Research Journal of Biotechnology

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Vol. 9(2) February 2014

Biopesticidal effect of Tinospora crispa extract against flea beetles, Phyliotera sinuata ateph

Nor Aziyah B., Norain I., Nor Aimi A.W., Lim B.T., Wan Zarina W.K. and Siti Nur Amirah Diana F.

Biopesticides have gained prominence as potential plant protecting agents. The stems of Tinospora crispa, a tropical plant, have been reported to possess insecticidal properties. However, study on the effect of T. crispa against flea beetles is very limited. Therefore, this study was carried out to investigate the insecticidal properties of the chloroform (CETC), ethanol (EETC), petroleum ether (PEETC) and ethyl acetate (EAETC) extracts of Tinospora crispa on Phyliotera sinuata ateph infesting mustard plant (Brassica juncea rugosa). The stems used in the experiment were collected from the locality and serially extracted with petroleum ether, followed by chloroform, ethanol, and ethyl acetate. A commercial pesticide, Cyperin (CCP) was included in the treatments as a standard check alongside the untreated (control). The experiment was laid out in a randomized complete block design (RCBD) with six treatments and four replicates. The efficacy of the treatments was based on the percentage reduction in Phyliotera sinuata ateph population. Results of the present study revealed that the application of 1.0 g/L of the ethanolic extract of T. crispa (EETC) could significantly (p<0.05) reduce the population of Phyliotera sinuata ateph by 30 to 89%. The preliminary phytochemical analysis of the most active extract, EETC showed the presence of alkaloids, glycosides and terpenoids compounds.

Characterization of castor (Ricinus communis L.) genotypes using different markers

Tomar Rukam S., Parakhia M. V., Kavani R. H., Dobariya K. L., Thakkar J. R., Rathod V. M., Dhingani R. M. and Golakiya B. A.

This work investigated the fingerprinting and phenotyping of 25 castor genotypes available in Gujarat and other States of India. An integrated approach based on the exploitation of morphological traits and molecular markers, such as RAPD and ISSR fingerprints was employed. Morphological trait analysis and statistical analysis of markers were useful for reconstructing a castor varietal dendrogram. The results of the morphological and molecular analyses allowed us to confirm a remarkable differentiation among castor genotypes. The UPGMA dendrogram obtained using morphological characters clearly separated the 25 genotypes of castor into three groups. Out of three groups, group I had fifteen genotypes while rest of the genotypes were divided among two groups. According to RAPD, highest genetic similarity (92%) was between SKI 336 and SKI 343. However, minimum similarity was found between JI 362 and SKI 336 (41%). Similarly, 81 polymorphic ISSR fragments generated by 27 primers divided castor genotypes into three main clusters; cluster one included two subclusters. The second main cluster contained five genotypes (JI 342, JI 372, JI 363 SKI 271 and DCS 78), while the third main cluster included only three genotypes.

Assessment of genetic diversity in wheat released from 1905 and breeding lines using microsatellite markers

Suneetha Kota, Mohapatra T. and Brajendra

Genetic diversity was assessed among 40 wheat genotypes consisting of new plant type (NPT) wheat derivatives, advanced breeding lines and released varieties since 1905 using 40 STMS markers. The total number of alleles amplified by 40 STMS markers was 113. The number of alleles amplified per STMS marker varied from one to six with an average of 3.32 alleles among the polymorphic markers. More alleles were detected for the B (3.55) genome derived STMS markers than A (3.18) and D (2.75) genomes. The maximum polymorphic information content (PIC) of 0.63 with an average of 0.35 was recorded while the PIC values for A, B, and D genomes were 0.243, 0.288 and 0.339 respectively. We identified 12 informative markers with three to six alleles and average of four alleles per marker and PIC values ranged from 0.31 to 0.63. Pair wise similarity coefficients ranged from 0.22 to 0.77 suggesting wider genetic diversity among wheat genotypes. The advanced breeding lines and NPT wheat derivatives exhibited different levels of diversity as compared with released cultivars since 1905 which could be utilized in widening the genetic base of wheat and as parental lines to tag as well as to mobilize useful traits for genetic improvement of wheat.

