Taxonomy and Distribution of Freshwater Pearl Mussels (Unionoida: Margaritiferidae) of the Russian Far East

The freshwater pearl mussel family Margaritiferidae includes 13 extant species, which are all listed by IUCN as endangered or vulnerable taxa. In this study, an extensive spatial sampling of Margaritifera spp. across the Russian Far East (Amur Basin, Kamchatka Peninsula, Kurile Archipelago and Sakhalin Island) was conducted for a revision of their taxonomy and distribution ranges. Based on their DNA sequences, shell and soft tissue morphology, three valid species were identified: Margaritifera dahurica (Middendorff, 1850), M. laevis (Haas, 1910) and M. middendorffi (Rosén, 1926). M. dahurica ranges across the Amur basin and some of the nearest river systems. M. laevis is distributed in Japan, Sakhalin Island and the Kurile Archipelago. M. middendorffi was previously considered an endemic species of the Kamchatka. However, it is widespread in the rivers of Kamchatka, Sakhalin Island, the Kurile Islands (across the Bussol Strait, which is the most significant biogeographical boundary within the archipelago), and, likely, in Japan. The Japanese species M. togakushiensis Kondo & Kobayashi, 2005 seems to be conspecific with M. middendorffi because of similar morphological patterns, small shell size (<100 mm long) and overlapped ranges, but it is in need of a separate revision. Phylogenetic analysis reveals that two NW Pacific margaritiferid species, M. laevis and M. middendorffi, formed a monophyletic 18S rDNA clade together with the North American species M. marrianae and M. falcata. The patterns that were found in these Margaritifera spp. are similar to those of freshwater fishes, indicating multiple colonizations of Eastern Asia by different mitochondrial lineages, including an ancient Beringian exchange between freshwater faunas across the Pacific.     

Microsporidian Parasites Found in the Hemolymph of Four Baikalian Endemic Amphipods

At present, approximately 187 genera and over 1300 species of Microsporidia have been described, among which almost half infect aquatic species and approximately 50 genera potentially infect aquatic arthropods. Lake Baikal is the deepest and one of the oldest lakes in the world, and it has a rich endemic fauna with a predominance of arthropods. Among the arthropods living in this lake, amphipods (Crustacea) are the most dominant group and are represented by more than 350 endemic species. Baikalian amphipods inhabit almost all depths and all types of substrates. The age and geographical isolation of this group creates excellent opportunities for studying the diversity, evolution and genetics of host-parasite relationships. However, despite more than 150 years of study, data investigating the microsporidia of Lake Baikal remain incomplete. In this study, we used molecular genetic analyses to detect microsporidia in the hemolymph of several endemic species of amphipods from Lake Baikal. We provide the first evidence that microsporidian species belonging to three genera (Microsporidium, Dictyocoela and Nosema) are present in the hemolymph of Baikalian endemic amphipods. In the hemolymph of Eulimnogammarus verrucosus, we detected SSU rDNA of microsporidia belonging to the genus Nozema. In the hemolymph of Pallasea cancellous, we found the DNA of Microsporidium sp. similar to that in other Baikalian endemic amphipods; Dictyocoela sp. was found in the hemolymph of Eulimnogammarus marituji and Acanthogammarus lappaceus longispinus.     

Endo-Lysosomal Dysfunction in Human Proximal Tubular Epithelial Cells Deficient for Lysosomal Cystine Transporter Cystinosin

Nephropathic cystinosis is a lysosomal storage disorder caused by mutations in the CTNS gene encoding cystine transporter cystinosin that results in accumulation of amino acid cystine in the lysosomes throughout the body and especially affects kidneys. Early manifestations of the disease include renal Fanconi syndrome, a generalized proximal tubular dysfunction. Current therapy of cystinosis is based on cystine-lowering drug cysteamine that postpones the disease progression but offers no cure for the Fanconi syndrome. We studied the mechanisms of impaired reabsorption in human proximal tubular epithelial cells (PTEC) deficient for cystinosin and investigated the endo-lysosomal compartments of cystinosin-deficient PTEC by means of light and electron microscopy. We demonstrate that cystinosin-deficient cells had abnormal shape and distribution of the endo-lysosomal compartments and impaired endocytosis, with decreased surface expression of multiligand receptors and delayed lysosomal cargo processing. Treatment with cysteamine improved surface expression and lysosomal cargo processing but did not lead to a complete restoration and had no effect on the abnormal morphology of endo-lysosomal compartments. The obtained results improve our understanding of the mechanism of proximal tubular dysfunction in cystinosis and indicate that impaired protein reabsorption can, at least partially, be explained by abnormal trafficking of endosomal vesicles.     

Surfactant Protein A in Exhaled Endogenous Particles Is Decreased in Chronic Obstructive Pulmonary Disease (COPD) Patients: A Pilot Study

Background

Exhaled, endogenous particles are formed from the epithelial lining fluid in small airways, where surfactant protein A (SP-A) plays an important role in pulmonary host defense. Based on the knowledge that chronic obstructive pulmonary disease (COPD) starts in the small airway epithelium, we hypothesized that chronic inflammation modulates peripheral exhaled particle SP-A and albumin levels. The main objective of this explorative study was to compare the SP-A and albumin contents in exhaled particles from patients with COPD and healthy subjects and to determine exhaled particle number concentrations.

