Survey of Legionella

Members of the Gram-negative genus Legionella are typically found in freshwater environments, with the exception of L. longbeachae, which is present in composts and potting mixes. When contaminated aerosols are inhaled, legionellosis may result, typically as either the more serious pneumonia Legionnaires’ disease or the less severe flu-like illness Pontiac fever. It is presumed that all species of the genus Legionella are capable of causing disease in humans. As a followup to a prior clinical study of legionellosis in rural Thailand, indigenous soil samples were collected proximal to cases’ homes and workplaces and tested for the presence of legionellae by culture. We obtained 115 isolates from 22/39 soil samples and used sequence-based methods to identify 12 known species of Legionella represented by 87 isolates

Introduction of Monochloramine into a Municipal Water System: Impact on Colonization of Buildings by Legionella spp.

Legionnaires' disease (LD) outbreaks are often traced to colonized potable water systems. We collected water samples from potable water systems of 96 buildings in Pinellas County, Florida, between January and April 2002, during a time when chlorine was the primary residual disinfectant, and from the same buildings between June and September 2002, immediately after monochloramine was introduced into the municipal water system. Samples were cultured for legionellae and amoebae using standard methods. We determined predictors of Legionella colonization of individual buildings and of individual sampling sites. During the chlorine phase, 19 (19.8%) buildings were colonized with legionellae in at least one sampling site. During the monochloramine phase, six (6.2%) buildings were colonized. In the chlorine phase, predictors of Legionella colonization included water source (source B compared to all others, adjusted odds ratio [aOR], 6.7; 95% confidence interval [CI], 2.0 to 23) and the presence of a system with continuously circulating hot water (aOR, 9.8; 95% CI, 1.9 to 51). In the monochloramine phase, there were no predictors of individual building colonization, although we observed a trend toward greater effectiveness of monochloramine in hotels and single-family homes than in county government buildings. The presence of amoebae predicted Legionella colonization at individual sampling sites in both phases (OR ranged from 15 to 46, depending on the phase and sampling site). The routine introduction of monochloramine into a municipal drinking water system appears to have reduced colonization by Legionella spp. in buildings served by the system. Monochloramine may hold promise as community-wide intervention for the prevention of LD

Recurrence of Legionnaires Disease at a Hotel in the United States Virgin Islands Over a 20-Year Period

We investigated 3 cases of legionnaires disease (LD) that developed in travelers who stayed at a hotel in the United States Virgin Islands where cases of LD occurred in 1981-1982 and in 1998. The temperature of the potable water at the hotel was in a range that could optimally support the growth of Legionella species, and the potable water was colonized with Legionella pneumophila in 1981-1982 and in 2002-2003

Virulence Factors Encoded by Legionella longbeachae Identified on the Basis of the Genome Sequence Analysis of Clinical Isolate D-4968▿ †

Legionella longbeachae causes most cases of legionellosis in Australia and may be underreported worldwide due to the lack of L. longbeachae-specific diagnostic tests. L. longbeachae displays distinctive differences in intracellular trafficking, caspase 1 activation, and infection in mouse models compared to Legionella pneumophila, yet these two species have indistinguishable clinical presentations in humans. Unlike other legionellae, which inhabit freshwater systems, L. longbeachae is found predominantly in moist soil. In this study, we sequenced and annotated the genome of an L. longbeachae clinical isolate from Oregon, isolate D-4968, and compared it to the previously published genomes of L. pneumophila. The results revealed that the D-4968 genome is larger than the L. pneumophila genome and has a gene order that is different from that of the L. pneumophila genome. Genes encoding structural components of type II, type IV Lvh, and type IV Icm/Dot secretion systems are conserved. In contrast, only 42/140 homologs of genes encoding L. pneumophila Icm/Dot substrates have been found in the D-4968 genome. L. longbeachae encodes numerous proteins with eukaryotic motifs and eukaryote-like proteins unique to this species, including 16 ankyrin repeat-containing proteins and a novel U-box protein. We predict that these proteins are secreted by the L. longbeachae Icm/Dot secretion system. In contrast to the L. pneumophila genome, the L. longbeachae D-4968 genome does not contain flagellar biosynthesis genes, yet it contains a chemotaxis operon. The lack of a flagellum explains the failure of L. longbeachae to activate caspase 1 and trigger pyroptosis in murine macrophages. These unique features of L. longbeachae may reflect adaptation of this species to life in soil

Samples with higher diversity of taxa correlate with higher levels of <i>Legionellaceae</i> abundance.

