Foliar Application of Rhizobium leguminosarum bv. viciae Strain 33504-Borg201 Promotes Faba Bean Growth and Enhances Systemic Resistance Against Bean Yellow Mosaic Virus Infection.

The bean yellow mosaic virus (BYMV) is one of the most serious economic diseases affecting faba bean crop production. Rhizobium spp., well known for its high nitrogen fixation capacity in legumes, has received little study as a possible biocontrol agent and antiviral. Under greenhouse conditions, foliar application of molecularly characterized Rhizobium leguminosarum bv. viciae strain 33504-Borg201 to the faba bean leaves 24 h before they were infected with BYMV made them much more resistant to the disease while also lowering its severity and accumulation. Furthermore, the treatment promoted plant growth and health, as evidenced by the increased total chlorophyll (32.75 mg/g f.wt.) and protein content (14.39 mg/g f.wt.), as well as the improved fresh and dry weights of the plants. The protective effects of 33504-Borg201 greatly lowered the levels of hydrogen peroxide (H2O2) (4.92 µmol/g f.wt.) and malondialdehyde (MDA) (173.72 µmol/g f.wt.). The antioxidant enzymes peroxidase (1.58 µM/g f.wt.) and polyphenol oxidase (0.57 µM/g f.wt.) inhibited the development of BYMV in plants treated with 33504-Borg201. Gene expression analysis showed that faba bean plants treated with 33504-Borg201 had higher amounts of pathogenesis-related protein-1 (PR-1) (3.28-fold) and hydroxycinnamoyl-CoA quinate hydroxycinnamoyltransferase (4.13-fold) than control plants. These findings demonstrate the potential of 33,504-Borg201 as a cost-effective and eco-friendly method to protect faba bean plants against BYMV. Implementing this approach could help develop a simple and sustainable strategy for protecting faba bean crops from the devastating effects of BYMV.

Enhancing the Antibiotic Production by Thermophilic Bacteria Isolated from Hot Spring Waters via Ethyl Methanesulfonate Mutagenesis.

Antibiotic-resistant bacteria represent a serious public health threat. For that reason, the development of new and effective antibiotics to control pathogens has become necessary. The current study aims to search for new microorganisms expressing antibiotic production capacity. Fifteen sites covering a wide range of harsh environmental conditions in Egypt were investigated. Two hundred and eighty bacterial isolates were obtained and then tested against pathogenic bacteria using the agar disk diffusion technique. Fifty-two (18.6% of the total) of the isolates exhibited antagonistic properties, which affected one or more of the tested pathogens. The isolate 113 was identified as Bacillus licheniformis and isolate 10 was identified as Brevibacillus borstelensis using the 16S rRNA technique. The B. licheniformis strain was stronger in antibiotic production against S. typhi, M. luteus, and P. ariginosa, whereas the strain Br. borstelensis was more efficient against B. cereus, E. coli, and Klebs. sp. The sensitivity of the strains to commercial antibiotics showed that B. licheniformis was highly sensitive to seven commercial antibiotics, whereas Br. borstelensis was sensitive to nine antibiotics. The two strains were subjected to ethyl methanesulfonate (EMS) mutagenesis to obtain mutants with a higher antibiotic production. The total bacterial count was measured after treatment with EMS mutagen and showed a significant gradual increase in the antimicrobial activity, which was achieved via shaking in the presence of EMS for 60 min. High antimicrobial activities were noted with 17 and 14 mutants from the B. licheniformis and Br. borstelensis strains, respectively. The mutant B. licheniformis (M15/Amo) was more active than the parent strain against S. aureus (212.5%), while the mutant Br. borstelensis (B7/Neo) was more effective against S. typhi (83.3%). The present study demonstrates the possibility of obtaining potent antibiotic-producing bacteria in hot spring waters and further improving the indigenous bacterial capacity to produce antibiotics by using EMS mutagenesis.

Protective Activity of Rhizobium leguminosarum bv. viciae Strain 33504-Mat209 against Alfalfa Mosaic Virus Infection in Faba Bean Plants.

