Traumatic spinal cord injury (SCI) disrupts spinal and supraspinal pathways, and this process is reflected in changes in surface electromyography (sEMG). sEMG is an informative complement to current clinical testing and can capture the residual motor command in great detail—including in muscles below the level of injury with seemingly absent motor activities. In this comprehensive review, we sought to describe how the sEMG properties are changed after SCI. We conducted a systematic literature search followed by a narrative review focusing on sEMG analysis techniques and signal properties post-SCI. We found that early reports were mostly focused on the qualitative analysis of sEMG patterns and evolved to semi-quantitative scores and a more detailed amplitude-based quantification. Nonetheless, recent studies are still constrained to an amplitude-based analysis of the sEMG, and there are opportunities to more broadly characterize the time- and frequency-domain properties of the signal as well as to take fuller advantage of high-density EMG techniques. We recommend the incorporation of a broader range of signal properties into the neurophysiological assessment post-SCI and the development of a greater understanding of the relation between these sEMG properties and underlying physiology. Enhanced sEMG analysis could contribute to a more complete description of the effects of SCI on upper and lower motor neuron function and their interactions, and also assist in understanding the mechanisms of change following neuromodulation or exercise therapy.The online version contains supplementary material available at 10.1186/s12984-021-00888-2. Traumatic spinal cord injury (SCI) may lead to severe sensorimotor dysfunction depending on the level and severity of injury. Changes in motor properties include muscle atrophy, muscle fiber type transformation, and increased passive stiffness in muscles and tendons , resultisEMG has been suggested to be a good marker for muscle health and function . ImportaIn this context, characterizing the impact of the SCI itself on sEMG is of interest for multiple reasons. First, injury-related changes and spontaneous recovery in the sEMG must be understood in order to isolate the impact of interventions. Second, neural interfaces that rely on sEMG require a good understanding of the expected signal properties. Third, a thorough understanding of what information can and cannot be obtained from non-invasive methods is essential to promoting the translation of electrophysiological techniques into routine clinical use. How are the sEMG properties changed after SCI? The answer to this question is not trivial, especially given that the sEMG signal reflects the net output of complex interactions between intrinsic spinal cord circuits, motor axon properties, and muscular mechanisms. sEMG is a well-established methodology to explore muscle activity after SCI; however, there is no consensus on data analysis and reporting strategies. The most common reporting methodology utilizes the signal amplitude, typically representing the average or maximal value recorded at a given time or window after some smoothing and normalization steps. However, the raw sEMG signal has the potential to be analyzed and expressed in many ways using temporal, spectral, and spatial techniques. A broad understanding of sEMG changes after SCI is of great interest in order to fully realize the potential of this measurement modality to characterize neurorecovery and support the development of neurorehabilitation technologies –16.This scoping review aims to summarize and critically appraise the existing literature on how SCI can alter the sEMG properties. We distilled the large SCI literature reporting sEMG properties during residual volitional movements or abnormal spontaneous activity. These two domains are clinically significant given that neurorehabilitative interventions after SCI are thought to increase volitional control and reduce spontaneous activity. Accordingly, our results are structured around the different properties of the sEMG that can be expected to change after SCI. Insights into the neurophysiological basis of these phenomena are discussed and a new evidence-based perspective on the use of sEMG after SCI is provided.https://www.crd.york.ac.uk/prospero/#aboutregpage; PROSPERO; Registration number CRD42020159040) and the Open Science Framework (https://osf.io/cg6yu/). Subsequently, a systematic review of the literature was conducted according to the checklist for the Preferred Reporting Items for Systematic reviews and Meta-Analyses extension for Scoping Reviews (PRISMA-ScR) Checklist [This is a scoping review using a systematic search (hybrid review). The systematic search was registered within the International Prospective Register of Systematic Reviews , Cochrane Central Register of Controlled Trials (116 entries), Cochrane Database of Systematic Reviews (17 entries), Embase (1680 entries), Emcare (555 entries), Cumulative Index to Nursing & Allied Health Literature , and PubMed . The searches were originally performed on December 23rd, 2019 with no time limitations and re-run on September 22nd, 2020. No limits were applied for language to avoid excluding references not assigned to a language. References were also searched manually by reviewing reference lists of the included studies. Refer to Additional file To be included, studies had to: (1) report sEMG properties in SCI participants ; (2) include participants with traumatic SCI ; and (3) be written in English. Regarding (1), we defined the sample size threshold a priori to accommodate the low prevalence characteristic of SCI while balancing the need to select studies that contain generalizable information. In (2), despite the obvious differences in the time profile of disease onset in traumatic and non-traumatic  SCI, it is thought that both lead to similar white matter degeneration . FurtherA PICO model  was used to build search criteria for the electronic databases. The PICO consisted of Population: “Spinal Cord Injury”, Intervention/Identifier: “sEMG”, and Outcome of interest: “Muscle/Motor Response”. Valid subject headings as appropriate for each database were utilized in the search strategies, as were free-text terms relevant to each topical concept.Duplicate references were removed manually in addition to using Covidence  and Mendeley software , and manually. Two authors (GB and GL) independently screened titles and abstracts to determine initial eligibility. Eligible references were included for full-text screening. Conflicts were resolved by a third reviewer (MJW).Data extraction from each full-text article was completed by the first author (GB) using a personalized spreadsheet, which was pilot-tested and refined using 5% of the references that passed the full-text screening. Extracted data included: (1) study identification information (author and year); (2) study design; (3) participant demographics: level of injury, American Spinal Injury Association Impairment Scale (AIS), time post-injury, sex, and age; (4) sample size; (5) the motives for using sEMG; (6) muscles evaluated; (7) sEMG equipment and electrodes; (8) electrodes placement and reliability information; (9) sEMG data analysis and reporting strategy; (10) main sEMG findings: means, standard deviations and p values for relevant outcome measures; (11) measures other than sEMG. If insufficient data were reported, the authors were contacted by email.Following the literature search and selection, a data score was assigned by the first author (GB) as follows to sEMG methodology description was rated as present or not using the following criteria: sEMG equipment/electrode and amplification/filtering description was considered present (yes) if the type and settings of equipment were described in sufficient details . Similarly, the electrode placement was considered as present if it was described in sufficient detail  or based on established references . We also rated the sEMG findings employed in the studies using a 3-point scale: qualitative , semi-quantitative scores  or quantitative (2).The PRISMA flowchart is described in Fig. Most of the references identified used the ASIA Impairment Scale (AIS) component of the ISNCSCI to describe their study sample (108 of 178 studies) when measuring sEMG properties at rest 32 studies) or during volitional effort (76 studies). Studies assessing sEMG at rest included more AIS A and B participants (≈ 71% of studies), while studies assessing volitional control included mostly AIS C and D (≈ 62% of studies). Most of the studies (≈ 67%) were conducted in cervical lesions, indicating sensorimotor impairments in the upper and lower limbs , a substantial amount in upper limbs muscles (≈ 26% of studies), and only a few studies in the trunk/head muscles (≈ 6% of studies) Fig. a. TibialThe interest in lower limb muscles, the predominance of males, and lack of AB control groups Fig. h can be The following sections summarize the currently available evidence about alterations in the characteristics of the sEMG signal after SCI. The focus of these investigations is summarized in Fig. Using qualitative sEMG graph analysis, early investigations explored the use of volitional muscle activation during postural movements, which described postural patterns and the tendency for clonus of hip flexors/extensors during the act of rising , as wellWith the progression of sEMG technology, researchers were able to assess multiple muscle groups simultaneously with higher resolution. This facilitated the use of sEMG as a means to control assistive devices, as different sites for myoelectric control in individuals with high-level quadriplegia were tested using integrated sEMG amplitude to detect the amount of volitional control over trunk/head muscles . It was Of 178 studies, only 18 were conducted during the acute and sub-acute phases of SCI (< 1-year post-injury) Fig. a. MuscleThe disruption of UMN and LMN activity at specific spinal cord segments results in muscle-specific weakness, which relates to the level of spinal cord lesion and determines muscle activity following SCI . UMN andThe solid relation between muscle strength and sEMG stemmed from the seminal work of Calancie and colleagues and opened new avenues in the application of sEMG in understanding neural plasticity and movement recovery following SCI . These sDietz and colleagues integrated electrophysiological evaluations into their description of the spontaneous recovery of locomotor muscles over 26 weeks post-injury. sEMG amplitude and reflex activity were absent for up to 14 weeks post-SCI; following resolution of the spinal shock, tendon tap response reappeared, and the sEMG amplitude gradually increased over the next 4 weeks—and plateaued at around 22 weeks post-SCI , 35. FurFinally, Sherwood and colleagues contributed to the understanding of muscle weakness and recovery following SCI through the introduction of the Brain Motor Control Assessment protocol. This comprehensive multichannel surface sEMG approach allowed a better characterization of motor control features in individuals with UMN dysfunction , 39. TheCo-activation in sub-acute SCI was also studied longitudinally using amplitude-based RMS sEMG properties from 3 to 5 months post-SCI. The results suggested an unbalanced recovery of UMN control over muscles (co-activation) and the attenuation of the recovery process by the presence of lower limb hypertonia and involuntary muscle activity . Muscle Early reports of volitional muscle activation during the chronic phase of SCI (> 1-year post-injury) aimed at determining the optimal myoelectric control sites  and lateThe ability of sEMG to capture residual impairments or muscle activity after SCI has been used to categorize muscles according to the presence or absence of detectable movement and determine the extent of preserved muscle activity , 48, incThe Brain Motor Control Assessment introduced in “Muscle properties may also change in chronic SCI. The pattern of central motor drive of the plantar flexors indicated that individuals with SCI generated greater activation of the plantar flexors during eccentric MVCs compared with isometric or concentric MVCs, likely related to increased efficacy of muscle spindles Ia-α motoneuron transmission during lengthening contractions , 58. FinNovel technologies take advantage of high-density sEMG and wearable electrodes. High-density sEMG can provide more detailed information on patterns of muscle activation, which can also be summarized, for example, using the center of gravity , 61. WeasEMG has been used extensively to gain insights into motor control alterations after