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Objective Next generation sequencing (NGS) has expanded the diagnostic paradigm turning the focus to the growth plate. The aim of the study was to determine the prevalence of variants in genes implicated in skeletal dysplasias in probands with short stature and mild skeletal anomalies. Design Clinical and radiological data were collected from 108 probands with short stature and mild skeletal anomalies. Methods A customized skeletal dysplasia NGS panel was performed. Variants were classified using ACMG recommendations and Sherloc. Anthropometric measurements and skeletal anomalies were subsequently compared in those with or without an identified genetic defect. Results Heterozygous variants were identified in 21/108 probands (19.4%). Variants were most frequently identified in ACAN (n = 10) and IHH (n = 7) whilst one variant was detected in COL2A1, CREBBP, EXT1, and PTPN11. Statistically significant differences (P < 0.05) were observed for sitting height/height (SH/H) ratio, SH/H ratio standard deviation score (SDS), and the SH/H ratio SDS >1 in those with an identified variant compared to those without. Conclusions A molecular defect was elucidated in a fifth of patients. Thus, the prevalence of mild forms of skeletal dysplasias is relatively high in individuals with short stature and mild skeletal anomalies, with variants in ACAN and IHH accounting for 81% of the cases. An elevated SH/H ratio appears to be associated with a greater probability in detecting a variant, but no other clinical or radiological feature has been found determinant to finding a genetic cause. Currently, we cannot perform extensive molecular studies in all short stature individuals so detailed clinical and radiological phenotyping may orientate which are the candidate patients to obtain worthwhile results. In addition, detailed phenotyping of probands and family members will often aid variant classification.

Spondyloepiphyseal dysplasia type Stanescu (SED-S) is a very rare type II collagenopathy. We describe an 8-year-old boy who presented with short trunk, C2-C3 vertebral fusion, hand, foot, leg and thigh pain, stiffness and limited joint mobility, and waddling gait. Radiographs showed platyspondyly with anterior wedging and endplate irregularities, broad femoral necks, and large epiphyses and epiphyseal equivalents. Differential diagnosis included progressive pseudorheumatoid dysplasia and SED-S. A skeletal dysplasia custom-designed NGS panel was performed and the heterozygous pathogenic variant c.620G>A; p.(Gly207Glu) in COL2A1 was detected, establishing the diagnosis of SED-S. Vertebral fusions, observed in our patient, have not been previously described in this dysplasia. This variant has not been previously associated with SED-S, but was reported in two other families with spondyloepiphyseal dysplasia. Thus, this case expands the clinical and mutational spectrum of SED-S and demonstrates that SED-S significantly overlaps with other skeletal dysplasias.

Hand hyperphalangism leading to shortened index fingers with ulnar deviation, hallux valgus, mild facial dysmorphism and respiratory compromise requiring assisted ventilation are the key features of Chitayat syndrome. This condition results from the recurrent heterozygous missense variant NM_006494.2:c.266A>G; p.(Tyr89Cys) in ERF on chromosome 19q13.2, encoding the ETS2 repressor factor (ERF) protein. The pathomechanism of Chitayat syndrome is unknown. To date, seven individuals with Chitayat syndrome and the recurrent pathogenic ERF variant have been reported in the literature. Here, we describe six additional individuals, among them only one presenting with a history of assisted ventilation, and the remaining presenting with variable pulmonary phenotypes, including one individual without any obvious pulmonary manifestations. Our findings widen the phenotype spectrum caused by the recurrent pathogenic variant in ERF, underline Chitayat syndrome as a cause of isolated skeletal malformations and therefore contribute to the improvement of diagnostic strategies in individuals with hand hyperphalangism.

Mesoaxial synostotic syndactyly with phalangeal reduction (MSSD) is an extremely rare autosomal recessive limb abnormality characterized by the fusion of third and fourth fingers. To date, only homozygous missense and frameshift mutations have been reported in BHLHA9 associated to MSSD. In this study, we report a patient who presented with clinical and radiological features of MSSD. A customized skeletal dysplasia NGS panel revealed the presence of two novel compounds heterozygous variants in BHLHA9: NM_001164405.1: c.[226A>T][269G>C]; p.[(Lys76*)][(Arg90Pro)]. Thus, this is the first case of MSSD in a nonconsanguineous family.

Hypochondroplasia (HCH), a skeletal dysplasia caused by mutations in the fibroblast growth factor receptor 3 (FGFR3) gene, is characterized by disproportionate short stature. The p.Asn540Lys (p.N540K) mutation accounts for ∼50 to 70% of cases of HCH, but novel FGFR3 mutations are described. We present a family with disproportionately short stature and mild radiologic findings seen in a major public pediatric hospital in Argentina. A previously undescribed heterozygous missense variant in FGFR3, NM_000142.4:667C > T; p.(Arg223Cys) was identified. The predicted phenotype correlates well with the mild auxologic and radiologic characteristics observed. In this case, disproportionately short stature raised the suspicion of skeletal dysplasia.

Background Evidence suggests that GABA may reduce pancreatic inflammation, protect β-cells from autoimmune destruction, and potentiate the regeneration of new β-cells in the setting of type 1 diabetes mellitus (T1DM). The enzyme GAD, also expressed in human pancreatic β-cells, is an antigenic target of reactive T cells. We hypothesized that treatment of children with recent onset T1DM with GABA or combination GABA with GAD will preserve β-cell function and ameliorate autoimmune dysregulation. Methods This is a one-year, prospective, randomized, double-blind, placebo-controlled trial. Ninety-nine patients aged 4–18 years with newly diagnosed T1DM are randomized into three treatment groups: 1) oral GABA twice daily in addition to two injections of recombinant GAD enzyme, 2) oral GABA plus placebo GAD injections, or 3) placebo GABA and placebo GAD. Patients are evaluated at baseline and months 1, 5, 8 and 12. Mixed meal tolerance testing is performed at all but the 8-month visit. Laboratory studies will assess indices of beta and alpha cell function, glycemic control, immunophenotyping, and diabetes-related autoantibodies. Results The primary outcome is the effect on pancreatic β-cell function as measured by meal-stimulated c-peptide secretion compared between the treatment groups before and after one year of treatment. Secondary outcomes include: 1) fasting and meal stimulated glucagon and proinsulin levels, 2) response in insulin usage by participants, 3) indices of immune cell function, and 4) effect on autoantibodies GAD65, ICA512, and ZnT8. Conclusions: This trial will determine the safety and efficacy of GABA and combination GABA/GAD therapy to delay T1DM progression in children.

