Pre-harvest host-resistance to Aspergillus infection and aflatoxin B1 contaminations in groundnut (Arachis hypogaea L.) genotypes.

Groundnut (Arachis hypogaea L.) is an important oil crop in the tropical and sub-tropical countries. Pod and seed coat crack-inducing factors favour Aspergillus species infections and aflatoxin B1 (AFB1) contamination of groundnut. Aflatoxin B1 (AFB1), a toxic secondary metabolite of Aspergillus species, remains a global concern due to its human and animal health, and economic impacts. Thus, the study was conducted at Babile in 2018 with the objective to identify groundnut genotypes resistant to pre-harvest fungal infections, aflatoxin contaminations and associated effects in crop physiology. Seventeen advanced groundnut breeding lines including one commercial cultivar (Werer-961), were evaluated using randomized complete block design and completely randomized design under field and with four replications for laboratory experiments, respectively. Aflatoxin B1 analysis was carried out using Enzyme-Linked Immunosorbent Assay (ELISA) kits. Appropriate statistical procedures, including regression, were employed for data analyses. Highly significant (p<0.01) variation existed among the genotypes for A. flavus and A. niger infections, and the AFB1 contamination ranged from 13.98 (G14) to 1990.86 ppb (G12). The more A. flavus infection, the more reduction in harvest yield and seedling vigour. Fortunately, 53 % of the test materials were found to be resistant to AFB1 production, and frighteningly, none of the AFB1 contaminated genotypes were within the acceptable limit of the lenient standard (10 ppb). All in all, the groundnut genotype (G4) was identified as a good source of pre-harvest resistance to A. flavus infection, AFB1 contamination and seedling vigour so that its inclusion in breeding programs is worthwhile utmost, specifically, in the test environment as pathogen-crop-environment interaction is natural. Since the experiment was employed at one location and for only one year, it is suggested to repeat the experiment across multiple locations and over seasons for reliable recommendation.

Bioinformatics study of phytase from Aspergillus niger for use as feed additive in livestock feed.

BACKGROUND: Phytase supplementation in rations can reduce their phytic acid composition in order to enhance their nutritional value. Aspergillus niger is a fungus that can encode phytase. This study aims to determine the characteristics of its DNA sequences and amino acid composition that encode the phytase enzyme, as well as to determine the primer designs. METHOD: This study used gene sequence data and protein-encoding phytase from Aspergillus niger that was collected manually from NCBI and PDB. The data was analyzed using SPDBV and then be aligned using the ClustalW Multiple Alignment features. The phylogenetic tree was built by Mega11 software. Primers were designed from selected candidate sequences that were analyzed. The designed primers were then simulated for PCR using FastPCR and SnapGene software. RESULTS: There are 18 Aspergillus niger phytases in NCBI which is 14.87% of the total Aspergillus. There are 14 Aspergillus niger phytases that have identity above 95%. Aspergillus niger 110. M94550.1 is the closest strain to the PDB template. Candidate sources of phytase genes are Aspergillus niger 110.M94550.1, 48.2.BCMY01000003.1, and 92.JQ654450.1. The primer design has 2 possibilities of self-annealing and high melting temperature on the reverse primer. PCR simulation shows that the primer design can attach completely but still has the possibility of mispriming. CONCLUSION: This study suggests promising results for the future development of phytase enzyme production from Aspergillus niger as a feed additive using genetic engineering to enhance the quality of livestock feed in Indonesia.

Statistical Optimization of Biosurfactant Production from Aspergillus niger SA1 Fermentation Process and Mathematical Modeling.

