Standardization of metagenomic DNA extraction protocol is a pre-requisite for a successful metagenomic study aiming to screen and exploit the variety of microorganisms inhabiting a particular soil environment. Six methods reported earlier were used for isolation of metagenomic DNA in the present study. These methods suffered with regard to either poor yield or quality of DNA. Therefore, we developed an improved method for isolation of high-molecular weight and good quality metagenomic DNA from different soil samples. Our protocol combines the enzymatic (lysozyme and proteinase K) and chemical (CTAB and CaCl) strategies to ensure efficient cell lysis and use of PEG and isopropanol for precipitation of humic impurities-free DNA. Our improved method gave high yield of good quality metagenomic DNA from diverse soils collected from garden, domestic waste dumping site, cellulose waste dumping site, sewage site, and tannery waste site. The good quality of the metagenomic DNA was evident by spectrophotometry data, PCR amplification of 16S rRNA gene and restriction digestion.
The prevalence of drug-resistant pathogenic fungi is a major global health challenge. There is an urgent need for novel drugs that can exert a potent antifungal activity and overcome resistance. Newly discovered anti-fungal properties of existing compounds can potentially offer a rapid solution to address this persistent threat. We rationalized that structures which disrupt the fungal cell membrane could address the above unmet need. As fatty acids underpin the formation and stability of cell membranes, we used computational simulations to evaluate the interactions between selected short chain fatty acids and a model cell membrane. Here, we report that caprylic acid could penetrate and perturb the membrane in silico. Based on the in silico findings, we identified a derivative of this fatty acid that disrupts fungal membranes as detected using steady-state fluorescence anisotropy. We show that this fatty acid derivative is potent against a variety of fungal pathogens like Candida and Trichophyton. We further demonstrated the ability of this fatty acid derivative to potentiate some azoles in vitro and enhance the efficacy of antifungal formulations in vivo. Our data suggests the emergence of a novel therapy for effective disease management and overcoming anti-fungal drug resistance.
histopathology and immune histochemical aspects of the ductal tumours-they did not mention the outcome of the disease in that patient. 2 Similarly, there are two other reports describing the orbital metastasis in case of a mixed parotid tumour and polymorphous low-grade adenocarcinoma of the parotid glands. 3,4 After orbital exenteration, the examination of the specimen under the microscope revealed a larger poorly differentiated tumour mass eroding the ocular coats with a posterior extension invading the proximal part of the optic nerve; however, the resected posterior end of the optic nerve and rest of the margins were free of tumour cells. Immunological stains for cytokeratin, epithelial membrane antigen, and cytokeratin 7 were positive, whereas reactivity for S-100 proteins, estrogen receptors, progesterone receptors were negative. To conclude, the primary salivary duct carcinoma of the salivary gland remained asymptomatic clinically as well as radiologically for 18 months. Thus orbital tumours with unusual presentations need a detailed evaluation with close follow-up to counter the problems related to a primary tumour elsewhere in the body at the earliest, thereby helping to reduce systemic morbidity and mortality.
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