Repair of leukemia-associated single nucleotide variants via interallelic gene conversion.

CRISPR-Cas9 is a useful tool for inserting precise genetic alterations through homology-directed repair (HDR), although current methods rely on provision of an exogenous repair template. Here, we tested the possibility of repairing heterozygous single nucleotide variants (SNVs) using the cell's own wild-type allele rather than an exogenous template. Using high-fidelity Cas9 to perform allele-specific CRISPR across multiple human leukemia cell lines as well as in primary hematopoietic cells from patients with leukemia, we find high levels of reversion to wild-type in the absence of exogenous template. Moreover, we demonstrate that bulk treatment to revert a truncating mutation in ASXL1 using CRISPR-mediated interallelic gene conversion (IGC) is sufficient to prolong survival in a human cell line-derived xenograft model (median survival 33 days vs 27.5 days; p = 0.0040). These results indicate that IGC can be applied to numerous types of leukemia and can meaningfully alter cellular phenotypes at scale. Because our method targets single-base mutations, rather than larger variants targeted by IGC in prior studies, it greatly expands the pool of risk-increasing genetic lesions which could potentially be targeted by IGC. This technique may reduce cost and complexity for experiments modeling phenotypic consequences of SNVs. The principles of SNV-specific IGC demonstrated in this proof-of-concept study could be applied to investigate the phenotypic effects of targeted clonal reduction of leukemogenic SNV driver mutations.

Phaseolus vulgaris STP13.1 is an H+-coupled monosaccharide transporter, present in source leaves and seed coats, with higher substrate affinity at depolarized potentials.

Sugar transport proteins (STPs) are high-affinity H+-coupled hexose symporters. Recently, the contribution of STP13 to bacterial and fungal pathogen resistance across multiple plant species has garnered significant interest. Quantitative PCR analysis of source leaves, developing embryos, and seed coats of Phaseolus vulgaris L. (common bean) revealed that PvSTP13.1 was expressed in source leaves and seed coats throughout seed development. In contrast, PvSTP13.1 transcripts were detected at exceedingly low levels in developing embryos. To characterize the transport mechanism, PvSTP13.1 was expressed in Xenopus laevis oocytes, and inward-directed currents were analyzed using two-electrode voltage clamping. PvSTP13.1 was shown to function as an H+-coupled monosaccharide symporter exhibiting a unique high affinity for hexoses and aldopentoses at depolarized membrane potentials. Specifically, of the 31 assessed substrates, which included aldohexoses, deoxyhexoses, fructose, 3-O-methyl-D-glucose, aldopentoses, polyols, glycosides, disaccharides, trisaccharides, and glucuronic acid, PvSTP13.1 displayed the highest affinity (K 0.5) for glucose (43 µM), mannose (92 µM), galactose (145 µM), fructose (224 µM), xylose (1.0 mM), and fucose (3.7 mM) at pH 5.6 at a depolarized membrane potential of -40 mV. The results presented here suggest PvSTP13.1 contributes to retrieval of hexoses from the apoplasmic space in source leaves and coats of developing seeds.

Enigmatic persistence of aerobic methanotrophs in oxygen-limiting freshwater habitats.

Methanotrophic bacteria mitigate emissions of the potent greenhouse gas methane (CH4) from a variety of anthropogenic and natural sources, including freshwater lakes, which are large sources of CH4 on a global scale. Despite a dependence on dioxygen (O2) for CH4 oxidation, abundant populations of putatively aerobic methanotrophs have been detected within microoxic and anoxic waters and sediments of lakes. Experimental work has demonstrated active aerobic methanotrophs under those conditions, but how they are able to persist and oxidize CH4 under O2 deficiency remains enigmatic. In this review, we discuss possible mechanisms that underpin the persistence and activity of aerobic methanotrophs under O2-limiting conditions in freshwater habitats, particularly lakes, summarize experimental evidence for microbial oxidation of CH4 by aerobic bacteria under low or no O2, and suggest future research directions to further explore the ecology and metabolism of aerobic methanotrophs in O2-limiting environments.

Barriers and facilitators to consuming pulses: a qualitative exploration including effects of trying recipes at home.

