Ultra-high pressure liquid chromatography-mass spectrometry for plant metabolomics: a systematic comparison of high-resolution quadrupole-time-of-flight and single stage Orbitrap mass spectrometers.

The response of Arabidopsis to stress caused by mechanical wounding was chosen as a model to compare the performances of high resolution quadrupole-time-of-flight (Q-TOF) and single stage Orbitrap (Exactive Plus) mass spectrometers in untargeted metabolomics. Both instruments were coupled to ultra-high pressure liquid chromatography (UHPLC) systems set under identical conditions. The experiment was divided in two steps: the first analyses involved sixteen unwounded plants, half of which were spiked with pure standards that are not present in Arabidopsis. The second analyses compared the metabolomes of mechanically wounded plants to unwounded plants. Data from both systems were extracted using the same feature detection software and submitted to unsupervised and supervised multivariate analysis methods. Both mass spectrometers were compared in terms of number and identity of detected features, capacity to discriminate between samples, repeatability and sensitivity. Although analytical variability was lower for the UHPLC-Q-TOF, generally the results for the two detectors were quite similar, both of them proving to be highly efficient at detecting even subtle differences between plant groups. Overall, sensitivity was found to be comparable, although the Exactive Plus Orbitrap provided slightly lower detection limits for specific compounds. Finally, to evaluate the potential of the two mass spectrometers for the identification of unknown markers, mass and spectral accuracies were calculated on selected identified compounds. While both instruments showed excellent mass accuracy (<2.5ppm for all measured compounds), better spectral accuracy was recorded on the Q-TOF. Taken together, our results demonstrate that comparable performances can be obtained at acquisition frequencies compatible with UHPLC on Q-TOF and Exactive Plus MS, which may thus be equivalently used for plant metabolomics.

Combining probabilistic land-use change and tree population dynamics modelling to simulate responses in mountain forests

Altitudinal tree lines are mainly constrained by temperature, but can also be influenced by factors such as human activity, particularly in the European Alps, where centuries of agricultural use have affected the tree-line. Over the last decades this trend has been reversed due to changing agricultural practices and land-abandonment. We aimed to combine a statistical land-abandonment model with a forest dynamics model, to take into account the combined effects of climate and human land-use on the Alpine tree-line in Switzerland. Land-abandonment probability was expressed by a logistic regression function of degree-day sum, distance from forest edge, soil stoniness, slope, proportion of employees in the secondary and tertiary sectors, proportion of commuters and proportion of full-time farms. This was implemented in the TreeMig spatio-temporal forest model. Distance from forest edge and degree-day sum vary through feed-back from the dynamics part of TreeMig and climate change scenarios, while the other variables remain constant for each grid cell over time. The new model, TreeMig-LAb, was tested on theoretical landscapes, where the variables in the land-abandonment model were varied one by one. This confirmed the strong influence of distance from forest and slope on the abandonment probability. Degree-day sum has a more complex role, with opposite influences on land-abandonment and forest growth. TreeMig-LAb was also applied to a case study area in the Upper Engadine (Swiss Alps), along with a model where abandonment probability was a constant. Two scenarios were used: natural succession only (100% probability) and a probability of abandonment based on past transition proportions in that area (2.1% per decade). The former showed new forest growing in all but the highest-altitude locations. The latter was more realistic as to numbers of newly forested cells, but their location was random and the resulting landscape heterogeneous. Using the logistic regression model gave results consistent with observed patterns of land-abandonment: existing forests expanded and gaps closed, leading to an increasingly homogeneous landscape.

Red blood cell structure and dynamics explored with digital holographic microspcopy

Digital holographic microscopy (DHM) is a technique that allows obtaining, from a single recorded hologram, quantitative phase image of living cell with interferometric accuracy. Specifically the optical phase shift induced by the specimen on the transmitted wave front can be regarded as a powerful endogenous contrast agent, depending on both the thickness and the refractive index of the sample. Thanks to a decoupling procedure cell thickness and intracellular refractive index can be measured separately. Consequently, Mean corpuscular volume (MCV) and mean corpuscular hemoglobin concentration (MCHC), two highly relevant clinical parameters, have been measured non-invasively at a single cell level. The DHM nanometric axial and microsecond temporal sensitivities have permitted to measure the red blood cell membrane fluctuations (CMF) on the whole cell surface. ©2009 COPYRIGHT SPIE--The International Society for Optical Engineering.

