Mercury remediation potential of mercury-resistant strain Rheinheimera metallidurans sp. nov. isolated from a municipal waste dumping site.

A novel mercury-resistant bacterium, designated strain DCL_24T, was isolated from the legacy waste at the Daddu Majra dumping site in Chandigarh, India. It showed resistance up to 300 µM of inorganic mercury (mercuric chloride). The isolate was found to be a Gram-negative, facultative anaerobic, motile, and rod-shaped bacterium that can grow at 4 - 30 °C (optimum 25 °C), pH 6.0 - 12.0 (optimum 7.0), and 0 - 4.0 % (w/v) NaCl (optimum 0.5 - 2.0 %). The 16 S rRNA gene-based phylogenetic analysis showed that DCL_ 24 T shared a 97.53 % similarity with itsºlosest type strain Rheinheimera muenzenbergensis E-49T. Insilico DNA-DNA hybridization and average nucleotide identity values were found to be 18.60 % and 73.77 %, respectively, between the genomes of DCL_24T and R. muenzenbergensis E-49T. The strain DCL_24T has 44.33 DNA G+C content (mol %). Based on the phenotypic, chemotaxonomic, and genotypic data, the strain DCL_24T represents a novel species within the genus Rheinheimera, for which the name Rheinheimera metallidurans sp. nov is proposed. The type strain is DCL_24T (MTCC13203T = NBRC115780T = JCM 35551 T). The isolate was found to volatilize and remove mercury efficiently, as demonstrated by X-ray film and dithizone-based colorimetric methods. Around 92 % of mercury removal was observed within 48 h. The mercury-resistant determinant mer operon consisting of merA, encoding the mercuric reductase enzyme, and transport and regulatory genes (merT, merP, merD, and merR) were found in the isolate. Relative expression analysis of merA at increasing concentrations of HgCl2 was confirmed by quantitative real-time PCR. These data indicate the merA-mediated reduction of toxic Hg2+ into a non-toxic volatile Hg0. The phytotoxicity assay performed using Arabidopsis thaliana seeds further demonstrated the mercury toxicity reduction potential of DCL_24T. The study shows that this novel isolate, DCL_24T, is an interesting candidate for mercury bioremediation. However, further studies are required to assess the bioremediation efficacy of the strain under the harsh environmental conditions prevailing in polluted sites.

Exploration of space to achieve scientific breakthroughs.

Living organisms adapt to changing environments using their amazing flexibility to remodel themselves by a process called evolution. Environmental stress causes selective pressure and is associated with genetic and phenotypic shifts for better modifications, maintenance, and functioning of organismal systems. The natural evolution process can be used in complement to rational strain engineering for the development of desired traits or phenotypes as well as for the production of novel biomaterials through the imposition of one or more selective pressures. Space provides a unique environment of stressors (e.g., weightlessness and high radiation) that organisms have never experienced on Earth. Cells in the outer space reorganize and develop or activate a range of molecular responses that lead to changes in cellular properties. Exposure of cells to the outer space will lead to the development of novel variants more efficiently than on Earth. For instance, natural crop varieties can be generated with higher nutrition value, yield, and improved features, such as resistance against high and low temperatures, salt stress, and microbial and pest attacks. The review summarizes the literature on the parameters of outer space that affect the growth and behavior of cells and organisms as well as complex colloidal systems. We illustrate an understanding of gravity-related basic biological mechanisms and enlighten the possibility to explore the outer space environment for application-oriented aspects. This will stimulate biological research in the pursuit of innovative approaches for the future of agriculture and health on Earth.

Transcriptome, proteome and draft genome of Euglena gracilis.

BACKGROUND: Photosynthetic euglenids are major contributors to fresh water ecosystems. Euglena gracilis in particular has noted metabolic flexibility, reflected by an ability to thrive in a range of harsh environments. E. gracilis has been a popular model organism and of considerable biotechnological interest, but the absence of a gene catalogue has hampered both basic research and translational efforts. RESULTS: We report a detailed transcriptome and partial genome for E. gracilis Z1. The nuclear genome is estimated to be around 500 Mb in size, and the transcriptome encodes over 36,000 proteins and the genome possesses less than 1% coding sequence. Annotation of coding sequences indicates a highly sophisticated endomembrane system, RNA processing mechanisms and nuclear genome contributions from several photosynthetic lineages. Multiple gene families, including likely signal transduction components, have been massively expanded. Alterations in protein abundance are controlled post-transcriptionally between light and dark conditions, surprisingly similar to trypanosomatids. CONCLUSIONS: Our data provide evidence that a range of photosynthetic eukaryotes contributed to the Euglena nuclear genome, evidence in support of the 'shopping bag' hypothesis for plastid acquisition. We also suggest that euglenids possess unique regulatory mechanisms for achieving extreme adaptability, through mechanisms of paralog expansion and gene acquisition.

Characterization and mode of action of a potent bio-preservative from food-grade Bacillus licheniformis MCC 2016.

