Microbial community organization designates distinct pulmonary exacerbation types and predicts treatment outcome in cystic fibrosis.

Polymicrobial infection of the airways is a hallmark of obstructive lung diseases such as cystic fibrosis (CF), non-CF bronchiectasis, and chronic obstructive pulmonary disease. Pulmonary exacerbations (PEx) in these conditions are associated with accelerated lung function decline and higher mortality rates. Understanding PEx ecology is challenged by high inter-patient variability in airway microbial community profiles. We analyze bacterial communities in 880 CF sputum samples collected during an observational prospective cohort study and develop microbiome descriptors to model community reorganization prior to and during 18 PEx. We identify two microbial dysbiosis regimes with opposing ecology and dynamics. Pathogen-governed PEx show hierarchical community reorganization and reduced diversity, whereas anaerobic bloom PEx display stochasticity and increased diversity. A simulation of antimicrobial treatment predicts better efficacy for hierarchically organized communities. This link between PEx, microbiome organization, and treatment success advances the development of personalized clinical management in CF and, potentially, other obstructive lung diseases.

Itaconic acid inhibits nontuberculous mycobacterial growth in pH dependent manner while 4-octyl-itaconic acid enhances THP-1 clearance of nontuberculous mycobacteria in vitro.

Increasingly prevalent, nontuberculous mycobacteria (NTM) infections affect approximately 20% of people with cystic fibrosis (CF). Previous studies of CF sputum identified lower levels of the host metabolite itaconate in those infected with NTM. Itaconate can inhibit the growth of M. tuberculosis (MTB) in vitro via the inhibition of the glyoxylate cycle enzyme (ICL), but its impact on NTM is unclear. To test itaconic acid's (IA) effect on NTM growth, laboratory and CF clinical strains of Mycobacterium abscessus and Mycobacterium avium were cultured in 7H9 minimal media supplemented with 1-10 mM of IA and short-chain fatty acids (SCFA). M. avium and M. abscessus grew when supplemented with SCFAs, whereas the addition of IA (≥ 10 mM) completely inhibited NTM growth. NTM supplemented with acetate or propionate and 5 mM IA displayed slower growth than NTM cultured with SCFA and ≤ 1 mM of IA. However, IA's inhibition of NTM was pH dependent; as similar and higher quantities (100 mM) of pH adjusted IA (pH 7) did not inhibit growth in vitro, while in an acidic minimal media (pH 6.1), 1 to 5 mM of non-pH adjusted IA inhibited growth. None of the examined isolates displayed the ability to utilize IA as a carbon source, and IA added to M. abscessus isocitrate lyase (ICL) decreased enzymatic activity. Lastly, the addition of cell-permeable 4-octyl itaconate (4-OI) to THP-1 cells enhanced NTM clearance, demonstrating a potential role for IA/itaconate in host defense against NTM infections.

Prevalence and Clinical Impact of Respiratory Viral Infections from the STOP2 Study of Cystic Fibrosis Pulmonary Exacerbations.

Rationale: Rates of viral respiratory infection (VRI) are similar in people with cystic fibrosis (CF) and the general population; however, the associations between VRI and CF pulmonary exacerbations (PEx) require further elucidation.Objectives: To determine VRI prevalence during CF PEx and evaluate associations between VRI, clinical presentation, and treatment response.Methods: The STOP2 (Standardized Treatment of Pulmonary Exacerbations II) study was a multicenter randomized trial to evaluate different durations of intravenous antibiotic therapy for PEx. In this ancillary study, participant sputum samples from up to three study visits were tested for respiratory viruses using multiplex polymerase chain reactions. Baselines and treatment-associated changes in mean lung function (percent predicted forced expiratory volume in 1 s), respiratory symptoms (Chronic Respiratory Infection Symptom Score), weight, and C-reactive protein were compared as a function of virus detection. Odds of PEx retreatment within 30 days and future PEx hazard were modeled by logistic and Cox proportional hazards regression, respectively.Results: A total of 1,254 sputum samples from 621 study participants were analyzed. One or more respiratory viruses were detected in sputum samples from 245 participants (39.5%). Virus-positive participants were more likely to be receiving CF transmembrane conductance regulator modulator therapy (45% vs. 34%) and/or chronic azithromycin therapy (54% vs. 44%) and more likely to have received treatment for nontuberculous Mycobacterium infection in the preceding 2 years (7% vs. 3%). At study visit 1, virus-positive participants were more symptomatic (mean Chronic Respiratory Infection Symptom Score, 53.8 vs. 51.1), had evidence of greater systemic inflammation (log10 C-reactive protein concentration, 1.32 log10 mg/L vs. 1.23 log10 mg/L), and had a greater drop in percent predicted forced expiratory volume in 1 second from the prior 6-month baseline (5.8 vs. 3.6). Virus positivity was associated with reduced risk of future PEx (hazard ratio, 0.82; 95% confidence interval, 0.69-0.99; P = 0.034) and longer median time to next PEx (255 d vs. 172 d; P = 0.021) compared with virus negativity.Conclusions: More than one-third of STOP2 participants treated for a PEx had a positive test result for a respiratory virus with more symptomatic initial presentation compared with virus-negative participants, but favorable long-term outcomes. More refined phenotyping of PEx, taking VRIs into account, may aid in optimizing personalized management of PEx.Clinical trial registered with www.clinicaltrials.gov (NCT02781610).