SCI. Lines of investigation have included abnormal synergies , completWhile these research efforts have yielded considerable insights into neural control following SCI, most of them rely on similar methodologies to quantify the sEMG signal. The emphasis has been placed on timing information to describe patterns of activation, and normalized envelope amplitudes  or frequency-domain . The amplitude metrics discussed above are time-domain metrics, but are dealt with separately in this article because of their widespread use. Other time-domain sEMG properties have been extensively used in the myoelectric control literature, and are thought to provide indirect information on motor unit activation , 117. CoOnly a few studies employed these analyses in SCI-related studies. Some studies performed detailed time-domain-based feature extraction to optimize myoelectric pattern recognition-based control systems, using, for example, the mean absolute value, zero crossings, waveform length, slope sign changes, and fourth-order autoregressive coefficients –126. WhiSCI decreases the motor unit firing during volitional drive. During maximal voluntary effort of the hand thenar muscles, the maximal motor unit firing rate of SCI participants was found to be roughly 15 Hz, much lower compared with 34 Hz in non-injured participants , 131. AlChanges in sEMG frequency-domain patterns have been associated with improvements in walking; for example, after training, the amount of power in the 7- to 9-Hz clonus band decreased . VariatiFinally, reduction in mean and median frequencies is a common indicator of increasing levels of muscle fatigue. This reflects that the mechanisms of central fatigue are likely related to the reduced firing of motor units. Early studies in SCI populations took advantage of these properties to test fatigability during wheelchair propulsion and showed how elite wheelchair athletes displayed improved endurance and a slower decline in these frequency-based sEMG properties . Similarβ-band activation in cortical motor networks leads to cortical UMN commands often being synchronized within this frequency. As such, β-band frequency is evident in the sEMG properties of intra- and intermuscular coherence.Norton and Gorassini have shown that changes in cortically related intermuscular coherence are associated with improvements in locomotor skills following treadmill training, likely mediated by increases in the corticospinal drive to muscles . IntramuFrequency-based sEMG analysis may also be used to investigate corticomuscular coherence following SCI using electroencephalography. Corticomuscular coherence at lower frequencies (≈ 10 Hz) may indicate decreased cortical influence on spinal centers, leading to increased muscle co-activation . InteresSCI reduces the UMN input to the spinal cord circuitry and creates an opportunity for plasticity and reorganization. Synaptic territory may be invaded by preserved connections not interrupted by the SCI, afferents arriving from the periphery , and/or pathways within the intrinsic spinal cord circuitry. As a consequence, the temporal and spatial summations of action potentials become unbalanced in the intrinsic spinal cord circuitry and more prone to excitation from the periphery, alongside the reduction of cortical inhibition. This leads to the emergence of hyperreflexia and long-lasting spontaneous firing of LMNs, and also suggests the loss of inhibitory interneurons within the spinal cord circuitry . AmplituHarnessing the residual brain influence on spinal cord function includes increasing volitional activation but also reducing involuntary spontaneous activity. sEMG activity below the lesion can be studied in great detail using sEMG amplitude-based properties at rest. For example, similar to volitional contractions but tailored to quantify the response of paralyzed muscles to tendon taps, vibration, and plantar stimulation, the Brain Motor Control Assessment score can reflect the presence or absence of reflexes or the ability of voluntary suppression . The resEarly reports of involuntary sEMG activity in SCI described how the deprivation of supraspinal control leads to an increase in LMN activities. This was manifested in sEMG response induced by passive knee movements, as sinusoidal torques can occasionally induced phasic sEMG and spasms in individuals with SCI . The exiReflex responses in the presence of patterned stimuli including rhythmic movements and vibrations have been investigated on multiple occasions , 152–162Some motor units may show prolonged, contraction-induced firing after the voluntary contraction. This induced motor unit firing may last for minutes—also denominated as unit spasms . The exaMultiple types of sEMG analyses were used to link muscle activity and spasticity measured through the Ashworth scale. Some positive correlations were found by linking sEMG amplitude-based and timing properties, e.g. mean, peak, and the time between onset and peak of electrical activity ; predictSubsequent studies reported weak correlations between the Ashworth scores and reflex activity, and a unique reflex mechanism in SCI was proposed . SimilarStudies on the time-domain sEMG properties at rest are lacking in SCI, which is understandable since time-domain features were first proposed for the active myoelectric control of upper-extremity prosthetic devices , 117.Frequency-domain sEMG properties are highly susceptible to noise given the low signal power in sEMG recordings at rest. For example, the investigation of sEMG activity during passive static stretch showed characteristics of “white noise” in the power density spectrums at rest with a median frequency of 400–548 Hz (AB controls) and 478–540 Hz (SCI) without a characteristic concentration of frequencies in the spectrum, and with strikingly low power .sEMG analysis at rest may provide information on the rate of involuntary motor unit firing and on whether medications can dampen such activity. Few methods have been proposed to identify spontaneous motor unit firing in SCI using sEMG. An automated multi-step classification algorithm of sEMG from paralyzed thenar muscles enabled the detection and classification of spontaneously firing motor units using sEMG . The motThe involuntary recruitment of populations of motor units during clonus has been described in detail in SCI. It was shown that the clonus frequency ranged from 4.7 to 7.0 Hz and the firing of motor units seemed to follow orderly recruitment during these involuntary contractions. This pattern of recruitment is consistent with motor unit recruitment seen during many voluntary contractions, and suggests the importance of spinal mechanisms in the control of motor unit behavior in SCI . AnotherIn contrast to the β-band synchronization of UMN signals to the spinal cord, the intrinsic spinal cord circuitry is thought to operate at lower frequencies. Lower frequency oscillation seems to be associated with spasticity during volitional contractions  but alsoThis scoping review aimed to examine the properties of the sEMG following SCI. Given the increasing role of technology and the possibility to assess neurorecovery in more detail, this review aims to support the interpretation of sEMG signals after SCI, facilitate the choice of sEMG methodology for planning and conducting research in SCI, support the development of assistive technologies, as well as highlight gaps in knowledge.Among the identified body of evidence, the minority of studies included sEMG measurements beyond amplitude-based analysis. Most studies employed amplitude-based analysis using RMS (57 studies), normalized sEMG , and qualitative or semi-quantitative scores of the sEMG pattern 16 studies). In contrast, only 19 studies used time-domain or frequency-domain analysis  is widely used to attempt to compensate for these issues, a limitation of most sEMG studies in SCI was the lack of reported electrode placement and between-days reliability of the sEMG measurements. Only 51/178 references reported detailed information about electrode placement, but the majority reported details about the sEMG equipment or filtering/amplification procedures , or with an insufficient number of SCI participants (less than 4 SCI) to be included in this scoping review. On the other hand, the SCI studies with larger samples mostly rely only on amplitude-based analysis. This creates an inverse relationship between the sample size and the variety of sEMG properties reported .In the new era of machine learning, the characterization of the volitional sEMG activity below the lesion has a potential application as a screening tool in the clinical settings , 187. Ad. and Merletti et al., there is a range of factors to consider when implementing sEMG assessments in clinical practice [The utility of sEMG is widely appreciated as it is seen to provide simple and easy-to-use assessments of motor impairments and rehabilitation after SCI. On the other hand, as recently described by Pilkar et alpractice , 189. Fopractice . More br. and Korupolu et al. have described the use and benefits of electrophysiological outcome measures, including sEMG, in the context of SCI clinical trials [From a research perspective, the availability of high-quality outcome measures is essential to the successful translation of new interventions. Recent reviews by Hubli et all trials , 184. ReOur study had some limitations concerning the exclusion of studies on treatment or interventions and the lack of studies encompassing recent advances in high-density sEMG. First, given our interest in how the sEMG properties are changed in response to an SCI, it was considered that any intervention or treatment would have the potential to interfere with the signal characteristics. For example, the use of epidural or transcutaneous spinal cord stimulation may generate electrically induced muscle activation that interferes with the understanding of how the sEMG is changed after an SCI. Other types of treatments were identified, such as pharmacological or rehabilitative . The most common interventions were the use of assistive devices . Although the review of these studies would be important, they are outside the scope of our present question and we suggest that future reviews should provide guidance on appropriate methods to use sEMG to answer specific research and clinical questions .Secondly, our review identified a lack of studies on high-density sEMG and motor unit decomposition in SCI. Studies on high-density sEMG were mostly conducted in AB participants or in individuals with an SCI but with insufficient sample size to be included in the present review. We believe that the identification of this knowledge gap is an important finding of this scoping review and should further well-sampled and comprehensive studies on how these novel techniques can be applied in SCI. Recently, novel methodological advancements in high-density sEMG preprocessing have been proposed to enhance the diagnostic power in individuals with an SCI, which will help to develop a standard sEMG preprocessing pipeline . In Fig.An additional limitation is the focus here on limb and trunk muscles. The inclusion of respiration and sphincter muscles would warrant further work, as the differences in neural control and muscle properties could be expected to alter the sEMG properties.The research on sEMG in SCI over the past seven decades has accumulated abundant evidence about changes in the sEMG properties after the injury. Most of the studies describe muscle weakness, coordination and spontaneous activity using sEMG amplitude properties. It is known that sEMG can capture the residual motor command in great detail, including in muscles below the level of injury with seemingly absent motor activities. Therapies promoting sensorimotor recovery aim at harnessing these residual supraspinal inputs to increase muscle strength and coordination while reducing spontaneous activity. Thus, the inclusion of sEMG assessments in the clinical setting affords important information on how novel therapies may engage and optimize the residual motor command. Nonetheless, current gaps include the lack of studies reporting changes in sEMG properties beyond the amplitude measurement. In order to advance the field, we suggest the incorporation of a broader range of signal properties into the neurophysiological assessment post-SCI and the development of a greater understanding of the relation between these sEMG properties and underlying physiology.Additional file 1: Figure S1. Additional instrumentation used in all studies (a), studies assessing sEMG properties at volitional effort (b) and rest (c). Table S1. Search strategy.


