Microbial Diversity of Bacteria Involved in Biomineralization Processes in Mine-Impacted Freshwaters

In order to increase the knowledge about geo-bio interactions in extreme metal-polluted mine waters, we combined microbiological, mineralogical, and geochemical analyses to study the indigenous sulfate-reducing bacteria (SRB) involved in the heavy metal (HM) biomineralization processes occurring in Iglesiente and Arburese districts (SW Sardinia, Italy). Anaerobic cultures from sediments of two different mining-affected streams of this regional framework were enriched and analyzed by 16S rRNA next-generation sequencing (NGS) technique, showing sequences closely related to SRB classified in taxa typical of environments with high concentrations of metals (Desulfovibrionaceae, Desulfosporosinus). Nevertheless, the most abundant genera found in our samples did not belong to the traditional SRB groups (i.e., Rahnella, Acinetobacter). The bio-precipitation process mediated by these selected cultures was assessed by anaerobic batch tests performed with polluted river water showing a dramatic (more than 97%) Zn decrease. Scanning electron microscopy (SEM) analysis revealed the occurrence of Zn sulfide with tubular morphology, suggesting a bacteria-mediated bio-precipitation. The inocula represent two distinct communities of microorganisms, each adapted to peculiar environmental conditions. However, both the communities were able to use pollutants in their metabolism and tolerating HMs by detoxification mechanisms. The Zn precipitation mediated by the different enriched cultures suggests that SRB inocula selected in this study have great potentialities for the development of biotechnological techniques to reduce contaminant dispersion and for metal recovery.

Sixteen Genome Sequences of Denitrifying Bacteria Assembled from Enriched Cultures of Anaerobic Pig Manure Digestate

We report 16 genomes assembled from the metagenome of pig manure digestate enriched with the addition of N 2 O. These denitrifying bacterial genomes all contain the nosZ gene, encoding N 2 O reductase. Their sizes range from 1,902,599 bp to 6,264,563 bp, with completeness of 75.03% to 98.89%, GC contents of 32.86% to 69.66%, and contamination of 0% to 8.4%.

Modulation of the Primary Astrocyte-Enriched Cultures’ Oxylipin Profiles Reduces Neurotoxicity

Recently, manipulations with reactive astrocytes have been viewed as a new therapeutic approach that will enable the development of treatments for acute brain injuries and neurodegenerative diseases. Astrocytes can release several substances, which may exert neurotoxic or neuroprotective effects, but the nature of these substances is still largely unknown. In the present work, we tested the hypothesis that these effects may be attributed to oxylipins, which are synthesized from n-3 or n-6 polyunsaturated fatty acids (PUFAs). We used astrocyte-enriched cultures and found that: (1) lipid fractions secreted by lipopolysaccharide (LPS)—stimulated rat primary astrocyte-enriched cultures—possessed neurotoxic activity in rat primary neuronal cultures; (2) both of the tested oxylipin synthesis inhibitors, ML355 and Zileuton, reduce the LPS-stimulated release of interleukin 6 (IL-6) by astrocyte cultures, but only ML355 can change lipid fractions from neurotoxic to non-toxic; and (3) oxylipin profiles, measured by ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) from neurotoxic and non-toxic lipid fractions, reveal a group of n-3 docosahexaenoic acid derivatives, hydroxydocosahexaenoic acids (HdoHEs)-4-HdoHE, 8-HdoHE, and 17-HdoHE, which may reflect the neuroprotective features of lipid fractions. Regulating the composition of astrocyte oxylipin profiles may be suggested as an approach for regulation of neurotoxicity in inflammatory processes.

Na+/Ca2+ exchanger isoform 1 takes part to the Ca2+-related prosurvival pathway of SOD1 in primary motor neurons exposed to beta-methylamino-L-alanine

Background: The cycad neurotoxin beta-methylamino-L-alanine (L-BMAA), causing the amyotrophic lateral sclerosis/Parkinson-dementia complex (ALS/PDC), may cause neurodegeneration by disrupting organellar Ca2+ homeostasis. By activating Akt/ERK1/2 pathway, the Cu,Zn-superoxide dismutase (SOD1) and its non-metallated form, ApoSOD1, prevent endoplasmic reticulum (ER) stress-induced cell death in motor neurons exposed to LBMAA. This occurs through the rapid increase of intracellular Ca2+ concentration ([Ca2+]i) in part flowing from the extracellular compartment and in part released from ER. However, the molecular components of this mechanism remain uncharacterized.Methods: By an integrated approach consisting on the use of siRNA strategy, Western blotting, confocal double labeling immunofluorescence, patch-clamp electrophysiology, and Fura 2- /SBFI-single-cell imaging, we explored in rat motor neuron-enriched cultures the involvement of plasma membrane Na+/Ca2+ exchanger (NCX) and the purinergic P2X7 receptor as well as of the intracellular cADP-ribose (cADPR) pathway in the rapid and neuroprotective mechanism of SOD1.Results: we showed that SOD1-induced [Ca2+]i rise was prevented by the pan inhibitor of NCX CB-DMB but not by A430879, a P2X7 receptor specific antagonist, or by 8-bromo-cADPR, a cell permeant antagonist of cADP-ribose. The same occurred for the ApoSOD1. Confocal double labeling immunofluorescence showed a huge expression of plasmalemmal NCX1 and intracellular NCX3 isoforms. Furthermore, we identified NCX1 reverse mode as the main mechanism responsible for the neuroprotective ER Ca2+ refilling elicited by SOD1 and ApoSOD1. Furthermore, SOD1 and ApoSOD1 promoted translocation of active Akt in some nuclei of primary motor neurons. Finally, the activation of NCX1 by the specific agonist CNPYB2 protected motor neurons from L-BMAA-induced cell death.Conclusion: collectively, our data indicate that SOD1 and ApoSOD1 exert their neuroprotective effect by modulating ER Ca2+ content through the activation of NCX1 reverse mode and Akt nuclear translocation in a subset of primary motor neurons.