The application of Rhizobium spp., nitrogen-fixing plant growth-promoting rhizobacteria, as biocontrol agents to enhance systemic disease resistance against plant viral infections is a promising approach towards achieving sustainable and eco-friendly agriculture. However, their potential as antivirals and biocontrol agents is less studied. Herein, the capability of Rhizobium leguminosarum bv. viciae strain 33504-Mat209 was evaluated to promote plant growth and enhance faba bean systemic resistance against alfalfa mosaic virus (AMV) infection. Under greenhouse conditions, the soil inoculation with 3504-Mat209 resulted in notable improvements in growth and an increase in chlorophyll content. This led to a marked decrease in the disease incidence, severity, and viral accumulation level by 48, 74, and 87%, respectively. The protective effect of 33504-Mat209 was linked to significant decreases in non-enzymatic oxidative stress indicators, specifically H2O2 and MDA. Additionally, there were significant increases in the activity of reactive oxygen species scavenging enzymes, such as peroxidase (POX) and polyphenol oxidase (PPO), compared to the virus treatment. The elevated transcript levels of polyphenolic pathway genes (C4H, HCT, C3H, and CHS) and pathogenesis-related protein-1 were also observed. Out of 18 detected compounds, HPLC analysis revealed that 33504-Mat209-treated plants increased the accumulation of several compounds, such as gallic acid, chlorogenic acid, catechin, pyrocatechol, daidzein, quercetin, and cinnamic acid. Therefore, the ability of 33504-Mat209 to promote plant growth and induce systemic resistance against AMV infection has implications for utilizing 33504-Mat209 as a fertilizer and biocontrol agent. This could potentially introduce a new strategy for safeguarding crops, promoting sustainability, and ensuring environmental safety in the agricultural sector. As far as we know, this is the first study of biological control of AMV mediated by Rhizobium spp. in faba bean plants.

Ocimum basilicum-Mediated Synthesis of Silver Nanoparticles Induces Innate Immune Responses against Cucumber Mosaic Virus in Squash.

Cucumber mosaic virus (CMV) causes a significant threat to crop output sustainability and human nutrition worldwide, since it is one of the most prevalent plant viruses infecting most kinds of plants. Nowadays, different types of nanomaterials are applied as a control agent against different phytopathogens. However, their effects against viral infections are still limited. In the current study, the antiviral activities of the biosynthesized silver nanoparticles (Ag-NPs) mediated by aqueous extract of Ocimum basilicum against cucumber mosaic virus in squash (Cucurbita pepo L.) were investigated. The prepared Ag-NPs were characterized using scanning electron microscopy (SEM), dynamic light scattering (DLS), transmission electron microscopy (TEM), energy-dispersive X-ray spectroscopy (EDX), Fourier transform infrared spectroscopy (FTIR) and zeta potential distribution techniques. DLS, SEM, and TEM analyses showed that the Ag-NPs were spherical, with sizes ranging from 26.3 to 83 nm with an average particle size of about 32.6 nm. FTIR identified different functional groups responsible for the capping and stability of Ag-NPs. The zeta potential was reported as being -11.1 mV. Under greenhouse conditions, foliar sprays of Ag-NPs (100 µg/mL) promoted growth, delayed disease symptom development, and significantly reduced CMV accumulation levels of treated plants compared to non-treated plants. Treatment with Ag-NPs 24 h before or after CMV infection reduced CMV accumulation levels by 92% and 86%, respectively. There was also a significant increase in total soluble carbohydrates, free radical scavenging activity, antioxidant enzymes (PPO, SOD, and POX), as well as total phenolic and flavonoid content. Furthermore, systemic resistance was induced by significantly increasing the expression levels of pathogenesis-related genes (PR-1 and PR-5) and polyphenolic pathway genes (HCT and CHI). These findings suggest that Ag-NPs produced by O. basilicum could be used as an elicitor agent and as a control agent in the induction and management of plant viral infections.

Enhancing systemic resistance in faba bean (Vicia faba L.) to Bean yellow mosaic virus via soil application and foliar spray of nitrogen-fixing Rhizobium leguminosarum bv. viciae strain 33504-Alex1.

