bluuebunny/crossref_metadata_2025 huggingface dataset:
Disseminating Education about HER2-Low Breast Cancer through Twitter Chats and Tweetorials
Authors: Kim J , Chivukula M , Feldman M , Wei S , Baskota S , Bui M , Khoury T , Akhtar I , Beumer K , Kelly M , O'Regan R , Krystel-Whittemore M
Published: November 2023

Introduction/Objective

Social media is often used to disseminate new information and as a teaching tool that reaches a broad audience. When practice-changing data about HER2-Low metastatic breast cancer (MBC) emerged in 2022, ASCP developed CME/CMLE Twitter chats and Tweetorials to educate pathologists and laboratory professionals about how these updates will impact the diagnosis and care of patients with HER2-Low MBC. These activities were funded by educational grants from AstraZeneca and Daiichi Sankyo, Inc.

Methods/Case Report

Working with faculty who were active on Twitter, ASCP organized Twitter chats and Tweetorials to increase awareness about HER2-Low MBC and address how these changes will impact clinical care. A Twitter chat is a live, interactive Tweet-based discussion held on Twitter. Also called Tweet chats, these events run for 1 hr and use hashtags to keep the discussion organized. A Tweetorial is a combination of threaded Tweets that teach about a specific topic. Activities were certified for continuing education credit and were designed to be short, practical, and relevant for pathologists and laboratory professionals.

Results (if a Case Study enter NA)

In 2022 and 2023, ASCP used the hashtag #HER2Low to organize Twitter chats and Tweetorials. Each chat had four faculty members responding to 6-8 questions and averaged 2.2 million impressions (an impression is the # of times a Tweet is served in a timeline or search result). Faculty covered the challenges differentiating an IHC score of 0 vs 1+, the emerging use of digital pathology, ways to improve reporting, and other challenges with IHC scoring. Tweets may be amplified by Retweets (a re-posting of a Tweet), so a Tweet sent by a faculty member often reached over 1,000 views. Tweetorials had polling questions, text, images, videos, and links to journal articles. Each Tweetorial achieved over 1,400 views and over 535,000 impressions during its first week. Collectively, the Twitter Chats and Tweetorials led to over 1,050 CME/CMLE certificates.

Conclusion

Twitter is an evolving platform that is actively used by pathologists (see #PathTwitter) and laboratory professionals to share knowledge to a broad audience and foster a community of learning. Since content posted on Twitter remains enduring and searchable, people may find old Tweets and see which topics achieved high impressions. When spreading education on new practice-changing topics like HER2-Low breast cancer, Twitter can be an effective platform for broad content dissemination.
Orthogonal biomarkers of Inflammation due to Aging Identified in Saliva and Serum
Authors: Jensen M M , Vu D , Vigerust D , Gupta R , Maecker H T , Nguyen T T
Published: November 2023

Introduction/Objective

The relationship between age and inflammation is well established, but little is known about the contribution of different biological fluids to this relationship. This study aims to compare inflammatory markers in young and old subjects using human saliva and serum samples.

Methods/Case Report

Saliva and serum were collected from young (18-27 years, n=15) and old (53-68 years, n=15) subjects without any reported pathologies and analyzed for inflammatory markers using an Olink® Target 96 Inflammation panel. Each group consisted of individuals with no known pathologies and in good health with 8 female and 7 male patients per group. Salvia samples were using a spectrum solutions Saliva Mulitiomic Device with associated preservation solution. Matched serum samples were taken within 15 minutes of saliva collection. Results from the link were analyzed using a two-way ANOVA with a Sidak correction for multiple comparisons.

Results (if a Case Study enter NA)

Results showed that there were significant differences in inflammatory marker levels between young and old subjects in both saliva and serum samples. However, some specific changes were only found in saliva such as changes to CD8A, IL7, CXCL11, TRAIL, IL20RA, NFG-β, CXCL10, CCL28, DNER CD40, and CXCL1. Other distinct changes were identified only serum samples for CDCP1, OPG, MCP4, FDG23, FDF5, LIFR, FLT3L, IL-33, CCL25, NT3, and TWEAK. Interestingly, only MCP1, CXCL9, CST5, showed statistically significant changes in both saliva and serum, indicating 89% orthogonality for the inflammatory markers analyzed. Only CXCL5 showed variation based on the subject’s sex assigned at birth in both serum and saliva.

Conclusion

These findings suggest that saliva and serum are both useful biological fluids for the assessment of age- related changes in inflammation and provide distinct sets of information. This data provides valuable information for understanding the mechanisms of aging and disease could benefit from the analysis of multiple biological fluid samples.
A novel GLCC1::BRAF fusion with independent MYCN and MYC amplifications in primary tumor and metastasis in pediatric pancreatic acinar carcinoma
Authors: Miller L J , Heider A , Shao L
Published: November 2023

Introduction/Objective

Pediatric pancreatic acinar cell carcinoma (PACC) is a rare malignancy, comprising 5-15% of pediatric pancreatic tumors. BRAF rearrangement is found in 20-30% of PACC cases, most commonly an inversion in chromosome 7. We report a case of PACC with GLCC1::BRAF fusion.

