Photon-efficient optical tweezers via wavefront shaping.

Optical tweezers enable noncontact trapping of microscale objects using light. It is not known how tightly it is possible to three-dimensionally (3D) trap microparticles with a given photon budget. Reaching this elusive limit would enable maximally stiff particle trapping for precision measurements on the nanoscale and photon-efficient tweezing of light-sensitive objects. Here, we customize the shape of light fields to suit specific particles, with the aim of optimizing trapping stiffness in 3D. We show, theoretically, that the confinement volume of microspheres held in sculpted optical traps can be reduced by one to two orders of magnitude. Experimentally, we use a wavefront shaping-inspired strategy to passively suppress the Brownian fluctuations of microspheres in every direction concurrently, demonstrating order-of-magnitude reductions in their confinement volumes. Our work paves the way toward the fundamental limits of optical control over the mesoscopic realm.

The Relationship Between the Anaerobic Speed Reserve and Acute Responses to High-Intensity Interval Training in Female Soccer Players.

ABSTRACT: Aspin, GL, Graham, M, Franklin, J, Hicks, KM, and Taylor, JM. The relationship between the anaerobic speed reserve and acute responses to high-intensity interval training in female soccer players. J Strength Cond Res XX(X): 000-000, 2024-The anaerobic speed reserve (ASR) is a popular method of profiling soccer players, often used to individualize training prescription. This study explored the reliability of ASR profiling, and the relationship between the ASR and acute physiological responses to high-intensity interval training (HIIT). Acute physiological responses to different HIIT types were also compared. Thirteen subelite female soccer players aged 20.2 ± 4.6 years completed 6 exercise sessions. In sessions 1-2, players completed a 40-m sprint to assess maximal sprint speed (MSS) and 1600-m time-trial to estimate maximal aerobic speed (MAS), which were used to calculate ASR and assess test-retest reliability. In sessions 3-6, players completed 4 HIIT sessions (repeated-sprint training, sprint interval training, long intervals, and short intervals HIIT). Intensities for long and short intervals HIIT were individualized according to MAS. Ratings of perceived exertion (RPE), heart rate (HR), and postsession blood lactates were recorded throughout. Relationships between the ASR and acute responses to HIIT, and between HIIT session comparisons in outcome measures were assessed. Anaerobic speed reserve (coefficient of variation ± 95% confidence limits; 3.1 ± 1.5%), MAS (1.8 ± 1.3%), and MSS (0.8 ± 0.6%) indicated acceptable reliability. Moderate correlations between ASR and RPE (r = 0.33), postsession blood lactate (r = 0.34), and HR (r = 0.37) were observed during long intervals HIIT. A strong correlation was observed between ASR and RPE during SIT (r = 0.50). Sprint interval training elicited higher RPE's and postsession blood lactate's than other HIIT sessions. Anaerobic speed reserve has good reliability and may influence acute physiological responses to HIIT in female soccer players.

The Effects of Exercise-Based Injury Prevention Programmes on Injury Risk in Adult Recreational Athletes: A Systematic Review and Meta-Analysis.