Rhizobium spp. manifests strong nitrogen fixation ability in legumes. However, their significance as biocontrol agents and antivirals has rarely been investigated. Under greenhouse conditions, the molecularly identified nitrogen-fixing plant growth-promoting rhizobacteria (PGPR), Rhizobium leguminosarum bv. viciae strain 33504-Alex1, isolated from the root nodules of faba bean plants, was tested as a soil inoculum or a foliar application to trigger faba bean plants' resistance against Bean yellow mosaic virus (BYMV) infection. Compared to the non-treated faba bean plants, the applications of 33504-Alex1 in either soil or foliar application significantly promoted growth and improved total chlorophyll content, resulting in a considerable reduction in disease incidence and severity and the inhibition index of BYMV in the treated faba bean plants. Furthermore, the protective activities of 33504-Alex1 were associated with significant reductions in non-enzymatic oxidative stress markers [hydrogen peroxide (H2O2) and malondialdehyde (MDA)] and remarkably increased DPPH free radical scavenging activity and total phenolic content compared to the BYMV treatment at 20 days post-inoculation. Additionally, an increase in reactive oxygen species scavenging enzymes [superoxide dismutase (SOD) and polyphenol oxidase (PPO)] and induced transcriptional levels of pathogenesis-related (PR) proteins (PR-1, PR-3, and PR-5) were observed. Of the 19 polyphenolic compounds detected in faba bean leaves by high-performance liquid chromatography (HPLC) analysis, gallic and vanillic acids were completely shut down in BYMV treatment. Interestingly, the 33504-Alex1 treatments were associated with the induction and accumulation of the most detected polyphenolic compounds. Gas chromatography-mass spectrometry (GC-MS) analysis showed hexadecanoic acid 2,3-dihydroxypropyl ester, tetraneurin-A-Diol, oleic acid, and isochiapin B are the major compounds in the ethyl acetate extract of 33504-Alex1 culture filtrate (CF), suggesting it acts as an elicitor for the induction of systemic acquired resistance (SAR) in faba bean plants. Consequently, the capacity of R. leguminosarum bv. viciae strain 33504-Alex1 to enhance plant growth and induce systemic resistance to BYMV infection will support the incorporation of 33504-Alex1 as a fertilizer and biocontrol agent and offer a new strategy for crop protection, sustainability, and environmental safety in agriculture production.

Induction of Systemic Resistance to Tobacco mosaic virus in Tomato through Foliar Application of Bacillus amyloliquefaciens Strain TBorg1 Culture Filtrate.

The application of microbe-derived products as natural biocontrol agents to boost systemic disease resistance to virus infections in plants is a prospective strategy to make agriculture more sustainable and environmentally friendly. In the current study, the rhizobacterium Bacillus amyloliquefaciens strain TBorg1 was identified based on 16S rRNA, rpoB, and gyrA gene sequences, and evaluated for its efficiency in conferring protection of tomato from infection by Tobacco mosaic virus (TMV). Under greenhouse circumstances, foliar sprays of TBorg1 culture filtrate (TBorg1-CF) promoted tomato growth, lowered disease severity, and significantly decreased TMV accumulation in systemically infected leaves of treated plants relative to untreated controls. TMV accumulation was reduced by 90% following the dual treatment, applied 24 h before and after TMV infection. Significant increases in levels of total soluble carbohydrates, proteins, and ascorbic acid were also found. In addition, a significant rise in activities of enzymes capable of scavenging reactive oxygen species (PPO and POX), as well as decreased levels of non-enzymatic oxidative stress markers (H2O2 and MDA) were observed, compared to untreated plants. Enhanced systemic resistance to TMV was indicated by significantly increased transcript accumulation of polyphenolic pathway (C4H, HCT, and CHI) and pathogenesis-related (PR-1 and PR-5) genes. Out of the 15 compounds identified in the GC-MS analysis, 1,2-benzenedicarboxylic acid mono(2-ethylhexyl) ester and phenol, 2,4-bis(1,1-dimethylethyl), as well as L-proline, N-valeryl-, and heptadecyl ester were present in the highest concentrations in the ethyl acetate extract of TBorg1-CF. In addition, significant amounts of n-hexadecanoic acid, pyrrolo [1,2-a] pyrazine-1,4-dione hexahydro-3-(2-methylpropyl)-, nonane, 5-butyl-, and eicosane were also detected. These compounds may act as inducers of systemic resistance to viral infection. Our findings indicate that the newly isolated B. amyloliquefaciens strain TBorg1 could be a potentially useful rhizobacterium for promoting plant growth and a possible source of biocontrol agents for combating plant virus infections.

