Critical Event Checklists for Simulated In-Hospital Dysrhythmias in Children with Heart Disease.

Children with heart disease are at increased risk of unstable dysrhythmias and in-hospital cardiac arrest (IHCA). Clinician adherence to lifesaving processes of care is an important contributor to improving patient outcomes. This study evaluated whether critical event checklists improve adherence to lifesaving processes during simulated acute events secondary to unstable dysrhythmias. A randomized controlled trial was conducted in a cardiac ward in a tertiary care, academic children's hospital. Unannounced simulated emergencies involving dysrhythmias in pediatric patients with underlying cardiac disease were conducted weekly. Responders were pediatric and anesthesiology residents, respiratory therapists, and bedside registered nurses. Six teams were randomized into two groups-three received checklists (intervention) and three did not (control). Each team participated in four simulated scenarios over a 4-week pediatric cardiology rotation. Participants received a brief slideshow presentation, which included a checklist orientation, at the start of their rotation. Simulations were video and audio recorded and those with three or more participants were included for analysis. The primary outcome was team adherence to lifesaving processes, expressed as the percentage of completed critical management steps. Secondary outcomes included participant perceptions of the checklist usefulness in identifying and managing dysrhythmias. We used generalized estimating equations (GEE) models, which accounted for clustering within groups, to evaluate the effects of the intervention. A total of 24 simulations were conducted; one of the 24 simulations was excluded due to an insufficient number of participants. In our GEE analysis, 81.21% (78.96%, 83.47%) of critical steps were completed with checklists available versus 68.06% (59.38%, 76.74%) without checklists (p = 0.004). Ninety-three percent of study participants reported that they would use the checklists during an unstable dysrhythmia of a child with underlying cardiac disease. Checklists were associated with improved adherence to lifesaving processes during simulated resuscitations for unstable pediatric dysrhythmias. These findings support the use of scenario specific checklists for the management of unstable dysrhythmias in simulations involving pediatric patients with underlying cardiac disease. Future studies should investigate whether checklists are as effective in actual pediatric in-hospital emergencies.

Association Between High Sensitivity Troponin Levels Following Pediatric Orthotopic Heart Transplantation and Intensive Care Unit Resource Utilization.

The utility of troponin levels, including high sensitivity troponin T (hs-TnT), after orthotopic heart transplant (OHT) is controversial. Conflicting data exist regarding its use as a marker of acute rejection. Few studies have examined possible associations of hs-TnT levels immediately after OHT with metrics of intensive care unit (ICU) resource utilization or risk of acute rejection. We performed a retrospective cohort chart review including all OHT recipients < 20 years of age at our center between June 2019 and December 2022. Patients were divided into two groups based on supra- or sub-median initial hs-TnT levels (median 3462.5 ng/L). Primary outcome was days requiring ICU-level care, secondary outcomes included days intubated, days requiring positive pressure ventilation (PPV), days on inotropic medications, actual ICU length of stay, Vasoactive Inotrope Scores (VIS) on postoperative days (POD) 0 through 7, and acute rejection at 30 days and one year after OHT. Patients with higher hs-TnT required ICU level care for longer [13.5 (10-17.5) vs. 9.5 (8-12) days, p = 0.01] and spent more days intubated [6 (4-7) vs. 3 (3-5) days, p < 0.001], on PPV [9 (6-15) vs. 6 (5-8.5) days, p = 0.02], and on inotropes [11 (9-14) vs. 8 (7-11) days, p = 0.025]. VIS was only different between groups on POD7 [5 (3-7) vs. 3 (0-5), p = 0.04]. There was no difference in rejection between the groups. Higher hs-TnT immediately following pediatric OHT may predict higher ICU resource utilization, despite no difference in VIS, although it does not predict acute rejection in the first year after OHT.

Ventricular Assist Device Training and Emergency Management Among Pediatric Cardiac Intensive Care Physicians - Multicenter Cross-Sectional Survey.

