Isolation and Molecular Identification of Fungi with Magnesite Enrichment Potential from KÜMAŞ Quarries in Turkey

Abstract

Magnesite is an important raw material used in various industrial applications, especially the production of high-temperature resistant materials. Due to its high reactant nature, magnesite ore is not found in pure form and it contains a great variety of pollutants such as calcium compounds, which restrict its use when exceeding 1% of the ore. Thus, the development of efficient strategies for the removal of pollutants remains a crucial step for magnesite utilization. In this regard, our present work was conducted to isolate and identify active fungal strains that remove calcium pollutants without changing the main magnesium content of the ore. For this aim, magnesite ore samples were collected from two quarries (Turanoca?? and Ortaocak) of KÜMA? Magnesite Inc. and fungal isolation studies were done by using the ore’s flora. Active isolates were chosen according to their CaCO₃ and MgCO₃ dissolving capabilities and identified by using conventional light microscopy and molecular characterization techniques. 71 fungal isolates were obtained from the isolation step and 14 of them were chosen as active isolates that solve calcium compounds while not affecting the magnesium component. The data of the microscopic examination and 18S rDNA gene sequence analysis showed that 14 active strains with magnesite enrichment potential grouped in Aspergillus alliaceus (3), Aspergillus flavus (2), Aspergillus leporis (1), Aspergillus nomius (1), Fusarium tricinctum (2), Penicillium chrysogenum (1) and Penicillium sp. (4).     

Molecular Characterization of Chitinase Genes from a Local Isolate of Serratia marcescens and Their Contribution to the Insecticidal Activity of Bacillus thuringiensis Strains

The chitinase B (chiB) and C (chiC) genes and flanking regions from a local isolate of Serratia marcescens were cloned individually and sequenced. Results showed that these chiB and chiC genes have a 96 % maximum similarity with chiB and chiC from different S. marcescens species (GenBank numbers Z36295.1 and AJ630582.1, respectively). The amplified chiB fragment, including some upstream and downstream regions, is 1,689-bp long with an open reading frame of 1,500 bp. The amplified fragment of chiC is 1,844 bp with an open reading frame of 1,443 bp. These sequences were submitted to the GenBank with accession numbers JX847796 (chiB) and JX847797 (chiC). Putative promoter regions and Shine–Dalgarno sequences were identified in both genes. The genes were cloned into a shuttle vector and the constructs were designated as pHYSB and pHYSC, respectively. Both plasmids were introduced separately into kurstaki and israelensis strains of Bacillus thuringiensis and the insecticidal activities of the engineered B. thuringiensis strains were assayed in larvae of Galleria mellonella and adult of Drosophila melanogaster. Engineered B. thuringiensis strains showed higher insecticidal activity than parental strain and the parental S. marcescens. In addition, pHYSB and pHYSC were stable over 16 daily passages under non-selective conditions in transformed B. t. israelensis 5724 strain.     

Antitumor Lipopolysaccharide from Heptoseless Mutant of Proteus mirabilis

Studies on lipopolysaccharide (LPS) from the cells of Proteus mirabilis RMS-203 were focused upon reduction of lethal toxicity and of pyrogenicity by biological and chemical modification. A heptoseless mutant, strain N-434, was isolated by the use of phage resistancy as a tool. LPS from that heptoseless mutant was completely deficient in neutral sugars and mainly composed of 2-keto-deoxy-octonic acid (KDO), glucosamine and fatty acids. It revealed almost the same antitumor activity as LPS of the wild type but it was less toxic and less pyrogenic.

Hydroxylaminolysis and reduction with LiAlH₄ resulted in removal of fatty acids from LPS accompanied with decrease in lethal toxicity and antitumor acitivity but not in pyrogenicity.

Lipid A fractions showed almost the same antitumor activity as intact LPS but less lethality and less pyrogenicity.     

Reconstructing changes in the genotype,phenotype, and climatic niche of an introduced species

An introduced species must contend with enormous environmental variation in its introduced range. In this study, we use niche models and ordination analyses to reconstruct changes in genotype, phenotype, and climatic niche of Johnsongrass Sorghum halepense, which is regarded as one of the world's most threatening invasive plants. In the United States, Johnsongrass has rapidly evolved within‐ and among‐population genetic diversity; our results show that genetic differentiation in expanding Johnsongrass populations has resulted in phenotypic variation that is consistent with habitat and climatic variation encountered during its expansion. Moreover, Johnsongrass expanded from agricultural to non‐agricultural habitat, and now, despite occupying overlapping ranges, extant agricultural and non‐agricultural populations are genetically and phenotypically distinct and manifest different plastic responses when encountering environmental variation. Non‐agricultural accessions are broadly distributed in climatic and geographic space and their fitness traits demonstrate plastic responses to common garden conditions that are consistent with local specialization. In contrast, agricultural accessions demonstrate ‘general purpose’ plastic responses and have more restricted climatic niches and geographic distributions. They also grow much larger than non‐agricultural accessions. If these differences are adaptive, our results suggest that adaptation to local habitat variation plays a crucial role in the ecology of this invader. Further, its success relates to its ability to succeed on dual fronts, by responding simultaneously to habitat and climate variability and by capitalizing on differential responses to these factors during its range expansion.     

