OptForce: An Optimization Procedure for Identifying All Genetic Manipulations Leading to Targeted Overproductions

Computational procedures for predicting metabolic interventions leading to the overproduction of biochemicals in microbial strains are widely in use. However, these methods rely on surrogate biological objectives (e.g., maximize growth rate or minimize metabolic adjustments) and do not make use of flux measurements often available for the wild-type strain. In this work, we introduce the OptForce procedure that identifies all possible engineering interventions by classifying reactions in the metabolic model depending upon whether their flux values must increase, decrease or become equal to zero to meet a pre-specified overproduction target. We hierarchically apply this classification rule for pairs, triples, quadruples, etc. of reactions. This leads to the identification of a sufficient and non-redundant set of fluxes that must change (i.e., MUST set) to meet a pre-specified overproduction target. Starting with this set we subsequently extract a minimal set of fluxes that must actively be forced through genetic manipulations (i.e., FORCE set) to ensure that all fluxes in the network are consistent with the overproduction objective. We demonstrate our OptForce framework for succinate production in Escherichia coli using the most recent in silico E. coli model, iAF1260. The method not only recapitulates existing engineering strategies but also reveals non-intuitive ones that boost succinate production by performing coordinated changes on pathways distant from the last steps of succinate synthesis

Payments and quality of care in private for-profit and public hospitals in Greece

<p>Abstract</p> <p>Background</p> <p>Empirical evidence on how ownership type affects the quality and cost of medical care is growing, and debate on these topics is ongoing. Despite the fact that the private sector is a major provider of hospital services in Greece, little comparative information on private versus public sector hospitals is available. The aim of the present study was to describe and compare the operation and performance of private for-profit (PFP) and public hospitals in Greece, focusing on differences in nurse staffing rates, average lengths of stay (ALoS), and Social Health Insurance (SHI) payments for hospital care per patient discharged.</p> <p>Methods</p> <p>Five different datasets were prepared and analyzed, two of which were derived from information provided by the National Statistical Service (NSS) of Greece and the other three from data held by the three largest SHI schemes in the country. All data referred to the 3-year period from 2001 to 2003.</p> <p>Results</p> <p>PFP hospitals in Greece are smaller than public hospitals, with lower patient occupancy, and have lower staffing rates of all types of nurses and highly qualified nurses compared with public hospitals. Calculation of ALoS using NSS data yielded mixed results, whereas calculations of ALoS and SHI payments using SHI data gave results clearly favoring the public hospital sector in terms of cost-efficiency; in all years examined, over all specialties and all SHI schemes included in our study, unweighted ALoS and SHI payments for hospital care per discharge were higher for PFP facilities.</p> <p>Conclusions</p> <p>In a mixed healthcare system, such as that in Greece, significant performance differences were observed between PFP and public hospitals. Close monitoring of healthcare provision by hospital ownership type will be essential to permit evidence-based decisions on the future of the public/private mix in terms of healthcare provision.</p

Comparative transcriptomics of drought responses in Populus: a meta-analysis of genome-wide expression profiling in mature leaves and root apices across two genotypes

<p>Abstract</p> <p>Background</p> <p>Comparative genomics has emerged as a promising means of unravelling the molecular networks underlying complex traits such as drought tolerance. Here we assess the genotype-dependent component of the drought-induced transcriptome response in two poplar genotypes differing in drought tolerance. Drought-induced responses were analysed in leaves and root apices and were compared with available transcriptome data from other <it>Populus </it>species.</p> <p>Results</p> <p>Using a multi-species designed microarray, a genomic DNA-based selection of probesets provided an unambiguous between-genotype comparison. Analyses of functional group enrichment enabled the extraction of processes physiologically relevant to drought response. The drought-driven changes in gene expression occurring in root apices were consistent across treatments and genotypes. For mature leaves, the transcriptome response varied weakly but in accordance with the duration of water deficit. A differential clustering algorithm revealed similar and divergent gene co-expression patterns among the two genotypes. Since moderate stress levels induced similar physiological responses in both genotypes, the genotype-dependent transcriptional responses could be considered as intrinsic divergences in genome functioning. Our meta-analysis detected several candidate genes and processes that are differentially regulated in root and leaf, potentially under developmental control, and preferentially involved in early and long-term responses to drought.</p> <p>Conclusions</p> <p>In poplar, the well-known drought-induced activation of sensing and signalling cascades was specific to the early response in leaves but was found to be general in root apices. Comparing our results to what is known in arabidopsis, we found that transcriptional remodelling included signalling and a response to energy deficit in roots in parallel with transcriptional indices of hampered assimilation in leaves, particularly in the drought-sensitive poplar genotype.</p

Do schistosome vaccine trials in mice have an intrinsic flaw that generates spurious protection data?

The laboratory mouse has been widely used to test the efficacy of schistosome vaccines and a long list of candidates has emerged from this work, many of them abundant internal proteins. These antigens do not have an additive effect when co-administered, or delivered as SWAP homogenate, a quarter of which comprises multiple candidates; the observed protection has an apparent ceiling of 40–50 %. We contend that the low level of maturation of penetrating cercariae (~32 % for Schistosoma mansoni) is a major limitation of the model since 68/100 parasites fail to mature in naïve mice due to natural causes. The pulmonary capillary bed is the obstacle encountered by schistosomula en route to the portal system. The fragility of pulmonary capillaries and their susceptibility to a cytokine-induced vascular leak syndrome have been documented. During lung transit schistosomula burst into the alveolar spaces, and possess only a limited capacity to re-enter tissues. The acquired immunity elicited by the radiation attenuated (RA) cercarial vaccine relies on a pulmonary inflammatory response, involving cytokines such as IFNγ and TNFα, to deflect additional parasites into the alveoli. A principal difference between antigen vaccine protocols and the RA vaccine is the short interval between the last antigen boost and cercarial challenge of mice (often two weeks). Thus, after antigen vaccination, challenge parasites will reach the lungs when both activated T cells and cytokine levels are maximal in the circulation. We propose that “protection” in this situation is the result of physiological effects on the pulmonary blood vessels, increasing the proportion of parasites that enter the alveoli. This hypothesis will explain why internal antigens, which are unlikely to interact with the immune response in a living schistosomulum, plus a variety of heterologous proteins, can reduce the level of maturation in a non-antigen-specific way. These proteins are “successful” precisely because they have not been selected for immunological silence. The same arguments apply to vaccine experiments with S. japonicum in the mouse model; this schistosome species seems a more robust parasite, even harder to eliminate by acquired immune responses. We propose a number of ways in which our conclusions may be tested

A922 Sequential measurement of 1 hour creatinine clearance (1-CRCL) in critically ill patients at risk of acute kidney injury (AKI)

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