scholarly journals New Insight regarding Legionella Non- Pneumophila Species Identification: Comparison between the Traditional mip Gene Classification Scheme and a Newly Proposed Scheme Targeting the rpoB Gene

Author(s):  
Maria Rosaria Pascale ◽  
Silvano Salaris ◽  
Marta Mazzotta ◽  
Luna Girolamini ◽  
Giulia Fregni Serpini ◽  
...  

Legionella spp. are a widely spread bacteria that cause a fatal form of pneumonia. While traditional laboratory techniques have provided valuable systems for Legionella pneumophila identification, the amplification of the mip gene has been recognized as the only useful tool for Legionella non- pneumophila species identification both in clinical and environmental samples.

2021 ◽  
Vol 9 (2) ◽  
pp. 224
Author(s):  
Ravleen Virdi ◽  
Melissa E. Lowe ◽  
Grant J. Norton ◽  
Stephanie N. Dawrs ◽  
Nabeeh A. Hasan ◽  
...  

Nontuberculous mycobacteria (NTM) are environmental organisms that can cause opportunistic pulmonary disease with species diversity showing significant regional variation. In the United States, Hawai’i shows the highest rate of NTM pulmonary disease. The need for improved understanding of NTM reservoirs led us to identify NTM from patient respiratory specimens and compare NTM diversity between outdoor and indoor locations in Hawai’i. A total of 545 water biofilm samples were collected from 357 unique locations across Kaua’i (n = 51), O’ahu (n = 202), Maui (n = 159), and Hawai’i Island (n = 133) and divided into outdoor (n = 179) or indoor (n = 366) categories. rpoB sequence analysis was used to determine NTM species and predictive modeling applied to develop NTM risk maps based on geographic characteristics between environments. M. chimaera was frequently identified from respiratory and environmental samples followed by M. chelonae and M. abscessus; yet significantly less NTM were consistently recovered from outdoor compared to indoor biofilms, as exemplified by showerhead biofilm samples. While the frequency of M. chimaera recovery was comparable between outdoor and indoor showerhead biofilms, phylogenetic analyses demonstrate similar rpoB gene sequences between all showerhead and respiratory M. chimaera isolates, supporting outdoor and indoor environments as possible sources for pulmonary M. chimaera infections.


2015 ◽  
Vol 53 (11) ◽  
pp. 3621-3623 ◽  
Author(s):  
Laurent Ferraris ◽  
Sophia Schönherr ◽  
Philippe Bouvet ◽  
Brunhilde Dauphin ◽  
Michel Popoff ◽  
...  

“Clostridium neonatale” sp. nov., previously involved in an outbreak of neonatal necrotizing enterocolitis, was recently proposed as a new species of theClostridiumgenussensu stricto. We developed a one-step multiplex colony PCR forC. neonataleidentification and investigatedC. neonataleintestinal colonization frequency in healthy preterm neonates.


2013 ◽  
Vol 82 (1) ◽  
pp. 275-285 ◽  
Author(s):  
Jens Jäger ◽  
Sebastian Marwitz ◽  
Jana Tiefenau ◽  
Janine Rasch ◽  
Olga Shevchuk ◽  
...  

ABSTRACTHistological and clinical investigations describe late stages of Legionnaires' disease but cannot characterize early events of human infection. Cellular or rodent infection models lack the complexity of tissue or have nonhuman backgrounds. Therefore, we developed and applied a novel model forLegionella pneumophilainfection comprising living human lung tissue. We stimulated lung explants withL. pneumophilastrains and outer membrane vesicles (OMVs) to analyze tissue damage, bacterial replication, and localization as well as the transcriptional response of infected tissue. Interestingly, we found that extracellular adhesion ofL. pneumophilato the entire alveolar lining precedes bacterial invasion and replication in recruited macrophages. In contrast, OMVs predominantly bound to alveolar macrophages. Specific damage to septa and epithelia increased over 48 h and was stronger in wild-type-infected and OMV-treated samples than in samples infected with the replication-deficient, type IVB secretion-deficient DotA−strain. Transcriptome analysis of lung tissue explants revealed a differential regulation of 2,499 genes after infection. The transcriptional response included the upregulation of uteroglobin and the downregulation of the macrophage receptor with collagenous structure (MARCO). Immunohistochemistry confirmed the downregulation of MARCO at sites of pathogen-induced tissue destruction. Neither host factor has ever been described in the context ofL. pneumophilainfections. This work demonstrates that the tissue explant model reproduces realistic features of Legionnaires' disease and reveals new functions for bacterial OMVs during infection. Our model allows us to characterize early steps of human infection which otherwise are not feasible for investigations.


