Detection of Diverse Sequence Types of Legionella pneumophila by Legiolert Enzymatic-Based Assay and the Development of a Long-Term Storage Protocol

Matthews, Sara; Trigui, Hana; Grimard-Conea, Marianne; Reyes, Elliston Vallarino; Villiard, Gabriel; Charron, Dominique; Bédard, Émilie; Faucher, Sébastien P.; Prévost, Michèle

doi:10.1128/spectrum.02118-22

Cited by 4 publications

(2 citation statements)

References 35 publications

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“…pneumophila clade shared the ancestral allele at this locus with Philadelphia-1. In contrast, all assemblies ( 71 – 74 ) representing the pascullei , raphaeli, fraseri , and D-7708 ( 71 ) subspecies cluster (comprising 64/302 or 21.2% of analyzed assemblies) contained the derived DnaK M94I allele. The uneven distribution of Legionella subspecies has been proposed to reflect an underlying occupation of separate ecological niches ( 71 ), so we chose to conduct follow-up experiments on the dnaK mutations observed in our heat-adapted lineages.…”

Section: Resultsmentioning

confidence: 95%

Development of heat-shock resistance in Legionella pneumophila modeled by experimental evolution

Liang,

Cameron,

Faucher

2023

Appl Environ Microbiol

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Because it can grow in buildings with complex hot water distribution systems (HWDS), healthcare facilities recognize the waterborne bacterium Legionella pneumophila as a major nosocomial infection threat and often try to clear the systems with a pasteurization process known as superheat-and-flush. After this treatment, many facilities find that the contaminating populations slowly recover, suggesting the possibility of in situ evolution favoring increased survival in high-temperature conditions. To mimic this process in a controlled environment, an adaptive laboratory evolution model was used to select a wild-type strain of L. pneumophila for survival to transient exposures to temperatures characteristic of routine hot water use or failed pasteurization processes in HWDS. Over their evolution, these populations became insensitive to exposure to 55°C and developed the ability to survive short exposures to 59°C heat shock. Heat-adapted lineages maintained a higher expression of heat-shock genes during low-temperature incubation in freshwater, suggesting a pre-adaptation to heat stress. Although there were distinct mutation profiles in each of the heat-adapted lineages, each acquired multiple mutations in the DnaJ/DnaK/ClpB disaggregase complex, as well as mutations in chaperone htpG and protease clpX . These mutations were specific to heat-shock survival and were not seen in control lineages included in the experimental model without exposure to heat shock. This study supports in situ observations of adaptation to heat stress and demonstrates the potential of L. pneumophila to develop resistance to control measures. IMPORTANCE As a bacterium that thrives in warm water ecosystems, Legionella pneumophila is a key factor motivating regulations on hot water systems. Two major measures to control Legionella are high circulating temperatures intended to curtail growth and the use of superheat-and-flush pasteurization processes to eliminate established populations. Facilities often suffer recolonization of their hot water systems; hospitals are particularly at risk due to the severe nosocomial pneumoniae caused by Legionella . To understand these long-term survivors, we have used an adaptive laboratory evolution model to replicate this process. We find major differences between the mutational profiles of heat-adapted and heat-naïve L. pneumophila populations including mutations in major heat-shock genes like chaperones and proteases. This model demonstrates that well-validated treatment protocols are needed to clear contaminated systems and—in an analog to antibiotic resistance—the importance of complete eradication of the resident population to prevent selection for more persistent bacteria.

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Section: Resultsmentioning

confidence: 95%

Development of heat-shock resistance in Legionella pneumophila modeled by experimental evolution

Liang,

Cameron,

Faucher

2023

Appl Environ Microbiol

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“…This additional step was not strictly necessary and was taken to enable collection of L. pneumophila isolates for sequencing. A recent study (Matthews et al, 2022) found that L. pneumophila isolates can also be obtained directly from the Legiolert ® tray for serotyping and/or storage for a year or more. If ISO Method 11731:2017 is used instead of Legiolert ® , follow-up analyses of positive samples should identify organisms to the species level and serogroup for L. pneumophila isolates.…”

Section: Protocolmentioning

confidence: 99%

Protocol for responding to the detection of Legionella pneumophila in drinking water distribution systems

Bartrand,

LeChevallier,

Clancy

et al. 2024

AWWA Water Science

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A protocol for responding to Legionella pneumophila detections in distribution system samples is presented and justified. The protocol was developed using existing protocols for building water systems, vetted in workshops, and provided to utilities participating in a research project. The protocol provides action levels and actions that utilities can take when L. pneumophila is detected in distribution system samples. Action levels are based on the best available science and expert judgment and are similar to those in protocols for assessing building water system monitoring results. Action levels should be reassessed as additional data and knowledge on the occurrence and growth potential of L. pneumophila in distribution systems become available. All positive detections trigger assessments that focus first on conditions local to the sample location and then extend further into the distribution system. Higher concentration or more frequent detections initiate more assertive responses and communication than lower concentration and frequency detections. The protocol provides a starting point for the development of protocols for purposes such as L. pneumophila operational surveillance or within an outbreak investigation.

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Assessment of monitoring approaches to control Legionella pneumophila within a complex cooling tower system

Trigui,

Matthews,

Bedard

et al. 2024

Science of The Total Environment

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Detection of Diverse Sequence Types of Legionella pneumophila by Legiolert Enzymatic-Based Assay and the Development of a Long-Term Storage Protocol

Abstract: Legionnaires’ disease is caused by the bacterium Legionella pneumophila , which can be found in a variety of water systems. When outbreaks of Legionnaires’ disease occur, it is necessary to find the water systems transmitting the bacterium to humans.

Cited by 4 publications

References 35 publications

Development of heat-shock resistance in Legionella pneumophila modeled by experimental evolution

Development of heat-shock resistance in Legionella pneumophila modeled by experimental evolution

Protocol for responding to the detection of Legionella pneumophila in drinking water distribution systems

Assessment of monitoring approaches to control Legionella pneumophila within a complex cooling tower system

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