Cryophilic algae thrive in liquid water within snow and ice in alpine and polar regions worldwide. Blooms of these algae lower albedo (reflection of sunlight), thereby altering melting patterns (Kohshima, Seko & Yoshimura, 1993; Lutz et al., 2014; Thomas & Duval, 1995). Here metagenomic DNA analysis and satellite imaging were used to investigate red snow in Franz Josef Land in the Russian Arctic. Franz Josef Land red snow metagenomes confirmed that the communities are composed of the autotroph Chlamydomonas nivalis that is supporting a complex viral and heterotrophic bacterial community. Comparisons with white snow communities from other sites suggest that white snow and ice are initially colonized by fungal-dominated communities and then succeeded by the more complex C. nivalis-heterotroph red snow. Satellite image analysis showed that red snow covers up to 80% of the surface of snow and ice fields in Franz Josef Land and globally. Together these results show that C. nivalis supports a local food web that is on the rise as temperatures warm, with potential widespread impacts on alpine and polar environments worldwide.
Red snow; Snow; Arctic; Watermelon snow; Viruses; Franz Josef Land; Phage; Metagenomics
A continuously mixed series of microbial communities inhabits various points of the respiratory tract, with community composition determined by distance from colonization sources, colonization rates, and extinction rates. Ecology and evolution theory developed in the context of biogeography is relevant to clinical microbiology and could reframe the interpretation of recent studies comparing communities from lung explant samples, sputum samples, and oropharyngeal swabs. We propose an island biogeography model of the microbial communities inhabiting different niches in human airways. Island biogeography as applied to communities separated by time and space is a useful parallel for exploring microbial colonization of healthy and diseased lungs, with the potential to inform our understanding of microbial community dynamics and the relevance of microbes detected in different sample types. In this perspective, we focus on the intermixed microbial communities inhabiting different regions of the airways of patients with cystic fibrosis.
We aim to understand the microbial ecology of noma (cancrum oris), a devastating ancient illness which causes severe facial disfigurement in>140,000 malnourished children every year. The cause of noma is still elusive. A chaotic mix of microbial infection, oral hygiene and weakened immune system likely contribute to the development of oral lesions. These lesions are a plausible entry point for unidentified microorganisms that trigger gangrenous facial infections. To catalog bacteria present in noma lesions and identify candidate noma-triggering organisms, we performed a cross-sectional sequencing study of 16S rRNA gene amplicons from sixty samples of gingival fluid from twelve healthy children, twelve children suffering from noma (lesion and healthy sites), and twelve children suffering from Acute Necrotizing Gingivitis (ANG) (lesion and healthy sites). Relative to healthy individuals, samples taken from lesions in diseased mouths were enriched with Spirochaetes and depleted for Proteobacteria. Samples taken from healthy sites of diseased mouths had proportions of Spirochaetes and Proteobacteria that were similar to healthy control individuals. Samples from noma mouths did not have a higher abundance of Fusobacterium, casting doubt on its role as a causative agent of noma. Microbial communities sampled from noma and ANG lesions were dominated by the same Prevotella intermedia OTU, which was much less abundant in healthy sites sampled from the same mouths. Multivariate analysis confirmed that bacterial communities in healthy and lesion sites were significantly different. Several OTUs in the Orders Erysipelotrichales, Clostridiales, Bacteroidales, and Spirochaetales were identified as indicators of noma, suggesting that one or more microbes within these Orders is associated with the development of noma lesions. Future studies should include longitudinal sampling of viral and microbial components of this community, before and early in noma lesion development.
Noma is a traumatic disease characterized by oral-facial lesions that often lead to severe disfigurement and ultimately shame and isolation from the community. Because the causes of noma are likely to be numerous, and reaching those who suffer from this illness is challenging, the etiology of noma remains ill-defined. Although it is known that oral hygiene and nutrition influence the development of noma, evidence suggests that one or more microbes play a crucial role in development of noma lesions. Previous studies have examined the DNA of microbes in lesions to determine which species are present and how their abundances differ between healthy mouth sites and noma lesions. These studies used techniques that were state-of-the-art at the time, though we know they likely only scratched the surface of the resident microbial diversity. Here we extend these studies by digging deeper to characterize a larger diversity of microbial species in noma and control samples, with the goal of better identifying which microbes are uniquely present or have altered abundances in noma lesions.
