scholarly journals Refining the Application of Microbial Lipids as Tracers of Staphylococcus aureus Growth Rates in Cystic Fibrosis Sputum

2018 ◽  
Author(s):  
Cajetan Neubauer ◽  
Ajay S. Kasi ◽  
Nora Grahl ◽  
Alex L. Sessions ◽  
Sebastian H. Kopf ◽  
...  
Keyword(s):  
Cystic Fibrosis ◽  
Fatty Acids ◽  
Fatty Acid ◽  
Microbial Community ◽  
Stable Isotope ◽  
Community Composition ◽  
Growth Rates ◽  
Microbial Community Composition ◽  
Stable Isotope Probing ◽  
Lung Infections

ABSTRACTChronic lung infections in cystic fibrosis (CF) could be treated more effectively if the effect of antimicrobials on pathogens in situ were known. Here, we compared changes in the microbial community composition and pathogen growth rates in longitudinal studies of CF patients undergoing intravenous antibiotic administration during pulmonary exacerbations. Microbial community composition was measured by NanoString DNA analysis and growth rates were obtained by incubating CF sputum with heavy water and tracing incorporation of deuterium into two different anteiso fatty acids (a-C15:0 and a-C17:0) using gas chromatography–mass spectrometry (GC/MS). Prior to this study, both lipids were thought to be specific for Staphylococcaceae and hence their isotopic enrichment was interpreted as a growth proxy for S. aureus. Our experiments revealed, however, that Prevotella is also a relevant microbial producer of a-C17:0 fatty acid in some CF patients, thus deuterium incorporation into these lipids is better interpreted as a more general pathogen growth rate proxy. Even accounting for a small non-microbial background source detected in some patient samples, a-C15:0 fatty acid still appear to be a relatively robust proxy for CF pathogens, revealing a median generation time of ~1.5 days, similar to prior observations. Contrary to our expectations, pathogen growth rates remained relatively stable throughout exacerbation treatment. We suggest two best practices for application of stable isotope probing in CF sputum: (1) parallel determination of microbial community composition in CF sputum using culture-independent tools, and (2) analysis of samples with a minimum a-C15:0 concentration of 0.1 weight percent of saturated fatty acids.IMPORTANCEIn chronic lung infections, populations of microbial pathogens change and mature in ways that are often unknown, which makes it challenging to identify appropriate treatment options. A promising tool to better understand the physiology of microorganisms in a patient is stable-isotope probing, which we previously developed to estimate the growth rates of S. aureus in cystic fibrosis (CF) sputum. Here, we tracked microbial communities in a cohort of CF patients and found that anteiso fatty acids can also originate from other sources in CF sputum. This awareness led us to develop an new workflow for the application of stable isotope probing in this context, improving our ability to estimate pathogen generation times in clinical samples.

scholarly journals Refining the Application of Microbial Lipids as Tracers of Staphylococcus aureus Growth Rates in Cystic Fibrosis Sputum

2018 ◽  
Vol 200 (24) ◽  
Author(s):  
Cajetan Neubauer ◽  
Ajay S. Kasi ◽  
Nora Grahl ◽  
Alex L. Sessions ◽  
Sebastian H. Kopf ◽  
...  
Keyword(s):  
Cystic Fibrosis ◽  
Fatty Acids ◽  
Microbial Community ◽  
Stable Isotope ◽  
Community Composition ◽  
Growth Rates ◽  
Microbial Community Composition ◽  
Stable Isotope Probing ◽  
Content Type ◽  
Lung Infections

ABSTRACT Chronic lung infections in cystic fibrosis (CF) could be treated more effectively if the effects of antimicrobials on pathogens in situ were known. Here, we compared changes in the microbial community composition and pathogen growth rates in longitudinal studies of seven pediatric CF patients undergoing intravenous antibiotic administration during pulmonary exacerbations. The microbial community composition was determined by counting rRNA with NanoString DNA analysis, and growth rates were obtained by incubating CF sputum with heavy water and tracing incorporation of deuterium into two branched-chain (“anteiso”) fatty acids (a-C15:0 and a-C17:0) using gas chromatography-mass spectrometry (GC/MS). Prior to this study, both lipids were thought to be specific for Staphylococcaceae; hence, their isotopic enrichment was interpreted as a growth proxy for Staphylococcus aureus. Our experiments revealed, however, that Prevotella is also a relevant microbial producer of a-C17:0 fatty acid in some CF patients; thus, deuterium incorporation into these lipids is better interpreted as a more general pathogen growth rate proxy. Even accounting for a small nonmicrobial background source detected in some patient samples, a-C15:0 fatty acid still appears to be a relatively robust proxy for CF pathogens, revealing a median generation time of ∼1.5 days, similar to prior observations. Contrary to our expectation, pathogen growth rates remained relatively stable throughout exacerbation treatment. We suggest two straightforward “best practices” for application of stable-isotope probing to CF sputum metabolites: (i) parallel determination of microbial community composition in CF sputum using culture-independent tools and (ii) assessing background levels of the diagnostic metabolite. IMPORTANCE In chronic lung infections, populations of microbial pathogens change and mature in ways that are often unknown, which makes it challenging to identify appropriate treatment options. A promising tool to better understand the physiology of microorganisms in a patient is stable-isotope probing, which we previously developed to estimate the growth rates of S. aureus in cystic fibrosis (CF) sputum. Here, we tracked microbial communities in a cohort of CF patients and found that anteiso fatty acids can also originate from other sources in CF sputum. This awareness led us to develop a new workflow for the application of stable-isotope probing in this context, improving our ability to estimate pathogen generation times in clinical samples.

