Subclinical and Device-Detected Atrial Fibrillation: Pondering the Knowledge Gap: A Scientific Statement From the American Heart Association.

The widespread use of cardiac implantable electronic devices and wearable monitors has led to the detection of subclinical atrial fibrillation in a substantial proportion of patients. There is evidence that these asymptomatic arrhythmias are associated with increased risk of stroke. Thus, detection of subclinical atrial fibrillation may offer an opportunity to reduce stroke risk by initiating anticoagulation. However, it is unknown whether long-term anticoagulation is warranted and in what populations. This scientific statement explores the existing data on the prevalence, clinical significance, and management of subclinical atrial fibrillation and identifies current gaps in knowledge and areas of controversy and consensus.

Type 8 long QT syndrome: pathogenic variants in CACNA1C-encoded Cav1.2 cluster in STAC protein binding site.

AIMS: Pathogenic gain-of-function variants in CACAN1C cause type-8 long QT syndrome (LQT8). We sought to describe the electrocardiographic features in LQT8 and utilize molecular modelling to gain mechanistic insights into its genetic culprits. METHODS AND RESULTS: Rare variants in CACNA1C were identified from genetic testing laboratories. Treating physicians provided clinical information. Variant pathogenicity was independently assessed according to recent guidelines. Pathogenic (P) and likely pathogenic (LP) variants were mapped onto a 3D modelled structure of the Cav1.2 protein. Nine P/LP variants, identified in 23 patients from 19 families with non-syndromic LQTS were identified. Six variants, found in 79% of families, clustered to a 4-residue section in the cytosolic II-III loop region which forms a region capable of binding STAC SH3 domains. Therefore, variants may affect binding of SH3-domain containing proteins. Arrhythmic events occurred in similar proportions of patients with II-III loop variants and with other P/LP variants (53% vs. 48%, P = 0.41) despite shorter QTc intervals (477 ± 31 ms vs. 515 ± 37 ms, P = 0.03). A history of sudden death was reported only in families with II-III loop variants (60% vs. 0%, P = 0.03). The predominant T-wave morphology was a late peaking T wave with a steep descending limb. Exercise testing demonstrated QTc prolongation on standing and at 4 min recovery after exercise. CONCLUSION: The majority of P/LP variants in patients with CACNA1C-mediated LQT8 cluster in an SH3-binding domain of the cytosolic II-III loop. This represents a 'mutation hotspot' in LQT8. A late-peaking T wave with a steep descending limb and QT prolongation on exercise are commonly seen.

Non-sustained microvolt level T-wave alternans in congenital long QT syndrome types 1 and 2.

BACKGROUND: Patients with long QT syndrome (LQTS) are predisposed to polymorphic ventricular tachycardia (VT) during adrenergic stimulation. Microvolt T-wave alternans (MTWA) is linked to vulnerability to VT in structural heart disease. The prevalence of non-sustained MTWA (NS-MTWA) in LQTS is unknown. METHODS: 31 LQT1, 42 LQT2, and 80 controls underwent MTWA testing during exercise. MTWA tests were classified per standardized criteria, and re-analyzed according to the modified criteria to account for NS-MTWA. RESULTS: LQT1 and LQT2 patients had a significantly higher frequency of late NS-MTWA (26% and 12%) compared to controls (0%). There was no significant difference between the groups with respect to sustained and early NS-MTWA. Late NS-MTWA was significantly associated with QTc. CONCLUSION: LQT1 and LQT2 patients had a higher prevalence of late NS-MTWA during exercise than matched controls. NS-MTWA likely reflects transient adrenergically mediated dispersion of repolarization, and could be a marker of arrhythmic risk in LQTS.

Clinical Aspects of Type 3 Long-QT Syndrome: An International Multicenter Study.

