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OBJECTIVES: Implementation of an antibiotic resistance detection tool in Legionella daily surveillance at the French National Reference Centre for Legionella. METHODS: Systematic WGS of Legionella pneumophila isolates and bioinformatics detection of specific mutations linked to antibiotic resistance. Phenotypic validation of antibiotic resistance detected by WGS was performed by the broth microdilution method. RESULTS: More than 3000 L. pneumophila strains were screened for antibiotic resistance. A macrolide resistance-associated A2052G mutation in the 23S rRNA gene was identified in the genome of eight isolates from a hotel water network. High-level macrolide resistance (i.e. MICs of 1024-2048â mg/L for azithromycin and erythromycin) with no cross-resistance to other antimicrobials was phenotypically confirmed by antimicrobial susceptibility testing for the eight isolates. CONCLUSIONS: Systematic WGS of L. pneumophila is a powerful tool for first-line high-throughput screening of antibiotic resistance before phenotypic validation.
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Legionella pneumophila , Antibacterianos/farmacología , Farmacorresistencia Bacteriana , Macrólidos/farmacología , Pruebas de Sensibilidad Microbiana , AguaRESUMEN
We describe a March 2020 co-occurrence of Legionnaires' disease (LD) and coronavirus disease in France. Severe acute respiratory syndrome coronavirus 2 co-infections were identified in 7 of 49 patients from LD case notifications. Most were elderly men with underlying conditions who had contracted severe pneumonia, illustrating the relevance of co-infection screening.
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COVID-19 , Coinfección , Legionella , Anciano , Coinfección/epidemiología , Francia/epidemiología , Humanos , Legionella/genética , Masculino , SARS-CoV-2RESUMEN
Legionella pneumophila, a Gram-negative bacillus, is the causative agent of Legionnaire's disease, a form of severe community-acquired pneumonia. Infection can have high morbidity, with a high proportion of patients requiring ICU admission, and up to 10% mortality, which is exacerbated by the lack of efficacy of typical empirical antibiotic therapy against Legionella spp. The fastidious nature of the entire Legionellaceae family historically required inclusion of activated charcoal in the solid medium to remove growth inhibitors, which inherently interferes with accurate antimicrobial susceptibility determination, an acknowledged methodological shortfall, now rectified by a new solid medium that gives results comparable to those of microbroth dilution. Here, as an international Legionella community (with authors representing various international reference laboratories, countries and clinical stakeholders for diagnosis and treatment of legionellosis), we set out recommendations for the standardization of antimicrobial susceptibility testing methods, guidelines and reference strains to facilitate an improved era of antibiotic resistance determination.
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Legionella pneumophila , Legionella , Enfermedad de los Legionarios , Antibacterianos/farmacología , Antibacterianos/uso terapéutico , Humanos , Enfermedad de los Legionarios/diagnóstico , Enfermedad de los Legionarios/tratamiento farmacológico , Estándares de ReferenciaRESUMEN
BACKGROUND: Legionella spp. are ubiquitous freshwater bacteria responsible for rare but potentially severe cases of Legionnaires' disease (LD). Legionella sainthelensi is a non-pneumophila Legionella species that was first isolated in 1980 from water near Mt. St-Helens (USA). Although rare cases of LD caused by L. sainthelensi have been reported, very little data is available on this pathogen. CASE PRESENTATION: We describe the first documented case of severe bilateral pleuropneumonia caused by L. sainthelensi. The patient was a 35-year-old woman with Sharp's syndrome treated with long-term hydroxychloroquine and corticosteroids who was hospitalized for an infectious illness in a university hospital in Reunion Island (France). The patient's clinical presentation was complicated at first (bilateral pneumonia, multiloculated pleural effusion, then bronchopleural fistula) but her clinical condition eventually improved with the reintroduction of macrolides (spiramycin) in intensive care unit. Etiological diagnosis was confirmed by PCR syndromic assay and culture on bronchoalveolar lavage. CONCLUSIONS: To date, only 14 documented cases of L. sainthelensi infection have been described worldwide. This pathogen is difficult to identify because it is not or poorly detected by urinary antigen and molecular methods (like PCR syndromic assays that primarily target L. pneumophila and that have only recently been deployed in microbiology laboratories). Pneumonia caused by L. sainthelensi is likely underdiagnosed as a result. Clinicians should consider the possibility of non-pneumophila Legionella infection in patients with a compatible clinical presentation when microbiological diagnostic tools targeted L. pneumophila tested negative.
