The parasite Plasmodium falciparum is responsible for hundreds of millions of cases of malaria, and kills more than one million African children annually. Here we report an analysis of the genome sequence of P. falciparum clone 3D7. The 23-megabase nuclear genome consists of 14 chromosomes, encodes about 5,300 genes, and is the most (A + T)-rich genome sequenced to date. Genes involved in antigenic variation are concentrated in the subtelomeric regions of the chromosomes. Compared to the genomes of free-living eukaryotic microbes, the genome of this intracellular parasite encodes fewer enzymes and transporters, but a large proportion of genes are devoted to immune evasion and host–parasite interactions. Many nuclear-encoded proteins are targeted to the apicoplast, an organelle involved in fatty-acid and isoprenoid metabolism. The genome sequence provides the foundation for future studies of this organism, and is being exploited in the search for new drugs and vaccines to fight malaria.
Elucidating the biogeography of bacterial communities on the human body is critical for establishing healthy baselines from which to detect differences associated with disease; however, little is known about the baseline bacterial profiles from various human habitats of healthy Chinese undergraduates.
Using parallel barcoded 454 pyrosequencing targeting on the 16S rRNA gene V3 region, the bacterial diversity of the nasopharynx, saliva, dominant hands, and feces were investigated from 10 healthy Chinese junior boarding undergraduates at Zhejiang University. The participants were 21–24 years of age with a body mass index (BMI) < 24 kg/m2. A total of 156,717 high-quality pyrosequencing reads were obtained for evaluating bacterial diversity, which represented 29,887 unique phylotypes. The overall taxonomic distribution of the 16S rRNA gene-based amplicons demonstrated that these 4 habitats of the human body harbored distinct microbiota and could be divided into different clusters according to anatomic site, while the established patterns of bacterial diversity followed the human body habitat (feces, hands, saliva, and nasopharynx). Although significant inter-individual variation was observed, the healthy microbiota still shared a large number of phylotypes in each habitat, but not among the four habitats, indicating that a core microbiome existed in each healthy habitat. The vast majority of sequences from these different habitats were classified into different taxonmies that became the predominant bacteria of the healthy microbiota.
We first established the framework of microbial communities from four healthy human habitats of the same participants with similar living environments for the Chinese undergraduates. Our data represent an important step for determining the diversity of Chinese healthy microbiota, and can be used for more large-scale studies that focus on the interactions between healthy and diseases states for young Chinese adults in the same age range.
Healthy microbiota; Saliva; Nasopharynx; Feces; Dominant hand; Pyrosequencing; Core microbiome
Background & Aims
Intestinal dysbiosis and bacterial translocation is common in patients with advanced liver disease, and there is strong evidence that the translocation of bacteria and their products across the epithelial barrier drives experimental liver disease progression. The aims of our study were to investigate dynamics of bacterial translocation and changes in the enteric microbiome in early stages of liver disease.
Cholestatic liver injury was induced by ligation of the common bile duct (BDL) and toxic liver injury by injection of carbon tetrachloride (CCl4) in mice.
Increased intestinal permeability and bacterial translocation occurred one day following liver injury in both disease models. This was accompanied by decreased intestinal expression of the tight junction protein occludin. Although BDL resulted in a rapid onset of intestinal bacterial overgrowth, bacterial overgrowth was observed in mice injected with CCl4 only in advanced stages of liver fibrosis. To further assess the qualitative changes in the intestinal microbiome, massively parallel pyrosequencing of 16S rRNA genes revealed minor microbial changes following BDL, while CCl4 administration resulted in a relative abundance of Firmicutes and Actinobacteria compared with oil injected mice. Four different liver disease models (cholestasis, toxic, alcohol, obesity) show few similarities in their intestinal microbiome.
Acute liver injury is associated with an early onset of increased intestinal permeability and bacterial translocation that precede changes in the microbiome. The enteric microbiome differs with respect to the etiology of liver disease.
bacterial overgrowth; microbiome; dysbiosis; bacterial translocation; 16S rRNA sequencing
Enterococcus faecalis is commonly isolated from the gastrointestinal tract of healthy infants and adults, where it contributes to host health and well-being. We describe here the draft genome sequence of E. faecalis PC1.1, a candidate probiotic strain isolated from human feces.