BACKGROUND: Risk stratification in patients with type 3 long-QT syndrome (LQT3) by clinical and genetic characteristics and effectiveness of ß-blocker therapy has not been studied previously in a large LQT3 population. METHODS: The study population included 406 LQT3 patients with 51 sodium channel mutations; 391 patients were known to be event free during the first year of life and were the focus of our study. Clinical, electrocardiographic, and genetic parameters were acquired for patients from 7 participating LQT3 registries. Cox regression analysis was used to evaluate the independent contribution of clinical, genetic, and therapeutic factors to the first occurrence of time-dependent cardiac events (CEs) from age 1 to 41 years. RESULTS: Of the 391 patients, 118 (41 males, 77 females) patients (30%) experienced at least 1 CE (syncope, aborted cardiac arrest, or long-QT syndrome-related sudden death), and 24 (20%) suffered from LQT3-related aborted cardiac arrest/sudden death. The risk of a first CE was directly related to the degree of QTc prolongation. Cox regression analysis revealed that time-dependent ß-blocker therapy was associated with an 83% reduction in CEs in females (P=0.015) but not in males (who had many fewer events), with a significant sex × ß-blocker interaction (P=0.04). Each 10-ms increase in QTc duration up to 500 ms was associated with a 19% increase in CEs. Prior syncope doubled the risk for life-threatening events (P<0.02). CONCLUSIONS: Prolonged QTc and syncope predispose patients with LQT3 to life-threatening CEs. However, ß-blocker therapy reduces this risk in females; efficacy in males could not be determined conclusively because of the low number of events.

Microvolt T-wave alternans amplifies spatial dispersion of repolarization in human subjects with ischemic cardiomyopathy.

INTRODUCTION: In experimental models, spatial dispersion of repolarization (DOR) due to discordant cellular alternans predisposes to ventricular fibrillation. To test the hypothesis that microvolt T-wave alternans (MTWA) in humans causes spatial DOR, we measured Tpeak-Tend interval (Tpe) and Tpe/QT ratio, electrocardiographic indices of spatial DOR. METHODS: Mean Tpe and Tpe/QT were compared in ischemic cardiomyopathy patients with positive and negative MTWA studies. RESULTS: MTWA was positive in 12 and negative in 24 patients. Tpe and Tpe/QT were higher in MTWA+ subjects compared to MTWA- subjects during exercise (64.5±6.8 vs. 54.9±8.7ms, p=0.001 and 0.218±0.03 vs. 0.177±0.02, p=0.001) but not at rest. CONCLUSION: Ischemic cardiomyopathy patients have increased Tpe and Tpe/QT when MTWA is induced during exercise, suggesting that MTWA causes increased spatial DOR in humans. Future studies are needed to determine if Tpe and Tpe/QT during exercise might predict increased risk of SCD alone or in combination with measurement of MTWA.

Does manual T-wave window adjustment affect microvolt T-wave alternans results in patients with structural heart disease?

INTRODUCTION: Microvolt T-wave alternans (MTWA) analysis can identify patients at low risk of sudden cardiac death who might not benefit from an implantable cardioverter-defibrillator (ICD). Current spectral methodology for performing MTWA analysis may "miss" part of the T-wave in patients with QT prolongation. The value of T-wave window adjustment in patients with structural heart disease has not been studied. METHODS: We assembled MTWA data from 5 prior prospective studies including 170 patients with reduced left ventricular ejection fraction, adjusted the T-wave window to include the entire T-wave, and reanalyzed MTWA. RESULTS: Of 170 patients, 43% required T-wave window adjustment. Only 3 of 170 patients (1.8%) had a clinically significant change in MTWA results. CONCLUSIONS: In 98.2% of patients, T-wave window adjustment did not improve the accuracy of MTWA analysis. Spectral MTWA as currently implemented remains effective for identifying patients with structural heart disease unlikely to benefit from ICD therapy.

Subclinical atrial fibrillation and the risk of stroke.

