Publications in peer reviewed journals
The unexpected versatility of the cellulosome2017 - Environmental Microbiology, 1: 13-14
Lifestyle and horizontal gene transfer-mediated evolution of Mucispirillum schaedleri, a core member of the murine gut microbiota2017 - mSystems, 2: e00171-16
Mucispirillum schaedleri is an abundant inhabitant of the intestinal mucus layer of rodents and other animals and has been suggested to be a pathobiont, a commensal that plays a role in disease. In order to gain insights into its lifestyle, we analyzed the genome and transcriptome of M. schaedleri ASF 457 and performed physiological experiments to test traits predicted by its genome. Although described as a mucus inhabitant, M. schaedleri has limited capacity for degrading host-derived mucosal glycans and other complex polysaccharides. Additionally, M. schaedleri reduces nitrate and expresses systems for scavenging oxygen and reactive oxygen species in vivo, which may account for its localization close to the mucosal tissue and expansion during inflammation. Also of note, M. schaedleri harbors a type VI secretion system and putative effector proteins and can modify gene expression in mucosal tissue, suggesting intimate interactions with its host and a possible role in inflammation. The M. schaedleri genome has been shaped by extensive horizontal gene transfer, primarily from intestinal Epsilon- and Deltaproteobacteria, indicating that horizontal gene transfer has played a key role in defining its niche in the gut ecosystem.
Hidden potential: Diet-driven changes in redox level shape the rumen microbiome2017 - Environmental Microbiology, 1: 19-20
Microbial nutrient niches in the gut2017 - Environ. Microbiol., in press
The composition and function of the mammalian gut microbiota has been the subject of much research in recent years, but the principles underlying the assembly and structure of this complex community remain incompletely understood. Processes that shape the gut microbiota are thought to be mostly niche-driven, with environmental factors such as the composition of available nutrients largely determining whether or not an organism can establish. The concept that the nutrient landscape dictates which organisms can successfully colonize and persist in the gut was first proposed in Rolf Freter's nutrient niche theory. In a situation where nutrients are perfectly mixed and there is balanced microbial growth, Freter postulated that an organism can only survive if it is able to utilize one or a few limiting nutrients more efficiently than its competitors. Recent experimental work indicates, however, that nutrients in the gut vary in space and time. We propose that in such a scenario, Freter's nutrient niche theory must be expanded to account for the co-existence of microorganisms utilizing the same nutrients but in distinct sites or at different times, and that metabolic flexibility and mixed-substrate utilization are common strategies for survival in the face of ever-present nutrient fluctuations.
A 12-week intervention with nonivamide, a TRPV1 agonist, prevents a dietary-induced body fat gain and increases peripheral serotonin in moderately overweight subjects2017 - Mol Nutr Food Res, in press
A bolus administration of 0.15 mg nonivamide has previously been demonstrated to reduce energy intake in moderately overweight men. This 12-week intervention investigated whether a daily consumption of nonivamide in a protein-based product formulation promotes a reduction in body weight in healthy overweight subjects, and affects outcome measures associated with mechanisms regulating food intake, e.g. plasma concentrations of an-/orexigenic hormones, energy substrates as well as changes in fecal microbiota .
Nineteen overweight subjects were randomly assigned to either a control (C) or a nonivamide group (NV). Changes in the body composition and plasma concentrations of satiating hormones were determined at fasting and 15, 30, 60, 90 and 120 min after a glucose load. Participants were instructed to consume 0.15 mg nonivamide per day in 450 mL of a milk shake additionally to their habitual diet. After treatment, a group difference in body fat mass change (- 0.61 ± 0.36 % in NV and + 1.36 ± 0.38 % in C), and an increase in postprandial plasma serotonin were demonstrated. Plasma metabolome and fecal microbiome read outs were not affected.
A daily intake of 0.15 mg nonivamide helps to support to maintain a healthy body composition. Nonivamide, a less-pungent capsaicin analog, was studied for its potential as an anti-obesity agent in a 12-week human intervention study. The body composition was determined at the beginning and in the end of intervention. Moreover, plasma concentrations of satiating hormones were measured Participants were instructed to drink the product formulation three times daily. After treatment, a group difference in body fat mass change, and an increase in postprandial plasma serotonin concentrations were determined in the nonivamide group. Thus, we suppose that a daily intake of nonivamide might help to maintain a healthy body composition. This article is protected by copyright. All rights reserved.
Pediatric obesity is associated with an altered gut microbiota and discordant shifts in Firmicutes populations2017 - Environ. Microbiol., 1: 95-105
An altered gut microbiota has been linked to obesity in adulthood, although little is known about childhood obesity. The aim of this study was to characterize the composition of the gut microbiota in obese (n = 42) and normal-weight (n = 36) children aged 6 to 16. Using 16S rRNA gene-targeted sequencing, we evaluated taxa with differential abundance according to age- and sex-normalized body mass index (BMI z-score). Obesity was associated with an altered gut microbiota characterized by elevated levels of Firmicutes and depleted levels of Bacteroidetes. Correlation network analysis revealed that the gut microbiota of obese children also had increased correlation density and clustering of operational taxonomic units (OTUs). Members of the Bacteroidetes were generally better predictors of BMI z-score and obesity than Firmicutes, which was likely due to discordant responses of Firmicutes OTUs. In accordance with these observations, the main metabolites produced by gut bacteria, short chain fatty acids (SCFAs), were higher in obese children, suggesting elevated substrate utilisation. Multiple taxa were correlated with SCFA levels, reinforcing the tight link between the microbiota, SCFAs and obesity. Our results suggest that gut microbiota dysbiosis and elevated fermentation activity may be involved in the etiology of childhood obesity.
Making It Stick: A Compelling Case for Precision Microbiome Reconstitution2016 - Cell Host & Microbe, 20: 415-417
Modification of the intestinal microbiome is an emerging target to improve health and prevent or treat a number of diseases. In this issue of Cell Host & Microbe, Maldonado-Gomez et al. (2016) uncover the basic principles that govern the successful establishment and persistence of an exogenously introduced gut bacterium.