Pulses are a healthy, sustainable, low cost food, but consumption levels are low for a variety of reasons, including practical and cooking concerns. This work aimed to explore barriers and facilitators towards pulse consumption and increasing consumption, and the potential value of cooking suggestions and recipes for changing these perceptions. Two qualitative studies were undertaken. In Study 1, 21 participants (10 males, 11 females, of a range of ages, cooking responsibilities, and experiences with pulses) were interviewed both before and after receiving cooking suggestions and recipes. In Study 2, 12 participants (2 males, 10 females, as above) were interviewed once after trying recipes. Interviews were audio-recorded, transcribed, and analysed using thematic analysis. Seven themes described barriers and facilitators towards pulse consumption: 'Enjoyment and Sensory properties'; 'Benefits and Recommendations'; 'Practical Concerns'; 'Cooking Concerns'; 'Compatibility with current diet'; 'Personal Influences'; and 'External Influences'. Some similar themes also referred to increasing consumption: 'Willingness'; 'Awareness, Knowledge of Benefits'; 'Knowledge of Cooking and Practical Concerns'; and 'Compatibility with current diet'. Cooking suggestions and recipe use resulted in themes on 'Awareness'; 'Willingness, Trying New Things'; 'Small Changes'; and facilitators associated with 'Enjoyment, Sensory Properties, Practical Concerns, Benefits' and 'Knowledge, Cooking Ideas and Confidence, Incorporation, Cooking Solutions'. Barriers related to 'Risk and Preconceptions'; 'Awareness, but' inaction and additional considerations were also found. Our findings demonstrate a positive role for pulse consumption for increased experience, familiarity, and confidence with preparing, cooking, and consuming these healthy and sustainable foods.

Severe epistaxis after posterior nasal nerve ablation requiring surgical intervention: A multi-center case series.

KEY POINTS: Severe epistaxis occurs in 2% of PNN ablation cases, independent of method or device type. Major epistaxis requiring intervention after PNN ablation can occur on average 20 days post-procedure.

Inhibition of ATR opposes glioblastoma invasion through disruption of cytoskeletal networks and integrin internalisation via macropinocytosis.

BACKGROUND: Glioblastomas have highly infiltrative growth patterns that contribute to recurrence and poor survival. Despite infiltration being a critical therapeutic target, no clinically useful therapies exist which counter glioblastoma invasion. Here, we report that inhibition of Ataxia telangiectasia and Rad 3 related kinase (ATR) reduces invasion of glioblastoma cells through dysregulation of cytoskeletal networks and subsequent integrin trafficking. METHODS: Glioblastoma motility and invasion were assessed in vitro and in vivo in response to ATR inhibition (ATRi) and ATR over-expression using timelapse microscopy, two orthotopic glioblastoma models and intravital imaging. Disruption to cytoskeleton networks and endocytic processing were investigated via high throughput, super resolution imaging and intravital imaging. RESULTS: High ATR expression was associated with significantly poorer survival in clinical datasets whilst histological, protein expression and spatial transcriptomics using glioblastoma tumour specimens revealed higher ATR expression at infiltrative margins. Pharmacological inhibition with two different compounds and RNAi targeting of ATR opposed invasion of glioblastoma, whereas overexpression of ATR drove migration. Subsequent investigation revealed that cytoskeletal dysregulation reduced macropinocytotic internalisation of integrins at growth cone-like structures, resulting in a tumour microtube retraction defect. The biological relevance and translational potential of these findings was confirmed using two orthotopic in vivo models of glioblastoma and intravital imaging. CONCLUSION: We demonstrate a novel role for ATR in determining invasion in glioblastoma cells and propose that pharmacological targeting of ATR could have far reaching clinical benefits beyond radiosensitisation.

Desulfoferula mesophilus gen. nov. sp. nov., a mesophilic sulfate-reducing bacterium isolated from a brackish lake sediment.

A novel sulfate-reducing bacterium (strain 12FAKT) was isolated from sediment sampled from a brackish lake in Japan. Respiratory growth was observed with formate and pyruvate as an electron donor. Sulfate, thiosulfate, elemental sulfur and dimethyl sulfoxide were utilized as an electron acceptor. The isolate grew over a temperature range of 18-42 °C (optimum 35-37 °C), a NaCl concentration range of 50-450 mM (optimum 150-300 mM) and a pH range of 6.6-7.5. The 12FAKT genome consists of a circular chromosome with a length of 4.5 Mbp and G + C content of 63.6%. Based on 16S rRNA gene sequence similarity, the closest cultured relative was Desulfarculus baarsii 2st14T (92.2%). Genome-based phylogenetic analysis placed strain 12FAKT within the family Desulfarculaceae but did not affiliate the strain with any existing genus. Taken together, we propose a novel species of a novel genus, Desulfoferula mesophilus gen. nov. sp. nov. with the type strain 12FAKT (= DSM 115219T = JCM 39399T).

Massive ossifying fibroma of mandible: A case report and review of the literature.

Key Clinical Message: Benign fibro-osseous neoplasm. Massive size is rarely reported. May be confused with other pathologies such as fibrous dysplasia or osteosarcoma. Aggressive nature and high recurrence pose management challenges. Treated by surgery. Abstract: Ossifying fibroma is a benign fibro-osseous neoplasm. It can affect both mandible and maxilla. Precise diagnosis can be challenging due to significant overlap of clinicopathological features with other neoplasms. Case reports with massive tumor sizes as presented in our case are uncommon. Huge tumor size can cause alarm for other pathologies such as osteosarcoma. The radiological tests should reassure the attending practitioner and histological examination confirm the diagnosis. The aim of the present report is to discuss a case of a giant ossifying fibroma in a 13-year-old male child. He presented with a progressive mandibular mass for 4 years. Clinical, radiological, and pathological characteristics and surgical treatment approaches are further discussed. This is one of the rare cases of massive ossifying fibroma in the English literature.