BACKGROUND: One quarter of strokes are of unknown cause, and subclinical atrial fibrillation may be a common etiologic factor. Pacemakers can detect subclinical episodes of rapid atrial rate, which correlate with electrocardiographically documented atrial fibrillation. We evaluated whether subclinical episodes of rapid atrial rate detected by implanted devices were associated with an increased risk of ischemic stroke in patients who did not have other evidence of atrial fibrillation. METHODS: We enrolled 2580 patients, 65 years of age or older, with hypertension and no history of atrial fibrillation, in whom a pacemaker or defibrillator had recently been implanted. We monitored the patients for 3 months to detect subclinical atrial tachyarrhythmias (episodes of atrial rate >190 beats per minute for more than 6 minutes) and followed them for a mean of 2.5 years for the primary outcome of ischemic stroke or systemic embolism. Patients with pacemakers were randomly assigned to receive or not to receive continuous atrial overdrive pacing. RESULTS: By 3 months, subclinical atrial tachyarrhythmias detected by implanted devices had occurred in 261 patients (10.1%). Subclinical atrial tachyarrhythmias were associated with an increased risk of clinical atrial fibrillation (hazard ratio, 5.56; 95% confidence interval [CI], 3.78 to 8.17; P<0.001) and of ischemic stroke or systemic embolism (hazard ratio, 2.49; 95% CI, 1.28 to 4.85; P=0.007). Of 51 patients who had a primary outcome event, 11 had had subclinical atrial tachyarrhythmias detected by 3 months, and none had had clinical atrial fibrillation by 3 months. The population attributable risk of stroke or systemic embolism associated with subclinical atrial tachyarrhythmias was 13%. Subclinical atrial tachyarrhythmias remained predictive of the primary outcome after adjustment for predictors of stroke (hazard ratio, 2.50; 95% CI, 1.28 to 4.89; P=0.008). Continuous atrial overdrive pacing did not prevent atrial fibrillation. CONCLUSIONS: Subclinical atrial tachyarrhythmias, without clinical atrial fibrillation, occurred frequently in patients with pacemakers and were associated with a significantly increased risk of ischemic stroke or systemic embolism. (Funded by St. Jude Medical; ASSERT ClinicalTrials.gov number, NCT00256152.).

Long-QT Syndrome and Therapy for Attention Deficit/Hyperactivity Disorder.

INTRODUCTION: Stimulants are the mainstay therapy for attention deficit/hyperactivity disorder (ADHD) and are associated with adrenergic side effects. There are limited data on the clinical course of patients treated for ADHD who have long-QT syndrome (LQTS), for which ß-blockade is the goal of therapy. METHODS: LQTS patients from the Rochester-based LQTS Registry (open-enrollment between 1979 and 2003; follow-up from 1979 to present) treated with stimulant or nonstimulant ADHD medications (n = 48) were compared to a 2:1 age-, gender-, and QTc-duration matched LQTS control group not exposed to ADHD medications (n = 96). Kaplan-Meier and Cox proportional hazards regression analyses were used to evaluate risk of cardiac events (syncope, aborted cardiac arrest, and sudden cardiac death) in LQTS patients treated with ADHD medications. RESULTS: During a mean follow-up of 7.9 ± 5.4 years after initiation of ADHD medication at a mean age 10.7 ±7.3 years, there was a 62% cumulative probability of cardiac events in the ADHD treatment group compared to 28% in the matched LQTS control group (P < 0.001). Time-dependent use of ADHD medication was associated with an increased risk for cardiac events (HR = 3.07; P = 0.03) in the multivariate Cox model adjusted for time-dependent ß-blocker use and prior cardiac events. Subgroup gender analyses showed that time-dependent ADHD medication was associated with an increased risk in male LQTS patients (HR = 6.80, P = 0.04). CONCLUSIONS: LQTS patients treated with ADHD medications have increased risk for cardiac events, particularly syncope, and this risk is augmented in males. The findings highlight the importance of heightened surveillance for LQTS patients on ADHD medications.

The Kv1.1 null mouse, a model of sudden unexpected death in epilepsy (SUDEP).

OBJECTIVE: Kv1.1 potassium channel null mouse (NULL) exhibits spontaneous seizure-related bradycardia, dies following seizure, and has been proposed as a model for vagus-mediated SUDEP. We characterized the cardiac events surrounding sudden unexpected death in epilepsy (SUDEP) in NULL during terminal asystole for comparison to patients with epilepsy who exhibit bradycardia and terminal or nonterminal asystole during/following seizure and explored the contribution of vagal-mediated bradycardia to SUDEP. METHODS: Electrocardiography (ECG) studies of 27 freely moving telemetered NULL mice was evaluated surrounding seizure-associated death. Chronic unilateral vagal section and, in a separate set of experiments, electrical stimulation of the cervical vagi in NULL and wild-type (WT) littermates assessed the role of the vagus nerve in seizure-related death. Seizure activity indicated by intense myogenic activity on the ECG recording correlated with visual and video recording. RESULTS: All NULL died following seizures, which were preceded by normal rhythm. Bradycardia followed seizure and led to slow ventricular escape rhythm (70-150 bpm) and asystole. The sequence from seizure to asystole was complete within approximately 3 min and was similar to that reported in individuals exhibiting ictal and postictal bradycardia/asystole. To address the singular role of vagus nerves in seizure-related asystole, cervical vagus nerves were stimulated in the absence of seizure. Heart rate was reduced 3 min to values similar to that following seizure but never produced asystole, suggesting activation of the vagi alone is insufficient for SUDEP. Nevertheless, unilateral chronic section of the vagus nerve increased survival time compared to nonsectioned NULL animals, supporting a role for the vagus nerve in seizure-associated death. SIGNIFICANCE: The Kv1.1 null mouse is a potential model for SUDEP in patients who experience ictal and postictal bradycardia. It offers the opportunity for evaluation of the combination of factors, in addition to vagal activation, necessary to produce a terminal asystole following seizure. It is notable that long-term studies that evaluate electroencephalography (EEG) and cardiorespiratory events surrounding nonfatal seizures may provide indices predictive of terminal seizure.

