High-density bubble echocardiography for patent foramen ovale assessment: sensitivity and relationship with invasively measured shunt severity and atrial septum morphology

High-density bubble echocardiography for patent foramen ovale assessment: sensitivity and relationship with invasively measured shunt severity and atrial septum morphology | Research Square window.SnipcartSettings = { analytics: { enabled: false } }; (function() { var accessVector = localStorage.getItem('access_vector') || ''; window.dataLayer = window.dataLayer || []; if (accessVector) { window.dataLayer.push({ user: { profile: { profileInfo: { snid: accessVector } } } }); } })(); (function(w,d,s,l,i){w[l]=w[l]||[];w[l].push({'gtm.start':new Date().getTime(),event:'gtm.js'});var f=d.getElementsByTagName(s)[0],j=d.createElement(s),dl=l!='dataLayer'?'&l='+l:'';j.async=true;j.src='https://www.googletagmanager.com/gtm.js?id='+i+dl;f.parentNode.insertBefore(j,f);})(window,document,'script','dataLayer','GTM-K279D39R'); Browse Preprints In Review Journals COVID-19 Preprints AJE Video Bytes Research Tools Research Promotion AJE Professional Editing AJE Rubriq About Preprint Platform In Review Editorial Policies Our Team Advisory Board Help Center Sign In Submit a Preprint Cite Share Download PDF Research Article High-density bubble echocardiography for patent foramen ovale assessment: sensitivity and relationship with invasively measured shunt severity and atrial septum morphology Karel Medilek, Tomas Ondrus, Tomas Mraz, Josef Bis, Jaroslav Dusek, and 6 more This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-8302087/v1 This work is licensed under a CC BY 4.0 License Status: Under Review Version 1 posted 8 You are reading this latest preprint version Abstract Background Transthoracic contrast echocardiography (cTTE) is a first-line method for patent foramen ovale assessment (PFO), but its sensitivity is reported as low and inconsistent. We aimed to investigate the sensitivity of high-density bubble study cTTE for PFO detection, and the relationship between left-heart contrast opacification grade, invasively measured shunt severity and atrial septum/PFO morphology. Methods Fifty-five consecutive patients with proven PFO on contrast transoesophageal echocardiography (cTOE) underwent contrast transthoracic echocardiography (cTTE) and invasive right-to-left shunt severity measurement. Echocardiographic shunt severity was classified according to the degree of left atrial (cTOE) or left atrial/ventricular (cTTE) opacification as grade 0: no bubbles; grade 1: mild; grade 2: moderate; grade 3: significant opacification. The thermodilution method was used for the right-to-left heart shunt measurement. Results For PFO detection, the sensitivity of rest + Valsalva cTOE was 100%, but rest cTOE vs rest + Valsalva cTOE was 77.3%. For transthoracic studies, the sensitivity of rest + Valsalva cTTE was 98.2%, Valsalva cTTE 97.6%, and rest cTTE 83.6% vs rest + Valsalva cTOE. No relationship was found between rest or Valsalva cTTE and cTOE left-heart bubble opacification grade and invasively measured right-to-left shunt < 10%, 10–19%, or ≥ 20% (p = 0.10–0.26). A moderate association only was revealed between rest + Valsalva cTOE opacification grade and < 10% and ≥ 10% right-to-left shunt (p = 0.024, Cramér's coefficient = 0.368). No relationship was also found between left-heart opacification grade and the presence of an atrial septal aneurysm, Eustachian valve/Chiari's network, or PFO channel width and length (p = 0.45–0.77). Rest cTTE demonstrated equal contrast opacification grade in 56% and higher in 40% vs rest cTOE. Valsalva cTTE showed equal contrast opacification grade in 29% and higher in 46% of studies vs Valsalva cTOE. Conclusions In bubble studies using high-density contrast, cTOE and cTTE with Valsalva, but not rest studies, are highly sensitive for PFO assessment. There is no clinically relevant relationship between the left-heart bubble opacification grade and invasively measured right-to-left shunt severity or atrial septum morphology. The left-heart opacification degree is equal to or higher in cTTE than in cTOE, especially in rest studies. high density contrast transthoracic echocardiography shunt severity atrial septum morphology Figures Figure 1 Figure 2 Background Patent foramen ovale (PFO) is a flap-like opening resulting from incomplete fusion of the septum primum and septum secundum at the fossa ovalis. Its overall reported prevalence on autopsy is 27%, with a positive relationship between PFO and cryptogenic stroke (CS) observed in up to 46% of individuals 55 years. ( 1 , 2 ) In the RESPECT trial, closure of a PFO was associated with a lower rate of recurrent ischaemic strokes than medical therapy alone, providing evidence for the benefit of PFO closure in selected patients with CS. ( 3 ) The contrast echocardiography effect was first described in an experimental in vitro model in 1968 with injected saline, and shortly afterwards with 5% dextrose and agitated patient’s blood. ( 4 – 6 ) Twelve years later, an experimental study confirmed that the echo reflection in a contrast study was primarily due to microbubbles created during mixing of these agents with gas. ( 7 ) Longer stability of contrast was observed in gelatine due to lower surface tension. The non-transpulmonary contrast agents currently in use for PFO detection contain bubbles generally too large to pass through the pulmonary circulation. ( 8 , 9 ) Contrast transcranial doppler (cTCD), transoesophageal (cTOE), and transthoracic (cTTE) echocardiography represent the recommended methods for right-to-left intracardiac shunt assessment. In most cases, precise PFO diagnosis requires the combined use of different techniques, with cTCD or cTTE recommended as the first-line method. ( 10 ) Due to technical improvements, the sensitivity of cTTE has increased over the last 30 years from 51% to 80%. ( 11 ) According to meta-analyses cited in the latest European Society of Cardiology PFO statement and European Stroke Organisation guidelines, the sensitivity of cTTE in comparison with cTOE is reported as 88% and 71%. ( 10 , 12 ) In the latter meta-analysis, colloid contrast was used in 4/20 studies, all of them published more than 20 years ago, with a reported sensitivity of 13–61%. ( 10 ) On the other hand, some papers have reported better sensitivity of colloid contrast in comparison with normal saline. ( 13 , 14 ) It is also reported that the diagnostic performance of cTOE might improve with larger volume and number of contrast boluses administered. ( 15 , 16 ) The risk of a PFO being a causative pathology in patients with CS is usually linked to the left-heart opacification degree, PFO tunnel width and length, presence of concomitant interatrial aneurysm or hypermobile septum, and a prominent Eustachian valve or Chiari’s network. ( 12 , 17 – 19 ) Severe shunt is usually defined by the appearance of more than 20–30 microbubbles in the left atrium within three cardiac cycles after opacification of the right atrium. ( 20 , 21 ) The interplay between PFO morphology and invasively measured shunt severity or left-heart bubble contrast opacification degree has not been clinically studied, apart from an experimental model investigating fluid dynamics to assess the impact of PFO tunnel geometry on PFO flow behaviour. ( 22 ) We aimed to investigate the relationship between left-heart bubble opacification degree using a high-density non-transpulmonary contrast agent, invasively measured shunt severity, and atrial septum morphology, and to assess the sensitivity of cTTE using high-density contrast with and without Valsalva manoeuvre for PFO detection. Methods Study population The MEASURE-PFO study (in print) included 151 consecutive patients with CS or systemic embolisation and PFO proven by cTOE in six tertiary centres in the Czech Republic. All patients underwent invasive measurement of the right-to-left shunt and successful PFO closure. For contrast echocardiography studies, high-density contrast (Gelofusine, mannitol), dextrose and normal saline (without blood) were used, and cTOE was performed. The indication for PFO assessment was at the discretion of the referring neurologist. All patients signed informed consent form. In our substudy, we enrolled 55 patients from three cardiac centres in whom high-density contrast was used and cTTE was available, Cardiac catheterisation Femoral access was used for the catheterisation procedure. The pressure in the aorta was measured with the Sensis® hemodynamic system (Siemens Healthineers AG, Erlangen, Germany). The systemic blood flow, pulmonary blood flow and right-to-left (R-L) shunt were determined using the thermodilution Inntherm® system at rest and during Valsalva manoeuvre, if necessary, facilitated by another person pressing their palm against the patient’s abdomen. At rest, 10 ml of ice-cold saline was injected into the inferior vena cava near its opening into the right atrium. During the Valsalva manoeuvre, the systemic pressure was monitored, and at the time of the short pressure drop to about 50% of the initial systemic pressure, the manoeuvre was stopped and the saline was injected. The temperature of the systemic blood was measured by a thermal probe positioned slightly below the aortic arch in the descending aorta. The presence and quantification of the R-L shunt were determined by the blood temperature change in the descending aorta as published previously ( 23 ). Echocardiography cTOE and cTTE were performed before PFO closure using Vivid E95 (GE®) with 6Vc-D TTE and 6CT-D TOE probes and EPIQ CVXi and CX50 with X5-1 or X51c TTE and X7-2t or X8-2t TOE probes (Philips®). All patients signed a TOE informed consent form. Peripheral line access, at least 20G size, was preferably placed in the cubital vein, with no side preference. Standard TTE protocol was performed according to recommendations. ( 24 , 25 ) For the contrast study, retrospective gated acquisition was chosen, with at least ten RR intervals set up. Nineteen mL of Gelofusine B Braun 4% or Mannitol Fresenius Kabi 20% and 1 mL of air were agitated using two 20 ml luer lock syringes connected by a stopcock until a white suspension was obtained. A resting image was obtained after a quick contrast bolus was administered into the straight iv line immediately after it was prepared. At least 5 RR intervals of the whole 10 RR loop were acquired after the opacification of the right atrium using retrospective acquisition. If no bubbles were visualised in the left heart, the test was repeated. In the next step, patients were trained to perform a Valsalva manoeuvre by taking in a deep breath, closing the mouth, and “bearing down” as much as they could against the echocardiographer’s fist pushed to the patient’s abdominal wall, followed by abrupt release. Afterwards, patients were asked to make a Valsalva manoeuvre with the same force but with no help from the operator, who was performing TTE and made sure to keep all four chambers in the apical four-chamber view visible during the Valsalva strain and release phases. This was practised several times. The contrast was quickly injected at the initiation of the Valsalva manoeuvre, which was abruptly released once the whole 20 mL of contrast had been administered, at the time of the appearance/contrast opacification of the right atrium (usually 7–8 s after initialisation of the Valsalva manoeuvre). If no bubbles appeared in the left heart or in the case of image deterioration during acquisition due to translation movements, the test was repeated. For TOE, pharyngeal anaesthesia with lidocaine spray was performed and standard protocol was followed. ( 26 , 27 ) When accomplished, the interatrial septum was visualized using the mid-oesophageal short axis 40–60° view and the rest contrast study using the same setting as in cTTE was performed. If negative, the test was repeated in bi-caval view. Next, the Valsalva study was performed in short axis 40–60° view. If negative, it was repeated in bi-caval view and also repeated if necessary. It had to be visualised that the contrast was passing to the left atrium via PFO canal to confirm its presence. If this was not the case, every effort was made to localize the site of the shunt (multiperforated PFO etc.) and to rule out contrast inflow from the pulmonary vein. The left heart contrast opacification grading was classified as follows: grade 0: no bubbles, grade 1: mild, grade 2: moderate, grade 3: severe left atrium contrast opacification in cTOE and left atrium/ventricle opacification in cTTE. (Fig. 1 ) The single frame with the largest number of bubbles in the left atrium (cTOE) or left heart (cTTE) was used for opacification grading. The length of the PFO channel and the septum primum and secundum separation width at rest in the mid-oesophageal short-axis 40–60° and bi-caval projections were measured. The larger diameter was considered. The presence of the Eustachian valve or Chiari’s network was recorded; atrial septal aneurysm was defined as a 10 mm bulging of the septum tissue to the left/right side. The aim was to avoid sedation to ensure a good-quality Valsalva manoeuvre during cTOE, confirmed by the presence of atrial shifting to the left. If needed, a minimum dose of midazolam allowing completion of the whole cTOE with reliable Valsalva manoeuvre was titrated in 1 mg increments. A positive study was defined as the presence of contrast in the left atrium (cTOE)/left heart (cTTE) within three cycles after the right heart opacification in the rest study, or after strain release in the Valsalva study. Low density bubbles coming into the left atrium/left ventricle later than 3 cycles after the contrast administration/strain relief were not considered as a positive study. All images were assessed in a core lab by an experienced echocardiographer holding EACVI accreditation in adult TTE with exposure to PFO contrast studies over a period of almost twenty years. Statistical analysis The statistical analysis in the study involved summarising continuous variables as means with standard deviations and categorical variables as frequencies and percentages. Normality was assessed using the Shapiro-Wilk test. For comparisons across multiple groups, ANOVA on ranks (Kruskal-Wallis test) was applied when normality assumptions were not met, followed by Dunn’s post hoc test with Bonferroni correction. Categorical variables were evaluated using the chi-squared test, with Fisher’s exact test applied when expected frequencies were below 5. Cramér's V was used to assess the strength of the association between categorical variables. The agreement between cTTE and cTOE for left heart opacification was assessed using Kendall’s τ coefficient, with statistical significance determined at p < 0.05. Systematic