The sanitary indoor environment-a potential source for intact human-associated anaerobes

Abstract

A healthy human microbiome relies on the interaction with and exchange of microbes that takes place between the human body and its environment. People in high-income countries spend most of their time indoors and for this reason, the built environment (BE) might represent a potent source of commensal microbes. Anaerobic microbes are of particular interest, as researchers have not yet sufficiently clarified how the human microbiome acquires oxygen-sensitive microbes. We sampled the bathrooms in ten households and used propidium monoazide (PMA) to assess the viability of the collected prokaryotes. We compared the microbiome profiles based on 16S rRNA gene sequencing and confirmed our results by genetic and cultivation-based analyses. Quantitative and qualitative analysis revealed that most of the microbial taxa in the BE samples are human-associated. Less than 25% of the prokaryotic signatures originate from intact cells, indicating that aerobic and stress resistant taxa display an apparent survival advantage. However, we also confirmed the presence of intact, strictly anaerobic taxa on bathroom floors, including methanogenic archaea. As methanogens are regarded as highly sensitive to aerobic conditions, oxygen-tolerance experiments were performed with human-associated isolates to validate their survival. These results show that human-associated methanogens can survive oxic conditions for at least 6 h. We collected strong evidence that supports the hypothesis that obligate anaerobic taxa can survive in the BE for a limited amount of time. This suggests that the BE serves as a potential source of anaerobic human commensals.

Fig. 1. Distribution and relative abundance of bacterial taxa in samples from different households/bathrooms.

Bar charts showing the microbial composition of non-PMA and PMA-treated bathroom floor samples at the (a) phylum and (b) genus levels. Genera with <2% rel. abundance are summarised in grey. Bubble plots display the 25 most-abundant genera (bubble size reflects the relative abundance): (c) Household samples (H1–H10) are depicted individually for non-PMA and PMA treatment together with human samples (grey background) that represent reads from several body sites: nasal cavity, skin, vagina, urine, stool and oral samples. d Human samples compared to non-PMA (blue background) and PMA (red background) bathroom samples. Genera are coloured according to their taxonomic phylum, and taxa that predominantly contain strict anaerobes are marked by an asterisk.

Fig. 2. A comparison between the indoor and human microbiome.

a Principal coordinates analysis plots based on Bray-Curtis dissimilarity and (b) alpha diversity indices (Shannon, left; richness, right) are depicted for all indoor samples (blue = untreated, red = PMA treated) together with representative human body site samples (green = nasal cavity, light blue = oral, yellow = skin, brown = stool, pink = urine, purple = vaginal). Significant differences are indicated by the different letters above the bars, as defined by a Mann–Whitney U test; P < 0.05, FDR-adjusted (samples that share the same letter do not significantly differ). The proportion of microbes in 10 different bathrooms (H1–H10) for (c) non-PMA and (d) PMA samples that can be explained by human body sites or are of unknown origin (grey). Each household is represented by a vertical bar, and values were fitted to 100%. e The proportion of microbes on different body sites that can be potentially explained by the bathroom microbiome; significant differences between treatments were defined by a Kruskal–Wallis test; **P < 0.01; ***P < 0.001. f The table shows mean values and standard deviations for (d) and (e); significant differences between treatments were defined by performing the Wilcoxon signed-rank test; NS not significant; *P < 0.05; **P < 0.01.

Fig. 3. The effects of PMA treatment on prokaryotic abundance and composition.

Quantitative PCR analysis of untreated and PMA-treated bathroom floor samples using (a) universal and (b) archaeal primer pairs. Wilcoxon signed-rank test analysis results, revealing the relative abundance of the 50 and 100 most-abundant bacterial (c) families and (d) genera, respectively, that were significantly affected by PMA treatment. Strict anaerobic taxa are marked by an asterix. Sign.: FDR-adjusted Wilcoxon signed-rank test; *P_adj < 0.05; ***P_adj < 0.01. e The relative abundance of different phenotypes as determined through BugBase. Significances were automatically defined by Mann–Whitney U test in BugBase.

Fig. 4. Archaeal community composition in indoor samples.

The relative abundance of archaeal taxa among 16S rRNA gene reads from bathroom floor surfaces at the (a) phylum and (b) genus levels. Bubble plot showing relative abundances of the 25 most-abundant archaeal genera found in bathroom and human samples: c Household samples (H1–H10) are depicted individually for non-PMA and PMA treatment of human samples (grey background) that represent reads from several body sites including nasal cavity, skin, vagina, urine, stool and oral samples. d Bubble plots of the 25 most-abundant archaeal genera, displayed according to their original samples. Relative abundance is reflected by the size of the bubbles. Genera are coloured according to their taxonomic phylum, and taxa that predominantly contain strict anaerobes are marked by an asterisk. Venn diagram of shared ASVs in (e) non-PMA indoor and (f) PMA indoor samples compared to nasal, oral, skin and stool samples (body sites that show the highest numbers of shared ASVs).

Fig. 5. Phylogenetic tree based on the sequences obtained from our study, NCBI and two other studies^,.

The circle indicates the origin of the sequences used to create the tree (see legend). The branches of the tree were coloured in different shades of green according to the taxa they represent.

Fig. 6. Observed growth of selected human-associated methanogens after exposure to 21% oxygen (air).

Plots indicate the growth as determined by (a) OD₆₀₀ measurements and (b) methane production of the tested strains after exposure to an aerobic (red) or anaerobic (blue) environment for different periods of time (n = 2). The error bars represent the standard deviation.