Biology subjects

Ravichandran, A.

Publications and source records attributed to Ravichandran, A..

3 recordsLinked to original sources

bioRxiv2018

A novel Versatile Peroxidase from Lentinus squarrosulus (MH172167)towards enhanced delignification and digestibility of crop residues

Scarcity of quality feed is a major constraint concerning livestock productivity with recalcitrant lignin hindering utilization of crop residues as quality animal feed. Degradation of lignin in nature is contributed by white-rot fungi through their enriched ligninolytic system. Versatile Peroxidase plays a key role in ligninolysis through its capability to oxidize diverse class of aromatics without mediators. In this study, wild isolates of wood rotting fungi were screened for potential peroxidases oxidizing manganese and aromatic compounds. The strain identified as Lentinus squarrosulus (TAMI004, BankIt2098576 MH172167) was monitored for enzyme activity in solid state and submerged fermentation. L. squarrosulus demonstrated predominant Versatile Peroxidase activity amongst the screened wild isolates displaying hybrid characteristic of manganese oxidation and manganese independent reactions on aromatic compounds. The manganese oxidizing peroxidase activity evidenced in submerged fermentation was 12 IU/L whereas in solid state fermentation it was 131 IU/L. This ability to act through manganese mediated and independent reactions on phenolics reveals its biotechnological and industrial significance. Treatment of common crop residues with crude extract of L. squarrosulus rich in Versatile Peroxidase obtained from both Solid state and submerged fermentations showed a decrease in their Neutral Detergent Fiber, Acid Detergent Fiber and Acid Detergent Lignin content showing biodegradation, substantiating the ligninolytic ability and more prominently increase in their digestibility. To the best of our knowledge, this is the first report describing Versatile Peroxidase from Lentinus squarrosulus with potential to augment the ruminant digestibility of crop residues.\n\nImportanceVersatile Peroxidase of White-rot fungi, a relatively less studied lignolytic enzyme, is very efficient in depolymerization of lignin macromolecule through its multivalent catalytic sites. Lignin degradation is very appealing from the application perspective as attack on lignin exposes the energy affluent polysaccharides for utilization in extensive biotechnological applications. Reports on relevance of Versatile Peroxidase for these purposes are still emerging, however the role of ligninolytic enzymes especially Versatile Peroxidase in enriching ruminant feed is yet unturned. Here, this work demonstrates the potential of Versatile Peroxidase from a novel species Lentinus squarrosulus in delignification thereby upgrading the digestibility and nutritive value of crop residues. The observations validate the importance of the enzyme in improvement of crop residues for feeding ruminants in the current scenario where, livestock productivity is severely impacted by lack of quality feed and demand for alternate feed resources is intensifying.

microbiology↗

bioRxiv2018

Occidiofungin, an actin binding antifungal with in vivo efficacy in a vulvovaginal candidiasis infection

Current antifungal treatment options are plagued with rapidly increasing occurrence of resistance, high degree of toxicity and a limited spectrum of activity. The need to develop a novel antifungal with a unique target, wider spectrum of activity, and reduced toxicity to the host, is urgent. We have identified and characterized one such compound named occidiofungin that is produced by the soil bacterium Burkholderia contaminans MS14. This study identifies the primary cellular target of the antifungal, which was determined to be actin. Actin binding metabolites are generally characterized by their ability to inhibit polymerization or depolymerization of actin filaments, which presumably accounts for their severe toxicity. Occidiofungin, instead, has a subtler effect on actin dynamics that triggers apoptotic cell death. We were able to demonstrate the effectiveness of the antifungal in treating a vulvovaginal yeast infection in a murine model. This discovery puts occidiofungin in a unique class of actin-binding antifungal compounds with minimal reported toxicity to the host. The results of this study are important for the development of a novel class of antifungals that could fill the existing gap in treatment options for fungal infections.\n\nAuthor summaryWidespread resistance to antifungal compounds currently in use has been alarming. Identification and development of a new class of antifungals with a novel cellular target is desperately needed. This study describes the assays carried out to determine the molecular target and evaluate efficacy of one such novel antifungal compound called occidiofungin. Occidiofungin modified with a functional alkyne group enabled affinity purification assays and localization studies in yeast. These studies led to the identification of the actin binding property of occidiofungin. Actin-binding by secondary metabolites often exhibit severe host toxicity, but this does not appear to be the case for occidiofungin. We have previously been able to administer occidiofungin to mice at concentrations in the range of 5 mg/kg without any serious complications. We were able to demonstrate the effectiveness of the antifungal in treating a vaginal fungal infection in a murine model. The results outlined in this manuscript establish that occidiofungin is an efficacious compound with a novel molecular target, putting it in a completely new class of antifungals.

microbiology↗

bioRxiv2018

A rapid microwave method for isolation of genomic DNA and identification of white rot fungi.

White rot fungi (WRF) produce lignolytic enzymes comprised by laccases and peroxidases responsible for mineralization of recalcitrant lignin. Because of the so-called lignin modifying enzymes(LMEs), these fungi have potential applications in biodegradation and bioremediation processes. Increased demand for lignolytic enzymes to exploit their various applications has sparked interest in identifying and characterizing new novel strains of WRF. Despite this undisputed biotechnological significance, molecular identification of WRF, remains a daunting task for researchers as genomic DNA isolation is a tedious process, unsuccessful many a times because of their rigid and resistant cell walls. A rapid, effective and efficient method to identify the innumerable fungal strains within no time is the need of the hour. The fungal mycelia of various unknown as well as know isolates of WRF, after alternative washing with TE buffer and sterile water were suspended in TE buffer. Fungi in solution were then exposed to microwave. The crude extract contained genomic DNA which was extracted and amplified using ITS primers for further identification. Based on sequencing results the identity of known cultures was confirmed, while the unknown cultures were identified as Clitopilus scyphoides (AGUM004, BankIt2098576 MH172163); Ganoderma rasinaceum (AGUM007, BankIt2098576 MH172163); Schizophyllum sp (KONA001 BankIt2098576 MH172164; AGUM011 BankIt2098576 MH172165and AGUM021 BankIt2098576 MH172166respectively), Coprinellus disseminatus (BANG001, BankIt2098576 MH172167) and Lentinus squarrosulus (TAMI004, BankIt2098576 MH172167). The microwave method described for isolating quality DNA of WRF without further purification steps proved a novel method requiring less than ten minutes and minimized the chances of the presence of PCR inhibitors.\n\nIMPORTANCEWhite rot fungi which decay wood, possess selective lignin degrading enzymes responsible for degrading a wide variety of environmental pollutants, xenobiotic compounds in addition to mineralizing chemicals that are insoluble and recalcitrant. Lignolytic enzymes hold potential towards replacing conventional chemical processes and their increased demand in the market has ignited interest in identifying and characterizing new strains of WRF. A rapid, efficient method capable of quickly identifying fungal isolates is a constraint. The microwave method is a novel quick method for isolating superior quality DNA. Its adoption circumvents the initial purification steps and /or interference of PCR inhibitors, which are encompassed in the use of conventional methods. The microwave method thus permits the thorough amplification of the ITS region thereby aiding in the easy identification of unknown species. Use of the microwave method will permit researchers to obtain DNA from fungi very quickly for further application in molecular studies.

molecular biology↗