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Legionella pneumophila , Legionella , Enfermedad de los Legionarios , Pleuroneumonía , Adulto , Femenino , Humanos , Legionella/genética , Legionella pneumophila/genética , Enfermedad de los Legionarios/diagnóstico , Enfermedad de los Legionarios/tratamiento farmacológico , Pleuroneumonía/diagnóstico , Pleuroneumonía/tratamiento farmacológicoRESUMEN
BACKGROUND: Rarely, Legionnaires' disease (LD) can progress into a slowly or nonresolving form. METHODS: A nationwide retrospective study was conducted by the French National Reference Center for Legionella (2013-2017) including cases of slowly or nonresolving LD defined as persistent clinical symptoms, computed tomography (CT) scan abnormalities, and Legionella detection in lower respiratory tract specimens by culture and/or real-time (RT) polymerase chain reaction (PCR) >30 days after symptom onset. RESULTS: Twelve cases of community-acquired slowly or nonresolving LD were identified among 1686 cases of culture-positive LD. Median (interquartile range [IQR]) age was 63 (29-82) years. Ten (83.3%) patients had ≥1 immunosuppressive factor. Clinically, 9 patients transiently recovered before further deterioration (median [IQR] symptom-free interval, 30 [18-55] days), 3 patients had uniformly persistent symptoms (median [IQR] time, 48 [41.5-54] days). Two patients had >2 recurrences. CT scan imagery found lung abscess in 5 (41.6%) cases. Slowly or nonresolving LD was diagnosed on positive Legionella cultures (n = 10, 83.3%) at 49.5 (IQR, 33.7-79) days. Two cases were documented through positive Legionella RT PCR at 52 and 53 days (cycle threshold detection of 21.5 and 33.7, respectively). No genomic microevolution and no Legionella resistance to antibiotics were detected. The median (IQR) duration of treatment was 46.5 (21-92.5) days. Two empyema cases required thoracic surgery. At a median (IQR) follow-up of 26 (14-41.5) months, LD-attributable mortality was 16.6% (n = 2). CONCLUSIONS: Slowly or nonresolving LD may occur in immunocompromised patients, possibly leading to lung abscess and empyema.
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Legionella pneumophila , Legionella , Enfermedad de los Legionarios , Antibacterianos/uso terapéutico , Humanos , Legionella/genética , Legionella pneumophila/genética , Enfermedad de los Legionarios/diagnóstico , Enfermedad de los Legionarios/tratamiento farmacológico , Reacción en Cadena en Tiempo Real de la Polimerasa , Estudios RetrospectivosRESUMEN
We describe 2 cases of healthcare-associated Legionnaires' disease in patients in France hospitalized 5 months apart in the same room. Whole-genome sequencing analyses showed that clinical isolates from the patients and isolates from the room's toilet clustered together. Toilet contamination by Legionella pneumophila could lead to a risk for exposure through flushing.
