The massive 340 megabase genome of Anisogramma anomala, a biotrophic ascomycete that causes eastern filbert blight of hazelnut.

BACKGROUND: The ascomycete fungus Anisogramma anomala causes Eastern Filbert Blight (EFB) on hazelnut (Corylus spp.) trees. It is a minor disease on its native host, the American hazelnut (C. americana), but is highly destructive on the commercially important European hazelnut (C. avellana). In North America, EFB has historically limited commercial production of hazelnut to west of the Rocky Mountains. A. anomala is an obligately biotrophic fungus that has not been grown in continuous culture, rendering its study challenging. There is a 15-month latency before symptoms appear on infected hazelnut trees, and only a sexual reproductive stage has been observed. Here we report the sequencing, annotation, and characterization of its genome. RESULTS: The genome of A. anomala was assembled into 108 scaffolds totaling 342,498,352 nt with a GC content of 34.46%. Scaffold N50 was 33.3 Mb and L50 was 5. Nineteen scaffolds with lengths over 1 Mb constituted 99% of the assembly. Telomere sequences were identified on both ends of two scaffolds and on one end of another 10 scaffolds. Flow cytometry estimated the genome size of A. anomala at 370 Mb. The genome exhibits two-speed evolution, with 93% of the assembly as AT-rich regions (32.9% GC) and the other 7% as GC-rich (57.1% GC). The AT-rich regions consist predominantly of repeats with low gene content, while 90% of predicted protein coding genes were identified in GC-rich regions. Copia-like retrotransposons accounted for more than half of the genome. Evidence of repeat-induced point mutation (RIP) was identified throughout the AT-rich regions, and two copies of the rid gene and one of dim-2, the key genes in the RIP mutation pathway, were identified in the genome. Consistent with its homothallic sexual reproduction cycle, both MAT1-1 and MAT1-2 idiomorphs were found. We identified a large suite of genes likely involved in pathogenicity, including 614 carbohydrate active enzymes, 762 secreted proteins and 165 effectors. CONCLUSIONS: This study reveals the genomic structure, composition, and putative gene function of the important pathogen A. anomala. It provides insight into the molecular basis of the pathogen's life cycle and a solid foundation for studying EFB.

First record of Ixodes keiransi (Acari: Ixodidae) in New Jersey, USA.

The hard tick, Ixodes keiransi Beati, Nava, Venzal, & Guglielmone, formerly the North American lineage of Ixodes affinis Neumann, is expanding its range northward along the US East Coast. In July 2023, we collected I. keiransi adult female and nymph in a single sampling event, suggesting its range now includes southern New Jersey. In this area, I. keiransi is sympatric with northern populations of Ixodes scapularis Say (Acari: Ixodidae), the primary vector of Lyme disease. Given its status as an enzootic vector of spirochaetes in the Borrelia burgdorferi sensu lato complex, proper differentiation of these 2 species will be critical for accurate estimates of entomological risk. Targeted surveillance should be implemented to monitor further I. keiransi expansion and to elucidate the phenology and enzootic role of this and other understudied Ixodes spp. in the northeastern United States.

Polycyclic aromatic hydrocarbon (PAH) source identification and a maternal transfer case study in threatened killer whales (Orcinus orca) of British Columbia, Canada.

The northeastern Pacific (NEP) Ocean spans the coast of British Columbia (Canada) and is impacted by anthropogenic activities including oil pipeline developments, maritime fossil fuel tanker traffic, industrial chemical effluents, agricultural and urban emissions in tandem with stormwater and wastewater discharges, and forest wildfires. Such events may expose surrounding marine environments to toxic polycyclic aromatic hydrocarbons (PAHs) and impact critical habitats of threatened killer whales (Orcinus orca). We analyzed skeletal muscle and liver samples from stranded Bigg's killer whales and endangered Southern Resident killer whales (SRKWs) for PAH contamination using LRMS. C3-phenanthrenes/anthracenes (mean: 632 ng/g lw), C4-dibenzothiophenes (mean: 334 ng/g lw), and C4-phenanthrenes/anthracenes (mean: 248 ng/g lw) presented the highest concentrations across all tissue samples. Diagnostic ratios indicated petrogenic-sourced contamination for SRKWs and pyrogenic-sourced burdens for Bigg's killer whales; differences between ecotypes may be attributed to habitat range, prey selection, and metabolism. A mother-fetus skeletal muscle pair provided evidence of PAH maternal transfer; low molecular weight compounds C3-fluorenes, dibenzothiophene, and naphthalene showed efficient and preferential exposure to the fetus. This indicates in-utero exposure of PAH-contamination to the fetus. Our results show that hydrocarbon-related anthropogenic activities are negatively impacting these top predators; preliminary data found here can be used to improve oil spill and other PAH pollution management and regulation efforts, and inform policy to conserve killer whale habitats in the NEP.

