Cadmium resistant Enterobacter cloacae and Klebsiella sp. isolated from industrial effluents and their possible role in cadmium detoxification

Three bacterial strains, two of Klebsiella sp. and one Enterobacter cloacae were isolated from industrial effluents of chemical and textile industries. They showed high efficiency in removing cadmium (Cd²⁺) from the medium. When 100 g/ml of Cd was added to the medium, the three isolates namely CMBL-Cd1, CMBL-Cd2 and CMBL-Cd3 removed or accumulated 86%, 87% and 85% of Cd, respectively, from the medium within 24h. Plasmids were detected in all the three strains. Plasmids of E. cloacae (pCBL1) and Klebsiella sp. (pCBL2 and pCBL3), estimated to be 6.6kb, were used to transform Escherichia coli C600. The transformed E. coli cells showed elevated resistance to Cd. Ethidium bromide curing indicated the presence of the Cd resistance gene on the plasmid. Resistance of the isolated strains against other metals like chromium (cr⁶⁺) and lead (pb²⁺) and a number of antibiotics was also checked. Cured strains showed lowered resistance against Cr and some antibiotics. This again supported the indication of the presence of Cd, Cr and some antibiotics resistance genes on plasmids.     

Protein proteinase inhibitor genes in combat against insects, pests, and pathogens: natural and engineered phytoprotection

The continual need to increase food production necessitates the development and application of novel biotechnologies to enable the provision of improved crop varieties in a timely and cost-effective way. A milestone in this field was the introduction of Bacillus thuringiensis (Bt) entomotoxic proteins into plants. Despite the success of this technology, there is need for development of alternative strategies of phytoprotection. Biotechnology offers sustainable solutions to the problem of pests, pathogens, and plant parasitic nematodes in the form of other insecticidal protein genes. A variety of genes, besides (Bt) toxins that are now available for genetic engineering for pest resistance are genes for vegetative insecticidal proteins, proteinase inhibitors, alpha-amylase inhibitors, and plant lectins. This review presents a comprehensive summary of research efforts that focus on the potential use and advantages of using proteinase inhibitor genes to engineer insect- and pest-resistance. Crop protection by means of PI genes is an important component of Integrated Pest Management programmes.     

Method for determining oxygen consumption rates of static cultures from microplate measurements of pericellular dissolved oxygen concentration

We describe a simple protocol for determining the oxygen consumption of cells in static culture. The protocol is based on a noninvasive oxygen-sensing microplate and a simple mathematical model derived from Fick's Law. The applicability of the model is confirmed by showing the correlation of computed oxygen consumption rate (OCR) values to actual cell densities ascertained by direct cell counting and/or MTT for HL60 and U937 cells cultured in suspension. Correlation between computed OCR and these other indications of cell number was quite good, as long as the cultures were not diffusion-limited for oxygen. The impact of the geometric factors of media depth and well size were confirmed to be consistent with the model. Based on this demonstrated correlation, we also developed a simple, completely noninvasive algorithm for ascertaining the per-cell oxygen utilization rate (OUR), which is the ratio of OCR to cell number, and a fundamental cell characteristic. This is accomplished by correlating the known seed densities to extrapolated determinations of OCR at time zero. Such determinations were performed for numerous cell types, in varying well sizes. Resulting OUR values are consistent with literature values acquired by far more painstaking methods, and ranged from <0.01 fmol.min(-1).cell(-1) for bacteria to 0.1-10 fmol.min(-1).cell(-1) for immortalized mammalian and insect cell lines to >10 fmol.min(-1).cell(-1) for primary hepatocytes. This protocol for determining OCR and OUR is extremely simple and broadly applicable and can afford rapid, informative, and noninvasive insight into the state of the culture.     

Inducive effect of cresoquinone on microbiological transformation of L-tyrosine to 3,4 dihydroxy phenyl L-alanine by Aspergillus oryzae NG-11(P1)

The present work describes the inducive effect of cresoquinone on microbiological transformation of L-tyrosine to 3,4 dihydroxy phenyl L-alanine ( L-DOPA) by Aspergillus oryzae NG-11(P1). Mould mycelium was used for biochemical conversion of L-tyrosine to L-DOPA because tyrosinases, beta-carboxylases and tyrosine hydroxylases are intracellular enzymes. The maximum conversion of L-tyrosine to L-DOPA (0.428 mg/ml) was achieved after 60 min of biochemical reaction. To enhance the production of L-DOPA, cresoquinone was added to the reaction mixture. Best L-DOPA biosynthesis results were observed when the concentration of cresoquinone was 3.5 x 10(-6) M (1.686 mg/ml L-DOPA produced with 1.525 mg/ml consumption of L-tyrosine). Cresoquinone not only increased enzyme activity but also enhanced cell membrane permeability to facilitate secretion of enzymes into the reaction broth. Comparison of kinetic parameters revealed the ability of the mutant to yield L-DOPA [Y(p/x) [i.e., mg L-DOPA formed (mg cells formed)(-1)] =7.360+/-0.04]. When the culture grown on various cresoquinone levels was monitored for Q(p), Q(s) and q(p) [ Q(p): mg L-DOPA produced ml(-1) x h(-1); Q(s): mg substrate consumed ml(-1) x h(-1); q(p): mg L-DOPA formed (mg cells)(-1) h(-1)], there was significant enhancement ( P<0.025) of these variables.     