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Aparatos Sanitarios , Infección Hospitalaria , Legionella pneumophila , Enfermedad de los Legionarios , Francia , Humanos , Legionella pneumophila/genética , Enfermedad de los Legionarios/diagnóstico , Enfermedad de los Legionarios/epidemiologíaRESUMEN
Detection of Legionella pneumophila serogroup 1 urinary antigens is the most widely used technique for the diagnosis of Legionnaires' disease (LD). The aim of this study was to evaluate the performance of the Legionella K-set® immunochromatographic test, in comparison with the BinaxNOW® Legionella urinary antigen card (UAC) on concentrated urine samples (US). A total of 250 concentrated US including 200 prospective US sent to the laboratory for urinary antigens' testing and 50 frozen US from patients with confirmed LD were tested. Positive US were retested after boiling (5 min, 100 °C). Each US leading to discordant results between the two tests was further tested using Binax™ Legionella EIA. Then, 10 additional positive non-concentrated US were tested using both tests. On concentrated US, Legionella K-set® test showed concordant results with that of BinaxNOW® Legionella. All negative US with BinaxNOW® were negative with Legionella K-set® test. For the 50 frozen US, all were positive with BinaxNOW® and 49 were positive with Legionella K-set®, all confirmed after boiling except 3 US which led to uninterpretable results with Legionella K-set®, due to a migration defect. Three of the 10 additional positive non-concentrated US were found negative with Legionella K-set® and only 1 US remained negative after concentration. All these positive non-concentrated US were positive with BinaxNOW® Legionella. The performance of the Legionella K-set® test is comparable to that of BinaxNOW® Legionella UAC, if performed on concentrated US.
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Antígenos Bacterianos/orina , Legionella pneumophila/inmunología , Enfermedad de los Legionarios/diagnóstico , Humanos , Pruebas Inmunológicas , Enfermedad de los Legionarios/microbiología , Enfermedad de los Legionarios/orina , Sensibilidad y Especificidad , UrinálisisRESUMEN
BACKGROUND: While Legionella is a common cause of pneumonia, extrapulmonary infections like arthritis are scarce. Here, we describe a case of monoarthritis due to Legionella bozemanii, with no history of pneumonia. We provide a literature review of the 9 previously published Legionella arthritis and highlight a dichotomous epidemiology suggesting different physiopathological pathways leading to joint infection. CASE PRESENTATION: A 56-year old woman under immunosuppressive treatment by oral and intra-articular corticosteroids, methotrexate, and tocilizumab for an anti-synthetase syndrome was hospitalized for worsening pain and swelling of the left wrist for 3 days. Clinical examination showed left wrist synovitis and no fever. The arthritis occurred a few days after an accidental fall on wet asphalt responsible for a cutaneous wound followed by a corticosteroid intra-articular injection. Due to both the negativity of conventional culture of articular fluid and suspicion of infection, 16S rRNA and specific PCRs were performed leading to the identification of L. bozemanii. Legionella-specific culture of the articular fluid was performed retrospectively and isolated L. bozemanii. The empiric antibiotic therapy was switched for oral levofloxacin and rifampin and the patient recovered after a 12-week treatment. CONCLUSION: We report a case of L. bozemanii monoarthritis in an immunosuppressed woman, following a fall on wet asphalt and intra-articular corticosteroid injection. The review of the literature found that the clinical presentation reveals the mode of infection and the bacterial species. Monoarthritis more likely occurred after inoculation in patients under immunosuppressive therapy and were associated with non-Legionella pneumophila serogroup 1 (Lp1) strains that predominate in the environment. Polyarthritis were more likely secondary legionellosis localizations after blood spread of Lp1, the most frequently found in pneumonia. In both settings, 16S rRNA and Legionella-specific PCR were key factors for the diagnosis.
