Farmers' perceptions, knowledge, and management of coffee pests and diseases and their natural enemies in Chiapas, Mexico

Small farmers' perceptions of coffee Coffea arabica L. herbivores and their natural enemies, how those perceptions relate to field infestation levels, and pest management practices being implemented by members from two organic and nonorganic coffee grower organizations in the Soconusco region, southeastern Mexico, were analyzed through an interview survey, diagnostic workshops, and field sampling. The terms pest, disease, and damage were commonly used as synonyms. The major phytophagous species, as perceived by the interviewees, were Hypothenemus hampei (Ferrari), and to a lesser extent the fungi Corticium koleroga Cooke (H?hnel) and Hemileia vastatrix Berkeley & Broome. Among the nonorganic farmers, other nonpest-related constraints were regarded as more important. Awareness of the existence of natural enemies was low, despite more organic farmers have used the ectoparasitoid bethylid Cephalonomia stephanoderis Betrem against H. hampei. Labor supplied by household members was most frequent for pest control; only organic farmers exchanged labor for this purpose. The levels of infestation by H. hampei, Leucoptera coffeella Guérin-Méneville, and C. koleroga were lower within the organic coffee stands. However, a low effectiveness for pest control was commonly perceived, probably due to a feeling, among the organic farmers, of a low impact of their pest management extension service, whereas a lack of motivation was prevalent among the nonorganic farmers, shown by a concern with their low coffee yields and the emigration of youth. The importance of understanding farmers' perceptions and knowledge of pests and their natural enemies and the need for participatory pest management approaches, are discussed.     

Stability studies of amphetamine and ephedrine derivatives in urine

Knowledge of the stability of drugs in biological specimens is a critical consideration for the interpretation of analytical results. Identification of proper storage conditions has been a matter of concern for most toxicology laboratories (both clinical and forensic), and the stability of drugs of abuse has been extensively studied. This concern should be extended to other areas of analytical chemistry like antidoping control. In this work, the stability of ephedrine derivatives (ephedrine, norephedrine, methylephedrine, pseudoephedrine, and norpseudoephedrine), and amphetamine derivatives (amphetamine, methamphetamine, 3,4-methylenedioxyamphetamine (MDA), and 3,4-methylenedioxymethamphetamine (MDMA)) in urine has been studied. Spiked urine samples were prepared for stability testing. Urine samples were quantified by GC/NPD or GC/MS. The homogeneity of each batch of sample was verified before starting the stability study. The stability of analytes was evaluated in sterilized and non-sterilized urine samples at different storage conditions. For long-term stability testing, analyte concentration in urine stored at 4 degrees C and -20 degrees C was determined at different time intervals for 24 months for sterile urine samples, and for 6 months for non-sterile samples. For short-term stability testing, analyte concentration was evaluated in liquid urine stored at 37 degrees C for 7 days. The effect of repeated freezing (at -20 degrees C) and thawing (at room temperature) was also studied in sterile urine for up to three cycles. No significant loss of the analytes under study was observed at any of the investigated conditions. These results show the feasibility of preparing reference materials containing ephedrine and amphetamine derivatives to be used for quality control purposes.     

Development of midline cell types and commissural axon tracts requires Fgfr1 in the cerebrum

The adult cerebral hemispheres are connected to each other by specialized midline cell types and by three axonal tracts: the corpus callosum, the hippocampal commissure, and the anterior commissure. Many steps are required for these tracts to form, including early patterning and later axon pathfinding steps. Here, the requirement for FGF signaling in forming midline cell types and commissural axon tracts of the cerebral hemispheres is examined. Fgfr1, but not Fgfr3, is found to be essential for establishing all three commissural tracts. In an Fgfr1 mutant, commissural neurons are present and initially project their axons, but these fail to cross the midline that separates the hemispheres. Moreover, midline patterning defects are observed in the mutant. These defects include the loss of the septum and three specialized glial cell types, the indusium griseum glia, midline zipper glia, and glial wedge. Our findings demonstrate that FGF signaling is required for generating telencephalic midline structures, in particular septal and glial cell types and all three cerebral commissures. In addition, analysis of the Fgfr1 heterozygous mutant, in which midline patterning is normal but commissural defects still occur, suggests that at least two distinct FGF-dependent mechanisms underlie the formation of the cerebral commissures.     