differences between cTTE and cTOE were evaluated using McNemar’s test. Statistical significance was set at p < 0.05. All statistical analyses were performed using SPSS Statistics software, version 26 (IBM Corp., Armonk, NY, USA). Graphical analyses were performed using SigmaPlot, version 14. Results Fifty-five patients fulfilling the inclusion criteria were enrolled in the study. Their baseline characteristics are summarised in Table 1 . Table 1 Baseline characteristics of the studied group n (% N) / mean ± SD age (years) 44.9 ± (7.6) gender male 38 (69.1%) female 17 (30.9%) BMI 28.3 ± 5.1 BSA 2.1 ± 0.2 creatinine (mmol/l) 78.3 ± 14.2 haemoglobin (g/l) 143 ± 14 height (cm) 178 ± 9 weight (kg) 89.9 ± 18 asthma bronchiale or COPD 3 (5.5%) diabetes 3 (5.5%) hyperlipidaemia 26 (47.3%) hypertension 16 (29.1%) contrast site injection peripheral line 53 (96.3%) inferior vena cava 2 (3.7%) type of contrast Gelofusine 4% 37 (67%) Mannitol 20% 18 (33%) sedation during cTOE 12 (18.5%) sedation dose (ml) 2.6 ± 0.7 Chiari network/Eustachian valve 24 (43.6%) PFO channel length (mm) 13.2 ± 4.7 PFO channel width (mm) 2.9 ± 2.0 atrial septum aneurysm 29 (52.7%) BMI - body mass index, BSA - body surface area, COPD - chronic obstructive pulmonary disease Sensitivity of rest + Valslava cTOE was 100%, as all patients underwent successful PFO occlusion. cTTE with Valsalva manoeuvre was found to be highly sensitive for PFO detection. By contrast, rest cTTE/cTOE bubble studies showed low sensitivity. (Table 2 ) Table 2 Sensitivity of high-density bubble studies for PFO detection method A method B sensitivity A vs B cTTE rest + Valsalva cTOE rest + Valsalva 98.2% cTTE Valsalva cTOE Valsalva 97.6% cTTE rest cTOE rest 83.6% cTOE rest cTOE rest + Valsalva 77.3% No relationship between the invasively measured shunt severity (0%, < 10%, 10–19%, ≥ 20%) and left heart bubble opacification grade was observed either for rest or Valsalva cTTE or cTOE studies (Table 3 ). When the shunt severity was categorised as < 10% and ≥ 10%, only a moderate association was revealed between the rest + Valsalva studies opacification grade and shunt severity in cTOE (p = 0.024, Cramér's coefficient 0.368), but not in the cTTE group (Table 4 ). Table 3 Shunt severity vs left heart contrast opacification grade for the rest and Valsalva cTTE/cTOE severity of R-L shunt not detected < 10% 10–19% ≥ 20% p bubbles left heart opacification rest cTTE n = 55 0.16 Grade 0: no bubbles 3 (50.0%) 4 (19.0%) 1 (6,7%) 1 (7.7%) Grade 1: mild opacification 1 (16.7%) 9 (42.9%) 4 (26.7%) 4 (30.8%) Grade 2: moderate opacification 0 (0.0%) 7 (33,3%) 6 (40.0%) 5 (38.5%) Grade 3: significant opacification 2 (33.3%) 1 (4,8%) 4 (26.7%) 3 (23.1%) bubbles left heart opacification Valslava cTTE n = 44 0.20 Grade 0: no bubbles 1 (25.0%) 2 (10.5%) 0 (0.0%) 1 (11.1%) Grade 1: mild opacification 1 (25.0%) 1 (5.3%) 1 (8.3%) 0 (0.0%) Grade 2: moderate opacification 0 (0.0%) 10 (52.6%) 2 (16.7%) 4 (44.4%) Grade 3: significant opacification 2 (50.0%) 6 (31.6%) 9 (75.0%) 4 (44.4%) bubbles left atrium opacification rest cTOE n = 55 0.10 Grade 0: no bubbles 2 (33.3%) 6 (28.6%) 1 (6,7%) 2 (15.4%) Grade 1: mild opacification 2 (33.3%) 9 (42.9%) 6 (40.0%) 7 (53.8%) Grade 2: moderate opacification 0 (0.0%) 6 (28.6%) 6 (40.0%) 4 (30.5%) Grade 3: significant opacification 2 (33.3%) 0 (0.0%) 2 (13.3%) 0 (0.0%) bubbles left atrium opacification Valsalva cTOE n = 41 0.26 Grade 0: no bubbles 0 (0.0%) 0 (0.0%) 0 (0.0%) 0 (0.0%) Grade 1: mild opacification 2 (50.0%) 9 (50.0%) 1 (10.0%) 2 (22.2%) Grade 2: moderate opacification 2 (50.0%) 7 (38.9%) 5 (50.0%) 5 (55.6%) Grade 3: significant opacification 0 (0.0%) 2 (11.1%) 4 (40.0%) 2 (22.2%) Chi-squared test. R-L - right to left Table 4 < 10% and ≥ 10% shunt severity vs contrast opacification grade for the rest + Valsalva cTTE/cTOE severity of R-L shunt < 10% ≥ 10% p bubbles left heart opacification rest + Valsalva cTTE 0.206 Grade 0: no bubbles 1 (3.7%) Grade 1: mild opacification 4 (14.8%) 1 (3.6%) Grade 2: moderate opacification 12 (44.5%) 11 (39.3%) Grade 3: significant opacification 10 (37.0%) 16 (57.1%) bubbles left heart opacification rest + Valsalva cTOE 0.024 Grade 1: mild opacification 11 (40.7%) 3 (10.7%) Grade 2: moderate opacification 12 (44.4%) 15 (53.6%) Grade 3: significant opacification 4 (14.8%) 10 (35.7%) Chi-squared test, R-L - right to left No significant association was also found between the left heart opacification grade and the presence of an atrial septal aneurysm, Eustachian valve/Chiari´s network, or PFO width and length. (Table 5 ). Table 5 Relationship between atrial septal morphology and the left heart opacification grade, p value. anatomical interatrial septum/PFO variations PFO width PFO length Chiari´s network/ Eustachian valve aneurysm left heart opacification grade rest cTTE 0.17 0.49 0.10 0.45 left heart opacification grade Valsalva cTTE 0.37 0.69 0.47 0.73 left atrium opacification grade rest cTOE 0.55 0.16 0.64 0.46 left atrium opacification grade Valsalva cTOE 0.81 0.43 0.29 0.77 Chi-squared and Kruskal-Wallis tests Finally, the left heart opacification grade between cTTE and cTOE was compared. In rest studies, cTTE produced the same degree of opacification as cTOE in 55.5% of cases and a higher degree in 40%, demonstrating similar or superior performance in 95.5% of cases. A moderate and statistically significant correlation and difference were observed between cTTE and cTOE opacification grade (Kendall’s τ = 0.53, p < 0.001 and χ² = 26.83, p = 0.002). In Valsalva studies, the left heart cTTE opacification grade matched cTOE in 29% and exceeded it in 45.5% of cases, showing comparable or higher opacification degree in 74.5% of cases. However, the correlation and difference between cTTE and cTOE opacification grade were weak and not statistically significant (Kendall’s τ = 0.14, p = 0.23 and χ² = 13.23, p = 0.15). Discussion This study of high-density contrast echocardiography in the investigation of the presence of PFO in CS patients demonstrated: (i) high sensitivity of Valsalva cTOE and cTTE for PFO detection, (ii) no clinically relevant relationship between the left heart bubble opacification grade and invasively measured left-to-right heart shunt severity, (iii) no significant relationship between the left heart opacification grade and anatomical features of the interatrial septum/PFO, (iv) equal or higher left heart bubble opacification degree in cTTE vs cTOE, especially in rest studies. In two studies with 91 patients included in the cTOE meta-analysis from Mojadidi, reflecting the usual practice (i.e. contrast injection into the peripheral line and Valsalva manoeuvre performance), the sensitivity of cTOE vs autopsy, cardiac surgery, and/or catheterization (ability to pass the catheter through PFO) as the reference was 89–91%. ( 28 ) In addition, cTOE studies yielded a significantly lower prevalence of PFO of 13% (95% CI: 8–18%) as compared to autopsy studies 25% (95% CI: 20–29%), p = 0.004. ( 12 ) The imperfect accuracy of cTOE is usually attributed to patient intolerance of the TOE probe or inadequate Valsalva manoeuvre during sedation, which might result in up to 50% of PFOs being missed. ( 29 , 30 ) Sedation use during cTOE is, however, often not reported in the studies. ( 31 – 33 ) The sensitivity of PFO detection also depends on the number of injections in Valsalva cTOE studies. ( 16 ) Despite all of these facts, cTOE is considered the gold standard for PFO detection. ( 12 ) In our study, the patients were trained for and rigorous operator´s adherence to the correct performance of the Valsalva manoeuvre was required. Visualisation of the contrast passing through the PFO canal was mandatory for its diagnosis and high-density contrast was used. Sedation was needed in < 20% of patients. All these facts likely contributed to the 100% sensitivity of cTOE against successful PFO occlusion in our study, highlighting the importance of an experienced imaging specialist performing cTOE for PFO detection. ( 34 ) There is heterogeneity in the type of contrast and Valsalva technique use in the published studies comparing cTTE to cTOE, with saline with or without blood prevailing in 80% and the contrast injected during the provocative manoeuvre in 66%. ( 10 , 35 ) The published sensitivity of saline and colloid cTTE ranges from 23% to 92% and 30% to 61%, respectively. ( 36 ) For the second harmonic cTTE, the sensitivity for PFO detection is reported to be higher at 91%, and similar to 96% for cTCD. ( 35 , 37 ) We used high-density contrast agents, mainly Gelofusine, showing excellent cTTE sensitivity > 95%. One of the reasons, according to our experience, is better Gelofusine bubble stability and opacification of the right and left heart during cTTE/cTOE in comparison with normal saline contrast. (Fig. 1 , 2 ) This finding is supported by previously published studies. ( 9 , 13 , 14 ) Indeed, adding blood to normal saline can stabilise microbubbles and increase their suspension time in the blood. ( 38 ) In our experience, the density of the right heart opacification using Gelofusine and normal saline with blood does not differ. The use of modified gelatine is safe, a potential risk of anaphylaxis associated with its use is low, although the intensive care data are conflicting. We found only one published case report on anaphylaxis during contrast echocardiography. ( 39 – 41 ) A relationship was found between allergic reactions to red meat and sensitisation to gelatine and galactose-α-1,3-galactose (α-Gal). ( 42 ) Gelofusine handling is also more practical and hygienic than mixing normal saline with blood, at least from the infection control point of view. For these reasons, the gelatine-based contrast has been the first line choice in our echo lab for almost two decades. In a recently published study by Takaya, normal saline cTTE was better for predicting large left to right shunts than cTOE (79% vs 28% of patients). ( 21 ) One of the explanations may be linked to our results demonstrating better opacification of the left heart in cTTE than in cTOE, although using a different contrast agent. (Fig. 1 ) Also, in cTTE, the whole left heart is visualised in apical four chamber view, in contrast to the cTOE study, which is focused on the presence of the contrast agent in a 2D often foreshortened image of the left atrium. The risk of PFO associated CS is usually assessed by the left heart bubble size and the presence of anatomical features such as PFO width, atrial septal aneurysm (ASA) or septum hypermobility and large Eustachian valve. It sounds intuitive that a large PFO means more right-to-left shunting, a mobile ASA may open the PFO with every heartbeat, and the Eustachian valve may direct thrombi from the inferior vena cava to the PFO. Indeed, in a metanalysis from Overell, the presence of ASA was associated with a higher risk of ischaemic stroke in comparison to healthy subjects (OR 2.35). Individual observational studies demonstrated even higher risk (OR 3.65). ( 43 , 44 ) We found no link between the severity of the bubble opacification grade and high risk PFO features. This might be attributed to the high prevalence of ASA (> 50%) in our group, reflecting the high-risk population studied, in which further risk stratification might be difficult. Scoring systems for high-risk PFO have been developed and include PFO size (≥ 2 mm), tunnel length (≥ 10 mm), the presence of ASA/hypermobile interatrial septum, prominent Eustachian/Chiari’s valve, straddling interatrial thrombus, concomitant pulmonary embolism or deep vein thrombosis, and a large shunt at rest and during the Valsalva manoeuvre (> 20 microbubbles in the left atrium). ( 42 , 43 ) Our study is the first to compare the invasively measured shunt severity and the left heart contrast opacification degree, showing no relevant relationship apart from a moderate association in rest + Valsalva cTOE studies with larger shunts > 10%. There were few patients with no invasively measurable shunt but with a right-to-left shunt on cTTE/cTOE in our study. The explanation might include relative hypovolaemia and difficulty in performing the Valsalva manoeuvre in some patients undergoing invasive measurements via the femoral access. Also, counting the number of bubbles in the left atrium within three cycles after it opacifies the right heart by number of bubbles as recommended is impractical for high-density contrast due to the high degree and often homogeneous left heart opacification that is often observed. This approach has its limitations in general, as bubbles do not necessarily travel linearly across the left atrium and the same bubbles may be present across multiple cardiac cycles, leading to multiple counting. On the other hand, the left atrium size on the tomographic TOE slice is often significantly foreshortened, so some bubbles are inevitably missed. We chose a pragmatic approach, i.e. use of one still left heart frame with the maximum contrast opacification for its grading and a semiquantitative opacification scale, which we propose for future studies with high-density contrast agents. Indeed, in the recent paper by Su from the USA, the quantified recommended ratings of PFO shunt severity were present in < 1 in every 6 TEE reports, reflecting real clinical practice. ( 46 ) Limitations Our study is of small sample size, and its results would need to be confirmed by larger studies. Also, its results are valid for younger population only, as all patients were aged < 55 years. We used a larger amount of contrast (20 mL) instead of the usual 10 mL per administration, which does not require a flush, thus simplifying the procedure, but leading to possibly higher opacification of the heart, which might have affected the results. We did not record the arm side of the venous access for the contrast administration and the inferior vena cava contrast injection site was reported in 2 (3.7%) patients, but this is unlikely to have affected the results. We also used different than usual approach to assess the left heart opacification grade as discussed. Anyway, the heart is a 3D structure, and any method using a single plane underestimates the true left heart bubble volume. ( 34 ) Counting the number of bubbles in a 3D image is however not currently feasible so far, but there are reports demonstrating better accuracy of 3D cTOE for PFO vs intrapulmonary shunt discrimination. ( 47 ) Despite every effort, there might have been some discrepancies in the Valsalva performance among the centres, especially in patients with cTOE performed under sedation, although this applied to only < 20% of the study population and midazolam doses were low. The same applies to the invasive shunt measurement, although appropriate Valsalva manoeuvre performance was monitored invasively by