HYSCORE and QM/MM Studies of Second Sphere Variants of the Type 1 Copper Site in Azurin: Influence of Mutations on the Hyperfine Couplings of Remote Nitrogens.

Secondary coordination sphere (SCS) interactions have been shown to play important roles in tuning reduction potentials and electron transfer (ET) properties of the Type 1 copper proteins, but the precise roles of these interactions are not fully understood. In this work, we examined the influence of F114P, F114N, and N47S mutations in the SCS on the electronic structure of the T1 copper center in azurin (Az) by studying the hyperfine couplings of (i) histidine remote Nε nitrogens and (ii) the amide Np using the two-dimensional (2D) pulsed electron paramagnetic resonance (EPR) technique HYSCORE (hyperfine sublevel correlation) combined with quantum mechanics/molecular mechanics (QM/MM) and DLPNO-CCSD calculations. Our data show that some components of hyperfine tensor and isotropic coupling in N47SAz and F114PAz (but not F114NAz) deviate by up to ∼±20% from WTAz, indicating that these mutations significantly influence the spin density distribution between the CuII site and coordinating ligands. Furthermore, our calculations support the assignment of Np to the backbone amide of residue 47 (both in Asn and Ser variants). Since the spin density distributions play an important role in tuning the covalency of the Cu-Scys bond of Type 1 copper center that has been shown to be crucial in controlling the reduction potentials, this study provides additional insights into the electron spin factor in tuning the reduction potentials and ET properties.

Moral Distress, Mental Health, and Risk and Resilience Factors Among Military Personnel Deployed to Long-Term Care Facilities During the COVID-19 Pandemic: Research Protocol and Participation Metrics.

BACKGROUND: The earliest days of the COVID-19 pandemic in Canada were marked by a significant surge in COVID-19 cases and COVID-19-related deaths among residents of long-term care facilities (LTCFs). As part of Canada's response to the COVID-19 pandemic, Canadian Armed Forces (CAF) personnel were mobilized for an initial emergency domestic deployment to the hardest-hit LTCFs (Operation LASER LTCF) to support the remaining civilian staff in ensuring the continued delivery of care to residents. Akin to what was observed following past CAF international humanitarian missions, there was an expected increased risk of exposure to multiple stressors that may be psychologically traumatic and potentially morally injurious in nature (ie, related to core values, eg, witnessing human suffering). Emerging data from health care workers exposed to the unprecedented medical challenges and dilemmas of the early pandemic stages also indicated that such experiences were associated with increased risk of adverse mental health outcomes. OBJECTIVE: This study aims to identify and quantify the individual-, group-, and organizational-level risk and resilience factors associated with moral distress, moral injury, and traditional mental health and well-being outcomes of Operation LASER LTCF CAF personnel. This paper aimed to document the methodology, implementation procedures, and participation metrics. METHODS: A multimethod research initiative was conducted consisting of 2 primary data collection studies (a quantitative survey and qualitative interviews). The quantitative arm was a complete enumeration survey with web-based, self-report questionnaires administered at 3 time points (3, 6, and 12 mo after deployment). The qualitative arm consisted of individual, web-based interviews with a focus on understanding the nuanced lived experiences of individuals participating in the Operation LASER LTCF deployment. RESULTS: CAF personnel deployed to Operation LASER LTCF (N=2595) were invited to participate in the study. Data collection is now complete. Overall, of the 2595 deployed personnel, 1088 (41.93%), 582 (22.43%), and 497 (19.15%) responded to the survey at time point 1 (3 mo), time point 2 (6 mo), and time point 3 (12 mo) after deployment, respectively. The target sample size for the qualitative interviews was set at approximately 50 considering resourcing and data saturation. Interest in participating in qualitative interviews surpassed expectations, with >200 individuals expressing interest; this allowed for purposive sampling across key characteristics, including gender, rank, Operation LASER LTCF role, and province. In total, 53 interviews were conducted. CONCLUSIONS: The data generated through this research have the potential to inform and promote better understanding of the well-being and mental health of Operation LASER LTCF personnel over time; identify general and Operation LASER LTCF-specific risk and protective factors; provide necessary support to the military personnel who served in this mission; and inform preparation and interventions for future missions, especially those more domestic and humanitarian in nature. INTERNATIONAL REGISTERED REPORT IDENTIFIER (IRRID): DERR1-10.2196/44299.