BACKGROUND: Injuries are common in adult recreational athletes. Exercise-based injury prevention programmes offer the potential to reduce the risk of injury and have been a popular research topic. Yet, syntheses and meta-analyses on the effects of exercise-based injury prevention programmes for adult recreational athletes are lacking. OBJECTIVES: We aimed to synthesise and quantify the pooled intervention effects of exercise-based injury prevention programmes delivered to adults who participate in recreation sports. METHODS: Studies were eligible for inclusion if they included adult recreational athletes (aged > 16 years), an exercise-based intervention and used a randomised controlled trial design. Exclusion criteria were studies without a control group, studies using a non-randomised design and studies including participants who were undertaking activity mandatory for their occupation. Eleven literature databases were searched from earliest record, up to 9 June, 2022. The Physiotherapy Evidence Database (PEDro) scale was used to assess the risk of bias in all included studies. Reported risk statistics were synthesised in a random-effects meta-analysis to quantify pooled treatment effects and associated 95% confidence intervals and prediction intervals. RESULTS: Sixteen studies met the criteria. Risk statistics were reported as risk ratios [RRs] (n = 12) or hazard ratios [HRs] (n = 4). Pooled estimates of RRs and HRs were 0.94 (95% confidence interval 0.80-1.09) and 0.65 (95% confidence interval 0.39-1.08), respectively. Prediction intervals were 0.80-1.09 and 0.16-2.70 for RR and HR, respectively. Heterogeneity was very low for RR studies, but high for HR studies (tau = 0.29, I2 = 81%). There was evidence of small study effects for RR studies, evidenced by funnel plot asymmetry and Egger's test for small study bias: - 0.99 (CI - 2.08 to 0.10, p = 0.07). CONCLUSIONS: Pooled point estimates were suggestive of a reduced risk of injury in intervention groups. Nevertheless, these risk estimates were insufficiently precise, too heterogeneous and potentially compromised by small study effects to arrive at any robust conclusion. More large-scale studies are required to clarify whether exercise-based injury prevention programmes are effective in adult recreational athletes. CLINICAL TRIAL REGISTRATION: The protocol for this review was prospectively registered in the PROSPERO database (CRD42021232697).

Fast algorithm for 3D volume reconstruction from light field microscopy datasets.

Light field microscopy can capture 3D volume datasets in a snapshot, making it a valuable tool for high-speed 3D imaging of dynamic biological events. However, subsequent computational reconstruction of the raw data into a human-interpretable 3D+time image is very time-consuming, limiting the technique's utility as a routine imaging tool. Here we derive improved equations for 3D volume reconstruction from light field microscopy datasets, leading to dramatic speedups. We characterize our open-source Python implementation of these algorithms and demonstrate real-world reconstruction speedups of more than an order of magnitude compared with established approaches. The scale of this performance improvement opens up new possibilities for studying large timelapse datasets in light field microscopy.

Snapshot volumetric imaging with engineered point-spread functions.

The biological world involves intracellular and intercellular interactions that occur at high speed, at multiple scales and in three dimensions. Acquiring 3D images, however, typically requires a compromise in either spatial or temporal resolution compared to 2D imaging. Conventional 2D fluorescence imaging provides high spatial resolution but requires plane-by-plane imaging of volumes. Conversely, snapshot methods such as light-field microscopy allow video-rate imaging, but at the cost of spatial resolution. Here we introduce 3D engineered point-spread function microscopy (3D-EPM), enabling snapshot imaging of real-world 3D extended biological structures while retaining the native resolution of the microscope in space and time. Our new computational recovery strategy is the key to volumetrically reconstructing arbitrary 3D structures from the information encapsulated in 2D raw EPM images. We validate our technique on both point-like and extended samples, and demonstrate its power by imaging the intracellular motion of chloroplasts undergoing cyclosis in a sample of Egeria densa. Our technique represents a generalised computational methodology for 3D image recovery which is readily adapted to a diverse range of existing microscopy platforms and engineered point-spread functions. We therefore expect it to find broad applicability in the study of rapid biological dynamics in 3D.

Macrophages trigger cardiomyocyte proliferation by increasing epicardial vegfaa expression during larval zebrafish heart regeneration.

Cardiac injury leads to the loss of cardiomyocytes, which are rapidly replaced by the proliferation of the surviving cells in zebrafish, but not in mammals. In both the regenerative zebrafish and non-regenerative mammals, cardiac injury induces a sustained macrophage response. Macrophages are required for cardiomyocyte proliferation during zebrafish cardiac regeneration, but the mechanisms whereby macrophages facilitate this crucial process are fundamentally unknown. Using heartbeat-synchronized live imaging, RNA sequencing, and macrophage-null genotypes in the larval zebrafish cardiac injury model, we characterize macrophage function and reveal that these cells activate the epicardium, inducing cardiomyocyte proliferation. Mechanistically, macrophages are specifically recruited to the epicardial-myocardial niche, triggering the expansion of the epicardium, which upregulates vegfaa expression to induce cardiomyocyte proliferation. Our data suggest that epicardial Vegfaa augments a developmental cardiac growth pathway via increased endocardial notch signaling. The identification of this macrophage-dependent mechanism of cardiac regeneration highlights immunomodulation as a potential strategy for enhancing mammalian cardiac repair.

