Person:

Kostic, Aleksandar

Background: Gut microbiome dysbiosis has been demonstrated in subjects with newly diagnosed and chronic inflammatory bowel disease (IBD). In this study we sought to explore longitudinal changes in dysbiosis and ascertain associations between dysbiosis and markers of disease activity and treatment outcome. Methods: We performed a prospective cohort study of 19 treatment-naïve pediatric IBD subjects and 10 healthy controls, measuring fecal calprotectin and assessing the gut microbiome via repeated stool samples. Associations between clinical characteristics and the microbiome were tested using generalized estimating equations. Random forest classification was used to predict ultimate treatment response (presence of mucosal healing at follow-up colonoscopy) or non-response using patients’ pretreatment samples. Results: Patients with Crohn’s disease had increased markers of inflammation and dysbiosis compared to controls. Patients with ulcerative colitis had even higher inflammation and dysbiosis compared to those with Crohn’s disease. For all cases, the gut microbial dysbiosis index associated significantly with clinical and biological measures of disease severity, but did not associate with treatment response. We found differences in specific gut microbiome genera between cases/controls and responders/non-responders including Akkermansia, Coprococcus, Fusobacterium, Veillonella, Faecalibacterium, and Adlercreutzia. Using pretreatment microbiome data in a weighted random forest classifier, we were able to obtain 76.5 % accuracy for prediction of responder status. Conclusions: Patient dysbiosis improved over time but persisted even among those who responded to treatment and achieved mucosal healing. Although dysbiosis index was not significantly different between responders and non-responders, we found specific genus-level differences. We found that pretreatment microbiome signatures are a promising avenue for prediction of remission and response to treatment. Electronic supplementary material The online version of this article (doi:10.1186/s13073-016-0331-y) contains supplementary material, which is available to authorized users.

The tumor microenvironment is a complex community consisting of neoplastic cells, surrounding stromal cells, a broad array of immune cells, and a microbiota. By sheer numbers, the microbiota has its greatest manifestation in colorectal cancer (CRC) because the colon contains up to 100 trillion bacteria, outnumbering human cells by a factor of 10 and encoding a gene-content that is 100-fold larger than that of the human genome. Indeed, previous studies using germ-free mice in a variety of genetic backgrounds have demonstrated that the microbiota can impact colorectal tumorigenesis. In addition, specific strains of enterotoxigenic bacteria have been shown to promote colitis-associated cancer in mice. Here, we explore the composition of the tissue-associated microbiota in human CRC and evaluate the role of tumor-enriched microbes in potentiating colorectal tumorigenesis in mice. Advances in DNA sequencing technology have fueled a renaissance in the microbiome field. Deep sequencing metagenomics enables rapid, culture-independent characterization of a microbial community. We present PathSeq, a highly scalable software tool that performs computational subtraction on high-throughput sequencing data to identify nonhuman nucleic acids. PathSeq makes it possible to analyze sequence datasets as large as human whole-genomes for the purpose of metagenomics and also to discover previously unsequenced microorganisms. We used PathSeq to characterize the composition of the microbiota in human CRCusing whole-genome sequencing on nine tumor/normal pairs and 16S rDNA sequencing on an additional 95 pairs. The genus Fusobacterium was highly enriched in tumors, while the Bacteroidetes and Firmicutes phyla were depleted.We show that in the (Apc^{Min/+}) mouse model of intestinal tumorigenesis, Fusobacterium nucleatum increases tumor multiplicity, selectively recruits tumor-infiltrating myeloid cells, and is associated with a pro-inflammatory expression signature that is shared with human fusobacteria-positive colorectal carcinomas. We find that Fusobacterium spp. are enriched in human colonic adenomas relative to surrounding tissues and fusobacterial abundance is increased in stool samples from patients with colorectal adenomas and carcinomas, compared to healthy subjects. Collectively, these data support that fusobacteria may be involved in early stages of intestinal tumorigenesis and, through recruitment of tumor-infiltrating immune cells, may generate a pro-inflammatory tissue microenvironment conducive to colorectal neoplasia progression.

The pattern recognition receptor RIG-I is critical for Type-I interferon production. However, the global regulation of RIG-I signaling is only partially understood. Using a human genome-wide RNAi-screen, we identified 226 novel regulatory proteins of RIG-I mediated interferon-β production. Furthermore, the screen identified a metabolic pathway that synthesizes the inositol pyrophosphate 1-IP7 as a previously unrecognized positive regulator of interferon production. Detailed genetic and biochemical experiments demonstrated that the kinase activities of IPPK, PPIP5K1 and PPIP5K2 (which convert IP5 to1-IP7) were critical for both interferon induction, and the control of cellular infection by Sendai and influenza A viruses. Conversely, ectopically expressed inositol pyrophosphate-hydrolases DIPPs attenuated interferon transcription. Mechanistic experiments in intact cells revealed that the expression of IPPK, PPIP5K1 and PPIP5K2 was needed for the phosphorylation and activation of IRF3, a transcription factor for interferon. The addition of purified individual inositol pyrophosphates to a cell free reconstituted RIG-I signaling assay further identified 1-IP7 as an essential component required for IRF3 activation. The inositol pyrophosphate may act by β-phosphoryl transfer, since its action was not recapitulated by a synthetic phosphonoacetate analogue of 1-IP7. This study thus identified several novel regulators of RIG-I, and a new role for inositol pyrophosphates in augmenting innate immune responses to viral infection that may have therapeutic applications.

