Flow cytometry, a powerful novel tool to assess viability of fungal conidia in metal working fluids

Fungal contamination of metal working fluids (MWF) is a dual problem in automatized processing plants because resulting fungal biofilms obstruct cutting, drilling and polishing machines. Moreover, some fungal species of MWF comprise pathogens such as Fusarium solani . Therefore, the development of an accurate analytical tool to evaluate conidia viability in MWF is important. We developed a flow cytometric method to measure fungal viability in MWF using F. solani as model organism. To validate this method, viable and dead conidia were mixed in several proportions and flow cytometrically analyzed. Subsequently, we assessed the fungicidal activity of two commercial MWF using flow cytometry (FCM) and compared it with microscopic analyses and plating experiments. We evaluated the fungal growth in both MWF after 7 days using qPCR to assess the predictive value of FCM. Our results show that FCM distinguishes live from dead conidia as early as 5 hours after exposure to MWF whereas the microscopic germination approach detected conidial viability much later and less accurately. At 24h, microscopic analyses of germinating conidia and live/dead analyses by FCM correlated well although the former consistently underestimated the proportion of viable conidia. In addition, reproducibility and sensitivity of the flow cytometric method was high and allowed to assess the fungicidal properties of two commercial MWF. Importantly, the obtained flow cytometric results on viability of F. solani conidia at both early time points (5h and 24h) correlated well with fungal biomass measurements assessed via a qPCR methodology 7 d after the start of the experiment. Importance This result depicts the predictive power of FCM in assessing the fungicidal capacity of MWF formulations. It also implies that FCM can be implemented as a rapid detection tool to estimate the viable fungal load in an industrial processing matrix (MWF).