Vol. 21, October, 2005

1/21 TECHNOLOGICAL STUDIES ON COMMON CARP FISH FILLETS DURING FROZEN STORAGE.

A.I. Ez-El-Rigal, M.I. Salama and S.I. Ali

Central Laboratory for Aquaculture Research, Abbassa, Agriculture Research Center, Ministry of Agriculture, Dokki, Giza.

The objective of this work was to evaluate the effect of filleting methods (skinned off and un skinned) of common carp Cyprinus carpio L. on fillet yield, chemical properties, freshness indices, the microbial count, sensory evaluation and histological structure, during storage at –20±2ºC for three months. Results obtained indicated that, the chemical and histological, freshness indices and the microbial count showed some changes in all treatments. Skinned off fillets showed much more stability than un skinned fillets during storage at –20±2ºC for three months. These results corresponded to the gradually decrease in sensory evaluation for all samples. From these results, it could be concluded that, skinned off fillets were much desirable than un skinned fillet of common carp during frozen storage.

 

 

2/21 SYNERGISTIC EFFECT OF GA3 AND BA ON GROWTH, PHOTOSYNTHETIC PIGMENTS, METABOLITES, PHYTOHORMONES AND FIBRE YIELD OF ROSELLE PLANTS

H.M. S. EL-Bassiouny;H.K.I. Khattab* andM.S. Sadak

Department of Botany, National Research Centre, Dokki, Cairo, Egypt.     *Department of Botany, Faculty of Science,Ain Shams University, Cairo,Egypt.

Two pot experiments were carried out during tow successive seasons, to study the effects of gibberellic (GA3) acid, benzyladenine (BA) and a mixture of both at a concentration of 100 and /or 200 mg L-1 in presence and absence of Fe-EDTA on growth criteria, photosynthetic pigments, carbohydrate fractions, protein-N, free amino acids as well as the fibre properties and composition of roselle plant. There was a positive relation between the growth criteria evident in terms of shoot height, number of branches, number of leaves per plant, stem circumference, leaf-area per plant, fresh and dry weights of shoot per plant and the applied plant hormones. The pigment levels concomitantly with carbohydrate fractions as well as free amino acids and protein-N contents of roselle plants were markedly increased. Moreover, the exogenous application of the investigated plant hormones significantly improved the fibre length (cm), strength (g/cm2) tensile and increased the percentage of cellulose in the cell wall whereas lignin content was reduced. On the other hand, endogenous hormones, as evident in terms of IAA, GA3, zeatin riboside and zeatin glucoside, were markedly increased particularly at 100 mg L-1 of both GA3 and BA in presence of Fe-EDTA. In contrast ABA levels were markedly declined in treated roselle plants.

3/21 INFLUENCE OF ALLELOPATHIC OF ACACIA RADDIANA LEAF EXTRACT ON GERMINATION AND SOME METABOLITES SEEDLING OF LUPINE TERMIS

S.E. Saffan and H.M. Salama

Botany Department, Faculty of Science, Zagazig University

The objective of this study was to determine the impact of allelopathic potential of aqueous extract of Acacia raddiana leaves on germination and metabolite accumulation in lupine termis seedlings. The results showed that the quantitative analysis of aqueous extract of Acacia contained the phenolic compounds and flavonoids that might be implicated as allelochemicals agent. The germinated seedling after 48 hrs. were inhibited with the increase in concentration of aqueous Acacia leaves, but the phenolic compounds of the Lupine seedling had stimulatory effects with high level of concentration of Acacia leaves extract. Degradation of storage carbohydrates of lupine seedlings was significantly retarded with increasing the concentration of Acacia leaves extract. This retardation was attributed to the inhibition of activity of amylase enzyme that reduced the contents of reducing and non reducing sugars. However, the polysaccharides remained at a high level when compared with those of the control. While, inhibition of protease activity led to accumulation of free amino acids and protein contents in germinated seedling of Lupine termis. Although aqueous extract of Acacia leaves contain allelopathic chemicals and phytotoxic, but the lipase activity during germinated seedling of lupine revealed the higher level at 2% concentration of Acacia leaves extract, but the lower level was accompanied with 4% concentration of Acacia leaves extract. This effects led to increase in unsaturated fatty acids than saturated fatty acids in germinated Lupine seeds. The treated germinated Lupine seeds with 4% Acacia leaves extract contain Erucic acid (C:22:0), but it is absent in another treatment.

 

4/21 ROLE OF HELICOBACTER PYLORI AMONG PATIENTS WITH DIFFERENT GASTROINTESTINAL DISODERS IN ISMILIA, EGYPT

S.M. Abdalla1, A.A. Abdelrahman2, K.A. Khalil3 and S.M. Enany4

Microbiology and Immunology Dept., Faculty of Pharmacy1,2,4, Internal Medicine Dept., Faculty of Medicine3, Suez Canal University1,2,3,4, Ismailia, Egypt.

Helicobacter pyloriinfection is now firmly established as a key etiological factor in peptic ulcer disease, gastric mucosa-associated lymphoid tissue lymphoma and gastric adenocarcinoma. The present study was carried out on 150 "out patients" referrals to the Upper Gastrointestinal Endoscopy Unit, Suez Canal University Hospital. Each patient was subjected to endoscopic examination. Biopsy Specimens were taken from the stomach of each patient for rapid urease test, isolation, identification, inoculation into Christensen's urea broth and culture on both Colombia blood agar and Brain heart infusion agar. Suspected H. pyloricolonies were subjected to colony morphology identification, microscopical examination and biochemical reactions. Furthermore, samples were subjected to PCR to detect ureA subunit of urease gene as one of the virulence factors.Our results showed that endoscopic examination of all patients revealed normal, gastric ulcer, duodenal ulcer, gastritis and gastric cancer with a rate of 20.7%, 20%, 24%, 33.3% and 2%, respectively. Direct smear exam revealed 52%H. pyloripositive while culture and rapid urease test showed 71.33%. Fifty four biopsies (36%) were urease positive after 1 hour at room temperature, 39 (62%) after 1 hour incubation at 37ºC and 14 (71.33%) after 24 hour incubation. Isolated H. pylori showed that they were catalase, oxidas, and urease positive. PCR results showed 411-bp fragment is indicative for the ureA subunit of urease gene using primers specific for this subunit. The presence of ureA gene is strong confirmation of H. pylori. We concluded that prevalence of theinfection is high. Strong association between H. pyloriand duodenal ulcer is noticed. H. pylorichanges from bacillary form to cocoid one in old culture were reported. Metronidazole resistance is a character of the isolates within most patients in Ismailia, while Amoxicillin resistance is rare. Finally, a 411-bp fragment indicative of the ureA subunit of urease gene is detected in all the tested isolates. UreA gene presence is a strong confirmation for H. pyloriidentification.