Background/Aim: Pediatric cardiac intensive care physicians practicing at centers that implant ventricular assist devices (VAD's) are exposed to increasing numbers of VAD patients, with a significant number of VAD-days. We aimed to delineate pediatric cardiac critical care practices surrounding routine and emergency management of VADs. Methodology: We administered a multicenter cross-sectional survey of pediatric cardiac intensive care unit (CICU) physicians in the United States and Canada. Survey distribution occurred between August 31st and October 26th 2021. Results: A total of 254 CICU physicians received a formal invitation to participate, with 108 returning completed surveys (42.5% response rate). Responses came from CICU attending physicians at 26 separate institutions. Respondents' level of experience was well distributed across junior, mid-level, and senior staff: less than 5 years (38%), 5-9 years (25%), and >/= 10 years (37%). Most respondents had received formal training in the management of VAD patients (n = 93, 86.1%), with training format including fellowship (61%), simulation (36%), and national/international conferences (26.5%). Dedicated advanced cardiac therapies teams were available at the institutions of 97.2% of respondents. A total of 78/108 (72.2%) described themselves as "comfortable" or "very comfortable" in pediatric VAD management. While 63% (68/108) of respondents reported that they had never performed (or overseen the performance of) chest compressions in a pediatric patient with a VAD, 37% (40/108) reported performing CPR at least once in a VAD patient. Conclusion: With no existing international guidelines for emergency cardiovascular care in the pediatric VAD population, our survey identifies an important gap in resuscitation recommendations.

Staged vs Complete Repair in Tetralogy of Fallot With Pulmonary Atresia.

BACKGROUND: We sought to compare outcomes for infants with tetralogy of Fallot with pulmonary atresia (TOF/PA) and confluent pulmonary arteries who underwent staged or primary complete surgical repair. METHODS: This retrospective study included infants undergoing initial surgical intervention between 0 and 60 days of age with TOF/PA without aortopulmonary collaterals from 2009 to 2018 at 20 centers. The primary outcome was days alive and out of the hospital in the first year of life (DAOH365). Secondary outcomes were mortality at 1 year of age and a composite major complication outcome. Multivariable modeling with generalized estimating equations were used to compare outcomes between groups. RESULTS: Of 221 subjects, 142 underwent staged repair and 79 underwent primary complete repair. There was no significant difference in median DAOH365 between the staged and primary repair groups (317 days [interquartile range, 278-336] vs 338 days [interquartile range, 314-348], respectively; adjusted P = .13). Nine staged repair patients (7%) died in the first year of life vs 5 primary repair patients (6%; adjusted odds ratio, 1.00; 95% CI, 0.25-3.95). At least 1 major complication occurred in 37% of patients who underwent staged repair vs 41% of patients who underwent primary complete repair (P = .75), largely driven by the need for unplanned cardiac reinterventions. CONCLUSIONS: For infants with TOF/PA with confluent pulmonary arteries, a surgical strategy of staged or primary complete repair resulted in statistically similar DAOH365, early mortality, and morbidity.

Post-operative troponin levels and left ventricular function in patients with d-transposition of the great arteries following the arterial switch operation.

The aim of this study was to assess the significance of post-operative troponin levels as a surrogate for left ventricular (LV) dysfunction measured by global longitudinal strain (GLS) in patients with dextro-transposition of the great arteries (d-TGA) who undergo an arterial switch operation (ASO), and to explore the LV GLS recovery in the mid-term follow-up period. Seventy-eight neonates were included, of whom 41 had troponin-I measurements and 37 had troponin-T measurements. The primary outcome of LV GLS was assessed and compared with healthy controls at the pre-operative stage and time of discharge, 3 months, 6 months and 12 months of age. Secondary outcomes included deaths or transplantations and other clinical markers such as length of hospital stay. D-TGA patients had worse LV GLS post-operatively compared to age-matched controls (p < 0.01) which improved by 12 months of age (p = 0.53). No association was found between changes in troponin-I or troponin-T levels and LV GLS at the time of discharge (r = 0.4, p = 0.64 and r = -0.5, p = 0.91, respectively). In addition, there were no deaths or transplantations in this cohort over a period of 12 months. LV GLS appears to worsen in the early post-operative period for d-TGA patients who undergo neonatal ASO but this recovers through the first post-operative year. Troponin levels have limited value in predicting early or midterm LV dysfunction and recovery.

