Immunoanalogue of Vertebrate β-adrenergic Receptor in the Unicellular Eukaryote Paramecium

Cell fractionation, SDS-PAGE, quantitative Western blot, confocal immunolocalization and immunogold labelling were performed to find an interpretation of the physiological response of the unicellular eukaryote Paramecium to β-adrenergic ligands. The 69?kDa polypeptide separated by SDS-PAGE in S2 and P2 Paramecium subcellular fractions cross-reacted with antibody directed against human β₂-adrenergic receptor. This was detected by Western blotting followed by chemiluminescent detection. Quantitative image analysis showed that β-selective adrenergic agonist (–)-isoproterenol – previously shown to enhance phagocytic activity – evoked redistribution of the adrenergic receptor analogue from membraneous (P2) to cytosolic (S2) fraction. The relative increase in immunoreactive band intensity in S2 reached 80% and was paralleled by a 59% decrease in P2 fraction. Confocal immunofluorescence revealed β₂-adrenergic receptor sites on the cell surface and at the ridge of the cytopharynx – where nascent phagosomes are formed. This localization was confirmed by immunoelectron microscopy. These results indicate that the 69?kDa Paramecium polypeptide immunorelated to vertebrate β₂-adrenergic receptor appeared in this evolutionary ancient cell as a nutrient receptor.     

Phylogenetic affiliations of members of the heterogeneous lichen-forming fungi of the genus Lecidea sensu Zahlbruckner (Lecanoromycetes, Ascomycota)

The genus Lecidea Ach. sensu lato (sensu Zahlbruckner) includes almost 1200 species, out of which only 100 species represent Lecidea sensu stricto (sensu Hertel). The systematic position of the remaining species is mostly unsettled but anticipated to represent several unrelated lineages within Lecanoromycetes. This study attempts to elucidate the phylogenetic placement of members of this heterogeneous group of lichen-forming fungi and to improve the classification and phylogeny of Lecanoromycetes. Twenty-five taxa of Lecidea sensu lato and 22 putatively allied species were studied in a broad selection of 268 taxa, representing 48 families of Lecanoromycetes. Six loci, including four ribosomal and two protein-coding genes for 315- and 209-OTU datasets were subjected to maximum likelihood and Bayesian analyses. The resulting well supported phylogenetic relationships within Lecanoromycetes are in agreement with published phylogenies, but the addition of new taxa revealed putative rearrangements of several families (e.g. Catillariaceae, Lecanoraceae, Lecideaceae, Megalariaceae, Pilocarpaceae and Ramalinaceae). As expected, species of Lecidea sensu lato and putatively related taxa are scattered within Lecanoromycetidae and beyond, with several species nested in Lecanoraceae and Pilocarpaceae and others placed outside currently recognized families in Lecanorales and orders in Lecanoromycetidae. The phylogenetic affiliations of Schaereria and Strangospora are outside Lecanoromycetidae, probably with Ostropomycetidae. All species referred to as Lecidea sensu stricto based on morphology (including the type species, Lecidea fuscoatra [L.] Ach.) form, with Porpidia species, a monophyletic group with high posterior probability outside Lecanorales, Peltigerales and Teloschistales, in Lecanoromycetidae, supporting the recognition of order Lecideales Vain. in this subclass. The genus name Lecidea must be redefined to apply only to Lecidea sensu stricto and to include at least some members of the genus Porpidia. Based on morphological and chemical similarities, as well as the phylogenetic relationship of Lecidea pullata sister to Frutidella caesioatra, the new combination Frutidella pullata is proposed here.     