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Artritis Infecciosa/inmunología , Artritis Infecciosa/microbiología , Legionellaceae/aislamiento & purificación , Legionelosis/microbiología , Accidentes por Caídas , Administración Oral , Corticoesteroides/administración & dosificación , Corticoesteroides/efectos adversos , Corticoesteroides/uso terapéutico , Artritis Infecciosa/tratamiento farmacológico , Femenino , Humanos , Hidrocarburos , Huésped Inmunocomprometido , Inyecciones Intraarticulares , Legionellaceae/genética , Levofloxacino/uso terapéutico , Persona de Mediana Edad , Reacción en Cadena de la Polimerasa , ARN Ribosómico 16S/genética , Rifampin/uso terapéutico , Resultado del TratamientoRESUMEN
trans-Translation is a ribosome-rescue system that is ubiquitous in bacteria. Small molecules defining a new family of oxadiazole compounds that inhibit trans-translation have been found to have broad-spectrum antibiotic activity. We sought to determine the activity of KKL-35, a potent member of the oxadiazole family, against the human pathogen Legionella pneumophila and other related species that can also cause Legionnaires' disease (LD). Consistent with the essential nature of trans-translation in L. pneumophila, KKL-35 inhibited the growth of all tested strains at submicromolar concentrations. KKL-35 was also active against other LD-causing Legionella species. KKL-35 remained equally active against L. pneumophila mutants that have evolved resistance to macrolides. KKL-35 inhibited the multiplication of L. pneumophila in human macrophages at several stages of infection. No resistant mutants could be obtained, even during extended and chronic exposure. Surprisingly, KKL-35 was not synergistic with other ribosome-targeting antibiotics and did not induce the filamentation phenotype observed in cells defective for trans-translation. Importantly, KKL-35 remained active against L. pneumophila mutants expressing an alternate ribosome-rescue system and lacking transfer-messenger RNA, the essential component of trans-translation. These results indicate that the antibiotic activity of KKL-35 is not related to the specific inhibition of trans-translation and its mode of action remains to be identified. In conclusion, KKL-35 is an effective antibacterial agent against the intracellular pathogen L. pneumophila with no detectable resistance development. However, further studies are needed to better understand its mechanism of action and to assess further the potential of oxadiazoles in treatment.
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Antibacterianos/farmacología , Benzamidas/farmacología , Farmacorresistencia Bacteriana/efectos de los fármacos , Legionella pneumophila/efectos de los fármacos , Legionella/efectos de los fármacos , Oxadiazoles/farmacología , Línea Celular , Humanos , Legionella/crecimiento & desarrollo , Legionella pneumophila/crecimiento & desarrollo , Enfermedad de los Legionarios , Macrólidos/farmacología , Macrófagos/efectos de los fármacos , Macrófagos/microbiología , Pruebas de Sensibilidad Microbiana , Biosíntesis de ProteínasRESUMEN
Identification and antimicrobial susceptibility testing (AST) are critical steps in the management of bloodstream infections. Our objective was to evaluate the performance of the Accelerate Pheno™ System, CE v1.2 software, for identification and AST of Gram-negative pathogens from positive blood culture bottles. A total of 104 bottles positive for Gram-negative bacteria collected from inpatients throughout our institution were randomly selected after Gram staining. The time-to-identification and AST results, and the raw AST results obtained by the Accelerate Pheno™ system and routine techniques (MALDI-TOF MS and VITEK®2, EUCAST guidelines) were compared. Any discrepant AST result was tested by microdilution. The Pheno™ significantly improved turn-around times for identification (5.3 versus 23.7 h; p < 0.0001) and AST (10.7 versus 35.1 h; p < 0.0001). Complete agreement between the Accelerate Pheno™ system and the MALDI-TOF MS for identification was observed for 96.2% of samples; it was 99% (98/99) for monomicrobial samples versus 40% (3/5) for polymicrobial ones. The overall categorical agreement for AST was 93.7%; it was notably decreased for beta-lactams (cefepime 84.4%, piperacillin-tazobactam 86.5%, ceftazidime 87.6%) or Pseudomonas aeruginosa (71.9%; with cefepime 33.3%, piperacillin-tazobactam 77.8%, ceftazidime 0%). Analysis of discrepant results found impaired performance of the Accelerate Pheno™ system for beta-lactams (except cefepime) in Enterobacteriales (six very major errors) and poor performance in P. aeruginosa. The Accelerate Pheno™ system significantly improved the turn-around times for bloodstream infection diagnosis. Nonetheless, improvements in the analysis of polymicrobial samples and in AST algorithms, notably beta-lactam testing in both P. aeruginosa and Enterobacteriales, are required for implementation in routine workflow.