What Is Targeted When We Train Working Memory? Evidence From a Meta-Analysis of the Neural Correlates of Working Memory Training Using Activation Likelihood Estimation.

Working memory (WM) is the system responsible for maintaining and manipulating information, in the face of ongoing distraction. In turn, WM span is perceived to be an individual-differences construct reflecting the limited capacity of this system. Recently, however, there has been some evidence to suggest that WM capacity can increase through training, raising the possibility that training can functionally alter the neural structures supporting WM. To address the hypothesis that the neural substrates underlying WM are targeted by training, we conducted a meta-analysis of functional magnetic resonance imaging (fMRI) studies of WM training using Activation Likelihood Estimation (ALE). Our results demonstrate that WM training is associated exclusively with decreases in blood oxygenation level-dependent (BOLD) responses in clusters within the fronto-parietal system that underlie WM, including the bilateral inferior parietal lobule (BA 39/40), middle (BA 9) and superior (BA 6) frontal gyri, and medial frontal gyrus bordering on the cingulate gyrus (BA 8/32). We discuss the various psychological and physiological mechanisms that could be responsible for the observed reductions in the BOLD signal in relation to WM training, and consider their implications for the construct of WM span as a limited resource.

Molecular understanding of heteronuclear active sites in heme-copper oxidases, nitric oxide reductases, and sulfite reductases through biomimetic modelling.

Heme-copper oxidases (HCO), nitric oxide reductases (NOR), and sulfite reductases (SiR) catalyze the multi-electron and multi-proton reductions of O2, NO, and SO32-, respectively. Each of these reactions is important to drive cellular energy production through respiratory metabolism and HCO, NOR, and SiR evolved to contain heteronuclear active sites containing heme/copper, heme/nonheme iron, and heme-[4Fe-4S] centers, respectively. The complexity of the structures and reactions of these native enzymes, along with their large sizes and/or membrane associations, make it challenging to fully understand the crucial structural features responsible for the catalytic properties of these active sites. In this review, we summarize progress that has been made to better understand these heteronuclear metalloenzymes at the molecular level though study of the native enzymes along with insights gained from biomimetic models comprising either small molecules or proteins. Further understanding the reaction selectivity of these enzymes is discussed through comparisons of their similar heteronuclear active sites, and we offer outlook for further investigations.

Blast in Context: The Neuropsychological and Neurocognitive Effects of Long-Term Occupational Exposure to Repeated Low-Level Explosives on Canadian Armed Forces' Breaching Instructors and Range Staff.

Currently, there is strong interest within the military to better understand the effects of long-term occupational exposure to repeated low-level blast on health and performance. To gain traction on the chronic sequelae of blast, we focused on breaching-a tactical technique for gaining entry into closed/blocked spaces by placing explosives and maintaining a calculated safe distance from the detonation. Using a cross-sectional design, we compared the neuropsychological and neurocognitive profiles of breaching instructors and range staff to sex- and age-matched Canadian Armed Forces (CAF) controls. Univariate tests demonstrated that breaching was associated with greater post-concussive symptoms (Rivermead Post Concussion Symptoms Questionnaire) and lower levels of energy (RAND SF-36). In addition, breaching instructors and range staff were slower on a test that requires moving and thinking simultaneously (i.e., cognitive-motor integration). Next, using a multivariate approach, we explored the impact of other possible sources of injury, including concussion and prior war-zone deployment on the same outcomes. Concussion history was associated with higher post-concussive scores and musculoskeletal problems, whereas deployment was associated with higher post-concussive scores, but lower energy and greater PTSD symptomatology (using PCL-5). Our results indicate that although breaching, concussion, and deployment were similarly correlated with greater post-concussive symptoms, concussion history appears to be uniquely associated with altered musculoskeletal function, whereas deployment history appears to be uniquely associated with lower energy and risk of PTSD. We argue that the broader injury context must, therefore, be considered when studying the impact of repetitive low-level explosives on health and performance in military members.

The relationship between methods of scoring the alternate uses task and the neural correlates of divergent thinking: Evidence from voxel-based morphometry.