Genome-guided design of a novel defined mouse microbiota that confers colonization resistance against Salmonella enterica serovar Typhimurium2016 - Nature Microbiol, 2: 16215
Protection against enteric infections, also termed colonization resistance, results from mutualistic interactions of the host and its indigenous microbes. The gut microbiota of humans and mice is highly diverse and it is therefore challenging to assign specific properties to its individual members. Here, we have used a collection of murine bacterial strains and a modular design approach to create a minimal bacterial community that, once established in germ-free mice, provided colonization resistance against the human enteric pathogen Salmonella enterica serovar Typhimurium (S. Tm). Initially, a community of 12 strains, termed Oligo-Mouse Microbiota (Oligo-MM12), representing members of the major bacterial phyla in the murine gut, was selected. This community was stable over consecutive mouse generations and provided colonization resistance against S. Tm infection, albeit not to the degree of a conventional complex microbiota. Comparative (meta)genome analyses identified functions represented in a conventional microbiome but absent from the Oligo-MM12. By genome-informed design, we created an improved version of the Oligo-MM community harbouring three facultative anaerobic bacteria from the Mouse Intestinal Bacterial Collection (miBC) that provided conventional-like colonization resistance. In conclusion, we have established a highly versatile experimental system that showed efficacy in an enteric infection model. Thus, in combination with exhaustive bacterial strain collections and systems-based approaches, genomeguided design can be used to generate insights into microbe–microbe and microbe–host interactions for the investigation of ecological and disease-relevant mechanisms in the intestine.
Bacterial nutrient foraging in a mouse model of enteral nutrient deprivation: Insight into the gut origin of sepsis2016 - Am J Physiol Gastrointest Liver Physiol, 311: G734-G743
Total parenteral nutrition (TPN) leads to a shift in small intestinal microbiota with a characteristic dominance of Proteobacteria. This study examined how metabolomic changes within the small bowel support an altered microbial community in enterally deprived mice.
C57BL/6 mice were given TPN or enteral chow. Metabolomic analysis of jejunal contents was performed by liquid chromatography/mass spectrometry (LC/MS). In some experiments, leucine in TPN was partly substituted with (13)C-leucine. Additionally, jejunal contents from TPN dependent and enterally fed mice were gavaged into germ-free mice to reveal if the TPN phenotype was transferrable.
Small bowel contents of TPN mice maintained an amino acid composition similar to that of the TPN solution. Mass spectrometry analysis of small bowel contents of TPN dependent mice showed increased concentration of (13)C compared to fed mice receiving saline enriched with (13)C-leucine. (13)C-leucine added to the serosal side of Ussing chambers showed rapid permeation across TPN-dependent jejunum, suggesting increased transmucosal passage. Single-cell analysis by fluorescence in situ hybridization (FISH) - NanoSIMS demonstrated uptake of (13)C-leucine by TPN-associated bacteria, with preferential uptake by Enterobacteriaceae. Gavage of small bowel effluent from TPN mice into germ-free, fed mice resulted in a trend toward the pro-inflammatory TPN-phenotype with loss of epithelial barrier function.
TPN-dependence leads to increased permeation of TPN-derived nutrients into the small intestinal lumen, where they are predominately utilized by Enterobacteriaceae. The altered metabolomic composition of the intestinal lumen during TPN promotes dysbiosis.
Behavior of platinum(iv) complexes in models of tumor hypoxia: cytotoxicity, compound distribution and accumulation2016 - Metallomics, 4: 422-33
Hypoxia in solid tumors remains a challenge for conventional cancer therapeutics. As a source for resistance, metastasis development and drug bioprocessing, it influences treatment results and disease outcome. Bioreductive platinum(iv) prodrugs might be advantageous over conventional metal-based therapeutics, as biotransformation in a reductive milieu, such as under hypoxia, is required for drug activation. This study deals with a two-step screening of experimental platinum(iv) prodrugs with different rates of reduction and lipophilicity with the aim of identifying the most appropriate compounds for further investigations. In the first step, the cytotoxicity of all compounds was compared in hypoxic multicellular spheroids and monolayer culture using a set of cancer cell lines with different sensitivities to platinum(ii) compounds. Secondly, two selected compounds were tested in hypoxic xenografts in SCID mouse models in comparison to satraplatin, and, additionally, (LA)-ICP-MS-based accumulation and distribution studies were performed for these compounds in hypoxic spheroids and xenografts. Our findings suggest that, while cellular uptake and cytotoxicity strongly correlate with lipophilicity, cytotoxicity under hypoxia compared to non-hypoxic conditions and antitumor activity of platinum(iv) prodrugs are dependent on their rate of reduction.
The emerging view of Firmicutes as key fibre degraders in the human gut2016 - Environ. Microbiol., 7: 2081-3
Activity and community structures of sulfate-reducing microorganisms in polar, temperate and tropical marine sediments2016 - ISME J, 10: 796–809
Temperature has a fundamental impact on the metabolic rates of microorganisms and strongly influences microbial ecology and biogeochemical cycling in the environment. In this study, we examined the catabolic temperature response of natural communities of sulfate-reducing microorganisms (SRM) in polar, temperate, and tropical marine sediments. In short-term sediment incubation experiments with 35S-sulfate, we demonstrated how the cardinal temperatures for sulfate reduction correlate with mean annual sediment temperatures, indicating specific thermal adaptations of the dominant SRM in each of the investigated ecosystems. The community structure of putative SRM in the sediments, as revealed by pyrosequencing of bacterial 16S rRNA gene amplicons and phylogenetic assignment to known SRM taxa, consistently correlated with in situ temperatures, but not with sediment organic carbon concentrations or C:N ratios of organic matter. Additionally, several species-level SRM phylotypes of the class Deltaproteobacteria tended to co-occur at sites with similar mean annual temperatures, regardless of geographic distance. The observed temperature adaptations of SRM imply that environmental temperature is a major controlling variable for physiological selection and ecological and evolutionary differentiation of microbial communities.
