Energy-Filtering Transmission Electron Microscopy of Biological Specimens

By energy-filtering transmission electron microscopy (EFTEM) electrons can be separated by their energy losses. An electron-energy filter, added to the microscope column allows the measurement of the energy distribution of transmitted electrons that have lost energy (\u3c 2,000 eV, with an energy resolution of ~ 1 eV). These filtered electrons, recorded either as a spectrum or as an image, are composed of two parts superimposed on top of each other: (a) the unspecific energy-loss population (= the continuum) and (b) the specific element-related energy-loss population (= the edges). At the edges, electron data in spectra and images are mathematically processed, to obtain the desired element-related net-intensity values or images. These data are related to the total transmitted electron intensity, from the zero-and low-loss spectral region giving the relative spectral-or image intensity ratios (SR*x, IR*x), which can be related to the element concentration. The acquisition of the zero-loss and low-loss data is hampered by the restricted dynamic range of the TV camera. By improvements through the introduction of calibrated attenuation filters in the optical path to the TV-camera, more reliable values for SR*x and IR*x can be acquired. By addition of Bio-standards adjacent to the tissue, a known and unknown concentration of the element present in the same ultrathin section and the bias in the concentration estimation, can be obtained. Some practical examples are given for the estimation of the iron concentration in siderosomes, boron in melanosomes and calcium in calcium oxalate monohydrate crystals

The KELT Follow-Up Network And Transit False-Positive Catalog: Pre-Vetted False Positives For TESS

The Kilodegree Extremely Little Telescope (KELT) project has been conducting a photometric survey of transiting planets orbiting bright stars for over 10 years. The KELT images have a pixel scale of ~23\u27\u27 pixel⁻¹—very similar to that of NASA\u27s Transiting Exoplanet Survey Satellite (TESS)—as well as a large point-spread function, and the KELT reduction pipeline uses a weighted photometric aperture with radius 3\u27. At this angular scale, multiple stars are typically blended in the photometric apertures. In order to identify false positives and confirm transiting exoplanets, we have assembled a follow-up network (KELT-FUN) to conduct imaging with spatial resolution, cadence, and photometric precision higher than the KELT telescopes, as well as spectroscopic observations of the candidate host stars. The KELT-FUN team has followed-up over 1600 planet candidates since 2011, resulting in more than 20 planet discoveries. Excluding ~450 false alarms of non-astrophysical origin (i.e., instrumental noise or systematics), we present an all-sky catalog of the 1128 bright stars (6 \u3c V \u3c 13) that show transit-like features in the KELT light curves, but which were subsequently determined to be astrophysical false positives (FPs) after photometric and/or spectroscopic follow-up observations. The KELT-FUN team continues to pursue KELT and other planet candidates and will eventually follow up certain classes of TESS candidates. The KELT FP catalog will help minimize the duplication of follow-up observations by current and future transit surveys such as TESS

ReCombine: A Suite of Programs for Detection and Analysis of Meiotic Recombination in Whole-Genome Datasets

In meiosis, the exchange of DNA between chromosomes by homologous recombination is a critical step that ensures proper chromosome segregation and increases genetic diversity. Products of recombination include reciprocal exchanges, known as crossovers, and non-reciprocal gene conversions or non-crossovers. The mechanisms underlying meiotic recombination remain elusive, largely because of the difficulty of analyzing large numbers of recombination events by traditional genetic methods. These traditional methods are increasingly being superseded by high-throughput techniques capable of surveying meiotic recombination on a genome-wide basis. Next-generation sequencing or microarray hybridization is used to genotype thousands of polymorphic markers in the progeny of hybrid yeast strains. New computational tools are needed to perform this genotyping and to find and analyze recombination events. We have developed a suite of programs, ReCombine, for using short sequence reads from next-generation sequencing experiments to genotype yeast meiotic progeny. Upon genotyping, the program CrossOver, a component of ReCombine, then detects recombination products and classifies them into categories based on the features found at each location and their distribution among the various chromatids. CrossOver is also capable of analyzing segregation data from microarray experiments or other sources. This package of programs is designed to allow even researchers without computational expertise to use high-throughput, whole-genome methods to study the molecular mechanisms of meiotic recombination

Cross-Species Transmission of a Novel Adenovirus Associated with a Fulminant Pneumonia Outbreak in a New World Monkey Colony

