Diversity and source-tracking of spoilage molds in bakery products using MALDI-TOF MS and molecular approaches

Filamentous fungi are significant contributors to global food losses and waste including bakery products. In the baking industry, mold contamination often occurs at the post-baking stage due to exposure to ambient air. The aim of this study was to investigate fungal diversity and to track the environmental sources of spoilage molds in finished products in a French industrial bakery plant, using MALDI-TOF MS and molecular approaches including barcoding and genotyping of microsatellites using next-generation sequencing (SSR-seq). Furthermore, we evaluated MALDI-TOF MS as a rapid tool for the source-tracking of two spoilage molds, i.e., Cladosporium sphaerospermum and Penicillium brevicompactum. Samples from air, food contact surfaces, work clothes and cakes were collected once every four weeks for 42 weeks on 1-4 consecutive days at 2 times. Results showed that areas with uncontrolled air quality such as the pre-baking area, had higher fungal loads compared to air-controlled areas such as post-baking zones. Products not sprayed with ethanol showed significantly higher mold contamination frequency highlighting the importance of surface treatment in reducing spoilage. Cladosporium sphaerospermum and Penicillium brevicompactum, known for their xerophilic and airborne dispersal capabilities, were identified as the main product spoilers. SSR-seq provided insights into the genetic diversity of these species and showed that certain haplotypes, particularly originating from post-baking areas, were more frequently found on spoiled cakes suggesting repeated contamination events from persistent sources. Using both unsupervised and supervised approaches, we showed that there was a good agreement between MALDI-TOF MS and genotyping data analysis at the population level. This study provides new insights into the fungal ecology encountered in the bakery industry.