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OBJECTIVES/SPECIFIC AIMS: Our primary objective is to determine whether the bacteria exerts its effect intra- or extra-cellularly. We have genomic and microscopy preliminary evidence indicating that the bacteria is capable of invading endometrial cells. Our secondary objective is to identify what type of impact the bacteria have on the host cells and whether they are capable of transforming the host cells from a benign into a malignant phenotype. We are currently testing a putative mechanism by which the bacteria may cause the overexpression of the hypoxia inducible factor (HIF), a hallmark of endometrial cancer. METHODS/STUDY POPULATION: We are utilizing our custom built optofluidics platform (microfluidics platform incorporated into an advanced microscope with optical laser tweezers) to isolate single cells from the endometrial tissues of 150 patients with and without endometrial cancer. We are utilizing single cell whole genome amplification followed by qPCR to identify if the bacteria is present intracellularly. We are coupling this procedure with standard microbiological invasion assays with endometrial cell line cultures and P.somerae. We are also utilizing our optofluidics platform to perform single cell transcriptomic amplification, followed by sequencing of cells invaded or in the presence of the bacteria to determine the impact in the transcriptome of the host cell. We are coupling this with western blots of factors hypothesized to be impacted by the bacteria in the overexpression of HIF. RESULTS/ANTICIPATED RESULTS: Based on our preliminary data we anticipate to find evidence that P.somerae is invading the host cells, in particular the cells in tumor tissues. We also expect to find that the intracellular presence of the bacteria is causing the overexpression of the HIF pathway, hence resulting in a cancerous phenotype. DISCUSSION/SIGNIFICANCE OF IMPACT: Our long-term goal is to develop primary prevention strategies that will reduce endometrial cancer incidence rates. A confirmation of our hypothesis could suggest that it is sufficient for endometrial cancer prevention efforts to eliminate P.somerae, in line with gastric and cervical cancer efforts. It could also mean that targeting P.somerae in cancer treatment is necessary to contain the disease and prevent recurrence.
To evaluate whole-genome sequencing (WGS) as a molecular typing tool for MRSA outbreak investigation.
Investigation of MRSA colonization/infection in a neonatal intensive care unit (NICU) over 3 years (2014–2017).
Single-center level IV NICU.
NICU infants and healthcare workers (HCWs).
Infants were screened for MRSA using a swab of the anterior nares, axilla, and groin, initially by targeted (ring) screening, and later by universal weekly screening. Clinical cultures were collected as indicated. HCWs were screened once using swabs of the anterior nares. MRSA isolates were typed using WGS with core-genome multilocus sequence typing (cgMLST) analysis and by pulsed-field gel electrophoresis (PFGE). Colonized and infected infants and HCWs were decolonized. Control strategies included reinforcement of hand hygiene, use of contact precautions, cohorting, enhanced environmental cleaning, and remodeling of the NICU.
We identified 64 MRSA-positive infants: 53 (83%) by screening and 11 (17%) by clinical cultures. Of 85 screened HCWs, 5 (6%) were MRSA positive. WGS of MRSA isolates identified 2 large clusters (WGS groups 1 and 2), 1 small cluster (WGS group 3), and 8 unrelated isolates. PFGE failed to distinguish WGS group 2 and 3 isolates. WGS groups 1 and 2 were codistributed over time. HCW MRSA isolates were primarily in WGS group 1. New infant MRSA cases declined after implementation of the control interventions.
We identified 2 contemporaneous MRSA outbreaks alongside sporadic cases in a NICU. WGS was used to determine strain relatedness at a higher resolution than PFGE and was useful in guiding efforts to control MRSA transmission.
The modern molecular biology movement was developed in the 1960s with the conglomeration of biology, chemistry, and physics. Today, molecular biology is an integral part of studies aimed at understanding the evolution and ecology of gastrointestinal microbial communities. Molecular techniques have led to significant gains in our understanding of the chicken gastrointestinal microbiome. New advances, primarily in DNA sequencing technologies, have equipped researchers with the ability to explore these communities at an unprecedented level. A reinvigorated movement in systems biology offers a renewed promise in obtaining a more complete understanding of chicken gastrointestinal microbiome dynamics and their contributions to increasing productivity, food value, security, and safety as well as reducing the public health impact of raising production animals. Here, we contextualize the contributions molecular biology has already made to our understanding of the chicken gastrointestinal microbiome and propose targeted research directions that could further exploit molecular technologies to improve the economy of the poultry industry.
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