Amna Shoaib

Antifungal activity of methanolic extracts of Sorghum halepense against Macrophomina phaseolina

Macrophomina phaseolina (Tassi) Goid., the cause of charcoal rot disease, is the major biotic factor that limits cowpea [Vigna unguiculata (L.) Walp.] productivity worldwide. The present study was designed to investigate the antifungal potential of an allelopathic grass Sorghum halepense Pers. for the management of M. phaseolina isolated from charcoal rot infected cowpea plants. In laboratory bioassays, different concentrations (0, 0.5, 1.0,…, 3.0 g/ml) of methanolic extracts of shoot, root and inflorescence of the test allelopathic grass were evaluated for their in vitro antifungal activity against M. phaseolina. Extracts of all the three parts of the grass exhibited variable antifungal activity. Shoot extract was found the most effective where all the extract concentrations significantly reduced the fungal biomass by 14 to 61% over control. Similarly, all concentrations of the root extract except 0.5% significantly suppressed the fungal biomass. Inflorescence extract exhibited the least antifungal activity where only 2% and higher concentrations showed the significant effect. There was 9 to 48% and 4 to 39% reduction in fungal biomass due to various concentrations of methanolic root and inflorescence extracts, respectively.

ANTIFUNGAL ACTIVITY OF Cirsium arvense EXTRACTS AGAINST PHYTOPATHOGENIC FUNGUS Macrophomina phaseolina

ABSTRACT Leaves, stems, roots and inflorescence of the asteraceous weed Circium arvense were extracted in methanol for two weeks. Methanol was evaporated in a rotary evaporator. Different concentrations (1, 2, 3, 4 and 5%) of methanolic extracts were prepared, and their antifungal activities were studied against Macrophomina phaseolina, using malt extract broth as growth medium. In general, extracts of all plant parts showed antifungal activities to variable extents. The highest antifungal activity occurred due to methanolic extract of leaves, followed by stem and root extracts, resulting in 10-74%, 6-57% and 11-39% reduction in fungal biomass over control, respectively. Inflorescence extract showed the least antifungal activity, resulting in 2-30% reduction in fungal biomass over control. There was a linear and inverse relationship between extract concentrations and fungal biomass for extracts of all the four parts. GC-MS analysis showed that there were 10 compounds in most effective methanolic leaf extract. Among these, 10-octadecanoic acid, methyl ester (26.442%), 2H-1-benzopyran, 6,7-dimethoxy-2-2-dimethyl (20.195%), hexadecanoic acid, methyl ester (15.752%) and 9,12-octadecadienoic acid (Z,Z)-, methyl ester (12.628%) were predominant compounds in the extract that may be responsible for antifungal activity. This study concludes that methanolic leaf extracts of C. arvense can be used for the management of M. phaseolina.

Effect of Incorporation of Leaf Biomass of Coronopus didymus on Management of Basal Rot Disease of Onion and its Physiology

Fusarium oxysporum f. sp. cepae is a soil-borne fungal pathogen that causes basal rot disease of onion (Allium cepa L.) resulting in significant yield losses of the crop. The present study was conducted to assess the disease management potential of Coronopus didymus (L.) Sm., and its effect on onion plant physiology. In laboratory bioassays, different concentrations ranging from 1% to 7% of methanolic extracts of leaves, stems and roots of C. didymus reduced fungal biomass by 60–84%, 51–85% and 73–87%, respectively. In a pot trial, dry leaf biomass of C. didymus was mixed in the soil at 1%, 2% and 3% in combination with F. oxysporum f. sp. cepae and their effect on disease development, plant growth and various physiological parameters was studied in comparison with a negative control (without any addition of pathogen or plant material) and a positive control inoculated only with the fungus. The highest disease incidence (47%) was recorded in positive control. Application of 2% and 3% dry leaf biomass of C. didymus as soil amendment significantly reduced disease incidence to 13% and 3%, respectively. Similarly, plant mortality was reduced to 0% due to 2% as well 3% dry leaf biomass treatments. The highest chlorophyll, sugar and protein content were recorded in treatment where F. oxysporum f. sp. cepae was inoculated with 1% dry leaf biomass incorporation. On the other hand, the highest phenolic content and peroxidase activity were recorded due to combined application of the pathogen and 3% dry biomass incorporation. Present study concludes that soil amendment with 2% leaf dry biomass of C. didymus can effectively manage the basal rot of onion without any adverse effect on plant growth.