Bacterial vaginosis (BV) is the most common vaginal disorder of reproductive-age women. Yet the cause of BV has not been established. To uncover key determinants of BV, we employed a multi-omic, systems-biology approach, including both deep 16S rRNA gene-based sequencing and metabolomics of lavage samples from 36 women. These women varied demographically, behaviorally, and in terms of health status and symptoms.
16S rRNA gene-based community composition profiles reflected Nugent scores, but not Amsel criteria. In contrast, metabolomic profiles were markedly more concordant with Amsel criteria. Metabolomic profiles revealed two distinct symptomatic BV types (SBVI and SBVII) with similar characteristics that indicated disruption of epithelial integrity, but each type was correlated to the presence of different microbial taxa and metabolites, as well as to different host behaviors. The characteristic odor associated with BV was linked to increases in putrescine and cadaverine, which were both linked to Dialister spp. Additional correlations were seen with the presence of discharge, 2-methyl-2-hydroxybutanoic acid, and Mobiluncus spp., and with pain, diethylene glycol and Gardnerella spp.
The results not only provide useful diagnostic biomarkers, but also may ultimately provide much needed insight into the determinants of BV.
Targeted drugs have significantly improved the therapeutic options for advanced renal cell carcinoma (RCC). However, resistance often develops, negating the benefit of these agents. In the present study, the molecular mechanisms of acquired resistance towards the histone deacetylase (HDAC) inhibitor valproic acid (VPA) in a RCC in vivo model were investigated. NMRI:nu/nu mice were transplanted with Caki-1 RCC cells and then treated with VPA (200 mg/kg/day). Controls remained untreated. Based on tumor growth dynamics, the mice were divided into “responders” and “non-responders” to VPA. Histone H3 and H4 acetylation and expression of cell signaling and cell cycle regulating proteins in the RCC mouse tumors were evaluated by Western blotting. Tumor growth of VPA responders was significantly diminished, whereas that of VPA non-responders even exceeded control values. Cdk1, 2 and 4 proteins were strongly enhanced in the non-responders. Importantly, Akt expression and activity were massively up-regulated in the tumors of the VPA non-responders. Chronic application (12 weeks) of VPA to Caki-1 cells in vitro evoked a distinct elevation of Akt activity and cancer cells no longer responded with cell growth reduction, compared to the short 2 week treatment. We assume that chronic use of an HDAC-inhibitor is associated with (re)-activation of Akt, which may be involved in resistance development. Consequently, combined blockade of both HDAC and Akt may delay or prevent drug resistance in RCC.
The pathophysiologic mechanisms behind urologic disease are increasingly being elucidated. The object of this investigation was to evaluate the publication policies of urologic journals during a period of progressively better understanding and management of urologic disease. Based on the ISI Web of Knowledge Journal Citation Reports and the PubMed database, the number and percentage of original experimental, original clinical, review or commentarial articles published between 2002–2010 in six leading urologic journals were analyzed. “British Journal of Urology International”, “European Urology”, “Urologic Oncology-Seminars and Original Investigations” (“Urologic Oncology”), “Urology”, “The Journal of Urology”, and “World Journal of Urology” were chosen, because these journals publish articles in all four categories. The publication policies of the six journals were very heterogeneous during the time period from 2002 to 2010. The percentage of original experimental and original clinical articles, related to all categories, remained the same in “British Journal of Urology International”, “Urologic Oncology”, “Urology” and “The Journal of Urology”. The percentage of experimental reports in “World Journal of Urology” between 2002–2010 significantly increased from 10 to 20%. A distinct elevation in the percentage of commentarial articles accompanied by a reduction of clinical articles became evident in “European Urology” which significantly correlated with a large increase in the journal’s impact factor. No clearly superior policy could be identified with regard to a general increase in the impact factors from all the journals. The publication policy of urologic journals does not expressly reflect the increase in scientific knowledge, which has occurred over the period 2002–2010. One way of increasing the exposure of urologists to research and expand the interface between experimental and clinical research, would be to enlarge the percentage of experimental articles published. There is no indication that such policy would be detrimental to a journal’s impact factor.