A cohort study on the growth of 19 Argentinean children, aged 0 to 18 years, and 11 of their first-degree relatives with alterations in the SHOX gene or its regulatory regions is reported. Children are born shorter and experience a growth delay during childhood with a stunted pubertal growth spurt. Body disproportion, with a sitting height/height ratio above +2 standard deviation score (SDS), was already present as early as 2 years old. Hand length was normal. Shortening of the radius, with a length below –1.9 SDS, was the earliest and most frequent radiological sign detected as early as 45 days old. We found a previously unreported mutation in a family with a highly variable phenotype, the boy had a severe phenotype with a milder presentation in other affected members of the family. We conclude that body disproportion and a shorter radius length on X-ray are useful tools for selecting children to undergo SHOX molecular studies.

Acoustic indices derived from environmental soundscape recordings are being used to monitor ecosystem health and vocal animal biodiversity. Soundscape data can quickly become very expensive and difficult to manage, so data compression or temporal down‐sampling are sometimes employed to reduce data storage and transmission costs. These parameters vary widely between experiments, with the consequences of this variation remaining mostly unknown.We analyse field recordings from North‐Eastern Borneo across a gradient of historical land use. We quantify the impact of experimental parameters (MP3 compression, recording length and temporal subsetting) on soundscape descriptors (Analytical Indices and a convolutional neural net derived AudioSet Fingerprint). Both descriptor types were tested for their robustness to parameter alteration and their usability in a soundscape classification task.We find that compression and recording length both drive considerable variation in calculated index values. However, we find that the effects of this variation and temporal subsetting on the performance of classification models is minor: performance is much more strongly determined by acoustic index choice, with Audioset fingerprinting offering substantially greater (12%–16%) levels of classifier accuracy, precision and recall.We advise using the AudioSet Fingerprint in soundscape analysis, finding superior and consistent performance even on small pools of data. If data storage is a bottleneck to a study, we recommend Variable Bit Rate encoded compression (quality = 0) to reduce file size to 23% file size without affecting most Analytical Index values. The AudioSet Fingerprint can be compressed further to a Constant Bit Rate encoding of 64 kb/s (8% file size) without any detectable effect. These recommendations allow the efficient use of restricted data storage whilst permitting comparability of results between different studies., Soundscapes were recorded from different forest structures in Malaysian Borneo. Data collection variation was simulated, and all data groups were analyzed via usual acoustic indices and a CNN‐derived AudioSet Fingerprint. The effect of variation in data collection was compared between the two types of soundscape descriptor, finding the AudioSet Fingerprint to be a stronger and more robust descriptor of soundscapes.

Metaphyseal anadysplasia is a very rare hereditary skeletal dysplasia with onset occurring normally during the second and third years of life, but unlike many other dysplasias, symptoms appear to resolve by adolescence. Two types exist, the more severe form, type 1, with both autosomal dominant and recessive inheritance due to pathogenic variants in MMP13, whilst type 2, an even rarer form is due to biallelic MMP9 variants. To date, only two metaphyseal anadysplasia type 2 families have been reported. We describe a third family, a young boy, born to consanguineous parents, referred at 19 months old for abnormal gait due to bowed legs. Clinical and radiological examination revealed scoliosis, genu varum and metaphyseal abnormalities. A homozygous MMP9 nonsense variant, NM_004994.2:c.1764G>A; p.(Trp588*) was identified. By the age of 39 months, lower limb alignment and metaphyseal features had already significantly improved and scoliosis had disappeared. This case confirms that biallelic MMP9 variants cause this very rare skeletal dysplasia, metaphyseal anadysplasia type 2 but also shows that the skeletal manifestations can improve within a short period time and at an early age.

Osteoporosis should be considered in children with severe chronic diseases or in association with some genetic diseases that bear an increased risk of bone fragility. Primary osteoporosis is an entity in which emerging aetiologies are being recognized. Its association with congenital retinal folds should guide the diagnosis to the Osteoporosis-Pseudoglioma syndrome (OMIM 259770), a rare disease (prevalence of 1/2 000 000), caused by the loss of function of the protein LRP5 (low-density lipoprotein receptor-related protein 5) resulting in the alteration of the Wnt/β-catenin signalling pathway. We report the case of a child with congenital retinal folds, progressive loss of vision and multiple fractures whose clinical, biochemical and genetic studies confirmed the diagnosis of primary osteoporosis due to a novel homozygous inactivating variant in LRP5.

There is concern that climate change will lead to expansion of vector-borne diseases as, of all disease types, they are the most sensitive to climate drivers1. Such expansion may threaten human health, and food security via effects on animal and crop health. Here we quantify the potential impact of climate change on a vector-borne disease of livestock, bluetongue, which has emerged in northern Europe in response to climate change2–4, affecting tens of thousands of farms at huge financial cost and causing the deaths of millions of animals5. We derive future disease risk trends for northern Europe, and use a detailed spatial transmission model6 to simulate outbreaks in England and Wales under future climatic conditions, using an ensemble of five downscaled general circulation models7. By 2100, bluetongue risk extends further north, the transmission season lengthens by up to three months and outbreaks are larger on average. A 1 in 20-year outbreak at present-day temperatures becomes typical by the 2070s under the highest greenhouse gas emission scenario. However, animal movement restrictions are sufficient to prevent truly devastating outbreaks. Disease transmission uncertainty dominates over climate uncertainty, even at the longest prediction timescales. Our results suggest that efficient detection and control measures to limit the spread of vector-borne diseases will be increasingly vital in future, warmer climates. Bluetongue risk to livestock across northern Europe is projected to extend further north, with a longer transmission season and larger outbreaks on average. As a result, disease detection and control measures will be increasingly important.