5/21 INFLUENCE OF WET AND DRY INOCULUM OF THE ENTOMOPATHOGENIC FUNGUS METARHIZIUM ANISPOLIAE (METSCHNIKOFF) AGAINST THE MATURE LARVAE OF THE POTATO TUBERMOTH, PHTHORIMAEA OPERCULELLA (ZELLER) COMBINED WITH NEEM AND GAMMA IRRADIATION

N.H. El-Sinary

Natural Products Department, National Center for Radiation Research and Technology, Atomic Energy Authority, Nasr City, Cairo Egypt

       Wet and dry inoculum of the entomopathogenic fungus Metarhizium anispoliae (Metschnikoff) were used against the mature larvae (4th larval instar) of the potato tubermoth (PTM), Phthorimaea operculella (Zeller). Results indicated that wet inoculum (5 x 105 , 5 x 106 and 5 x 107 spores/ ml saline) were more effective than dry inoculum; (5: 100, 10: 100 and 20: 100, (inoculated rice: sand / gm)) in reducing the viability of the mature larvae, pupation and adult emergence percentages. In both wet and dry inoculum treatments there was positive correlation between the fungus concentrations and the reduction in larval development. Combined effect of neem and gamma irradiation were also studied. Results indicated that addition of neem leaves powder to the sand where the mature larvae pupate in ratios; (1: 10 and 1: 20 (neem: sand/ gm)) and rearing neonate larvae on irradiated potato tubers with different gamma irradiation doses (40, 80 and 120 Gy) reducing the viability of (PTM) larvae when combined with M. anispoliae treatments the best results were recorded with wet treatments (5 x 107) M. anispoliae combined with (1: 10 (neem: sand) ) and 120 Gy gamma irradiation dose.

 

 

6/21 IN-VITRO STUDY OF THE EFFECT OF SOME BIOTECHNOLOGY DRUGS "r.h. ERYTHROPOIETIN; r.h. GROWTH HORMONE AND R.H. INSULIN" ON O2- CONSUMPTION AND GENOMIC DNA, RNA AND PROTEIN OF ISOLATED RAT LIVER MITOCHONDRIA AND THE TISSUE HOMOGENATE

Z.A. Teleb, K.M. El-Deib, N.Z. Ahmed; M.M. Ahmed and M.I. Ibrahim

National Organization For Drug Control & Research, "NODCAR" Department of Molecular Drug Evaluation

The effect of recombinant human Erythropoietin "r.h.EPO"; recombinant human Growth hormone "r.h. GH"; and recombinant human Insulin "r.h. Insulin" on O2-consumption rate of isolated rat liver homogenate and mitochondrial suspension incubated in-vitro at 37°C over two hours were performed. Two concentrations of each drug were used "0.9 U/ml & 1.8 U/ml" for Erythropoietin; "0.0003U/ml & 0.0006 U/ml" for Growth hormone, and" 0.026 U/ml &0.052 U/ml" for Insulin. In addition to, the genomic DNA, RNA and protein were extracted and purified, then the determination of the purity and concentration were investigated. Identification of these macromolecules on agarose and acrylamide were performed. The total protein content was increased after the first half hour either in mitochondria or homogenate of rat liver under the effect of the two concentration of each drug while the protein was decreased after one and half hour incubation period in either rat liver homogenate or mitochondria. EPO, GH and Insulin exerted on a non significant decrease by the two doses after 1 and 2.06 hours incubation period in rat liver homogenate while the effect was significantly decreased in rat liver mitochondria. Identification of protein on SDS/PAGE after the drug treatments in-vitro for incubation periods exerted unchanged pattern. Isolated rat liver mitochondria and homogenate were assayed for in-vitro oxygen consumption by O2 - monitor inolab Ox1 level 2 instrument. The addition of the biotechnology drugs by the two concentration levels to the mitochondria fractions resulted in activation of O2 - uptake by the two doses of EPO after the four hours of incubation time, as well as the O2 -uptake after the first hour of incubation period under the effect of the two doses of r.h. GH and r.h. Insulin. Growth hormone by the two doses decreases the O2-uptake after the second, third, and fourth hours of incubation period. Insulin by the two doses exerted an inhibitory effect after the second hour incubation while after the third and fourth hours of incubation time the O2-consumption was increased. Comparative studies of these biotechnology drugs by the two doses on the O2-uptake of rat liver homogenate showed an inhibitory effect after four hours incubation. The purity and concentration of DNA of mitochondria and homogenate isolated from rat liver after treatment in vitro by the three drugs by the two doses were investigated. The purified genomic DNA was isolated on agarose 1% and compared against Ladder DNA. RNA concentration was varied according to the dose of drug and the incubation periods. The results of RNA molecular size exerted no alteration between the treated and non treated samples by the two doses of these drugs

 

 

7/21 Enterovirus Assessmentof WastewaterTreatment Plants in Damietta Governorate, Egypt

N.A. El-Esnawy, W.M. El-Senousy and A.K. Allayeh

Virology Laboratory- Water Pollution Research Department, Environment Research Division, National Research Centre, Cairo- Dokki, Egypt

This study was carried out to detect enteroviruses genome by RT-PCR in three wastewater treatment plants, to compare the presence of infectious enteroviruses to the presence of enteroviruses genome, and to investigate the genetic diversity of isolated enteroviruses strains. The experiments were performed on 54 wastewater samples collected from wastewater treatment plants (WWTPs) in Damietta city. Enterovirus was detected using RT-PCR to amplify a 236 bp fragment in the 5` non-coding region (NCR) using P1 (sense) and PV444 (antisense) pair of primers. Enteroviruses were detected in 16.66% (9/54) of total collected samples. While, they were 29.62 % (8 /27), and 3.70 % (1/27) in the inlet and outlet; of (WWTP) respectively. A sequenceanalysis of an amplified 236 bp fragment was occurred and compared with sequences derived from the correspondingenterovirus genome region deposited in GenBank. Phylogentic analysis revealed that our sequences were closely related to human poliovirus 1 Sabin strain (98% homology). It may, return to the intensive campaigns of poliovaccination in Egypt for poliomyelitis eradication.

