Rasmus Oestergaard Nielsen

A transcriptome map of perennial ryegrass (Lolium perenne L.)

BackgroundSingle nucleotide polymorphisms (SNPs) are increasingly becoming the DNA marker system of choice due to their prevalence in the genome and their ability to be used in highly multiplexed genotyping assays. Although needed in high numbers for genome-wide marker profiles and genomics-assisted breeding, a surprisingly low number of validated SNPs are currently available for perennial ryegrass.ResultsA perennial ryegrass unigene set representing 9,399 genes was used as a reference for the assembly of 802,156 high quality reads generated by 454 transcriptome sequencing and for in silico SNP discovery. Out of more than 15,433 SNPs in 1,778 unigenes fulfilling highly stringent assembly and detection parameters, a total of 768 SNP markers were selected for GoldenGate genotyping in 184 individuals of the perennial ryegrass mapping population VrnA, a population being previously evaluated for important agronomic traits. A total of 592 (77%) of the SNPs tested were successfully called with a cluster separation above 0.9. Of these, 509 (86%) genic SNP markers segregated in the VrnA mapping population, out of which 495 were assigned to map positions. The genetic linkage map presented here comprises a total of 838 DNA markers (767 gene-derived markers) and spans 750 centi Mogan (cM) with an average marker interval distance of less than 0.9 cM. Moreover, it locates 732 expressed genes involved in a broad range of molecular functions of different biological processes in the perennial ryegrass genome.ConclusionsHere, we present an efficient approach of using next generation sequencing (NGS) data for SNP discovery, and the successful design of a 768-plex Illumina GoldenGate genotyping assay in a complex genome. The ryegrass SNPs along with the corresponding transcribed sequences represent a milestone in the establishment of genetic and genomics resources available for this species and constitute a further step towards molecular breeding strategies. Moreover, the high density genetic linkage map predominantly based on gene-associated DNA markers provides an important tool for the assignment of candidate genes to quantitative trait loci (QTL), functional genomics and the integration of genetic and physical maps in perennial ryegrass, one of the most important temperate grassland species.

SNP Discovery Using Next Generation Transcriptomic Sequencing in Atlantic Herring (Clupea harengus)

The introduction of Next Generation Sequencing (NGS) has revolutionised population genetics, providing studies of non-model species with unprecedented genomic coverage, allowing evolutionary biologists to address questions previously far beyond the reach of available resources. Furthermore, the simple mutation model of Single Nucleotide Polymorphisms (SNPs) permits cost-effective high-throughput genotyping in thousands of individuals simultaneously. Genomic resources are scarce for the Atlantic herring (Clupea harengus), a small pelagic species that sustains high revenue fisheries. This paper details the development of 578 SNPs using a combined NGS and high-throughput genotyping approach. Eight individuals covering the species distribution in the eastern Atlantic were bar-coded and multiplexed into a single cDNA library and sequenced using the 454 GS FLX platform. SNP discovery was performed by de novo sequence clustering and contig assembly, followed by the mapping of reads against consensus contig sequences. Selection of candidate SNPs for genotyping was conducted using an in silico approach. SNP validation and genotyping were performed simultaneously using an Illumina 1,536 GoldenGate assay. Although the conversion rate of candidate SNPs in the genotyping assay cannot be predicted in advance, this approach has the potential to maximise cost and time efficiencies by avoiding expensive and time-consuming laboratory stages of SNP validation. Additionally, the in silico approach leads to lower ascertainment bias in the resulting SNP panel as marker selection is based only on the ability to design primers and the predicted presence of intron-exon boundaries. Consequently SNPs with a wider spectrum of minor allele frequencies (MAFs) will be genotyped in the final panel. The genomic resources presented here represent a valuable multi-purpose resource for developing informative marker panels for population discrimination, microarray development and for population genomic studies in the wild.

