A Programmable Escherichia coli Consortium via Tunable Symbiosis

Synthetic microbial consortia that can mimic natural systems have the potential to become a powerful biotechnology for various applications. One highly desirable feature of these consortia is that they can be precisely regulated. In this work we designed a programmable, symbiotic circuit that enables continuous tuning of the growth rate and composition of a synthetic consortium. We implemented our general design through the cross-feeding of tryptophan and tyrosine by two E. coli auxotrophs. By regulating the expression of genes related to the export or production of these amino acids, we were able to tune the metabolite exchanges and achieve a wide range of growth rates and strain ratios. In addition, by inverting the relationship of growth/ratio vs. inducer concentrations, we were able to “program” the co-culture for pre-specified attributes with the proper addition of inducing chemicals. This programmable proof-of-concept circuit or its variants can be applied to more complex systems where precise tuning of the consortium would facilitate the optimization of specific objectives, such as increasing the overall efficiency of microbial production of biofuels or pharmaceuticals

Utility of a multiplex reverse transcriptase-polymerase chain reaction assay (HemaVision) in the evaluation of genetic abnormalities in Korean children with acute leukemia: a single institution study

PurposeIn children with acute leukemia, bone marrow genetic abnormalities (GA) have prognostic significance, and may be the basis for minimal residual disease monitoring. Since April 2007, we have used a multiplex reverse transcriptase-polymerase chain reaction tool (HemaVision) to detect of GA.MethodsIn this study, we reviewed the results of HemaVision screening in 270 children with acute leukemia, newly diagnosed at The Catholic University of Korea from April 2007 to December 2011, and compared the results with those of fluorescence in situ hybridization (FISH), and G-band karyotyping.ResultsAmong the 270 children (153 males, 117 females), 187 acute lymphoblastic leukemia and 74 acute myeloid leukemia patients were identified. Overall, GA was detected in 230 patients (85.2%). HemaVision, FISH, and G-band karyotyping identified GA in 125 (46.3%), 126 (46.7%), and 215 patients (79.6%), respectively. TEL-AML1 (20.9%, 39/187) and AML1-ETO (27%, 20/74) were the most common GA in ALL and AML, respectively. Overall sensitivity of HemaVision was 98.4%, with false-negative results in 2 instances: 1 each for TEL-AML1 and MLL-AF4. An aggregate of diseasesspecific FISH showed 100% sensitivity in detection of GA covered by HemaVision for actual probes utilized. G-band karyotype revealed GA other than those covered by HemaVison screening in 133 patients (49.3%). Except for hyperdiplody and hypodiploidy, recurrent GA as defined by the World Health Organizationthat were not screened by HemaVision, were absent in the karyotype.ConclusionHemaVision, supported by an aggregate of FISH tests for important translocations, may allow for accurate diagnosis of GA in Korean children with acute leukemia

Comparison of Quantitative Cytomegalovirus Real-time PCR in Whole Blood and pp65 Antigenemia Assay: Clinical Utility of CMV Real-time PCR in Hematopoietic Stem Cell Transplant Recipients

Successful preemptive therapy for cytomegalovirus (CMV) infection in transplant patients depends on the availability of sensitive, specific, and timely diagnostic tests for CMV infection. Although the pp65 antigenemia assay has been widely used for this purpose, real-time quantification of CMV DNA has recently been recognized as an alternative diagnostic approach. However, the guidelines for antiviral therapy based on real-time quantitative polymerase chain reaction (RQ-PCR) have yet to be established. From November 2004 to March 2005, a total of 555 whole blood samples from 131 hematopoietic stem cell transplant (HSCT) recipients were prospectively collected. RQ-PCR was conducted using an Artus® CMV LC PCR kit (QIAGEN). Both qualitative and quantitative correlations were drawn between the two methods. Exposure to the antiviral agent influenced the results of the two assays. Additionally, the discrepancy was observed at low levels of antigenemia and CMV DNA load. Via ROC curve analysis, the tentative cutoff value for preemptive therapy was determined to be approximately 2×104 copies/mL (sensitivity, 80.0%; specificity, 50.0%) in the high risk patients, and approximately 3×104 copies/mL (sensitivity, 90.0%; specificity, 70.0%) in the patients at low risk for CMV disease. Further study to validate the optimal cutoff value for the initiation of preemptive therapy is currently underway

