Development of Sensitive Methods for the Detection of Minimum Concentrations of DNA on Martian Soil Simulants.

Several methods used for the quantification of DNA are based on UV absorbance or the fluorescence of complexes with intercalator dyes. Most of these intercalators are used in gels to visualize DNA and its structural integrity. Due to many extraterrestrial samples, such as meteorites or comets, which are likely to contain very small amounts of biological material, and because the ability to detect this material is crucial for understanding the origin and evolution of life in the universe, the development of assays that can detect DNA at low limits and withstand the rigors of space exploration is a pressing need in the field of astrobiology. In this study, we present a comparison of optimized protocols used for the fast and accurate quantification of DNA using common intercalator dyes. The sensitivity of assays exceeded that generated by any commercial kit and allowed for the accurate quantification of minimum concentrations of DNA. The methods were successful when applied to the detection and measurement of DNA spiked on soil samples. Furthermore, the impact of UV radiation as a harsh condition on the surface of Mars was assessed by DNA degradation and this was also confirmed by gel electrophoresis. Overall, the methods described provide economical, simple-step, and efficient approaches for the detection of DNA and can be used in future planetary exploration missions as tests used for the extraction of nucleic acid biosignatures.

Enhancement effect of l-cysteine on lactic acid fermentation production.

BACKGROUND: Environmental stress resistance is still a bottleneck for economical process for l-lactic acid fermentation. Chronological lifespan (CLS) extension has represented a promising strategy for improving stress resistance of microbial cell factories. MAIN METHODS AND MAJOR RESULTS: In this study, addition of anti-aging drug cysteine, a kind of extending CLS of microbial cell factories, was systematically evaluated on cell viability and l-lactic acid production in Bacillus coagulans CICC 23843. The results revealed that 16 mm l-cysteine supplement significantly improved l-lactic acid titer in B. coagulans. The enhanced total antioxidant capacity (T-AOC) and key enzymes activities involving in glycolytic pathway as well as differentially expressed genes involved in cysteine synthesize and cysteine precursor synthesize pathways, and fatty acid degradation pathway may help to further understand the relative mechanism of l-cysteine effect on improving l-lactic acid accumulation. Finally, based on 16 mm l-cysteine supplement, a final l-lactic acid titer of 130.5 g L-1 with l-lactic acid productivity of 4.07 g L-1  h-1 and the conversion rate of 0.94 g g-1 total sugar was achieved in a 5 L bioreactor. CONCLUSIONS AND IMPLICATIONS: This study provided a valuable option for engineering lactic acid bacteria lifespan for enhancement of lactic acid yield.

A simple approach for the detection of Escherichia coli as a model bacterium on Martian soil simulants: A proof of concept study.

The possibility of microbial life beyond Earth presents a fundamental question in astrobiology. Given the likelihood that any extra-terrestrial life will be microbial in nature, the development of sensitive and specific confirmatory tests is crucial for the identification of potential habitats for life. Here, we describe a novel methodology for the detection of microorganisms in Martian soil simulants through spiking and recovery experiments. Our approach employs miniaturised techniques that enable the rapid and sensitive assessment of microbial presence in soil samples. The results of our study suggest that this methodology could be a valuable tool for the identification of potential habitats for microbial life on Mars and other extraterrestrial bodies.

A Simple Biochemical Method for the Detection of Proteins as Biomarkers of Life on Martian Soil Simulants and the Impact of UV Radiation.

The search for life on other planets relies on the detection of biosignatures of life. Many macromolecules have been suggested as potential targets, among which are proteins that are considered vital components of life due to their essential roles in forming cellular structures, facilitating cellular communication and signaling, and catalyzing metabolic reactions. In this context, accurate quantification of protein signatures in soil would be advantageous, and while several proposed methods exist, which are limited by their sensitivity and specificity, their applicability needs further testing and validation. To this aim, we optimized a Bradford-based assay with high sensitivity and reproducibility and a simple protocol to quantify protein extracted from a Martian soil simulant. Methods for protein spiking, extraction, and recovery were optimized, using protein standards and bacterial proteins as representative models. The proposed method achieved high sensitivity and reproducibility. Taking into account that life remains could exist on the surface of Mars, which is subjected to UV radiation, a simulation of UV exposure was performed on a spiked soil simulant. UV radiation degraded the protein spike, thus highlighting the importance of searching for the remaining signal from degraded proteins. Finally, the applicability of the method was explored in relation to the storage of the reagent which was stable even up to 12 months, thus making its application possible for future planetary exploration missions.