Protective and Curative Activities of Paenibacillus polymyxa against Zucchini yellow mosaic virus Infestation in Squash Plants.

The use of microbial products as natural biocontrol agents to increase a plant's systemic resistance to viral infections is a promising way to make agriculture more sustainable and less harmful to the environment. The rhizobacterium Paenibacillus polymyxa has been shown to have strong biocontrol action against plant diseases, but its antiviral activity has been little investigated. Here, the efficiency of the culture filtrate of the P. polymyxa strain SZYM (Acc# ON149452) to protect squash (Cucurbita pepo L.) plants against a Zucchini yellow mosaic virus (ZYMV, Acc# ON159933) infection was evaluated. Under greenhouse conditions, the foliar application of the culture filtrate of SZYM either in protective or curative treatment conditions enhanced squash growth, reduced disease severity, and decreased ZYMV accumulation levels in the treated plants when compared to the non-treated plants. The protective treatment group exhibited the highest inhibitory effect (80%), with significant increases in their total soluble carbohydrates, total soluble protein content, ascorbic acid content, and free radical scavenging activity. Furthermore, a considerable increase in the activities of reactive oxygen species scavenging enzymes (superoxide dismutase, polyphenol oxidase, and peroxidase) were also found. In addition, the induction of systemic resistance with a significant elevation in the transcriptional levels of polyphenolic pathway genes (CHS, PAL, and C3H) and pathogenesis-related genes (PR-1 and PR-3) was observed. Out of the 14 detected compounds in the GC-MS analysis, propanoic acid, benzenedicarboxylic acid, tetradecanoic acid, and their derivatives, as well as pyrrolo [1,2-a] pyrazine-1,4-dione, hexahydro-3-(2-methylpropyl) were the primary ingredient compounds in the ethyl acetate extract of the SZYM-culture filtrate. Such compounds may act as elicitor molecules that induce systemic resistance against viral infection. Consequently, P. polymyxa can be considered a powerful plant growth-promoting bacterium (PGPB) in agricultural applications as well as a source of bioactive compounds for sustainable disease management. As far as we know, this is the first time that P. polymyxa has been shown to fight viruses in plants.

Rhizobium leguminosarum bv. viciae-Mediated Silver Nanoparticles for Controlling Bean Yellow Mosaic Virus (BYMV) Infection in Faba Bean Plants.

The faba bean plant (Vicia faba L.) is one of the world's most important legume crops and can be infected with various viral diseases that affect its production. One of the more significant viruses in terms of economic impact is bean yellow mosaic virus (BYMV). The current study used the molecularly identified Rhizobium leguminosarum bv. viciae strain 33504-Borg1, a nitrogen-fixing bacteria, to biosynthesize silver nanoparticles (AgNPs) to control BYMV disease in faba bean plants. Scanning electron microscopy (SEM), a particle size analyzer (PSA) with dynamic light scattering (DLS), transmission electron microscopy (TEM), energy dispersive X-ray spectroscopy (EDX), and Fourier transform infrared spectroscopy (FTIR) were used to characterize the prepared AgNPs. The DLS, SEM, and TEM analyses revealed that the AgNPs were spherical and rough, with sizes ranging from 13.7 to 40 nm. The FTIR analysis recognized various functional groups related to AgNP capping and stability. Under greenhouse conditions, spraying faba bean leaves with the AgNPs (100 µg/mL) 24 h before BYMV inoculation induced plant resistance and reduced plant disease severity and virus concentration levels. Contrarily, the AgNP treatment enhanced plant health by raising photosynthetic rates, increasing the fresh and dry weight of the faba bean plants, and increasing other measured metrics to levels comparable to healthy controls. Antioxidant enzymes (peroxidase and polyphenol oxidase) inhibited the development of BYMV in the faba bean plants treated with the AgNPs. The AgNPs decreased oxidative stress markers (H2O2 and MDA) in the faba bean plants. The plants treated with the AgNPs showed higher expression levels of PR-1 and HQT than the control plants. The study findings could be used to develop a simple, low-cost, and environmentally friendly method of protecting the faba bean plant from BYMV.

