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Article

Inflammatory Patterns Associated with Legionella in HIV and Pneumonia Coinfections

by
Breanne M. Head
1,
Adriana Trajtman
1,
Ruochen Mao
1,
Kathryn Bernard
2,
Lázaro Vélez
3,4,
Diana Marin
5,
Lucelly López
5,
Zulma Vanessa Rueda
1,5 and
Yoav Keynan
1,6,7,*
1
Department of Medical Microbiology and Infectious Diseases, University of Manitoba, Winnipeg, MB R3E 0J9, Canada
2
National Microbiology Laboratory, Public Health Agency of Canada, Winnipeg, MB R3E 3P6, Canada
3
School of Medicine, Universidad de Antioquia, Medellin 050010, Colombia
4
Infectious Diseases Section, Hospital Universitario San Vicente Fundación, Medellin 050010, Colombia
5
School of Medicine, Universidad Pontificia Bolivariana, Medellin 050010, Colombia
6
Department of Internal Medicine, University of Manitoba, Winnipeg, MB R3A 1R9, Canada
7
Department of Community Health Sciences, University of Manitoba, Winnipeg, MB R3E 0W3, Canada
*
Author to whom correspondence should be addressed.
Pathogens 2024, 13(2), 173; https://doi.org/10.3390/pathogens13020173
Submission received: 4 January 2024 / Revised: 7 February 2024 / Accepted: 12 February 2024 / Published: 14 February 2024
(This article belongs to the Special Issue Immunity to Respiratory Infections)
Versions Notes

Abstract

:
Legionella infections have a propensity for occurring in HIV-infected individuals, with immunosuppressed individuals tending to present with more severe disease. However, understanding regarding the Legionella host response in immune compromised individuals is lacking. This study investigated the inflammatory profiles associated with Legionella infection in patients hospitalized with HIV and pneumonia in Medellín, Colombia from February 2007 to April 2014, and correlated these profiles with clinical outcomes. Sample aliquots from the Colombian cohort were shipped to Canada where Legionella infections and systemic cytokine profiles were determined using real-time PCR and bead-based technology, respectively. To determine the effect of Legionella coinfection on clinical outcome, a patient database was consulted, comparing laboratory results and outcomes between Legionella-positive and -negative individuals. Principal component analysis revealed higher plasma concentrations of eotaxin, IP-10 and MCP-1 (p = 0.0046) during Legionella infection. Individuals with this immune profile also had higher rates of intensive care unit admissions (adjusted relative risk 1.047 [95% confidence interval 1.027–1.066]). Results demonstrate that systemic markers of monocyte/macrophage activation and differentiation (eotaxin, MCP-1, and IP-10) are associated with Legionella infection and worse patient outcomes. Further investigations are warranted to determine how this cytokine profile may play a role in Legionella pneumonia pathogenesis or immunity.

1. Introduction

Legionella, and in particular L. pneumophila serogroup 1, are increasingly recognized as a major cause of pneumonia, accounting for $434 million in hospitalization claims annually [1,2]. Legionella pneumonia often presents as nonspecific symptoms (cough, fever, malaise, dyspnea, etc.) [3], accompanied by pulmonary infiltrates, pleural effusions, and radiographic deterioration [3,4]. Swift and accurate diagnosis and treatment are crucial to prevent significant morbidity and mortality. In cases of severe infection, individuals with Legionella pneumonia frequently need hospitalization [4], particularly when complications arise such as acute lung injury, hypoxemia, and pulmonary fibrosis, necessitating admission to the intensive care unit (ICU) [5,6].
Amidst the rising trends in Legionella infections [7,8,9], knowledge of the host response to these infections is not well understood. During infection, Legionella infiltrates pulmonary macrophages and evades the phagosomal maturation process by establishing a specialized replication structure known as the Legionella containing vacuole [10]. As the bacterium multiplies, its Dot/Icm type IV secretion system discharges effector proteins into the host cytosol, manipulating the cell to favor its intercellular growth and survival [11]. Despite playing a role in the progression of Legionella infections, macrophages also assume a pivotal function in instigating the host inflammatory response [12,13]. Upon bacterial detection, resident macrophages prompt the release of inflammatory mediators from epithelial cells which, in turn, leads to the recruitment of neutrophil and monocyte cells, triggering a Th1-predominant response and eventual cell death [14], albeit understanding of the precise mechanisms and cytokines involved are still incomplete.
In response to Legionella infection, there is an upregulation of the macrophage mitogen-activated protein kinase signaling pathway [15,16], a process crucial for regulating the immune response and producing immunomodulatory cytokines such as tumor necrosis factor (TNF)-α, interleukin (IL)-1, IL-10, and IL-12 [17]. Other cytokines thought to contribute to the defense against Legionella infection also include interferon (IFN)-γ, IL-2, IL-4, IL-6, IL-8, IL-12, IL-18, and monocyte chemoattractant protein 1 (MCP-1) [13,15,18].
In populations with compromised immune systems, such as those with chronic lung disease, or those on TNF-α inhibitors or immunomodulatory drugs, higher rates of Legionella infection have been reported [13,18]. Recent studies also highlight a propensity for Legionella infections to occur in people living with HIV [19,20]. Moreover, immunosuppressed HIV-infected individuals have been shown to present with a more severe course of disease, resulting in higher mortality rates [20,21]. Despite these findings, there is a notable lack of studies investigating the role that Legionella infections play in the causation and consequences of pneumonia among people living with HIV. Additionally, an understanding regarding the host cytokine secretion during Legionella spp. in HIV-infected individuals is lacking. Thus, the aim of this study was to investigate the inflammatory profile associated with Legionella spp. infections in HIV and pneumonia coinfected individuals and to correlate these profiles with clinical outcomes.

