Evaluation of a method to fluorescently label platelets for in-human recovery and survival studies.

BACKGROUND AND OBJECTIVES: Platelets for transfusion are evaluated for in vivo quality using recovery and survival measurements in healthy human subjects. Radiolabelling is the standard for tracing platelets post-transfusion but imposes logistical and technical limitations. This study investigates the in vitro feasibility of labelling platelets with the calcein family of fluorescent dyes as an alternative to radioisotopes or biotin. MATERIALS AND METHODS: Protocols for radiolabelling were adapted for use with calcein acetoxymethyl ester (CAM) and biotin. Labelled platelets were analysed by flow cytometry and evaluated for activation and function. We tested feasibility for labelling without manipulation of platelets and for multiplexing of samples. RESULTS: Labelling at 2 µg CAM/1010 platelets resulted in >99% of CAM+ platelets. There was no significant difference in activation or aggregation between CAM-labelled or biotinylated platelets and vehicle controls although %CD62P+ was significantly lower in platelets that were not processed for labelling. Addition of CAM to the platelet storage bag labelled >95% of platelets. Platelet populations labelled with different dyes could be distinguished by flow cytometry. CONCLUSION: These data provide a rationale for further development of CAM and other fluorescent dyes as tools for measuring post-transfusion kinetics of platelets.

A review and analysis of outcomes in randomized clinical trials of plasma transfusion in patients with bleeding or for the prevention of bleeding: The BEST collaborative study.

BACKGROUND: Previous systematic reviews have revealed an inconsistency of outcome definitions as a major barrier in providing evidence-based guidance for the use of plasma transfusion to prevent or treat bleeding. We reviewed and analyzed outcomes in randomized controlled trials (RCTs) to provide a methodology for describing and classifying outcomes. STUDY DESIGN AND METHODS: RCTs involving transfusion of plasma published after 2000 were identified from a prior review (Yang 2012) and combined with an updated systematic literature search of multiple databases (July 1, 2011 to January 17, 2023). Inclusion of publications, data extraction, and risk of bias assessments were performed in duplicate. (PROSPERO registration number is: CRD42020158581). RESULTS: In total, 5579 citations were identified in the new systematic search and 22 were included. Six additional trials were identified from the previous review, resulting in a total of 28 trials: 23 therapeutic and five prophylactic studies. An increasing number of studies in the setting of major bleeding such as in cardiovascular surgery and trauma were identified. Eighty-seven outcomes were reported with a mean of 11 (min-max. 4-32) per study. There was substantial variation in outcomes used with a preponderance of surrogate measures for clinical effect such as laboratory parameters and blood usage. CONCLUSION: There is an expanding literature on plasma transfusion to inform guidelines. However, considerable heterogeneity of reported outcomes constrains comparisons. A core outcome set should be developed for plasma transfusion studies. Standardization of outcomes will motivate better study design, facilitate comparison, and improve clinical relevance for future trials of plasma transfusion.

Risk factors for T-cell lymphopenia in frequent platelet donors: The BEST collaborative study.

BACKGROUND: Severe T-cell lymphopenia of uncertain clinical significance has been observed in frequent apheresis platelet donors. Two commonly used plateletpheresis instruments are the Trima Accel, which uses a leukoreduction system (LRS) chamber to trap leukocytes and the Fenwal Amicus, which does not use an LRS chamber. STUDY DESIGN AND METHODS: We performed an international, multicenter, observational study comparing T-cell populations in frequent platelet donors collected exclusively using the Trima instrument (n = 131) or the Amicus instrument (n = 77). Age- and sex-matched whole blood donors (n = 126) served as controls. RESULTS: CD4+ T-cell counts <200 cells/µL were found in 9.9% of frequent Trima (LRS+) platelet donors, 4.4% of frequent Amicus (LRS-) platelet donors, and 0 whole blood donors (p < .0001). CD4+ T-cell counts <200 cells/µL were only seen in platelet donors with ≥200 lifetime donations. In multivariable analysis, age, lifetime donations, and instrument (Trima vs. Amicus) were independent risk factors for lymphopenia. In 40 Trima platelet donors, a plasma rinseback procedure was routinely performed following platelet collections. No Trima platelet donors receiving plasma rinseback had a CD4+ T-cell count <200 cells/µL versus 13/91 Trima platelet donors not receiving plasma rinseback (p = .01). DISCUSSION: Recurrent bulk lymphocyte removal appears to contribute to the development of T-cell lymphopenia in frequent, long-term platelet donors. Lymphopenia is more common when an LRS chamber is used during platelet collection but can occur without an LRS chamber. Blood centers using LRS chambers can mitigate donor lymphopenia by performing plasma rinseback.

