The relationship between eDNA particle concentration and organism abundance in nature is strengthened by allometric scaling.

Organism abundance is a critical parameter in ecology, but its estimation is often challenging. Approaches utilizing eDNA to indirectly estimate abundance have recently generated substantial interest. However, preliminary correlations observed between eDNA concentration and abundance in nature are typically moderate in strength with significant unexplained variation. Here, we apply a novel approach to integrate allometric scaling coefficients into models of eDNA concentration and organism abundance. We hypothesize that eDNA particle production scales nonlinearly with mass, with scaling coefficients < 1. Wild populations often exhibit substantial variation in individual body size distributions; we therefore predict that the distribution of mass across individuals within a population will influence population-level eDNA production rates. To test our hypothesis, we collected standardized body size distribution and mark-recapture abundance data using whole-lake experiments involving nine populations of brook trout. We correlated eDNA concentration with three metrics of abundance: density (individuals/ha), biomass (kg/ha) and allometrically scaled mass (ASM) (∑(individual mass0.73 )/ha). Density and biomass were both significantly positively correlated with eDNA concentration (adj. r2  = 0.59 and 0.63, respectively), but ASM exhibited improved model fit (adj. r2  = 0.78). We also demonstrate how estimates of ASM derived from eDNA samples in "unknown" systems can be converted to biomass or density estimates with additional size-structure data. Future experiments should empirically validate allometric scaling coefficients for eDNA production, particularly where substantial intraspecific size distribution variation exists. Incorporating allometric scaling may improve predictive models to the extent that eDNA concentration may become a reliable indicator of abundance in nature.

Novel SBF2 mutations and clinical spectrum of Charcot-Marie-Tooth neuropathy type 4B2.

Biallelic SBF2 mutations cause Charcot-Marie-Tooth disease type 4B2 (CMT4B2), a sensorimotor neuropathy with autosomal recessive inheritance and association with glaucoma. Since the discovery of the gene mutation, only few additional patients have been reported. We identified seven CMT4B2 families with nine different SBF2 mutations. Revisiting genetic and clinical data from our cohort and the literature, SBF2 variants were private mutations, including exon-deletion and de novo variants. The neuropathy typically started in the first decade after normal early motor development, was predominantly motor and had a rather moderate course. Electrophysiology and nerve biopsies indicated demyelination and excess myelin outfoldings constituted a characteristic feature. While neuropathy was >90% penetrant at age 10 years, glaucoma was absent in ~40% of cases but sometimes developed with age. Consequently, SBF2 mutation analysis should not be restricted to individuals with coincident neuropathy and glaucoma, and CMT4B2 patients without glaucoma should be followed for increased intraocular pressure. The presence of exon-deletion and de novo mutations demands comprehensive mutation scanning and family studies to ensure appropriate diagnostic approaches and genetic counseling.

Long-term safety and efficacy of bimatoprost solution 0·03% application to the eyelid margin for the treatment of idiopathic and chemotherapy-induced eyelash hypotrichosis: a randomized controlled trial.

BACKGROUND: Bimatoprost ophthalmic solution 0·03% is approved in several countries for the treatment of eyelash hypotrichosis. Previous trials were limited to 4 months of treatment and primarily idiopathic hypotrichosis. OBJECTIVES: To evaluate the long-term safety and efficacy of bimatoprost in patients with idiopathic or chemotherapy-induced hypotrichosis. METHODS: This multicentre, double-masked, randomized, parallel-group study included two 6-month treatment periods [treatment period 1 (TP1) and treatment period 2 (TP2)]. Patients with idiopathic hypotrichosis were randomized to three treatment groups: (i) bimatoprost (TP1 and TP2); (ii) bimatoprost (TP1) and vehicle (TP2); and (iii) vehicle (TP1) and bimatoprost (TP2). Patients with chemotherapy-induced hypotrichosis were randomized to two treatment groups: (i) bimatoprost or vehicle (TP1) and (ii) bimatoprost (TP2). Primary end point was a composite of at least a one-grade improvement in investigator-assessed Global Eyelash Assessment and at least a three-point improvement in patient-reported Eyelash Satisfaction Questionnaire Domain 2 at month 4. Secondary measures included digitally assessed eyelash characteristics. RESULTS: The primary efficacy end point was met in both populations (idiopathic responder rate was 40·2% for bimatoprost vs. 6·8% for vehicle; postchemotherapy responder rate was 37·5% for bimatoprost vs. 18·2% for vehicle). Efficacy by month 6 was maintained (idiopathic) or enhanced (postchemotherapy) at 12 months. Treatment effects were maintained for approximately 2 months but markedly diminished 4-6 months following treatment cessation in patients with idiopathic hypotrichosis. No drug-related serious adverse events were reported. CONCLUSIONS: Daily treatment with bimatoprost ophthalmic solution 0·03% for 1 year was effective and well tolerated in patients with idiopathic and chemotherapy-induced hypotrichosis.