2. Materials and Methods

2.1. Population, Samples, and Ethics

This investigation utilized bronchoalveolar lavage (BAL) and plasma samples obtained from a cohort of individuals with HIV and pneumonia. The cohort consisted of patients admitted to Hospital Universitario San Vicente Fundación in Medellín, Colombia, between February 2007 and April 2014 [22,23]. Inclusion criteria for the original cohort included adults aged 18 years and older, diagnosed with both HIV and pneumonia, and those that underwent bronchoscopy with BAL sample collection [24] at the time of hospital admission. Exclusion criteria involved individuals with underlying lung conditions other than pneumonia, hospitalization within the last 14 days, recent antibiotic exposure (within 3 days of admission), or any other non-HIV-related immunodeficiencies. Informed consent was obtained from all study participants and ethics approvals were received from both the University of Manitoba (Winnipeg, Canada) and the Universidad de Antioquia (Medellín, Colombia).

2.2. In-Hospital Procedures

In hospital, BAL samples underwent microbiological investigations (culture and staining) based on findings from the clinical assessment. For culture, samples were plated on standard agar for bacterial quantification [25]. Additionally, samples were plated on Mycosel, Girasol, and Sabouraud’s agar for fungal assessment, and on OgawaKudoh medium and thin-layer agar [26] for mycobacteria cultivation. Stains of interest included Wright, Gram, Ziehl–Neelsen, modified Kinyoun, and modified toluidine blue, according to the Kahn–Jones’ protocol [27].
During care, patients were admitted to the ICU based on patient monitoring, need for mechanical ventilation, and hospital recommendation, and details were recorded in patient clinical charts. Patient data, including past medical history, symptoms, physical findings, laboratory and microbiology results, and outcomes (ICU admission, intubation, mechanical ventilation, and mortality), were collected from hospital clinical charts and collated into a patient database.
Aliquots of BAL and plasma samples were shipped to Winnipeg, Canada for further analysis.

2.3. Legionella Identification and Immunological Array Assay

Legionella identification and cytokine analysis were determined retrospectively and independently. Researchers were blinded to results as well as the patient database until all sample assays were completed.
To identify Legionella infections, bacterial DNA was extracted from patient BAL samples and Legionella-positive controls using the QIAamp DNA Microbiome Kit (QIAGEN, Hilden, Germany) following the manufacturer’s instructions. The DNA was aliquoted and stored at −20 °C. For Legionella qPCR, primer and probe sequences and assay conditions were adapted from Benitez et al. [28] and Cross et al. [29]. Primer and probe oligonucleotides (Integrated DNA Technologies, Coralville, IA, USA) were aliquoted and stored at −20 °C until required.
BAL DNA and positive control DNA were screened for Legionella by targeting a section of the 23 S–5 S intergenic spacer region, which encompasses both conserved and variable regions. Each PCR reaction contained the following: 12.5 μL of (2X) PrimeTime Gene Expression Mastermix (Integrated DNA Technologies, Coralville, IA, USA), 5 μL of template, 150 nM of each primer, and 100 nM of probe. All reactions were run using a C1000 Touch Thermal Cycler (Bio-Rad, Mississauga, ON, Canada). Cycling conditions consisted of an initial denaturation step of 95 °C for 300 s then 45 PCR cycles of 15 s at 95 °C and 60 s at 60 °C. Following qPCR, data analysis was performed using the CFX Manager System Software, Version 3.1 (Bio-Rad, Mississauga, ON, Canada). Samples were considered positive if they amplified with a sigmoidal curve and had a crossing threshold value greater than 10. All samples were run in duplicate and any discordant results were resolved using a third replicate.
Immunological testing was performed on plasma, cell-free supernatants using a Bio-Plex Pro Human Cytokine Magnetic 19-Plex Panel (Bio-Rad, Mississauga, ON, Canada) on the Bio-Plex platform (version 6.0 software) and was carried out according to the manufacturers’ instructions. Targets of interest included: IFN-γ, TNF-α, IL-6, IL-7, IL-8, IL-10, IL-12, IL-13, IL-17, IL-1 receptor antagonist (IL-1RA), regulated on activation, normal T-cell expressed and secreted (RANTES), vascular endothelial growth factor (VEGF), granulocyte colony-stimulating factor (G-CSF), recombinant human platelet-derived growth factor-BB (PDGF-BB), eotaxin, interferon gamma-induced protein 10 (IP-10), macrophage inflammatory protein (MIP)-1α, MIP-1β, and MCP-1. All testing was performed in duplicate. Results are reported as mean fluorescence intensity and converted to pictogram/mL concentrations using the Bio-Plex Manager version 6.0 software.

2.4. Data Analysis

Data were analysed using STATA® version 14. Frequencies, median plus interquartile range (IQR), or mean plus standard deviation were calculated for patient characteristics. Patients were divided into two groups based on whether they had a Legionella infection. Categorical variables were analysed between the two groups using the Pearson’s Chi-square test, or Fisher’s exact test if n < 5. Normally distributed continuous variables were compared using the T-test, while non-normal data were compared using the Wilcoxon test. Differences between groups were considered statistically significant if p ≤ 0.05.
Systemic immunological factors were analysed to determine whether they differed between the two groups (Legionella-positive vs. -negative) and if they correlated with clinical outcome (defined as ICU admission). Cytokines and chemokines are reported as median (IQR) and were compared using a two-sample Wilcoxon rank-sum test.
Next, to determine the systemic cytokine profile associated with Legionella infection, a principal component analysis (PCA) was conducted. PCA is a widely employed statistical technique used to discern patterns within data, revealing specific constellations of markers that contribute significantly to variations between groups, such as Legionella-infected and uninfected individuals [30]. In this analysis, components were extracted using the principal axis method and rotated using the varimax rotation, a linear transformation facilitating a more straightforward interpretation of the data. The rotated factor pattern that accounted for at least 10% of the total variance was chosen. This factor pattern represents a set of cytokines collectively explaining a substantial portion of the differences between Legionella-infected and uninfected individuals. A cytokine was considered to load onto a specific factor component wherever the loading value was the highest (i.e., the cytokine/chemokine had a strong association with that component). Subsequently, the inflammatory markers that loaded onto each rotated component were compared between the Legionella-positive and -negative groups using the two-sample Wilcoxon rank-sum test. A Poisson regression was then performed to identify which factors associated with poor clinical outcome (ICU admission) or those associated with Legionella infection. Covariates selected for the regression analysis were those with p < 0.25 in the univariate analysis, or those deemed biologically relevant based on the literature or prior analyses from our group. Categorical variables were directly integrated into the model, while continuous variables were recoded prior to the analysis. Interaction was evaluated using the Mantel–Haenszel test of homogeneity, and if p ≤ 0.05, an interaction term was introduced for the corresponding variables. The model was adjusted for confounders and interaction terms, and results are presented as crude and adjusted relative risk (RR) with 95% CI, alongside the corresponding p values.

