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Legionella pneumophila pangenome reveals strain-specific virulence factors
Giuseppe D'Auria, Nuria Jiménez-Hernández, Francesc Peris-Bondia, Andrés Moya, Amparo Latorre
BMC Genomics , 2010, DOI: 10.1186/1471-2164-11-181
Abstract: We have sequenced the genome of the particularly persistent L. pneumophila strain Alcoy 2300/99 and have compared it with four previously sequenced strains known as Philadelphia (USA), Lens (France), Paris (France) and Corby (England).Pangenome analysis facilitated the identification of strain-specific features, as well as some that are shared by two or more strains. We identified: (1) three islands related to anti-drug resistance systems; (2) a system for transport and secretion of heavy metals; (3) three systems related to DNA transfer; (4) two CRISPR (Clustered Regularly Interspaced Short Palindromic Repeats) systems, known to provide resistance against phage infections, one similar in the Lens and Alcoy strains, and another specific to the Paris strain; and (5) seven islands of phage-related proteins, five of which seem to be strain-specific and two shared.The dispensable genome disclosed by the pangenomic analysis seems to be a reservoir of new traits that have mainly been acquired by horizontal gene transfer and could confer evolutionary advantages over strains lacking them.Legionella pneumophila is a gram-negative facultative intracellular pathogen, identified as the infectious agent of the Legionnaire's disease (LD) or Legionellosis in 1977 [1]. It is found in aquatic environments parasitizing its natural hosts, amoebae and protozoa. From this environment, Legionella can colonize water treatment plants, such as refrigeration towers, potable water pipes, etc., and can cause infections in humans, when infected aerosols are inhaled [2,3]. Despite efforts to keep water systems free of Legionella, this pathogen is still causing infection throughout the world, including Spain, where it is endemic in some areas. From 1989 to 2005, around 310 outbreaks with 2,974 cases were recorded worldwide. In 2002 and 2005 there were two important epidemic events with 1,461 and 1,292 cases respectively. In Alcoy, an industrial town in the Valencian Region (Spain), a large outbre
Development of a miniaturized DNA microarray for identification of 66 virulence genes of Legionella pneumophila
Mariusz ?ak,Piotr Zaborowski,Milena Baczewska-Rej,Aleksandra A. Zasada
Post?py Higieny i Medycyny Do?wiadczalnej , 2011,
Abstract: Introduction:For the last five years, Legionella sp. infections and legionnaire’s disease in Poland have been receiving a lot of attention, because of the new regulations concerning microbiological quality of drinking water. This was the inspiration to search for and develop a new assay to identify many virulence genes of Legionella pneumophila to better understand their distribution in environmental and clinical strains. The method might be an invaluable help in infection risk assessment and in epidemiological investigations.Material/Methods:The microarray is based on Array Tube technology. It contains 3 positive and 1 negative control. Target genes encode structural elements of T4SS, effector proteins and factors not related to T4SS. Probes were designed using OligoWiz software and data analyzed using IconoClust software. To isolate environmental and clinical strains, BAL samples and samples of hot water from different and independent hot water distribution systems of public utility buildings were collected.Results.We have developed a miniaturized DNA microarray for identification of 66 virulence genes of L. pneumophila. The assay is specific to L. pneumophila sg 1 with sensitivity sufficient to perform the assay using DNA isolated from a single L. pneumophila colony. Seven environmental strains were analyzed. Two exhibited a hybridization pattern distinct from the reference strain.Discussion:The method is time- and cost-effective. Initial studies have shown that genes encoding effector proteins may vary among environmental strains. Further studies might help to identify set of genes increasing the risk of clinical disease and to determine the pathogenic potential of environmental strains.