the pressure curve changes. Conclusion Our study demonstrated that in bubble studies using high-density contrast, cTOE and cTTE with Valsalva, but not rest studies, are highly sensitive for PFO assessment. There is no clinically relevant relationship between the left-heart bubble opacification grade and invasively measured right-to-left shunt severity or atrial septum/PFO morphology. The left-heart opacification degree is equal to or higher in cTTE than in cTOE, especially in rest studies. Declarations Authors´contribution Dr Medilek was responsible for performing the echocardiography studies, interpreting and analysing the data, and compiling the manuscript. Dr Ondrus, Dr Mraz, Dr Praus and Dr Ballon performed echocardiography studies at their respective centres. Dr. Stasek, Dr Bis, Dr Dusek, Dr Poloczek and Dr Mates performed the invasive measurements. Dr Stasek was also responsible for the design of the MEASURE - PFO trial. All authors contributed to the editing of the manuscript, and approved its final version and content. Funding The trial was supported by a research grant from the Ministry of Health, Czech Republic, No. AZV NU20-02-00310 and by and by the Cooperatio Program, Charles University, Czech Republic, research area CARD. Acknowledgement We thank David Belcak for English language proofreading. Data availability The datasets used and analysed during the current study are available from the corresponding author on reasonable request. Ethics approval The study has been carried out in accordance with the Code of Ethics of the World Medical Association (Declaration of Helsinki) and approved by the Ethics Committee, Faculty of Medicine Hradec Kralove, Czech Republic (ref. No 201906S19P). The authors are responsible for the design and conduct of this study, all study analyses, drafting and editing of the manuscript, and its final contents. Consent for publication Not applicable. Competing interests The authors declare no competing interests. References Hagen PT, Scholz DG, Edwards WD. Incidence and Size of Patent Foramen Ovale During the First 10 Decades of Life: An Autopsy Study of 965 Normal Hearts. Mayo Clin Proc. 1984;59:17–20. Homma S, Sacco RL. Patent Foramen Ovale and Stroke. Circulation. 2005;112:1063–172. Saver JL, Carroll JD, Thaler DE, Smalling RW, MacDonald LA, Marks DS, et al. Long-Term Outcomes of Patent Foramen Ovale Closure or Medical Therapy after Stroke. N Engl J Med. 2017;377:1022–32. Gramiak R, Shah PM. Echocardiography of the Aortic Root. Invest Radiol. 1968;3:356–66. Gramiak R, Shah PM, Kramer DH. Ultrasound Cardiography: Contrast Studies in Anatomy and Function. Radiology. 1969;92:939–48. Kremkau FW, Gramiak R, Carstensen EL, Shah PM, Kramer DH. ULTRASONIC DETECTION OF CAVITATION AT CATHETER TIPS. Am J Roentgenol. 1970;110:177–83. Meltzer RS, Tickner EG, Sahines TP, Popp RL. The source of ultrasound contrast effect. J Clin Ultrasound. 1980;8:121–7. Nanda NC. History of echocardiographic contrast agents. Clin Cardiol. 1997;20(Suppl 1):7–11. Smith MD, Kwan OL, Reiser HJ, DeMaria AN. Superior intensity and reproducibility of SHU-454, a new right heart contrast agent. J Am Coll Cardiol. 1984;3:992–8. Caso V, Turc G, Abdul-Rahim AH, Castro P, Hussain S, Lal A, et al. European Stroke Organisation (ESO) Guidelines on the diagnosis and management of patent foramen ovale (PFO) after stroke. Eur Stroke J. 2024;9:800–34. Khan R, Karim MN, Hosseini F, Fine N. Diagnostic Accuracy of Transthoracic Echocardiography With Contrast for Detection of Right-to-Left Shunt: A Systematic Review and Meta-analysis. Can J Cardiol. 2022;38:1948–58. Pristipino C, Sievert H, D’Ascenzo F, Mas JL, Meier B, Scacciatella P, et al. European position paper on the management of patients with patent foramen ovale. General approach and left circulation thromboembolism. EuroIntervention. 2019;14:1389–402. Cukon Buttignoni S, Khorsand A, Mundigler G, Bergler-Klein J, Heger M, Zehetgruber M, et al. Agitated saline versus polygelatine for the echocardiographic assessment of patent foramen ovale. J Am Soc Echocardiogr. 2004;17:1059–65. Puledda F, Toscano M, Pieroni A, Veneroso G, Di Piero V, Vicenzini E. Right-to-left shunt detection sensitivity with air–saline and air–succinil gelatin transcranial Doppler. Int J Stroke. 2016;11:229–38. Li H, Huang Y, Luo D, Wen S, Fei H, Zhang C. Optimizing agitated saline volume for contrast echocardiography: Balancing diagnostic performance and operator fatigue. Med Adv. 2024;2:254–61. Johansson MC, Helgason H, Dellborg M, Eriksson P. Sensitivity for Detection of Patent Foramen Ovale Increased with Increasing Number of Contrast Injections: A Descriptive Study with Contrast Transesophageal Echocardiography. J Am Soc Echocardiogr. 2008;2:419–24. Turc G, Calvet D, Guérin P, Sroussi M, Chatellier G, Mas J, et al. Closure, Anticoagulation, or Antiplatelet Therapy for Cryptogenic Stroke With Patent Foramen Ovale: Systematic Review of Randomized Trials, Sequential Meta-Analysis, and New Insights From the CLOSE Study. J Am Heart Assoc. 2018;7:e008356. Mas JL, Derumeaux G, Guillon B, Massardier E, Hosseini H, Mechtouff L, et al. Patent Foramen Ovale Closure or Anticoagulation vs. Antiplatelets after Stroke. N Engl J Med. 2017;377:1011–21. Lee PH, Song JK, Kim JS, Heo R, Lee S, Kim DH, et al. Cryptogenic Stroke and High-Risk Patent Foramen Ovale. J Am Coll Cardiol. 2018;71:2335–42. Mas JL, Derumeaux G, Guillon B, Massardier E, Hosseini H, Mechtouff L, et al. Patent Foramen Ovale Closure or Anticoagulation vs. Antiplatelets after Stroke. N Engl J Med. 2017;377:1011–21. Takaya Y, Nakayama R, Akagi T, Yokohama F, Miki T, Nakagawa K, et al. Importance of saline contrast transthoracic echocardiography for evaluating large right-to-left shunt in patent foramen ovale associated with cryptogenic stroke. Int J Cardiovasc Imaging. 2022;38:515–20. Morelli F, Guadagno V, Mencarini T, Brambilla M, Fusini L, Bozzi S, et al. Morphotype-based risk stratification in patients with patent foramen ovale using computational fluid dynamics. Sci Rep. 2025;15:34557. Stasek J, Bis J, Dusek J, Medilek K, Dostal J, Branny M, et al. Evaluation of the severity of right-to-left shunt in PFO patients after systemic embolism (MEASURE-PFO study): Study design. Pol Heart J. 2024;82:774–6. Mitchell C, Rahko PS, Blauwet LA, Canaday B, Finstuen JA, Foster MC, et al. Guidelines for Performing a Comprehensive Transthoracic Echocardiographic Examination in Adults: Recommendations from the American Society of Echocardiography. J Am Soc Echocardiogr. 2019;32:1–64. Robinson S, Rana B, Oxborough D, Steeds R, Monaghan M, Stout M, et al. A practical guideline for performing a comprehensive transthoracic echocardiogram in adults: the British Society of Echocardiography minimum dataset. Echo Res Pract. 2020;7:G59–93. Hahn RT, Abraham T, Adams MS, Bruce CJ, Glas KE, Lang RM, et al. Guidelines for Performing a Comprehensive Transesophageal Echocardiographic Examination: Recommendations from the American Society of Echocardiography and the Society of Cardiovascular Anesthesiologists. J Am Soc Echocardiogr. 2013;26:921–694. Wheeler R, Steeds R, Rana B, Wharton G, Smith N, Allen J, et al. A minimum dataset for a standard transoesophageal echocardiogram: a guideline protocol from the British Society of Echocardiography. Echo Res Pract. 2015;2:G29–45. Mojadidi MK, Bogush N, Caceres JD, Msaouel P, Tobis JM. Diagnostic Accuracy of Transesophageal Echocardiogram for the Detection of Patent Foramen Ovale: A Meta-Analysis. Echocardiography. 2014;31:752–8. Ghazal SN. Valsalva maneuver in echocardiography. J Echocardiogr. 2017;15:1–5. Rodrigues AC, Picard MH, Carbone A, Arruda AL, Flores T, Klohn J, et al. Importance of Adequately Performed Valsalva Maneuver to Detect Patent Foramen Ovale during Transesophageal Echocardiography. J Am Soc Echocardiogr. 2013;26:1337–43. Belkin RN, Pollack BD, Ruggiero ML, Alas LL, Tatini U. Comparison of transesophageal and transthoracic echocardiography with contrast and color flow Doppler in the detection of patent foramen ovale. Am Heart J. 1994;128:520–5. Maffe S, Dellavesa P, Zenone F, Paino AM, Paffoni P, Perucca A, et al. Transthoracic second harmonic two- and three-dimensional echocardiography for detection of patent foramen ovale. Eur J Echocardiogr. 2010;11:57–63. Yang X, Wang H, Wei Y, Zhai N, Liu B, Li X. Diagnosis of Patent Foramen Ovale: The Combination of Contrast Transcranial Doppler, Contrast Transthoracic Echocardiography, and Contrast Transesophageal Echocardiography. BioMed Res Int. 2020. 10.1155/2020/8701759 . Song JK. Pearls and Pitfalls in the Transesophageal Echocardiographic Diagnosis of Patent Foramen Ovale. J Am Soc Echocardiogr. 2023;36:895–e9053. Mojadidi MK, Winoker JS, Roberts SC, Msaouel P, Gevorgyan R, Zolty R. Two-dimensional echocardiography using second harmonic imaging for the diagnosis of intracardiac right-to-left shunt: a meta-analysis of prospective studies. Int J Cardiovasc Imaging. 2014;30:911–23. Mojadidi MK, Winoker JS, Roberts SC, Msaouel P, Zaman MO, Gevorgyan R, et al. Accuracy of Conventional Transthoracic Echocardiography for the Diagnosis of Intracardiac Right-to‐Left Shunt: A Meta‐Analysis of Prospective Studies. Echocardiography. 2014;31:1036–48. Katsanos AH, Psaltopoulou T, Sergentanis TN, Frogoudaki A, Vrettou A, Ikonomidis I, et al. Transcranial Doppler versus transthoracic echocardiography for the detection of patent foramen ovale in patients with cryptogenic cerebral ischemia: A systematic review and diagnostic test accuracy meta-analysis. Ann Neurol. 2016;79:625–35. Gentile M, De Vito A, Azzini C, Tamborino C, Casetta I. Adding Blood to Agitated Saline Significantly Improves Detection of Right-to-Left Shunt by Contrast-Transcranial Color-Coded Duplex Sonography. Ultrasound Med Biol. 2014;40:2637–41. Moeller C, Fleischmann C, Thomas-Rueddel D, Vlasakov V, Rochwerg B, Theurer P, et al. How safe is gelatin? A systematic review and meta-analysis of gelatin-containing plasma expanders vs crystalloids and albumin. J Crit Care. 2016;35:75–83. Saw MM, Chandler B, Ho KM. Benefits and Risks of Using Gelatin Solution as a Plasma Expander for Perioperative and Critically Ill Patients: A Meta-Analysis. Anaesth Intensive Care. 2012;40:17–32. Dubrey S, Dahdal G, Grocottmason R. Severe anaphylaxis to Gelofusine during a transthoracic echo bubble study. Eur J Echocardiogr. 2006. 10.1016/j.euje.2006.11.003 . Mullins RJ, James H, Platts-Mills TAE, Commins S. Relationship between red meat allergy and sensitization to gelatin and galactose-α-1,3-galactose. J Allergy Clin Immunol. 2012;129:1334–e13421. Agmon Y, Khandheria BK, Meissner I, Gentile F, Whisnant JP, Sicks JD, et al. Frequency of Atrial Septal Aneurysms in Patients With Cerebral Ischemic Events. Circulation. 1999;99:1942–4. Overell JR, Bone I, Lees KR. Interatrial septal abnormalities and stroke: A meta-analysis of case-control studies. Neurology. 2000;55:1172–9. Nakayama R, Takaya Y, Akagi T, Watanabe N, Ikeda M, Nakagawa K, et al. Identification of High-Risk Patent Foramen Ovale Associated With Cryptogenic Stroke: Development of a Scoring System. J Am Soc Echocardiogr. 2019;32:811–6. Sun PY, Tobis JM, Daneshvar SA, Alfonso RC, Liebeskind DS, Saver JL. Quantification of patent foramen ovale shunt severity by transesophageal echocardiogram and transcranial doppler in routine clinical practice. J Stroke Cerebrovasc Dis. 2025;34:108306. Gwak SY, Kim K, Lee HJ, Cho I, Hong GR, Ha JW, et al. Three-dimensional agitated saline contrast transesophageal echocardiography for the diagnosis of patent foramen ovale. Sci Rep. 2025;15:29136. Additional Declarations No competing interests reported. Cite Share Download PDF Status: Under Review Version 1 posted Editorial decision: Revision requested 06 Feb, 2026 Reviews received at journal 06 Feb, 2026 Reviewers agreed at journal 28 Jan, 2026 Reviewers agreed at journal 15 Dec, 2025 Reviewers invited by journal 11 Dec, 2025 Editor assigned by journal 11 Dec, 2025 Submission checks completed at journal 09 Dec, 2025 First submitted to journal 07 Dec, 2025 You are reading this latest preprint version Research Square lets you share your work early, gain feedback from the community, and start making changes to your manuscript prior to peer review in a journal. As a division of Research Square Company, we’re committed to making research communication faster, fairer, and more useful. We do this by developing innovative software and high quality services for the global research community. Our growing team is made up of researchers and industry professionals working together to solve the most critical problems facing scientific publishing. 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08:43:27","extension":"xml","order_by":7,"title":"","display":"","copyAsset":false,"role":"acdc-reference","size":129990,"visible":true,"origin":"","legend":"","description":"","filename":"583f9f22bee2429a8ce30e49fc5911e41structuring.xml","url":"https://assets-eu.researchsquare.com/files/rs-8302087/v1/c0a1603daf6adb6a440bc166.xml"},{"id":98746372,"identity":"43f85e0d-c535-4717-8600-9a84bfee04b0","added_by":"auto","created_at":"2025-12-22 08:43:27","extension":"html","order_by":8,"title":"","display":"","copyAsset":false,"role":"acdc-reference","size":141626,"visible":true,"origin":"","legend":"","description":"","filename":"earlyproof.html","url":"https://assets-eu.researchsquare.com/files/rs-8302087/v1/661e9b6290fa7cab7fe29919.html"},{"id":98746361,"identity":"c355f47c-2a00-4c2d-ba68-3fbe72a4f48f","added_by":"auto","created_at":"2025-12-22 08:43:27","extension":"png","order_by":1,"title":"Figure 1","display":"","copyAsset":false,"role":"figure","size":481022,"visible":true,"origin":"","legend":"\u003cp\u003eLeft heart contrast opacification grading. A - cTTE grade 1, B cTTE grade 2, C - cTTE grade 3, D - cTOE grade 1, E - cTOE grade 2, F - cTOE grade 3. Small images in D-E document visualisation of the contrast passing through the PFO before the left heart opacification in cTOE.