 


8/21 CHEMICAL CONTROL OF EDIBLE MUSHROOM MYCOPATHOGENS (COMPETITIVE SAPROPHYTIC AND PARASITIC WEED FUNGI)

M.E.A. Dawoud and M. Eweis

Department of Botany, Faculty of Science, Cairo Univ., Giza, Egypt

For chemical control of mushroom mycopathogen (competitive weed fungi), the growth response Pleurotus ostreatus (mushroom) and it’s competitive weeds (Pencillium chrysogenum, Trichoderma viride and Chaetomium olivarum) was studied under different concentrations (0.00 – 104 mM) of Li+ , Cr6+ , Mn2+ , Cu2+ , Co2+ , Zn2+ , Sr2+ , Mo2+ , La2+ and Ba2+. The results of the main effect showed that, increased heavy metal concentrations decreased the linear growth and dry weight gain of mushroom, Trichoderma and Chaetomium, but some heavy metals increased these criteria in Pencillium chrysogenum. Nevertheless, the data of interaction between heavy metals and test fungi showed that some low concentrations (10mM) favoured the fungal growth as in the following profile; mushroom Mo2+ > Zn2+ > Li+ > Cu2+. Trichoderma Mn2+ > Co2+> Mo2+ > La2+ > Zn2+ > Li+ > Ba2+; Chaetomium Zn2+ > Mo2+ > Li+ > Cr6+ and Penicillium chrysogenum Mo2+ > Sr2+ > Co2+> Zn2+ > Mn2+ > Li+. These results led to the conclusion that, Penicillium chrysogenum was the potent heavy metal-resistant one among the tested fungi followed by Trichoderma then Chaetomium. Also the result showed that, Cr6+ was lethal to Trichoderma and Penicillium but stimulated the growth of Chaetomium and had a little suppressive effect on mushroom. On the other hand, Cu2+ was toxic to Penicillium, Trichoderma and Chaetomium but more suppressive (at higher concentration) to mushroom growth than Cr6+. Mixture of Cu2+, Cr2+ at 10mM inhibited the growth of competitive weeds and did not harm mushroom. Addition of siapton (amino acids biostimulant) favoured protein and decreased lipid, carbohydrate in addition to Cu2+, Cr6+ contents and energy contents of mushroom mat. These results clarify a conclusion that, a mixture of Cu2+ and Cr6+ is promising potant chemical agent for controlling mushroom competitive weed fungi with the least harm to mushroom growth.

 

 

9/21 IDENTIFICATION AND CHARACTERIZATION OF SOME MEDICINAL PLANTS BELONGING TO FAMILY "LABIATAE" USING SPECIFIC MOLECULAR TECHNIQUES

Z.A. Teleb, W.W. Mohamed, S.S. Abdel-Fattah and E.M. El-Garawani

National Organization for Drug Control and Research "NODCAR"

Applied Research Center of Medicinal Plants "ARCMP"., Biotechnology Laboratory

The four species of Labiatae “Melissa officinalis; Rosmarinus officinalis; Salvia officinalis; and Ocimum basilicum" were selected and evaluated using molocular techniques. Here we describe the essential steps of total RNA, protein, and rapid DNA isolation protocols that can be used for diverse medicinal and aromatic plants, which produce essential oils and secondary metabolites. The protein concentration and the identification on SDS/PAGE were performed. Total RNA was extracted and the purity and concentrations were carried out. The purified RNA was isolated on agarose 1.3%. In addition, the genomic DNA was purified and its purity and concentration were determined. The purified DNA & RNA were isolated on agarose 0.7% &1.3% respectively. RAPD-PCR technique of the four medicinal plants was performed. The results obtained from the molecular analysis revealed two main proteins in the four species having molecular weight 197 KD & 31KD. It also revealed no alteration in the molecular size of RNA in the species studied. The molecular weight of RNA was less than 74 bp. The genomic DNA of these medicinal plants was similar having a molecular weight about 19.329 Kbp. DNA marker technique have been used to investigate the genetic relationships between the types of species of Labiatae. Random amplified polymorphic DNA (RAPD) was used to characterize genomic alterations of these species.

 

 

10/21 PURIFICATION AND CHARACTERIZATION OF THE MAJOR GLUTATHIONE TRANSFERASE ISOENZYME

FROM PHYSA ACUTA.

A.A. Abdalla

Molecular Biology Department, National Research Centre, Dokki, Cairo, Egypt

The major glutathione transferase isoenzyme (GST3) was purified from the cytosolic fraction of the fresh water snails Physa acuta to electrophoretic homogeneity by ammonium sulfate fractionation, anion exchange chromatography and glutathione-Sepharose affinity chromatography with 47.2 % recovery of the total activity. The specific activity toward 1-chloro-2, 4-dinitrobenzene (CDNB) was increased to 33.5 µmol/min/mg protein with 192 purification fold. The enzyme is a homo-dimeric protein, with a subunit molecular weight of approximately 23.5 kDa. The Km for CDNB was 75 µM and for GSH was 105 µM. I50 values for cibacron blue and bromosulphophthalein were 325 nM and 60 nM, respectively. The isoenzyme showed optimum pH value at 7.5 and optimum temperature at 50°C.The half life time at 50°C was 40 min. Addition of 5 mM GSH to the incubation buffer increased the half life to more than 100 min. The acid-induced inactivation of GST3 showed that the isoenzyme was completely inactive at pH 3.0 but retains most of its activity at pH above 5.4. Unfolding/refolding of the P. acuta GST3 monitored by activity were investigated using guanidinium chloride (GdmCl) and urea as denaturants. The midpoint was 0.8 M and 3.75 M for GdmCl and urea, respectively. The reactivation was at least 80 % and 90 % for GdmCl   and urea, respectively.


