The Democratic Republic of the Congo (DRC) declared an Ebola virus disease (EVD) outbreak in North Kivu in August 2018. By June 2019, the outbreak had spread to 26 health zones in northeastern DRC, causing >2,000 reported cases and >1,000 deaths. On June 10, 2019, three members of a Congolese family with EVD-like symptoms traveled to western Uganda's Kasese District to seek medical care. Shortly thereafter, the Viral Hemorrhagic Fever Surveillance and Laboratory Program (VHF program) at the Uganda Virus Research Institute (UVRI) confirmed that all three patients had EVD. The Ugandan Ministry of Health declared an outbreak of EVD in Uganda's Kasese District, notified the World Health Organization, and initiated a rapid response to contain the outbreak. As part of this response, UVRI and the United States Centers for Disease Control and Prevention, with the support of Uganda's Public Health Emergency Operations Center, the Kasese District Health Team, the Superintendent of Bwera General Hospital, the United States Department of Defense's Makerere University Walter Reed Project, and the United States Mission to Kampala's Global Health Security Technical Working Group, jointly established an Ebola Field Laboratory in Kasese District at Bwera General Hospital, proximal to an Ebola Treatment Unit (ETU). The laboratory consisted of a rapid containment kit for viral inactivation of patient specimens and a GeneXpert Instrument for performing Xpert Ebola assays. Laboratory staff tested 76 specimens from alert and suspect cases of EVD; the majority were admitted to the ETU (89.3%) and reported recent travel to the DRC (58.9%). Although no EVD cases were detected by the field laboratory, it played an important role in patient management and epidemiological surveillance by providing diagnostic results in <3 hours. The integration of the field laboratory into Uganda's National VHF Program also enabled patient specimens to be referred to Entebbe for confirmatory EBOV testing and testing for other hemorrhagic fever viruses that circulate in Uganda.

BACKGROUND: The contribution of alternative splicing and isoform expression to cellular response is emerging as an area of considerable interest, and the newly developed exon arrays allow for systematic study of these processes. We use this pilot study to report on the feasibility of exon array implementation looking to replace the 3' in vitro transcription expression arrays in our laboratory. One of the most widely studied models of cellular response is T-cell activation from exogenous stimulation. Microarray studies have contributed to our understanding of key pathways activated during T-cell stimulation. We use this system to examine whole genome transcription and alternate exon usage events that are regulated during lymphocyte proliferation in an attempt to evaluate the exon arrays. RESULTS: Peripheral blood mononuclear cells form healthy donors were activated using phytohemagglutinin, IL2 and ionomycin and harvested at 5 points over a 7 day period. Flow cytometry measured cell cycle events and the Affymetrix exon array platform was used to identify the gene expression and alternate exon usage changes. Gene expression changes were noted in a total of 2105 transcripts, and alternate exon usage identified in 472 transcript clusters. There was an overlap of 263 transcripts which showed both differential expression and alternate exon usage over time. Gene ontology enrichment analysis showed a broader range of biological changes in biological processes for the differentially expressed genes, which include cell cycle, cell division, cell proliferation, chromosome segregation, cell death, component organization and biogenesis and metabolic process ontologies. The alternate exon usage ontological enrichments are in metabolism and component organization and biogenesis. We focus on alternate exon usage changes in the transcripts of the spliceosome complex. The real-time PCR validation rates were 86% for transcript expression and 71% for alternate exon usage. CONCLUSIONS: This study illustrates that the Exon array technology has the potential to provide information on both transcript expression and isoform usage, with very little increase in expense.

Understanding the biologic significance of alternative splicing has been impeded by the difficulty in systematically identifying and validating transcript isoforms. Current exon array workflows suggest several different filtration steps to reduce the number of tests and increase the detection of alternative splicing events. In this study, we examine the effects of the suggested pre-analysis filtration by detection above background P value or signal intensity. This is followed post-analytically by restriction of exon expression to a fivefold change between groups, limiting the analysis to known alternative splicing events, or using the intersection of the results from different algorithms. Combinations of the filters are also examined. We find that none of the filtering methods reduces the number of technical false-positive calls identified by visual inspection. These include edge effects, nonresponsive probe sets, and inclusion of intronic and untranslated region probe sets into transcript annotations. Modules for filtering the exon microarray data on the basis of annotation features are needed. We propose new approaches to data filtration that would reduce the number of technical false-positives and therefore, impact the time spent performing visual inspection of the exon arrays.

We investigated peripheral blood mononuclear cell gene expression responses to acute psychosocial stress to identify molecular pathways relevant to the stress response. Blood samples were obtained from 10 healthy male subjects before, during and after (at 0, 30, and 60 min) a standardized psychosocial laboratory stressor. Ribonucleic acid (RNA) was extracted and gene expression measured by hybridization to a 20,000-gene microarray. Gene Set Expression Comparisons (GSEC) using defined pathways were used for the analysis. Forty-nine pathways were significantly changed from baseline to immediately after the stressor (p<0.05), implicating cell cycle, cell signaling, adhesion and immune responses. The comparison between stress and recovery (measured 30 min later) identified 36 pathways, several involving stress-responsive signaling cascades and cellular defense mechanisms. These results have relevance for understanding molecular mechanisms of the physiological stress response, and might be used to further study adverse health outcomes of psychosocial stress.