Genetic Manipulations in Dermatophytes

Dermatophytes are a group of closely related fungi that nourish on keratinized materials for their survival. They infect stratum corneum, nails, and hair of human and animals, accounting the largest portion of fungi causing superficial mycoses. Huge populations are suffering from dermatophytoses, though the biology of these fungi is largely unknown yet. Reasons are partially attributed to the poor amenability of dermatophytes to genetic manipulation. However, advancements in this field over the last decade made it possible to conduct genetic studies to satisfying extents. These included genetic transformation methods, indispensable molecular tools, i.e., dominant selectable markers, inducible promoter, and marker recycling system, along with improving homologous recombination frequency and gene silencing. Furthermore, annotated genome sequences of several dermatophytic species have recently been available, ensuring an optimal recruitment of the molecular tools to expand our knowledge on these fungi. In conclusion, the establishment of basic molecular tools and the availability of genomic data will open a new era that might change our understanding on the biology and pathogenicity of this fungal group.     

A first insight into the intestinal microbiota of snow trout (<Emphasis Type="Italic">Schizothorax zarudnyi</Emphasis>)

This study addresses the biodiversity profile of bacterial community in the intestinal lumen and mucosa of snow trout fish by applying 16S rRNA gene 454-pyrosequencing. A total of 209,106 sequences with average length 689 (±53) were filtered, denoised, trimmed, and then sorted into OTUs based on 97 % sequence similarity using the USEARCH software pipeline. Bacteria representing 10 phyla were found in the samples investigated. Fimicutes ribotypes were present in intestinal-mucosa and lumen in all fish and often dominated the libraries (average 43 and 38 %, respectively). Proteobacteria were also prevalent, but at a lower relative abundance, at 22 and 29 % in mucosa and lumen, respectively. The autochthonous microbiota was dominated by sequences belonging to the Bacilli (mean sequence abundance 24 %), in particular the Lactobacillaceae, with Lactobacillus and Pediococcous being the most abundant genera. Fewer Bacilli (mean sequence abundance 22 %) and Actinobacteria (2 %) were present in the lumen, and allochthonous communities consisted of a more even split among the bacterial classes, with increases in sequences assigned to members of the γ-Proteobacteria (16 %) and Fusobacteriia (8 %). The principal bacterial genera recorded in the lumen belonged to the lactic acid bacteria group, Cetobacterium, Clostridium and Synechococcus. Results obtained suggest that the lumen and mucosal layer of the snow trout intestine may host different microbial communities. Moreover, both regions harbour a diverse microbiome with a greater microbial diversity in the intestinal mucus compared with the luminal communities of the fish. Many of these microbes might be of high physiological relevance for the fish and may play key roles in the functioning of its gut.     

Solution structure of monomeric and trimeric photosystem I of <Emphasis Type="Italic">Thermosynechococcus elongatus</Emphasis> investigated by small-angle X-ray scattering

The structure of monomeric and trimeric photosystem I (PS I) of Thermosynechococcus elongatus BP1 (T. elongatus) was investigated by small-angle X-ray scattering (SAXS). The scattering data reveal that the protein–detergent complexes possess radii of gyration of 58 and 78 Å in the cases of monomeric and trimeric PS I, respectively. The results also show that the samples are monodisperse, virtually free of aggregation, and contain empty detergent micelles. The shape of the protein–detergent complexes can be well approximated by elliptical cylinders with a height of 78 Å. Monomeric PS I in buffer solution exhibits minor and major radii of the elliptical cylinder of about 50 and 85 Å, respectively. In the case of trimeric PS I, both radii are equal to about 110 Å. The latter model can be shown to accommodate three elliptical cylinders equal to those describing monomeric PS I. A structure reconstitution also reveals that the protein–detergent complexes are larger than their respective crystal structures. The reconstituted structures are larger by about 20 Å mainly in the region of the hydrophobic surfaces of the monomeric and trimeric PS I complexes. This seeming contradiction can be resolved by the addition of a detergent belt constituted by a monolayer of dodecyl-β-D-maltoside molecules. Assuming a closest possible packing, a number of roughly 1024 and 1472 detergent molecules can be determined for monomeric and trimeric PS I, respectively. Taking the monolayer of detergent molecules into account, the solution structure can be almost perfectly modeled by the crystal structures of monomeric and trimeric PS I.     