Endogenous Viral Elements in Ixodid Tick Genomes.

The documentation of endogenous viral elements (EVEs; virus-derived genetic material integrated into the genome of a nonviral host) has offered insights into how arthropods respond to viral infection via RNA interference pathways. Small non-coding RNAs derived from EVE loci serve to direct RNAi pathways in limiting replication and infection from cognate viruses, thus benefiting the host's fitness and, potentially, vectorial capacity. Here we use informatic approaches to analyze nine available genome sequences of hard ticks (Acari: Ixodidae; Rhipicephalus sanguineus, R. microplus, R. annulatus, Ixodes ricinus, I. persulcatus, I. scapularis, Hyalomma asiaticum, Haemaphysalis longicornis, and Dermacentor silvarum) to identify endogenous viral elements and to illustrate the shared ancestry of all elements identified. Our results highlight a broad diversity of viral taxa as having given rise to 1234 identified EVEs in ticks, with Mononegavirales (specifically Rhabdoviridae) well-represented in this subset of hard ticks. Further investigation revealed extensive adintovirus integrations in several Ixodes species, the prevalence of Bunyavirales EVEs (notably not observed in mosquitoes), and the presence of several elements similar to known emerging human and veterinary pathogens. These results will inform subsequent work on current and past associations with tick species with regard to the viruses from which their "viral fossils" are derived and may serve as a reference for quality control of various tick-omics data that may suffer from misidentification of EVEs as viral genetic material.

The development, content and response process validation of a caregiver-reported severity measure for CDKL5 deficiency disorder.

BACKGROUND: CDKL5 Deficiency Disorder (CDD) is a severe X-linked developmental and epileptic encephalopathy. Existing developmental outcome measures have floor effects and cannot capture incremental changes in symptoms. We modified the caregiver portion of a CDD clinical severity assessment (CCSA) and assessed content and response-process validity. METHODS: We conducted cognitive interviews with 15 parent caregivers of 1-39-year-old children with CDD. Caregivers discussed their understanding and concerns regarding appropriateness of both questions and answer options. Item wording and questionnaire structure were adjusted iteratively to ensure questions were understood as intended. RESULTS: The CCSA was refined during three rounds of cognitive interviews into two measures: (1) the CDD Developmental Questionnaire - Caregiver (CDQ-Caregiver) focused on developmental skills, and (2) the CDD Clinical Severity Assessment - Caregiver (CCSA-Caregiver) focused on symptom severity. Branching logic was used to ensure questions were age and skill appropriate. Initial pilot data (n = 11) suggested no floor effects. CONCLUSIONS: This study modified the caregiver portion of the initial CCSA and provided evidence for its content and response process validity.

Detection of multiple tick-borne pathogens in Ixodes scapularis from Hunterdon County, NJ, USA.

Several human pathogens vectored by the blacklegged tick (Ixodes scapularis Say; Acari: Ixodidae) are endemic in the state of New Jersey. Disease incidence data suggest that these conditions occur disproportionately in the northwestern portion of the state, including in the county of Hunterdon. We conducted active surveillance at three forested sites in Hunterdon County during 2020 and 2021, collecting 662 nymphal and adult I. scapularis. Ticks were tested for five pathogens by qPCR/qRT-PCR: Anaplasma phagocytophilum, Babesia microti, Borrelia burgdorferi, Borrelia miyamotoi, and Powassan virus (POWV) lineage 2. Over 2 years, 25.4% of nymphs and 58.4% of adults were found infected with at least one pathogen, with 10.6% of all ticks infected with more than one pathogen. We report substantial spatial and temporal variability of A. phagocytophilum and B. burgdorferi, with high relative abundance of the human-infective A. phagocytophilum variant Ap-ha. Notably, POWV was detected for the first time in Hunterdon, a county where human cases have not been reported. Based on comparisons with active surveillance initiatives in nearby counties, further investigation of non-entomological factors potentially influencing rates of tick-borne illness in Hunterdon is recommended.

