Cutting edge Legionella testing and treatment technologies
By Fabian Basker RSci MRSB, Dr Kenneth Scally MSc FRSC, and Dr Vicente Catalán
In the ever-evolving landscape of public health, staying one step ahead of emerging threats is crucial. Legionella, a persistent bacterium known for its association with legionnaires' disease, demands our attention now more than ever.
Legionella is a potential threat to man-made water systems, including cooling towers, hotel and healthcare water networks; and to natural bodies of water such as rivers, streams, and lakes. The more stagnant the water systems (in temperatures ranging from 20-50°C), the more these potentially life-threatening bacteria can proliferate.
Traditional approaches to Legionella prevention and detection have often fallen short in combating this elusive pathogen due to a variety of factors: inconsistency or a lack of testing, inadequate laboratory equipment, and post-antibiotics testing, among others. Breakthrough technologies, however, may now be paving the way for a paradigm shift in Legionella management.
Contemporary Analytical Methods for Legionella Testing
The culture and Polymerase Chain Reaction (PCR) methods are current common ways of determining the presence of Legionella. Traditional culture methods primarily follow ISO 11731:2017 guidelines, which are considered the gold standard in terms of environmental quality monitoring worldwide.
When cultured and observed in the lab, Legionella from water samples can take between 10-13 days to grow and confirm, after which final results can be obtained. Some of this method's limitations include the slow growth rate of bacteria, which results in a long incubation time; non-detection of viable but non-culturable bacteria, which could pose a public health risk; and the challenge of isolating Legionella from other microorganisms in samples.
The PCR method, on the other hand, is very useful in detecting Legionella spp. including L. pneumophila in both clinical and environmental samples. Apart from providing high sensitivity and specificity, the method also has low detection limits and is relatively fast, with results available in just a few hours. Moreover, quantitative PCR (qPCR) can quantify samples with greater accuracy.
On the other hand, PCR is not immune to its limitations. These include the potential presence of PCR inhibitors, which can increase sample limit of detection, and the method's inability to differentiate between live and dead cells due to DNA persisting long after cell death.
Most Probable Number (MPN) testing provides a culture-based alternative for L. pneumophila detection only, delivering results in seven days instead of 10-13 days. It is user-friendly, cost-effective, and can be processed by those with minimal technical experience, potentially eliminating the need for laboratory testing.
Amoeba co-culture, employing Acanthamoeba spp., improves the ability to cultivate bacteria, enabling detection even when bacteria reside within other organisms and fail to grow using conventional culture techniques. However, it should be noted that the results obtained from this method are primarily qualitative.
Advanced Molecular-Based Methodologies for Legionella Detection
In the realm of innovation, advanced molecular-based methodologies are transforming the way we detect Legionella. The advent of Digital Droplet PCR (ddPCR), for instance, has provided greater sensitivity and precision to Legionella testing. While more costly than standard thermocyclers used in qPCR, it enables absolute DNA quantification, requires minimal technical expertise and fewer reagents.
One other emerging technique is Lateral Flow Testing, a simple yet effective method that offers rapid, on-site water systems screening. It is user-friendly and is combined with additional sample concentration steps. A downside is that many of these tests have high detection limits, making it challenging to identify low concentrations effectively.
Fluorescence in situ hybridisation (FISH), another innovative approach, utilises fluorescently labelled probes to detect Legionella cells directly. This technique enhances the visualisation and allows for the quantification of Legionella populations, including L. pneumophila.
Alternative Legionella Detection Techniques
Flow cytometry (FC), in combination with immunomagnetic separation, is another useful technique for environmental testing. It offers better recovery rates compared to standard culture methods. While FC allows for real-time detection and quantification of bacteria, it is typically used to detect a wide range of viable bacteria, not just Legionella. There are, however, specific systems available in the market designed specifically for Legionella testing.
Next-Generation Sequencing (NGS) is a promising method for analysing bacterial diversity, including Legionella, and tracing infection sources. However, interpreting NGS data requires expertise due to its high learning curve. Yet, it is potentially a valuable tool in unravelling the mysteries of bacterial populations.
While it shares certain limitations inherent to other PCR methods, Loop-Mediated Isothermal Amplification (LAMP) offers an affordable alternative to PCR, allowing amplification at a constant temperature without requiring costly equipment.
Microcolony Enumeration (MICA) is a method that relies on detecting tiny colonies before they become visible, offering advantages in reducing errors, saving resources, and cutting costs while improving efficiency. It has its limitations, however, including low recovery rates and the inability to detect non-culturable cells that can still cause disease.
Viability PCR uses chemicals to detect dead cells and free DNA, providing a safe way to identify non-viable bacteria. However, it may still count bacteria killed by methods that keep the cell intact, such as UV treatment.
Legionella testing, the SUEZ way
Latis Scientific, a SUEZ company, performs the culture method for a majority of its Legionella sample testing, or 15,000-20,000 samples processed monthly. If a client wishes to, Latis Scientific can also perform qPCR, alongside culture testing which is a regulatory compliance requirement.
Samples are promptly collected and transported to Latis Scientific within 24 hours, and carefully stored at between 6-20°C to maintain their integrity. After assigning a unique sample number, the samples are then subjected to rigorous testing using the Laboratory Information Management System (LIMS). This ensures accurate and organised data management where test results are meticulously entered into the system, thoroughly checked for quality assurance, and promptly reported back to the client.
To expedite the overall process, Latis Scientific employs SamplefyTM, a digitised logistics system that proactively allocates relevant information before the samples even arrive at the laboratory. This results in streamlined operations and reduction of turnaround time.
SUEZ also offers expert consultancy drawing upon 40 years of experience in managing and remediating risks associated with Legionella and other bacteria in various settings. The Latis Scientific methodologies are driven by industry regulations and guidelines, including accredited ISO 17025:2017 culture and qPCR methods, ensuring the highest quality standards.
The dedicated laboratory team develops, tests, and validates new Legionella methodologies such as amoeba co-culture and flow cytometry, among others. By staying at the forefront of technological advancements, SUEZ ensures the laboratory team are equipped with the most effective tools to address emerging challenges in Legionella management.