Pathological non-response to chemotherapy in a neoadjuvant setting of breast cancer: an inter-institutional study.

To identify markers of non-response to neoadjuvant chemotherapy (NAC) that could be used in the adjuvant setting. Sixteen pathologists of the European Working Group for Breast Screening Pathology reviewed the core biopsies of breast cancers treated with NAC and recorded the clinico-pathological findings (histological type and grade; estrogen, progesterone receptors, and HER2 status; Ki67; mitotic count; tumor-infiltrating lymphocytes; necrosis) and data regarding the pathological response in corresponding surgical resection specimens. Analyses were carried out in a cohort of 490 cases by comparing the groups of patients showing pathological complete response (pCR) and partial response (pPR) with the group of non-responders (pathological non-response: pNR). Among other parameters, the lobular histotype and the absence of inflammation were significantly more common in pNR (p < 0.001). By ROC curve analyses, cut-off values of 9 mitosis/2 mm(2) and 18% of Ki67-positive cells best discriminated the pNR and pCR + pPR categories (p = 0.018 and < 0.001, respectively). By multivariable analysis, only the cut-off value of 9 mitosis discriminated the different response categories (p = 0.036) in the entire cohort. In the Luminal B/HER2- subgroup, a mitotic count <9, although not statistically significant, showed an OR of 2.7 of pNR. A lobular histotype and the absence of inflammation were independent predictors of pNR (p = 0.024 and <0.001, respectively). Classical morphological parameters, such as lobular histotype and inflammation, confirmed their predictive value in response to NAC, particularly in the Luminal B/HER2- subgroup, which is a challenging breast cancer subtype from a therapeutic point of view. Mitotic count could represent an additional marker but has a poor positive predictive value.

Chapter 7: Assessing Habitability Beyond Earth.

All known life on Earth inhabits environments that maintain conditions between certain extremes of temperature, chemical composition, energy availability, and so on (Chapter 6). Life may have emerged in similar environments elsewhere in the Solar System and beyond. The ongoing search for life elsewhere mainly focuses on those environments most likely to support life, now or in the past-that is, potentially habitable environments. Discussion of habitability is necessarily based on what we know about life on Earth, as it is our only example. This chapter gives an overview of the known and presumed requirements for life on Earth and discusses how these requirements can be used to assess the potential habitability of planetary bodies across the Solar System and beyond. We first consider the chemical requirements of life and potential feedback effects that the presence of life can have on habitable conditions, and then the planetary, stellar, and temporal requirements for habitability. We then review the state of knowledge on the potential habitability of bodies across the Solar System and exoplanets, with a particular focus on Mars, Venus, Europa, and Enceladus. While reviewing the case for the potential habitability of each body, we summarize the most prominent and impactful studies that have informed the perspective on where habitable environments are likely to be found.

Chapter 11: Astrobiology Education, Engagement, and Resources.

Although astrobiology is a relatively new field of science, the questions it seeks to answer (e.g., "What is life?" "What does life require?") have been investigated for millennia. In recent decades, formal programs dedicated specifically to the science of astrobiology have been organized at academic, governmental, and institutional scales. Constructing educational programs around this emerging science relies on input from broad expertise and backgrounds. Because of the interdisciplinary nature of this field, career pathways in astrobiology often begin in more specific fields such as astronomy, geology, or biology, and unlike many other sciences, typically involve substantial training outside one's primary discipline. The recent origin of astrobiology as a field of science has led to strong collaborations with education research in the development of astrobiology courses and offers a unique instructional laboratory for further pedagogical studies. This chapter is intended to support students, educators, and early career scientists by connecting them to materials and opportunities that the authors and colleagues have found advantageous. Annotated lists of relevant programs and resources are included as a series of appendices in the supplementary material.

