Assessment of bacteria and archaea in metalworking fluids using massive parallel 16S rRNA gene tag sequencing

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Abstract

Determination of the bacterial diversity in industry-based liquid in-use water-miscible metalworking fluid (MWF) samples was targeted by massive parallel multiplex DNA sequencing, either directly or upon pretreatment with propidium monoazide (PMA) that allows differentiation between intact and physically damaged cells. As MWFs provide a suitable basis of life for micro-organisms, the majority is preserved by biocides. ‘Bio-concept’ fluids on the other hand are bactericide free, which intentionally leads to substantial bacterial populations. Samples from both fluid types were chosen: A median of 51 operational taxonomic units at genera level (OTUs) were detected per sample, but only 13 were present at or above 1·0% of the total population in any PMA-treated sample analysed. As both fluid types were mainly dominated by Pseudomonas spp., we resolved this genus on the species level and found the Pseudomonas oleovorans/pseudoalcaligenes group to predominate. We also looked for archaea and detected Methanobrevibacter spp., albeit in <3% of all samples analysed.

Significance and Impact of the Study:

Water-miscible metalworking fluids provide a suitable base of life for micro-organisms, mainly bacteria and fungi. Earlier publications suggested that the diversity is rather low, but these studies were largely based on heterotrophic plate counts. This might have resulted in underestimation of population density and microbial diversity as some organisms might just refuse to grow. This study used high-throughput sequencing in the absence and presence of propidium monoazide to explore bacterial and archaeal presence in metalworking fluids. We established that diversity is low and bacterial populations are dominated by the genus Pseudomonas spp.

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