Novel Tools for Conservation Genomics: Comparing Two High-Throughput Approaches for SNP Discovery in the Transcriptome of the European Hake

The growing accessibility to genomic resources using next-generation sequencing (NGS) technologies has revolutionized the application of molecular genetic tools to ecology and evolutionary studies in non-model organisms. Here we present the case study of the European hake (Merluccius merluccius), one of the most important demersal resources of European fisheries. Two sequencing platforms, the Roche 454 FLX (454) and the Illumina Genome Analyzer (GAII), were used for Single Nucleotide Polymorphisms (SNPs) discovery in the hake muscle transcriptome. De novo transcriptome assembly into unique contigs, annotation, and in silico SNP detection were carried out in parallel for 454 and GAII sequence data. High-throughput genotyping using the Illumina GoldenGate assay was performed for validating 1,536 putative SNPs. Validation results were analysed to compare the performances of 454 and GAII methods and to evaluate the role of several variables (e.g. sequencing depth, intron-exon structure, sequence quality and annotation). Despite well-known differences in sequence length and throughput, the two approaches showed similar assay conversion rates (approximately 43%) and percentages of polymorphic loci (67.5% and 63.3% for GAII and 454, respectively). Both NGS platforms therefore demonstrated to be suitable for large scale identification of SNPs in transcribed regions of non-model species, although the lack of a reference genome profoundly affects the genotyping success rate. The overall efficiency, however, can be improved using strict quality and filtering criteria for SNP selection (sequence quality, intron-exon structure, target region score).

Foot pronation is not associated with increased injury risk in novice runners wearing a neutral shoe: a 1-year prospective cohort study

Objective To investigate if running distance to first running-related injury varies between foot postures in novice runners wearing neutral shoes. Design A 1-year epidemiological observational prospective cohort study. Setting Denmark. Participants A total of 927 novice runners equivalent to 1854 feet were included. At baseline, foot posture on each foot was evaluated using the foot-posture index and categorised into highly supinated (n=53), supinated (n=369), neutral (n=1292), pronated (n=122) or highly pronated (n=18). Participants then had to start running in a neutral running shoe and to use global positioning system watch to quantify the running distance in every training session. Main outcome measure A running-related injury was defined as any musculoskeletal complaint of the lower extremity or back caused by running, which restricted the amount of running for at least 1 week. Results During 1 year of follow-up, the 1854 feet included in the analyses ran a total of 326 803 km until injury or censoring. A total of 252 participants sustained a running-related injury. Of these, 63 were bilateral injuries. Compared with a neutral foot posture, no significant body mass index-adjusted cumulative risk differences (RD) were found after 250 km of running for highly supinated feet (RD=11.0% (−10% to 32.1%), p=0.30), supinated feet (RD=−1.4% (−8.4% to 5.5%), p=0.69), pronated feet (RD=−8.1% (−17.6% to 1.3%), p=0.09) and highly pronated feet (RD=9.8% (−19.3% to 38.8%), p=0.51). In addition, the incidence-rate difference/1000 km of running, revealed that pronators had a significantly lower number of injuries/1000 km of running of −0.37 (−0.03 to −0.70), p=0.03 than neutrals. Conclusions The results of the present study contradict the widespread belief that moderate foot pronation is associated with an increased risk of injury among novice runners taking up running in a neutral running shoe. More work is needed to ascertain if highly pronated feet face a higher risk of injury than neutral feet.

A step towards understanding the mechanisms of running-related injuries

OBJECTIVES To investigate the association between training-related characteristics and running-related injury using a new conceptual model for running-related injury generation, focusing on the synergy between training load and previous injuries, short-term running experience or body mass index (> or < 25 kg m(-2)). DESIGN Prospective cohort study with a 9-month follow-up. METHODS The data of two previous studies using the same methodology were revisited. Recreational runners (n = 517) reported information about running training characteristics (weekly distance, frequency, speed), other sport participation and injuries on a dedicated internet platform. Weekly volume (dichotomized into < 2h and ≥ 2 h) and session frequency (dichotomized into < 2 and ≥ 2) were the main exposures because they were considered necessary causes for running-related injury. Non-training-related characteristics were included in Cox regression analyses as effect-measure modifiers. Hazard ratio was the measure of association. The size of effect-measure modification was calculated as the relative excess risk due to interaction. RESULTS One hundred sixty-seven runners reported a running-related injury. Crude analyses revealed that weekly volume < 2h (hazard ratio = 3.29; 95% confidence intervals = 2.27; 4.79) and weekly session frequency < 2 (hazard ratio = 2.41; 95% confidence intervals = 1.71; 3.42) were associated with increased injury rate. Previous injury was identified as an effect-measure modifier on weekly volume (relative excess risk due to interaction = 4.69; 95% confidence intervals = 1.42; 7.95; p = 0.005) and session frequency (relative excess risk due to interaction = 2.44; 95% confidence intervals = 0.48; 4.39; p = 0.015). A negative synergy was found between body mass index and weekly volume (relative excess risk due to interaction = -2.88; 95% confidence intervals = -5.10; -0.66; p = 0.018). CONCLUSIONS The effect of a runners training load on running-related injury is influenced by body mass index and previous injury. These results show the importance to distinguish between confounding and effect-measure modification in running-related injury research.

