Diversity and phylogenetic profiling of niche-specific Bacilli from extreme environments of India

The diversity of culturable, aerobic and heterotrophic Bacillus and Bacillus-derived genera (BBDG) was investigated in various extreme environments (including thermal springs, cold deserts, mangroves, salt lakes, arid regions, salt pans and acidic soils) of India. Heat treatment followed by enrichment in different media led to a total of 893 bacterial isolates. Amplified ribosomal DNA restriction analysis (ARDRA) using three restriction enzymes AluI, MspI and HaeIII led to the clustering of these isolates into 12–74 groups for the different sites at 75 % similarity index, adding up to 559 groups. Phylogenetic analysis based on 16S rRNA gene sequencing led to the identification of 392 bacilli, grouped in two families, Bacillaceae (89.03 %) and Paenibacillaceae (10.97 %), and included 13 different genera with 75 distinct species. It was found that among the thirteen genera, nine (Bacillus, Halobacillus, Lysinibacillus, Oceanobacillus, Pontibacillus, Salinibacillus, Sediminibacillus, Thalassobacillus and Virgibacillus) belonged to Bacillaceae and four (Ammoniphilus, Aneurinibacillus, Brevibacillus and Paenibacillus) belonged to Paenibacillaceae. Novel isolates tolerant to low and high pH and temperature, salt and low moisture were identified. The major outcome of the present investigation was the identification of niche-specific species and also the ubiquitous presence of selected species of BBDG, which illustrate the diversity and pervasive nature of BBDG in extreme environments.     

Cell disruption methods for improving lipid extraction efficiency in unicellular microalgae

Identification of cost‐effective cell disruption methods to facilitate lipid extraction from microalgae represents a crucial step in identifying promising biofuel‐producing species. Various cell disruption methods including autoclaving, microwave, osmotic shock, and pasteurization were tested in the microalgae Chlorococcum sp. MCC30, Botryococcus sp. MCC31, Botryococcus sp. MCC32, and Chlorella sorokiniana MIC‐G5. Lipid content (on dry weight basis) from the four cultures on day 7 ranged from 11.15 to 48.33%, and on day 14 from 11.42 to 44.26%. Among the methods tested, enhanced lipid extraction was achieved through osmotic shock (15% NaCl) for Botryococcus sp. MCC32, microwave (6 min) for Botryococcus sp. MCC31, osmotic shock (5% NaCl) for Chlorella sorokiniana MIC‐G5 and microwave (2 min) for Chlorococcum sp. MCC30. The highest palmitate (16:0) contents (25.64% and 34.20%) were recorded with osmotic shock (15% NaCl) treatment for Botryococcus sp. MCC32 and microwave (6 min) for Botryococcus sp. MCC31, respectively. Two strains, along with their respective cell disruption methods, were identified as promising oil blends or nutraceuticals due to their high unsaturated fatty acid (UFA) content: Botryococcus sp. MCC31 (37.6% oleic acid content; 39.37% UFA) after autoclaving and Botryococcus sp. MCC32 after osmotic shock of 15% NaCl treatment (19.95% oleic acid content; 38.17% UFA).     

Differential activation of IFN regulatory factor (IRF)-3 and IRF-5 transcription factors during viral infection

Members of the IFN regulatory factor (IRF) family regulate gene expression critical to immune response, hemopoiesis, and proliferation. Although related by homology at their N-terminal DNA-binding domain, they display individual functional properties. The distinct properties result from differences in regulated expression, response to activating signals, and interaction with DNA regulatory elements. IRF-3 is expressed ubiquitously and is activated by serine phosphorylation in response to viral infection or TLR signaling. Evidence indicates that the kinases TANK-binding kinase 1 and inhibitor of NF-kappaB kinase-epsilon specifically phosphorylate and thereby activate IRF-3. We evaluated the contribution of another member of the IRF family, IRF-5, during viral infection since prior studies provided varied results. Analysis of phosphorylation, nuclear translocation, dimerization, binding to CREB-binding protein, recognition of DNA, and induction of gene expression were used comparatively with IRF-3 as a measure of IRF-5 activation. IRF-5 was not activated by viral infection; however, expression of TANK-binding kinase 1 or inhibitor of NF-kappaB kinase-epsilon did provide clear activation of IRF-5. IRF-5 is therefore distinct in its activation profile from IRF-3. However, similar to the biological effects of IRF-3 activation, a constitutively active mutation of IRF-5 promoted apoptosis. The apoptosis was inhibited by expression of Bcl-x(L) but not a dominant-negative mutation of the Fas-associated death domain. These studies support the distinct activation profiles of IRF-3 in comparison to IRF-5, but reveal a potential shared biological effect.     

Effect of Aluminum Phosphate on Alkaline Phosphatase Activity of Polyurethane Foam Immobilized Cyanobacteria

The impact of insoluble phosphorus such as aluminum and rock phosphate on alkaline phosphatase activity of polyurethane foam immobilized cyanobacteria was assessed. Polyurethane foam immobilized Nodularia recorded the highest alkaline phosphatase activity of 9.04 (m. mol p-nitrophenol released h^–1 mg^–1 protein) in vitro. A higher concentration of aluminum phosphate was recorded a 25% reduction in alkaline phosphatase activity, ammonia content, and available phosphorus in culture filtrate of polyurethane foam immobilized cyanobacteria. In general, immobilized cyanobacteria exhibited a higher alkaline phosphatase activity in rock phosphate than aluminum phosphate.     

