Revolutionising High Resolution HLA Genotyping for Transplant Assessment: Validation, Implementation and Challenges of Oxford Nanopore Technologies' Q20+ Sequencing.

The advent of third-generation sequencing (TGS) represents a significant shift in the field of genetic sequencing, enabling single-molecule sequencing to overcome limitations of short-read NGS platforms. Several studies have assessed the utilisation of TGS in HLA genotyping, though many of these studies have described the high error rate as an obstacle to achieving a robust and highly accurate HLA typing assay. In 2021, Oxford Nanopore Technologies (ONT) released the high-accuracy sequencing Kit 14 and the MinION flow cell model R10.4.1, which were reported to achieve sequencing accuracies up to 99%. The aim of this study was to validate this novel high-accuracy sequencing kit for HLA genotyping coupled with a full-gene HLA PCR assay. Comparison with historical data obtained using legacy flow cell models such as R9.4, R10.3 and R10.4 was also done to assess progressive improvement in sequencing performance with each sequential release. The workflow was validated based on data throughput, sequence quality and accuracy, and HLA genotyping resolution. An initial validation was performed using an internal reference panel of 42 samples representing common HLA allele groups, followed by an analysis of data obtained from 111 sequencing batch runs since the implementation, to assess assay performance and define quality control metrics to assess inter-run variability and monitor quality. Furthermore, challenges arising from MinION flow cell stability and use, and assessment of barcode contamination are discussed. The findings of this study highlight advantages of ONT sequencing kit 14/R10.4.1 for HLA genotyping and the implementation considerations for the routine diagnostic HLA laboratory.

"The Hotel of 10,000 Stars": The Impact of Social-Structural Determinants of Health Among Im/migrant Shrimpers in the Gulf of Mexico.

Objectives. To identify appropriate interventions to prevent injury, we conducted a qualitative study among commercial shrimp fishermen in the Gulf of Mexico. Methods. Using qualitative and participatory research methods, including interviews, photovoice, and workplace observations in southeast Texas and the Rio Grande Valley in Texas, we examined the social‒structural dimensions that contribute to physical and psychological injury. Results. We found that multiple layers of vulnerability and danger exist among shrimpers with interacting themes: (1) recognizing risk, (2) precarious employment, and (3) psychological distress. Conclusions. Our results add to the growing body of knowledge that emphasizes the negative health impacts of underregulated, high-risk, and physically demanding work performed primarily by im/migrants. Public Health Implications. Our findings highlight the larger social‒structural conditions and context of hardships endemic to migrant labor and suggest implications for practice and policy interventions. (Am J Public Health. 2024;114(8):824-832. https://doi.org/10.2105/AJPH.2024.307696).

Best practices for implementation of Kamishibai cards in the healthcare setting to improve nursing-sensitive indicator associated outcomes: An integrative review.

BACKGROUND: Many adverse events are identified as nursing-sensitive indicators (NSIs) and have evidence-based care bundles known to reduce risk of occurrence. Kamishibai cards are a tool from the manufacturing industry used for practice auditing and improvements. Use of Kamishibai cards is believed to be common in the healthcare setting, but true evidence-based guidelines do not yet exist to guide their implementation. AIMS: The aim of this integrative review was to identify best practices around the implementation of Kamishibai cards in the healthcare setting for improvement in NSI-associated outcomes. METHODS: Eleven nurses at three facilities worked through the evidence using the Johns Hopkins Evidence-Based Practice Model. RESULTS: Ten articles were included for this review. Broad themes included direct observation with non-punitive and timely feedback, clearly visualized results, use of evidence-based care bundles, pre-implementation education, and both leadership and frontline-staff involvement. All facilities showed improvement in NSI-associated outcomes after the implementation of K-cards. LINKING ACTION TO ACTION: In health care, K-cards can be implemented and designed with additional focus on the bundles of care they are intended to audit and staff support, but further evidence would better define guidelines around implementation.

Epilepsy self-management mobile health application: A needs assessment in people with epilepsy and caregivers in Viet Nam.