3. Results

Forty-seven patients were included in this study, all of which were diagnosed in hospital with HIV and pneumonia (Table 1; see reference [23] for in-depth details). Individuals consisted predominantly of immunosuppressed males (CD4 cell count < 50 cells/μL, IQR 16-101), with a mean age of 35 years, with the most common in-hospital diagnoses consisting of Mycobacterium tuberculosis or Pneumocystis jirovecii infection.
Patients were divided into two groups based on our retrospective analysis: (1) Legionella-positive individuals with HIV and pneumonia (n = 17); and (2) Legionella-negative individuals with HIV and pneumonia (n = 30). Based on patient clinical charts, individuals with Legionella infections had higher rates of ICU admissions compared to Legionella-negative individuals (n = 9 [53%] vs. n = 7 [23%], respectively; p = 0.04) as well as higher mortality rates (n = 9 [53%] vs. n = 6 [20%], respectively; p = 0.02).
Of the 19 measured systemic inflammatory markers, the majority were higher in the Legionella-positive patients, of which only 2 were significantly different between the patient groups, namely IL-7 and eotaxin (Table 2). Individuals with Legionella infection had higher IL-7 plasma levels compared to Legionella-negative individuals (0.50 [IQR 0.24–1.91] and 0.24 [IQR 0.24–0.25] pg/mL, respectively, p = 0.04). Likewise, plasma eotaxin was also higher among the Legionella-infected population (5.27 [IQR 3.58–17.13] vs. 3.08 [IQR 2.06–3.86] pg/mL, p < 0.001).
The PCA of plasma immune markers in Legionella-positive and -negative patient samples at hospital admission can be seen in Table 3. PCA identified five principal components which had eigenvalues greater than one that were included in the analysis. Together, the five factors accounted for 83.9% of the total variance (22.2%, 20.6%, 20.2%, 10.7% and 10.3% for Factors 1, 2, 3, 4, and 5, respectively). In the rotated factor analysis, six molecules loaded onto component 1 (IL-7, IL-10, IL-12, IL-13, RANTES, and VEGF), four onto component 2 (IL-6, G-CSF, IFN-γ, and TNF-α), four onto component 3 (IL-17, MIP-1α, PDGF-BB, and MIP-1β), three onto component 4 (eotaxin, IP-10, and MCP-1), and two loaded onto the last component (IL-1RA and IL-8).
Of the five component factors, Factor 4 was the only one that was significantly different between the Legionella positive and negative groups (p = 0.622, p = 0.376, p > 0.999, p = 0.005, and p = 0.757, for factors 1, 2, 3, 4, and 5, respectively).
In a binomial logistic regression analysis, higher plasma concentrations of Factor 4 (eotaxin, IP-10 and MCP-1) associated with higher rates of ICU admission. Moreover, individuals with this immune profile had a 5% higher risk of ICU admission, even after controlling for potential confounders such as Legionella infection, viral load, CD4 count, and TB coinfection (crude RR 1.021 [CI 1.011–1.031] vs. adjusted RR 1.047 [CI 1.027–1.066]).