Putative Glycosyltransferases and Other Plant Golgi Apparatus Proteins Are Revealed by LOPIT Proteomics

The Golgi apparatus is the central organelle in the secretory pathway and plays key roles in glycosylation, protein sorting, and secretion in plants. Enzymes involved in the biosynthesis of complex polysaccharides, glycoproteins, and glycolipids are located in this organelle, but the majority of them remain uncharacterized. Here, we studied the Arabidopsis (Arabidopsis thaliana) membrane proteome with a focus on the Golgi apparatus using localization of organelle proteins by isotope tagging. By applying multivariate data analysis to a combined data set of two new and two previously published localization of organelle proteins by isotope tagging experiments, we identified the subcellular localization of 1,110 proteins with high confidence. These include 197 Golgi apparatus proteins, 79 of which have not been localized previously by a high-confidence method, as well as the localization of 304 endoplasmic reticulum and 208 plasma membrane proteins. Comparison of the hydrophobic domains of the localized proteins showed that the single-span transmembrane domains have unique properties in each organelle. Many of the novel Golgi-localized proteins belong to uncharacterized protein families. Structure-based homology analysis identified 12 putative Golgi glycosyltransferase (GT) families that have no functionally characterized members and, therefore, are not yet assigned to a Carbohydrate-Active Enzymes database GT family. The substantial numbers of these putative GTs lead us to estimate that the true number of plant Golgi GTs might be one-third above those currently annotated. Other newly identified proteins are likely to be involved in the transport and interconversion of nucleotide sugar substrates as well as polysaccharide and protein modification.The Golgi apparatus is the central organelle in the secretory pathway, and in higher plants it is involved in the biosynthesis and transport of cell wall matrix polysaccharides, glycoproteins, proteoglycans, and glycolipids as well as in protein trafficking to different subcellular compartments. The last decade has produced substantial findings on the function of the Golgi apparatus: insights into the protein trafficking at the endoplasmic reticulum (ER)/Golgi interface, Golgi structural maintenance, its involvement in endocytosis, and its behavior during cell division (for review, see Faso et al., 2009). However, despite its importance, only a small proportion of the Golgi proteome has been studied: relatively few Golgi proteins have been localized, and even fewer have been functionally characterized.The Golgi apparatus is thought to contain a large and diverse group of membrane-bound glycosyltransferases (GTs). The current view is that different GT activities are required for synthesis of the linkage between different donor and acceptor sugars. Having in mind the diversity of linkage types found in cell wall polysaccharides, the number of different GTs involved is likely to be very large. For instance, it has been estimated that for the biosynthesis of pectin alone, the action of 65 different enzymatic activities is needed (Caffall and Mohnen, 2009). By the end of the year 2011, 468 Arabidopsis (Arabidopsis thaliana) sequences had been annotated in the Carbohydrate-Active EnZymes (CAZy) GT database (Cantarel et al., 2009; http://www.cazy.org). We estimate that two-thirds of these CAZy-classified GTs may be targeted to the Golgi. The remaining one-third are cytosolic or plastidic enzymes involved in processes including, secondary metabolism or starch synthesis. The reported sequences are classified into 43 CAZy families based on amino acid sequence similarities within which at least one member has been biochemically characterized. Each family is likely to have a common structural fold, and three-dimensional (3-D) structures have been resolved for 20 of these 43 families. These are divided mostly into two structural classes, having either a GT-A fold or a GT-B fold (Unligil and Rini, 2000; Bourne and Henrissat, 2001). Moreover, most of the structurally uncharacterized GT families are predicted to adopt either the GT-A or GT-B fold based on 3-D structural homology modeling (Coutinho et al., 2003; Lairson et al., 2008). Despite this conserved 3-D structure, different GT families have very low or undetectable sequence similarities. Consequently, predicting novel GTs based solely on their amino acid sequence similarities is not always achievable, and structural homology searches have also proven useful (Hansen et al., 2009).The length and properties of the transmembrane domain (TMD) of endomembrane proteins appear to play a role in protein sorting and location within the secretory pathway and can be used to predict protein localization (Hanton et al., 2005; Sharpe et al., 2010). In order to perform such predictions, a high number of experimentally localized proteins is required, but only limited data sets have been available for plants to date.In order to identify the most abundant CAZy-classified GTs as well as novel putative GTs, in this work we rigorously extended our proteomic studies of the Golgi apparatus. We have previously developed a high-throughput mass spectrometry (MS)-based quantitative proteomics technique for localization of organelle proteins by isotope tagging (LOPIT; Dunkley et al., 2004, 2006). Here, we report new LOPIT data sets and apply a new method of combining them with published LOPIT data sets, localizing an unprecedented number of plant organelle proteins. We have analyzed the TMD properties of the proteins assigned to the ER, Golgi, and plasma membrane (PM) and determined the organelle-specific features. Structural prediction analysis of the Golgi-localized proteins with unknown functions assessed the protein sequences for the potential to fold similarly to known GT structures. We found that the Golgi contains a substantial number of candidate GT families that have no characterized functions. These results yield a broader understanding of the Golgi function and its biochemical properties.     