Methods/Case Report

A 10-year-old male presented with six months of weight loss, back pain, and loose stools. Imaging demonstrated concentric soft tissue thickening around the celiac axis and superior mesenteric artery, clinically interpreted as vasculitis. Follow-up imaging showed extension of the soft tissue thickening, prompting biopsy of a periaortic lymph node showing metastatic PACC. Pancreaticduodenectomy revealed a 3.8 cm pancreatic neck mass with perineural invasion and metastatic deposits in multiple lymph nodes and retroperitoneal soft tissue.

Fluorescence in situ hybridization of both specimens demonstrated BRAF gene rearrangement, with the partner identified as GLCC1 by next generation sequencing and fusion assays. Microarray on the lymph node demonstrated amplification of MYC on chromosome 8, but not in the resection specimen, which showed gain of chromosome 2 and amplification of MYCN.

Results (if a Case Study enter NA)

NA

Conclusion

The GLCC1::BRAF fusion has previously been reported once in a case of pheochromocytoma with peritoneal seeding. The MYC gene family members, c-MYC and MYCN, are involved many cancer types; amplifications are often associated with unfavorable prognosis in tumors such as neuroblastoma. However, the amplification of both genes in the same tumor at different loci has not been reported. The combination of the primary GLCC1::BRAF fusion with secondary amplification of MYC and MYCN is likely to drive the aggressive behavior and metastasis in this case of PACC. Treatment for PACC includes chemoradiation and BRAF inhibition. However, BRAF rearrangements without the V600E mutation show variable response to BRAF inhibitors. Potential alternatives include inhibitors to other targets within the BRAF signaling pathway. Further investigation is needed to determine the clinicopathologic impact of these novel genetic aberrancies.
Evaluating Genetic Alterations in Metastatic Pancreatic Neuroendocrine Tumors with Potential Therapeutic and Prognostic Applications: A Comparative In Silico Study with Metastatic Neuroendocrine Tumors Originating from the Small Intestine and Rectum
Authors: Mousavi S , Mirzaei Baboli K , Arabi M , Bereimipour A
Published: November 2023

Introduction/Objective

Pancreatic neuroendocrine tumors (P-NET) are diverse tumors with slow growth and varying levels of potential malignant behavior. The overall 5-year survival rates for P-NET are approximately 95%, 72%, and 23% if localized, and in cases of regional and distant metastasis respectively. These tumors exhibit a low rate of mutations but show varied genomic abnormalities, contributing to their unpredictable nature. In this study, we aimed to find the genetic alterations observed in metastatic P-NET for their potential therapeutic and prognostic applications and compared these tumors to metastatic neuroendocrine tumors originating from the small intestine (SI-NET) and rectum (R-NET).

Methods/Case Report

We initially extracted RNA-seq datasets from the GEO database. Next, we analyzed the datasets in six defined groups and differentially expressed genes were obtained by comparison to controls. Either of the groups of P-NET, SI-NET, or R-NET cases were evaluated individually and altogether (primary vs metastatic; 4 groups) and in 2 groups comparing primary/metastatic P-NET cases to primary/metastatic cases of SI-NET plus R- NET. Using a Venn diagram, we isolated the commonality between gene expression profiles. Afterward, we obtained the ontology of genes and signaling pathways from KEGG and Enrichr. Finally, we drew the protein network and evaluated the proteins in GEPIA and TCGA clinical databases.

Results (if a Case Study enter NA)

A total of 212 cases including 113 P-NET (83 primary and 30 metastatic), 81 SI- NET (44 primary and 37 metastatic), and 18 R-NET (3 primary and 15 metastatic) were selected. We found 8 genes (ORM1, HP, APOA2, HRG, KNG1, ALB, SERPINC1, and AHSG) are mutually markedly upregulated in cases of metastatic P-NET and SI-NET but were not identified in metastatic R-NET. The products of these genes were predominantly involved in complement cascades. CABYR, MYCN, and CALU genes were among the most upregulated genes specifically isolated in the metastatic P-NET that played roles in activating intracellular signaling pathways, transcription, and protein folding.

Conclusion

The shared upregulated genes in metastatic P-NET and SI-NET may support the relatively better prognosis of these tumors in comparison to the metastatic R-NET due to the activation of the complement cascade (immune system). Furthermore, the identified genetic alterations in the metastatic P-NET cases may have potential therapeutic and prognostic applications for future use.
Qualitopix: Artificial intelligence-based quantitative quality assurance of immunohistochemistry staining-The Henry Ford Health experience
Authors: Baba O
Published: November 2023

Introduction/Objective

Immunohistochemistry (IHC) offers crucial patient data for diagnosis, prognosis, and treatment. Pathologists currently assess stain quality subjectively, comparing control sections to patient tissue. Qualitopix (Visiopharm, Denmark) is a cloud-based platform that enables objective, quantitative analysis of staining consistency through external cell line controls.