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Antibacterianos/farmacología , Bacteriemia/diagnóstico , Bacteriemia/microbiología , Técnicas de Tipificación Bacteriana , Bacterias Gramnegativas/efectos de los fármacos , Infecciones por Bacterias Gramnegativas/diagnóstico , Infecciones por Bacterias Gramnegativas/microbiología , Pruebas de Sensibilidad Microbiana , Adolescente , Adulto , Anciano , Anciano de 80 o más Años , Antibacterianos/uso terapéutico , Bacteriemia/tratamiento farmacológico , Niño , Preescolar , Bacterias Gramnegativas/clasificación , Bacterias Gramnegativas/aislamiento & purificación , Infecciones por Bacterias Gramnegativas/tratamiento farmacológico , Humanos , Lactante , Recién Nacido , Pruebas de Sensibilidad Microbiana/métodos , Persona de Mediana Edad , Espectrometría de Masa por Láser de Matriz Asistida de Ionización Desorción , Factores de Tiempo , Adulto JovenRESUMEN
Monitoring the emergence of antibiotic resistance is a recent issue in the treatment of Legionnaires' disease. Macrolides are recommended as first-line therapy, but resistance mechanisms have not been studied in Legionella species. Our aim was to determine the molecular basis of macrolide resistance in L. pneumophila Twelve independent lineages from a common susceptible L. pneumophila ancestral strain were propagated under conditions of erythromycin or azithromycin pressure to produce high-level macrolide resistance. Whole-genome sequencing was performed on 12 selected clones, and we investigated mutations common to all lineages. We reconstructed the dynamics of mutation for each lineage and demonstrated their involvement in decreased susceptibility to macrolides. The resistant mutants were produced in a limited number of passages to obtain a 4,096-fold increase in erythromycin MICs. Mutations affected highly conserved 5-amino-acid regions of L4 and L22 ribosomal proteins and of domain V of 23S rRNA (G2057, A2058, A2059, and C2611 nucleotides). The early mechanisms mainly affected L4 and L22 proteins and induced a 32-fold increase in the MICs of the selector drug. Additional mutations related to 23S rRNA mostly occurred later and were responsible for a major increase of macrolide MICs, depending on the mutated nucleotide, the substitution, and the number of mutated genes among the three rrl copies. The major mechanisms of the decreased susceptibility to macrolides in L. pneumophila and their dynamics were determined. The results showed that macrolide resistance could be easily selected in L. pneumophila and warrant further investigations in both clinical and environmental settings.
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Proteínas Bacterianas/genética , Farmacorresistencia Bacteriana/genética , Legionella pneumophila/genética , Mutación , ARN Ribosómico 23S/genética , Proteínas Ribosómicas/genética , Antibacterianos/farmacología , Azitromicina/farmacología , Proteínas Bacterianas/metabolismo , Células Clonales , Eritromicina/farmacología , Secuenciación de Nucleótidos de Alto Rendimiento , Legionella pneumophila/efectos de los fármacos , Legionella pneumophila/crecimiento & desarrollo , Pruebas de Sensibilidad Microbiana , Isoformas de Proteínas/genética , Isoformas de Proteínas/metabolismo , ARN Ribosómico 23S/metabolismo , Proteínas Ribosómicas/metabolismoRESUMEN
Objectives: A previous study on 12 in vitro -selected azithromycin-resistant Legionella pneumophila lineages showed that ribosomal mutations were major macrolide resistance determinants. In addition to these mechanisms that have been well described in many species, mutations upstream of lpeAB operon, homologous to acrAB in Escherichia coli , were identified in two lineages. In this study, we investigated the role of LpeAB and of these mutations in macrolide resistance of L. pneumophila . Methods: The role of LpeAB was studied by testing the antibiotic susceptibility of WT, deleted and complemented L. pneumophila Paris strains. Translational fusion experiments using GFP as a reporter were conducted to investigate the consequences of the mutations observed in the upstream sequence of lpeAB operon. Results: We demonstrated the involvement of LpeAB in an efflux pump responsible for a macrolide-specific reduced susceptibility of L. pneumophila Paris strain. Mutations in the upstream sequence of lpeAB operon were associated with an increased protein expression. Increased expression was also observed under sub-inhibitory macrolide concentrations in strains with both mutated and WT promoting regions. Conclusions: LpeAB are components of an efflux pump, which is a macrolide resistance determinant in L. pneumophila Paris strain. Mutations observed in the upstream sequence of lpeAB operon in resistant lineages led to an overexpression of this efflux pump. Sub-inhibitory concentrations of macrolides themselves participated in upregulating this efflux and could constitute a first step in the acquisition of a high macrolide resistance level.