Microbial tolerance engineering for boosting lactic acid production from lignocellulose.

Lignocellulosic biomass is an attractive non-food feedstock for lactic acid production via microbial conversion due to its abundance and low-price, which can alleviate the conflict with food supplies. However, a variety of inhibitors derived from the biomass pretreatment processes repress microbial growth, decrease feedstock conversion efficiency and increase lactic acid production costs. Microbial tolerance engineering strategies accelerate the conversion of carbohydrates by improving microbial tolerance to toxic inhibitors using pretreated lignocellulose hydrolysate as a feedstock. This review presents the recent significant progress in microbial tolerance engineering to develop robust microbial cell factories with inhibitor tolerance and their application for cellulosic lactic acid production. Moreover, microbial tolerance engineering crosslinking other efficient breeding tools and novel approaches are also deeply discussed, aiming to providing a practical guide for economically viable production of cellulosic lactic acid.

Serum Alkaline Phosphatase Level is Associated with Angiographic Vasospasm, Delayed Cerebral Ischemia-Caused Clinical Deterioration, and Functional Outcome After Aneurysmal Subarachnoid Hemorrhage.

BACKGROUND: Alkaline phosphatase (ALP) has been implicated to be associated with poor outcome in ischemic stroke patients, yet its role in aneurysmal subarachnoid hemorrhage (aSAH) patients is unknown. The current study aimed to investigate the on-admission and short-term variation trend of ALP levels in aSAH patients as well as its associations with vasospasm, delayed cerebral ischemia (DCI), and outcome after aSAH. METHODS: Between January 2014 and May 2018, all consecutive aSAH patients were prospectively enrolled. Blood samples from patients and 78 healthy individuals were obtained. Baseline information, clinical data, and radiologic data were collected, and serum ALP levels during hospitalization were measured. Patients were followed up for 6 months. RESULTS: One hundred and ninety-six aSAH patients were included. The serum ALP levels in aSAH patients were significantly higher compared to controls (71 vs. 61 U/L, p = 0.0002), yet did not differ significantly between patients with severe (WFNS 4-5) and mild clinical condition (72 vs. 63 U/L, p = 0.3362). However, ALP was significantly higher in patients with severe radiologic status (modified Fisher 3-4) compared to those with mild radiologic status (77 vs. 61.5 U/L, p = 0.0005). A significant correlation emerged between modified Fisher score and ALP level (r = 0.246, p = 0.001). Multivariable analysis found that higher ALP level was associated with angiographic vasospasm (OR 1.019, 95% CI 1.002-1.036, p = 0.026) and DCI-caused clinical deterioration (OR 1.019, 95% CI 1.001-1.037, p = 0.037), while higher WFNS score, modified Fisher score, and ALP level were independently associated with unfavorable outcome (serum ALP level, OR 1.083, 95% CI 1.041-1.127, p < 0.001). Trend analysis of ALP level based on 103 patients' data revealed a significant decrease in ALP level on post-admission day 7-9 (median; on-admission day vs. post-admission day 7-9, 72 vs. 60 U/L, p = 0.0012; post-admission day 3-5 vs. day 7-9, 70 vs. 60 U/L, p = 0.0052) and subsequent increase in ALP level on post-admission day 12-14 (median, 84 U/L, p < 0.0001). Higher ALP levels were observed in patients with unfavorable outcome on on-admission day, post-admission day 3-5, and 12-14 (median; unfavorable vs. favorable; on-admission day, 86 vs. 67 U/L, p = 0.0122; post-admission day 3-5, 80 vs. 64 U/L, p = 0.0044; post-admission day 7-9, 75 vs. 53.5 U/L, p < 0.0001) but not on post-admission day 12-14. CONCLUSIONS: Elevated serum ALP level is associated with vasospasm, DCI-caused clinical deterioration, and functional outcome after aSAH. Further studies are required to examine the potential role of serum ALP as an outcome predictor for aSAH patients.