Response of the bacterial community associated with a cosmopolitan marine diatom to crude oil shows a preference for the biodegradation of aromatic hydrocarbons.2016 - Environ. Microbiol., 6: 1817-33
Emerging evidence shows that hydrocarbonoclastic bacteria (HCB) may be commonly found associated with phytoplankton in the ocean, but the ecology of these bacteria and how they respond to crude oil remains poorly understood. Here, we used a natural diatom-bacterial assemblage to investigate the diversity and response of HCB associated with a cosmopolitan marine diatom, Skeletonema costatum, to crude oil. Pyrosequencing analysis and qPCR revealed a dramatic transition in the diatom-associated bacterial community, defined initially by a short-lived bloom of Methylophaga (putative oil degraders) that was subsequently succeeded by distinct groups of HCB (Marinobacter, Polycyclovorans, Arenibacter, Parvibaculum, Roseobacter clade), including putative novel phyla, as well as other groups with previously unqualified oil-degrading potential. Interestingly, these oil-enriched organisms contributed to the apparent and exclusive biodegradation of substituted and non-substituted polycyclic aromatic hydrocarbons (PAHs), thereby suggesting that the HCB community associated with the diatom is tuned to specializing in the degradation of PAHs. Furthermore, the formation of marine oil snow (MOS) in oil-amended incubations was consistent with its formation during the Deepwater Horizon oil spill. This work highlights the phycosphere of phytoplankton as an underexplored biotope in the ocean where HCB may contribute importantly to the biodegradation of hydrocarbon contaminants in marine surface waters.
Intestinal microbiota signatures associated with inflammation history in mice experiencing recurring colitis2015 - Front Microbiol, 6: 1408
Acute colitis causes alterations in the intestinal microbiota, but the microbiota is thought to recover after such events. Extreme microbiota alterations are characteristic of human chronic inflammatory bowel diseases, although alterations reported in different studies are divergent and sometimes even contradictory. To better understand the impact of periodic disturbances on the intestinal microbiota and its compositional difference between acute and relapsing colitis, we investigated the beginnings of recurrent inflammation using the dextran sodium sulfate (DSS) mouse model of chemically induced colitis. Using bacterial 16S rRNA gene-targeted pyrosequencing as well as quantitative fluorescence in situ hybridization, we profiled the intestinal and stool microbiota of mice over the course of three rounds of DSS-induced colitis and recovery. We found that characteristic inflammation-associated microbiota could be detected in recovery-phase mice. Successive inflammation episodes further drove the microbiota into an increasingly altered composition post-inflammation, and signatures of colitis history were detectable in the microbiota more sensitively than by pathology analysis. Bacterial indicators of murine colitis history were identified in intestinal and stool samples, with a high degree of consistency between both sample types. Stool may therefore be a promising non-invasive source of bacterial biomarkers that are highly sensitive to inflammation state and history.
Intestinal epithelial cell tyrosine kinase 2 transduces interleukin-22 signals to protect from acute colitis2015 - J Immunol., 195: 5011-5024
In the intestinal tract, IL-22 activates signal transducer and activator of transcription 3 (STAT3) to promote intestinal epithelial cell (IEC) homeostasis and tissue healing. The mechanism has remained obscure but we demonstrate that IL-22 acts via tyrosine kinase 2 (Tyk2), a member of the Janus kinase (Jak) family. Using a mouse model for colitis, we show that Tyk2 deficiency is associated with an altered composition of the gut microbiota and exacerbates inflammatory bowel disease (IBD). Colitic Tyk2-/- mice have less phosphorylated STAT3 (pY-STAT3) in colon tissue and their IECs proliferate less efficiently. Tyk2-deficient primary IECs show reduced pY-STAT3 in response to IL-22 stimulation and expression of IL-22-STAT3 target genes is reduced in IECs from healthy and colitic Tyk2-/- mice. Experiments with conditional Tyk2-/- mice reveal that IEC-specific depletion of Tyk2 aggravates colitis. Disease symptoms can be alleviated by administering high doses of recombinant IL-22-Fc, indicating that Tyk2 deficiency can be rescued via the IL-22 receptor complex. The pivotal function of Tyk2 in IL-22-dependent colitis was confirmed in Citrobacter rodentium-induced disease. Thus, Tyk2 protects against acute colitis in part by amplifying inflammation-induced epithelial IL-22 signaling to STAT3.
A flexible and economical barcoding approach for highly multiplexed amplicon sequencing of diverse target genes.2015 - Front Microbiol, 6: 731
High throughput sequencing of phylogenetic and functional gene amplicons provides tremendous insight into the structure and functional potential of complex microbial communities. Here, we introduce a highly adaptable and economical PCR approach to barcoding and pooling libraries of numerous target genes. In this approach, we replace gene- and sequencing platform-specific fusion primers with general, interchangeable barcoding primers, enabling nearly limitless customized barcode-primer combinations. Compared to barcoding with long fusion primers, our multiple-target gene approach is more economical because it overall requires lower number of primers and is based on short primers with generally lower synthesis and purification costs. To highlight our approach, we pooled over 900 different small-subunit rRNA and functional gene amplicon libraries obtained from various environmental or host-associated microbial community samples into a single, paired-end Illumina MiSeq run. Although the amplicon regions ranged in size from approximately 290 to 720 bp, we found no significant systematic sequencing bias related to amplicon length or gene target. Our results indicate that this flexible multiplexing approach produces large, diverse, and high quality sets of amplicon sequence data for modern studies in microbial ecology.
Cyanate as an energy source for nitrifiers.2015 - Nature, 524: 105-108
Ammonia- and nitrite-oxidizing microorganisms are collectively responsible for the aerobic oxidation of ammonia via nitrite to nitrate and have essential roles in the global biogeochemical nitrogen cycle. The physiology of nitrifiers has been intensively studied, and urea and ammonia are the only recognized energy sources that promote the aerobic growth of ammonia-oxidizing bacteria and archaea. Here we report the aerobic growth of a pure culture of the ammonia-oxidizing thaumarchaeote Nitrososphaera gargensis using cyanate as the sole source of energy and reductant; to our knowledge, the first organism known to do so. Cyanate, a potentially important source of reduced nitrogen in aquatic and terrestrial ecosystems, is converted to ammonium and carbon dioxide in Nitrososphaera gargensis by a cyanase enzyme that is induced upon addition of this compound. Within the cyanase gene family, this cyanase is a member of a distinct clade also containing cyanases of nitrite-oxidizing bacteria of the genus Nitrospira. We demonstrate by co-culture experiments that these nitrite oxidizers supply cyanase-lacking ammonia oxidizers with ammonium from cyanate, which is fully nitrified by this microbial consortium through reciprocal feeding. By screening a comprehensive set of more than 3,000 publically available metagenomes from environmental samples, we reveal that cyanase-encoding genes clustering with the cyanases of these nitrifiers are widespread in the environment. Our results demonstrate an unexpected metabolic versatility of nitrifying microorganisms, and suggest a previously unrecognized importance of cyanate in cycling of nitrogen compounds in the environment.