Chapter 1: The Astrobiology Primer 3.0.

The Astrobiology Primer 3.0 (ABP3.0) is a concise introduction to the field of astrobiology for students and others who are new to the field of astrobiology. It provides an entry into the broader materials in this supplementary issue of Astrobiology and an overview of the investigations and driving hypotheses that make up this interdisciplinary field. The content of this chapter was adapted from the other 10 articles in this supplementary issue and thus represents the contribution of all the authors who worked on these introductory articles. The content of this chapter is not exhaustive and represents the topics that the authors found to be the most important and compelling in a dynamic and changing field.

[Diagnosis and therapy of late onset Pompe disease]. / Diagnose und Therapie des Late-onset-Morbus-Pompe.

As Pompe disease glycogen storage disease type 2 with a severely reduced life expectancy is now a treatable disorder, accurate diagnostic procedures and evidence-based indications for therapy are mandatory. We screened the literature for consensus reports and published trial data of late-onset Pompe disease. These data were summarized in a Delphi consensus method approach. The clinical suspicion of late-onset Pompe disease should be substantiated by the validated dry blood spot test measurement for acid α-glucosidase activity. Alternatively, enzyme activity analysis in lymphocytes is also feasible. Glucosidase α gene sequencing for verifying the diagnosis is recommended. A muscle biopsy including measurements of acid α-glucosidase activity and glycogen concentration is warranted for differential diagnosis in selected cases. The confirmed diagnosis should lead to a multidisciplinary treatment approach, possibly including enzyme replacement therapy.

Coupling habitat-specific temperature scenarios with tolerance landscape to predict the impacts of climate change on farmed bivalves.

Due to climate change, heatwaves are likely to become more frequent, prolonged and characterized by higher peak values, compared with climatological averages. However, the thermal tolerance of organisms depends on the actual exposure, which can be modulated by environmental context and microhabitat characteristics. This study investigated the frequency of occurrence of mass mortality events in the next decades for two species of farmed bivalves, the mussel Mytilus galloprovincialis and the clam Ruditapes philippinarum, in a shallow coastal lagoon, characterised by marked diurnal oscillations of water temperature. The effect of heatwaves was estimated by means of tolerance landscape models, which predict the occurrence of 50% mortality based on the exposure intensity and duration. Scenarios of water temperature up to the year 2100 were modelled by combining two mechanistic components, namely: 1) monthly mean water temperatures, simulated using a hydrodynamic model including the heat budget; 2) daily oscillations, estimated from the harmonic analysis of a twenty year-long site-specific time series of water temperature. Scenarios of mean daily sediment temperature were estimated by means of a cross-correlation model, using as input the water temperature one: the model parameters were estimated based on a comprehensive set of site-specific water and sediment temperature observations. The results indicate that for both species the risk of mass mortality rapidly increases starting from the 2060s. Furthermore, the daily patterns of water temperature seemed to be relevant, as overnight it falls below the predicted mortality thresholds for a few hours. These findings suggest that further studies should address: 1) the improvement of tolerance landscape models, in order to take into account the integrated effect of repeated non-lethal stress events on mortality rate; 2) the prediction of environmental temperature in specific habitat, by means of both process-based and data driven models.

Generation of a human iPSC line (HIMRi001-A) from a patient with filaminopathy.

Here we introduce the human induced pluripotent stem cell (hiPSC) line HIMRi001-A generated from cultured dermal fibroblasts of a 60-year-old male patient with a myofibrillar myopathy, carrying a heterozygous c.4984C > T [p.Q1662X] mutation in the filamin C (FLNC)-gene, via lentiviral expression of OCT4, SOX2, KLF4 and c-MYC. HIMRi001-A displays typical embryonic stem cell-like morphology, carries the c.4984C > T FLNC gene mutation, expressed several pluripotent stem cell makers, retained normal karyotype (46, XY) and holds the potential to differentiate in all three germ layers. We postulate that HIMRi001-A can be used for the elucidation of FLNC-associated pathomechanisms and for developing new therapeutic options.

Estimating biofuel contaminant concentration from 4D ERT with mixing models.