4. Discussion

Despite being a major cause of respiratory-associated drinking water outbreaks [32], an understanding of host immunity to Legionella spp. infections, particularly in HIV-infected individuals, is lacking. To expand our knowledge base, we studied the plasma cytokine response in Legionella-infected individuals with HIV-associated pneumonias.
Previous studies looking at cytokines involved in Legionella immunity in immune-competent individuals found that Legionella-positive individuals had high serum levels of TNF-α, IFN-γ, and IL-12 during infection [13], while in vitro and in vivo studies reported elevated levels of TNF-α, IFN-γ, IL-2, IL-4, IL-6, IL-8, IL-12, IL-18, and MCP-1 [15,18,33]. The present research builds on previously reported work by adding evidence on the systemic involvement of MCP-1, IP-10, and eotaxin. As mononuclear phagocytes are important cells in Legionella infection, it is not surprising that chemokines that attract or stimulate the formation of these cell types associated with Legionella infection. Indeed, MCP-1 is a key marker regularly involved in monocyte and macrophage recruitment and infiltration [34]. Although it is primarily secreted by phagocytic inflammatory cells, it is also produced by lymphocytes, fibroblasts, and airway endothelial and epithelial cells, following monocyte chemotaxis and oxidative stress [35,36,37,38]. In a study by Matsunaga et al., MCP-1 was found to be upregulated in macrophage cells following stimulation with Legionella lipopolysaccharide [33]. This chemokine has also been linked to several pulmonary comorbidities including interstitial lung disease [36], acute lung injury [38], and intracellular infections [35,36,37]. Furthermore, a recent publication by Allam et al. [39] involving 84 hospitalized patients with Legionella pneumonia reported that individuals admitted to the ICU with mechanical ventilation had increased levels of plasmatic MCP-1, MIP-1β, IL-6, IL-8, IFN-γ, TNF-α, and IL-17. They also highlighted that upon cell stimulation, Legionella-positive patients exhibited elevated release of IL-18 and MCP-1 compared to healthy individuals.
Similarly, a previous study by Lettinga and co-workers which also looked at IP-10 and its association with Legionella infection found that IP-10 release following whole blood stimulation with IFN-γ weakly correlated with the severity of Legionella pneumonia [40]. IP-10, expressed by Th1 cells, monocytes, fibroblasts, and endothelial cells, plays a crucial role in inflammation by recruiting T lymphocytes and monocytes to the site of infection [41]. It activates T-cells, inducing IFN-γ release and enhancing macrophage activity through a positive feedback loop mechanism [42]. Earlier research has highlighted the essential role of the IP-10 signaling pathway in eradicating intracellular infections [41,43]. During infection, IP-10 orchestrates the adaptive immune response by triggering the mitogen-activated protein kinase and phosphoinositide 3-kinase pathways, thereby affecting chemotaxis, apoptosis, and cell proliferation as well as various cell types (natural killer cells, dendritic cells, and B cells) [43].
In contrast, an association between eotaxin and Legionella has not yet been described; however, higher levels have been reported in severe community-acquired pneumonia and M. tuberculosis infections [22,44]. Eotaxin upregulation has also been reported in other intracellular infections as well [45,46,47]. Induced by cell stimulation with IFN-γ, eotaxin is produced by endothelial and epithelial cells, eosinophils, monocytes, and macrophages [48]. Once produced, this small protein binds to the CCR3 receptor present on eosinophils, basophils, Th1 and Th2 cells, and airway epithelium [48], initiating a cascade that activates the mitogen-activated protein kinase pathway, increases oxygen radical species production, induces monocytes, recruits eosinophils, basophils, and Th2 lymphocytes, and triggers eosinophil granule release [48,49]. Considering the insights gained in this study and the role that eotaxin, IP-10, and MCP-1 play in the immune response, the evidence suggests that markers of monocyte and macrophage activation and differentiation may play a role in Legionella pneumonia pathogenesis and host immunity, warranting further investigation.
An additional intriguing finding is the higher ICU admission rates among Legionella-infected individuals, with those having a Factor 4 profile facing an increased risk of being admitted to the ICU. These findings are in agreement with Wolter et al., which showed that Legionella patients had case-fatality ratios twice that of individuals who had other severe respiratory illnesses [50]. Moreover, as was seen in our patient population, they found that Legionella-positive cases often occurred in coinfected individuals (67%) with HIV, TB, or both HIV and TB.
This work contributes to the existing knowledge surrounding Legionella infection and describes for the first time, to the best of our knowledge, systemic cytokine profiles in the context of Legionella and HIV coinfections. Another strength of this research lies in the fact that Legionella identification and cytokine analysis were performed independently by two researchers who were blinded to the results until all samples were completed. However, a major limitation of this study is that, since it was based on a previously established cohort, we were unable to include individuals with Legionella monoinfection (in HIV-infected and HIV-uninfected individuals) in the analysis. Without comparison with these groups, the observations herein cannot definitely state whether the association between Factor 4 and Legionella-infected individuals is based on the Legionella and HIV coinfection or due to the Legionella-induced pneumonia irrespective of HIV. Additionally, due to the retrospective nature of the study, we cannot establish a causal relationship between Legionella infection and the Factor 4 profile. Thirdly, as we had a finite volume of BAL samples to which we had access, cytokine analysis could only be performed on patient plasma samples; therefore, we were only able to look at systemic cytokine response which may differ from that seen at the local level. Lastly, our patient population was small and fairly homogenous in that the majority of patients were highly immunosuppressed males with similar coinfections (all with HIV, and most with either tuberculosis or Pneumocystis jirovecii pneumonia); therefore, data may not be extrapolated to other patient populations. Other researchers are encouraged to verify our findings as very few groups have studied Legionella coinfection among people living with HIV.
While acknowledging its limitations and exploratory nature, this study provides valuable insights, suggesting that plasmatic markers of monocyte and macrophage activation and differentiation (eotaxin, IP-10 and MCP-1) are associated with Legionella infection in individuals coinfected with HIV and M. tuberculosis or P. jirovecii infection. Furthermore, it indicates that this systemic cytokine pattern is linked to more severe outcomes. Although further work is needed to explore and validate these findings in additional, larger, and prospective cohorts, this study marks an important step towards better understanding cytokines associated with Legionella infection in immunocompromised individuals. The insights gained from this work can serve as a foundation for future studies investigating the role that Legionella spp. play in inflammation and disease in people living with HIV.

Author Contributions

Conceptualization, B.M.H. and Y.K.; methodology, B.M.H., A.T., R.M.; formal analysis, B.M.H., D.M., and L.L.; investigation, B.M.H.; resources, K.B.; data curation, B.M.H.; writing—original draft preparation, B.M.H.; writing—review and editing, B.M.H., K.B., L.V., D.M., L.L., Z.V.R., and Y.K.; supervision, Z.V.R. and Y.K.; project administration, Z.V.R. and Y.K.; funding acquisition, Y.K. All authors have read and agreed to the published version of the manuscript.

Funding

This project was funded by Minciencias (Ministerio de Ciencia, Tecnologia e Innovacion; Grant number: 111580763362), Universidad de Antioquia, Universidad Pontificia Bolivariana, and University of Manitoba, which provided materials and reagents for sample processing. This research was also supported, in part, by the Canada Research Chairs Program for ZVR (Award # 950-232963).