2018 ◽  
Vol 6 (15) ◽  
pp. e00295-18
Author(s):  
Alexander Fortuna ◽  
Ricardo Ramnarine ◽  
Aimin Li ◽  
Nahuel Fittipaldi ◽  
Christine Frantz ◽  
...  

ABSTRACT Legionella pneumophila outbreak investigations require the development of reliable typing methods to better understand the genetic relationships of the isolates involved. Here, we report the draft genome sequences of four clinical Legionella pneumophila isolates obtained between 2000 and 2012 in Ontario, Canada.


mBio ◽  
2011 ◽  
Vol 2 (4) ◽  
Author(s):  
Christopher L. Case ◽  
Craig R. Roy

ABSTRACTNucleotide-binding domain, leucine-rich repeat containing proteins (NLRs) activate caspase-1 in response to a variety of bacterium-derived signals in macrophages. NLR-mediated activation of caspase-1 byLegionella pneumophilaoccurs through both an NLRC4/NAIP5-dependent pathway and a pathway requiring the adapter protein Asc. Both pathways are needed for maximal activation of caspase-1 and for the release of the cytokines interleukin-1β (IL-1β) and IL-18. Asc is not required for caspase-1-dependent pore formation and cell death induced upon infection of macrophages byL. pneumophila. Here, temporal and spatial localization of caspase-1-dependent processes was examined to better define the roles of Asc and NLRC4 during infection. Imaging studies revealed that caspase-1 localized to a single punctate structure in infected cells containing Asc but not in cells lacking this adapter. Both endogenous Asc and ectopically produced NLRC4 tagged with green fluorescent protein (GFP) were found to localize to caspase-1 puncta followingL. pneumophilainfection, suggesting that NLRC4 and Asc coordinate signaling through this complex during caspase-1 activation. Formation of caspase-1-containing puncta correlated with caspase-1 processing, suggesting a role for the Asc/NLRC4/caspase-1 complex in caspase-1 cleavage. In cells deficient for Asc, NLRC4 did not assemble into discrete puncta, and pyroptosis occurred at an accelerated rate. These data indicate that Asc mediates integration of NLR components into caspase-1 processing platforms and that recruitment of NLR components into an Asc complex can dampen pyroptotic responses. Thus, a negative feedback role of complexes containing Asc may be important for regulating caspase-1-mediated responses during microbial infection.IMPORTANCECaspase-1 is a protease activated during infection that is central to the regulation of several innate immune pathways. Studies examining the macromolecular complexes containing this protein, known as inflammasomes, have provided insight into the regulation of this protease. This work demonstrates that the intracellular bacteriumLegionella pneumophilainduces formation of complexes containing caspase-1 by multiple mechanisms and illustrates that an adapter molecule called Asc integrates signals from multiple independent upstream caspase-1 activators in order to assemble a spatially distinct complex in the macrophage. There were caspase-1-associated activities such as cytokine processing and secretion that were controlled by Asc. Importantly, this work uncovered a new role for Asc in dampening a caspase-1-dependent cell death pathway called pyroptosis. These findings suggest that Asc plays a central role in controlling a distinct subset of caspase-1-dependent activities by both assembling complexes that are important for cytokine processing and suppressing processes that mediate pyroptosis.


2012 ◽  
Vol 78 (12) ◽  
pp. 4420-4433 ◽  
Author(s):  
Brian D. Sauders ◽  
Jon Overdevest ◽  
Esther Fortes ◽  
Katy Windham ◽  
Ynte Schukken ◽  
...  