The airways of cystic fibrosis (CF) patients are chronically colonized by patient-specific polymicrobial communities. The conditions and nutrients available in CF lungs affect the physiology and composition of the colonizing microbes. Recent work in bioreactors has shown that the fermentation product 2,3-butanediol mediates cross-feeding between some fermenting bacteria and Pseudomonas aeruginosa, and that this mechanism increases bacterial current production. To examine bacterial fermentation in the respiratory tract, breath gas metabolites were measured and several metagenomes were sequenced from CF and non-CF volunteers. 2,3-butanedione was produced in nearly all respiratory tracts. Elevated levels in one patient decreased during antibiotic treatment, and breath concentrations varied between CF patients at the same time point. Some patients had high enough levels of 2,3-butanedione to irreversibly damage lung tissue. Antibiotic therapy likely dictates the activities of 2,3-butanedione-producing microbes, which suggests a need for further study with larger sample size. Sputum microbiomes were dominated by P. aeruginosa, Streptococcus spp. and Rothia mucilaginosa, and revealed the potential for 2,3-butanedione biosynthesis. Genes encoding 2,3-butanedione biosynthesis were disproportionately abundant in Streptococcus spp, whereas genes for consumption of butanedione pathway products were encoded by P. aeruginosa and R. mucilaginosa. We propose a model where low oxygen conditions in CF lung lead to fermentation and a decrease in pH, triggering 2,3-butanedione fermentation to avoid lethal acidification. We hypothesize that this may also increase phenazine production by P. aeruginosa, increasing reactive oxygen species and providing additional electron acceptors to CF microbes.
breath gas; cystic fibrosis; metagenomics; polymicrobial infection; metabolomics; biomarker
The cystic fibrosis (CF) lung contains thick mucus colonized by opportunistic pathogens which adapt to the CF lung environment over decades. The difficulty associated with sampling airways has impeded a thorough examination of the biochemical microhabitats these pathogens are exposed to. An indirect approach is to study the responses of microbial communities to these microhabitats, facilitated by high-throughput sequencing of microbial DNA and RNA from sputum samples. Microbial metagenomes and metatranscriptomes were sequenced from multiple CF patients, and the reads were assigned taxonomy and function through sequence homology to NCBI and the Kyoto Encyclopedia of Genes and Genomes (KEGG) database hierarchies. For a comparison, saliva microbial metagenomes from the Human Microbiome Project (HMP) were also analyzed. These analyses identified that functions encoded and expressed by CF microbes were significantly enriched for amino acid catabolism, folate biosynthesis, and lipoic acid biosynthesis. The data indicate that the community uses oxidative phosphorylation as a major energy source but that terminal electron acceptors were diverse. Nitrate reduction was the most abundant anaerobic respiratory pathway, and genes for nitrate reductase were largely assigned to Pseudomonas and Rothia. Although many reductive pathways of the nitrogen cycle were present, the cycle was incomplete, because the oxidative pathways were absent. Due to the abundant amino acid catabolism and incomplete nitrogen cycle, the CF microbial community appears to accumulate ammonia. This finding was verified experimentally using a CF bronchiole culture model system. The data also revealed abundant sensing and transport of iron, ammonium, zinc, and other metals along with a low-oxygen environment. This study reveals the core biochemistry and physiology of the CF microbiome.
The cystic fibrosis (CF) microbial community is complex and adapts to the environmental conditions of the lung over the lifetime of a CF patient. This analysis illustrates the core functions of the CF microbial community in the context of CF lung biochemistry. There are many studies of the metabolism and physiology of individual microbes within the CF lung, but none that collectively analyze data from the whole microbiome. Understanding the core metabolism of microbes that inhabit the CF lung can provide new targets for novel therapies. The fundamental processes that CF pathogens rely on for survival may represent an Achilles heel for this pathogenic community. Novel therapies that are designed to disrupt understudied survival strategies of the CF microbial community may succeed against otherwise untreatable or antibiotic-resistant microbes.
As DNA sequencing becomes faster and cheaper, genomics-based approaches are being explored for their use in personalized diagnoses and treatments. Here, we provide a proof of principle for disease monitoring using personal metagenomic sequencing and traditional clinical microbiology by focusing on three adults with cystic fibrosis (CF). The CF lung is a dynamic environment that hosts a complex ecosystem composed of bacteria, viruses, and fungi that can vary in space and time. Not surprisingly, the microbiome data from the induced sputum samples we collected revealed a significant amount of species diversity not seen in routine clinical laboratory cultures. The relative abundances of several species changed as clinical treatment was altered, enabling the identification of the climax and attack communities that were proposed in an earlier work. All patient microbiomes encoded a diversity of mechanisms to resist antibiotics, consistent with the characteristics of multidrug-resistant microbial communities that are commonly observed in CF patients. The metabolic potentials of these communities differed by the health status and recovery route of each patient. Thus, this pilot study provides an example of how metagenomic data might be used with clinical assessments for the development of treatments tailored to individual patients.