\u003c/p\u003e","description":"","filename":"1.png","url":"https://assets-eu.researchsquare.com/files/rs-8302087/v1/b5404b5f28aca00211b31808.png"},{"id":98746362,"identity":"6694f69e-083a-454f-9e4e-8e8e2cd64c14","added_by":"auto","created_at":"2025-12-22 08:43:27","extension":"png","order_by":2,"title":"Figure 2","display":"","copyAsset":false,"role":"figure","size":785179,"visible":true,"origin":"","legend":"\u003cp\u003eContrast density immediately after agitation. A - normal saline, B - 10% dextrose, C – Mannitol 20%, D - Gelofusine 4%\u003c/p\u003e","description":"","filename":"2.png","url":"https://assets-eu.researchsquare.com/files/rs-8302087/v1/6bbb4d333ddb5b18323893e4.png"},{"id":100406019,"identity":"3d8a4bf7-09e8-4461-94d5-cf110e446f70","added_by":"auto","created_at":"2026-01-16 12:33:31","extension":"pdf","order_by":0,"title":"","display":"","copyAsset":false,"role":"manuscript-pdf","size":2596607,"visible":true,"origin":"","legend":"","description":"","filename":"manuscript.pdf","url":"https://assets-eu.researchsquare.com/files/rs-8302087/v1/4dc3bb27-12f3-4d84-98d7-a147a7c678ad.pdf"}],"financialInterests":"No competing interests reported.","formattedTitle":"High-density bubble echocardiography for patent foramen ovale assessment: sensitivity and relationship with invasively measured shunt severity and atrial septum morphology","fulltext":[{"header":"Background","content":"\u003cp\u003ePatent foramen ovale (PFO) is a flap-like opening resulting from incomplete fusion of the septum primum and septum secundum at the fossa ovalis. Its overall reported prevalence on autopsy is 27%, with a positive relationship between PFO and cryptogenic stroke (CS) observed in up to 46% of individuals\u0026thinsp;\u0026lt;\u0026thinsp;55 years of age and 21% in those\u0026thinsp;\u0026gt;\u0026thinsp;55 years. (\u003cspan citationid=\"CR1\" class=\"CitationRef\"\u003e1\u003c/span\u003e, \u003cspan citationid=\"CR2\" class=\"CitationRef\"\u003e2\u003c/span\u003e) In the RESPECT trial, closure of a PFO was associated with a lower rate of recurrent ischaemic strokes than medical therapy alone, providing evidence for the benefit of PFO closure in selected patients with CS. (\u003cspan citationid=\"CR3\" class=\"CitationRef\"\u003e3\u003c/span\u003e)\u003c/p\u003e \u003cp\u003eThe contrast echocardiography effect was first described in an experimental in vitro model in 1968 with injected saline, and shortly afterwards with 5% dextrose and agitated patient\u0026rsquo;s blood. (\u003cspan additionalcitationids=\"CR5\" citationid=\"CR4\" class=\"CitationRef\"\u003e4\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR6\" class=\"CitationRef\"\u003e6\u003c/span\u003e) Twelve years later, an experimental study confirmed that the echo reflection in a contrast study was primarily due to microbubbles created during mixing of these agents with gas. (\u003cspan citationid=\"CR7\" class=\"CitationRef\"\u003e7\u003c/span\u003e) Longer stability of contrast was observed in gelatine due to lower surface tension. The non-transpulmonary contrast agents currently in use for PFO detection contain bubbles generally too large to pass through the pulmonary circulation. (\u003cspan citationid=\"CR8\" class=\"CitationRef\"\u003e8\u003c/span\u003e, \u003cspan citationid=\"CR9\" class=\"CitationRef\"\u003e9\u003c/span\u003e)\u003c/p\u003e \u003cp\u003eContrast transcranial doppler (cTCD), transoesophageal (cTOE), and transthoracic (cTTE) echocardiography represent the recommended methods for right-to-left intracardiac shunt assessment. In most cases, precise PFO diagnosis requires the combined use of different techniques, with cTCD or cTTE recommended as the first-line method. (\u003cspan citationid=\"CR10\" class=\"CitationRef\"\u003e10\u003c/span\u003e) Due to technical improvements, the sensitivity of cTTE has increased over the last 30 years from 51% to 80%. (\u003cspan citationid=\"CR11\" class=\"CitationRef\"\u003e11\u003c/span\u003e) According to meta-analyses cited in the latest European Society of Cardiology PFO statement and European Stroke Organisation guidelines, the sensitivity of cTTE in comparison with cTOE is reported as 88% and 71%. (\u003cspan citationid=\"CR10\" class=\"CitationRef\"\u003e10\u003c/span\u003e, \u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e12\u003c/span\u003e) In the latter meta-analysis, colloid contrast was used in 4/20 studies, all of them published more than 20 years ago, with a reported sensitivity of 13\u0026ndash;61%. (\u003cspan citationid=\"CR10\" class=\"CitationRef\"\u003e10\u003c/span\u003e) On the other hand, some papers have reported better sensitivity of colloid contrast in comparison with normal saline. (\u003cspan citationid=\"CR13\" class=\"CitationRef\"\u003e13\u003c/span\u003e, \u003cspan citationid=\"CR14\" class=\"CitationRef\"\u003e14\u003c/span\u003e) It is also reported that the diagnostic performance of cTOE might improve with larger volume and number of contrast boluses administered. (\u003cspan citationid=\"CR15\" class=\"CitationRef\"\u003e15\u003c/span\u003e, \u003cspan citationid=\"CR16\" class=\"CitationRef\"\u003e16\u003c/span\u003e)\u003c/p\u003e \u003cp\u003eThe risk of a PFO being a causative pathology in patients with CS is usually linked to the left-heart opacification degree, PFO tunnel width and length, presence of concomitant interatrial aneurysm or hypermobile septum, and a prominent Eustachian valve or Chiari\u0026rsquo;s network. (\u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e12\u003c/span\u003e, \u003cspan additionalcitationids=\"CR18\" citationid=\"CR17\" class=\"CitationRef\"\u003e17\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR19\" class=\"CitationRef\"\u003e19\u003c/span\u003e) Severe shunt is usually defined by the appearance of more than 20\u0026ndash;30 microbubbles in the left atrium within three cardiac cycles after opacification of the right atrium. (\u003cspan citationid=\"CR20\" class=\"CitationRef\"\u003e20\u003c/span\u003e, \u003cspan citationid=\"CR21\" class=\"CitationRef\"\u003e21\u003c/span\u003e) The interplay between PFO morphology and invasively measured shunt severity or left-heart bubble contrast opacification degree has not been clinically studied, apart from an experimental model investigating fluid dynamics to assess the impact of PFO tunnel geometry on PFO flow behaviour. (\u003cspan citationid=\"CR22\" class=\"CitationRef\"\u003e22\u003c/span\u003e)\u003c/p\u003e \u003cp\u003eWe aimed to investigate the relationship between left-heart bubble opacification degree using a high-density non-transpulmonary contrast agent, invasively measured shunt severity, and atrial septum morphology, and to assess the sensitivity of cTTE using high-density contrast with and without Valsalva manoeuvre for PFO detection.\u003c/p\u003e"},{"header":"Methods","content":"\u003cdiv id=\"Sec3\" class=\"Section2\"\u003e \u003ch2\u003eStudy population\u003c/h2\u003e \u003cp\u003eThe MEASURE-PFO study (in print) included 151 consecutive patients with CS or systemic embolisation and PFO proven by cTOE in six tertiary centres in the Czech Republic. All patients underwent invasive measurement of the right-to-left shunt and successful PFO closure. For contrast echocardiography studies, high-density contrast (Gelofusine, mannitol), dextrose and normal saline (without blood) were used, and cTOE was performed. The indication for PFO assessment was at the discretion of the referring neurologist. All patients signed informed consent form. In our substudy, we enrolled 55 patients from three cardiac centres in whom high-density contrast was used and cTTE was available,\u003c/p\u003e \u003c/div\u003e\n\u003ch3\u003eCardiac catheterisation\u003c/h3\u003e\n\u003cp\u003eFemoral access was used for the catheterisation procedure. The pressure in the aorta was measured with the Sensis\u0026reg; hemodynamic system (Siemens Healthineers AG, Erlangen, Germany). The systemic blood flow, pulmonary blood flow and right-to-left (R-L) shunt were determined using the thermodilution Inntherm\u0026reg; system at rest and during Valsalva manoeuvre, if necessary, facilitated by another person pressing their palm against the patient\u0026rsquo;s abdomen. At rest, 10 ml of ice-cold saline was injected into the inferior vena cava near its opening into the right atrium. During the Valsalva manoeuvre, the systemic pressure was monitored, and at the time of the short pressure drop to about 50% of the initial systemic pressure, the manoeuvre was stopped and the saline was injected. The temperature of the systemic blood was measured by a thermal probe positioned slightly below the aortic arch in the descending aorta. The presence and quantification of the R-L shunt were determined by the blood temperature change in the descending aorta as published previously (\u003cspan citationid=\"CR23\" class=\"CitationRef\"\u003e23\u003c/span\u003e).\u003c/p\u003e\n\u003ch3\u003eEchocardiography\u003c/h3\u003e\n\u003cp\u003ecTOE and cTTE were performed before PFO closure using Vivid E95 (GE\u0026reg;) with 6Vc-D TTE and 6CT-D TOE probes and EPIQ CVXi and CX50 with X5-1 or X51c TTE and X7-2t or X8-2t TOE probes (Philips\u0026reg;). All patients signed a TOE informed consent form. Peripheral line access, at least 20G size, was preferably placed in the cubital vein, with no side preference.\u003c/p\u003e \u003cp\u003eStandard TTE protocol was performed according to recommendations. (\u003cspan citationid=\"CR24\" class=\"CitationRef\"\u003e24\u003c/span\u003e, \u003cspan citationid=\"CR25\" class=\"CitationRef\"\u003e25\u003c/span\u003e) For the contrast study, retrospective gated acquisition was chosen, with at least ten RR intervals set up. Nineteen mL of Gelofusine B Braun 4% or Mannitol Fresenius Kabi 20% and 1 mL of air were agitated using two 20 ml luer lock syringes connected by a stopcock until a white suspension was obtained. A resting image was obtained after a quick contrast bolus was administered into the straight iv line immediately after it was prepared. At least 5 RR intervals of the whole 10 RR loop were acquired after the opacification of the right atrium using retrospective acquisition. If no bubbles were visualised in the left heart, the test was repeated. In the next step, patients were trained to perform a Valsalva manoeuvre by taking in a deep breath, closing the mouth, and \u0026ldquo;bearing down\u0026rdquo; as much as they could against the echocardiographer\u0026rsquo;s fist pushed to the patient\u0026rsquo;s abdominal wall, followed by abrupt release. Afterwards, patients were asked to make a Valsalva manoeuvre with the same force but with no help from the operator, who was performing TTE and made sure to keep all four chambers in the apical four-chamber view visible during the Valsalva strain and release phases. This was practised several times. The contrast was quickly injected at the initiation of the Valsalva manoeuvre, which was abruptly released once the whole 20 mL of contrast had been administered, at the time of the appearance/contrast opacification of the right atrium (usually 7\u0026ndash;8 s after initialisation of the Valsalva manoeuvre). If no bubbles appeared in the left heart or in the case of image deterioration during acquisition due to translation movements, the test was repeated.\u003c/p\u003e \u003cp\u003eFor TOE, pharyngeal anaesthesia with lidocaine spray was performed and standard protocol was followed. (\u003cspan citationid=\"CR26\" class=\"CitationRef\"\u003e26\u003c/span\u003e, \u003cspan citationid=\"CR27\" class=\"CitationRef\"\u003e27\u003c/span\u003e) When accomplished, the interatrial septum was visualized using the mid-oesophageal short axis 40\u0026ndash;60\u0026deg; view and the rest contrast study using the same setting as in cTTE was performed. If negative, the test was repeated in bi-caval view. Next, the Valsalva study was performed in short axis 40\u0026ndash;60\u0026deg; view. If negative, it was repeated in bi-caval view and also repeated if necessary. It had to be visualised that the contrast was passing to the left atrium via PFO canal to confirm its presence. If this was not the case, every effort was made to localize the site of the shunt (multiperforated PFO etc.) and to rule out contrast inflow from the pulmonary vein.\u003c/p\u003e \u003cp\u003eThe left heart contrast opacification grading was classified as follows: grade 0: no bubbles, grade 1: mild, grade 2: moderate, grade 3: severe left atrium contrast opacification in cTOE and left atrium/ventricle opacification in cTTE. (Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003e) The single frame with the largest number of bubbles in the left atrium (cTOE) or left heart (cTTE) was used for opacification grading.\u003c/p\u003e \u003cp\u003eThe length of the PFO channel and the septum primum and secundum separation width at rest in the mid-oesophageal short-axis 40\u0026ndash;60\u0026deg; and bi-caval projections were measured. The larger diameter was considered. The presence of the Eustachian valve or Chiari\u0026rsquo;s network was recorded; atrial septal aneurysm was defined as a 10 mm bulging of the septum tissue to the left/right side. The aim was to avoid sedation to ensure a good-quality Valsalva manoeuvre during cTOE, confirmed by the presence of atrial shifting to the left. If needed, a minimum dose of midazolam allowing completion of the whole cTOE with reliable Valsalva manoeuvre was titrated in 1 mg increments.\u003c/p\u003e \u003cp\u003eA positive study was defined as the presence of contrast in the left atrium (cTOE)/left heart (cTTE) within three cycles after the right heart opacification in the rest study, or after strain release in the Valsalva study. Low density bubbles coming into the left atrium/left ventricle later than 3 cycles after the contrast administration/strain relief were not considered as a positive study. All images were assessed in a core lab by an experienced echocardiographer holding EACVI accreditation in adult TTE with exposure to PFO contrast studies over a period of almost twenty years.\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003cdiv id=\"Sec6\" class=\"Section2\"\u003e \u003ch2\u003eStatistical analysis\u003c/h2\u003e \u003cp\u003eThe statistical analysis in the study involved summarising continuous variables as means with standard deviations and categorical variables as frequencies and percentages. Normality was assessed using the Shapiro-Wilk test. For comparisons across multiple groups, ANOVA on ranks (Kruskal-Wallis test) was applied when normality assumptions were not met, followed by Dunn\u0026rsquo;s post hoc test with Bonferroni correction. Categorical variables were evaluated using the chi-squared test, with Fisher\u0026rsquo;s exact test applied when expected frequencies were below 5. Cram\u0026eacute;r's V was used to assess the strength of the association between categorical variables. The agreement between cTTE and cTOE for left heart opacification was assessed using Kendall\u0026rsquo;s τ coefficient, with statistical significance determined at p\u0026thinsp;\u0026lt;\u0026thinsp;0.05. Systematic differences between cTTE and cTOE were evaluated using McNemar\u0026rsquo;s test. Statistical significance was set at p\u0026thinsp;\u0026lt;\u0026thinsp;0.05. All statistical analyses were performed using SPSS Statistics software, version 26 (IBM Corp., Armonk, NY, USA). Graphical analyses were performed using SigmaPlot, version 14.\u003c/p\u003e \u003c/div\u003e"},{"header":"Results","content":"\u003cp\u003eFifty-five patients fulfilling the inclusion criteria were enrolled in the study. Their baseline characteristics are summarised in Table\u0026nbsp;\u003cspan refid=\"Tab1\" class=\"InternalRef\"\u003e1\u003c/span\u003e.