The cell aggregating propensity of probiotic actinobacterial isolates: isolation and characterization of the aggregation inducing peptide pheromone

The auto-aggregating ability of a probiotic is a prerequisite for colonization and protection of the gastrointestinal tract, whereas co-aggregation provides a close interaction with pathogenic bacteria. Peptide pheromone mediated signaling has been studied in several systems. However, it has not yet been explored in prokaryotes, especially actinobacteria. Hence, in the present study, the diffusible aggregation promoting factor was purified from the culture supernatant of a potent actinobacterial probiont and characterized using 20 different actinobacterial cultures isolated from the gut region of chicken and goat. The results showed that the pheromone-like compound induces the aggregation propensity of treated isolates. The factor was found to be a heat stable, acidic pH resistant, low molecular weight peptide which enhances the biofilm forming ability of other actinobacterial isolates. The aggregation promoting factor represents a bacterial sex factor (pheromone) and its characterization confirms its usage in the probiotic formulation     

Fluorescence detection of intracellular cadmium with Leadmium Green

Leadmium Green is a commercially available, small molecule, fluorescent probe advertised as a detector of free intracellular cadmium (Cd²⁺) and lead (Pb²⁺). Leadmium Green has been used in various paradigms, such as tracking Cd²⁺ sequestration in plant cells, heavy metal export in protozoa, and Pb²⁺ absorption by vascular endothelial cells. However very little information is available regarding its affinity and selectivity for Cd²⁺, Pb²⁺, and other metals. We evaluated the in vitro selectivity of Leadmium Green using spectrofluorimetry. Consistent with manufacturer’s claims, Leadmium Green was sensitive to Cd²⁺ (K_D ~600 nM) and also Pb²⁺ (K_D ~9.0 nM) in a concentration-dependent manner, and furthermore proved insensitive to Ca²⁺, Co²⁺, Mn²⁺ and Ni²⁺. Leadmium Green also responded to Zn²⁺ with a K_D of ~82 nM. Using fluorescence microscopy, we evaluated Leadmium Green in live mouse hippocampal HT22 cells. We demonstrated that Leadmium Green detected ionophore-mediated acute elevations of Cd²⁺ or Zn²⁺ in a concentration-dependent manner. However, the maximum fluorescence produced by ionophore-delivered Zn²⁺ was much less than that produced by Cd²⁺. When tested in a model of oxidant-induced liberation of endogenous Zn²⁺, Leadmium Green responded weakly. We conclude that Leadmium Green is an effective probe for monitoring intracellular Cd²⁺, particularly in models where Cd²⁺ accumulates rapidly, and when concomitant fluctuations of intracellular Zn²⁺ are minimal.     

Humpback whale singing activity off the Goan coast in the Eastern Arabian Sea

For over two decades, passive acoustic monitoring (PAM) methods have been successfully employed around the world for studying aquatic megafauna. PAM-driven studies in Indian waters have so far been relatively very scarce. Furthermore, cetacean populations inhabiting the north western Indian Ocean are far less studied than those in many other regions around the world. This work likely constitutes the first systematic study of the vocal repertoire of humpback whales (Megaptera novaeangliae) at a near-shore site along the western coast of India. Analysis of the observed vocalizations provides an insight into the behaviour of the species. This is significant as it assists in developing a better understanding of the habitat use of the non-migratory Arabian Sea humpback whale population. In contrast, other breeding populations such as those around the North Atlantic, South Pacific and Australia have been relatively well studied. Underwater passive acoustic data were collected during March 2017 using an autonomous logger at a shallow-water site off the eastern edge of Grande Island off the coast of Goa. Humpback whale vocalizations were found to occur over multiple days in the recordings. Time–frequency contours of individual units of vocalization were extracted with the aid of an automatic detection technique and the characteristics of the units were measured. Further, successive units were analysed for formation of phrases and themes. Reconstruction of putative songs from the identified units and themes was not possible due to the limitations imposed by the nature of data collection. Detailed analyses of units, phrases and themes are presented.     

Optical analysis of RE3+ (RE = Nd or Er):B2O3–P2O5–CaF2–ZnO–(Li2O/Na2O/K2O) glasses

Calcium boro fluoro zinc phosphate glasses modified using alkali oxide and doped with Nd³⁺ and Er³⁺ ions with the chemical composition of 69.5 (B₂O₃) + 10 (P₂O₅) + 10 (CaF₂) + 5 (ZnO) + 5 (Na₂O/Li₂O/K₂O) + 0.5 (Er₂O₃/Nd₂O₃) were prepared using a conventional melt quenching technique. The results of X-ray diffraction patterns indicated the amorphous nature of all the prepared glasses. The visible–near-infrared red (NIR) absorption spectra of these glasses were analyzed systematically. The NIR emission spectra of Er³⁺ and Nd³⁺:calcium boro fluoro zinc phosphate glasses showed prominent emission bands at 1536 nm (⁴I_13/2→⁴I_15/2) and 1069 nm (⁴F_3/2→⁴I_11/2) respectively with λ_exci = 514.5 nm (Ar⁺ laser) as the excitation source.     