Partial identification of antimicrobial compound produced by thermotolerant Bacillus subtilis KFSB5 isolated from compost soil

Kanse Omkar Shankarrao, Kadam Tukaram Angadrao and Dnyanoba Kodgire Vasudha

The infectious and disease causing fungal and bacterial pathogens of plants and animals causes high rate of mortality and reduction of agricultural productivity. Therefore search for new antimicrobial compound is essential to overcome the problems. In the present study, we tried for isolation of thermophilic bacteria having antimicrobial potential against bacterial and fungal pathogens. A total 12 bacterial isolates were isolated and screened for antimicrobial activity. Among these, the Bacillus KFSB5 isolate was selected and identified as Bacillus subtilis KFSB5 by cultural, biochemical and 16S rRNA sequencing. It exhibited activity against Candida albicans, Ustilago maydis, Cryptococcus neoformans, Staphylococcus aureus and Escherichia coli. This antimicrobial activity was found to be due to production of amocaumacin-A like secondary metabolite, determined by using UV, IR, 1H NMR and Mass (m/z 424 (M+1)+) spectroscopic analysis. These results indicated that compost teak forest soil environment contains thermotolerant B. subtilis strains and can be rich source of new bioactive compound.

Molecular Characterization in Grain Amaranthus (Amaranthus spp.) using RAPD marker

Patel K. S., Parihar A., Chaurasia Pratibha, Patel P. J. and Pachchigar Karen

RAPD assessment on genetic variation of total 58 accessions comprising 15 parental genotypes and their 43 F1 hybrids of grain Amaranthus were analyzed. Among the 40 tested primers, four primers viz. OPA-06, OPA-10, OPA-14 and OPA-15 were able to discriminate all the genotypes sufficiently. In RAPD analysis, at the intraspeciﬁc level, the percentages of RAPD polymorphism were found to be 71.43 (A. hypochondriacus), 100 (A. cruentus), 100 (A. caudatus), 75.76 [IC-1733 (A. edulis)] and 84.85 [SKGPA-144 (A. tricolor)] and hybrids showed varied degree of polymorphism ranging from 42.86 to 71.43 per cent. The population matrix showed that allele at SKGPA-144 (Amaranthus tricolor) and F1 hybrids have maximum genetic distance of 0.742 to 0.962 in all populations while population IC-1733 (Amaranthus edulis) has minimum genetic distance among all the germplasm. The lowest and the maximum genetic distance of IC-1733(Amaranthus edulis) was 0.663 and 0.720 respectively to all populations. It further matches the pattern of allelic frequency distribution where SKGPA-144(Amaranthus tricolor) and F1 have remarkable high polymorphism.

Intergenic spacer region based marker for identification and quantification of Fusarium oxysporum f. sp. ciceris in chickpea plant using real time PCR assay

Priyanka K., Dubey S.C. and Singh A.K.

Wilt caused by Fusarium oxysporum f. sp. ciceris (Foc) is an important disease of chickpea worldwide. For rapid, reliable and specific detection of Foc, a set of markers PS15 F1 and R1was designed from the sequences of partial IGS region of Foc. The marker gave 197 bp amplicon in Foc isolates as well as in infected chickpea plants. Sensitivity of the marker was evaluated using conventional and real-time PCR assays. In conventional PCR assay the marker could amplify 0.5 ng and 1 ng of genomic DNA of Foc and chickpea plant infected with Foc respectively. Real time PCR assay was found more sensitive to amplify DNA at 0.01 pg extracted from Foc and 0.1 ng from Foc infected plant. The marker could detect Foc infection starting from 3 days up to 45 days post inoculation in chickpea roots as well as in lower half and upper parts of the plant. The marker was validated against 14 isolates of Foc collected from different parts of India and did not amplify the DNA extracted from Fusarium solani, Fusarium udum, Rhizoctonia solani, Rhizoctonia bataticola and Sclerotinia sclerotiorum. Therefore, a set of markers designed in the present study was found specific to Foc and can be used consistently for detection and identification of Foc isolates.

In Vitro Anti-Diabetic Activity of Aqueous Extract of the Medicinal Plants Nigella sativa, Eugenia jambolana, Andrographis paniculata and Gymnema sylvestre

Rastogi Archit, Mythili S. and Sathiavelu A.

Aqueous extracts of medicinal plants Nigella sativa, Eugenia jambolana and Andrographis paniculata, Gymnema sylvestre were prepared by maceration. They were then tested for inhibition of α-amylase activity by DNSA colour reagent. They were tested for their ability to hinder diffusion of glucose across a dialysis membrane. The aqueous extract of Nigella sativa showed maximum inhibition of α-amylase activity and a strong hindrance to diffusion of glucose across a dialysis membrane. Andrographis paniculata showed both a strong inhibition of α-amylase and a significant hindrance to the diffusion of glucose across the dialysis membrane. Gymnema sylvestre showed low inhibition of α-amylase activity but it showed maximum hindrance to the diffusion of glucose across the dialysis membrane. Nigella sativa was found to possess maximum anti-diabetic properties. All the above plants possess anti-diabetic properties to varying degrees. They can be used to develop natural drugs which may be used in lieu of commonly used strong allopathic drugs which possess a number of harmful side effects.