Potential of tailored amorphous multiporous calcium silicate glass for pulp capping regenerative endodontics-A preliminary assessment.

INTRODUCTION/OBJECTIVE: The tailored amorphous multi-porous (TAMP) material fabrication technology has led to a new class of bioactive materials possessing versatile characteristics. It has not been tested for dental applications. Thus, we aimed to assess its biocompatibility and ability to regenerate dental mineral tissue. METHODS: 30CaO-70SiO2 model TAMP discs were fabricated by a sol-gel method followed by in vitro biocompatibility testing with isolated human or mini-swine dental pulp stem cells (DPSCs). TAMP scaffolds were tested in vivo as a pulp exposure (pin-point, 1 mm, 2 mm, and entire pulp chamber roof) capping material in the molar teeth of mini-swine. RESULTS: The in vitro assays showed that DPSCs attached well onto the TAMP discs with comparable viability to those attached to culture plates. Pulp capping tests on mini-swine showed that after 4.5 months TAMP material was still present at the capping site, and mineral tissue (dentin bridge) had formed in all sizes of pulp exposure underneath the TAMP material. CONCLUSIONS: TAMP calcium silicate is biocompatible with both human and swine DPSCs in vitro and with pulp in vivo, it may help regenerate the dentin bridge after pulp exposure.

Process development for scale-up production of a therapeutic L-asparaginase by Streptomyces brollosae NEAE-115 from shake flasks to bioreactor.

L-asparaginase is a promising enzyme that has a wide range of significant applications including cancer therapy and starchy food industries. The statistical design of Plackett-Burman and face centered central composite design were employed to optimize L-asparaginase production by Streptomyces brollosae NEAE-115. As a result, a medium of the following formula is the optimum for producing L-asparaginase in the culture filtrate of Streptomyces brollosae NEAE-115: Dextrose 2 g, starch 20 g, L-asparagine 10 g, KNO3 1 g, K2HPO4 1 g, MgSO4.7H2O 0.5 g, NaCl 0.1 g, pH 7, fermentation period 7 days, temperature 30 °C, inoculum size 4%, v/v, agitation speed 150 rpm and inoculum age 48 h. The kinetics of cell growth, carbohydrates consumption and L- asparaginase production were studied in 7-L stirred tank bioreactor under different cultivation conditions. A significant increase in both cell growth and carbohydrate consumption was observed as the stirring speed increased from 200 to 600 rpm under uncontrolled pH. The highest L- asparaginase activity of 108.46 U/mL was obtained after 96 h at 400 rpm. On the other hand, the specific enzyme production (Yp/x) under uncontrolled pH reached its maximal value of about 20.3 U/mg cells. Further improvement of enzyme production was attained by controlling pH at 7 using the selected stirring speed of 400 rpm. Enzyme production of 162.11 U/mL obtained from the controlled pH cultures exceeded this value gained from uncontrolled pH (108.46 U/mL) by about 50%.

Optimization of conditions for decolorization of azo-based textile dyes by multiple fungal species.

Wastewater from textile industries contains azo dye residues that negatively affect most environmental systems. The biological treatment of these wastes is the best option due to safety and cost concerns. Here we isolated and identified 19 azo dye-degrading fungi and optimized conditions resulting in enhanced degradation. The fungi belonged to five species of Aspergillus and a single Lichtheimia sp. All fungi were evaluated for their ability to decolorize 20 azo dyes. While the most easily transformable azo dye was direct violet (decolorization ranged from 71.1 to 93.3%), the most resistant to decolorization was fast green azo dye. The greatest degradation potential of azo dyes (direct violet and methyl red) was optimized using the most promising four fungal strains and changing media glucose concentration, nitrogen source, and micronutrients. Biomass, lignin peroxidase, and laccases production were also determined in the optimization studies. The decolorization of both azo dyes by the four fungal strains was greatly enhanced by glucose supplementation. The fungal strains were not able to produce lignin peroxidases in the absence of organic nitrogen source. Both yeast extract and casamino acid supplementation enhanced decolorization of direct violet and methyl red dyes and production of lignin peroxidase by the fungal strains. In contrast, the laccases were absent in the similar medium enriched with the same organic nitrogen sources.