Microbial community organization designates distinct pulmonary exacerbation types and predicts treatment outcome in cystic fibrosis.

Polymicrobial infection of the airways is a hallmark of obstructive lung diseases such as cystic fibrosis (CF), non-CF bronchiectasis, and chronic obstructive pulmonary disease (COPD). Intermittent pulmonary exacerbations (PEx) in these conditions are associated with lung function decline and higher mortality rates. An understanding of the microbial underpinnings of PEx is challenged by high inter-patient variability in airway microbial community profiles. We analyzed 880 near-daily CF sputum samples and developed non-standard microbiome descriptors to model community reorganization prior and during 18 PEx. We identified two communal microbial regimes with opposing ecology and dynamics. Whereas pathogen-governed dysbiosis showed hierarchical community organization and reduced diversity, anaerobic bloom dysbiosis displayed stochasticity and increased diversity. Microbiome organization modulated the relevance of pathogens and a simulation of antimicrobial treatment predicted better efficacy for hierarchically organized microbiota. This causal link between PEx, microbiome organization, and treatment success advances the development of personalized dysbiosis management in CF and, potentially, other obstructive lung diseases.

Genomic Analyses of Longitudinal Mycobacterium abscessus Isolates in a Multicenter Cohort Reveal Parallel Signatures of In-Host Adaptation.

BACKGROUND: Nontuberculous mycobacteria (NTM) are ubiquitous in the environment and an increasingly frequent cause of opportunistic infections. Mycobacterium abscessus complex (MABC) is one of the major NTM lung pathogens that disproportionately colonize and infect the lungs of individuals with cystic fibrosis (CF). MABC infection can persist for years, and antimicrobial treatment is frequently ineffective. METHODS: We sequenced the genomes of 175 isolates longitudinally collected from 30 patients with MABC lung infection. We contextualized our cohort amidst the broader MABC phylogeny and investigated genes undergoing parallel adaptation across patients. Finally, we tested the phenotypic consequences of parallel mutations by conducting antimicrobial resistance and mercury-resistance assays. RESULTS: We identified highly related isolate pairs across hospital centers with low likelihood of transmission. We further annotated nonrandom parallel mutations in 22 genes and demonstrated altered macrolide susceptibility co-occurring with a nonsynonymous whiB1 mutation. Finally, we highlighted a 23-kb mercury-resistance plasmid whose loss during chronic infection conferred phenotypic susceptibility to organic and nonorganic mercury compounds. CONCLUSIONS: We characterized parallel genomic processes through which MABC is adapting to promote survival within the host. The within-lineage polymorphisms we observed have phenotypic effects, potentially benefiting fitness in the host at the putative detriment of environmental survival.

Quantifying variation in home spirometry in people with cystic fibrosis during baseline health, and associations with clinical outcomes.