We present the results of a lab-scaled feasibility study to assess the performance of electrical resistivity tomography for detection, characterization, and monitoring of fuel grade ethanol releases to the subsurface. Further, we attempt to determine the concentration distribution of the ethanol from the electrical resistivity tomography data using mixing-models. Ethanol is a renewable fuel source as well as an oxygenate fuel additive currently used to replace the known carcinogen methyl tert-butyl ether; however, ethanol is preferentially biodegraded and a cosolvent. When introduced to areas previously impacted by nonethanol-based fuels, it will facilitate the persistence of carcinogenic fuel compounds like benzene and ethylbenzene, as well as remobilize them to the ground water. These compounds would otherwise be retained in the soil column undergoing active or passive remediation processes such as soil vapor extraction or natural attenuation. Here, we introduce ethanol to a saturated Ottawa sand in a tank instrumented for four-dimensional geoelectrical measurements. Forward model results suggest pure phase ethanol released into a water saturated silica sand should present a detectable target for electrical resistivity tomography relative to a saturated silica sand only. We observe the introduction of ethanol to the closed hydraulic system and subsequent migration over the duration of the experiment. One-dimensional and three-dimensional temporal data are assessed for the detection, characterization, and monitoring of the ethanol release. Results suggest one-dimensional geoelectrical measurements may be useful for monitoring a release, while three-dimensional geoelectrical field imaging would be useful to characterize, monitor, and design effective remediation approaches for an ethanol release, assuming field conditions do not preclude the application of geoelectrical methods. We then attempt to use predictive mixing models to calculate the distribution of ethanol concentration within the measurement domain. For this study we examine four different models: a nested parallel mixing model, a nested cubic mixing model, the complex refractive index model (CRIM), and the Lichtenecker-Rother (L-R) model. The L-R model, modified to include an electrical formation factor geometry term, provided the best agreement with expected EtOH concentrations.

[Muscular dystrophy due to mutations in anoctamin 5: clinical and molecular genetic findings]. / Muskeldystrophie infolge Anoctamin-5-Mutationen : Klinische und molekulargenetische Befunde.

Recessive mutations in the anoctamin 5 (ANO5) gene have been recently identified in families with limb girdle muscular dystrophy (LGMD2L) and distal non-dysferlin Miyoshi myopathy. Anoctamin 5 is supposed to be a putative calcium-activated chloride channel. We report five German patients (four index patients) with muscle dystrophy due to mutations in the ANO5 gene. Sequencing of the ANO5 exons 5, 13 and 20 was performed to screen for a common c.191dupA mutation and two other reported mutations (c.1295C>G and p.R758C). The whole coding region of the ANO5 gene was sequenced to identify new mutations. Phenotypically, three patients showed LGMD and one patient Miyoshi type distal myopathy. One sibling had asymptomatic hyperCKemia. The age at onset was 64, 38 and 40 years in patients with LGMD and 23 years in the patient with distal myopathy. The four symptomatic patients showed remarkable asymmetric muscle involvement. There was marked CK elevation (11 to 30 times). Electron microscopy showed multifocal gaps in the sarcolemmal membrane. All patients harboured the common c.191dupA mutation in at least one allele. Two patients with LGMD were homozygous and the third patient and his asymptomatic sister were compound heterozygous for the c.191dupA mutation and a novel p.T548I mutation. The patient with distal myopathy harboured the p.R758C mutation in the second allele. Mutations in the ANO5 gene seem to be a relatively common cause of muscular dystrophy in Germany. Cases with late onset or asymptomatic hyperCKemia can occur. Clinically, asymmetric manifestation is typical.

An exploratory study of the association between reactive attachment disorder and attachment narratives in early school-age children.

OBJECTIVE: To explore attachment narratives in children diagnosed with reactive attachment disorder (RAD). METHOD: We compared attachment narratives, as measured by the Manchester Child Attachment Story Task, in a group of 33 children with a diagnosis of RAD and 37 comparison children. RESULTS: The relative risk (RR) for children with RAD having an insecure attachment pattern was 2.4 (1.4-4.2) but 30% were rated as securely attached. Within the RAD group, children with a clear history of maltreatment were more likely to be Insecure-Disorganised than children without a clear history of maltreatment. CONCLUSIONS: Reactive attachment disorder is not the same as attachment insecurity, and questions remain about how attachment research informs clinical research on attachment disorders.

Development and validation of the Axillary Sweating Daily Diary: a patient-reported outcome measure to assess axillary sweating severity.

