Efficacy of bacterial auxin on in vitro growth of Brassica oleracea L.

Murashige and Skoog’s (MS) medium was supplemented with supernatant of Halomonas desiderata RE1 in different combinations to observe the impact of bacterial auxin on in vitro growth of Brassica oleracea L. Three groups of combinations MS + BS (Bacterial supernatant), MS + BS + 10% CW (coconut water) and MS + BS + 4 mg l⁻¹ 2,4-dichlorophenoxyacetic acid (2,4-D) were considered. Different amounts of BS used in each combination were 50, 100, 150 and 200 μl in 5 ml MS medium. Media combinations inoculated with seeds, internodal explants and callus of B. oleracea L. were incubated in a growth chamber at 25 ± 1°C and exposed to 16-h cool fluorescent light. Seeds inoculated on MS + BS and MS + BS + 10% CW, shoot elongation was observed over control whereas this response was suppressed in 2,4-D-containing media. In explants inoculated on MS + BS, MS + BS + 10% CW and MS + BS + 4 mg l⁻¹ 2,4-D different responses such as callus induction, adventitious shoot induction and hypertrophy were observed at different supernatant treatments. In callus inoculation, callus proliferation was observed in most of the treatments at different media combinations.     

Rhizobacterial potential to alter auxin content and growth of <Emphasis Type="Italic">Vigna radiata</Emphasis> (L.)

Potential of non-symbiotic plant growth promoting rhizobacteria (PGPR) to influence the endogenous indole-3-acetic acid (IAA) content and growth of Vigna radiata (L.) was evaluated. The bacterial strains used belonged to Pseudomonas, Escherichia, Micrococcus and Staphylococcus genera. All strains were able to produce IAA (1.16–8.22 μg ml⁻¹) in the presence of 1,000 μg ml⁻¹ of l-tryptophan as revealed by gas chromatography and mass spectrometric (GC–MS) analysis. However, strains exhibited variable results for other growth promoting traits such as phosphate solubilization and siderophore or hydrogen cyanide production. Bacterial IAA production showed significant positive correlation with endogenous IAA content of roots (r = 0.969; P = 0.01) and leaves (r = 0.905; P = 0.01) under axenic conditions. Bacterization of V. radiata seeds significantly enhanced shoot length (up to 48.10%) and shoot fresh biomass (up to 43.80%) under fully axenic conditions. Bacterial strains applied under wire-house conditions also improved shoot length, number of pods, and grain weight up to 58, 65, and 17.15% respectively, over control. Hence, free living (non-symbiotic) PGPR have the ability to influence endogenous IAA content and growth of leguminous plants.     

Aeromonas punctata PNS-1: a promising candidate to change the root morphogenesis of Arabidopsis thaliana in MS and sand system

A model system of sand, comprising Arabidopsis plants inoculated with Aeromonas punctata PNS-1 strain, was used to evaluate the bacterial effect in modulation of plant root structure at second-order lateral root level. In MS media, the root morphogenesis was changed only at first-order lateral root level when inoculated with PNS-1 strain. Inoculation with PNS-1 strain was subjected to significant (P < 0.01) increase in primary root length and lateral root density in both MS and sand system. However, this strain modulated the root structure in the sand environment in a complex manner that may be helpful for incitation of the plant–microbe interaction close to natural environment. In order to determine whether this change in root morphology was due to bacterial auxin, Arabidopsis transgenic line (DR5:GUS) was used to reveal the change in homeostasis of endogenous auxin. In PNS-1 inoculated transgenic seedlings of Arabidopsis plant (DR5:GUS), endogenous auxin in primary root apices and lateral roots was enhanced. For confirmation, PNS-1 was evaluated for auxin production in vitro, showed an increase in auxin production after supplementation of l-tryptophan. The presence of ACC deaminase activity in PNS-1 showed its possible involvement in primary root elongation. In the present study Aeromonas punctata PNS-1 is the potential candidate for triggering the change in root morphogenesis of Arabidopsis thaliana with the involvement of auxin and ACC deaminase production.     