Wild goat’s habitat suitability modeling by ENFA in Agh-Dagh protected area, Ardabil, Iran

Gholamreza Naderi, Hossein Saadati, Mansureh Khalatbari, Aliakbar Imani and Borhan Riazi

We used Ecological niche factor analysis (ENFA) to describe wild goat’s niche in Agh-Dagh protected area and to identify key factors shaping its distribution and to describe its niche structure. Habitat suitability modeling showed that the species distribution is restricted to the marginal habitats principally because of repulsive factors such as human settlements and road networks. Meanwhile we found that the occurrence of the species was positively correlated with variables related to topographical features including elevation, scape terrain and slope. Since this area is the most important protected area in the province, immediate conservational actions should be done for managing repulsive factors.

Kinetics of biodegradation of carbonaceous organic matter through wet oxidation for environmental effluents released into marine waters

Dhage S. S., Kelkar P. S. and Prabhu D. V.

Industrialization and urbanization have twisted marine pollution as a serious problem. Coastal mega cities use ocean for waste disposal resulting in ecological stress on marine environment. Temperature, Biochemical oxygen demand (BOD) and Dissolved Oxygen (DO) are the most relevant parameters considered in water pollution. The reactions related to the biodegradation, decomposition and ratification of pollutants are expected to be altered due to high chlorides. Researchers had observed that the “k” values for complex decay reaction differ greatly with the diversity of effluents, temperatures and chemical composition of receiving waters. Reaction kinetics is the quantitative study of the reaction mechanism covering rates of chemical reactions and the different factors that affect the reaction rate during conversion of reactants into products. The decay of carbonaceous organic matter measured as BOD is a complex “Redox” reaction, termed as “wet oxidation” with first order kinetics. The activation complex theory for chemical reactions states that an activated complex is formed by the rearrangement of the atoms and bonds of the reactant molecule as an intermediate transient species which breaks into products as and when the reactant molecules possessing the energy of activation, come together and undergo effective collisions. A study on biodegradation of organic substrates commonly discharged into the creeks and near shore marine waters by wet oxidation was undertaken at selected chloride concentrations at two temperatures. The kinetics for decomposition of organic waste was assessed through calculation of the rate constant “k”. Effect of temperature on Reaction rate and thermodynamic activation parameters was also evaluated.

The reduction feed on shrimp vaname (Litopenaues vannamae) replaced by the addition biofloc in Ciamis District

Rostika R.

The increase in density and feeding level to increase the production of vaname shrimp culture (Litopenaeus vanamae) can cause the accumulation of of organic matter on the farming environment. Accumulation of organic matter leads to the water quality decrease and causes of organic Nitrogen compound from metabolic waste (excretion), unfed feed, shrimp feaces, dead algae and others organic matter. Heterotrophic system has the potential to utilize ammonia waste in the shrimp culture. Bacteria community accumulates inside aquaculture heterotrofik systems which can be utilized as a feed source. Bioflok contains high protein content and can be used as a source of feed for shrimp. This research has been carried out in the Centrol of Brackish and Sea Water Fish Seed Development (BPBIAPL), Ciamis District, West Java Province. The length of time of the study was 8 weeks February 2012 - May 2012. The material used were PL-21 vaname shrimp, molase, feed pellet shaped crumble with minimum 35% protein content. Flake bacteria contain Bacillus .sp, trash fish with nitrogen content of 9.25%. This study used a completely randomized design consisting of five treatments and three replicates: feed reduction 5 % (A), 10 % (B), 15 % (C), 20 % (D) and 25 % (E). The results of this study are vanamae shrimp fed 25% reduction in feed has 10.04 % daily growth rate, 8.31 grams absolute growth and 94% survival rate.

Comparative Studies on Antioxidant and Antipyretic Activities of Leaf Extracts of Cassia fistula, Psida cordifolia and Aegel marmelos

Kulkarni Harshal C. and Lohar Prakash S.

Cassia fistula, Psida cordifolia and Aegel marmelos were assessed for antioxidant and antipyretic activities. Scavenging free radical activity of each plant extract was quantified by in vitro decolorization of 2,2‐Diphenyl‐1‐picrylhydrazyl (DPPH) by spectrophotometric assay. The half maximal inhibitory concentration (IC50) values for each test samples were calculated and compared with ascorbic acid as a positive control. Yeast induced pyrexia was used to evaluate the antipyretic activity of the plant leaf extracts in albino rats. Test plants showed statistically significant antioxidant and antipyretic abilities which were dose dependant. Leaf extract of A.marmelos shown highest free radical scavenging activity as compared to C. fistula and then P. cordifolia while methanolic leaf extract of P. cordifolia showed statistically significant antipyretic effect in yeast-induced elevated body temperature in rats in comparison to that of C.fistula and A.marmelos.

