scholarly journals Multi-Omics Study of Keystone Species in a Cystic Fibrosis Microbiome

2021 ◽  
Vol 22 (21) ◽  
pp. 12050
Author(s):  
Cynthia B. Silveira ◽  
Ana G. Cobián-Güemes ◽  
Carla Uranga ◽  
Jonathon L. Baker ◽  
Anna Edlund ◽  
...  
Keyword(s):  
Cystic Fibrosis ◽  
Lung Function ◽  
Sequence Data ◽  
Anaerobic Bacteria ◽  
Keystone Species ◽  
Clinical Diagnostics ◽  
Epifluorescence Microscopy ◽  
Fermentation Products ◽  
Degrading Bacteria

Ecological networking and in vitro studies predict that anaerobic, mucus-degrading bacteria are keystone species in cystic fibrosis (CF) microbiomes. The metabolic byproducts from these bacteria facilitate the colonization and growth of CF pathogens like Pseudomonas aeruginosa. Here, a multi-omics study informed the control of putative anaerobic keystone species during a transition in antibiotic therapy of a CF patient. A quantitative metagenomics approach combining sequence data with epifluorescence microscopy showed that during periods of rapid lung function loss, the patient’s lung microbiome was dominated by the anaerobic, mucus-degrading bacteria belonging to Streptococcus, Veillonella, and Prevotella genera. Untargeted metabolomics and community cultures identified high rates of fermentation in these sputa, with the accumulation of lactic acid, citric acid, and acetic acid. P. aeruginosa utilized these fermentation products for growth, as indicated by quantitative transcriptomics data. Transcription levels of P. aeruginosa genes for the utilization of fermentation products were proportional to the abundance of anaerobic bacteria. Clindamycin therapy targeting Gram-positive anaerobes rapidly suppressed anaerobic bacteria and the accumulation of fermentation products. Clindamycin also lowered the abundance and transcription of P. aeruginosa, even though this patient’s strain was resistant to this antibiotic. The treatment stabilized the patient’s lung function and improved respiratory health for two months, lengthening by a factor of four the between-hospitalization time for this patient. Killing anaerobes indirectly limited the growth of P. aeruginosa by disrupting the cross-feeding of fermentation products. This case study supports the hypothesis that facultative anaerobes operated as keystone species in this CF microbiome. Personalized multi-omics may become a viable approach for routine clinical diagnostics in the future, providing critical information to inform treatment decisions.

scholarly journals Multi-omics study of keystone species in the cystic fibrosis lung microbiome

2020 ◽  
Author(s):  
Cynthia Silveira ◽  
Ana Georgina Cobian-Guemes ◽  
Carla Uranga ◽  
Jonathon Baker ◽  
Anna Edlund ◽  
...  
Keyword(s):  
Cystic Fibrosis ◽  
Microbial Community ◽  
Lung Function ◽  
Respiratory Health ◽  
Keystone Species ◽  
Fermentation Products ◽  
Community Function ◽  
Degrading Bacteria

Abstract Background: Ecological networking and in vitro studies have predicted that anaerobic, mucus-degrading bacteria are keystone species in cystic fibrosis (CF) microbiomes by sustaining the growth of canonical CF pathogens. Here, a multi-omics approach was deployed to test this hypothesis in vivo and in real time during a transition in antibiotic therapy of a CF patient with a hypervariable lung function phenotype . Results: Quantitative meta-omics and community culturing demonstrated that the use of a non-traditional clindamycin therapy targeting gram-positives and gram-negative anaerobes re-structured the entire CF microbial community. During rapid lung function loss, when the patient was off antibiotics, the microbial community was dominated by anaerobic mucus-degrading Streptococcus sp., Veilonella sp., and Prevotella sp. that produced fermentation gas and led to the accumulation of fermentation products in sputum. The rise of anaerobes was followed within 6 days by an increase in Pseudomonas aeruginosa transcripts encoding the acquisition of fermentation products from anaerobes and the production of virulence factors. The initiation of clindamycin treatment reduced the fermentation and the abundance of anaerobes. Clindamycin also lowered the abundance and transcription of P. aeruginosa, which is resistant to this antibiotic. The treatment stabilized the patient’s lung function and improved respiratory health for two months, lengthening by a factor of four the between-hospitalization time for this patient. Conclusions: The results presented here show that killing anaerobes, the weakest link in the community in terms of antibiotic resistance, effectively limited the growth of classic CF pathogen by disrupting community cross-feeding. The role that anaerobic, mucus-degrading bacteria played in structuring the CF microbiome corroborates in vivo their position as keystone bacteria, with high impact on community function despite lower relative abundances.