Methods

Patients with stable COPD ranging from moderate to very severe (n = 13), and healthy non-smoking subjects (n = 12) were studied. Subjects performed repeated breath maneuvers allowing for airway closure and re-opening, and exhaled particles were optically counted and collected on a membrane using the novel PExA^® instrument setup. Immunoassays were used to quantify SP-A and albumin.

Results

COPD patients exhibited significantly lower SP-A mass content of the exhaled particles (2.7 vs. 3.9 weight percent, p = 0.036) and lower particle number concentration (p<0.0001) than healthy subjects. Albumin mass contents were similar for both groups.

Conclusions

Decreased levels of SP-A may lead to impaired host defense functions of surfactant in the airways, contributing to increased susceptibility to COPD exacerbations. SP-A in exhaled particles from small airways may represent a promising non-invasive biomarker of disease in COPD patients.     

Proteomic and functional analysis of the mitotic Drosophila centrosome

Regulation of centrosome structure, duplication and segregation is integrated into cellular pathways that control cell cycle progression and growth. As part of these pathways, numerous proteins with well‐established non‐centrosomal localization and function associate with the centrosome to fulfill regulatory functions. In turn, classical centrosomal components take up functional and structural roles as part of other cellular organelles and compartments. Thus, although a comprehensive inventory of centrosome components is missing, emerging evidence indicates that its molecular composition reflects the complexity of its functions. We analysed the Drosophila embryonic centrosomal proteome using immunoisolation in combination with mass spectrometry. The 251 identified components were functionally characterized by RNA interference. Among those, a core group of 11 proteins was critical for centrosome structure maintenance. Depletion of any of these proteins in Drosophila SL2 cells resulted in centrosome disintegration, revealing a molecular dependency of centrosome structure on components of the protein translation machinery, actin‐ and RNA‐binding proteins. In total, we assigned novel centrosome‐related functions to 24 proteins and confirmed 13 of these in human cells.     

Maintaining high anaerobic succinic acid productivity by product removal

During dual-phase fermentations using Escherichia coli engineered for succinic acid production, the productivity and viable cell concentration decrease as the concentration of succinic acid increases. The effects of succinic acid on the fermentation kinetics, yield, and cell viability were investigated by resuspending cells in fresh media after selected fermentation times. The cellular succinic acid productivity could be restored, but cell viability continuously decreased throughout the fermentations by up to 80% and subsequently the volumetric productivity was reduced. Omitting complex nutrients in the resuspension media had no significant effect on cellular succinate productivity and yield, although the viable cell concentration and thus the volumetric productivity was reduced by approximately 20%. By resuspending the cells, the amount of succinate produced during a 100-h fermentation was increased by more than 60%. The results demonstrate that by product removal succinic acid productivity can be maintained at high levels for extended periods of time.     

Redox potential of the Rieske iron-sulfur protein: Quantum-chemical and electrostatic study

Quantum-chemical study of structures, energies, and effective partial charge distribution for several models of the Rieske protein redox center is performed in terms of the B3LYP density functional method in combination with the broken symmetry approach using three different atomic basis sets. The structure of the redox complex optimized in vacuum differs markedly from that inside the protein. This means that the protein matrix imposes some stress on the active site resulting in distortion of its structure. The redox potentials calculated for the real active site structure are in a substantially better agreement with the experiment than those calculated for the idealized structure. This shows an important role of the active site distortion in tuning its redox potential. The reference absolute electrode potential of the standard hydrogen electrode is used that accounts for the correction caused by the water surface potential. Electrostatic calculations are performed in the framework of the polarizable solute model. Two dielectric permittivities of the protein are employed: the optical permittivity for calculation of the intraprotein electric field, and the static permittivity for calculation of the dielectric response energy. Only this approach results in a reasonable agreement of the calculated and experimental redox potentials.     

Polymer Partitioning and Ion Selectivity Suggest Asymmetrical Shape for the Membrane Pore Formed by Epsilon Toxin

Using poly-(ethylene glycol)s of different molecular weights, we probe the channels formed in planar lipid bilayers by epsilon toxin secreted by the anaerobic bacterium Clostridium perfringens. We find that the pore is highly asymmetric. The cutoff size of polymers entering the pore through its opening from the cis side, the side of toxin addition, is ∼500 Da, whereas the cutoff size for the polymers entering from the trans side is ∼2300 Da. Comparing these characteristic molecular weights with those reported earlier for OmpF porin and the α-Hemolysin channel, we estimate the radii of cis and trans openings as 0.4 nm and 1.0 nm, respectively. The simplest geometry corresponding to these findings is that of a truncated cone. The asymmetry of the pore is also confirmed by measurements of the reversal potential at oppositely directed salt gradients. The moderate anionic selectivity of the channel is salted-out more efficiently when the salt concentration is higher at the trans side of the pore.