| Summary: | In two previous surveys, the U.S. Food and Drug Administration (FDA) identified microbial contamination in 53 of 112 (47%) unopened tattoo inks and tattoo-ink-related products (e.g., diluents) from 15 manufacturers in the U.S. In this study, we primarily focused our microbiological survey on permanent makeup (PMU) inks. We conducted a survey of 47 unopened PMU inks from nine manufacturers and a comparative species-centric co-occurrence network (SCN) analysis using the survey results. Aerobic plate count and enrichment culture methods using the FDA’s <i>Bacteriological Analytical Manual</i> (BAM) Chapter 23 revealed that 9 (19%) inks out of 47, from five manufacturers, were contaminated with microorganisms. The level of microbial contamination was less than 250 CFU/g in eight inks and 980 CFU/g in one ink. We identified 26 bacteria that belong to nine genera and 21 species, including some clinically relevant species, such as <i>Alloiococcus otitis</i>, <i>Dermacoccus nishinomiyaensis</i>, <i>Kocuria rosea</i>, and <i>Pasteurella canis</i>. Among the identified microorganisms, the SCN analysis revealed dominance and a strong co-occurrence relation of spore-forming extreme environment survivors, <i>Bacillus</i> spp., with close phylogenetic/phenotypic relationships. These results provide practical insights into the possible microbial contamination factors and positive selection pressure of PMU inks.
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