pinene, d-limonene, l-limonene, α-terpinene, γ-terpinene, terpinolene, Δ3-carene, and β-phellandrene) and three monoterpenes (α-pinene, d-limonene, and l-limonene), respectively. Organosulfates were characterized using liquid chromatographic techniques coupled to electrospray ionization combined with both linear ion trap and high-resolution time-of-flight mass spectrometry. Organosulfates are formed only when monoterpenes are oxidized in the presence of acidified sulfate seed aerosol, a result consistent with prior work. Archived laboratory-generated isoprene SOA and ambient filter samples collected from the southeastern U.S. were reexamined for organosulfates. By comparing the tandem mass spectrometric and accurate mass measurements collected for both the laboratory-generated and ambient aerosol, previously uncharacterized ambient organic aerosol components are found to be organosulfates of isoprene, α-pinene, β-pinene, and limonene-like monoterpenes (e.g., myrcene), demonstrating the ubiquity of organosulfate formation in ambient SOA. Several of the organosulfates of isoprene and of the monoterpenes characterized in this study are ambient tracer compounds for the occurrence of biogenic SOA formation under acidic conditions. Furthermore, the nighttime oxidation experiments conducted under highly acidic conditions reveal a viable mechanism for the formation of previously identified nitrooxy organosulfates found in ambient nighttime aerosol samples. We estimate that the organosulfate contribution to the total organic mass fraction of ambient aerosol collected from K-puszta, Hungary, a field site with a similar organosulfate composition as that found in the present study for the southeastern U.S., can be as high as 30%

Towards the metabolic engineering of myrcene pathway of pseudomonas sp. M1 using an integrated omic approach

Pseudomonas sp. M1, isolated from the Rhine River, is able to utilize a large variety of toxic and/or recalcitrant compounds as sole carbon and energy sources, including phenols, benzene and monoterpenes like myrcene [1-3]. Therefore, M1 strain holds great potential as a source of novel biomolecules and cell factories for various biotechnological applications namely in biocatalysis, biosensors, bioremediation and biomedicine. However, the full exploitation of its enzymatic repertoire requires detailed and integrated information about the biomolecular catalog of M1 strain, including genes, proteins and metabolites. In this context, the genome of Pseudomonas sp. M1 was sequenced by NGS technologies, using Illumina Genome Analyser IIx and Roche 454 FLX. The resulting raw data was assembled into 41 contigs and annotated using different pipelines. The current genome draft of Pseudomonas sp. M1 has an estimated GC content of 67%, a size of about 6.9 Mbps and includes 6214 CDS. Importantly, in silico genome analysis predicted a number of metabolic pathways involved in utilization/biotransformation of several unusual carbons sources (e.g. biphenyls, halophenols and different monoterpenes). Proteomic and transcriptomic approaches have been setup envisaging the elucidation of the myrcene stimulon. In 2009, a set of myrcene-dependent proteins has been described using subproteome analysis of the cytoplasmic fraction [3]. More recently, a RNA-seq transcriptome analysis led to the identification of a 28kb genomic island of key importance in the catabolism of myrcene. This island includes genes involved in: i) myrcene oxidation and bioconversion of myrcene derivatives via a beta-oxidation like pathway; ii) regulation of myrcene pathway; iii) myrcene sensing. In addition several other gene clusters spread in the genome of Pseudomonas sp. M1 have been found to be myrcene-dependently expressed and are currently being characterized. Integration of genomic, transcriptomic, proteomic and metabolic data (which is currently being setup) will deliver a very solid and detailed description of the myrcene catabolism (and other monoterpenes), and on the associated molecular mechanisms of adaptation, providing the adequate support for the application of M1 as a biocatalyst in whole-cell biotransformations of plant-derived volatiles.Fundação para a Ciência e a Tecnologia (FCT

Odorants for surveillance and control of the Asian Citrus Psyllid (Diaphorina citri).

BackgroundThe Asian Citrus Psyllid (ACP), Diaphorina citri, can transmit the bacterium Candidatus Liberibacter while feeding on citrus flush shoots. This bacterium causes Huanglongbing (HLB), a major disease of citrus cultivation worldwide necessitating the development of new tools for ACP surveillance and control. The olfactory system of ACP is sensitive to variety of odorants released by citrus plants and offers an opportunity to develop new attractants and repellents.ResultsIn this study, we performed single-unit electrophysiology to identify odorants that are strong activators, inhibitors, and prolonged activators of ACP odorant receptor neurons (ORNs). We identified a suite of odorants that activated the ORNs with high specificity and sensitivity, which may be useful in eliciting behavior such as attraction. In separate experiments, we also identified odorants that evoked prolonged ORN responses and antagonistic odorants able to suppress neuronal responses to activators, both of which can be useful in lowering attraction to hosts. In field trials, we tested the electrophysiologically identified activating odorants and identified a 3-odor blend that enhances trap catches by ∼230%.ConclusionThese findings provide a set of odorants that can be used to develop affordable and safe odor-based surveillance and masking strategies for this dangerous pest insect