11/21 Investigation of virulence genes in Yersinia   enterocolitica O: 8 using subtractive hybridization

M.I. Abou-Dobara, A. Ismail*, F. A. Mansour, M.M. Zaky*, A. Rakin**

and J. Heesemann**

Botany Department, Faculty of Science, Mansoura University, Mansoura,

Egypt.

*Department of biological science, Faculty of Education, Suez Canal University,

Port-Said, Egypt.

**Max von Pettenkofer-Institute, fűr Hygiene und Medizinische Mikrobiologie,

Munich University, Germany.

The pathogenic Yersinia enterocolitica, the causative agent of a broad range of gastrointestinal syndromes always harbours the important virulence factors, such as the virulence 70-kbp plasmid which encodes the Yop virulon and the high-pathogenicity island (HPI) which encodes the Yersiniabactin iron responsible genes, but other virulence genes still exist in Yersinia enterocolitica need to be identified and characterized. Subtractive hybridization is one of the most powerful tools for the identification of virulence genes in a wide range of bacterial pathogens. In this study the hybridization of highly pathogenic Yersinia enterocolitica O:8 and low pathogenic Yersinia enterocolitica O:5 was successful to identify the two novel genes which are probably have relation to virulence. The first is pripiline peptidase, which has been demonstrated by Polymerase Chain Reaction (PCR) in most pathogenic Yersinia species, is suggested to be responsible for the formation of faimbriae and pilli. The second gene is an invasin Inv homolog sequence which has an opening reading frame (ORF) of invasin Inv of pathogenic Yersinia enterocolitica which could be named as Inv2.

 

12/21 ENHANCEMENT OF EXTRACELLULAR GLUCOSE OXIDASE PRODUCTION BY ASPERGILLUS NIGER THROUGH FERMENTATION- OPTIMIZATION PROCESS.

W.A. Bazaraa

Food Science Department, Faculty of Agriculture, Cairo University, Giza, Egypt.

Factors (carbon source, carbon source concentration, nitrogen source, nitrogen source concentration and calcium carbonate concentration) affecting the synthesis of the extracellular glucose oxidase (GOD, b-D-glucose: O2 1-oxido reductase, EC 1.1.3.4) by Aspergillus niger (FS-3) in batch shaking system were investigated. The enzyme synthesis was strongly influenced by calcium carbonate and glucose. The highest GOD production (28.5 U ml-1) was achieved when the organism was allowed to grow in medium containing 14 % glucose, 0.4 % peptone and 5 % CaCO3. Consecutive optimization of growth media improved GOD production by 179.3 %. When the optimized conditions were applied on the A. niger mutant (E-407) or the fusant (C-18), increments of 559.1 and 613.8 % in GOD production were obtained. Time course studies of growth and GOD production by A. niger strain FS-3 before and after optimization as well as strains E-407 and C-18 in the optimized medium are also reported.

13/21 BIOLOGICAL AND CHEMICAL STUDY OF NIGELLA SATIVA L. SEEDS. I

H.A. Kadry, S.E. El-Dondaity and A. I. Muhammad.

Department of Pharmacognosy, Faculty of Pharmacy (Boys)

Al-Azhar University, Cairo, Egypt.

Thebiological activity of Nigella sativa L. expressed seeds oil was evaluated on human volunteers complaining of Candida albicans (oral thrush), otomycosis, inflammatory conditions of the   mucous membranes of the mouth, throat and gums (aphthous stomatitis or common mouth ulcers, alveolar pyorrhea, teething troubles or denture rubbing, sore gums ..etc), tineaversicolor (tinea pityriasis) tinea pedis (Athlete's foot) and Onychomycosis (tinea of nails).Good results were obtained. Chemical study of n-butanol fraction of Nigella sativa L. seeds marc lead to isolation and identification of kaempferol, astragallin, a-hederin and 3-O-[β-D-xylopyranosyl (1® 3) -a-L-rhamnopyranosyl (1 ® 2) -a-L-arabinosyl) hederagenin. Their structures were determined by spectral analysis. This represents the first report for isolation of kaempferol and astragallin from Nigella sativa L. seeds.


14/21 BIOSYSTEMATIC STUDY OF GENUS NICOTIANA IN EGYPT BASED ON FINGERPRINTS OF RAPD AND ISOZYME

W.M. Amer and A.M. Fawzy*

Herbarium, Botany Deptartment, Faculty of Science, Cairo University, Giza, Egypt.

* Department of Flora and Phyto-Taxonomy Research, Horticultural Research Institute, Agricultural Research Centre, Egypt.

The loss of biodiversity has become an issue of great global concern, especially the wild relatives of the cultivated economic species. Genus Nicotiana (Solanaceae) is among the genera of high potentiality in the Egyptian flora. The genus represented in Egypt by four sections; each of them represented by one species: three wild Nicotiana species namely: N. glauca Graham, N. plumbaginifolia Viviani and N. rustica L. While, N. tabacum L. (tobacco) is the only cultivated species. Sixteen RAPD primers and two isozymes were used to assess the genetic variation and inter-specific relationships among tobacco and its wild relatives in Egypt. The polymorphism in RAPD and isozyme bands were used to construct the phylogenetic dendrogram. The inter-specific relationship reflects that: (1) The genetic improvement of the cultivated N. tabacum can be achieved using its closely wild relative (N. plumbaginifolia). (2) Conservation of wild crop-relatives is an urgent topic to secure the gene pool of the economic species.

 

15/21 GIBBERELLIC ACID PRODUCTION BY SOLID STATE FERMENTATION USING CORNCOBS.