We evaluated psychometric properties (validity, reliability, and responsiveness) of a modified Faces Pain Scale-Revised (FPS-R) in 257 patients with sickle cell anemia (SCA) 7 to below 18 years old in a randomized, multinational clinical study. The modified FPS-R asks patients to report, by daily diary, their worst intraday SCA-related pain. Intraclass correlation coefficient assessed test-retest reliability between month 1 and month 2. Pearson correlations between monthly mean SCA-related pain intensity, activity interference score, analgesic use, and opioid use assessed convergent validity. Responsiveness was assessed with correlations of changes of monthly pain rate or intensity and changes in analgesic use or activity interference score from month 1 to month 9. Intraclass correlation coefficients for pain intensity and pain rate were 0.777 and 0.820, respectively, indicating agreement among stable patients. Moderate associations were shown between mean pain intensity and analgesic use (r=0.39) and opioid use (r=0.44), and between monthly pain rate and analgesic use (r=0.38). Moderate-to-large associations were observed between change in mean pain rate or intensity and changes in analgesic use (r=0.38 to 0.39, both P

Resumen Introduccion y objetivos La displasia valvular cardiaca ligada al cromosoma X es una cardiopatia congenita rara especifica del sexo masculino y caracterizada principalmente por una degeneracion mixomatosa de las valvulas auriculoventriculares con consecuencias hemodinamicas variables. Se debe a defectos geneticos en la filamina A (codificada por FLNA), una proteina de union a actina de expresion ubicua que regula la organizacion del citoesqueleto. La perdida de funcion de la filamina A tambien se ha asociado con manifestaciones neurologicas y del tejido conectivo a menudo simultaneas, y aparentemente las mutaciones en la primera mitad del dominio Rod 1 expresan el fenotipo cardiaco completo. En esta familia de nueva descripcion, se ha contribuido a las correlaciones genotipo-fenotipo previas con un enfoque multidisciplinario. Metodos La evaluacion cardiologica, dismorfologica y genetica de los miembros disponibles se complemento con estudios de la transcripcion y de la inactivacion del cromosoma X. Resultados La nueva mutacion de FLNA c.1066-3C>G cosegregaba con un fenotipo cardiaco aparentemente aislado y expresado en los varones, sin que hubiera un sesgo en el patron de inactivacion del cromosoma X en las mujeres portadoras. Esta variante resulto en una delecion dentro del marco de lectura de 8 residuos de aminoacidos cercanos a la region N-terminal de la proteina. Conclusiones La perdida de funcion parcial y no sometida a impronta del dominio Rod 1 proximal de la filamina A parece ser el mecanismo patogenico de la displasia valvular cardiaca, expresada en algunos casos con manifestaciones extracardiacas.

Introduction and objectives X-linked cardiac valvular dysplasia is a rare form of male-specific congenital heart defect mainly characterized by myxomatous degeneration of the atrioventricular valves with variable hemodynamic consequences. It is caused by genetic defects in FLNA-encoded filamin A, a widely expressed actin-binding protein that regulates cytoskeleton organization. Filamin A loss of function has also been associated with often concurring neurologic and connective tissue manifestations, with mutations in the first half of the Rod 1 domain apparently expressing the full cardiac phenotype. We contribute to previous genotype-phenotype correlations with a multidisciplinary approach in a newly-described family. Methods Cardiologic, dysmorphologic, and genetic evaluation of available members were complemented with transcriptional and X-chromosome inactivation studies. Results A novel FLNA mutation c.1066-3C>G cosegregated with a male-expressed, apparently isolated, cardiac phenotype with no skewed X-inactivation pattern in female carriers. This variant was shown to result in an in-frame deletion of 8 amino acid residues near the N-terminal region of the protein. Conclusions A nonimprinted, partial loss of function of filamin A proximal Rod 1 domain seems to be the pathogenetic mechanism of cardiac valvular dysplasia, with some cases occasionally expressing associated extracardiac manifestations.

OBJECTIVE: Mutations in the aggrecan gene (ACAN) have been identified in two autosomal dominant skeletal dysplasias, spondyloepiphyseal dysplasia, Kimberley type (SEDK), and osteochondritis dissecans, as well as in a severe recessive dysplasia, spondyloepimetaphyseal dysplasia, aggrecan type. Next-generation sequencing (NGS) has aided the identification of heterozygous ACAN mutations in individuals with short stature, minor skeletal defects and mild facial dysmorphisms, some of whom have advanced bone age (BA), poor pubertal spurt and early growth cessation as well as precocious osteoarthritis. DESIGN AND METHODS: This study involves clinical and genetic characterization of 16 probands with heterozygous ACAN variants, 14 with short stature and mild skeletal defects (group 1) and two with SEDK (group 2). Subsequently, we reviewed the literature to determine the frequency of the different clinical characteristics in ACAN-positive individuals. RESULTS: A total of 16 ACAN variants were located throughout the gene, six pathogenic mutations and 10 variants of unknown significance (VUS). Interestingly, brachydactyly was observed in all probands. Probands from group 1 with a pathogenic mutation tended to be shorter, and 60% had an advanced BA compared to 0% in those with a VUS. A higher incidence of coxa valga was observed in individuals with a VUS (37% vs 0%). Nevertheless, other features were present at similar frequencies. CONCLUSIONS: ACAN should be considered as a candidate gene in patients with short stature and minor skeletal defects, particularly those with brachydactyly, and in patients with spondyloepiphyseal dysplasia. It is also important to note that advanced BA and osteoarticular complications are not obligatory conditions for aggrecanopathies/aggrecan-associated dysplasias. info:eu-repo/semantics/publishedVersion

Marine renewable energy devices require mooring and foundation systems that suitable in terms of device operation and are also robust and cost effective. In the initial stages of mooring and founda...

Tissue engineering and regenerative medicine (TERM) is a field that applies biology and engineering principles to "restore, maintain or repair a tissue after injury". Besides the potential to treat various diseases, these endeavours increase our understanding of fundamental cell biology. Although TERM has progressed rapidly, engineering a whole organ is still beyond our skills, primarily due to the complexity of tissues. Material science and current manufacturing methods are not capable of mimicking this complexity. Therefore, many researchers explore the use of naturally derived materials that maintain important biochemical, structural and mechanical properties of tissues. Consequently, employing non-cellular components of tissues, particularly the extracellular matrix, has emerged as an alternative to synthetic materials. Because of their complexity, decellularized tissues are not as well defined as synthetic materials but they provide cells with a microenvironment that resembles their natural niche. Decellularized tissues are produced from a variety of sources, among which the fetal membranes are excellent candidates since their supply is virtually unlimited, they are readily accessible with minimum ethical concerns and are often discarded as a biological waste. In this review, we will discuss various applications of decellularized fetal membranes as substrates for the expansion of stem cells, their use as two and three-dimensional scaffolds for tissue regeneration, and their use as cell delivery systems. We conclude that fetal membranes have great potential for use in TERM.