Delayed Leukoreduction of whole blood with a platelet-sparing filter to increase low-titer group O whole blood production in the United States.

BACKGROUND: Demand for low-titer Group O whole blood (LTOWB) is increasing for trauma. The whole blood (WB) platelet-sparing (WB-SP) filter enables leukoreduction (LR) while retaining platelet quantity and function; however, in the United States WB must be filtered and placed in the cold within 8 h of collection. A longer processing window would facilitate improved logistics and supply of LR-WB to meet the growing medical need. This study evaluated the impact of increasing filtration timing from <8 h to <12 h on the quality of LR-WB. STUDY DESIGN AND METHODS: Thirty WB units were collected from healthy donors. Control units were filtered within 8 h and test units within 12 h of collection. WB was tested throughout 21 days of storage. Hemolysis, WBC content, component recovery, and 25 additional markers of WB quality were tested including hematologic and metabolic markers, RBC morphology, aggregometry, thromboelastography, and p-selectin. RESULTS: There were 0 failures for residual WBC content, hemolysis, or pH, and no differences in component recovery between arms. Few differences in metabolic parameters were observed, but the small effect size suggests these are not clinically significant. Trends throughout storage were similar and filtration timing did not impact hematological parameters, platelet activation and aggregation, or hemostatic capacity. CONCLUSION: Our studies showed that extending filtration timing from 8 to 12 h from the collection does not significantly impact the quality of LR-WB. Characterization of the platelets demonstrated that storage lesions were not exacerbated. Extending the time from collection to filtration will improve LTOWB inventory in the United States.

Reduction of anti-A and anti-B isoagglutinin titers of group O whole blood units employing an ABO antibody immune adsorption column.

BACKGROUND: Massive hemorrhage is a leading cause of death from trauma. There is growing interest in group O whole blood transfusions to mitigate coagulopathy and hemorrhagic shock. Insufficient availability of low-titer group O whole blood is a barrier to routine use. We tested the efficacy of the Glycosorb® ABO immunoadsorption column to reduce anti-A/B titers in group O whole blood. METHODS: Six group O whole blood units were collected from healthy volunteers, and centrifuged to separate platelet poor plasma. Platelet-poor plasma was filtered through a Glycosorb® ABO antibody immunoabsorption column, then reconstituted to prepare post-filtration whole blood. Anti-A/B titers, CBC, free hemoglobin, and thromboelastography (TEG) assays were performed on pre-and post-filtration whole blood. RESULTS: Mean( ± SEM) anti-A (224 ± 65 pre vs 13 ± 4 post) and anti-B (138 ± 38 pre vs 11 ± 4 post) titers were significantly reduced (p = 0.004) in post-filtration whole blood. No significant changes were detected in CBC, free hemoglobin, and TEG parameters on day 0. Free hemoglobin increased throughout storage (48 mg/dl ± 24 Day 0 vs 73 ± 35 Day 7 vs 96 ± 44 Day 14; p = 0.14). CONCLUSIONS: The Glycosorb® ABO column can significantly reduce anti-A/B isoagglutinin titers of group O whole blood units. Glycosorb® ABO could be employed to provide whole blood with lower risk of hemolysis and other consequences of infusing ABO incompatible plasma. Preparation of group O whole blood with substantially reduced anti-A/B would also increase the supply of low-titer group O whole blood for transfusion.

