Hypothesis-driven weight-of-evidence analysis of endocrine disruption potential: a case study with triclosan

Triclosan is an antimicrobial agent used in a range of consumer products, such as deodorants, oral care, clothing, and household items. As with many consumer products, triclosan can be rinsed down the drain and transported to wastewater treatment plants. While most is eliminated during activated sludge sewage treatment by biodegradation and adsorption, some triclosan enters the aquatic environment and may expose wildlife. Given the potential for exposure to both humans and wildlife, resolving whether triclosan is endocrine active is important due to growing concerns about potential adverse public health and environmental effects of endocrine-disrupting substances. A weight of evidence (WoE) analysis focusing on specific hypotheses related to interaction with estrogen, androgen, and thyroid hormone pathways, and steroidogenesis was applied to triclosan. This WoE procedure involved systematic consideration of each endpoint, focused on screening level studies in the US Endocrine Disruptor Screening Program, as well as those in levels 1 through 5 of the OECD Conceptual Framework. This was followed by a semiquantitative relevance weighting of each endpoint to a given hypothesis to reach scientifically justified conclusions. Use of all relevant and reliable information and consistent observations in multiple studies strengthen support for or against each mode of action hypothesis. Using data from multiple animal species and in vitro systems, this systematic and transparent WoE assessment indicated that triclosan is not acting as an agonist or antagonist within the estrogen, androgen, thyroid, or steroidogenic pathways and is not impacting endocrine pathways as a lead or primary mode of toxicity.     

Managing operating room budget variances

Credibility, power, and the ability to obtain greater departmental resources are three benefits of managing the operating room (OR) department's operating budget effectively. Still, few resources exist to help novice as well as seasoned OR directors grapple with the practicalities of maintaining their budget after the annual budget process is completed. The authors examine how astutely controlling personnel, materials, and services budget variances will result in hospital administrators "hearing" and approving an OR director's requests for resources more readily, staff and physicians who enjoy the benefits of better-staffed services and new technology, and an OR director with a reputation as an effective department head among peers in the hospital.     

Limitations of TaqMan PCR for detecting divergent viral pathogens illustrated by hepatitis A,B, C,and E viruses and human immunodeficiency virus

Recent events illustrate the imperative to rapidly and accurately detect and identify pathogens during disease outbreaks, whether they are natural or engineered. Particularly for our primary goal of detecting bioterrorist releases, detection techniques must be both species-wide (capable of detecting all known strains of a given species) and species specific. Due to classification restrictions on the publication of data for species that may pose a bioterror threat, we illustrate the challenges of finding such assays using five nonthreat organisms that are nevertheless of public health concern: human immunodeficiency virus (HIV) and four species of hepatitis viruses. Fluorogenic probe-based PCR assays (TaqMan; Perkin-Elmer Corp., Applied Biosystems, Foster City, Calif.) may be sensitive, fast methods for the identification of species in which the genome is conserved among strains, such as hepatitis A virus. For species such as HIV, however, the strains are highly divergent. We use computational methods to show that nine TaqMan primer and probe sequences, or signatures, are needed to ensure that all strains will be detected, but this is an unfeasible number, considering the cost of TaqMan probes. Strains of hepatitis B, C, and E viruses show intermediate divergence, so that two to three TaqMan signatures are required to detect all strains of each virus. We conclude that for species such as hepatitis A virus with high levels of sequence conservation among strains, signatures can be found computationally for detection by the TaqMan assay, which is a sensitive, rapid, and cost-effective method. However, for species such as HIV with substantial genetic divergence among strains, the TaqMan assay becomes unfeasible and alternative detection methods may be required. We compare the TaqMan assay with some of the alternative nucleic acid-based detection techniques of microarray, chip, and bead technologies in terms of sensitivity, speed, and cost.     