Microdroplet-Enabled Highly Parallel Co-Cultivation of Microbial Communities

Microbial interactions in natural microbiota are, in many cases, crucial for the sustenance of the communities, but the precise nature of these interactions remain largely unknown because of the inherent complexity and difficulties in laboratory cultivation. Conventional pure culture-oriented cultivation does not account for these interactions mediated by small molecules, which severely limits its utility in cultivating and studying “unculturable” microorganisms from synergistic communities. In this study, we developed a simple microfluidic device for highly parallel co-cultivation of symbiotic microbial communities and demonstrated its effectiveness in discovering synergistic interactions among microbes. Using aqueous micro-droplets dispersed in a continuous oil phase, the device could readily encapsulate and co-cultivate subsets of a community. A large number of droplets, up to ∼1,400 in a 10 mm×5 mm chamber, were generated with a frequency of 500 droplets/sec. A synthetic model system consisting of cross-feeding E. coli mutants was used to mimic compositions of symbionts and other microbes in natural microbial communities. Our device was able to detect a pair-wise symbiotic relationship when one partner accounted for as low as 1% of the total population or each symbiont was about 3% of the artificial community

Derivation of preliminary derived concentration guideline levels for surface soil at Kori Unit 1 by RESRAD probabilistic analysis

Preliminary surface soil Derived Concentration Guideline Levels (DCGLs) were derived conforming to the Multi-Agency Radiation Site Survey and Investigation Manual (MARSSIM) procedure for the site release and reuse of Kori Unit 1 in Korea. Based on the decommissioning experiences of the U.S. nuclear power plants, a suite of residual radionuclides was determined, and uncertainties contributed to the resultant dose by the input parameters were quantified via the sensitivity analysis of parameters. The peak of the mean dose was obtained via the probabilistic analysis of the RESRAD (RESidual RADioactivity)-ONSITE code. Consequently, DCGLw of Kori Unit 1 in accordance with two scenarios, industrial worker and residential farmer scenario, were derived and the results were compared respectively with other NPPs. It could be used as a basic guideline for establishing regulatory standards for reuse planning, designing the site characterization surveys and implementing final status survey (FSS)

XN. A programmable Escherichia coli consortium via tunable symbiosis. PLoS One

Abstract Synthetic microbial consortia that can mimic natural systems have the potential to become a powerful biotechnology for various applications. One highly desirable feature of these consortia is that they can be precisely regulated. In this work we designed a programmable, symbiotic circuit that enables continuous tuning of the growth rate and composition of a synthetic consortium. We implemented our general design through the cross-feeding of tryptophan and tyrosine by two E. coli auxotrophs. By regulating the expression of genes related to the export or production of these amino acids, we were able to tune the metabolite exchanges and achieve a wide range of growth rates and strain ratios. In addition, by inverting the relationship of growth/ratio vs. inducer concentrations, we were able to ''program'' the co-culture for pre-specified attributes with the proper addition of inducing chemicals. This programmable proof-of-concept circuit or its variants can be applied to more complex systems where precise tuning of the consortium would facilitate the optimization of specific objectives, such as increasing the overall efficiency of microbial production of biofuels or pharmaceuticals. Citation: Kerner A, Park J, Williams A, Lin XN (2012) A Programmable Escherichia coli Consortium via Tunable Symbiosis. PLoS ONE 7(3): e34032

Co-culture growth and ratio dynamics: baseline and with tuning.

<p>(<b>A</b>) Co-culture density (measured by OD₆₀₀) and Y3∶W3 ratio during growth in minimal medium without inducers. The ratio measurement is shown only for the exponential growth phase because the YFP calibration is not reliable after the cells enter the stationary phase. (<b>B</b>) An example of Y3∶W3 ratio dynamics at various arabinose concentrations. Propionate concentration was held at 20 mM. Only the exponential growth phase is shown. Each curve represents the mean of 4 replicates. Note: the secondary y-axis label in (<b>A</b>), Ratio Y3∶W3, is also the label for the primary y-axis in (<b>B</b>).</p

Relationship between comorbidity and health outcomes in patients with heart failure: a systematic review and meta-analysis

