A benign approach to the preparation of freshwater bryozoan statoblasts for scanning electron microscopy (SEM) imaging

Abstract

Several different species of freshwater Bryozoa, belonging to the genera Plumatella, Rumarcanella and Fredericella, were detected within the Northern Mallee Pipeline (NMP) system in Victoria, Australia, that required definitive identification. These organisms produce asexual buds called statoblasts, with valves composed of sclerotised chitin that bear minute micro-ornamentations of considerable taxonomical significance. Imaging and analysis of these distinctive micro-ornamentations using scanning electron microscopy (SEM) is often employed for species identification. Meticulous preparation of statoblast samples is therefore required that necessitates the removal of adhering debris, dehydration and drying—whilst mitigating specimen damage and distortion. This technical note describes an approach whereby each of these three steps have been individually designed to be as benign as possible, using mild detergent/sonication to remove debris, a gradual and gentle dehydration procedure using ethanol, and critical point drying. For the overall process, these methods are chosen to optimise control and to minimise the use of harsh and hazardous chemicals.     

How do salinity and water stress affect transport of water,assimilates and ions to tomato fruits?

The study was conducted in order to determine whether water stress affects the accumulation of dry matter in tomato fruits similarly to salinity, and whether the increase in fruit dry matter content is solely a result of the decrease in water content. Although the rate of water transport to tomato fruits decreased throughout the entire season in saline water irrigated plants, accumulation rates of dry matter increased significantly. Phloem water transport contributed 80–85% of the total water transport in the control and water-stressed plants, and over 90% under salinity. The concentration of organic compounds in the phloem sap was increased by 40% by salinity. The rate of ions transported via the xylem was also significantly increased by salinity, but their contribution to fruit osmotic adjustment was less. The rate of fruit transpiration was also markedly reduced by salinity. Water stress also decreased the rate of water transport to the tomato fruit and increased the rate of dry matter accumulation, but much less than salinity. The similar changes, 10–15%, indicate that the rise in dry matter accumulation was a result of the decrease in water transport. Other parameters such as fruit transpiration rates, phloem and xylem sap concentration, relative transport via phloem and xylem, solutes contributing to osmotic adjustment of fruits and leaves, were only slightly affected by water stress. The smaller response of these parameters to water stress as compared to salinity could not be attributed to milder stress intensity, as leaf water potential was found to be more negative. Measuring fruit growth of girdled trusses, in which phloem flow was inactive, and comparing it with ungirdled trusses validated the mechanistic model. The relative transport of girdled as compared to ungirdled fruits resembled the calculated values of xylem transport.     

Flexibility in crosstalk between H2B ubiquitination and H3 methylation in vivo

Histone H2B ubiquitination is a dynamic modification that promotes methylation of histone H3K79 and H3K4. This crosstalk is important for the DNA damage response and has been implicated in cancer. Here, we show that in engineered yeast strains, ubiquitins tethered to every nucleosome promote H3K79 and H3K4 methylation from a proximal as well as a more distal site, but only if in a correct orientation. This plasticity indicates that the exact location of the attachment site, the native ubiquitin-lysine linkage and ubiquitination cycles are not critical for trans-histone crosstalk in vivo. The flexibility in crosstalk also indicates that other ubiquitination events may promote H3 methylation.     

Immunoprophylaxis of respiratory syncytial virus: global experience

In sarcoidosis, host genetic factors are discussed as contributing to disease susceptibility and course. Since tumor necrosis factor (TNF)-α is a central mediator of granuloma formation and since elevated TNF-α levels are found during active phases of sarcoidosis, genetic polymorphisms correlating with influences on TNF-α levels are of special interest. The complete sequencing of the MHC region and the increase in the number of identified gene polymorphisms in this locus associated with TNF-α production offer the opportunity of detecting new genes associated with sarcoidosis and perhaps of defining disease-associated haplotypes that bear the potential of serving as predictive markers for this disease.     

Clustering of nuclei in multinucleated hyphae is prevented by dynein-driven bidirectional nuclear movements and microtubule growth control in Ashbya gossypii

During filamentous fungus development, multinucleated hyphae employ a system for long-range nuclear migration to maintain an equal nuclear density. A decade ago the microtubule motor dynein was shown to play a central role in this process. Previous studies with Ashbya gossypii revealed extensive bidirectional movements and bypassings of nuclei, an autonomous cytoplasmic microtubule (cMT) cytoskeleton emanating from each nucleus, and pulling of nuclei by sliding of cMTs along the cortex. Here, we show that dynein is the sole motor for bidirectional movements and bypassing because these movements are concomitantly decreased in mutants carrying truncations of the dynein heavy-chain DYN1 promoter. The dynactin component Jnm1, the accessory proteins Dyn2 and Ndl1, and the potential dynein cortical anchor Num1 are also involved in the dynamic distribution of nuclei. In their absence, nuclei aggregate to different degrees, whereby the mutants with dense nuclear clusters grow extremely long cMTs. As in budding yeast, we found that dynein is delivered to cMT plus ends, and its activity or processivity is probably controlled by dynactin and Num1. Together with its role in powering nuclear movements, we propose that dynein also plays (directly or indirectly) a role in the control of cMT length. Those combined dynein actions prevent nuclear clustering in A. gossypii and thus reveal a novel cellular role for dynein.     