Comparison between double-contrast barium enema and colonoscopy to investigate lower gastrointestinal bleeding

A retrospective study was performed to compare the diagnostic accuracy of high-quality double-contrast barium enema (DCBE) against gold standard colonoscopy in 288 patients with suspected lower gastrointestinal bleeding who went through both examinations. Colonoscopy detected the potential cause of bleeding in 99 patients (100%); in order of frequency: polyps1 cm (N=47; 48%), carcinoma (N=21; 21%), inflammatory bowel disease (IBD) (N=15; 15%), solitary ulcers (N=6; 6%), other types of colitis (N=5; 5%), angiodysplasia (N=3; 3%), and stenosis (N=2; 2%). DCBE diagnosed 88 cases (89%) and missed 11 consisting of IBD (N=4), angiodysplasia (N=3), solitary ulcers (N=3), and polyps (N=1). The overall sensitivity and specificity of DCBE was 0.89 and 0.97, respectively. The sensitivity for carcinoma, polyps, and IBD was 1.00, 0.98, and 0.73, respectively. We conclude that DCBE is very effective to diagnose carcinoma and polyps1 cm, the most frequent causes of bleeding, but less effective to diagnose IBD and other nonfrequent causes. If a high-quality DCBE does not reveal the cause of bleeding, the contribution of a following colonoscopy will be to diagnose causes of bleeding other than carcinoma and polyps < 1 cm and to offer therapeutic possibilities.     

Extranodal extension in lymph node-positive prostate cancer.

Evaluation of extranodal tumor extension may provide prognostic information for patients with epithelial malignancies. However, its importance for the patient who has prostate cancer with regional lymph node metastasis requires further investigation and clarification. This study was performed to evaluate the prognostic significance of extranodal extension (ENE) in a large series of node-positive patients. The study group included 212 node-positive patients who were treated by bilateral pelvic lymphadenectomy, radical retropubic prostatectomy, and androgen deprivation between 1987 and 1992 at the Mayo Clinic. ENE was defined as cancer perforating through the lymph node capsule into perinodal tissue. Nodal cancer volume was measured by the grid method. Univariate and multivariate risk ratios (RR) for distant metastasis-free and cancer-specific survival were estimated using the Cox proportional model. The mean follow-up was 6.3 years (median, 6.1 years). Distant metastasis-free and cancer-specific survival at 5 years for all patients was 91% and 95%, respectively. ENE was found in 126 of 212 patients (59%). The presence of ENE was not significantly associated with distant metastasis-free (RR = 1.6; 95% confidence interval [CI], 0.7 to 3.9) or cancer-specific survival (RR = 2.2; 95% CI, 0.7 to 6.8). Among 98 patients with a single positive node, there was no significant difference in distant metastasis or cancer-specific survival according to the presence of ENE (P = .88 and P = .36, respectively). After adjusting for Gleason score, DNA ploidy, and ENE, only nodal cancer volume was significantly associated with adverse distant metastasis-free (RR = 1.9; 95% CI, 1.5 to 2.8) and cancer-specific survival (RR = 1.4; 95% CI, 1.1 to 1.9). Our data indicate that the presence of ENE is not associated with unfavorable survival in patients with node-positive prostate cancer treated by radical retropubic prostatectomy, bilateral pelvic lymphadenectomy, and androgen deprivation therapy. In contrast, nodal cancer volume was predictive of distant metastasis-free survival and cancer-specific survival.     

Association of germ-free mice with a simplified human intestinal microbiota results in a shortened intestine

Genetic, nutritional, and gut microbiota-derived factors have been proposed to play a role in the development of the whole intestine that is around 40% longer in PRM/Alf mice compared with other mouse strains. The PRM/Alf genotype explains 60% of this length difference. The remaining 40% are due to a maternal effect that could depend on the gut microbiota transmitted by the mother to their pups. Germ-free PRM/Alf mice and C3H/He mice were associated with a simplified human microbiota (SIHUMI) to study its impact on gut length. The small intestines of the SIHUMI-associated mice were 16.4% (PRM/Alf) and 9.7% (C3H/He) shorter than those of the corresponding germ-free counterparts. Temporal temperature gradient gel electrophoresis and quantitative real-time PCR revealed differences in microbiota composition between both SIHUMI-associated mouse strains. Anaerostipes caccae was one log lower in PRM/Alf mice than in C3H/He mice. Since polyamines and short-chain fatty acids (SCFAs) are important intestinal growth factors, their concentrations were explored. Cecal concentrations of putrescine, spermine, spermidine, and N-acetylspermine were 1.5-fold, 3.7-fold, 2.2-fold, and 1.4-fold higher, respectively, in the SIHUMI-C3H/He mice compared with the SIHUMI-PRM/Alf mice. In addition, cecal acetate, propionate, and butyrate concentrations in SIHUMI-C3H/He mice were 1.4-fold, 1.1-fold, and 2.1-fold higher, respectively, than in SIHUMI-PRM/Alf mice. These results indicate that polyamines and SCFAs did not promote gut lengthening in any of the two mouse strains. This suggests that as yet unknown factors provided by the SIHUMI prevented gut lengthening in the SIHUMI-associated mice compared with the germfree mice.     