Divergent thinking tests have been used extensively in neuroscientific studies of creativity. However, output from tests of divergent thinking can be scored in different ways, and those scores can influence assessments of divergent thinking performance and its relationship with brain activation. Here we sought to investigate the relationship between various methods of scoring the Alternate Uses Task (AUT)-a well-known test of divergent thinking-and regional grey matter volume (GMV) using voxel-based morphometry (VBM). We assessed AUT performance based on (a) traditional approaches that involve scoring participants' output on fluency, flexibility, originality, and elaboration, (b) a subjective approach that involves scoring output directly on "snapshot" creativity, and (c) the definitional approach that involves scoring output separately on novelty and usefulness-the two criteria deemed necessary and jointly sufficient to categorize an idea as creative. Correcting for age, sex, intracranial volume, verbal IQ and working memory capacity, we found negative correlations between regional GMV in the left inferior temporal gyrus (ITG) and novelty and usefulness scores, but no correlation involving other scoring approaches. As part of the brain's core semantic system, this region is involved in concept retrieval and integration. We discuss the implications of these findings for our understanding of the neural bases of divergent thinking, and how ITG could be related to the generation of novel and useful responses.

The Reflective Mind: Examining Individual Differences in Susceptibility to Base Rate Neglect with fMRI.

Performance on heuristics and bias tasks has been shown to be susceptible to bias. In turn, susceptibility to bias varies as a function of individual differences in cognitive abilities (e.g., intelligence) and thinking styles (e.g., propensity for reflection). Using a classic task (i.e., lawyer-engineer problem), we conducted two experiments to examine the differential contributions of cognitive abilities versus thinking styles to performance. The results of Experiment 1 demonstrated that the Cognitive Reflection Test (CRT)-a well-established measure of reflective thinking-predicted performance on conflict problems (where base rates and intuition point in opposite directions), whereas STM predicted performance on nonconflict problems. Experiment 2 conducted in the fMRI scanner replicated this behavioral dissociation and enabled us to probe their neural correlates. As predicted, conflict problems were associated with greater activation in the ACC-a key region for conflict detection-even in cases when participants responded stereotypically. In participants with higher CRT scores, conflict problems were associated with greater activation in the posterior cingulate cortex (PCC), and activation in PCC covaried in relation to CRT scores during conflict problems. Also, CRT scores predicted activation in PCC in conflict problems (over and above nonconflict problems). Our results suggest that individual differences in reflective thinking as measured by CRT are related to brain activation in PCC-a region involved in regulating attention between external and internal foci. We discuss the implications of our findings in terms of PCC's possible involvement in switching from intuitive to analytic mode of thought.

Structural correlates of Openness and Intellect: Implications for the contribution of personality to creativity.

Openness/Intellect (i.e., openness to experience) is the Big Five personality factor most consistently associated with individual differences in creativity. Recent psychometric evidence has demonstrated that this factor consists of two distinct aspects-Intellect and Openness. Whereas Intellect reflects perceived intelligence and intellectual engagement, Openness reflects engagement with fantasy, perception, and aesthetics. We investigated the extent to which Openness and Intellect are associated with variations in brain structure as measured by cortical thickness, area, and volume (N = 185). Our results demonstrated that Openness was correlated inversely with cortical thickness and volume in left middle frontal gyrus (BA 6), middle temporal gyrus (MTG, BA 21), and superior temporal gyrus (BA 41), and exclusively with cortical thickness in left inferior parietal lobule (BA 40), right inferior frontal gyrus (IFG, BA 45), and MTG (BA 37). When age and sex were statistically controlled for, the inverse correlations between Openness and cortical thickness remained statistically significant for all regions except left MTG, whereas the correlations involving cortical volume remained statistically significant only for left middle frontal gyrus. There was no statistically significant correlation between Openness and cortical area, and no statistically significant correlation between Intellect and cortical thickness, area, or volume. Our results demonstrate that individual differences in Openness are correlated with variation in brain structure-particularly as indexed by cortical thickness. Given the involvement of the above regions in processes related to memory and cognitive control, we discuss the implications of our findings for the possible contribution of personality to creative cognition.

Topographical memory for newly-learned maps is differentially affected by route-based versus landmark-based learning: a functional MRI study.