Mean biceps femoris eccentric and concentric sEMG ranged from 27 to 50%, although without significant differences between drop heights. Mean gastrocnemius medialis eccentric and concentric sEMG remained relatively constant (∼60–80% MVIC) across DJs heights, although eccentric values reached 90–120% MVIC from DJ75 to DJ90. Mean variations of ∼50–100% MVIC for eccentric and ∼50–70% MVIC for concentric sEMG activations were observed in the vastus medialis across DJs heights. The biceps femoris eccentric/concentric sEMG ratio during DJ45  was lower (p = 0.03) compared to the ratio observed after DJ90 . The gastrocnemius medialis and vastus medialis eccentric/concentric sEMG ratio were not significantly different between drop heights. In conclusion, jumping performance and most neuromuscular markers were not sensitive to DJ height (intensity) in amateur female volleyball athletes.The aim of this study was to assess drop jump (DJ) performance variables  concomitant to surface electromyography (sEMG) of lower limb muscles during DJs from different drop heights (intensities). The eccentric and concentric phase sEMG from the gastrocnemius medialis, biceps femoris, and vastus medialis muscles were assessed during all tests, with sEMG activity normalized to maximal voluntary isometric contraction (MVIC). In a cross-sectional, study, 10 amateur female volleyball players  completed DJs from six heights [15–90 cm (DJ15 to DJ90)]. During DJs there was no jump-target box to rebound on to. Results of one-way analysis of variance (ANOVA) showed that the jump height, contact time, and reactive strength index were not significantly ( Volleyball is a team sport characterized by periods of short duration , high-intensity activities, interspersed with periods  of recovery . Althougvia stimulation of the muscle spindle , consequently allowing more work to be performed in the concentric phase , libero (n = 1), spiker (n = 5), and opposite-spiker (n = 2). Athletes were recruited during the competitive period, where they usually completed one regional-level competition per week. Athletes participated in regular volleyball training sessions for 2 h per day, 3 days per week, during 3 months prior to inclusion in this study. All athletes had ≥2 years of regular training and competition experience in volleyball. At recruitment in this study, most of the volleyball training practice was devoted to technical and tactical drills. Moreover, the participants had no regular experience with resistance training or structured plyometric training in the 3 months preceding this study, particularly with DJs. Inclusion criteria were: (i) healthy by self-report ; (ii) completion of an exhaustive health questionnaire; (iii) without injury history in the past 3 months prior to testing (confirmed by checking their training logs with the team head coach). Exclusion criteria were: (i) any condition considered to affect muscle function  or the measurement protocol ; and (ii) being on medications considered to affect dependent variables . The methods and procedures used were approved by the local university (CODE: N°155/2018-code 195.18) and were based on the latest version of the Helsinki declaration.Ten female amateur volleyball players  participated in this study. The participants positions were as follow: middle-blocker  test on day one and a series of DJ tests on the next familiarization session, with ≥48 h from previous MVIC testing. During testing sessions, the participants performed the MVIC and (after 5 min of recovery) DJs on the same day (avoiding repositioning of the electrodes). The day before a testing session, participants were instructed to perform a low-intensity workout and maintain their dietary routine. A standardized warm-up was completed before each testing session including 5 min of free running and 5 min of dynamic jumps and dynamic stretching . All jum–2).Standing height (m) and body mass (kg) were assessed using a stadiometer/mechanical scale  with precisions of 0.1 cm and 0.1 kg, respectively. Subjects were tested in light clothing (shoes were removed). The body mass index was calculated from the two measurements   see , with 1 tes) see . For all–1), as previously described (p > 0.05 between DJ heights). Considering that maximal jumping performance may be more ecologically valid for volleyball players, the best performance trial  was used for statistical analysis as previously suggested  see , and parescribed . Playersescribed . They haescribed . Of noteuggested .1 The electrodes were secured with adhesive tape . The sEMG signals from each electrode were amplified  . All data were centered and filtered with a 10 Hz high-pass and a 500 Hz low-pass second order infinite impulse response (IIR) Butterworth filter.The sEMG data was acquired  and used to quantify muscle activity. After warm-up the skin was carefully shaved, abraded and cleansed with alcohol prior to application of EMG + IMU sensor , whose sample rate of the EMG signal is 1950 and 148 Hz for the accelerometer. As previously suggested  the elecf 10 mm)  and gainThe root mean square (RMS) was used to assess sEMG recorded during jump testing . Data wewhere Xn is the value of the sEMG signal and n is the sample number. During DJ the eccentric and concentric sEMG activity was recorded, coupled with an inertial measurement unit to detect each phase . To this aim, the IMU was placed at 1/3 on the line between the tip of the fibula and the tip of the medial malleolus. Behavior analysis of the IMU acceleration signal during a DJ was conducted in a pilot session before the testing sessions. After the application of 10 Hz finite-impulse-response (FIR) filter, signal rectification, and low-pass filtered with a 5 Hz FIR filter, both using a Blackman window , an initial peak at the beginning of the DJ was noted, corresponding to initial DJ ground contact (start of the eccentric phase), visually determined as the increase in the amplitude of the acceleration (above baseline). Thereafter, the peak progressively descended until return to baseline . Another peak was observed after the flight phase of the DJ. Using data from contact time and flight time, calculations allowed the identification of the eccentric and concentric phases. From these recordings, eccentric/concentric ratio was calculated. Each RMS sEMG data was expressed as a percentage of MVIC  using thThe data used in the statistical analyses is available from the corresponding author upon reasonable request.n < 50) and Levene tests, respectively analyzed the normality and homoscedasticity of the outcome variables. When the data did not confront to the test for normality and/or homogeneity of variance, they were log transformed and were back transformed for presentation purposes.The Shapiro–Wilk  reliabilities of all dependent variables were assessed by calculating intra-class correlation coefficients and coefficient of variation and the associated 95% confidence intervals. As previously suggested, intra-class correlation coefficients values <0.5, between 0.5 and 0.75, between 0.75 and 0.9, and >0.9 were interpreted as poor, moderate, good, and excellent reliability, respectively, based on the lower bound 95% confidence interval .f) = 0.6–1.2  were determined by calculating partial eta-squared .A series of repeated measures one-way ANOVA tests were used to compare dependent variables collected during the DJ. When a significant outlined  and inteoutlined . SignifiFor the DJ performance variables contact time, jump height, and reactive strength index, the intra-class coefficients were calculated, for each of the six drop heights, yielding 18 calculations, and 8 demonstrated good reliability, 7 moderate reliability, and 3 poor reliability, based on the lower bound 95% confidence intervals for intra-class correlation coefficients . The gen–1 for reactive strength index, without significant differences between any drop heights (The lowest contact time was achieved in the DJ30 (375.2 ± 41.7 ms), and the highest was achieved in the DJ90 (460.6 ± 50.0 ms), with a difference of ∼23%, although no significant  differences were observed between drop heights . Further heights .The sEMG of the biceps femoris during the eccentric phase varied from 32% (DJ75) to 50% (DJ90), although without significant differences between any drop heights . Similarp = 0.03; d = 1.0, moderate) greater ratio was observed after DJ90  , a significant  . The gase., 3.2) . The vase., 3.2) .The main aim of this study was to assess jumping performance and neuromuscular activity in lower limb muscles of female volleyball athletes after DJ from different drop heights, using parameters such as reactive strength index, jump height, contact time, and sEMG. Based on a previous study , we hypoNo significant differences in contact time were noted between DJ15 to DJ90 box heights, whereas a ∼23% increase  in contact time was noted in the DJ90 compared to previous drop heights. Relatedly, the reactive strength index, although did not varied significantly across the different drop heights, the lowest value  was achieved after DJ90, a ∼22% reduction compared to the highest reactive strength index value achieved in the DJ30 . Such reduction almost perfectly mimics the increase in the contact time observed in the DJ90. Athletes commonly perform DJs at increased heights for a greater training-intensity stimulus , with soOf note, although no significant concentric or eccentric sEMG differences were shown between DJ box heights, the highest values of sEMG activity usually occurred from greater DJ box heights, although the jumping parameters  achieved relatively better performance at lower DJs box heights. Reduced jumping performance from higher DJ box drop heights might be related to neuromuscular inhibition, which serves as a protective mechanism to prevent muscle and tendon injury from excessive stress in the muscle-tendon unit . HoweverTaken together, our results suggest that motor unit recruitment does not change across DJ box heights during the eccentric and concentric phases. This observation remains for different muscles  and different muscle actions (agonist and antagonist). Therefore, it is debatable if a greater DJ drop height is indicative of greater intensity. Based on recent results, the 50–100% of maximal countermovement jump height may be the appropriate individual relative drop height for the DJ among volleyball players . Future Some limitations should be acknowledged. Firstly, it would be risky to extrapolate current findings to other highly trained female volleyball groups, especially those with extensive strength and conditioning preparation, particularly resistance training preparation. Secondly, during the DJ testing session, all the players (except one) achieved contact time >250 ms. On one side, such contact time are commonly observed in the jumping actions of volleyball competitions, meaning high ecological validity for the current findings. However, if current findings may be extrapolated to DJ drills involving fast SSC muscle action, or ground foot contact time <250 ms, also referred as bounce DJ , should Our results show that contact time, jump height, reactive strength index, as well as biceps femoris, medial gastrocnemius, and vastus medialis muscles sEMG  were not significantly affected by DJs performed from different drop heights , except for a greater biceps femoris eccentric/concentric ratio from DJ90 compared to DJ45. Therefore, DJ performance variables  concomitant to sEMG of lower limb muscles during DJs from different box heights (intensities) are not sensitive markers to DJ box height (intensity) in amateur female volleyball athletes. In other words, increasing the box heights for DJs may not induce greater training stimulus .The original contributions presented in the study are included in the article/supplementary material, further inquiries can be directed to the corresponding author/s.The studies involving human participants were reviewed and approved by the local university  and were based on the latest version of the Helsinki declaration. The patients/participants provided their written informed consent to participate in this study.All authors made significant contributions, including preparation of the first draft of the manuscript, data collection, analysis of data, interpretation of data, and provided meaningful revision and feedback. All authors read and approved the final manuscript.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article, or claim that may be made by its manufacturer, is not guaranteed or endorsed by the publisher.


