Absence Of Genetic Information Research Articles

Multiset correlation and factor analysis enables exploration of multi-omics data

Multi-omics datasets are becoming more common, necessitating better integration methods to realize their revolutionary potential. Here, we introduce multi-set correlation and factor analysis (MCFA), an unsupervised integration method tailored to the unique challenges of high-dimensional genomics data that enables fast inference of shared and private factors. We used MCFA to integrate methylation markers, protein expression, RNA expression, and metabolite levels in 614 diverse samples from the Trans-Omics for Precision Medicine/Multi-Ethnic Study of Atherosclerosis multi-omics pilot. Samples cluster strongly by ancestry in the shared space, even in the absence of genetic information, while private spaces frequently capture dataset-specific technical variation. Finally, we integrated genetic data by conducting a genome-wide association study (GWAS) of our inferred factors, observing that several factors are enriched for GWAS hits and trans-expression quantitative trait loci. Two of these factors appear to be related to metabolic disease. Our study provides a foundation and framework for further integrative analysis of ever larger multi-modal genomic datasets.

Sensitivity and Specificity of Different Prognostic Systems in Guiding Surveillance for Metastases in Uveal Melanoma.

Uveal melanoma (UM) metastasises in ~50% of patients, most frequently to the liver. Surveillance imaging can provide early detection of hepatic metastases; however, guidance regarding UM patient risk stratification for surveillance is unclear. This study compared sensitivity and specificity of four current prognostic systems, when used for risk stratification for surveillance, on patients treated at the Liverpool Ocular Oncology Centre (LOOC) between 2007-2016 (n = 1047). It found that the Liverpool Uveal Melanoma Prognosticator Online III (LUMPOIII) or Liverpool Parsimonious Model (LPM) offered greater specificity at equal levels of sensitivity than the American Joint Committee on Cancer (AJCC) system or monosomy 3 alone, and suggests guidance to achieve 95% sensitivity and 51% specificity (i.e., how to detect the same number of patients with metastases, while reducing the number of negative scans). For example, 180 scans could be safely avoided over 5 years in 200 patients using the most specific approach. LUMPOIII also offered high sensitivity and improved specificity over the AJCC in the absence of genetic information, making the result relevant to centres that do not perform genetic testing, or where such testing is inappropriate or fails. This study provides valuable information for clinical guidelines for risk stratification for surveillance in UM.

Comparing estimates of census and effective population size in an endangered amphibian

AbstractThe field of conservation has seen a shift in focus from monitoring trends in census population size to trends in ‘effective’ population size. Numerous genetic methods exist for estimating effective population size, resulting in uncertainty among conservation practitioners as to which methods are most appropriate when conducting population assessments or evaluating recovery efforts. Demographic approaches offer a promising avenue to provide a link between census and effective population size using life‐history information, but rarely do studies have all three sources of data (genetic, demographic, life history) necessary to perform an explicit evaluation of their performance. Using data from a long‐term study of reticulated flatwoods salamanders (Ambystoma bishopi) in western Florida, USA, we assessed the magnitude of temporal variation in census population sizes and the effective number of breeders of two breeding populations to (1) document changes in the number of breeding adults over the 9‐year study duration, (2) determine whether and provide similar information about population size and trends and (3) compare alternative demographic and genetic approaches for estimating . We found that genetic estimates of , particularly if averaged across multiple estimation methods, closely tracked spatiotemporal variation in . Demographic estimates of also closely tracked but were sensitive to the assumed variance in reproductive success. In the absence of genetic information, detailed knowledge of mating systems and the environmental factors that skew reproductive contributions appear necessary for demographic to reliably inform management decisions. In these populations, appears too small (<40 individuals) to confer long‐term genetic resilience, highlighting the importance of restoring landscape connectivity and indicating that caution must be taken when sourcing animals for reintroduction efforts. More generally, our study reveals insights into the utility of alternative estimation methods in guiding recovery efforts of threatened and endangered species.

