FSH: fast spaced seed hashing exploiting adjacent hashes

Background

Patterns with wildcards in specified positions, namely spaced seeds, are increasingly used instead of k-mers in many bioinformatics applications that require indexing, querying and rapid similarity search, as they can provide better sensitivity. Many of these applications require to compute the hashing of each position in the input sequences with respect to the given spaced seed, or to multiple spaced seeds. While the hashing of k-mers can be rapidly computed by exploiting the large overlap between consecutive k-mers, spaced seeds hashing is usually computed from scratch for each position in the input sequence, thus resulting in slower processing.

Results

The method proposed in this paper, fast spaced-seed hashing (FSH), exploits the similarity of the hash values of spaced seeds computed at adjacent positions in the input sequence. In our experiments we compute the hash for each positions of metagenomics reads from several datasets, with respect to different spaced seeds. We also propose a generalized version of the algorithm for the simultaneous computation of multiple spaced seeds hashing. In the experiments, our algorithm can compute the hashing values of spaced seeds with a speedup, with respect to the traditional approach, between 1.6\(\times\) to 5.3\(\times\), depending on the structure of the spaced seed.

Conclusions

Spaced seed hashing is a routine task for several bioinformatics application. FSH allows to perform this task efficiently and raise the question of whether other hashing can be exploited to further improve the speed up. This has the potential of major impact in the field, making spaced seed applications not only accurate, but also faster and more efficient.

Availability

The software FSH is freely available for academic use at: https://bitbucket.org/samu661/fsh/overview.

     

Probing the structure-activity relationship of Escherichia coli LT-A by site-directed mutagenesis

Computer analysis of the crystallographic structure of the A subunit of Escherichia coil heat-labile toxin (LT) was used to predict residues involved in NAD binding, catalysis and toxicity. Following site-directed mutagenesis, the mutants obtained could be divided into three groups. The first group contained fully assembled, non-toxic new molecules containing mutations of single amino acids such as Val-53 → Glu or Asp, Ser-63 → Lys, Val-97 → Lys, Tyr-104 → Lys or Asp, and Ser-14 → Lys or Glu. This group also included mutations in amino acids such as Arg-7, Glu-110 and Glu-112 that were already known to be important for enzymatic activity. The second group was formed by mutations that caused the collapse or prevented the assembly of the A subunit: Leu-41 → Phe, Ala-45 → Tyr or Glu, Val-53 → Tyr, Val-60 → Gly, Ser-68 → Pro, His-70 → Pro, Val-97 → Tyr and Ser-114 → Tyr. The third group contained those molecules that maintained a wild-type level of toxicity in spite of the mutations introduced: Arg-54 → Lys or Ala, Tyr-59 → Met, Ser-68 → Lys, Ala-72 → Arg, His or Asp and Arg-192 → Asn. The results provide a further understanding of the structure–function of the active site and new, non-toxic mutants that may be useful for the development of vaccines against diarrhoeal diseases.     

Strontium stratigraphy of the upper Miocene Lithothamnion Limestone in the Majella Mountain,central Italy,and its palaeoenvironmental implications

The ⁸⁷Sr/⁸⁶Sr isotope ratio has been widely used as a physical tool to date and correlate carbonate successions due to the long Sr residence time in comparison with the ocean mixing time. If this method works on oceanic successions, marginal basins may show different Sr isotope records in comparison with the coeval ocean one due to sea‐level variations, continental run‐off and restricted water exchanges. In this work, we present the ⁸⁷Sr/⁸⁶Sr isotope record of the upper Miocene carbonate ramp of the Lithothamnion Limestone (Majella Mountain, central Apennines), as an example of the onset of restricted water exchanges between a marginal basin and the ocean water masses. The overall latemost Tortonian–early Messinian Sr isotope record of the Lithothamnion Limestone fits below the global reference line. This deviation has been interpreted as due to the strong control that freshwater input and enhanced continental run‐off, linked to the migration of the Apennine accretionary wedge and foredeep system, have had on the central Adriatic water chemistry. These results imply that an accurate oceanographic and geodynamic framework along with diagenetic overprint investigation has to be taken into consideration prior to apply SIS on carbonate successions on marginal basins, even when facies analyses indicate fully marine conditions. This seems to be the case for the upper Miocene Central Mediterranean carbonate successions, but may have more general validity and be extended to other recent or past marginal basins.     

Festuca valesiaca Schleich. ex Gaudin newly discovered in the Central Apennines (Italy): a further example of steppe relict in the Abruzzo “dry valleys”

Abstract

Festuca valesiaca Schleich. ex Gaudin subsp. valesiaca is acknowledged as one of the most characteristic grasses of the continental grasslands and steppes of Eurasia. In Italy, the species was known almost only from the continental valleys of the Alpine chain (with the exception of two small populations at the northernmost edge of the Apennines). Here, we report on the first confirmed finding of F. valesiaca subsp. valesiaca in the surroundings of the Fucino Basin, Central Apennines. This finding expands by c. 400?km southwards the species’ range in Italy. The identity of the species was first ascertained by means of leaf cross-sections and stomata measurements, and then confirmed by chromosome count. Its habitat preferences within the area were found to match well with those described in the literature for C-Europe. After discussing some identification issues and the population’s ecological niche, we present the main biogeographic implications of our finding. By recalling and discussing similar examples, we conclude that during the Holocene forest spread some Apennine “dry valleys” performed as refugia for a number of Pleistocene steppic taxa. The grasslands of these areas may thus be ancient and valuable ecosystems maintained by climate and disturbance throughout the Holocene.     