Evaluation of antifungal activity of methanolic extracts of Dicanthium annulatum for the management of Macrophomina phaseolina

The present study was carried out to assess the antifungal potential of an allelopathic grass Dicanthium annulatum (Forssk.) Stapf. against Macrophomina phaseolina (Tassi) Goid. isolated from cowpea [Vigna unguiculata (L.) Walp.] plants suffering from charcoal rot disease. Different parts of this grass namely shoot, root and inflorescence were extracted in methanol. After evaporation of methanol, different concentrations (0, 0.5, 1.0, …, 3.0 g ml -1 ) of the extracts were prepared and their antifungal activity was studied. In general, extracts of all the three parts exhibited antifungal activity. However, a marked variation in antifungal activity among the extracts of different parts of the test grass was observed. There was 7 to 51%, 29 to 71% and 33 to 81% reduction in fungal biomass due to different concentrations of shoot, root and inflorescence extracts of D. annulatum, respectively.

Macrophomina phaseolina alters the biochemical pathway in Vigna radiata chastened by Zn2+ and FYM to improve plant growth

ABSTRACT Mung bean [Vigna radiata (L.) Wilczek] is an important cash pulse crop extensively cultivated in the arid region of Pakistan, which encounters intimidating charcoal rot disease caused by Macrophomina phaseolina (Tassi) Goid. The current research was conducted to check the potential of Zn (1.25, 2.44 and 5 mg kg−1) and FYM [farmyard manure (1% and 2%)] in mono-, bi- and trilateral interaction in managing disease and improving yield. Suppression of plant immunity by M. phaseolina was indicated by the change in activities of antioxidant enzymes (CAT and SOD) and cell wall strengthening enzymes (POX and PAL) that revealed inability of the protein receptor to identify the pathogen elicitor. FYM improved soil physicochemical properties and beneficial microbes activity, which released antimicrobial protein- and plant defense-stimulating protein and in response to ROS (reactive oxygen species) signaling molecules plant susceptibility was reduced. However, Zn as a co-factor chastened the ROS in stressed cells by upregulation of antioxidant enzymes in favor of the plant. The complex interaction of FYM and Zn potentially hijacked the further multiplication of pathogen. Finally, soil amendment improved biological attributes and grain yield to profitable farming in terms of harvest index percentage and benefit–cost ratio.

Allelopathy for the Management of Phytopathogens

Plant pathogens including fungi, bacteria, viruses and nematodes are responsible for huge yield losses in many economically important crops. Use of synthetic agrochemicals as soil fumigation, foliar spray or seed dressing is the most popular strategy for the management of plant diseases in the recent days. However, due to adverse effects of these chemicals on health and environment, consumers are currently demanding produce, which is free of these chemicals. Natural compounds derived from plants are more environmentally safe than synthetic chemicals. Many recent studies have shown that allelochemicals can effectively be used for the management of plant pathogens. Members of plant families like Acanthaceae, Amranthaceae, Chenopodiacea, Brassicaceae and Magnoliaceae are famous for their antifungal properties while those of Asteraceae, Poaceae and Papillionaceae are known for their nematicidal properties. These plants secondary metabolites can be exploited for the management of plant pathogens following crop rotation, green manuring and cultivation of allelopathic plants as cover crops or using crude plant extracts. In addition, the structures of novel allelochemicals can be used as analogue for the synthesis of new natural product-based pesticides.