\u003c/p\u003e \u003cp\u003e \u003cdiv class=\"gridtable\"\u003e\u003ctable float=\"Yes\" id=\"Tab1\" border=\"1\"\u003e \u003ccaption language=\"En\"\u003e \u003cdiv class=\"CaptionNumber\"\u003eTable 1\u003c/div\u003e \u003cdiv class=\"CaptionContent\"\u003e \u003cp\u003eBaseline characteristics of the studied group\u003c/p\u003e \u003c/div\u003e \u003c/caption\u003e \u003ccolgroup cols=\"2\"\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e \u003cthead\u003e \u003ctr\u003e \u003cth align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/th\u003e \u003cth align=\"left\" colname=\"c2\"\u003e \u003cp\u003en (% N) / mean\u0026thinsp;\u0026plusmn;\u0026thinsp;SD\u003c/p\u003e \u003c/th\u003e \u003c/tr\u003e \u003c/thead\u003e \u003ctbody\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eage\u0026nbsp;(years)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e44.9 \u0026plusmn; (7.6)\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003egender\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e\u0026nbsp;\u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003emale\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e38 (69.1%)\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003efemale\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e17 (30.9%)\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eBMI\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e28.3\u0026thinsp;\u0026plusmn;\u0026thinsp;5.1\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eBSA\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e2.1\u0026thinsp;\u0026plusmn;\u0026thinsp;0.2\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003ecreatinine\u0026nbsp;(mmol/l)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e78.3\u0026thinsp;\u0026plusmn;\u0026thinsp;14.2\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003ehaemoglobin\u0026nbsp;(g/l)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e143\u0026thinsp;\u0026plusmn;\u0026thinsp;14\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eheight\u0026nbsp;(cm)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e178\u0026thinsp;\u0026plusmn;\u0026thinsp;9\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eweight\u0026nbsp;(kg)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e89.9\u0026thinsp;\u0026plusmn;\u0026thinsp;18\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003easthma bronchiale or COPD\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e3 (5.5%)\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003ediabetes\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e3 (5.5%)\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003ehyperlipidaemia\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e26 (47.3%)\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003ehypertension\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e16 (29.1%)\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003econtrast site injection\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e\u0026nbsp;\u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eperipheral line\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e53 (96.3%)\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003einferior vena cava\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e2 (3.7%)\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003etype of contrast\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e\u0026nbsp;\u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eGelofusine 4%\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e37 (67%)\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eMannitol 20%\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e18 (33%)\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003esedation during cTOE\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e12 (18.5%)\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003esedation dose (ml)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e2.6\u0026thinsp;\u0026plusmn;\u0026thinsp;0.7\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eChiari network/Eustachian valve\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e24 (43.6%)\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003ePFO\u0026nbsp;channel\u0026nbsp;length\u0026nbsp;(mm)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e13.2\u0026thinsp;\u0026plusmn;\u0026thinsp;4.7\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003ePFO\u0026nbsp;channel\u0026nbsp;width\u0026nbsp;(mm)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e2.9\u0026thinsp;\u0026plusmn;\u0026thinsp;2.0\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eatrial septum aneurysm\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e29 (52.7%)\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003c/tbody\u003e \u003c/colgroup\u003e \u003c/table\u003e\u003c/div\u003e \u003c/p\u003e \u003cp\u003eBMI - body mass index, BSA - body surface area, COPD - chronic obstructive pulmonary disease\u003c/p\u003e \u003cp\u003eSensitivity of rest\u0026thinsp;+\u0026thinsp;Valslava cTOE was 100%, as all patients underwent successful PFO occlusion. cTTE with Valsalva manoeuvre was found to be highly sensitive for PFO detection. By contrast, rest cTTE/cTOE bubble studies showed low sensitivity. (Table\u0026nbsp;\u003cspan refid=\"Tab2\" class=\"InternalRef\"\u003e2\u003c/span\u003e)\u003c/p\u003e \u003cp\u003e \u003cdiv class=\"gridtable\"\u003e\u003ctable float=\"Yes\" id=\"Tab2\" border=\"1\"\u003e \u003ccaption language=\"En\"\u003e \u003cdiv class=\"CaptionNumber\"\u003eTable 2\u003c/div\u003e \u003cdiv class=\"CaptionContent\"\u003e \u003cp\u003eSensitivity of high-density bubble studies for PFO detection\u003c/p\u003e \u003c/div\u003e \u003c/caption\u003e \u003ccolgroup cols=\"3\"\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e \u003cdiv align=\"char\" char=\".\" class=\"colspec\" colname=\"c3\" colnum=\"3\"\u003e\u003c/div\u003e \u003cthead\u003e \u003ctr\u003e \u003cth align=\"left\" colname=\"c1\"\u003e \u003cp\u003emethod A\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c2\"\u003e \u003cp\u003emethod B\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c3\"\u003e \u003cp\u003esensitivity A vs B\u003c/p\u003e \u003c/th\u003e \u003c/tr\u003e \u003c/thead\u003e \u003ctbody\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003ecTTE rest\u0026thinsp;+\u0026thinsp;Valsalva\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003ecTOE rest\u0026thinsp;+\u0026thinsp;Valsalva\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e \u003cp\u003e98.2%\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003ecTTE Valsalva\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003ecTOE Valsalva\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e \u003cp\u003e97.6%\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003ecTTE rest\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003ecTOE rest\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e \u003cp\u003e83.6%\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003ecTOE rest\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003ecTOE rest\u0026thinsp;+\u0026thinsp;Valsalva\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e \u003cp\u003e77.3%\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003c/tbody\u003e \u003c/colgroup\u003e \u003c/table\u003e\u003c/div\u003e \u003c/p\u003e \u003cp\u003eNo relationship between the invasively measured shunt severity (0%, \u0026lt;\u0026thinsp;10%, 10\u0026ndash;19%, \u0026ge;\u0026thinsp;20%) and left heart bubble opacification grade was observed either for rest or Valsalva cTTE or cTOE studies (Table\u0026nbsp;\u003cspan refid=\"Tab3\" class=\"InternalRef\"\u003e3\u003c/span\u003e). When the shunt severity was categorised as \u0026lt;\u0026thinsp;10% and \u0026ge;\u0026thinsp;10%, only a moderate association was revealed between the rest\u0026thinsp;+\u0026thinsp;Valsalva studies opacification grade and shunt severity in cTOE (p\u0026thinsp;=\u0026thinsp;0.024, Cram\u0026eacute;r's coefficient 0.368), but not in the cTTE group (Table\u0026nbsp;\u003cspan refid=\"Tab4\" class=\"InternalRef\"\u003e4\u003c/span\u003e).\u003c/p\u003e \u003cp\u003e \u003cdiv class=\"gridtable\"\u003e\u003ctable float=\"Yes\" id=\"Tab3\" border=\"1\"\u003e \u003ccaption language=\"En\"\u003e \u003cdiv class=\"CaptionNumber\"\u003eTable 3\u003c/div\u003e \u003cdiv class=\"CaptionContent\"\u003e \u003cp\u003eShunt severity vs left heart contrast opacification grade for the rest and Valsalva cTTE/cTOE\u003c/p\u003e \u003c/div\u003e \u003c/caption\u003e \u003ccolgroup cols=\"6\"\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c3\" colnum=\"3\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c4\" colnum=\"4\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c5\" colnum=\"5\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c6\" colnum=\"6\"\u003e\u003c/div\u003e \u003ctbody\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colspan=\"4\" nameend=\"c5\" namest=\"c2\"\u003e \u003cp\u003eseverity of R-L shunt\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e\u0026nbsp;\u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003enot detected\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e\u0026lt;\u0026nbsp;10%\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e10\u0026ndash;19%\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e\u0026ge;\u0026thinsp;20%\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003ep\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cb\u003ebubbles left heart opacification rest cTTE\u003c/b\u003e n\u0026thinsp;=\u0026thinsp;55\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\" morerows=\"4\" rowspan=\"5\"\u003e \u003cp\u003e0.16\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eGrade 0: no bubbles\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e3 (50.0%)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e4 (19.0%)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e1 (6,7%)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e1 (7.7%)\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eGrade 1: mild opacification\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e1 (16.7%)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e9 (42.9%)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e4 (26.7%)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e4 (30.8%)\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eGrade 2: moderate opacification\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e0 (0.0%)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e7 (33,3%)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e6 (40.0%)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e5 (38.5%)\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eGrade 3: significant opacification\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e2 (33.3%)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e1 (4,8%)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e4 (26.7%)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e3 (23.1%)\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cb\u003ebubbles left heart opacification Valslava cTTE\u003c/b\u003e n\u0026thinsp;=\u0026thinsp;44\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\" morerows=\"4\" rowspan=\"5\"\u003e \u003cp\u003e0.20\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eGrade 0: no bubbles\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e1 (25.0%)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e2 (10.5%)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e0 (0.0%)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e1 (11.1%)\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eGrade 1: mild opacification\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e1 (25.0%)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e1 (5.3%)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e1 (8.3%)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e0 (0.0%)\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eGrade 2: moderate opacification\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e0 (0.0%)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e10 (52.6%)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e2 (16.7%)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e4 (44.4%)\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eGrade 3: significant opacification\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e2 (50.0%)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e6 (31.6%)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e9 (75.0%)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e4 (44.4%)\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cb\u003ebubbles left atrium opacification rest cTOE\u003c/b\u003e n\u0026thinsp;=\u0026thinsp;55\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\" morerows=\"4\" rowspan=\"5\"\u003e \u003cp\u003e0.10\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eGrade 0: no bubbles\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e2 (33.3%)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e6 (28.6%)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e1 (6,7%)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e2 (15.4%)\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eGrade 1: mild opacification\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e2 (33.3%)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e9 (42.9%)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e6 (40.0%)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e7 (53.8%)\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eGrade 2: moderate opacification\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e0 (0.0%)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e6 (28.6%)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e6 (40.0%)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e4 (30.5%)\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eGrade 3: significant opacification\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e2 (33.3%)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e0 (0.0%)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e2 (13.3%)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e0 (0.0%)\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cb\u003ebubbles left atrium opacification Valsalva cTOE\u003c/b\u003e n\u0026thinsp;=\u0026thinsp;41\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\" morerows=\"4\" rowspan=\"5\"\u003e \u003cp\u003e0.26\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eGrade 0: no bubbles\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e0 (0.0%)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e0 (0.0%)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e0 (0.0%)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e0 (0.0%)\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eGrade 1: mild opacification\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e2 (50.0%)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e9 (50.0%)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e1 (10.0%)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e2 (22.2%)\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eGrade 2: moderate opacification\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e2 (50.0%)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e7 (38.9%)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e5 (50.0%)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e5 (55.6%)\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eGrade 3: significant opacification\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e0 (0.0%)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e2 (11.1%)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003e4 (40.0%)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e2 (22.2%)\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003c/tbody\u003e \u003c/colgroup\u003e \u003ctfoot\u003e \u003ctr\u003e\u003ctd colspan=\"6\"\u003eChi-squared test. R-L - right to left\u003c/td\u003e\u003c/tr\u003e \u003c/tfoot\u003e \u003c/table\u003e\u003c/div\u003e \u003c/p\u003e \u003cp\u003e \u003cdiv class=\"gridtable\"\u003e\u003ctable float=\"Yes\" id=\"Tab4\" border=\"1\"\u003e \u003ccaption language=\"En\"\u003e \u003cdiv class=\"CaptionNumber\"\u003eTable 4\u003c/div\u003e \u003cdiv class=\"CaptionContent\"\u003e \u003cp\u003e\u0026thinsp;\u0026lt;\u0026thinsp;10% and \u0026ge;\u0026thinsp;10% shunt severity vs contrast opacification grade for the rest\u0026thinsp;+\u0026thinsp;Valsalva cTTE/cTOE\u003c/p\u003e \u003c/div\u003e \u003c/caption\u003e \u003ccolgroup cols=\"4\"\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c3\" colnum=\"3\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c4\" colnum=\"4\"\u003e\u003c/div\u003e \u003ctbody\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colspan=\"2\" nameend=\"c3\" namest=\"c2\"\u003e \u003cp\u003eseverity of R-L shunt\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e\u0026nbsp;\u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e\u0026lt;\u0026nbsp;10%\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e\u0026ge;\u0026thinsp;10%\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\"\u003e \u003cp\u003ep\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cb\u003ebubbles left heart opacification rest\u0026thinsp;+\u0026thinsp;Valsalva cTTE\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\" morerows=\"4\" rowspan=\"5\"\u003e \u003cp\u003e0.206\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eGrade 0: no bubbles\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e1 (3.7%)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e\u0026nbsp;\u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eGrade 1: mild opacification\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e4 (14.8%)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e1 (3.6%)\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eGrade 2: moderate opacification\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e12 (44.5%)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e11 (39.3%)\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eGrade 3: significant opacification\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e10 (37.0%)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e16 (57.1%)\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cb\u003ebubbles left heart opacification rest\u0026thinsp;+\u0026thinsp;Valsalva cTOE\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e\u0026nbsp;\u003c/td\u003e \u003ctd align=\"left\" colname=\"c4\" morerows=\"3\" rowspan=\"4\"\u003e \u003cp\u003e0.024\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eGrade 1: mild opacification\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e11 (40.7%)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e3 (10.7%)\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eGrade 2: moderate opacification\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e12 (44.4%)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e15 (53.6%)\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003eGrade 3: significant opacification\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003e4 (14.8%)\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003e10 (35.7%)\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003c/tbody\u003e \u003c/colgroup\u003e \u003ctfoot\u003e \u003ctr\u003e\u003ctd colspan=\"4\"\u003eChi-squared test, R-L - right to left\u003c/td\u003e\u003c/tr\u003e \u003c/tfoot\u003e \u003c/table\u003e\u003c/div\u003e \u003c/p\u003e \u003cp\u003eNo significant association was also found between the left heart opacification grade and the presence of an atrial septal aneurysm, Eustachian valve/Chiari\u0026acute;s network, or PFO width and length. (Table\u0026nbsp;\u003cspan refid=\"Tab5\" class=\"InternalRef\"\u003e5\u003c/span\u003e).\u003c/p\u003e \u003cp\u003e \u003cdiv class=\"gridtable\"\u003e\u003ctable float=\"Yes\" id=\"Tab5\" border=\"1\"\u003e \u003ccaption language=\"En\"\u003e \u003cdiv class=\"CaptionNumber\"\u003eTable 5\u003c/div\u003e \u003cdiv class=\"CaptionContent\"\u003e \u003cp\u003eRelationship between atrial septal morphology and the left heart opacification grade, p value.\u003c/p\u003e \u003c/div\u003e \u003c/caption\u003e \u003ccolgroup cols=\"5\"\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e \u003cdiv align=\"char\" char=\".\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e \u003cdiv align=\"char\" char=\".\" class=\"colspec\" colname=\"c3\" colnum=\"3\"\u003e\u003c/div\u003e \u003cdiv align=\"char\" char=\".\" class=\"colspec\" colname=\"c4\" colnum=\"4\"\u003e\u003c/div\u003e \u003cdiv align=\"char\" char=\".\" class=\"colspec\" colname=\"c5\" colnum=\"5\"\u003e\u003c/div\u003e \u003cthead\u003e \u003ctr\u003e \u003cth align=\"left\" colname=\"c1\" morerows=\"1\" rowspan=\"2\"\u003e\u0026nbsp;\u003c/th\u003e \u003cth align=\"left\" colspan=\"4\" nameend=\"c5\" namest=\"c2\"\u003e \u003cp\u003eanatomical interatrial septum/PFO variations\u003c/p\u003e \u003c/th\u003e \u003c/tr\u003e \u003ctr\u003e \u003cth align=\"left\" colname=\"c2\"\u003e \u003cp\u003ePFO width\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c3\"\u003e \u003cp\u003ePFO length\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c4\"\u003e \u003cp\u003eChiari\u0026acute;s network/ Eustachian valve\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c5\"\u003e \u003cp\u003eaneurysm\u003c/p\u003e \u003c/th\u003e \u003c/tr\u003e \u003c/thead\u003e \u003ctbody\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cb\u003eleft heart opacification grade rest cTTE\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e0.17\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e \u003cp\u003e0.49\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e0.10\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c5\"\u003e \u003cp\u003e0.45\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cb\u003eleft heart opacification grade Valsalva cTTE\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e0.37\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e \u003cp\u003e0.69\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e0.47\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c5\"\u003e \u003cp\u003e0.73\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cb\u003eleft atrium opacification grade rest cTOE\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e0.55\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e \u003cp\u003e0.16\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e0.64\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c5\"\u003e \u003cp\u003e0.46\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e\u003cb\u003eleft atrium opacification grade Valsalva cTOE\u003c/b\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c2\"\u003e \u003cp\u003e0.81\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c3\"\u003e \u003cp\u003e0.43\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e0.29\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c5\"\u003e \u003cp\u003e0.77\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003c/tbody\u003e \u003c/colgroup\u003e \u003ctfoot\u003e \u003ctr\u003e\u003ctd colspan=\"5\"\u003eChi-squared and Kruskal-Wallis tests\u003c/td\u003e\u003c/tr\u003e \u003c/tfoot\u003e \u003c/table\u003e\u003c/div\u003e \u003c/p\u003e \u003cp\u003eFinally, the left heart opacification grade between cTTE and cTOE was compared. In rest studies, cTTE produced the same degree of opacification as cTOE in 55.5% of cases and a higher degree in 40%, demonstrating similar or superior performance in 95.5% of cases. A moderate and statistically significant correlation and difference were observed between cTTE and cTOE opacification grade (Kendall\u0026rsquo;s τ\u0026thinsp;=\u0026thinsp;0.53, p\u0026thinsp;\u0026lt;\u0026thinsp;0.001 and χ\u0026sup2; = 26.83, p\u0026thinsp;=\u0026thinsp;0.002).\u003c/p\u003e \u003cp\u003eIn Valsalva studies, the left heart cTTE opacification grade matched cTOE in 29% and exceeded it in 45.5% of cases, showing comparable or higher opacification degree in 74.5% of cases. However, the correlation and difference between cTTE and cTOE opacification grade were weak and not statistically significant (Kendall\u0026rsquo;s τ\u0026thinsp;=\u0026thinsp;0.14, p\u0026thinsp;=\u0026thinsp;0.23 and χ\u0026sup2; = 13.23, p\u0026thinsp;=\u0026thinsp;0.15).\u003c/p\u003e"},{"header":"Discussion","content":"\u003cp\u003eThis study of high-density contrast echocardiography in the investigation of the presence of PFO in CS patients demonstrated: (i) high sensitivity of Valsalva cTOE and cTTE for PFO detection, (ii) no clinically relevant relationship between the left heart bubble opacification grade and invasively measured left-to-right heart shunt severity, (iii) no significant relationship between the left heart opacification grade and anatomical features of the interatrial septum/PFO, (iv) equal or higher left heart bubble opacification degree in cTTE vs cTOE, especially in rest studies.\u003c/p\u003e \u003cp\u003eIn two studies with 91 patients included in the cTOE meta-analysis from Mojadidi, reflecting the usual practice (i.e. contrast injection into the peripheral line and Valsalva manoeuvre performance), the sensitivity of cTOE vs autopsy, cardiac surgery, and/or catheterization (ability to pass the catheter through PFO) as the reference was 89\u0026ndash;91%. (\u003cspan citationid=\"CR28\" class=\"CitationRef\"\u003e28\u003c/span\u003e) In addition, cTOE studies yielded a significantly lower prevalence of PFO of 13% (95% CI: 8\u0026ndash;18%) as compared to autopsy studies 25% (95% CI: 20\u0026ndash;29%), p\u0026thinsp;=\u0026thinsp;0.004. (\u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e12\u003c/span\u003e) The imperfect accuracy of cTOE is usually attributed to patient intolerance of the TOE probe or inadequate Valsalva manoeuvre during sedation, which might result in up to 50% of PFOs being missed. (\u003cspan citationid=\"CR29\" class=\"CitationRef\"\u003e29\u003c/span\u003e, \u003cspan citationid=\"CR30\" class=\"CitationRef\"\u003e30\u003c/span\u003e) Sedation use during cTOE is, however, often not reported in the studies. (\u003cspan additionalcitationids=\"CR32\" citationid=\"CR31\" class=\"CitationRef\"\u003e31\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR33\" class=\"CitationRef\"\u003e33\u003c/span\u003e) The sensitivity of PFO detection also depends on the number of injections in Valsalva cTOE studies. (\u003cspan citationid=\"CR16\" class=\"CitationRef\"\u003e16\u003c/span\u003e) Despite all of these facts, cTOE is considered the gold standard for PFO detection. (\u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e12\u003c/span\u003e) In our study, the patients were trained for and rigorous operator\u0026acute;s adherence to the correct performance of the Valsalva manoeuvre was required. Visualisation of the contrast passing through the PFO canal was mandatory for its diagnosis and high-density contrast was used. Sedation was needed in \u0026lt;\u0026thinsp;20% of patients. All these facts likely contributed to the 100% sensitivity of cTOE against successful PFO occlusion in our study, highlighting the importance of an experienced imaging specialist performing cTOE for PFO detection. (\u003cspan citationid=\"CR34\" class=\"CitationRef\"\u003e34\u003c/span\u003e)\u003c/p\u003e \u003cp\u003eThere is heterogeneity in the type of contrast and Valsalva technique use in the published studies comparing cTTE to cTOE, with saline with or without blood prevailing in 80% and the contrast injected during the provocative manoeuvre in 66%. (\u003cspan citationid=\"CR10\" class=\"CitationRef\"\u003e10\u003c/span\u003e, \u003cspan citationid=\"CR35\" class=\"CitationRef\"\u003e35\u003c/span\u003e) The published sensitivity of saline and colloid cTTE ranges from 23% to 92% and 30% to 61%, respectively. (\u003cspan citationid=\"CR36\" class=\"CitationRef\"\u003e36\u003c/span\u003e) For the second harmonic cTTE, the sensitivity for PFO detection is reported to be higher at 91%, and similar to 96% for cTCD. (\u003cspan citationid=\"CR35\" class=\"CitationRef\"\u003e35\u003c/span\u003e, \u003cspan citationid=\"CR37\" class=\"CitationRef\"\u003e37\u003c/span\u003e) We used high-density contrast agents, mainly Gelofusine, showing excellent cTTE sensitivity\u0026thinsp;\u0026gt;\u0026thinsp;95%. One of the reasons, according to our experience, is better Gelofusine bubble stability and opacification of the right and left heart during cTTE/cTOE in comparison with normal saline contrast. (Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003e, \u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003e) This finding is supported by previously published studies. (\u003cspan citationid=\"CR9\" class=\"CitationRef\"\u003e9\u003c/span\u003e, \u003cspan citationid=\"CR13\" class=\"CitationRef\"\u003e13\u003c/span\u003e, \u003cspan citationid=\"CR14\" class=\"CitationRef\"\u003e14\u003c/span\u003e) Indeed, adding blood to normal saline can stabilise microbubbles and increase their suspension time in the blood. (\u003cspan citationid=\"CR38\" class=\"CitationRef\"\u003e38\u003c/span\u003e) In our experience, the density of the right heart opacification using Gelofusine and normal saline with blood does not differ. The use of modified gelatine is safe, a potential risk of anaphylaxis associated with its use is low, although the intensive care data are conflicting. We found only one published case report on anaphylaxis during contrast echocardiography. (\u003cspan additionalcitationids=\"CR40\" citationid=\"CR39\" class=\"CitationRef\"\u003e39\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR41\" class=\"CitationRef\"\u003e41\u003c/span\u003e) A relationship was found between allergic reactions to red meat and sensitisation to gelatine and galactose-α-1,3-galactose (α-Gal). (\u003cspan citationid=\"CR42\" class=\"CitationRef\"\u003e42\u003c/span\u003e) Gelofusine handling is also more practical and hygienic than mixing normal saline with blood, at least from the infection control point of view. For these reasons, the gelatine-based contrast has been the first line choice in our echo lab for almost two decades.