Humans rely on topographical memory to encode information about spatial aspects of environments. However, even though people adopt different strategies when learning new maps, little is known about the impact of those strategies on topographical memory, and their neural correlates. To examine that issue, we presented participants with 40 unfamiliar maps, each of which displayed one major route and three landmarks. Half were instructed to memorize the maps by focusing on the route, whereas the other half memorized the maps by focusing on the landmarks. One day later, the participants were tested on their ability to distinguish previously studied 'old' maps from completely unfamiliar 'new' maps under conditions of high and low working memory load in the functional MRI scanner. Viewing old versus new maps was associated with relatively greater activation in a distributed set of regions including bilateral inferior temporal gyrus - an important region for recognizing visual objects. Critically, whereas the performance of participants who had followed a route-based strategy dropped to chance level under high working memory load, participants who had followed a landmark-based strategy performed at above chance levels under both high and low working memory load - reflected by relatively greater activation in the left inferior parietal lobule (i.e. rostral part of the supramarginal gyrus known as area PFt). Our findings suggest that landmark-based learning may buffer against the effects of working memory load during recognition, and that this effect is represented by the greater involvement of a brain region implicated in both topographical and working memory.

One-way traffic: The inferior frontal gyrus controls brain activation in the middle temporal gyrus and inferior parietal lobule during divergent thinking.

Contrary to earlier approaches that focused on the contributions of isolated brain regions to the emergence of creativity, there is now growing consensus that creative thought emerges from the interaction of multiple brain regions, often embedded within larger brain networks. Specifically, recent evidence from studies of divergent thinking suggests that kernel ideas emerge in posterior brain regions residing within the semantic system and/or the default mode network (DMN), and that the prefrontal cortex (PFC) regions within the executive control network (ECN) constrain those ideas for generating outputs that meet task demands. However, despite knowing that regions within these networks exhibit interaction, to date the direction of the relationship has not been tested directly. By applying Dynamic Causal Modeling (DCM) to fMRI data collected during a divergent thinking task, we tested the hypothesis that the PFC exerts unidirectional control over the middle temporal gyrus (MTG) and the inferior parietal lobule (IPL), vs. the hypothesis that these two sets of regions exert bidirectional control over each other (in the form of feedback loops). The data were consistent with the former model by demonstrating that the right inferior frontal gyrus (IFG) exerts unidirectional control over MTG and IPL, although the evidence was somewhat stronger in the case of the MTG than the IPL. Our findings highlight potential causal pathways that could underlie the neural bases of divergent thinking.

Task switching following 24 h of total sleep deprivation: a functional MRI study.

Task switching is a ubiquitous feature of many human activities that involve multitasking. In addition, owing to occupational demands, many individuals are required to engage in task switching under various levels of sleep deprivation, such as those who work in military and medical contexts. Nevertheless, little is known about the effects that sleep loss has on the neural bases of task switching. To address this shortcoming, we administered a cued switching task to participants following a night of normal sleep and also following a night of total sleep deprivation - in counterbalanced order. The behavioral results demonstrated a cost (i.e. longer reaction time) both as a function of sleep deprivation and task switching. Sleep deprivation resulted in greater activation in the frontoparietal network, whereas task switching was correlated with greater activation in the thalamus and superior temporal gyrus. However, despite increases in fatigue and sleepiness and a reduction in cognitive effectiveness (computed from actigraphic data), the reaction time cost associated with switching (i.e. switch cost) was not exacerbated by sleep deprivation. The results are discussed in terms of the involvement of executive functions in mitigating the effects of sleep deprivation on task switching.

Sbp1 modulates the translation of Pab1 mRNA in a poly(A)- and RGG-dependent manner.

RNA-binding protein Sbp1 facilitates the decapping pathway in mRNA metabolism and inhibits global mRNA translation by an unclear mechanism. Here we report molecular interactions responsible for Sbp1-mediated translation inhibition of mRNA encoding the polyadenosine-binding protein (Pab1), an essential translation factor that stimulates mRNA translation and inhibits mRNA decapping in eukaryotic cells. We demonstrate that the two distal RRMs of Sbp1 bind to the poly(A) sequence in the 5'UTR of the Pab1 mRNA specifically and cooperatively while the central RGG domain of the protein interacts directly with Pab1. Furthermore, methylation of arginines in the RGG domain abolishes the protein-protein interaction and the inhibitory effect of Sbp1 on translation initiation of Pab1 mRNA. Based on these results, the underlying mechanism for Sbp1-specific translational regulation is proposed. The functional differences of Sbp1 and RGG repeats alone on transcript-specific translation were observed, and a comparison of the results suggests the importance of remodeling the 5'UTR by RNA-binding proteins in mRNA translation.