BACKGROUND: Home spirometry is increasingly used to monitor lung function in people with cystic fibrosis (pwCF). Although decreases in lung function in the setting of increased respiratory symptoms are consistent with a pulmonary exacerbation (PEx), the interpretation of home spirometry during asymptomatic periods of baseline health is unclear. The aims of this study were to determine the variation in home spirometry in pwCF during asymptomatic periods of baseline health and to identify associations between this variation and PEx. METHODS: Near-daily home spirometry measurements were obtained from a cohort of pwCF enrolled in a long-term study of the airway microbiome. Associations between the degree of variation in home spirometry and the time to next PEx were evaluated. RESULTS: Thirteen subjects (mean age of 29 years and mean percent predicted forced expiratory volume in one second [ppFEV1] of 60) provided a median of 204 spirometry readings taken during 40 periods of baseline health. The mean week-to-week within-subject level of variation in ppFEV1 was 15.2 ± 6.2%. The degree of variation in ppFEV1 during baseline health was not associated with time to PEx. CONCLUSIONS: Variation in ppFEV1 measured with near-daily home spirometry in pwCF during periods of baseline health exceeded the variation in ppFEV1 expected in clinic spirometry (based on ATS guidelines). The degree of variation in ppFEV1 during baseline health was not associated with time to PEx. These data are relevant for guiding interpretation of home spirometry.

Airway bacterial community composition in persons with advanced cystic fibrosis lung disease.

BACKGROUND: The progression of lung disease in people with cystic fibrosis (pwCF) has been associated with a decrease in the diversity of airway bacterial communities. How often low diversity communities occur in advanced CF lung disease and how they may be associated with clinical outcomes is not clear, however. METHODS: We sequenced a region of the bacterial 16S ribosomal RNA gene to characterize bacterial communities in sputum from 190 pwCF with advanced lung disease (FEV1≤40% predicted), with particular attention to the prevalence and relative abundance of dominant genera. We evaluated relationships between community diversity and clinical outcomes. RESULTS: Although most of the 190 pwCF with advanced lung disease had airway bacterial communities characterized by low diversity with a dominant genus, a considerable minority (40%) did not. The absence of a dominant genus, presence of methicillin-susceptible Staphylococcus aureus, and greater bacterial richness positively correlated with lung function. Higher relative abundance of the dominant genus and greater antimicrobial use negatively correlated with lung function. PwCF with a low diversity community and dominant genus had reduced lung transplant-free survival compared to those without (median survival of 1.6 vs 2.9 years). CONCLUSIONS: A considerable proportion of pwCF with advanced lung disease do not have airway bacterial communities characterized by low diversity and a dominant genus and these individuals had better survival. An understanding of the antecedents of low diversity airway communities- and the impact these may have on lung disease trajectory - may provide avenues for improved management strategies.

The Impact of Highly Effective Modulator Therapy on Cystic Fibrosis Microbiology and Inflammation.

Highly effective cystic fibrosis (CF) transmembrane conductance regulator (CFTR) modulator therapy (HEMT) corrects the underlying molecular defect causing CF disease. HEMT decreases symptom burden and improves clinical metrics and quality of life for most people with CF (PwCF) and eligible cftr mutations. Improvements in measures of pulmonary health suggest that restoration of function of defective CFTR anion channels by HEMT not only enhances airway mucociliary clearance, but also reduces chronic pulmonary infection and inflammation. This article reviews the evidence for how HEMT influences the dynamic and interdependent processes of infection and inflammation in the CF airway, and what questions remain unanswered.

The Elusive Role of Airway Infection in Cystic Fibrosis Exacerbation.

Cystic fibrosis (CF) pulmonary exacerbations (PEx) are clinical events that commonly result in increased treatment burden, decreased quality of life, and accelerated lung disease progression. CF PEx have historically been approached as though dealing with acute infections, and antibiotic treatments have been associated with improved outcomes. In this review, we discuss data supporting a causal role of CF airway infection in PEx as well studies that highlight our knowledge gaps in regard to PEx definitions, pathophysiology, and optimal treatment approaches. In the era of highly effective cystic fibrosis transmembrane conductance regulator modulator therapy, and the continually increasing health and longevity of persons with CF, a better understanding of PEx and further optimization of PEx antibiotic treatment approaches are needed.

Changes in airway bacterial communities occur soon after initiation of antibiotic treatment of pulmonary exacerbations in cystic fibrosis.