Purpose Short stature is a common condition of great concern to patients and their families. Mostly genetic in origin, the underlying cause often remains elusive due to clinical and genetic heterogeneity. Methods We systematically phenotyped 565 patients where common nongenetic causes of short stature were excluded, selected 200 representative patients for whole-exome sequencing, and analyzed the identified variants for pathogenicity and the affected genes regarding their functional relevance for growth. Results By standard targeted diagnostic and phenotype assessment, we identified a known disease cause in only 13.6% of the 565 patients. Whole-exome sequencing in 200 patients identified additional mutations in known short-stature genes in 16.5% of these patients who manifested only part of the symptomatology. In 15.5% of the 200 patients our findings were of significant clinical relevance. Heterozygous carriers of recessive skeletal dysplasia alleles represented 3.5% of the cases. Conclusion A combined approach of systematic phenotyping, targeted genetic testing, and whole-exome sequencing allows the identification of the underlying cause of short stature in at least 33% of cases, enabling physicians to improve diagnosis, treatment, and genetic counseling. Exome sequencing significantly increases the diagnostic yield and consequently care in patients with short stature.

Background/Aims: Patients with sickle cell anemia can experience recurrent pain episodes, which affect quality of life. The reported prevalence of pain is higher in studies using patient diaries than in healthcare facility utilization data. Determining Effects of Platelet Inhibition on Vaso-Occlusive Events was a multinational study that assessed the efficacy and safety of prasugrel in reducing the rate of vaso-occlusive events in children with sickle cell anemia (NCT01794000) and included an electronic patient-reported outcome diary to record pain occurrence. We aimed to capture diary completion rates and compliance in children who used the electronic patient-reported outcome diary during the Determining Effects of Platelet Inhibition on Vaso-Occlusive Events study and examine factors contributing to diary completion rates and compliance. Methods: Daily electronic patient-reported outcome diary data were collected for up to 9 months in Determining Effects of Platelet Inhibition on Vaso-Occlusive Events participants aged 4 to Results: A total of 311 participants received a diary; 268 provided diary data through Month 9. Diary completion rates and compliance were high throughout the collection period and across all groups and regions, despite no games being included on the device. For subjective data, the overall completion rate was 94.4%, and 92.6% of participants were compliant. For objective data, the overall completion rate was 93.3%, and 89.7% of participants were compliant. Completion rates and compliance differed significantly by age and region and were higher for 4 to Conclusion: With appropriate design, participant training, and sufficient monitoring, an electronic patient-reported outcome diary can capture daily sickle cell–related pain data in large multinational studies. Providing a mechanism for caregiver reporting is particularly valuable for participants

Summary Cryptorchidism is the most common congenital disorder in boys, but the cause for most cases remains unknown. Patients with Noonan Syndrome are characterized by a typical face, growth retardation, congenital heart defects, learning disabilities and cryptorchidism. Copy number variations of Ras/MAPK pathway genes are unusual in patients with several clinical features of Noonan Syndrome; however, they have not been studied in patients with only one feature of this condition, such as cryptorchidism. Our aim was to determine whether patients with isolated cryptorchidism exhibit Ras/MAPK pathway gene copy number variations (CNVs). Fifty-nine patients with isolated cryptorchidism and negative for mutations in genes associated with Noonan Syndrome were recruited. Determination of Ras/MAPK pathway gene CNVs was performed by Comparative Genome Hybridization array. A CNV was identified in two individuals, a ~175 kb microduplication at 3p25.2, partially including RAF1. A similar RAF1 microduplication has been observed in a patient with testicular aplasia. This suggests that some patients with isolated cryptorchidism may harbor Ras/MAPK pathway gene CNVs.

For Marine Renewable Energy (MRE) to become a viable alternative energy source, it must encompass large arrays of devices. Arrays may include 1000s of devices. The associated foundations or anchors may encounter a range of seafloor sediment types and geotechnical properties. Wave and tidal energy convertors induce unique loads on foundations and anchors that are different from other seafloor engineering applications. Thus, there is a need for a combination of advanced site analysis and performance assessment. Geotechnical engineering plays the vital role of ensuring that foundation and anchor systems perform successfully for MRE devices. Our paper reviews the unique frequency and magnitude of loading regimes experienced by MRE arrays. We examine potential loading conditions on the foundation-anchor systems. Loading regimes include environmental and system loads from single devices or arrays of devices. We present specific load examples from field data. We explore the applicable geotechnical approaches to address these conditions, including constitutive models that may or may not adequately capture the response of the seafloor sediments to the MRE loads. Partially to fully dynamic constitutive model formulations may be necessary to properly model sediment-fluid hydromechanical response to MRE loading. Spacing of full MRE arrays and spatial variability in sediment properties may require multiple foundation types.

Hypophosphatasia (HPP) is an inborn error of metabolism characterized by low levels of serum alkaline phosphatase (ALP). Scarce evidence exists about features that should signal the potential association between hypophosphatasaemia and HPP in adults. The aim of this study is to estimate the prevalence of ALPL variants in subjects with persistent hypophosphatasaemia and determine the associated clinical and laboratory features. For this cross-sectional study, laboratory records of 386,353 subjects were screened by measurement of ALP activity. A total of 85 (0.18%) subjects with persistent hypophosphatasaemia (≥2 serum alkaline phosphatase-ALP-measurements ≤35 IU/L and none > 45 IU/L) were included (secondary causes previously discarded). ALPL genetic testing and a systematized questionnaire to retrieve demographic, clinical and laboratory data were performed. Descriptive analysis and logistic regression models were employed to identify the clinical and laboratory characteristics associated with ALPL variants. Results: Forty subjects (47%) had a variant(s) in ALPL. With regard to clinical characteristics, the presence of an ALPL variant was significantly associated only with musculoskeletal pain (OR: 7.6; 95% IC: 1.9-30.9). Nevertheless, a trend to present more dental abnormalities (OR: 3.6; 95% IC: 0.9-13.4) was observed. Metatarsal stress fractures were also more frequent (4 vs 0; p < 0.05) in this group. Regarding laboratory features, median ALP levels were lower in subjects with ALPL variants (26 vs 29 IU/L; p < 0.005). Interestingly, the threshold of ALP levels < 25 IU/L showed a specificity, positive predictive value and positive likelihood ratio of 97.8, 94.4% and 19.8 to detect a positive ALPL test, respectively. Conclusions: In subjects with persistent hypophosphatasaemia -secondary causes excluded- one out of two presented ALPL variants. Musculoskeletal pain and ALP levels < 25 IU/L are associated with this variant(s). In this scenario, ALP levels < 25 IU/L seem to be very useful to identify individuals with the presence of an ALPL variant, Genetic testing was supported by a grant from Alexion Pharmaceuticals Inc., which had no role in the study design or data analysis

