Black yeast-like fungi associated with Lethargic Crab Disease (LCD) in the mangrove-land crab, Ucides cordatus (Ocypodidae)

Lethargic Crab Disease (LCD) caused extensive epizootic mortality of the mangrove land crab Ucides cordatus (Brachyura: Ocypodidae) along the Brazilian coast, mainly in the Northeastern region. The disease was named after the symptoms of slow movement of infected crabs. Causative agents were suspected to be two black yeast-like fungi of the family Herpotrichiellaceae (ascomycete order Chaetothyriales), judged by infected tissue biopsies from moribund U. cordatus. The aim of the present study is to prove that two species are involved in the disease: the recently described black yeast Exophiala cancerae, but also a less virulent, hitherto undescribed fonsecaea-like species, introduced here as the novel species Fonsecaea brasiliensis. Strains were identified by ITS rDNA sequencing, and species borderlines were established by multilocus sequencing and AFLP analysis. Fonsecaea brasiliensis proved to be closely related to the pathogenic species Cladophialophora devriesii which originally was isolated from a systemic infection in a human patient. The virulence of F. brasiliensis is lower than that of E. cancerae, as established by artificial inoculation of mangrove crabs.     

Immunoproteomic analyses of outer membrane antigens of Actinobacillus pleuropneumoniae grown under iron-restricted conditions

Actinobacillus pleuropneumoniae, a bacterial pathogen of swine and agent of porcine pneumonia, causes a highly infectious disease of economic importance in the pig industry. Commercial vaccines for A. pleuropneumoniae include whole-cell bacterins and second generation subunit vaccines but they only confer partial protective immunity. Our search for new vaccine candidates identified antigens that are expressed during conditions that mimic infection; the outer membrane (OM) proteome of A. pleuropneumoniae serotype 5b was examined under iron restriction. Quantitative profiling by 2D-DiGE technology revealed that iron restriction induced expression of previously described transferrin binding proteins (TbpA, TbpB) plus four lipoproteins including spermidine/putrescine binding periplasmic protein 1 precursor (PotD2). Immunoproteomic analyses with antisera from na?ve animals and from infected pigs were able to differentiate antigens within the OM proteome that were specifically recognized during A. pleuropneumoniae infection. Immunoblots of iron-restricted profiles detected PotD2, heme-binding protein A (HbpA), and capsule polysaccharide export protein (CpxD) as well as surface antigens TbpA, TbpB, and OmlA. These data identify OM proteins that demonstrate immunogenicity and upregulation under conditions mimicking infection, providing emphasis on lipoproteins as an important class of antigens to exploit for vaccine development for A. pleuropneumoniae.     

Dating branch growth units in a tropical tree using morphological and anatomical markers: the case of Parkia velutina Benoist (Mimoso?deae)

Context

In tropical areas, studies based on the retrospective analysis of tree development have focused principally on growth ring research. The interpretation of primary growth markers is overlooked although it opens perspectives to provide long time-series on tree-crown development.

Aims

This study focused on Parkia velutina, an emergent tree of neotropical rain forests. Our objectives were (1) to characterize the phenological cycle of this species, and (2) to identify temporally interpretable morphological and anatomical markers.

Methods

We collected dominant branches in 14 adult trees and identified growth markers that limit longitudinal and radial increments. We coupled this approach with a 2-year phenological survey of 20 trees.

Results

Leaf shedding, growth unit elongation and growth ring formation define the phenological cycle. At tree scale, this cycle is synchronous and affects all axes. At population scale, trees can be desynchronized. This cycle is annual despite some slight variability. Successive growth units and growth rings are easily identifiable.

Conclusion

Dating a branch by counting the number of growth units or growth rings is possible in many years with a reasonable error. Nevertheless, estimating their precise month of formation in order to study climatic influences remains difficult.     

Description of a new procedure to estimate the carbon stocks of all forest pools and impact assessment of methodological choices on the estimates

Forest ecosystems play a major role in atmospheric carbon sequestration and emission. Comparable organic carbon stock estimates at temporal and spatial scales for all forest pools are needed for scientific investigations and political purposes. Therefore, we developed a new carbon stock (CS) estimation procedure that combines forest inventory and soil and litter geodatabases at a regional scale (southern Belgium). This procedure can be implemented in other regions and countries on condition that available external carbon soil and litter data can be linked to forest inventory plots. The presented procedure includes a specific CS estimation method for each of the following forest pools and subpools (in brackets): living biomass (aboveground and belowground), deadwood (dead trees and snags, coarse woody debris and stumps), litter, and soil. The total CS of the forest was estimated at 86 Tg (185 Mg ha^?1). Soil up to 0.2 m depth, living biomass, litter, and deadwood CSs account, respectively, for 48, 47, 4, and 1 % of the total CS. The analysis of the CS variation within the pools across ecoregions and forest types revealed in particular that: (1) the living biomass CS of broadleaved forests exceeds that of coniferous forests, (2) the soil and litter CSs of coniferous forest exceed those of broadleaved forests, and (3) beech stands come at the top in carbon stocking capacity. Because our estimates differ sometimes significantly from the previous studies, we compared different methods and their impacts on the estimates. We demonstrated that estimates may vary highly, from ?16 to +12 %, depending on the selected methods. Methodological choices are thus essential especially for estimating CO₂ fluxes by the stock change approach. The sources of error and the accuracy of the estimates were discussed extensively.     