Genetic Consequences of Multiple Translocations of the Banded Hare-Wallaby in Western Australia

Many Australian mammal species now only occur on islands and fenced mainland havens free from invasive predators. The range of one species, the banded hare-wallaby (Lagostrophus fasciatus), had contracted to two offshore islands in Western Australia. To improve survival, four conservation translocations have been attempted with mixed success, and all occurred in the absence of genetic information. Here, we genotyped seven polymorphic microsatellite markers in two source (Bernier Island and Dorre Island), two historic captive, and two translocated L. fasciatus populations to determine the impact of multiple translocations on genetic diversity. Subsequently, we used population viability analysis (PVA) and gene retention modelling to determine scenarios that will maximise demographic resilience and genetic richness of two new populations that are currently being established. One translocated population (Wadderin) has undergone a genetic bottleneck and lost 8.1% of its source population’s allelic diversity, while the other (Faure Island) may be inbred. We show that founder number is a key parameter when establishing new L. fasciatus populations and 100 founders should lead to high survival probabilities. Our modelling predicts that during periodic droughts, the recovery of source populations will be slower post-harvest, while 75% more animals—about 60 individuals—are required to retain adequate allelic diversity in the translocated population. Our approach demonstrates how genetic data coupled with simulations of stochastic environmental events can address central questions in translocation programmes.

Abstract 13743: A Tale of Syncope, Prolonged QT and ICD

Introduction: Long QT syndrome (LQTS) was first described in the 1960s. It manifests clinically as syncope, cardiac arrest or sudden cardiac death. LQTS can be caused by 15 different genes. These mutations lead to action potential prolongation by causing impaired repolarizing currents. Case Discussion: A 29-year-old previously healthy Caucasian woman was admitted after recurrent episodes of syncope that happened within 1-month prior to the presentation. She was hemodynamically stable with normal vitals. Her ECG showed normal sinus rhythm with corrected QT (QTc) of 598ms. In the ED, she suffered an episode of sustained monomorphic ventricular tachycardia (VT) and underwent cardioversion. She was started on amiodarone infusion. Serial ECGs showed prolonged QTc. She had another episode of pulseless VT that terminated without defibrillation. She was transferred to our facility for further care. Her family history was significant for paternal aunt who had died unexpectedly at the age of 39. All her lab work including electrolytes, thyroid panel, cardiac enzymes, inflammatory markers and extended drug screen was unrevealing. Transthoracic echocardiogram showed normal biventricular size and function. Decision making: She was started on propranolol for possible LQTS. Cardiac MR did not show any evidence of structural abnormalities. Genetic panel was sent. Since myocarditis or familial LQTS could not be ruled out, we proceeded with implantable cardioverter defibrillator (ICD) implantation for secondary prevention. She was discharged home on nadolol. Conclusion: In the absence of genetic information, LQTS can be diagnosed in symptomatic patients with QTc >480msec on serial ECGs after excluding secondary causes. Schwartz score comprising of ECG findings, symptoms, clinical & family history is diagnostic when greater than 3.5. Beta-blockers are indicated in all patients with a clinical diagnosis. Patients must avoid any QT prolonging agents and strenuous exercise. An ICD is indicated in patients who suffered cardiac arrest. ICD may also be considered for primary prevention in high risk patients.

Risk Stratification and Transfusion Dependence Are Independent Risk Factors for Cardiovascular Disease (CVD) in Myelodysplastic Syndrome (MDS)