Rotavirus and adenovirus in Rondônia

Acute gastroenteritis is one of the most common diseases in humans worldwide. Viral gastroenteritis is a global problem in infants and young children. In this study the incidence of diarrhea was assessed in 877 hospitalized children under five years old, over a period of 24 months and distributed in 470 cases of diarrhea and 407 age-matched group with other pathologies, as control group. Two antigen detection techniques based on enzyme immunoassay (EIA) and latex particles were used for detection of rotavirus and adenovirus. Rotavirus A was a major cause of gastroenteritis with 23.6% of cases, being 90% of these cases in young children. Adenovirus infections was detected by EIA with frequency of 6.4%. Rotavirus and adenovirus were detected in 10.1 and 1.7% of stools from control group, respectively. Interestingly, the frequency of the youngest children in the control group excreting Rotavirus A was comparable to that detected in stools from diarrheic children. We cannot rule out the existence of other enteric viruses because the etiology of 171 cases of diarrhea was not determined and active search for astrovirus and calicivirus was not done. This is the first study that shows the presence of enteric viruses in the infantile population from Western Brazilian Amazonia and it was important to help physicians in the treatment of viral gastroenteritis.     

Hydraulic resistance components of mature apple trees on rootstocks of different vigours

Dwarfing of fruit trees is often achieved through the use of dwarfing rootstocks. Dwarf trees are characterized by sustained reductions in vegetative growth during the lifetime of the tree. The dwarfing mechanism is not well understood, but it has been hypothesized that hydraulic properties of the rootstock and the graft union are involved. It is hypothesized here that leaf- or stem-specific resistance of at least one hydraulic component of the water transport system would be negatively correlated with rootstock 'vigour', and this could be useful for selection of rootstocks. Hydraulic resistance (R) of fully grown apple trees on a variety of rootstocks of different 'vigours' was measured. Most measurements were with the evaporative flux (EF) method, where water uptake measured with sap flow sensors was related to the pressure gradient from soil (taken as pre-dawn leaf) and midday root (taken as covered root-sucker), stem (from covered leaf), and exposed and shaded leaf water potentials (Psi(l)). R of trees on dwarfing M9 rootstock was compared with that of more vigorous MM106 and MM111 rootstocks in Israel and Vermont, USA. In Israel, M9 consistently had higher leaf-specific hydraulic resistance (R(l)) in the soil to scion stem pathway, but this difference was only significant for one summer. R was larger in M9 between the root and stem, implicating the graft union as the site of increased resistance. In Vermont, R(l) of 9- and 10-year-old trees on six rootstocks of various vigours was not consistently related to vigour, and stem-specific resistance (R(s)) increased with increasing vigour. High pressure flow meter (HPFM) measurements gave a lower R than the EF method in all but one case, perhaps indicating a significant amount of xylem dysfunction in these trees, and demonstrated the increased resistivity of stem sections that included dwarf graft unions as compared with non-graft stem sections. It is concluded that stem- and leaf-specific R are not consistently positively correlated with dwarfing, although the increased resistivity of the graft union in dwarfing rootstocks may influence the transport of water and other elements across the graft union, and therefore be involved in the dwarfing mechanism.     

Asymmetric loading of Kar9 onto spindle poles and microtubules ensures proper spindle alignment

Spindle alignment is the process in which the two spindle poles are directed toward preselected and opposite cell ends. In budding yeast, the APC-related molecule Kar9 is required for proper alignment of the spindle with the mother-bud axis. We find that Kar9 localizes to the prospective daughter cell spindle pole. Kar9 is transferred from the pole to cytoplasmic microtubules, which are then guided in a myosin-dependent manner to the bud. Clb4/Cdc28 kinase phosphorylates Kar9 and accumulates on the pole destined to the mother cell. Mutations that block phosphorylation at Cdc28 consensus sites result in localization of Kar9 to both poles and target them both to the bud. Thus, Clb4/Cdc28 prevents Kar9 loading on the mother bound pole. In turn, asymmetric distribution of Kar9 ensures that only one pole orients toward the bud. Our results indicate that Cdk1-dependent spindle asymmetry ensures proper alignment of the mitotic spindle with the cell division axis.     

The AtRbx1 protein is part of plant SCF complexes,and its down-regulation causes severe growth and developmental defects

Recently in yeast and animal cells, one particular class of ubiquitin ligase (E3), called the SCF, was demonstrated to regulate diverse processes including cell cycle and development. In plants SCF-dependent proteolysis is also involved in different developmental and hormonal regulations. To further investigate the function of SCF, we characterized at the molecular level the Arabidopsis RING-H2 finger protein AtRbx1. We demonstrated that the plant gene is able to functionally complement a yeast knockout mutant strain and showed that AtRbx1 protein interacts physically with at least two members of the Arabidopsis cullin family (AtCul1 and AtCul4). AtRbx1 also associates with AtCul1 and the Arabidopsis SKP1-related proteins in planta, indicating that it is part of plant SCF complexes. AtRbx1 mRNAs accumulate in various tissues of the plant, but at higher levels in tissues containing actively dividing cells. Finally to study the function of the gene in planta, we either overexpressed AtRbx1 or reduced its expression by a dsRNA strategy. Down-regulation of AtRbx1 impaired seedling growth and development, indicating that the gene is essential in plants. Furthermore, the AtRbx1-silenced plants showed a reduced level of AtCul1 protein, but accumulated higher level of cyclin D3.