ABSTRACTA total of 442Listeriaisolates, including 234Listeria seeligeri, 80L. monocytogenes, 74L. welshimeri, 50L. innocua, and 4L. marthiiisolates, were obtained from 1,805 soil, water, and other environmental samples collected over 2 years from four urban areas and four areas representing natural environments.Listeriaspp. showed similar prevalences in samples from natural (23.4%) and urban (22.3%) environments. WhileL. seeligeriandL. welshimeriwere significantly associated with natural environments (P≤ 0.0001),L. innocuaandL. monocytogeneswere significantly associated with urban environments (P≤ 0.0001). Sequencing ofsigBfor all isolates revealed 67 allelic types with a higher level of allelic diversity among isolates from urban environments. SomeListeriaspp. andsigBallelic types showed significant associations with specific urban and natural areas. Nearest-neighbor analyses also showed that certainListeriaspp. andsigBallelic types were spatially clustered within both natural and urban environments, and there was evidence that these species and allelic types persisted over time in specific areas. Our data show that members of the genusListerianot only are common in urban and natural environments but also show species- and subtype-specific associations with different environments and areas. This indicates thatListeriaspecies and subtypes within these species may show distinct ecological preferences, which suggests (i) that molecular source-tracking approaches can be developed forListeriaand (ii) that detection of someListeriaspecies may not be a good indicator forL. monocytogenes.


2018 ◽  
Vol 87 (1) ◽  
Author(s):  
J. Rasch ◽  
C. M. Ünal ◽  
A. Klages ◽  
Ü. Karsli ◽  
N. Heinsohn ◽  
...  

ABSTRACTThe gammaproteobacteriumLegionella pneumophilais the causative agent of Legionnaires’ disease, an atypical pneumonia that manifests itself with severe lung damage.L. pneumophila, a common inhabitant of freshwater environments, replicates in free-living amoebae and persists in biofilms in natural and man-made water systems. Its environmental versatility is reflected in its ability to survive and grow within a broad temperature range as well as its capability to colonize and infect a wide range of hosts, including protozoa and humans. Peptidyl-prolyl-cis/trans-isomerases (PPIases) are multifunctional proteins that are mainly involved in protein folding and secretion in bacteria. InL. pneumophilathe surface-associated PPIase Mip was shown to facilitate the establishment of the intracellular infection cycle in its early stages. The cytoplasmic PpiB was shown to promote cold tolerance. Here, we set out to analyze the interrelationship of these two relevant PPIases in the context of environmental fitness and infection. We demonstrate that the PPIases Mip and PpiB are important for surfactant-dependent sliding motility and adaptation to suboptimal temperatures, features that contribute to the environmental fitness ofL. pneumophila. Furthermore, they contribute to infection of the natural hostAcanthamoeba castellaniias well as human macrophages and human explanted lung tissue. These effects were additive in the case of sliding motility or synergistic in the case of temperature tolerance and infection, as assessed by the behavior of the double mutant. Accordingly, we propose that Mip and PpiB are virulence modulators ofL. pneumophilawith compensatory action and pleiotropic effects.


2018 ◽  
Vol 87 (1) ◽  
Author(s):  
Yuta Nanjo ◽  
Michael W. Newstead ◽  
Tetsuji Aoyagi ◽  
Xianying Zeng ◽  
Kazuhisa Takahashi ◽  
...  

ABSTRACTLegionella pneumophilacauses life-threatening pneumonia culminating in acute lung injury. Innate and adaptive cytokines play an important role in host defense againstL. pneumophilainfection. Interleukin-36 (IL-36) cytokines are recently described members of the larger IL-1 cytokine family known to exert potent inflammatory effects. In this study, we elucidated the role for IL-36 cytokines in experimental pneumonia caused byL. pneumophila. Intratracheal (i.t.) administration ofL. pneumophilainduced the upregulation of both IL-36α and IL-36γ mRNA and protein production in the lung. Compared to the findings forL. pneumophila-infected wild-type (WT) mice, the i.t. administration ofL. pneumophilato IL-36 receptor-deficient (IL-36R−/−) mice resulted in increased mortality, a delay in lung bacterial clearance, increasedL. pneumophiladissemination to extrapulmonary organs, and impaired glucose homeostasis. Impaired lung bacterial clearance in IL-36R−/−mice was associated with a significantly reduced accumulation of inflammatory cells and the decreased production of proinflammatory cytokines and chemokines.Ex vivo, reduced expression of costimulatory molecules and impaired M1 polarization were observed in alveolar macrophages isolated from infected IL-36R−/−mice compared to macrophages from WT mice. WhileL. pneumophila-induced mortality in IL-36α- or IL-36γ-deficient mice was not different from that in WT animals, antibody-mediated neutralization of IL-36γ in IL-36α−/−mice resulted in mortality similar to that observed in IL-36R−/−mice, indicating redundant and overlapping roles for these cytokines in experimental murineL. pneumophilapneumonia.