Adenoviruses are DNA viruses that naturally infect many vertebrates, including humans and monkeys, and cause a wide range of clinical illnesses in humans. Infection from individual strains has conventionally been thought to be species-specific. Here we applied the Virochip, a pan-viral microarray, to identify a novel adenovirus (TMAdV, titi monkey adenovirus) as the cause of a deadly outbreak in a closed colony of New World monkeys (titi monkeys; Callicebus cupreus) at the California National Primate Research Center (CNPRC). Among 65 titi monkeys housed in a building, 23 (34%) developed upper respiratory symptoms that progressed to fulminant pneumonia and hepatitis, and 19 of 23 monkeys, or 83% of those infected, died or were humanely euthanized. Whole-genome sequencing of TMAdV revealed that this adenovirus is a new species and highly divergent, sharing <57% pairwise nucleotide identity with other adenoviruses. Cultivation of TMAdV was successful in a human A549 lung adenocarcinoma cell line, but not in primary or established monkey kidney cells. At the onset of the outbreak, the researcher in closest contact with the monkeys developed an acute respiratory illness, with symptoms persisting for 4 weeks, and had a convalescent serum sample seropositive for TMAdV. A clinically ill family member, despite having no contact with the CNPRC, also tested positive, and screening of a set of 81 random adult blood donors from the Western United States detected TMAdV-specific neutralizing antibodies in 2 individuals (2/81, or 2.5%). These findings raise the possibility of zoonotic infection by TMAdV and human-to-human transmission of the virus in the population. Given the unusually high case fatality rate from the outbreak (83%), it is unlikely that titi monkeys are the native host species for TMAdV, and the natural reservoir of the virus is still unknown. The discovery of TMAdV, a novel adenovirus with the capacity to infect both monkeys and humans, suggests that adenoviruses should be monitored closely as potential causes of cross-species outbreaks

KELT-21b: A Hot Jupiter Transiting The Rapidly Rotating Metal-Poor Late-A Primary Of A Likely Hierarchical Triple System

We present the discovery of KELT-21b, a hot Jupiter transiting the V = 10.5 A8V star HD 332124. The planet has an orbital period of P = 3.6127647 ± 0.0000033 days and a radius of 1.586 (+0.039)/(-0.040) RJ. We set an upper limit on the planetary mass of MP \u3c 3.91 MJ at 3σ confidence. We confirmed the planetary nature of the transiting companion using this mass limit and Doppler tomographic observations to verify that the companion transits HD 332124. These data also demonstrate that the planetary orbit is well-aligned with the stellar spin, with a sky-projected spin–orbit misalignment of λ = -5.6(+1.7o)/(-1.9). The star has Teff = 7598 (+81)/(-84) K, M* = 1.458 (+0.029)/(-0.028) M⊙, R* = 1.638 ± 0.034 R⊙, and v sin I* = 146 km s⁻¹, the highest projected rotation velocity of any star known to host a transiting hot Jupiter. The star also appears to be somewhat metal poor and α-enhanced, with [Fe/H] = -0.405 (+0.032)/(-0.033) and [α/Fe] = 0.145 ± 0.053; these abundances are unusual, but not extraordinary, for a young star with thin-disk kinematics like KELT-21. High-resolution imaging observations revealed the presence of a pair of stellar companions to KELT-21, located at a separation of 1farcs2 and with a combined contrast of ΔKS = 6.39 ± 0.06 with respect to the primary. Although these companions are most likely physically associated with KELT-21, we cannot confirm this with our current data. If associated, the candidate companions KELT-21 B and C would each have masses of ∼0.12 M⊙, a projected mutual separation of ∼20 au, and a projected separation of ∼500 au from KELT-21. KELT-21b may be one of only a handful of known transiting planets in hierarchical triple stellar systems

Transcriptome Analysis of the Model Protozoan, Tetrahymena thermophila, Using Deep RNA Sequencing

Background: The ciliated protozoan Tetrahymena thermophila is a well-studied single-celled eukaryote model organism for cellular and molecular biology. However, the lack of extensive T. thermophila cDNA libraries or a large expressed sequence tag (EST) database limited the quality of the original genome annotation. Methodology/Principal Findings: This RNA-seq study describes the first deep sequencing analysis of the T. thermophila transcriptome during the three major stages of the life cycle: growth, starvation and conjugation. Uniquely mapped reads covered more than 96 % of the 24,725 predicted gene models in the somatic genome. More than 1,000 new transcribed regions were identified. The great dynamic range of RNA-seq allowed detection of a nearly six order-of-magnitude range of measurable gene expression orchestrated by this cell. RNA-seq also allowed the first prediction of transcript untranslated regions (UTRs) and an updated (larger) size estimate of the T. thermophila transcriptome: 57 Mb, or about 55 % of the somatic genome. Our study identified nearly 1,500 alternative splicing (AS) events distributed over 5.2 % of T. thermophila genes. This percentage represents a two order-of-magnitude increase over previous EST-based estimates in Tetrahymena. Evidence of stage-specific regulation of alternative splicing was also obtained. Finally, our study allowed us to completely confirm about 26.8 % of the genes originally predicted by the gene finder, to correct coding sequence boundaries an

Correspondence analysis for quantification in electron energy loss spectroscopy and imaging.

Electron energy loss spectroscopy (EELS) is a technique to investigate the physical properties of material. Using this technique it is possible to detect the presence of a specific element in a specimen. When used in combination with an electron microscope, energy filtered images may be obtained, which in principle may be used to quantify the local element concentration. This involves a process of background correction, conventionally performed assuming a specific parametric behavior of the spectral intensity as a function of electron energy loss. In this article a parameter-free method is described for background correction based on the formalism of correspondence analysis. Such a method may be used in parts of the spectrum where the functional dependence of the spectral intensity is unknown. Use of this method for element detection has been suggested before. This article reports simulation experiments suggesting its suitability for quantitative determination of element distributions and element concentrations