Institutional Review Board Statement

The study was conducted according to the guidelines of the Declaration of Helsinki, and approved by the Ethics Committees at the University of Manitoba (H2014:077; 30 September 2014) and the University of Antioquia (initial study approved by the “Comité de Ética del Centro de Investigaciones de la Facultad de Medicina” [10 May 2006, no. 003]; with an amended approval for conducting experiments in Canada received from the “Comite de Bioetica del Instituto de Investigaciones Médicas de la Facultad de Medicina” [14 February 2014, no. 002]). All study participants provided written informed consent. Sample shipment was approved by the Ministerio de Salud y Protección Social de Colombia.

Informed Consent Statement

Informed consent was obtained from all subjects involved in the study.

Data Availability Statement

The data supporting the findings of this study are available within the article as well as in Head et al. [23].

Acknowledgments

The authors would like to thank their Colombian collaborators for their help in organizing the cohort and providing the samples. Additionally, they would like to acknowledge Kimberly Nichol from the Diagnostics Services Manitoba at the Health Sciences Centre (Winnipeg, Canada) for providing the Legionella-positive controls.

Conflicts of Interest

The authors declare no conflicts of interest. The funders had no role in the design of the study; in the collection, analyses, or interpretation of data; in the writing of the manuscript; or in the decision to publish the results.

