In vitro rapid regeneration of plantlets from nodal explants of Mucuna pruriens– a valuable medicinal plant

A rapid and efficient protocol for the large‐scale propagation of a potential medicinal plant, Mucuna pruriens, through in vitro culture of nodal segment explants obtained from 15‐day‐old aseptic seedlings is described. Of the three different cytokinins, 6‐benzyladenine (BA), kinetin (Kin) and 2‐isopentenyl adenine (2‐iP) evaluated as supplements to Murashige and Skoog (MS) medium, BA at an optimal concentration of 5.0 μM was effective in inducing multiple shoots. Strength of the basal media also influenced the efficiency of shoot regeneration. The frequency of shoot regeneration tended to increase when the salt concentration in the basal media was reduced. Highest number of multiple shoots (23.3) and maximum average length (5.6 cm) were standardised on half‐strength MS medium supplemented with 5.0 μM BA along with 0.5 μM α‐naphthalene acetic acid (NAA) at pH 5.8. Rooting was best induced in shoots excised from proliferated shoot cultures on MS medium augmented with an optimal concentration of 1.0 μM indole‐3‐butyric acid (IBA). The in vitro‐raised plantlets with well‐developed shoots and roots were successfully established in earthen pots containing garden soil and were grown in greenhouse with 90% survival rate. The results of this study provide the first report on in vitro plant regeneration of M. pruriens.     

Effect of Transport at Ambient Temperature on Detection and Isolation of Vibrio cholerae from Environmental Samples

It has long been assumed that prolonged holding of environmental samples at the ambient air temperature prior to bacteriological analysis is detrimental to isolation and detection of Vibrio cholerae, the causative agent of pandemic cholera. The present study was aimed at understanding the effect of transporting environmental samples at the ambient air temperature on isolation and enumeration of V. cholerae. For water and plankton samples held at ambient temperatures ranging from 31°C to 35°C for 20 h, the total counts did not increase significantly but the number of culturable V. cholerae increased significantly compared to samples processed within 1 h of collection, as measured by culture, acridine orange direct count, direct fluorescent-antibody-direct viable count (DFA-DVC), and multiplex PCR analyses. For total coliform counts, total bacterial counts, and DFA-DVC counts, the numbers did not increase significantly, but the culturable plate counts for V. cholerae increased significantly after samples were held at the ambient temperature during transport to the laboratory for analysis. An increase in the recovery of V. cholerae O1 and improved detection of V. cholerae O1 rfb and ctxA also occurred when samples were enriched after they were kept for 20 h at the ambient temperature during transport. Improved detection and isolation of toxigenic V. cholerae from freshwater ecosystems can be achieved by holding samples at the ambient temperature, an observation that has significant implications for tracking this pathogen in diverse aquatic environments.     

MazF-induced Growth Inhibition and Persister Generation in Escherichia coli

Toxin-antitoxin systems are ubiquitous in nature and present on the chromosomes of both bacteria and archaea. MazEF is a type II toxin-antitoxin system present on the chromosome of Escherichia coli and other bacteria. Whether MazEF is involved in programmed cell death or reversible growth inhibition and bacterial persistence is a matter of debate. In the present work the role of MazF in bacterial physiology was studied by using an inactive, active-site mutant of MazF, E24A, to activate WT MazF expression from its own promoter. The ectopic expression of E24A MazF in a strain containing WT mazEF resulted in reversible growth arrest. Normal growth resumed on inhibiting the expression of E24A MazF. MazF-mediated growth arrest resulted in an increase in survival of bacterial cells during antibiotic stress. This was studied by activation of mazEF either by overexpression of an inactive, active-site mutant or pre-exposure to a sublethal dose of antibiotic. The MazF-mediated persistence phenotype was found to be independent of RecA and dependent on the presence of the ClpP and Lon proteases. This study confirms the role of MazEF in reversible growth inhibition and persistence.     

Protective role of allicin and L-ascorbic acid against the genotoxic damage induced by chlormadinone acetate in cultured human lymphocytes

In our present study, different doses of allicin and L-ascorbic acid were tested against the genotoxic damage induced by chlormadinone acetate (CMA; 40 microM) using chromosomal aberrations (CAs) and sister chromatid exchanges (SCEs) as the parameters. Treatment with allicin and L-ascorbic acid resulted in reduction of CAs and SCEs. The results suggested a protective role of allicin and L-ascorbic acid against CMA induced genotoxic damage.     