The impaired mucociliary clearance in individuals with Cystic Fibrosis (CF) enables opportunistic pathogens to colonize CF lungs. Here we show that Rothia mucilaginosa is a common CF opportunist that was present in 83% of our patient cohort, almost as prevalent as Pseudomonas aeruginosa (89%). Sequencing of lung microbial metagenomes identified unique R. mucilaginosa strains in each patient, presumably due to evolution within the lung. The de novo assembly of a near-complete R. mucilaginosa (CF1E) genome illuminated a number of potential physiological adaptations to the CF lung, including antibiotic resistance, utilization of extracellular lactate, and modification of the type I restriction-modification system. Metabolic characteristics predicted from the metagenomes suggested R. mucilaginosa have adapted to live within the microaerophilic surface of the mucus layer in CF lungs. The results also highlight the remarkable evolutionary and ecological similarities of many CF pathogens; further examination of these similarities has the potential to guide patient care and treatment.
Multiple studies have demonstrated that early-life exposure to pets or siblings affords protection against allergic disease; these associations are commonly attributed to the “hygiene hypothesis”. Recently, low diversity of the infant gut microbiota has also been linked to allergic disease. In this study, we characterize the infant gut microbiota in relation to pets and siblings.
The study population comprised a small sub-sample of 24 healthy, full term infants from the Canadian Healthy Infant Longitudinal Development (CHILD) birth cohort. Mothers reported on household pets and siblings. Fecal samples were collected at 4 months of age, and microbiota composition was characterized by high-throughput signature gene sequencing.
Microbiota richness and diversity tended to be increased in infants living with pets, whereas these measures were decreased in infants with older siblings. Infants living with pets exhibited under-representation of Bifidobacteriaceae and over-representation of Peptostreptococcaceae; infants with older siblings exhibited under-representation of Peptostreptococcaceae.
This study provides new evidence that exposure to pets and siblings may influence the early development of the gut microbiota, with potential implications for allergic disease. These two traditionally protective “hygiene hypothesis” factors appear to differentially impact gut microbiota composition and diversity, calling into question the clinical significance of these measures. Further research is required to confirm and expand these findings.
Infants; Gut microbiota; Gut microbiome; Hygiene hypothesis; Microflora hypothesis; Pets; Siblings; Atopy; Allergic disease; Environmental exposures
Exogenous glucagon-like peptide-2 receptor (GLP-2R) activation elicits proliferative and cytoprotective responses in the gastrointestinal mucosa and ameliorates experimental small and large bowel gut injury. Nevertheless, the essential physiological role(s) of the endogenous GLP-2R remain poorly understood. We studied the importance of the GLP-2R for gut growth, epithelial cell lineage allocation, the response to mucosal injury, and host-bacterial interactions in Glp2r−/− and littermate control Glp2r+/+ mice. Glp2r−/− mice exhibit normal somatic growth and preserved small and large bowel responses to IGF-I and keratinocyte growth factor. However, Glp2r−/− mice failed to up-regulate intestinal epithelial c-fos expression in response to acute GLP-2 administration and do not exhibit changes in small bowel conductance or small or large bowel growth after administration of GLP-2R agonists. The crypt and villus compartment and the numbers and localization of Paneth, enteroendocrine, and goblet cells were comparable in Glp2r+/+ vs. Glp2r−/− mice. Although the severity and extent of colonic mucosal injury in response to 3% oral dextran sulfate was similar across Glp2r genotypes, Glp2r−/− mice exhibited significantly increased morbidity and mortality and increased bacterial translocation after induction of enteritis with indomethacin and enhanced mucosal injury in response to irinotecan. Moreover, bacterial colonization of the small bowel was significantly increased, expression of Paneth cell antimicrobial gene products was reduced, and mucosal bactericidal activity was impaired in Glp2r−/− mice. Although the Glp2r is dispensable for gut development and the response to colonic injury, Glp2r−/− mice exhibit enhanced sensitivity to small bowel injury, and abnormal host-bacterial interactions in the small bowel.
PMID: 22253424 CAMSID: cams2742
The gut microbiota is essential to human health throughout life, yet the acquisition and development of this microbial community during infancy remains poorly understood. Meanwhile, there is increasing concern over rising rates of cesarean delivery and insufficient exclusive breastfeeding of infants in developed countries. In this article, we characterize the gut microbiota of healthy Canadian infants and describe the influence of cesarean delivery and formula feeding.
