Research Journal of Biotechnology

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Vol. 11(9) September 2016

Gener cloning and expression characterization of a novel JcBCCPII gene from Jatropha curcas

Wen Jinfen, Deng Minghua, Gong Ming and Chen Kai

In the present study, jatropha JcBCCPII gene was cloned and characterized. This novel gene was deposited into NCBI (KC875846). The results of cDNA sequences of the JcBCCPII gene demonstrated that it is a gene with a complete open reading frame of 642 bp encoding a JcBCCPII protein of 213 amino acids. Similarly comparisons for amino acid sequences reveal that the JcBCCPII protein has high homology with the BCCPII proteins of six other species. The phylogenetic tree analysis revealed that the JcBCCPII protein has a closer genetic relationship with the JcBCCPII protein of Ricinus communis and Populus trichocarpa than with those of Vitis vinifera, Glycine max, Lotus japonicas and Arabidopsis thaliana. Real-time PCR analysis shows that JcBCCPII gene is expressed in various tissues but at different levels. The expression levels of this gene are highly expressed in leaf, flower and seed, moderately expressed in stem and pericarp, weakly expressed in root. The JcBCCPII gene displayed a bell-shaped pattern of expression which had peak expression at 30 and 40 DAF but low at the first two stages of seed development, i.e. 10 and 20 DAP. The JcBCCPII expression can be induced by MeJA and BRs. These data provide a foundation for further insight into this gene.

Impact of UV treatment on Exotoxin production by Bacillus species

Akhtar Mahvish, Siddiqa Ayesha and Faisal Muhammad

Four bacterial strains Bacillus cereus, Bacillus subtilis, Bacillus licheniformis and Bacillus pumilis already isolated were studied for the optimal exotoxins production after 24 hrs incubation. Lecithin, hemolysin and casein production was optimal during 6, 18 and 24 hours incubation respectively. These strains were resistant against many heavy metals at various concentrations. These Bacillus species were plasmid bearing giving a hint of the presence of their pathogenic markers on plasmids. After 30, 60 and 90 min exposure of UV radiations, mutagenesis study showed their reduced ability to form casein only by B. subtilis and B. cereus. However all strains lost their ability to show lecithinase activity. Reduced resistance at different concentration of heavy metals was observed after UV radiations exposure at various time periods.

Replacement of Antibiotics by Feeding Probiotic Lactobacilli and Medical Plant Herba Houttuyniae in Broiler Chickens

Li Qingzhi and Zhang Fan

To investigate the combined effects of a probiotic strain Lactobacillus acidophilus NCFM and a medical plant Herba Houttuyniae on meat quality under healthy and growth performance under infection of broilers, 120 one-d-old broiler chicks were randomly assigned to 4 experimental groups with two separate pens. Each pen was composed by 4 cages and each cage was allocated with 15 birds. Four groups in each pen were treatments as 1) basal diet supplemented with 0.05% antibiotics, 2) basal diets supplemented with 1% probiotics, 3) basal diets supplemented with 1% medical plant extract and 4) basal diets supplemented with both 1% probiotics and 1% medical plant extract. Under healthy condition, supplementation of medical plant can significantly decrease the feed conversion ratio (FCR) during whole feeding period comparing to antibiotics or probiotics (p<0.05). Meanwhile, medical plant alone cannot increase weight gain. Under Salmonella enteritidis infection, only the combination of probiotics and medical plant can maintain survive ratio to comparable level to antibiotics. Even they both show anti-Salmonella activity in vitro respectively; neither alone can effectively cure infection in the study. Collectively, the study indicates that antibiotics, exactly chlortetracycline can be replaced in broiler farming by feeding probiotic L. acidophilus NCFM and medical plant Herba Houttuyniae in combination from the perspectives of both promoting productivity and preventing infectious diseases.

Biodegradation study of Bisphenol –A by Cornybacterium glutamicum isolated from effluent of thermal paper industry

Vijayalakshmi V.

A bacterial strain with bisphenol - A (BPA) degrading ability was isolated from thermal paper industry effluent and identified as Cornybacterium glutamicum (99% similarity with species) based on their 16s rRNA gene sequence analysis. The bacterial cells were rod shaped, gram positive and motile. The bacteria were able to resist 30mM BPA in a nutrient medium and degrade up to 25mM in BSM medium. The half maximal inhibitory concentration (IC50) values of bacteria in nutrient and basal salt medium were observed as 24.89 and 17.62 respectively. The Monad model was also applied for BPA degradations in nutrient and basal salt medium. High Performance Liquid Chromatography (HPLC) chromatograms revealed that 25mM concentrations of BPA gradually decreased within 9 days of incubation observed in a corresponding reduction in the peak area from 100% (initial concentration) to 19.17% (final concentration). The present study was reported as an efficient protocol for BPA degradation.