In this study, we sought to investigate the production and optimization of biosurfactants by soil fungi isolated from petroleum oil-contaminated soil in Saudi Arabia. Forty-four fungal isolates were isolated from ten petroleum oil-contaminated soil samples. All isolates were identified using the internal transcribed spacer (ITS) region, and biosurfactant screening showed that thirty-nine of the isolates were positive. Aspergillus niger SA1 was the highest biosurfactant producer, demonstrating surface tension, drop collapsing, oil displacement, and an emulsification index (E24) of 35.8 mN/m, 0.55 cm, 6.7 cm, and 70%, respectively. This isolate was therefore selected for biosurfactant optimization using the Fit Group model. The biosurfactant yield was increased 1.22 times higher than in the nonoptimized medium (8.02 g/l) under conditions of pH 6, temperature 35°C, waste frying oil (5.5 g), agitation rate of 200 rpm, and an incubation period of 7 days. Model significance and fitness analysis had an RMSE score of 0.852 and a p-value of 0.0016. The biosurfactant activities were surface tension (35.8 mN/m), drop collapsing (0.7 cm), oil displacement (4.5 cm), and E24 (65.0%). The time course of biosurfactant production was a growth-associated phase. The main outputs of the mathematical model for biomass yield were Yx/s (1.18), and µmax (0.0306) for biosurfactant yield was Yp/s (1.87) and Yp/x (2.51); for waste frying oil consumption the So was 55 g/l, and Ke was 2.56. To verify the model's accuracy, percentage errors between biomass and biosurfactant yields were determined by experimental work and calculated using model equations. The average error of biomass yield was 2.68%, and the average error percentage of biosurfactant yield was 3.39%.

Enzymatic synthesis of fructooligosaccharides: From carrot discards to prebiotic juice.

A great volume of carrots is discarded daily worldwide because they do not meet the required shape and size standards. However, they have the same nutritional characteristics as those commercialized, and can be used in different food products. Carrot juice is an excellent matrix for the development of functional foods with prebiotic compounds, such as fructooligosaccharides (FOS). In this work, the production of FOS in situ in carrot juice was evaluated using a fructosyltransferase from Aspergillus niger, produced by solid-state fermentation on carrot bagasse. The enzyme was partially purified 12.5-fold with a total yield of 93 %, and specific activity of 59 U/mg of protein by Sephadex G-105 molecular exclusion chromatography. It was identified by nano LC-MS/MS as a ß-fructofuranosidase with a 63.6 kDa MW and it allowed obtaining a FOS yield of 31.6 % in carrot juice. The result was a prebiotic juice with a final concentration of 32.4 mg/mL of FOS. Using the commercial enzyme Viscozyme L a higher yield of FOS (39.8 %) was obtained in carrot juice, corresponding to a total amount of FOS of 54.6 mg/mL. This circular economy scheme allowed the obtention of a functional juice, that may contribute to improve health of consumers.

Stress-driven metabolites of desert soil fungi.

Microorganisms produce secondary metabolites to survive under stressful conditions. The effect of drought and heat stress on fungi isolated from Arabian desert soil during the hot (ca 40°C) and cool (ca 10°C) seasons was studied using the genome mining approach. The presence of three stress-related genes (calmodulin, polyketide synthase and beta tubulin) was analyzed molecularly using specific primers. The presence of the genes in desert fungi was compared to their antimicrobial (ten bacterial or fungal pathogens) and anticancer (liver, cervical and breast) properties and the production of thermostable enzymes (phytase and xylanase). The genes appeared to be present in the fungal sequence obtained during the summer, while none of the genes were present during winter. Appreciable differences were observed in enzyme activities, with summer activities high and winter low. The antagonistic activities of A. niger were relatively stable and varying, while those of P. chrysogenum were consistently higher in summer than in winter. The presence of the three genes seemed to correlate with the highly antagonistic activities of P. chrysogenum, while A. niger had relatively active winter isolates without any of the genes. The hot season in deserts yields fungal isolates with biological activities useful in biotechnological solutions.

A contrast of Pb(II), Cd(II), and Cu(II) toxicities to Aspergillus niger through biochemical, morphological, and genetic investigations.

The toxicity of metals to microorganisms is highly correlated with the type of metal used. However, the differences in the resistance mechanisms of filamentous fungi to multiple metals remain unclear. In this study, we investigated the responses of Aspergillus niger to three toxic metals, i.e., Pb2+, Cd2+, and Cu2+. Fungal growth and metabolism indices showed that A. niger had a higher tolerance to Pb2+ (>1000 mg L-1) than to Cu2+ (300 mg L-1) and Cd2+ (50 mg L-1). An appropriate Pb2+ concentration (<500 mg L-1) stimulated fungal growth and metabolic activity, whereas Cd2+ and Cu2+ stress showed continuously negative influences on fungal physiological parameters, such as biomass and secretion of oxalic acid. A. niger responded to Pb stress by constructing a new border layer around its cell wall. This pathway was also confirmed using RNA-seq analysis, i.e., the gene encoding cell wall α-1,3-glucan synthase was upregulated. This upregulation subsequently promoted the production of polysaccharides, which are the main components that support fungal cell walls. In contrast, the expression of genes encoding both AAA family ATPase and efflux pump antibiotic resistance proteins for Cd2+ and Cu2+ was significantly downregulated. Therefore, these findings elucidated the relatively complete fungal responses to different metal stresses.