Lone star ticks (Acari: Ixodidae) infected with Bourbon virus in New Jersey, USA.

Lone star ticks (Amblyomma americanum L.) are expanding within the northeast United States, a region historically focused on Ixodes scapularis-transmitted diseases. In Monmouth County, NJ, the shift has been dramatic, and lone star ticks now vastly outnumber blacklegged ticks. As a result, there is an enhanced need to focus on the potential health risks of A. americanum-transmitted pathogens, such as the emerging Heartland (HRTV) and Bourbon (BRBV) viruses. We screened 1,205 nymphal lone star ticks for HRTV and BRBV using RT-qPCR assays and detected BRBV in 3 ticks collected in Monmouth County, NJ, in 2021. Additionally, we sequenced a complete BRBV genome from a single infected specimen, finding 99.4% identity with human pathogenic isolates from the eastern-central United States. Our results have important public health implications for a region only recently becoming aware of public health risks posed by lone star ticks. Of note, we report successful detection of viral RNA in samples that were stored and intended for DNA preservation, for example, kept in ethanol at room temperature, which may reduce barriers for public health agencies seeking to expand their tick testing to include viruses.

Alberta Collaborative Quality Improvement Strategies to Improve Outcomes of Moderate and Late Preterm Infants (ABC-QI) Trial: a protocol for a multicentre, stepped-wedge cluster randomized trial.

BACKGROUND: Evidence-based Practice for Improving Quality (EPIQ) is a collaborative quality improvement method adopted by the Canadian Neonatal Network that led to decreased mortality and morbidity in very preterm neonates. The Alberta Collaborative Quality Improvement Strategies to Improve Outcomes of Moderate and Late Preterm Infants (ABC-QI) Trial aims to evaluate the impact of EPIQ collaborative quality improvement strategies in moderate and late preterm neonates in Alberta, Canada. METHODS: In a 4-year, multicentre, stepped-wedge cluster randomized trial involving 12 neonatal intensive care units (NICUs), we will collect baseline data with the current practices in the first year (all NICUs in the control arm). Four NICUs will transition to the intervention arm at the end of each year, with 1 year of follow-up after the last group transitions to the intervention arm. Neonates born at 32 + 0 to 36 + 6 weeks' gestation with primary admission to NICUs or postpartum units will be included. The intervention includes implementation of respiratory and nutritional care bundles using EPIQ strategies, including quality improvement team building, quality improvement education, bundle implementation, quality improvement mentoring and collaborative networking. The primary outcome is length of hospital stay; secondary outcomes include health care costs and short-term clinical outcomes. Neonatal intensive care unit staff will complete a survey in the first year to assess quality improvement culture in each unit, and a sample will be interviewed 1 year after implementation in each unit to evaluate the implementation process. INTERPRETATION: The ABC-QI Trial will assess whether collaborative quality improvement strategies affect length of stay in moderate and late preterm neonates. It will provide detailed population-based data to support future research, benchmarking and quality improvement. TRIAL REGISTRATION: ClinicalTrials.gov, no. NCT05231200.

Rickettsia and relapsing fever Borrelia in Alectorobius kelleyi (Ixodida: Argasidae) from peri domestic bats in the northeastern United States.