Objective To assess the diagnostic accuracy of prenatal ultrasound for detecting fetal skeletal dysplasias and to describe its role in orienting genetic studies. Study design Observational study of pregnant women surveyed in our hospital, between 2011 and 2018, with fetal long bones below the 3rd centile (shortened long bones), either as an isolated finding or associated to other skeletal anomalies. We used a systematic protocol for the ultrasound evaluation and selection of those fetuses suspected of having a skeletal dysplasia. We report the demographics of these patients along with the sonographic follow-up of their fetuses, the genetic results and the outcome of the pregnancies and the newborn in the entire group and also compare data between the two sub-groups (isolated shortened long bones vs shortened long bones associated to other anomalies). Results A total of 81 pregnancies with a suspected fetal skeletal dysplasia were included, with a complete follow-up available in 75 cases, 22 with isolated shortened long bones and 53 cases that presented shortened long bones with other skeletal anomalies. In the shortened long bones sub-group, a total of five (23 %) were born healthy neonates, 10 (45 %) were small for gestational age or intrauterine growth restricted (one of them of genetic origin) and seven (32 %) had a skeletal dysplasia (6 of them with genetic diagnosis). Whilst among the 53 cases that presented with shortened long bones + other skeletal anomalies, three (6%) were healthy neonates, five (9%) were small for gestational age/intrauterine growth restricted (two of genetic origin) and 45 (85 %) had a skeletal dysplasia (19 genetically confirmed and 26 with a clinical diagnosis). These differences in frequencies between the two sub-groups were determined to be statistically significant (χ2: p = 0.02). Conclusion Around one third of fetuses with isolated shortened long bones will have a skeletal dysplasia. If abnormal skeletal ultrasound findings are associated with shortened long bones, the risk for skeletal dysplasia is significantly increased (85 %). Prenatal systematic approach in a multidisciplinary unit is useful in the orientation of genetic studies.

Background: DOVE (Determining Effects of Platelet Inhibition on Vaso-Occlusive Events) was a Phase 3, randomized, double-blind, placebo-controlled study conducted in children with sickle cell anemia at 51 sites in 13 countries across four continents. Procedure: Data from DOVE were assessed for regional differences in subject phenotype and treatment. Demographics, baseline clinical and laboratory data, hydroxyurea (HU) use, vaso-occlusive crisis (VOCs, composite endpoint of painful crisis or acute chest syndrome (ACS, Beijing, China)), serious adverse events (SAEs, Florence, Italy), hospitalization, and treatments were compared across the Americas, Europe, North Africa/Middle East, and Sub-Saharan Africa (SSA). Results: Race, body mass index, and blood pressures differed by region. Pre-enrollment VOCs were highest in the Americas. For subjects not on HU, baseline hemoglobin was lowest in SSA, reticulocyte count was lowest in the Americas. Within SSA, Kenya subjects presented higher baseline hemolysis. Painful crisis was the most common SAE, followed by ACS in the Americas and infections in other regions. VOC rate and percentage of VOC hospitalizations were highest in Europe. Regardless of region, most VOCs were treated with analgesics, approximately half were treated with intravenous fluids. The proportion of VOC-related transfusions was greatest in Europe. Lengths of hospital stay were similar across regions. Conclusions: Overall differences in SAEs and hospitalization for VOCs may be due to cultural diversities, resource utilization, disease severity, or a combination of factors. These data are of importance for the planning of future trials in SCA in a multinational setting.

Hypophosphatasia (HPP) is a rare autosomal dominant or recessive metabolic disorder caused by mutations in the tissue nonspecific alkaline phosphatase gene (ALPL). To date, over 300 different mutations in ALPL have been identified. Disease severity is widely variable with severe forms usually manifesting during perinatal and/or infantile periods while mild forms are sometimes only diagnosed in adulthood or remain undiagnosed. Common clinical features of HPP are defects in bone and tooth mineralization along with the biochemical hallmark of decreased serum alkaline phosphatase activity. The incidence of severe HPP is approximately 1 in 300,000 in Europe and 1 in 100,000 in Canada. We present the clinical and molecular findings of 83 probands and 28 family members, referred for genetic analysis due to a clinical and biochemical suspicion of HPP. Patient referrals included those with isolated low alkaline phosphatase levels and without any additional clinical features, to those with a severe skeletal dysplasia. Thirty-six (43.3%) probands were found to have pathogenic ALPL mutations. Eleven previously unreported mutations were identified, thus adding to the ever increasing list of ALPL mutations. Seven of these eleven were inherited in an autosomal dominant manner while the remaining four were observed in the homozygous state. Thus, this study includes a large number of well-characterized patients with hypophosphatasemia which has permitted us to study the genotype:phenotype correlation. Accurate diagnosis of patients with a clinical suspicion of HPP is crucial as not only is the disease life-threatening but the patients may be offered bone targeted enzymatic replacement therapy. © 2017 Wiley Periodicals, Inc.

Interstitial deletions in the short arm of chromosome 1 are infrequent. We report a female with a 1p31.1p31.3 deletion and cloverleaf skull, who presented with renal and central nervous system malformations, cleft palate, severe ocular anomalies, and cutis laxa, in addition to the previously described clinical data present in other cases with deletions encompassing this region, such as developmental delay, seizures, round face with a prominent nose, micro/retrognathia, half-opened mouth, short neck, hand/foot malformations, hernia, congenital heart malformations, and abnormal external genitalia. The deletion spanned ∼18.6 Mb and included a total of 68 OMIM protein coding genes. We have reviewed 17 cases previously described in the literature and in DECIPHER involving the chromosomal region 1p31.1p31.3. Only 3 of these affect the whole region, 9 are partial deletions of this region, and 5 are much smaller deletions. Taking into account the MORBID ID and the haploinsufficiency score of the genes, we go on to propose which genes may explain particular clinical features observed in the patient. IL23R may be responsible for the craniosynostosis, FOXD2 for the renal anomalies, LHX8 for closure defects of the palate, and ST6GALNAC3 for skin anomalies. In summary, we have identified a chromosome 1p31.1p31.3 deletion in a patient with an atypical presentation of craniosynostosis amongst other more typical features observed in individuals with similar deletions.