References

  1. Collier, S.A.; Stockman, L.J.; Hicks, L.A.; Garrison, L.E.; Zhou, E.J.; Beach, M.J. Direct healthcare costs of selected diseases primarily or partially transmitted by water. Epidemiol. Infect. 2012, 140, 2003–2013. [Google Scholar] [CrossRef]
  2. Herwaldt, L.A.; Marra, A.R. Legionella: A reemerging pathogen. Curr. Opin. Infect. Dis. 2018, 31, 325–333. [Google Scholar] [CrossRef]
  3. Murder, R.R.; Yu, V.L. Infection due to Legionella species other Than L. pneumophila. Emerg. Infect. Dis. 2002, 35, 990–999. [Google Scholar] [CrossRef]
  4. Mondino, S.; Schmidt, S.; Rolando, M.; Escoll, P.; Gomez-Valero, L.; Buchrieser, C. Legionnaires’ Disease: State of the Art Knowledge of Pathogenesis Mechanisms of Legionella. Annu. Rev. Pathol. Mech. Dis. 2020, 15, 439–466. [Google Scholar] [CrossRef]
  5. Nara, C.; Tateda, K.; Matsumoto, T.; Ohara, A.; Miyazaki, S.; Standiford, T.J.; Yamaguchi, K. Legionella-induced acute lung injury in the setting of hyperoxia: Protective role of tumour necrosis factor-alpha. J. Med. Microbiol. 2004, 53, 727–733. [Google Scholar] [CrossRef] [PubMed]
  6. Chastre, J.; Raghu, G.; Soler, P.; Brun, P.; Basset, F.; Gibert, C. Pulmonary fibrosis following pneumonia due to acute legionnaires’ disease. Chest 1987, 91, 57–62. [Google Scholar] [CrossRef] [PubMed]
  7. European Centre for Disease Prevention and Control. Legionnaires’ Disease—Annual Epidemiological Report for 2017; European Centre for Disease Prevention and Control: Stockholm, Sweden, 2019; Available online: https://www.ecdc.europa.eu/sites/default/files/documents/AER_for_2017-Legionnaires-disease_1.pdf (accessed on 5 February 2024).
  8. Public Health Investigating Legionella Cases: 2017. Available online: https://www.hamilton.ca/government-information/news-centre/news-releases/publichealth-investigating-legionella-cases (accessed on 6 March 2019).
  9. Centers for Disease Control and Prevention. Legionellosis—United States, 2000–2009. Morb. Mort. Week. Rep. 2011, 60, 1083–1086. [Google Scholar]
  10. Moffat, J.F.; Tompkins, L.S. A quantitative model of intracellular growth of Legionella pneumophila in Acanthamoeba castellanii. Infect. Immun. 1992, 60, 296–301. [Google Scholar] [CrossRef] [PubMed]
  11. Luo, Z. Striking a balance: Modulation of host cell death pathways by Legionella pneumophila. Front. Microbiol. 2011, 2, 1–6. [Google Scholar] [CrossRef] [PubMed]
  12. Ziltener, P.; Reinheckel, T.; Oxenius, A. Neutrophil and alveolar macrophage- mediated innate immune control of Legionella pneumophila lung infection via TNF and ROS. PLoS Pathog. 2016, 12, e1005591. [Google Scholar] [CrossRef] [PubMed]
  13. Tateda, K.; Matsumoto, T.; Ishii, Y.; Furuya, N.; Ohno, A.; Miyazaki, S.; Yamaguchi, K. Serum cytokines in patients with Legionella pneumonia: Relative predominance of Th1-type cytokines. Clin. Diagn. Lab. Immunol. 1998, 5, 401–403. [Google Scholar] [CrossRef] [PubMed]
  14. Pereira, M.S.F.; Marques, G.G.; Dellama, J.E.; Zamboni, D.S.; Voth, D.E. The Nlrc4 inflammasome contributes to restriction of pulmonary infection by flagellated Legionella spp. that trigger pyroptosis. Front. Microbiol. 2011, 2, 33. [Google Scholar] [CrossRef] [PubMed]
  15. Newton, H.J.; Ang, D.K.Y.; Van Driel, I.R.; Hartland, E.L. Molecular pathogenesis of infections caused by Legionella pneumophila. Clin. Microbiol. Rev. 2010, 23, 274–298. [Google Scholar] [CrossRef] [PubMed]
  16. Morrison, D.K. MAP Kinase Pathways. Cold. Spring. Harb. Perspect. Biol. 2012, 4, a011254. [Google Scholar] [CrossRef] [PubMed]
  17. Soares-Silva, M.; Diniz, F.F.; Gomes, G.N.; Bahia, D. The Mitogen-Activated Protein Kinase (MAPK) Pathway: Role in Immune Evasion by Trypanosomatids. Front. Microbiol. 2016, 7, 183. [Google Scholar] [CrossRef]
  18. Brieland, J.K.; Jackson, C.; Hurst, S.; Loebenberg, D.; Muchamuel, T.; Debets, R.; Kastelein, R.; Churakova, T.; Abrams, J.; Hare, R.; et al. Immunomodulatory role of endogenous interleukin- 18 in gamma interferon-mediated resolution of replicative Legionella pneumophila lung infection. Infect. Immun. 2000, 68, 6567–6573. [Google Scholar] [CrossRef]
  19. Head, B.M.; Trajtman, A.; Rueda, Z.V.; Vélez, L.; Keynan, Y. Atypical bacterial pneumonia in the HIV-infected population. Pneumonia 2017, 9, 12. [Google Scholar] [CrossRef]
  20. Weiss, D.; Boyd, C.; Rakeman, J.L.; Greene, S.K.; Fitzhenry, R.; McProud, T.; Musser, K.; Huang, L.; Kornblum, J.; Nazarian, E.J.; et al. A Large Community Outbreak of Legionnaires’ Disease Associated With a Cooling Tower in New York City, 2015. Public Health Rep. 2017, 132, 241–250. [Google Scholar] [CrossRef]
  21. Durrance, R.J.; Min, A.K.; Fabbri, M.; McGarry, T. Cavitary Legionella Pneumonia in AIDS: When Intracellular Immunity Failure Leads to Rapid Intrapulmonary Cavitation. Case Rep. Pulmon. 2021, 2021, 6754094. [Google Scholar] [CrossRef] [PubMed]
  22. Keynan, Y.; Rueda, Z.V.; Aguilar, Y.; Trajtman, A.; Vélez, L. Unique cytokine and chemokine patterns in bronchoalveolar lavage are associated with specific causative pathogen among HIV infected patients with pneumonia, in Medellin, Colombia. Cytokine 2015, 73, 295–301. [Google Scholar] [CrossRef] [PubMed]
  23. Head, B.M.; Trajtman, A.; Bernard, K.; Burdz, T.; Vélez, L.; Herrera, M.; Rueda, Z.V.; Keynan, Y. Legionella co-infection in HIV-associated pneumonia. Diagn. Microbiol. Infect. Dis. 2019, 95, 71–76. [Google Scholar] [CrossRef] [PubMed]
  24. Goldstein, R.A.; Rohatgi, P.K.; Bergofsky, E.H.; Block, E.R.; Daniele, R.P.; Dantzker, D.R.; Davis, G.S.; Hunninghake, G.W.; King Jr, T.E.; Metzger, W.J. Clinical role of bronchoalveolar lavage in adults with pulmonary disease. Am. Rev. Respir. Dis. 1990, 142, 481–486. [Google Scholar] [CrossRef] [PubMed]