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Antibacterianos/farmacología , Farmacorresistencia Bacteriana Múltiple/genética , Legionella pneumophila/efectos de los fármacos , Macrólidos/farmacología , Proteínas de Transporte de Membrana/genética , Azitromicina , Proteínas Bacterianas/genética , Proteínas Bacterianas/metabolismo , Eritromicina/farmacología , Genes Bacterianos , Legionella pneumophila/genética , Legionella pneumophila/metabolismo , Proteínas de Transporte de Membrana/metabolismo , Pruebas de Sensibilidad Microbiana , Mutación , Operón , ARN Ribosómico 23SRESUMEN
Rapid antigen detection tests (RADTs) for group A streptococci (GAS) are widely used for diagnosing acute pharyngitis, which has led to a considerable reduction in antibiotic prescriptions over the past decade. Beyond this intended use, their reassessment on invasive samples may be relevant in the management of life-threatening GAS infections. To this end, we evaluated the performances of three RADTs, culture, GAS PCR, and 16S rRNA gene PCR assays, and compared them with a composite gold standard (GAS-PCR assay and/or culture) for the diagnosis of severe GAS infection. A total of 192 specimens from deep-tissue (mostly normally sterile) sites enriched for 75 GAS-positive samples were enrolled in the study. The three evaluated RADTs showed sensitivities ranging from 88.0% to 94.7% versus 98.7% for GAS PCR, 84% for 16S rRNA gene PCR, and 77.3% for culture. The sensitivities of the ImmunoCardSTAT! Strep A test (Meridian Bioscience) and the NADAL Strep A strip (Nal Von Minden) were similar to that of GAS PCR (P= 0.25 and 0.03, respectively) and higher than that of culture (P= 0.001 and 0.006, respectively), whereas the SD Bioline Strep A test strip (Standard Diagnostics) showed a performance similar to that of culture (P= 0.02). The three RADTs detected 10 distinctemmtypes, including a predominance ofemm1 (33.3%),emm89 (10.6%), andemm12 (7.6%). No false-positive results were observed, leading to a specificity of 100% for all the evaluated RADTs. The GAS RADTs turned out to be sensitive, specific, and easy-to-use tools that may aid in the management of invasive GAS infections in 24/7 point-of-care laboratories by enabling early diagnosis and focused therapy.
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Antígenos Bacterianos/análisis , Cromatografía de Afinidad/métodos , Infecciones Estreptocócicas/diagnóstico , Streptococcus pyogenes/aislamiento & purificación , Adolescente , Adulto , Anciano , Anciano de 80 o más Años , Técnicas Bacteriológicas/métodos , Niño , Preescolar , Femenino , Humanos , Lactante , Masculino , Persona de Mediana Edad , Reacción en Cadena de la Polimerasa/métodos , Sensibilidad y Especificidad , Factores de Tiempo , Adulto JovenRESUMEN
No consensus exists about the techniques to use for microbiological diagnosis of bone and joint infections (BJIs). The objective herein was to define an algorithm to optimize BJI diagnosis in adults using various bacteriological methods on synovial fluid samples. This prospective multi-center study included 423 synovial fluids collected from adult patients with suspected BJIs. Culture (using five solid media, an enrichment broth, and blood culture bottles), universal 16S rRNA PCR followed by Sanger sequencing, and seven specific bacterial PCRs were systematically performed. Combinations of methods were compared to arrive at the optimized algorithm. Among 423 synovial fluids, 242 infections were diagnosed (57.2â¯%): 213 mono- and 29 poly-microbial for a total of 284 bacteria (staphylococci at 54.6â¯%, streptococci-enterococci at 16.5â¯%, Gram-negative bacilli at 15.5â¯%, anaerobic species at 8.8â¯%). Comparing culture techniques, blood culture bottles had the highest sensitivity (67.6â¯% for pediatric and 63.9â¯% for anaerobic bottles) but are not sufficient alone and require being combined with solid media. The 16S rDNA PCR detected only 52.3â¯% of the bacteria, whereas specific PCRs had a higher sensitivity (Staphylococcus spp. at 66.2â¯%, S. aureus at 85.2â¯%, Streptococcus spp. at 91.2â¯%). Based on these results, an algorithm was proposed associating three solid media; inoculation into blood culture bottles; and 16S, Staphylococcus spp., and Streptococcus spp. PCRs, which would have detected 90.5â¯% of bacteria in the present cohort versus 79.2â¯% using all culture techniques on synovial fluid. This prospective study shows that a combination of culture and molecular methods on synovial fluids allows the optimization of bacterial detection.