Insights into an efficient light-driven hybrid P450 BM3 enzyme from crystallographic, spectroscopic and biochemical studies.

BACKGROUND: In order to perform selective CH functionalization upon visible light irradiation, Ru(II)-diimine functionalized P450 heme enzymes have been developed. The sL407C-1 enzyme containing the Ru(bpy)2PhenA (bpy=2,2'-bipyridine and PhenA=5-acetamido-1,10-phenanthroline) photosensitizer (1) covalently attached to the non-native single cysteine L407C of the P450BM3 heme domain mutant, displays high photocatalytic activity in the selective CH bond hydroxylation of several substrates. METHODS: A combination of X-ray crystallography, site-directed mutagenesis, transient absorption measurements and enzymatic assays was used to gain insights into its photocatalytic activity and electron transfer pathway. RESULTS: The crystal structure of the sL407C-1 enzyme was solved in the open and closed conformations revealing a through-space electron transfer pathway involving highly conserved, F393 and Q403, residues. Several mutations of these residues (F393A, F393W or Q403W) were introduced to probe their roles in the overall reaction. Transient absorption measurements confirm rapid electron transfer as heme reduction is observed in all four hybrid enzymes. Compared to the parent sL407C-1, photocatalytic activity was negligible in the dF393A-1 enzyme while 60% increase in activity with total turnover numbers of 420 and 90% product conversion was observed with the dQ403W-1 mutant. CONCLUSIONS: In the sL407C-1 enzyme, the photosensitizer is ideally located to rapidly deliver electrons, using the naturally occurring electron transfer pathway, to the heme center in order to activate molecular dioxygen and sustain photocatalytic activity. GENERAL SIGNIFICANCE: The results shed light on the design of efficient light-driven biocatalysts and the approach can be generalized to other members of the P450 superfamily.

Ru(II)-diimine functionalized metalloproteins: From electron transfer studies to light-driven biocatalysis.

The unique photochemical properties of Ru(II)-diimine complexes have helped initiate a series of seminal electron transfer studies in metalloenzymes. It has thus been possible to experimentally determine rate constants for long-range electron transfers. These studies have laid the foundation for the investigation of reactive intermediates in heme proteins and for the design of light-activated biocatalysts. Various metalloenzymes such as hydrogenase, carbon monoxide dehydrogenase, nitrogenase, laccase and cytochrome P450 BM3 have been functionalized with Ru(II)-diimine complexes. Upon visible light-excitation, these photosensitized metalloproteins are capable of sustaining photocatalytic activity to reduce small molecules such as protons, acetylene, hydrogen cyanide and carbon monoxide or activate molecular dioxygen to produce hydroxylated products. The Ru(II)-diimine photosensitizers are hence able to deliver multiple electrons to metalloenzymes buried active sites, circumventing the need for the natural redox partners. In this review, we will highlight the key achievements of the light-driven biocatalysts, which stem from the extensive electron transfer investigations. This article is part of a Special Issue entitled Biodesign for Bioenergetics--the design and engineering of electronic transfer cofactors, proteins and protein networks, edited by Ronald L. Koder and J.L. Ross Anderson.

Chromogenic nitrophenolate-based substrates for light-driven hybrid P450 BM3 enzyme assay.

The incorporation of a p-nitrophenoxy moiety in substrates has enabled the development of colorimetric assays to rapidly screen for O-demethylation activity of P450 enzymes. For the light-driven hybrid P450 BM3 enzymes, where a Ru(II) photosensitizer powers the enzyme upon visible light irradiation, we have investigated a family of p-nitrophenoxy derivatives as useful chromogenic substrates compatible with the light-driven approach. The validation of this assay and its adaptability to a 96-well plate format will enable the screening of the next generation of hybrid P450 BM3 enzymes towards C-H bond functionalization of non-natural substrates.

Transfer of training from one working memory task to another: behavioural and neural evidence.