Methods/Case Report

Over six months, we tested 1121 slides from five cell-line blocks with epitopes of different intensities for ER, PR, Ki-67, Her-2 Neu, and PD-L1. Slides were stained using Ventana Benchmark Ultra (Roche, Basel, Switzerland) for ER, PR, and Her-2 and Dako Omnis (Agilent, Santa Clara, California, United states) for Ki-67 and PD-L1. They were scanned with DP 200 scanner (Roche) and uploaded to Qualitopix for image analysis. Cells were detected using artificial intelligence and classified based on diaminobenzidine (DAB) staining intensity, reported as H-scores (0-100). Outliers were determined at 1 standard deviation. a selection of outliers was rescanned. Inter- scanner comparisons and repeatability studies were performed for ER and 50 within and between the DP 200 and HT scanners (Roche).

Results (if a Case Study enter NA)

Our analysis revealed a 26-34% rate of outliers at 1 SD for the five stains, 10 times higher than the currently reported number from our lab. Most outliers (70-100%) remained out of range at rescanning, suggesting pre-analytical technical issues. 5-33% of slides failed analysis due to technical errors. Poor discrimination between sequential cores of varying intensity for ER and Ki-67 was noted. Inter-scanner comparison and repeatability studies demonstrated precise and consistent H-scores within and across both scanning platforms for ER and PR.

Conclusion

Qualitopix offers automated, objective, and quantitative quality assurance for IHC stain quality assessment, a hitherto manual, subjective and qualitative-driven process. Our findings emphasize the need for continuous IHC quality monitoring. The origin of outliers, technical failures, and their correlation with pathologist's subjective findings require further investigation.
Use of Plasmapheresis in Heparin Induced Thrombocytopenia in Patients Undergoing Urgent Cardiac Surgery
Authors: Bava Ejas Palathingal , Epelman Mathew , Yeldo Nicholas , Uribe-Marquez Santiago , Lopez-Plaza Ileana
Published: November 2023

Heparin Induced Thrombocytopenia (HIT) is a potentially life-threatening but uncommon disease process characterized by IgG antibodies that recognize platelet factor 4/heparin immune complexes causing thrombocytopenia and thrombosis. Although therapeutic plasmapheresis (PLEX) appears to be effective, protocols regarding how and when to institute PLEX remain elusive. At our institution, we put together a standardized approach to assess the interventional success across patients that included 1 plasmapheresis procedure per day over 4 days, processing one plasma volume and using thawed plasma as the replacement fluid. Treatment effectiveness was evaluated by pre- and post-interventional HIPA OD levels and platelet count. Treatment aim was to decrease the HIPA OD to less than 0.6 and obtain a normal platelet count.

The aim of this study was to design a protocol for PLEX to address urgent need for patients with HIT for which heparin is planned for intraoperative cardiac surgery requiring cardiopulmonary bypass (CPB) anticoagulation.

We retrospectively examined the medical records of 2 HIT-positive patients who underwent PLEX prior to cardiac surgery at our large quaternary care hospital. Both patients had a 4Ts score of 5, with a medium probability of disease. One patient's HIT was confirmed by a positive serotonin release assay (SRA). The second patient had persistently negative confirmatory test but was deemed clinical HIT.

The first patient is a 53-year-old female, who was admitted for mitral valve replacement, aortic valve replacement, and tricuspid valve repair. She was confirmed HIT by positive SRA. Three sessions of PLEX were performed on consecutive days. Her HIPA (heparin induced platelet antibody) O.D. value decreased from 1.925 to 0.534 and platelet counts normalized. The second patient is a 73-year-old male, who was admitted for LVAD implantation and LV thrombus resection. Two sessions of PLEX were performed on consecutive days. His HIPA O.D. value deceased from 1.261 to < 0.4 (negative) and platelet counts normalized. For both patients, the last PLEX procedure was performed the day of surgery, and the cardiac surgical procedure was completed successfully.

Based on these 2 patients, we have proposed a protocol for PLEX in HIT patients as follows: daily plasmapheresis x 4 days will be performed until the OD has decreased to less than 0.6 and platelet count has normalized. Each plasmapheresis will consist of the processing of one plasma volume, using thawed plasma as the replacement fluid. The last plasmapheresis will occur on the day of surgery prior to surgery. Further assessment will be performed if the targeted parameters have not been met by the 4th plasmapheresis procedure.

In conclusion, we present 2 patients with HIT who benefitted from PLEX prior to cardiac surgery under CPB. We will adopt the protocol used for these 2 patients as our standard of practice moving forward.
Evaluating the utility and challenges associated with "anonymous" patients in the electronic medical record
Authors: Rogers Kai , Krasowski Matthew
Published: November 2023