Cryoprecipitated anti-hemophiliac factor manufactured from plasma frozen within 24 h after phlebotomy meets AABB quality standards.

BACKGROUND: Cryoprecipitated antihemophiliac factor (CryoAHF) manufacturing in the US has not kept pace with the increasing demand for hospital transfusion services. Association for Advancement of Blood and Biologics (AABB) and Food and Drug Administration (FDA) require that CryoAHF be manufactured from fresh frozen plasma within 8 h (FFP). We evaluated whether CryoAHF manufactured from plasma frozen within 24 h (PF24) met regulatory quality control (QC) requirements to increase available plasma for CryoAHF. STUDY DESIGN AND METHODS: In a "worst-case scenario" feasibility study, we produced 21 single units of CryoAHF from type-O whole blood-derived PF24 frozen between 20 and 24 h after collection. A follow-up QC validation was conducted wherein 69 PF24 units across three sites were manufactured into CryoAHF. Factor VIII (FVIII) and fibrinogen levels were measured. RESULTS: CryoAHF manufactured in our feasibility study from PF24 contained FVIII levels of 208 ± 61 IU (mean ± SD) and 509 ± 152 mg of fibrinogen levels per unit. CryoAHF manufactured in our QC validation from PF24 yielded FVIII levels of 214 ± 58 IU and 607 ± 176 mg of fibrinogen levels per unit. The coagulation factor levels from each of the individual CryoAHF units exceeded the minimum AABB and FDA requirement of ≥80 IU of FVIII per unit and ≥150 mg of fibrinogen per unit. There was no decrease in FVIII or fibrinogen levels in CryoAHF produced from PF24 as compared to historic QC results of CryoAHF produced from FFP. CONCLUSION: These studies demonstrated that CryoAHF produced from PF24 meets AABB and FDA QC requirements. FDA approved the American Red Cross request to manufacture CryoAHF singles and pools from PF24 as source material.

Lessons learned from the use of convalescent plasma for the treatment of COVID-19 and specific considerations for immunocompromised patients.

Coronavirus disease 2019 (COVID-19) convalescent plasma (CovCP) infusions have been widely used for the treatment of hospitalized patients with COVID-19. The aims of this narrative review were to analyze the safety and efficacy of CovCP infusions in the overall population and in immunocompromised patients with COVID-19 and to identify the lessons learned concerning the use of convalescent plasma (CP) to fill treatment gaps for emerging viruses. Systematic searches (PubMed, Scopus, and COVID-19 Research) were conducted to identify peer-reviewed articles and pre-prints published between March 1, 2020 and May 1, 2021 on the use of CovCP for the treatment of patients with COVID-19. From 261 retrieved articles, 37 articles reporting robust controlled studies in the overall population of patients with COVID-19 and 9 articles in immunocompromised patients with COVID-19 were selected. While CovCP infusions are well tolerated in both populations, they do not seem to improve clinical outcomes in critically-ill patients with COVID-19 and no conclusion could be drawn concerning their potential benefits in immunocompromised patients with COVID-19. To be better prepared for future epidemics/pandemics and to evaluate potential benefits of CP treatment, only CP units with high neutralizing antibodies (NAbs) titers should be infused in patients with low NAb titers, patient eligibility criteria should be based on the disease pathophysiology, and measured clinical outcomes and methods should be comparable across studies. Even if CovCP infusions did not improve clinical outcomes in patients with COVID-19, NAb-containing CP infusions remain a safe, widely available and potentially beneficial treatment option for future epidemics/pandemics.

Time from apheresis platelet donation to cold storage: Evaluation of platelet quality and bacterial growth.