Tracking heavy water (D2O) incorporation for identifying and sorting active microbial cells2015 - Proc Natl Acad Sci USA, 112: E194-203
Microbial communities are essential to the function of virtually all ecosystems and eukaryotes, including humans. However, it is still a major challenge to identify microbial cells active under natural conditions in complex systems. In this study, we developed a new method to identify and sort active microbes on the single-cell level in complex samples using stable isotope probing with heavy water (D2O) combined with Raman microspectroscopy. Incorporation of D2O-derived D into the biomass of autotrophic and heterotrophic bacteria and archaea could be unambiguously detected via C-D signature peaks in single-cell Raman spectra, and the obtained labeling pattern was confirmed by nanoscale-resolution secondary ion MS. In fast-growing Escherichia coli cells, label detection was already possible after 20 min. For functional analyses of microbial communities, the detection of D incorporation from D2O in individual microbial cells via Raman microspectroscopy can be directly combined with FISH for the identification of active microbes. Applying this approach to mouse cecal microbiota revealed that the host-compound foragers Akkermansia muciniphila and Bacteroides acidifaciens exhibited distinctive response patterns to amendments of mucin and sugars. By Raman-based cell sortingof active (deuterated) cells with optical tweezers and subsequent multiple displacement amplification and DNA sequencing, novel cecal microbes stimulated by mucin and/or glucosamine were identified, demonstrating the potential of the nondestructive D2O-Raman approach for targeted sortingof microbial cells with defined functional properties for single-cell genomics.
Type I interferons have opposing effects during the emergence and recovery phases of colitis.2014 - Eur J Immunol., 44: 2749-60
The contribution of the innate immune system to inflammatory bowel disease (IBD) is under intensive investigation. Research in animal models has demonstrated that type I interferons (IFN-Is) protect from IBD. In contrast, studies of patients with IBD have produced conflicting results concerning the therapeutic potential of IFN-Is. Here, we present data suggesting that IFN-Is play dual roles as regulators of intestinal inflammation in dextran sodium sulfate (DSS)-treated C57BL/6 mice. Though IFN-Is reduced acute intestinal damage and the abundance ofcolitis-associated intestinal bacteria caused by treatment with a high dose of DSS, they also inhibited the resolution of inflammation after DSS treatment. IFN-Is played an anti-inflammatory role by suppressing the release of IL-1β from the colon MHC class II(+) cells. Consistently, IL-1 receptor blockade reduced the severity of inflammation in IFN-I receptor-deficient mice and myeloid cell-restricted ablation of the IFN-I receptor was detrimental. The proinflammatory role of IFN-Is during recovery from DSS treatment was caused by IFN-I-dependent cell apoptosis as well as an increase in chemokine production and infiltrating inflammatory monocytes and neutrophils. Thus, IFN-Is play opposing roles in specificphases of intestinal injury and inflammation, which may be important for guiding treatment strategies in patients.
Deciphering microbial interactions and detecting keystone species with co-occurrence networks2014 - Front Microbiol, 5: 219
Co-occurrence networks produced from microbial survey sequencing data are frequently used to identify interactions between community members. While this approach has potential to reveal ecological processes, it has been insufficiently validated due to the technical limitations inherent in studying complex microbial ecosystems. Here, we simulate multi-species microbial communities with known interaction patterns using generalized Lotka-Volterra dynamics. We then construct co-occurrence networks and evaluate how well networks reveal the underlying interactions and how experimental and ecological parameters can affect network inference and interpretation. We find that co-occurrence networks can recapitulate interaction networks under certain conditions, but that they lose interpretability when the effects of habitat filtering become significant. We demonstrate that networks suffer from local hot spots of spurious correlation in the neighborhood of hub species that engage in many interactions. We also identify topological features associated with keystone species in co-occurrence networks. This study provides a substantiated framework to guide environmental microbiologists in the construction and interpretation of co-occurrence networks from microbial survey datasets.
High-fat diet alters gut microbiota physiology in mice2014 - ISME J., 8: 295-308
The intestinal microbiota is known to regulate host energy homeostasis and can be influenced by high-calorie diets. However, changes affecting the ecosystem at the functional level are still not well characterized. We measured shifts in cecal bacterial communities in mice fed a carbohydrate orhigh-fat (HF) diet for 12 weeks at the level of the following: (i) diversity and taxa distribution by high-throughput 16S ribosomal RNA gene sequencing; (ii) bulk and single-cell chemical composition by Fourier-transform infrared- (FT-IR) and Raman micro-spectroscopy and (iii) metaproteome and metabolome via high-resolution mass spectrometry. High-fat diet caused shifts in the diversity of dominant gut bacteria and altered the proportion of Ruminococcaceae (decrease) and Rikenellaceae (increase). FT-IR spectroscopy revealed that the impact of the diet on cecal chemical fingerprints is greater than the impact of microbiota composition. Diet-driven changes in biochemical fingerprints of members of the Bacteroidales and Lachnospiraceae were also observed at the level of single cells, indicating that there were distinct differences in cellular composition of dominant phylotypes under different diets. Metaproteome and metabolome analyses based on the occurrence of 1760 bacterial proteins and 86 annotated metabolites revealed distinct HF diet-specific profiles. Alteration of hormonal and anti-microbial networks, bile acid and bilirubin metabolism and shifts towards amino acid and simple sugars metabolism were observed. We conclude that a HF diet markedly affects the gut bacterial ecosystem at the functional level.
