Unbiased Collections Research Articles

Reusing a prepaid health plan’s fecal immunochemical tests for microbiome associations with colorectal adenoma

An altered colonic microbiota probably increases colorectal adenoma (CRA) and cancer (CRC) risk, but large, unbiased fecal collections are needed to examine the relationship of gut microbiota diversity and composition to colorectal carcinogenesis. This study assessed whether fecal immunochemical tests (FITs) from CRA/CRC screening may fulfill this requirement. Using FIT, self-collected by members of Kaiser Permanente Hawaii (KPH), as well as interspersed quality control (QC) specimens, DNA was extracted and amplified to generate 16S rRNA microbiome profiles rarified at 10,000 reads. CRA/CRC were diagnosed by colonoscopy and histopathology. Covariates were from electronic KPH records. Of 921 participants’ FIT devices, 538 (58%) yielded at least 10,000 rRNA reads and 1016 species-level variants mapped to 46 genera. Of the 538 evaluable participants, 63 (11.7%) were FIT-negative per protocol, and they were considered negative for CRA/CRC. Of the 475 FIT + participants, colonoscopy and pathologic review revealed that 8 (1.7%) had CRC, 71 (14.9%) had high-risk CRA, 107 (22.5%) had low-risk CRA, and 289 (60.8%) did not have CRA/CRC. Men were 2.27-fold [95% confidence interval (CI) 1.32–3.91] more likely than women to be FIT+ . Men also had 1.96-fold (CI 1.24–3.07) higher odds of low-risk CRA, with similar trends for high-risk CRA and CRC. CRA/CRC were not associated with overweight, obesity, diabetes, or antibiotic prescriptions in this study. QC analysis across 24 batches of FIT devices revealed QC outliers in four batches. With or without exclusion of the four QC-outlier batches, as well as lenient (1000-read) rarefaction, CRA/CRC had no consistent, statistically significant associations with fecal microbiome alpha diversity, beta diversity or genera relative abundance. CRA/CRC had expected associations with male sex but not with microbiome metrics. Fecal microbiome profiling using DNA extracted from at-home collected, re-used FIT devices is feasible, albeit with substantial challenges. Using FITs for prospective microbiome studies of CRA/CRC risk should consider the impact of the current findings on statistical power and requisite sample sizes.

Reply to the comment by M. Genge and M. Van Ginneken on paper entitled “Unmelted cosmic metal particles from the Indian Ocean”

Reply to the comment by M. Genge and M. Van Ginneken on paper entitled “Unmelted cosmic metal particles from the Indian Ocean”

Comparison of Biting Midges of the Early Eocene Cambay Amber (India) and Late Eocene European Ambers Supports the Independent Origin of European Ambers

Abstract Of the three genera of Ceratopogoninae dominant in Late Eocene ambers of Europe, the two known are absent in the Early Eocene Cambay amber (India): the Holarctic genus Ceratopogon and the worldwide distributed genus Culicoides, known since the Late Cretaceous. Tropical biting midge genera (Meunierohelea and Leptoconops and genera with feminized male antennae: Camptopterohelea, Eohelea, and Gedanohelea) are abundantly represented in Cambay amber. The proportion of Ceratopogonini with feminized male antennae among all Ceratopogonidae dramatically increases from north to south: from 1.4-2.4 % in unbiased collections of Bitterfeld amber (Humboldt Museum collection) and Baltic amber (the Giecewicz collection) to 7 % in Danish amber and 12.7 % in Rovno amber; their proportion in Cambay amber is 17 %. The proportion of tropical specimens among specimens Ceratopogoninae in unbiased collections is 6.4 % in Baltic amber, 5.3 % in Bitterfeld amber, 21 % in Rovno and Danish amber, and 58 % in Cambay amber. Strong differences in the proportion of tropical components among Ceratopogoninae from different European ambers are in agreement with data on Cambay amber and so are indicative of origin of the European ambers under different climatic and hence geographic environments.

