Nature of stress: differential effects on brain acetylcholinesterase activity and memory in rats

Effect of acute, chronic-predictable and chronic-unpredictable stress on memory and acetylcholinesterase (AChE) was investigated in rats. The animals were subjected to 3 type of stressors--(1) acute immobilization stress, (2) chronic-predictable stress i.e., immobilization daily for 5 consecutive days and (3) chronic-unpredictable stress that included reversal of light/dark cycle, over-night fasting, forced-swimming, immobilization and forced exercise in random unpredictable manner daily for 5 consecutive days. Learning and memory function was studied by single trial Passive avoidance test. AChE activity was assayed spectrophotometrically in the detergent (DS) and salt (SS) soluble fractions in different brain regions. Learning was obtained in acute and chronic-predictable stress groups but not in chronic-unpredictable group. Acute, chronic-predictable and chronic-unpredictable stress caused significant decrease in AChE activity in the DS fraction of cortex, hippocampus and hypothalamus as compared to control. Results indicate that AChE in DS fraction is predominantly affected in stressed and stressed-trained group but cognition is affected only by chronic-unpredictable stress. In acute and chronic-predictable groups the decreased AChE activity in the hippocampal DS fraction during learning may be responsible to maintain cognitive function by enhancing the cholinergic activity.     

Expressing multiple genes in a single open reading frame with the 2A region of foot-and-mouth disease virus as a linker

The Food-and-mouth disease virus (FMDV) 2A proteinis only 16–20 amino acid long. It is responsible for thecleavage of the FMDV polyprotein at its own carboxyl-terminus. Weused the cleavage property of the 2A protein to processartificial polyproteins produced in transgenic plants. In our system, single or multiplecopies of the reporter CAT and GUS genes were fused into a single open readingframe (ORF) with a copy of the FMDV 2A protein gene placed between the reportergenes. Expression of various constructs in transgenic tobacco resulted inconsistent detection of freed CAT and/or GUS proteins, suggesting that FMDV 2Aprotein functioned properly in plant cells. Cleavage efficiencyranged from 80% to 100% depending on the constructs. The variability incleavage efficiency suggested that the contexts flanking a 2Aprotein might modulate its activity. We further expressed constructs wheremultiple copies of the 2A and reporter genes were fused into one ORF. Thepresence of freed GUS protein together with partially processed polyproteinintermediates in the transgenic plants indicated that multiple copies of the 2Aprotein in a single ORF function independently. Our data demonstrate that usingthe FMDV 2A protease as a linker, multiple genes could be easily expressed in asingle ORF.     

Effects of physical, ionic, and structural factors on the binding of repressor of mycobacteriophage L1 to its cognate operator DNA

To determine the factors influencing the binding of L1 repressor to its cognate operator DNA, several gel shift as well as bioinformatic analyses have been carried out. The data show that time, temperature, salt, and pH each greatly affect the binding. In order to achieve optimum operator binding of L1 repressor in Tris buffer, the minimum requirements of time, temperature, salt, and pH were estimated to be 1 min, 32 degrees C, NaCl (50 mM), and 7.9, respectively. Interestingly Na+ but not NH4+, K+, or Li+ was found to augment significantly the binding activity of CI protein above the basal level. Anions like Cl-, citrate-, acetate-, and H2PO4- do not alter the binding of L1 repressor to its operator. We also show that an in frame deletion mutant of L1 repressor which does not carry the putative HTH motif (at its N-terminal end) fails to bind to its cognate operator DNA even at very high concentrations. The putative HTH motif was found highly conserved and evolutionarily very close to that of regulatory proteins of Y. pestis, H. marismortui, A. tumefaciens, etc. Taken together we suggest that N-terminal end of L1 repressor carries a HTH motif. Further analysis of the putative secondary structures of mycobacteriophage repressors reveals that two common regions encompassing more than 90% of primary sequence are present in all the four repressor molecules studied here. The results suggest that these common regions are utilized for carrying out identical functions.     

Gene-centric metegenome analysis reveals diversity of Pseudomonas aeruginosa biofilm gene orthologs in fresh water ecosystem

Metagenomic analysis of biofilm forming bacteria in environmental samples remains challenging due to the non-availability of gene sequences of most of the uncultivable bacteria. Sequences of Pseudomonas aeruginosa PAO1-UW genes involved either directly or indirectly in biofilm formation were analyzed using BLASTn to obtain matching sequences from different strain, species and genus. Conserved regions in the functional domain of the amino acid sequences were used to design common primers for direct PCR analysis of freshwater metagenomes. Seven key genes such as aceA, clpP, typA, cbrA, phoR, rpoS and gacA involved in biofilm formation were validated. The ortholog genes belonged to wide range of Pseudomonas sp. indicating the diversity of biofilm genes and the conservation of protein functional domains. The approach would also help in analyzing the expression of biofilm genes in different bacteria of freshwater systems for monitoring toxic contaminations such as organic or inorganic pollutants.     

Distribution-free simultaneous tests for location–scale and Lehmann alternative in two-sample problem

The paper deals with the classical two-sample testing problem for the equality of two populations, one of the most fundamental problems in biomedical experiments and case–control studies. The most familiar alternatives are the difference in location parameters or the difference in scale parameters or in both the parameters of the population density. All the tests designed for classical location or scale or location–scale alternatives assume that there is no change in the shape of the distribution. Some authors also consider the Lehmann-type alternative that addresses the change in shape. Two-sample tests under Lehmann alternative assume that the location and scale parameters are invariant. In real life, when a shift in the distribution occurs, one or more of the location, scale, and shape parameters may change simultaneously. We refer to change of one or more of the three parameters as a versatile alternative. Noting the dearth of literature for the equality two populations against such versatile alternative, we introduce two distribution-free tests based on the Euclidean and Mahalanobis distance. We obtain the asymptotic distributions of the two test statistics and study asymptotic power. We also discuss approximating p-values of the proposed tests in real applications with small samples. We compare the power performance of the two tests with several popular existing distribution-free tests against various fixed alternatives using Monte Carlo. We provide two illustrations based on biomedical experiments. Unlike existing tests which are suitable only in certain situations, proposed tests offer very good power in almost all types of shifts.     

Phytoremediation of Parboiled Rice Mill Wastewater Using Water Lettuce (Pistia Stratiotes)

Phytoremediation is an emerging technology applied for treatment of wastewater. It is a suitable option notably in developing countries as it is simple, sustainable and cost effective. In the present lab-based batch study the free floating aquatic plant water lettuce (Pistia stratiotes) is used for treatment of parboiled rice mill wastewater having low pH, high chemical oxygen demand (COD), nitrogen, and phosphate. In raw rice mill wastewater (undiluted) growth of water lettuce is found to be inhibited. Later on, two different dilution approaches (raw and facultative pond effluent 1:1; raw and tap water 1:1) are applied in order to effectively use this technology. In all cases a control (without plant) is maintained to compare the performance with the Aquatic Plant based Treatment (APT) system. In the APT system results reveal that removal of soluble COD (SCOD), ammoniacal nitrogen (NH₄-N), nitrate nitrogen (NO₃-N), and soluble phosphorus (sol. P) are upto 65%, 98%, 70%, and 65% respectively. The study highlights the efficacy of water lettuce in removing organics and nutrients from parboiled rice mill wastewater.