OBJECTIVES: This study aimed to determine (1) the needsof Vietnamese people with epilepsy (PWE) and their caregivers for self-management mobile health applications and (2) the self-management features expected to be included in an application. METHODS: The survey consisted of an anonymous self-administered questionnaire that was distributed to PWE and caregivers from the age of 18 in Vietnam through online platforms and onsite at Nguyen Tri Phuong Hospital and University Medical Center, Ho Chi Minh City, from February 2022 to May 2022. The questionnaire assessed the participants' attitudes toward epilepsy self-management mobile applications, their willingness to use applications, and their expectations of the contents of an application. RESULTS: Responses from 103 participants were submitted. Eighty-one participants (78.6%) reported using a smartphone, but only 50.6% of those claimed to know about self-management applications. Most respondents (70.9%) thought the applications would be useful for disease self-management, and 68.9% were willing to use epilepsy self-management applications. In addition, the most expected features to be included in self-management applications were epilepsy information, seizure first aid, connecting with medical professionals, and a seizure diary. CONCLUSION: Most Vietnamese PWE and caregivers had a willingness to use epilepsy self-management applications.The expected features are related to all aspects of self-management, including information, seizure, medication, and safety management.

How neurologists are viewed by their colleagues: exploring stereotypes and social representations of neurologists.

INTRODUCTION: The choice of medical specialization is influenced by various factors, including personal, educational, and interpersonal aspects. However, stereotypes and social representations (SRs) can also play a significant role in biasing the choice of a particular medical specialty. The aim of this study is to describe and understand the social representation (SR) of French Neurologist among medical peers, and factors explaining stereotypes about neurology. METHODS: A nationwide web-based survey was sent to the French medical community (students, residents, and graduated physicians) to collect sociodemographic and professional data, status, experience, and acquaintance in Neurology as well as qualitative hierarchical evocation question to assess the SR of French Neurologists. RESULTS: Overall, 367 people participated in the survey, including 112 medical students, 170 residents, and 85 graduated physicians. Only 14.3% of students listed neurology among their top 5 specialty choices, and 63.8% disagreed with the statement "I could have chosen (or I will choose) neurology after the validation of my 6th year of medical studies." Qualitative analysis revealed that the most frequently occurring words used to describe neurologists were "stroke", "complicated", "no treatment," "clinical", and "brain" and five themes corresponded to SRs of neurologist: his/her personal and professional traits (36.4% of the corpus), his/her daily practice (18.1%), the negative aspects of the neurological practice (15.3%), and the neurological pathology and daily skills (30.2%). CONCLUSION: The perception of neurologists by other physicians is nuanced. Neurologists are described as rigorous specialists, maybe excessively so but the cliché of a contemplative specialty with no effective treatment remains. The specialty and neurological patients suffer also from a reputation of complexity. Further interventions among medical students and better information are required to increase the attractiveness of our specialty.

Medical oncologist stereotypes among medical students, residents and physicians: a national cross-sectional study.

OBJECTIVES: The perception of oncologists could impact the attractiveness of the specialty and dialogue between oncologists and other physicians. The aim of the study was to describe and understand the stereotypes and social representation (SR) associated with oncologists among medical students, residents and physicians in France. METHODS: This nationwide web-based survey conducted in 2021 was based on hierarchical evocation methods. Qualitative analyses were based on the Reinert method with factorial analyses. Each respondent's SR was graded from 1 to 5 (from 1: very positive SR to 5: very negative SR). RESULTS: Oncologists suffer from a rather negative SR. The negative representation was mostly related to difficulties in practising and the proximity with death and end of life. Oncologists were also associated with more positive notions like interdisciplinarity or intellectual complexity. Attendance to an oncology course was associated with a better SR of oncology (p=0.036), whereas having someone in the family practising oncology had a negative impact (p=0.028). CONCLUSIONS: SR of oncologists is rather contrasted. It was positively influenced by attendance to an oncology course, which could be an option to correct stereotypes and update on this rapidly evolving specialty.

De novo variants of dominant monogenic disorders in Vietnam detected by a noninvasive prenatal test: a case series.