Combined Application of Tacrolimus with Cyproconazole, Hymexazol and Novel {2-(3-R-1H-1,2,4-triazol-5-yl)phenyl}amines as Antifungals: In Vitro Growth Inhibition and In Silico Molecular Docking Analysis to Fungal Chitin Deacetylase.

Agents with antifungal activity play a vital role as therapeutics in health care, as do fungicides in agriculture. Effectiveness, toxicological profile, and eco-friendliness are among the properties used to select suitable substances. Furthermore, a steady supply of new agents with different modes of action is required to counter the well-known potential of human and phyto-pathogenic fungi to develop resistance against established antifungals. Here, we use an in vitro growth assay to investigate the activity of the calcineurin inhibitor tacrolimus in combination with the commercial fungicides cyproconazole and hymexazol, as well as with two earlier reported novel {2-(3-R-1H-1,2,4-triazol-5-yl)phenyl}amines, against the fungi Aspergillus niger, Colletotrichum higginsianum, Fusarium oxysporum and the oomycete Phytophthora infestans, which are notoriously harmful in agriculture. When tacrolimus was added in a concentration range from 0.25 to 25 mg/L to the tested antifungals (at a fixed concentration of 25 or 50 mg/L), the inhibitory activities were distinctly enhanced. Molecular docking calculations revealed triazole derivative 5, (2-(3-adamantan-1-yl)-1H-1,2,4-triazol-5-yl)-4-chloroaniline), as a potent inhibitor of chitin deacetylases (CDA) of Aspergillus nidulans and A. niger (AnCDA and AngCDA, respectively), which was stronger than the previously reported polyoxorin D, J075-4187, and chitotriose. The results are discussed in the context of potential synergism and molecular mode of action.

[99m Tc]Tc-HYNIC-EcgDf21: A defensin short analogue with potential application in infection foci imaging.

Opportunistic infections are a problem of great relevance in public health and the precise detection and localization of infection in the early stages of the disease is of great importance for patient management as well as cost containment. Our proposal seeks to contribute to developing a new agent that meets the needs of diagnosis and follow-up of fungal and bacterial infections, focused on the design of a radiotracer with the potential for recognition of hidden infection foci. Defensins are plant antimicrobial peptides that not only show activity against plant pathogens but also against human ones. A short analogue of EcgDf1 defensin, EcgDf21d (NH2 -ERFTGGHCRGFRRRCFCTKHC-COOH), was labelled through the formation of a 99m Tc-HYNIC complex which was assessed for physicochemical and biological behaviour both in vitro and in vivo. The [99m Tc]Tc-HYNIC-EcgDf21 labelling procedure rendered a single product with remarkably high RCP and stability in the labelling milieu. The Log p value indicated that [99m Tc]Tc-HYNIC-EcgDf21 has a hydrophilic behaviour, confirmed by the biodistribution profiles. The optimal uptake value was obtained for Candida albicans infection model reaching a lesion/muscle ratio of 3, this correlates with in vitro binding studies, and the lesion can be definitely observed in the scintigraphic images.

Molecular detection of Aspergilli from commercial chicken in selected areas of Bangladesh.