<p>PCR positive samples were analyzed by 16S rRNA amplicon sequencing and the relative <i>Legionellaceae</i> abundance of each was compared to the number of bacterial families (relative abundance >1%) detected and organized by initial sample multiplex PCR result (A). The relative <i>Legionellaceae</i> abundance (B) and number of taxa (C) of samples that were positive or negative for multiplex PCR and culture were analyzed. Bars represent the geometric (B) or arithmetic (C) mean and error bars indicate standard deviation.</p

Comparison of Pinellas County CT bacterial family compositions between spring and summer 2016.

<p>Multiplex PCR results, culture results, and five highest abundance Proteobacteria family compositions were compared between spring (A) and summer (B) 2016. These samples were PCR-positive for Lspp marker only (Lspp) or all three Lspp, Lp, and Lp1 markers (Lp1). Samples were culture-positive for non-Lp species (Lspp), Lp1 isolates only (Lp1), or were culture-negative [(-)]. In samples where <i>Legionella</i> spp. were detected by PCR, <i>Legionellaceae</i> was not always among the bacterial families with the highest abundance.</p

Geographic distribution of results.

<p>Samples from eight US climate regions [Central (C), East North Central (ENC), Northeast (NE), Northwest (NW), South [Central] (SC), Southeast (SE), Southwest (SW), and West (W)] were assayed by PCR (A) and culture (B). Specimens that were positive for any of the targets by PCR underwent culture. Isolates were identified to the species (C) and <i>L</i>. <i>pneumophila</i> serogroup (D) level.</p

Phyla and Proteobacteria family profiles related to PCR and culture results.

<p>The composition of the five most abundant phyla (A, B) and five most abundant Proteobacteria families (C, D) present in cooling tower water samples were compared by overall <i>Legionella</i> PCR result (A, C) and culture result (B, D). Note that only samples that were positive by PCR underwent culture.</p

Distribution of <i>Legionella</i> and bacterial community composition among regionally diverse US cooling towers

<div><p>Cooling towers (CTs) are a leading source of outbreaks of Legionnaires’ disease (LD), a severe form of pneumonia caused by inhalation of aerosols containing <i>Legionella</i> bacteria. Accordingly, proper maintenance of CTs is vital for the prevention of LD. The aim of this study was to determine the distribution of <i>Legionella</i> in a subset of regionally diverse US CTs and characterize the associated microbial communities. Between July and September of 2016, we obtained aliquots from water samples collected for routine <i>Legionella</i> testing from 196 CTs located in eight of the nine continental US climate regions. After screening for <i>Legionella</i> by PCR, positive samples were cultured and the resulting <i>Legionella</i> isolates were further characterized. Overall, 84% (164) were PCR-positive, including samples from every region studied. Of the PCR-positive samples, <i>Legionella</i> spp were isolated from 47% (78), <i>L</i>. <i>pneumophila</i> was isolated from 32% (53), and <i>L</i>. <i>pneumophila</i> serogroup 1 (Lp1) was isolated from 24% (40). Overall, 144 unique <i>Legionella</i> isolates were identified; 53% (76) of these were <i>Legionella pneumophila</i>. Of the 76 <i>L</i>. <i>pneumophila</i> isolates, 51% (39) were Lp1. <i>Legionella</i> were isolated from CTs in seven of the eight US regions examined. 16S rRNA amplicon sequencing was used to compare the bacterial communities of CT waters with and without detectable <i>Legionella</i> as well as the microbiomes of waters from different climate regions. Interestingly, the microbial communities were homogenous across climate regions. When a subset of seven CTs sampled in April and July were compared, there was no association with changes in corresponding CT microbiomes over time in the samples that became culture-positive for <i>Legionella</i>. <i>Legionella</i> species and Lp1 were detected frequently among the samples examined in this first large-scale study of <i>Legionella</i> in US CTs. Our findings highlight that, under the right conditions, there is the potential for CT-related LD outbreaks to occur throughout the US.</p></div