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Introduction: Severe Legionnaires' disease (LD) can lead to multi-organ failure or death in 10%-30% of patients. Although hyper-inflammation and immunoparalysis are well described in sepsis and are associated with high disease severity, little is known about the immune response in LD. This study aimed to evaluate the immune status of patients with LD and its association with disease severity. Methods: A total of 92 hospitalized LD patients were included; 19 plasmatic cytokines and pulmonary Legionella DNA load were measured in 84 patients on the day of inclusion (day 0, D0). Immune functional assays (IFAs) were performed from whole blood samples collected at D2 and stimulated with concanavalin A [conA, n = 19 patients and n = 21 healthy volunteers (HV)] or lipopolysaccharide (LPS, n = 14 patients and n = 9 HV). A total of 19 cytokines (conA stimulation) and TNF-α (LPS stimulation) were quantified from the supernatants. The Sequential Organ Failure Assessment (SOFA) severity score was recorded at D0 and the mechanical ventilation (MV) status was recorded at D0 and D8. Results: Among the 84 patients, a higher secretion of plasmatic MCP-1, MIP1-ß, IL-6, IL-8, IFN-γ, TNF-α, and IL-17 was observed in the patients with D0 and D8 MV. Multiparametric analysis showed that these seven cytokines were positively associated with the SOFA score. Upon conA stimulation, LD patients had a lower secretion capacity for 16 of the 19 quantified cytokines and a higher release of IL-18 and MCP-1 compared to HV. IL-18 secretion was higher in D0 and D8 MV patients. TNF-α secretion, measured after ex vivo LPS stimulation, was significantly reduced in LD patients and was associated with D8 MV status. Discussion: The present findings describe a hyper-inflammatory phase at the initial phase of Legionella pneumonia that is more pronounced in patients with severe LD. These patients also present an immunoparalysis for a large number of cytokines, except IL-18 whose secretion is increased. An assessment of the immune response may be relevant to identify patients eligible for future innovative host-directed therapies.
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Interleucina-18 , Enfermedad de los Legionarios , Humanos , Factor de Necrosis Tumoral alfa , Lipopolisacáridos , Enfermedad de los Legionarios/complicaciones , CitocinasRESUMEN
A macrolide antimicrobial drug was administered to a newborn with cough. On day 23 of hospitalization, macrolide-resistant Bordetella pertussis was isolated from nasopharyngeal aspirates. DNA sequencing and PCR-restriction fragment length polymorphism showed a 2047 A-to-G mutation in the 3 copies of the 23S rRNA gene. Monitoring for macrolide resistance is essential in infants <6 months of age.