scholarly journals Switch of fungal to bacterial degradation in natural, drained and rewetted oligotrophic peatlands reflected in δ<sup>15</sup>N and fatty acid composition

2020 ◽  
Author(s):  
Miriam Groß-Schmölders ◽  
Pascal von Sengbusch ◽  
Jan Paul Krüger ◽  
Kristy Woodard ◽  
Axel Birkholz ◽  
...  
Keyword(s):  
Fatty Acid ◽  
Microbial Community ◽  
Stable Isotope ◽  
Community Composition ◽  
Turning Point ◽  
Phospholipid Fatty Acid ◽  
Microbial Community Composition ◽  
Microscopic Analysis ◽  
Bacterial Degradation ◽  
Ecosystem Functions

Abstract. During the last centuries major parts of European peatlands were degraded along with drainage and land use changes. Peatland biodiversity and essential ecosystem functions (e.g. flood prevention, groundwater purification and CO2 sink) were dramatically impaired. Moreover, climate change threatens peatlands in the near future. Increasing pressure to peatland ecosystems calls for a more cost-efficient method to indicate the current state of peatlands and the success of restoration effort. Metabolism processes in peatland soils are imprinted in stable isotope signatures due to differences in microorganism communities and their metabolic pathways. Therefore we hypothesize that depth profiles of nitrogen stable isotope values provide a promising opportunity to detect peatland decomposition or restoration. We studied five peatlands: Degerö Stormyr (Northern Sweden), Lakkasuo (Central Finland) and three mires in the Black Forest (Southern Germany). At all locations cores were taken from adjacent drained (or rewetted) and natural sites to identify δ15N trends that could indicate changes due to drainage and restoration. At all drained (and rewetted) sites we found a distinct peak (turning point) of the δ15N values in the center of the drained horizon. To verify our interpretation δ13C, the C / N ratio and the bulk density were measured and a microscopic analysis of the macro residuals in the peat cores was made. In addition we did a phospholipid fatty acid (PLFAs) analysis to link our results to microbial community composition. We distinguished between fungal and bacterial-derived PLFAs. In accordance with other studies, our results suggest, that fungi dominate the microbial metabolism in the upper, aerobic peat horizon. This is reflected by depleted δ15N values. Downwards the drained horizon conditions slowly switch to oxygen limitation. In consequence fungal-derived PLFAs decreases whereas bacterial-derived PLFAs are rising. The highest diversity of microbial-derived PLFAs is indicated by the δ15N turning point. Below the δ15N turning point, oxygen is increasingly limited and concentrations of all microbial-derived PLFAs are decreasing down to the onset of the permanently waterlogged, anaerobic horizon. Peatland cores with restoration success show, above the formerly drainage-affected horizon, again no depth trend of the isotopic values. Hence, we conclude that δ15N stable isotope values reflect microbial community composition, which differ between drained and natural peatlands.

scholarly journals Shotgun Isotope Array for Rapid, Substrate-Specific Detection of Microorganisms in a Microbial Community

2011 ◽  
Vol 77 (20) ◽  
pp. 7430-7432 ◽  
Author(s):  
Tomohiro Tobino ◽  
Futoshi Kurisu ◽  
Ikuro Kasuga ◽  
Hiroaki Furumai
Keyword(s):  
Microbial Community ◽  
Stable Isotope ◽  
Activated Sludge ◽  
Prior Knowledge ◽  
Community Composition ◽  
Stable Isotope Probing ◽  
Proof Of Concept ◽  
Specific Detection