Clinical Management of Brugada Syndrome: Commentary From the Experts.

Although there is consensus on the management of patients with Brugada Syndrome with high risk for sudden cardiac arrest, asymptomatic or intermediate-risk patients present clinical management challenges. This document explores the management opinions of experts throughout the world for patients with Brugada Syndrome who do not fit guideline recommendations. Four real-world clinical scenarios were presented with commentary from small expert groups for each case. All authors voted on case-specific questions to evaluate the level of consensus among the entire group in nuanced diagnostic and management decisions relevant to each case. Points of agreement, points of controversy, and gaps in knowledge are highlighted.

Mutations in cytoplasmic loops of the KCNQ1 channel and the risk of life-threatening events: implications for mutation-specific response to ß-blocker therapy in type 1 long-QT syndrome.

BACKGROUND: ß-Adrenergic stimulation is the main trigger for cardiac events in type 1 long-QT syndrome (LQT1). We evaluated a possible association between ion channel response to ß-adrenergic stimulation and clinical response to ß-blocker therapy according to mutation location. METHODS AND RESULTS: The study sample comprised 860 patients with genetically confirmed mutations in the KCNQ1 channel. Patients were categorized into carriers of missense mutations located in the cytoplasmic loops (C loops), membrane-spanning domain, C/N terminus, and nonmissense mutations. There were 27 aborted cardiac arrest and 78 sudden cardiac death events from birth through 40 years of age. After multivariable adjustment for clinical factors, the presence of C-loop mutations was associated with the highest risk for aborted cardiac arrest or sudden cardiac death (hazard ratio versus nonmissense mutations=2.75; 95% confidence interval, 1.29-5.86; P=0.009). ß-Blocker therapy was associated with a significantly greater reduction in the risk of aborted cardiac arrest or sudden cardiac death among patients with C-loop mutations than among all other patients (hazard ratio=0.12; 95% confidence interval, 0.02-0.73; P=0.02; and hazard ratio=0.82; 95% confidence interval, 0.31-2.13; P=0.68, respectively; P for interaction=0.04). Cellular expression studies showed that membrane spanning and C-loop mutations produced a similar decrease in current, but only C-loop mutations showed a pronounced reduction in channel activation in response to ß-adrenergic stimulation. CONCLUSIONS: Patients with C-loop missense mutations in the KCNQ1 channel exhibit a high risk for life-threatening events and derive a pronounced benefit from treatment with ß-blockers. Reduced channel activation after sympathetic activation can explain the increased clinical risk and response to therapy in patients with C-loop mutations.

Risk of recurrent cardiac events after onset of menopause in women with congenital long-QT syndrome types 1 and 2.