Electronic medical records (EMRs) allow for the creation of "test" patients that can be used for training new users, testing changes before they go "live", simulating the placement of orders, or for a range of similar applications. Test patients are commonly utilized in EMR validation, support, or “playground” environments that might use a copy of the production EMR database but do not connect to the official medical record to prevent billing errors or patient harm. However, there are a variety of cases where it is useful to have test or anonymous patients within the EMR production environment as well. One example of this is infectious disease testing for employee blood-borne pathogen exposure (e.g., needlestick injury) that may need to be stored in a de-identified or restricted manner to comply with workplace privacy regulations. There are also instances, generally in emergency situations, where a medical record is created for a patient about whom little identifying information is known. In this example, these so-called "anonymous" patients are assigned random or arbitrarily pre-determined birthdates (e.g., 1/1/1900) and on occasion have no sex assigned in their chart. This poses a variety of challenges from a laboratory standpoint as it can be difficult to provide accurate normal ranges for laboratory results that are age and/or gender specific. Additionally, this has on occasion resulted in an inability to identify appropriate blood products as these recommendations are similarly based on age and gender. Another layer of complexity comes as additional information regarding the patient is made available, and health care information services merges data from a previously anonymous account to a specific patient record. Such mergers can result in cancellation of pending orders, errors in reporting laboratory results, or other complications depending on the specific process utilized. Surprisingly, there is little to no literature regarding the use cases for these “test” and “anonymous” patients or the challenges associated with their existence in the EMR. To explore this topic, we identified three classes of patients in our institutional EMR: true virtual “test” patients, “anonymous” patients that were subsequently identified, and “anonymous” patients that were never identified. Basic characteristics of these patients (sex and date of birth) and available laboratory results were compiled. Along with this, we present a series of instructional cases adapted from actual patient safety events at our institution involving anonymous patient records. These illustrative cases highlight the utility of these “test” and “anonymous” patients, as well as the challenges these pose on an institutional and individual level. Lastly, we provide recommendations on how best to manage similar scenarios that may arise.
Multilocus Sequence Typing of Emerging Azole-Resistant Candida auris Isolates from Critically Ill Patients in a New York City Medical Center
Authors: Weidmann Maxwell , Srivastava-Malick Mahavrat , Shi Jun , Wu Fann
Published: November 2023

Candida auris has developed into a slow-moving global pandemic since its emergence in 2009, particularly as a source of nosocomial infection amongst immunocompromised and critically-ill patients. According to CDC data, C. auris emergence has accelerated in the US during the COVID-19 pandemic and has garnered particular attention for its high level of resistance to multiple classes of antifungals. C. auris has been categorized into five genetically diverse clades, each with unique antifungal drug resistance patterns. We have observed a sharp increase in cases identified at our tertiary medical center in New York City over the past three years. In this study, we designed a set of C. auris-specific oligonucleotides to determine the clonality of isolates through Multilocus Sequence Typing (MLST) of four genetic loci known to have a high level of divergence amongst Candida species. From 2020 through 2022 we identified 22 patients with C. auris bloodstream infection at our institution by Matrix-assisted laser desorption/ionization-time of flight (MALDI-TOF) and performed MLST on isolates that were available from 19 patients. We assessed four genetic loci: ITS1/ITS2, D1/D2, RPB1 and RPB2; ERG11 was included to assess for mutations associated with fluconazole resistance. We found that the number of blood stream infections increased from 2 cases in 2020 to 12 in 2022. Our MLST results showed that all C. auris isolates from blood and body fluids belong to CDC-defined Clade 1c, which carries the ERG11 gene mutation resulting the amino acid substitution K143R in 14-α-Demethylase associated with fluconazole resistance. These isolates also demonstrated fluconazole resistance by in vitro testing (19/19), amphotericin resistance was present in 44% of isolates with available data (7/16), and all isolates were susceptible to micafungin (19/19). Of 19 patients with blood cultures positive for C. auris, 13 (68%) were in a critical care setting at time of collection, 17 (89%) had been in an ICU during the same admission, and 8 (42%) expired within 3 months of C. auris positivity. MLST is considered a relatively fast and inexpensive method of sequence-based strain typing and has recently been applied to assessing the clonality of C. auris isolates. We show that the use of C. auris-specific oligonucleotides for sequencing, rather than those derived from other Candida species, produces high quality data for MLST, as well as the detection of specific mutations involved in antifungal resistance, such as ERG11. Further assessment is needed by the gold standard in sequence typing, Whole Genome Sequencing (WGS), to confirm our analysis of clonality here. MLST may therefore represent a useful tool in understanding the interrelatedness of invasive C. auris infections in hospitalized patients as well as inform clinical decision-making around infection control and empiric antifungal therapy.
Detection of Prostate Cancer Screening Disparities Using Geospatial Analysis of Laboratory Results
Authors: Azimi Vahid , Zaydman Mark
Published: November 2023