Abstract Background The prevalence of heart failure (HF) is expected to rise due to increased survivorship and life expectancy of patients with acute heart conditions. Patients with HF and other multiple comorbid conditions are likely to have poor health outcomes. This study aimed to assimilate the current body of knowledge and to provide the pooled effect of HF patients’ comorbid conditions on health outcomes. Methods A systematic search was performed using MEDLINE, EMBASE and CINAHL databases. Observational studies evaluating the relationship between comorbid conditions and the health outcomes of HF were included. The pooled effect sizes of comorbidity on the identified health outcomes were calculated using a random effects model, and the heterogeneity was evaluated using I2 statistics. Results A total of 42 studies were included in this review, and a meta-analysis was performed using the results of 39 studies. In the pooled analysis, the presence of a comorbid condition showed a significant pooled effect size in relation to the prognostic health outcomes: all-cause mortality (HR 1.31; 95% CI 1.18, 1.45), all-cause readmission (HR 1.16; 95% CI 1.09, 1.23), HF-related readmission (HR 1.13; 95% CI 1.05, 1.23), and non-HF-related readmission (HR 1.17; 95% CI 1.07, 1.27). Also, comorbidity was significantly associated with health-related quality of life and self-care confidence. Furthermore, we identified a total of 32 comorbid conditions from included studies. From these, 16 individual conditions were included in the meta-analyses, and we identified 10 comorbid conditions to have negative effects on overall prognostic outcomes: DM (HR 1.16, 95% CI 1.11, 1.22), COPD (HR 1.31, 95% CI 1.23, 1.39), CKD (HR 1.18, 95% CI 1.14, 1.23, stroke (HR 1.25, 95% CI 1.17, 1.31), IHD (HR 1.17, 95% CI 1.11, 1.23), anemia (HR 1.42, 95% CI 1.14, 1.78), cancer (HR 1.17, 95% CI 1.04, 1.32), atrial fibrillation (HR 1.25, 95% CI 1.01, 1.54), dementia (HR 1.19, 95% CI 1.03, 1.36) and depression (HR 1.17, 95% CI 1.04, 1.31). Conclusions Comorbid conditions have significantly negative pooled effects on HF patient health outcomes, especially in regard to the prognostic health outcomes. Clinicians should carefully identify and manage these conditions when implementing HF interventions to improve prognostic outcomes

Design space and testing.

<p>(<b>A, B</b>) By inverting the relationships of growth rate/strain ratio vs. inducer concentrations, a design space was generated to represent the two-dimensional space of achievable growth rates and strain ratios, and to determine the arabinose (<b>A</b>) and propionate (<b>B</b>) concentrations for a desired growth rate and end-exponential ratio combination. The colored circles are “prediction” points and the asterisks of the same color are the “actual” results of using that combination of arabinose and propionate in the co-culture. The colors denote inducer combinations: purple (0.11%, 8 mM); pink (0.02%, 5 mM); orange (0.13%, 30 mM); red (0.06%, 12 mM); yellow (0.06%, 20 mM); black (0.11%, 5 mM). (<b>C, D</b>) Comparing the predicted and actual outcome for growth rate (<b>C</b>) and end-exponential ratio (<b>D</b>) in bar graph form; the predictions are in darker colors and the actual (experimental) results are in lighter ones. Error bars: ± standard deviation. The mid-exponential ratio design space is shown in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0034032#pone.0034032.s004" target="_blank">Figure S4</a>. Experimental data are given <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0034032#pone.0034032.s009" target="_blank">Data S1</a>.</p

Basic schematic of the tunable cross-feeding circuit.

<p>(<b>A</b>) In this general design, inducer 1 and inducer 2 control the export of metabolites 1 and 2, respectively. The two auxotrophs must cross-feed in order to survive in the minimal medium. (<b>B</b>) In our specific implementation, two <i>E. coli</i> auxotrophic strains exchange tryptophan (Trp) and tyrosine (Tyr). The forced symbiosis is controlled by plasmids pAK1 (in the Trp auxotroph, W3) and pAK5 (in the Tyr auxotroph, Y3). Plasmid pAK1 contains gene <i>yddG</i> behind the tunable promoter P_BAD, and pAK5 contains <i>trpEDfbr</i> behind P_prpB (Methods). Strain Y3 is tagged with yellow fluorescent protein (YFP).</p