BACKGROUND: Cold storage reduces posttransfusion survival of platelets; however, it can improve platelet activation, lower risk of bacterial contamination, and extend shelf-life compared to room temperature (RT) storage. To facilitate large-scale availability, manufacturing process optimization is needed, including understanding the impact of variables on platelet potency and safety. Short time requirements from collection to storage is challenging for large blood centers to complete resuspension and qualify platelets for production. This study evaluated the impact of time from platelet component collection to cold storage on in vitro properties and bacterial growth. STUDY DESIGN AND METHODS: Double-apheresis platelet components were collected from healthy donors, suspended in 65% PAS-III/35% plasma, and split into 2 equal units. One unit was placed into cold storage within 2 h and the other unit after 8 h. Eight matched pairs were evaluated for 12 in vitro parameters. Twenty-four matched pairs were evaluated with 8 bacterial strains tested in triplicate. Samples were tested throughout 21 days of storage. RESULTS: In vitro properties were not different between 2 and 8 h units, and trends throughout storage were similar between arms. Time to cold storage did not significantly impact bacterial growth, with <1 log10 difference at all timepoints between units. DISCUSSION: Our studies showed that extending time to cold storage from 2 to 8 h from collection did not significantly increase the bacterial growth, and the platelet component quality and function is maintained. The ability to extend the time required from collection to storage will improve blood center logistics to feasibly produce CSPs.

Systematic literature review evaluating evidence and mechanisms of action for platelet-rich plasma as an antibacterial agent.

Platelet rich plasma or PRP is a supraphysiologic concentrate of platelets derived by centrifugation and separation of whole blood components. Along with platelets and plasma, PRP contains various cell types including white blood cells (WBC)/leukocytes, both granulocytes (neutrophils, basophils, eosinophils) and agranulocytes (monocytes, lymphocytes). Researchers and clinicians have explored the application of PRP in wound healing and prevention of surgical wound infections, such as deep sternal wounds. We conducted this systematic literature review to evaluate the preclinical and clinical evidence for the antibacterial effect of PRP and its potential mechanism of action. 526 records were identified for screening. 34 unique articles were identified to be included in this literature review for data summary. Overall, the quality of the clinical trials in this review is low, and collectively qualify as Oxford level C. Based on the available clinical data, there is a clear trend towards safety of autologous PRP and potential efficacy in deep sternal wound management. The preclinical and bench data is very compelling. The application of PRP in treatment of wounds or prevention of infection with PRP is promising but there is a need for foundational bench and preclinical animal research to optimize PRP as an antibacterial agent, and to provide data to aid in the design and conduct of well-designed RCTs with adequate power to confirm antimicrobial efficacy of PRP in specific disease states and wound types.

Treatment for emerging viruses: Convalescent plasma and COVID-19.

Use of convalescent plasma transfusions could be of great value in the current pandemic of coronavirus disease (COVID-19), given the lack of specific preventative and therapeutic options. This convalescent plasma therapy is of particular interest when a vaccine or specific therapy is not yet available for emerging viruses, such as severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), which causes COVID-19. This report summarizes existing literature around convalescent plasma as a therapeutic option for COVID-19. It also includes recommendations for establishing a convalescent plasma program, enhancement considerations for convalescent plasma, and considerations around pathogen reduction treatment of convalescent plasma. Time is of the essence to set up protocols for collection, preparation, and administration of apheresis-collected convalescent plasma in response to the current pandemic. The immediate use of convalescent plasma provides prompt availability of a promising treatment while specific vaccines and treatments are evaluated and brought to scale. Further development of improved convalescent plasma, vaccines and other therapeutics depends on quick generation of additional data on pathogenesis and immune response. Additionally, given the lack of information around the natural history of this disease, PRT should be considered to add a layer of safety to protect recipients of convalescent plasma.

Application of monoclonal antibodies in quantifying fungal growth dynamics during aerobic spoilage of silage.