Micrometeorite flux on Earth during the last ~50,000 years

AbstractFlux of micrometeorites is estimated by using cosmic spherule counts from a seafloor area of 2.50 m2 from the Indian Ocean. The spherules are recovered from sediment samples in close‐spaced locations from the Indian Ocean after sieving 293 kg of sediment. The terrestrial age of the spherules has a range of 0–~50,000 years. The spherules have a size range of 57–750 µm (average size 265 ± 92 µm). The diameter of the spherules increases from scoriaceous‐barred‐cryptocrystalline‐glassy types. The time‐averaged flux of the spherules is 160 t/yr, a sizeable mass (>60%) resides in the >300 µm fraction; the slope of distribution is similar to that of Deep‐Sea Spherules but significantly different from other collections which have lower average diameters. It is observed here, a significant population of cosmic dust resides in the larger sizes which can be recovered by sampling large areas in time and space. The spherule textures are similar to that of unbiased collections from the polar regions, indicating that the textural types of cosmic dust that have been raining on the Earth during the last 50 kyr have been constant regardless of size. Major element chemistry of a majority of the spherules show elemental ratios that are close to a CM or CI chondritic parent body; a single spherule (0.2% of the population) suggests an achondritic parent body. Unbiased collections spanning large areas temporally and spatially enlarge the inventory of the Earth‐crossing meteoroid complex and provide valuable inputs for models on cosmic dust accretion.

Fractionation and fragmentation of glass cosmic spherules during atmospheric entry

Glass cosmic spherules are a sub-type of S-type cosmic spherules that are almost at the end phase of fractionation and volatilization during atmospheric entry. A focused investigation of 36 glass spherules out of several hundred spherules reveals fractionation and fragmentation phenomena. During atmospheric entry, some of the glass spherules largely preserve relict pyroxene normative crystals within. The spherules appear to have experienced temperatures of ∼1700°C that resulted in vaporization of Fe and other volatile elements such as S, Na, and K from the silicate spherule, resulting in the enrichment of SiO2, MgO, and CaO in weight percent. Calculations with a chemical ablation model suggest an entry velocity of ∼11–16km/s to achieve the temperature required for glass spherule formation under these conditions the micrometeoroid undergoes a mass of loss of ∼20%. Any increase in entry velocity will increase the mass loss drastically and change its chemical composition. Most glass spherules beyond a certain threshold diameter of ∼300μm appear to undergo fragmentation resulting in two different spherules, which could lead to an apparent enhancement of the percentage of glass spherules in unbiased collections. Siderophile and volatile element loss and subsequent development of a platinum group element (PGE) nugget are observed in one of the spherules. This appears to be the first reported observation of a platinum group nugget (PGN) in a glass spherule. The nugget has a diameter of ∼3μm and contains all the six PGE, although some of the more volatile PGE are depleted relative to an average chondritic composition. Interestingly, the PGE data of the nugget seem to have a trend which is similar to that of a PGE nugget from a calcium aluminium rich inclusion (CAI). The trace element data of these glass spherules suggest that their parent bodies were carbonaceous chondrites.

In vivo RNAi screening to genetically dissect malignant progression from hepatocytes harboring constitutive active AKT–1

The neoplastic evolution of hepatocellular carcinoma (HCC) proceeds through a multi-step process which is still poorly characterized on the molecular level. The discovery of RNA interference (RNAi) has revolutionized loss of function genetics and RNAi based functional genomic screens allow for the identification of unbiased and comprehensive collections of new gene targets in virtually every biological system. Through the combination of intraheptic gene transfer via transposable elements with miRNA based shRNA technology, we developed a system allowing to conduct in vivo RNAi screens for new tumor suppressor genes whose loss cooperate with defined oncogenic lesions in HCC. For example, we took the advantage of a focused shRNA library targeting genes found in focal genomic deletions of 98 human HCCs (Zender et al., Cell, 2008) to screen for tumor suppressor genes whose loss allow for malignant progression of hepatocytes expressing a constitutive active form of AKT–1. The focused shRNA library was divided into several low complexity pools (n=250) and subjected to an in vivo positive selection screen in combination with AKT–1 in p19Arf deficient mouse livers. Arising tumors were analyzed by deep sequencing to identify shRNAs which had allowed for malignant progression of AKT–1 expressing hepatocytes. Identified candidate genes were subjected to thorough functional validation using multiple independent shRNAs against each target gene.