Flight metabolism in Panstrongylus megistus (Hemiptera: Reduviidae): the role of carbohydrates and lipids

The metabolism of lipids and carbohydrates related to flight activity in Panstrongylus megistus was investigated. Insects were subjected to different times of flight under laboratory conditions and changes in total lipids, lipophorin density and carbohydrates were followed in the hemolymph. Lipids and glycogen were also assayed in fat body and flight muscle. In resting insects, hemolymph lipids averaged 3.4 mg/ml and significantly increased after 45 min of flight (8.8 mg/ml, P < 0.001). High-density lipophorin was the sole lipoprotein observed in resting animals. A second fraction with lower density corresponding to low-density lipophorin appeared in insects subjected to flight. Particles from both fractions showed significant differences in diacylglycerol content and size. In resting insects, carbohydrate levels averaged 0.52 mg/ml. They sharply declined more than twofold after 15 min of flight, being undetectable in hemolymph of insects flown for 45 min. Lipid and glycogen from fat body and flight muscle decreased significantly after 45 min of flight. Taken together, the results indicate that P. megistus uses carbohydrates during the initiation of the flight after which, switching fuel for flight from carbohydrates to lipids.

Population dynamics of intermediate snail hosts of Fasciola hepatica and some environmental factors in San Juan y Martinez municipality, Cuba

The variation of abundances of intermediate snail hosts of Fasciola hepatica in Cuba (Fossaria cubensis and Pseudosuccinea columella) was studied during one year under natural conditions at five sampling sites in San Juan y Martinez municipality, Pinar del Rio province, Cuba. The effect of some environmental variables on the lymnaeid abundances was also studied. A canonical correspondence analysis showed that both species do not generally occur together in the same habitat and that most factors affect them in an opposite fashion, although both of them correlate positively through time to the diversity of the habitats. F. cubensis prefers the sites that are in or closer to the city whereas P. columella is more abundant in rural sites. Lymnaeid abundances are mainly affected by nitrite and nitrate concentrations as well as by the abundance of the thiarid Tarebia granifera. F. cubensis is more abundant in polluted habitats with low densities (or absence) of T. granifera whereas P. columella prefers cleaner habitats and can coexist with the thiarid, even at its higher densities. The implications of divergent preferences of the two lymnaeids for the control of fasciolosis are discussed.

How local excitation-inhibition ratio impacts the whole brain dynamics.

The spontaneous activity of the brain shows different features at different scales. On one hand, neuroimaging studies show that long-range correlations are highly structured in spatiotemporal patterns, known as resting-state networks, on the other hand, neurophysiological reports show that short-range correlations between neighboring neurons are low, despite a large amount of shared presynaptic inputs. Different dynamical mechanisms of local decorrelation have been proposed, among which is feedback inhibition. Here, we investigated the effect of locally regulating the feedback inhibition on the global dynamics of a large-scale brain model, in which the long-range connections are given by diffusion imaging data of human subjects. We used simulations and analytical methods to show that locally constraining the feedback inhibition to compensate for the excess of long-range excitatory connectivity, to preserve the asynchronous state, crucially changes the characteristics of the emergent resting and evoked activity. First, it significantly improves the model's prediction of the empirical human functional connectivity. Second, relaxing this constraint leads to an unrealistic network evoked activity, with systematic coactivation of cortical areas which are components of the default-mode network, whereas regulation of feedback inhibition prevents this. Finally, information theoretic analysis shows that regulation of the local feedback inhibition increases both the entropy and the Fisher information of the network evoked responses. Hence, it enhances the information capacity and the discrimination accuracy of the global network. In conclusion, the local excitation-inhibition ratio impacts the structure of the spontaneous activity and the information transmission at the large-scale brain level.

Fast analysis of doping agents in urine by ultra-high-pressure liquid chromatography-quadrupole time-of-flight mass spectrometry. II: Confirmatory analysis.

For doping control, analyses of samples are generally achieved in two steps: a rapid screening and, in the case of a positive result, a confirmatory analysis. A two-step methodology based on ultra-high-pressure liquid chromatography coupled to a quadrupole time-of-flight mass spectrometry (UHPLC-QTOF-MS) was developed to screen and confirm 103 doping agents from various classes (e.g., beta-blockers, stimulants, diuretics, and narcotics). The screening method was presented in a previous article as part I (i.e., Fast analysis of doping agents in urine by ultra-high-pressure liquid chromatography-quadrupole time-of-flight mass spectrometry. Part I: screening analysis). For the confirmatory method, basic, neutral and acidic compounds were extracted by a dedicated solid-phase extraction (SPE) in a 96-well plate format and detected by MS in the tandem mode to obtain precursor and characteristic product ions. The mass accuracy and the elemental composition of precursor and product ions were used for compound identification. After validation including matrix effect determination, the method was considered reliable to confirm suspect results without ambiguity according to the positivity criteria established by the World Anti-Doping Agency (WADA). Moreover, an isocratic method was developed to separate ephedrine from its isomer pseudoephedrine and cathine from phenylpropanolamine in a single run, what allowed their direct quantification in urine.