Elevated cardiac biomarkers and outcomes in children and adolescents with acute COVID-19.

Cardiac involvement associated with multi-system inflammatory syndrome in children has been extensively reported, but the prevalence of cardiac involvement in children with SARS-CoV-2 infection in the absence of inflammatory syndrome has not been well described. In this retrospective, single centre, cohort study, we describe the cardiac involvement found in this population and report on outcomes of patients with and without elevated cardiac biomarkers. Those with multi-system inflammatory syndrome in children, cardiomyopathy, or complex CHD were excluded. Inclusion criteriaz were met by 80 patients during the initial peak of the pandemic at our institution. High-sensitivity troponin T and/or N-terminal pro-brain type natriuretic peptide were measured in 27/80 (34%) patients and abnormalities were present in 5/27 (19%), all of whom had underlying comorbidities. Advanced respiratory support was required in all patients with elevated cardiac biomarkers. Electrocardiographic abnormalities were identified in 14/38 (37%) studies. Echocardiograms were performed on 7/80 patients, and none demonstrated left ventricular dysfunction. Larger studies to determine the true extent of cardiac involvement in children with COVID-19 would be useful to guide recommendations for standard workup and management.

Left Heart Decompression on Veno-Arterial Extracorporeal Membrane Oxygenation in Children With Dilated Cardiomyopathy and Myocarditis: An Extracorporeal Life Support Organization Registry Review.

OBJECTIVES: To describe the association between left heart decompression on veno-arterial extracorporeal membrane oxygenation and survival in patients with myocarditis and dilated cardiomyopathy. The secondary outcome is to study association of left heart decompression with survival in children with myocarditis compared with those with dilated cardiomyopathy. DESIGN: Retrospective study of a multicenter registry database. SETTING: Data reported to Extracorporeal Life Support Organization from international extracorporeal membrane oxygenation centers. PATIENTS: Patients less than or equal to 18 years old with a diagnosis of myocarditis or dilated cardiomyopathy receiving extracorporeal membrane oxygenation. INTERVENTIONS: None. MEASUREMENTS AND MAIN RESULTS: A total of 1,438 pediatric extracorporeal membrane oxygenation runs were identified. Thirty-seven percent of the patients had myocarditis (n = 532), whereas the rest had dilated cardiomyopathy. Survival to hospital discharge was 63%. Median extracorporeal membrane oxygenation duration was 148 hours with interquartile range (84-248 hr). Nineteen percent of patients (n = 274) had left heart decompression. Multivariable analysis revealed using left heart decompression (adjusted odds ratio, 1.42; 95% CI, 1.06-1.89; p = 0.02), e-cardiopulmonary resuscitation (adjusted odds ratio, 0.63; 95% CI, 0.51-0.79; p < 0.001), higher pH (adjusted odds ratio, 3.69; 95% CI, 1.80-7.53; p < 0.001), and diagnosis of myocarditis (adjusted odds ratio, 1.69; 95% CI, 1.35-2.08; p < 0.001) were associated with greater odds of survival. In the multivariable analysis for patients with dilated cardiomyopathy, left heart decompression failed to reveal a significant association with survival (20% among survivors vs 17% among nonsurvivors, 95% CI, -2.2% to 8.0%). Meanwhile in patients with myocarditis, the multivariable analysis failed to exclude the possibility that left heart decompression was associated with up to a three-fold greater odds of survival (adjusted odds ratio, 1.77; 95% CI, 0.99-.15). CONCLUSIONS: Retrospective review of the Extracorporeal Life Support Organization registry revealed an association between left heart decompression and greater odds of survival in children with myocarditis and dilated cardiomyopathy on extracorporeal membrane oxygenation. When comparing patients with dilated cardiomyopathy against those with myocarditis, we could not exclude a three-fold greater odds of survival associated with the use of left heart decompression. This finding warrants further prospective evaluation.

Simulation as an Educational Tool in the Pediatric Cardiac Intensive Care Unit.