Transfusion of granulocyte concentrates (GC) is an alternative therapy for neutropenic patients with life-threatening infections. While neutrophils are the main source of antimicrobial activity, only neutrophil numbers are used to certify GCs. The objective of this study was thus to functionally characterize neutrophils in GCs prepared by leukapheresis from G-CSF-stimulated donors and compare to the less characterized prednisone GCs. GCs prepared from healthy donors stimulated with prednisone and then G-CSF after a 6-month washout period were analyzed prior to and after leukapheresis, and after storage. Leukocyte composition, neutrophil viability, calcium mobilization, chemotaxis, phagocytosis, reactive oxygen species, cytokine production and metabolites were determined. G-CSF GCs contained significantly more neutrophils than prednisone GCs of which 40% were immature. In comparison to non-stimulated healthy donor neutrophils, prednisone GC neutrophils exhibited enhanced phagocytosis and G-CSF GC neutrophils showed decreased chemotaxis but increased IL-8 production. Leukapheresis altered prednisone GC neutrophil responses. Storage had a significant, negative impact on G-CSF GC neutrophils compared to prednisone GC neutrophils. G-CSF and prednisone GC neutrophils thus differ in maturity and function, and G-CSF GC neutrophils are more sensitive to storage. Functional testing of GC neutrophils and better storage conditions would improve the quality of this blood product. Life-threatening infections are a major health concern due to growing resistance to antimicrobial and antifungal therapies . This is10/transfusion) healthy donors are stimulated with G-CSF and/or a corticosteroid. In Canada, all GCs are prepared from prednisone-stimulated donors by leukapheresis, whereas in the US GC donors are stimulated with G-CSF (Granulocyte transfusions (GTXs) temporarily increase neutrophil count until the bone marrow restores granulopoiesis . To harvth G-CSF . In EuroNeutrophil anti-microbial activity encompasses several effector functions including chemotaxis, phagocytosis and pathogen destruction by antimicrobial peptides and reactive oxygen species (ROS). In addition, neutrophils sequester and destroy pathogens by releasing extracellular traps (NETs) composed of chromatin, histones and intracellular proteins , 10. CytWhile GCs are transfused to temporarily provide antimicrobial defenses, only the absolute neutrophil count (ANC) is used to certify GCs for transfusion . This is® 6% B. Braun Medical Inc.) to enhance red blood cell sedimentation at a ratio of 13:1 (product:anticoagulant solution). GCs were sent by the Globule Laval collection center to Québec City Héma-Québec for irradiation at 25 Gy prior to delivery to our laboratory for analysis on the day of collection (D1) as well as 24 h (D2) and 48 h (D3) post-collection. GCs were stored at room temperature without agitation. A pre-leukapheresis blood sample was drawn from donors in acid-citrate-dextrose (ACD) (BD Vacutainer) tubes. Blood sample and GC composition was determined at the Héma-Québec collection center with a cell counter . One donation was excluded from the functional analysis because of a change in the sedimentation agent during the first GC donation. For comparison, blood donations from unstimulated frequency matched healthy donors was drawn in ACD tubes by the Clinical Research Platform at the CHU de Québec-Laval University and were used as control.Ten healthy donors were recruited by Héma-Québec for two GC donations (the first after prednisone stimulation and the second after G-CSF stimulation) separated by a 6-months wash out period prior to leukapheresis with SpectraOptia (Terumo) according to Héma-Québec guidelines. All donations were collected in blood collection bags containing sodium citrate (46.7%) and hydroxyethyl starch . Fura-2- acetoxymethyl ester (Fura-2AM), CM-H2DCFDA, calcein-AM, the 7AAD viability staining solution and the pHRodo Red Zymosan A Bioparticles kit were obtained from Thermo Fisher ; dextran T500, cytochrome C, Hemacolor ® Rapid staining kit from Sigma-Aldrich ; lymphocyte separation medium, RPMI-1640, BSA  and FBS  from Wisent Bioproducts ; and the ChemoTx microplates (101–8) from NeuroProbe .2+-free HBSS containing 1.6 mM CaCl2 at 10 x 106 ml−1. Neutrophils isolated from GCs did not require dextran sedimentation as they already contain hydroxyethyl starch, a sedimenting agent. The mononuclear cell layer was harvested after density gradient separation, washed and resuspended in PBS at 10 x 106 ml−1.Pre-leukapheresis circulating neutrophils were isolated by density gradient from peripheral blood at room temperature under sterile conditions as described in Fernandes et al. . Neutrop6 ml−1, stained at room temperature and kept on ice until analysis. All assays are explained in detail in the A LSRII flow cytometer was used for immunophenotyping with a twelve antibody panel and for fluorescent non-opsonized zymosan phagocytosis analysis. A CantoII flow cytometer was used for viability and intracellular ROS production assays. Cells were always used at 10 x 10Ten healthy donors that met the exclusion and inclusion criteria in 9.L−1 and 109.5 x 109.L−1 leukocytes, respectively; per transfusion of 1010 neutrophils was thus observed in 6/9 (67%) of prednisone GCs compared to 10/10 G-CSF GCs  and immature (40% of CD10low) neutrophils into the circulation  compared to their CD16high mature counterparts. The expression of the other cell-surface markers tested in prednisone and G-CSF-mobilized neutrophils was not altered . An. Anhigh) altered . TogetheSince neutrophils are the main source of antimicrobial activity in GCs, we assayed the key neutrophil functions required to fight infections in GC neutrophils 6 h post-leukapheresis. For comparative purposes, the assays were also performed on neutrophils from unstimulated, healthy donors. Of these neutrophil responses, the function that was significantly altered in prednisone GC neutrophils was the phagocytosis of non-opsonized zymosan. While considerable variability in the phagocytosis was observed amongst non-stimulated healthy donors, prednisone leveled this response to 90% phagocytosis in all donors . A diminG-CSF GC neutrophils also exhibited functional differences with non-stimulated, healthy donor neutrophils albeit for different functions . ChemotaTo determine whether leukapheresis contributes to the functional alterations observed in GC neutrophils, their responses were compared before and after leukapheresis. A significant diminution in the fMLF-induced increase in cytoplasmic calcium  and an iWhile the negative effect of storage on G-CSF GC neutrophil viability and function was reported in several studies , 26, 27,(data not shown).As observed for prednisone GC neutrophils, G-CSF GC neutrophils were also affected by storage. The most striking effect was the difficulty in isolating neutrophils from these GCs. The neutrophil yield dropped by 40% after 24 h storage and 80%, 48 h post-leukapheresis . The funA change in metabolite concentration and/or pH could explain the observed negative effects of storage on GCs. Data for two prednisone GC donations and for all G-CSF GCs was generated as the metabolite measurements began later in the study. We consistently observed a decrease in pH  24 h post-collection in G-CSF GCs as well as glucose with a concomitant 5 to 8-fold increase in lactate concentration during storage . TogetheA diminution in most neutrophil responses including calcium mobilization, fMLF-induced chemotaxis and the phagocytosis of non-opsonized zymosan by prednisone GC neutrophils was observed after 24 h storage but did not reach significance . In contComparison of neutrophil functional studies performed on GCs prepared by different blood centers is challenging due to differences in GC preparation and analysis. Moreover, there are few reports on prednisone GCs. To address these gaps in our knowledge of GCs, we compared prednisone and G-CSF GC neutrophil viability and function in a single center study and on the same donors to minimize differences in GC production and inter-donor variability. Major differences in ANC, neutrophil maturity, functional capacity and response to storage were observed between prednisone and G-CSF GCs. To our knowledge, this is the first comprehensive study that compares the antimicrobial defenses of prednisone- and G-CSF GC neutrophils on the day of collection and during storage.per GC unit for transfusion  and by the Héma-Québec research Ethics Committee (# 2018-012). Written informed consent was obtained from all the participants.AM: significant contribution to performing the research, data compilation and analysis, participated in writing the manuscript, and prepared the figures. M-ÈA, GP, MV, LB, M-PC, JV, PL, and M-ML: all these co-authors contributed to performing the research and data compilation and part of the data analysis. NR: contributed to the data analysis and manuscript review. DB: contributed to the conceptualization, data analysis and manuscript review as well as funding acquisition. MG: contributed to the supervision of the personnel, conceptualization and methodology of the study, data analysis, manuscript review, and funding acquisition. MF: funding acquisition, resources and project administration, major contribution to the conceptualization and methodology of the study, the manuscript writing as well as review, editing, visualization, and supervised the personnel. All authors contributed to the article and approved the submitted version.This research was supported by funding from Héma-Québec and the Canadian Blood Services awarded to MF, DB, and MG. A MITACS/ Héma-Québec scholarship was awarded to AM (IT5795).The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article, or claim that may be made by its manufacturer, is not guaranteed or endorsed by the publisher.