Acarine regulators of water hyacinth in Kerala (India)

Screening of the mite fauna on water hyacinth, Eichhornia crassipes, at 25 localities throughout Kerala (India) yielded a list of 21 phytophagous species from nine families. Distribution of mites at these localities and type of injury produced suggested that Oligonychus biharensis and Orthogalumna terebrantis warrant further study. In an experimental set-up the injury caused by these mites was quantified. Infestation by Ol. biharensis lead to reduced photosynthesis by the host (significant loss in chlorophyll content). Injurious effects of Or. terebrantis involved leaf mining by the developing instars leading to wilting of the leaves resulting in reduced dry weight. The relative merits of the two species as potential control agents of water hyacinth are discussed.     

Deletion of cytosolic phospholipase A2 promotes striated muscle growth

Generation of arachidonic acid by the ubiquitously expressed cytosolic phospholipase A2 (PLA2) has a fundamental role in the regulation of cellular homeostasis, inflammation and tumorigenesis. Here we report that cytosolic PLA2 is a negative regulator of growth, specifically of striated muscle. We find that normal growth of skeletal muscle, as well as normal and pathologic stress-induced hypertrophic growth of the heart, are exaggerated in Pla2g4a-/- mice, which lack the gene encoding cytosolic PLA2. The mechanism underlying this phenotype is that cytosolic PLA2 negatively regulates insulin-like growth factor (IGF)-1 signaling. Absence of cytosolic PLA2 leads to sustained activation of the IGF-1 pathway, which results from the failure of 3-phosphoinositide-dependent protein kinase (PDK)-1 to recruit and phosphorylate protein kinase C (PKC)-zeta, a negative regulator of IGF-1 signaling. Arachidonic acid restores activation of PKC-zeta, correcting the exaggerated IGF-1 signaling. These results indicate that cytosolic PLA2 and arachidonic acid regulate striated muscle growth by modulating multiple growth-regulatory pathways.     

Influence of selected physical parameters on the biodegradation of acrylamide by immobilized cells of Rhodococcus sp.

The influences of concentration of acrylamide, pH, temperature, duration of storage of encapsulated cells and presence of different metals and chelators on the ability of immobilized cells of a Rhodococcus sp. to degrade acrylamide were evaluated. Immobilized cells (3 g) rapidly degraded 64 and 128 mM acrylamide in 3 and 5 h, espectively, whereas free cells took more than 24 h to degrade 64 mM acrylamide. An acrylamide concentration of 128 mM inhibited the growth of the free cells. Immobilized bacteria were slow to degrade acrylamide at 10 °C. Less than 60% of acrylamide was degraded in 4 h. However, 100% of the compound was degraded in less than 3 h at 28 °C and 45 °C. The optimum pH for the degradation of acrylamide by encapsulated cells was pH 7.0. Less than 10% of acrylamide was degraded at pH 6.0, while ca. 60% of acrylamide was degraded at pH 8.0 and 8.5. Copper and nickel inhibited the degradation, suggesting the presence of sulfhydryl (-SH) groups in the active sites of the acrylamide degrading amidase. Iron enhanced the rates of degradation and chelators (EDTA and 1,10 phenanthroline) reduced the rates of degradation suggesting the involvement of iron in its active site(s) of the acrylamide-degrading-amidase. Immobilized cells could be stored up to 10 days without any detectable loss of acrylamide-degrading activity.     

Toxic and Antifeedant Effects of Different Pesticidal Plant Extracts against Beet Armyworm (<i>Spodoptera exigua</i>)

The beet armyworm (BAW), Spodoptera exigua (Lepidoptera: Noctuidae) is a highly destructive pest of vegetables and field crops. Management of beet armyworm primarily relies on synthetic pesticides, which is threatening the beneficial community and environment. Most importantly, the BAW developed resistance to synthetic pesticides with making it difficult to manage. Therefore, alternative and environment-friendly pest management tactics are urgently required. The use of pesticidal plant extracts provides an effective way for a sustainable pest management program. To evaluate the use of pesticidal plant extracts against BAW, we selected six plant species (Lantana camara, Aloe vera, Azadirachta indica, Cymbopogon citratus, Nicotianatabacum, and Ocimum basilicum) for initial screening experiment. Four out of six plant species such as A. indica, N. tabacum, C. citratus and O. basilicum showed promising mortality of more than 50%. Therefore, we selected these four plant extracts for the subsequent experiments. Through contact bioassay, A. indica showed high mortality 66.63%, followed by the N. tabacum 53.33%, at 10% w/v concentration. Similarly, N. tabacum showed the highest mortality rate, 66% at 10% w/v concentration, followed by the A. indica 46% through feeding bioassay. Furthermore, the feeding deterrence assay showed that C. citratus had a high antifeedant index (−50) followed by A. indica (−39), and N. tabacum (−28). In living plant assay, the N. tabacum extract showed a low mean damage score 3.6 on living cotton plant followed by C. citratus 4.5 and A. indica 5.5. Hence, extracts of three plant species provided promising results against the BAW, which can minimize the use of synthetic chemicals, particularly for small landholding farmers. Further studies are also required to evaluate the effects of these plant extract against BAW on cotton plants under field conditions to optimize the further use.