PURPOSE OF REVIEW: This review highlights the use of simulation as an educational tool in the highly specialized pediatric cardiac intensive care unit (PCICU). RECENT FINDINGS: Healthcare simulation is used in high acuity medical environments to test healthcare systems. Healthcare simulation can improve team training, patient safety, and improve medical decision-making. Complex physiologies in the PCICU demand effective teamwork to consistently deliver high-quality patient care. Simulation-based PCICU learning objectives depend on a structured cognitive load framework to account for individual learner abilities, team constructs, and healthcare resources. SUMMARY: PCICU simulation programs are strengthened by utilizing traditional education theory, with careful consideration of complex physiologies, interprofessional personnel, and center-specific resources. Virtual platforms should continue to evolve to provide additional, more convenient venues for individual learners and teams. Healthcare systems should frequently intersect with simulation educators to create relevant learning objectives that will contribute to patient safety, improve team performance, and patient outcomes.

Early Thromboprophylaxis Initiation is Associated With Reduced Fontan Thromboses in the Early Postoperative Period.

Fontan circuit thrombosis is a significant cause of early postoperative morbidity and mortality. Thrombosis incidence and relationship to thromboprophylaxis choice and timing of initiation are not well established. We sought to evaluate the incidence of Fontan circuit thrombosis in the first 30 postoperative days and its relationship to thromboprophylaxis choice and timing. Patients undergoing Fontan surgery, 2006-2016, were reviewed. Fontan circuit thrombosis was defined by sonographic detection of intracardiac or deep venous thrombi. Logistic regression was used to assess relationships between thromboprophylaxis characteristics and thrombosis. One hundred ninety-two patients underwent Fontan. Fontan thrombosis occurred in 19 (10%) patients. 54% were started on aspirin, 27% coumadin, 4% heparin, and 7% none. There was no relationship between thrombosis and baseline anatomy, Fontan type or fenestration. Median time to thromboprophylaxis initiation was 4 days (interquartile range 2-6). Patients not started on thromboprophylaxis had 44.8 times the odds of thrombosis as those on thromboprophylaxis (confidence interval 6.4-311.7, P < 0.01); no children starting thromboprophylaxis before postoperative day 2 developed thromboses. For every day that thromboprophylaxis was delayed, odds of thrombosis increased by 30% (odds ratio 1.3; CI 1.1-1.6, P < 0.01). There was no difference in the odds of thrombosis between children taking aspirin vs other thromboprophylaxis types. Odds of early postoperative Fontan circuit thrombosis are increased in patients in whom thromboprophylaxis is delayed beyond the second postoperative day, with no difference in the odds of thrombosis between patients initiated on aspirin vs other thromboprophylaxis. Early aspirin institution post Fontan is recommended to reduce morbidity. Ultra-mini-Abstract: Odds of early postoperative Fontan circuit thrombosis are increased in patients in whom thromboprophylaxis is delayed beyond the second postoperative day, with no difference in the odds of thrombosis between patients initiated on aspirin vs other thromboprophylaxis. Early aspirin institution post Fontan is recommended to reduce morbidity and resource utilization.

Cardiac workup and monitoring in hospitalised children with COVID- 19.

Approximately, 1.7 million individuals in the United States have been infected with SARS-CoV-2, the virus responsible for the novel coronavirus disease-2019 (COVID-19). This has disproportionately impacted adults, but many children have been infected and hospitalised as well. To date, there is not much information published addressing the cardiac workup and monitoring of children with COVID-19. Here, we share the approach to the cardiac workup and monitoring utilised at a large congenital heart centre in New York City, the epicentre of the COVID-19 pandemic in the United States.

A Case of Mitral Valve Endocarditis Complicated by Multiple Embolic Phenomena: Leaping from Adult Guidelines to Pediatric Critical Care Decisions.

Early surgical intervention for children with infective endocarditis (IE) and cerebrovascular sequelae has significant risks, resulting in practice variation amongst pediatric cardiologists, intensivists, and cardiothoracic surgeons. The limited pediatric consensus recommendations make decision making for practitioners challenging. The added risk of multiorgan dysfunction syndrome can make these decisions even more difficult. We present the case of a 14-year-old with IE and resultant multiorgan dysfunction syndrome including cerebrovascular complication, successfully treated by primary valve repair within the 1st week of diagnosis.