Nesaecrepida infuscata: a biological control agent of the invasive plant Mimosa pigra

Mimosa pigra L. is a serious weed of wetlands of Australia, Asia and Africa. A suite of established biocontrol agents have been introduced in Australia and some Asian countries, but better control is needed. Nesaecrepida infuscata (Schaeffer) (Coleoptera: Chrysomelidae) is a common insect on M. pigra in tropical America. The larvae develop on the roots while the adults feed on the leaves. As both roots and leaves of M. pigra are relatively undamaged in the introduced range, this species has potential to limit the growth, survival and seed production. Furthermore, it is abundant in the dry season and so inflicts damage when most other agents are not active. In host specificity tests, larvae did not develop on any of the 65 test plant species other than M. pigra. Adult feeding on test plant species other than M. pigra was minimal. Based on these results, this insect has been released in Australia.     

Gametic, morphometric, and physiological variables influencing clutch size in the Chilean oyster, Ostrea chilensis (Philippi, 1845)

A number of marine bivalve taxa, including species of the genus Ostrea, have adopted brooding of the young in the mantle cavity as a reproductive mechanism. In spite of the importance of brooding in the reproductive success of such species, little is known about the most important variables influencing the process, including those limiting clutch size. This study addresses the regulation of brood size in the hermaphroditic oyster Ostrea chilensis. During spawning, oysters released all their oocytes into the mantle cavity. No residual oocytes remained in the gonad, so a second spawning during the same brooding season was not possible. There was a weak correlation between the number of embryos incubated during the early phase of brooding and the dry tissue weight of the brooding oyster, and between the number incubated and the shell length of the brooding adult during the later phases of brooding. The number of embryos was also correlated with the area of the labial palps of the brooder during the later stages, suggesting that the loss of veligers observed at this time may be at least partially attributable to a limitation of space around the palps, which manipulate the larvae and with which the larvae are closely associated. The oxygen consumption rate of brooders incubating a normal clutch of embryos was not significantly different from that of oysters in which clutch size had been experimentally reduced by 50%. Experimental increase of the normal clutch size by 100% significantly increased the oxygen consumption of the brooder, suggesting that there is a physiological as well as a spatial limit to brood size. Thus the number of embryos brooded by an oyster is initially dependent on its production of oocytes and secondarily by the high metabolic costs of incubating large numbers of embryos. As development proceeds, space available for brooding apparently becomes a limiting factor as the larvae grow. The fate of the excess larvae is not known at present, but any larvae released prematurely cannot be competent to settle, since development is synchronous and there is a complete release of all pediveligers at the end of the brooding period.