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All sourcesNASA NTRSDOE OSTI
DOE OSTI2022

Science Requirements and Detector Concepts for the Electron-Ion Collider: EIC Yellow Report

Here, this report describes the physics case, the resulting detector requirements, and the evolving detector concepts for the experimental program at the Electron-Ion Collider (EIC). The EIC will be a powerful new high-luminosity facility in the United States with the capability to collide high-energy electron beams with high-energy proton and ion beams, providing access to those regions in the nucleon and nuclei where their structure is dominated by gluons. Moreover, polarized beams in the EIC will give unprecedented access to the spatial and spin structure of the proton, neutron, and light ions. The studies leading to this document were commissioned and organized by the EIC User Group with the objective of advancing the state and detail of the physics program and developing detector concepts that meet the emerging requirements in preparation for the realization of the EIC. The effort aims to provide the basis for further development of concepts for experimental equipment best suited for the science needs, including the importance of two complementary detectors and interaction regions. This report consists of three volumes. Volume I is an executive summary of our findings and developed concepts. In Volume II we describe studies of a wide range of physics measurements and the emerging requirements on detector acceptance and performance. Volume III discusses general-purpose detector concepts and the underlying technologies to meet the physics requirements. These considerations will form the basis for a world-class experimental program that aims to increase our understanding of the fundamental structure of all visible matter.

73 NUCLEAR PHYSICS AND RADIATION PHYSICS↗
DOE OSTI2016

Predicting environmental mitigation requirements for hydropower projects through the integration of biophysical and socio-political geographies

Uncertainty about environmental mitigation needs at existing and proposed hydropower projects makes it difficult for stakeholders to minimize environmental impacts. Hydropower developers and operators desire tools to better anticipate mitigation requirements, while natural resource managers and regulators need tools to evaluate different mitigation scenarios and order effective mitigation. Here we sought to examine the feasibility of using a suite of multidisciplinary explanatory variables within a spatially explicit modeling framework to fit predictive models for future environmental mitigation requirements at hydropower projects across the conterminous U.S. Using a database comprised of mitigation requirements from more than 300 hydropower project licenses, we were able to successfully fit models for nearly 50 types of environmental mitigation and to apply the predictive models to a set of more than 500 non-powered dams identified as having hydropower potential. The results demonstrate that mitigation requirements have been a result of a range of factors, from biological and hydrological to political and cultural. Furthermore, project developers can use these models to inform cost projections and design considerations, while regulators can use the models to more quickly identify likely environmental issues and potential solutions, hopefully resulting in more timely and more effective decisions on environmental mitigation.

13 HYDRO ENERGY↗
DOE OSTI2013

Requirement of B-Raf, C-Raf, and A-Raf for the growth and survival of mouse embryonic stem cells

Extracellular signal-regulated kinases (ERKs) have been implicated to be dispensable for self-renewal of mouse embryonic stem (ES) cells, and simultaneous inhibition of both ERK signaling and glycogen synthase kinase 3 (GSK3) not only allows mouse ES cells to self-renew independent of extracellular stimuli but also enables more efficient derivation of naïve ES cells from mouse and rat strains. Interestingly, some ERKs stay active in mouse ES cells which are maintained in regular medium containing leukemia inhibitory factor (LIF) and bone morphogenetic protein (BMP). Yet, the upstream signaling for ERK activation and their roles in mouse ES cells, other than promoting or priming differentiation, have not been determined. Here we found that mouse ES cells express three forms of Raf kinases, A-Raf, B-Raf, and C-Raf. Knocking-down each single Raf member failed to affect the sustained ERK activity, neither did A-Raf and B-Raf double knockdown or B-Raf and C-Raf double knockdown change it in ES cells. Interestingly, B-Raf and C-Raf double knockdown, not A-Raf and B-Raf knockdown, inhibited the maximal ERK activation induced by LIF, concomitant with the slower growth of ES cells. On the other hand, A-Raf, B-Raf, and C-Raf triple knockdown markedly inhibited both the maximal and sustained ERK activity in ES cells. Moreover, Raf triple knockdown, similar to the treatment of U-0126, an MEK inhibitor, significantly inhibited the survival and proliferation of ES cells, thereby compromising the colony propagation of mouse ES cells. In summary, our data demonstrate that all three Raf members are required for ERK activation in mouse ES cells and are involved in growth and survival of mouse ES cells. - Highlights: ●Mouse ES (mES) cells express all three Raf members, A-Raf, B-Raf, and C-Raf. ●Leukemia inhibitory factor (LIF) temporally activates ERKs in mES cells. ●B-Raf and C-Raf are required for LIF-induced maximal ERKs activity in mES cells. ●All Raf members are required for LIF-induced sustained ERK activity in mES cells. ●All Raf members are required the survival and proliferation of mES cells.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2026