Early Eocene plant diversity and dynamics in the Falkland flora, Okanagan Highlands, British Columbia, Canada

The early Eocene fossil localities of the Okanagan Highlands in British Columbia, Canada, and Washington State, USA, span the Early Eocene Climatic Optimum, the warmest period of the Cenozoic, and reflect mild but equable upland climates (mean annual temperature 0°C). The Okanagan Highlands region has been identified as a centre of temperate plant family diversification in the northern hemisphere during the early Eocene. Here, we test the hypothesis of mid-latitude high diversity through rarefaction analysis of unbiased census collections from the Okanagan Highlands Falkland fossil locality, demonstrating levels of diversity similar to those documented at hyperdiverse Eocene sites in South America when adjusted for sample size. An explanation for this diversity may lie in the upland character of the Falkland site, as altitudinal gradients provide a mosaic of microhabitats through interacting effects of topography and climate. Fine-scale trends are also examined within the Falkland site, demonstrating a shift in plant community composition over time to a more diverse flora, although the dominant taxa persist through the section in varying levels of abundance. Intra-site patterns in plant community structure and composition are attributed to a combination of environmental factors, including disturbance and microhabitat diversity.

Genome-Wide siRNA-Based Functional Genomics of Pigmentation Identifies Novel Genes and Pathways That Impact Melanogenesis in Human Cells

Melanin protects the skin and eyes from the harmful effects of UV irradiation, protects neural cells from toxic insults, and is required for sound conduction in the inner ear. Aberrant regulation of melanogenesis underlies skin disorders (melasma and vitiligo), neurologic disorders (Parkinson's disease), auditory disorders (Waardenburg's syndrome), and opthalmologic disorders (age related macular degeneration). Much of the core synthetic machinery driving melanin production has been identified; however, the spectrum of gene products participating in melanogenesis in different physiological niches is poorly understood. Functional genomics based on RNA-mediated interference (RNAi) provides the opportunity to derive unbiased comprehensive collections of pharmaceutically tractable single gene targets supporting melanin production. In this study, we have combined a high-throughput, cell-based, one-well/one-gene screening platform with a genome-wide arrayed synthetic library of chemically synthesized, small interfering RNAs to identify novel biological pathways that govern melanin biogenesis in human melanocytes. Ninety-two novel genes that support pigment production were identified with a low false discovery rate. Secondary validation and preliminary mechanistic studies identified a large panel of targets that converge on tyrosinase expression and stability. Small molecule inhibition of a family of gene products in this class was sufficient to impair chronic tyrosinase expression in pigmented melanoma cells and UV-induced tyrosinase expression in primary melanocytes. Isolation of molecular machinery known to support autophagosome biosynthesis from this screen, together with in vitro and in vivo validation, exposed a close functional relationship between melanogenesis and autophagy. In summary, these studies illustrate the power of RNAi-based functional genomics to identify novel genes, pathways, and pharmacologic agents that impact a biological phenotype and operate outside of preconceived mechanistic relationships.

Synthetic lethal screen identification of chemosensitizer loci in cancer cells

Abundant evidence suggests that a unifying principle governing the molecular pathology of cancer is the co-dependent aberrant regulation of core machinery driving proliferation and suppressing apoptosis. Anomalous proteins engaged in support of this tumorigenic regulatory environment most probably represent optimal intervention targets in a heterogeneous population of cancer cells. The advent of RNA-mediated interference (RNAi)-based functional genomics provides the opportunity to derive unbiased comprehensive collections of validated gene targets supporting critical biological systems outside the framework of preconceived notions of mechanistic relationships. We have combined a high-throughput cell-based one-well/one-gene screening platform with a genome-wide synthetic library of chemically synthesized small interfering RNAs for systematic interrogation of the molecular underpinnings of cancer cell chemoresponsiveness. NCI-H1155, a human non-small-cell lung cancer line, was employed in a paclitaxel-dependent synthetic lethal screen designed to identify gene targets that specifically reduce cell viability in the presence of otherwise sublethal concentrations of paclitaxel. Using a stringent objective statistical algorithm to reduce false discovery rates below 5%, we isolated a panel of 87 genes that represent major focal points of the autonomous response of cancer cells to the abrogation of microtubule dynamics. Here we show that several of these targets sensitize lung cancer cells to paclitaxel concentrations 1,000-fold lower than otherwise required for a significant response, and we identify mechanistic relationships between cancer-associated aberrant gene expression programmes and the basic cellular machinery required for robust mitotic progression.