Integrated genetic map and genetic analysis of a region associated with root traits on the short arm of rye chromosome 1 in bread wheat

A rye–wheat centric chromosome translocation 1RS.1BL has been widely used in wheat breeding programs around the world. Increased yield of translocation lines was probably a consequence of increased root biomass. In an effort to map loci-controlling root characteristics, homoeologous recombinants of 1RS with 1BS were used to generate a consensus genetic map comprised of 20 phenotypic and molecular markers, with an average spacing of 2.5 cM. Physically, all recombination events were located in the distal 40% of the arms. A total of 68 recombinants was used and recombination breakpoints were aligned and ordered over map intervals with all the markers, integrated together in a genetic map. This approach enabled dissection of genetic components of quantitative traits, such as root traits, present on 1S. To validate our hypothesis, phenotyping of 45-day-old wheat roots was performed in five lines including three recombinants representative of the entire short arm along with bread wheat parents ‘Pavon 76’ and Pavon 1RS.1BL. Individual root characteristics were ranked and the genotypic rank sums were subjected to Quade analysis to compare the overall rooting ability of the genotypes. It appears that the terminal 15% of the rye 1RS arm carries gene(s) for greater rooting ability in wheat.     

3D-QSAR analysis of antimalarial farnesyltransferase inhibitors based on a 2,5-diaminobenzophenone scaffold

With annual death tolls in the millions and emerging resistance to existing drugs, novel therapies are needed against malaria. Wiesner et al. recently developed a novel class of antimalarials derived from farnesyltransferase inhibitors based on a 2,5-diaminobenzophenone scaffold. The compounds displayed a wide range of activity, including submicromolar, against the multi-drug resistant Plasmodium falciparum strain Dd2. In order to investigate quantitatively the local physicochemical properties involved in the interaction between drug and biotarget, we used the 3D-QSAR methods CoMFA and CoMSIA to study some of the series, including the screened lead compound 2,5-bis-acylaminobenzophenone, 28 cinnamic acid derivatives, 29 N-(3-benzoyl-4-tolylacetylaminophenyl)-3-(5-aryl-2-furyl)acrylic acid amides, and 34 N-(4-substituted-amino-3-benzoylphenyl)-[5-(4-nitrophenyl)-2-furyl]acrylic acid amides. We found that steric, electrostatic, and hydrophobic properties of substituent groups play key roles in the bioactivity of the series of compounds, while hydrogen bonding interactions show no obvious impact. We built several highly predictive 3D-QSAR models, including a CoMSIA one composed of steric, electrostatic, and hydrophobic fields, with r(2)=0.94, q(2)=0.63, and r(pred)(2)=0.63. The results provide insight for optimization of this class of antimalarials for better activity and may prove helpful for further lead optimization.     

Developing biochemical and molecular markers for cyanobacterial inoculants

Markers for evaluating the establishment of cyanobacteria based on their sensitivity or resistance to antibiotics, saccharide utilization patterns and PCR generated fingerprints were developed. Four selected strains (isolates from rhizosphere soils of diverse agro-ecosystems) have shown potential as diazotrophs and exhibited plant growth promoting abilities. Different responses were obtained on screening against 40 antibiotics, which aided in developing selectable antibiotic markers for each strain. Biochemical profiles generated using standardized chromogenic identification system (including saccharide utilization tests) revealed that 53 % of the saccharides tested were not utilized by any strain, while some strains exhibited unique ability for utilization of saccharides such as melibiose, cellobiose, maltose and glucosamine. PCR based amplification profiles developed using a number of primers based on repeat sequences revealed the utility of 3 primers in providing unique fingerprints for the strains.     

Analysis of phenotypic and microsatellite-based diversity of maize landraces in India,especially from the North East Himalayan region

The maize landraces in the North East Himalayan (NEH) region in India, especially in the Sikkim state, are morphologically highly diverse. The present study provides details of phenotypic and molecular characterization of a set of 48 selected maize landrace accessions, including the ‘Sikkim Primitives’ which have a unique habit of prolificacy (5–9 ears on a single stalk). Multi-location phenotypic evaluation of these 48 accessions revealed significant genetic variability for grain yield and its components, leading to identification of several promising accessions. Cluster analysis and PCA using nine morpho-agronomic characters clearly separated ‘Sikkim Primitives’ from the rest of the accessions. PCA revealed two principal components describing 90% of the total variation, with hundred kernel weight, ear length, ear diameter, number of kernels per ear and flowering behaviour forming the most discriminatory traits. The accessions were genotyped using 42 microsatellite or simple sequence repeat (SSR) markers using a ‘population bulk DNA fingerprinting strategy’, with allele resolution using an automated DNA Sequencer. The study revealed a high mean number of alleles per SSR locus (13.0) and high Polymorphism Information Content (PIC) value of 0.60. The analysis also led to identification of 163 private/unique alleles, differentiating 44 out of 48 accessions. Six highly frequent SSR alleles were detected at different loci (phi014, phi062, phi090, umc1266, umc1367 and umc2250) with individual frequencies ≥0.75. Some of these SSR loci were reported to tag specific genes/QTL for some important traits, indicating that chromosomal regions harboring these SSR alleles were not selectively neutral. Cluster analysis using Rogers’ genetic distance also revealed distinct genetic identity of the ‘Sikkim Primitives’ from the rest of the accessions in India, including Sikkim. Mantel’s test revealed significant and positive correlation between the phenotypic and molecular genetic dissimilarity matrices. The study was the first to portray the patterns of phenotypic and molecular diversity in the maize landraces from the NEH region in India.