\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003cp\u003eIn a recently published study by Takaya, normal saline cTTE was better for predicting large left to right shunts than cTOE (79% vs 28% of patients). (\u003cspan citationid=\"CR21\" class=\"CitationRef\"\u003e21\u003c/span\u003e) One of the explanations may be linked to our results demonstrating better opacification of the left heart in cTTE than in cTOE, although using a different contrast agent. (Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003e) Also, in cTTE, the whole left heart is visualised in apical four chamber view, in contrast to the cTOE study, which is focused on the presence of the contrast agent in a 2D often foreshortened image of the left atrium.\u003c/p\u003e \u003cp\u003eThe risk of PFO associated CS is usually assessed by the left heart bubble size and the presence of anatomical features such as PFO width, atrial septal aneurysm (ASA) or septum hypermobility and large Eustachian valve. It sounds intuitive that a large PFO means more right-to-left shunting, a mobile ASA may open the PFO with every heartbeat, and the Eustachian valve may direct thrombi from the inferior vena cava to the PFO. Indeed, in a metanalysis from Overell, the presence of ASA was associated with a higher risk of ischaemic stroke in comparison to healthy subjects (OR 2.35). Individual observational studies demonstrated even higher risk (OR 3.65). (\u003cspan citationid=\"CR43\" class=\"CitationRef\"\u003e43\u003c/span\u003e, \u003cspan citationid=\"CR44\" class=\"CitationRef\"\u003e44\u003c/span\u003e) We found no link between the severity of the bubble opacification grade and high risk PFO features. This might be attributed to the high prevalence of ASA (\u0026gt;\u0026thinsp;50%) in our group, reflecting the high-risk population studied, in which further risk stratification might be difficult.\u003c/p\u003e \u003cp\u003eScoring systems for high-risk PFO have been developed and include PFO size (\u0026ge;\u0026thinsp;2 mm), tunnel length (\u0026ge;\u0026thinsp;10 mm), the presence of ASA/hypermobile interatrial septum, prominent Eustachian/Chiari\u0026rsquo;s valve, straddling interatrial thrombus, concomitant pulmonary embolism or deep vein thrombosis, and \u003cb\u003ea\u003c/b\u003e large shunt at rest and during the Valsalva manoeuvre (\u0026gt;\u0026thinsp;20 microbubbles in the left atrium). (\u003cspan citationid=\"CR42\" class=\"CitationRef\"\u003e42\u003c/span\u003e, \u003cspan citationid=\"CR43\" class=\"CitationRef\"\u003e43\u003c/span\u003e) Our study is the first to compare the invasively measured shunt severity and the left heart contrast opacification degree, showing no relevant relationship apart from \u003cb\u003ea\u003c/b\u003e moderate association in rest\u0026thinsp;+\u0026thinsp;Valsalva cTOE studies with larger shunts\u0026thinsp;\u0026gt;\u0026thinsp;10%. There were few patients with no invasively measurable shunt but with a right-to-left shunt on cTTE/cTOE in our study. The explanation might include relative hypovolaemia and difficulty in performing the Valsalva manoeuvre in some patients undergoing invasive measurements via the femoral access. Also, counting the number of bubbles in the left atrium within three cycles after it opacifies the right heart by number of bubbles as recommended is impractical for high-density contrast due to the high degree and often homogeneous left heart opacification that is often observed. This approach has its limitations in general, as bubbles do not necessarily travel linearly across the left atrium and the same bubbles may be present across multiple cardiac cycles, leading to multiple counting. On the other hand, the left atrium size on the tomographic TOE slice is often significantly foreshortened, so some bubbles are inevitably missed. We chose a pragmatic approach, i.e. use of one still left heart frame with the maximum contrast opacification for its grading and a semiquantitative opacification scale, which we propose for future studies with high-density contrast agents. Indeed, in the recent paper by Su from the USA, the quantified recommended ratings of PFO shunt severity were present in \u0026lt;\u0026thinsp;1 in every 6 TEE reports, reflecting real clinical practice. (\u003cspan citationid=\"CR46\" class=\"CitationRef\"\u003e46\u003c/span\u003e)\u003c/p\u003e"},{"header":"Limitations","content":"\u003cp\u003eOur study is of small sample size, and its results would need to be confirmed by larger studies. Also, its results are valid for younger population only, as all patients were aged\u0026thinsp;\u0026lt;\u0026thinsp;55 years. We used a larger amount of contrast (20 mL) instead of the usual 10 mL per administration, which does not require a flush, thus simplifying the procedure, but leading to possibly higher opacification of the heart, which might have affected the results. We did not record the arm side of the venous access for the contrast administration and the inferior vena cava contrast injection site was reported in 2 (3.7%) patients, but this is unlikely to have affected the results. We also used different than usual approach to assess the left heart opacification grade as discussed. Anyway, the heart is a 3D structure, and any method using a single plane underestimates the true left heart bubble volume. (\u003cspan citationid=\"CR34\" class=\"CitationRef\"\u003e34\u003c/span\u003e) Counting the number of bubbles in a 3D image is however not currently feasible so far, but there are reports demonstrating better accuracy of 3D cTOE for PFO vs intrapulmonary shunt discrimination. (\u003cspan citationid=\"CR47\" class=\"CitationRef\"\u003e47\u003c/span\u003e) Despite every effort, there might have been some discrepancies in the Valsalva performance among the centres, especially in patients with cTOE performed under sedation, although this applied to only\u0026thinsp;\u0026lt;\u0026thinsp;20% of the study population and midazolam doses were low. The same applies to the invasive shunt measurement, although appropriate Valsalva manoeuvre performance was monitored invasively by the pressure curve changes.\u003c/p\u003e"},{"header":"Conclusion","content":"\u003cp\u003eOur study demonstrated that in bubble studies using high-density contrast, cTOE and cTTE with Valsalva, but not rest studies, are highly sensitive for PFO assessment. There is no clinically relevant relationship between the left-heart bubble opacification grade and invasively measured right-to-left shunt severity or atrial septum/PFO morphology. The left-heart opacification degree is equal to or higher in cTTE than in cTOE, especially in rest studies.\u003c/p\u003e "},{"header":"Declarations","content":"\u003cp\u003e\u003cstrong\u003eAuthors\u0026acute;contribution\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eDr Medilek was responsible for performing the echocardiography studies, interpreting and analysing the data, and compiling the manuscript. Dr Ondrus, Dr Mraz, Dr Praus and Dr Ballon performed echocardiography studies at their respective centres. Dr. Stasek, Dr Bis, Dr Dusek, Dr Poloczek and Dr Mates performed the invasive measurements. Dr Stasek was also responsible for the design of the MEASURE - PFO trial. All authors contributed to the editing of the manuscript, and approved its final version and content.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eFunding\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe trial was supported by a research grant from the Ministry of Health, Czech Republic, No. AZV NU20-02-00310 and by and by\u0026nbsp;the\u0026nbsp;Cooperatio\u0026nbsp;Program,\u0026nbsp;Charles University, Czech Republic, research\u0026nbsp;area\u0026nbsp;CARD.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eAcknowledgement\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eWe thank David Belcak for English language proofreading.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eData availability\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe datasets used and analysed during the current study are available from the corresponding author on reasonable request.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eEthics approval\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe study has been carried out in accordance with the Code of Ethics of the World Medical Association (Declaration of Helsinki) and approved by the Ethics Committee, Faculty of Medicine Hradec Kralove, Czech Republic (ref. No 201906S19P). The authors are responsible for the design and conduct of this study, all study analyses, drafting and editing of the manuscript, and its final contents.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eConsent for publication\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eNot applicable.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eCompeting interests\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe authors declare no competing interests.\u003c/p\u003e"},{"header":"References","content":"\u003col\u003e\u003cli\u003e\u003cspan\u003eHagen PT, Scholz DG, Edwards WD. Incidence and Size of Patent Foramen Ovale During the First 10 Decades of Life: An Autopsy Study of 965 Normal Hearts. Mayo Clin Proc. 1984;59:17\u0026ndash;20.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eHomma S, Sacco RL. Patent Foramen Ovale and Stroke. Circulation. 2005;112:1063\u0026ndash;172.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eSaver JL, Carroll JD, Thaler DE, Smalling RW, MacDonald LA, Marks DS, et al. Long-Term Outcomes of Patent Foramen Ovale Closure or Medical Therapy after Stroke. N Engl J Med. 2017;377:1022\u0026ndash;32.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eGramiak R, Shah PM. Echocardiography of the Aortic Root. Invest Radiol. 1968;3:356\u0026ndash;66.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eGramiak R, Shah PM, Kramer DH. Ultrasound Cardiography: Contrast Studies in Anatomy and Function. Radiology. 1969;92:939\u0026ndash;48.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eKremkau FW, Gramiak R, Carstensen EL, Shah PM, Kramer DH. ULTRASONIC DETECTION OF CAVITATION AT CATHETER TIPS. Am J Roentgenol. 1970;110:177\u0026ndash;83.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eMeltzer RS, Tickner EG, Sahines TP, Popp RL. The source of ultrasound contrast effect. J Clin Ultrasound. 1980;8:121\u0026ndash;7.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eNanda NC. History of echocardiographic contrast agents. Clin Cardiol. 1997;20(Suppl 1):7\u0026ndash;11.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eSmith MD, Kwan OL, Reiser HJ, DeMaria AN. Superior intensity and reproducibility of SHU-454, a new right heart contrast agent. J Am Coll Cardiol. 1984;3:992\u0026ndash;8.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eCaso V, Turc G, Abdul-Rahim AH, Castro P, Hussain S, Lal A, et al. European Stroke Organisation (ESO) Guidelines on the diagnosis and management of patent foramen ovale (PFO) after stroke. Eur Stroke J. 2024;9:800\u0026ndash;34.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eKhan R, Karim MN, Hosseini F, Fine N. Diagnostic Accuracy of Transthoracic Echocardiography With Contrast for Detection of Right-to-Left Shunt: A Systematic Review and Meta-analysis. Can J Cardiol. 2022;38:1948\u0026ndash;58.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003ePristipino C, Sievert H, D\u0026rsquo;Ascenzo F, Mas JL, Meier B, Scacciatella P, et al. European position paper on the management of patients with patent foramen ovale. General approach and left circulation thromboembolism. EuroIntervention. 2019;14:1389\u0026ndash;402.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eCukon Buttignoni S, Khorsand A, Mundigler G, Bergler-Klein J, Heger M, Zehetgruber M, et al. Agitated saline versus polygelatine for the echocardiographic assessment of patent foramen ovale. J Am Soc Echocardiogr. 2004;17:1059\u0026ndash;65.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003ePuledda F, Toscano M, Pieroni A, Veneroso G, Di Piero V, Vicenzini E. Right-to-left shunt detection sensitivity with air\u0026ndash;saline and air\u0026ndash;succinil gelatin transcranial Doppler. Int J Stroke. 2016;11:229\u0026ndash;38.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eLi H, Huang Y, Luo D, Wen S, Fei H, Zhang C. Optimizing agitated saline volume for contrast echocardiography: Balancing diagnostic performance and operator fatigue. Med Adv. 2024;2:254\u0026ndash;61.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eJohansson MC, Helgason H, Dellborg M, Eriksson P. Sensitivity for Detection of Patent Foramen Ovale Increased with Increasing Number of Contrast Injections: A Descriptive Study with Contrast Transesophageal Echocardiography. J Am Soc Echocardiogr. 2008;2:419\u0026ndash;24.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eTurc G, Calvet D, Gu\u0026eacute;rin P, Sroussi M, Chatellier G, Mas J, et al. Closure, Anticoagulation, or Antiplatelet Therapy for Cryptogenic Stroke With Patent Foramen Ovale: Systematic Review of Randomized Trials, Sequential Meta-Analysis, and New Insights From the CLOSE Study. J Am Heart Assoc. 2018;7:e008356.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eMas JL, Derumeaux G, Guillon B, Massardier E, Hosseini H, Mechtouff L, et al. Patent Foramen Ovale Closure or Anticoagulation vs. Antiplatelets after Stroke. N Engl J Med. 2017;377:1011\u0026ndash;21.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eLee PH, Song JK, Kim JS, Heo R, Lee S, Kim DH, et al. Cryptogenic Stroke and High-Risk Patent Foramen Ovale. J Am Coll Cardiol. 2018;71:2335\u0026ndash;42.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eMas JL, Derumeaux G, Guillon B, Massardier E, Hosseini H, Mechtouff L, et al. Patent Foramen Ovale Closure or Anticoagulation vs. Antiplatelets after Stroke. N Engl J Med. 2017;377:1011\u0026ndash;21.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eTakaya Y, Nakayama R, Akagi T, Yokohama F, Miki T, Nakagawa K, et al. Importance of saline contrast transthoracic echocardiography for evaluating large right-to-left shunt in patent foramen ovale associated with cryptogenic stroke. Int J Cardiovasc Imaging. 