Background: MDS has been largely excluded from epidemiological studies that demonstrate an increased risk of CVD in older adults with cancer. Nevertheless, somatic mutations leading to clonally restricted hematopoiesis are associated with increased risk of CVD and CVD constitutes the most common non-disease-related cause of death among patients with MDS. Using the SEER Medicare tumor registry, we previously demonstrated that MDS was independently associated with increased risk of CVD and cardiovascular mortality in older adults, thus constituting an important but underestimated cardiovascular hazard. In this study, we sought to determine the influence of MDS-specific factors on the risk of CVD among MDS patients. Methods: We used the SEER Medicare database to identify individuals 66 years or older with an incident diagnosis of MDS between 2004 and 2014. Incident CVD was defined as new myocardial infraction and/or ischemic stroke occurring after diagnosis of MDS. Events were identified with ICD9 and HCPCS codes in Medicare claims from the date of MDS diagnosis to death or study end. Subjects with CVD prior to MDS diagnosis were excluded. Comorbidity burden was estimated with the Charlson comorbidity index (CCI), calculated using claims from the year before MDS diagnosis. MDS was classified into low, intermediate and high risk using ICD-O morphology codes. Subjects requiring 2 or more blood product transfusions within 60 days from MDS diagnosis were considered transfusion dependent. Cumulative incidence functions were used to assess the risk of new events. Cox proportional hazards models, extended for time dependent covariates and coefficients, were used to evaluate clinical variables associated with CVD. Results: The cohort included 13,972 subjects with MDS, most were male (55.2%), non-hispanic white (82.1%), and from metropolitan areas (82.5%). Median age at MDS diagnosis was 82 years (IQR, 78 - 87). MDS was of low, intermediate and high risk in 18.1%, 68.8% and 13.1% respectively. The median CCI score was 0 (IQR, 0 - 2). Red blood cells (RBC) and platelet transfusion dependence occurred in 29% and 6.1% of subjects. The 10-year cumulative incidence of CVD was 47%, 54% and 65% in subjects with low, intermediate and high-risk MDS, respectively (p<0.016; Fig. 1). Similarly, the 10-year cumulative incidence of CVD was 57.5% in RBC transfusion-dependent subjects vs. 49% in non-dependent individuals (p<0.001; Fig 2A). Other factors that significantly influenced the hazard of CVD were male sex, older age and higher CCI. The effect of higher MDS risk (HR 1.1, 95%CI, 1.02 - 1.2) and RBC transfusion dependence (HR 1.3, 95%CI, 1.2 - 1.5) on increased incidence of CVD persisted after accounting for these factors in a multivariate Cox model adjusted for age, gender and CCI. Platelet transfusion dependence did not significantly influence the cumulative incidence of CVD (p=0.252; Fig 2B) or the hazard for CVD in multivariate analysis (HR 0.8, 95%CI, 0.6 - 1.2). Both higher risk MDS (HR 1.6, 95%CI, 1.5 - 1.7) and blood product transfusion dependence (HR 1.9, 95%CI 1.8 - 2) were associated with increased mortality in subjects with MDS. Conclusion: In a large cohort of elderly Medicare beneficiaries with incident diagnosis of MDS, the cumulative incidence of CVD was greater in subjects with higher-risk MDS as determined by bone marrow morphology and in those who were RBC transfusion dependent. Both MDS risk and RBC transfusion dependence were independent risk factors for CVD in multivariate analysis accounting for demographic factors and comorbidity burden. These factors have been previously associated with increased mortality and risk of leukemia transformation in MDS but their role in risk of CVD has not been previously reported. CVD is an important cause of mortality among older adults with MDS and these results can help identify high risk patients for whom strict cardiovascular screening and preventive strategies could impact mortality. Though we used validated methods, study limitations include relying on administrative claims, absence of genetic information and unmeasured confounding factors. Prospective studies and further preclinical research is needed to clarify the mechanisms of CVD in patients with MDS. Figure 1 Disclosures Verma: Janssen: Research Funding; Stelexis: Equity Ownership, Honoraria; Acceleron: Honoraria; Celgene: Honoraria; BMS: Research Funding.