Mycobacterium tuberculosis (M. tuberculosis) uses multiple strategies to escape host defenses. The M. tuberculosis Rv0927c protein is predicted to be a short-chain dehydrogenase/reductase related to bacterial metabolism. However, the role of Rv0927c during M. tuberculosis infection remains unclear. Here, we observed that Rv0927c inhibited the expression of IL-6, TNF-α, and IL-1β, an effect dependent on NF-κB and p38 pathways. Western blot analysis of macrophages infected with recombinant Mycobacterium smegmatis strains showed that Rv0927c attenuated NF-κB activation by downregulating the phosphorylation of IκBα. Additionally, Rv0927c enhanced intracellular survival of M. smegmatis and pathological effects in mice. In conclusion, our findings demonstrate that Rv0927c functions as a regulator of inflammatory genes and enhances the survival of M. smegmatis.Through long-term coevolution with its host,  Mycobacterium tuberculosis (M. tuberculosis) complex (Tuberculosis (TB) is a communicable disease caused by  complex . TB led  complex . Macroph complex , 4. TherM. tuberculosis infected mice  . When lited mice –13.M. tuberculosis infection (M. tuberculosis can suppress the expression of proinflammatory cytokines by interfering with a range of processes in NF-κB and MAPKs pathways. Rv0222 utilizes E3 ubiquitin ligase anaphase promoting complex (APC) subunit 2 (ANAPC2) to induce K11-linked ubiquitination. K11-linked ubiquitination of Rv0222 promotes the recruitment of protein tyrosine phosphatase SHP1 to TRAF6, which prevents TRAF6 activation, followed by the inhibition of proinflammatory cytokine gene expression  were obtained from C57BL/6 mouse bone marrow as described previously  was cultured in LB medium for DNA cloning. M. smegmatis strain mc2 155 (Our laboratory) and recombinant M. smegmatis were cultured in Middlebrook 7H9  liquid medium containing 0.05% (v/v) Tween 80 with shaking or on Middlebrook 7H10  agar supplemented with 0.5% (v/v) glycerol.RAW264.7 cells  were cultured in complete DMEM  containing 10% Fetal Bovine Serum , 100 U/ml streptomycin and 100 U/μl penicillin (Gibco) at 37°C and 5% COeviously . BMDMs w7 CFU of recombinant M. smegmatis strains per animal (five mice per group). Spleens, livers and lungs of mouse were collected after infection at 3 days or 6 days. Tissues were weighted and homogenized for CFU analysis. The samples of mouse livers were fixed in 10% neutral formalin fix solution and stained with hematoxylin and eosin (H&E) for evaluation of pathologic changes.C57BL/6 mice were obtained from the Comparative Medical Center of Yangzhou University  and kept in specific pathogen-free conditions, in the mouse isolators . All animal experiments were approved by the Animal Welfare and Ethics Committees of Yangzhou University and complied with the guidelines of the Institutional Administrative Committee and Ethics Committee of Laboratory Animals (IACUC license number: YZUDWLL-201811-001). Six to eight weeks old C57BL/6 mice were used for BMDMs preparation. Six weeks old C57BL/6 mice were intraperitoneally infected with 5×10Rv0927c gene was amplified from the genomic DNA of M. tb H37Rv by PCR using appropriate primers listed in Rv0927c and pCMV-Rv0927c. Then, the recombinant plasmid pMV261-Rv0927c was electroporated into M. smegmatis mc2155, while pCMV-Rv0927c was transformed into Escherichia coli (E. coli) DH5α cells by heat shock according standard procure. Expression of Rv0927c was confirmed by immunoblotting.The full-length of Rv0927c gene was synthesized and ligated into pET30a(+) expression vector . Recombinant plasmid was transformed into E. coli BL21(DE3) cells and transformed E. coli cells were induced to express recombinant protein. The rHis-Rv0927c protein was purified with a His-binding purification kit . Endotoxins were removed from Rv0927c protein by ToxinEraser™ . Endotoxin level of Rv0927c was <0.05 EU/μg determined by ToxinSensor™ (GenScript).Rv0927c were shaken at 180 rpm and 37°C in 7H9 medium with a starting absorbance (OD600) of 0.02. The OD600 of the culture was continuously monitored at an interval of 6 h over 60 h growth period. Subcellular localization of the Rv0927c protein was determined by protocols previously described  of 10 or with M. smegmatis (MOI 1/10) and treated with recombinant Rv0927c protein (5 μg/ml). After 2 h, cells were washed three times with PBS and incubated again with the DMEM medium supplemented with 10 μg/μl gentamicin to kill the bacteria outside macrophages. After 12, 24 and 48 h infection, cells were lysed by 0.05% Triton X-100 for intracellular bacterial counting. The cell lysates were serially ten-fold diluted and then plated on 7H10 agar plates and the colonies were numerated after 3 days.RAW264.7 cells and BMDMs were seeded into 24-well plates at 5×10After infection for 6, 24 and 48 h, cell culture media were collected for quantitative determination of the lactate dehydrogenase (LDH) activities by LDH Cytotoxicity Assay Kit  and the proinflammatory cytokines IL-6, TNF-α and IL-1β by employing BD OptEIA™ Mouse IL-6 ELISA Kit (BD), BD OptEIA™ Mouse TNF ELISA Kit (BD) and Mouse IL-1 beta/IL-1F2 DuoSet ELISA  according to the manufacturer’s manual. Total mRNA from infected mice tissues was extracted using the RNeasy Plus Mini kit , followed by reverse transcription. mRNA levels of proinflammatory cytokines IL-6, TNF-α and IL-1β were determined by RT-PCR using primers listed in Myc, pCMV-Rv0927c and NF-κB or AP-1 reporter plasmid (250 ng), as well as the control plasmid pRL-TK (50 ng) by using Lipofectamin® 3000. Cells were harvested to determine the luciferase activities by Dual-Luciferase Reporter Assay System , following infection with M. smegmatis. Results were normalized by pRL-TK-derived Renilla luminescence.We transfected RAW264.7 cells with pCMV-Myc or pCMV-Rv0927c. Following transfection for 24 h, cells were stimulated by LPS (100 ng/ml) for 15 min. In the protein assay, HEK293-TLR4 cells were treated with Rv0927c (5 μg/ml) for 2 h followed by LPS for 15 min. Then p65 was stained using primary rabbit anti-p65 antibody  and secondary Alexa 488-conjugated antibody (Abcam), while nuclei were stained using DAPI (BD). All samples were observed using Leica TCS SP8 STED fluorescence microscope .HEK293-TLR4 cells  were transfected with pCMV-M. smegmatis strains at MOI of 10 for indicated times and lysed in Western blot sample buffer. Cell lysates were loaded onto SDS-PAGE and the separated protein were transferred to a polyvinyldifluoride membrane. Membranes were blocked with 5% skim milk for 1 h at room temperature, followed by overnight incubation with primary antibody at 4°C. Next day, the membranes were incubated at room temperature for 1 h with HRP-conjugated secondary antibodies. Images were visualized with ECL chemiluminescence substrate  using Amersham Imager 600 Imaging System . The intensity of the bands was analyzed by Image-J and normalized by β-actin.Macrophages were infected with recombinant The primary antibodies used in this study were as follows: anti-Phospho-IκBα antibody , anti-p65 antibody , anti-Phospho-p65 antibody , anti-SAPK/Jnk antibody (9252S), anti-Phospho-SAPK/Jnk antibody , anti-p44/42 MAPK (ERK1/2) antibody , anti-Phospho-p44/42 MAPK (ERK1/2) antibody , anti-p38 MAPK antibody , anti-Phospho-p38 MAPK antibody , anti-Phospho-TAK1 antibody , anti-TAK1 antibody , anti-IRAK1 antibody  and anti-β-actin antibody . The secondary antibodies were goat anti-mouse IgG-HRP  and goat anti-rabbit IgG-HRP .t-test was used to assess differences between groups. Statistical significance was determined at p values of < 0.05 (*), < 0.01 (**).Prism V6.01 software  was adopted for statistical analyses. Data were shown as mean ± SEM and a two-tailed unpaired M. tuberculosis gene Rv0927c encodes a 27 kDa protein. This study, we successfully constructed an Rv0927c-overexpressing M. smegmatis strain (rMS::pMV261-Rv0927c)  and rMS::pMV261-Rv0927c were monitored. We found no significant difference in growth kinetics between the two recombinant strains  luciferase activity in macrophages stimulated by Rv0927c  were determined. The bacterial load of macrophages infected with rMS::pMV261-Rv0927c was higher than that of macrophages infected with rMS::pMV261 , the Science and Technology Innovation Cultivation Program of Yangzhou University (2019CXJ158), the independent program of Jiangsu Key Laboratory of Zoonosis (RZZ202003) and the Qinglan Project and Priority Academic Development Program of Jiangsu Higher Education Institutions (PADP).The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article, or claim that may be made by its manufacturer, is not guaranteed or endorsed by the publisher.