Selective Cdk9 inhibition resolves neutrophilic inflammation and enhances cardiac regeneration in larval zebrafish.

Sustained neutrophilic inflammation is detrimental for cardiac repair and associated with adverse outcomes following myocardial infarction (MI). An attractive therapeutic strategy to treat MI is to reduce or remove infiltrating neutrophils to promote downstream reparative mechanisms. CDK9 inhibitor compounds enhance the resolution of neutrophilic inflammation; however, their effects on cardiac repair/regeneration are unknown. We have devised a cardiac injury model to investigate inflammatory and regenerative responses in larval zebrafish using heartbeat-synchronised light-sheet fluorescence microscopy. We used this model to test two clinically approved CDK9 inhibitors, AT7519 and flavopiridol, examining their effects on neutrophils, macrophages and cardiomyocyte regeneration. We found that AT7519 and flavopiridol resolve neutrophil infiltration by inducing reverse migration from the cardiac lesion. Although continuous exposure to AT7519 or flavopiridol caused adverse phenotypes, transient treatment accelerated neutrophil resolution while avoiding these effects. Transient treatment with AT7519, but not flavopiridol, augmented wound-associated macrophage polarisation, which enhanced macrophage-dependent cardiomyocyte number expansion and the rate of myocardial wound closure. Using cdk9-/- knockout mutants, we showed that AT7519 is a selective CDK9 inhibitor, revealing the potential of such treatments to promote cardiac repair/regeneration.

Live Imaging of Heart Injury in Larval Zebrafish Reveals a Multi-Stage Model of Neutrophil and Macrophage Migration.

Neutrophils and macrophages are crucial effectors and modulators of repair and regeneration following myocardial infarction, but they cannot be easily observed in vivo in mammalian models. Hence many studies have utilized larval zebrafish injury models to examine neutrophils and macrophages in their tissue of interest. However, to date the migratory patterns and ontogeny of these recruited cells is unknown. In this study, we address this need by comparing our larval zebrafish model of cardiac injury to the archetypal tail fin injury model. Our in vivo imaging allowed comprehensive mapping of neutrophil and macrophage migration from primary hematopoietic sites, to the wound. Early following injury there is an acute phase of neutrophil recruitment that is followed by sustained macrophage recruitment. Both cell types are initially recruited locally and subsequently from distal sites, primarily the caudal hematopoietic tissue (CHT). Once liberated from the CHT, some neutrophils and macrophages enter circulation, but most use abluminal vascular endothelium to crawl through the larva. In both injury models the innate immune response resolves by reverse migration, with very little apoptosis or efferocytosis of neutrophils. Furthermore, our in vivo imaging led to the finding of a novel wound responsive mpeg1+ neutrophil subset, highlighting previously unrecognized heterogeneity in neutrophils. Our study provides a detailed analysis of the modes of immune cell migration in larval zebrafish, paving the way for future studies examining tissue injury and inflammation.

Systematic Reductions in Differential Ratings of Perceived Exertion Across a 2-Week Repeated-Sprint-Training Intervention That Improved Soccer Players' High-Speed-Running Abilities.