Background: Noninvasive prenatal tests for monogenic diseases (NIPT-SGG) have recently been reported as helpful in early-stage antenatal screening. Our study describes the clinical and genetic features of cases identified by NIPT-SGG. Materials & methods: In a cohort pregnancy with abnormal sonograms, affected cases were confirmed by invasive diagnostic tests concurrently, with NIPT-SGG targeting 25 common dominant single-gene diseases. Results: A total of 13 single-gene fetuses were confirmed, including Noonan and Costello syndromes, thanatophoric dysplasia, achondroplasia, osteogenesis imperfecta and Apert syndrome. Two novel variants seen were tuberous sclerosis complex (TSC2 c.4154G>A) and Alagille syndrome (JAG1 c.3452del). Conclusion: NIPT-SGG and standard tests agree on the results for 13 fetuses with monogenic disorders. This panel method of screening can benefit high-risk Vietnamese pregnancies, but further research is encouraged to expand on the causative gene panel.

How do medical students, residents and graduated physicians really perceive radiologists? A cross-national study.

BACKGROUND: Radiology has always been an attractive specialty for residents, but its attractiveness has recently decreased in France regarding the median choice rank after at the National Residency Board. AIM: To study Radiologists' perceptions and social representations (SRs) among a group of medical students, residents and graduated physicians in France, to better understand the view of Radiologists to debunk stereotypes. METHODS: The nationwide web-based survey was based on valid hierarchical evocation methods. We determined the corpus's central core and SRs' principal themes with prototypical and correspondence factor analysis (CFA), respectively. RESULTS: Overall, 419 answers were analyzed. Radiologists' SRs were divided into 3 classes: negative stereotypes of Radiologists, negative stereotypes of the Radiologists' daily practice and Radiologists' skills. After multivariate analysis, variables that seemed to have a positive influence on Radiologists' SRs were considering radiology as a potential choice of specialty (p < 0.001) and the existence of practical experience in Radiology (p = 0.008). Women seemed to have a more negative SR of Radiologists than men (p = 0.035). DISCUSSION: This was the largest qualitative study on the subject and the only one among medical students, residents and graduated physicians, allowing a global picture. SRs of Radiologists seemed to be negative, potentially caused by poor knowledge of the Radiologists' profession. CONCLUSION: SRs of Radiologists among medical students and graduated physicians appears to be negative. Promoting the specialty among medical students and encouraging their immersion in a Radiology department could help to debunk many stereotypes about the daily life and missions of Radiologists.

What do medical students and graduated physicians think about infectious disease specialists?

OBJECTIVES: This study evaluated the social representation and stereotypes on infectious disease (ID) specialists among medical students and physicians in France after the COVID-19 pandemic. METHODS: A survey applying the hierarchical evocation model assessed the social representations (SRs) of ID specialists. RESULTS: All in all, 372 answers were analyzed. The positive elements related to the personal and professional qualities of ID specialists ('intellectual prestige", "open-mindedness"), in contrast with negative stereotypes related to their perceived daily life and practice characteristics ("hospital-based", "intense", "overspecialized"). Variables such as "I would not have chosen (or I won't choose) ID after the national ranking exam" and "I know someone who is an ID specialist" were associated with worse SR scores (p < 0.001 and p = 0.022 respectively). CONCLUSIONS: These findings provide insights into the attractiveness of ID as a specialty. Rounds in ID departments may enhance the interest of the specialty as a possible residency choice.

Serum CA-125 as a predictor in the early diagnosis of ectopic pregnancy in Vietnam - A case-control study.