Removal of pharmaceuticals and personal care products during water recycling: microbial community structure and effects of substrate concentration.2014 - Appl Environ Microbiol., 80: 2440-50
Many pharmaceuticals and personal care products (PPCPs) have been shown to be biotransformed in water treatment systems. However, little research exists on the effect of initial PPCP concentration on PPCP biotransformation or on the microbial communities treating impacted water. In this study, biological PPCP removal at various concentrations was assessed using laboratory columns inoculated with wastewater treatment plant effluent. Pyrosequencing was used to examine microbial communities in the columns and in soil from a soil aquifer treatment (SAT; a method ofwater treatment prior to reuse) site. Laboratory columns were supplied with different concentrations (0.25, 10, 100, or 1,000 μg liter(-1)) of each of 15 PPCPs. Five PPCPs (4-isopropyl-3-methylphenol [biosol], p-chloro-m-xylenol, gemfibrozil, ketoprofen, and phenytoin) were not removed at any tested concentrations. Two PPCPs (naproxen and triclosan) exhibited removals independent of PPCP concentration. PPCP removal efficiencies were dependent on initial concentrations for biphenylol, p-chloro-m-cresol, chlorophene, diclofenac, 5-fluorouracil, ibuprofen, and valproic acid, showing that PPCP concentration can affect biotransformation. Biofilms from sand samples collected from the 0.25- and 10-μg liter(-1) PPCP columns were pyrosequenced along with SAT soil samples collected on three consecutive days of a wetting and drying cycle to enable comparison of these two communities exposed to PPCPs. SAT communities were similar to column communities in taxonomy and phylotype composition, and both were found to contain close relatives of known PPCP degraders. The efficiency of biological removal of PPCPs was found to be dependent on the concentration at which the contamination occurs for some, but not all, PPCPs.
Longitudinal study of murine microbiota activity and interactions with the host during acute inflammation and recovery2014 - ISME J., 8(5):1101-14
Although alterations in gut microbiota composition during acute colitis have been repeatedly observed, associated functional changes and therecovery from dysbiosis received little attention. In this study, we investigated structure and function of the gut microbiota during acute inflammationand recovery in a dextran sodium sulfate (DSS)-colitis mouse model using metatranscriptomics, bacterial 16S rRNA gene amplicon sequencing and monitoring of selected host markers. Parallel to an increase of host markers of inflammation during acute colitis, we observed relative abundance shifts and alterations in phylotype composition of the dominant bacterial orders Clostridiales and Bacteroidales, and an increase of the low abundant Enterobacteriales, Deferribacterales, Verrucomicrobiales and Erysipelotrichales. During recovery, the microbiota began to resume, but did not reach its original composition until the end of the experiment. Microbial gene expression was more resilient to disturbance, with pre-perturbation-type transcript profiles appearing quickly after acute colitis. The decrease of Clostridiales during inflammation correlated with a reduction of transcripts related to butyrate formation, suggesting a disturbance in host-microbe signalling and mucosal nutrient provision. The impact of acute inflammationon the Clostridiales was also characterized by a significant downregulation of their flagellin-encoding genes. In contrast, the abundance of members of the Bacteroidales increased along with an increase in transcripts related to mucin degradation. We propose that acute inflammation triggered a selective reaction of the immune system against flagella of commensals and temporarily altered murine microbiota composition and functions relevant for the host. Despite changes in specific interactions, the host-microbiota homeostasis revealed a remarkable ability for recovery.
Intestinal microbiota reduces genotoxic endpoints induced by high-energy protons.2014 - Radiat Res, 181: 45-53
Ionizing space radiation causes oxidative DNA damage and triggers oxidative stress responses, and compromised DNA repair mechanisms can lead to increased risk of carcinogenesis. Young adult mice with developed innate and adaptive immune systems that harbored either a conventionalintestinal microbiota (CM) or an intestinal microbiota with a restricted microbial composition (RM) were irradiated with a total dose of 1 Gy delivered by high-energy protons (2.5 GeV/n, LET = 0.2-2 keV/μm) or silicon or iron ions (850 MeV/n, LET ≈ 50 keV/μm and 1 GeV/n, LET = 150 keV/μm, respectively). Six hours after whole-body irradiation, acute chromosomal DNA lesions were observed for RM mice but not CM mice. High-throughput rRNA gene sequencing of intestinal mucosal bacteria showed that Barnesiella intestinihominis and unclassified Bacterodiales were significantly more abundant in male RM mice than CM mice, and phylotype densities changed in irradiated mice. In addition, Helicobacter hepaticus and Bacteroides stercoris were higher in CM than RM mice. Elevated levels of persistently phosphorylated γ-H2AX were observed in RM mice exposed to high-energy protons compared to nonirradiated RM mice, and they also were associated with a decrease of the antioxidant glutathione in peripheral blood measured at four weeks after irradiation. After radiation exposure, CM mice showed lower levels of γ-H2AX phosphorylation than RM mice and an increase in specific RM-associated phylotypes, indicating a down-regulating force on DNA repair by differentially abundant phylotypes in RM versus a radiation-sensitive complex CM.
Endospores of thermophilic bacteria as tracers of microbial dispersal by ocean currents2014 - ISME J., 8: 1153-65
Microbial biogeography is influenced by the combined effects of passive dispersal and environmental selection, but the contribution of either factor can be difficult to discern. As thermophilic bacteria cannot grow in the cold seabed, their inactive spores are not subject to environmental selection. We therefore conducted a global experimental survey using thermophilic endospores that are passively deposited by sedimentation to the cold seafloor as tracers to study the effect of dispersal by ocean currents on the biogeography of marine microorganisms. Our analysis of 81 different marine sediments from around the world identified 146 species-level 16S rRNA phylotypes of endospore-forming, thermophilic Firmicutes. Phylotypes showed various patterns of spatial distribution in the world oceans and were dispersal-limited to different degrees. Co-occurrence of several phylotypes in locations separated by great distances (west of Svalbard, the Baltic Sea and the Gulf of California) demonstrated a widespread but not ubiquitous distribution. In contrast, Arctic regions with water masses that are relatively isolated from global ocean circulation (Baffin Bay and east of Svalbard) were characterized by low phylotype richness and different compositions of phylotypes. The observed distribution pattern ofthermophilic endospores in marine sediments suggests that the impact of passive dispersal on marine microbial biogeography is controlled by the connectivity of local water masses to ocean circulation.