BACKGROUND: Hyperhidrosis is estimated to affect ~ 4.8% of the US population, and most patients experience a negative psychological impact. Here, we describe development and psychometric evaluation of a patient-reported outcome (PRO) measure to assess severity of axillary hyperhidrosis in clinical trials that meets current U.S. regulatory standards to support product approvals. METHODS: Three rounds of hybrid concept-elicitation/cognitive-debriefing qualitative interviews were conducted in adults with clinician-diagnosed primary axillary hyperhidrosis, followed by similar interviews in children/adolescents. The draft measure included diary items for presence, severity, impact and bothersomeness (basis of the Axillary Sweating Daily Diary [ASDD]), exploratory weekly impact items, and a single-item Patient Global Impression of Change (PGIC). Phase 2 (adults only) and phase 3 (adults and children ≥9 years) clinical trial data were utilized to evaluate measurement properties of the resulting draft measure: floor/ceiling effects, nonresponse bias, test-retest reliability, construct validity, and responsiveness were assessed. The primary concept of interest was axillary sweating severity (ASDD Item 2); however, additional supportive concepts were explored to allow for development of a comprehensive hyperhidrosis measure. RESULTS: Twenty-nine patient interviews were conducted (N = 21 adult and N = 8 children/adolescents), resulting in the ASDD (4 items, patients ≥16y) and child-specific ASDD-C (2 items ≥9y to <16y), as well as 6 Weekly Impact items and the PGIC (patients ≥16y). No floor/ceiling effects or response biases were identified. Consistency between hypothesized and observed correlation patterns between ASDD/ASDD-C items and other efficacy measures supported construct validity. Intraclass correlation coefficients supported test-retest reliability (0.91-0.93; Item 2). Large effect sizes (- 2.2 to - 2.4) demonstrated that the ASDD/ASDD-C Item 2 could detect changes in hyperhidrosis severity, supporting the measure's responsiveness. Patients perceiving a moderate improvement in symptoms on the PGIC experienced an average 3.8-point improvement on ASDD axillary sweating severity (Item 2); thus, a 4-point responder threshold was defined as a clinically meaningful change. CONCLUSIONS: Qualitative and quantitative evidence support the reliability and validity of the ASDD/ASDD-C and its use in the clinical evaluation of axillary hyperhidrosis treatments. Further evaluation of this measure in future research studies is warranted to demonstrate consistent performance across different axillary hyperhidrosis populations and in different study contexts.

Imaging cortical dynamics at high spatial and temporal resolution with novel blue voltage-sensitive dyes.

Conventional imaging techniques have provided high-resolution imaging either in the spatial domain or in the temporal domain. Optical imaging utilizing voltage-sensitive dyes has long had the unrealized potential to achieve high resolution in both domains simultaneously, providing subcolumnar spatial detail with millisecond precision. Here, we present a series of developments in voltage-sensitive dyes and instrumentation that make functional imaging of cortical dynamics practical, in both anesthetized and awake behaving preparations, greatly facilitating exploration of the cortex. We illustrate this advance by analyzing the millisecond-by-millisecond emergence of orientation maps in cat visual cortex.

Molecular diagnosis of German patients with late-onset glycogen storage disease type II.

UNLABELLED: In patients with late-onset glycogen storage disease type II, one mutation, c.-32-13T>G, in the α-glucosidase (GAA) gene is identified frequently in European populations from different regions along with many rarer mutations. We have performed molecular genetic investigations in 18 German index patients with late-onset disease. The c.-32-13T>G, c.525delT (p.Glu176fsX45), and c.2481+102_2646+31del mutations were detected by PCR/restriction enzyme digest. Other mutations were detected by sequencing. All patients were compound heterozygous and 17 patients harboured the c.-32-13T>G mutation. Seven other previously described mutations (including the c.-32-13T>G) were identified, of which the p.C103G (c.307T>G) and the c.2481+102_2646+31del mutations were present each in three unrelated patients. Sequencing revealed five novel mutations. CONCLUSIONS: Genetic testing was able to identify the genetic defects in all patients and screening of the c.-32-13T>G mutation identified 94% of the cases. This is important for quick and reliable diagnosis, especially in view of enzyme replacement. Among the rarer mutations, c.2481+102_2646+31del and p.C103G are rather frequent in Germany.

