CRISPR-based homing gene drives have sparked both enthusiasm and deep concerns due to their potential for genetically altering entire species. This raises the question about our ability to prevent the unintended spread of such drives from the laboratory into a natural population. Here, we experimentally demonstrate the suitability of synthetic target site drives as well as split drives as flexible safeguarding strategies for gene drive experiments by showing that their performance closely resembles that of standard homing drives in Drosophila melanogaster. Using our split drive system, we further find that maternal deposition of both Cas9 and gRNA is required to form resistance alleles in the early embryo and that maternally-deposited Cas9 alone can power germline drive conversion in individuals that lack a genomic source of Cas9.
Objective: To determine whether the incidence and clinical features of pediatric multiple sclerosis (MS) and other forms of pediatric acquired demyelinating syndromes (ADS) vary by race/ethnicity in a population-based cohort. Methods:We used a combination of electronic database searches followed by complete medical records review to identify all children diagnosed with MS and ADS in the multiethnic membership of Kaiser Permanente Southern California from January 1, 2004, to December 31, 2009. Incidence rates were standardized to the US census by age and gender. Results:We identified 81 incident cases of ADS from 4.87 million person-years of observation in children 0-18 years of age. The incidence rate of pediatric MS was 0.51 per 100,000 personyears (95% confidence interval [CI] 0.33-0.75) and incidence of other forms of ADS including optic neuritis, transverse myelitis, other forms of clinically isolated syndrome (CIS), and acute disseminated encephalomyelitis (ADEM) was 1.56 (95% CI 1.23-1.95) for an overall incidence of ADS of 1.66 per 100,000 person-years (95% CI 1.32-2.06). Incidence of ADS was higher in black (4.4 per 100,000 person-years, 95% CI 2.5-7.2, p Ͻ 0.001) and Asian/Pacific Islander (2.8, 95% CI 1.2-5.2, p ϭ 0.02) than white (1.03, 95% CI 0.6-1.7) and Hispanic (1.5, 95% CI 1.1-2.1, per 100,000 person-years) children. Black children were also significantly more likely to have MS than white children (p ϭ 0.001). Children who presented with ADEM were significantly younger than children with other types of ADS clinical presentations (mean age 5.6, range 0.7-17.6 years vs 14.6, range 2.7-18.5, respectively).
CRISPR-based gene drives have sparked both enthusiasm and deep concerns due to their potential for genetically altering entire species. This raises the question about our ability to prevent the unintended spread of such drives from the laboratory into a natural population.Here, we experimentally demonstrate the suitability of synthetic target sites and split drives as flexible safeguarding strategies for gene drive experiments.
Gene drives could allow for control of vector-borne diseases by directly suppressing vector populations or spreading genetic payloads designed to reduce pathogen transmission. Clustered regularly interspaced short palindromic repeat (CRISPR) homing gene drives work by cleaving wild-type alleles, which are then converted to drive alleles by homology-directed repair, increasing the frequency of the drive in a population over time. However, resistance alleles can form when end-joining repair takes place in lieu of homology-directed repair. Such alleles cannot be converted to drive alleles, which would eventually halt the spread of a drive through a population. To investigate the effects of natural genetic variation on resistance formation, we developed a CRISPR homing gene drive in Drosophila melanogaster and crossed it into the genetically diverse Drosophila Genetic Reference Panel (DGRP) lines, measuring several performance parameters. Most strikingly, resistance allele formation postfertilization in the early embryo ranged from 7 to 79% among lines and averaged 42 6 18%. We performed a genome-wide association study using our results in the DGRP lines, and found that the resistance and conversion rates were not explained by common alleles of large effect, but instead there were several genetic polymorphisms showing weak association. RNA interference knockdown of several genes containing these polymorphisms confirmed their effect, but the small effect sizes imply that their manipulation would likely yield only modest improvements to the efficacy of gene drives.
Estimating fitness differences between allelic variants is a central goal of experimental evolution. Current methods for inferring such differences from allele frequency time series typically assume that the effects of selection can be described by a fixed selection coefficient. However, fitness is an aggregate of several components including mating success, fecundity, and viability. Distinguishing between these components could be critical in many scenarios. Here, we develop a flexible maximum likelihood framework that can disentangle different components of fitness from genotype frequency data, and estimate them individually in males and females. As a proof-of-principle, we apply our method to experimentally evolved cage populations of Drosophila melanogaster, in which we tracked the relative frequencies of a loss-of-function and wild-type allele of yellow. This X-linked gene produces a recessive yellow phenotype when disrupted and is involved in male courtship ability. We find that the fitness costs of the yellow phenotype take the form of substantially reduced mating preference of wild-type females for yellow males, together with a modest reduction in the viability of yellow males and females. Our framework should be generally applicable to situations where it is important to quantify fitness components of specific genetic variants, including quantitative characterization of the population dynamics of CRISPR gene drives.