The Mount Simon Sandstone and Eau Claire Formation represent a potential reservoir-caprock system for wastewater disposal, geologic CO 2 storage, and compressed air energy storage (CAES) in the Midwestern United States. A primary concern to site performance is heterogeneity in rock properties that could lead to nonideal injectivity and distribution of injected fluids (e.g., poor sweep efficiency). Using core samples from the Dallas Center domal structure, Iowa, we investigate pore characteristics that govern flow properties of ­major lithofacies of these formations. Methods include gas porosimetry and perme­ametry, mercury intrusion porosimetry, thin section petrography, and X-ray diffraction. The lithofacies exhibit highly variable intraformational and interformational distributions of pore throat and body sizes. Based on pore-throat size, there are four distinct sample groups. Micropore-throat–dominated samples are from the Eau Claire Formation, whereas the macropore-dominated, mesopore-dominated, and uniform-dominated samples are from the Mount Simon Sandstone. Complex paragenesis governs the high degree of pore and pore-throat size hetero­geneity, due to an interplay of precipitation, non­uniform compaction, and later dissolution of cements. The cement dissolution event probably accounts for much of the current porosity in the unit. Mercury intrusion porosimetry data demonstrate that the heterogeneous nature of the pore networks in the Mount Simon Sandstone results in a greater than normal opportunity for reservoir capillary trapping of nonwetting fluids, as quantified by CO 2 and air column heights that vary over three orders of magnitude, which should be taken into account when assessing the potential of the reservoir-caprock system for waste disposal (CO 2 or produced water) and resource storage (natural gas and compressed air). Our study quantitatively demonstrates the significant impact of millimeter-scale to micron-scale porosity heterogeneity on flow and transport in reservoir sandstones.

Progressive pseudorheumatoid dysplasia (PPD) is a rare autosomal recessive disorder characterized by spondyloepiphyseal dysplasia associated with pain and stiffness of multiple joints, enlargement of the interphalangeal joints, normal inflammatory parameters, and absence of extra-skeletal manifestations. Homozygous or compound heterozygous WISP3 mutations cause PPD. We report two siblings from a non-consanguineous Ecuadorian family with a late-onset spondyloepiphyseal dysplasia. Mutation screening was undertaken in the two affected siblings using a customized skeletal dysplasia next generation sequencing (NGS) panel and confirmed by Sanger sequencing. Two compound heterozygous mutations were identified in WISP3 exon 2, c.[190G>A];[197G>A] (p.[(Gly64Arg)];[(Ser66Asn)]) in the two siblings, both of which had been inherited. The p. (Gly64Arg) mutation has not been previously described whilst the p. (Ser66Asn) mutation has been reported in two PPD families. The two siblings presented with atypical PPD, as they presented during late childhood, yet the severity was different between them. The progression was particularly aggressive in the male sibling who suffered severe scoliosis by the age of 13 years. This case reaffirms the clinical heterogeneity of this disorder and the clinical utility of NGS to genetically diagnose skeletal dysplasias, enabling adequate management, monitorization, and genetic counseling. © 2016 Wiley Periodicals, Inc.

Background Patient diaries and pain scales can capture the course and complications of pain managed at home in children. The Faces Pain Scale-Revised (FPS-R) is a validated scale showing reliability in children, but it has not been validated in children with sickle cell disease (SCD). Objective The purpose of this study was to evaluate comprehension and usability of an electronic modified version of the FPS-R among pediatric patients with SCD. Methods This was a cross-sectional, qualitative study involving in-person interviews with children/adolescents from the USA and their parents/legal guardians. Interviews involved cognitive debriefing and usability testing of the FPS-R. Results In total, 22 children with SCD aged 4–17 years participated. Children aged 4–6 were generally unable to demonstrate clear understanding of the FPS-R and its response scale. Overall, children aged ≥7 years understood the instrument and could complete it on the electronic device, although children aged 7–8 often needed assistance from the parent. Children aged 9–17 years were able to read and complete the instrument independently. Most participants considered the electronic device easy to use. Conclusions The FPS-R was shown to be a comprehensible and usable pain measure for children aged 7–17 with SCD and to be beneficial for future clinical studies.

Purpose To assess outcomes of computed tomography (CT)-guided methylene blue/collagen marking of preoperative lung nodules before video-assisted thoracoscopic surgery (VATS) and robotic-assisted thoracic surgery (RATS). Materials and Methods A retrospective cohort study assessing 25 methylene blue/collagen solution CT-guided lung nodule localization procedures on 26 nodules in 25 patients was performed. The procedures were performed by a fellowship-trained radiologist 1–2 hours before scheduled surgery under local anesthesia. Approximately 4–6 ml of methylene blue/collagen solution was injected in a perinodular location under CT guidance with a 19-gauge trocar needle and along the track to the visceral pleural surface. Post-procedural CT images confirmed appropriate lung nodule location marking. Results Perinodular CT-guided trocar needle placement was achieved in all marking procedures (n = 26/26). Increased consolidation near the target nodule was also demonstrated in all patients on the post-procedural localized CT scans. One patient with moderate emphysema developed a small to moderate-sized pneumothorax (∼20%–30%), and an 8-Fr thoracentesis catheter was placed under CT guidance before surgery. There was no bleeding or hemoptysis in any patient. Methylene blue/collagen solution was readily visible by the thoracic surgeon in association with all target nodules. One patient required conversion to open procedure due to the proximal portion of the right lower lobe pulmonary artery segmental branch. Of the 26 identified nodules, pathology specimens confirmed the adequacy of nodule resection in all cases. Conclusions Preoperative CT-guided methylene blue/collagen solution injection offers a safe and highly effective technique for marking subpleural lung nodules undergoing VATS or RATS.

Summary Normal faults in clastic sequences are segmented at a range of scales, but often only a single localised fault can be identified on seismic reflection data due to limited lateral resolution. Seismically invisible fault segmentation and associated displacement partitioning can cause unexpected across-fault communication. In this presentation we describe new algorithms and software for modelling subs-seismic fault segmentation in reservoir geomodels, and for risking its effect on across-fault juxtaposition (FaultMaker).