2016 ◽  
Vol 83 (4) ◽  
Author(s):  
Danielle D. Cloutier ◽  
Sandra L. McLellan

ABSTRACT Alternative indicators have been developed that can be used to identify host sources of fecal pollution, yet little is known about how their distribution and fate compare to traditional indicators. Escherichia coli and enterococci were widely distributed at the six beaches studied and were detected in almost 95% of water samples (n = 422) and 100% of sand samples (n = 400). Berm sand contained the largest amount of E. coli (P < 0.01), whereas levels of enterococci were highest in the backshore (P < 0.01). E. coli and enterococci were the lowest in water, using a weight-to-volume comparison. The gull-associated Catellicoccus marimammalium (Gull2) marker was found in over 80% of water samples, regardless of E. coli levels, and in 25% of sand samples. Human-associated Bacteroides (HB) and Lachnospiraceae (Lachno2) were detected in only 2.4% of water samples collected under baseflow and post-rain conditions but produced a robust signal after a combined sewage overflow, despite low E. coli concentrations. Burdens of E. coli and enterococci in water and sand were disproportionately high in relation to alternative indicators when comparing environmental samples to source material. In microcosm studies, Gull2, HB, and Lachno2 quantitative PCR (qPCR) signals were reduced twice as quickly as those from E. coli and enterococci and approximately 20% faster than signals from culturable E. coli. High concentrations of alternative indicators in source material illustrated their high sensitivity for the identification of fecal sources; however, differential survival and the potential for long-term persistence of traditional fecal indicators complicate the use of alternative indicator data to account for the levels of E. coli and enterococci in environmental samples. IMPORTANCE E. coli and enterococci are general indicators of fecal pollution and may persist in beach sand, making their use problematic for many applications. This study demonstrates that gull fecal pollution is widespread at Great Lakes beaches, whereas human and ruminant contamination is evident only after major rain events. An exploration of sand as a reservoir for indicators found that E. coli was ubiquitous, while gull host markers were detected in only 25% of samples. In situ sand beach microcosms provided decay rate constants for E. coli and enterococci relative to alternative indicators, which establish comparative benchmarks that would be helpful to distinguish recent from past pollution. Overall, alternative indicators are useful for identifying sources and assessing potentially high health risk contamination events; however, beach managers should be cautious in attempting to directly link their detection to the levels of E. coli or enterococci.


2018 ◽  
Vol 56 (11) ◽  
Author(s):  
David R. Greig ◽  
Ulf Schaefer ◽  
Sophie Octavia ◽  
Ebony Hunter ◽  
Marie A. Chattaway ◽  
...  

ABSTRACT Epidemiological and microbiological data on Vibrio cholerae strains isolated between April 2004 and March 2018 (n = 836) and held at the Public Health England culture archive were reviewed. The traditional biochemical species identification and serological typing results were compared with the genome-derived species identification and serotype for a subset of isolates (n = 152). Of the 836 isolates, 750 (89.7%) were from a fecal specimen, 206 (24.6%) belonged to serogroup O1, and 7 (0.8%) were serogroup O139; 792 (94.7%) isolates were from patients reporting recent travel abroad, most commonly to India (n = 209) and Pakistan (n = 104). Of the 152 V. cholerae isolates identified by use of kmer, 149 (98.1%) were concordant with those identified using the traditional biochemical approach. Traditional serotyping results were 100% concordant with those of the whole-genome sequencing (WGS) analysis for the identification of serogroups O1 and O139 and classical and El Tor biotypes. ctxA was detected in all isolates of V. cholerae O1 El Tor and O139 belonging to sequence type 69 (ST69) and in V. cholerae O1 classical variants belonging to ST73. A phylogeny of isolates belonging to ST69 from U.K. travelers clustered geographically, with isolates from India and Pakistan located on separate branches. Moving forward, WGS data from U.K. travelers will contribute to global surveillance programs and the monitoring of emerging threats to public health and the global dissemination of pathogenic lineages. At the national level, these WGS data will inform the timely reinforcement of direct public health messaging to travelers and mitigate the impact of imported infections and the associated risks to public health.


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