PURPOSE: To quantify changes in differential ratings of perceived exertion (dRPE) across a 2-wk repeated-sprint-training intervention that improved high-intensity intermittent-running ability and linear speed of semiprofessional soccer players. METHODS: Thirteen players completed 3 (sessions 1-3) or 4 (sessions 4-6) sets of 7 sprints (group 1 [n = 7]: 30-m straight; group 2 [n = 6]: 2 × 10-m shuttle), with 20 s and 4 min of recovery between sprints and sets, respectively. Postset perceptions of breathlessness (RPE-B) and leg-muscle exertion (RPE-L) were rated using the CR100 scale. RESULTS: Overall, RPE-B (mean [SD]: 46 [13] arbitrary units [AU], "hard") was most likely higher than RPE-L (39 [13] AU, "somewhat hard," mean difference: 8 AU; 90% confidence limits [CLs]: ±2). Set-to-set increases in dRPE (in AU; 90% CL: approximately ±2) were large in session 1 (RPE-B: 15; RPE-L: 14), moderate in sessions 2-5 (RPE-B: 7-10; RPE-L: 7-8), and small (RPE-B: 6) to moderate (RPE-L: 7) in session 6. Across the intervention, RPE-B reduced moderately in sets 3 (-13; 90% CL: ±4) and 4 (-12; 90% CL: ±12) and RPE-L reduced by a small magnitude in set 3 (-5; 90% CL: ±6). The set 4 change in RPE-L was unclear (-11; 90% CL: ±13). CONCLUSIONS: The authors observed systematic intrasession and intersession changes in dRPE across a 2-wk repeated-sprint-training intervention, with a fixed prescription of external load that improved semiprofessional soccer players' high-speed-running abilities. These findings could support dRPE as a measure of internal load and highlight its usefulness in evaluating repeated-sprint-training dose-response.

Author Correction: Adaptive prospective optical gating enables day-long 3D time-lapse imaging of the beating embryonic zebrafish heart.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

Twin-Airy Point-Spread Function for Extended-Volume Particle Localization.

The localization of point sources in optical microscopy enables nm-precision imaging of single-molecules and biological dynamics. We report a new method of localization microscopy using twin Airy beams that yields precise 3D localization with the key advantages of extended depth range, higher optical throughput, and potential for imaging higher emitter densities than are possible using other techniques. A precision of better than 30 nm was achieved over a depth range in excess of 7 µm using a 60×, 1.4 NA objective. An illustrative application to extended-depth-range blood-flow imaging in a live zebrafish is also demonstrated.

Rapid functionalisation and detection of viruses via a novel Ca2+-mediated virus-DNA interaction.

Current virus detection methods often take significant time or can be limited in sensitivity and specificity. The increasing frequency and magnitude of viral outbreaks in recent decades has resulted in an urgent need for diagnostic methods that are facile, sensitive, rapid and inexpensive. Here, we describe and characterise a novel, calcium-mediated interaction of the surface of enveloped viruses with DNA, that can be used for the functionalisation of intact virus particles via chemical groups attached to the DNA. Using DNA modified with fluorophores, we have demonstrated the rapid and sensitive labelling and detection of influenza and other viruses using single-particle tracking and particle-size determination. With this method, we have detected clinical isolates of influenza in just one minute, significantly faster than existing rapid diagnostic tests. This powerful technique is easily extendable to a wide range of other enveloped pathogenic viruses and holds significant promise as a future diagnostic tool.

Adaptive prospective optical gating enables day-long 3D time-lapse imaging of the beating embryonic zebrafish heart.

Three-dimensional fluorescence time-lapse imaging of the beating heart is extremely challenging, due to the heart's constant motion and a need to avoid pharmacological or phototoxic damage. Although real-time triggered imaging can computationally "freeze" the heart for 3D imaging, no previous algorithm has been able to maintain phase-lock across developmental timescales. We report a new algorithm capable of maintaining day-long phase-lock, permitting routine acquisition of synchronised 3D + time video time-lapse datasets of the beating zebrafish heart. This approach has enabled us for the first time to directly observe detailed developmental and cellular processes in the beating heart, revealing the dynamics of the immune response to injury and witnessing intriguing proliferative events that challenge the established literature on cardiac trabeculation. Our approach opens up exciting new opportunities for direct time-lapse imaging studies over a 24-hour time course, to understand the cellular mechanisms underlying cardiac development, repair and regeneration.