Sulfite Is Not Required for N 2 Reduction Catalyzed by Mo-Nitrogenase

Mo-nitrogenase catalyzes the reduction of dinitrogen (N 2 ) to two ammonia (NH 3 ) at the active-site FeMo-cofactor. Substrate activation requires the accumulation of three or four electrons and protons as two Fe-bound hydrides and is coupled to obligatory H 2 release through reductive hydride elimination. Subsequent delivery of four or five additional electrons and protons to the bound N 2 yields two NH 3 molecules. Increasing evidence suggests that at least one belt sulfide within FeMo-cofactor is dynamically involved in the catalytic cycle. A recent report further proposed that sulfite (SO 3 2− ) is required for N 2 reduction, with sulfite binding required for NH 3 release and a subsequent six-electron reduction of the bound sulfite to regenerate the resting cofactor. To test this proposal, we conducted turnover studies of Mo-nitrogenase under sulfite-free conditions using a reduced viologen as reductant and protein preparations devoid of dithionite or sulfite. Under these conditions, nitrogenase effectively catalyzed both N 2 reduction and proton reduction, exhibiting steady-state turnover under N 2 for 6 min, with a turnover number exceeding 150, approaching that observed with dithionite as reductant. The same H 2 -formed/N 2 -reduced ratio was observed whether dithionite or the viologen species was used as reductant. Further, EPR spectroscopic analyses showed that the FeMo-cofactor returned to its resting state after multiple catalytic cycles in the absence of sulfite. Finally, physiological bypass of sulfite formation does not affect the capacity for diazotrophic growth of the model nitrogen-fixing organism Azotobacter vinelandii . Finally, these results demonstrate that sulfite is not required for Mo-nitrogenase-catalyzed N 2 reduction either in vitro or in vivo .

37 INORGANIC, ORGANIC, PHYSICAL, AND ANALYTICAL CH↗
DOE OSTI2015

The study on knowledge transferring incentive for information system requirement development

Information system requirement development is a process of users’ knowledge sharing and transferring. However the tacit requirements developing is a main problem during requirement development process, for the reason of difficult to encoding, express, and communicate. Knowledge fusion and corporate effort is needed to finding tacit requirements. Under this background, our paper try to find out the rule of effort dynamic evolutionary of software developer and user by building an evolutionary game model on the condition of incentive system. And in addition this paper provides an in depth discussion at the end of this paper.

97 MATHEMATICS AND COMPUTING↗
DOE OSTI2020

Investigating the minimum post-burst energy required to function an exploding bridgewire detonator

The minimum energy required after bridge-burst to function a commercially available RP-1 exploding bridgewire detonator with a gold bridgewire was investigated using two capacitor discharge units (CDUs) making use of state-of-the-art on/off high-voltage high-current switches. Both switches are capable of shutting off the current flow in ≈100ns and the conduction time was varied systematically to limit the duration of electrical energy delivered from the CDU until a detonation go/no-go threshold was reached. It was found that the minimal extra electrical energy required for a soft-fire (a statistically marginal go) was approximately half that required for a hard-fire (a definite go). Furthermore, the measured energy required to function the RP-1 detonator from the 11.5 kV CDU was approximately 20 mJ greater than for the 600 V CDU, and it is postulated that this was due to arcing across the detonator terminals and an associated current shunt that was not efficient at depositing energy conducive to detonator functioning. Measurements of the bridgewire temperatures bursting in air showed that the maximum temperature reached was consistent (≈18kK) when sufficient energy was delivered that the detonator functioned (either soft- or hard-fire), but that the temperature for a sub-threshold bridgewire burst was significantly cooler at approximately 8 kK. Furthermore, it was demonstrated that the intensity of the light emitted at burst scaled with deposited energy. A related finding is that the light emission from the bridgewire peaks some tens of nanoseconds after both peak voltage and peak electrical power.