Black men experience higher prevalence and mortality from prostate cancer (PCa) but are less likely to undergo PCa screening with prostate-specific antigen (PSA). The reasons for this disparity are multifactorial and include lack of access, PCa education, and mistrust in the healthcare system. Identifying geographic regions with low PSA screening and high positivity rates could potentially help inform targeted interventions. This study aimed to examine racial, socioeconomic, and geospatial disparities in PSA screening and positivity rates in the Siteman Cancer Center (St. Louis, MO) catchment region. The results of all PSA screening tests ordered in the Barnes-Jewish Healthcare Corporation (St. Louis, MO) and performed from 9/2019-12/2022 were retrieved from the laboratory information system with patient demographics. Patient addresses were geocoded using ArcGIS Pro (ESRI, Redlands, CA) and assigned to a census tract of residence. The PSA screening-eligible population was defined as the number of persons ≥50 years of age based on U.S. Census data. Social Vulnerability Index (SVI) values were retrieved from the Centers for Disease Control. The PSA screening rate was calculated by census tract as the total number of PSA tests performed/PSA screening-eligible population. The PSA positivity rate was calculated by census tract as the number of PSA positive tests/total PSA tests. All census tracts with fewer than 10 PSA tests performed were omitted from PSA positivity analysis. Hotspot analysis was performed by calculating Local Moran’s I statistic per census tract and comparing with 999 randomly-generated maps. A total of 90,994 screening tests for 60,005 patients (48,024 White [80%], 10,425 Black [17%], and 1,556 Other [3%]) were performed for patients in the catchment region. Black patients were 78% more likely to receive a positive PSA screen than White patients (p<0.001). Of those screening positive, Black patients had a significantly higher median[IQR] (7.94[6.2-13.7 ng/mL]) PSA result compared to White patients (7.4[6.2-10.1 ng/mL]) (p<0.001) and a median[IQR] age of 67[61-74 years] vs. 69[63-76 years] (p<0.001). In a multivariate logistic regression using age, race, and census tract of residence SVI to predict PSA positivity, only census tract SVI was not statistically significant. A model using only age and race demonstrated that every 10 years of age and Black race were associated with 11% and 7% increased odds of receiving a PSA positive test, respectively (p<0.001). 75 and 191 of 789 census tracts in the catchment region were identified as PSA positivity hotspots and PSA screening cold spots, respectively. In conclusion, this study confirms previous reports of higher PSA values among Black patients and suggests that geospatial analysis of PSA testing can identify geographic regions with low testing and high positivity rates.
Simplifying molecular testing through electronic order entry
Authors: Kwon Regina , Krumm Niklas , Hoffman Noah , Mathias Patrick , Konnick Eric
Published: November 2023

Our solid-tumor molecular laboratory receives more than 3,000 orders each year, with about two-thirds coming from internal providers. Before this project, molecular tests were ordered using one of two different paper forms, which the care team then faxed or e-mailed to the Genetics Preanalytical Service (GPS). These requests led to frequent modifications and cancellations due to the complexity of molecular testing. A small fraction of requisitions were lost due to transmission errors. We undertook a project to design, develop, and implement computerized physician order entry (CPOE) in Epic (Epic Systems Corp., Verona, WI) and Sunquest (Clinisys, Tucson, AZ). We conducted extensive interviews with the directors of the molecular lab, laboratory medicine IT, GPS, anatomic pathology staff, laboratory scientists, laboratory genetic counselors, oncologists, clinicians, and nurse coordinators. The order itself and other technical changes underwent iterative review and testing by a subset of interviewees. Five oncologists participated in formal beta-testing. The user interviews allowed us to produce a detailed process diagram that drew attention to specific areas of repetition or risk. We proposed a new workflow that would eliminate repetition, provide redundant order-tracking, and automate some lab processes. Once approved, we built the electronic order in Epic and modified existing laboratory programs and interfaces as needed. The process of placing an order, which previously typically involved both nurses and doctors over several calendar days, was now being completed in fewer than 5 minutes by a single practitioner. Requisition status events were no longer created manually in Sunquest by GPS. Certain data-entry tasks were eliminated through the incorporation of label printers. Within one month of launch, more than 50% of somatic molecular orders arrived via CPOE, increasing to 90% within six months. At a rate of 125 CPOE orders per month, time savings range from 20 hours per month for GPS staff, 50 hours per month for nursing, and 20 hours per month for physicians. The success of this project demonstrates the feasibility of replacing a paper-based process for a complex test menu with CPOE. Due to the intricacies of molecular testing, such a change will undoubtedly involve multiple divisions. We attribute the success of this project to early involvement by key stakeholder groups, including developers; an extended testing phase to ensure reliability at roll-out; creation of straightforward reference documents; and rapid response to user questions, especially during the first weeks post-launch.
Anticoagulation management in patients tested for heparin-induced thrombocytopenia in the cardiothoracic intensive care unit diverges from expert guidelines
Authors: Spies Nicholas , Jackups Ronald , Zaydman Mark
Published: November 2023

