Targeted Mutagenesis in the Malaria Mosquito Using TALE Nucleases

Anopheles gambiae, the main mosquito vector of human malaria, is a challenging organism to manipulate genetically. As a consequence, reverse genetics studies in this disease vector have been largely limited to RNA interference experiments. Here, we report the targeted disruption of the immunity gene TEP1 using transgenic expression of Transcription-Activator Like Effector Nucleases (TALENs), and the isolation of several TEP1 mutant A. gambiae lines. These mutations inhibited protein production and rendered TEP1 mutants hypersusceptible to Plasmodium berghei. The TALEN technology opens up new avenues for genetic analysis in this disease vector and may offer novel biotechnology-based approaches for malaria control

Transgenic Expression of the Anti-parasitic Factor TEP1 in the Malaria Mosquito Anopheles gambiae

Mosquitoes genetically engineered to be resistant to Plasmodium parasites represent a promising novel approach in the fight against malaria. The insect immune system itself is a source of anti-parasitic genes potentially exploitable for transgenic designs. The Anopheles gambiae thioester containing protein 1 (TEP1) is a potent anti-parasitic protein. TEP1 is secreted and circulates in the mosquito hemolymph, where its activated cleaved form binds and eliminates malaria parasites. Here we investigated whether TEP1 can be used to create malaria resistant mosquitoes. Using a GFP reporter transgene, we determined that the fat body is the main site of TEP1 expression. We generated transgenic mosquitoes that express TEP1r, a potent refractory allele of TEP1, in the fat body and examined the activity of the transgenic protein in wild-type or TEP1 mutant genetic backgrounds. Transgenic TEP1r rescued loss-of-function mutations, but did not increase parasite resistance in the presence of a wild-type susceptible allele. Consistent with previous reports, TEP1 protein expressed from the transgene in the fat body was taken up by hemocytes upon a challenge with injected bacteria. Furthermore, although maturation of transgenic TEP1 into the cleaved form was impaired in one of the TEP1 mutant lines, it was still sufficient to reduce parasite numbers and induce parasite melanization. We also report here the first use of Transcription Activator Like Effectors (TALEs) in Anopheles gambiae to stimulate expression of endogenous TEP1. We found that artificial elevation of TEP1 expression remains moderate in vivo and that enhancement of endogenous TEP1 expression did not result in increased resistance to Plasmodium. Taken together, our results reveal the difficulty of artificially influencing TEP1-mediated Plasmodium resistance, and contribute to further our understanding of the molecular mechanisms underlying mosquito resistance to Plasmodium parasites

Thermal Infrared MMTAO Observations of the HR 8799 Planetary System

We present direct imaging observations at wavelengths of 3.3, 3.8 (L',band), and 4.8 (M band) microns, for the planetary system surrounding HR 8799. All three planets are detected at L'. The c and d component are detected at 3.3 microns, and upper limits are derived from the M band observations. These observations provide useful constraints on warm giant planet atmospheres. We discuss the current age constraints on the HR 8799 system, and show that several potential co-eval objects can be excluded from being co-moving with the star. Comparison of the photometry is made to models for giant planet atmospheres. Models which include non-equilibrium chemistry provide a reasonable match to the colors of c and d. From the observed colors in the thermal infrared we estimate T_eff < 960 K for b, and T_eff=1300 and 1170 K for c and d, respectively. This provides an independent check on the effective temperatures and thus masses of the objects from the Marois 2008 results.Comment: 16 pages, 6 figures, accepted to Ap

The InfraRed Imaging Spectrograph (IRIS) for TMT: latest science cases and simulations