Polycyclic aromatic hydrocarbon degradation of a phytoplankton-associated Arenibacter spp. and description of Arenibacter algicola sp. nov., an aromatic hydrocarbon-degrading bacterium2014 - Appl Environ Microbiol., 80: 618-28
Pyrosequencing of the bacterial community associated with a cosmopolitan marine diatom during enrichment with crude oil revealed severalArenibacter phylotypes, of which one (OTU-202) had become significantly enriched by the oil. Since members of the genus Arenibacter have not been previously shown to degrade hydrocarbons, we attempted to isolate a representative strain of this genus in order to directly investigate itshydrocarbon-degrading potential. Based on 16S rRNA sequencing, one isolate (designated strain TG409(T)) exhibited >99% sequence identity to three type strains of this genus. On the basis of phenotypic and genotypic characteristics, strain TG409(T) represents a novel species in the genusArenibacter, for which the name Arenibacter algicola sp. nov. is proposed. We reveal for the first time that polycyclic aromatic hydrocarbon (PAH)degradation is a shared phenotype among members of this genus, indicating that it could be used as a taxonomic marker for this genus. Kinetic data for PAH mineralization rates showed that naphthalene was preferred to phenanthrene, and its mineralization was significantly enhanced in the presence of glass wool (a surrogate for diatom cell surfaces). During enrichment on hydrocarbons, strain TG409(T) emulsified n-tetradecane and crude oil, and cells were found to be preferentially attached to oil droplets, indicating an ability by the strain to express cell surface amphiphilic substances (biosurfactants or bioemulsifiers) as a possible strategy to increase the bioavailability of hydrocarbons. This work adds to our growing knowledge on the diversity of bacterial genera in the ocean contributing to the degradation of oil contaminants and of hydrocarbon-degrading bacteria found living in association with marine eukaryotic phytoplankton.
NxrB encoding the beta subunit of nitrite oxidoreductase as functional and phylogenetic marker for nitrite-oxidizing Nitrospira2014 - Environ Microbiol, 16: 3055-3071
Nitrospira are the most widespread and diverse known nitrite-oxidizing bacteria and key nitrifiers in natural and engineered ecosystems. Nevertheless, their ecophysiology and environmental distribution are understudied due to the recalcitrance of Nitrospira to cultivation and the lack of a molecular functional marker, which would allow the detection of Nitrospira in the environment. Here we introduce nxrB, the gene encoding subunit beta of nitrite oxidoreductase, as a functional and phylogenetic marker for Nitrospira. Phylogenetic trees based on nxrB of Nitrospira were largely congruent to 16S rRNA-based phylogenies. By using new nxrB-selective PCR primers, we obtained almost full-length nxrB sequences from Nitrospira cultures, two activated sludge samples, and several geographically and climatically distinct soils. Amplicon pyrosequencing of nxrB fragments from 16 soils revealed a previously unrecognized diversity of terrestrial Nitrospira with 1,801 detected species-level OTUs (using an inferred species threshold of 95% nxrB identity). Richness estimates ranged from 10 to 946 co-existing Nitrospira species per soil. Comparison to an archaeal amoA dataset obtained from the same soils [Environ. Microbiol. 14: 525-539 (2012)] uncovered that ammonia-oxidizing archaea and Nitrospira communities were highly correlated across the soil samples, possibly indicating shared habitat preferences or specific biological interactions among members of these nitrifier groups.
Polycyclic Aromatic Hydrocarbon Degradation of Phytoplankton-Associated Arenibacter spp. and Description of Arenibacter algicola sp. nov., an Aromatic Hydrocarbon-Degrading Bacterium2013 - Appl. Environ. Microbiol., 80:2 618-628
Temporal bacterial community dynamics vary among ulcerative colitis patients after fecal microbiota transplantation2013 - Am. J. Gastroenterol., 108: 1620-1630
OBJECTIVES: Fecal microbiota transplantation (FMT) from healthy donors, which is an effective alternative for treatment of Clostridium difficile-associated disease, is being considered for several disorders such as inflammatory bowel disease, irritable bowel syndrome, and metabolic syndrome. Disease remission upon FMT is thought to be facilitated by an efficient colonization of healthy donor microbiota, but knowledge of the composition and temporal stability of patient microbiota after FMT is lacking.METHODS:Five patients with moderately to severely active ulcerative colitis (Mayo score ≥6) and refractory to standard therapy received FMT via nasojejunal tube and enema. In addition to clinical activity and adverse events, the patients' fecal bacterial communities were monitored at multiple time points for up to 12 weeks using 16S rRNA gene-targeted pyrosequencing. RESULTS:FMT elicited fever and a temporary increase of C-reactive protein. Abundant bacteria from donors established in recipients, but the efficiency and stability of donor microbiota colonization varied greatly. A positive clinical response was observed after 12 weeks in one patient whose microbiota had been effectively augmented by FMT. This augmentation was marked by successive colonization of donor-derived phylotypes including the anti-inflammatory and/or short-chain fatty acid-producing Faecalibacterium prausnitzii, Rosebura faecis, and Bacteroides ovatus. Disease severity (as measured by the Mayo score) was associated with an overrepresentation of Enterobacteriaceae and an underrepresentation of Lachnospiraceae. CONCLUSIONS:This study highlights the value of characterizing temporally resolved microbiota dynamics for a better understanding of FMT efficacy and provides potentially useful diagnostic indicators for monitoring FMT success in the treatment of ulcerative colitis.
Intestinal bacteria modify lymphoma incidence and latency by affecting systemic inflammatory state, oxidative stress, and leucocyte genotoxicity2013 - Cancer Res., 73: 4222-4232
Ataxia-telangiectasia is a genetic disorder associated with high incidence of B-cell lymphoma. Using an ataxia-telangiectasia mouse model, we compared lymphoma incidence in several isogenic mouse colonies harboring different bacterial communities, finding that intestinal microbiota are a major contributor to disease penetrance and latency, lifespan, molecular oxidative stress, and systemic leukocyte genotoxicity. High-throughput sequence analysis of rRNA genes identified mucosa-associated bacterial phylotypes that were colony-specific. Lactobacillus johnsonii, which was deficient in the more cancer-prone mouse colony, was causally tested for its capacity to confer reduced genotoxicity when restored by short-term oral transfer. This intervention decreased systemic genotoxicity, a response associated with reduced basal leukocytes and the cytokine-mediated inflammatory state, and mechanistically linked to the host cell biology of systemic genotoxicity. Our results suggest that intestinal microbiota are a potentially modifiable trait for translational intervention in individuals at risk for B-cell lymphoma, or for other diseases that are driven by genotoxicity or the molecular response to oxidative stress.