Loss of Shp2 within radial glia is associated with cerebral cortical dysplasia, glial defects of cerebellum and impaired sensory­motor development in newborn mice.

Radial glia are key neural progenitors involved in the development of the central nervous system. Tyrosine-protein phosphatase non­receptor type 11 (Shp2) is a widely expressed intracellular enzyme with multiple cellular functions. Previous studies have revealed the critical role of Shp2 in a variety of neural cell types; however, further investigation into the function of Shp2 within radial glia is required. In the present study, a conditional knockout mouse was generated using a human glial fibrillary acidic protein (hGFAP)­Cre driver, in which the Shp2 genes were deleted within radial glia. Loss of Shp2 within radial glia was associated with developmental retardation, postnatal lethality, reduced brain size and thinner cerebral cortices in newborn mice. Deletion of Shp2 also led to an increase in gliogenesis, a reduction in neural genesis and extracellular signal­regulated kinase signaling within the cerebral cortex. Furthermore, glial cell defects within the cerebellum of Shp2 mutants were observed, with abnormal granular cell retention and glial cell alignment in the external granular layer. In addition, Shp2 mutants exhibited impaired sensory­motor development. The results of the present study suggested that Shp2 may have an important role within radial glia, and regulate cerebral cortical and cerebellar development in newborn mice.

Macrophage-Restricted Shp2 Tyrosine Phosphatase Acts as a Rheostat for MMP12 through TGF-ß Activation in the Prevention of Age-Related Emphysema in Mice.

Persistent activation of macrophages in lungs plays a critical role in the production of matrix metalloproteinases (MMPs) that contributes to the destruction of alveolar walls, a hallmark for pulmonary emphysema. Dysregulated TGF-ß1 signaling has been an essential determinant in the elevation of MMPs during the development of emphysema. Nevertheless, the mechanism for this MMP-dependent pathogenesis has yet to be clearly investigated. Recently, we identified an important role for tyrosine phosphatase Src homology domain-containing protein tyrosine phosphatase 2 (Shp2) in regulating the activation of alveolar macrophages. Over a long-term observation period, mice with Shp2 deletion in macrophages (LysMCre:Shp2fl/fl ) develop spontaneous, progressive emphysema-like injury in the lungs, characterized by massive destruction of alveolar morphology, interstitial extracellular matrix degradation, and elevated levels of MMPs, particularly, significant increases of macrophage elastase (MMP12) in aged mice. Further analysis demonstrated that MMP12 suppression by TGF-ß1 activation was apparently abrogated in LysMCre:Shp2fl/fl mice, whereas the TGF-ß1 concentration in the lungs was relatively the same. Mechanistically, we found that loss of Shp2 resulted in attenuated SMAD2/3 phosphorylation and nuclear translocation in response to TGF-ß activation, thereby upregulating MMP12 expression in macrophages. Together, our findings define a novel physiological function of Shp2 in TGF-ß1/MMP12-dependent emphysema, adding insights into potential etiologies for this chronic lung disorder.

[Synthesis and fluorescence of KznF3 : Ce3+, Tb3+ nanoparticles].

Phosphors of KZnF3 : Ce3+, KZnF3 : Tb3+ and KZnF3 : Ce3+, Tb3+ nanoparticles were synthesized in a cetyltrimethylammonium bromide (CTAB)/2-octanol/water microemulsion system. X-ray diffraction (XRD) pattern was used to identify the formation of KZnF3 phase without detectable impurity. Environment scanning electron microscopy (ESEM) image showed that the average sizes of the KZnF3 : Ce3+ , Tb3+ nanocrystals were 30 nm in diameter. Photoluminescence characteristics of the rare earth ions doped nanoparticles were investigated and compared with that of the sample prepared by solid state reaction at a high temperature. The emission peak of the KZnF3 : Ce3+ nanoparticles showed an obvious red shift as compared to that of polycrystalline powder. In a co-doped system of KZnF3 nanoparticles, the emission band of Ce3+ even could hardly be observed and the luminescence intensity of Tb3+ was increased much compared with that Tb-doped singly. This showed that the emission of the Tb3+ was sensitized by Ce3+. The experimental results indicated that there was an effective energy transfer from Ce3+ to Tb3+ in KZnF3 nanoparticles.