Methylation of histone H3 lysine-79 by Dot1p plays multiple roles in the response to UV damage in Saccharomyces cerevisiae

Various proteins have been found to play roles in both the repair of UV damaged DNA and heterochromatin-mediated silencing in the yeast Saccharomyces cerevisiae. In particular, factors that are involved in the methylation of lysine-79 of histone H3 by Dot1p have been implicated in both processes, suggesting a bipartite function for this modification. We find that a dot1 null mutation and a histone H3 point mutation at lysine-79 cause increased sensitivity to UV radiation, suggesting that lysine-79 methylation is important for efficient repair of UV damage. Epistasis analysis between dot1 and various UV repair genes indicates that lysine-79 methylation plays overlapping roles within the nucleotide excision, post-replication and recombination repair pathways, as well as RAD9-mediated checkpoint function. In contrast, epistasis analysis with the H3 lysine-79 point mutation indicates that the lysine-to-glutamic acid substitution exerts specific effects within the nucleotide excision repair and post-replication repair pathways, suggesting that this allele only disrupts a subset of the functions of lysine-79 methylation. The overall results indicate the existence of distinct and separable roles of histone H3 lysine-79 methylation in the response to UV damage, potentially serving to coordinate the various repair processes.     

Heat shock prevents simulated ischemia-induced apoptosis in renal tubular cells via a PKC-dependent mechanism

Heat shock produces cellular tolerance to a variety of adverse conditions; however, the protective effect of heat shock on renal cell ischemic injury remains unclear. Protein kinase C (PKC) has been implicated in the signaling mechanisms of acute preconditioning, yet it remains unknown whether PKC mediates heat shock-induced delayed preconditioning in renal cells. To study this, renal tubular cells (LLC-PK1) were exposed to thermal stress (43 degrees C) for 1 h and heat shock protein (HSP) 72 induction was confirmed by Western blot analysis. Cells were subjected to simulated ischemia 24 h after thermal stress, and the effect of heat shock (delayed preconditioning) on ischemia-induced apoptosis (terminal deoxynucleotidyl transferase dUTP nick-end labeling) and B cell lymphoma 2 (Bcl(2)) expression (Western) was determined. Subsequently, the effect of PKC inhibition on HSP72 induction and heat stress-induced ischemic tolerance was evaluated. Thermal stress induced HSP72 production, increased Bcl(2) expression, and prevented simulated ischemia-induced renal tubular cell apoptosis. PKC inhibition abolished thermal induction of HSP72 and prevented heat stress-induced ischemic tolerance. These data demonstrate that thermal stress protects renal tubular cells from simulated ischemia-induced apoptosis through a PKC-dependent mechanism.     

Antiviral activity of an N-allyl acridone against dengue virus

Background

Dengue virus (DENV), a member of the family Flaviviridae, is at present the most widespread causative agent of a human viral disease transmitted by mosquitoes. Despite the increasing incidence of this pathogen, there are no antiviral drugs or vaccines currently available for treatment or prevention. In a previous screening assay, we identified a group of N-allyl acridones as effective virus inhibitors. Here, the antiviral activity and mode of action targeted to viral RNA replication of one of the most active DENV-2 inhibitors was further characterized.

Results

The compound 10-allyl-7-chloro-9(10H)-acridone, designated 3b, was active to inhibit the in vitro infection of Vero cells with the four DENV serotypes, with effective concentration 50% (EC₅₀) values in the range 12.5-27.1 μM, as determined by virus yield inhibition assays. The compound was also effective in human HeLa cells. No cytotoxicity was detected at 3b concentrations up to 1000 μM. Mechanistic studies demonstrated that virus entry into the host cell was not affected, whereas viral RNA synthesis was strongly inhibited, as quantified by real time RT-PCR. The addition of exogenous guanosine together with 3b rescued only partially the infectivity of DENV-2.

Conclusions

The acridone derivative 3b selectively inhibits the infection of Vero cells with the four DENV serotypes without a direct interaction with the host cell or the virion but interfering specifically with the intracellular virus multiplication. The mode of antiviral action for this acridone apparently involves the cellular enzyme inosine-monophospahe dehydrogenase together with another still unidentified target related to DENV RNA synthesis.     

Effect of extracellular enzyme activity on digestion performance of mesophilic UASB reactor treating high-strength municipal wastewater

Effect of extracellular enzyme activity on digestion performance of up-flow anaerobic sludge blanket (UASB) reactor was investigated for enhancement of anaerobic treatability of municipal wastewater. Two identical UASB reactors (9 L), namely Reactor-A (without enzyme addition) and Reactor-B (with enzyme addition), were simultaneously operated at mesophilic conditions (32 ± 2 °C) with a hydraulic retention time of 24 h. Preliminary test results showed that the highest total chemical oxygen demand (TCOD) removal were achieved with an extracellular enzyme dosage of 0.2 mL/L. In the activation period of the extracellular enzyme (on days 186–212), while Reactor-A removed up to 69.3% of TCOD and 55.9% of soluble chemical oxygen demand (SCOD), Reactor-B effectively removed up to 81.9% of TCOD and 72.2% of SCOD. The average VFA/alkalinity ratios were determined to be about 0.40 (±0.03) and 0.28 (±0.08) for Reactor-A and Reactor-B, respectively.     

Cry3Aa11: A New Cry3Aa δ-Endotoxin from a Local Isolate of <Emphasis Type="Italic">Bacillus thuringiensis</Emphasis>

A local isolate of Bacillus thuringiensis Mm2 had insecticidal activity against the larvae of Melolontha melolontha, Agelastica alni, Leptinotarsa decemlineata and Amphimallon solstitiale and produced a 65 kDa protein. SDS-PAGE profile of B. thuringiensis Mm2 was compared with those of 29 different Cry3Aa producers which verified Cry3Aa biosynthesis by the isolate. The cry3Aa gene of Mm2 was cloned, sequenced and the deduced amino acid sequence was compared with the cry3Aa sequences of ten different quaternary ranks. Its identity to these sequences ranged between 97.4% and 99.2%. The gene was next cloned into E. coli-Bacillus shuttle vector pNW33N and expressed at a low level in B. subtilis 168.