RESUMO
A distinct strain named Micromonospora sp. Rc5 was isolated from Sinai desert of Egypt and recorded high antagonistic activities against some food and bloodborne pathogens. Morphological and chemotaxonomy characterization confirmed that this isolate belongs to genus Micromonospora. Sequencing of partial 16S rDNA and BLASTN showed that isolate Rc5 is identical to Micromonospora haikouensis (99%) but with low bootstrap value in NJ phylogenetic tree. Comprehensive optimization of several growth factors was performed including initial pH, incubation periods, and different sources of carbon and nitrogen. The highest yield of antimicrobial agent production was obtained after 8 days of incubation at 30°C, pH 6.0, 3 x 105CFU/ml in soya bean meal broth media with agitation of 150 rpm. A dramatic proportional decrease occurred with 0.3, 0.6, 0.9 µg active fraction /ml against Staphylococcus aureus culture and reached to complete inhibition at a minimum inhibitory concentration of (1.5 µg /ml). The physicochemical characterization of the purified fraction was identified as phthalate derivative. Our results indicated that Rc5 generated potential antimicrobial compounds against foodborne pathogens and may combat resistant strains of Staphylococcus aureus.
RESUMO
Aims: Design and assembly of an inexpensive microfluidic PDMS chip for visual detection of cell adhesion and biofilm formation. Study Design: Three different styles of microchannels (2.6, 5.0, and 11.5 µl volumes) were designed, fabricated and tested for adhesion and biofilm formation in a microfluidic system. The pressure drop measurements system includes a bio-Ferrograph connected to the PDMS microchannel via a syringe and a pressure transducer. Methodology: Microfluidic chips were fabricated using Polydimethylsiloxane (PDMS) by means of soft lithography. Different cell densities of E.coli K12 cells were introduced to investigate adhesion and biofilm formation at different time intervals. Stabilization time and hydraulic resistance were obtained via a Bio-Ferrograph connected to a pressure transducer. Results: PDMS microfluidic volume (2.6 µl) failed to generate noticeable biofilm, while slight and greatest yield occurred with PDMS microchannels (5.0, and 11.5 µl), respectively, and could detect as low as 26 cells in 11.5 µl microchannel. As incubation time and/or initial cell density increases, cell adhesion increased, illustrated by crystal violet color intensity. High stabilization time (3 h) didn’t allow for bacterial attachment and cultivation inside the microchannel (2.6 µl) while lower stabilization time (10 min) yielded the highest capacity of cell adhesion in microchannel (11.5 µl). Conclusions: We developed a microfluidic chip with low stabilization time and hydraulic resistance, thus offering more volume for adhesion of bacterial cells and biofilm formation. It allowed bacterial cultivation without any addition of nutrients. The microfluidic chip provides a platform to monitor biofilm growth and can be integrated in situ investigations for biological systems, food biotechnology and other industrial biotechnology applications. This would allow a non-destructive and non-invasive monitoring of the biofilm-forming bacteria inside the PDMS microfluidic chip. This work opens opportunities for further investigations of pressure drop phenomena in microchannels that would otherwise go unnoticed in macro scale measurements.