  25. Velez, L.; Corread, L.T.; Mayad, M.A.; Mejıad, P.; Ortegab, J.; Bedoyac, V.; Ortega, H. Diagnostic accuracy of bronchoalveolar lavage samples in immunosuppressed patients with suspected pneumonia: Analysis of a protocol. Resp. Med. 2007, 101, 2160–2167. [Google Scholar] [CrossRef] [PubMed]
  26. Mejia, G.I.; Castrillon, L.; Trujillo, H.; Robledo, J.A. Microcolony detection in 7H11 thin layer culture is an alternative for rapid diagnosis of Mycobacterium tuberculosis infection. Int. J. Tuberc. Lung. Dis. 1999, 3, 138–142. [Google Scholar] [PubMed]
  27. Kahn, F.W.; Jones, J.M. Analysis of bronchoalveolar lavage specimens from immunocompromised patients with a protocol applicable in the microbiology laboratory. J. Clin. Microbiol. 1988, 26, 1150–1155. [Google Scholar] [CrossRef] [PubMed]
  28. Benitez, A.J.; Winchell, J.M. Clinical application of a multiplex real-time PCR assay for simultaneous detection of Legionella species, Legionella pneumophila, and Legionella pneumophila serogroup 1. J. Clin. Microbiol. 2013, 51, 348–351. [Google Scholar] [CrossRef] [PubMed]
  29. Cross, K.E.; Mercante, J.W.; Benitez, A.J.; Brown, E.W.; Diaz, M.H.; Winchell, J.M. Simultaneous detection of Legionella species and L. anisa, L. bozemanii, L. longbeachae and L. micdadei using conserved primers and multiple probes in a multiplex real-time PCR assay. Diagn. Microbiol. Infect. Dis. 2016, 85, 295–301. [Google Scholar] [CrossRef] [PubMed]
  30. Smith, L. A tutorial on Principal Components Analysis. Commun. Stat. Theory. Methods 2002, 17, 1–27. Available online: https://www.iro.umontreal.ca/~pift6080/H09/documents/papers/pca_tutorial.pdf (accessed on 25 January 2024).
  31. Head, B. Legionella- an underappreciated but important pathogen in HIV. Ph.D. Thesis, University of Manitoba, Winnipeg, MB, Canada, 2020. [Google Scholar]
  32. Benedict, K.; Reses, H.; Vigar, M.; Al, E. Surveillance for waterborne disease outbreaks associated with drinking water- United States, 2013–2014. Morb. Mortal. Wkly. Rep. 2017, 66, 1216–1221. [Google Scholar] [CrossRef]
  33. Matsunaga, K.; Klein, T.W.; Newton, C.; Friedman, H. Legionella pneumophila Suppresses Interleukin-12 Production by Macrophages. Infect. Immun. 2001, 69, 1929–1933. [Google Scholar] [CrossRef]
  34. Singh, S.; Anshita, D.; Ravichandiran, V. MCP-1: Function, regulation, and involvement in disease. Int. Immunopharmacol. 2021, 101, 107598. [Google Scholar] [CrossRef]
  35. Lin, Y.; Gong, J.; Zhang, M.; Xue, W.; Barnes, P.F. Production of monocyte chemoattractant protein 1 in tuberculosis patients. Infect. Immun. 1998, 66, 2319–2322. [Google Scholar] [CrossRef]
  36. Deshmane, S.L.; Kremlev, S.; Amini, S.; Sawaya, B.E. Monocyte chemoattractant protein-1 (MCP-1): An overview. J. Interf. Cytokine. Res. 2009, 29, 313–325. [Google Scholar] [CrossRef] [PubMed]
  37. Weiss, G.; Schaible, U.E. Macrophage defense mechanisms against intracellular bacteria. Immunol. Rev. 2015, 264, 182–203. [Google Scholar] [CrossRef] [PubMed]
  38. Hartl, D.; Griese, M.; Nicolai, T.; Zissel, G.; Prell, C.; Reinhardt, D.; Schendel, D.J.; Krauss-Etschmann, S. A role for MCP-1/CCR2 in interstitial lung disease in children. Respir. Res. 2005, 6, 93. [Google Scholar] [CrossRef] [PubMed]
  39. Allam, C.; Mouton, W.; Testaert, H.; Ginevra, C.; Fessy, N.; Ibranosyan, M.; Descours, G.; Beraud, L.; Guillemot, J.; Chapalain, A.; et al. Hyper-inflammatory profile and immunoparalysis in patients with severe Legionnaires’ disease. Front. Cell. Infect. Microbiol. 2023, 13, 1252515. [Google Scholar] [CrossRef] [PubMed]
  40. Lettinga, K.; Weijer, S.; Speelman, P.; Prins, J.; Van der Poll, T.; Verbon, A. Reduced interferon-γ release in patients recovered from Legionnaires’ disease. Thorax 2003, 58, 63–67. [Google Scholar] [CrossRef]
  41. Antonia, A.L.; Gibbs, K.D.; Trahair, E.D.; Pittman, K.J.; Martin, A.T.; Schott, B.H.; Smith, J.S.; Rajagopal, S.; Thompson, J.W.; Reinhardt, R.L.; et al. Pathogen evasion of chemokine response through suppression of CXCL10. Front. Cell. Infect. Microbiol. 2019, 9, 280. [Google Scholar] [CrossRef]
  42. Madhurantakam, S.; Lee, Z.L.; Naqvi, A.; Prasad, S. Importance of IP-10 as a biomarker of host immune response: Critical perspective as a target for biosensing. Curr. Res. Biotechnol. 2023, 5, 100130. [Google Scholar] [CrossRef]
  43. Liu, M.; Guo, S.; Hibbert, J.M.; Jain, V.; Singh, N.; Wilson, N.O.; Stiles, J.K. CXCL10/IP-10 in infectious diseases pathogenesis and potential therapeutic implications. Cytokine Growth Factor Rev. 2011, 22, 121–130. [Google Scholar] [CrossRef]
  44. Haugen, J.; Chandyo, R.K.; Brokstad, K.A.; Mathisen, M. Cytokine Concentrations in Plasma from Children with Severe and Non-Severe Community Acquired Pneumonia. PLoS ONE 2015, 10, e0138978. [Google Scholar] [CrossRef]
  45. Dávila, H.; Almeida, P.E.; Roque, N.R.; Castro-Faria-Neto, H.C.; Bozza, P.T. Toll-like receptor-2- mediated C-C chemokine receptor 3 and eotaxin-driven eosinophil influx induced by Mycobacterium bovis BCG pleurisy. Infect. Immun. 2007, 75, 1507–1511. [Google Scholar] [CrossRef]
  46. Raqib, R.; Moly, P.K.; Sarker, P.; Qadri, F.; Alam, N.H.; Mathan, M.; Andersson, J. Persistence of mucosal mast cells and eosinophils in Shigella-infected children. Infect. Immun. 2003, 71, 2684–2692. [Google Scholar] [CrossRef] [PubMed]
  47. Medeiros, A.I.; Malheiro, A.; Jose, P.J.; Conroy, D.M.; Williams, T.J.; Faccioli, L.H. Differential release of MIP-1α and eotaxin during infection of mice by Histoplasma capsulatum or inoculation of β-glucan. Inflamm. Res. 2004, 53, 351–354. [Google Scholar] [CrossRef] [PubMed]
