Identification of individuals by trait prediction using whole-genome sequencing data.

Prediction of human physical traits and demographic information from genomic data challenges privacy and data deidentification in personalized medicine. To explore the current capabilities of phenotype-based genomic identification, we applied whole-genome sequencing, detailed phenotyping, and statistical modeling to predict biometric traits in a cohort of 1,061 participants of diverse ancestry. Individually, for a large fraction of the traits, their predictive accuracy beyond ancestry and demographic information is limited. However, we have developed a maximum entropy algorithm that integrates multiple predictions to determine which genomic samples and phenotype measurements originate from the same person. Using this algorithm, we have reidentified an average of >8 of 10 held-out individuals in an ethnically mixed cohort and an average of 5 of either 10 African Americans or 10 Europeans. This work challenges current conceptions of personal privacy and may have far-reaching ethical and legal implications.

Comparison of microarray-predicted closest genomes to sequencing for poliovirus vaccine strain similarity and influenza A phylogeny.

We evaluate sequence data from the PathChip high-density hybridization array for epidemiological interpretation of detected pathogens. For influenza A, we derive similar relative outbreak clustering in phylogenetic trees from PathChip-derived compared to classical Sanger-derived sequences. For a positive polio detection, recent infection could be excluded based on vaccine strain similarity.

Detection of chromosomal breakpoints in patients with developmental delay and speech disorders.

Delineating candidate genes at the chromosomal breakpoint regions in the apparently balanced chromosome rearrangements (ABCR) has been shown to be more effective with the emergence of next-generation sequencing (NGS) technologies. We employed a large-insert (7-11 kb) paired-end tag sequencing technology (DNA-PET) to systematically analyze genome of four patients harbouring cytogenetically defined ABCR with neurodevelopmental symptoms, including developmental delay (DD) and speech disorders. We characterized structural variants (SVs) specific to each individual, including those matching the chromosomal breakpoints. Refinement of these regions by Sanger sequencing resulted in the identification of five disrupted genes in three individuals: guanine nucleotide binding protein, q polypeptide (GNAQ), RNA-binding protein, fox-1 homolog (RBFOX3), unc-5 homolog D (C.elegans) (UNC5D), transmembrane protein 47 (TMEM47), and X-linked inhibitor of apoptosis (XIAP). Among them, XIAP is the causative gene for the immunodeficiency phenotype seen in the patient. The remaining genes displayed specific expression in the fetal brain and have known biologically relevant functions in brain development, suggesting putative candidate genes for neurodevelopmental phenotypes. This study demonstrates the application of NGS technologies in mapping individual gene disruptions in ABCR as a resource for deciphering candidate genes in human neurodevelopmental disorders (NDDs).

Pathogen chip for respiratory tract infections.

Determining the viral etiology of respiratory tract infections (RTI) has been limited for the most part to specific primer PCR-based methods due to their increased sensitivity and specificity compared to other methods, such as tissue culture. However, specific primer approaches have limited the ability to fully understand the diversity of infecting pathogens. A pathogen chip system (PathChip), developed at the Genome Institute of Singapore (GIS), using a random-tagged PCR coupled to a chip with over 170,000 probes, has the potential to recognize all known human viral pathogens. We tested 290 nasal wash specimens from Filipino children <2 years of age with respiratory tract infections using culture and 3 PCR methods-EraGen, Luminex, and the GIS PathChip. The PathChip had good diagnostic accuracy, ranging from 85.9% (95% confidence interval [CI], 81.3 to 89.7%) for rhinovirus/enteroviruses to 98.6% (95% CI, 96.5 to 99.6%) for PIV 2, compared to the other methods and additionally identified a number of viruses not detected by these methods.

Oseltamivir ring prophylaxis for containment of 2009 H1N1 influenza outbreaks.

BACKGROUND: From June 22 through June 25, 2009, four outbreaks of infection with the pandemic influenza A (H1N1) virus occurred in Singapore military camps. We report the efficacy of ring chemoprophylaxis (geographically targeted containment by means of prophylaxis) with oseltamivir to control outbreaks of 2009 H1N1 influenza in semiclosed environments. METHODS: All personnel with suspected infection were tested and clinically isolated if infection was confirmed. In addition, we administered postexposure ring chemoprophylaxis with oseltamivir and segregated the affected military units to contain the spread of the virus. All personnel were screened three times weekly both for virologic infection, by means of nasopharyngeal swabs and reverse-transcriptase-polymerase-chain-reaction assay with sequencing, and for clinical symptoms, by means of questionnaires. RESULTS: A total of 1175 personnel were at risk across the four sites, with 1100 receiving oseltamivir prophylaxis. A total of 75 personnel (6.4%) were infected before the intervention, and 7 (0.6%) after the intervention. There was a significant reduction in the overall reproductive number (the number of new cases attributable to the index case), from 1.91 (95% credible interval, 1.50 to 2.36) before the intervention to 0.11 (95% credible interval, 0.05 to 0.20) after the intervention. Three of the four outbreaks showed a significant reduction in the rate of infection after the intervention. Molecular analysis revealed that all four outbreaks were derived from the New York lineage of the 2009 H1N1 virus and that cases within each outbreak were due to transmission rather than unrelated episodes of infection. Of the 816 personnel treated with oseltamivir who were surveyed, 63 (7.7%) reported mild, nonrespiratory side effects of the drug, with no severe adverse events. CONCLUSIONS: Oseltamivir ring chemoprophylaxis, together with prompt identification and isolation of infected personnel, was effective in reducing the impact of outbreaks of 2009 H1N1 influenza in semiclosed settings.