This phase 2 study was designed to characterize the relationship among prasugrel dose, prasugrel's active metabolite (Pras-AM), and platelet inhibition while evaluating safety in children with sickle cell disease. It was open-label, multicenter, adaptive design, dose ranging, and conducted in 2 parts. Part A: Patients received escalating single doses leading to corresponding increases in Pras-AM exposure and VerifyNow®P2Y12 (VN) platelet inhibition and decreases in VNP2Y12 reaction units and vasodilator-stimulated phosphoprotein platelet reactivity index. Part B: Patients were assigned daily doses (0.06, 0.08, and 0.12 mg/kg) based on VN pharmacodynamic measurements at the start of 2 dosing periods, each 14±4 days. Platelet inhibition was significantly higher at 0.12 mg/kg (56.3%±7.4%; least squares mean±SE) compared with 0.06 mg/kg (33.8%±7.4%) or 0.08 mg/kg (37.9%±5.6%). Patients receiving 0.12 mg/kg achieved ≥30% platelet inhibition; only 1 patient receiving 0.06 mg/kg exceeded 60% platelet inhibition. High interpatient variability in response to prasugrel and the small range of exposures precluded rigorous characterization of the relationship among dose, Pras-AM, and platelet inhibition.No hemorrhagic events occurred in Part A; 3 occurred in Part B, all mild and self-limited.Most children with sickle cell disease may achieve clinically relevant platelet inhibition with titration of daily-dose prasugrel.

Purpose: To quantify dose mapping errors (DMEs) of a point‐based dose mapping method for 4D lung treatment plans. Methods: Point‐based dose mapping methods utilize deformation vector fields (DVFS) to interpolate dose from a deformed image. Volume‐based dose mapping methods consider the volume overlap between deformed and reference voxels; defining dose as the integral energy divided by the integral mass of the voxel, and conserving integral dose . DME is defined as the dose differences between volume‐based and point‐based mapped dose (DME=( DpointBased‐DvolumeBased)/DRx). The DME for a 4D lung case is compared with a bitmap DME method, both using a Pinnacle research version 8.1y DVF. DME is computed for ten 4D lung cases (five 10 phases, five 3 phase) with Pinnacle research version 9.100 DVFs. Multi‐phase accumulated 4D DMEs are also evaluated. Results: For all cases, the largest DMEs are located in the dose/density gradient regions. With Pinnacle 8.1y DVF, mapping dose from phase 9 to phase 0, results in a DME=−0.2%±6.1% (range of −76%∼112%). The same case with Pinnacle 9.100 DVFs, DME=0.3%±4.8%(−41%∼32%). Locations of large DME are consistent with those from the bitmap method. For the ten 4D lung cases, accumulated mean DME are within ±0.07% (std. deviations: 1∼5%, range −102%∼64%). Maximum tumor DMEs are less than 30cGy (DRx=7200cGy) for all patients. Conclusions: Due to its inherent integral dose conservation, volume‐based dose mapping methods can quantify errors in point‐based dose mapping methods. While mean DME values are small for the cases tested, standard deviations near 5% indicate that a substantial number of voxels have ∼5% dose mapping errors, however these dose errors do not occur in the target structures. Work supported by NIH P01CA116602

Background Prolonged sitting is a risk factor for low-back pain. The primary purpose of this study is to determine if prolonged active sitting will result in increased trunk motion. Methods Fifteen healthy female participants volunteered to sit for 30 min on each of three surfaces including an air-cushion, a stability ball, and a hard surface. Trunk motion was monitored using a Vicon motion capture system, and foot center of pressure was collected with two AMTI force plates. Results Our findings indicated that the average speed of the trunk center of mass significantly increased with seating surface compliance. There were significant differences in right and left foot centers of pressure in the antero-posterior direction between the ball and air-cushion conditions and the ball and chair conditions. Conclusion Active sitting results in increased trunk motion and could have a positive effect on low-back health.

[Background]: Osteogenesis imperfecta (OI) is a heterogeneous bone disorder characterized by recurrent fractures. Although most cases of OI have heterozygous mutations in COL1A1 or COL1A2 and show autosomal dominant inheritance, during the last years there has been an explosion in the number of genes responsible for both recessive and dominant forms of this condition. Herein, we have analyzed a cohort of patients with OI, all offspring of unaffected parents, to determine the spectrum of variants accounting for these cases. Twenty patients had nonrelated parents and were sporadic, and 21 were born to consanguineous relationships., [Methods]: Mutation analysis was performed using a next‐generation sequencing gene panel, homozygosity mapping, and whole exome sequencing (WES)., [Results]: Patients offspring of nonconsanguineous parents were mostly identified with COL1A1 or COL1A2 heterozygous changes, although there were also a few cases with IFITM5 and WNT1 heterozygous mutations. Only one sporadic patient was a compound heterozygote for two recessive mutations. Patients offspring of consanguineous parents showed homozygous changes in a variety of genes including CRTAP,FKBP10,LEPRE1,PLOD2,PPIB,SERPINF1,TMEM38B, and WNT1. In addition, two patients born to consanguineous parents were found to have de novo COL1A1 heterozygous mutations demonstrating that causative variants in the collagen I structural genes cannot be overlooked in affected children from consanguineous couples. Further to this, WES analysis in probands lacking mutations in OI genes revealed deleterious variants in SCN9A,NTRK1, and SLC2A2, which are associated with congenital indifference to pain (CIP) and Fanconi–Bickel syndrome (FBS)., [Conclusion]: This work provides useful information for clinical and genetic diagnosis of OI patients with no positive family history of this disease. Our data also indicate that CIP and FBS are conditions to be considered in the differential diagnosis of OI and suggest a positive role of SCN9A and NTRK1 in bone development., Spanish Ministry of Economy and Competitiveness (Grant/Award Number: ‘SAF2013-43365-R’), CIBERER (Grant/Award Number: ‘ER14-PR04-ACCI13-760’).

Deep (> ∼800 m) saline water-bearing formations in the United States have substantial pore volume that is targeted for storage of carbon dioxide (CO2) and the associated saline water can be extracted to increase CO2 storage efficiency, manage pressure build up, and create a new water source that, once treated, can be used for power-plant cooling or other purposes. Extraction, treatment and disposal costs of saline formation water to meet added water demands from CO2 capture and storage (CCS) are discussed. This underutilized water source may be important in meeting new water demand associated with CCS. For a representative natural gas combined-cycle (NGCC) power plant, simultaneous extraction of brine from the storage formation could provide enough water to meet all CCS-related cooling demands for 177 out of the 185 (96 %) saline formations analyzed in this study. Calculated total cost of water extraction, treatment and disposal is less than 4.00 US Dollars (USD) m−3 for 93 % of the 185 formations considered. In 90 % of 185 formations, treated water costs are less than 10.00 USD tonne−1 of CO2 injected. On average, this represents approximately 6 % of the total CO2 capture and injection costs for the NGCC scenario.

