Antifungal Potential of Plant Growth Promoting Bacillus Species Against Blossom Blight of Rose

Blossom blight caused by Botrytis cinerea is one among the most devastating diseases that cause complete post-harvest loss in flower crops. The present study focuses on the development of effective bioformulation towards suppression of blossom blight and plant growth promotion in rose. Bacillus amyloliquefaciens (VB2) and Bacillus subtilis (AP) effectively inhibited mycelial growth of B. cinerea in vitro. Genome screening of VB2 and AP revealed the presence of antimicrobial peptide genes including, ituD, ipa14, bacA, bacD, srfA, sfP, spaC, spaS responsible for the biosynthesis of antibiotics such as iturin, bacilysin, bacillomycin, surfactin and subtilin. Further, the presence of volatile antifungal compounds in the bacterial secretome was identified through gas chromatography–mass spectrometry (GC/MS) analysis. Upon treatment, AP accelerated the metabolite profile of the plants and a rise in peak area of antifungal compounds such as, pentadecanoic acid, n-hexadecanoic acid, octadecanoic acid (stearic acid) and tetradecanoic acid was observed. In vitro, VB2 produced maximum indole acetic acid (9.17 µg/ml) and gibberellic acid (8.20 µg/ml) in nutrient broth. Under field conditions, foliar spray of VB2 at 0.5% (5 ml/l), four times at weekly interval suppressed blossom blight incidence (64% reduction over control) and also promoted yield. Future research towards development of an effective bioformulation with extended shelf life will aid in the management of various fungal, bacterial and viral diseases in different crop plants.

     

Thiol protease and cathepsin D activities in selected tissues and cultured cells from normal and dystrophic mice

Thiol protease and cathepsin D activities were studied in extracts from hindlimb muscle of 60-day-old normal and dystrophic mice, strain 129 ReJ, and from cultured normal and dystrophic cells. Total thiol protease activity in dystrophic muscle extracts was 3.5 times higher than in normal muscle extracts, while cathepsin D, activity was 2.2 times greater in dystrophic muscle compared with normal muscle. Activation (pH 4.5, 30 degrees C) of latent thiol protease activity in extracts of muscle occurred concomitant with the inactivation or dissociation of endogenous protease inhibitors. Thiol protease assays revealed a higher ratio of active to inactive protease activity in extracts from dystrophic muscle than from normal muscle. Cultured myoblasts (L69/1) were found to contain 30-fold more thiol protease(s) and 6-fold more cathepsin D activity than whole muscle. Cells established from dystrophic muscle and grown in culture for periods up to 6 months were more responsive to thiol protease activation conditions than similar cultures derived from normal muscle. From data on the rate and extent of thiol protease activation in extracts from dystrophic cells and hindlimb muscle compared with normal tissue, it appears that cells and tissues from dystrophic mice contain a lower level of protease inhibitors than cells and tissues from normal mice.     

Role of exogenous hemin in the synthesis of hemoproteins and nonheme proteins during glucose repression in Saccharomyces cerevisiae

Exogenous addition of hemin to glucose-repressed cells of Saccharomyces cerevisiae stimulates the incorporation of amino acid into cytoplasmic proteins twofold. There was no significant change in the synthesis of total cytoplasmic RNA whereas a 40% increase in the synthesis of poly(A)-containing RNA was observed upon hemin treatment. Cell-free translation of cytoplasmic mRNAs and immunoprecipitation analysis of the translated products with antibodies against subunit V of cytochrome oxidase and the alpha and beta subunits of F1-ATPase reveals that there is an eightfold enrichment of the mRNA for subunit V of cytochrome oxidase upon hemin treatment. The effect of hemin on the alpha and beta subunits of F1-ATPase is only marginal, suggesting a differential role for heme in the synthesis of hemoproteins and nonheme proteins during glucose repression.     