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Estimating fitness differences between allelic variants is a central goal of experimental evolution. Current methods for inferring selection from allele frequency time series typically assume that evolutionary dynamics at the locus of interest can be described by a fixed selection coefficient. However, fitness is an aggregate of several components including mating success, fecundity, and viability, and distinguishing between these components could be critical in many scenarios. Here we develop a flexible maximum likelihood framework that can disentangle different components of fitness and estimate them individually in males and females from genotype frequency data. As a proof-of-principle, we apply our method to experimentally-evolved cage populations of Drosophila melanogaster, in which we tracked the relative frequencies of a loss-of-function and wild-type allele of yellow. This X-linked gene produces a recessive yellow phenotype when disrupted and is involved in male courtship ability. We find that the fitness costs of the yellow phenotype take the form of substantially reduced mating preference of wild-type females for yellow males, together with a modest reduction in the viability of yellow males and females. Our framework should be generally applicable to situations where it is important to quantify fitness components of specific genetic variants, including quantitative characterization of the population dynamics of CRISPR gene drives.
establishing the diagnosis. However, the final diagnosis relied on pathological investigations. In spite of the benign nature of myxomas, they may inevitably cause valvular dysfunction and secondary pulmonary hyperten-sion and have a significant propensity to embolize the pulmonary artery (1). Pulmonary artery and pulmonary valve myxomas have the common features of right heart system, such as predilections of right ventricular obstruction, right-sided valve insufficiencies, and pulmonary embolism; however, their special characteristic is a smaller size. Pulmonary myxomas could occur isolated or in association with other congenital heart defects or acquired disorders. There were more myxomas arising from the pulmonary valve than from the pulmonary artery or from pulmonary valve and pulmonary artery. Most pulmonary valve myxomas arose from the valve leaflets, and most pulmonary artery myxomas arose from the main pulmonary artery. Because of potential hazards and occasional misdiagnosis, the patients endow an early surgical treatment upon diagnosis (5). Most patients warranted a surgical resection of the myxoma under standard cardiopulmonary bypass, while some patients were operated under normothermic cardiopulmonary bypass or deep hypothermic circulatory arrest. Concurrent procedures to myxoma resection, such as pulmonary valve repair or replacement, or right ventricular outflow tract reconstruction should be performed simultaneously. An early surgical treatment is warranted upon diagnosis because of potential hemodynamic disturbances and predilection of embolization. Most patients have a good prognosis following surgical treatment. References 1. Huang CY, Huang CH, Yang AH, Wu MH, Ding YA, Yu WC. Solitary pulmonary artery myxoma manifesting as pulmonary embolism and subacute cor pulmonale. Am J Med 2003; 115: 680-1. Crossref 2. Blodorn M. Myxoma of the pulmonary valve, respectively of the pulmonary artery. Zentralbl Allg Pathol 1955; 94: 283-9. 3. Restrepo CS, Betancourt SL, Martinez-Jimenez S, Gutierrez FR. Tumors of the pulmonary artery and veins. Semin Ultrasound CT MR 2012; 33: 580-90. Crossref 4. Barış VO, Uslu A, Gerede DM, Kılıçkap M. Rare cause of dyspnoea: pulmonary artery myxoma. Eur Heart J We report a 63-year-old patient with prior coronary artery bypass surgery and recent history of recurrent hospital admissions for refractory heart failure because of ischemic car-diomyopathy and sustained ventricular tachycardia. The patient underwent ICD implantation through left subclavian vein approach (Medtronic, single chamber, model-Maximo II VR, D284VRC, with 6947 ventricular active fixation lead). This case report describes the first patient, to our knowledge, with defi-brillator lead dislocation after manual lumbar traction for low back pain. The patient was admitted to the emergency service with severe chest pain, and electrocardiography revealed 0.5-1 mm ST-segment elevation in leads DII, DIII, and aVF. After initial evaluation, coronary angiography and percutaneous coronary intervention were immediately ...
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