Dynamics of feeding and defecation in Triatoma vitticeps (Stal, 1859) (Hemiptera, Reduviidae, Triatominae) and its potential in the transmission of Trypanosoma cruzi

Adults of Triatoma vitticeps infected by flagellates similar to Trypanosoma cruzi are frequently captured by the inhabitants of rural areas in the Brazilian state of Espírito Santo. The dynamics of feeding and defecation were observed in three groups of adult triatomines, consisting of sylvatic T. vitticeps and laboratory-reared specimens of this species and T. infestans. Triatomines were observed from the moment they were presented with an immobilized chicken as a bloodmeal source until 240 min after feeding had ended. Mean times between the end of feeding and defecation for T. infestans, wild T. vitticeps and laboratory-reared specimens of the latter species were 1.2, 21.1, and 64 min respectively. All T. infestans defecated within 10 min of feeding, while only 29.9 of wild and 52.8% laboratory-reared specimens of T. vitticeps did so within this period. These results may explain the low efficiency of T. vitticeps in T. cruzi transmission to man. The shorter time between feeding and defecation in laboratory-reared T. vitticeps may indicate a change in behaviour of this species as a result of adaptation to an artificial environment.

Biomphalaria tenagophila: dynamics of populations of resistant and susceptible strains to Schistosoma mansoni, with or without pressure of the parasite

Resistant (Taim, RS) and susceptible albino (Joinville, SC) Biomphalaria tenagophila populations were kept together, at different proportions, throughout a 18-month-period. Some of the snail groups were submitted to Schistosoma mansoni infection. The targets of this study were (a) to analyze the populational dynamics among resistant and susceptible individuals to S. mansoni; (b) to study the resistance phenotype in descendants of cross-breeding; (c) to observe whether the parasite could exert any kind of selection in those snail populations. Throughout the experiment it could be observed that the susceptible B. tenagophila strain (Joinville) underwent a selective pressure of the parasite that was negative, since the individuals showed a high mortality rate. Although B. tenagophila (Taim) population presented a higher mortality rate without pressure of the parasite, this event was compensated by a reproductive capacity. B. tenagophila Taim was more fecund than B. tenagophila Joinville and was able to transmit the resistance character to their descendants. F1 generation obtained by cross-breeding between resistant and susceptible lineages was completely resistant to S. mansoni infection, irrespective of the Taim proportion. Moreover, less than 5% of F2 progeny were susceptible to S. mansoni infection.

Diverse population dynamics of three Anopheles species belonging to the Triannulatus Complex (Diptera: Culicidae)

The Triannulatus Complex of Anopheles (Nyssorhynchus) consists of at least three sibling species, namely Anopheles triannulatus s.s., Anopheles halophylus and a third undescribed member herein referred to as An. triannulatus "C". Sympatric anophelines belonging to species complexes, even though closely related, may exploit different environments such as larval habitats. In this paper we hypothesize that rainfall and seasonal flooding would distinctly influence the availability of larval habitats and consequently the seasonal population dynamics of sympatric members of the Triannulatus Complex. A reflection of this is distinct seasonal biting frequencies exhibited by three members of the Triannulatus Complex at a site in Central Brazil. Population dynamics seem to be influenced by the water level in the local rivers, although biting frequency of all three species was negatively influenced by rainfall. An. triannulatus s.s. was more abundant following the end of the rainy season, but notably 30 to 60 days after flooding. On the other hand, An. halophylus and An. triannulatus C peaked during the middle of the dry season, when water impoundments have no inflow, are somewhat reduced in size and the water becomes brackish. Differences in population dynamics were greater between An. triannulatus s.s. and An. halophylus and An. triannulatus C than between An. halophylus and An. triannulatus C. This might reflect differences in larval habitat exploitation and therefore spatial segregation among these members of the complex.

Population dynamics of Triatoma vitticeps (Stål, 1859) in Itanhomi, Minas Gerais, Brazil

Taxonomic markers (head structure morphometry, isoenzymes and randon amplified polymorphism of DNA - RAPD) were used to understand the population dynamics of Triatoma vitticeps, predominant triatomine species in Itanhomi district, using samples obtained from domestic, peridomiciliary and sylvatic habitats. Morphometric analysis revealed sexual dimorphism within the three samples although specimens could not be separated according to the habitat in which they were captured. Forty-two bands were analyzed from RAPD profiles generated using four primers. A dendrogram constructed from Dice's similarity coefficient values showed that migration of the insects between the habitats has occurred, without structuring of populations. Moreover, the dendrogram obtained from the genetic distance values showed an important gene flow between the sylvatic and domestic habitats. No polymorphism was found in the electrophoretic mobility of proteins for the ten enzymes studied. Our results revealed movement of triatomines between the three habitats, suggesting that the presence of T. vitticeps in houses should not be ignored. As invasion of houses by sylvatic insects is frequent and the natural infection indices of this species are among the highest known, epidemiological vigilance studies may reveal possible changes in T. vitticeps behaviour which could present future risks to public health.