Association of allele diversity and polymorphism of microsatellites markers in the tropical goat

Raed M. Al-Atiyat

The association relationship between allelic diversity and polymorphism is still not covered by good examples in goat genome. Seven tropical goat breeds, two from Jordan, three from Saudi Arabia, one from Syria and one from Somalia, were genotyped at 17 microsatelliate loci in order to give better insights on the relationship. The average allele number (N) of all breeds was 9.65 whereas the average polymorphic information content (PIC) of all loci in all breeds was 0.721. The N and PIC values per breed were 6.12, 4.706, 4.647, 6.88, 6, 5.059 and 5 and 0.665, 0.614, 0.618, 0.656, 0.617, 0.646 and 0.614 in the Black Bedouin and Black Mountain from Jordan, the Damascus from Syria, the Jabali, Bishi, Tohami from Saudi Arabia and the Somali from Somalia. Both values at the studied loci were considered very high. The N was strongly correlated (r2=0.83) with the PIC whereas it was largely correlation with an allele size range (r2=0.60). However, a low correlation (r2=0.21) was observed between PIC and the allele size range. The majority of alleles were found in all breeds, except the alleles with either low frequencies or extreme sizes that were probably considered alleles come from gene flow or mutation. To sum up, this study concludes that the studied goat breeds were showing good evidence of the positive and strong relationship between allele diversity and polymorphism.

Association of melanin content with pathogenicity and virulence in Bipolaris oryzae

Singh Kartar, Valarmathi P., Sharma Sapna, Bashyal Bishnu Maya, Gurjar Malkhan Singh and Aggarwal Rashmi

Brown spot disease caused by Bipolaris oryzae (Cochliobolus miyabeanus) has been associated with two major epidemics in India and one of the strongest yield reducers among rice diseases in recent years. In the present study, morphological and pathological characterization has been investigated. Isolates of B. oryzae were collected from NWPZ (North western plain zone) and NEPZ (North eastern plain zone) of India. Out of 27 isolates, those of group I with black fluffy growth had the highest frequency (30%) in the population, whereas group V with pink fluffy growth had the lowest frequency (6%). Based on pathogenicity studies, isolate BO 1 from Ludhiana, Punjab was found to be most virulent and BO 9 from Sonipat, Haryana was the least virulent. The melanin content was quantified in all the isolates and a positive correlation between melanin concentration and pathogenicity in rice host was observed.

Green synthesis of silver nanoparticles using Barleria cristata

Meenakshi K. and George Indu A.

This is a simple, cost effective and reproducible green method to synthesize stable silver nanoparticles from silver nitrate through bioreduction method using Barleria cristata leaf extract. The phytosynthesis of silver nanoparticles was demonstrated first by visual observation and then by spectral methods [UV-Visible absorption spectroscopy, Fourier Transform Infrared Spectroscopy (FTIR) and Dynamic Light Scattering technique (DLS)]. The Energy dispersive X-ray spectrum and TEM analysis revealed that the metal was indeed silver and the nanoparticles were spherical in shape with a particle size that ranged from 7.61nm to 16.46 nm respectively.

The effects of propolis to anti-inflammatory in tumor necrosis factor-α-stimulated human periodontal ligament fibroblasts

Huang Wen-Chien, Tsai Hsin-Chi, Chen Young-Fa, Hsu Tsui-Kang, Wang Cheng-Chun, Chen Jyh-Larng, Shih Feng-Cheng and Chiu Yi-Chou

The study investigated whether propolis has inhibitory effects on tumor necrosis factor (TNF)-α-stimulated inflammatory response in human periodontal ligament fibroblasts (hPDLFs). Human periodontal ligament fibroblasts were exposed to TNF-α and treated with propolis. Cytotoxicity was assessed by the MTT assay. mRNA expression levels of proinflammatory mediators including interleukin (IL)-1β, IL-6, TNF-α, and cyclooxygenase (COX)-2 were assessed by quantitative polymerase chain reaction (qPCR). Proinflammatory cytokine production was measured by enzyme-linked immunosorbent assay (ELISA). Nuclear localization of nuclear factor (NF)-κB was examined by western blotting. Exposure to TNF-α resulted in significant elevation of mRNA expression of IL-1β, IL-6, TNF-α, and COX-2 in hPDLFs, increased the production of proinflammatory cytokines, IL-1β and IL-6, and enhanced nuclear translocation of NF-κB. Treatment with propolis downregulated the mRNA levels of IL-1β, IL-6, TNF-α, and COX-2 in TNF-α -stimulated hPDLFs in a dose-dependent manner. TNF-α-induced NF-κB nuclear translocation was reduced in presence of propolis. Propolis treatment inhibited mRNA expression of inflammatory mediators and suppressed NF-κB activation in hPDLFs exposed to TNF-α. Propolis suppresses TNF-α -induced inflammatory responses, including the expression of IL-1β, IL-6, TNF-α and COX-2, suggesting that propolis treatment significantly inhibits the upregulation of inflammatory mediators induced by TNF-α in hPDLFs.