De novo assembly and functional annotation of the emma field cricket (Teleogryllus emma) transcriptome

Abstract In recent years, insects have been used in animal-based foods for sustainable live-stock management and in the human diet as therapeutic foods. These insects are classified as edible and are widely consumed around the world for their broad medicinal and nutritional values. Antimicrobial peptides/proteins (AMPs) confer resistance to microbial pathogens and contribute to the immune function of insects. The emma field cricket (Teleogryllus emma) is widely consumed around the world for its broad-spectrum therapeutic effects. However, the absence of genetic information on T. emma is an obstacle to understanding the diversity and activity of its AMPs. Hence, deep sequencing of the T. emma transcriptome and expression profiling are essential. We report whole-body de novo transcriptome profiling of T. emma and analysis of its potential bacterial immunization-induced AMPs. We assembled 18.2 Gb of Illumina reads, generated 102,311 transcripts (minimum length: >500 bp, N50: 2585 bp), and identified 67,838 unigenes. Of these unigenes, 39,937 (58.87%) matched known proteins in a Basic Local Alignment Search Tool search of the National Center for Biotechnology Information database. Functional analysis demonstrated that the 39,937 unigenes clustered under 5707 Gene Ontology terms. In addition, we identified 10 cDNAs encoding new AMPs via an in-silico approach using public databases. Our transcriptome data set and AMP profile greatly improve the understanding of T. emma genetics and provide a huge number of gene sequences, including those for genes of known and unknown function, for further study.

De novo transcriptome analysis of the egg parasitoid Trichogramma chilonis Ishii (Hymenoptera: Trichogrammatidae): A biological control agent

Trichogramma chilonis Ishii is an endoparasitic wasp widely used as a biological control agent of economically important pests. However, absence of genetic information on T. chilonis hampers further research, especially at the molecular level. Here, we performed the first RNA-Seq of the adult T. chilonis. A total of 18,372,639 high-quality reads were generated using Illumina pair-ended sequencing platform. De novo assembly resulted into 24,488 transcripts, with mean length and N50 values of 1444.2 bp and 2667 bp, respectively. Of the transcripts, 14,643 exhibited putative homology with class Insecta proteins from UniPROT database. To understand the contig functions, gene descriptions, gene ontology and cluster of orthologous group annotations were analyzed. Following functional annotation, a number of transcripts encoding detoxification, target receptor action and olfaction- related proteins were identified and analyzed. In addition, 4520 transcripts containing simple sequence repeats, ranging from dinucleotide to hexanucleotide repeats were discovered, with trinucleotide being the most enriched repeats. This new sequence resource will shed light on various molecular mechanisms in T. chilonis and related species.

Proton Nuclear Magnetic Resonance-Spectroscopic Discrimination of Wines Reflects Genetic Homology of Several Different Grape (V. vinifera L.) Cultivars.

Background and AimsProton nuclear magnetic resonance spectroscopy coupled multivariate analysis (1H NMR-PCA/PLS-DA) is an important tool for the discrimination of wine products. Although 1H NMR has been shown to discriminate wines of different cultivars, a grape genetic component of the discrimination has been inferred only from discrimination of cultivars of undefined genetic homology and in the presence of many confounding environmental factors. We aimed to confirm the influence of grape genotypes in the absence of those factors.Methods and ResultsWe applied 1H NMR-PCA/PLS-DA and hierarchical cluster analysis (HCA) to wines from five, variously genetically-related grapevine (V. vinifera) cultivars; all grown similarly on the same site and vinified similarly. We also compared the semi-quantitative profiles of the discriminant metabolites of each cultivar with previously reported chemical analyses. The cultivars were clearly distinguishable and there was a general correlation between their grouping and their genetic homology as revealed by recent genomic studies. Between cultivars, the relative amounts of several of the cultivar-related discriminant metabolites conformed closely with reported chemical analyses.ConclusionsDifferences in grape-derived metabolites associated with genetic differences alone are a major source of 1H NMR-based discrimination of wines and 1H NMR has the capacity to discriminate between very closely related cultivars.Significance of the StudyThe study confirms that genetic variation among grape cultivars alone can account for the discrimination of wine by 1H NMR-PCA/PLS and indicates that 1H NMR spectra of wine of single grape cultivars may in future be used in tandem with hierarchical cluster analysis to elucidate genetic lineages and metabolomic relations of grapevine cultivars. In the absence of genetic information, for example, where predecessor varieties are no longer extant, this may be a particularly useful approach.