The animal and bacterial kingdoms have coevolved and coadapted in response to environmental selective pressures over hundreds of millions of years. The meta'omics revolution in both sequencing and its analytic pipelines is fostering an explosion of interest in how the gut microbiome impacts physiology and propensity to disease. Gut microbiome studies are inherently interdisciplinary, drawing on approaches and technical skill sets from the biomedical sciences, ecology, and computational biology. Central to unraveling the complex biology of environment, genetics, and microbiome interaction in human health and disease is a deeper understanding of the symbiosis between animals and bacteria. Experimental model systems, including mice, fish, insects, and the Hawaiian bobtail squid, continue to provide critical insight into how host-microbiota homeostasis is constructed and maintained. Here we consider how model systems are influencing current understanding of host-microbiota interactions and explore recent human microbiome studies.

The tumor microenvironment of colorectal carcinoma is a complex community of genomically altered cancer cells, nonneoplastic cells, and a diverse collection of microorganisms. Each of these components may contribute to carcinogenesis; however, the role of the microbiota is the least well understood. We have characterized the composition of the microbiota in colorectal carcinoma using whole genome sequences from nine tumor/normal pairs. Fusobacterium sequences were enriched in carcinomas, confirmed by quantitative PCR and 16S rDNA sequence analysis of 95 carcinoma/normal DNA pairs, while the Bacteroidetes and Firmicutes phyla were depleted in tumors. Fusobacteria were also visualized within colorectal tumors using FISH. These findings reveal alterations in the colorectal cancer microbiota; however, the precise role of Fusobacteria in colorectal carcinoma pathogenesis requires further investigation.

Abstract Background: More extensive use of metagenomic shotgun sequencing in microbiome research relies on the development of high-throughput, cost-effective sequencing. Here we present a comprehensive evaluation of the performance of the new high-throughput sequencing platform BGISEQ-500 for metagenomic shotgun sequencing and compare its performance with that of 2 Illumina platforms. Findings: Using fecal samples from 20 healthy individuals, we evaluated the intra-platform reproducibility for metagenomic sequencing on the BGISEQ-500 platform in a setup comprising 8 library replicates and 8 sequencing replicates. Cross-platform consistency was evaluated by comparing 20 pairwise replicates on the BGISEQ-500 platform vs the Illumina HiSeq 2000 platform and the Illumina HiSeq 4000 platform. In addition, we compared the performance of the 2 Illumina platforms against each other. By a newly developed overall accuracy quality control method, an average of 82.45 million high-quality reads (96.06% of raw reads) per sample, with 90.56% of bases scoring Q30 and above, was obtained using the BGISEQ-500 platform. Quantitative analyses revealed extremely high reproducibility between BGISEQ-500 intra-platform replicates. Cross-platform replicates differed slightly more than intra-platform replicates, yet a high consistency was observed. Only a low percentage (2.02%–3.25%) of genes exhibited significant differences in relative abundance comparing the BGISEQ-500 and HiSeq platforms, with a bias toward genes with higher GC content being enriched on the HiSeq platforms. Conclusions: Our study provides the first set of performance metrics for human gut metagenomic sequencing data using BGISEQ-500. The high accuracy and technical reproducibility confirm the applicability of the new platform for metagenomic studies, though caution is still warranted when combining metagenomic data from different platforms.

Abstract Motivation Across biology, we are seeing rapid developments in scale of data production without a corresponding increase in data analysis capabilities. Results: Here, we present Aether (http://aether.kosticlab.org), an intuitive, easy-to-use, cost-effective and scalable framework that uses linear programming to optimally bid on and deploy combinations of underutilized cloud computing resources. Our approach simultaneously minimizes the cost of data analysis and provides an easy transition from users’ existing HPC pipelines. Availability and implementation Data utilized are available at https://pubs.broadinstitute.org/diabimmune and with EBI SRA accession ERP005989. Source code is available at (https://github.com/kosticlab/aether). Examples, documentation and a tutorial are available at http://aether.kosticlab.org. Contact chirag_patel@hms.harvard.edu or aleksandar.kostic@joslin.harvard.edu Supplementary information Supplementary data are available at Bioinformatics online.

Objectives: Evidence suggests a possible role of Fusobacterium nucleatum in colorectal carcinogenesis, especially in right-sided proximal colorectum. Considering a change in bowel contents and microbiome from proximal to distal colorectal segments, we hypothesized that the proportion of colorectal carcinoma enriched with F. nucleatum might gradually increase along the bowel subsites from rectum to cecum. Methods: A retrospective, cross-sectional analysis was conducted on 1,102 colon and rectal carcinomas in molecular pathological epidemiology databases of the Nurses’ Health Study and the Health Professionals Follow-up Study. We measured the amount of F. nucleatum DNA in colorectal tumor tissue using a quantitative PCR assay and equally dichotomized F. nucleatum-positive cases (high vs. low). We used multivariable logistic regression analysis to examine the relationship of a bowel subsite variable (rectum, rectosigmoid junction, sigmoid colon, descending colon, splenic flexure, transverse colon, hepatic flexure, ascending colon, and cecum) with the amount of F. nucleatum. Results: The proportion of F. nucleatum-high colorectal cancers gradually increased from rectal cancers (2.5% 4/157) to cecal cancers (11% 19/178), with a statistically significant linear trend along all subsites (P<0.0001) and little evidence of non-linearity. The proportion of F. nucleatum-low cancers was higher in rectal, ascending colon, and cecal cancers than in cancers of middle segments. Conclusions: The proportion of F. nucleatum-high colorectal cancers gradually increases from rectum to cecum. Our data support the colorectal continuum model that reflects pathogenic influences of the gut microbiota on neoplastic and immune cells and challenges the prevailing two-colon (proximal vs. distal) dichotomy paradigm.