ABSTRACT
The expanding interest in bioremediation of poorly degradable wastes has led to the discovery of many microbial enzymes capable of degrading recalcitrant substances such as keratinaceous wastes that are produced in vast quantities on daily basis. Such enzymes don't only work as a bioremediation tool but also have multiple beneficial applications. Hence, environmental samples were collected from sewage water, soils, animal bodies and feces in order to isolate keratinase producing organisms. Keratinolytic isolates were isolated from sewage water; soils; animal bodies; animal feces, and identified both traditionally and molecularly through 16S-rRNA sequencing to be Bacillus cereus strain. Produced keratinase was purified by centrifugation, ammonium sulfate precipitation, and HPLC, then assayed using Azokeratine based analysis. keratinase quantification yielded a 420 ± 1.63 U/mL. Optimum production was obtained at 40 °C, pH 7, 3 days incubation, 0.5 % substrate, 0.4 g/l magnesium ion, 2% v/v inoculum, 0.5 g/l NaCl, 0.4 g/l K2HPO4, and 0.3 g/l KH2PO4. Production was increased by 1.9 fold after acclimatization to reach 809 ± 2.49 U/mL in only 2 days. Thermal and pH stability testing revealed the effectiveness of the isolated keratinase over a wide range of temperatures at neutral pH. Finally, isolated keratinase enhanced fusidic acid topical penetration to treat induced deep skin bacterial infection in mice. A 1.4 fold decrease in treatment period and a 2 log cycle reduction in the viable count of Staphylococcus aureus were noticed in keratinase/fusidic acid treated mice compared to mice treated with fusidic acid alone. This study shed some light on a simple keratinase production optimization technique and suggested a promising medical application of this enzyme as a drug delivery agent.
ABSTRACT
Autoimmune hepatitis (AIH) is a chronic inflammatory disorder with complex immunopathogenesis. Dysbiosis has been linked to many autoimmune diseases, but its detailed role in autoimmune hepatitis (AIH) still needs rigorous evaluation, especially in Egypt. We aimed to identify the shift in the gut microbiota profile and resultant metabolic pathways in AIH Egyptian patients compared to healthy individuals. Stool samples were collected from 15 AIH-naive patients and from 10 healthy individuals. The V3-V4 hyper-variable regions in16S rRNA gene was amplified and sequenced using Illumina MiSeq platform. Significantly lower bacterial diversity in AIH patients was found compared to the controls. A phylum-level analysis showed the overrepresentation of Firmicutes, Bacteroides, and Proteobacteria. At the genus level, AIH-associated enrichment of Faecalibacterium, Blautia, Streptococcus, Haemophilus, Bacteroides, Veillonella, Eubacterium, Lachnospiraceae and Butyricicoccus was reported in contrast to Prevotella, Parabacteroides and Dilaster, which were significantly retracted in such patients. Overall, the predicted metabolic pathways associated with dysbiosis in AIH patients could orchestrate the potential pathogenic roles of gut microbiota in autoimmune disease, though not in a disease-specific manner, calling for future large-scale studies.