Internal eliminated sequences interrupting the Oxytricha 81 locus: allelic divergence, conservation, conversions, and possible transposon origins

Internal eliminated sequences (IESs) often interrupt ciliate genes in the silent germline nucleus but are exactly excised and eliminated from the developing somatic nucleus from which genes are then expressed. Some long IESs are transposons, supporting the hypothesis that short IESs are ancient transposon relics. In light of that hypothesis and to explore the evolutionary history of a collection of IESs, we have compared various alleles of a particular locus (the 81 locus) of the ciliated protozoa Oxytricha trifallax and O. fallax. Three short IESs that interrupt two genes of the locus are found in alleles from both species, and thus must be relatively ancient, consistent with the hypothesis that short IESs are transposon relics. In contrast, TBE1 transposon interruptions of the locus are allele-specific and probably the results of recent transpositions. These IESs (and the TBE1s) are precisely excised from the DNA of the developing somatic macronucleus. Each IES interrupts a highly conserved sequence. A few nucleotides at the ends of each IES are also conserved, suggesting that they interact critically with IES excision machinery. However, most IES nucleotide positions have evolved at high rates, showing little or no selective constraint for function. Nonetheless, the length of each IES has been maintained (+/- 3 bp). While one IES is approximately 33 bp long, three other IESs have very similar sizes, approximately 70 bp long. Two IESs are surrounded by direct repeats of the sequence TTCTT. No other sequence similarities were found between any of the four IESs. However, the ends of one IES do match the inverted terminal repeat consensus sequence of the "TA" IESs of Paramecium. Three O. trifallax alleles appear to have been recipients in recent conversion events that could have been provoked by double-strand breaks associated with IES ends subsequent to IES transposition. Our findings support the hypothesis that short IESs evolved from ancient transposons that have lost most of their sequences, except those necessary for precise excision during macronuclear development.      

Optimization of conscious sedation in plastic surgery.

The administration of conscious sedation by the plastic surgeon must be safe, efficient, and consistent. In the proper setting, with trained staff and appropriate backup, conscious sedation can allow optimal patient satisfaction with expedient recovery in addition to cost containment. The highly effective local anesthesia afforded by dilute, high-volume ("tumescent") infiltration extends the use of conscious sedation to cases previously performed under general anesthesia or deep sedation. The purpose of this analysis was to identify variables in conscious sedation that affect traditional outcome parameters in ambulatory surgery, particularly the duration of recovery and adverse events such as nausea and emesis. All perioperative and operative records of 300 consecutive patients having plastic surgical procedures under conscious sedation were carefully reviewed. Patients were ASA class I or II by requisite. Conscious sedation followed a standardized administration protocol, using incremental doses of two agents: midazolam (0.25 to 1 mg) and fentanyl (12.5 to 50 mcg). A subset of patients received preoperative oral sedation. Multivariate statistical analysis was conducted using SPSS 8.0 for Windows (SPSS Inc., Chicago, Ill.). Of the 300 patients, same-day discharge was intended for 281. Eight procedure categories were defined. No anesthetic complications occurred. As expected, recovery time was significantly correlated with the duration and type of procedure (p < 0.001) and the total dosage of both intraoperative sedative agents (p < 0.001). Interestingly, a negative correlation with advancing age existed (p < 0.001), likely reflecting the significantly higher intraoperative sedative dosing in younger patients (p < 0.001). When controlled for the effects of procedure duration and intraoperative sedative dosing, two other variables-use of preoperative oral sedation and postoperative nausea/emesis-significantly lengthened recovery time (p = 0.0001 for each). Fifteen unintended admissions occurred secondary to nausea, prolonged drowsiness, or pain control needs. Conscious sedation is an effective anesthetic choice for routine plastic surgical procedures, many of which would commonly be performed under general anesthesia. In our experience with a carefully structured and controlled conscious sedation protocol, the technique has proven to be safe and effective. This analysis of outcome parameters identified two important and potentially avoidable causes of recovery delay following conscious sedation-oral premedication and nausea/emesis. Nausea and emesis were particularly problematic in that they were responsible for 11 of 15 (73 percent) unintended admissions. Preoperative sedation is valuable in certain circumstances, and its use is not discouraged; however, its benefits must be weighed against its unwanted effects, which can include a prolongation of recovery.     

Efficient linkage mapping using exome capture and extreme QTL in schistosome parasites

Background

Identification of parasite genes that underlie traits such as drug resistance and host specificity is challenging using classical linkage mapping approaches. Extreme QTL (X-QTL) methods, originally developed by rodent malaria and yeast researchers, promise to increase the power and simplify logistics of linkage mapping in experimental crosses of schistosomes (or other helminth parasites), because many 1000s of progeny can be analysed, phenotyping is not required, and progeny pools rather than individuals are genotyped. We explored the utility of this method for mapping a drug resistance gene in the human parasitic fluke Schistosoma mansoni.

Results

We staged a genetic cross between oxamniquine sensitive and resistant parasites, then between two F1 progeny, to generate multiple F2 progeny. One group of F2s infecting hamsters was treated with oxamniquine, while a second group was left untreated. We used exome capture to reduce the size of the genome (from 363 Mb to 15 Mb) and exomes from pooled F2 progeny (treated males, untreated males, treated females, untreated females) and the two parent parasites were sequenced to high read depth (mean = 95-366×) and allele frequencies at 14,489 variants compared. We observed dramatic enrichment of alleles from the resistant parent in a small region of chromosome 6 in drug-treated male and female pools (combined analysis: = 11.07, p = 8.74 × 10^-29). This region contains Smp_089320 a gene encoding a sulfotransferase recently implicated in oxamniquine resistance using classical linkage mapping methods.

Conclusions

These results (a) demonstrate the utility of exome capture for generating reduced representation libraries in Schistosoma mansoni, and (b) provide proof-of-principle that X-QTL methods can be successfully applied to an important human helminth. The combination of these methods will simplify linkage analysis of biomedically or biologically important traits in this parasite.

Electronic supplementary material

The online version of this article (doi:10.1186/1471-2164-15-617) contains supplementary material, which is available to authorized users.     

Profiled support vector machines for antisense oligonucleotide efficacy prediction

Background  

This paper presents the use of Support Vector Machines (SVMs) for prediction and analysis of antisense oligonucleotide (AO) efficacy. The collected database comprises 315 AO molecules including 68 features each, inducing a problem well-suited to SVMs. The task of feature selection is crucial given the presence of noisy or redundant features, and the well-known problem of the curse of dimensionality. We propose a two-stage strategy to develop an optimal model: (1) feature selection using correlation analysis, mutual information, and SVM-based recursive feature elimination (SVM-RFE), and (2) AO prediction using standard and profiled SVM formulations. A profiled SVM gives different weights to different parts of the training data to focus the training on the most important regions.