A variety of microbial communities and their genes (microbiome) exist throughout the human body, playing fundamental roles in human health and disease. The NIH funded Human Microbiome Project (HMP) Consortium has established a population-scale framework which catalyzed significant development of metagenomic protocols resulting in a broad range of quality-controlled resources and data including standardized methods for creating, processing and interpreting distinct types of high-throughput metagenomic data available to the scientific community. Here we present resources from a population of 242 healthy adults sampled at 15 to 18 body sites up to three times, which to date, have generated 5,177 microbial taxonomic profiles from 16S rRNA genes and over 3.5 Tb of metagenomic sequence. In parallel, approximately 800 human-associated reference genomes have been sequenced. Collectively, these data represent the largest resource to date describing the abundance and variety of the human microbiome, while providing a platform for current and future studies.
A combination of Sanger and 454 sequences of small subunit rRNA loci were used to interrogate microbial diversity in the bovine rumen of 12 cows consuming a forage diet. Observed bacterial species richness, based on the V1–V3 region of the 16S rRNA gene, was between 1,903 to 2,432 species-level operational taxonomic units (OTUs) when 5,520 reads were sampled per animal. Eighty percent of species-level OTUs were dominated by members of the order Clostridiales, Bacteroidales, Erysipelotrichales and unclassified TM7. Abundance of Prevotella species varied widely among the 12 animals. Archaeal species richness, also based on 16S rRNA, was between 8 and 13 OTUs, representing 5 genera. The majority of archaeal OTUs (84%) found in this study were previously observed in public databases with only two new OTUs discovered. Observed rumen fungal species richness, based on the 18S rRNA gene, was between 21 and 40 OTUs with 98.4–99.9% of OTUs represented by more than one read, using Good’s coverage. Examination of the fungal community identified numerous novel groups. Prevotella and Tannerella were overrepresented in the liquid fraction of the rumen while Butyrivibrio and Blautia were significantly overrepresented in the solid fraction of the rumen. No statistical difference was observed between the liquid and solid fractions in biodiversity of archaea and fungi. The survey of microbial communities and analysis of cross-domain correlations suggested there is a far greater extent of microbial diversity in the bovine rumen than previously appreciated, and that next generation sequencing technologies promise to reveal novel species, interactions and pathways that can be studied further in order to better understand how rumen microbial community structure and function affects ruminant feed efficiency, biofuel production, and environmental impact.
Comparative genomic hybridization was used to compare genetic diversity of five strains of Leptospira (Leptospira interrogans serovars Bratislava, Canicola, and Hebdomadis and Leptospira kirschneri serovars Cynopteri and Grippotyphosa). The array was designed based on two available sequenced Leptospira reference genomes, those of L. interrogans serovar Copenhageni and L. interrogans serovar Lai. A comparison of genetic contents showed that L. interrogans serovar Bratislava was closest to the reference genomes while L. kirschneri serovar Grippotyphosa had the least similarity to the reference genomes. Cluster analysis indicated that L. interrogans serovars Bratislava and Hebdomadis clustered together first, followed by L. interrogans serovar Canicola, before the two L. kirschneri strains. Confirmed/potential virulence factors identified in previous research were also detected in the tested strains.
Infections of the vaginal tract result from perturbations in the complex interactions between the microbiome and the host vaginal ecosystem. Recent data have linked specific vaginal microbes and urogenital infection with pre-term birth. Here we discuss how next generation sequencing-based approaches to study the vaginal microbiome will be important for defining what constitutes an imbalance of the microbiome and the associated host conditions that lead to subsequent infection and disease states. These studies will provide clinicians reliable diagnostic tools and treatments for women who are at increased risk for vaginal infections, preterm birth, HIV and other sexually acquired diseases, and will provide opportunities for intervention.