Trends in the research profile of matched independent plastic surgery fellows

The independent plastic surgery pathway recruits candidates with 5 years of surgical training who are typically more advanced in research than their integrated counterparts. Research productivity helps to discriminate between applicants. However, no studies exist detailing the academic attributes of matched independent plastic surgery candidates.We performed a cohort study of 161 independent plastic surgery fellows from accredited residency programs from the 2015 to 2017 application cycles. We performed a bibliometric analysis utilizing Scopus, PubMed, and Google Scholar to identify research output measures at the time of application.The cohort was predominantly men (66%) with a median of 3 articles and a H-index of 1 at the time of application. Interestingly, 16% of successful candidates had no published articles at the time of application, and this did not change significantly over time (P = .0740). Although the H-index remained stable (R 0.13, P = .1095), the number of published journal articles per candidate significantly decreased over 3 consecutive application cycles (R −0.16, P = .0484). Analysis of article types demonstrated a significant increase in basic science articles (R 0.18, P = .0366) and a concurrent decrease in editorial-type publications (R = −0.18, P = .0374).Despite the decline in publication volume of matched independent plastic surgery fellows, the quality of their research portfolio has remained constant. Matched applicants appear to be shifting focus from faster-to-publish articles to longer but higher impact projects. In selecting a training route, applicants must weigh the highly competitive integrated path against the dwindling number of independent positions.     

Using populations of human and microbial genomes for organism detection in metagenomes