Screening of different contaminated environments for polyhydroxyalkanoates-producing bacterial strains

Total sixteen bacterial strains were isolated and purified from the samples collected from sugarcane molasses soil, sewage water and long-chain-hydrocarbon-contaminated area of the Punjab University, Lahore, Pakistan. Tolerance to different antibiotics was studied and strains showed multiple antibiotic resistance. All strains were characterized for Gram stain, biochemical reactions and polyhydroxyalkanoate (PHA) production. Total fourteen strains were Gram negative and two were Gram positive, while biochemically nine PHA producers showed affiliation to Pseudomonas, Enterobacter, Citrobacter, Bacillus and Escherichia. Screening for PHA production was done by Sudan black staining and nine out of sixteen strains exhibited PHA producing ability. PHA production was optimized for different growth parameters, like nitrogen concentration, pH and temperature. PHA extraction was done by solvent extraction method. Bacterial strains US1 and M1 accumulated up to 30% PHA of their cell dry weight on PHA extraction by solvent extraction method. Bacterial strain US1 was identified by 16S rRNA gene analysis as P. aeruginosa (DQ455691). PHA production was confirmed by PCR amplification of 500 bp fragment from PHA polymerase (Pha C) gene; five strains from nine PHA producers gave positive results on PCR. Pha C gene fragment of US1 was sequenced and submitted to Gene Bank under the accession number DQ455690. The amino acid sequence showed homology using the protein BLAST at 129–132 sites with different PHA synthases of the Pseudomonas sp.     

Targeting of 5-aza-2′-deoxycytidine residues by chromatin-associated DNMT1 induces proteasomal degradation of the free enzyme

5-Aza-2′-deoxycytidine (5-aza-dC) is a nucleoside analogue with cytotoxic and DNA demethylating effects. Here we show that 5-aza-dC induces the proteasomal degradation of free (non-chromatin bound) DNMT1 through a mechanism which is dependent on DNA synthesis and the targeting of incorporated 5-aza-dC residues by DNMT1 itself. Thus, 5-aza-dC induces Dnmt1 degradation in wild-type mouse ES cells, but not in Dnmt [3a^–/–, 3b^–/–] mouse ES cells which express Dnmt1 but lack DNA methylation (<0.7% of CpG methylated) and contain few hemi-methylated CpG sites, these being the preferred substrates for Dnmt1. We suggest that adducts formed between DNMT1 and 5-aza-dC molecules in DNA induce a ubiquitin-E3 ligase activity which preferentially targets free DNMT1 molecules for degradation by the proteasome. The proteasome inhibitor MG132 prevents DNMT1 degradation and reduces hypomethylation induced by 5-aza-dC.     

Comparative study of Cr(VI) uptake and reduction in industrial effluent by Ochrobactrum intermedium and Brevibacterium sp.

Two chromium-resistant bacterial strains, CrT-1 and CrT-13, tolerant up to 40 mg K2CrO4 ml(-1) on nutrient agar, 25 mg ml(-1) in nutrient broth, and up to 10 mg ml(-1) in acetate-minimal media, were identified as Ochrobactrum intermedium and Brevibacterium sp., respectively, on the basis of 16S rRNA gene sequencing. Uptake of chromate was greater in living cells than in heat-killed on dried cells. CrT-1 reduced 82%, 28% and 16% of Cr(VI) at 100, 500, and 1000 microg ml(-1) after 24 h while CrT-13 reduced 41%, 14% and 9%. Other heavy metals at low concentrations did not affect these reductions. At 150 and 300 microg ml(-1) in an industrial effluent sample Cr(VI) was reduced by 87% and 71%, respectively, with CrT-1 and by 68% and 47% with CrT-13.     

Fluorescent vesicles for signal amplification in reverse phase protein microarray assays

Developments in microarray technology promise to lead to great advancements in the biomedical and biological field. However, implementation of these analytical tools often relies on signal amplification strategies that are essential to reach the sensitivity levels required for a variety of biological applications. This is true especially for reverse phase arrays where a complex biological sample is directly immobilized on the chip. We present a simple and generic method for signal amplification based on the use of antibody-tagged fluorescent vesicles as labels for signal generation. To assess the gain in assay sensitivity, we performed a model assay for the detection of rabbit immunoglobulin G (IgG) and compared the limit of detection (LOD) of the vesicle assay with the LOD of a conventional assay performed with fluorescent reporter molecules. We evaluated the improvements for two fluorescence-based transduction setups: a high-sensitivity microarray reader (ZeptoREADER) and a conventional confocal scanner. In all cases, our strategy led to an increase in sensitivity. However, gain in sensitivity widely depended on the type of illumination; whereas an approximately 2-fold increase in sensitivity was observed for readout based on evanescent field illumination, the contribution was as high as more than 200-fold for confocal scanning.     

Report of a parasitic wasp (Hymenoptera: Encyrtidae) parasitizing cotton mealybug (Hemiptera: Pseudococcidae) in Pakistan and use of PCR for estimating parasitism levels

A parasitic wasp, Aenasius bambawalei, was studied for its biological parameters and parasitism levels in the cotton mealybug (Phenacoccus solenopsis). Biological parameters including parasitism efficiency, time to pupation, time to eclosion and adult sex ratio were studied under lab conditions. Parasitism levels in field collected mealybug were determined using PCR. Results showed an increase in parasitism over the study period, with higher parasitism levels in 2009 compared to the preceding 2 years.