Objectives: This study was designed to isolate, identify, and determine the prevalence of Aspergilli in commercial chicken in selected areas of Bangladesh. Materials and Methods: A total of 50 lung samples from suspected dead chickens, comprising broilers (n = 32) and layers (n = 18), aged between 5 days and 45 weeks, were collected from poultry farms located in the Gazipur district in Bangladesh. Fungi were primarily identified based on the colony morphology using potato dextrose agar (PDA). DNA was extracted from the suspected colonies. Aspegillus spp. was detected by genus-specific ASAP-1 and ASAP-2. Aspergillus spp. were then screened by polymerase chain reaction targeting Aspergillus flavus (FLA-1 and FLA-2), Aspergillus fumigatus (ASPU and Af3r), and Aspergillus niger (ASPU and Nilr). Results: The overall prevalence of Aspergillus spp. was 44% (n = 22/50; p < 0.05). Among the Aspergilli, A. flavus was detected in 10% (n = 5/50) of the samples. Similarly, A. fumigatus and A. niger were detected at 26% (n = 13/50) and 8% (n = 4/50) respectively. Three samples were associated with more than one fungus; two fungi (A. flavus and A. niger) were in two samples, and three fungi (A. flavus, A. fumigatus, and A. niger) were in one sample. Conclusion: Isolation and prevalence of Aspergillus spp. in commercial chicken were studied for the first time in Bangladesh.

Secretory Vesicle and Glucoamylase Distribution in Aspergillus niger and Macromorphology in Regions of Varying Shear Stress.

In technical fermentations, filamentous microorganisms are exposed to different forms of mechanical stress, among which shear stress is prevalent in turbulent broths. Whereas small-scale bioreactors allow for realistic turbulent flow field conditions, they are not well-suited to investigate the fungal response to shear stress in more detail, as they only reveal the integral effect of a highly dynamic stress stimulus. Therefore, the widely used model system for producing constant, but rather low shear forces, the parallel plate flow chamber, is extended in this work by adding a backward-facing step (BFS). The BFS induces vortex shedding in the wake of the step and brings out distinct areas of different shear stress levels at the bottom of the chamber where mycelia grow. This allows for a stress-dependent analysis of growing cells using a confocal laser-scanning microscope. As the real stress cannot be measured in the experiment, the wall shear stress is estimated numerically using computational fluid dynamics (CFD). As a first application of the experimental setup, the relative biomass concentration, the relative amount of secretory vesicles and the relative amount of the chosen product glucoamylase produced by the filamentous fungus Aspergillus niger were measured. The obtained area scans show homogeneous mycelia growth in areas of low stress and cloud-like patterns downstream of the predicted flow reattachment length where high shear stress dominates. Quantitative analysis of the time course suggests that the amount of available secretory vesicles inside of A. niger decreases when the shear stress is increased, despite that no significant differences in biomass production could be found. In contrast, the highest level of glucoamylase was reached for intermediate volumetric flow rates, i.e., levels of shear stress.

Assessing the intracellular primary metabolic profile of Trichoderma reesei and Aspergillus niger grown on different carbon sources.

Trichoderma reesei and Aspergillus niger are efficient biological platforms for the production of various industrial products, including cellulases and organic acids. Nevertheless, despite the extensive research on these fungi, integrated analyses of omics-driven approaches are still missing. In this study, the intracellular metabolic profile of T. reesei RUT-C30 and A. niger N402 strains grown on glucose, lactose, carboxymethylcellulose (CMC), and steam-exploded sugarcane bagasse (SEB) as carbon sources for 48 h was analysed by proton nuclear magnetic resonance. The aim was to verify the changes in the primary metabolism triggered by these substrates and use transcriptomics data from the literature to better understand the dynamics of the observed alterations. Glucose and CMC induced higher fungal growth whereas fungi grown on lactose showed the lowest dry weight. Metabolic profile analysis revealed that mannitol, trehalose, glutamate, glutamine, and alanine were the most abundant metabolites in both fungi regardless of the carbon source. These metabolites are of particular interest for the mobilization of carbon and nitrogen, and stress tolerance inside the cell. Their concomitant presence indicates conserved mechanisms adopted by both fungi to assimilate carbon sources of different levels of recalcitrance. Moreover, the higher levels of galactose intermediates in T. reesei suggest its better adaptation in lactose, whereas glycolate and malate in CMC might indicate activation of the glyoxylate shunt. Glycerol and 4-aminobutyrate accumulated in A. niger grown on CMC and lactose, suggesting their relevant role in these carbon sources. In SEB, a lower quantity and diversity of metabolites were identified compared to the other carbon sources, and the metabolic changes and higher xylanase and pNPGase activities indicated a better utilization of bagasse by A. niger. Transcriptomic analysis supported the observed metabolic changes and pathways identified in this work. Taken together, we have advanced the knowledge about how fungal primary metabolism is affected by different carbon sources, and have drawn attention to metabolites still unexplored. These findings might ultimately be considered for developing more robust and efficient microbial factories.