A small mobilizable IncP group plasmid vector packageable into bacteriophage lambda capsids in vitro

A mobilizable cosmid derivative of an IncP group plasmid was constructed by cloning the oriT region of RK2, a wide host-range plasmid, and the minimal DNA sequence of bacteriophage lambda required for efficient packaging in vitro. This cosmid is 13 kb in size and has unique restriction sites for EcoRI, XhoI, HindIII, and SalI. The XhoI and HindIII sites are within the kanamycin-resistance gene and the SalI site is in the tetracycline-resistance gene. This plasmid was mobilizable from an Escherichia coli donor to a number of diverse gram-negative bacteria at a frequency of 0.8 to 10 per 100 donors. This vector is one of the smallest of all wide host-range cosmids described in the literature. As part of this study, another mobilizable IncP group plasmid vector has also been constructed which, in addition to the sites listed above, has a unique BglII site, but which lacks the packager sequence.     

Results and recommendations

In the first phase of a collaborative study by the International Programme on Chemical Safety (PRCS), four coded chemicals, i.e. azidoglycerol (AG, 3-azido-1,2-propanediol), methyl nitrosurea (MNU), sodium azide (NaN₃) and maleic hydrazide (MH), and ethyl methanesulfonate (EMS) as a positive control were tested in four plant bioassays, namely the Arabidopsis embryo and chlorophyll mutation assay, the Tradescantia stamen hair assay (Trad-SH assay), the Tradescantia micronucleus assay (Trade-MCN), and the Vicia faba root tip assay. Seventeen laboratories from diverse regions of the world participated with four to six laboratories each using one plant assay. For the Arabidopsis assay, laboratories were in agreement with MNU and AG giving positive responses and NaN₃ giving a negative response. With the exception of one laboratory which reported MH as weakly mutagenic, no mutagenic response was reported for MH by the other laboratories. For the Vicia faba assay, all laboratories reported a positive response for MNU, AG, and MH, whereas two of the six laboratories reported a negative response for NaN₃. For the Trad-SH assay, MH was reported as giving a positive response and a positive response was also observed for MNU with the exception of one laboratory. NaN₃, which exhibited a relatively high degree of toxicity, elicited a positive response in three of the five laboratories. AG was found positive in only one of the two laboratories which tested this chemical. For the Trad-MCN assay, MNU and MH were reported as positive by all laboratories, while four out of five laboratories reported NaN₃ to be positive. Only one of three laboratories reported AG to be positive. The major sources of variability were identified and considered to be in the same range as found in similar studies on other test systems. Recommendations were made for minor changes in methodology and for initiating the second phase of this study.     

Genetic studies of water buffalo blood markers. I. Red cell acid phosphatase,albumin, catalase,red cell α-esterase-3, group-specific component,and protease inhibitor

We have developed the methodologies for typing and family studies to establish the modes of inheritance of water buffalo red cell acid phosphatase (Acp), protease inhibitor (Pi), and group-specific component (Gc) on isoelectric focusing and albumin (Alb), red cell -esterase-3 (Est-3), and catalase (Cat) on polyacrylamide gel electrophoresis. Family studies showed that Pi, Gc, Alb, and Cat are coded by autosomal genes with two codominant alleles, while Est-3 is autosomal with two codominant alleles and a recessive null allele and Acp exhibits three codominant alleles.This project was funded by the Australian Centre for International Agricultural Research through Grant PN 8364 and the Malaysian programme for Intensification of Research in Priority Areas through Grant IRPA 1-07-05-057.     

Stereochemical control of yeast reductions. 2. Quantitative treatment of the kinetics of competing enzyme systems for a single substrate

Quantitative expressions have been developed for systems such as yeast reductions where competing enzymes act on one substrate to yield two enantiomeric products. These expressions relate the observed stereochemical variables, the extent of conversion (C), the optical purity expressed as enantiomeric excess (ee), and the initial substrate concentration (A₀) to the kinetic parameters K_R and K_S (apparent Michaelis constants) and y ($\text{V}{}_{\mathrm{R}}\text{V}{}_{\mathrm{S}}$, the ratio of maximal velocities) of such competing enzymes. The expressions have been experimentally verified using a purified competing enzyme system of l- and d-lactic dehydrogenases. Furthermore, the enantioselective reduction of β-keto esters by intact yeast cells has been examined by means of this kinetic analysis.