Indirect optical trapping using light driven micro-rotors for reconfigurable hydrodynamic manipulation.

Optical tweezers are a highly versatile tool for exploration of the mesoscopic world, permitting non-contact manipulation of nanoscale objects. However, direct illumination with intense lasers restricts their use with live biological specimens, and limits the types of materials that can be trapped. Here we demonstrate an indirect optical trapping platform which circumvents these limitations by using hydrodynamic forces to exert nanoscale-precision control over aqueous particles, without directly illuminating them. Our concept is based on optically actuated micro-robotics: closed-loop control enables highly localised flow-fields to be sculpted by precisely piloting the motion of optically-trapped micro-rotors. We demonstrate 2D trapping of absorbing particles which cannot be directly optically trapped, stabilise the position and orientation of yeast cells, and demonstrate independent control over multiple objects simultaneously. Our work expands the capabilities of optical tweezers platforms, and represents a new paradigm for manipulation of aqueous mesoscopic systems.

Multi-purpose SLM-light-sheet microscope.

By integrating a phase-only Spatial Light Modulator (SLM) into the illumination arm of a cylindrical-lens-based Selective Plane Illumination Microscope (SPIM), we have created a versatile system able to deliver high quality images by operating in a wide variety of different imaging modalities. When placed in a Fourier plane, the SLM permits modulation of the microscope's light-sheet to implement imaging techniques such as structured illumination, tiling, pivoting, autofocusing and pencil beam scanning. Previous publications on dedicated microscope setups have shown how these techniques can deliver improved image quality by rejecting out-of-focus light (structured illumination and pencil beam scanning), reducing shadowing (light-sheet pivoting), and obtaining a more uniform illumination by moving the highest-resolution region of the light-sheet across the imaging Field of View (tiling). Our SLM-SPIM configuration is easy to build and use, and has been designed to allow all of these techniques to be employed on an easily reconfigurable optical setup, compatible with the OpenSPIM design. It offers the possibility to choose between three different light-sheets, in thickness and height, which can be selected according to the characteristics of the sample and the imaging technique to be applied. We demonstrate the flexibility and performance of the system with results obtained by applying a variety of different imaging techniques on samples of fluorescent beads, zebrafish embryos, and optically cleared whole mouse brain samples. Thus our approach allows easy implementation of advanced imaging techniques while retaining the simplicity of a cylindrical-lens-based light-sheet microscope.

3D + time blood flow mapping using SPIM-microPIV in the developing zebrafish heart.

We present SPIM-µPIV as a flow imaging system, capable of measuring in vivo flow information with 3D micron-scale resolution. Our system was validated using a phantom experiment consisting of a flow of beads in a 50 µm diameter FEP tube. Then, with the help of optical gating techniques, we obtained 3D + time flow fields throughout the full heartbeat in a ∼3 day old zebrafish larva using fluorescent red blood cells as tracer particles. From this we were able to recover 3D flow fields at 31 separate phases in the heartbeat. From our measurements of this specimen, we found the net pumped blood volume through the atrium to be 0.239 nL per beat. SPIM-µPIV enables high quality in vivo measurements of flow fields that will be valuable for studies of heart function and fluid-structure interaction in a range of small-animal models.

High-speed extended-volume blood flow measurement using engineered point-spread function.

Experimental characterization of blood flow in living organisms is crucial for understanding the development and function of cardiovascular systems, but there has been no technique reported for snapshot imaging of thick samples in large volumes with high precision. We have combined computational microscopy and the diffraction-free, self-bending property of Airy-beams to track fluorescent beads with sub-micron precision through an extended axial range (up to 600 µm) within the flowing blood of 3 days post-fertilization (dpf) zebrafish embryos. The spatial trajectories of the tracer beads within flowing blood were recorded during transit through both cardinal and intersegmental vessels, and the trajectories were found to be consistent with the segmentation of the vasculature recorded using selective-plane illumination microscopy (SPIM). This method provides sufficiently precise spatial and temporal measurement of 3D blood flow that has the potential for directly probing key biomechanical quantities such as wall shear stress, as well as exploring the fluidic repercussions of cardiovascular diseases. Although we demonstrate the technique for blood flow, the ten-fold better enhancement in the depth range offers improvements in a wide range of applications of high-speed precision measurement of fluid flow, from microfluidics through measurement of cell dynamics to macroscopic aerosol characterizations.