45 MILITARY TECHNOLOGY, WEAPONRY, AND NATIONAL DEF↗
DOE OSTI2022

Polarization angle requirements for CMB B-mode experiments. Application to the LiteBIRD satellite

A methodology to provide the polarization angle requirements for different sets of detectors, at a given frequency of a CMB polarization experiment, is presented. The uncertainties in the polarization angle of each detector set are related to a given bias on the tensor-to-scalar ratio r parameter. The approach is grounded in using a linear combination of the detector sets to obtain the CMB polarization signal. In addition, assuming that the uncertainties on the polarization angle are in the small angle limit (lower than a few degrees), it is possible to derive analytic expressions to establish the requirements. The methodology also accounts for possible correlations among detectors, that may originate from the optics, wafers, etc. The approach is applied to the LiteBIRD space mission. We show that, for the most restrictive case (i.e., full correlation of the polarization angle systematics among detector sets), the requirements on the polarization angle uncertainties are of around 1 arcmin at the most sensitive frequency bands (i.e., ≈ 150 GHz) and of few tens of arcmin at the lowest (i.e., ≈ 40 GHz) and highest (i.e., ≈ 400 GHz) observational bands. Conversely, for the least restrictive case (i.e., no correlation of the polarization angle systematics among detector sets), the requirements are ≈ 5 times less restrictive than for the previous scenario. At the global and the telescope levels, polarization angle knowledge of a few arcmins is sufficient for correlated global systematic errors and can be relaxed by a factor of two for fully uncorrelated errors in detector polarization angle. The reported uncertainty levels are needed in order to have the bias on r due to systematics below the limit established by the LiteBIRD collaboration.

79 ASTRONOMY AND ASTROPHYSICS↗
DOE OSTI2018

Improving BA Control Performance Through Advanced Regulation Requirements Prediction

The paper presents a comprehensive approach to predict balancing authority (BA) regulation requirements in order to improve BAs control performance. The proposed probabilistic approach takes into account multiple uncertain and time dependent factors affecting the system balance (e.g., wind and solar generation, electrical loads). A set of methods to predict the regulation has been developed: (1) probabilistic analysis of regulation requirements with respect to time of the day, (2) linear or nonlinear regression models linking regulation requirements to different influencing factors, (3) adopting time series forecasting techniques (e.g., autoregressive integrated moving average (ARIMA)) to account for temporal continuity and auto-correlation in the regulation requirements data. Proposed methodology has been tested and validated using actual California Independent System Operator (ISO) data.

Solar Generation, Wind Generation, Area Control Er↗
DOE OSTI2020

Probabilistic Zonal Reserve Requirements for Improved Energy Deliverability with Wind Power

In power systems with high penetration of renewable energy resources, uncertainty and variability of these stochastic resources introduce additional challenges for the operation of power systems. To improve the power system's reliability in the face of uncertainty, reserves are required as additional generation capacity to rebalance the power system following random disturbances. However, reserve deliverability is not guaranteed, because it may encounter potential transmission line congestion. Zonal reserve requirements can address this issue, but operators lack efficient ways to allocate reserves to zones while accounting for wind power forecast uncertainty. Herein we propose a methodology for probabilistic zonal reserve requirements to address wind power forecast uncertainties. This method estimates the probability distribution of line flows based on the system generation margin and injection shift factor. This estimate is then used to construct pre-defined and post-zonal reserve requirements. Case studies demonstrate that the proposed method efficiently schedules energy and reserves to balance energy and manage deliverability with wind power forecast uncertainty.We also discuss operational implications of the proposed method.

17 WIND ENERGY↗
DOE OSTI2020

Indoor air quality in California homes with code-required mechanical ventilation

Data were collected in 70 detached houses built in 2011-2017 in compliance with the mechanical ventilation requirements of California's building energy efficiency standards. Each home was monitored for a 1-week period with windows closed and the central mechanical ventilation system operating. Pollutant measurements included time-resolved fine particulate matter (PM 2.5 ) indoors and outdoors and formaldehyde and carbon dioxide (CO 2 ) indoors. Time-integrated measurements were made for formaldehyde, NO 2 , and nitrogen oxides (NO X ) indoors and outdoors. Operation of the cooktop, range hood, and other exhaust fans was continuously recorded during the monitoring period. Onetime diagnostic measurements included mechanical airflows and envelope and duct system air leakage. All homes met or were very close to meeting the ventilation requirements. On average, the dwelling unit ventilation fan moved 50% more airflow than the minimum requirement. Pollutant concentrations were similar to or lower than those reported in a 2006-2007 study of California new homes built in 2002-2005. Mean and median indoor concentrations were lower by 44% and 38% for formaldehyde and 44% and 54% for PM 2.5 . Ventilation fans were operating in only 26% of homes when first visited, and the control switches in many homes did not have informative labels as required by building standards.