Evaluation of Bcr-Abl Gene Rearrangement among Bangladeshi Chronic Myeloid Leukaemia Patients.

Chronic Myeloid Leukaemia (CML) is a clonal myeloproloferative disorder. Presence of molecular translocation t (9; 22) in CML patients can be confirmed by Reverse Transcriptase Polymerase Chain Reaction (RT-PCR). Among haematological malignancies CML is the commonest leukaemia of adults in Asia. Despite this, there are very few studies published from Bangladesh, documenting the frequency of bcr-abl fusion transcripts. So, we would like to perform this observational study to evaluate bcr-abl fusion transcripts and demographic status among RT-PCR positive chronic phase CML patients at BIRDEM & other two centers of Dhaka city, Bangladesh where patients admitted from different districts of Bangladesh in the period of January 2010 to June 2012. Twenty patients from three centers willing to perform RT-PCR test and showed positive PCR result, were enrolled under written consent before starting treatment with targeted molecular therapy. The age range of the sample was 18 to 52 years, with mean age of 34.80 and male - female ratio of 4:1. This study showed among chronic phase CML patients bcr-abl transcript rearrangement frequency was b3a2 (e14a 2) 45% while b2a2 (e13a2) was 55% by RT-PCR testing method. Cytogenetic and molecular analysis has become mandatory in order to make a correct diagnosis and monitoring response to newer molecular targeted treatment modalities. So our health sector should be enriched with laboratories with modern technology to diagnose haemato-oncological diseases as per WHO guideline.

African horse sickness in The Gambia: circulation of a live-attenuated vaccine-derived strain.

African horse sickness virus serotype 9 (AHSV-9) has been known for some time to be circulating amongst equids in West Africa without causing any clinical disease in indigenous horse populations. Whether this is due to local breeds of horses being resistant to disease or whether the AHSV-9 strains circulating are avirulent is currently unknown. This study shows that the majority (96%) of horses and donkeys sampled across The Gambia were seropositive for AHS, despite most being unvaccinated and having no previous history of showing clinical signs of AHS. Most young horses (<3 years) were seropositive with neutralizing antibodies specific to AHSV-9. Eight young equids (<3 years) were positive for AHSV-9 by serotype-specific RT-PCR and live AHSV-9 was isolated from two of these horses. Sequence analysis revealed the presence of an AHSV-9 strain showing 100% identity to Seg-2 of the AHSV-9 reference strain, indicating that the virus circulating in The Gambia was highly likely to have been derived from a live-attenuated AHSV-9 vaccine strain.

Bluetongue virus serotype 26: infection kinetics and pathogenesis in Dorset Poll sheep.

Bluetongue virus serotype 26 (BTV-26) has recently been isolated from sheep in Kuwait. The aim of this study was to assess the pathogenicity and infection kinetics of BTV-26 in Dorset Poll sheep. Six sheep were experimentally infected with BTV-26 and samples taken throughout the study were used to determine the kinetics of infection using a pan specific BTV real time RT-PCR assay and two group specific ELISAs. Five of the six sheep showed mild clinical signs characteristic of bluetongue including conjunctivitis, reddening of the mouth mucosal membranes, slight oedema of the face and nasal discharge. Viral RNA was detected in 5 of the 6 sheep by real time RT-PCR, however the levels of viral RNA detected in the samples were lower and of shorter duration than seen with other field strains of BTV. Virus was isolated from the blood of infected animals at the peak of viraemia at around 9 dpi. Antibodies against BTV were first detected by 7 dpi using the early detection BTV ELISA and a little later (7-14 dpi) using a BTV specific competitive ELISA. Four of the five remaining sheep developed neutralising antibodies to BTV-26, measured by a serum neutralisation test (SNT), with titres (log(10)) ranging from 1.40 to 2.08.

Infection kinetics of Epizootic Haemorrhagic Disease virus serotype 6 in Holstein-Friesian cattle.