INTRODUCTION: This study aimed to determine the predictive value of cancer antigen-125 (CA-125) in combination with serum beta-human chorionic gonadotropin (ß-hCG) and progesterone in the early detection of ectopic pregnancy (EP). METHODS: Between May 2019 and May 2020, the cross-sectional study recruited 42 cases of EP and 42 cases of IUP at the same gestational age who visited the Department of Obstetrics and Gynecology, Hospital of Hue University of Medicine and Pharmacy. EP was diagnosed based on surgical (laparoscopy) and postoperative pathology examination. RESULTS: There were significant differences of mean level of ß-hCG (2570 mUI/mL vs. 18357.7 mUI/mL), progesterone (10.79 ± 8.16 ng/ml vs. 27.42 ± 4.17 ng/ml) and CA-125 (26.90 ± 10.26 U/mL vs. 70.61 ± 20.89 U/mL) between the EP and the IUP groups (p < 0.001). In the prediction of early diagnosis of EP, the cut-off value of CA-125 at 30.94 U/mL has a sensitivity of 89.3% and a specificity of 87,9%; the cut-off value of ß hCG at 2750mIU/ml has the sensitivity of 75%, specificity of 78,8%; the cut-off value of progesterone at 10.24 ng/mL has the sensitivity of 85.7%, specificity of 81.8%. A combination of CA-125, ß hCG, and progesterone had a sensitivity of 92.8% and a specificity of 90.9% in early diagnosis of EP. DISCUSSION: Serum CA-125 levels can be used independently or in combination with other markers in the early diagnosis of EP.

Developing and validating noninvasive prenatal testing for de novo autosomal dominant monogenic diseases in Vietnam.

Background: Over 60% of single-gene diseases in newborns are autosomal dominant variants. Noninvasive prenatal testing for monogenic conditions (NIPT-SGG) is cost-effective and timesaving, but not widely applied. This study introduces and validates NIPT-SGG in detecting 25 monogenic conditions. Methods: NIPT-SGG with a 30-gene panel applied next-generation sequencing and trio assays to confirm de novo variants. Diagnostic tests confirmed NIPT-detected cases. Results: Among 93 pregnancies with ultrasound findings, 11 (11.8%) fetuses were screened and diagnosed with monogenic diseases, mostly with Noonan syndrome. NIPT-SGG determined >99.99% of actual positive and negative cases, confirmed by diagnostic tests. No false-negatives or false-positives were reported. Conclusion: NIPT-SGG effectively identifies the fetuses affected with monogenic diseases, which is a promisingly safe and timely antenatal screening option for high-risk pregnancies.

Report from the extended HLA-DPA1 ~ promoter ~ HLA-DPB1 haplotype of the 18th international HLA and immunogenetics workshop.

The primary goal of the HLA-DPA1 ~ promoter ~ HLA-DPB1 haplotype component of the 18th IHIWS was to characterise the extended haplotypes within the HLA-DP region and survey the extent of genetic diversity in this region across human populations. In this report, we analysed single-nucleotide polymorphisms (SNPs) in 255 subjects from 6 different cohorts. The results from the HLA-DP haplotype component have validated findings from the initial pilot study. SNPs in this region were inherited in strong linkage, particularly HLA-DPA1, SNP-linked promoter haplotypes and motifs in exon 2 of HLA-DPB1. We reported 17 SNP-linked haplotypes in the promoter region. Together with HLA-DPA1 and HLA-DPB1 alleles, they formed 74 distinct extended HLA-DP haplotypes in 438 sequences. We also observed the presence of region-specific alleles and promoter haplotypes. Our approach involved phasing extended SNPs including promoter SNPs, HLA-DPA1 and HLA-DPB1 alleles, in a 22 kb region, GRCh38/hg38 (chr6:33,064,111-33,086,679), followed by clustering of these SNPs as one extended haplotype. This hierarchical clustering revealed four major clades, suggesting that haplotypes within each clade may have diverged from a common ancestral haplotype and undergone similar evolutionary processes. The correlation between HLA-DPA1 and the promoter region raises questions about the role of HLA-DPA1 antigen in the heterodimer. This finding requires validation on a larger sample size specifically designed for anthropological analysis. Nevertheless, the results from this study highlight the clinical potential of selecting better-matched donors for patients awaiting haematopoietic stem cell transplants from genetically overlapping groups that share common ancestral haplotypes.

Short communication: Antibacterial effects of essential oils from Cinnamomum cassia bark and Eucalyptus globulus leaves-The involvements of major constituents.