The soft ticks (Argasidae) are known vectors of human and animal pathogens around the globe and are relatively understudied. Our aim was to assess the presence of Rickettsia and Borrelia bacteria in Alectorobius kelleyi (Argasidae) parasitizing synanthropic bats in the highly urbanized northeastern United States. By collaborating with parasitologists, bat scientists and wildlife rehabilitators we were successful in obtaining A. kelleyi from five states. Since Argasid larvae will attach to their hosts for many days, most A. kelleyi examined (92%) were larvae collected from sick or injured big brown bats, Eptesicus fuscus, undergoing care at rehabilitation centers. In addition, we obtained adult A. kelleyi captured in residential living areas and trapped in attics. An in-depth analysis of a A. kelleyi found to be infected with a spotted fever group Rickettsia (SFGR) revealed a dual infection with a R. belli-like taxon (ancestral group) as well as an SFGR closely related to R. peacockii, likely the same previously found in A. kelleyi from Iowa and Kansas. We found that 36% of the A. kelleyi tested carried the SFGR. Furthermore, we detected a relapsing fever spirochete, likely Candidatus Borrelia johnsonii, in 25% of the A. kelleyi from Pennsylvania. While it is unclear if these bacteria constitute a health risk to either bats or humans, our study indicates that human exposure to ectoparasites infesting peridomestic wildlife should be considered in the epidemiology of tick-borne diseases.

Intensive Cardiac Rehabilitation Attenuates the Gender Gap in Cardiac Rehabilitation Participation.

Traditional cardiac rehabilitation (CR) programs historically have had considerable gender gaps in CR outcomes. Intensive-Cardiac-Rehabilitation (ICR) has nonexercise components such as stress management, peer support and diet modification, in addition to exercise, which may reduce barriers to CR completion. However, there is a paucity of data in ICR outcomes based on gender. We conducted a nationwide retrospective cohort study of 15,613 patients enrolled in 46 Ornish-ICR programs across the United States from January 2016 to December 2020 and compared ICR physical and psychological outcomes in women vs men. Primary endpoints were depression scores (CESD), health status (SF-36 physical and mental composite-scores) and exercise minutes per week. Of 15,613 ICR participants who participated in the program, 6788 (44%) were women. ICR completion rates were 64.7% overall and nearly equal between men and women (63.3% women vs 65.9% men). There were a few small differences in outcomes after ICR completion: women had slightly more improvement in depression scores (women: -6.10 ± 8.79, 49.6% reduction vs men: -4.92 ± 7.80, 46.4% reduction; P < 0.01) and SF-36 physical (women: 5.95 ± 6.86 vs men: 5.28 ± 6.71, P < 0.01) and SF-36 mental (women: 5.76 ± 8.11 vs men: 4.60 ± 7.33, P < 0.01) composite scores than men. There was no significant difference in exercise-minutes per week between the 2 groups. ICR attenuates the gender gap in CR completion. Further, of the completers, women achieved slightly higher depression and quality-of-life improvements compared to men. As such, ICR can be a good option to reduce gender-based differences in CR participation.

Microhabitat modeling of the invasive Asian longhorned tick (Haemaphysalis longicornis) in New Jersey, USA.

The Asian longhorned tick (Haemaphysalis longicornis) is a vector of multiple arboviral and bacterial pathogens in its native East Asia and expanded distribution in Australasia. This species has both bisexual and parthenogenetic populations that can reach high population densities under favorable conditions. Established populations of parthenogenetic H. longicornis were detected in the eastern United States in 2017 and the possible range of this species at the continental level (North America) based on climatic conditions has been modeled. However, little is known about factors influencing the distribution of H. longicornis at geographic scales relevant to local surveillance and control. To examine the importance of local physiogeographic conditions such as geology, soil characteristics, and land cover on the distribution of H. longicornis we employed ecological niche modeling using three machine learning algorithms - Maxent, Random Forest (RF), and Generalized Boosting Method (GBM) to estimate probability of finding H. longicornis in a particular location in New Jersey (USA), based on environmental predictors. The presence of H. longicornis in New Jersey was positively associated with Piedmont physiogeographic province and two soil types - Alfisols and Inceptisols. Soil hydraulic conductivity was the most important predictor explaining H. longicornis habitat suitability, with more permeable sandy soils with higher hydraulic conductivity being less suitable than clay or loam soils. The models were projected over the state of New Jersey creating a probabilistic map of H. longicornis habitat suitability at a high spatial resolution of 90×90 meters. The model's sensitivity was 87% for locations sampled in 2017-2019 adding to the growing evidence of the importance of soil characteristics to the survival of ticks. For the 2020-2022 dataset the model fit was 57%, suggestive of spillover to less optimal habitats or, alternatively, heterogeneity in soil characteristics at the edges of broad physiographic zones. Further modeling should incorporate abundance and life-stage information as well as detailed characterization of the soil at collection sites. Once critical parameters that drive the survival and abundance of H. longicornis are identified they can be used to guide surveillance and control strategies for this invasive species.

