SINCERA: A Pipeline for Single-Cell RNA-Seq Profiling Analysis
A major challenge in developmental biology is to understand the genetic and cellular processes/programs driving organ formation, and differentiation of the diverse cell types that comprise the embryo. While recent studies using single cell transcriptome analysis illustrate the power to measure and understand cellular heterogeneity in complex biological systems, processing large amounts of RNA-seq data from heterogeneous cell populations creates the need for readily accessible tools for the analysis of single-cell RNA-seq (scRNA-seq) profiles. The present study presents a generally applicable analytic pipeline (SINCERA: a computational pipeline for SINgle CEll RNA-seq profiling Analysis) for processing scRNA-seq data from a whole organ or sorted cells. The pipeline supports the analysis for: 1) the distinction and identification of major cell types; 2) the identification of cell type specific gene signatures; and 3) the determination of driving forces of given cell types. We applied this pipeline to the RNA-seq analysis of single cells isolated from embryonic mouse lung at E16.5. Through the pipeline analysis, we distinguished major cell types of fetal mouse lung, including epithelial, endothelial, smooth muscle, pericyte, and fibroblast-like cell types, and identified cell type specific gene signatures, bioprocesses, and key regulators. Implemented in R, and licensed under the GNU General Public License v3, SINCERA is freely available from Cincinnati Children's PBGE website
Metagenomic Sequencing with Strain-Level Resolution Implicates Uropathogenic E. coli in Necrotizing Enterocolitis and Mortality in Preterm Infants
(NEC) afflicts approximately 10% of extremely preterm infants with high fatality. There is an implication of inappropriate bacterial colonization with Enterobacteriaceae, but there has been no specific pathogen identified. We identify uropathogenic E. coli (UPEC) colonization as a significant risk factor for the development of NEC and subsequent mortality. We describe a large-scale deep shotgun metagenomic sequence analysis of the early intestinal microbiome of 144 preterm and 22 term infants. Using a pan-genomic approach to functionally subtype the E. coli, we identify genes associated with NEC and mortality that indicate colonization by UPEC. Metagenomic multilocus sequence typing analysis further defined NEC-associated strains as sequence types often associated with urinary tract infections, including ST69, ST73, ST95, ST127, ST131, and ST144. Although other factors associated with prematurity may also contribute, this report suggests a link between UPEC and NEC, and indicates the need for further attention to these sequence types as potential causal agents.
Functional and Structural Connectivity of the Visual System in Infants with Perinatal Brain Injury
In this study, we evaluated term and preterm infants with perinatal brain injury and term controls in the first 8 weeks of life using task-based functional MRI, functional connectivity during a visual task, and structural connectivity using diffusion tensor imaging. We found that infants with brain injury had reduced functional and structural connectivity compared to term control infants. Specifically, infants with brain injury had reduced activation in the expected area of the occipital cortex, weaker connectivity between visual areas and other areas of the brain during the visual task, and reduced fractional anisotropy (a measure of white matter integrity) in white matter tracts projecting to visual regions. Our next steps will be to correlate these early neuroimaging findings with later visual outcomes in this cohort.