Molecular phylogeny of the fungus gnat subfamilies Gnoristinae and Mycomyinae,and their position within Mycetophilidae (Diptera)

The phylogeny of the fungus gnat family Mycetophilidae (Diptera) is reconstructed with a focus on the species‐rich and taxonomically difficult subfamilies Gnoristinae and Mycomyinae. The multigene phylogenetic analyses are based on five nuclear (18S, 28S, CAD, MCS, ITS2) and four mitochondrial (12S, 16S, COI, CytB) gene markers. The analyses strongly support the monophyly of Mycetophilidae and the subfamilies Manotinae, Sciophilinae, Leiinae, and Mycomyinae, although Gnoristinae is paraphyletic with respect to Mycetophilinae. All the genera and groups of genera included are supported as monophyletic, except for Acomoptera Vockeroth, Boletina Staeger, Dziedzickia Johannsen, Ectrepesthoneura Enderlein, and Neoempheria Osten Sacken. Ancestral character state reconstructions were applied to two morphological features present in Gnoristinae and Mycomyinae (i.e. presence of setae on wing membrane and wing vein R₄) in order to assess their evolution. The wing vein R₄ appears as an unstable character, spread throughout different clades. A dated phylogeny of the family Mycetophilidae showed that most of the subfamilies of Mycetophilidae originated and diversified during the Cretaceous. The youngest subfamilies, originated in the Paleogene, appear to be Mycomyinae and Mycetophilinae.     

Testing the accuracy of methods for reconstructing ancestral states of continuous characters.

Many methods are available for estimating ancestral values of continuous characteristics, but little is known about how well these methods perform. Here we compare six methods: linear parsimony, squared-change parsimony, one-parameter maximum likelihood (Brownian motion), two-parameter maximum likelihood (Ornstein-Uhlenbeck process), and independent comparisons with and without branch-length information. We apply these methods to data from 20 morphospecies of Pleistocene planktic Foraminifera in order to estimate ancestral size and shape variables, and compare these estimates with measurements on fossils close to the phylogenetic position of 13 ancestors. No method produced accurate estimates for any variable: estimates were consistently less good as predictors of the observed values than were the averages of the observed values. The two-parameter maximum-likelihood model consistently produces the most accurate size estimates overall. Estimation of ancestral sizes is confounded by an evolutionary trend towards increasing size. Shape showed no trend but was still estimated very poorly: we consider possible reasons. We discuss the implications of our results for the use of estimates of ancestral characteristics.     

Isolation of fungi from bats of the Amazon basin

A total of 2,886 bats captured in the Amazon Basin of Brazil were processed for the isolation of fungi. From the livers, spleens, and lungs of 155 bats (5.4%), 186 fungal isolates of the genera Candida (123 isolates), Trichosporon (26 isolates), Torulopsis (25 isolates), Kluyveromyces (11 isolates), and Geotrichum (1 isolate) were recovered. Seven known pathogenic species were present: Candida parapsilosis, C. guilliermondii, C. albicans, C. stellatoidea, C. pseudotropicalis, Trichosporon beigelii, and Torulopsis glabrata. Twenty-three culture-positive bats showed identical fungal colonization in multiple organs or mixed colonization in a single organ. The fungal isolation rates for individual bat species varied from 1 fungus per 87 bats to 3 fungi per 13 bats, and the mycoflora diversity for members of an individual fungus-bearing bat species varied from 16 fungi per 40 bats to 7 fungi per 6 bats. Of the 38 fungal species isolated, 36 had not been previously described as in vivo bat isolates. Of the 27 culture-positive bat species, 21 had not been previously described as mammalian hosts for medically or nonmedically important fungi.     

Determination of duodenogastric reflux by quantification of bile acids in the gastric juice

The existence of duodeno-gastric reflux was evaluated in a group of 15 healthy subjects, in fasting and for 24 hours. The assessment of duodeno-gastric reflux was made by quantitation of the bile acids (BA) present in the gastric juice. The individual free and conjugated BA were separated and quantified by means of thin-layer chromatography and in situ spectrofluorometry. In 7 of the subjects studied no BA were detected, and in the other 8 subjects the BA levels were below 6 mumol reflux/hour. There were no free BA detected in any of the subjects. The levels of BA in gastric juice increased progressively with age, but there were no differences between sex. The chromatographic technique used is highly sensitive for the analysis of BA in biological samples. The study of BA in the gastric juice provides a quantitative and reliable assessment of the degree of duodeno-gastric reflux.     

Genome-wide analysis of the SET DOMAIN GROUP family in grapevine

The SET DOMAIN GROUP (SDG) proteins represent an evolutionarily-conserved family of epigenetic regulators present in eukaryotes and are putative candidates for the catalysis of lysine methylation in histones. Plant genomes analyses of this family have been performed in arabidopsis, maize, and rice and functional studies have shown that SDG genes are involved in the control of plant development. In this work, we describe the identification and structural characterization of SDG genes in the Vitis vinifera genome. This analysis revealed the presence of 33 putative SDG genes that can be grouped into different classes, as it has been previously described for plants. In addition to the SET domain, the proteins identified possessed other domains in the different classes. As part of our study regarding the growth and development of grapevine, we selected eight genes and their expression levels were analyzed in representative vegetative and reproductive organs of this species. The selected genes showed different patterns of expression during inflorescence and fruit development, suggesting that they participate in these processes. Furthermore, we showed that the expression of selected SDGs changes during viral infection, using as a model Grapevine Leafroll Associated Virus 3-infected symptomatic grapevine leaves and fruits. Our results suggest that developmental changes caused by this virus could be the result of alterations in SDG expression.     