A.S. Gad* and M. Ewis

* Chemistry of Natural and Microbial Products lab, NRC, Giza, Egypt.

Botany Dept, Faculty of Science, Cairo University, Giza, Egypt.

Desirable production of Gibberellic acid in solid state culture by Aspergillus niger using corncobs or wheat bran as carbon sources were evaluated. Maximum GA3 production, 248mg/kg corncobs was found to depend on the coarse particle size after addition of 1 g % glucose and 0.25 mg % urea after 8 days. The optimum conditions studied were 2x 106 spore/ml. (75.5%) initial moisture content and autoclaving time for 75 min on using the same concentration of glucose and urea.

16/21 Integrated APPROACH for Rapid Mass Propagation of sugarcane

M.E. Wagih, Y. Musa*, A. Ala* and O.M. Badawy**

Biotechnology Centre, University of Technology, Lae, PNG, North Australia

*University of Hasanuddin, Makassar, Indonesia

**Department of Breeding and Genetics, Agriculture Research Station (Sabahia), Sugar Crops Research Institute, Agriculture Research Centre, Egypt

         A strategy aiming at rapid mass propagation of sugarcane (Saccharum spp.) cultivars Q77, Cadmus and Co 997 was developed. The strategy involved in vitro methods including embryogenic callus (EC) and one-eye microset (1.0 cm) shoot cultures combined with nursery methods involving decapitation/stumping, planting one-eye microset in soil and splitting of multi-shoot, were developed. ECs were initiated and grown on a medium composed of MS salt (Murashige and Skoog, 1962), 1 mg L-1 thiamin, 50 mg L-1 arginine, 100 mg L-1 myoinositol, 100 ml L-1 coconut water, 30 g L-1 sucrose and 8 g L-1 agar supplemented with 4.0 mg L-1 2,4-D 0.5 mg L-1 Nicotinic acid and 0.5 mg L-1 pyridoxine. This medium, when lacked 2,4-D and BAP was suitable for the regeneration of plantlets from ECs and microsets. Microset planting in soil was superior and more vigorous compared to microset shoot culture, while germination rate was 100% in both cases. Regeneration rate by EC culture from all varieties was in average 9,000 plantlets per plant per annum. Decapitation/stumping of greenhouse grown plants together with microset planting in soil resulted in multi-shooting, which, when singled-out, accounted for up to 2 fold increase in the multiplication rate by EC alone. Similarly, while traditional propagation in the field produce in average 25 plants per one-eye set per annum, decapitation/stumping, splitting and one-eye set planting practices on plants grown in the field accounted for 45 fold increase in the multiplication rate compared to the traditional methods. These methods combined offer valuable optional strategy for rapid clonal mass propagation of sugarcane.

 

17/21 REGENERATION OF GUS-TRANSFORMANT PLANTS FROM CELL SUSPENSION OF LICORICE GLYCYRRHIZA GLABRA

N. Mousa, P. Siaguru, S. Wiryowidagdo*, O.M. Badawy** and M.E. Wagih

Biotechnology Centre, University of Technology, Lae, Papua New Guinea

*Department of Pharmacology, University of Hasanuddin, Makassar, Indonesia

**Department of Breeding and Genetics, Agriculture ResearchStation (Sabahia), Sugar Crops Research Institute, Agriculture Research Centre, Alexandria, Egypt

Actively-dividing embryogenic cell suspension (ES) of a low-in-Peroxidase cell line (L58) of Glycyrrhiza glabra was initiated from embryogenic callus (EC) originated from coleoptiles of one week-old in vitro plantlets of sprouted seeds. The EC was developed on modified Gamborg’s B5 medium supplemented with 2,4-D, and Kinetin, (B5DK), at concentrations of 1.0 and 0.1 mg L-1, respectively, and the ES of EC was developed in liquid B5DK medium under orbital shaking at 120 rpm with 6cm stroke for 4 weeks in the dark at 25 ± 2 C°. Under the same conditions, regenerative ECs of 120 cell lines (CLs) were developed and maintained on B5DK medium, and subcultured at 2 weeks interval. Morphogenesis of CLs and regeneration of plantlets from EC was achieved on B5 medium and under illumination of 350 µmol E m-2 s-1 for 16 hrs. Homogenous cell suspension at a yield of 6.3 x 106 ml-1 from the ES of L58 was used for the standardization of heat shock-aided direct DNA transformation system. The treated cell suspensions successfully allowed for plant regeneration and efficient expression of the GUS reporter gene in the shoot and root tissues. Having the process completed in almost 50 weeks, the system developed should be qualified as a model for successful genetic transformation of G. glabra for the engineering of its secondary metabolites leading to possible enhancement of the production of the important secondary metabolite “Glycyrrhizin” in cell suspension.

18/21 BIOLOGICAL AND CHEMICAL STUDY OF NIGELLA SATIVA L. SEEDS. II

H.A. Kadry, S.E. El-Dondaity and A. I. Muhammad.

Department of Pharmacognosy, Faculty of Pharmacy (Boys)

Al-Azhar University, Cairo, Egypt.

The acute toxicity (LD50) of 70 % alcohol extract of expressed Nigella sativa L. seeds (marc) is 9.29 g/kg (mice) with fiducial limit of 8.13 and 10.63 g/kg (mice) which revealed the safety margin of the drug. Nigella sativa L. expressed seeds oil was more effective than placebo and 0.2% flumethasone pivalate in management of induced eczema in mice. In treatment of chronic eczema in humans, Nigella sativa L. expressed seeds oil and 0.2% w/w of flumethasone pivalate ointment are statistically comparable and show a significant difference from placebo. Subjectively, it appears that, the Nigella sativa L. seeds oil is better than 0.2% w/w of flumethasone pivalate ointment. In treatment of benign prostatic hyperplasia (BPH), Nigella sativa L. expressed seeds oil and LSE-Sr (Lipido-sterolic extract of Serenoa repens) are statistically comparable and show a high significant difference from the placebo. Subjectively, it appears that Nigella sativa L. expressed seeds oil is better than LSE-Sr (lipido-sterolic extract of Serenoa repens), which revealed the clinical efficacy of the Nigella sativa L. expressed seeds oil. Chemical study of n-butanol fraction of seeds marc lead to isolation and identification of kaempferol, astragallin, a-hederin, 3-O-[β-D-xylopyranosyl (1®3) -a-L-rhamnopyranosyl (1® 2) -a-L-arabinosyl) hederagenin, 3-O-[b-D-xylopyranosyl-(1®3)-a-L-rhamnopyranosyl-(1-2)-a-L-arabinosyl]-28-O-[a-L rhamno-pyranosyl(1®4)b-D-glucopyranosyl-(1®6)-b-D-glucopyronosyl] hederagenin and kaempferol-3-O-b-D-gluco-pyranosyol (1®2)-b-D-glucopyranosyol (1®2)-b-D-glucopyranoside.