Role of bacterial exopolymers (EPS) in the fate of the oil released during the Deepwater Horizon oil spill2013 - PLoS One, 8(6):e67717
Halomonas species are recognized for producing exopolysaccharides (EPS) exhibiting amphiphilic properties that allow these macromolecules to interface with hydrophobic substrates, such as hydrocarbons. There remains a paucity of knowledge, however, on the potential of Halomonas EPS to influence the biodegradation of hydrocarbons. In this study, the well-characterized amphiphilic EPS produced by Halomonas species strain TG39 was shown to effectively increase the solubilization of aromatic hydrocarbons and enhance their biodegradation by an indigenous microbial community from oil-contaminated surface waters collected during the active phase of the Deepwater Horizon oil spill. Three Halomonas strains were isolated from the Deepwater Horizon site, all of which produced EPS with excellent emulsifying qualities and shared high (97-100%) 16S rRNA sequence identity with strain TG39 and other EPS-producing Halomonas strains. Analysis of pyrosequence data from surface water samples collected during the spill revealed several distinct Halomonas phylotypes, of which some shared a high sequence identity (≥97%) to the Halomonas isolates. Other bacterial groups comprising members with well-characterized EPS-producing qualities, such as Alteromonas, Colwellia and Pseudoalteromonas, were also found enriched in surface waters, suggesting that the total pool of EPS in the Gulf during the spill may have been supplemented by these organisms. Roller bottle incubations with one of the Halomonas isolates from Deepwater Horizon demonstrated its ability to effectively produce oil aggregates and emulsify the oil. The enrichment of EPS-producing bacteria during the spill coupled with their capacity to produce amphiphilic EPS is likely to have contributed to the ultimate removal of the oil and in the formation of oil aggregates, which were a dominant feature observed in contaminated surface waters.
Colonization resistance and microbial ecophysiology: Using gnotobiotic mouse models and single-cell technology to explore the intestinal jungle2013 - FEMS Microbiol. Rev., 37: 793-829
The highly diverse intestinal microbiota forms a structured community engaged in constant communication with itself and its host and is characterized by extensive ecological interactions. A key benefit that the microbiota affords its host is its ability to protect against infections in a process termed colonization resistance (CR), which remains insufficiently understood. In this review, we connect basic concepts of CR with new insights from recent years and highlight key technological advances in the field of microbial ecology. We present a selection of statistical and bioinformatics tools used to generate hypotheses about synergistic and antagonistic interactions in microbial ecosystems from metagenomic datasets. We emphasize the importance of experimentally testing these hypotheses and discuss the value of gnotobiotic mouse models for investigating specific aspects related to microbiota-host-pathogen interactions in a well-defined experimental system. We further introduce new developments in the area of single-cell analysis using fluorescence in situ hybridization in combination with metabolic stable isotope labeling technologies for studying the in vivo activities of complex community members. These approaches promise to yield novel insights into the mechanisms of CR and intestinal ecophysiology in general, and give researchers the means to experimentally test hypotheses in vivo at varying levels of biological and ecological complexity.
Host-compound foraging by intestinal microbiota revealed by single-cell stable isotope probing2013 - Proc. Natl. Acad. Sci. USA, 110: 4720-4725
The animal and human intestinal mucosa secretes an assortment of compounds to establish a physical barrier between the host tissue and intestinal contents, a separation that is vital for health. Some pathogenic microorganisms as well as members of the commensal intestinal microbiota have been shown to be able to break down these secreted compounds. Our understanding of host-compound degradation by the commensal microbiota has been limited to knowledge about simplified model systems because of the difficulty in studying the complex intestinal ecosystem in vivo. In this study, we introduce an approach that overcomes previous technical limitations and allows us to observe which microbial cells in the intestine use host-derived compounds. We added stable isotope-labeled threonine i.v. to mice and combined fluorescence in situ hybridization with high-resolution secondary ion mass spectrometry imaging to characterize utilization of host proteins by individual bacterial cells. We show that two bacterial species, Bacteroides acidifaciens and Akkermansia muciniphila, are important host-protein foragers in vivo. Using gnotobiotic mice we show that microbiota composition determines the magnitude and pattern of foraging by these organisms, demonstrating that a complex microbiota is necessary in order for this niche to be fully exploited. These results underscore the importance of in vivo studies of intestinal microbiota, and the approach presented in this study will be a powerful tool to address many other key questions in animal and human microbiome research.
Intestinal microbiota: a source of novel biomarkers in inflammatory bowel disease?2013 - Best Pract. Res. Clin. Gastroenterol., 27: 47-58
The human intestine harbors a complex microbial ecosystem that performs manifold functions important to the nutrition and health of its host. Extensive study has revealed that the composition of the intestinal microbiota is altered in individuals with inflammatory bowel disease (IBD). The IBD associated intestinal microbiota generally has reduced species richness and diversity, lower temporal stability, and disruption of the secreted mucus layer structure. Multiple studies have identified certain bacterial taxa that are enriched or depleted in IBD including Enterobacteriaceae, Ruminococcus gnavus, and Desulfovibrio (enriched) and Faecalibacterium prausnitzii, Lachnospiraceae, and Akkermansia (depleted). Additionally, the relative abundance of some taxa appears to correlate with established markers of disease activity such as Enterobacteriaceae (enriched) and Lachnospiraceae (depleted). Signature shifts in fecal microbial community composition may therefore prove to be valuable as diagnostic biomarkers, particularly for longitudinal monitoring of disease activity and response to treatments.