An integrated omic approach towards the metabolic engineering of myrcene pathway of pseudomonas sp. M1

Best Poster AwardPseudomonas sp. M1 is able to utilize a large variety of toxic and/or recalcitrant compounds as sole carbon and energy sources, including phenols, benzene and monoterpenes like myrcene [1-3]. Therefore, M1 strain holds great potential as a source of novel biomolecules and cell factories for various biotechnological applications namely in biocatalysis, biosensors, bioremediation and biomedicine. However, the full exploitation of its enzymatic repertoire requires detailed and integrated information about the biomolecular catalog of M1 strain, including genes, proteins and metabolites. In this context, the genome of Pseudomonas sp. M1 was sequenced by NGS technologies, using Illumina GA IIx and Roche 454 FLX. The resulting raw data was assembled and annotated using different pipelines. The current genome draft of Pseudomonas sp. M1 has an estimated GC content of 67%, a size of about 7.1 Mbps and includes 6276 CDS. Importantly, in silico genome analysis predicted a number of metabolic pathways involved in utilization/biotransformation of several unusual carbons sources (e.g. biphenyls, halophenols and different monoterpenes). Proteomic and transcriptomic approaches have been setup envisaging the elucidation of the myrcene stimulon. In 2009, a set of myrcene-dependent proteins has been described using subproteome analysis of the cytoplasmic fraction [3]. In this work, a RNA-seq transcriptome analysis led to the identification of a 28kb genomic island of key importance in the catabolism of myrcene. This island includes genes involved in: i) myrcene oxidation and bioconversion of myrcene derivatives via a beta-oxidation like pathway; ii) regulation of myrcene pathway; iii) myrcene sensing. In addition several other gene clusters spread in the genome of Pseudomonas sp. M1 have been found to be myrcene-dependently expressed and are under investigation. Integration of genomic, transcriptomic, proteomic and metabolic data will deliver a very solid and detailed description of the myrcene catabolism (and other monoterpenes), and on the associated molecular mechanisms of adaptation, providing the adequate support for the application of M1 as a biocatalyst in whole-cell biotransformations of plantderived volatiles.Fundação para a Ciência e a Tecnologia (FCT

The oxygen-independent metabolism of cyclic monoterpenes in Castellaniella defragrans 65Phen

BACKGROUND: The facultatively anaerobic betaproteobacterium Castellaniella defragrans 65Phen utilizes acyclic, monocyclic and bicyclic monoterpenes as sole carbon source under oxic as well as anoxic conditions. A biotransformation pathway of the acyclic β-myrcene required linalool dehydratase-isomerase as initial enzyme acting on the hydrocarbon. An in-frame deletion mutant did not use myrcene, but was able to grow on monocyclic monoterpenes. The genome sequence and a comparative proteome analysis together with a random transposon mutagenesis were conducted to identify genes involved in the monocyclic monoterpene metabolism. Metabolites accumulating in cultures of transposon and in-frame deletion mutants disclosed the degradation pathway. RESULTS: Castellaniella defragrans 65Phen oxidizes the monocyclic monoterpene limonene at the primary methyl group forming perillyl alcohol. The genome of 3.95 Mb contained a 70 kb genome island coding for over 50 proteins involved in the monoterpene metabolism. This island showed higher homology to genes of another monoterpene-mineralizing betaproteobacterium, Thauera terpenica 58Eu(T), than to genomes of the family Alcaligenaceae, which harbors the genus Castellaniella. A collection of 72 transposon mutants unable to grow on limonene contained 17 inactivated genes, with 46 mutants located in the two genes ctmAB (cyclic terpene metabolism). CtmA and ctmB were annotated as FAD-dependent oxidoreductases and clustered together with ctmE, a 2Fe-2S ferredoxin gene, and ctmF, coding for a NADH:ferredoxin oxidoreductase. Transposon mutants of ctmA, B or E did not grow aerobically or anaerobically on limonene, but on perillyl alcohol. The next steps in the pathway are catalyzed by the geraniol dehydrogenase GeoA and the geranial dehydrogenase GeoB, yielding perillic acid. Two transposon mutants had inactivated genes of the monoterpene ring cleavage (mrc) pathway. 2-Methylcitrate synthase and 2-methylcitrate dehydratase were also essential for the monoterpene metabolism but not for growth on acetate. CONCLUSIONS: The genome of Castellaniella defragrans 65Phen is related to other genomes of Alcaligenaceae, but contains a genomic island with genes of the monoterpene metabolism. Castellaniella defragrans 65Phen degrades limonene via a limonene dehydrogenase and the oxidation of perillyl alcohol. The initial oxidation at the primary methyl group is independent of molecular oxygen