Molecular Design,Expression and Evaluation of PASylated Human Recombinant Erythropoietin with Enhanced Functional Properties

Erythropoietin (EPO) is the principal hormone which, has somewhat short half-life involved in the differentiation and regulation of circulating red blood cells. The present study was carried out to evaluate the capability of a polyethylene glycol mimetic technology as a biological alternative to improve pharmaceutical properties of human recombinant EPO. In silico models of EPO fused to 200 amino acids of proline, alanine, and serine (PAS) were initially generated and assessed by molecular dynamic (MD) simulation. The fluctuations of the modeled structure reached a plateau after 6000 ps of MD simulation. The Phi and psi analysis showed >99.2% of residues were located in the allowed regions. An expression vector consisting of EPO cDNA tagged to PAS coding sequences was synthesized and expressed in CHO-K1 Cells. The produced PASylated molecule was purified and characterized by standard analytical methods. The molecular weight of fusion protein was expanded to 70 kDa using sodium dodecyl sulfate polyacrylamide gel electrophoresis method. Analytical size exclusion chromatography revealed an approximately sevenfold increase in apparent size of produced protein. Although the in vitro potency of the fusion protein was significantly reduced (1.26?±?0.05 vs. 0.24?±?0.03 ng/ml) but, the in vivo activity was considerably increased up to 1.58?×?10⁵ IU/ml in normocythemic mice assay. Pharmacokinetic animal studies revealed strongly 15.6-fold plasma half-life extension for the PASylated EPO (83.16?±?13.28 h) in comparison to epoetin α (8.5?±?2.4 h) and darbepoetin α (25.3?±?2.2h).     

Multigene phylogeny reveals a new species and novel records and hosts in the genus <Emphasis Type="Italic">Ramularia</Emphasis> from Iran

Ramularia is a species-rich genus in the order Capnodiales (Dothideomycetes, Ascomycota) that includes numerous phytopathogenic taxa, several of which are economically important plant pathogens. In this study, six isolates of Ramularia were recovered from leaf spot symptoms on six herbaceous and woody plants from Guilan, East and West Azarbaijan provinces in the north and northwest of Iran. The isolates were studied by a polyphasic approach involving morphological and cultural data, and multi-gene phylogeny (ITS, TEF1-α, ACT, HIS, RPB2 and GAPDH). The isolates were grouped in three species clades of the R. eucalypti species complex. Of these, R. mali is recorded for the first time in Asia and R. glennii represents a new record for the mycobiota of Iran. Ramularia taleshina on Alnus subcordata is described as a new species. Ramularia taleshina is phylogenetically related to R. mali, but they can be differentiated by morphological and cultural characters as well as molecular data. Acalypha australis, Ficus carica and Platanus sp. are reported as new hosts of R. glennii, and Prunus cerasus and Vitis vinifera as new hosts of R. mali.     

Aluminum-mediated metabolic changes in rat serum and urine: a proton nuclear magnetic resonance study

The toxic effects of Al(3+) have been studied in 90-days AlCl(3) orally treated male albino rats (n = 7) using (1)H NMR spectroscopy-based metabolic profile of rat serum and urine, serum enzyme tests, behavioral impairment, and histopathology of kidney and liver. Metabolic profile of 90-days Al(3+)-treated rat sera showed significantly elevated levels of alanine, glutamine, beta-hydroxy-butyrate, and acetoacetate and significantly decreased level of acetone when compared with that of control rats. However, metabolic profile of 90-days Al(3+)-treated rat urine showed significantly decreased levels of citrate, creatinine, allantoin, trans-aconitate, and succinate and significantly increased level of acetate when compared to control rats. The overall perturbations observed in the metabolic profile of serum and urine demonstrate the impairment in the tricarboxylic acid cycle, liver and kidney metabolism, which was further reinstated by clinical chemistry and histopathological observations. Moreover, "in vivo" behavioral impairment has also been observed as the indication of aluminum neurotoxicity.     

Calcium salicylate-mediated apoptosis in human HT-1080 fibrosarcoma cells

Salicylates are novel biologically active compounds that exhibit multiple therapeutic activities. The anti-cancer effectiveness of calcium salicylate has been investigated on human HT-1080 fibrosarcoma cell lines at relatively low concentrations (predominantly 0.4 mM) compared to those previously reported. Although low calcium salicylate concentrations did not retard tumour growth progression significantly, as measured by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) and time-lapse assays, its cytotoxic characteristics were proven to be prominent by various morphological and immunocytological techniques. The results here demonstrate evidence for approximately 25% apoptosis after treatment with calcium salicylate, which up-regulatd the expression of p53, p21 and Bax, and down-regulated Bcl-2 in HT-1080 cells.