Hyper-inflammatory profile and immunoparalysis in patients with severe Legionnaires' disease.

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.

TNF-α response in macrophages depends on clinical Legionella pneumophila isolates genotypes.

Legionnaires' Disease (LD) is a severe pneumonia mainly caused in Europe by Legionella pneumophila serogroup 1 (Lp1). Sequence-based typing methods reveal that some sequence types (ST) are overrepresented in clinical samples such as ST1 and ST47, suggesting that some strains are more fit for infection than others. In the present study, a collection of 108 Lp1 clinical isolates were used to evaluate the strain-dependent immune responses from human macrophages. Clinical Lp1 isolates induced differential TNFα secretion from macrophages. ST1 isolates induced a significantly higher TNF-α secretion than non-ST1, whereas ST47 isolates induced a significantly lower TNF-α secretion than non-ST47 isolates. ST1 isolates induced a significantly higher cell death than ST47 isolates evaluated by lactate dehydrogenase activity (cytotoxicity) and caspase-3 activity (apoptosis). Treatment of macrophages with anti-TNF-α antibodies significantly reduced the cell death in macrophages infected with ST1 or ST47 strains. The TNF-α secretion was neither explained by a differential bacterial replication nor by the number or type (bystander or infected) of TNF-α producing cells following infection but by a differential response from macrophages. The Paris ST1 reference strain elicited a significantly higher TNF-α gene transcription and a higher induction of NF-κB signaling pathway than the Lorraine ST47 reference strain.Clinical Lp1 isolates induce a diverse immune response and cell death, which could be related to the genotype. The two predominant sequence-types ST1 and ST47 trigger opposite inflammatory response that could be related to the host susceptibility.

Optimizing Strategies for Chlamydia trachomatis and Neisseria gonorrhoeae Screening in Men Who Have Sex With Men: A Modeling Study.

BACKGROUND: International guidelines recommend the systematic screening for Neisseria gonorrhoeae (NG) and Chlamydia trachomatis (CT) infections in all men who have sex with men (MSM) who have engaged in unprotected sex. However, the optimal screening strategy remains unclear. We developed a modeling approach to optimize NG/CT screening strategy in MSM. METHODS: A compartmental model of NG/CT screening and infection was implemented. NG/CT anal, pharyngeal, and urine (APU) samples from MSM attending the sexually transmitted infections clinic were used to estimate the screening rate, prevalence, and incidence in a base case scenario. Different screening strategies (scenarios; S) were then evaluated: APU samples every 12 months (S1); APU samples every 3 months (S2); APU samples every 6 months (S3); anal and pharyngeal (AP) samples every 6 months (S4); and AP samples every 3 months (S5). RESULTS: We analyzed 2973 triplet APU samples from 1255 patients. We observed 485 NG and 379 CT diagnoses. NG/CT prevalence and incidence estimates were 12.0/11.1% and 40/29 per 100 person-years, respectively, in the base case scenario. As compared to S2, the reference strategy, the proportions of missed NG/CT diagnoses were 42.0/41.2% with S1, 21.8/22.5% with S3, 25.6/28.3% with S4, and 6.3/10.5% with S5, respectively. As compared to S2, S1 reduced the cost of the analysis by 74%, S3 by 50%, S4 by 66%, and S5 by 33%. The numbers needed to screen for catching up the missed NG/CT diagnoses were 49/67 with S1, 62/82 with S3, 71/87 with S4, and 143/118 with S5. CONCLUSIONS: S5 appears to be the best strategy, missing only 6.3/10.5% of NG/CT diagnoses, for a cost reduction of 33%.

Decreased intra-lymphocyte cytokines measurement in septic shock patients: A proof of concept study in whole blood.

Functional testing protocols are thought to be the gold standard for the exploration of the immune system. However, in terms of routine analysis, they present numerous drawbacks and consequently their use is mainly limited to research applications. In the clinical context of septic shock, characterized by marked lymphocyte alterations, a new approach for lymphocyte intracellular cytokine measurement in whole blood upon was evaluated in a proof-of-concept study. Following lymphocyte activation, simultaneous intracellular labeling of Interferon-γ (IFN-γ), Tumor Necrosis Factor-α (TNF-α), and Interleukin-2 (IL-2) was performed in CD4+ and CD8+ T cells (identified by surface marking). The analysis was carried out by flow cytometry (6 colors). Results obtained in septic patients (n=22) were compared to those of healthy volunteers (n=8). Independently of lymphopenia, there were significant differences between groups. In particular there was significant decrease in the production of IL-2 and TNF-α in septic patients, while the production of IFN-γ was not significantly altered. Polyfunctional results showed that patients presented with increased percentages of triple negative lymphocytes. In contrast, volunteers had higher proportions of triple positive cells. The approach could be performed in a robust and consistent way, taking 4.5h to complete. Moreover, clear differences could be observed between clinical groups with this modified method. These characteristics illustrate the potential of this novel whole blood protocol for clinical applications. However, further research is required to determine the applicability compared to alternative test and to evaluate clinical performances in larger cohorts of patients.