ABSTRACTThe shotgun isotope array method has been proposed to be an effective new tool for use in substrate-specific microbe exploration without any prior knowledge of the community composition. Proof of concept was demonstrated by detection of acetate-degrading microorganisms in activated sludge and further verified by independent stable isotope probing (SIP).

scholarly journals EPS4.01 Microbial community composition in explanted cystic fibrosis lungs

2018 ◽  
Vol 17 ◽  
pp. S43
Author(s):  
C.D. Spence ◽  
S. Verleden ◽  
G.G. Einarsson ◽  
J. Mcdonough ◽  
A.J. Lee ◽  
...  
Keyword(s):  
Cystic Fibrosis ◽  
Microbial Community ◽  
Community Composition ◽  
Microbial Community Composition

scholarly journals P139 Microbial community composition and patient reported symptoms in cystic fibrosis

2019 ◽  
Vol 18 ◽  
pp. S96-S97
Author(s):  
K. O'Neill ◽  
G.G. Einarsson ◽  
E. Johnson ◽  
C. McGettigan ◽  
L. Sherrard ◽  
...  
Keyword(s):  
Cystic Fibrosis ◽  
Microbial Community ◽  
Community Composition ◽  
Microbial Community Composition ◽  
Patient Reported

scholarly journals Switch of fungal to bacterial degradation in natural, drained and rewetted oligotrophic peatlands reflected in <i>δ</i><sup>15</sup>N and fatty acid composition

SOIL ◽  
2020 ◽  
Vol 6 (2) ◽  
pp. 299-313
Author(s):  
Miriam Groß-Schmölders ◽  
Pascal von Sengbusch ◽  
Jan Paul Krüger ◽  
Kristy Klein ◽  
Axel Birkholz ◽  
...  
Keyword(s):  
Microbial Community ◽  
Stable Isotope ◽  
Community Composition ◽  
Metabolic Pathways ◽  
Turning Point ◽  
Microbial Community Composition ◽  
Bacterial Degradation ◽  
Gram Positive Bacteria ◽  
Current State ◽  
Cost Efficient

Abstract. For centuries European peatlands have been degrading along with drainage, land use and climate changes. Increasing pressure on peatland ecosystems calls for a more cost-efficient method to indicate the current state of peatlands and the success of restoration efforts. Metabolic pathways in peatland soils are imprinted in stable isotope compositions due to differences in microorganism communities and their metabolic pathways. Therefore, we hypothesize that depth profiles of nitrogen stable isotope values provide a promising opportunity to detect peatland decomposition or restoration. We studied five peatlands, namely Degerö Stormyr (northern Sweden), Lakkasuo (central Finland) and three mires in the Black Forest (southern Germany). At all locations, cores were taken from adjacent drained (or rewetted) and natural sites to identify δ15N trends that could indicate changes due to drainage and restoration. At all drained (and rewetted) sites we found a distinct peak (“turning point”) of the δ15N values in the center of the drained horizon. We did a fatty acids (FAs) analysis to link our results to microbial community composition. As markers, we distinguished between one fungal-derived FA (C18:2ω9c) and four bacterial-derived FAs. For bacteria, we looked for one general bacterial-derived FA (C14:0), two FAs for gram-positive bacteria (i-C15:0; a-C15:0), and one FA for gram-negative bacteria (C16:1ω9c). In accordance with other studies, our results suggest that fungi dominate the microbial metabolism in the upper aerobic peat horizon. This is reflected by depleted δ15N values. Moving downwards, the drained horizon conditions slowly switch to oxygen limitation. Consequently, fungal-derived FAs decrease whereas bacterial-derived FAs rise. The highest diversity of microbial-derived FAs is indicated by the δ15N turning point. Below the δ15N turning point, oxygen is increasingly limited and concentrations of all microbial-derived FAs are decreasing down to the onset of the permanently waterlogged anaerobic horizon. Peatland cores with restoration successes again show, above the formerly drained horizon, no depth trend of the isotopic values. Hence, we conclude that δ15N stable isotope values reflect microbial community composition, which differs between drained and natural peatlands.

scholarly journals S103 Microbial community composition in the lungs of patients with Cystic Fibrosis and non-CF bronchiectasis

Thorax ◽  
2011 ◽  
Vol 66 (Suppl 4) ◽  
pp. A48-A48
Author(s):  
G. G. Einarsson ◽  
E. Klem ◽  
A. A. Fodor ◽  
L. Wei ◽  
M. Drain ◽  
...  
Keyword(s):  
Cystic Fibrosis ◽  
Microbial Community ◽  
Community Composition ◽  
Microbial Community Composition
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