We included a subset of 24 term infants from the Canadian Healthy Infant Longitudinal Development (CHILD) birth cohort. Mode of delivery was obtained from medical records, and mothers were asked to report on infant diet and medication use. Fecal samples were collected at 4 months of age, and we characterized the microbiota composition using high-throughput DNA sequencing.
We observed high variability in the profiles of fecal microbiota among the infants. The profiles were generally dominated by Actinobacteria (mainly the genus Bifidobacterium) and Firmicutes (with diverse representation from numerous genera). Compared with breastfed infants, formula-fed infants had increased richness of species, with overrepresentation of Clostridium difficile. Escherichia–Shigella and Bacteroides species were underrepresented in infants born by cesarean delivery. Infants born by elective cesarean delivery had particularly low bacterial richness and diversity.
These findings advance our understanding of the gut microbiota in healthy infants. They also provide new evidence for the effects of delivery mode and infant diet as determinants of this essential microbial community in early life.
The characterization of bacterial communities using DNA sequencing has revolutionized our ability to study microbes in nature and discover the ways in which microbial communities affect ecosystem functioning and human health. Here we describe Serial Illumina Sequencing (SI-Seq): a method for deep sequencing of the bacterial 16S rRNA gene using next-generation sequencing technology. SI-Seq serially sequences portions of the V5, V6 and V7 hypervariable regions from barcoded 16S rRNA amplicons using an Illumina short-read genome analyzer. SI-Seq obtains taxonomic resolution similar to 454 pyrosequencing for a fraction of the cost, and can produce hundreds of thousands of reads per sample even with very high multiplexing. We validated SI-Seq using single species and mock community controls, and via a comparison to cystic fibrosis lung microbiota sequenced using 454 FLX Titanium. Our control runs show that SI-Seq has a dynamic range of at least five orders of magnitude, can classify >96% of sequences to the genus level, and performs just as well as 454 and paired-end Illumina methods in estimation of standard microbial ecology diversity measurements. We illustrate the utility of SI-Seq in a pilot sample of central airway secretion samples from cystic fibrosis patients.
Background. Recurrent bacterial infections play a key role in the pathogenesis of bronchiectasis, but conventional microbiologic methods may fail to identify pathogens in many cases. We characterized and compared the pulmonary bacterial communities of cystic fibrosis (CF) and non-CF bronchiectasis patients using a culture-independent molecular approach. Methods. Bacterial 16S rRNA gene libraries were constructed from lung tissue of 10 non-CF bronchiectasis and 21 CF patients, followed by DNA sequencing of isolates from each library. Community characteristics were analyzed and compared between the two groups. Results. A wide range of bacterial diversity was detected in both groups, with between 1 and 21 bacterial taxa found in each patient. Pseudomonas was the most common genus in both groups, comprising 49% of sequences detected and dominating numerically in 13 patients. Although Pseudomonas appeared to be dominant more often in CF patients than in non-CF patients, analysis of entire bacterial communities did not identify significant differences between these two groups. Conclusions. Our data indicate significant diversity in the pulmonary bacterial community of both CF and non-CF bronchiectasis patients and suggest that this community is similar in surgically resected lungs of CF and non-CF bronchiectasis patients.
Five batch cultures of Bacillus subtilis were subjected to evolution in the laboratory for 6,000 generations under conditions repressing sporulation in complex liquid medium containing glucose. Between generations 1,000 and 2,000, variants with a distinct small-colony morphology arose and swept through four of the five populations that had been previously noted for their loss of sporulation (H. Maughan et al., Genetics 177:937-948, 2007). To better understand the nature of adaptation in these variants, individual strains were isolated from one population before (WN715) and after (WN716) the sweep. In addition to colony morphology, strains WN715 and WN716 differed in their motility, aerotaxis, and cell morphology. Competition experiments showed that strain WN716 had evolved a distinct fitness advantage over the ancestral strain and strain WN715 during growth and the transition to the postexponential growth phase, which was more pronounced when WN715 was present in the coculture. Microarray analyses revealed candidate genes in which mutations may have produced some of the observed phenotypes. For example, loss of motility in WN716 was accompanied by decreased transcription of all flagellar, motility, and chemotaxis genes on the microarray. Transcription of alsS and alsD was also lower in strain WN716, and the predicted loss of acetoin production and enhanced acetate production was confirmed by high-performance liquid chromatography (HPLC) analysis. The results suggested that the derived colony morphology of strain WN716 was associated with increased fitness, the alteration of several metabolic pathways, and the loss of a typical postexponential-phase response.