BACKGROUND: Women with congenital long-QT syndrome experience an increased risk for cardiac events after the onset of adolescence that is more pronounced among carriers of the LQT2 genotype. We hypothesized that the hormonal changes associated with menopause may affect clinical risk in this population. METHODS AND RESULTS: We used a repeated-events analysis to evaluate the risk for recurrent syncope during the menopause transition and postmenopausal periods (5 years before and after the age at onset of menopause, respectively) among 282 LQT1 (n=151) and LQT2 (n=131) women enrolled in the Long-QT Syndrome Registry. Multivariate analysis showed that the risk for recurrent syncope (n=150) among LQT2 women was significantly increased during both menopause transition (hazard ratio, 3.38; P=0.005) and the postmenopausal period (hazard ratio, 8.10; P<0.001) compared with the reproductive period. The risk increase was evident among women who did or did not receive estrogen therapy. In contrast, among LQT1 women, the onset of menopause was associated with a reduction in the risk for recurrent syncope (hazard ratio, 0.19; P=0.05; P=0.02 for genotype-by-menopause interaction). Only 22 women (8%) experienced aborted cardiac arrest or sudden cardiac death during follow-up. The frequency of aborted cardiac arrest/sudden cardiac death showed a similar genotype-specific association with the onset of menopause. CONCLUSIONS: The onset of menopause is associated with a significant increase in the risk of cardiac events (dominated by recurrent episodes of syncope) in LQT2 women, suggesting that careful follow-up and continued long-term therapy are warranted in this population.

Microvolt-level T-wave alternans determination using the spectral method in patients with QT prolongation: value of adjusting the T-wave window.

BACKGROUND: Microvolt-level T-wave alternans (MTWA) measured by the spectral method is a useful risk predictor for sudden cardiac death because of its high negative predictive value. MTWA analysis software selects a segment of the ECG that encompasses the T-wave in most individuals, but may miss the T-wave end in patients with QT prolongation. HYPOTHESES: (1) In patients with QT prolongation, adjustment of the T-wave window will increase the sensitivity of MTWA detection. (2) The extent of T-wave window adjustment needed will correspond to the degree of QT prolongation. METHODS: Using data from long-QT syndrome patients, including QTc <0.45 s (normal), 0.45-0.49 s (moderate prolongation), and ≥ 0.50s (severe prolongation), MTWA analysis was performed before and after T-wave window adjustment. RESULTS: Of 119 patients, 74% required T-wave window adjustment. There was a stronger association between the magnitude of the T-wave offset and the unadjusted QT than between the magnitude of the T-wave offset and QTc (Spearman correlation coefficient 0.690 vs. 0.485 respectively, P<.05). Of 99 initially negative MTWA results, 4 became non-negative after adjustment of the T-wave window (P<.05). All 8 initially positive studies and 12 initially indeterminate studies remained positive and indeterminate, respectively. CONCLUSIONS: T-wave window adjustment can enable detection of abnormal MTWA that otherwise would be classified as "negative" or "normal." Newly developed T-wave window adjustment software may further improve the negative predictive value of MTWA testing and should be validated in a structural heart disease population.

ß-blockers protect against dispersion of repolarization during exercise in congenital long-QT syndrome type 1.

INTRODUCTION: ß-Blocker therapy reduces syncope and sudden death in long-QT syndrome type 1 (LQT1), but the mechanism of protection is incompletely understood. This study tested the hypothesis that ß-blockade reduces QT prolongation and dispersion of repolarization, measured as the T peak-to-end interval (T(pe) ), during exercise and recovery in LQT1 patients. METHODS AND RESULTS: QT and T(pe) were measured in 10 LQT1 patients (33 ± 13 years) and 35 normal subjects (32 ± 12 years) during exercise tests on and off ß-blockade. In LQT1 patients, ß-blockade reduced QT (391 ± 25 milliseconds vs 375 ± 26 milliseconds, P = 0.04 during exercise; 419 ± 41 milliseconds vs 391 ± 39 milliseconds, P = 0.02 during recovery) and markedly reduced T(pe) (91 ± 26 milliseconds vs 67 ± 19 milliseconds, P = 0.03 during exercise; 103 ± 26 milliseconds vs 78 ± 11 milliseconds, P = 0.02 during recovery). In contrast, in normal subjects, ß-blockade had no effect on QT (320 ± 17 milliseconds vs 317 ± 16 milliseconds, P = 0.29 during exercise; 317 ± 13 milliseconds vs 315 ± 14 milliseconds, P = 0.15 during recovery) and mildly reduced T(pe) (69 ± 13 milliseconds vs 61 ± 11 milliseconds, P = 0.01 during exercise; 77 ± 19 milliseconds vs. 68 ± 14 milliseconds, P < 0.001 during recovery). CONCLUSION: In LQT1 patients, ß-blockers reduced QT and T(pe) during exercise and recovery, supporting the theory that ß-blocker therapy protects LQT1 patients by reducing dispersion of repolarization during exercise and recovery.