scholarly journals Prevalence and impact of oprD mutations in Pseudomonas aeruginosa strains in cystic fibrosis

2019 ◽  
Author(s):  
Laura J. Sherrard ◽  
Bryan A. Wee ◽  
Christine Duplancic ◽  
Kay A. Ramsay ◽  
Keyur A. Dave ◽  
...  
Keyword(s):  
Cystic Fibrosis ◽  
Pseudomonas Aeruginosa ◽  
Lung Function ◽  
Hazard Rate ◽  
Carbapenem Resistance ◽  
Adverse Outcomes ◽  
Nonsense Mutations ◽  
Allelic Variants ◽  
Novel Variants

ABSTRACTDefective OprD porins contribute to carbapenem resistance and may be important in Pseudomonas aeruginosa adaptation to cystic fibrosis airways. It is unclear whether oprD mutations are fixed in populations of shared strains that are transmitted between patients or whether novel variants arise during infection. We investigated oprD sequences and antimicrobial resistance of two common Australian shared strains, constructed P. aeruginosa mutants with the most common oprD allelic variants and compared characteristics between patients with or without evidence of infection with strains harbouring these variants. Our data show that three independently acquired nonsense mutations arising from a 1-base pair substitution are fixed in strain sub-lineages. These nonsense mutations are likely to contribute to reduced carbapenem susceptibility in the sub-lineages without compromising in vitro fitness. Not only was lung function worse among patients infected with strains harbouring the nonsense mutations than those without, but they also had an increased hazard rate of lung transplantation/death. Our findings further highlight that understanding adaptive changes may help to distinguish patients with greater adverse outcomes despite infection with the same strain.

scholarly journals Disruption of Cross-Feeding Inhibits Pathogen Growth in the Sputa of Patients with Cystic Fibrosis

mSphere ◽  
2020 ◽  
Vol 5 (2) ◽  
Author(s):  
Jeffrey M. Flynn ◽  
Lydia C. Cameron ◽  
Talia D. Wiggen ◽  
Jordan M. Dunitz ◽  
William R. Harcombe ◽  
...  
Keyword(s):  
Cystic Fibrosis ◽  
Pseudomonas Aeruginosa ◽  
Antibiotic Susceptibility ◽  
Susceptibility Testing ◽  
Anaerobic Bacteria ◽  
Content Type ◽  
Growth Environment ◽  
Polymicrobial Infections

ABSTRACT A critical limitation in the management of chronic polymicrobial infections is the lack of correlation between antibiotic susceptibility testing (AST) and patient responses to therapy. Underlying this disconnect is our inability to accurately recapitulate the in vivo environment and complex polymicrobial communities in vitro. However, emerging evidence suggests that, if modeled and tested accurately, interspecies relationships can be exploited by conventional antibiotics predicted to be ineffective by standard AST. As an example, under conditions where Pseudomonas aeruginosa relies on cocolonizing organisms for nutrients (i.e., cross-feeding), multidrug-resistant P. aeruginosa may be indirectly targeted by inhibiting the growth of its metabolic partners. While this has been shown in vitro using synthetic bacterial communities, the efficacy of a “weakest-link” approach to controlling host-associated polymicrobial infections has not yet been demonstrated. To test whether cross-feeding inhibition can be leveraged in clinically relevant contexts, we collected sputa from cystic fibrosis (CF) subjects and used enrichment culturing to isolate both P. aeruginosa and anaerobic bacteria from each sample. Predictably, both subpopulations showed various antibiotic susceptibilities when grown independently. However, when P. aeruginosa was cultured and treated under cooperative conditions in which it was dependent on anaerobic bacteria for nutrients, the growth of both the pathogen and the anaerobe was constrained despite their intrinsic antibiotic resistance profiles. These data demonstrate that the control of complex polymicrobial infections may be achieved by exploiting obligate or facultative interspecies relationships. Toward this end, in vitro susceptibility testing should evolve to more accurately reflect in vivo growth environments and microbial interactions found within them. IMPORTANCE Antibiotic efficacy achieved in vitro correlates poorly with clinical outcomes after treatment of chronic polymicrobial diseases; if a pathogen demonstrates susceptibility to a given antibiotic in the lab, that compound is often ineffective when administered clinically. Conversely, if a pathogen is resistant in vitro, patient treatment with that same compound can elicit a positive response. This discordance suggests that the in vivo growth environment impacts pathogen antibiotic susceptibility. Indeed, here we demonstrate that interspecies relationships among microbiotas in the sputa of cystic fibrosis patients can be targeted to indirectly inhibit the growth of Pseudomonas aeruginosa. The therapeutic implication is that control of chronic lung infections may be achieved by exploiting obligate or facultative relationships among airway bacterial community members. This strategy is particularly relevant for pathogens harboring intrinsic multidrug resistance and is broadly applicable to chronic polymicrobial airway, wound, and intra-abdominal infections.

