Premise plumbing bacterial communities in four European cities and their association with Legionella

Abstract

IntroductionLegionella species are Gram negative, facultative, intracellular bacteria found in natural and engineered water systems. Understanding the bacterial interactions underlying the success of Legionella in aquatic environments could be beneficial for control.Materials and methodsWe aimed to profile, by 16S rRNA gene amplicon sequencing (V3-V4), the bacterial communities in premise plumbing systems of buildings in four European cities (Copenhagen, Warsaw, Rome, Athens), and identify positive and negative associations of specific community members to culturable Legionella.ResultsThe coarse taxonomic composition was similar across the four cities, but Copenhagen and Warsaw had richer, more diverse communities than Athens and Rome, with a greater number of city-specific amplicon sequence variants (ASVs). The cities had statistically significant differences in bacterial communities at the ASV level, with relatively few shared ASVs. Out of 5,128 ASVs, 73 were classified as Legionella, and one or more of these were detected in most samples from each city (88.1% overall). Interestingly, the relative abundance of Legionella ASVs did not correlate with Legionella culture status. Overall, 44.2% of samples were Legionella culture positive: 71.4% in Warsaw, 62.2% in Athens, 22.2% in Rome, and 15.2% in Copenhagen. 54 specific ASVs and 42 genera had significant positive or negative associations with culturable Legionella. Negative associations included Staphylococcus, Pseudomonas, and Acinetobacter. Positive associations included several Nitrospira ASVs and one classified as Nitrosomodaceae oc32, ASVs in the amoeba-associated genera Craurococcus-Caldovatus and Reyranella, and the predatory genus Bdellovibrio.DiscussionSome of these associations are well supported by laboratory studies, but others are the opposite of what was expected. This highlights the difficulties in translating pure culture results to in complex real-life scenarios. However, these positive and negative associations held across the four cities, across multiple buildings and plumbing compartments. This is important because developing better control measures, including probiotic approaches, will require an understanding of ecological relationships that can be generalised across different engineered water systems.

Legionella relative abundance in shower hose biofilms is associated with specific microbiome members

“…4 ). A recent investigation on plumbing systems across four cities of Europe revealed associations between culturable Legionella and the microbial communities detected in the water samples (Scaturro et al 2022 ). While reinforcing the negative association between Legionella and Pseudomonas , the authors also described a negative association with the genera Staphylococcus and, consistently with our analysis, Sphingopyxis .…”

“…Moreover, both these methods would amplify and detect non-viable Legionella , of which the importance in ecological-association studies is questionable. Previous studies with a similar focus used cultivation-based quantification methods (Ma et al 2020 , Scaturro et al 2022 ), which also has known bias towards L. pneumophila and is limited by only analysing cells that grow on agar plates (Toplitsch et al 2021 ). Clearly, the method of Legionella quantification would considerably affect the outcome of any correlation/association analysis.…”

“…(3) Biofilm vs water phase: In our study, the biofilm phase was specifically collected and analysed, compared to other investigations where the water phase was analysed (Ma et al 2020 , Paranjape et al 2020 , Scaturro et al 2022 ). While noting the challenges in biofilm sampling, we do believe that this choice is critical for the determination of associations between microorganisms.…”

“…Paranjape and colleagues (Paranjape et al 2020 , Paranjape et al 2020 ) conducted two studies on cooling tower water and found that the genus Pseudomonas exhibited a strong negative correlation with Legionella , which, in turn, positively correlated with the genus Brevundimonas ; the latter positive interaction has been further demonstrated in laboratory experiments with pure culture isolates. Scaturro and co-workers (Scaturro et al 2022 ) investigated building plumbing water samples from several European cities and found positive correlations with bacteria of the genus Nitrospira , while observing negative associations with the genera Pseudomonas and Sphingopyxis .…”