19/21 INULINASE PRODUCTION BY KLUYVEROMYCES MARXIANUS NRRL Y-8281 GROWN ON CHICORY JUICE

A.E. Mahmoud

Biochemistry Department, National Research Centre, Dokki, Cairo, Egypt

Kluyveromyces marxianus NRRL Y-8281 was the most potent yeast strain among five yeast strains grown on either synthetic medium containing inulin as carbon source or chicory juice medium without any additives for producing extracellular inulinase enzyme (E C 3.2.1.7). Growing the yeast K. marxianus NRRL Y-8281 on chicory juice medium was selected for a study of the parameters relevant to the commercial production of inulinase. This yeast produced high levels of extracellular inulinase (38.70 U/ml) with specific activity (133.45 U/mg protein) after 30 h. at pH 6.0 and temperature 35oC in the presence of 1% sucrose with 4% (v/v) inoculum size. The addition of different nitrogen sources and inorganic salts to the chicory juice medium were repressed the inulinase production. The optimum pH for enzyme activity was 5.0 and the enzyme was stable in the pH range from 3.0 to 6.0. The enzyme exhibited optimum temperature at 45oC and full activity of its was retained up to 40oC for 1h. Also, it was heat stable up to 50oC for 1h. with only 25% loss activity. All these conditions make the inulinase from Kluyveromyces marxianus NRRL Y-8281, a potential candidate for industrial enzymatic production of fructose from inulin and chicory juice.

 

20/21 ADAPTIVE TOLERANCE BEHAVIOUR OF TRICHODERMA HAMATUM IN CADMIUM, COPPER AND LEAD HEAVY METALS

S.A. El-Aasar

Botany Department, Faculty of Science, Zagazig University, Egypt.

A strain of Trichoderma hamatum, isolated from rhizosphere soil of cultivated tomato field, has been studied for growth and its tolerance to heavy metals; cadmium, copper and lead. The metabolic activities as; nitrogenous, carbohydrate constituents, oxalic acid, total organic acids and nucleic acid (RNA and DNA) were determined. Adapted and non-adapted T. hamatum, were studied for their tolerance index and metal uptake in the presence of a single metal and in the presence of a combination of the three cations. Moreover, chitinase activity has been assayed for the adapted and non-adapted fungus. Effectiveness of T. hamatum treated with metal ions applied as tomato seed dressing against pathogenic fungus Fusarium oxysporum (the casual agent of tomato wilt disease), revealed that metal adapted T. hamatum showed relatively similar effect as control on the antagonistic activity towards F. oxysporum.

 

21/21 FERMENTATIVE PRODUCTION OF PHYTASE AND ACID PHOSPHATASE FROM A LOCAL ISOLATE OF BACILLUS SP. 48

E.N. Danial; M.A. Abdel-Hadi* and A.M. Aboul-Enein**

* Chemistry of Natural and Microbial Products Department National Research Center, Dokki, Cairo, Egypt.

** Biochemistry Department Faculty of Agriculture Cairo University Egypt.

A bacterial strain that produced dephosphorylating enzymes (phytase and acid phosphatase) was isolated from soil and identified to be a genus of Bacillus sp. 48. Among the various carbon and nitrogen sources tested, xylose and potassium nitrate prove to be optimum for two enzymes production as sole carbon and nitrogen sources respectivety. The effect of inoculum age, inoculume size, initial pH, and incubation temperature were also tested. Phytase appeared in the early stage of cultivation and reached its maximum after 78 hours, while acid phosphatase reached its maximum after 96 hours of cultivation.

 

 

22/21 FLORAL MORPHOLOGY OF SOME TAXA OF   POLYGONACEAE IN EGYPT

M.E. Tantawy; K.A. Hamed and U.I. El-Magliy

Department of Botany, Faculty of Science, Ain Shams University, Cairo, Egypt

The macro- and micro-floral characters of 20 taxa of Polygonaceae (representing 17 species, two subspecies and one variety) were carried out to investigate the different conditions of perianth, the high number and the paired condition of stamens and the varied number of carpels with basal ovule. The obtained macro- and micro-floral criteria revealed three main conditions of perianth viz. trimery, pentamery, and tetramery. The former condition was the fundamental plan through the studied taxa and the other two cases were derived from it. The high number and the paired condition of stamens did not result through dedoublement or splitting. The vascular supply to the ovary whatever the number of carpels in all the studied taxa is the same (one dorsal and two ventral bundles for each carpel). All ventrals were fused forming one prominent ventral cord and this considered a step toward advancement. The obtained macro- and micro-floral data facilitate the construction of key for the studied taxa.

 

 


23/21 PHYLOGENETIC POSITION OF BRASSICA TOURNEFORTII USING RAPD MARKERS

H. Y. Hijazy

Botany Department, Faculty of Science, Zagazig University, Egypt

Based on the recent molecular PCR biotechnology, two evolutionary lineages for Brassica diploid species and its related genera have been proposed. These are (I) the “nigra” lineage and (II) the “rapa/oleracea” lineage. The phylogenetic relationship of Brassica tournefortii species to these two lineages and genetic distance between its related species is not studied by this recent molecular technology. Here random amplified polymorphic DNA (RAPD) markers are used successfully to prove that B. tournefortii is more closely related to the “nigra” lineage than to “rapa” lineage. Also genetic polymorphism and genetic distance between the wild species, B. tournefortii, B.nigra & S.alba and one cultivated species, B.rapa also have been discussed.  