scholarly journals Pseudomonas aeruginosa sabotages the generation of host proresolving lipid mediators

2016 ◽  
Vol 114 (1) ◽  
pp. 136-141 ◽  
Author(s):  
Becca A. Flitter ◽  
Kelli L. Hvorecny ◽  
Emiko Ono ◽  
Taylor Eddens ◽  
Jun Yang ◽  
...  
Keyword(s):  
Cystic Fibrosis ◽  
Pseudomonas Aeruginosa ◽  
Lung Function ◽  
Epoxide Hydrolase ◽  
Pulmonary Inflammation ◽  
Critical Role ◽  
Lavage Fluid ◽  
Lipid Mediator ◽  
Epoxyeicosatrienoic Acid

Recurrent Pseudomonas aeruginosa infections coupled with robust, damaging neutrophilic inflammation characterize the chronic lung disease cystic fibrosis (CF). The proresolving lipid mediator, 15-epi lipoxin A4 (15-epi LXA4), plays a critical role in limiting neutrophil activation and tissue inflammation, thus promoting the return to tissue homeostasis. Here, we show that a secreted P. aeruginosa epoxide hydrolase, cystic fibrosis transmembrane conductance regulator inhibitory factor (Cif), can disrupt 15-epi LXA4 transcellular biosynthesis and function. In the airway, 15-epi LXA4 production is stimulated by the epithelial-derived eicosanoid 14,15-epoxyeicosatrienoic acid (14,15-EET). Cif sabotages the production of 15-epi LXA4 by rapidly hydrolyzing 14,15-EET into its cognate diol, eliminating a proresolving signal that potently suppresses IL-8–driven neutrophil transepithelial migration in vitro. Retrospective analyses of samples from patients with CF supported the translational relevance of these preclinical findings. Elevated levels of Cif in bronchoalveolar lavage fluid were correlated with lower levels of 15-epi LXA4, increased IL-8 concentrations, and impaired lung function. Together, these findings provide structural, biochemical, and immunological evidence that the bacterial epoxide hydrolase Cif disrupts resolution pathways during bacterial lung infections. The data also suggest that Cif contributes to sustained pulmonary inflammation and associated loss of lung function in patients with CF.

scholarly journals Oxygen Affects Gut Bacterial Colonization and Metabolic Activities in a Gnotobiotic Cockroach Model

2015 ◽  
Vol 82 (4) ◽  
pp. 1080-1089 ◽  
Author(s):  
Dorothee Tegtmeier ◽  
Claire L. Thompson ◽  
Christine Schauer ◽  
Andreas Brune
Keyword(s):  
Bacterial Colonization ◽  
Anaerobic Bacteria ◽  
Close Relative ◽  
Bacterial Strains ◽  
Fermentation Products ◽  
Content Type ◽  
Facultatively Anaerobic ◽  
Pure Cultures ◽  
Metabolic Activities

ABSTRACTThe gut microbiota of termites and cockroaches represents complex metabolic networks of many diverse microbial populations. The distinct microenvironmental conditions within the gut and possible interactions among the microorganisms make it essential to investigate how far the metabolic properties of pure cultures reflect their activities in their natural environment. We established the cockroachShelfordella lateralisas a gnotobiotic model and inoculated germfree nymphs with two bacterial strains isolated from the guts of conventional cockroaches. Fluorescence microscopy revealed that both strains specifically colonized the germfree hindgut. In diassociated cockroaches, the facultatively anaerobic strain EbSL (a new species ofEnterobacteriaceae) always outnumbered the obligately anaerobic strain FuSL (a close relative ofFusobacterium varium), irrespective of the sequence of inoculation, which showed that precolonization by facultatively anaerobic bacteria does not necessarily favor colonization by obligate anaerobes. Comparison of the fermentation products of the cultures formedin vitrowith those accumulatedin situindicated that the gut environment strongly affected the metabolic activities of both strains. The pure cultures formed the typical products of mixed-acid or butyrate fermentation, whereas the guts of gnotobiotic cockroaches accumulated mostly lactate and acetate. Similar shifts toward more-oxidized products were observed when the pure cultures were exposed to oxygen, which corroborated the strong effects of oxygen on the metabolic fluxes previously observed in termite guts. Oxygen microsensor profiles of the guts of germfree, gnotobiotic, and conventional cockroaches indicated that both gut tissue and microbiota contribute to oxygen consumption and suggest that the oxygen status influences the colonization success.