Hepatoprotective role of Spirulina fusiformis on Methotrexate induced liver injury in Wistar Rats

Asha Devi S. and Blossom Benny

Several clinically useful drugs in addition to its beneficial action when used for long periods undergo biotransformation in the liver resulting in the production of reactive oxygen species (ROS) which lead to oxidative stress. The action of antioxidant enzymes present in the liver gets debilitated due to the excess ROS production. Therefore protective measures relevant to liver are of great interest. The present study was performed to evaluate the efficiency of Spirulina fusiformis in protecting against Methotrexate (MTX) induced hepatotoxicity in female wistar albino rats. Animals were divided into 5 groups and first group was treated with normal saline, second group with MTX 20 mg/ kg body weight, third group Spirulina fusiformis (300 mg/kg) + MTX, fourth group Spirulina fusiformis (500 mg/kg) + MTX and fifth group Spirulina fusiformis alone (500mg/kg). The study was carried out for 14 days following which rats were sacrificed and processed for histopathological, antioxidant enzymatic assays. Significantly (p<0.05) reduced antioxidant levels and significantly (p<0.05) increased lipid peroxidation levels were observed in MTX treated group which was ameliorated by the action of Spirulina fusiformis. Histopathological study also supported the finding. From the results obtained, it is conceivable that pretreatment of Spirulina fusiformis protected the species from MTX induced hepatotoxicity.

Cytotoxicity and apoptotic effect of biogenic silver nanoparticles on human colorectal cell line HT-29

Manimegalai Sengani and Devi Rajeswari V.

One of the most deliberate aggravation of ulcerative colitis commences to the development of colorectal cancer (CRC) owing to its high prevalence and mortality rates. The aim of our present study is to comprehensively explore the efficacy of the use of biogenic silver nanoparticles (AgNPs) cytotoxicity in primary colon cells and colorectal malignant cell line (HT-29). In addition, induction of apoptotic cell death by the influence of AgNPs was evaluated on HT-29 and primary cells in a dose-dependent manner. Formerly, the AgNPs were synthesised using Couroupita guianensis and its characterization was accomplished. We detail the execution of anti-proliferative effect of biogenic AgNPs at Ic50 of 20µg/ml on human colorectal malignant cell line HT-29 whereas the primary colon cells proliferation was inhibited at IC50 of 40µg/ml at 24 h incubation. Collectively the results intended that AgNPs expend their anti-proliferative activity towards HT-29 controlling cell cycle phases by inducing apoptosis.

The Fatty Acid Profiles in the Muscle Tissues of Four Benthic Fish Species from Northeastern Mediterranean Sea of Turkey

İlkan Ali Olgunoğlu and Engin Artar

The total fat contents and the fatty acid compositions in muscle tissue of four benthic marine fish species namely Merluccius merluccius, Lophius budegassa, Helicolenus dactylopterus, and Chlorophthalmus agassizi collected from the Mediterranean Sea of Turkey were determined. The results show that these species were particularly rich in protein while they were poor in fat as well. The highest crude protein value was observed in C. agassizi (22.34%) while the lowest was observed in L. budegassa (20.21%). The fat content was identified to differ between 0.44-0.82% and the highest value was observed in H. dactylopterus while the lowest in L. budegassa. Major saturated fatty acid (SFA), monounsaturated fatty acid (MUFA) and polyunsaturated fatty acid (PUFA) were palmitic acid, oleic acid and docosahexaenoic acid (DHA) followed by EPA in four species respectively. In conclusion, these species are desirable item for the human diet.