Arrhythmic risk in congenital long QT syndrome.

One of the most important and challenging aspects of caring for patients with congenital long QT syndrome (LQTS) is assessing an individual's risk of sudden cardiac death (SCD) because of torsades de pointes. Current risk assessment integrates clinical and genetic features known to be associated with SCD, but more accurate methods of risk assessment could lead to more appropriate use of therapies, potentially saving lives and avoiding overtreatment. Conventional indices of risk include sex, age, extent of QT prolongation, history of symptoms (syncope or aborted SCD), and genetic subtype. The biophysical properties of specific mutations (eg, those that affect transmembrane segments of the ion channel protein or those that cause a dominant negative effect on ion channel function vs haplotype insufficiency) also contribute to risk. A growing body of basic mechanistic and clinical evidence points to heterogeneity of repolarization as a potent determinant of risk in LQTS patients. Mechanistically, heterogeneities of repolarization provide substrate for reentry, which likely causes perpetuation of torsades de pointes. Clinical markers that reflect heterogeneity of repolarization include abnormal microvolt-level T wave alternans, increased Tpeak-end interval, and dispersion of mechanical contraction time. The optimal methodology for using these indices as risk predictors in LQTS remains under active investigation. Further studies are needed to determine how indices of heterogeneity such as microvolt-level T wave alternans, Tpeak-end interval, and dispersion of mechanical contraction can be incorporated into models of risk prediction in LQTS, both for initial risk stratification and for assessment of efficacy of therapies.

Lessons Learned During the Pandemic: Correlation of QT Intervals Between Telemetry and 12-Lead Electrocardiogram.

Background and objective QT prolongation is associated with an increased risk of ventricular arrhythmias. Since some patients on contact or droplet precautions require QT-prolonging medications, monitoring the QT interval may become imperative to prevent fatal arrhythmias. To limit the exposure of staff to patients during and even after the coronavirus disease 2019 (COVID-19) pandemic and judiciously use personal protective equipment (PPE), it is important to find alternatives to frequent 12-lead electrocardiograms (ECG). The objective of this study was to compare QT intervals measured on telemetry to those measured on 12-lead ECG to determine whether telemetry QT interval measurements could be used in place of 12-lead measurements. Methods Simultaneous telemetry recordings via a Philips telemetry monitoring system (Philips Healthcare, Eindhoven, Netherlands) and 12-lead ECGs were obtained from 50 patients. Patients were from cardiac telemetry and cardiac intensive care units. QT interval from the telemetry system was compared to the QT interval on the 12-lead ECG. QT intervals on two telemetry strips were uninterpretable as the termination of the T-wave could not be defined appropriately; therefore, these patients were excluded. Results In 33 of 48 patients (69%), QT intervals from the telemetry studies matched the QT intervals measured by 12-lead ECG. The intraclass correlation coefficient (ICC) between telemetry QT and 12-lead ECG QT was 0.887 (95% CI: 0.809-0.934; p<0.001). In 15 of 48 patients (31%), the QT intervals measured from telemetry were different from those measured by 12-lead ECG. These patients either had an abnormal rhythm, conduction abnormalities, or repolarization abnormalities at baseline. Conclusion Telemetry is a suitable alternative for measuring QT intervals in the majority of patients. However, those with baseline ECG abnormalities should have serial 12-lead ECGs. This can reduce the risk of staff exposure to pathogens and prevent overuse of PPE during the COVID-19 pandemic and for other patients in isolation.

Management of Congenital Long-QT Syndrome: Commentary From the Experts.

While published guidelines are useful in the care of patients with long-QT syndrome, it can be difficult to decide how to apply the guidelines to individual patients, particularly those with intermediate risk. We explored the diversity of opinion among 24 clinicians with expertise in long-QT syndrome. Experts from various regions and institutions were presented with 4 challenging clinical scenarios and asked to provide commentary emphasizing why they would make their treatment recommendations. All 24 authors were asked to vote on case-specific questions so as to demonstrate the degree of consensus or divergence of opinion. Of 24 authors, 23 voted and 1 abstained. Details of voting results with commentary are presented. There was consensus on several key points, particularly on the importance of the diagnostic evaluation and of ß-blocker use. There was diversity of opinion about the appropriate use of other therapeutic measures in intermediate-risk individuals. Significant gaps in knowledge were identified.