A proteomic analysis of the wound response in Medicago leaves reveals the early activation of a ROS-sensitive signal pathway

Wounded Medicago truncatula leaves produce a burst of O(2)(-) (phase I) between 1 and 15 min, then of O(2)(-) and H(2)O(2) (phase II) between 1 and 3 h. Our previous results suggest reactive oxygen species (ROS) may provide signals to mobilise early (6 h), apoplastic, wound-responsive proteins (WRPs). 2DE and MALDI-TOF/TOF were used to analyse how the suppression of ROS production at different time points by diphenyleneiodonium (DPI), affects the expression of WRPs. Rapid (≤3 min) DPI inhibition of phase I O(2)(-) production suppressed the differential regulation of 7 out of 19 WRPs, which were consequently classified as ROS-dependent WRPs. DPI inhibition of only phase II ROS production failed to suppress the wound regulation of 18 out of 19 WRPs, but led to the altered expression of 1 ROS-dependent WRP and 2 non-WRPs (Group B). The data indicates Group B proteins are alternatively targeted via the modulation of phase II ROS production. This reinforces an important role for phase I O(2)(-) signalling in the early wound response, but indicates that this response is partly regulated by phase II of the oxidative burst. This data provides an informed basis for further proteomic studies aimed at identifying early activated O(2)(-) signalling components in wounded Medicago.     

Genomic stability over 9 years of an isoniazid resistant Mycobacterium tuberculosis outbreak strain in Sweden

In molecular epidemiological studies of drug resistant Mycobacterium tuberculosis (TB) in Sweden a large outbreak of an isoniazid resistant strain was identified, involving 115 patients, mainly from the Horn of Africa. During the outbreak period, the genomic pattern of the outbreak strain has stayed virtually unchanged with regard to drug resistance, IS6110 restriction fragment length polymorphism and spoligotyping patterns. Here we present the complete genome sequence analyses of the index isolate and two isolates sampled nine years after the index case as well as experimental data on the virulence of this outbreak strain. Even though the strain has been present in the community for nine years and passaged between patients at least five times in-between the isolates, we only found four single nucleotide polymorphisms in one of the later isolates and a small (4 amino acids) deletion in the other compared to the index isolate. In contrast to many other evolutionarily successful outbreak lineages (e.g. the Beijing lineage) this outbreak strain appears to be genetically very stable yet evolutionarily successful in a low endemic country such as Sweden. These findings further illustrate that the rate of genomic variation in TB can be highly strain dependent, something that can have important implications for epidemiological studies as well as development of resistance.     

Microorganisms participating in the biodegradation of modified polyethylene films in different soils under laboratory conditions

The degree of biodegradation of low-density polyethylene (LDPE) films modified with Bionolle^® polyester in different soils under laboratory conditions was evaluated. Films were incubated in soils from waste coal, a forest and an extinct volcano crater. Prior to degradation studies, soils underwent chemical and microbiological analysis. Film weight loss and mechanical properties, as well as the surface of the polymeric samples determined via scanning electron microscopy, were evaluated after 75, 150 and 225 days of biodegradation. Important chemical changes in the polymeric chains were detected by Fourier Transform Infrared Spectroscopy (FTIR). Fungal and bacterial species that were able to grow on the film surfaces were monitored in order to see whether the films were easily colonised by autochthonous microorganisms (i.e., typical to each soil). Identification of microorganisms was based on their cellular fatty acid methyl ester (FAME) profiles. Biodegradation of modified polyethylene films in soils led to significant changes (i.e., elongation at brake of 98%) in their mechanical properties that were caused by biochemical modifications of both polyester and polyethylene. Compared to waste coal soil, films underwent rapid biodegradation in soils that were rich in organic matter. Bacteria belonging to the genus, Bacillus, and the fungi, Gliocladium viride, Aspergillus awamori and Mortierella subtilissima, were easily able to colonise both polyethylene and polyethylene modified with Bionolle^®.     

A novel mutation (c. 341A>G) in the SRY gene in a 46,XY female patient with gonadal dysgenesis

SRY (sex-determining region Y) gene, MIM 480000, NM_005634) is crucial for sex differentiation which encodes the protein responsible for initiating testis differentiation. SRY mutations are associated with the presence of XY gonadal dysgenesis symptoms.