Chronic polymicrobial airway infections are a hallmark of cystic fibrosis (CF) lung disease. Antibiotic therapy is a primary treatment of CF pulmonary exacerbations (PEx); however, the impact of episodic antibiotic treatment on airway bacterial communities has not been well described. We analyzed sputum samples from adults with CF obtained immediately before and during antibiotic treatment of PEx. Sequencing of the V4 region of the bacterial 16S ribosomal RNA gene was used to assess changes in bacterial community structure during antibiotic treatment. The peak impact of antibiotic treatment was observed by day four or five of treatment. These findings advance our understanding of bacterial community dynamics during antibiotic treatment of PEx and complement recent and ongoing studies evaluating the optimal duration of antibiotic therapy for PEx.

Sputum Metabolites Associated with Nontuberculous Mycobacterial Infection in Cystic Fibrosis.

Nontuberculous mycobacterial (NTM) pulmonary infections in people with cystic fibrosis (CF) are associated with significant morbidity and mortality and are increasing in prevalence. Host risk factors for NTM infection in CF are largely unknown. We hypothesize that the airway microbiota represents a host risk factor for NTM infection. In this study, 69 sputum samples were collected from 59 people with CF; 42 samples from 32 subjects with NTM infection (14 samples collected before incident NTM infection and 28 samples collected following incident NTM infection) were compared to 27 samples from 27 subjects without NTM infection. Sputum samples were analyzed with 16S rRNA gene sequencing and metabolomics. A supervised classification and correlation analysis framework (sparse partial least-squares discriminant analysis [sPLS-DA]) was used to identify correlations between the microbial and metabolomic profiles of the NTM cases compared to the NTM-negative controls. Several metabolites significantly differed in the NTM cases compared to controls, including decreased levels of tryptophan-associated and branched-chain amino acid metabolites, while compounds involved in phospholipid metabolism displayed increased levels. When the metabolome and microbiome data were integrated by sPLS-DA, the models and component ordinations showed separation between the NTM and control samples. While this study could not determine if the observed differences in sputum metabolites between the cohorts reflect metabolic changes that occurred as a result of the NTM infection or metabolic features that contributed to NTM acquisition, it is hypothesis generating for future work to investigate host and bacterial community factors that may contribute to NTM infection risk in CF. IMPORTANCE Host risk factors for nontuberculous mycobacterial (NTM) infection in people with cystic fibrosis (CF) are largely unclear. The goal of this study was to help identify potential host and bacterial community risk factors for NTM infection in people with CF, using microbiome and metabolome data from CF sputum samples. The data obtained in this study identified several metabolic profile differences in sputum associated with NTM infection in CF, including 2-methylcitrate/homocitrate and selected ceramides. These findings represent potential risk factors and therapeutic targets for preventing and/or treating NTM infections in people with CF.

Tenets of a holistic approach to drinking water-associated pathogen research, management, and communication.

In recent years, drinking water-associated pathogens that can cause infections in immunocompromised or otherwise susceptible individuals (henceforth referred to as DWPI), sometimes referred to as opportunistic pathogens or opportunistic premise plumbing pathogens, have received considerable attention. DWPI research has largely been conducted by experts focusing on specific microorganisms or within silos of expertise. The resulting mitigation approaches optimized for a single microorganism may have unintended consequences and trade-offs for other DWPI or other interests (e.g., energy costs and conservation). For example, the ecological and epidemiological issues characteristic of Legionella pneumophila diverge from those relevant for Mycobacterium avium and other nontuberculous mycobacteria. Recent advances in understanding DWPI as part of a complex microbial ecosystem inhabiting drinking water systems continues to reveal additional challenges: namely, how can all microorganisms of concern be managed simultaneously? In order to protect public health, we must take a more holistic approach in all aspects of the field, including basic research, monitoring methods, risk-based mitigation techniques, and policy. A holistic approach will (i) target multiple microorganisms simultaneously, (ii) involve experts across several disciplines, and (iii) communicate results across disciplines and more broadly, proactively addressing source water-to-customer system management.

Testing the effects of combining azithromycin with inhaled tobramycin for P. aeruginosa in cystic fibrosis: a randomised, controlled clinical trial.