Deregulated direct targets of the hepatitis B virus (HBV) protein, HBx, identified through chromatin immunoprecipitation and expression microarray profiling.

The hepatitis B-X (HBx) protein is strongly associated with hepatocellular carcinoma. It is implicated not to directly cause cancer but to play a role in hepatocellular carcinoma as a co-factor. The oncogenic potential of HBx primarily lies in its interaction with transcriptional regulators resulting in aberrant gene expression and deregulated cellular pathways. Utilizing ultraviolet irradiation to simulate a tumor-initiating event, we integrated chip-based chromatin immunoprecipitation (ChIP-chip) with expression microarray profiling and identified 184 gene targets directly deregulated by HBx. One-hundred forty-four transcription factors interacting with HBx were computationally inferred. We experimentally validated that HBx interacts with some of the predicted transcription factors (pTF) as well as the promoters of the deregulated target genes of these pTFs. Significantly, we demonstrated that the pTF interacts with the promoters of the deregulated HBx target genes and that deregulation by HBx of these HBx target genes carrying the pTF consensus sequences can be reversed using pTF small interfering RNAs. The roles of these deregulated direct HBx target genes and their relevance in cancer was inferred via querying against biogroup/cancer-related microarray databases using web-based NextBio(TM) software. Six pathways, including the Jak-STAT pathway, were predicted to be significantly deregulated when HBx binds indirectly to direct target gene promoters. In conclusion, this study represents the first ever demonstration of the utilization of ChIP-chip to identify deregulated direct gene targets from indirect protein-DNA binding as well as transcriptional factors directly interacting with HBx. Increased knowledge of the gene/transcriptional factor targets of HBx will enhance our understanding of the role of HBx in hepatocellular carcinogenesis and facilitate the design of better strategies in combating hepatitis B virus-associated hepatocellular carcinoma.

Global mapping of c-Myc binding sites and target gene networks in human B cells.

The protooncogene MYC encodes the c-Myc transcription factor that regulates cell growth, cell proliferation, cell cycle, and apoptosis. Although deregulation of MYC contributes to tumorigenesis, it is still unclear what direct Myc-induced transcriptomes promote cell transformation. Here we provide a snapshot of genome-wide, unbiased characterization of direct Myc binding targets in a model of human B lymphoid tumor using ChIP coupled with pair-end ditag sequencing analysis (ChIP-PET). Myc potentially occupies > 4,000 genomic loci with the majority near proximal promoter regions associated frequently with CpG islands. Using gene expression profiles with ChIP-PET, we identified 668 direct Myc-regulated gene targets, including 48 transcription factors, indicating that Myc is a central transcriptional hub in growth and proliferation control. This first global genomic view of Myc binding sites yields insights of transcriptional circuitries and cis regulatory modules involving Myc and provides a substantial framework for our understanding of mechanisms of Myc-induced tumorigenesis.

The Oct4 and Nanog transcription network regulates pluripotency in mouse embryonic stem cells.

Oct4 and Nanog are transcription factors required to maintain the pluripotency and self-renewal of embryonic stem (ES) cells. Using the chromatin immunoprecipitation paired-end ditags method, we mapped the binding sites of these factors in the mouse ES cell genome. We identified 1,083 and 3,006 high-confidence binding sites for Oct4 and Nanog, respectively. Comparative location analyses indicated that Oct4 and Nanog overlap substantially in their targets, and they are bound to genes in different configurations. Using de novo motif discovery algorithms, we defined the cis-acting elements mediating their respective binding to genomic sites. By integrating RNA interference-mediated depletion of Oct4 and Nanog with microarray expression profiling, we demonstrated that these factors can activate or suppress transcription. We further showed that common core downstream targets are important to keep ES cells from differentiating. The emerging picture is one in which Oct4 and Nanog control a cascade of pathways that are intricately connected to govern pluripotency, self-renewal, genome surveillance and cell fate determination.