Adaptive foveated single-pixel imaging with dynamic supersampling.

In contrast to conventional multipixel cameras, single-pixel cameras capture images using a single detector that measures the correlations between the scene and a set of patterns. However, these systems typically exhibit low frame rates, because to fully sample a scene in this way requires at least the same number of correlation measurements as the number of pixels in the reconstructed image. To mitigate this, a range of compressive sensing techniques have been developed which use a priori knowledge to reconstruct images from an undersampled measurement set. Here, we take a different approach and adopt a strategy inspired by the foveated vision found in the animal kingdom-a framework that exploits the spatiotemporal redundancy of many dynamic scenes. In our system, a high-resolution foveal region tracks motion within the scene, yet unlike a simple zoom, every frame delivers new spatial information from across the entire field of view. This strategy rapidly records the detail of quickly changing features in the scene while simultaneously accumulating detail of more slowly evolving regions over several consecutive frames. This architecture provides video streams in which both the resolution and exposure time spatially vary and adapt dynamically in response to the evolution of the scene. The degree of local frame rate enhancement is scene-dependent, but here, we demonstrate a factor of 4, thereby helping to mitigate one of the main drawbacks of single-pixel imaging techniques. The methods described here complement existing compressive sensing approaches and may be applied to enhance computational imagers that rely on sequential correlation measurements.

Repeated Sprints: An Independent Not Dependent Variable.

The ability to repeatedly perform sprints has traditionally been viewed as a key performance measure in team sports, and the relationship between repeated-sprint ability (RSA) and performance has been explored extensively. However, when reviewing the repeated-sprint profile of team-sports match play it appears that the occurrence of repeated-sprint bouts is sparse, indicating that RSA is not as important to performance as commonly believed. Repeated sprints are, however, a potent and time-efficient training strategy, effective in developing acceleration, speed, explosive leg power, aerobic power, and high-intensity-running performance--all of which are crucial to team-sport performance. As such, we propose that repeated-sprint exercise in team sports should be viewed as an independent variable (eg, a means of developing fitness) as opposed to a dependent variable (eg, a means of assessing fitness/performance).

Contrasting effects of a mixed-methods high-intensity interval training intervention in girl football players.

Little is known about the responses of girl athletes to training interventions throughout maturation. This study evaluated group and individual responses to an 8-week, mixed-methods, high-intensity interval training (HIIT) programme in girl football players. Thirty-seven players (age 13.4 ± 1.5 years) were tested for 20-m speed, repeated-sprint ability, change-of-direction speed and level 1 yo-yo intermittent recovery (YYIR). Players were subcategorised into before-, at- and after-PHV (peak height velocity) based on maturity offset. Very likely moderate (25%; ±90% confidence limits = 9.2) improvements occurred in YYIR, but data were unclear in players before-PHV with moderate individual differences in response. Decrements in repeated-sprint ability were most likely very large (6.5%; ±3.2) before-PHV, and likely moderate (1.7%; ±1.0) at-PHV. Data were unclear after-PHV. A very likely moderate (2.7%; ±1.0) decrement occurred in change-of-direction speed at-PHV while there was a very likely increase (-2.4%; ±1.3) in after-PHV players. Possibly small (-1.1%; ±1.4) improvements in 20-m speed occurred before-PHV but the effect was otherwise unclear with moderate to large individual differences. These data reflect specific responses to training interventions in girls of different biological maturity, while highlighting individual responses to HIIT interventions. This can assist practitioners in providing effective training prescription.