scholarly journals Localization of Burkholderia cepacia Complex Bacteria in Cystic Fibrosis Lungs and Interactions with Pseudomonas aeruginosa in Hypoxic Mucus

2014 ◽  
Vol 82 (11) ◽  
pp. 4729-4745 ◽  
Author(s):  
Ute Schwab ◽  
Lubna H. Abdullah ◽  
Olivia S. Perlmutt ◽  
Daniel Albert ◽  
C. William Davis ◽  
...  
Keyword(s):  
Cystic Fibrosis ◽  
Pseudomonas Aeruginosa ◽  
Burkholderia Cepacia ◽  
Single Cells ◽  
Burkholderia Cenocepacia ◽  
Burkholderia Cepacia Complex ◽  
Test Medium ◽  
Fermentation Products ◽  
Content Type

ABSTRACTThe localization ofBurkholderia cepaciacomplex (Bcc) bacteria in cystic fibrosis (CF) lungs, alone or during coinfection withPseudomonas aeruginosa, is poorly understood. We performed immunohistochemistry for Bcc andP. aeruginosabacteria on 21 coinfected or singly infected CF lungs obtained at transplantation or autopsy. Parallelin vitroexperiments examined the growth of two Bcc species,Burkholderia cenocepaciaandBurkholderia multivorans, in environments similar to those occupied byP. aeruginosain the CF lung. Bcc bacteria were predominantly identified in the CF lung as single cells or small clusters within phagocytes and mucus but not as “biofilm-like structures.” In contrast,P. aeruginosawas identified in biofilm-like masses, but densities appeared to be reduced during coinfection with Bcc bacteria. Based on chemical analyses of CF and non-CF respiratory secretions, a test medium was defined to study Bcc growth and interactions withP. aeruginosain an environment mimicking the CF lung. When test medium was supplemented with alternative electron acceptors under anaerobic conditions,B. cenocepaciaandB. multivoransused fermentation rather than anaerobic respiration to gain energy, consistent with the identification of fermentation products by high-performance liquid chromatography (HPLC). Both Bcc species also expressed mucinases that produced carbon sources from mucins for growth. In the presence ofP. aeruginosain vitro, both Bcc species grew anaerobically but not aerobically. We propose that Bcc bacteria (i) invade aP. aeruginosa-infected CF lung when the airway lumen is anaerobic, (ii) inhibitP. aeruginosabiofilm-like growth, and (iii) expand the host bacterial niche from mucus to also include macrophages.

scholarly journals Pseudomonas aeruginosaCan Inhibit Growth of Streptococcal Species via Siderophore Production

2018 ◽  
Author(s):  
Jessie E. Scott ◽  
Kewei Li ◽  
Laura M. Filkins ◽  
Bin Zhu ◽  
Sherry L. Kuchma ◽  
...  
Keyword(s):  
Cystic Fibrosis ◽  
Lung Function ◽  
Response To Treatment ◽  
Oral Streptococci ◽  
Interspecies Interactions ◽  
Viridans Streptococci ◽  
Lung Function Decline ◽  
Streptococcus Milleri ◽  
Polymicrobial Infections

AbstractCystic Fibrosis (CF) is a genetic disease that causes patients to accumulate thick, dehydrated mucus in the lung and develop chronic, polymicrobial infections due to reduced mucociliary clearance. These chronic polymicrobial infections and subsequent decline in lung function are significant factors in the morbidity and mortality of CF.Pseudomonas aeruginosaandStreptococcusspp. are among the most prevalent organisms in the CF lung; the presence ofP. aeruginosacorrelates with lung function decline and theStreptococcus millerigroup (SMG), a subgroup of the viridans streptococci, is associated with exacerbations in patients with CF. Here we characterize the interspecies interactions that occur between these two genera. We demonstrated that multipleP. aeruginosalaboratory strains and clinical CF isolates promote the growth of multiple SMG strains and oral streptococci in anin vitrococulture system. We investigated the mechanism by whichP. aeruginosaenhances growth of streptococci by screening for mutants ofP. aeruginosaPA14 unable to enhanceStreptococcusgrowth, and we identified theP. aeruginosa pqsL::TnMmutant, which failed to promote growth ofS. constellatusandS. sanguinis. Characterization of theP. aeruginosaΔpqsLmutant revealed that this strain cannot promoteStreptococcusgrowth. Our genetic data and growth studies support a model whereby theP. aeruginosaΔpqsLmutant overproduces siderophores, and thus likely outcompetesStreptococcus sanguinisfor limited iron. We propose a model whereby competition for iron represents one important means of interaction betweenP. aeruginosaandStreptococcusspp.ImportanceCystic fibrosis (CF) lung infections are increasingly recognized for their polymicrobial nature. These polymicrobial infections may alter the biology of the organisms involved in CF-related infections, leading to changes in growth, virulence and/or antibiotic tolerance, and could thereby affect patient health and response to treatment. In this study, we demonstrate interactions betweenP. aeruginosaand streptococci using a coculture model, and show that one interaction between these microbes is likely competition for iron. Thus, these data indicate that one CF pathogen may influence the growth of another and add to our limited knowledge of polymicrobial interactions in the CF airway.