Adsorption of Amoxicillin on Surfactant Modified Zeolites and their Antibacterial Activity

Hoda Hosseinpour Nasirmahaleh, Sher Zaman Safi, Mohammad Reza Sazegar, Rajes Qvist, Ikram Shah Bin Ismail, Muhammad Aqeel Ashraf and Nik Ahmad Nizam Nik Malek

This research was divided into three main parts: loading of HDTMA and amoxicillin on the natural clinoptilolite, characterization of the product using Fourier Transform Infrared (FTIR) spectroscopy and the antibacterial activity using minimum inhibitory concentration (MIC). IR spectra of natural zeolites, clinoptilolite and the modified ones with HDTMA and amoxicillin proved that the band position was same even at the highest surfactant loading level which suggests no changes in the structure of the samples after modification with amoxicillin and surfactant molecules (HDTMA). No changes in the peaks of the IR spectra in the regions of 3400 to 2600 cm-1 for HDTMA-zeolite were seen when compared with the respective parent zeolites. More over our results demonstrate two significant peaks at around 2849 cm−1 and 2916 cm−1 which mean that the surfactant had been successfully attached on clinoptilolite. In addition, loaded amoxicillin on SMZ did not show any special peaks due to similar groups such as C-H with HDTMA-clinoptilolite. FTIR spectra of Amoxicilin-surfactant-clinoptilolite and surfactant modified zeolite were same. MIC results revealed that the antibacterial activity of the modified clinoptilolite with surfactant and amoxicilin was significantly higher as compared to the parent clinoptilolite and amoxicilin alone.

Use of the red winged sticky traps for collecting bark and ambrosia beetles [Scolytinae (Coleoptera: Curculionidae)] on deciduous trees of Kasnak oak nature protection area in Isparta, Turkey

Sarıkaya Oguzhan and Sayın Huseyin

Kasnak Oak (Quercus vulcanica) Nature Protection Area is situated in Yukarıgökdere province of Isparta city in the South-western of Turkey. This area contains Quercus vulcanica (Boiss. and Heldr.ex) Kotschy) which is endemic for Turkey. In this study bark and ambrosia beetle species (Col.: Curculionidae, Scolytinae) distributed on deciduous trees of the Kasnak Oak Nature Protection Area were determined. For this aim, the Red wing sticky traps (by mixing of ethyl alcohol 96% and toluene 1%) were set in stand and were checked periodically. Also, specimens were collected from weak trees and also broken and fallen trees by snow. As a result, 12 Scolytinae species were determined. These species are; Hylesinus crenatus (Fabricius, 1787); Hylesinus varius Fabricius, 1775;Dryocoetes villosus (Fabricius 1792); Taphrorychus ramicola (Reitter, 1894); Taphrorychus villifrons (Dufour, 1843); Scolytus intricatus (Ratzeburg, 1837); Scolytus koenigi (Schevyrew 1890); Scolytus mali (Bechstein 1805); Scolytus rugulosus (Müller, 1818); Anisandrus dispar (Fabricius, 1792); Trypodendron signatum (Fabricius 1787) and Xyleborinus saxesenii (Ratzeburg, 1837). Among these species, X. saxesenii is the most abundant species.

Biosynthesis of Bacterial Cellulose imparting Antibacterial Property through Novel Bio-Agents

Sakthivel K., Periyasamy Siddhan and Hungund Basavaraj

Biosynthesis of bacterial cellulose was carried out under ambient conditions by gram negative bacterium Acetobacter xylinum using standard medium and modified media. The yield of cellulose in these media was compared. Bacterial cellulose was characterized using infrared spectroscopy and scanning electron microscopy. Its water holding capacity was assessed and results were compared with that of plant cellulose. Bacterial cellulose and plant cellulose were treated with extracts of novel natural herbs such as bitter gourd (Momordica charantia), tridax daisy (Tridax procumbens) and with chitosan to impart antibacterial activity. The treated samples demonstrated good antibacterial property against Staphylococcus aureus and Escherichia coli. Modified bacterial cellulose demonstrated better antibacterial activity than that of modified plant cellulose.

Microbes using PAHs as Energy source: Relationship with diseases

Yasir Anwar, Amr A EL Hanafy, Jamal SM Sabir, Saleh Mohamed Al-Garni and Mohamed Morsi M. Ahmed

Bioremediation is known as a natural process which relies on bacteria and fungi to degrade, to breakdown and to remove various contaminants such as petroleum hydrocarbons from soil and water. These microorganisms possess the capability to use petroleum hydrocarbons as energy source and consequently render the contaminants harmless or less toxic. Various substances with known toxic properties are continuously being introduced into the environment due to human activities. These contaminants pose great risk to human health and cause serious damage to environment and biodiversity. Polycyclic aromatic hydrocarbons (PAHs) have been detected in high concentrations at sites that use or process petroleum products. PAHs are found to be carcinogens and very recalcitrant and persist in natural environment for several years. In this review our main focus is on biodegradation of PAHs (pollutant) by different bacterial and fungal groups and the different pathways that are used by these microbial groups to degrade PAHs. PAHs are not only an environmental hazard but are found to be a good source of energy to these microbes and hence their degradation not only relieves the environment of this environmental pollutant but provides the basic building blocks for microbes to survive. Bioremediation has tremendous potential to provide a cost-effective natural process to render toxic substances in environment harmless overtime.