Spore swelling and germination as a bioassay for the rapid screening of crude biological extracts for antifungal activity

Screening for bioactivity is commonly performed in vivo in a bioassay purposefully designed for revealing a defined bioactivity (e.g. fungicide or antibacterial activity). This allows the testing of many crude extracts. In the present work a new method (bioassay) targeting spore swelling and germination to assess antifungal susceptibility is developed and evaluated. Traditionally, antifungal activity has been investigated using disk diffusion assays or micro-well plates. Inhibition is measured as a function of radial growth, inhibition zone or turbidity. The construction of a bioassay composed of germinating fungal spores bears the prospect of being a more rapid method, allowing more extracts to be screened within a shorter time frame. It can also be used to reveal antifungal action at an early state in the prospecting process. Suppression of spore swelling provides early indication of inhibitory potential and the type of swelling curve produced might indicate the mechanism of fungistasis. A strain of Absidia glauca Hagem served as model organism. A Beckman Coulter Multiziser™ 3 particle analyser was applied for the determination of bioactivity and investigation of the sporangiospores. Inhibition was standardized against two known fungicides (sorbic and benzoic acid). Four biological extract solvents were also tested; where DMSO was found to be the best candidate as extract solvent in the assay. Inhibition was investigated as changes in volumes of the germinating spores using germination as endpoint target. The new bioassay was found to be a simple and rapid method for detection of antifungal activity of extracts.