First report of Powdery Mildew Caused by Golovinomyces ambrosiae on Industrial Hemp in New Jersey.

In December 2019, New Jersey became one of the first states to have its industrial hemp (Cannabis sativa L.) plan approved by the U.S. Department of Agriculture (USDA) following enactment of the 2018 Farm Bill that authorized the production of hemp. Following this approval, hemp was legally grown for the first time in 2020. During the growing seasons of 2020 and 2021, powdery mildew-like symptoms were observed during the summer months (Jun to Aug) in greenhouse hemp research and fall months (Aug to Oct) in field production plots on Rutgers Agricultural Experiment Station farms in southern and northern New Jersey. Symptoms were observed on leaves and stems of hemp cultivars 'CB Genius', 'Cherry Wine' and 'Bay Mist'. Symptoms initially appeared as small white patches of mycelia and conidia on the adaxial surface of leaves that gradually spread to entire leaves and stems. Leaf discoloration (e.g., chlorosis) and premature leaf drop were observed. More severe symptoms and damage were observed in the greenhouse than outdoor cultivation. A voucher specimen was deposited in the U.S National Fungus Collections, USDA-ARS, Beltsville, MD (accession number 929187). Morphological examination of the white colonies from the cultivar 'Baymist' was carried out using light microscopy and further characterized by sequencing. This isolate was labelled PMH2. Hyphae were septate, conidiophores were hyaline, unbranched, measuring 130 to 240 µm in length and produced 1 to 4 conidia in chains. Conidia were hyaline, ellipsoid to ovoid in shape and measured 20 to 36 ×10 to 18 µm (n=30). Oil-like drops were present within conidia, although no distinct fibrosin bodies were observed. Chasmothecia were not observed. Morphological observations were consistent with those of Golovinomyces spp. as described by Braun and Cook (2012). Morphological observations (conidiophore and conidial measurements) were also similar to the description of G. ambrosiae on Hemp, as described in Wiseman et al, 2021. Sequencing of internal transcribed spacer (ITS), large ribosomal subunit (28S), intergenic spacer (IGS), beta- tubulin (TUB2) and chitin synthase 1(CHS1) region, were carried out with the primer sets ITS5/ITS4, LSU1/LSU2, IGS-12a/NS1R, TubF1/TubR1 and gCS1a1/gCS1b respectively, as shown by Qiu et al. (2020). Maximum-likelihood phylogenetic analysis confirmed the grouping of the PMH2 isolate within the G. ambrosiae accessions. Each individual gene alignment was treated as a separate partition. Sequences were not concatenated for maximum -likelihood phylogenetic analysis. Sequence data were deposited in GenBank under the accessions OK626453 (ITS), OK626454 (28S), OL456201 (IGS), OL415512 (TUB2) and OL415513(CHS1). To fulfill Koch's postulates, two mature, potted plants of C. sativa cv. 'Alpha Explorer' were inoculated by gently pressing symptomatic hemp leaves onto their leaf surface. They were incubated in an indoor grow room at 23°C and relative humidity of 50%. Non-inoculated healthy plants of C. sativa cv. 'Alpha Explorer' served as control. Inoculated plants developed powdery mildew symptoms within 10 to 12 days, while all control plants were asymptomatic. The powdery mildew on inoculated plants was found to be morphologically similar to the original. G. ambrosiae has been reported on C.sativa in Oregon (Wiseman et al. 2021) and G. ambrosiae (as G. spadiceus) has been reported on Cannabis in Kentucky (Szarka et al. 2019), Ohio (Farinas and Hand 2020) and New York (Weldon et al. 2020). This is the first known report of Golovinomyces ambrosiae causing powdery mildew on hemp in New Jersey. With the recent opening ( Dec15, 2021) of cultivation licensing and retailing of recreational marijuana, the acreage of Hemp production in New Jersey is expected to significantly increase, particularly for greenhouse production. It is important to document the species to develop management strategies to control this disease.

Prevalence and Risk Factors of Neurologic Manifestations in Hospitalized Children Diagnosed with Acute SARS-CoV-2 or MIS-C.