Bioprocess Optimisation for High Cell Density Endoinulinase Production from Recombinant Aspergillus niger.

Endoinulinase gene was expressed in recombinant Aspergillus niger for selective and high-level expression using an exponential fed-batch fermentation. The effects of the growth rate (µ), glucose feed concentration, nitrogen concentration and fungal morphology on enzyme production were evaluated. A recombinant endoinulinase with a molecular weight of 66 kDa was secreted. Endoinulinase production was growth associated at µ> 0.04 h-1, which is characteristic of the constitutive gpd promoter used for the enzyme production. The highest volumetric activity (670 U/ml) was achieved at a growth rate of 93% of µmax (0.07 h-1), while enzyme activity (506 U/ml) and biomass substrate yield (0.043 gbiomassDW/gglucose) significantly decreased at low µ (0.04 h-1). Increasing the feed concentration resulted in high biomass concentrations and viscosity, which necessitated high agitation to enhance the mixing efficiency and oxygen. However, the high agitation and low DO levels (ca. 8% of saturation) led to pellet disruption and growth in dispersed morphology. Enzyme production profiles, product (Yp/s) and biomass (Yx/s) yield coefficients were not affected by feed concentration and morphological change. The gradual increase in the concentration of nitrogen sources showed that, a nitrogen limited culture was not suitable for endoinulinase production in recombinant A. niger. Moreover, the increase in enzyme volumetric activity was still directly related to an increase in biomass concentration. An increase in nitrogen concentration, from 3.8 to 12 g/L, resulted in volumetric activity increase from 393 to 670 U/ml, but the Yp/s (10053 U/gglucose) and Yx/s (0.049 gbiomasDWs/gglucose) did not significantly change. The data demonstrated the potential of recombinant A. niger and high cell density fermentation for the development of large-scale endoinulinase production system.

The chemical profile of activated secondary metabolites by overexpressing LaeA in Aspergillus niger.

Although the mechanisms of regulating secondary metabolism by LaeA remains unclear, the synthesis of many secondary metabolites (SMs) in Aspergilli could be activated by LaeA mutation. In our previous sutdy, RNA-seq data has showed that the transcriptional level of many SM backbone genes could be upregulated by overexpressing LaeA. Herein, we analyzed the chemical profile of activated secondary metabolites in the variant of A. niger FGSC A1279 by overexpressing LaeA (OElaeA). 14 compounds were activated in A. niger FGSC A1279 OElaeA variant in the WATM medium. Chemical workup of organic extracts of the culture broth from the A. niger OElaeA mutant identified three pure compounds, flaviolin, orlandin and kotanin. The structures of these compounds were confirmed by HR-ESIMS, 1D/2D NMR, and computer assisted structure elucidation (CASE). Based on homologous alignment and comparison of literatures, the biosynthetic gene cluster (fla) of flaviolin was identified. The in vivo function of the backbone gene, flaA, encoding a multidomain non-reducing polyketide synthase (SAT-KS-AT-PT-ACP), was verified via gene knockout and chemical analysis. Finally, a biosynthetic model for fungal flaviolin was proposed.

Purification and partial characterization of carbohydrate-recognition protein C-type lectin from Hemifusus pugilinus.

A mannose binding lectin (C-type lectin) was detected in a molluscan snail Hemifusus pugilinus, this lectin molecule was isolated and purified from the plasma using mannose-fixed sepharose CL-4B column affinity chromatography. The purified protein corresponds to the molecular weight of 118 kDa on an SDS-PAGE gel. The divalent cation-dependent nature of the H. pugilinus lectin (Hp-Lec) evidenced through pH and thermal stability analysis using Circular Dichroism (CD) and Surface Plasmon Resonance (SPR) respectively. Functional investigations of the Hp-Lec reveal a broad spectrum of bacterial agglutination activity against wide range of Gram-positive and Gram-negative bacterial strains. Furthermore, Hp-Lec displayed the haemo agglutination activity against vertebrate red blood cells (RBCs) and its titers were recorded. Excitingly, microbial virulent pathogens such as fungal strains tested against the purified Hp-Lec (25 and 50 µg/ml), which exhibits the effective antifungal activity against tested fungal pathogens such as Aspergillus niger and A. flavus.

