LEGIONELLA - World Health Organization

During outbreak investigations, swabs should be taken in conjunction with water samples from
sites where biofilms are likely to form. These swabs can be taken from various points within
plumbing systems, from surfaces such as biofilms, and from areas that are difficult to reach,
such as within the jets of hot tubs (see Chapter 8), thermostatic mixer valves or showers. The
swabs can be submerged in a small volume of water taken at the same time, or in Pages’s
saline to prevent drying during transportation to the laboratory.
All samples should be transported to the laboratory in dark, insulated containers to protect
them from extreme temperatures and from light.
Information should be gathered to help interpret the results. As a minimum, the following
information should be included on the request form:
• the site and sample point
• the sample references and date
• the reason for sampling
• the temperature of the sample source (e.g. the temperature of a hot-water system at one
minute after turning on the tap, and at two minutes after turning on the cold tap)
• any biocide used
• the timing of the dosage in relation to sampling
• the concentration detected at the time of sampling
• any other risk factors of importance (e.g. closed system opened for maintenance)
• high risk of nutrient present, such as in plastics manufacturing plants
• any cases associated with the site.
11.4.5 Sample preparation and isolation
Isolation methods for clinical and environmental samples differ. Legionellae are usually a very
minor component of the total bacterial population in environmental samples, and are rarely
present in high numbers. Thus, when working with environmental samples, it is usually necessary
to first concentrate the microfloras. In the case of clinical specimens such as sputa and tissue
biopsies, these may need to be homogenised before culture; in contrast, the organisms in
fluids such as bronchiolar lavages will need to be concentrated by centrifuging. For both
environmental concentrates and clinical samples, it is necessary to eliminate or suppress the
competing background flora during primary culture.
Legionellae and background bacteria can be concentrated from water samples by centrifugation
or membrane filtration, or by a combination of the two. Recovery in the presence of other
bacterial species present in the sample can be improved by heating, usually at 50 ºC for
LEGIONELLA AND THE PREVENTION OF LEGIONELLOSIS