Clinical dogma is that healthy urine is sterile and the presence of bacteria with an inflammatory response is indicative of urinary tract infection (UTI). Asymptomatic bacteriuria (ABU) represents the state in which bacteria are present but the inflammatory response is negligible. Differentiating ABU from UTI is diagnostically challenging, but critical because overtreatment of ABU can perpetuate antimicrobial resistance while undertreatment of UTI can result in increased morbidity and mortality. In this study, we describe key characteristics of the healthy and ABU urine microbiomes utilizing 16S rRNA gene (16S rDNA) sequencing and metaproteomics, with the future goal of utilizing this information to personalize the treatment of UTI based on key individual characteristics.
A cross-sectional study of 26 healthy controls and 27 healthy subjects at risk for ABU due to spinal cord injury-related neuropathic bladder (NB) was conducted. Of the 27 subjects with NB, 8 voided normally, 8 utilized intermittent catheterization, and 11 utilized indwelling Foley urethral catheterization for bladder drainage. Urine was obtained by clean catch in voiders, or directly from the catheter in subjects utilizing catheters. Urinalysis, urine culture and 16S rDNA sequencing were performed on all samples, with metaproteomic analysis performed on a subsample.
A total of 589454 quality-filtered 16S rDNA sequence reads were processed through a NextGen 16S rDNA analysis pipeline. Urine microbiomes differ by normal bladder function vs. NB, gender, type of bladder catheter utilized, and duration of NB. The top ten bacterial taxa showing the most relative abundance and change among samples were Lactobacillales, Enterobacteriales, Actinomycetales, Bacillales, Clostridiales, Bacteroidales, Burkholderiales, Pseudomonadales, Bifidobacteriales and Coriobacteriales. Metaproteomics confirmed the 16S rDNA results, and functional human protein-pathogen interactions were noted in subjects where host defenses were initiated.
Counter to clinical belief, healthy urine is not sterile. The healthy urine microbiome is characterized by a preponderance of Lactobacillales in women and Corynebacterium in men. The presence and duration of NB and method of urinary catheterization alter the healthy urine microbiome. An integrated approach of 16S rDNA sequencing with metaproteomics improves our understanding of healthy urine and facilitates a more personalized approach to prevention and treatment of infection.
Bacteriuria; Urine; Catheter; Neuropathic; Bladder; Microbiome; Metaproteome; Next-generation; Personalized; rRNA
This manuscript describes the NIH Human Microbiome Project, including a brief review of human microbiome research, a history of the project, and a comprehensive overview of the consortium's recent collection of publications analyzing the human microbiome.
This study is the first to use a metagenomics approach to characterize the phylogeny and functional capacity of the canine gastrointestinal microbiome. Six healthy adult dogs were used in a crossover design and fed a low-fiber control diet (K9C) or one containing 7.5% beet pulp (K9BP). Pooled fecal DNA samples from each treatment were subjected to 454 pyrosequencing, generating 503 280 (K9C) and 505 061 (K9BP) sequences. Dominant bacterial phyla included the Bacteroidetes/Chlorobi group and Firmicutes, both of which comprised ∼35% of all sequences, followed by Proteobacteria (13–15%) and Fusobacteria (7–8%). K9C had a greater percentage of Bacteroidetes, Fusobacteria and Proteobacteria, whereas K9BP had greater proportions of the Bacteroidetes/Chlorobi group and Firmicutes. Archaea were not altered by diet and represented ∼1% of all sequences. All archaea were members of Crenarchaeota and Euryarchaeota, with methanogens being the most abundant and diverse. Three fungi phylotypes were present in K9C, but none in K9BP. Less than 0.4% of sequences were of viral origin, with >99% of them associated with bacteriophages. Primary functional categories were not significantly affected by diet and were associated with carbohydrates; protein metabolism; DNA metabolism; cofactors, vitamins, prosthetic groups and pigments; amino acids and derivatives; cell wall and capsule; and virulence. Hierarchical clustering of several gastrointestinal metagenomes demonstrated phylogenetic and metabolic similarity between dogs, humans and mice. More research is required to provide deeper coverage of the canine microbiome, evaluate effects of age, genetics or environment on its composition and activity, and identify its role in gastrointestinal disease.
canine gut; gastrointestinal bacteria; metagenomics; pyrosequencing
Thermotoga sp. strain RQ2 is probably a strain of Thermotoga maritima. Its complete genome sequence allows for an examination of the extent and consequences of gene flow within Thermotoga species and strains. Thermotoga sp. RQ2 differs from T. maritima in its genes involved in myo-inositol metabolism. Its genome also encodes an apparent fructose phosphotransferase system (PTS) sugar transporter. This operon is also found in Thermotoga naphthophila strain RKU-10 but no other Thermotogales. These are the first reported PTS transporters in the Thermotogales.