32 ENERGY CONSERVATION, CONSUMPTION, AND UTILIZATI↗
DOE OSTI2014

Sample size requirements for estimating effective dose from computed tomography using solid-state metal-oxide-semiconductor field-effect transistor dosimetry

Purpose: Effective dose (ED) is a widely used metric for comparing ionizing radiation burden between different imaging modalities, scanners, and scan protocols. In computed tomography (CT), ED can be estimated by performing scans on an anthropomorphic phantom in which metal-oxide-semiconductor field-effect transistor (MOSFET) solid-state dosimeters have been placed to enable organ dose measurements. Here a statistical framework is established to determine the sample size (number of scans) needed for estimating ED to a desired precision and confidence, for a particular scanner and scan protocol, subject to practical limitations. Methods: The statistical scheme involves solving equations which minimize the sample size required for estimating ED to desired precision and confidence. It is subject to a constrained variation of the estimated ED and solved using the Lagrange multiplier method. The scheme incorporates measurement variation introduced both by MOSFET calibration, and by variation in MOSFET readings between repeated CT scans. Sample size requirements are illustrated on cardiac, chest, and abdomen–pelvis CT scans performed on a 320-row scanner and chest CT performed on a 16-row scanner. Results: Sample sizes for estimating ED vary considerably between scanners and protocols. Sample size increases as the required precision or confidence is higher and also as the anticipated ED is lower. For example, for a helical chest protocol, for 95% confidence and 5% precision for the ED, 30 measurements are required on the 320-row scanner and 11 on the 16-row scanner when the anticipated ED is 4 mSv; these sample sizes are 5 and 2, respectively, when the anticipated ED is 10 mSv. Conclusions: Applying the suggested scheme, it was found that even at modest sample sizes, it is feasible to estimate ED with high precision and a high degree of confidence. As CT technology develops enabling ED to be lowered, more MOSFET measurements are needed to estimate ED with the same precision and confidence.

61 RADIATION PROTECTION AND DOSIMETRY↗
DOE OSTI2014

SU-E-P-03: Implementing a Low Dose Lung Screening CT Program Meeting Regulatory Requirements

Purpose: To provide information pertaining to IROC Houston QA Center's (RPC) credentialing process for institutions participating in NCI-sponsored clinical trials. Purpose: Provide guidance to the Radiology Departments with the intent of implementing a Low Dose CT Screening Program using different CT Scanners with multiple techniques within the framework of the required state regulations. Method: State Requirements for the purpose of implementing a Low Dose CT Lung Protocol required working with the Radiology and Pulmonary Department in setting up a Low Dose Screening Protocol designed to reduce the radiation burden to the patients enrolled. Radiation dose measurements (CTDIvol) for various CT manufacturers (Siemens16, Siemens 64, Philips 64, and Neusoft128) for three different weight based protocols. All scans were reviewed by the Radiologist. Prior to starting a low dose lung screening protocol, information had to be submitted to the state for approval. Performing a Healing Arts protocol requires extensive information. This not only includes name and address of the applicant but a detailed description of the disease, the x-ray examination and the population to be examined. The unit had to be tested by a qualified expert using the technique charts. The credentials of all the operators, the supervisors and the Radiologists had to be submitted to the state. Results: All the appropriate documentation was sent to the state for review. The measured results between the Low Dose Protocol versus the default Adult Chest Protocol showed that there was a dose reduction of 65% for small (100-150 lb.) patient, 75% for the Medium patient (151-250 lbs.), and a 55% reduction for the Large patient ( over 250 lbs.). Conclusion: Measured results indicated that the Low Dose Protocol indeed lowered the screening patient's radiation dose and the institution was able to submit the protocol to the State's regulators.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2016

WE-AB-206-02: ACR Ultrasound Accreditation: Requirements and Pitfalls

The involvement of medical physicists in diagnostic ultrasound imaging service is increasing due to QC and accreditation requirements. The goal of this ultrasound hands-on workshop is to demonstrate quality control (QC) testing in diagnostic ultrasound and to provide updates in ACR ultrasound accreditation requirements. The first half of this workshop will include two presentations reviewing diagnostic ultrasound QA/QC and ACR ultrasound accreditation requirements. The second half of the workshop will include live demonstrations of basic QC tests. An array of ultrasound testing phantoms and ultrasound scanners will be available for attendees to learn diagnostic ultrasound QC in a hands-on environment with live demonstrations and on-site instructors. The targeted attendees are medical physicists in diagnostic imaging. Learning Objectives: Gain familiarity with common elements of a QA/QC program for diagnostic ultrasound imaging dentify QC tools available for testing diagnostic ultrasound systems and learn how to use these tools Learn ACR ultrasound accreditation requirements Jennifer Walter is an employee of American College of Radiology on Ultrasound Accreditation.