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Bordetella pertussis/efectos de los fármacos , Farmacorresistencia Bacteriana , Tos Ferina/diagnóstico , Azitromicina/farmacología , Azitromicina/uso terapéutico , Bordetella pertussis/genética , Claritromicina/farmacología , Claritromicina/uso terapéutico , Análisis Mutacional de ADN , Recambio Total de Sangre , Femenino , Humanos , Recién Nacido , ARN Ribosómico 23S/genética , Resultado del Tratamiento , Tos Ferina/terapiaRESUMEN
Endemic strains of Legionella pneumophila sequence type 1 (ST1), in particular the ST1/Paris pulsotype, are dispersed worldwide and represent about 10% of culture-proven clinical cases of Legionnaires' disease in France. The high rate of isolation of this strain from both clinical and environmental samples makes identification of the source of infection difficult during epidemiological investigations. The full-length genome sequence of this strain was recently determined, and it revealed the presence of a CRISPR/cas complex. The aim of this study was to develop and evaluate a spoligotyping tool based on the diversity of this CRISPR locus that would allow the accurate subtyping of the L. pneumophila serogroup 1 ST1/Paris pulsotype. The CRISPR loci of 28 L. pneumophila ST1/Paris pulsotype isolates were sequenced, and 42 different spacers regions were characterized. A membrane-based spoligotyping method was developed and used to determine the subtypes of 406 L. pneumophila isolates, including 233 with the ST1/Paris pulsotype profile that were collected in France from 2000 to 2011. A total of 46 different spoligotypes were detected, and 41 of these were specifically identified in the ST1/Paris pulsotype isolates. In 27 of 33 epidemiological investigations, the environmental source of contamination was confirmed by comparing spoligotypes of clinical isolates with those of environmental isolates. With an index of discrimination of 79.72% (95% confidence interval, 75.82 to 83.63), spoligotyping of the L. pneumophila ST1/Paris pulsotype has the potential to be a useful complementary genotyping tool for discriminating isolates with undistinguishable pulsed-field gel electrophoresis (PFGE) and ST genotypes, which could help to identify environmental sources of infection.
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Legionella pneumophila/clasificación , Legionella pneumophila/genética , Enfermedad de los Legionarios/epidemiología , Enfermedad de los Legionarios/microbiología , Tipificación Molecular/métodos , Análisis por Conglomerados , Dermatoglifia del ADN/métodos , Electroforesis en Gel de Campo Pulsado/métodos , Francia/epidemiología , Genotipo , Humanos , Epidemiología Molecular/métodosRESUMEN
We report a nosocomial case of Legionella pneumophila pneumonia caused by a serogroup 10 strain diagnosed with the Biofire® Pneumonia plus panel. Molecular investigations of the environment of the patient allowed us to identify the source of contamination.
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Legionnaires' disease is a severe pneumonia mainly caused by Legionella pneumophila that is treated by antibiotics. The purpose of this study was to describe the susceptibility of clinical strains of L. pneumophila to eight antibiotics used for treatment of legionellosis. The minimum inhibitory concentrations (MICs) of 109 well-characterised clinical strains of L. pneumophila serogroup 1 were determined by the broth microdilution method without charcoal and were compared with antibiotic-resistant strains selected in vitro. All strains were inhibited by low concentrations of fluoroquinolones, macrolides and rifampicin. The epidemiological cut-off values (ECOFFs) were 0.064 mg/L for ciprofloxacin, 0.064 mg/L for moxifloxacin, 0.032 mg/L for levofloxacin, 1 mg/L for erythromycin, 2 mg/L for azithromycin, 0.064 mg/L for clarithromycin, 2 mg/L for doxycycline and 0.001 mg/L for rifampicin. However, MIC distributions revealed a subpopulation of strains displaying reduced susceptibility to some macrolides (especially azithromycin), which correlated with the presence of the lpeAB genes encoding a macrolide efflux pump found specifically in sequence type (ST) ST1, ST701 and closely related STs. Thus, all isolates could be considered susceptible to the tested antibiotics, although macrolides were less active against some strains harbouring a specific efflux system.
Asunto(s)
Antibacterianos/farmacología , Farmacorresistencia Bacteriana , Genes Bacterianos , Legionella pneumophila/efectos de los fármacos , Legionella pneumophila/genética , Macrólidos/farmacología , Fluoroquinolonas/farmacología , Humanos , Legionella pneumophila/aislamiento & purificación , Enfermedad de los Legionarios/microbiología , Pruebas de Sensibilidad Microbiana , Rifampin/farmacologíaRESUMEN
Three urinary antigen tests were compared using purified Legionella pneumophila (Lp) LPS. For Lp serogroup1, Sofia®FIA and Binax®EIA limits of detection (LOD) were similar; that of BinaxNOW® lower. For all tests the LOD was higher with LPS from non-Pontiac compared to Pontiac-strains. The LOD was variable for other Lp serogroups.