With growing experience, the indications for salvage extracorporeal membrane oxygenation continue to expand. We describe a successful application of extracorporeal support in a polytrauma patient presenting with profound hypothermia, respiratory failure, and whom was later found to have an intracranial hemorrhage. We advocate the role of salvage therapy even in patients with complex pathophysiology despite perceived relative or absolute contraindications to extracorporeal support.
A steep drop in the cost of next-generation sequencing during recent years has made the technology affordable to the majority of researchers, but downstream bioinformatic analysis still poses a resource bottleneck for smaller laboratories and institutes that do not have access to substantial computational resources. Sequencing instruments are typically bundled with only the minimal processing and storage capacity required for data capture during sequencing runs. Given the scale of sequence datasets, scientific value cannot be obtained from acquiring a sequencer unless it is accompanied by an equal investment in informatics infrastructure.
Cloud BioLinux is a publicly accessible Virtual Machine (VM) that enables scientists to quickly provision on-demand infrastructures for high-performance bioinformatics computing using cloud platforms. Users have instant access to a range of pre-configured command line and graphical software applications, including a full-featured desktop interface, documentation and over 135 bioinformatics packages for applications including sequence alignment, clustering, assembly, display, editing, and phylogeny. Each tool's functionality is fully described in the documentation directly accessible from the graphical interface of the VM. Besides the Amazon EC2 cloud, we have started instances of Cloud BioLinux on a private Eucalyptus cloud installed at the J. Craig Venter Institute, and demonstrated access to the bioinformatic tools interface through a remote connection to EC2 instances from a local desktop computer. Documentation for using Cloud BioLinux on EC2 is available from our project website, while a Eucalyptus cloud image and VirtualBox Appliance is also publicly available for download and use by researchers with access to private clouds.
Cloud BioLinux provides a platform for developing bioinformatics infrastructures on the cloud. An automated and configurable process builds Virtual Machines, allowing the development of highly customized versions from a shared code base. This shared community toolkit enables application specific analysis platforms on the cloud by minimizing the effort required to prepare and maintain them.
Although Bacteroides vulgatus is one of the most prevalent microorganisms in the human gastrointestinal tract, little is known about the genetic potential of this species. Here, we describe the annotated draft genome sequence of B. vulgatus PC510 isolated from human feces.
The translocation of bacteria and bacterial products into the circulation contributes to alcoholic liver disease. Intestinal bacterial overgrowth is common in patients with alcoholic liver disease. The aims of our study were to investigate bacterial translocation, changes in the enteric microbiome, and its regulation by mucosal antimicrobial proteins in alcoholic liver disease. We used a mouse model of continuous intragastric feeding of alcohol or an isocaloric diet. Bacterial translocation occurred prior to changes observed in the microbiome. Quantitative changes in the intestinal microflora of these animals were assessed first by conventional culture techniques in the small and large intestine. Although we found no difference after 1 day or 1 week, intestinal bacterial overgrowth was observed in the gastrointestinal tract of mice fed alcohol for 3 weeks as compared to control liquid diet fed mice. Because less than 20% of all gastrointestinal bacteria are able to be cultured by conventional methodologies, we performed massively parallel pyrosequencing to further assess the qualitative changes in the intestinal microbiome following alcohol exposure. Sequencing of 16S rRNA genes revealed a relative abundance of Bacteroidetes and Verrucomicrobia bacteria in mice fed alcohol compared with a relative predominance of Firmicutes bacteria in control mice. With respect to the host’s transcriptome, alcohol feeding was associated with downregulation in gene and protein expression of bactericidal c-type lectins Reg3b and Reg3g in the small intestines. Treatment with prebiotics partially restored Reg3g protein levels, reduced bacterial overgrowth and lessened alcoholic steatohepatitis. In conclusion, alcohol feeding is associated with intestinal bacterial overgrowth and enteric dysbiosis. Intestinal antimicrobial molecules are dysregulated following chronic alcohol feeding contributing to changes in the enteric microbiome and to alcoholic steatohepatitis.