The symbiovar trifolii of Rhizobium bangladeshense and Rhizobium aegyptiacum sp. nov. nodulate Trifolium alexandrinum in Egypt.

In the present work we analyzed the taxonomic status of several Rhizobium strains isolated from Trifolium alexandrinum L. nodules in Egypt. The 16S rRNA genes of these strains were identical to those of Rhizobium bangladeshense BLR175(T) and Rhizobium binae BLR195(T). However, the analyses of recA and atpD genes split the strains into two clusters. Cluster II strains are identified as R. bangladeshense with >98% similarity values in both genes. The cluster I strains are phylogenetically related to Rhizobium etli CFN42(T) and R. bangladeshense BLR175(T), but with less than 94% similarity values in recA and atpD genes. DNA-DNA hybridization analysis showed 42% and 48% average relatedness between the strain 1010(T) from cluster I with respect to R. bangladeshense BLR175(T) and R. etli CFN42(T), respectively. Phenotypic characteristics of cluster I strains also differed from those of their closest related Rhizobium species. Analysis of the nodC gene showed that the strains belong to two groups within the symbiovar trifolii which was identified in Egypt linked to the species R. bangladeshense. Based on the genotypic and phenotypic characteristics, the group I strains belong to a new species for which the name Rhizobium aegyptiacum sp. nov. (sv. trifolii) is proposed, with strain 1010(T) being designated as the type strain (= USDA 7124(T)=LMG 29296(T)=CECT 9098(T)).

High cell density cultivation of six fungal strains efficient in azo dye bioremediation.

This work aims at optimizing the high cell density fungal cultivation for producing large quantities of fungal biomass to be used in azo dye residues bioremediation. In our previous studies the efficacy of using certain fungal strains to decolorize a range of commercial textile dyes of different structures (azo, disazo) were investigated. Several promising fungal strains belonging to Aspergillus tubigenesis, Aspergillus niger, Aspergillus terreus, and Aspergillus fumigates demonstrated high capacity in decolorizing various azo dyes. This study focuses on the high cell density cultivation of the fungal strains identified as potential bioremediation agents. The study includes the optimization of all parameters involved in bioprocess development for high cell density cultivation of six promising fungal strains. The growth of the fungal strains was tested on the sucrose medium in 7 l-fermenter. The growth of these fungal strains having the capacity to accumulate large quantities of biomass was also tested in medium containing molasses as a cheap substrate. The residual molasses, biomass dry weight and protein content of the six fungal strains showed that the strains 20 and 2 were marked by the highest protein content. In this study a comparative analysis between the results of dry weight, residual molasses and protein content of geowth of the strains 20, 5 and 2 under uncontrolled and controlled pH of media in batch fermentation was studied to follow the accumulation of biomass and protein production in the growth media. The results indicate that the dry weight accumulated by strains No. 20, 5 and 2 grown on molasses was better than those of strains grown on sucrose. Fungal strain No. 5 had the highest biomass dry weight accumulation. The study shows that the molasses as cheaper sugar sources were better than sucrose for growing fungal biomass.

Optimization of Culture Conditions for Production of the Anti-Leukemic Glutaminase Free L-Asparaginase by Newly Isolated Streptomyces olivaceus NEAE-119 Using Response Surface Methodology.

Among the antitumor drugs, bacterial enzyme L-asparaginase has been employed as the most effective chemotherapeutic agent in pediatric oncotherapy especially for acute lymphoblastic leukemia. Glutaminase free L-asparaginase producing actinomycetes were isolated from soil samples collected from Egypt. Among them, a potential culture, strain NEAE-119, was selected and identified on the basis of morphological, cultural, physiological, and biochemical properties together with 16S rRNA sequence as Streptomyces olivaceus NEAE-119 and sequencing product (1509 bp) was deposited in the GenBank database under accession number KJ200342. The optimization of different process parameters for L-asparaginase production by Streptomyces olivaceus NEAE-119 using Plackett-Burman experimental design and response surface methodology was carried out. Fifteen variables (temperature, pH, incubation time, inoculum size, inoculum age, agitation speed, dextrose, starch, L-asparagine, KNO3, yeast extract, K2HPO4, MgSO4·7H2O, NaCl, and FeSO4·7H2O) were screened using Plackett-Burman experimental design. The most positive significant independent variables affecting enzyme production (temperature, inoculum age, and agitation speed) were further optimized by the face-centered central composite design-response surface methodology.

