Biased estimates of clonal evolution and subclonal heterogeneity can arise from PCR duplicates in deep sequencing experiments

Accurate allele frequencies are important for measuring subclonal heterogeneity and clonal evolution. Deep-targeted sequencing data can contain PCR duplicates, inflating perceived read depth. Here we adapted the Illumina TruSeq Custom Amplicon kit to include single molecule tagging (SMT) and show that SMT-identified duplicates arise from PCR. We demonstrate that retention of PCR duplicate reads can imply clonal evolution when none exists, while their removal effectively controls the false positive rate. Additionally, PCR duplicates alter estimates of subclonal heterogeneity in tumor samples. Our method simplifies PCR duplicate identification and emphasizes their removal in studies of tumor heterogeneity and clonal evolution.

Electronic supplementary material

The online version of this article (doi:10.1186/s13059-014-0420-4) contains supplementary material, which is available to authorized users.     

Sustained rheumatoid arthritis remission is uncommon in clinical practice

Introduction

Remission is an important goal of therapy in rheumatoid arthritis (RA), but data on duration of remission are lacking. Our objective was to describe the duration of remission in RA, assessed by different criteria.

Methods

We evaluated patients from the Brigham and Women''s Rheumatoid Arthritis Sequential Study (BRASS) not in remission at baseline with at least 2 years of follow-up. Remission was assessed according to the Disease Activity Score 28-C-reactive protein (DAS28-CRP4), Simplified Disease Activity Index (SDAI), and Clinical Disease Activity Index (CDAI) scores, and the recently proposed American College of Rheumatology (ACR)/European League against Rheumatism (EULAR) criteria for remission. Analyses were performed by using Kaplan-Meier survival curves.

Results

We identified 871 subjects with ≥2 years of follow-up. Of these subjects, 394 were in remission at one or more time-points and not in remission at baseline, according to at least one of the following criteria: DAS28-CRP < 2.6 (n = 309), DAS28-CRP < 2.3 (n = 275), SDAI (n = 168), CDAI (n = 170), and 2010 ACR/EULAR (n = 158). The median age for the 394 subjects at entrance to BRASS was 56 years; median disease duration was 8 years; 81% were female patients; and 72% were seropositive. Survival analysis performed separately for each remission criterion demonstrated that < 50% of subjects remained in remission 1 year later. Median remission survival time was 1 year. Kaplan-Meier curves of the various remission criteria did not significantly differ (P = 0.29 according to the log-rank test).

Conclusions

This study shows that in clinical practice, a minority of RA patients are in sustained remission.     

Aniseed oil increases glucose absorption and reduces urine output in the rat

Anise (Pimpinella anisum) has been used as a traditional aromatic herb in many drinks and baked foods because of the presence of volatile oils in its fruits commonly known as seeds. Hot water extracts of the seeds have been used also in folk medicine for their diuretic and laxative effect, expectorant and anti-spasmodic action, and their ability to ease intestinal colic and flatulence. The aim of this work was to study the effect of aniseed oil on transport processes through intestinal and renal epithelia and determine its mechanism of action. The essential oils were extracted from the seeds by hydrodistillation and analyzed by gas chromatography. Aniseed oil enhanced significantly glucose absorption from the rat jejunum and increased the Na+-K+ ATPase activity in a jejunal homogenate in a dose dependent manner. The oil, however, exerted no effect on water absorption from the colon and did not alter the activity of the colonic Na+-K+ ATPase. When added to drinking water, it reduced the volume of urine produced in the rat and increased the activity of the renal Na+-K+ ATPase even at extremely low concentrations. It was concluded that aniseed oil increases glucose absorption by increasing the activity of the Na+-K+ ATPase and consequently the sodium gradient needed for the sugar transport. Its anti-diuretic effect is also mediated through a similar mechanism in the kidney whereby a stimulation of the Na+-K+ pump increases tubular sodium reabsorption and osmotic water movement. The colonic Na+-K+ ATPase was however, resistant to the oil.     

Identifying differentially methylated genes using mixed effect and generalized least square models

Background  

DNA methylation plays an important role in the process of tumorigenesis. Identifying differentially methylated genes or CpG islands (CGIs) associated with genes between two tumor subtypes is thus an important biological question. The methylation status of all CGIs in the whole genome can be assayed with differential methylation hybridization (DMH) microarrays. However, patient samples or cell lines are heterogeneous, so their methylation pattern may be very different. In addition, neighboring probes at each CGI are correlated. How these factors affect the analysis of DMH data is unknown.     

Genotypic comparison ofPantoea agglomeransplant and clinical strains

Background  

Pantoea agglomeransstrains are among the most promising biocontrol agents for a variety of bacterial and fungal plant diseases, particularly fire blight of apple and pear. However, commercial registration ofP. agglomeransbiocontrol products is hampered because this species is currently listed as a biosafety level 2 (BL2) organism due to clinical reports as an opportunistic human pathogen. This study compares plant-origin and clinical strains in a search for discriminating genotypic/phenotypic markers using multi-locus phylogenetic analysis and fluorescent amplified fragment length polymorphisms (fAFLP) fingerprinting.     

Historical biogeography and the evolution of the latitudinal gradient of species richness in the Papionini (Primata: Cercopithecidae)

We apply historical biogeography techniques to the macaques, baboons and their relatives (Primata: Papionini) and relate the inferred history of range shifts, and associated evolutionary events, to the latitudinal distribution of extant species, which is strongly tropical. The results of reversible parsimony, weighted ancestral area and dispersal-vicariance analyses all agree that Central Africa was part of the range of the ancestor of the tribe. Tropical regions with high current species richness (Central Africa, South-east Asia, Indonesia) have: (1) had a relatively long history of occupation, (2) experienced both a greater number and a greater average rate of speciation events and (3) given rise to more dispersal events to other regions. However, nested sister-taxon comparisons across the tribe show no overall association between differences in latitude and differences in rates of cladogenesis. Our historical reconstructions are largely consistent with previous hypotheses and fossil data, and suggest that both the passage of time since colonization and rates of cladogenesis have enhanced tropical species richness. Historical biogeography may thus considerably aid understanding of this and other spatial problems in macroecology.  © 2005 The Linnean Society of London, Biological Journal of the Linnean Society , 2005, 85 , 235–246.     

A histopathological and stereological study of liver damage in female rats caused by mercury vapor

We examined the possible effects of elemental mercury vapor on the liver of the female rats. We divided the animals into an untreated control group and an experimental group that was exposed to mercury vapor for 45 days. Liver samples were obtained for histological and stereological analysis. The total liver, parenchyma and sinusoid volumes were increased significantly in the mercury vapor treated group compared to controls. Also, the mean density, total number and mean nuclear diameter of hepatocytes, except for binucleated hepatocytes, was decreased in the experimental group compared to controls. Light and electron microscopy revealed alterations of liver structure of the experimental animals compared to controls.     

Sorption of penicillium-cellulase on cellulose and lignin

Sorption of Penicillium cellulase onto cotton linters samples differing in physical structure and onto KLASON - lignin from spruce has been investigated by determining cellobiase, CMCL- and ‘linters-activity’ as well as protein content of a cellulase culture filtrate before and after contact with the sorbent at different temperature for different time intervals. CMCL- and cellobiase-activity were found to be reduced much less by the sorption process onto the cellulose samples than ‘linters-activity’ and protein content. Sorption increased in the order: untreated linters < mercerized linters < wet fibrillated (‘colloidal milled’) linters. With the KLASON -lignin also a considerable sorption of the cellulase system was observed. Results are discussed with respect to preferential sorption of enzyme components.