alcoholic liver disease; microbiome; dysbiosis; bacterial translocation
The primate order began radiating ∼ 55 million years ago and is today represented by more than 300 extant species. These species have adapted to inhabit a vast gradation of environments, consume a broad range of diets, come in a wide spectra of anatomical designs and exhibit myriad social structures. Determining the relationships among these variables, primate health and the taxonomic and functional compositions of their associated microbial ecosystems holds enormous potential to understand the forces shaping or perturbing our own microbiomes, and may offer clues to the evolved relationship between humans and these ecosystems. In addition, it may help to better direct conservation efforts toward the 48% of primates currently endangered. With these goals in mind we have spent the past few years cataloguing and exploring the gastrointestinal and vaginal microbial ecosystems of mostly wild primates. Functionally and compositionally microbiomes show clear delineation between host species and for the GI system between captive and wilds. Habitat quality affects foraging behavior impacting the microbiome with strong ties to host health. Several observations also hint at evolved traits that would benefit the host physiology.
The bovine rumen maintains a diverse microbial community that serves to break down indigestible plant substrates. However, those bacteria specifically adapted to degrade cellulose, the major structural component of plant biomass, represent a fraction of the rumen microbiome. Previously, we proposed scaC as a candidate for phylotyping Ruminococcus flavefaciens, one of three major cellulolytic bacterial species isolated from the rumen. In the present report we examine the dynamics and diversity of scaC-types both within and between cattle temporally, following a dietary switch from corn-silage to grass-legume hay. These results were placed in the context of the overall bacterial population dynamics measured using the 16S rRNA.
As many as 117 scaC-types were estimated, although just nineteen were detected in each of three rumens tested, and these collectively accounted for the majority of all types present. Variation in scaC populations was observed between cattle, between planktonic and fiber-associated fractions and temporally over the six-week survey, and appeared related to scaC phylogeny. However, by the sixth week no significant separation of scaC populations was seen between animals, suggesting enrichment of a constrained set of scaC-types. Comparing the amino-acid translation of each scaC-type revealed sequence variation within part of the predicted dockerin module but strong conservation in the N-terminus, where the cohesin module is located.
The R. flavefaciens species comprises a multiplicity of scaC-types in-vivo. Enrichment of particular scaC-types temporally, following a dietary switch, and between fractions along with the phylogenetic congruence suggests that functional differences exist between types. Observed differences in dockerin modules suggest at least part of the functional heterogeneity may be conferred by scaC. The polymorphic nature of scaC enables the relative distribution of R. flavefaciens strains to be examined and represents a gene-centric approach to investigating the intraspecific adaptation of an important specialist population.
While the microbiota resident in the human gut is now known to provide a range of functions relevant to host health, many of the microbial members of the community have not yet been cultured or are represented by a limited number of isolates. We describe here the draft genome sequence of Turicibacter sanguinis PC909, isolated from a pooled healthy human fecal sample as part of the Australian Human Gut Microbiome Project.
The human intestinal microbiota is composed of 1013 to 1014 microorganisms whose collective genome (“microbiome”) contains at least 100 times as many genes as our own genome. We analyzed ~78 million base pairs of unique DNA sequence and 2062 polymerase chain reaction–amplified 16S ribosomal DNA sequences obtained from the fecal DNAs of two healthy adults. Using metabolic function analyses of identified genes, we compared our human genome with the average content of previously sequenced microbial genomes. Our microbiome has significantly enriched metabolism of glycans, amino acids, and xenobiotics; methanogenesis; and 2-methyl-d-erythritol 4-phosphate pathway–mediated biosynthesis of vitamins and isoprenoids. Thus, humans are superorganisms whose metabolism represents an amalgamation of microbial and human attributes.