Echinocandin B (ECB) biosynthesis in Aspergillus nidulans is primarily governed by multiple genes located within the biosynthetic echinocandin (ecd) gene cluster. The contributory functions of many genes, including transcription factors and tailoring enzymes of the ecd gene cluster, have been previously studied. The present study focused on determining the role of transporter proteins, EcdLp, EcdCp, and EcdDp, in ECB efflux using in silico and biochemical approaches. The molecular docking analysis revealed that ECB relatively showed higher binding affinity for EcdLp than the other co-clustered MFS transporters EcdCp and EcdDp, suggesting a preferred substrate of EcdLp. These results were further confirmed by heterologous integration of the ecdL gene in the ABC transporters-deficient Saccharomyces cerevisiae AD1-8u⁻, confirming active efflux. However, the binding of ECB in EcdLp is distinct from the R6G binding, overlapping the promiscuous site of farnesol, resulting in inhibition of R6G efflux in a dose-dependent manner. In conclusion, these results decipher the ECB binding and efflux mechanism and unveil the evolutionarily specialized architecture of EcdLp that permits targeted metabolite export in addition to environmental responsiveness, and lay the groundwork for optimizing ECB production via transporter engineering.

The present study was aimed to evaluate the antifungal activity of solvent based extracts of Lentinus edodes mushroom on the phytopathogen Alternaria triticina, the causual agent of leaf blight disease in wheat plant by Poisoned Food Technique. Result showed that the aqueous extracts of this mushroom was not so much effective while methanolic extract significantly reduced their growth. Only ethanolic extract of Lentinus edodes mushroom was found to inhibit the growth of this pathogen completely. These studies suggest that Lentinus edodes mushroom contain potential compound which may be used for controlling leaf blight disease of wheat and it is profitable for economy of the country.

Fungi are well known for their ability to produce a multitude of secondary metabolites (SMs) which act as a weapon of defense to protect themselves against parasites and predators. A variety of fungal SMs have proved to serve as an important factor for decades. The synthesis of SMs in fungi is a complex, multi-step process and is stage-specific under specialized conditions. SMs are primarily synthesized by non-ribosomal peptide synthetase (NRPS) or polyketide synthase (PKS) enzymes. The genes encoded for SM synthesis are often located in the cluster form at the sub-telomere region. The SM biosynthetic gene cluster comprises of genes encoding for NRPS/PKS, a transcription factor, and other accessory genes essential for assembly and maturation of SM. The regulation of SM synthesis in fungi can be achieved by pathway-specific (in-clustered transcription factor), global regulatory proteins and chromatin remodeling. The regulatory protein-encoding gene present in each gene cluster is considered to be a crucial regulatory circuit of the SM biosynthetic pathway. Moreover, the regulation of fungal SM biosynthesis is also guided by global regulatory proteins responsive to pH, carbon, nitrogen, light/dark, and other environmental cues. Histone modifications by methylation and acetylation often altered the chromatin structure; as a result, these changes repress or express the genes of SM biosynthetic pathway. Taken together, we conclude that fungal SM ability as antibiotics to toxins is useful to mankind.

The ecdB is a transcription factor, located in the echinocandin B biosynthetic gene cluster of Emericella rugulosa NRRL11440. Here, we validated the ecdB mRNA sequence for functional expression and to explore the role of EcdB protein in the echinocandin B regulation. The sequence alignment study revealed that the ecdB coding sequence was found 75 bp shorter than the reference mRNA sequence. This coding sequence encodes for EcdB protein and comprises three conserved domains; DNA binding domain (DBD), coiled-coil domain, and signature middle homology region. The full-length and DBD (truncated) DNA sequences were expressed in Escherichia coli BL21(DE3) under different tested conditions. The expression of EcdB protein was found to be toxic, which curbs the cell growth. In contrast to truncated protein (GST:EcdB1-54), the full-length (GST:EcdB) protein was expressed at very low titer and not detectable in SDS-PAGE under the varying isopropyl β-d-1-thiogalactopyranoside (IPTG), temperature, and media conditions. However, GST:EcdB1-54 was successfully purified under standard conditions (0.5 mM IPTG at 0.5OD) with 33 kDa expected size. The functionality of GST:EcdB1-54 was attained by electrophoretic mobility shift assay study as a clear band shifting showed with ecdA promoter. Taken together, we conclude that EcdB interacts with the ecdA promoter that reflected to require for echinocandin B regulation.

Staphylococcus aureus is an opportunistic pathogen, responsible for superficial and invasive infections both in nosocomial and community-acquired settings. The incidences of infection have become more problematic attributable to emerging drug resistance and biofilm formation. These challenges suggest the need for new antimicrobial agents against S. aureus. In present work, we purified a fungal xenobiotic (FI3) which elicits a potent antimicrobial activity against a list of tested microbes including methicillin sensitive (MSSA) and methicillin resistance (MRSA) S. aureus. The cell growth of MSSA and MRSA were completely ceased with the 1× minimum inhibitory concentration (MIC); 32 µg/mL and 128 µg/mL, respectively. The cell viability severely decreased within 90 min, due to disturbance of membrane homeostasis. This bactericidal effect was enhanced at lower pH (pH 4) with a speculation to retain positive charge. The FI3 potently disrupts biofilm adherence at 64 µg/mL and found to be a safe with no toxic effect on mammalian tissue. FI3 also leads to increase the potency of tested antibiotics. Taken together, we established that FI3 has a potent antimicrobial activity against tested microbes and safer to human tissue. It may be proven a leading molecule for the treatment of bacterial infections.