Heparin-induced thrombocytopenia (HIT) is a rare, life-threatening complication in patients receiving heparin products. Clinical scoring tools, such as the ‘4T’ score, aim to help identify patients at risk for HIT and are recommended by expert guidelines. For example, the American Society of Hematology (ASH) recommends that patients with an intermediate or high-risk 4T score are presumptively treated for HIT by withdrawing heparin and starting an alternative anticoagulant. At our institution, laboratory HIT testing involves an in-house latex immunoassay (LIA) that, if positive, reflexes to a send-out serotonin release assay (SRA) at the Blood Center of Wisconsin. This confirmation may take several days to result, underscoring the importance of presumptive treatment. Anecdotally, we have observed that clinicians have expressed reservations about the use of the 4T score in the cardiothoracic intensive care unit (CTICU). These reservations may lead to increased HIT testing in patients with low 4T scores and delays in the initiation of alternative anticoagulants. We set out to objectively determine if clinical practice in the CTICU is aligned with ASH guidelines. To do so, we used anticoagulant usage as a proxy for medical decisions made at the time of order for HIT laboratory testing. Data was extracted from the electronic medical records of all patients who were admitted to the CTICU between 5/2018 and 7/2022 and administered a heparin product (n = 1,602). Patients that were not tested for HIT (n = 1,449) were excluded. The remaining patients’ (n = 153) medication record timings were normalized such that the time of their first LIA was set to zero to allow for relative comparisons. 25 (16%) of the LIAs were positive, of which 5 (20%) were confirmed by SRA. Of the 153 patients, 136 (89%) received heparin within 24 hours prior to LIA order, while 6 (4%) were already on alternative anticoagulation (bivalirudin). Of the patients on heparin prior to LIA, 18 (13%) were switched to bivalirudin upon LIA order, 66 (49%) had heparin discontinued without an alternative, and 52 (39%) were continued on heparin. Our results suggest that the prevalence of HIT in this cohort is 3.3% by SRA, much lower than the expected pretest probability estimated for intermediate- and high-risk 4T scores in published meta-analyses. In addition, the observed anticoagulation practices varied widely and diverged from ASH guidelines, where only 17% of patients were on alternative anticoagulation when the testing was ordered. Future work will investigate these patterns across LIA positivity and 4T scores while attempting to address the clinical outcomes of these treatment decisions.
Delayed Hemolytic Transfusion Reactions Could be Prevented by a National Alloimmunization Registry
Authors: Stanek Caroline , Miller Brianna , Marques Marisa
Published: November 2023

Evanescence of alloantibodies to RBC antigens places patients at risk of delayed hemolytic transfusion reactions (DHTRs) unless the transfusion service is aware of their alloantibody history. In the past year, three patients with sickle cell disease (SCD) developed a DHTR at our institution because their pre-transfusion testing did not reflect their full alloimmunezation. We sought to determine how often the pre-transfusion antibody screen is negative or fails to show the full alloimmunization profile. We reviewed the electronic records of all current adult patients with SCD receiving chronic outpatient RBC transfusions (including red cell exchange) to collect demographic and laboratory results to answer this question. Our analysis included 162 patients of African-American descent: 80 (49%) males and 82 (51%) females, with an average age of 34 years. The majority, 92 or 56%, had a history of alloimmunization with a mean number of 3 antibodies (range: 1-10). The most common were to the C, E, and K antigens. The next two most common alloantibodies were nonspecific, representing alloimmunization to an antigen that could not be further determined, followed by the D antigen. In 81 patients, the most recent antibody screen did not reflect their history, with most being completely negative (n=69), and the rest (n=12) unable to detect all alloantibodies of which they had a history. The antibodies most often undetectable were: Anti-E, Anti-C, and Anti-D. These are simultaneously the second, first, and fifth most common antibodies in this population, all of which are considered clinically significant and capable of causing a hemolytic transfusion reaction. Of note, the alloantibodies implicated in the recent DHTRs we observed, were to the JKa, Jkb, Fya and the S antigens. In addition, 28 individuals (17% of the total) had at least one antibody to a low incidence antigen (LIA) such as Goa, V, Vs, Jsa, Kpa and Lua. Five (3%) patients were exclusively sensitized to one or more low incidence antigens. Considering that commercially available reagent red cells utilized to perform pre-transfusion testing may lack LIAs, knowing the history of alloimmunization is the only way to prevent hemolytic reactions. In conclusion, evanescence and limitations of the commercially available reagents to test patients with SCD prior to a transfusion place them at increased risk of transfusion reactions. One potential way to minimize risk from the latter is to perform serologic crossmatching of all units intended for patients with SCD, independent of the result of the antibody screen. This is the practice we have instituted several years ago in our transfusion service. However, the solution to both problems is a national repository of alloimmunization histories accessible to all transfusion services. Such informatics tools would help avoid DHTRs in patients with and without SCD.
Monitoring Infected Reticulocytes During Murine Malaria Infection
Authors: Gordy Dominique , Hudson Krystalyn , Thomas Tiffany , Shah Shivang , Stone Elizabeth
Published: November 2023