Efficient light‐induced floral stimulation plays a key role in energy conservation and maintaining stable productivity during off‐season periods of dragon fruit plants. In this study, we first reported on results of a survey on dragon fruit farmers regarding use of lamps in performing artificially induced flowering process in Vietnam. It was found that the use of incandescent lamp was prevalent in dragon fruit cultivation practices, resulting in heavy electricity consumption, and that low‐power compact fluorescent light (CFL) bulbs were not extensively utilized, possibly due to low floral induction performance of domestic CFL bulbs. Arguing that emission spectra of currently used lamps were not consistent with adsorption spectra of phytochromes, whose transformation is responsible for flowering process of dragon fruit, we then proposed three improved CFL lamps (power capacity of 20 W) having emission spectra focused on red and far‐red regions. New lamp prototypes were tested in 7 field experiments in three different provinces in Vietnam. One improved CFL bulb (treatment 2) performed relatively well in comparison with the incandescent control lamp (60 W) in six out of seven experiments with regard to some growth indicators  and fruit yield. Recent success on commercialization of the improved CFL lamp demonstrates the potential of CFL lamps in floral stimulating irradiation of other crops and plants and in alleviating electricity burden in dragon fruit growing areas. The study first surveyed on dragon fruit farmers on measures to stimulate flowering in commercial dragon fruit farming in three provinces of Vietnam. We then modified emission spectra of Compact Fluorescent Lamps and used them in field experiments. One improved low‐power CFL bulb showed flowering induction capability comparable to the incandescent bulb. It was concluded that even 9 hr of lighting extension after sunset induced no effect on flowering, possibly due to the subtropical climate in the study area (Israel). This explanation was further confirmed by Jiang et al.  and was conducted in different regions having varying temperature. As a result, obtained results are contradictory and carry little implication when being implemented in regions with different climatic conditions. Driven by such shortcomings and the high electricity consumption in areas with intensive dragon fruit cultivation in Vietnam, this study aims to rationalize a low‐capacity compact fluorescent lighting (CFL) lamp that is specifically designed for controlling off‐season flowering of dragon fruit. We first presented the results on the current measures for controlling off‐season dragon fruit flowering in three typical areas in Vietnam, namely Tien Giang, Tay Ninh, and Binh Thuan Province. Then, some CFL configurations were proposed and analyzed for their light spectra. Lastly, the selected CFL lamps were then used in experiments carried out in these areas to assess their stimulation effectiveness on some growth indicators of dragon fruits of both red‐flesh and white‐flesh varieties. Current results are expected to contribute to alleviate electricity burden for agricultural activities in those areas and aid in further development of CFL lamps specialized for fruit irradiation.22.1A questionnaire was designed aiming at collecting relevant information on dragon fruit farming and techniques for flowering stimulation from farmers in Vietnam. Some typical categories include type of lamp, irradiation process, irradiation duration, and bulb density per hectare. A total of 100 respondents were selected from the farmer lists prepared by the local authorities in three provinces, including Binh Thuan (50 households), Tien Giang (25 households), and Tay Ninh (25 households) according to probabilistic randomized sampling procedure. We also collected lamp samples that were being used for floral stimulation from local farmers for further analysis.2.2A PG100N Handheld Spectral PAR Meter  connected to a computer was used to measure emission spectra and determine photosynthetically active photon flux density (PPFD), photon flux density in the red region , and photon flux density in the far‐red region  of lamps. The testing lamp was positioned 30 cm in front of the instrument sensor in the dark. The procedure was carried out at Laboratory of Agricultural Biotechnology, Nguyen Tat Thanh University, Vietnam.2.3Based on the survey results and spectrum analysis, we designed new CFL bulbs with spectra focused in the red and far‐red regions. The light bulbs were manufactured by Rang Dong Light Source & Vacuum Flask Joint Stock Company (Vietnam) and were screened before being used in field experiment.In the field experiment, three experimental CFL bulbs (20 W) selected from the spectra analysis results were used as treatments (denoted from 1 to 3) and one commercial incandescent lamp (60 W) acted as the control bulb (denoted as control). No lighting and domestic CFL lighting were not included as controls due to zero or very low productivity resulted from the adoption of the two options. In addition, Yamada et al.  also maiHylocereus undatus) and red‐flesh pitaya (Hylocereus polyrhizus) that were normally bearing fruits with the age ranging from 5 to 8 years. The experiment was carried out in household fields located in three provinces with largest dragon fruit production in Vietnam, namely Binh Thuan, Tien Giang, and Tay Ninh  confirmed that, from September to March of the following year, dragon fruit plants cultivated in these areas were unable to bloom under natural conditions. In fact, this is also the period in which the daytime is shorter than 12 hr. As a result, a majority of farming households adopted artificial irradiation as a measure to stimulate flowering of dragon fruit crops. Prior to 2012, off‐season floral stimulation of dragon fruit often involved incandescent bulbs with the capacity of either 100 W, 75 W, or 60 W to supplement lighting during nighttime. In the years 2013–2014, there was a shift in the use of bulbs for dragon fruit flowering induction toward incandescent bulbs with lower capacity (40 W) and compact fluorescent bulbs (20 W) to save electricity. To gain further insights into farmers' behavior in dragon fruit farming, a typical sample of 100 farming households located in the three regions was surveyed for their use of light bulbs in off‐season harvests. Table On the whole, it was showed that all surveyed household relied on artificial lighting to control blossoming of both white and red‐flesh dragon fruits. Incandescent light bulbs with power capacity of 40 W and 60 W were the dominantly used lamps for the floral stimulation while the households that used CFL bulbs (20 W) only accounted for around 10% of the total surveyed respondents. Moreover, no other types of bulbs were recorded for dragon fruit irradiation purpose. Measures aimed to improve energy efficiency of irradiation, such as intermittent lighting were not adopted . Tay Ninh is also the province in which many households adopted the combinational irradiation with both incandescent and CFL light bulbs. In addition, there were 5 out of 25 surveyed households in Tay Ninh reporting the use of incandescent bulbs with lower capacity of 40 W. Higher adoption of CFL in Tay Ninh could be explained by the recently emerged inclination to commercially grow dragon fruit in the province, as opposed to the other two regions where cultivation traditions have already been established; thus, the local farmers are hesitant to endorse new, improved growing techniques.Another notable feature is that growers in Binh Thuan province started their artificial lighting earlier than those in Tay Ninh and Tien Giang. Specifically, the latest time in Binh Thuan in which irradiation started was 19:00 p.m. meanwhile 100% of farmers in Tien Giang and 92% of surveyed Tay Ninh growers commenced their lighting 2 hr later, at 21:00 p.m. Two households who were able to begin early irradiation in Tay Ninh also reported the use of CFL bulbs, which are less energy consuming than incandescent counterparts, in their fields. This is reportedly due to delayed provision of electricity source reserved for agricultural activities and possibly reflects the inadequacy between grid capacity and production readiness of the electricity supplier and the recent, rising energy needs for dragon fruit cultivation in these areas.Regarding lamp density, most growers allocated around 1,000–1,200 bulbs per hectare of cultivation. Higher distribution densities were only implemented in the harvest season from January to March where dragon fruits face difficulties in blooming. Farmers also reported that, during this time period, white‐flesh dragon fruit plants were less susceptible to light‐induced floral stimulation than red‐flesh plants and in some cases in Binh Thuan, a denser distribution of more than 2,000 bulbs per hectare was required. With respect to lamp use specific to dragon fruit variety, there was no indication suggesting light bulb preference against certain variety. In Binh Thuan, 45 out of 50 surveyed growers specializing in white‐flesh variety mainly utilized incandescent lamps for floral stimulation. Of which, incandescent bulbs with capacity of 60 W and 40 W accounted for nearly 90% and 6%, respectively. In two areas specialized in cultivating red‐flesh dragon fruit, the percentages are also similar