In vivo resistance of secondary antitumor immune response to cyclophosphamide: effects on T cell subsets

Summary We have analyzed the effects of high doses of cyclophosphamide (Cy) on primary and secondary antitumor immune response against immunogenic (tum^–) variants of Lewis lung carcinoma (3LL) treated in vitro with UV light. Normal mice and mice previously immunized with tum^– clones were inoculated i.p. with Cy (200 mg/kg body weight) and 24 h later challenged intrafootpad with tum^– or parental 3LL cells. Cy treatment suppressed the primary immune response of normal animals and allowed the growth of tum^– cells. In contrast, Cy-treated immune mice rejected the tumor challenge. The in vivo treatment with Cy decreased the total number of lymphoid cells in the spleens, as well as the proportion of B lymphocytes; however, it increased the percentage of both Lyt2⁺ and L3T4⁺ lymphocytes. Thus, the immunosuppressive effects of Cy on the primary antitumor response could not be attributed to elimination of major T lymphocyte subpopulations. Although the treatment of immune mice with Cy did not significantly impair their antitumor resistance, nor the proportion of Lyt2⁺ and L3T4⁺ lymphocytes in their spleens, the in vitro generation of cytotoxic T lymphocytes (CTL) was markedly reduced.After Cy treatment, the proliferative ability of spleen cells in response to interleukin-2 (IL-2) was substantially impaired. Using monoclonal antibodies to the IL-2 receptor, we found that Cy-treated T lymphocytes failed to fully express the IL-2 receptor following in vitro stimulation with irradiated tumor cells. In line with these findings, the in vitro generation of CTL was not restored by addition of recombinant IL-2 to the cultures. In vivo experiments using purified functional subsets of immune T cells showed that Lyt1⁺, but not Lyt2⁺ lymphocytes were able to transfer antitumor immunity in normal irradiated recipients.Therefore, since Ly1⁺ T lymphocytes were responsible for the antitumor resistance in vivo, the Cy-induced impairment of CTL generation did not affect the ability of immune mice to reject a secondary tumor challenge.This project has been funded at least in part with Federal funds from the Department of Health and Human Services, under contract number NO1-CO-23910 with Resources, Inc. The contents of this publication do not necessarily reflect the view or policies of the Department of Health and Human Services, nor does mention of trade names, commercial products, or organizations imply endorsement by the U.S. Government     

Isolation and structure elucidation of cytotoxic polyacetylenes and polyenes from Echinacea pallida

Bioassay-guided fractionation of n-hexane extracts of Echinacea pallida (Asteraceae) roots led to the isolation and structure elucidation of two polyacetylenes (1, 3) and three polyenes (2, 4, 5). Two are known hydroxylated compounds, namely 8-hydroxy-pentadeca-(9E)-ene-11,13-diyn-2-one (1) and 8-hydroxy-pentadeca-(9E,13Z)-dien-11-yn-2-one (2). Two dicarbonylic constituents, namely pentadeca-(9E)-ene-11,13-diyne-2,8-dione (3) and pentadeca-(9E,13Z)-dien-11-yne-2,8-dione (4), were isolated and characterized for the first time. Furthermore, the structure elucidation of pentadeca-(8Z,13Z)-dien-11-yn-2-one (5) is described. The structure of the compounds isolated was determined on the basis of UV, IR, NMR (including 1D and 2D NMR experiments, such as 1H-1H gCOSY, gHSQC-DEPT, gHMBC, gNOESY) and MS spectroscopic data. The cytotoxic activity of the isolated constituents against MIA PaCa-2 human pancreatic adenocarcinoma cells was evaluated in the concentration range 1-100 microg/ml. Results show that the hydroxylated compounds (1, 2) have low cytotoxicity, while the more hydrophobic polyacetylenes (3) and polyenes (4, 5) displayed moderate activity.     

The protein "mycoarray": a novel serological assay for the laboratory diagnosis of primitive endemic mycoses

A protein microarray containing fungal antigens (the "mycoarray") has been set up to provide rapid and appropriate serodiagnosis of primitive endemic mycoses, an important cause of morbidity and mortality in an increasingly high number of patients. The mycoarray consists of three antigen extracts (histoplasmin, coccidioidin and Coccidioides "TP") and antibody dilution curves were spotted on microarray slides. The arrays were processed with coccidioidomycosis and histoplasmosis patients? sera or with control sera and the occurring immunocomplexes were detected by indirect immunofluorescence. In agreement with clinical and microbiological diagnosis, the results distinguished between histoplasmosis and coccidioidomycosis patients. In addition, the assay could clearly discriminate between IgM and IgG antibody reactivity. No reactivity was ever observed in the arrays processed with negative control sera. Therefore, this pilot study demonstrates that the "mycoarray" is sensitive and specific enough to discriminate between healthy individuals and patients with histoplasmosis or coccidioidomycosis. Because of miniaturization and multiparametricity, the new assay cuts costs and processing time. Thus, once clinically validated and implemented as a large-scale array, the "mycoarray" will be ready to be applied to the daily clinical practice.