Epizootic Haemorrhagic Disease virus serotype 6 (EHDV-6) has recently caused serious outbreaks of Epizootic Haemorrhagic Disease (EHD) on the edges of Europe, in Turkey, Israel and Morocco. The aim of this study was to assess the pathogenicity and infection kinetics of EHD in Holstein-Friesian cattle infected with the two distinct strains of EHDV-6 isolated from the recent Turkish and Moroccan outbreaks. Samples taken throughout the study were used to validate two recently developed diagnostic assays that detect EHDV antibodies and viral genome. Two groups of five Holstein-Friesian cattle were experimentally infected with either the Moroccan or the Turkish isolate of EHDV-6. Cattle in both groups remained clinically unaffected throughout the study, but displayed high levels of viral RNA and virus in their blood, confirming that sub-clinical infection of cattle is likely to play an important role in EHDV transmission. A recently developed and commercialised real-time RT-PCR assay detected viral RNA as early as 2 days post infection (dpi) in both infection studies and viral RNA persisted for the course of the study. Antibodies against EHDV were first detected by 9dpi using a recently developed EHDV blocking ELISA and antibodies persisted up to the end of the study. All animals developed high levels of neutralising antibodies to EHDV-6, measured by a serum neutralisation test (SNT), with titres (log(10)) ranging from 2.20 to 2.38 at the end of the study. Virus was isolated from the blood of infected animals from as early as 2dpi up to 28dpi.

Colostral antibody protection and interference with immunity in lambs born from sheep vaccinated with an inactivated Bluetongue serotype 8 vaccine.

Widespread vaccination programmes against Bluetongue virus serotype 8 (BTV-8), using inactivated vaccines, are being carried out across many countries in northern, western and southern Europe. This study investigates the extent and length of colostral antibody protection, as well as the degree of colostral antibody induced interference of the immune response to BTV-8, in sheep. Significantly lower titres of neutralising antibodies were transferred in colostrum to lambs born from sheep vaccinated once as opposed those vaccinated twice (single vaccine in the first year and a booster vaccine in the second year). On BTV-8 challenge, lambs born from sheep vaccinated on two occasions, with the second booster vaccine given approximately 1 month prior to lambing, were protected from clinical disease for up to 14 weeks. BTV-8 was isolated from 5 of the 22 challenged lambs, although only one of these lambs showed a transient rise in body temperature with no other clinical signs. Lambs born from ewes given a second booster vaccine 1 month prior to lambing, are likely to be protected from clinical disease for at least 14 weeks, whereas lambs born from ewes vaccinated once are likely to be protected for a shorter time. Colostral antibodies present in the 13-14-week-old lambs appeared to interfere with the humoral response to challenge virus. These results suggest that colostral antibodies may interfere with vaccination in lambs up to at least 14 weeks of age.

Emergence of bluetongue serotypes in Europe, part 2: the occurrence of a BTV-11 strain in Belgium.

An EDTA-blood sample from a cow without clinical signs, which gave early birth to a newborn calf that died soon after delivery, was shown to be positive for bluetongue virus (BTV)-RNA using a group-specific real-time RT-PCR (RT-qPCR). In-house serotype-specific RT-qPCR assays for bluetongue virus serotype 1 (BTV-1), -6 and -8 all gave negative results. Subsequent assays were carried out using conventional (gel-based) RT-PCR primers for all 25 BTV serotypes and only two primer sets, both specific for BTV-11, gave bands of the expected size. The cDNAs generated were sequenced and comparisons of the genome segment 2 sequence with that of the modified 'live' vaccine strain of BTV-11 from South Africa showed 100% identity. A survey of all ruminants in a 1-km area around the first positive farm using a BTV-11 serotype-specific RT-qPCR revealed five other holdings with in total nine BTV-11 positive animals. A cross-sectional monitoring of dairy cattle in Belgium showed an overall prevalence of 3.8% on herd level and 0.2% on animal level. A BTV-11 has been introduced into the Belgian cattle herd during the 2008 vector season. The source of the infection and the way by which the virus was introduced are unknown.

Seroconversion, neutralising antibodies and protection in bluetongue serotype 8 vaccinated sheep.