in which only 4% of households adopted CFL lamps.Overall, from the survey results, it is indicated that efficient control of off‐season blossoming is an important concern in commercial dragon fruit farming and that possible improvements should be called for in efforts to conserve energy for such processes. Currently, all dragon fruit farmers adopted artificial irradiation as the main measure for controlling off‐season flowering and ruled out other stimulation methods such as chemical intervention and foliar fertilization. Moreover, the irradiation process was carried out based on farmers' experience, thus lacking considerations on standardizing critical aspects including bulb type, height of bulb hanging, bulb density, irradiation duration, and implementation of intermittent lighting.3.2We collected three types of lamps commonly used by dragon fruit farmers  and measured their emission spectra. The results are shown as in Figure 660 and P730 , followed by the 40 W incandescent lamp (38.0 µmol m−2 s−1). The 20 W CFL lamps usually have total PFD at the lowest value (9.69 µmol m−2 s−1).In terms of PPFD, the 60 W incandescent type exhibited the highest value, at 44.5 µmol mClearly, currently used lamps all had emission spectra focused on red and far‐red regions. Of which, two incandescent lamps showed that the density of the far‐red and red regions were proportional to the bulb capacity and the CFL lamp usually have much lower red and far‐red photon density than incandescent lamps. As a result, the use of domestic CFL lamps might induce weaker floral stimulation effect on dragon fruit cactus than incandescent counterparts, even at similar power capacities. This is in part illustrated by the previous survey results where only 9% of surveyed farmers with CFL irradiation reported the productivity of higher than 10 kg per plant, as opposed to 92% of growers using incandescent lamps reported the same yield. Current results suggest possible improvements in stimulation efficiency could be achieved by manipulating spectral intensity of CFL lamps toward increased far‐red and far‐red photon density.3.3We obtained various improved CFL specimen lamps from Rang Dong Light Source & Vacuum Flask Joint Stock Company, one of leading manufacturers of light bulbs in Vietnam. All bulbs had the power capacity of 20 W and were specifically designed with increased intensity in red and far‐red regions and reduced green and yellow intensity in their emission spectra. After measuring their spectra, we selected three CFL bulbs with highest total PFD and used them in the following field experiment. The spectra and analysis results are shown as in Figure Three improved CFL bulbs were used in field experiments conducted in three main fields located in three typical provinces with commercial dragon fruit farming activities. The experiments spanned across multiple harvest seasons of the year from 2013–2014 and were conducted on the two varieties of dragon fruit. Incandescent lamp with the capacity of 60 W was used as the control. The results are presented in Table Plant height, canopy diameter, and the number of stems per plant before and after the experiment 1 were not significantly different  is statistically comparable to that of the control (19.3) and is higher than those illuminated by two other CFL lamps (15.1 and 12.5). For the indicator of number of buds per plant, a similar trend was found where the control and the treatment 2 both gave almost equal results  and treatments 1 and 3 showed inferior bud‐stimulating efficacy . The better flowering of plants treated with the incandescent and treatment 2 bulb also led to higher fruit counts in those plants compared with the rest . Some flowers were unable to develop into fruits, evidenced by the fact that the number of fruits is lower than the number of buds per plant.Flowering indicators of dragon fruits in the experiment 2 are considerably lower than those in the experiment 1. This is possibly due to the lower temperature in this harvesting season (24.4–24.6°C) compared with the last season (26.9–27.5°C) and is consistent with the experiments of Kim et al.  and of PThe experiment 3 was carried out at the same location and under climatic conditions similar to the experiment 1. The experiment results again confirmed the stimulus efficiencies of newly designed light bulbs in the experiment 1 in which plants treated with the incandescent bulbs and CFL bulb 2 showed higher number of floral stems, buds, and fruits per plant compared with those of other plants. Strikingly, plants illuminated with the control and treatment 2 displayed exceptionally large number of bubs per plant , approximately 46% higher than plants treated with other treatments and were therefore subjected to bud trimming (to around 25–30 buds/plant) to ensure productivity. Ultimately, trimmed plants showed high fruit yield and were among best white‐flesh dragon fruits plants in terms of yield per plant throughout the three examined harvesting seasons.Some interesting observations could be made from experiment conducted on red‐flesh dragon fruit plants in Tien Giang province (experiments 4 and 5). Firstly, dragon fruit plants treated with the incandescent light (control) displayed higher number of floral stems, buds, and in turn fruits per plant than those treated with CFL bulbs and even higher than those indicators of the control group in the experiment 3. This suggests that the influence of lamp type on the floral stimulation efficiency is more profound in red‐flesh dragon fruit than in white‐flesh plants and that the development from bud to flowers of the red‐flesh variety is expedited compared with the white‐flesh counterpart under incandescent irradiation. To date, the mechanism explaining for lower supplemented irradiation required for flowering of the red‐flesh dragon fruit has not been clearly elaborated. Nevertheless, current findings on variety‐specific response to artificial irradiation of dragon fruit are in agreement with previous experimental studies asserting that off‐season flowering of red‐flesh dragon fruit could be more easily stimulated than of white‐flesh variety . To be specific, the number of buds, fruits, and productivity between control and treatment 2 plants displayed no statistical differences. In addition, unlike experiment 4, all plants in this experiment did not require trimming and virtually all buds of dragon fruit plants developed into fruits. This result may carry implication that controlling the number of buds is essential in the light‐induced stimulation process for off‐season fruiting.Similar to experiment 2, experiment 6 was conducted under colder growing condition (24.9–25.9°C), resulting in comparatively poor growth and productivity. The treatment 2 gave plants with higher number of buds than other groups. In some plants in the control group, no budding was observed. Despite that, productivities and bud numbers of the control and treatment groups in this experiment are still higher than those in the experiment 2, confirming that red‐flesh dragon fruit is more responsive to floral stimulation by artificial lighting. Both thermal and light sensitivity of the flowering process in red‐flesh dragon fruit plant have been confirmed by a previous study where a temperature of at least 15°C was recommended for night lighting for the red‐flesh cultivar . Given that the tested CFL lamps have power capacity almost threefold lower than the control, adoption of improved CFL lamps is recommended in energy‐scarce areas with intensive dragon fruit farming activities. In fact, treatment 2 has been commercialized under the code name of CFL‐20 W NN R for flowering stimulation of agricultural crops and ornamental plants in various provinces including Binh Thuan, Long An, Tien Giang, Tay Ninh, Lam Dong, and Ha Noi city since 2016. The product earned good acceptance from farmers due to its affordability and short payback period.4In an effort to reduce energy consumption in dragon fruit farming, this study aimed to propose and evaluate new improved CFL lamps specialized for inducing light‐stimulated off‐season flowering in dragon fruit fields. The preliminary survey results indicated that the use of incandescent light bulbs was prevalent among farmers and that domestic CFL lights were generally inefficient in stimulating off‐season flowering in both white‐flesh and red‐flesh dragon fruit cultivars. Emission spectrum measurement of currently used lamps suggested that the intensity in red and far‐red regions of the lamp might be responsible for inducing phytochrome transformation, which is critical in flowering process of the plant. Field experiments adopting three improved CFL light bulbs in three provinces showed that one CFL light (treatment 2) resulted in better flowering in dragon fruit plants than others, which were demonstrated by better growth, blossoming indicators, and productivity. Future studies on combinational strategies in stimulating flowering in dragon fruits are recommended.The authors declare no conflict of interest.This study was funded by Ministry of Science and Technology of Vietnam under the project “Design and Manufacture of Specialized Irradiation System and Usage Guidelines for Propagation and Flowering Control of Some Crops at Industrial Scale” (Code: ĐM.06.DN/13) belonging to the National Programme for Technology Innovation to 2020.