Characterization of craft beer through flavour component analysis by GC-MS and multivariate statistical tools

Beer is a rather popular drink and represents the most widely consumed alcoholic beverage in the world.The present research aims at characterizing the flavour profile of lager pilsner, the category of low fermentation beers most common in Europe. The largest portion of the global market is dominated by a few multinational companies, but in the last years the number of independent craft breweries has increased very rapidly also in countries where there weren’t an established craft brewing tradition. According to the Italian Brewers Association, in Italy there are eight brewing companies which operate 14 industrial breweries, which in the years have standardized the product to increase their slice of market. The craft-beer sector represents a niche market, about 3% of total production (1% in 2011) [EU Report, 2016]. Italy has a relatively young craft brewing tradition, but the data together with the new ways of consumption, can be considered promising for the sector development. In 2016, the “craft beer” has been defined for the first time in Italy with DDL 1328-B (art.35). Legislation does not consider the quality of the raw materials, but only the manufacturing processes: the artisanal beer-making is a beer obtained without microfiltration and pasteurization steps, unlike industrial products. In this contest, the aim of the study was to characterize the beers (all lager style) purchased on the market through the analysis of the aromatic profile of samples produced under different processes (craft methods or industrial processes). In fact, in addition to smaller production scale and independent, the main characteristic of craft beer is to put the emphasis on flavour and brewing techniques. Flavour, consisting of a large number of volatile compounds, has a great influence on consumer acceptability and, when safety and nutritional value are guaranteed, sensory parameters become the discriminating factor in the product quality assessment which determines the differentiation on the market. The identification of specific compounds, which confer a particular aroma, suitable to be used as potential quality/process markers in order to discriminate beer samples according to their production method. A headspace solid-phase microextraction coupled to gas chromatography-mass spectrometry (HS-SPME-GC-MS) was performed to evaluate the beer samples fingerprint. Multivariate statistical methods were then applied to the collected profiles to built model which could allow differentiating craft beers from all the others. Hence, the proposed method may represent an interesting tool to authenticate craft beer by verification of the compliance with their label description which, at the same time, can entail brand protection

Seasonal measurements of total OH reactivity emission rates from Norway spruce in 2011

Numerous reactive volatile organic compounds (VOCs) are emitted into the atmosphere by vegetation. Most biogenic VOCs are highly reactive towards the atmosphere's most important oxidant, the hydroxyl (OH) radical. One way to investigate the chemical interplay between biosphere and atmosphere is through the measurement of total OH reactivity, the total loss rate of OH radicals. This study presents the first determination of total OH reactivity emission rates (measurements via the comparative reactivity method) based on a branch cuvette enclosure system mounted on a Norway spruce (Picea abies) throughout spring, summer and autumn 2011. In parallel VOC emission rates were monitored by a second proton-transfer-reaction mass spectrometer (PTR-MS), and total ozone (O3) loss rates were obtained inside the cuvette. Total OH reactivity emission rates were in general temperature and light dependent, showing strong diel cycles with highest values during daytime. Monoterpene emissions contributed most, accounting for 56–69% of the measured total OH reactivity flux in spring and early summer. However, during late summer and autumn the monoterpene contribution decreased to 11–16%. At this time, a large missing fraction of the total OH reactivity emission rate (70–84%) was found when compared to the VOC budget measured by PTR-MS. Total OH reactivity and missing total OH reactivity emission rates reached maximum values in late summer corresponding to the period of highest temperature. Total O3 loss rates within the closed cuvette showed similar diel profiles and comparable seasonality to the total OH reactivity fluxes. Total OH reactivity fluxes were also compared to emissions from needle storage pools predicted by a temperature-only-dependent algorithm. Deviations of total OH reactivity fluxes from the temperature-only-dependent emission algorithm were observed for occasions of mechanical and heat stress. While for mechanical stress, induced by strong wind, measured VOCs could explain total OH reactivity emissions, during heat stress they could not. The temperature-driven algorithm matched the diel variation of total OH reactivity emission rates much better in spring than in summer, indicating a different production and emission scheme for summer and early autumn. During these times, unmeasured and possibly unknown primary biogenic emissions contributed significantly to the observed total OH reactivity flux