Proliferation of filamentous fungi following ingress of oxygen to silage is an important cause of dry matter losses, resulting in significant waste. In addition, the production of mycotoxins by some filamentous fungi poses a risk to animal health through mycotoxicosis. Quantitative assessment of fungal growth in silage, through measurement of ergosterol content, colony-forming units or temperature increase is limiting in representing fungal growth dynamics during aerobic spoilage due to being deficient in either representing fungal biomass or being able to identify specific genera. Here, we conducted a controlled environment aerobic exposure experiment to test the efficacy of a monoclonal antibody-based enzyme-linked immunosorbent assay (ELISA) to detect the proliferation of fungal biomass in six silage samples. We compared this to temperature which has been traditionally deployed in such experiments and on-farm to detect aerobic deterioration. In addition, we quantified ergosterol, a second marker of fungal biomass. After 8 days post-aerobic exposure, the ergosterol and ELISA methods indicated an increase in fungal biomass in one of the samples with a temperature increase observed after 16 days. A comparison of the methods with Pearson's correlation coefficient showed a positive association between temperature and ergosterol and both markers of fungal biomass. This work indicates that the technology has potential to be used as an indicator of microbial degradation in preserved forage. Consequently, if it developed as an on-farm technique, this could inform forage management decisions made by farmers, with the goal of decreasing dry matter losses, improving resource and nutrient efficiency and reducing risks to animal health.

Exposure to Violence and Posttreatment Substance Use Among Patients With Opioid Use Disorder.

Exposure to violence can lead to a dramatic increase in the likelihood of the development of a substance use disorder (SUD). Given the overlap between the two, substance use for survivors of violence, then, can be a coping mechanism to manage the traumatic effects of abuse and persistent use can evolve into a diagnosable SUD. This study was designed to examine the posttreatment substance use among adults who have a history of exposure to violence and sought treatment for opioid use disorder. Data for this study were drawn from the Comprehensive Addiction Treatment Outcome Research system. Among the 13,105 patients included in the study, 444 (3.4%) received a formal diagnosis for opioid use disorder. Female victims of violence are at a greater risk of suffering injuries related to violence, resulting in increased levels of medical care utilization, which may prompt the initiation and prolonged use of prescription pain relief medication. Related to this important finding is another indicating that exposure to violence at multiple points in the past was associated with more severe indicators of substance use. These data show that there is a relationship between exposure to violence, SUDs, and relapse among patients seeking treatment. Not only must patients and treatment providers address these past violent experiences as important psychological factors in recovery, but in the context of opioid use disorder, physical injuries contributing to chronic pain may also trigger persistent substance use.

Infectious causes and outcomes in patients presenting with cerebral spinal fluid pleocytosis.

To evaluate the infectious etiologies, clinical features, and outcomes of patients with CNS infections at a tertiary care center. Patients that present with a pleocytosis in the cerebral spinal fluid (CSF), defined as a CSF WBC count > 5 cells/mm3, from July 2015 to June 2016 at a tertiary care hospital were analyzed for this report. Data from patients with confirmed (n = 43) and presumed (n = 51) CNS infections were analyzed. CNS infection was the leading known cause of CSF pleocytosis (n = 43, 18% of all patients with a pleocytosis in the CSF), and HSV-2 was identified as the leading causative pathogen (n = 10) followed by varicella zoster virus (n = 5). Fifty-three percent of patients with a pleocytosis in the CSF did not receive a diagnosis. In the patients that did not receive a diagnosis, CNS infection was presumed to be the cause in 51 patients (21% of patients with CSF pleocytosis). The mean time to diagnosis for patients with confirmed CNS infection was 16 days, but time to diagnosis was highly variable depending on the causative pathogen. There was a significant overlap in CSF parameters and peripheral white blood cell counts in patients diagnosed with a viral, bacterial, or fungal infection. Neuroimaging changes were present in only 44% of CNS infections. The overall mortality was 7% for CNS infections, and 17% of patients with a CNS infection had a severe neurologic deficit at presentation while only 3% had a severe deficit at the last neurologic assessment. This study provides new insights into the infectious causes of disease in a cohort of patients with pleocytosis in the CSF. The study provides new insights into the time to diagnosis and outcomes in patients that present with pleocytosis in the CSF.

Meta-analysis of the visuospatial aftereffects of prism adaptation, with two novel experiments.