PRNP genetic variants. CSF PrP was lowered in the presence of rare pathogenic PRNP variants, with heterozygous carriers of P102L displaying 55%, and D178N just 31%, of the CSF PrP concentration of mutation-negative controls. In rodents, pharmacologic reduction of brain Prnp RNA was reflected in brain parenchyma PrP and, in turn in CSF PrP, validating CSF as a sampling compartment for the effect of PrP-lowering therapy. Our findings support the use of CSF PrP as a pharmacodynamic biomarker for PrP-lowering drugs and suggest that relative reduction from individual baseline CSF PrP concentration may be an appropriate marker for target engagement.Prion protein (PrP) concentration controls the kinetics of prion replication and is a genetically and pharmacologically validated therapeutic target for prion disease. In order to evaluate PrP concentration as a pharmacodynamic biomarker and assess its contribution to known prion disease risk factors, we developed and validated a plate-based immunoassay reactive for PrP across 6 species of interest and applicable to brain and cerebrospinal fluid (CSF). PrP concentration varied dramatically across different brain regions in mice, cynomolgus macaques, and humans. PrP expression did not appear to contribute to the known risk factors of age, sex, or common   Prion disease is a fatal neurodegenerative disease caused by misfolding of the prion protein (PrP) leading to a gain of toxic function . LowerinIn previous studies, PrP in human CSF was quantified using a commercially available ELISA assay specific to human PrP –11, as wPRNP genetic variants are risk factors for prion disease , we developed a final assay protocol  using monoclonals EP1802Y for capture and 8H4 for detection, with C-terminal epitopes mapped approximately to residues 218–227 and 182–196 (human codon numbering)  –22. The mbering)  (Supplemmbering) , except mbering) . In braimbering) , and alsmbering) , and plambering) . The assmbering) .n = 28 human brain samples , with PrP almost 10 times higher in parietal cortex (BA7) than in olivary nuclei , and correcting for multiple testing, only minor salivary gland and skeletal muscle showed any evidence of age-dependent expression  from our cohort study at Massachusetts General Hospital  , CSF PrP measured in cross-species ELISA exhibited tight test-retest reliability (mean CV = 11.1%). We therefore focused on each individual’s mean CSF PrP value observed across all visits. We found no evidence for CSF PrP association with sex  . Exquisin = 47 cohort study participants with at least 1 CSF sample available, we examined genotypic differences in CSF PrP by comparing mean CSF PrP levels averaged, for each individual, across all study visits (n = 21), CSF PrP was lower for carriers of P102L  and D178N ; the trend was preserved but non-significant for E200K .In y visits . ComparePRNP mutations.ELISA relies on the presence of 2 intact epitopes on the same protein, so non-reactivity of 1 of our antibodies for 1 of these mutations could give rise to artifactual genotypic differences. The 8H4 epitope has been mapped to a region adjacent to D178N and E200K , and somPRNP: M129V (rs1799990) and a non-coding variant 72 kb upstream of PRNP (rs17327121) implicated as the lead variant for an expression quantitative trait locus (eQTL) in cerebellum . Antibody pair and other key assay configuration parameters were established, and the assay was subjected to a full validation study for rat CSF  compliant with World Health Organization Good Clinical Laboratory Practice Regulations (GCLP) . The assThe exact assay protocol and checklist referred to at the bench while running the assay are provided as Supplemental Appendices 1 and 2. The method is briefly summarized as follows.To prepare biotinylated detection antibody, 1 mg of EZ-Link Sulfo-NHS-SS-Biotin (Thermo Fisher Scientific A39258) was combined with 0.09 mg of 8H4 antibody (Abcam ab61409). Conjugated antibody was purified using Zeba spin desalting columns (Thermo Fisher Scientific 89889) and quantified by NanoDrop. Assay buffer of 0.05% wt/vol Tween (Teknova T0710) and 5% BSA in 1X CSHL PBS was 0.22 μm vacuum-filtered and stored at 4°C. Wash buffer was 0.1% Tween in 1X CSHL PBS, stored at room temperature.Clear flat-bottom MaxiSorp plates (Thermo Fisher Scientific 439454) were coated overnight at 4°C with 2.0 μg/mL EP1802Y capture antibody (Abcam ab52604) in PBS, sealed with clear adhesive MicroAmp Film (Life 4306311) and then washed 3 times with 300 μL wash buffer and tapped dry (subsequent washes followed this same procedure). Plates were blocked with 250 μL assay buffer , sealed at room temperature for 1–3 hours and then washed. A fresh aliquot of recombinant PrP was thawed to make a new standard curve for every plate . Standards, quality control samples (QCs), and samples were diluted into assay buffer in microcentrifuge tubes and 100 μL was added per well. Plates were sealed and incubated with sample for 60–75 minutes and then washed. Biotinylated 8H4 detection antibody was added at 0.25 μg/mL in 100 μL assay buffer; plates were sealed, incubated 60–75 minutes, and then washed. Pierce High Sensitivity Streptavidin-HRP (Thermo Fisher Scientific 21130) was added at 24.69 ng/mL in 100 μL assay buffer; plates were sealed, incubated for 30 minutes, and then washed. 100 μL TMB  was added; plates were incubated in darkness but monitored periodically for absorbance at 605 nm. After 30 minutes or when absorbance for the 5 ng/mL standard reached 0.8, whichever came sooner, 100 μL of stop solution  was added; plates were shaken briefly and then read at 450 nm with 630 nm background subtraction on a Spectramax M5 plate reader (Molecular Devices). Standard curves were fit with a 4-point hill slope curve using the minpack.lm package  in R. FDFor plates prepared in the prion laboratory, the protocol was modified as follows. All reagents, standard curves, and QCs were diluted to working concentrations in the morning before beginning the protocol and were kept at 4°C throughout the day. Instead of tapping dry, wells were washed with 190 μL of wash buffer using a multichannel pipette ejecting waste into a bleach bath. Plates were read at 450 nm with 620 nm background subtraction on a Fluostar Optima plate reader (BMG Labtech).E. coli and purified from inclusion bodies as described previously  peak area ratio to estimate the concentration of PrP in each sample.MRM was performed as described . For rat15N-labeled HuPrP23-231 as the reference standard and light:15N (L/15N) peak area ratio used to calculate PrP concentration. Test-retest analysis utilized CSF pairs taken 2–4 months apart from 5 individuals deliberately selected to include 2 individuals with, and 3 without, CSF processing anomalies, as this affects test-retest reliability for PrP , except where otherwise indicated, in cold 0.2% wt/vol CHAPS, 1X PBS, and 1 tablet protease inhibitor  per 10 mL, using 3 × 40 second pulses on a Bertin MiniLys homogenizer in 7 mL tubes pre-loaded with zirconium oxide beads (Precellys KT039611307.7). Human CSF was collected as described , E200K (n = 12), D178N (n = 6), P102L (n = 4), or other PRNP mutation (n = 4), and each made 1–5 study visits (mean: 2.3) spanning a time period of up to approximately 3.5 years. Immediately after CSF sample collection, 0.03% CHAPS  was added.CSF from asymptomatic escribed . Particin = 5 control individuals without dementia, aged from approximately 50 to 90 years, postmortem intervals of 8–86 hours, n = 4 male and n = 1 female.Postmortem human brain samples were obtained from the Massachusetts Alzheimer’s Disease Research Center (MADRC). Samples were from All mice were C57BL/6. PrP ZH3 knockout mice  on a C57Rats were Sprague-Dawley males  and females . Rat CSF collection was performed under terminal anesthesia as follows. Occipital and nuchal areas were trimmed of hair and wiped with 70% ethanol. The heads of the rats were immobilized in a stereotaxic instrument (ASI SAS-4100) while being maintained on 3% isoflurane and warmed on a heating pad (Physitemp HP-1M). The nose was rotated down 45° and held in this position with the nose bar of the stereotax. A 90° hemostatic forceps (Roboz RS-7291) was depressed against the skin to locate the space between the trapezii and the base of the skull, and a 27G butterfly needle  was held in a custom stereotaxic needle holder and attached to a 1 mL syringe, then inserted through the nuchal skin by lowering the dorsal/ventral knob of the stereotaxic instrument. The plunger of the syringe was withdrawn to create vacuum, and then the needle was lowered further, into the cisterna magna, until CSF began flowing into the butterfly tubing. When CSF flow ceased or blood was observed, the tubing was clamped with a hemostat and, if necessary, the tube was clipped at the meniscus of blood. The syringe was plunged to eject CSF into a low protein binding microcentrifuge tube (Eppendorf 022431081), and 3% wt/vol CHAPS stock solution was added at a 1:100 dilution to yield a final concentration of 0.03% CHAPS.n = 3 male, n = 3 female, age 2–4 years) tissue punches were obtained from tissue archived at –80°C from control animals treated with artificial CSF as part of previous ASO studies.Cynomolgus macaque (Rats were shaved and maintained at 3% isoflurane while being warmed with a heating pad (Physitemp HP-1M). They were placed in a stereotaxic instrument  with 27° atraumatic ear bars , with the rat gas adapter riser set to –6 mm to set the lambda and bregma landmarks flat. The scalp was swabbed with betadine and ethanol and a 1.5 cm midline scalpel incision was made, centered between the nose and occipital ridge. Sterile cotton-tipped applicators were used to retract the subcutaneous and periosteal tissues. A sterile 1 mm × 33 mm drill bit  in a hanging-style handpiece  was positioned above the bregma in a stereotactic handpiece holder  and then moved 1 mm caudal and 1.5 mm lateral. A bore hole was drilled at low speed and then a gastight 1710 small RN syringe (Hamilton 81030) was lowered through the skull hole, 3.7 mm from the surface of the brain into the lateral ventricle; 30 μL of injection solution was then ejected gradually over 10 seconds, and the needle was retracted after 3 minutes. The incision was closed with 5-O monofilament suture (Ethilon 661G-RL), and rats recovered in their home cages.P values were nominal except for GTEx analyses, which were Bonferroni-corrected for the number of tissues studied. P values less than 0.05 were considered significant. All code and all raw data, except for potentially sensitive patient data from the clinical cohort, are available in a public git repository and can be used to reproduce the analyses herein: https://github.com/ericminikel/cns_prp_quant . Skyline files for mass spectrometry data have been uploaded to Panorama at https://panoramaweb.org/mortprpjci2202fz.url under ProteomeXchange submission PXD031432.All analyses utilized custom scripts in R 4.0.4. All statistical tests and all confidence intervals were 2-sided, including those for ANOVA  and 3, SCollection and analysis of human clinical samples were approved by the Partners Institutional Review Board (protocol #2017P000214). Animal studies were conducted under approved Institutional Animal Care and Use Committee protocols .EVM and SMV conceived and designed the experiments. MAM, HTZ, AGR, JEG, EK, JO, PMD, TRC, CM, SWA, BAT, AM, MRM, and DEC performed the experiments. EVM, SMV, HTZ, JL, HBK, JM, DEC, and SEA supervised the research. EVM performed statistical analyses and drafted the manuscript. All authors reviewed and approved the manuscript.