BACKGROUND: Our objective was to characterize the frequency, early impact, and risk factors for neurological manifestations in hospitalized children with acute severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection or multisystem inflammatory syndrome in children (MIS-C). METHODS: Multicenter, cross-sectional study of neurological manifestations in children aged <18 years hospitalized with positive SARS-CoV-2 test or clinical diagnosis of a SARS-CoV-2-related condition between January 2020 and April 2021. Multivariable logistic regression to identify risk factors for neurological manifestations was performed. RESULTS: Of 1493 children, 1278 (86%) were diagnosed with acute SARS-CoV-2 and 215 (14%) with MIS-C. Overall, 44% of the cohort (40% acute SARS-CoV-2 and 66% MIS-C) had at least one neurological manifestation. The most common neurological findings in children with acute SARS-CoV-2 and MIS-C diagnosis were headache (16% and 47%) and acute encephalopathy (15% and 22%), both P < 0.05. Children with neurological manifestations were more likely to require intensive care unit (ICU) care (51% vs 22%), P < 0.001. In multivariable logistic regression, children with neurological manifestations were older (odds ratio [OR] 1.1 and 95% confidence interval [CI] 1.07 to 1.13) and more likely to have MIS-C versus acute SARS-CoV-2 (OR 2.16, 95% CI 1.45 to 3.24), pre-existing neurological and metabolic conditions (OR 3.48, 95% CI 2.37 to 5.15; and OR 1.65, 95% CI 1.04 to 2.66, respectively), and pharyngeal (OR 1.74, 95% CI 1.16 to 2.64) or abdominal pain (OR 1.43, 95% CI 1.03 to 2.00); all P < 0.05. CONCLUSIONS: In this multicenter study, 44% of children hospitalized with SARS-CoV-2-related conditions experienced neurological manifestations, which were associated with ICU admission and pre-existing neurological condition. Posthospital assessment for, and support of, functional impairment and neuroprotective strategies are vitally needed.

Rapid Spread of Severe Fever with Thrombocytopenia Syndrome Virus by Parthenogenetic Asian Longhorned Ticks.

Severe fever with thrombocytopenia syndrome virus (SFTSV) is spreading rapidly in Asia. This virus is transmitted by the Asian longhorned tick (Haemaphysalis longicornis), which has parthenogenetically and sexually reproducing populations. Parthenogenetic populations were found in ≥15 provinces in China and strongly correlated with the distribution of severe fever with thrombocytopenia syndrome cases. However, distribution of these cases was poorly correlated with the distribution of populations of bisexual ticks. Phylogeographic analysis suggested that the parthenogenetic population spread much faster than bisexual population because colonization is independent of sexual reproduction. A higher proportion of parthenogenetic ticks was collected from migratory birds captured at an SFTSV-endemic area, implicating the contribution to the long-range movement of these ticks in China. The SFTSV susceptibility of parthenogenetic females was similar to that of bisexual females under laboratory conditions. These results suggest that parthenogenetic Asian longhorned ticks, probably transported by migratory birds, play a major role in the rapid spread of SFTSV.

Comparative hologenomics of two Ixodes scapularis tick populations in New Jersey.

Tick-borne diseases, such as those transmitted by the blacklegged tick Ixodes scapularis, are a significant and growing public health problem in the US. There is mounting evidence that co-occurring non-pathogenic microbes can also impact tick-borne disease transmission. Shotgun metagenome sequencing enables sampling of the complete tick hologenome-the collective genomes of the tick and all of the microbial species contained therein, whether pathogenic, commensal or symbiotic. This approach simultaneously uncovers taxonomic composition and allows the detection of intraspecific genetic variation, making it a useful tool to compare spatial differences across tick populations. We evaluated this approach by comparing hologenome data from two tick samples (N = 6 ticks per location) collected at a relatively fine spatial scale, approximately 23 km apart, within a single US county. Several intriguing variants in the data between the two sites were detected, including polymorphisms in both in the tick's own mitochondrial DNA and that of a rickettsial endosymbiont. The two samples were broadly similar in terms of the microbial species present, including multiple known tick-borne pathogens (Borrelia burgdorferi, Babesia microti, and Anaplasma phagocytophilum), filarial nematodes, and Wolbachia and Babesia species. We assembled the complete genome of the rickettsial endosymbiont (most likely Rickettsia buchneri) from both populations. Our results provide further evidence for the use of shotgun metagenome sequencing as a tool to compare tick hologenomes and differentiate tick populations across localized spatial scales.