 

24/21 NON-ENZYMATIC METHOD FOR DNA PREPARATION FROM DIFFERENT MICROBIAL STRAINS

A.A. Amara, I.K. Afifi*, M.A. Younis**, M.M. Sharaf and M.S. Shabeb**

Mubarak City for Scientific Research and Technology Applications,

Alexandria, Egypt.

*Faculty of Medicine, Tanta University, Tanta, Egypt.

**Botany Department, Faculty of Science, Aswan, South Valley University.

Aswan. Egypt

The increasing importance of molecular biological techniques in microbial studies have made the efficient extraction and purification of microbial DNA very important. DNA isolation and purification using simple methods is considered being one of the researchers main target. The most efficient methods should result in high DNA quality for further using in various molecular biology applications. This method was used for DNA isolation and purification of thirty-eight microbial strains including five strains of E. coli competent cells are represented by the following strains: XL1-Blue, JM109, LS1298, BL21StarTM(DE) and XL1-Red; eight strains of purple non-sulfur bacteria (Rhodospirillaceae) represented by two isolates of Rhodopseudomonas palustris ASEC12, two isolates of Rhodospirillum rubrum ASEC36, two isolates of Rhodospirillum rubrum ASEC39, and two isolates of Rhodopseudomonas palustris ASEC47; ten strains of Pseudomonas represented by one strain of P. putida GPP104, eight isolates of Pseudomonas aeruginosa, and one isolate of P. putida. Fungi are represented by three isolates of Saccharomyces cerevisiae, two isolates of Candida albicans, three isolates of Candida tropicalis, one isolate of Candida glabrata and one isolate of Aspergillus sp.Blue green algae are represented by three isolates of Anabaena sp, two isolates of Synechocystis sp. The purity of the DNA has been examined by the A260/A280 ratio, agarose gel electrophoresis and PCR. It can be recommended that this protocol can be used in Egypt research institutes working with DNA to reduce, the time including within this protocol steps and secondly, the cost coming from using the expensive enzymatic and DNA isolation kits.

 

25/21 PURIFICATION AND CHARACTERIZATION OF ACTIVE SUBSTANCES WITH PROMISING ANTIBACTERIAL ACTIVITY EXTRACTED FROM MYRTUS COMMUNIS

B.M. Refaat, B.M. Haruon, M.H. El-Sehrawi, A.A. Mira* and Y.A. El-Mrakby

Botany and Microbiology Department, Faculty of Science, Al-Azhar University, Madenit Nasr, Cairo, Egypt

*Faculty of Medicine (For Girls), Al-Azhar University, Madenit Nasr, Cairo, Egypt

Pathogenic bacteria were isolated from certain patients. They were subjected to identification through morphological, physiological and biochemical investigations. They were investigated for their activity in presence of certain antibiotics, the resistant bacterial isolates were subjected to the action of certain plant secondary metabolites. Certain metabolites exhibited antibacterial activities and were subjected to purification and characterization which revealed the structural formula for each metabolite.

 

26/21 COMPARATIVE EFFICACY OF AZADIRACHTA INDICA AND FUNGICIDES AGAINST FUNGAL DETERIORATION ON ARCHAEOLOGICALTURKISH CARPET

N.S. Geweely, Y.E. Zidan* and D.A. Zeid*

Faculty of Science, Botany Department (Microbiology), Cairo University,

Giza, Egypt.

*Faculty of Archaeology, Conservation Department, Cairo University, Giza, Egypt.

In this study, the productivity of the cellulolytic enzymes by deteriorating fungi was tested in absence and presence of the fungicides. Archaeological Turkish carpet dated back at the end of the 17th century at Islamic time, obtained from El-Manyal palas museum in Cairo, Egypt, was treated by 4 different fungicides namely Rizolex 50, Vitavax -200, Diniconazole and botanical fungicide called Azadirachta indica (Neem azal). Three fungal species namely: Aspergillus flavus, A. niger and Cladosporium cladosporioides were isolated from different sites on carpet, the dominant one was Aspergillus flavus. The toxicity of fungicides concentrations against spore germination of isolated fungi revealed that Neem azal is the most effective one in stopping fungal deterioration followed by Diniconazole fungicide which retarded fungal growth, while Rhizolex fungicide was the least effective one. The most tolerant species was Aspergillus flavus, while the most sensitive one was Cladosporium cladosporioides. The loss of initial weight was completely lost at 3% of Diniconazole and Azadirachta indica after three weeks incubation for all tested species. Tensile strength of the biodeteriorating samples was assessed. Aspergillus flavus led to higher loss in tensile strength of the carpet fabric.

27/21 CHEMICAL AND BIOLOGICAL STUDY OF RHAMNUS LYCIOIDES L. ROOTS GROWING IN EGYPT.

H.A. Kadry, S.E. El-Dondaity, A.I. Muhammad.

Department of Pharmacognosy, Faculty of Pharmacy (boys)

Al-Azhar University, Cairo, Egypt.

Chemical study of Rhamnus lycioides L. roots lead to isolation and identification of 11 compounds viz., chrysophanol, physcion, emodin, rhamnazin, rhamnocitrin (7-methoxy kaempferol) kaempferol,lycioidin A (kaempferol-3-O-[2,3,4,-tri-O-acetyl-a-L-rhamnopyranosyl-(1 ® 3) – 2,4,- di-O-acetyl-a-L- rhamnopyranosyl-(1 ® 6) ]-b -D-galactopyranoside),lycioidin B (kaempferol-3-O-[ 3,4,-di-O-acetyl-a-L-rhamnopyranosyl-(1 ® 3) – 2,4,- di-O-acetyl-a-L- rhamnopyranosyl-(1 ® 6) ]-b -D-galactopyranoside),nubigenol catharticin (rhamnocitrin -3-O-[a-L-rhamnopyranosyl- (1 ® 3) – a-L- rhamnopyranosyl-(1 ® 6)]-b -D-galactopyranoside) and kaempferol-3-O-triosides( kaempferol-3-O-[a-L-rhamnopyranosyl-(1 ® 3) –a-L- rhamnopyranosyl-(1 ® 6)]-b -D-galactopyranoside). Their structures were determined by spectral data. This represents the first report for isolation of rhamnazin from Rhamnus lycioides L.growing in Egypt, of lycioidin A, B, catharticin and kaempferol-3-O- rhamninosidefrom Rhamnus lycioides L. roots and of nubigenol from the genus Rhamnus. The LD50 of 70 % alcohol extract of Rhamnus lycioides L. roots revealed that the drug is significantly safe . A double-blind trial comparing different concentrations of ointments prepared from 70 % alcohol extracts of Rhamnus lycioides L. roots with, standard therapy, flumethasone pivalate ointment and a placebo showed that, the extracts of Rhamnus lycioides L. roots were effective in treatment of induced eczema in mice.