Virulence factor rtx in Legionella pneumophila, evidence suggesting it is a modular multifunctional protein
Giuseppe D'Auria, Núria Jiménez, Francesc Peris-Bondia, Carmen Pelaz, Amparo Latorre, Andrés Moya
BMC Genomics , 2008, DOI: 10.1186/1471-2164-9-14
Abstract: The comparative analysis of rtx gene among 6 strains of L. pneumophila showed modularity in their structures. Among compared genomes, the N-terminal region of the protein presents highly dissimilar repeats with functionally similar domains. On the contrary, the C-terminal region is maintained with a fashionable modular configuration, which gives support to its proposed role in adhesion and pore formation. Despite the variability of rtx among the considered strains, the flanking genes are maintained in synteny and similarity.In contrast to the extracellular bacteria Vibrio cholerae, in which the rtx gene is highly conserved and flanking genes have lost synteny and similarity, the gene region coding for the Rtx toxin in the intracellular pathogen L. pneumophila shows a rapid evolution. Changes in the rtx could play a role in pathogenicity. The interplay of the Rtx toxin with host membranes might lead to the evolution of new variants that are able to escape host cell defences.Legionella pneumophila is a gram negative, gamma-proteobacteria organism whose natural hosts are amoebae and protozoa. This bacterium can infect humans by inhalation of aerosols [1,2] entering alveolar macrophages causing the well-known, and often lethal, Legionnaires' disease (LD) or Legionellosis. Despite the great number of isolates of L. pneumophila, the ones belonging to serogroup 1 are responsible of about 80 to 90% of cases of Legionellosis [3]. The first critical event during infection by L. pneumophila involves the macrophages by the action of the type IV secretion system, which prevents the fusion of the phagosome with the lysosome and its acidification [4,5]. It has been demonstrated that these events start very early after the infection [6]. Several mechanisms play an important role in the formation of infection vacuoles. Legionella enters the macrophages by vacuoles that are morphologically similar to macropinosomes by an unusual mechanisms called "coiling phagocytosis" [6,7]. The vac
Molecular diversity and high virulence of Legionella pneumophila strains isolated from biofilms developed within a warm spring of a thermal spa
Chaabna Zineddine,Forey Fran?oise,Reyrolle Monique,Jarraud Sophie
BMC Microbiology , 2013, DOI: 10.1186/1471-2180-13-17
Abstract: Background Several cases of legionellosis have been diagnosed in the same French thermal spa in 1986, 1994 and 1997. L. pneumophila serogroup 1 (Lp1) strains have been isolated from several patients, but the source of contamination was not identified despite the presence of different Lp1 in water samples of the three natural springs feeding the spa at this period. Results Our strategy was to investigate L. pneumophila (Lp) strains from natural biofilms developed in a sulphur-rich warm spring of this contaminated site. Biofilm analysis revealed the presence of three Lp serogroups (Lp1, Lp10 and Lp12). Surprisingly, Lp10 and Lp12 were not reported in the previous described studies from water samples. Besides, the new seven Lp1 we isolated exhibit a high molecular diversity and have been differentiated in five classes according to their DNA genome patterns obtained by PFGE and mip sequences. It must be noted that these DNA patterns are original and unknown in databases. Interestingly, the 27 Lp environmental strains we isolated display a higher cytotoxicity and virulence towards the amoeba Acanthamoeba castellanii than those of known Lp1 epidemic strains. Conclusion The characteristics of Legionella pneumophila Lp1 strains isolated from the warm spring are in agreement with their presence in biofilms and their probable long-term persistence in this ecosystem.
The ClpP protease homologue is required for the transmission traits and cell division of the pathogen Legionella pneumophila
Xiang-hui Li, Yong-lun Zeng, Ye Gao, Xiao-cong Zheng, Qin-fen Zhang, Shi-ning Zhou, Yong-jun Lu
BMC Microbiology , 2010, DOI: 10.1186/1471-2180-10-54