A predictive model for freshwater bioassessment (Mondego River,Portugal)

We sampled macroinvertebrates at 75 locations in the Mondego river catchment, Central Portugal, and developed a predictive model for water quality assessment of this basin, based on the Reference Condition Approach. Sampling was done from June to September 2001. Fifty-five sites were identified as “Reference sites” and 20 sites were used as “Test sites” to test the model. At each site we also measured 40 habitat variables to characterize water physics and chemistry, habitat type, land use, stream hydrology and geographic location. Macroinvertebrates were generally identified to species or genus level; a total of 207 taxa were found. By Unweighted Pair Group Method with Arithmetic mean (UPGMA) clustering and analysis of species contribution to similarities percentage (SIMPER), two groups of reference sites were established. Using Discriminant Analysis (stepwise forward), four variables correctly predicted 78% of the reference sites to the appropriate group: stream order, pool quality, substrate quality and current velocity. Test sites’ environmental quality was established from their relative distance to reference sites, in MDS ordination space, using a series of bands (BEAST methodology). The model performed well at upstream sites, but at downstream sites it was compromised by the lack of reference sites. As with the English RIVPACS predictive model, the Mondego model should be continually improved with the addition of new reference sites. The adaptation of the Mondego model methodology to the Water Framework Directive is possible and would consist mainly of the integration of the WFD typology and increasing the number of ellipses that define quality bands. Handling editor: K. Martens     

The Peculiar Facets of Nitric Oxide as a Cellular Messenger: From Disease-Associated Signaling to the Regulation of Brain Bioenergetics and Neurovascular Coupling

In this review, we address the regulatory and toxic role of ^·NO along several pathways, from the gut to the brain. Initially, we address the role on ^·NO in the regulation of mitochondrial respiration with emphasis on the possible contribution to Parkinson’s disease via mechanisms that involve its interaction with a major dopamine metabolite, DOPAC. In parallel with initial discoveries of the inhibition of mitochondrial respiration by ^·NO, it became clear the potential for toxic ^·NO-mediated mechanisms involving the production of more reactive species and the post-translational modification of mitochondrial proteins. Accordingly, we have proposed a novel mechanism potentially leading to dopaminergic cell death, providing evidence that NO synergistically interact with DOPAC in promoting cell death via mechanisms that involve GSH depletion. The modulatory role of NO will be then briefly discussed as a master regulator on brain energy metabolism. The energy metabolism in the brain is central to the understanding of brain function and disease. The core role of ^·NO in the regulation of brain metabolism and vascular responses is further substantiated by discussing its role as a mediator of neurovascular coupling, the increase in local microvessels blood flow in response to spatially restricted increase of neuronal activity. The many facets of NO as intracellular and intercellular messenger, conveying information associated with its spatial and temporal concentration dynamics, involve not only the discussion of its reactions and potential targets on a defined biological environment but also the regulation of its synthesis by the family of nitric oxide synthases. More recently, a novel pathway, out of control of NOS, has been the subject of a great deal of controversy, the nitrate:nitrite:NO pathway, adding new perspectives to ^·NO biology. Thus, finally, this novel pathway will be addressed in connection with nitrate consumption in the diet and the beneficial effects of protein nitration by reactive nitrogen species.

     

Dual Cyclin-Binding Domains Are Required for p107 To Function as a Kinase Inhibitor

The retinoblastoma (pRB) family of proteins includes three proteins known to suppress growth of mammalian cells. Previously we had found that growth suppression by two of these proteins, p107 and p130, could result from the inhibition of associated cyclin-dependent kinases (cdks). One important unresolved issue, however, is the mechanism through which inhibition occurs. Here we present in vivo and in vitro evidence to suggest that p107 is a bona fide inhibitor of both cyclin A-cdk2 and cyclin E-cdk2 that exhibits an inhibitory constant (K_i) comparable to that of the cdk inhibitor p21/WAF1. In contrast, pRB is unable to inhibit cdks. Further reminiscent of p21, a second cyclin-binding site was mapped to the amino-terminal portions of p107 and p130. This amino-terminal domain is capable of inhibiting cyclin-cdk2 complexes, although it is not a potent substrate for these kinases. In contrast, a carboxy-terminal fragment of p107 that contains the previously identified cyclin-binding domain serves as an excellent kinase substrate although it is unable to inhibit either kinase. Clustered point mutations suggest that the amino-terminal domain is functionally important for cyclin binding and growth suppression. Moreover, peptides spanning the cyclin-binding region are capable of interfering with p107 binding to cyclin-cdk2 complexes and kinase inhibition. Our ability to distinguish between p107 and p130 as inhibitors rather than simple substrates suggests that these proteins may represent true inhibitors of cdks.