The salinity preference of members of the genus Macroplea (Coleoptera, Chrysomelidae, Donaciinae), fully aquatic leaf beetles that occur in brackish water

Reed beetles (Donaciinae) of the genus Macroplea Samouelle, 1819 live permanently submerged. Literature indicates that Macroplea mutica occurs in brackish water, whereas Macroplea appendiculata is restricted to freshwater. The salinity preference of these two species was tested in a linear and a circular device that offered a continuous salinity gradient. The distribution of animals in the devices was monitored over at least 3 h in each of the 21 experiments. Both species preferred freshwater (salinity 0) over brackish water (salinity 10). In particular, this holds true for specimens collected in brackish water. Likewise, immediate reactions could be observed when during such experiments the direction of the gradient was reversed. While M. mutica can be regarded as a truly marine insect, this marine environment does not strictly reflect its fundamental niche with respect to salinity preference. This is in line with accumulating evidence that M. mutica can be found in freshwater habitats (and M. appendiculata in brackish water). This indicates that the species’ distribution might be influenced by other factors like host plant preference or dispersal mechanisms. It is discussed if—in spite of similar fundamental niches—differences in salinity tolerance (and hence performance in brackish water) may have contributed to speciation in the genus Macroplea.     

High Prevalence of VanA-Type Vancomycin-Resistant Enterococci in Austrian Poultry

Fecal samples from humans and food-producing animals were analyzed for the presence of vancomycin-resistant enterococci (VRE). The VRE carriage rate in humans was 6%, and there was a predominance of VanC-type resistance. Enterococcus faecium with vanA-mediated resistance was frequent in broiler chickens (42%) but rare in cattle and pig samples.     

Validated age and growth of the leopard shark,Triakis semifasciata,with comments on reproduction

Synopsis The age, growth, and sexual maturation of the leopard shark, Triakis semifasciata, from central California were studied. Growth band counts in vertebral centra of 162 leopard sharks produced von Bertalanffy growth curves with L, K. and t_o parameters of 1536 mm. 0.082, and -2.31, respectively, for both sexes combined. The L₈ value for females (1602 mm TL) was slightly but insignificantly higher than for males (1499 mm TL), but the K and t_o values were almost identical. Seasonal changes in size modes of young-of-the-year leopard sharks, centrum edge characteristics, and growth and tetracycline mark-recapture from the field were used to validate annual deposition of vertebral centrum band pairs. Sexual maturity was evaluated by the gonads and presence of sperm and eggs; males mature at 7 yr and at about 63% of asymptotic length, and females mature at 10 yr, and at about 72% of asymptotic length. This slow growth, late maturity, and relatively low fecundity may increase their susceptibility to over-exploitation.     

Translocation of the Drosophila transient receptor potential-like (TRPL) channel requires both the N- and C-terminal regions together with sustained Ca2+ entry

In Drosophila photoreceptors the transient receptor potential-like (TRPL), but not the TRP channels undergo light-dependent translocation between the rhabdomere and cell body. Here we studied which of the TRPL channel segments are essential for translocation and why the TRP channels are required for inducing TRPL translocation. We generated transgenic flies expressing chimeric TRP and TRPL proteins that formed functional light-activated channels. Translocation was induced only in chimera containing both the N- and C-terminal segments of TRPL. Using an inactive trp mutation and overexpressing the Na(+)/Ca(2+) exchanger revealed that the essential function of the TRP channels in TRPL translocation is to enhance Ca(2+)-influx. These results indicate that motifs present at both the N and C termini as well as sustained Ca(2+) entry are required for proper channel translocation.     

Two Alternative Ways of Start Site Selection in Human Norovirus Reinitiation of Translation

The calicivirus minor capsid protein VP2 is expressed via termination/reinitiation. This process depends on an upstream sequence element denoted termination upstream ribosomal binding site (TURBS). We have shown for feline calicivirus and rabbit hemorrhagic disease virus that the TURBS contains three sequence motifs essential for reinitiation. Motif 1 is conserved among caliciviruses and is complementary to a sequence in the 18 S rRNA leading to the model that hybridization between motif 1 and 18 S rRNA tethers the post-termination ribosome to the mRNA. Motif 2 and motif 2* are proposed to establish a secondary structure positioning the ribosome relative to the start site of the terminal ORF. Here, we analyzed human norovirus (huNV) sequences for the presence and importance of these motifs. The three motifs were identified by sequence analyses in the region upstream of the VP2 start site, and we showed that these motifs are essential for reinitiation of huNV VP2 translation. More detailed analyses revealed that the site of reinitiation is not fixed to a single codon and does not need to be an AUG, even though this codon is clearly preferred. Interestingly, we were able to show that reinitiation can occur at AUG codons downstream of the canonical start/stop site in huNV and feline calicivirus but not in rabbit hemorrhagic disease virus. Although reinitiation at the original start site is independent of the Kozak context, downstream initiation exhibits requirements for start site sequence context known for linear scanning. These analyses on start codon recognition give a more detailed insight into this fascinating mechanism of gene expression.     