61 RADIATION PROTECTION AND DOSIMETRY↗
DOE OSTI2017

An Amoebal Grazer of Cyanobacteria Requires Cobalamin Produced by Heterotrophic Bacteria

Amoebae are unicellular eukaryotes that consume microbial prey through phagocytosis, playing a role in shaping microbial food webs. Many amoebal species can be cultivated axenically in rich media or monoxenically with a single bacterial prey species. Here, we characterize heterolobosean amoeba LPG3, a recent natural isolate, which is unable to grow on unicellular cyanobacteria, its primary food source, in the absence of a heterotrophic bacterium, a Pseudomonas species coisolate. To investigate the molecular basis of this requirement for heterotrophic bacteria, we performed a screen using the defined nonredundant transposon library of Vibrio cholerae , which implicated genes in corrinoid uptake and biosynthesis. Furthermore, cobalamin synthase deletion mutations in V. cholerae and the Pseudomonas species coisolate do not support the growth of amoeba LPG3 on cyanobacteria. While cyanobacteria are robust producers of a corrinoid variant called pseudocobalamin, this variant does not support the growth of amoeba LPG3. Instead, we show that it requires cobalamin that is produced by the Pseudomonas species coisolate. The diversity of eukaryotes utilizing corrinoids is poorly understood, and this amoebal corrinoid auxotroph serves as a model for examining predator-prey interactions and micronutrient transfer in bacterivores underpinning microbial food webs.Cyanobacteria are important primary producers in aquatic environments, where they are grazed upon by a variety of phagotrophic protists and, hence, have an impact on nutrient flux at the base of microbial food webs. Here, we characterize amoebal isolate LPG3, which consumes cyanobacteria as its primary food source but also requires heterotrophic bacteria as a source of corrinoid vitamins. Amoeba LPG3 specifically requires the corrinoid variant produced by heterotrophic bacteria and cannot grow on cyanobacteria alone, as they produce a different corrinoid variant. This same corrinoid specificity is also exhibited by other eukaryotes, including humans and algae. This amoebal model system allows us to dissect predator-prey interactions to uncover factors that may shape microbial food webs while also providing insight into corrinoid specificity in eukaryotes.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2022

Macro Analysis to Estimate Electric Vehicles Fast-Charging Infrastructure Requirements in Small Urban Areas

Electric vehicles (EVs) are known to reduce emissions and fossil fuel dependency. However, the limited range, long charging time, and inadequate charging infrastructure have hampered the adoption of EVs. The current EV charging infrastructure planning studies and tools require detailed information, extensive resources, and skills that can be a significant barrier to urban areas for finding the required charging infrastructure to support a targeted EV market share. This study generates regression models to estimate the number of direct current fast charging stations and the chargers to support the EV charging demand for urban areas. These models provide macro-level estimates of the required infrastructure investment in urban areas, which can be easily implemented by policy-makers and city planners. This study incorporates data obtained from applying a disaggregate optimization-based charger placement model, developed recently by the same authors, for multiple case studies to generate the required data to calibrate the macro-level models, in the state of Michigan. This simulated data set includes the number of charging stations and chargers for each market share, technology advancement scenario, and the transportation network topology. The results show that the number of charging stations reduces with battery size and charging power and increases with EV market share and the road network lane length. The number of chargers reduces with charging power, whereas it increases with battery size, EV market share, and vehicle miles traveled in the system. The model developed here can be applied to any state having urban characteristics and weather conditions similar to Michigan.