Bluetongue virus serotype 8 (BTV-8) has caused a major outbreak of disease in cattle and sheep in several countries across northern and western Europe from 2006 to 2008. In 2008 the European Union instigated a mass-vaccination programme in affected countries using whole virus inactivated vaccines. We evaluated vaccinal responses in sheep and the ability of the vaccine to protect against experimental challenge. Sheep vaccinated 10 months previously under field conditions were challenged with BTV-8. One of 7 vaccinated sheep became infected, as evidenced by detection of viral RNA by real-time RT-PCR and by virus isolation. The remaining 6 sheep appeared fully protected from virus replication. None of the vaccinated sheep showed clinical signs of BTV and there was a good correlation between the presence of neutralising antibodies on challenge and protection. Commercially available ELISAs were evaluated for their ability to detect antibodies in sheep vaccinated on a single occasion. The sandwich (double antigen) ELISA assays were found to be more sensitive at detecting antibodies in vaccinated sheep than the competitive ELISAs.

Bluetongue virus: European Community proficiency test (2007) to evaluate ELISA and RT-PCR detection methods with special reference to pooling of samples.

Bluetongue virus European Community national reference laboratories (BTV-EC-NRLs) participated in an inter-laboratory proficiency test in 2007. The aim of the inter-laboratory proficiency test was to determine the ability of laboratories to detect antibodies to a series of BTV serotypes by cELISA and to detect viral RNA in animals infected with the European strain of BTV-8 by RT-PCR. Both serum and EDTA blood sample were diluted in order to determine the sensitivity of the assays. All the cELISAs were 'fit-for purpose' to detect antibodies to the common BTV serotypes circulating in Europe and the real time RT-PCR assays were all capable of detecting BTV-8 RNA albeit with varying sensitivities. There were however inconsistencies in the ability of the gel-based PCR assays to detect BTV RNA. In addition, samples taken on the first day of viraemia and at the peak of viraemia from animals experimentally infected with BTV-8, were diluted to determine if the diluting of samples affected the ability of the Shaw et al. (Shaw, A.E., M., P., Alpar, H.O., Anthony, S., Darpel, K.E., Batten, C.A., Carpenter, S., Jones, H., Oura, C.A.L., King, D.P., Elliott, H., Mellor, P.S., Mertens, P.P.C., 2007. Development and validation of a real-time RT-PCR assay to detect genome bluetongue virus segment 1. Journal of Virological Methods) RT-PCR assay to detect BTV-RNA at these time-points. Results indicated that, if samples were taken at the onset of viraemia, diluting at 1/5 resulted in a reduced ability of the assay to detect BTV RNA in the diluted compared to the neat samples. Diluting samples taken at the peak of viraemia at 1/10 however resulted in no loss in sensitivity.

Bluetongue virus: European Community inter-laboratory comparison tests to evaluate ELISA and RT-PCR detection methods.

European Community national reference laboratories participated in two inter-laboratory comparison tests in 2006 to evaluate the sensitivity and specificity of their 'in-house' ELISA and RT-PCR assays for the detection of bluetongue virus (BTV) antibodies and RNA. The first ring trial determined the ability of laboratories to detect antibodies to all 24 serotypes of BTV. The second ring trial, which included both antisera and EDTA blood samples from animals experimentally infected with the northern European strain of BTV-8, determined the ability of laboratories to detect BTV-8 antibodies and RNA, as well as the diagnostic sensitivity of the assays. A total of six C-ELISAs, six real-time RT-PCR and three conventional RT-PCR assays were used. All C-ELISAs were capable of detecting the BTV serotypes currently circulating in Europe (BTV-1, 2, 4, 8, 9 and 16), however some assays displayed inconsistencies in the detection of other serotypes, particularly BTV-19. All C-ELISAs detected BTV-8 antibodies in cattle and sheep by 21 dpi, while the majority of assays detected antibodies by 9 dpi in cattle and 8 dpi in sheep. All the RT-PCR assays were able to detect BTV-8, although the real-time assays were more sensitive compared to the conventional assays. The majority of real-time RT-PCR assays detected BTV RNA as early as 2 dpi in cattle and 3 dpi in sheep. These two ring trails provide evidence that national reference laboratories within the EC are capable of detecting BTV antibodies and RNA and provide specificity and sensitivity information on the detection methods currently available.