Can GPS Be Used to Detect Deleterious Progression in Training Volume Among Runners

Abstract Nielsen, RO, Cederholm, P, Buist, I, Sørensen, H, Lind, M, and Rasmussen, S. Can GPS be used to detect deleterious progression in training volume among runners? J Strength Cond Res 27(6): 1471–1478, 2013—There is a need to ascertain if an association exists between excessive progression in weekly volume and development of running-related injuries (RRI). The purpose of this study was to investigate if GPS can be used to detect deleterious progression in weekly training volume among 60 novice runners included in a 10-week prospective study. All participants used GPS to quantify training volume while running. In case of injury, participants attended a clinical examination. The 13 runners who sustained injuries during follow-up had a significantly higher weekly progression in total training volume in the week before the injury origin (86% [95% confidence interval: 12.9–159.9], p = 0.026) compared with other weeks. Although not significant, participants with injuries had an increase in weekly training volume of 31.6% compared with a 22.1% increase among the healthy participants. The error of the GPS measurements in open landscape, forest, and urban area of volume was ⩽6.2%. To conclude, no clinically relevant measurement errors of the GPS devices were found for training volume. Based on this, GPS has a potential to detect errors in training volume, which may be associated with development of RRI. Based on the results from the current study, increases in weekly training progression may become deleterious at a weekly increase above 30%, which is more than the 10% rule currently used as a guideline for correct progression in weekly volume by runners and coaches. Still, no clear evidence for safe progression of weekly volume exists. But it seems likely that some individuals may tolerate weekly progressions around 20–25%, at least for a short period of time.

Predictors of Running-Related Injuries Among 930 Novice Runners A 1-Year Prospective Follow-up Study

Background: To identify persons at high risk of sustaining running-related injuries, an evidence-based understanding of the risk factors associated with injury is needed. Purpose: To identify demographic and behavioral risk factors associated with running-related injuries. Study Design: Observational prospective cohort study with a 1-year follow-up. Methods: Exposures including sex, age, body mass index (BMI), behavior (Type A Self-Rating Inventory [TASRI]), running experience, other sports activity, previous running-related injuries, and other injuries not related to running were assessed prior to or at baseline. The outcome of interest was a running-related injury, defined as any musculoskeletal complaint of the lower extremity or back caused by running that restricted the amount of running (volume, duration, pace, or frequency) for at least 1 week. All participants quantified their running volume by global positioning system (GPS) and used a neutral running shoe. Time to first injury for each exposure variable was analyzed using a generalized linear model, with cumulative kilometers of the training sessions as the time scale. Results: A total of 930 individuals were included in the study, of which 254 sustained a running-related injury during a total of 155.318 km of running. By calculating the cumulative injury risk differences (cIRDs) [95% confidence intervals] after 500 km of running, the TASRI Type B behavior (cIRD, 11.9% [−0.5%; 23.3%]; P = .04) was found to be a significant predictor of injury, while age between 45 and 65 years (cIRD, 14.7% [−2.1%; 31.5%]; P = .08) and previous injuries not related to running (cIRD, 11.1% [−0.2%; 22.4%]; P = .05) were considered clinically interesting, although not statistically significant. In addition, χ2 test results across 4 BMI groups also revealed a borderline significant relationship (P = .06). No significant or clinically relevant relationships were found for sex (P = .42), previous running-related injury (P = .47), running experience (P = .30), and other sports activities (P = .30). Conclusion: The findings of the present study suggest BMI >30 kg/m2, age between 45 and 65 years, noncompetitive behavior, and previous injuries not related to running are associated with increased risk of injury among novice runners, while BMI <20 kg/m2 was protective. Still, the role of the risk factors in the causal mechanism leading to injury needs to be investigated.