Engineering↗
DOE OSTI2022

An initial investigation of accuracy required for the identification of small molecules in complex samples using quantum chemical calculated NMR chemical shifts

The majority of primary and secondary metabolites in nature have yet to be identified, representing a major challenge for metabolomics studies that currently require reference libraries from analyses of authentic compounds. Using currently available analytical methods, complete chemical characterization of metabolomes is infeasible for both technical and economic reasons. For example, unambiguous identification of metabolites is limited by the availability of authentic chemical standards, which, for the majority of molecules, do not exist. Computationally predicted or calculated data are a viable solution to expand the currently limited metabolite reference libraries, if such methods are shown to be sufficiently accurate. For example, determining nuclear magnetic resonance (NMR) spectroscopy spectra in silico has shown promise in the identification and delineation of metabolite structures. Many researchers have been taking advantage of density functional theory (DFT), a computationally inexpensive yet reputable method for the prediction of carbon and proton NMR spectra of metabolites. However, such methods are expected to have some error in predicted 13 >C and 1 H NMR spectra with respect to experimentally measured values. This leads us to the question–what accuracy is required in predicted 13 C and 1 H NMR chemical shifts for confident metabolite identification? Using the set of 11,716 small molecules found in the Human Metabolome Database (HMDB), we simulated both experimental and theoretical NMR chemical shift databases. We investigated the level of accuracy required for identification of metabolites in simulated pure and impure samples by matching predicted chemical shifts to experimental data. We found 90% or more of molecules in simulated pure samples can be successfully identified when errors of 1 H and 13 C chemical shifts in water are below 0.6 and 7.1 ppm, respectively, and below 0.5 and 4.6 ppm in chloroform solvation, respectively. In simulated complex mixtures, as the complexity of the mixture increased, greater accuracy of the calculated chemical shifts was required, as expected. However, if the number of molecules in the mixture is known, e.g., when NMR is combined with MS and sample complexity is low, the likelihood of confident molecular identification increased by 90%.

37 INORGANIC, ORGANIC, PHYSICAL, AND ANALYTICAL CH↗
DOE OSTI2021

Computation of condition-dependent proteome allocation reveals variability in the macro and micro nutrient requirements for growth

Sustaining a robust metabolic network requires a balanced and fully functioning proteome. In addition to amino acids, many enzymes require cofactors (coenzymes and engrafted prosthetic groups) to function properly. Extensively validated resource allocation models, such as genome-scale models of metabolism and gene expression (ME-models), have the ability to compute an optimal proteome composition underlying a metabolic phenotype, including the provision of all required cofactors. Here we apply the ME-model for Escherichia coli K-12 MG1655 to computationally examine how environmental conditions change the proteome and its accompanying cofactor usage. We found that: (1) The cofactor requirements computed by the ME-model mostly agree with the standard biomass objective function used in models of metabolism alone (M-models); (2) ME-model computations reveal non-intuitive variability in cofactor use under different growth conditions; (3) An analysis of ME-model predicted protein use in aerobic and anaerobic conditions suggests an enrichment in the use of peroxyl scavenging acids in the proteins used to sustain aerobic growth; (4) The ME-model could describe how limitation in key protein components affect the metabolic state of E . coli . Genome-scale models have thus reached a level of sophistication where they reveal intricate properties of functional proteomes and how they support different E . coli lifestyles.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2013

Diverse Secreted Effectors Are Required for Salmonella Persistence in a Mouse Infection Model

Salmonella enterica serovar Typhimurium causes typhoid-like disease in mice and is a model of typhoid fever in humans. One of the hallmarks of typhoid is persistence, the ability of the bacteria to survive in the host weeks after infection. Virulence factors called effectors facilitate this process by direct transfer to the cytoplasm of infected cells thereby subverting cellular processes. Secretion of effectors to the cell cytoplasm takes place through multiple routes, including two separate type III secretion (T3SS) apparati as well as outer membrane vesicles. The two T3SS are encoded on separate pathogenicity islands, SPI-1 and -2, with SPI-1 more strongly associated with the intestinal phase of infection, and SPI-2 with the systemic phase. Both T3SS are required for persistence, but the effectors required have not been systematically evaluated. In this study, mutations in 48 described effectors were tested for persistence. We replaced each effector with a specific DNA barcode sequence by allelic exchange and co-infected with a wild-type reference to calculate the ratio of wild-type parent to mutant at different times after infection. The competitive index (CI) was determined by quantitative PCR in which primers that correspond to the barcode were used for amplification. Mutations in all but seven effectors reduced persistence demonstrating that most effectors were required. One exception was CigR, a recently discovered effector that is widely conserved throughout enteric bacteria. Deletion of cigR increased lethality, suggesting that it may be an anti-virulence factor. The fact that almost all Salmonella effectors are required for persistence argues against redundant functions. This is different from effector repertoires in other intracellular pathogens such as Legionella.

Environmental Molecular Sciences Laboratory↗