Abstract: In this study, we showed that ClpP was required for optimal growth of L. pneumophila at high temperatures and under several other stress conditions. We also observed that cells devoid of clpP exhibited cell elongation, incomplete cell division and compromised colony formation. Furthermore, we found that the clpP-deleted mutant was more resistant to sodium stress and failed to proliferate in the amoebae host Acanthamoeba castellanii.The data present in this study illustrate that the ClpP protease homologue plays an important role in the expression of transmission traits and cell division of L. pneumophila, and further suggest a putative role of ClpP in virulence regulation.Legionella pneumophila, a Gram-negative, intracellular bacterial pathogen, is the opportunistic agent responsible for a severe form of pneumonia named Legionnaires' disease and the less severe flu-like Pontiac fever [1,2]. The remarkable capability of L. pneumophila to colonize a wide range of natural protozoa and mammalian host cells is mostly attributed to its unique Type IVB secretory system (T4BSS) whose components are encoded by the dot (defect in organelle trafficking) and icm (intracellular multiplication) genes [3-6]. L. pneumophila uses the Dot/Icm apparatus to inject effectors into the host cells to promote invasion and to modulate organelle trafficking, which in turn leads to formation of replication-permissive endosomes [7-9].Similar to a variety of microbes, L. pneumophila undergoes a life cycle characterized by a biphasic conversion between a vegetative replicative form and a non-replicating, infectious and stress resistant transmissive form. On one hand, bacteria cultured in broth to either exponential or stationary phase display many similar attributes shared by the replicative and transmissive forms, respectively [10,11]. For example, upon the transition from exponential phase to stationary phase, L. pneumophila becomes more infectious and more resistant to various stresses [12]. F
Biofilms: The Stronghold of Legionella pneumophila  [PDF]
Mena Abdel-Nour,Carla Duncan,Donald E. Low,Cyril Guyard
International Journal of Molecular Sciences , 2013, DOI: 10.3390/ijms141121660
Abstract: Legionellosis is mostly caused by Legionella pneumophila and is defined as a severe respiratory illness with a case fatality rate ranging from 5% to 80%. L. pneumophila is ubiquitous in natural and anthropogenic water systems. L. pneumophila is transmitted by inhalation of contaminated aerosols produced by a variety of devices. While L. pneumophila replicates within environmental protozoa, colonization and persistence in its natural environment are also mediated by biofilm formation and colonization within multispecies microbial communities. There is now evidence that some legionellosis outbreaks are correlated with the presence of biofilms. Thus, preventing biofilm formation appears as one of the strategies to reduce water system contamination. However, we lack information about the chemical and biophysical conditions, as well as the molecular mechanisms that allow the production of biofilms by L. pneumophila. Here, we discuss the molecular basis of biofilm formation by L. pneumophila and the roles of other microbial species in L. pneumophila biofilm colonization. In addition, we discuss the protective roles of biofilms against current L. pneumophila sanitation strategies along with the initial data available on the regulation of L. pneumophila biofilm formation.
Legionella pneumophila serogroup 5 infection in the presence of multiple environmental contamination. The importance of a bacteriological diagnosis
Maria Teresa Montagna,Maria Luisa Ricci,Christian Napoli,Daniela Tat
Italian Journal of Public Health , 2007, DOI: 10.2427/5906
Abstract: Legionella pneumophila is a pathogen that causes severe pneumonia in humans; L. pneumophila serogroup 1 accounts for at least 90% of infections. This is not linked to an environmental predominance of Legionella pneumophila 1, but may be due to a greater virulence of the strain. L. pneumophila sg 5 has also been reported, albeit less frequently, to be a cause of the disease. We report a case of L. pneumophila sg 5 occurring in a large hospital in southern Italy (Apulia region), where both L. pneumophila sg 1 and sg 5 were detected in the water supply; the nosocomial origin was demonstrated by molecular subtyping (PFGE). An environmental investigation, performed immediately after diagnosis of the case of legionellosis, identified a ow L. pneumophila sg 5 contamination level. Our experience highlights that in hospital, risk assessment, in rder to institute control measures for Legionella, should be carried out not only in response to a case of the disease and/or in risk wards only, as described in the Italian Guidelines, but periodically in every ward. The present study confirms that, although in the community L. pneumophila sg 1 is the most frequent strain isolated in both outbreaks and isolated cases, in hospital other serogroups and species may often cause infection because of the high susceptibility of the hosts.