Correlating calcium binding, Förster resonance energy transfer, and conformational change in the biosensor TN-XXL

Genetically encoded calcium indicators have become instrumental in imaging signaling in complex tissues and neuronal circuits in vivo. Despite their importance, structure-function relationships of these sensors often remain largely uncharacterized due to their artificial and multimodular composition. Here, we describe a combination of protein engineering and kinetic, spectroscopic, and biophysical analysis of the Förster resonance energy transfer (FRET)-based calcium biosensor TN-XXL. Using fluorescence spectroscopy of engineered tyrosines, we show that two of the four calcium binding EF-hands dominate the FRET output of TN-XXL and that local conformational changes of these hands match the kinetics of FRET change. Using small-angle x-ray scattering and NMR spectroscopy, we show that TN-XXL changes from a flexible elongated to a rigid globular shape upon binding calcium, thus resulting in FRET signal output. Furthermore, we compare calcium titrations using fluorescence lifetime spectroscopy with the ratiometric approach and investigate potential non-FRET effects that may affect the fluorophores. Thus, our data characterize the biophysics of TN-XXL in detail and may form a basis for further rational engineering of FRET-based biosensors.     

Detection of p24 in HIV-1 infected cells embedded in LR White and Lowicryl K4M

Summary In this study we present a postembedding on-grid immunogold labelling procedure for the ultrastructural localization of the HIV-1 core protein p24. HIV-1 infected cells were fixed in 0.1% glutaraldehyde, incompletely dehydrated and embedded in LR White or in Lowicryl K4M. Antigenic sites were detected by incubation of ultrathin sections with primary mouse monoclonal antibody anti-HIV-1 p24, followed by the secondary antibody goat anti-mouse IgG coupled to 10nm gold particles. Antigenicity of p24 was found to withstand the applied fixation and was shown to be preserved in LR White as well as in Lowicryl. The described procedure permits the uncomplicated and easy detection of p24 in HIV-1 infected cells and tissues.     

Individual differences in migratory behavior shape population genetic structure and microhabitat choice in sympatric blackcaps (Sylvia atricapilla)

In migratory birds, traits such as orientation and distance are known to have a strong genetic background, and they often exhibit considerable within‐population variation. How this variation relates to evolutionary responses to ongoing selection is unknown because the underlying mechanisms that translate environmental changes into population genetic changes are unclear. We show that within‐population genetic structure in southern German blackcaps (Sylvia atricapilla) is related to individual differences in migratory behavior. Our 3‐year study revealed a positive correlation between individual migratory origins, denoted via isotope (δ²H) values, and genetic distances. Genetic diversity and admixture differed not only across a recently established migratory polymorphism with NW‐ and SW‐migrating birds but also across δ²H clusters within the same migratory route. Our results suggest assortment based on individual migratory origins which would facilitate evolutionary responses. We scrutinized arrival times and microhabitat choice as potential mechanisms mediating between individual variation in migratory behavior and assortment. We found significant support that microhabitat choice, rather than timing of arrival, is associated with individual variation in migratory origins. Moreover, examining genetic diversity across the migratory divide, we found migrants following the NW route to be genetically more distinct from each other compared with migrants following the traditional SW route. Our study suggests that migratory behavior shapes population genetic structure in blackcaps not only across the migratory divide but also on an individual level independent of the divide. Thus, within‐population variation in migratory behavior might play an important role in translating environmental change into genetic change.     

Myofibrillar creatine kinase activity inferred from a 3D model

Myofibrillar creatine kinase (CK) that buffers ATP during fluctuating muscle energy metabolism has been selected for studies of conformational changes underlying the cellular control of enzyme activity. The force field was computed for three energetic states, namely for the substrate-free CK molecule, for the molecule conjugated with the MgATP complex, and for the molecule conjugated with the pair of reactants MgATP-creatine. Without its substrates, the enzyme molecule assumes an inactive "open" form. Upon binding of the MgATP complex, the CK molecule takes up a reactive "closed" conformation. Subsequent binding of creatine yields a nonreactive "intermediary" conformation. Acid-base catalysis is considered to be the basic principle for the reversible transfer of the phosphoryl group between the substrates. The results indicate that the substrate-induced energy minimizing conformational changes do not represent a sufficient condition for CK activity and that some other essential component of physiological control at the cellular level is involved in the transition from the intermediary to the closed structure of the molecule.