Malaria infection is a major cause of mortality worldwide. According to the 2022 World Health Organization Malaria Report, malaria caused approximately 247 million infections and 619,000 deaths in 2021, with more than 470,000 children dying of malaria. The gold-standard for measuring parasitemia is counting infected red blood cells (RBCs) by eye using Giemsa staining whole blood smears. This method is time consuming and is subject to error by both who is counting, the number of cells counted, and which frames are selected for counting. Automating the process of measuring malaria infection by flow cytometry decreases both the time required to obtain accurate parasitemia counts and the possible error on behalf of the researcher. Hundreds of thousands of cells can be counted by flow cytometry in a shorter period of time. Animal models of malaria infection are widespread and well-characterized. The rodent malaria strain P. berghei has shown preferential infection of reticulocytes over mature RBCs. Here we outline a method of measuring P. berghei blood stage infection in reticulocytes and mature RBCs and comparing the results to Giemsa stained whole blood smears. Using commercially available P. berghei ANKA expressing green-fluorescent protein (GFP), we infected wild type mice with 1x106 parasitized RBCs by intraperitoneal injection. Whole blood samples were collected daily starting day 3 post infection through day 7 by tail vein collection. Random and representative fields of Giemsa stained peripheral smears were selected to measure the percentage of parasitemia in mature RBCs and reticulocytes for a total cell count of approximately 1000. For flow cytometry, whole blood was incubated in 500uL of 1X SYBR Green I Nucleic Acid Gel Stain in PBS at 37°C for 20 minutes in the dark, washed with PBS, and stained with anti-CD71 to label reticulocytes. During the early stages of infection, the reticulocyte percentage of infected animals is not significant from uninfected animals, approximately 1.86%. As parasitemia increases over time, although the percentage of total reticulocytes decreases, the percentage of infected reticulocytes increases. On day 3 through 7 for one experimental replicate, day 3 demonstrated 1.86% total reticulocytes, and of these 2.56% reticulocytes were infected; day 4 had 0.66% total reticulocytes, and of these 32.92% were infected; day 5 showed 0.18% reticulocytes with 71.81% infected reticulocytes; on day 6 there were 0.04% reticulocytes with 75.44% infected reticulocytes; and day 7 had 0.01% reticulocytes with 69.56% infected reticulocytes. There was a strong correlation (R2 = 0.9147) when comparing flow cytometry to measure parasitemia with counting parasitemia by peripheral smear. This flow cytometry panel is an excellent tool to assess malarial infection of reticulocytes compared to mature RBCs and can be applied to other malarial species while decreasing bias, error and time needed to conduct the experiment.
‘Reputation, reputation, reputation! Oh, I have lost my reputation!’; A literature review on alcohol addiction in the British Sikh and/or Punjabi community and the barriers to accessing support
Authors: Kaur Karamdeep
Published: November 2023

Aims

The literature review aimed to identify the barriers the Punjabi and/or Sikh community have in accessing support for alcohol addiction.

Method

A systematic review of the literature was undertaken of four scholarly databases, Google Scholar and grey literature on UK-based research conducted after 1980 into alcohol addiction in the Punjabi and/or Sikh community. Fourteen papers met the inclusion criteria outlined in the paper and were included in the review.

Results

Ten main barriers to accessing support were found; stigma, religion, lack of understanding of addiction, over reliance on a medical model of treatment and disregard of therapy, cultural implications of being a member of the Punjabi and/or Sikh community and the addiction community, gender and generational differences and a lack of government commitment to alcohol support for BME communities with a lack of culturally specific services.

Conclusion

Despite the many barriers explored, the role of stigma remained a powerful theme throughout often underpinning other barriers. A key recommendation across many papers was the need for culturally sensitive support services. Several areas for future research were identified.
Assessing protein model quality based on deep graph coupled networks using protein language model
Authors: Liu Dong , Zhang Biao , Liu Jun , Li Hui , Song Le , Zhang Guijun
Published: November 2023

Model quality evaluation is a crucial part of protein structural biology. How to distinguish high-quality models from low-quality models, and to assess which high-quality models have relatively incorrect regions for improvement, are remain a challenge. More importantly, the quality assessment of multimer models is a hot topic for structure prediction. In this study, we propose GraphCPLMQA, a novel approach for evaluating residue-level model quality that combines graph coupled networks and embeddings from protein language models. The GraphCPLMQA consists of a graph encoding module and a transform-based convolutional decoding module. In encoding module, the underlying relational representations of sequence and high-dimensional geometry structure are extracted by protein language models with Evolutionary Scale Modeling. In decoding module, the mapping connection between structure and quality is inferred by the representations and low-dimensional features. Specifically, the triangular location and residue level contact order features are designed to enhance the association between the local structure and the overall topology. Experimental results demonstrate that GraphCPLMQA using single-sequence embedding achieves the best performance compared with the CASP15 residue-level interface evaluation methods among 9108 models in the local residue interface test set of CASP15 multimers. In CAMEO blind test (20 May 2022 to 13 August 2022), GraphCPLMQA ranked first compared with other servers (https://www.cameo3d.org/quality-estimation). GraphCPLMQA also outperforms state-of-the-art methods on 19, 035 models in CASP13 and CASP14 monomer test set.
FireProt 2.0: web-based platform for the fully automated design of thermostable proteins
Authors: Musil Milos , Jezik Andrej , Horackova Jana , Borko Simeon , Kabourek Petr , Damborsky Jiri , Bednar David
Published: November 2023

Thermostable proteins find their use in numerous biomedical and biotechnological applications. However, the computational design of stable proteins often results in single-point mutations with a limited effect on protein stability. However, the construction of stable multiple-point mutants can prove difficult due to the possibility of antagonistic effects between individual mutations. FireProt protocol enables the automated computational design of highly stable multiple-point mutants. FireProt 2.0 builds on top of the previously published FireProt web, retaining the original functionality and expanding it with several new stabilization strategies. FireProt 2.0 integrates the AlphaFold database and the homology modeling for structure prediction, enabling calculations starting from a sequence. Multiple-point designs are constructed using the Bron–Kerbosch algorithm minimizing the antagonistic effect between the individual mutations. Users can newly limit the FireProt calculation to a set of user-defined mutations, run a saturation mutagenesis of the whole protein or select rigidifying mutations based on B-factors. Evolution-based back-to-consensus strategy is complemented by ancestral sequence reconstruction. FireProt 2.0 is significantly faster and a reworked graphical user interface broadens the tool’s availability even to users with older hardware. FireProt 2.0 is freely available at http://loschmidt.chemi.muni.cz/fireprotweb.
Phanto-IDP: compact model for precise intrinsically disordered protein backbone generation and enhanced sampling
Authors: Zhu Junjie , Li Zhengxin , Tong Haowei , Lu Zhouyu , Zhang Ningjie , Wei Ting , Chen Hai-Feng
Published: November 2023