We present a meta-analysis of the effects of visuomotor adaptation to leftward displacing prisms on visuospatial judgements in healthy people, as assessed by perceptual (landmark) and manual versions of the line bisection task. To supplement previously published datasets, we report two novel experiments: Experiment 1 (n = 12) found null effects of adaptation to 10° leftward prisms on spatial bias in the landmark task, and Experiment 2 (n = 24) found null effects of 12° leftward prisms on spatial bias in a computerised line bisection task. Including these data, we considered 17 experiments for the landmark task (total n = 256), and 12 experiments for line bisection (total n = 172), in which participants were adapted for between 7 and 20 min to prism strengths from 8 to 17°. A random-effects meta-analysis, with prism strength and exposure duration as moderators, confirmed robust rightward shifts in visuospatial judgements following leftward prism adaptation. The average standardised effect sizes (Cohen's d) were similar between tasks, increasing by around .1 per degree of prismatic displacement, and being boosted by a long (10 min +) period of prism exposure. However, the quality of evidence and precision of prediction was superior for the landmark task, with a higher signal-to-noise ratio within studies, and less heterogeneity between studies. We suggest that line bisection responses may be contaminated by sensorimotor aftereffects, and that the landmark task is a more suitable method for measuring true visuospatial aftereffects of prism adaptation. To harness these effects, we recommend that researchers should expose participants to 15° (or higher) leftward prisms for more than ten minutes, with upwards of 250 pointing movements. Power calculations should take account of heterogeneity in the true effect size between studies; and further investigation of the factors underlying this heterogeneity will help to refine optimally-effective methods.

Defining diagnostic approaches and outcomes in patients with inflammatory CSF: A retrospective cohort study.

OBJECTIVE: Define the etiologies, clinical features, time to diagnosis, and outcomes of patients that present with cerebral spinal fluid (CSF) pleocytosis. PATIENTS AND METHODS: This is retrospective cohort study of patients with CSF pleocytosis, defined as WBC count >5 cells/mm3 in the CSF, from July 2015 to June 2016 at a large tertiary care hospital. The proportion of patients within specific diagnostic categories were analyzed for differences in diagnostic testing and outcomes. RESULTS: 53% of patients had CSF pleocytosis due to an unknown etiology. The leading known cause of neuroinflammation was CNS infection (n = 43/244, 18%), followed by malignancy (n = 28/244, 11%). Mean time to diagnosis was 125 days in patients with autoimmune neuroinflammation and was 16 days in patients with an infection or malignancy. CSF parameters and peripheral white blood cell counts did not distinguish between categories of disease. The presence of CSF oligoclonal bands or a positive biopsy result most commonly supported a diagnosis of an autoimmune disease or malignancy, respectively. Neuroimaging changes were present in only 44% of infections but were found in 80-90% of other categories of neuroinflammation. Patients presenting with a severe neurologic deficit had 22.29 higher odds of a severe deficit at the last neurologic assessment, and mortality was highest (29%) in patients with malignancy-associated neuroinflammation. CONCLUSIONS: This study to defines general diagnostic categories of neuroinflammatory disease in patients and provides new insight on the value of specific diagnostic testing, time to diagnosis, and outcomes in these patient populations.

Self-perceived uselessness is associated with lower likelihood of successful aging among older adults in China.