FIRST REPORT OF ASPERGILLUS MINISCLEROTIGENES AS A POSTHARVEST PATHOGEN OF SOYBEAN SEEDS FROM PAKISTAN

In August 2014, seeds of Glycine max in storage rooms at Lahore, Pakistan, were found to be colonized by an unknown fungus. For pathogen isolation, seeds were incubated on moist blotters, developing a green mass of fungal spores. Spores were transferred to Czapek Dox agar medium and incubated at 25±2°C. After 7 days of incubation colonies were velvety, green and floccose, consisting of white vegetative mycelium and acquired the diameter of 3-4 cm. Globose dark brown sclerotia were produced, smaller than A. flavus. Radiate conidial heads were observed that were mostly biseriate however uniseriate heads were also present. Conidiophores were hyaline, coarsely roughened and 0.9-1.2 mm in length. Vesicles were subglobose to globose, 25-40 μm in diameter, while metulae were 8-12 μm and phialides 5-8 μm long. Conidia were pale or olive green, 3.5-5 μm in diameter. Pathogen was identified as Aspergillus minisclerotigenes (FCBP1353) (Pildain et al., 2008) and differentiated from A. flavus and A. parvisclerotigenus by studying aflatoxin profiles (B1, B2, G1, and G2) using thin-layer chromatography. Amplification of DNA fragment of 650 bp was done using the universal primers ITS1/ITS4 (Batista et al., 2008) and total genomic DNA as a template (White et al., 1990). BLAST analysis of this strain (GenBank accession No. KJ564033) showed 99% similarity to different strains (JX292091, JF412776, KF841549, JF412775). Phylogenetic analysis of identified pathogen and closely related species of A. flavus group by the Maximum Likelihood Hood tree method and profiling of DNA bands examined in the members of Aspergillus section Flavi amplified by three different Inter Simple Sequence Repeat (ISSR) primers clearly confirmed the pathogen identity. Surface sterilized healthy seeds were soaked in spore suspension (104 spores ml-1) from a one week old pathogen culture for 30 s, dried and transferred onto moist blotting paper in Petri plates. Control seeds were treated with sterilized distilled water. After 7 days incubation at 25°C, 90% of seeds were colonized and re-isolation of the same pathogen confirmed the Kochs postulates. To our knowledge, this is the first report of A. minisclerotigenes seed rot from Pakistan.

ANTIFUNGAL POTENTIAL OF LEAF EXTRACTS OF LEGUMINOUS TREES AGAINST SCLEROTIUM ROLFSII

Background: Sclerotium rolfsii Sacc. is a destructive soil-borne plant pathogen that infects over 500 plant species and causes significant yield losses in many economically important plant species. Synthetic fungicides used to combat the menace also pollute the environment and cause health hazards. In order to search environmental friendly alternatives from natural resources, methanolic extracts of three leguminous tree species namely Acacia nilotica (L.) Willd. ex Delile subsp. indica (Benth.) Brenan, Prosopis juliflora (Sw.) DC. and Albizia lebbeck (L.) Benth. were evaluated for their antifungal activity against S. rolfsii and A. nilotica subsp. indica exhibited the maximum fungicidal potential. Materials and Methods: Two hundred grams dried leaf material of each of the three test plant species were extracted with methanol for two weeks. After filtration, methanol was evaporated on a rotary evaporator. Malt extract broth was used to make various concentrations of the crude methanolic extracts and their antifungal potential was determined by comparing the fungal biomass in various treatments with control. Chemical composition of methanolic leaf extract of A. nilotica subsp. indica was determined through GC-MS analysis. Results: Methanolic leaf extract of A. nilotica subsp. indica showed the highest fungicidal activity. Fungal biomass was decreased by 17-55% due to various concentrations of this extract over control. Different concentrations of P. juliflora reduced fungal biomass by 3-52%. Fourteen compounds were identified in methanolic extract of A. nilotica subsp. indica. 9,12,15-octadecatrienoic acid, methyl ester, (Z,Z,Z,)- (16.59%) was the most abundant compound followed by 1-pentanol, 2 methyl-, acetate (14.80%); hexanedioic acid, dimethyl ester (13.10%) and cyclotriaconta- 1, 7, 16, 22-tetraone (10.28%). Conclusion: This study concludes that methanolic leaf extract of A. nilotica subsp. indica can be used for management of S. rolfsii.

Mycoremediation of Cu(II) and Ni(II)

In the present study, Cu(II) and Ni(II) removal capacity of two filamentous fungi namely, Aspergillus niger van. Tieghem and Rhizopus arrhizus Fischer was evaluated through batch adsorption assays. Batch sorption experiments were performed at 30°C, pH 5.0 and 150 rpm taking biomass amount of 0.2 g. Adsorption trials were carried out as function of initial metal ion concentration within range of 100 to 500 mg/L -1 for both single and binary metal solutions. Results revealed the biosorption capacity of both fungal species significantly increased as the initial concentration of metal ions was raised in the medium. Whereas, adsorption capacity of fungal species significantly decline in binary metal in comparison with single metal solution, desorption and reusability of the biosorbents biomass demonstrated a significantly higher desorption capability of R. arrhizus (> 65%) as compared to A. niger (> 60%) for both metal ions after 4th adsorption/desorption cycle. Batch experiments conducted with real industrial effluents reveled 5 to 10% reduction in sorption potential of test fungi in comparison to capacity obtained with synthetic solution of metal ions.