The biological function of proteins is determined not only by their static structures but also by the dynamic properties of their conformational ensembles. Numerous high-accuracy static structure prediction tools have been recently developed based on deep learning; however, there remains a lack of efficient and accurate methods for exploring protein dynamic conformations. Traditionally, studies concerning protein dynamics have relied on molecular dynamics (MD) simulations, which incur significant computational costs for all-atom precision and struggle to adequately sample conformational spaces with high energy barriers. To overcome these limitations, various enhanced sampling techniques have been developed to accelerate sampling in MD. Traditional enhanced sampling approaches like replica exchange molecular dynamics (REMD) and frontier expansion sampling (FEXS) often follow the MD simulation approach and still cost a lot of computational resources and time. Variational autoencoders (VAEs), as a classic deep generative model, are not restricted by potential energy landscapes and can explore conformational spaces more efficiently than traditional methods. However, VAEs often face challenges in generating reasonable conformations for complex proteins, especially intrinsically disordered proteins (IDPs), which limits their application as an enhanced sampling method. In this study, we presented a novel deep learning model (named Phanto-IDP) that utilizes a graph-based encoder to extract protein features and a transformer-based decoder combined with variational sampling to generate highly accurate protein backbones. Ten IDPs and four structured proteins were used to evaluate the sampling ability of Phanto-IDP. The results demonstrate that Phanto-IDP has high fidelity and diversity in the generated conformation ensembles, making it a suitable tool for enhancing the efficiency of MD simulation, generating broader protein conformational space and a continuous protein transition path.
Possible ‘silver-spoon’ effect on adult terrestrial jumping in the mangrove rivulus fish (Kryptolebias marmoratus) (Cyprinodontiformes: Rivulidae)
Authors: Styga Joseph M , Brown Isobel G , McMullen Margaret V
Published: November 2023

Survival in complex environments is dependent on whole-animal performance, a target of natural and sexual selection. Variation in whole-animal performance among mature individuals can become broadened by phenotypic plasticity driven by early life experiences. Thus, detailing the environmental factors that contribute to phenotypic plasticity, and the associated phenotypic responses governed by those environmental factors, is essential if we are to understand the evolution of whole-animal performance. Here, using isogenic lines established from the hermaphroditic mangrove rivulus fish (Kryptolebias marmoratus) housed in a ‘common garden’, we investigate the link between variation in adult terrestrial jumping performance and early life experience (i.e. egg density). First, we hypothesize that individuals from different isogenic lineages should vary in jumping performance independent of egg density, suggesting a strict genetic source of variation. Second, we hypothesize that phenotypic plasticity should also introduce variation in adult jumping performance. Specifically, we expect that hatchlings incubated at higher egg densities in early development should adopt better jumping performances as adults, because higher egg density might be a cue for greater intraspecific competition upon hatching.
Genetic and morphological differentiation within Euphorbia japygica (Euphorbiaceae) suggests divergence of populations from the south-eastern Apennine Peninsula
Authors: Boschin Micol , Schönswetter Peter , Frajman Božo
Published: November 2023

The Mediterranean Basin is a hotspot of animal and plant diversity. Contrary to the Balkan and Iberian Peninsulas that were subject of many phylogeographic studies in past decades, the Apennine Peninsula and, in particular, the diversification of its flora has been neglected in contemporary phylogenetic studies. The few available studies showed a complex pattern of north–south differentiation of genetic diversity in Italy, both among the Alps, the Apuan Alps, and the Apennines, as well as within the Apennines. Here, we explore phylogeographic patterns within recently described Euphorbia adriatica, distributed in the central and northern Apennine Peninsula, the southern margin of the Eastern Alps and the north-western Balkan Peninsula, and its relationship to the southern Italian E. japygica. Our integrative approach using nuclear ITS sequences, amplified fragment length polymorphisms, relative genome size estimation combined with chromosome counting, as well as multivariate morphometrics inferred a weak genetic differentiation that only partly corresponds to the morphological differentiation. Whereas all southern populations have hairy capsules characteristic for E. japygica, only those in south-eastern Italy (Puglia and Basilicata) are genetically divergent. There are, however, additional morphological characters that differentiate them from other populations. Our data are thus in favour of recognizing a single species, E. japygica, which includes E. adriatica that should be treated as a subspecies, named E. japygica subsp. prostrata. We provide a revised taxonomic treatment for E. japygica, including the typification. Our study shows the necessity of further in-depth investigations of diversification of Italian biota.