Hydrocarbon-degrading bacteria enriched by the Deepwater Horizon oil spill identified by cultivation and stable isotope probing2013 - ISME J., 7: 2091-2104
The massive influx of crude oil into the Gulf of Mexico during the Deepwater Horizon disaster triggered dramatic microbial community shifts in surface oil slick and deep plume waters. Previous work had shown several taxa, notably DWH Oceanospirillales, Cycloclasticus and Colwellia, were found enriched in these waters based on their dominance in conventional clone and pyrosequencing libraries and were thought to have played a significant role in the degradation of the oil. However, this type of community analysis data failed to provide direct evidence on the functional properties, such as hydrocarbon degradation of organisms. Using DNA-based stable-isotope probing (DNA-SIP) with uniformly 13C-labelled hydrocarbons, we identified several aliphatic- (Alcanivorax, Marinobacter) and polycyclic aromatic hydrocarbon (PAH)- (Alteromonas, Cycloclasticus, Colwellia) degrading bacteria. We also isolated several strains (Alcanivorax, Alteromonas, Cycloclasticus, Halomonas, Marinobacter and Pseudoalteromonas) with demonstrable hydrocarbon-degrading qualities from surface slick and plume water samples collected during the active phase of the spill. Some of these organisms accounted for the majority of sequence reads representing their respective taxa in a pyrosequencing dataset constructed from the same and additional water column samples. Hitherto, Alcanivorax was not identified in any of the previous water column studies analysing the microbial response to the spill and we discuss its failure to respond to the oil. Collectively, our data provides unequivocal evidence on the hydrocarbon-degrading qualities for some of the dominant taxa enriched in surface and plume waters during the Deepwater Horizon oil spill, and a more complete understanding of their role in the fate of the oil.
A dynamic and complex monochloramine stress response in Escherichia coli revealed by transcriptome analysis2013 - Water Res., 47: 4978-4985
Despite the widespread use of monochloramine in drinking water treatment, there is surprisingly little information about its mode of action. In this study, DNA microarrays were used to investigate the global gene expression of Escherichia coli cells exposed to sub-lethal concentrations of monochloramine, with a focus on temporal dynamics. Genes induced by monochloramine were associated with several stress response functions, including oxidative stress, DNA repair, multidrug efflux, biofilm formation, antibiotic resistance, and cell wall repair. The diversity of functional associations supports a model of monochloramine action involving multiple cellular targets including cell membranes, nucleic acids, and proteins. These data suggest that E. coli responds to monochloramine exposure by activating diverse defense responses rather than a single antioxidant system and the exposure may also induce biofilm formation. The induction of multidrug efflux pumps and specific antibiotic resistance genes further suggests that exposure to monochloramine may contribute to reduced susceptibility to some antibiotics.
Phylotype-level 16S rRNA analysis reveals new bacterial indicators of health state in acute murine colitis2012 - ISME J., 6: 2091-106
Human inflammatory bowel disease and experimental colitis models in mice are associated with shifts in intestinal microbiota composition, but it is unclear at what taxonomic/phylogenetic level such microbiota dynamics can be indicative for health or disease. Here, we report that dextran sodium sulfate (DSS)-induced colitis is accompanied by major shifts in the composition and function of the intestinal microbiota of STAT1(-/-) and wild-type mice, as determined by 454 pyrosequencing of bacterial 16S rRNA (gene) amplicons, metatranscriptomics and quantitative fluorescence in situ hybridization of selected phylotypes. The bacterial families Ruminococcaceae, Bacteroidaceae, Enterobacteriaceae, Deferribacteraceae and Verrucomicrobiaceae increased in relative abundance in DSS-treated mice. Comparative 16S rRNA sequence analysis at maximum possible phylogenetic resolution identified several indicator phylotypes for DSS treatment, including the putative mucin degraders Akkermansia and Mucispirillum. The analysis additionally revealed strongly contrasting abundance changes among phylotypes of the same family, particularly within the Lachnospiraceae. These extensive phylotype-level dynamics were hidden when reads were grouped at higher taxonomic levels. Metatranscriptomic analysis provided insights into functional shifts in the murine intestinal microbiota, with increased transcription of genes associated with regulation and cell signaling, carbohydrate metabolism and respiration and decreased transcription of flagellin genes during inflammation. These findings (i) establish the first in-depth inventory of the mouse gut microbiota and its metatranscriptome in the DSS colitis model, (ii) reveal that family-level microbial community analyses are insufficient to reveal important colitis-associated microbiota shifts and (iii) support a scenario of shifting intra-family structure and function in the phylotype-rich and phylogenetically diverse Lachnospiraceae in DSS-treated mice.
Barcoded primers used in multiplex amplicon pyrosequencing bias amplification2011 - Appl. Environ. Microbiol., 77: 7846-7849
"Barcode-tagged" PCR primers used for multiplex amplicon sequencing generate a thus-far-overlooked amplification bias that produces variable terminal restriction fragment length polymorphism (T-RFLP) and pyrosequencing data from the same environmental DNA template. We propose a simple two-step PCR approach that increases reproducibility and consistently recovers higher genetic diversity in pyrosequencing libraries.
Systematic spatial bias in DNA microarray hybridization is caused by probe spot position-dependent variability in lateral diffusion2011 - PLoS One, 6: e23727
The hybridization of nucleic acid targets with surface-immobilized probes is a widely used assay for the parallel detection of multiple targets in medical and biological research. Despite its widespread application, DNA microarray technology still suffers from several biases and lack of reproducibility, stemming in part from an incomplete understanding of the processes governing surface hybridization. In particular, non-random spatial variations within individual microarray hybridizations are often observed, but the mechanisms underpinning this positional bias remain incompletely explained.
This study identifies and rationalizes a systematic spatial bias in the intensity of surface hybridization, characterized by markedly increased signal intensity of spots located at the boundaries of the spotted areas of the microarray slide. Combining observations from a simplified single-probe block array format with predictions from a mathematical model, the mechanism responsible for this biasis found to be a position-dependent variation in lateral diffusion of target molecules. Numerical simulations reveal a strong influence of microarraywell geometry on the spatial bias.
Reciprocal adjustment of the size of the microarray hybridization chamber to the area of surface-bound probes is a simple and effective measure to minimize or eliminate the diffusion-based bias, resulting in increased uniformity and accuracy of quantitative DNA microarrayhybridization.
Mycobacterium avium infections of Acanthamoeba strains: host strain variability, grazing-acquired infections, and altered dynamics of inactivation with monochloramine2010 - Appl. Environ. Microbiol., 76: 6685-6688
Stable Mycobacterium avium infections of several Acanthamoeba strains were characterized by increased infection resistance of recent environmental isolates and reduced infectivity in the presence of other bacteria. Exposure of M. avium in co-culture with Acanthamoeba castellanii to monochloramine yielded inactivation kinetics strikingly similar to those observed for A. castellanii alone.