Composition and Structure of Bacterial Communities in Waste Water of Aquatic Products Processing Factories

Wang Zhonghua, Yang Jianqiang, Zhou Jun, Zhang Chundan, Su Xiurong and Li Taiwu

The wastewater from aquatic products processing factories contains large amount of organics, pathogenic bacteria, virus, parasite, and inormanic salt. In this study, we examined the bacterial diversity associated with wastewater outfalls in aquatic processing industrial zone in Ningbo, China over a seasonal cycle using 454-pyrosequencing technology. α-diversity indices were high in all of the samples indicating that the species diversity was also likely to be very high. In the samples analyzed, the most abundant taxonomic phylum was Proteobacteria which represented 73.9%, 70.1%, 68.5% and 69.3% of the total sequences obtained in March, May, August and October. The second most abundant phylum was Firmicutes. The composition of proteobacterial assemblages changed over the seasonal cycle with sequences affiliated with γ-proteobacteria representing 91.7% of the total proteobacterial sequences in March but only 21.3%, 45.9% and 39.8% of the total in May, August and October respectively. The abundance of sequences affiliated with β-proteobacteria represented only 2.5% and 2.6% of the total in March and October respectively, but increased to 20.7% and 11.5% of the total in May and August. Moreover, another notable change was ε-proteobacteria which represented only 3.4% of the total sequences obtained in March but dramatically increased in May (48.0%), August (38.5%) and October (56.3%). These results suggest that the taxonomic composition of this sewage outfall was stable when considered at high taxonomic ranks (e.g. phylum level of taxonomic classification) over a seasonal cycle but shifted dramatically when lower taxonomic ranks (e.g. class level of taxonomic classification) were considered.

Proteomic profiling of the serum from stage I lung squamous cell carcinoma patients

Xin-Ju Li, Da-Lin He and Jun-Ke Fu

The aim of this study is to employ proteomic profiling to identify specific tumor markers that could improve early diagnosis of lung squamous cell carcinoma. Serum samples were isolated from 30 patients with stage I lung squamous cell carcinoma and 30 age-and gender-matched healthy controls and the serum proteomic profiles were obtained by matrix-assisted laser desorption ionization time of flight (MALDI-TOF) mass spectrometry (MS). In comparison to healthy controls, three highly expressed potential tumor markers were identified in the sera of stage I lung squamous cell carcinoma patients, with the molecular weights 3261.69, 3192.07 and 2556.92 Da (P < 0.001). A model was established based on the patterns of these three markers and evaluated by an independent test of blind serum samples from 22 patients with lung squamous cell carcinoma and 19 healthy individuals. The analysis yielded a sensitivity of 95.5%, and a specificity of 94.7%. In addition, one protein peak with molecular weight 3261.69 Da was chosen as the candidate biomarker and was identified as fibrinogen alpha chain through a search of the IPI, NCBI or SWISS-PROT protein databases. Fibrinogen alpha chain may be a potential tumor biomarker for the early diagnosis and prognosis of lung squamous cell carcinoma.

Coat protein-mediated resistance to Turnip mosaic virus in Chinese cabbage (B. rapa ssp. pekinensis) by the pollen-tube pathway

Han Li Wei, Ge Yu, Rao Lei, Cui Chong Shi and Qu Shu Ping

In this research we established a plant transformation system of coat protein-mediated resistance to Turnip mosaic virus. The gene TuMV-CP was introduced into Chinese cabbage through the pollen-tube pathway using the plasmid pBBBast-TuMV-CP (9.79 kb) which was composed of the gene TuMV-CP, expression regulatory sequence (CaMV 35S promoter, OCS terminator), the herbicide resistant gene Bar and other elements. A total of 0.15% of T0 plants was positive for the gene TuMV-CP by specific PCR. Further PCR Southern blot analyses showed that TuMV-CP insertions were harbored in T0 plants.

Factors Influencing the Salt Stress Tolerance in Plants – An Overview

Vaibhav Kumar Maurya and Gothandam K. M.

Salinity stress is one of the adverse stresses among abiotic stress which mitigates ion toxicity, water unavailability and oxidative stress apart from the hampering growth and productivity of plants. Various anthropogenic activities also indulge in limiting the cultivatable land thus minimizing the food productivity. For sustainable food supply to world population which is increasing at its peril, it is important to develop stress tolerant crops. Salt adaptation involves various trait of gene which comes across in an integrated manner to lessen the adverse effect of salt. This salt signaling cascade activates various mechanisms after perceiving the signal at membrane level receptor which leads to activate the various genes and transcription factors involved in production of osmolytes and ion sequestration. In this review we tried to accentuate the impacts of salinity on plant, role of calcium ion, transcription factor, osmolytes and plant hormone in salt tolerance.