Abstract
Pseudomonas species are frequent inhabitants of freshwater environments and colonizers of water supply networks via bioadhesion and biofilm formation. P. aeruginosa is the species most commonly associated with human disease, causing a wide variety of infections with links to its presence in freshwater systems. Though several other Pseudomonas species are of ecological and public health importance, little knowledge exists regarding environmental abundances of these species. In the present study, an Illumina-based next-generation sequencing (NGS) approach using Pseudomonas-specific primers targeting the 16S rRNA gene was evaluated and applied to a set of freshwater samples from different environments including a cooling tower sampled monthly during 2 years. Our approach showed high in situ specificity and accuracy. NGS read counts revealed a precise quantification of P. aeruginosa and a good correlation with the absolute number of Pseudomonas genome copies in a validated genus-specific qPCR assay, demonstrating the ability of the NGS approach to determine both relative and absolute abundances of Pseudomonas species and P. aeruginosa. The characterization of Pseudomonas communities in cooling tower water allowed us to identify 43 phylotypes, with P. aeruginosa being the most abundant. A shift existed within each year from a community dominated by phylotypes belonging to P. fluorescens and P. oleovorans phylogenetic groups to a community where P. aeruginosa was highly abundant. Co-occurrence was observed between P. aeruginosa and other phylotypes of P. aeruginosa group as well as the potentially pathogenic species P. stutzeri, but not with phylotypes of the P. fluorescens group, indicating the need to further investigate the metabolic networks and ecological traits of Pseudomonas species. This study demonstrates the potential of deep sequencing as a valuable tool in environmental diagnostics and surveillance of health-related pathogens in freshwater environments.
Highlights
The genus Pseudomonas, currently comprising about 150 species, contains several species of high ecological importance that are linked to a large array of catabolic functions, i.e., bioremediation, biodegradation, and biosorption (Sayler et al, 1990; Banat, 1995; Zhang et al, 2005; Wasi et al, 2013)
We aimed to develop a robust next-generation sequencing (NGS) assay to be applied for surveillance and monitoring of Pseudomonas species, including the most potentially pathogenic species, P. aeruginosa, in freshwater environments
Freshwater is the natural reservoir for Pseudomonas species and the main source of human exposure
Summary
The genus Pseudomonas, currently comprising about 150 species, contains several species of high ecological importance that are linked to a large array of catabolic functions, i.e., bioremediation, biodegradation, and biosorption (Sayler et al, 1990; Banat, 1995; Zhang et al, 2005; Wasi et al, 2013). P. aeruginosa is a major pathogen in a wide variety of hospital-acquired infections and the main causative agent of respiratory tract infections in patients with bronchiectasis and cystic fibrosis, causing high health-care costs, clinical morbidity and mortality (Jones et al, 2002; Lyczak et al, 2002; Al-Aloul et al, 2004; Moradali et al, 2017). The exposure of personnel and patients in hospital facilities to P. aeruginosa occurs by means of bacterial colonization, with biofilm formation in the plumbing system and transmission occurring initially through contact and aerosol generation in contaminated areas, and posteriorly by horizontal acquisition (Morrison and Wenzel, 1984; Döring et al, 1991; Bergmans et al, 1998; Trautmann et al, 2005). Less frequent modes of transmission such as drinking water consumption and aerosol breathing have been described (Mena and Gerba, 2009)
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