Spontaneous vegetable fermentations, with their rich flavors and postulated health benefits, are regaining popularity. However, their microbiology is still poorly understood, therefore raising concerns about food safety. In addition, such spontaneous fermentations form interesting cases of man-made microbial ecosystems. Here, samples from 38 carrot juice fermentations were collected through a citizen science initiative, in addition to three laboratory fermentations. Culturing showed that Enterobacteriaceae were outcompeted by lactic acid bacteria (LAB) between 3 and 13 days of fermentation. Metabolite-target analysis showed that lactic acid and mannitol were highly produced, as well as the biogenic amine cadaverine. High-throughput 16S rRNA gene sequencing revealed that mainly species of Leuconostoc and Lactobacillus (as identified by 8 and 20 amplicon sequence variants [ASVs], respectively) mediated the fermentations in subsequent order. The analyses at the DNA level still detected a high number of Enterobacteriaceae, but their relative abundance was low when RNA-based sequencing was performed to detect presumptive metabolically active bacterial cells. In addition, this method greatly reduced host read contamination. Phylogenetic placement indicated a high LAB diversity, with ASVs from nine different phylogenetic groups of the Lactobacillus genus complex. However, fermentation experiments with isolates showed that only strains belonging to the most prevalent phylogenetic groups preserved the fermentation dynamics. The carrot juice fermentation thus forms a robust man-made microbial ecosystem suitable for studies on LAB diversity and niche specificity.IMPORTANCE The usage of fermented food products by professional chefs is steadily growing worldwide. Meanwhile, this interest has also increased at the household level. However, many of these artisanal food products remain understudied. Here, an extensive microbial analysis was performed of spontaneous fermented carrot juices which are used as nonalcoholic alternatives for wine in a Belgian Michelin star restaurant. Samples were collected through an active citizen science approach with 38 participants, in addition to three laboratory fermentations. Identification of the main microbial players revealed that mainly species of Leuconostoc and Lactobacillus mediated the fermentations in subsequent order. In addition, a high diversity of lactic acid bacteria was found; however, fermentation experiments with isolates showed that only strains belonging to the most prevalent lactic acid bacteria preserved the fermentation dynamics. Finally, this study showed that the usage of RNA-based 16S rRNA amplicon sequencing greatly reduces host read contamination.

Background: The popularity of Galleria mellonella as invertebrate model is increasing rapidly, because it forms an attractive alternative to study bacterial, fungal and viral infections, toxin biology, and to screen antimicrobial drugs. For a number of vertebrate and invertebrate animal and plant models, it has been established that the commensals present within the microbial communities on various host surfaces will influence the host's immune and growth development state and the colonization capacity of newly introduced microorganisms. The microbial communities of Galleria mellonella larvae have, however, not yet been well characterized. Results: In this study, we present the bacterial communities that were found by 16S rRNA amplicon sequencing on different body sites of G. mellonella larvae. These communities showed very little diversity and were mostly dominated by one Enterococcus taxon. In addition, we found that the production conditions (as 'bait' for fishing or under more controlled 'research grade' conditions-with or without hormones and antibiotics) appear to have little impact on the microbiota of the larvae. Conclusions: Establishment of the simplicity of the microbiota of G. mellonella larvae underlines the potential of the larvae as a model host system for microbiome-host interactions.

During the current COVID-19 pandemic, the use of face masks has become increasingly recommended and even mandatory in community settings. To evaluate the risk of bacterial cross-contamination, this study analyzed the bacterial bioburden of disposable surgical masks and homemade cotton masks, and surveyed the habits and face mask preferences of the Flemish population. Using culture approaches and 16S rRNA gene amplicon sequencing, we analyzed the microbial community on surgical and/or cotton face masks of 13 healthy volunteers after 4 h of wearing. Cotton and surgical masks contained on average 1.46 × 105 CFU/mask and 1.32 × 104 CFU/mask, respectively. Bacillus, Staphylococcus, and Acinetobacter spp. were mostly cultured from the masks and 43% of these isolates were resistant to ampicillin or erythromycin. Microbial profiling demonstrated a consistent difference between mask types. Cotton masks mainly contained Roseomonas, Paracoccus, and Enhydrobacter taxa and surgical masks Streptococcus and Staphylococcus. After 4 h of mask wearing, the microbiome of the anterior nares and the cheek showed a trend toward an altered beta-diversity. According to dedicated questions in the large-scale Corona survey of the University of Antwerp with almost 25,000 participants, only 21% of responders reported to clean their cotton face mask daily. Laboratory results indicated that the best mask cleaning methods were boiling at 100°C, washing at 60°C with detergent or ironing with a steam iron. Taken together, this study suggests that a considerable number of bacteria, including pathobionts and antibiotic resistant bacteria, accumulate on surgical and even more on cotton face masks after use. Based on our results, face masks should be properly disposed of or sterilized after intensive use. Clear guidelines for the general population are crucial to reduce the bacteria-related biosafety risk of face masks, and measures such as physical distancing and increased ventilation should not be neglected when promoting face mask use.

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Allergic asthma is a highly prevalent inflammatory disease of the lower airways, clinically characterized by airway hyperreactivity and deterioration of airway function. Immunomodulatory probiotic bacteria are increasingly being explored to prevent asthma development, alone or in combination with other treatments. In this study, wild-type and recombinant probiotic Lactobacillus rhamnosus GR-1 were tested as preventive treatment of experimental allergic asthma in mice. Recombinant L. rhamnosus GR-1 was designed to produce the major birch pollen allergen Bet v 1, to promote allergen-specific immunomodulation. Administration of wild-type and recombinant L. rhamnosus GR-1 prevented the development of airway hyperreactivity. Recombinant L. rhamnosus GR-1 also prevented elevation of airway total cell counts, lymphocyte counts and lung IL-1β levels, while wild-type L. rhamnosus GR-1 inhibited airway eosinophilia. Of note, a shift in gut microbiome composition was observed after asthma development, which correlated with the severity of airway inflammation and airway hyperreactivity. In the groups that received L. rhamnosus GR-1, this asthma-associated shift in gut microbiome composition was not observed, indicating microbiome-modulating effects of this probiotic. These data demonstrate that L. rhamnosus GR-1 can prevent airway function deterioration in allergic asthma. Bet v 1 expression by L. rhamnosus GR-1 further contributed to lower airway inflammation, although not solely through the expected reduction in T helper 2-associated responses, suggesting involvement of additional mechanisms. The beneficial effects of L. rhamnosus GR-1 correlate with increased gut microbiome resilience, which in turn is linked to protection of airway function, and thus further adds support to the existence of a gut-lung axis.