Characterization and inhibition of four fungi producing citrinin in various culture media.

PURPOSE: This study aimed to investigate the effects of different fermentation conditions (culture medium, temperature, incubation time, pH value and additive) on citrinin production by four fungi. RESULTS: Among the culture media, potato dextrose medium had lowest citrinin production, followed by yeast sucrose medium and monosodium glutamate medium. The lowest citrinin contents were produced by Monascus anka (M. anka) in potato dextrose medium and yeast sucrose medium, Aspergillus oryzae AS3.042 (A. oryzae) produced the lowest citrinin production in monosodium glutamate medium. The optimum fermentation temperatures for citrinin production by Aspergillus niger (A. niger) and Penicillium citrinum (P. citrinum) were at 30 °C, whereas those by M. anka and A. oryzae were at 35 °C. Citrinin synthesis by four fungi were completely inhibited with a pH value of less than 5.4. By adding ethylene diamine tetraacetic acid (EDTA) or triammonium citrate into monosodium glutamate medium, citrinin production by A. oryzae and A. niger were totally inhibited. Ammonium sulfate completely inhibited citrinin production by A. oryzae, M. anka and P. citrinum, and ammonium nitrate completely inhibited citrinin production by A. oryzae. CONCLUSIONS: These results indicated that the suitable fermentation conditions could make considerable contributions to the reduction of citrinin production. This study provided an effective way for decreasing the citrinin production.

Influences of phosphate addition on fungal weathering of carbonate in the red soil from karst region.

Carbonate in soil from karst region is a substantial carbon sink on Earth. Many karst regions are covered by P-deficient soil. This study evaluated the influences of phosphate addition on fungal weathering (by typical phosphate-solubilizing fungus Aspergillus niger) of carbonate in the soil with red color from karst region. Two weathering pathways were recognized, i.e., biochemical and biomechanical deterioration. The biochemical pathway was performed by dissolving carbonate via secreting organic acids. Meanwhile, the dominant organic acid, i.e., oxalic acid, induced the formation of calcium oxalate, which prevented the loss of Ca2+ cations. It was estimated that the ideal carbonate solubilization driven by geological fluorapatite and fungal weathering is up to 3.3% per year, based on the equation of 12 × (RBase + RPSF) × m × (Areal/APSF). Moreover, fungal weathering of carbonate is very sensitive to the solubility of phosphates. Phosphates supply essential P source for the fungal growth and subsequently raise water-soluble P content in the soil. The addition of bioapatite (a variety of natural apatite with relatively high solubility) elevated the value to 4.6% (a ~ 40% enhancement compared with FAp). This research hence elucidated the tight correlation between carbonate weathering and P supply. Inorganic C release driven by P availability and microbial weathering should be addressed in karst region.

New morphological criteria and molecular characterization of black aspergilli aggregate from corn, sorghum and wheat grains.

Corn, sorghum and wheat grains are used as livestock feed in the world. Identification of black aspergilli associated with these grains is necessary to make sure of the safety of the grains because its occurrence is an indicator of mycotoxin production. Forty-five isolates were isolated from the samples collected from Upper Egypt's markets and identified morphologically based on colony color, conidia, stipe and vesicle size and molecularly by using ß-tubulin and calmodulin genes. Isolates were divided into 30 strains of Aspergillus welwitschiae and 15 strains of A. niger. We have found new criteria in the morphological identification of A. welwitschiae as its colony color was black to brown with yellow edge, but in A. niger was black with white edge, also A. welwitschiae sometimes produced finely-to-distinctly roughened brownish conidia on malt extract agar (MEA) media. Thirteen isolates of A. welwitschiae and six of A. niger were recognized as potential producers for ochratoxin A.

Aspergillus niger Spores Are Highly Resistant to Space Radiation.