Near-full length sequencing of 16S rDNA and RFLP indicates that Rhizobium etli is the dominant species nodulating Egyptian winter Berseem clover (Trifolium alexandrinum L.).

Egyptian winter Berseem clover (EWBC) is one of the main important forage legume crops in Egypt that is used for animal feeding in winter and it occupies about 2.5 million feddans (Feddan=4200m(2)) in winter agricultural rotation systems. Forty-eight rhizobial isolates that nodulated this legume host from different geographical regions within Egypt were isolated. RFLP analyses of 16S rDNA (1.5kb) and whole ribosomal DNA (5kb), the sequencing of 16S rDNA, and the sequencing of nodC, nifH and house keeping genes were used to identify these isolates. The RFLP analysis of 16S rDNA (1.5kb) among 15 representative strains with three enzymes generated two genotypes. The largest genotype was similar to Rhizobium etli CFN42T (93.33%) except for strain 902 that failed to re-nodulate EWBC. RFLP analysis of complete ribosomal DNA (5kb) produced five genotypes. The majority of tested strains shared the genotype with R. etli CFN42T (53.33%). Only one strain (1002) shared the genotype with Rhizobium leguminosarum sv. trifolii 3023. The other four strains were comprised of two unique genotypes. Phylogenetic analysis of 16S rDNA sequences revealed that seven representative strains could be divided into two genetic clusters sharing the ancestral clad with R. etli CFN42T. A phylogenetic tree based on nodC gene sequence confirmed that all the examined strains shared the genetic lineage with R. leguminosarum sv. trifolii WSM1325. The phylogenetic trees of house keeping genes are supported strongly the identification of majority of strains as a novel symbiovar of R. etli with new lineages.

Beta-glucanase productivity improvement via cell immobilization of recombinant Escherichia coli cells in different matrices.

The studies have been performed to analyze the production of beta-glucanase by a recombinant strain of Escherichia coli immobilized in different matrices. Porous sintered glass SIRAN, Ceramic supporting matrices and Broken Pumice stone as well as SIRAN Raschig-rings were examined for the immobilization of whole bacterial cells. The beta-glucanase activity of bacteria immobilized in CeramTec PST 5 (4-5 mm) was very low. CeramTec PST 5 (1.5-2.5 mm) was found to be the best carrier compared to all other matrices regarding glucanase production (630 U/ml) and compared to enzyme activity produced by free cells (500 U/ml). Different doses of matrices were applied (2, 5, 7, 10 g/lask) in the form of "matrix weight". Using 2 g/flask of CeramTec PST 5 (1.5-2.5 mm) yielded enzyme activity of 630 U/ml). CeramTec gives highest operational stability of beta-glucanase by repeated batch fermentation to 5 cycles, and activity reached 660 U/ml. Scanning electron microscopy observations showed a high number of vegetative cells that continued growth inside the matrices, indicating that beta-glucanase activity improvement was due to the immobilization of the cells.

Persistence of two Rhizobium etli inoculant strains in clay and silty loam soils.

The persistence of Rhizobium etli strains CE3 and Ph 163 was studied in two soil types representing major French bean growing areas in Egypt. Clay soil from MENOUFIA and silty loam soil from ISMAILIA were planted by bean Cultivars; Bronco and Giza 6. The inoculation with strain Ph 163 in the first bean cultivation was significantly higher in nodule biomass and number; whereas, the strain CE3 was significantly higher in plant biomass accumulation (Moawad et al. 2004). The persisting inocula strains seem to perform differently in the two soils in terms of nodulation, biomass accumulation and N-uptake by the two cultivars as compared with their performance with the first inoculation. CE3 strain persisting in the soil performed better than Ph.163 strain. The nodule occupancy by the persisting inoculant rhizobial was determined by two approaches; fluorescent antibody (FA) technique and other Rep-PCR fingerprinting. Both techniques were close in the evaluation of persisting inoculant strains which nodulated beans in the second planting season without inoculation. The results obtained showed that both strains are good survivors in the two soils.