2022;38:515\u0026ndash;20.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eMorelli F, Guadagno V, Mencarini T, Brambilla M, Fusini L, Bozzi S, et al. Morphotype-based risk stratification in patients with patent foramen ovale using computational fluid dynamics. Sci Rep. 2025;15:34557.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eStasek J, Bis J, Dusek J, Medilek K, Dostal J, Branny M, et al. Evaluation of the severity of right-to-left shunt in PFO patients after systemic embolism (MEASURE-PFO study): Study design. Pol Heart J. 2024;82:774\u0026ndash;6.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eMitchell C, Rahko PS, Blauwet LA, Canaday B, Finstuen JA, Foster MC, et al. Guidelines for Performing a Comprehensive Transthoracic Echocardiographic Examination in Adults: Recommendations from the American Society of Echocardiography. J Am Soc Echocardiogr. 2019;32:1\u0026ndash;64.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eRobinson S, Rana B, Oxborough D, Steeds R, Monaghan M, Stout M, et al. A practical guideline for performing a comprehensive transthoracic echocardiogram in adults: the British Society of Echocardiography minimum dataset. Echo Res Pract. 2020;7:G59\u0026ndash;93.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eHahn RT, Abraham T, Adams MS, Bruce CJ, Glas KE, Lang RM, et al. Guidelines for Performing a Comprehensive Transesophageal Echocardiographic Examination: Recommendations from the American Society of Echocardiography and the Society of Cardiovascular Anesthesiologists. J Am Soc Echocardiogr. 2013;26:921\u0026ndash;694.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eWheeler R, Steeds R, Rana B, Wharton G, Smith N, Allen J, et al. A minimum dataset for a standard transoesophageal echocardiogram: a guideline protocol from the British Society of Echocardiography. Echo Res Pract. 2015;2:G29\u0026ndash;45.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eMojadidi MK, Bogush N, Caceres JD, Msaouel P, Tobis JM. Diagnostic Accuracy of Transesophageal Echocardiogram for the Detection of Patent Foramen Ovale: A Meta-Analysis. Echocardiography. 2014;31:752\u0026ndash;8.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eGhazal SN. Valsalva maneuver in echocardiography. J Echocardiogr. 2017;15:1\u0026ndash;5.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eRodrigues AC, Picard MH, Carbone A, Arruda AL, Flores T, Klohn J, et al. Importance of Adequately Performed Valsalva Maneuver to Detect Patent Foramen Ovale during Transesophageal Echocardiography. J Am Soc Echocardiogr. 2013;26:1337\u0026ndash;43.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eBelkin RN, Pollack BD, Ruggiero ML, Alas LL, Tatini U. Comparison of transesophageal and transthoracic echocardiography with contrast and color flow Doppler in the detection of patent foramen ovale. Am Heart J. 1994;128:520\u0026ndash;5.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eMaffe S, Dellavesa P, Zenone F, Paino AM, Paffoni P, Perucca A, et al. Transthoracic second harmonic two- and three-dimensional echocardiography for detection of patent foramen ovale. Eur J Echocardiogr. 2010;11:57\u0026ndash;63.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eYang X, Wang H, Wei Y, Zhai N, Liu B, Li X. Diagnosis of Patent Foramen Ovale: The Combination of Contrast Transcranial Doppler, Contrast Transthoracic Echocardiography, and Contrast Transesophageal Echocardiography. BioMed Res Int. 2020. \u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003e10.1155/2020/8701759\u003c/span\u003e\u003cspan address=\"10.1155/2020/8701759\" targettype=\"DOI\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eSong JK. Pearls and Pitfalls in the Transesophageal Echocardiographic Diagnosis of Patent Foramen Ovale. J Am Soc Echocardiogr. 2023;36:895\u0026ndash;e9053.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eMojadidi MK, Winoker JS, Roberts SC, Msaouel P, Gevorgyan R, Zolty R. Two-dimensional echocardiography using second harmonic imaging for the diagnosis of intracardiac right-to-left shunt: a meta-analysis of prospective studies. Int J Cardiovasc Imaging. 2014;30:911\u0026ndash;23.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eMojadidi MK, Winoker JS, Roberts SC, Msaouel P, Zaman MO, Gevorgyan R, et al. Accuracy of Conventional Transthoracic Echocardiography for the Diagnosis of Intracardiac Right-to‐Left Shunt: A Meta‐Analysis of Prospective Studies. Echocardiography. 2014;31:1036\u0026ndash;48.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eKatsanos AH, Psaltopoulou T, Sergentanis TN, Frogoudaki A, Vrettou A, Ikonomidis I, et al. Transcranial Doppler versus transthoracic echocardiography for the detection of patent foramen ovale in patients with cryptogenic cerebral ischemia: A systematic review and diagnostic test accuracy meta-analysis. Ann Neurol. 2016;79:625\u0026ndash;35.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eGentile M, De Vito A, Azzini C, Tamborino C, Casetta I. Adding Blood to Agitated Saline Significantly Improves Detection of Right-to-Left Shunt by Contrast-Transcranial Color-Coded Duplex Sonography. Ultrasound Med Biol. 2014;40:2637\u0026ndash;41.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eMoeller C, Fleischmann C, Thomas-Rueddel D, Vlasakov V, Rochwerg B, Theurer P, et al. How safe is gelatin? A systematic review and meta-analysis of gelatin-containing plasma expanders vs crystalloids and albumin. J Crit Care. 2016;35:75\u0026ndash;83.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eSaw MM, Chandler B, Ho KM. Benefits and Risks of Using Gelatin Solution as a Plasma Expander for Perioperative and Critically Ill Patients: A Meta-Analysis. Anaesth Intensive Care. 2012;40:17\u0026ndash;32.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eDubrey S, Dahdal G, Grocottmason R. Severe anaphylaxis to Gelofusine during a transthoracic echo bubble study. Eur J Echocardiogr. 2006. \u003cspan class=\"ExternalRef\"\u003e\u003cspan class=\"RefSource\"\u003e10.1016/j.euje.2006.11.003\u003c/span\u003e\u003cspan address=\"10.1016/j.euje.2006.11.003\" targettype=\"DOI\" class=\"RefTarget\"\u003e\u003c/span\u003e\u003c/span\u003e.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eMullins RJ, James H, Platts-Mills TAE, Commins S. Relationship between red meat allergy and sensitization to gelatin and galactose-α-1,3-galactose. J Allergy Clin Immunol. 2012;129:1334\u0026ndash;e13421.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eAgmon Y, Khandheria BK, Meissner I, Gentile F, Whisnant JP, Sicks JD, et al. Frequency of Atrial Septal Aneurysms in Patients With Cerebral Ischemic Events. Circulation. 1999;99:1942\u0026ndash;4.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eOverell JR, Bone I, Lees KR. Interatrial septal abnormalities and stroke: A meta-analysis of case-control studies. Neurology. 2000;55:1172\u0026ndash;9.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eNakayama R, Takaya Y, Akagi T, Watanabe N, Ikeda M, Nakagawa K, et al. Identification of High-Risk Patent Foramen Ovale Associated With Cryptogenic Stroke: Development of a Scoring System. J Am Soc Echocardiogr. 2019;32:811\u0026ndash;6.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eSun PY, Tobis JM, Daneshvar SA, Alfonso RC, Liebeskind DS, Saver JL. Quantification of patent foramen ovale shunt severity by transesophageal echocardiogram and transcranial doppler in routine clinical practice. J Stroke Cerebrovasc Dis. 2025;34:108306.\u003c/span\u003e\u003c/li\u003e \u003cli\u003e\u003cspan\u003eGwak SY, Kim K, Lee HJ, Cho I, Hong GR, Ha JW, et al. Three-dimensional agitated saline contrast transesophageal echocardiography for the diagnosis of patent foramen ovale. Sci Rep. 2025;15:29136.\u003c/span\u003e\u003c/li\u003e\u003c/ol\u003e"}],"fulltextSource":"","fullText":"","funders":[],"hasAdminPriorityOnWorkflow":false,"hasManuscriptDocX":true,"hasOptedInToPreprint":true,"hasPassedJournalQc":"","hasAnyPriority":false,"hideJournal":false,"highlight":"","institution":"","isAcceptedByJournal":true,"isAuthorSuppliedPdf":false,"isDeskRejected":"","isHiddenFromSearch":false,"isInQc":false,"isInWorkflow":false,"isPdf":false,"isPdfUpToDate":true,"isWithdrawnOrRetracted":false,"journal":{"display":true,"email":"[email protected]","identity":"echo-research-and-practice","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":false,"externalIdentity":"","sideBox":"Learn more about [Echo Research \u0026 Practice](https://echo.biomedcentral.com/)","snPcode":"44156","submissionUrl":"https://submission.nature.com/new-submission/44156/3","title":"Echo Research \u0026 Practice","twitterHandle":"@BioMedCentral","acdcEnabled":true,"dfaEnabled":true,"editorialSystem":"stoa","reportingPortfolio":"BMC/SO AJ","inReviewEnabled":true,"inReviewRevisionsEnabled":true},"keywords":"high density contrast, transthoracic echocardiography, shunt severity, atrial septum morphology","lastPublishedDoi":"10.21203/rs.3.rs-8302087/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-8302087/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"\u003cp\u003e \u003cb\u003eBackground\u003c/b\u003e \u003c/p\u003e \u003cp\u003eTransthoracic contrast echocardiography (cTTE) is a first-line method for patent foramen ovale assessment (PFO), but its sensitivity is reported as low and inconsistent. We aimed to investigate the sensitivity of high-density bubble study cTTE for PFO detection, and the relationship between left-heart contrast opacification grade, invasively measured shunt severity and atrial septum/PFO morphology.\u003c/p\u003e \u003cp\u003e \u003cb\u003eMethods\u003c/b\u003e \u003c/p\u003e \u003cp\u003eFifty-five consecutive patients with proven PFO on contrast transoesophageal echocardiography (cTOE) underwent contrast transthoracic echocardiography (cTTE) and invasive right-to-left shunt severity measurement. Echocardiographic shunt severity was classified according to the degree of left atrial (cTOE) or left atrial/ventricular (cTTE) opacification as grade 0: no bubbles; grade 1: mild; grade 2: moderate; grade 3: significant opacification. The thermodilution method was used for the right-to-left heart shunt measurement.\u003c/p\u003e \u003cp\u003e \u003cb\u003eResults\u003c/b\u003e \u003c/p\u003e \u003cp\u003eFor PFO detection, the sensitivity of rest\u0026thinsp;+\u0026thinsp;Valsalva cTOE was 100%, but rest cTOE vs rest\u0026thinsp;+\u0026thinsp;Valsalva cTOE was 77.3%. For transthoracic studies, the sensitivity of rest\u0026thinsp;+\u0026thinsp;Valsalva cTTE was 98.2%, Valsalva cTTE 97.6%, and rest cTTE 83.6% vs rest\u0026thinsp;+\u0026thinsp;Valsalva cTOE. No relationship was found between rest or Valsalva cTTE and cTOE left-heart bubble opacification grade and invasively measured right-to-left shunt\u0026thinsp;\u0026lt;\u0026thinsp;10%, 10\u0026ndash;19%, or \u0026ge;\u0026thinsp;20% (p\u0026thinsp;=\u0026thinsp;0.10\u0026ndash;0.26). A moderate association only was revealed between rest\u0026thinsp;+\u0026thinsp;Valsalva cTOE opacification grade and \u0026lt;\u0026thinsp;10% and \u0026ge;\u0026thinsp;10% right-to-left shunt (p\u0026thinsp;=\u0026thinsp;0.024, Cram\u0026eacute;r's coefficient\u0026thinsp;=\u0026thinsp;0.368). No relationship was also found between left-heart opacification grade and the presence of an atrial septal aneurysm, Eustachian valve/Chiari's network, or PFO channel width and length (p\u0026thinsp;=\u0026thinsp;0.45\u0026ndash;0.77). Rest cTTE demonstrated equal contrast opacification grade in 56% and higher in 40% vs rest cTOE. Valsalva cTTE showed equal contrast opacification grade in 29% and higher in 46% of studies vs Valsalva cTOE.\u003c/p\u003e \u003cp\u003e \u003cb\u003eConclusions\u003c/b\u003e \u003c/p\u003e \u003cp\u003eIn bubble studies using high-density contrast, cTOE and cTTE with Valsalva, but not rest studies, are highly sensitive for PFO assessment. There is no clinically relevant relationship between the left-heart bubble opacification grade and invasively measured right-to-left shunt severity or atrial septum morphology. The left-heart opacification degree is equal to or higher in cTTE than in cTOE, especially in rest studies.\u003c/p\u003e","manuscriptTitle":"High-density bubble echocardiography for patent foramen ovale assessment: sensitivity and relationship with invasively measured shunt severity and atrial septum morphology","msid":"","msnumber":"","nonDraftVersions":[{"code":1,"date":"2025-12-22 08:43:22","doi":"10.21203/rs.3.rs-8302087/v1","editorialEvents":[{"type":"communityComments","content":0},{"type":"decision","content":"Revision requested","date":"2026-02-06T11:02:48+00:00","index":"","fulltext":""},{"type":"editorInvitedReview","content":"","date":"2026-02-06T09:00:02+00:00","index":"hide","fulltext":""},{"type":"reviewerAgreed","content":"305438704741628998674178816407744208938","date":"2026-01-28T10:09:00+00:00","index":"hide","fulltext":""},{"type":"reviewerAgreed","content":"336640423457278884347133896628038142081","date":"2025-12-15T23:11:36+00:00","index":"hide","fulltext":""},{"type":"reviewersInvited","content":"","date":"2025-12-11T07:27:54+00:00","index":"","fulltext":""},{"type":"editorAssigned","content":"","date":"2025-12-11T07:20:48+00:00","index":"","fulltext":""},{"type":"checksComplete","content":"","date":"2025-12-09T10:15:32+00:00","index":"","fulltext":""},{"type":"submitted","content":"Echo Research \u0026 Practice","date":"2025-12-07T22:03:48+00:00","index":"","fulltext":""}],"status":"published","journal":{"display":true,"email":"[email protected]","identity":"echo-research-and-practice","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":false,"externalIdentity":"","sideBox":"Learn more about [Echo Research \u0026 Practice](https://echo.biomedcentral.com/)","snPcode":"44156","submissionUrl":"https://submission.nature.com/new-submission/44156/3","title":"Echo Research \u0026 Practice","twitterHandle":"@BioMedCentral","acdcEnabled":true,"dfaEnabled":true,"editorialSystem":"stoa","reportingPortfolio":"BMC/SO AJ","inReviewEnabled":true,"inReviewRevisionsEnabled":true}}],"origin":"","ownerIdentity":"5e4b8f86-afbf-4869-8b39-e7ea08928a27","owner":[],"postedDate":"December 22nd, 2025","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"under-review","subjectAreas":[],"tags":[],"updatedAt":"2026-03-10T07:11:39+00:00","versionOfRecord":[],"versionCreatedAt":"2025-12-22 08:43:22","video":"","vorDoi":"","vorDoiUrl":"","workflowStages":[]},"version":"v1","identity":"rs-8302087","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-8302087","identity":"rs-8302087","version":["v1"]},"buildId":"8U1c8b4HqxoKbykW_rLl7","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}