Extensive recombination events and horizontal gene transfer shaped the Legionella pneumophila genomes
Laura Gomez-Valero, Christophe Rusniok, Sophie Jarraud, Benoit Vacherie, Zoé Rouy, Valerie Barbe, Claudine Medigue, Jerome Etienne, Carmen Buchrieser
BMC Genomics , 2011, DOI: 10.1186/1471-2164-12-536
Abstract: We show that L. pneumophila Sg1 has a highly conserved and syntenic core genome that comprises the many eukaryotic like proteins and a conserved repertoire of over 200 Dot/Icm type IV secreted substrates. However, recombination events and horizontal gene transfer are frequent. In particular the analyses of the distribution of nucleotide polymorphisms suggests that large chromosomal fragments of over 200 kbs are exchanged between L. pneumophila strains and contribute to the genome dynamics in the natural population. The many secretion systems present might be implicated in exchange of these fragments by conjugal transfer. Plasmids also play a role in genome diversification and are exchanged among strains and circulate between different Legionella species.Horizontal gene transfer among bacteria and from eukaryotes to L. pneumophila as well as recombination between strains allows different clones to evolve into predominant disease clones and others to replace them subsequently within relatively short periods of time.Legionella pneumophila is the etiologic agent of Legionnaires' disease, an atypical pneumonia, which is often fatal if not treated promptly. However, it is principally an environmental bacterium that inhabits fresh water reservoirs worldwide where it parasitizes within free-living protozoa but also survives in biofilms [1-3]. Since L. pneumophila does not spread from person-to-person, humans have been inconsequential for the evolution of this pathogen. Instead, the virulence strategies of L. pneumophila have been shaped by selective pressures in aquatic ecosystems. Indeed, the co-evolution of L. pneumophila with fresh-water amoebae is reflected in its genome sequence. The analysis of two L. pneumophila genomes identified the presence of an unexpected high number and variety of eukaryotic-like proteins and proteins containing motifs mainly found in eukaryotes [4]. These proteins were predicted to interfere in different steps of the infectious cycle by mimick
Metabolism of the vacuolar pathogen Legionella and implications for virulence  [PDF]
Christian Manske,Hubert Hilbi
Frontiers in Cellular and Infection Microbiology , 2014, DOI: 10.3389/fcimb.2014.00125
Abstract: Legionella pneumophila is a ubiquitous environmental bacterium that thrives in fresh water habitats, either as planktonic form or as part of biofilms. The bacteria also grow intracellularly in free-living protozoa as well as in mammalian alveolar macrophages, thus triggering a potentially fatal pneumonia called “Legionnaires' disease.” To establish its intracellular niche termed the “Legionella-containing vacuole” (LCV), L. pneumophila employs a type IV secretion system and translocates ~300 different “effector” proteins into host cells. The pathogen switches between two distinct forms to grow in its extra- or intracellular niches: transmissive bacteria are virulent for phagocytes, and replicative bacteria multiply within their hosts. The switch between these forms is regulated by different metabolic cues that signal conditions favorable for replication or transmission, respectively, causing a tight link between metabolism and virulence of the bacteria. Amino acids represent the prime carbon and energy source of extra- or intracellularly growing L. pneumophila. Yet, the genome sequences of several Legionella spp. as well as transcriptome and proteome data and metabolism studies indicate that the bacteria possess broad catabolic capacities and also utilize carbohydrates such as glucose. Accordingly, L. pneumophila mutant strains lacking catabolic genes show intracellular growth defects, and thus, intracellular metabolism and virulence of the pathogen are intimately connected. In this review we will summarize recent findings on the extra- and intracellular metabolism of L. pneumophila using genetic, biochemical and cellular microbial approaches. Recent progress in this field sheds light on the complex interplay between metabolism, differentiation and virulence of the pathogen.
Investigation of Legionella Pneumophila serogroup 1 Population in Morocco by Monoclonal Antibody subtyping ,  [PDF]
Mariam Mekkour,El Khalil Ben Driss,Sophie Jarraud,Jalila Tai
International Journal of Emerging Sciences , 2013,
Abstract: Legionella Pneumophila is a common cause of hospital and community-acquired pneumonia, being transmitted by inhalation of aqueous aerosols. Most legionellosis outbreaks are linked to contaminated hot water systems and cooling towers. The aim of this preliminary study was to determine the population diversity of (n=40) environmental strains of L. pneumophila serogroup 1 recovered from the hot water distribution systems of nine establishments in four Moroccan towns during the period 2009–2011. Four subgroups were detected by monoclonal antibody technique. A total of 60% of isolates (n=24) had the virulence-associated epitope that is recognized by MAb 3/1 (32.5% of isolates belong to France/Allentown and 27.5% to Benidorm MAb subgroups). Forty percent (40%) of isolates, are not recognized by Mab 3/1 (27.5% of isolates belong to Olda and 12.5 % to Bellingham MAb subgroups). Our results showed the existence of various L. pneumophila serogroup 1 subgroups circulating in Morocco. The MAb patterns are thus a valuable adjunct to genotyping methods in defining subgroups inside a genotypic cluster of L. pneumophila serogroup 1.
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