  48. Conroy, D.M.; Williams, T.J. Eotaxin and the attraction of eosinophils to the asthmatic lung. Respir. Res. 2001, 2, 150–156. [Google Scholar] [CrossRef] [PubMed]
  49. Ogilvie, P.; Paoletti, S.; Clark-lewis, I.; Uguccioni, M. Eotaxin-3 is a natural antagonist for CCR2 and exerts a repulsive effect on human monocytes. Chemokines 2003, 102, 789–794. [Google Scholar] [CrossRef] [PubMed]
  50. Wolter, N.; Carrim, M.; Cohen, C.; Tempia, S.; Walaza, S.; Sahr, P.; de Gouveia, L.; Treurnicht, F.; Hellferscee, O.; Cohen, A.L.; et al. Legionnaires’ disease in South Africa, 2012–2014. Emerg. Infect. Dis. 2016, 22, 131–133. [Google Scholar] [CrossRef]
Table 1. Demographics and clinical characteristics of all HIV and pneumonia patients with and without Legionella infection.
Table 1. Demographics and clinical characteristics of all HIV and pneumonia patients with and without Legionella infection.
All Patients
(n = 47)		Legionella-Negative
(n = 30)		Legionella-Positive
(n = 17)		p Value 3
Demographics
  Age, mean (SD 2), years35.7 (8.8)36.7 (8.4)33.9(9.5)0.279
  Male, n (%)38 (81)27 (90)11 (65)0.054
  CD4 cell count (median cells/μL, IQR 1)36.5 (16–101)25 (12–92.8)32.5 (16–90.5)0.957
Pneumonia (n,%)
   Pneumocystis jirovecii15 (32)9 (30)6 (35)0.708
   Mycobacterium tuberculosis19 (40)9 (30)10 (59)0.054
  Other13 (28)12 (40)1 (5.9)0.012
Outcome
  ICU admission, n (%)16 (34)7 (23)9 (53)0.040
  Intubation, n (%)6 (13)3 (10)3 (18)>0.999
  Mechanical ventilation, n (%)4 (8.5)2 (6.7)2 (12)0.613
  Death, n (%)15 (32)6 (20)9 (53)0.020
1 IQR: interquartile range; 2 SD: standard deviation; 3 p values comparing Legionella positive and negative groups; table adapted from Head et al. [23] and reference [31].
Table 2. Plasma inflammatory mediator concentrations for Legionella-positive and -negative coinfected individuals at hospital admission.
Table 2. Plasma inflammatory mediator concentrations for Legionella-positive and -negative coinfected individuals at hospital admission.
Legionella-Positive
Median (IQR 1) pg/mL		Legionella-Negative
Median (IQR) pg/mL 		p Value 2
IL-1Ra59.9 (34.41–216.72)63.72 (30.28–166.87)0.65
IL-611.43 (1.68–69.61)4.13 (0.94–11.47)0.22
IL-70.50 (0.24–1.91)0.24 (0.24–0.25)0.04
IL-8235.7 (91.54–438.18)137.71 (27.59–436.19)0.45
IL-101.66 (1.12–2.39)1.29 (1.09–1.67)0.28
IL-124.45 (3.48–9.31)4.55 (2.85–6.98)0.77
IL-132.01 (1.45–3.26)1.43 (0.99–3.81)0.27
IL-174.03 (2.28–5.09)2.87 (2.20–3.65)0.43
eotaxin5.27 (3.58–17.13)3.08 (2.60–3.86)<0.001
GCSF44.27 (11.29–73.41)20.39 (9.66–48.18)0.39
IFN-y1.11 (1.1–8.75)1.10 (1.10–2.22)0.18
IP-102017.09 (385.88–7902.06)496.58 (49.56–1618.3)0.07
MCP-1133.84 (64.57–464.29)98.94 (46.7–211.93)0.12
MIP-1a3.66 (1.83–5.85)1.66 (1.14–3.92)0.07
PDGFBB2.87 (2.02–6.48)2.55 (1.6–7.08)0.50
MIP-1b52.97 (15.86–82.19)18.03 (8.46–46.59)0.14
RANTES182.35 (80.08–293.91)125.5 (22.87–269.71)0.44
TNF-a2.49 (1.63–3.46)2.12 (1.43–3.22)0.46
VEGF38.08 (23.69–94.31)59.0 (23.56–129.79)0.54
1 IQR: interquartile range; 2 Two-sample Wilcoxon rank-sum (Mann–Whitney) test.
Table 3. Principal component analysis of plasma immunological markers in Legionella-positive and -negative patient samples at hospital admission.
Table 3. Principal component analysis of plasma immunological markers in Legionella-positive and -negative patient samples at hospital admission.
Rotated Component Matrix
VariableFactor1Factor2Factor3Factor4Factor5
IL-70.8669−0.13460.17150.17480.0641
IL-100.77100.37060.22380.28950.2582
IL-120.90910.08970.13070.11460.0667
IL-130.65620.23330.40580.0078−0.1972
RANTES0.7304−0.08630.37110.0116−0.2910
VEGF0.65760.04450.0447−0.19780.6067
IL-60.03220.9334−0.04110.2984−0.0054
G-CSF0.07270.95460.05450.1163−0.0033
IFN-γ0.04250.88670.32150.25800.0208
TNF-α−0.00230.70500.0396−0.07680.3273
IL-170.28060.43280.78310.17160.0922
MIP-1α0.12880.02620.93550.03490.0392
PDGF-BB0.62810.02820.72000.0162−0.1241
MIP-1β0.24360.03870.91200.13040.0418
eotaxin0.24570.31610.00720.82400.2004
IP-100.04460.42560.38260.74810.0394
MCP-10.16470.36250.14120.5934 −0.1809
IL-1RA0.01710.02850.00900.13100.9320
IL-8−0.03240.14520.56220.16970.5624
Note: Molecules that loaded onto each component are shaded. Extraction method: principal component analysis. Rotation method: varimax with Kaiser normalization.
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Head, B.M.; Trajtman, A.; Mao, R.; Bernard, K.; Vélez, L.; Marin, D.; López, L.; Rueda, Z.V.; Keynan, Y. Inflammatory Patterns Associated with Legionella in HIV and Pneumonia Coinfections. Pathogens 2024, 13, 173. https://doi.org/10.3390/pathogens13020173

AMA Style

Head BM, Trajtman A, Mao R, Bernard K, Vélez L, Marin D, López L, Rueda ZV, Keynan Y. Inflammatory Patterns Associated with Legionella in HIV and Pneumonia Coinfections. Pathogens. 2024; 13(2):173. https://doi.org/10.3390/pathogens13020173

Chicago/Turabian Style

Head, Breanne M., Adriana Trajtman, Ruochen Mao, Kathryn Bernard, Lázaro Vélez, Diana Marin, Lucelly López, Zulma Vanessa Rueda, and Yoav Keynan. 2024. "Inflammatory Patterns Associated with Legionella in HIV and Pneumonia Coinfections" Pathogens 13, no. 2: 173. https://doi.org/10.3390/pathogens13020173

APA Style

Head, B. M., Trajtman, A., Mao, R., Bernard, K., Vélez, L., Marin, D., López, L., Rueda, Z. V., & Keynan, Y. (2024). Inflammatory Patterns Associated with Legionella in HIV and Pneumonia Coinfections. Pathogens, 13(2), 173. https://doi.org/10.3390/pathogens13020173

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