The Thirty Meter Telescope (TMT) first light instrument IRIS (Infrared Imaging Spectrograph) will complete its preliminary design phase in 2016. The IRIS instrument design includes a near-infrared (0.85 - 2.4 micron) integral field spectrograph (IFS) and imager that are able to conduct simultaneous diffraction-limited observations behind the advanced adaptive optics system NFIRAOS. The IRIS science cases have continued to be developed and new science studies have been investigated to aid in technical performance and design requirements. In this development phase, the IRIS science team has paid particular attention to the selection of filters, gratings, sensitivities of the entire system, and science cases that will benefit from the parallel mode of the IFS and imaging camera. We present new science cases for IRIS using the latest end-to-end data simulator on the following topics: Solar System bodies, the Galactic center, active galactic nuclei (AGN), and distant gravitationally-lensed galaxies. We then briefly discuss the necessity of an advanced data management system and data reduction pipeline.Comment: 15 pages, 7 figures, SPIE (2016) 9909-0

Dynamical Mass Measurement of the Young Spectroscopic Binary V343 Normae AaAb Resolved With the Gemini Planet Imager

We present new spatially resolved astrometry and photometry from the Gemini Planet Imager of the inner binary of the young multiple star system V343 Normae, which is a member of the beta Pictoris moving group. V343 Normae comprises a K0 and mid-M star in a ~4.5 year orbit (AaAb) and a wide 10" M5 companion (B). By combining these data with archival astrometry and radial velocities we fit the orbit and measure individual masses for both components of M_Aa = 1.10 +/- 0.10 M_sun and M_Ab = 0.290 +/- 0.018 M_sun. Comparing to theoretical isochrones, we find good agreement for the measured masses and JHK band magnitudes of the two components consistent with the age of the beta Pic moving group. We derive a model-dependent age for the beta Pic moving group of 26 +/- 3 Myr by combining our results for V343 Normae with literature measurements for GJ 3305, which is another group member with resolved binary components and dynamical masses.Comment: 12 pages, 7 figures. Accepted to A

Improving and Assessing Planet Sensitivity of the GPI Exoplanet Survey with a Forward Model Matched Filter

We present a new matched filter algorithm for direct detection of point sources in the immediate vicinity of bright stars. The stellar Point Spread Function (PSF) is first subtracted using a Karhunen-Lo\'eve Image Processing (KLIP) algorithm with Angular and Spectral Differential Imaging (ADI and SDI). The KLIP-induced distortion of the astrophysical signal is included in the matched filter template by computing a forward model of the PSF at every position in the image. To optimize the performance of the algorithm, we conduct extensive planet injection and recovery tests and tune the exoplanet spectra template and KLIP reduction aggressiveness to maximize the Signal-to-Noise Ratio (SNR) of the recovered planets. We show that only two spectral templates are necessary to recover any young Jovian exoplanets with minimal SNR loss. We also developed a complete pipeline for the automated detection of point source candidates, the calculation of Receiver Operating Characteristics (ROC), false positives based contrast curves, and completeness contours. We process in a uniform manner more than 330 datasets from the Gemini Planet Imager Exoplanet Survey (GPIES) and assess GPI typical sensitivity as a function of the star and the hypothetical companion spectral type. This work allows for the first time a comparison of different detection algorithms at a survey scale accounting for both planet completeness and false positive rate. We show that the new forward model matched filter allows the detection of $50\%$ fainter objects than a conventional cross-correlation technique with a Gaussian PSF template for the same false positive rate.Comment: ApJ accepte

GPI spectra of HR 8799 c, d, and e from 1.5 to 2.4μ\mum with KLIP Forward Modeling

We explore KLIP forward modeling spectral extraction on Gemini Planet Imager coronagraphic data of HR 8799, using PyKLIP and show algorithm stability with varying KLIP parameters. We report new and re-reduced spectrophotometry of HR 8799 c, d, and e in H & K bands. We discuss a strategy for choosing optimal KLIP PSF subtraction parameters by injecting simulated sources and recovering them over a range of parameters. The K1/K2 spectra for HR 8799 c and d are similar to previously published results from the same dataset. We also present a K band spectrum of HR 8799 e for the first time and show that our H-band spectra agree well with previously published spectra from the VLT/SPHERE instrument. We show that HR 8799 c and d show significant differences in their H & K spectra, but do not find any conclusive differences between d and e or c and e, likely due to large error bars in the recovered spectrum of e. Compared to M, L, and T-type field brown dwarfs, all three planets are most consistent with mid and late L spectral types. All objects are consistent with low gravity but a lack of standard spectra for low gravity limit the ability to fit the best spectral type. We discuss how dedicated modeling efforts can better fit HR 8799 planets' near-IR flux and discuss how differences between the properties of these planets can be further explored.Comment: Accepted to AJ, 25 pages, 16 Figure