Cloning and characterization of a norbelladine 4'-O-methyltransferase involved in the biosynthesis of the Alzheimer's drug galanthamine in Narcissus sp. aff. pseudonarcissus.

Galanthamine is an Amaryllidaceae alkaloid used to treat the symptoms of Alzheimer’s disease. This compound is primarily isolated from daffodil (Narcissus spp.), snowdrop (Galanthus spp.), and summer snowflake (Leucojum aestivum). Despite its importance as a medicine, no genes involved in the biosynthetic pathway of galanthamine have been identified. This absence of genetic information on biosynthetic pathways is a limiting factor in the development of synthetic biology platforms for many important botanical medicines. The paucity of information is largely due to the limitations of traditional methods for finding biochemical pathway enzymes and genes in non-model organisms. A new bioinformatic approach using several recent technological improvements was applied to search for genes in the proposed galanthamine biosynthetic pathway, first targeting methyltransferases due to strong signature amino acid sequences in the proteins. Using Illumina sequencing, a de novo transcriptome assembly was constructed for daffodil. BLAST was used to identify sequences that contain signatures for plant O-methyltransferases in this transcriptome. The program HAYSTACK was then used to identify methyltransferases that fit a model for galanthamine biosynthesis in leaf, bulb and inflorescence tissues. One candidate gene for the methylation of norbelladine to 4′-O-methylnorbelladine in the proposed galanthamine biosynthetic pathway was identified. This methyltransferase cDNA was expressed in E. coli and the protein purified by affinity chromatography. The resulting protein was found to be a norbelladine 4′-O-methyltransferase (NpN4OMT) of the proposed galanthamine biosynthetic pathway.

Exome Sequencing of Index Patients with Retinal Dystrophies as a Tool for Molecular Diagnosis

BackgroundRetinal dystrophies (RD) are a group of hereditary diseases that lead to debilitating visual impairment and are usually transmitted as a Mendelian trait. Pathogenic mutations can occur in any of the 100 or more disease genes identified so far, making molecular diagnosis a rather laborious process. In this work we explored the use of whole exome sequencing (WES) as a tool for identification of RD mutations, with the aim of assessing its applicability in a diagnostic context.Methodology/Principal FindingsWe ascertained 12 Spanish families with seemingly recessive RD. All of the index patients underwent mutational pre-screening by chip-based sequence hybridization and resulted to be negative for known RD mutations. With the exception of one pedigree, to simulate a standard diagnostic scenario we processed by WES only the DNA from the index patient of each family, followed by in silico data analysis. We successfully identified causative mutations in patients from 10 different families, which were later verified by Sanger sequencing and co-segregation analyses. Specifically, we detected pathogenic DNA variants (∼50% novel mutations) in the genes RP1, USH2A, CNGB3, NMNAT1, CHM, and ABCA4, responsible for retinitis pigmentosa, Usher syndrome, achromatopsia, Leber congenital amaurosis, choroideremia, or recessive Stargardt/cone-rod dystrophy cases.Conclusions/SignificanceDespite the absence of genetic information from other family members that could help excluding nonpathogenic DNA variants, we could detect causative mutations in a variety of genes known to represent a wide spectrum of clinical phenotypes in 83% of the patients analyzed. Considering the constant drop in costs for human exome sequencing and the relative simplicity of the analyses made, this technique could represent a valuable tool for molecular diagnostics or genetic research, even in cases for which no genotypes from family members are available.

