Cell viability assays require expensive commercial testing kits and sophisticated cell culture techniques. Thus, a laboratory activity for testing the cellular proliferative and cytotoxic effects of certain substances using baker’s yeast (Saccharomyces cerevisiae) in resazurin reduction test (RRT) was developed. This activity is based on the fact that a laboratory-prepared resazurin has identical effects to a resazurin-based commercial kits and that S. cerevisiae is an established model system for eukaryotic organisms. Here, a complete procedure based on optimized conditions was described in detail, from preparation of test materials and solutions up to the gathering and analysis of data. This laboratory activity provides a simple and cheap alternative to conventional cell viability assays and is therefore recommended for use in institutions with limited budget for the acquisition of materials for instruction purposes.
The use of microorganisms in decontaminating the environment encumbered with heavy metal pollutants through biosorption is considered as a good option for bioremediation. This study was conducted to isolate Cadmium (Cd) tolerant fungi from coastal waters and sediments, compare their biosorption capabilities, and identify the isolates with the highest Cd uptake. Water and sediment samples were collected near the effluent sites of industrial belt in Ibo, Lapu-lapu City, Cebu, Philippines. Potato dextrose agar (PDA) plates containing Cd (25, 50, 75, and 100 ppm) were used to isolate Cd tolerant fungi from the samples. The distinct colonies that grew on the highest Cd concentration (100 ppm) were then isolated into pure cultures. The pure cultures of Cd tolerant fungi served as a source of inocula for in vitro biosorption assay using Cd dissolved in potato dextrose broth (PDB) as the substrate. Cd tolerant fungal isolates with the highest Cd uptake were finally identified up to the lowest possible taxon based on their colonial and microscopic characteristics. Most filamentous fungal colonies have grown most at the lower Cd concentrations and least at the higher concentrations. From the characteristics of the fungal growth on the plate with the highest Cd concentration, eight distinct colonies from both sediment and water samples were isolated into pure cultures. Among the eight fungal isolates, only three had significant Cd biosorption efficiency, these were fungal isolate 3 (13.87 %), fungal isolate 6 (11.46 %), and fungal isolate 4 (10.71 %). Two of them (fungal isolates 3 and 4) belong to genus Aspergillus while the other (fungal isolate 6) is a species of Penicillium. The results of this study showed that Cd tolerant fungi with biosorption capacity could be isolated from coastal water and sediments in the vicinity of areas suspected of heavy metal contamination.
Brown macroalgae species are constantly reported as potential sources of bioactive compounds useful in inhibiting cell proliferation and vascular formation. Thus, this study was conducted to determine and compare the in vitro cytotoxic activities of Turbinaria ornata Agardh and Padina australis Hauck ethanolic extracts against baker's yeast (Saccharomyces cerevisiae) using the resazurin reduction test (RRT) and investigate their in vivo antiangiogenic activity through duck (Anas platyrhynchos) chorioallantoic membrane (CAM) assay. Both T. ornata and P. australis ethanolic extracts exhibited cytotoxic activities at IC50 of 530.53 ppm and 528.78 ppm, respectively, and significant cytotoxicity was determined in 750 ppm and 1000 ppm concentrations of T. ornata and 1000 ppm concentration of P. australis. Also, both T. ornata and P. australis ethanolic extracts exhibited antiangiogenic activity (100% vascular inhibition) as all the concentrations of both species caused severe vascular damage in all the duck CAM samples treated. These results show the potential future application of these species for cytotoxic activities and vascular inhibition. The conduct of further tests using other model systems is recommended.
Heavy metals are frequently reported for their mutagenic and teratogenic effects on benthic organisms. Thus, this study aimed to determine the toxicity of cadmium (Cd) and zinc (Zn) in the gametes of T. gratilla and to compare its fertilization and embryonic development under the highest nongametotoxic concentrations of these heavy metals. Gamete viability of T. gratilla under CdCl2 and ZnSO4 treatments was assayed through resazurin reduction test (RRT) and was confirmed through gamete morphology assay. ZnSO4 was more toxic to T. gratilla gametes than CdCl2 and egg cells were more sensitive to both than the sperm cells. Higher concentrations of CdCl2 and ZnSO4 induced gamete apoptosis and necrosis while highest nongametotoxic concentrations were determined at 1 × 10−3 M and 1 × 10−4 M, respectively, and were used in an in vitro fertilization and embryonic development experiment. ZnSO4 treatment inhibited fertilization more than CdCl2 and yielded more deformed embryos, while both induced abnormalities and hindered further embryonic development. This study gives the first report on the specific concentrations of Cd and Zn that are toxic to T. gratilla gametes and has confirmed the teratogenic effects of these heavy metals.
The use of microorganisms to decontaminate the environment loaded with oil pollutants is a beneficial option for bioremediation. This study aimed to isolate, characterize and identify oil-degrading fungi from the coastal sediments of the port area of Ormoc City, Philippines, and to evaluate and compare the oil-biodegrading capabilities of these fungi. Fungal isolates were subjected to an in-vitro biodegradation assay using used engine oil as substrate. Four species of filamentous fungi were isolated and identified down to genus level. The fungal isolates include three Aspergillus spp. and one Penicillium sp. Aspergillus spp. 1 & 2 were the most efficient in degrading and utilizing the used engine oil for growth. Aspergillus sp. 1 & 2 and Penicillium sp.had mean pH values of 6.286, 6.136 and 6.32, respectively. Highest mean percent fat loss on the media and highest mean dry weights were exhibited by Aspergillus sp. 2 (mean fat loss = 77.12% and mean dry weight= 0.24g) and Aspergillus sp. 1 (mean fat loss = 69.4% and mean dry weight = 0.21g). Filamentous fungi from the coastal sediments in the vicinity of Ormoc City port area could therefore be used as potential bioremediation agents in areas contaminated with petroleum oils.
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