Transgenic Expression of the Anti-parasitic Factor TEP1 in the Malaria Mosquito Anopheles gambiae

Mosquitoes genetically engineered to be resistant to Plasmodium parasites represent a promising novel approach in the fight against malaria. The insect immune system itself is a source of anti-parasitic genes potentially exploitable for transgenic designs. The Anopheles gambiae thioester containing protein 1 (TEP1) is a potent anti-parasitic protein. TEP1 is secreted and circulates in the mosquito hemolymph, where its activated cleaved form binds and eliminates malaria parasites. Here we investigated whether TEP1 can be used to create malaria resistant mosquitoes. Using a GFP reporter transgene, we determined that the fat body is the main site of TEP1 expression. We generated transgenic mosquitoes that express TEP1r, a potent refractory allele of TEP1, in the fat body and examined the activity of the transgenic protein in wild-type or TEP1 mutant genetic backgrounds. Transgenic TEP1r rescued loss-of-function mutations, but did not increase parasite resistance in the presence of a wild-type susceptible allele. Consistent with previous reports, TEP1 protein expressed from the transgene in the fat body was taken up by hemocytes upon a challenge with injected bacteria. Furthermore, although maturation of transgenic TEP1 into the cleaved form was impaired in one of the TEP1 mutant lines, it was still sufficient to reduce parasite numbers and induce parasite melanization. We also report here the first use of Transcription Activator Like Effectors (TALEs) in Anopheles gambiae to stimulate expression of endogenous TEP1. We found that artificial elevation of TEP1 expression remains moderate in vivo and that enhancement of endogenous TEP1 expression did not result in increased resistance to Plasmodium. Taken together, our results reveal the difficulty of artificially influencing TEP1-mediated Plasmodium resistance, and contribute to further our understanding of the molecular mechanisms underlying mosquito resistance to Plasmodium parasites

Performance of the Gemini Planet Imager Non-Redundant Mask and spectroscopy of two close-separation binaries HR 2690 and HD 142527

The Gemini Planet Imager (GPI) contains a 10-hole non-redundant mask (NRM), enabling interferometric resolution in complement to its coronagraphic capabilities. The NRM operates both in spectroscopic (integral field spectrograph, henceforth IFS) and polarimetric configurations. NRM observations were taken between 2013 and 2016 to characterize its performance. Most observations were taken in spectroscopic mode with the goal of obtaining precise astrometry and spectroscopy of faint companions to bright stars. We find a clear correlation between residual wavefront error measured by the AO system and the contrast sensitivity by comparing phase errors in observations of the same source, taken on different dates. We find a typical 5-$\sigma$ contrast sensitivity of $2-3~\times~10^{-3}$ at $\sim\lambda/D$. We explore the accuracy of spectral extraction of secondary components of binary systems by recovering the signal from a simulated source injected into several datasets. We outline data reduction procedures unique to GPI's IFS and describe a newly public data pipeline used for the presented analyses. We demonstrate recovery of astrometry and spectroscopy of two known companions to HR 2690 and HD 142527. NRM+polarimetry observations achieve differential visibility precision of $\sigma\sim0.4\%$ in the best case. We discuss its limitations on Gemini-S/GPI for resolving inner regions of protoplanetary disks and prospects for future upgrades. We summarize lessons learned in observing with NRM in spectroscopic and polarimetric modes.Comment: Accepted to AJ, 22 pages, 14 figure