Towards a sensorimotor aesthetics of performing art.

The field of neuroaesthetics attempts to identify the brain processes underlying aesthetic experience, including but not limited to beauty. Previous neuroaesthetic studies have focussed largely on paintings and music, while performing arts such as dance have been less studied. Nevertheless, increasing knowledge of the neural mechanisms that represent the bodies and actions of others, and which contribute to empathy, make a neuroaesthetics of dance timely. Here, we present the first neuroscientific study of aesthetic perception in the context of the performing arts. We investigated brain areas whose activity during passive viewing of dance stimuli was related to later, independent aesthetic evaluation of the same stimuli. Brain activity of six naïve male subjects was measured using fMRI, while they watched 24 dance movements, and performed an irrelevant task. In a later session, participants rated each movement along a set of established aesthetic dimensions. The ratings were used to identify brain regions that were more active when viewing moves that received high average ratings than moves that received low average ratings. This contrast revealed bilateral activity in the occipital cortices and in right premotor cortex. Our results suggest a possible role of visual and sensorimotor brain areas in an automatic aesthetic response to dance. This sensorimotor response may explain why dance is widely appreciated in so many human cultures.

Continual near-infrared spectroscopy monitoring in the injured lower limb and acute compartment syndrome: an FDA-IDE trial.

Aims: The aim of this study was to evaluate near-infrared spectroscopy (NIRS) as a continuous, non-invasive monitor for acute compartment syndrome (ACS). Patients and Methods: NIRS sensors were placed on 86 patients with, and 23 without (controls), severe leg injury. NIRS values were recorded for up to 48 hours. Longitudinal data were analyzed using summary and graphical methods, bivariate comparisons, and multivariable multilevel modelling. Results: Mean NIRS values in the anterior, lateral, superficial posterior, and deep posterior compartments were between 72% and 78% in injured legs, between 69% and 72% in uninjured legs, and between 71% and 73% in bilaterally uninjured legs. In patients without ACS, the values were typically > 3% higher in injured compartments. All seven limbs with ACS had at least one compartment where NIRS values were 3% or more below a reference uninjured control compartment. Missing data were encountered in many instances. Conclusion: NIRS oximetry might be used to aid the assessment and management of patients with ACS. Sustained hyperaemia is consistent with the absence of ACS in injured legs. Loss of the hyperaemic differential warrants heightened surveillance. NIRS values in at least one injured compartment(s) were > 3% below the uninjured contralateral compartment(s) in all seven patients with ACS. Additional interventional studies are required to validate the use of NIRS for ACS monitoring. Cite this article: Bone Joint J 2018;100-B:787-97.

Analysis of spectrum and frequencies of mutations in McArdle disease. Identification of 13 novel mutations.

BACKGROUND: McArdle disease, a common metabolic myopathy with autosomal recessive inheritance, is caused by a frequent R50X mutation and many rare mutations in the myophosphorylase gene. OBJECTIVES: To identify spectrum and frequencies of myophosphorylase gene mutations in a large cohort of patients with McArdle disease, to discuss diagnostic implications, and to analyse genotype-phenotype relationship. METHODS: Molecular genetic analysis of 56 index patients with muscle biopsy-proven myophosphorylase deficiency from Germany (n = 35), UK (n = 13), and several other countries (n = 8) was performed using direct sequencing. RESULTS: Allele frequency of the R50X mutation was 58%, and 71% of the patients carried this mutation at least on one allele. We detected 26 other less common mutations, 13 of which are novel: G157V, R161C, Q337R, E384K, S450L, G486D, R570W, K575E, IVS6-2A>T, IVS10+1G>A, R650X, c.1354insC, c.1155_1156delGG. There was no genotype-phenotype correlation with respect to age of onset and severity. R270X was the most frequent mutation among the less common mutations reaching an allele frequency of 5% followed by R94W and G686R representing a frequency of 4% each. CONCLUSIONS: The study further extends the genetic heterogeneity of myophosphorylase gene mutations showing no mutational hotspot and no genotype-phenotype correlation. Most novel missense mutations were located in secondary structures or active sites of the enzyme. Some of the less common mutations are recurrent with different frequencies within Europe. Ethnic origin and frequency of less common mutations must be considered to establish efficient strategies in molecular genetic testing. Performing molecular testing can avoid muscle biopsy.