 

 

28/21 EFFECT OF CYMBOPOGON CITRATUS L. ESSENTIAL OIL ON THE GROWTH AND MORPHOGENESIS OF SACCHAROMYCES CEREVISIAE

G.A. Helal, M.M. Sarhan, A.N.K. Abu Shahla* and E.K. Abou El-Khair*

Botany Department, Faculty of Science, Zagazig University,

Sharkia Governorate, Egypt.

*Biology Department, Faculty of Science, Al Azhar University - Gaza,

Gaza Strip, Palestine.

Addition of 4.0 ul/ml of C. citratus L. essential oil to the Sabouraud's broth medium completely inhibited growth of Saccharomyces cerevisiae. Using the oil as fumigating agent was highly effective in liquid medium than using it contact. Fumigation by 2.0 ul/ml completely inhibited yeast growth in liquid medium. LM and SEM observation showed morphogenic changes in the fumigated cells including decrease in cell size. Also TEM appeared changes in the ultrastructure of cells including; cell wall, plasma membrane and mitochondria. Oil fumigation increases leakage of ions from yeast cells, decreases lipid content and alters its fatty acid composition. E-citral, a`-Myrcene and z-citral, were the major among 19 components constituting about 75% of this oil.

 

 

29/21 EFFECT OF NICLOSAMIDE ON THE DETOXIFICATION ENZYMES IN SCHISTOSOMA TARGET SNAILS BULINUS TRUNCATUS.

A. Abdalla and M. El-Mogy.

Molecular Biology Department, National Research Centre, Dokki, Cairo, Egypt.

Niclosamide (2`, 5-dichloro-4`-nitrosalicyl-anilide) a widely used molluscicide, is the only commercially available molluscicide for large-scale use in schistosomiasis control programs. This work aimed to evaluate the toxicity of niclosamide (NA) on the fresh water snail Bulinus truncatus as well as its effects on glutathione (GSH) and some detoxification enzymes (glutathione transferase, GST; glutathione reductase, GR; glutathione peroxidase, GPx; catalase, CAT).The 24 h LC50 ofNA was 0.095 mg/L, whereas, 100% mortality was achieved at 0.18 mg/L.The overall results indicated that NA concentration up to 0.4 mg/L did not significantly affect the level of GSH. Whereas, GST activity was significantly increased at any of NA concentration used (0.05, 0.1, 0.2 or 0.4 mg/L). On the other hand, GPx was significantly increased or decreased, while GR activity was insignificantly increased or decreased. The significant increase or decrease in CAT depends on the exposure time and the concentration of NA used. Antisera obtained against the major B. truncatus GST isoenzyme (GST3) do not react with the other B. truncatus GSTs. Western blot analysis using GST3 antisera showed that there was no significant difference in snail GST3 levels ofsnail exposed to different concentrations of NA for different time's intervals.

 

30/21 IMPACT OF PHENOTYPIC VARIATION ON RUBBER DEGRADATION AND METAL RESISTANCE IN GORDONIA WESTFALICA VARIANTS

M.M. Berekaa and H. Hussein*

Environmental Sciences Department, Faculty of Science, Alexandria University, Alexandria, Egypt.

*Environmental Biotechnology Department, Genetic Engineering and Biotechnology Reseach Institute (GEBRI), Mubark City for Scientific Research & Technology Applications, New Burg El-Arab City, Alexandria, Egypt.

            The change in colony morphology was recognized among a member of the nocardioform actinomycete, Gordonia wesfalica. During growth o wild type on complex LB medium, tw types of colonies were observed, snooth form colonies G. westfalica strain kb1 and rough form colonies G. westfalica strain kb2, the rough form colonies produced in low frequency. While, subsequent cultivation of the cells of the rough colonies gave rough colonies. In contrast to G. westfalica strain Kb1, strain Kb2 showed greater adhesivity to the rubber substrate, this was evident by higher mineralization rate (expressed as % CO2 released/day). The optimum rate of natural rubber (NR) degradation by G. westfalica strain Kb2 and strain Kb1 was recognized after 7 and 25 days of incubation, respectively, with approximately more than 2-fold increase in mineralization efficiency for G. westfalica strain Kb2. While, both variants were able to colonize the rubber substrate, characteristic high molecular weight proteins were enriched during growth of Kb2 variant on natural rubber (NR) granules while, few proteins of lower molecular weight were enriched during growth of Kb1 variant on the granules due to lower colonization efficency. From the minimum inhibitory concentration (MIC) and maximum tolerable concentration (MIC) results, it was found that resistance pattern of both strains to lower concentrations of all investigated metals (around 2 mmole/L of each) was similar. This resistance decreased with the increase of metal concentration more than 2 mmole/L. Furthermore, no growth was observed when each variant was cultivated on LB medium supplemented with more than 4 mmol/L of Ni and Cr. Both variants gave a similar resistance pattern, with little growth, when cultivated on LB medium supplemented with Cu concentrations more than 8 mmole/L. On the other hand, there was clear difference in growth and resistance patterns of each variant in presence of 8 mmole/L of lead. Furthermore, no growth was observed on metal concentrations higher than 10 mmole/L.
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