Essential oils from Cinnamomum cassia bark and Eucalyptus globulus leaves have been traditionally applied for bacterial infections, through both of aromatherapy and oral application. (E)-cinnamaldehyde and 1,8 cineole have been identified as their major secondary metabolites, and are also generally considered as the main active ingredients responsible for their medicinal applications. However, ethnobotanical doctors still prefer to use whole essentials oils over purified compounds in bacterial infections. We therefore hypothesized that multi-compound extracts might exert better effects than isolated ingredients. In order to verify the hypothesis about advantages of whole materials, we examined antibacterial properties of the 2 plant essential oils in the comparison with their isolated major compounds, such as (E)-cinnamaldehyde and 1,8 cineole. Effects of liquid- and vapor-phase were examined on a set of 6 gram-positive and -negative bacteria, applying broth dilution, agar well diffusion and disc volatilization methods. In all 3 investigations, we observed that whole cinnamon and eucalyptus oils, with the lower concentrations of (E)-cinnamaldehyde (89.1%) and 1,8 cineole (61.2%), were able to induce better effects than the purified active compounds (≥ 99%). These results partly explain the advantages of using whole essential oils over isolated ingredients, and therefore support the application of traditional dosage forms for bacterial infections in ethnomedicine.

Current state of RNA delivery using lipid nanoparticles to extrahepatic tissues: A review towards clinical translation.

Genetic medicine, including ribonucleic acid (RNA) therapy, has delivered numerous progresses to the treatment of diseases thanks to the development of lipid nanoparticles (LNPs) as a delivery vehicle. However, RNA therapeutics are still limited by the lack of safe, precise, and efficient delivery outside of the liver. Thus, to fully realize the potential of genetic medicine, strategies to arm LNPs with extrahepatic targeting capabilities are urgently needed. This review explores the current state of next-generation LNPs that can bring RNA biomolecules to their targeted organ. The main approaches commonly used are described, including the modulation of internal lipid chemistries, the use of conjugated targeting moieties, and the designs of clinical administration. This work will demonstrate the advances in each approach and the remaining challenges in the field, focusing on clinical translation.

Assessment of antibody levels to SARS-CoV-2 in patients with idiopathic inflammatory myopathies receiving treatment with intravenous immunoglobulin.

Antibodies to Severe Acute Respiratory Syndrome-Coronavirus 2 (SARS-CoV-2) have been reported in pooled healthy donor plasma and intravenous immunoglobulin products (IVIG). It is not known whether administration of IVIG increases circulating anti-SARS-CoV-2 antibodies (COVID ab) in IVIG recipients. COVID ab against the receptor binding domain of the spike protein were analyzed using a chemiluminescent microparticle immunoassay in patients with idiopathic inflammatory myopathies (IIM) both receiving and not receiving IVIG (IVIG and non-IVIG group, respectively). No significant differences in COVID ab levels were noted between IVIG and non-IVIG groups (417 [67-1342] AU/mL in IVIG vs 5086 [43-40,442] AU/mL in non-IVIG, p = 0.11). In linear regression models including all post-vaccination patient samples, higher number of vaccine doses was strongly associated with higher COVID ab levels (2.85 [1.21, 4.48] log AU/mL, regression coefficient [Formula: see text] [95% CI], p = 0.001), while use of RTX was associated with lower ab levels (2.73 [- 4.53, - 0.93] log AU/mL, [Formula: see text][95%CI], p = 0.004). In the IVIG group, higher total monthly doses of IVIG were associated with slightly higher COVID ab levels (0.02 [0.002-0.05] log AU/mL, p = 0.04). While patients on IVIG did not have higher COVID ab levels compared to the non-IVIG group, higher monthly doses of IVIG were associated with higher circulating levels of COVID ab in patients receiving IVIG, particularly in patients concomitantly receiving RTX. Our findings suggest that IIM patients, especially those at increased risk of COVID infection and worse COVID outcomes due to RTX therapy may have protective benefits when on concurrent IVIG treatment.

C3d-binding assay for the detection of complement activating HLA antibodies: A useful tool for allocation to highly sensitised recipients in the post-CDC era?