Consequences of Biofilm and Sessile Growth in the Large Intestine

1997 ◽  
Vol 11 (1) ◽  
pp. 59-68 ◽  
Author(s):  
S. Macfarlane ◽  
A.J. McBain ◽  
G.T. Macfarlane
Keyword(s):  
Particulate Matter ◽  
Gut Microbiota ◽  
Product Formation ◽  
Complex Environment ◽  
Short Term ◽  
Fermentation Products ◽  
Degrading Bacteria ◽  
Food Particles ◽  
Different Strains

The human colonic ecosystem is an extremely complex environment comprised of several hundred different strains of bacteria. Studies were undertaken to determine whether these organisms formed metabolic or genotypically distinct assemblages in the gut microbiota in relation to polysaccharide fermentation. Measurements of depolymerizing enzymes (4 polysac-charidases, 6 glycosidases) showed that specific amylase and pectinase activities were comparable in bacteria desorbed from the surfaces of food particles and in non-particulate organisms. However, xylanase, β-xylosidase, arabinogalac-tanase, a-arabinofuranosidase, and β-galacturonidase activities were always significantly greater in particulate bacteria. Short-term in vitro fermentations with both groups of bacteria showed marked differences in relative rates of starch, arabinogalactan, and mucin metabolism, while rates of fermentation product formation with pectin and xylan were broadly comparable. Significant differences were observed with respect to formation of individual fermentation products, especially when mucin or pectin were substrates, where particulate bacteria produced proportionally higher amounts of acetate. Bacteriological studies showed that communities of polymer-degrading bacteria and other groups of intestinal anaerobes growing on particulate matter were essentially similar to those occurring elsewhere in the gut lumen, at genus and species levels. In vitro colonization experiments demonstrated that a variety of polysaccharide-fermenting bifidobacteria and bacteroides-together with other cross-feeding organisms such as peptostreptococci, fusobacteria, and coliforms-rapidly attached to particulate intestinal materials.

scholarly journals An observational study of anaerobic bacteria in cystic fibrosis lung using culture dependant and independent approaches

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Claudie Lamoureux ◽  
Charles-Antoine Guilloux ◽  
Clémence Beauruelle ◽  
Stéphanie Gouriou ◽  
Sophie Ramel ◽  
...  
Keyword(s):  
Cystic Fibrosis ◽  
Lung Function ◽  
Observational Study ◽  
Bacterial Species ◽  
Anaerobic Bacteria ◽  
Forced Expiratory Volume ◽  
Molecular Approach ◽  
Anaerobic Conditions ◽  
Potential Impact ◽  
Pcr Targeting

AbstractStrict anaerobes are undeniably important residents of the cystic fibrosis (CF) lung but are still unknowns. The main objectives of this study were to describe anaerobic bacteria diversity in CF airway microbiota and to evaluate the association with lung function. An observational study was conducted during eight months. A hundred and one patients were enrolled in the study, and 150 sputum samples were collected using a sterile sample kit designed to preserve anaerobic conditions. An extended-culture approach on 112 sputa and a molecular approach (quantitative PCR targeting three of the main anaerobic genera in CF lung: Prevotella, Veillonella, and Fusobacterium) on 141 sputa were developed. On culture, 91.1% of sputa were positive for at least one anaerobic bacterial species, with an average of six anaerobic species detected per sputum. Thirty-one anaerobic genera and 69 species were found, which is the largest anaerobe diversity ever reported in CF lungs. Better lung function (defined as Forced Expiratory Volume in one second > 70%) was significantly associated with higher quantification of Veillonella. These results raise the question of the potential impact of anaerobes on lung function.

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