The antibacterial peptide of Bacillus licheniformis MCC 2016 have potential biopreservative efficacy. Here, we report the purification process, properties, and mode of action of this antibacterial peptide for its potential application in the food industry. The antibacterial peptide from the cell-free supernatant was purified using a sequence of purification steps. The purified antibacterial peptide showed a specific activity of 68817 AU mg-1 and 0.4% yield. Liquid chromatography-mass spectroscopy analysis showed an mz-1 value of 279.28 for the active peptide. The SDS-PAGE analysis confirmed the antibacterial peptide is low-molecular weight and the size is between 3.0 and 3.5 kDa. Scanning electron microscopy, Fourier transform infrared spectroscopy, ß-gal induction assay and release of UV-absorbing materials indicated that the antibacterial peptide targets the cell wall of pathogens. Minimum inhibitory concentration of the antibacterial peptide against Listeria monocytogenes Scott A and others (Kocuria rhizophila ATCC 9341, Staphylococcus aureus FRI 722 and Salmonella typhimurium MTCC 1251) was found to be 1600 and 800 AU mL-1, respectively. The antibacterial peptide is temperature and pH stable, proteolytic-enzyme-sensitive, low-molecular weight, cell wall active class I bacteriocin and exhibits remarkable antibacterial activity against pathogens, suggesting its application as a potential biopreservative in the food industry.

Stable nuclear transformation of rhodophyte species Porphyridium purpureum: advanced molecular tools and an optimized method.

A mutated phytoene desaturase (pds) gene, pds-L504R, conferring resistance to the herbicide norflurazon has been reported as a dominant selectable marker for the genetic engineering of microalgae (Steinbrenner and Sandmann in Appl Environ Microbiol 72:7477-7484, 2006; Prasad et al. in Appl Microbiol Biotechnol 98(20):8629-8639, 2014). However, this mutated genomic clone harbors several introns and the entire expression cassette including its native promoter and terminator has a length > 5.6 kb, making it unsuitable as a standard selection marker. Therefore, we designed a synthetic, short pds gene (syn-pds-int) by removing introns and unwanted internal restriction sites, adding suitable restriction sites for cloning purposes, and introduced the first intron from the Chlamydomonas reinhardtii RbcS2 gene close to the 5'end without changing the amino acid sequence. The syn-pds-int gene (1872 bp) was cloned into pCAMBIA 1380 under the control of a short sequence (615 bp) of the promoter of pds (pCAMBIA 1380-syn-pds-int). This vector and the plasmid pCAMBIA1380-pds-L504R hosting the mutated genomic pds were used for transformation studies. To broaden the existing transformation portfolio, the rhodophyte Porphyridium purpureum was targeted. Agrobacterium-mediated transformation of P. purpureum with both the forms of pds gene, pds-L504R or syn-pds-int, yielded norflurazon-resistant (NR) cells. This is the first report of a successful nuclear transformation of P. purpureum. Transformation efficiency and lethal norflurazon dosage were determined to evaluate the usefulness of syn-pds-int gene and functionality of the short promoter of pds. PCR and Southern blot analysis confirmed transgene integration into the microalga. Both forms of pds gene expressed efficiently as evidenced by the stability, tolerance and the qRT-PCR analysis. The molecular toolkits and transformation method presented here could be used to genetically engineer P. purpureum for fundamental studies as well as for the production of high-value-added compounds.

Agrobacterium tumefaciens-mediated genetic transformation of haptophytes (Isochrysis species).

Isochrysis galbana and Isochrysis sp. are economically important microalgae from the division of haptophytes. Here, we report Agrobacterium-mediated stable DNA transfer into their nuclear genomes. Initial studies were performed to standardize co-cultivation media and determine the sensitivity of the microalgae to selective agents. Up to 1 mg/ml of the antibiotic hygromycin did not inhibit growth, whereas both the haptophytes bleached in artificial seawater (ASW) medium containing micromolar concentrations of the herbicide norflurazon. Co-cultivation of Isochrysis sp. and I. galbana with Agrobacterium tumefaciens strain LBA 4404 harboring the binary vector pCAMBIA 1380-pds-L504R yielded norflurazon-resistant (NR) colonies visible on selective plates after 20-30 days. pCAMBIA 1380-pds-L540R was constructed by cloning a mutated genomic phytoene desaturase (pds) gene from Haematococcus pluvialis as a selectable marker gene into the binary vector system pCAMBIA 1380. Co-cultivation of Isochrysis sp. with A. tumefaciens in ASW medium containing 200 µM of acetosyringone for 72 h produced the highest number of NR cells. For I. galbana, 100 µM of acetosyringone, ASW medium, and 48 h co-cultivation period appeared to be optimum co-cultivation parameters. The NR colonies kept their resistance phenotype for at least 24 months, even in the absence of selective pressure. The transfer of the pds gene in NR cells was shown by PCR amplification of the T-DNA sequences from the genomic DNA of NR cells and Southern blot analysis using T-DNA sequences as probes. The genetic manipulation described here will allow metabolic engineering and a better understanding of several biochemical pathways in the future.