Genetic engineering of indica rice with AtDREB1A gene for enhanced abiotic stress tolerance

Plant Cell, Tissue and Organ Culture (PCTOC) - Adventitious root (AR) culturing is an effective approach for obtaining bioactive compounds from the endangered plant species of Oplopanax elatus...     

Evolutionary history of the SARS-CoV-2 Gamma variant of concern (P.1): a perfect storm

Our goal was to describe in more detail the evolutionary history of Gamma and two derived lineages (P.1.1 and P.1.2), which are part of the arms race that SARS-CoV-2 wages with its host. A total of 4,977 sequences of the Gamma strain of SARS-CoV-2 from Brazil were analyzed. We detected 194 sites under positive selection in 12 genes/ORFs: Spike, N, M, E, ORF1a, ORF1b, ORF3, ORF6, ORF7a, ORF7b, ORF8, and ORF10. Some diagnostic sites for Gamma lacked a signature of positive selection in our study, but these were not fixed, apparently escaping the action of purifying selection. Our network analyses revealed branches leading to expanding haplotypes with sites under selection only detected when P.1.1 and P.1.2 were considered. The P.1.2 exclusive haplotype H_5 originated from a non-synonymous mutational step (H3509Y) in H_1 of ORF1a. The selected allele, 3509Y, represents an adaptive novelty involving ORF1a of P.1. Finally, we discuss how phenomena such as epistasis and antagonistic pleiotropy could limit the emergence of new alleles (and combinations thereof) in SARS-COV-2 lineages, maintaining infectivity in humans, while providing rapid response capabilities to face the arms race triggered by host immuneresponses.     

Enhanced cellular uptake of a triplex-forming oligonucleotide by nanoparticle formation in the presence of polypropylenimine dendrimers

We used polypropylenimine dendrimers for delivering a 31 nt triplex-forming oligonucleotide (ODN) in breast, prostate and ovarian cancer cell lines, using ³²P-labeled ODN. Dendrimers enhanced the uptake of ODN by ~14-fold in MDA-MB-231 breast cancer cells, compared with control ODN uptake. Dendrimers exerted their effect in a concentration- and molecular weight-dependent manner, with generation 4 (G-4) dendrimer having maximum efficacy. A similar increase in ODN uptake was found with MCF-7 and SK-BR-3 (breast), LNCaP (prostate) and SK-OV-3 (ovarian) cancer cells. The dendrimers had no significant effect on cell viability at concentrations at which maximum ODN uptake occurred. [³H]Thymidine incorporation showed that complexing the ODN with G-4 significantly increased the growth-inhibitory effect of the ODN. Western blot analysis showed a significant 65% reduction of c-myc protein level in ODN–G-4 treated cells compared with that of ODN-treated/control cells. Gel electrophoretic analysis showed that ODN remained intact in cells even after 48 h of treatment. The hydrodynamic radii of nanoparticles formed from ODN in the presence of the dendrimers were in the range of 130–280 nm, as determined by dynamic laser light scattering. Taken together, our results indicate that polypropylenimine dendrimers might be useful vehicles for delivering therapeutic oligonucleotides in cancer cells.     

Aging-related attenuation of EGF receptor signaling is mediated in part by increased protein tyrosine phosphatase activity

As fibroblasts near senescence, their responsiveness to external signals diminishes. This well-documented phenomenon likely underlies physiological deterioration and limited tissue regeneration in aging individuals. Understanding the underlying molecular mechanisms would provide opportunities to ameliorate these situations. A key stimulus for human dermal fibroblasts are ligands for the epidermal growth factor receptor (EGFR). We have shown earlier that EGFR expression decreases by about half in near senescent fibroblasts (Shiraha et al., 2000, J. Biol. Chem. 275 (25), 19343-19351). However, as the cell responses are nearly absent near senescence, other aging-related signal attenuation changes must also occur. Herein, we show that EGFR signaling as determined by receptor autophosphorylation is diminished over 80%, with a corresponding decrease in the phosphorylation of the immediate postreceptor adaptor Shc. Interestingly, we found that this was due at least in part to increased dephosphorylation of EGFR. The global cell phosphotyrosine phosphatase activity increased some threefold in near senescent cells. An initial survey of EGFR-associated protein tyrosine phosphatases (PTPases) showed that SHP-1 (PTPIC, HCP, SHPTP-1) and PTPIB levels are increased in parallel in these cells. Concomitantly, we also discovered an increase in expression of receptor protein tyrosine phosphatase alpha (RPTPalpha). Last, inhibition of protein tyrosine phosphatases by sodium orthovanadate in near senescent cells resulted in increased EGFR phosphorylation. These data support a model in which, near senescence, dermal fibroblasts become resistant to EGFR-mediated stimuli by a combination of receptor downregulation and increased signal attenuation.