Classification of the height and flexibility of the medial longitudinal arch of the foot

BackgroundThe risk of developing injuries during standing work may vary between persons with different foot types. High arched and low arched feet, as well as rigid and flexible feet, are considered to have different injury profiles, while those with normal arches may sustain fewer injuries. However, the cut-off values for maximum values (subtalar position during weight-bearing) and range of motion (ROM) values (difference between subtalar neutral and subtalar resting position in a weight-bearing condition) for the medial longitudinal arch (MLA) are largely unknown. The purpose of this study was to identify cut-off values for maximum values and ROM of the MLA of the foot during static tests and to identify factors influencing foot posture.MethodsThe participants consisted of 254 volunteers from Central and Northern Denmark (198 m/56 f; age 39.0 ± 11.7 years; BMI 27.3 ± 4.7 kg/m2). Navicular height (NH), longitudinal arch angle (LAA) and Feiss line (FL) were measured for either the left or the right foot in a subtalar neutral position and subtalar resting position. Maximum values and ROM were calculated for each test. The 95% and 68% prediction intervals were used as cut-off limits. Multiple regression analysis was used to detect influencing factors on foot posture.ResultsThe 68% cut-off values for maximum MLA values and MLA ROM for NH were 3.6 to 5.5 cm and 0.6 to 1.8 cm, respectively, without taking into account the influence of other variables. Normal maximum LAA values were between 131 and 152° and normal LAA ROM was between -1 and 13°. Normal maximum FL values were between -2.6 and -1.2 cm and normal FL ROM was between -0.1 and 0.9 cm. Results from the multivariate linear regression revealed an association between foot size with FL, LAA, and navicular drop.ConclusionsThe cut-off values presented in this study can be used to categorize people performing standing work into groups of different foot arch types. The results of this study are important for investigating a possible link between arch height and arch movement and the development of injuries.

A prospective study on time to recovery in 254 injured novice runners

Objectives Describe the diagnoses and the time to recovery of running-related injuries in novice runners. Design Prospective cohort study on injured runners. Method This paper is a secondary data analysis of a 933-person cohort study (DANO-RUN) aimed at characterizing risk factors for injury in novice runners. Among those sustaining running-related injuries, the types of injuries and time to recovery is described in the present paper. All injured runners were diagnosed after a thorough clinical examination and then followed prospectively during their recovery. If they recovered completely from injury, time to recovery of each injury was registered. Results A total of 254 runners were injured. The proportion of runners diagnosed with medial tibial stress syndrome was 15%, 10% for patellofemoral pain, 9% for medial meniscal injury, 7% for Achilles tendinopathy and 5% for plantar fasciitis. Among the 220 runners (87%) recovering from their injury, the median time to recovery was 71 days (minimum  = 9 days, maximum  = 617 days). Conclusions Medial tibial stress syndrome was the most common injury followed by patellofemoral pain, medial meniscal injury and Achilles tendinopathy. Half of the injured runners were unable to run 2×500 meters without pain after 10 weeks. Almost 5% of the injured runners received surgical treatment.

A framework for the etiology of running-related injuries.

The etiology of running‐related injury is important to consider as the effectiveness of a given running‐related injury prevention intervention is dependent on whether etiologic factors are readily modifiable and consistent with a biologically plausible causal mechanism. Therefore, the purpose of the present article was to present an evidence‐informed conceptual framework outlining the multifactorial nature of running‐related injury etiology. In the framework, four mutually exclusive parts are presented: (a) Structure‐specific capacity when entering a running session; (b) structure‐specific cumulative load per running session; (c) reduction in the structure‐specific capacity during a running session; and (d) exceeding the structure‐specific capacity. The framework can then be used to inform the design of future running‐related injury prevention studies, including the formation of research questions and hypotheses, as well as the monitoring of participation‐related and non‐participation‐related exposures. In addition, future research applications should focus on addressing how changes in one or more exposures influence the risk of running‐related injury. This necessitates the investigation of how different factors affect the structure‐specific load and/or the load capacity, and the dose‐response relationship between running participation and injury risk. Ultimately, this direction allows researchers to move beyond traditional risk factor identification to produce research findings that are not only reliably reported in terms of the observed cause‐effect association, but also translatable in practice.