Integrated real-time PCR for detection and monitoring of Legionella pneumophila in water systems

We evaluated a ready-to-use real-time quantitative Legionella pneumophila PCR assay system by testing 136 hot-water-system samples collected from 55 sites as well as 49 cooling tower samples collected from 20 different sites, in parallel with the standard culture method. The PCR assay was reproducible and suitable for routine quantification of L. pneumophila. An acceptable correlation between PCR and culture results was obtained for sanitary hot-water samples but not for cooling tower samples. We also monitored the same L. pneumophila-contaminated cooling tower for 13 months by analyzing 104 serial samples. The culture and PCR results were extremely variable over time, but the curves were similar. The differences between the PCR and culture results did not change over time and were not affected by regular biocide treatment. This ready-to-use PCR assay for L. pneumophila quantification could permit more timely disinfection of cooling towers.     

Carbonic anhydrase XIV in skeletal muscle: subcellular localization and function from wild-type and knockout mice

The expression of carbonic anhydrase (CA) XIV was investigated in mouse skeletal muscles. Sarcoplasmic reticulum (SR) and sarcolemmal (SL) membrane fractions were isolated from wild-type (WT) and CA XIV knockout (KO) mice. The CA XIV protein of 54 kDa was present in SR and SL membrane fractions as shown by Western blot analysis. CA activity measurements of WT and KO membrane fractions showed that CA XIV accounts for 50% and 66% of the total CA activities determined in the SR and SL fractions, respectively. This indicates the presence of at least one other membrane-associated CA isoform in these membranes, e.g., CA IV, CA IX, or CA XII. Muscle fibers of the extensor digitorum longus (EDL) muscle were immunostained with anti-CA XIV/FITC and anti-sarco(endo)plasmic reticulum Ca²⁺-ATPase 1/TRITC, with anti-CA XIV/FITC and anti-ryanodine receptor/TRITC, or with anti-CA XIV/FITC and anti-monocarboxylate transporter-4/TRITC. CA XIV was expressed in the plasma membrane and in the longitudinal SR but not in the terminal SR. Isometric contraction measurements of single twitches and tetani and a fatigue protocol applied to fiber bundles of the fast-twitch EDL and of the slow-twitch soleus muscle from WT and KO mice showed that the lack of SR membrane-associated CA XIV did not affect maximum force, rise and relaxation times, and fatigue behavior. Thus, it is concluded that a reduction of the total SR CA activity by 50% in CA XIV KO mice does not lead to an impairment of SR function. sarcoplasmic reticulum; sarcolemma; isometric contraction; Ca²⁺-ATPase; ryanodine receptor     

Obligate groundwater fauna of France: diversity patterns and conservation implications

We examined taxonomic and geographic patterns of the obligate groundwater fauna (i.e. stygobiotic fauna) by assembling in a distributional data base all species occurrences reported from France since 1805. A simulated annealing algorithm was used to identify conservation targets. Until the 60s, biological surveys were restricted to caves but the proportion of sampling sites in unconsolidated sediments increased from 1 to 16% over the last 40 years. A total of 380 species and subspecies in 40 families were collected, 70% of which being restricted to France. As observed in other temperate regions, the stygobiotic fauna was dominated by crustaceans (65% of species) and molluscs (22%). The cumulative number of species did not level off over time, clearly showing that biodiversity was underestimated. Temporal trends in the cumulative number of obligate groundwater and surface water species suggested that groundwater comprised more crustaceans than surface freshwater. Endemism was high although the geographic range size of species increased as distributional data accumulated. Of 380 species, 156 were known from a single 400-km² cell, among which 73% were located in the southern third of France. The distribution map of species richness changed dramatically over time, indicating that the location of richness hotspots was sensitive to sampling effort. Less than 2% of the French landscape was needed to capture 60% of known species. Thus, a large proportion of species could be protected by focusing habitat conservation efforts on a few complementary species-rich aquifers located in distinct regions.     

Membrane effects of 2,4-dichlorophenoxyacetic acid in motor cells of Mimosa pudica L.

2,4-dichlorophenoxyacetic acid applied to excised leaves of Mimosa pudica L. inhibited in a dose-dependent manner the shock-induced pulvinar movement. This inhibition was negatively correlated with the amount of [(14)C] 2,4-dichlorophenoxyacetic acid present in the vicinity of the motor cells. Although 2,4-dichlorophenoxyacetic acid is a weak acid, its greatest physiological efficiency was obtained with pH values close to neutrality. This observation opens the question of its mode of action which may be through external signaling or following internal transport by a specific anionic form transporter. The effect was related to molecular structure since 2,4-dichlorophenoxyacetic acid>3,4-dichlorophenoxyacetic acid>2,3-dichlorophenoxyacetic acid. An essential target of 2,4-dichlorophenoxyacetic acid action lies at the plasmalemma as indicated by the induced hyperpolarization of the cell membrane. Compared to indole-3-acetic acid and fusicoccin, it induced a complex effect on H(+) fluxes. Applied to plasma membrane vesicles purified from motor organs, 2,4-dichlorophenoxyacetic acid enhanced proton pumping, but, unlike fusicoccin, it did not increase the H(+)-ATPase catalytic activity in our experimental conditions. Taken together, the data suggest that 2,4-dichlorophenoxyacetic acid acts on cell turgor variation and the concomittant ion migration, in particular K(+), by a mechanism involving specific steps compared to indole-3-acetic acid and fusicoccin.     

Ultrastructure of cell types within the scolex and cephalic peduncle of the litobothriidean tapeworm,Litobothrium aenigmaticum

Litobothrium aenigmaticum Caira , Jensen , Waeschenbach , & Littlewood 2014 is a cestode species that parasitizes pelagic thresher sharks in Taiwan and the Gulf of California. A previous study using light microscopy suggested the scolex contains four types of unusual tissues, one of which was considered to aid in adhesion of the worm to the mucosa of the spiral intestine. The function of the other tissues was unknown. Also unknown was the function of two laterally paired ducts found extending throughout the length of the worm. The goal of the present study was to use transmission electron microscopy (TEM) to examine the internal anatomy of the scolex and cephalic peduncle of this tapeworm in more detail. Three specimens from Taiwan were examined with TEM and two with light microscopy. Transmission electron microscopy revealed that the laterally paired ducts were excretory, since they were closely associated with numerous protonephridia. The number of cell types, previously referred to as tissues, in the scolex was expanded to 11 based on a combination of features including cell size, nucleus size, ratio of the maximum diameter of the nucleus to maximum diameter of the whole cell, and types of observed organelles, as well as the presence or absence of cytoplasmic electron‐dense and electron‐lucent vesicles. The cytoplasm of all 11 cell types was found to include electron‐dense vesicles, and all types were periodic acid‐Schiff (PAS) positive. This suggests that the electron‐dense vesicles may contain glycoproteins and/or mucoproteins. As all 11 cell types occurred within the region of the scolex that is surrounded by a papilla of host mucosa at the site of tapeworm attachment, we hypothesize that one or more of these cell types may produce a substance that induces this inflammatory reaction. The specific products in the secretory vesicles and how those products are released are still unclear, as are any relationships between or among the cell types in terms of products produced. Further research will be necessary to fully understand the complexity of this organism.     

Probing whole‐stream metabolism: influence of spatial heterogeneity on rate estimates

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