SummaryPatients with sickle cell anaemia (SCA) have vaso-occlusive crises resulting from occlusive hypoxic-ischaemic injury. Prasugrel inhibits platelet activation and aggregation involved in SCA pathophysiology. Determining Effects of Platelet Inhibition on Vaso-Occlusive Events (DOVE) was a phase 3, double-blind, randomised, placebo-controlled trial assessing prasugrel efficacy. DOVE sought to bring patients’ P2Y12 reaction unit (PRU) value within a targeted range via prasugrel dose adjustments using encrypted VerifyNow P2Y12® (VN-P2Y12) point-of-care testing and an interactive voice-response system (IVRS). After PRU determination, randomised patients received 0.08 mg/kg/day prasugrel or placebo. Encrypted PRUs and IVRS provided double-blind dose adjustments to achieve a defined PRU target range of 136–231; placebo patients had mock titrations. Of 341 randomised patients, 166 placebo and 160 prasugrel patients reached the fully titrated dose (FTD). Most prasugrel patients (n=104, 65 %) remained on the initial 0.08 mg/kg dose; doses escalations occurred in 23 % of patients (n=36). Mean PRUs for the pharmacodynamic population at baseline were similar in the prasugrel (273 ± 44.9) and placebo groups (273 ± 51.7), with significant reductions in PRU (p

Background Heterozygous loss-of-function mutations in the natriuretic peptide receptor B gene (NPR2) are responsible for short stature in patients without a distinct phenotype. Some of these patients have been treated with recombinant human growth hormone (rhGH) therapy with a variable response. Case presentation The proband was a healthy boy who presented at the age of 5.1 years with familial short stature (height SDS of -3.1). He had a prominent forehead, a depressed nasal bridge, centripetal fat distribution and a high-pitched voice resembling that of children with GH deficiency. His hormonal evaluation showed low insulin-like growth factor-1 (IGF-1) but a normal GH peak at a stimulation test. During the first year of rhGH treatment, his growth velocity increased from 3.4 to 10.4 cm/year (height SDS change of +1.1). At the last visit, he was 8.8 years old and still on treatment, his growth velocity was 6.4 cm/year and height SDS was -1.8. Results We identified through exome sequencing a novel heterozygous loss-of-function NPR2 mutation (c.2905G>C; p.Val969Leu). Cells cotransfected with the p.Val969Leu mutant showed a significant decrease in cyclic guanosine monophosphate (cGMP) production compared to the wild type (WT), suggesting a dominant negative effect. Conclusions This case reveals a novel heterozygous loss-of-function NPR2 mutation responsible for familial short stature and the good response of rhGH therapy in this patient.

outbreak, will have to be managed during a disease outbreak to minimize adverse impact on animal welfare. To put animal welfare in disease outbreaks into context, the obvious solution to obtaining optimal animal welfare is to prevent animal disease outbreaks in the fi rst place, as has been recently accomplished with the global eradication of Rinderpest. It also helps to appreciate that, although the focus of this article is on the impact of disease outbreaks on animal welfare, the number of animals suffering from substandard welfare as a result of production-related disease and poor management under non-disaster conditions dwarfs the number of animals suffering from geophysical, technological, and biological disasters combined. Finally, the reader should be aware of the extensive literature and experience showing that preparedness before disaster strikes as well as the reliance on proven management techniques during an event provide the greatest returns on investment in emergency management (Disaster Research Center, 2012; FEMA, 2012a; National Hazards Center, 2012). When the principles of emergency management are applied to the care of animals, animal welfare in disease outbreaks will be optimized.

Summary The Pump Canyon CO2-enhanced coalbed methane (ECBM)/ sequestration demonstration in New Mexico has the primary objective of demonstrating the feasibility of CO2 sequestration in deep, unmineable coal seams through a small-scale geologic sequestration pilot. This project is not the first of its kind; several small- or large-scale pilots were already conducted previously in the United States [Allison Unit (Reeves et al. 2003) in the San Juan, Appalachian, and Warrior basins] as well as internationally [the Recopol (Reeves and Oudinot 2002) project in Poland, and the Yubari project in Japan, Canada, and Australia]. Additional pilots are currently under way. At the project site, a new CO2-injection well was drilled within an existing pattern of coalbed-methane-production wells. Primarily operated by ConocoPhillips, these wells produce from the Late Cretaceous Fruitland coals. CO2 injection into these coal seams was initiated in late July 2008 and ceased in August 2009. A variety of monitoring, verification, and accounting (MVA) methods were employed to track the movement of the CO2 in order to determine the occurrence of leakage. Within the injection well, MVA methods included continuous measurement of injection volumes, pressures, and temperatures. The offset production wells sampled gas-production rates, pressures, and gas composition through CO2 sensors, tracers in the injected CO2, time-lapse vertical seismic profiling, and surface tiltmeter arrays. A detailed study of the overlying Kirtland shale was also conducted to investigate the integrity of this primary caprock. This information was used to develop a detailed geologic characterization and reservoir model that has been used to further understand the behavior of this reservoir. The CO2-injection pilot has ended with no significant CO2 buildup occurring in the offset production wells. However, a small but steady increase in CO2 and N2 at two of the offset wells may have been an indication of imminent breakthrough. More recent gas samples are, however, showing a decrease in CO2 and N2 content at those wells. This paper describes the project, covering the regulatory process and injection-well construction, the different techniques used to monitor for CO2 leakage, and the results of the modeling work.

Materials science needs to study dynamic properties of high-Z materials lead to a unique and challenging set of requirements for future X-ray free-electron lasers (XFELs), with single-pulse fluxes of up to 1012 50 keV X-rays that are both transversely and longitudinally coherent. These parameters cannot be met through an extension of the beam and FEL technologies used at existing and currently planned X-ray FEL facilities. We describe a novel technique to achieve higher fluxes by reducing the transverse beam emittance of high bunch charges and another to achieve longitudinal coherency by pre-modulating the electron beam current before it reaches the undulator. These techniques are investigated numerically and analytically, and also hold potential for increasing performance and decreasing cost of soft X-ray FELs.