A Paleocene lowland macroflora from Patagonia reveals significantly greater richness than North American analogs

Few South American macrofloras of Paleocene age are known, and this limits our knowledge of diversity and composition between the end-Cretaceous event and the Eocene appearance of high floral diversity. We report new, unbiased collections of 2516 compression specimens from the Paleocene Salamanca Formation (ca. 61.7 Ma) from two localities in the Palacio de los Loros exposures in southern Chubut, Patagonia, Argentina. Our samples reveal considerably greater richness than was previously known from the Paleocene of Patagonia, including 36 species of angiosperm leaves as well as angiosperm fruits, flowers, and seeds; ferns; and conifer leaves, cones, and seeds. The floras, which are from siltstone and sandstone channel-fills deposited on low-relief floodplain landscapes in a humid, warm temperate climate, are climatically and paleoenvironmentally comparable to many quantitatively collected Paleocene floras from the Western Interior of North America. Adjusted for sample size, there are >50% more species at each Palacio de los Loros quarry than in any comparable U.S. Paleocene sample. These results indicate more vibrant terrestrial ecosystems in Patagonian than in North American floodplain environments ∼4 m.y. after the end-Cretaceous extinction, and they push back the time line 10 m.y. for the evolution of high floral diversity in South America. The cause of the dis parity is unknown but could involve reduced impact effects because of greater distance from the Chicxulub site, higher latest Cretaceous diversity, or faster recovery or immigration rates.

Eocene Plant Diversity at Laguna del Hunco and Río Pichileufú, Patagonia, Argentina

The origins of South America's exceptional plant diversity are poorly known from the fossil record. We report on unbiased quantitative collections of fossil floras from Laguna del Hunco (LH) and Río Pichileufú (RP) in Patagonia, Argentina. These sites represent a frost-free humid biome in South American middle latitudes of the globally warm Eocene. At LH, from 4,303 identified specimens, we recognize 186 species of plant organs and 152 species of leaves. Adjusted for sample size, the LH flora is more diverse than comparable Eocene floras known from other continents. The RP flora shares several taxa with LH and appears to be as rich, although sampling is preliminary. The two floras were previously considered coeval. However, (40)Ar/(39)Ar dating of three ash-fall tuff beds in close stratigraphic association with the RP flora indicates an age of 47.46+/-0.05 Ma, 4.5 million years younger than LH, for which one tuff is reanalyzed here as 51.91+/-0.22 Ma. Thus, diverse floral associations in Patagonia evolved by the Eocene, possibly in response to global warming, and were persistent and areally extensive. This suggests extraordinary richness at low latitudes via the latitudinal diversity gradient, corroborated by published palynological data from the Eocene of Colombia.

Screening and synthesis: high throughput technologies applied to parasitology

High throughput technologies continue to develop in response to the challenges set by the genome projects. This article discusses how the techniques of both high throughput screening (HTS) and synthesis can influence research in parasitology. Examples of the use of targeted and phenotype-based HTS using unbiased compound collections are provided. The important issue of identifying the protein target(s) of bioactive compounds is discussed from the synthetic chemist's perspective. This article concludes by reviewing recent examples of successful target identification studies in parasitology.

The Text REtrieval Conferences (TRECs): Providing a Test‐Bed for Information Retrieval Systems

Les congres de la TREC (Text REtrieval Conferences) ont pour sujet d'etude les methodes de recherche d'information ainsi que l'evaluation de ces methodes. La TREC fournit des collection-test