N-back working memory (WM) tasks necessitate the maintenance and updating of dynamic rehearsal sets during performance. The delayed matching-to-sample (dMTS) task is another WM task, which in turn involves the encoding, maintenance, and retrieval of stimulus representations in sequential order. Because both n-back and dMTS engage WM function, we hypothesized that compared to a control task not taxing WM, training on the n-back task would be associated with better performance on dMTS by virtue of training a shared mental capacity. We tested this hypothesis by randomly assigning subjects (N = 43) to train on either the n-back (including 2-back and 3-back levels) or an active control task. Following training, dMTS was administered in the fMRI scanner. The n-back group performed marginally better than the active control group on dMTS. In addition, although the n-back group improved more on the less difficult 2-back level than the more difficult 3-back level across training sessions, it was improvement on the 3-back level that accounted for 21% of the variance in dMTS performance. For the control group, improvement in training across sessions was unrelated to dMTS performance. At the neural level, greater activation in the left inferior frontal gyrus, right posterior parietal cortex, and the cerebellum distinguished the n-back group from the control group in the maintenance phase of dMTS. Degree of improvement on the 3-back level across training sessions was correlated with activation in right lateral prefrontal and motor cortices in the maintenance phase of dMTS. Our results suggest that although n-back training is more likely to improve performance in easier blocks, it is improvement in more difficult blocks that is predictive of performance on a target task drawing on WM. In addition, the extent to which training on a task can transfer to another task is likely due to the engagement of shared cognitive capacities and underlying neural substrates-in this case WM.

The effects of a single night of sleep deprivation on fluency and prefrontal cortex function during divergent thinking.

The dorsal and ventral aspects of the prefrontal cortex (PFC) are the two regions most consistently recruited in divergent thinking tasks. Given that frontal tasks have been shown to be vulnerable to sleep loss, we explored the impact of a single night of sleep deprivation on fluency (i.e., number of generated responses) and PFC function during divergent thinking. Participants underwent functional magnetic resonance imaging scanning twice while engaged in the Alternate Uses Task (AUT) - once following a single night of sleep deprivation and once following a night of normal sleep. They also wore wrist activity monitors, which enabled us to quantify daily sleep and model cognitive effectiveness. The intervention was effective, producing greater levels of fatigue and sleepiness. Modeled cognitive effectiveness and fluency were impaired following sleep deprivation, and sleep deprivation was associated with greater activation in the left inferior frontal gyrus (IFG) during AUT. The results suggest that an intervention known to temporarily compromise frontal function can impair fluency, and that this effect is instantiated in the form of an increased hemodynamic response in the left IFG.

Right inferior frontal gyrus activation as a neural marker of successful lying.

There is evidence to suggest that successful lying necessitates cognitive effort. We tested this hypothesis by instructing participants to lie or tell the truth under conditions of high and low working memory (WM) load. The task required participants to register a response on 80 trials of identical structure within a 2 (WM Load: high, low) × 2 (Instruction: truth or lie) repeated-measures design. Participants were less accurate and responded more slowly when WM load was high, and also when they lied. High WM load activated the fronto-parietal WM network including dorsolateral prefrontal cortex (PFC), middle frontal gyrus, precuneus, and intraparietal cortex. Lying activated areas previously shown to underlie deception, including middle and superior frontal gyrus and precuneus. Critically, successful lying in the high vs. low WM load condition was associated with longer response latency, and it activated the right inferior frontal gyrus-a key brain region regulating inhibition. The same pattern of activation in the inferior frontal gyrus was absent when participants told the truth. These findings demonstrate that lying under high cognitive load places a burden on inhibition, and that the right inferior frontal gyrus may provide a neural marker for successful lying.

Sound attenuation of the indoor/outdoor range E-A-R plug.

The sound attenuation provided by the AOSafety Indoor/Outdoor Range E-A-R Plug was examined. This device, currently used in military operations in several countries, is comprised of two plugs that provide conventional ("indoor" plug) and level-dependent sound reduction (the "outdoor" plug), respectively. The effects of the user's gender and repeated fittings were explored. Eight men and eight women were tested on two separate occasions. Unoccluded and protected hearing thresholds were measured for each of nine one-third octave noise bands centered at 0.125 to 8 kHz. Attenuation was calculated as the difference between these two measures. The indoor plug provided 21 to 40 dB of sound reduction across the frequencies tested, closely matching the manufacturer's specification. The outdoor plug provided 5 to 22 dB of conventional attenuation, suggesting that it might serve as a safe means of conventional and level-dependant attenuation in hearing-impaired users. No differences were found in relation to gender or repeated fittings.