We used microarrays to identify the causes of sporulation deficiencies in Bacillus subtilis after 6,000 generations of evolution. We found that sporulation loss did not result from large-scale deletions; therefore, it must have resulted from smaller indels and/or substitutions. Transcription patterns of one strain versus its ancestor showed that sporulation was not initiated and suggested that sporulation loss may be part of an overall decline in plasticity.
Natural competence is the genetically encoded ability of some bacteria to take up DNA from the environment. Although most of the incoming DNA is degraded, occasionally intact homologous fragments can recombine with the chromosome, displacing one resident strand. This potential to use DNA as a source of both nutrients and genetic novelty has important implications for the ecology and evolution of competent bacteria. However, it is not known how frequently competence changes during evolution, or whether non-competent strains can persist for long periods of time. We have previously studied competence in H. influenzae and found that both the amount of DNA taken up and the amount recombined varies extensively between different strains. In addition, several strains are unable to become competent, suggesting that competence has been lost at least once. To investigate how many times competence has increased or decreased during the divergence of these strains, we inferred the evolutionary relationships of strains using the largest datasets currently available. However, despite the use of three datasets and multiple inference methods, few nodes were resolved with high support, perhaps due to extensive mixing by recombination. Tracing the evolution of competence in those clades that were well supported identified changes in DNA uptake and/or transformation in most strains. The recency of these events suggests that competence has changed frequently during evolution but the poor support of basal relationships precludes the determination of whether non-competent strains can persist for long periods of time. In some strains, changes in transformation have occurred that cannot be due to changes in DNA uptake, suggesting that selection can act on transformation independent of DNA uptake.
Adaptive plasticity allows populations to adjust rapidly to environmental
change. If this is useful only rarely, plasticity may undergo mutational
degradation and be lost from a population. We consider a population of constant
size N undergoing loss of plasticity at functional mutation
rate m and with selective advantage s
associated with loss. Environmental change events occur at rate θ
per generation, killing all individuals that lack plasticity. The expected time
until loss of plasticity in a fluctuating environment is always at least τ¯, the expected time until loss of plasticity in a
static environment. When mN > 1 and
Nθ >> 1, we find that plasticity
will be maintained for an average of at least 108 generations in a
single population provided τ¯ > 18/θ. In a metapopulation, plasticity is
retained under the more lenient condition τ¯ > 1.3/θ, irrespective of
mN, for a modest number of demes. We calculate both exact and
approximate solutions for τ¯ and find that it is linearly dependent on only the logarithm
of N, and so surprisingly, both the population size and the
number of demes in the metapopulation make little difference to the retention of
plasticity. Instead, τ¯ is dominated by the term 1/(m +
population genetics; Moran model; fluctuating environment; phenotypic plasticity; regressive evolution
Previously, spontaneous rifampin resistance mutations were isolated in cluster I of the rpoB gene, resulting in amino acid replacements (Q469R, H482R, H482Y, or S487L) in the Bacillus subtilis RNA polymerase β subunit (W. L. Nicholson and H. Maughan, J. Bacteriol. 184:4936-4940, 2002). In this study, each amino acid change in the β subunit was observed to result in its own unique spectrum of effects on growth and various developmental events, including sporulation, germination, and competence for transformation. The results thus establish the important role played by the RNA polymerase β subunit, not only in the catalytic aspect of transcription, but also in the regulation of major developmental events in B. subtilis.
It has recently been proposed that phenotypic variation in clonal populations of bacterial species results from intracellular “noise,” i.e., random fluctuations in levels of cellular molecules, which would be predicted to be insensitive to selective pressure. To test this notion, we propagated five populations of Bacillus subtilis for 5,000 generations with selection for one phenotype: the decision to sporulate. In support of the noise hypothesis, we report that none of the populations responded to selection by improving their efficiency of sporulation, indicating that intracellular noise is independent of heritable genotype.
Mutations causing rifampin resistance in vegetative cells of Bacillus subtilis 168 have thus far been mapped to a rather restricted set of alterations at either Q469 or H482 within cluster I of the rpoB gene encoding the β subunit of RNA polymerase. In this study, we demonstrated that spores of B. subtilis 168 exhibit a spectrum of spontaneous rifampin resistance mutations distinct from that of vegetative cells. In addition to the rpoB mutations Q469K, Q469R, and H482Y previously characterized in vegetative cells, we isolated a new mutation of rpoB, H482R, from vegetative cells. Additional new rifampin resistance mutations arising from spores were detected at A478N and most frequently at S487L. The S487L change is the predominant change found in rpoB mutations sequenced from rifampin-resistant clinical isolates of Mycobacterium tuberculosis. The observations are discussed in terms of the underlying differences of the DNA environment within dormant cells and vegetatively growing cells.