No abstract

Legionellae are gram-negative bacteria most commonly found in freshwater ecosystems and purpose-built water systems. In humans, the bacterium causes Legionnaires’ disease (LD) or a Pontiac fever. In this study, the different waters (drinking water, pool water, cooling towers) in which Legionella pneumophila has been isolated were studied to assess the possible risk of bacterial spreading in the population. The influence of physical and chemical parameters, and interactions with Acanthamoeba castellanii on L. pneumophila, were analyzed by Heterotrophic Plate Count, the Colony-forming units (CFU) methods, transmission electron microscopy (TEM), and Sequence-Based Typing (SBT) analysis. During the study period (2013–2019), a total of 1932 water samples were analyzed, with the average annual rate of Legionella-positive water samples of 8.9%, showing an increasing trend. The largest proportion of Legionella-positive samples was found in cooling towers and rehabilitation centers (33.9% and 33.3%, respectively). Among the isolates, L. pneumophila SGs 2–14 was the most commonly identified strain (76%). The survival of Legionella was enhanced in the samples with higher pH values, while higher electrical conductivity, nitrate, and free residual chlorine concentration significantly reduced the survival of Legionella. Our results show that growth in amoeba does not affect the allelic profile, phenotype, and morphology of the bacterium but environmental L. pneumophila becomes more resistant to pasteurization treatment.

Research background. Legionella are Gram-negative bacteria that are ubiquitous in the natural environment. Contaminated water in manmade water systems is a potential source of transmission of Legionnaires’ disease (LD). The aim of this study was to explore the prevalence of Legionella pneumophila (L. pneumophila) in the drinking water distribution system (DWDS) of Primorje-Gorski Kotar County (PGK County), Croatia, for the period 2013-2019, coupled with the incidence of LD. A number of L. pneumophila-positive samples (>100 CFU/L), serogroup distribution, and the degree of contamination of specific facilities (health & aged care, tourism, sports) were assessed. Based on the results obtained, the reasoning for the implementation of a mandatory Legionella environmental surveillance program was assessed. Experimental approach. Sample testing for Legionella was carried out according to ISO 1173. A Heterotrophic Plate Count (HPC) and P. aeruginosa were analysed along with the basic physico-chemical indicators of drinking water quality. The research period was divided into two parts, namely, the 2013-2018 period (before implementation of the prevention program, after the outbreak of LD), and year 2019 (proactive approach, no LD cases recorded). Results and conclusion. During the 7-year observation period in PGK County, an increase in the number of samples tested for Legionella was found. An increase in Legionella-positive samples (particularly pronounced during the warmer part of the year) was recorded, along with a growing trend in the number of reported LD cases. In addition to hot water systems, the risk of Legionella colonization also applies to cold water systems. Health & aged care facilities appear to be at highest risk. In addition to the higher proportion of positive samples and a higher degree of microbiological load at these facilities, the highest proportion of L. pneumophila SGs 2-14 was identified. Due to the diagnostic limitations of the applied tests, the number of LD cases is underdiagnosed. Novelty and scientific contribution. The introduction of a mandatory preventive approach to monitoring Legionella in DWDS water samples, along with the definition of national criteria for the interpretation of results, will create the preconditions for diagnosis and adequate treatment of larger numbers of LD cases.