De Novo Sequencing and Transcriptome Analysis of the Central Nervous System of Mollusc Lymnaea stagnalis by Deep RNA Sequencing

The pond snail Lymnaea stagnalis is among several mollusc species that have been well investigated due to the simplicity of their nervous systems and large identifiable neurons. Nonetheless, despite the continued attention given to the physiological characteristics of its nervous system, the genetic information of the Lymnaea central nervous system (CNS) has not yet been fully explored. The absence of genetic information is a large disadvantage for transcriptome sequencing because it makes transcriptome assembly difficult. We here performed transcriptome sequencing for Lymnaea CNS using an Illumina Genome Analyzer IIx platform and obtained 81.9 M of 100 base pair (bp) single end reads. For de novo assembly, five programs were used: ABySS, Velvet, OASES, Trinity and Rnnotator. Based on a comparison of the assemblies, we chose the Rnnotator dataset for the following blast searches and gene ontology analyses. The present dataset, 116,355 contigs of Lymnaea transcriptome shotgun assembly (TSA), contained longer sequences and was much larger compared to the previously reported Lymnaea expression sequence tag (EST) established by classical Sanger sequencing. The TSA sequences were subjected to blast analyses against several protein databases and Aplysia EST data. The results demonstrated that about 20,000 sequences had significant similarity to the reported sequences using a cutoff value of 1e-6, and showed the lack of molluscan sequences in the public databases. The richness of the present TSA data allowed us to identify a large number of new transcripts in Lymnaea and molluscan species.

Rapid growth of a Eurasian haplotype of Phragmites australis in a restored brackish marsh in Louisiana, USA

While numerous studies have documented patterns of invasion by non-indigenous plant species, few have considered the invasive properties of non-native genotypes of native species. Characteristics associated with specific genotypes, such as tolerance to disturbance, may mistakenly be applied to an entire species in the absence of genetic information, which consequently may affect management decisions. We report here on the incidence and growth of an introduced lineage of Phragmites australis in the Gulf of Mexico coastal zone of Louisiana. P. australis was collected from nine separate locations for inclusion in a series of growth experiments. Chloroplast DNA analysis indicated that specimens collected from four locations in the Mississippi River Delta represented the introduced Eurasian haplotype; the remainder represented the gulf coast haplotype. Three distinct genotypes, or clones, were identified within each haplotype via analysis using amplified fragment length polymorphisms, which also revealed reduced genetic diversity of the gulf coast clones compared to the Eurasian clones. Clones of each haplotype were planted along with three other native macrophytes at similar densities in a restored brackish marsh and monitored for growth. After 14 months, the Eurasian haplotype had spread vegetatively to cover about 82% of the experimental plots, more than four times the coverage (18%) of the gulf coast haplotype. Thus, the use of P. australis plantings for wetland restoration should consider the genetic lineage of plants used since our results indicate the potential of the Eurasian haplotype to grow rapidly at newly restored sites. This rapid growth may limit the establishment of more slowly growing native species.

Rapid detection of Panton–Valentine leukocidin-positive Staphylococcus aureus by real-time PCR targeting the lukS-PV gene

In order to assess the speed and accuracy of a real-time PCR assay targeting the lukS-PV gene of Panton-Valentine leukocidin (PVL)-positive Staphylococcus aureus, 700 S. aureus strains were tested and the results were compared to those achieved with block cycler PCR. Cross-reactivity was tested with 166 other bacterial species. Using this homogeneous real-time PCR assay format, the presence or absence of genetic information for PVL, which is also found in community-associated methicillin-resistant S. aureus, was correctly identified from pure culture and directly in various types of clinical specimens.

Evidence for low-molecular weight antibodies in the serum of a urodele amphibian, Ambystoma mexicanum

Adult axolotls ( A. mexicanum) were hyperimmunized with the haptenic determinant, azobenzene-arsonate (ARS). Specific antibodies were isolated from their serum by immune-affinity chromatography on immobilized ARS columns. Analysis of the specific ARS-binding molecules by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) indicated that these animals were capable of producing both high molecular-weight (presumably IgM) and low molecular-weight antibodies to the ARS hapten. The low molecular-weight anti-ARS antibody produced by one inbred colony of axolotls did not show any restricted heterogeneity, as assessed by isoelectric focusing. Our results suggest that regulatory events, not the absence of genetic information, may be responsible for the apparent lack of Ig isotype diversity in this species.