Echinocandin B is a potent antifungal against the majority of fungal pathogens and its biosynthesis occurred by ecd and hty gene clusters in Emericella rugulosa NRRL 11440. We elucidated the functional necessity of in-clustered transcription factor; ecdB in the production of echinocandin B. We deleted the ecdB gene and found that ΔecdB mutant has no significant effect on echinocandin B production. The expression level of most of the ecd and hty cluster genes was not significantly altered except few of them up-regulated in knockout strain. The complete abrogation in ecdB gene expression was observed in ΔecdB strain. However, the interactions of purified EcdB protein with DNA sequence of ecdA, ecdH, ecdK and ecdI promoter was confirmed in-vitro. Our results conclude that EcdB protein in-vitro binds to the ecdA, ecdH, ecdK and ecdI promoter but in-vivo, it could not significantly affect the gene expression and echinocandin B production in Emericella rugulosa.

ABSTRACT Objectives: Insect pest problems in agriculture have a considerable shift during first decade of 21st century due to global climate changes. Various synthetic insecticides were used from last 50 years to overcome the pest problem, but major concerns in using these pesticides were: Public health and pesticide resistance in pests. To abolish the catastrophic effect of synthetic insecticides, urgent need to develop new drugs for pest control. In this study we evaluate the insecticidal and genotoxic activity of ethanolic extract of Acorus calamus rhizome against Drosophila melanogaster. Methods: Different concentration of ethanolic extracts of A. calamus, fed to different developmental stages of D. melanogaster to determine insecticidal activity and genotoxicity; genotoxicity was determined by the comet assay in male flies. Results: Lethal concentration 50 (LC50) values of ethanolic extract against larvae, adult males and females of D. melanogaster were 109.54, 52.51 and 41.11 mg/L respectively. Genotoxicity of adult’s flies were determined at 30 and 55 mg/L ethanolic extract of A. calamus, The mean comet tail length was 4.24±0.653 μm and 6.13±0.721 μm and the respective DNA damage was 5.1% and 7.3% with reference to controls. Conclusion: The present finding suggests that ethanolic extract of A. calamus rhizome showed higher insecticidal potential in adults than larvae of D. melanogaster. Results from genotoxicity studies further support the insecticidal activity against adult stages of D. melanogaster. The study reflects ethanolic extract of A. calamus could be used as an alternative pest control negotiator for minimizing the noxious effects of pesticides in the environment. Keywords: Insecticidal activity, Genotoxicity, Ethanolic extract, Botanical pesticides, Acorus calamus, Drosophila melanogaster.

BACKGROUND:Indiscriminate use of synthetic insecticides to eradicate mosquitoes has caused physiological resistance. Plants provide a reservoir of biochemical compounds; among these compounds some have inhibitory effect on mosquitoes. In the present study the larvicidal, adulticidal and genotoxic activity of essential oil of Psoralea corylifolia Linn. against Culex quinquefasciatus Say was explored.METHODS:Essential oil was isolated from the seeds of P. corylifolia Linn. Larvicidal and adulticidal bioassay of Cx. quinquefasciatus was carried out by WHO method. Genotoxic activity of samples was determined by comet assay. Identification of different compounds was carried out by gas chromatography- mass spectrometry analysis.RESULTS:LC50 and LC90 values of essential oil were 63.38±6.30 and 99.02±16.63 ppm, respectively against Cx. quinquefasciatus larvae. The LD50 and LD90 values were 0.057±0.007 and 0.109±0.014 mg/cm2 respectively against adult Cx. quinquefasciatus,. Genotoxicity of adults was determined at 0.034 and 0.069 mg/cm2. The mean comet tail length was 6.2548±0.754 μm and 8.47±0.931 μm and the respective DNA damage was significant i.e. 6.713% and 8.864% in comparison to controls. GCMS analysis of essential oil revealed 20 compounds. The major eight compounds were caryophyllene oxide (40.79%), phenol,4-(3,7-dimethyl-3-ethenylocta-1,6-dienyl) (20.78%), caryophyllene (17.84%), α-humulene (2.15%), (+)- aromadendrene (1.57%), naphthalene, 1,2,3,4-tetra hydro-1,6-dimethyle-4-(1-methyl)-, (1S-cis) (1.53%), trans- caryophyllene (0.75%), and methyl hexadecanoate (0.67%).CONCLUSION:Essential oil obtained from the seeds of P. corylifolia showed potent toxicity against larvae and adult Cx. quinquefasciatus. The present work revealed that the essential oil of P. corylifolia could be used as environmentally sound larvicidal and adulticidal agent for mosquito control.