Content Validation of Clinician-Reported Items for a Severity Measure for CDKL5 Deficiency Disorder.

CDKL5 deficiency disorder (CDD) results in early-onset seizures and severe developmental impairments. A CDD clinical severity assessment (CCSA) was previously developed with clinician and parent-report items to capture information on a range of domains. Consistent with US Food and Drug Administration (FDA) guidelines, content validation is the first step in evaluating the psychometric properties of an outcome measure. The aim of this study was to validate the content of the clinician-reported items in the CCSA (CCSA-Clinician). Eight neurologists leading the USA CDD Center of Excellence clinics were interviewed using the "think aloud" technique to critique 26 clinician-reported items. Common themes were aggregated, and a literature search of related assessments informed item modifications. The clinicians then participated in 2 consensus meetings to review themes and finalize the items. A consensus was achieved for the content of the CCSA-Clinician. Eight of the original items were omitted, 11 items were added, and the remaining 18 items were revised. The final 29 items were classified into 2 domains: functioning and neurologic impairments. This study enabled refinement of the CCSA-Clinician and provided evidence for its content validity. This preliminary validation is essential before field testing and further validation, in order to advance the instrument toward clinical trial readiness.

Effect of fenfluramine on convulsive seizures in CDKL5 deficiency disorder.

CDKL5 deficiency disorder (CDD) is an X-linked pharmacoresistant neurogenetic disorder characterized by global developmental delays and uncontrolled seizures. Fenfluramine (FFA), an antiseizure medication (ASM) indicated for treating convulsive seizures in Dravet syndrome, was assessed in six patients (five female; 83%) with CDD whose seizures had failed 5-12 ASMs or therapies. Median age at enrollment was 6.5 years (range: 2-26 years). Mean FFA treatment duration was 5.3 months (range: 2-9 months) at 0.4 mg/kg/day (n = 2) or 0.7 mg/kg/day (n = 4; maximum: 26 mg/day). One patient had valproate added for myoclonic seizures. The ASM regimens of all other patients were stable. Among five patients with tonic-clonic seizures, FFA treatment resulted in a median 90% reduction in frequency (range: 86%-100%). Tonic seizure frequency was reduced by 50%-60% in two patients with this seizure type. One patient experienced fewer myoclonic seizures; one patient first developed myoclonic seizures on FFA, which were controlled with valproate. Adverse events were reported in two patients. The patient with added valproate experienced lethargy; one patient had decreased appetite and flatus. No patient developed valvular heart disease or pulmonary arterial hypertension. Our preliminary results suggest that FFA may be a promising ASM for CDD. Randomized clinical trials are warranted.

Amoeba Genome Reveals Dominant Host Contribution to Plastid Endosymbiosis.

Eukaryotic photosynthetic organelles, plastids, are the powerhouses of many aquatic and terrestrial ecosystems. The canonical plastid in algae and plants originated >1 Ga and therefore offers limited insights into the initial stages of organelle evolution. To address this issue, we focus here on the photosynthetic amoeba Paulinella micropora strain KR01 (hereafter, KR01) that underwent a more recent (∼124 Ma) primary endosymbiosis, resulting in a photosynthetic organelle termed the chromatophore. Analysis of genomic and transcriptomic data resulted in a high-quality draft assembly of size 707 Mb and 32,361 predicted gene models. A total of 291 chromatophore-targeted proteins were predicted in silico, 208 of which comprise the ancestral organelle proteome in photosynthetic Paulinella species with functions, among others, in nucleotide metabolism and oxidative stress response. Gene coexpression analysis identified networks containing known high light stress response genes as well as a variety of genes of unknown function ("dark" genes). We characterized diurnally rhythmic genes in this species and found that over 49% are dark. It was recently hypothesized that large double-stranded DNA viruses may have driven gene transfer to the nucleus in Paulinella and facilitated endosymbiosis. Our analyses do not support this idea, but rather suggest that these viruses in the KR01 and closely related P. micropora MYN1 genomes resulted from a more recent invasion.