Identifying causative disease agents in human patients from shotgun metagenomic sequencing (SMS) presents a powerful tool to apply when other targeted diagnostics fail. Numerous technical challenges remain, however, before SMS can move beyond the role of research tool. Accurately separating the known and unknown organism content remains difficult, particularly when SMS is applied as a last resort. The true amount of human DNA that remains in a sample after screening against the human reference genome and filtering nonbiological components left from library preparation has previously been underreported. In this study, we create the most comprehensive collection of microbial and reference-free human genetic variation available in a database optimized for efficient metagenomic search by extracting sequences from GenBank and the 1000 Genomes Project. The results reveal new human sequences found in individual Human Microbiome Project (HMP) samples. Individual samples contain up to 95% human sequence, and 4% of the individual HMP samples contain 10% or more human reads. Left unidentified, human reads can complicate and slow down further analysis and lead to inaccurately labeled microbial taxa and ultimately lead to privacy concerns as more human genome data is collected.Metagenomic analysis of human clinical samples is often confounded by the abundance of host genome present in the sample. This can be particularly challenging when trying to uncover a poorly characterized etiologic agent that may or may not be recoverable from a diagnostic next-generation sequencing run (Yozwiak et al. 2012; Wilson et al. 2014). Ideally each sequencer read should be examined and its relationship relative to all previously sequenced organisms accurately reported. Unfortunately, the most sensitive search needed to detect highly divergent organisms does not scale when naively examining an entire sequencing run that may total many gigabases (Zhao et al. 2012). A complementary approach is to use a metagenomic assembler; however, low coverage of individual microbial genomes can limit the potential for good quality assemblies in complex samples (Howe et al. 2014) or samples dominated by host background. Marker-based approaches present an important option for fast estimation of microbial content but only tag a small fraction of the input leaving potentially interesting fragments hidden in a large collection of unlabeled reads (Liu et al. 2011; Berendzen et al. 2012; Segata et al. 2012; Sunagawa et al. 2013; Minot et al. 2014; Tu et al. 2014). Recent efforts have demonstrated substantial progress in labeling all reads by applying ordered searches that attempt to reserve the most computationally expensive analysis for the fewest reads (Nakamura et al. 2009; Zhao et al. 2013; Byrd et al. 2014; Cotten et al. 2014; Takeuchi et al. 2014). A recent example is SURPI (Naccache et al. 2014), which maps reads to the human reference genome to subtract host reads prior to search of the GenBank NT database. DeconSeq is another tool that identifies human reads as contaminants by aligning reads to seven assembled human genomes (Schmieder and Edwards 2011) to detect human variants beyond the single reference genome. A limitation of these approaches, however, is the use of read mapping tools that organize their respective database search against each reference gene/genome independently. This presents a fundamental scaling problem as more organisms in the population are added to the database. For example, the reference database used to build taxonomic profiles for Human Microbiome Project (HMP) samples (Martin et al. 2012) used a microbial reference database totaling 7.3 gigabases (Gb), which represents only a small fraction of the sequenced microbial strains available. In the case of SURPI, although a 42-Gb GenBank NT database is searched, a best first match approach is taken, which requires careful downstream analysis to avoid overly specific calls that fail to recognize highly conserved elements.Nonredundant search of a population of human genomes was recently shown to improve detection sensitivity using a new data structure to map reads to multiple human genomes simultaneously (Huang et al. 2013). This approach used assembled human genomes for reference but excluded microbial genomes. A related approach uses a large k-mer index as implemented in software tools Kraken (Wood and Salzberg 2014) and Livermore Metagenomics Analysis Toolkit (LMAT) (Ames et al. 2013). Kraken supports efficient search by mapping k-mers to each source genome and storing the lowest common ancestor. A highly conserved genetic region found in hundreds of genomes is efficiently identified with the lowest common ancestor (LCA) in a single search step. Recent improvements to LMAT present a similar approach but place a greater emphasis on storing more data in two ways. First, rather than store the LCA for each k-mer, the list of source genomes and the taxonomic hierarchy are stored—up to a user specified limited number of taxonomic identifiers (set to 200)—to improve discrimination at lower taxonomic ranks while placing an upper bound on overall runtime. When a k-mer maps to more genomes than allowed by the threshold, a pruning procedure retains only the higher rank taxonomy identifiers. Second, LMAT uses a larger microbial genome database, which totals >115 gigabases of genomic sequence—assembled draft and complete genomes for virus, bacteria, fungi, protozoa and mitochondrial genomes of eukaryotes. The original database required at least 620 gigabytes (GB) of DRAM, which limited its use to researchers with access to large memory machines. To improve accessibility, the software was recently tuned to exploit a combination of DRAM and NVRAM (e.g., flash drives) (Ames et al. 2014) and in addition reduced the database size to 458 GB for the full and 17 GB for the marker databases, thus allowing the software to run on substantially lower cost machines. These recent optimizations additionally enable the use of a larger cluster, where NVRAM is used as a supplemental memory resource. Scaling is supported in part by the use of NVRAM, which allows use of a much larger local single address space than is possible with DRAM and at a lower cost. For example, the 800 GB of NVRAM available per compute node in this study is commercially available at $5 per GB compared to $10 per GB of DRAM (assuming 1 terabyte of DRAM). A key benefit of NVRAM over traditional high performance disks is the ability to randomly access any part of flash memory similarly to traditional memory at orders of magnitude higher input output operations per second (IOPS) than is possible with traditional single high performance disks. RAID disk configurations and parallel file systems with multiple storage servers increase IOPS, but also increase cost and complexity. Additionally, parallel file systems based on storage area networks (SAN) greatly increase latency compared to node local NVRAM and are not suitable for our extended memory use case. NVRAM represents an important middle memory tier that allows the NVRAM to act as a supplemental memory resource in a way that is not possible with traditional spinning disk technology.