The filamentous fungus Aspergillus niger is one of the main contaminants of the International Space Station (ISS). It forms highly pigmented, airborne spores that have thick cell walls and low metabolic activity, enabling them to withstand harsh conditions and colonize spacecraft surfaces. Whether A. niger spores are resistant to space radiation, and to what extent, is not yet known. In this study, spore suspensions of a wild-type and three mutant strains (with defects in pigmentation, DNA repair, and polar growth control) were exposed to X-rays, cosmic radiation (helium- and iron-ions) and UV-C (254 nm). To assess the level of resistance and survival limits of fungal spores in a long-term interplanetary mission scenario, we tested radiation doses up to 1000 Gy and 4000 J/m2. For comparison, a 360-day round-trip to Mars yields a dose of 0.66 ± 0.12 Gy. Overall, wild-type spores of A. niger were able to withstand high doses of X-ray (LD90 = 360 Gy) and cosmic radiation (helium-ion LD90 = 500 Gy; and iron-ion LD90 = 100 Gy). Drying the spores before irradiation made them more susceptible toward X-ray radiation. Notably, A. niger spores are highly resistant to UV-C radiation (LD90 = 1038 J/m2), which is significantly higher than that of other radiation-resistant microorganisms (e.g., Deinococcus radiodurans). In all strains, UV-C treated spores (1000 J/m2) were shown to have decreased biofilm formation (81% reduction in wild-type spores). This study suggests that A. niger spores might not be easily inactivated by exposure to space radiation alone and that current planetary protection guidelines should be revisited, considering the high resistance of fungal spores.

Genetic transformation of tropical maize (Zea mays L.) inbred line with a phytase gene from Aspergillus niger.

A full-length cDNA of phyA gene of Aspergillus niger, encoding phytase enzyme, was cloned and expressed in E. coli BL21 cells and assayed for its activity. The phyA cDNA consisted of 1404 bp, which encoded 467 amino acid residues. The phytase activity of purified phytase was 826.33 U/mL. The phyA gene under the control of endosperm-specific promoters was transformed into an Indian maize inbred line, UMI29, using particle bombardment-mediated transformation method to generate transgenic maize plants over-expressing phytase in seeds. PCR and GUS analyses demonstrated the presence of transgenes in T0 transgenic plants and their stable inheritance in the T1 progenies. Three transgenic events expressing detectable level of A. niger phytase were characterized by western blot analysis. Phytase activity of 463.158 U/kg of seed was observed in one of the events, JB-UMI29-Z17/2. The phytase activity of transgenic maize seeds was 5.5- to 7-fold higher than the wild-type UMI29 seeds and, consequently, the seeds had 0.6- to 5-fold higher inorganic phosphorus content.

Production of nitrogen fixing Azotobacter (SR-4) and phosphorus solubilizing Aspergillus niger and their evaluation on Lagenaria siceraria and Abelmoschus esculentus.

AIMS: Current study was aimed to produce nitrogen fixing Azotobacter sp. (SR-4) and phosphorus solubilizers Aspergillus niger (A. niger) and to evaluate their efficiency as biofertilizers for agricultural practices. METHODS: Two biofertilizer including nitrogen fixing and phosphorus solubilizing were grown. The nitrogen fixing efficiency of Azotobacter (SR-4) was determined by Kjeldahl method. Similarly, Vanadomoybdate method was used to measure the soluble phosphorus while Heinonen method was used to analyze concentration of phytase and phosphatase in the cultures. Furthermore, both biofertilizers were tested in a field trail on Lagenaria siceraria (bottle gourd) and Abelmoschus esculentus (okra). RESULTS: The Azotobacter (SR-4) strain was found efficient nitrogen fixer as 35.08 mg of nitrogen per gram of carbon was produced after 72 h of fermentation. Similarly, A. niger strain excrete extracellular phosphate solubilizing enzymes such as phytase (133UI in 48 h of fermentation) and phosphatase (170UI in 48 h of fermentation) which can solubilize the rock phosphate and make it available to plants. In field trials on selected plants (L. siceraria and A. esculentus), both biofertilizers showed significant increase in plant height, leaf length/width, fruit size and number of fruits per plant when compared with controls/untreated plants. Furthermore, plants co-inoculated with both the N fixing Azotobacter and phosphorus solubilizing A. niger have enhanced performance than those treated with each biofertilizer alone. CONCLUSION: The inoculation of seeds with A. niger and Azotobacter may replace costly and environment toxic chemical fertilizers with environment friendly and cost effective biofertilizers.