Comparative toxic potential of market formulation of two organophosphate pesticides in transgenic Drosophila melanogaster (hsp70-lacZ)

This study investigated the working hypothesis that two widely used organophosphate pesticides; Nuvan and Dimecron, exert toxic effects in Drosophila. Transgenic D. melanogaster (hsp70-lacZ) was used as a model for assaying stress gene expression and AchE activity as an endpoint for toxicity and also to evaluate whether stress gene expression is sufficient to protect against toxic insult of the chemicals and to prevent tissue damage. The study was extended to investigate the effect of the pesticides on the life cycle and reproduction of the organism. The study showed that Nuvan affected emergence of the exposed flies more drastically than Dimecron and the effect was lethal at the highest tested concentration (0.075 ppm). While Nuvan at 0.0075 and 0.015 ppm concentrations affected reproduction of the flies significantly, the effect of Dimecron was significant only at 0.015 and 0.075 ppm. Nuvan-exposed third-instar larvae exhibited a 1.2-fold to 1.5-fold greater hsp70 expression compared to Dimecron at concentrations ranging from 0.0075 to 0.075 ppm following 12 and 18 h exposure. While maximum expression of hsp70 was observed in Nuvan-exposed third-instar larval tissues following 18 h exposure at 0.075 ppm, Dimecron at the same dietary concentration induced a maximum expression of hsp70 following 24 h exposure. Further, concomitant with a significant induction of hsp70, significant inhibition of AchE was observed following chemical exposure and temperature shock. Concurrent with a significant decline in hsp70 expression in Nuvan-exposed larvae after 48 h at 0.075 ppm, tissue damage was evident. Dimecron-exposed larvae exhibited a plateau in hsp70 induction even after 48 h exposure and moderate tissue damage was observed in these larvae. The present study suggests that Nuvan is more cytotoxic than Dimecron in transgenic Drosophila melanogaster.     

Antibodies to citrullinated proteins and differences in clinical progression of rheumatoid arthritis

Antibodies to citrullinated proteins (anti-cyclic-citrullinated peptide [anti-CCP] antibodies) are highly specific for rheumatoid arthritis (RA) and precede the onset of disease symptoms, indicating a pathogenetic role for these antibodies in RA. We recently showed that distinct genetic risk factors are associated with either anti-CCP-positive disease or anti-CCP-negative disease. These data are important as they indicate that distinct pathogenic mechanisms are underlying anti-CCP-positive disease or anti-CCP-negative disease. Likewise, these observations raise the question of whether anti-CCP-positive RA and anti-CCP-negative RA are clinically different disease entities. We therefore investigated whether RA patients with anti-CCP antibodies have a different clinical presentation and disease course compared with patients without these autoantibodies. In a cohort of 454 incident patients with RA, 228 patients were anti-CCP-positive and 226 patients were anti-CCP-negative. The early symptoms, tender and swollen joint count, and C-reactive protein level at inclusion, as well as the swollen joint count and radiological destruction during 4 years of follow-up, were compared for the two groups. There were no differences in morning stiffness, type, location and distribution of early symptoms, patients' rated disease activity and C-reactive protein at inclusion between RA patients with and without anti-CCP antibodies. The mean tender and swollen joint count for the different joints at inclusion was similar. At follow-up, patients with anti-CCP antibodies had more swollen joints and more severe radiological destruction. Nevertheless, the distribution of affected joints, for swelling, bone erosions and joint space narrowing, was similar. In conclusion, the phenotype of RA patients with or without anti-CCP antibodies is similar with respect to clinical presentation but differs with respect to disease course.     

Proteome analysis of tobacco bright yellow-2 (BY-2) cell culture plastids as a model for undifferentiated heterotrophic plastids

We analyzed the proteome of undifferentiated plastids from a tobacco BY-2 cell culture by shotgun proteomics following multidimensional protein fractionation. The fractionation strategy initiated with the serial extraction of proteins from membranes which allowed us to distinguish soluble, peripheral, and integral membrane proteins. The majority of the identified proteins have a function in the cellular metabolism and most of them are active in amino acid synthesis pathways. A significant number of the identified proteins was not identified in chloroplast proteome analyses before. This suggests BY-2 plastid specific functions that differ from the major activities of chloroplasts. We have used the BY-2 plastid proteins reported here to assess the metabolic activities of undifferentiated heterotrophic plastids and compared the functional profile with that of differentiated heterotrophic amyloplasts. Comparative shotgun proteome analyses as reported here provide information about prevalent metabolic activities of different plastid types.