BACKGROUND: Plenty of evidence has shown that self-perceived uselessness among older adults is negatively associated with successful aging in terms of good health in Western societies. It is unclear whether these findings are valid in China where living into older age is more selective due to high mortality at younger ages. METHODS: Using five waves (2000, 2002, 2005, 2008/2009 and 2011/2012) of a large nationally representative survey in China with 29,954 observations from 19,070 older adults aged 65 and older, this study aimed to investigate the association between self-perceived uselessness and successful aging. Self-perceived uselessness was measured by a single item "with age, do you feel more useless?" with six answers: always, often, sometimes, seldom, never, and unable to answer. Successful aging was measured by independence in activities of daily living (ADL), independence in instrumental activities of daily living (IADL), unimpaired cognition, good life satisfaction, and good self-rated health. Logistic regression models were applied to each successful aging indicator after controlling for a rich set of covariates that included demographics, socioeconomic status, family/social support, and health practices. The models also adjusted for intraperson correlations across waves. RESULTS: We found that self-perceived uselessness was negatively associated with successful aging among older adults aged 65 or older. Specifically, compared to never having self-perceived uselessness, always having such a perception was associated with 16-42 % lower odds of being ADL independent, IADL independent, cognitively unimpaired, and having good life satisfaction and good self-rated health. Often or sometimes having such a perception also reduced odds of aging successfully, although such reductions were less pronounced. The associations were similar among the oldest-old aged 80 or older with one exception for the case of IADL independence. CONCLUSIONS: Self-perceived uselessness is negatively associated with successful aging among Chinese older adults as well as among the oldest-old. Our findings could be informative for China in the development of public health programs that aim to improve self-perceptions about aging and promote successful aging.

Analytical Validation of the ReEBOV Antigen Rapid Test for Point-of-Care Diagnosis of Ebola Virus Infection.

BACKGROUND: Ebola virus disease (EVD) is a severe viral illness caused by Ebola virus (EBOV). The 2013-2016 EVD outbreak in West Africa is the largest recorded, with >11 000 deaths. Development of the ReEBOV Antigen Rapid Test (ReEBOV RDT) was expedited to provide a point-of-care test for suspected EVD cases. METHODS: Recombinant EBOV viral protein 40 antigen was used to derive polyclonal antibodies for RDT and enzyme-linked immunosorbent assay development. ReEBOV RDT limits of detection (LOD), specificity, and interference were analytically validated on the basis of Food and Drug Administration (FDA) guidance. RESULTS: The ReEBOV RDT specificity estimate was 95% for donor serum panels and 97% for donor whole-blood specimens. The RDT demonstrated sensitivity to 3 species of Ebolavirus (Zaire ebolavirus, Sudan ebolavirus, and Bundibugyo ebolavirus) associated with human disease, with no cross-reactivity by pathogens associated with non-EBOV febrile illness, including malaria parasites. Interference testing exhibited no reactivity by medications in common use. The LOD for antigen was 4.7 ng/test in serum and 9.4 ng/test in whole blood. Quantitative reverse transcription-polymerase chain reaction testing of nonhuman primate samples determined the range to be equivalent to 3.0 × 105-9.0 × 108 genomes/mL. CONCLUSIONS: The analytical validation presented here contributed to the ReEBOV RDT being the first antigen-based assay to receive FDA and World Health Organization emergency use authorization for this EVD outbreak, in February 2015.

Chromosomal rearrangements in holocentric organisms lead to reproductive isolation by hybrid dysfunction: The correlation between karyotype rearrangements and germination rates in sedges.

PREMISE OF THE STUDY: Understanding the drivers of speciation is a central task of evolutionary biology. Chromosomal rearrangements are known to play an important role in species diversification, but the role of rearrangements of holocentric chromosomes-chromosomes without localized centromeres-is poorly understood. METHODS: We made numerous artificial crosses between Carex scoparia individuals of different diploid chromosome numbers and, for comparison, between individuals of the same chromosome number. We studied chromosome pairing and chromosomal rearrangements in the F1 individuals using light microscopy. We then estimated germination rates as a function of geographic distance, genetic distance, chromosome number differences in parents, and pairing irregularities in F1 individuals, using generalized least squares to fit alternative regression models. KEY RESULTS: The most informative predictors of germination rates in the F1 generation are chromosome number differences and minimum number of chromosome pairing irregularities in the F1 individuals. Genetic and geographic distances between parents are not significant predictors. CONCLUSIONS: Holocentric chromosomal rearrangements play an important role in postzygotic reproductive isolation in Carex through F1 hybrid inviability and sterility. Hybrid dysfunction seems to be a suitable model for chromosomal speciation when there are several chromosomal rearrangements between parents. However, we have not tested the hypothesis that genome rearrangements may also play an important role in suppressing recombination between cytogenetically divergent populations.