Performance of phaseolus bean rhizobia in soils from the major production sites in the Nile Delta.

The symbiotic and competitive performances of two highly effective rhizobia nodulating French bean P. vulgaris were studied in silty loam and clayey soils. The experiments were carried out to address the performance of two rhizobia strains (CE3 and Ph. 163] and the mixture thereof with the two major cultivated bean cultivars in two soil types from major growing French bean areas in Egypt. Clay and silty loam soils from Menoufia and Ismailia respectively were planted with Bronco and Giza 6 phaseolus bean cultivars. The data obtained from this study indicated that rhizobial inoculation of Giza 6 cultivar in clayey soil showed a positive response to inoculation in terms of nodule numbers and dry weight. This response was also positive in dry matter and biomass accumulation by the plants. The inoculant of strain CE3 enhanced plant growth and N-uptake relative to Ph. 163. However, the mixed inoculant strains were not always as good as single strain inoculants. The competition for nodulation was assessed using two techniques namely fluorescent antibody testing (FA) and REP-PCR fingerprinting. The nodule occupancy by inoculant strain Ph. 163 in both soils occupied 30-40% and 38-50 of nodules of cultivar Bronco. The mixed inocula resulted in higher proportions of nodules containing CE3 in silty loam soil and Ph. 163 in clayey soil. The native rhizobia occupied at least 50% of the nodules on the Bronco cultivar. For cultivar Giza 6, the native rhizobia were more competitive with the inoculant strains. Therefore, we suggest using the studied strains as commercial inocula for phaseolus bean.

Biosorption of heavy metals from waste water using Pseudomonas sp

Biosorption experiments for Cr(VI), Cu(II), Cd(II) and Ni(II) were investigated in this study using nonliving biomass of different Pseudomonas species. The applicability of the Langmuir and Freundlich models for the different biosorbent was tested. The coefficient of determination (R²) of both models were mostly greater than 0.9. In case of Ni(II) and Cu(II), their coefficients were found to be close to one. This indicates that both models adequately describe the experimental data of the biosorption of these metals. The maximum adsorption capacity was found to be the highest for Ni followed by Cd(II), Cu(II) and Cr(VI). Whereas the Freundlich constant k in case of Cd(II) was found to be greater than the other metals. Maximum Cr(VI) removal reached around 38 percent and its removal increased with the increase of Cr(VI) influent. Cu(II) removal was at its maximum value in presence of Cr(VI) as a binary metal, which reached 93 percent of its influent concentration. Concerning to Cd(II) and Ni(II) similar removal ratios were obtained, since it was ranged between 35 to 88 percent and their maximum removal were obtained in the case of individual Cd(II) and Ni(II).

Enhancing bioremoval of textile dyes by eight fungal strains from media supplemented with gelatine wastes and sucrose.

Eight Aspergillus strains were found to be successful in removing textile dyes from liquid media. These fungal strains were grown on medium containing: gelatine wastes and sucrose, as sources of nitrogen and carbon to test the possible speed up of the dyes removing while fungus biomass is building up in the media. The growth of fungal strains ranged from 10 to 110 mg biomass dry weight/100 ml medium. This growth induced high decolorization percentages, which ranged 33-95% within eight days. Two textile dyes Direct brown and Polar red were included in the study. The growth of the fungal strains as well as decolorization percentage of the dyes increased after 5, 6, and 8 days from incubation time with most tested strains. With Direct brown dye the strains number 2, 5, 31 and 37 recorded the highest percentage of decolorization (91, 92, 93 and 95 respectively) after incubation for 6 days. Fungal strains Aspergillus 5 and 31 gave the highest mycelium dry weight being 110 mg. Most of fungal strains induced 86 to 95 percentage of decolorization after 6 days incubation with Polar red dye. The possible toxicity of the remaining supernatant media after fungal biomass removal was tested by Ames test to assess the residual mutagenic agents remaining after dye removal, using three strains of Salmonella typhimurium (TA 1535, TA 1537, TA 1538). The results showed that the toxicity of the dyes, measured by Ames test could be removed by the dye absorption on the fungal biomass.