Biocementation mediated by native microbes from Brahmaputra riverbank for mitigation of soil erodibility

Riverbank erosion is a global problem with significant socio-economic impacts. Microbially induced calcite precipitation (MICP) has recently emerged as a promising technology for improving the mechanical properties of soils. The present study investigates the potential of selectively enriched native calcifying bacterial community and its supplementation into the riverbank soil of the Brahmaputra river for reducing the erodibility of the soil. The ureolytic and calcium carbonate cementation abilities of the enriched cultures were investigated with reference to the standard calcifying culture of Sporosarcina pasteurii (ATCC 11859). 16S rRNA analysis revealed Firmicutes to be the most predominant calcifying class with Sporosarcina pasteurii and Pseudogracilibacillus auburnensis as the prevalent strains. The morphological and mineralogical characterization of carbonate crystals confirmed the calcite precipitation potential of these communities. The erodibility of soil treated with native calcifying communities was examined via needle penetration and lab-scale hydraulic flume test. We found a substantial reduction in soil erosion in the biocemented sample with a calcite content of 7.3% and needle penetration index of 16 N/mm. We report the cementation potential of biostimulated ureolytic cultures for minimum intervention to riparian biodiversity for an environmentally conscious alternative to current erosion mitigation practices.

High Glucose Shifts the Oxylipin Profiles in the Astrocytes towards Pro-Inflammatory States

Hyperglycemia is associated with several complications in the brain, which are also characterized by inflammatory conditions. Astrocytes are responsible for glucose metabolism in the brain and are also important participants of inflammatory responses. Oxylipins are lipid mediators, derived from the metabolism of polyunsaturated fatty acids (PUFAs) and are generally considered to be a link between metabolic and inflammatory processes. High glucose exposure causes astrocyte dysregulation, but its effects on the metabolism of oxylipins are relatively unknown and therefore, constituted the focus of our work. We used normal glucose (NG, 5.5 mM) vs. high glucose (HG, 25 mM) feeding media in primary rat astrocytes-enriched cultures and measured the extracellular release of oxylipins (UPLC-MS/MS) in response to lipopolysaccharide (LPS). The sensitivity of HG and NG growing astrocytes in oxylipin synthesis for various serum concentrations was also tested. Our data reveal shifts towards pro-inflammatory states in HG non-stimulated cells: an increase in the amounts of free PUFAs, including arachidonic (AA), docosahexaenoic (DHA) and eicosapentaenoic (EPA) acids, and cyclooxygenase (COX) mediated metabolites. Astrocytes cultivated in HG showed a tolerance to the LPS, and an imbalance between inflammatory cytokine (IL-6) and oxylipins release. These results suggest a regulation of COX-mediated oxylipin synthesis in astrocytes as a potential new target in treating brain impairment associated with hyperglycemia.

Detection of INVA Gene and Cytotoxin of Salmonella entertidis in Food Samples Using Molecular Methods

Salmonella entertidis is a foodborne pathogen that causes various diseases in human beings worldwide. The toxin of Salmonella can cause infectious diseases. In this research project, Salmonella was detected through various microbial, biochemical and molecular tests in diverse food samples collected from highly populated, moderately populated and less populated areas of Lahore, Pakistan. Enriched cultures of all food samples such as apples, tomatoes, yogurt and mayonnaise was streaked on violet-red bile glucose agar, Simmon’s citrate agar and eosin-methylene blue agar (EMB). Salmonella isolates were screened for the presence of toxin encoding gene through PCR. 27% apples, 19% tomatoes, 5% mayonnaise and 7% yogurt were found to be positive for INVA genes (invasion protein genes). In medical and pharmaceutical point of views the INVA gene can also help to develop specific medicines against salmonella. The cytotoxin that is protein in nature was confirmed by SDS PAGE in mayonnaise samples. This study illustrates that foods of highly populated areas are reservoir for Salmonella entertidis in Pakistan. There is need to develop specific drugs, precautionary measures to control salmonella and its disease.

Biocementation Mediated By Stimulated Ureolytic Microbes From Brahmaputra Riverbank For Mitigation of Soil Erosion

Riverbank erosion is a global problem with significant socio-economic impacts. Microbially induced carbonate precipitation (MICP) has recently emerged as a promising technology for improving the mechanical properties of soils. The presented study investigates the potential of native calcifying bacterial communities of the Brahmaputra riverbank for the first time via biostimulation and explores its effect on the mitigation of soil erosion. The ureolytic and calcium carbonate cementation ability of the enriched cultures were investigated with reference to the standard calcifying culture of Sporosarcina pasteurii (ATCC 11859). 16S rRNA analysis revealed Firmicutes to be the most predominant calcifying class with Sporosarcina pasteurii and Pseudogracilibacillus auburnensis as the prevalent strains. The morphological and mineralogical characterization of carbonate crystals confirmed the calcite precipitation potential of these communities. The erosion resistance of soil treated with native calcifying communities was examined via needle penetration and lab-scale flume erosion test. We found a substantial reduction in soil erosion in the biocemented sample with calcite content of 7.3% and needle penetration index of 16 N/mm. We report cementation potential of biostimulated ureolytic cultures for a cost-competitive and environmentally-conscious alternative to current erosion mitigation practices.