The CDC crossmatch test is being phased out in solid organ donor allocation, and standard luminex single antigen bead assays do not differentiate complement activating function of HLA antibodies. The current study investigated the LIFECODES C3d-binding assay to determine if it could accurately predict actual T and B cell CDC results in a cohort of highly sensitised patients. Nineteen serum samples from different highly sensitised solid organ patients were crossmatched against cells from 62 unique donors, with 174 total T and B cell crossmatches performed. The sera also underwent SAB assay using OLI and LC platforms, and C3d-binding assay. Complement activating ability of each unique HLA antibody specificity detected using SAB was assigned based on the actual CDC results, which was then used to determine the accuracy of the C3d-binding assay. The C3d-binding assay was found to be highly accurate, with sensitivity of 95%, specificity 89% and negative predictive value 97% for class I DSA and the T cell CDC crossmatch results. Furthermore, we found 100% accuracy for prediction of the complement activating function of HLA-C antibodies. Negative predictive value of above 90% was also found for HLA class II DSA. C3d-binding proved more accurate than virtual crossmatch alone to predict CDC results. This study confirms that the C3d-binding assay predicts actual CDC crossmatch results accurately. In particular, the high negative predictive value of the C3d-binding assay may be extremely useful to define HLA antibodies that do not activate complement in highly sensitised recipients.

Anorectal manometry findings in relation with long-term functional outcomes of the patients operated on for Hirschsprung's disease compared to the reference-based population.

PURPOSE: This study investigated anorectal manometry (AM) findings and bowel function of patients operated on for Hirschsprung's disease (HD). METHODS: A cross-sectional study was conducted at Children's Hospital 2. Patients operated on for HD from January 2015 to January 2020 were reviewed. Their clinical characteristics, bowel function, and manometric findings were investigated and compared with the references. RESULTS: Ninety-five patients and 95 references were enrolled. Mean ages were 6.6 ± 2.2 years and 7.2 ± 2.9 years,; fecal incontinence rates were 25.3% and 2.1%, and constipation rates were 12.6% and 4.2 for the patients versus the references, respectively. Anal resting pressures were significantly decreased in the patients compared to the references (53.2 ± 16.1 mmHg versus 62.2 ± 14.0 mmHg; p < 0.05). Among the patients, the anal resting pressure was significantly decreased in the incontinents than in the continents (46.0 ± 10.6 mmHg versus 55.6 ± 16.9 mmHg, p < 0.05). During the sensation test, the value of maximum tolerated volume was significantly decreased in the incontinents than in the continents (135.9 ± 47.9 mL versus 166.6 ± 58.3 mL, p < 0.05). CONCLUSION: AM is an objective method providing beneficial information that could guide a more adapted management in HD patients with defecation disorders.

Nanopore sequencing data analysis using Microsoft Azure cloud computing service.

Genetic information provides insights into the exome, genome, epigenetics and structural organisation of the organism. Given the enormous amount of genetic information, scientists are able to perform mammoth tasks to improve the standard of health care such as determining genetic influences on outcome of allogeneic transplantation. Cloud based computing has increasingly become a key choice for many scientists, engineers and institutions as it offers on-demand network access and users can conveniently rent rather than buy all required computing resources. With the positive advancements of cloud computing and nanopore sequencing data output, we were motivated to develop an automated and scalable analysis pipeline utilizing cloud infrastructure in Microsoft Azure to accelerate HLA genotyping service and improve the efficiency of the workflow at lower cost. In this study, we describe (i) the selection process for suitable virtual machine sizes for computing resources to balance between the best performance versus cost effectiveness; (ii) the building of Docker containers to include all tools in the cloud computational environment; (iii) the comparison of HLA genotype concordance between the in-house manual method and the automated cloud-based pipeline to assess data accuracy. In conclusion, the Microsoft Azure cloud based data analysis pipeline was shown to meet all the key imperatives for performance, cost, usability, simplicity and accuracy. Importantly, the pipeline allows for the on-going maintenance and testing of version changes before implementation. This pipeline is suitable for the data analysis from MinION sequencing platform and could be adopted for other data analysis application processes.