Extraction of grape seed oil using compressed carbon dioxide and propane: extraction yields and characterization of free glycerol compounds

The main objective of this work was to compare the extraction of grape seed oil with compressed carbon dioxide and propane on the extraction yields and chemical characteristics of free glycerol compounds. The experiments were performed in a laboratory scale unit in the temperature range of 30 to 60 degrees C and pressures from 60 to 254 bar. The results showed that propane is a more suitable solvent for grape seed oil extraction than carbon dioxide, as higher extractions yields and a very fast kinetic of extraction were achieved with this solvent. In relation to compressed carbon dioxide extractions, both temperature and density presented a very pronounced and positive effect on the extraction yield. The oils extracted were analyzed qualitatively and quantitatively with regard to the free glycerol compounds, mainly fatty acids, ethyl, and methyl esters. The results showed that these compounds are present in low concentration in vegetable oil (<3%) and that, in general, samples extracted with propane present a smaller amount of peaks of free glycerol compounds in the oil than samples extracted with carbon dioxide.     

Worldwide genotyping of castor bean germplasm (<Emphasis Type="Italic">Ricinus communis</Emphasis> L.) using AFLPs and SSRs

Worldwide genetic diversity in 200 individuals comprising 41 castor bean accessions was assessed using amplified fragment polymorphisms (AFLPs) and simple sequence repeats (SSRs). We found that, despite surveying five continents and 35 countries, genetic diversity in castor bean germplasm is relatively low (overall H _e = 0.126 for AFLPs and 0.188 for SSRs) compared to estimates of genetic diversity in other plant species. Our data also show no geographic structuring of genotypes across continents or countries within continents. An assessment of the congruence between AFLP and SSRs indicates a low correlation (R ² = 0.19) between the two data sets, but each marker class nonetheless shows similar patterns of low-genetic diversity and a lack of geographic structure. Our data do suggest that SSRs yield a higher percentage of polymorphic loci, higher heterozyosity and a greater range of genetic distances, and are therefore more informative than are AFLPs on a locus-by-locus basis. Based on comparisons with numerous other plant species, we suggest that the lower genetic variation in this worldwide collection may be due to one or more factors including: sampling strategies that have not captured the full extent of genetic variation in the species; artifactual variation due to long-term germplasm storage and seed regeneration; or intense selection followed by domestic cultivation of a limited number of castor bean genotypes, which are widely propagated for their horticultural and agro-economic value.     

Excretion of BSE and scrapie prions in stools from murine models

Faeces from infected animals have been suggested as a potential source of contamination and transmission of prion diseases in the environment. This work describes the development of a procedure for the detection of PrP(res) in stools which is based on a detergent-based extraction and immunoprecipitation (IP). The procedure was evaluated by analyzing TSE-spiked sheep and mice faeces, and proved to be specific for PrP(res) with sensitivities of 5-10mug of infected brain tissue. In order to analyze the shedding of prions, we studied stools from orally inoculated mice over 4-days post-inoculation and also stools from terminally sick scrapie-infected mice. PrP(res) was only detected in stools shortly after the oral ingestion of TSE agents. The procedure described could be a useful tool for studying the excretion of prions and for evaluating potential environmental contamination by prions.     

Interactions of Haemophilus parasuis and its LOS with porcine brain microvascular endothelial cells

Haemophilus parasuis is a swine pathogen that causes Gl?sser's disease, which is characterized by polyserositis and meningitis. The pathogenesis of the H. parasuis infection is poorly understood. To cause meningitis, H. parasuis has to cross the blood-brain barrier (BBB) to gain access to the central nervous system (CNS). We recently showed that H. parasuis adheres to and invades porcine brain microvascular endothelial cells (PBMEC). The aim of this study was to evaluate the role of H. parasuis lipooligosaccharide (LOS) in the adhesion to PBMEC and to determine if H. parasuis (and/or its LOS) is able to induce apoptosis and activation of PBMEC. Results showed that adhesion of H. parasuis to PBMEC was partially mediated by LOS. Moreover, H. parasuis induces caspase-3-mediated apoptosis of PBMEC in a time--and dose--dependent manner, but its LOS did not seem to be involved in such a process. Furthermore, H. parasuis and, to a lesser extent, its LOS, was able to induce the release of IL-8 and IL-6 by PBMEC. Field strains of H. parasuis serotypes 4 and 5 induced similar levels of these inflammatory mediators. Our data suggest that H. parasuis uses cellular adhesion, induction of apoptosis and up-regulation of inflammatory mediators as mechanisms to invade the CNS via the BBB, and that LOS would play a certain but limited role in such pathological process.