RATIONALE: Inhaled tobramycin and oral azithromycin are common chronic therapies in people with cystic fibrosis and Pseudomonas aeruginosa airway infection. Some studies have shown that azithromycin can reduce the ability of tobramycin to kill P. aeruginosa. This trial was done to test the effects of combining azithromycin with inhaled tobramycin on clinical and microbiological outcomes in people already using inhaled tobramycin. We theorised that those randomised to placebo (no azithromycin) would have greater improvement in forced expiratory volume in one second (FEV1) and greater reduction in P. aeruginosa sputum in response to tobramycin. METHODS: A 6-week prospective, randomised, placebo-controlled, double-blind trial testing oral azithromycin versus placebo combined with clinically prescribed inhaled tobramycin in individuals with cystic fibrosis and P. aeruginosa airway infection. RESULTS: Over a 6-week period, including 4 weeks of inhaled tobramycin, the relative change in FEV1 did not statistically significantly differ between groups (azithromycin (n=56) minus placebo (n=52) difference: 3.44%; 95% CI: -0.48 to 7.35; p=0.085). Differences in secondary clinical outcomes, including patient-reported symptom scores, weight and need for additional antibiotics, did not significantly differ. Among the 29 azithromycin and 35 placebo participants providing paired sputum samples, the 6-week change in P. aeruginosa density differed in favour of the placebo group (difference: 0.75 log10 CFU/mL; 95% CI: 0.03 to 1.47; p=0.043). CONCLUSIONS: Despite having greater reduction in P. aeruginosa density in participants able to provide sputum samples, participants randomised to placebo with inhaled tobramycin did not experience significantly greater improvements in lung function or other clinical outcomes compared with those randomised to azithromycin with tobramycin.

Comparative Genomics of Mycobacterium avium Complex Reveals Signatures of Environment-Specific Adaptation and Community Acquisition.

Nontuberculous mycobacteria, including those in the Mycobacterium avium complex (MAC), constitute an increasingly urgent threat to global public health. Ubiquitous in soil and water worldwide, MAC members cause a diverse array of infections in humans and animals that are often multidrug resistant, intractable, and deadly. MAC lung disease is of particular concern and is now more prevalent than tuberculosis in many countries, including the United States. Although the clinical importance of these microorganisms continues to expand, our understanding of their genomic diversity is limited, hampering basic and translational studies alike. Here, we leveraged a unique collection of genomes to characterize MAC population structure, gene content, and within-host strain dynamics in unprecedented detail. We found that different MAC species encode distinct suites of biomedically relevant genes, including antibiotic resistance genes and virulence factors, which may influence their distinct clinical manifestations. We observed that M. avium isolates from different sources-human pulmonary infections, human disseminated infections, animals, and natural environments-are readily distinguished by their core and accessory genomes, by their patterns of horizontal gene transfer, and by numerous specific genes, including virulence factors. We identified highly similar MAC strains from distinct patients within and across two geographically distinct clinical cohorts, providing important insights into the reservoirs which seed community acquisition. We also discovered a novel MAC genomospecies in one of these cohorts. Collectively, our results provide key genomic context for these emerging pathogens and will facilitate future exploration of MAC ecology, evolution, and pathogenesis. IMPORTANCE Members of the Mycobacterium avium complex (MAC), a group of mycobacteria encompassing M. avium and its closest relatives, are omnipresent in natural environments and emerging pathogens of humans and animals. MAC infections are difficult to treat, sometimes fatal, and increasingly common. Here, we used comparative genomics to illuminate key aspects of MAC biology. We found that different MAC species and M. avium isolates from different sources encode distinct suites of clinically relevant genes, including those for virulence and antibiotic resistance. We identified highly similar MAC strains in patients from different states and decades, suggesting community acquisition from dispersed and stable reservoirs, and we discovered a novel MAC species. Our work provides valuable insight into the genomic features underlying these versatile pathogens.

Aminoglycoside dosing and monitoring for Pseudomonas aeruginosa during acute pulmonary exacerbations in cystic fibrosis.

Aminoglycosides are commonly used for the treatment of Pseudomonas aeruginosa (PsA) in the setting of acute pulmonary exacerbations (PEx) in pediatric patients with cystic fibrosis (CF). There are controversies and practice differences between institutions related to aminoglycoside dosing and monitoring strategies. The purpose of this review article is to summarize the currently available literature and identify gaps in the literature related to pharmacokinetic parameter goals, aminoglycoside dosing strategies, and methods for monitoring serum aminoglycoside concentrations for treatment of PsA in CF PEx, and throughout will discuss anticipated changes with the increasing availability of highly effective CF transmembrane conductance regulator modulators. This review focuses on tobramycin, as it is the most commonly used aminoglycoside in CF PEx, and will briefly discuss special circumstances surrounding use of amikacin and gentamicin.