Management of a giant pleomorphic adenoma of the soft palate: A case report.

Key Clinical Message: Benign mixed salivary gland tumor is comprised of epithelial and myoepithelial cells and represents up to 80% of tumors of the parotid gland. It is relatively rare in the soft palate and in other minor salivary glands. Surgery is the standard care. Abstract: Salivary gland tumors are relatively rare and morphologically diverse group of lesions. Pleomorphic adenoma (PA) is the most common salivary gland tumor, accounting for approximately 80% of all major and minor salivary gland tumors. PA usually affects the parotid gland. Huge PA occurring in soft palate is extremely rare. Patients with these tumors are usually between at the age of 40 and 60 years. The tumors exhibit pleomorphic nature microscopically that may pose diagnostic challenges to pathologists as may confuse PA histopathologically with other salivary gland tumors. Surgery is the standard treatment. The purpose of writing this case study is to describe unusual case of a giant PA of the soft palate found in a 44-year-old male successfully managed at our facility.

The prevalence of equine odontoclastic tooth resorption and hypercementosis and the role of interincisal angulation in disease severity in a representative cohort of horses in Switzerland.

INTRODUCTION: Equine odontoclastic tooth resorption and hypercementosis (EOTRH) is an increasingly diagnosed degenerative dental disease in aged horses. The primary aim of this retrospective study was to determine the prevalence of EOTRH in horses admitted to the Equine Hospital, University of Zurich, for dental procedures from 2004 to 2017. A secondary goal was to measure and compare interincisal angles on two-dimensional radiographs of horses with EOTRH to determine whether the interincisal angle is associated with age and severity of the disease. Radiographs were assessed for the presence of lysis and/or hypercementosis, and the number and position of the teeth affected were determined. Each tooth was also evaluated using the modified classification system introduced by Rehrl et al. (2018), in which stage 0 indicates no radiographic abnormalities and stage 3 denotes severe abnormalities. The overall stage was defined by the tooth with the most severe lesions. The interincisal angle was determined in horses that had suitable radiographs. The medical records of 838 horses admitted for dental procedures were evaluated, and 85 (10,1 %) had clinical evidence of EOTRH. The mean interincisal angle was 136,06 ° in horses with mild to moderate EOTRH and 135,10 ° (SD = 11,90 °) in severely affected patients. In conclusion, the angle measurements on lateral radiographs were highly reproducible. However, the interincisal angle was not associated with age or the severity of EOTRH. The interincisal angle and the disease pattern were not correlated.

INTRODUCTION: La résorption et l'hypercémentose odontoclastique des dents chez le cheval (EOTRH) est une maladie dentaire dégénérative de plus en plus diagnostiquée chez les animaux âgés. L'objectif principal de cette étude rétrospective était de déterminer la prévalence de l'EOTRH chez les chevaux admis à l'Hôpital équin de l'Université de Zurich pour des interventions dentaires entre 2014 et 2017. Un objectif secondaire était de mesurer et de comparer les angles inter-incisifs sur des radiographies bidimensionnelles de chevaux atteints d'EOTRH afin de déterminer si cet angle est associé à l'âge et à la gravité de la maladie. Les radiographies ont été évaluées pour la présence de lyse et/ou d'hypercémentose, et le nombre et la position des dents affectées ont été déterminés. Chaque dent a également été évaluée à l'aide du système de classification modifié introduit par Rehrl et al. (2018), dans lequel le stade 0 indique l'absence d'anomalies radiographiques et le stade 3 indique des anomalies graves. Le stade global a été défini par la dent présentant les lésions les plus sévères. L'angle inter-incisif a été déterminé chez les chevaux qui avaient des radiographies appropriées. Les dossiers médicaux de 838 chevaux admis pour des interventions dentaires ont été évalués et 85 (10,1 %) présentaient des signes cliniques d'EOTRH. L'angle inter-incisif moyen était de 136,06° chez les chevaux présentant une EOTRH légère à modérée et de 135,10° (écart-type = 11,90°) chez les patients gravement atteints. En conclusion, les mesures d'angle sur les radiographies latérales étaient très reproductibles. Cependant, l'angle inter-incisif n'était pas associé à l'âge ou à la sévérité de l'EOTRH. L'angle inter-incisif et le profil de la maladie n'étaient pas corrélés.

Discovery of the vector of visceral leishmaniasis, Phlebotomus (Artemievus) alexandri Sinton, 1928, in Kenya suggests complex transmission dynamics.

Visceral and cutaneous leishmaniasis are endemic to specific regions due to the ecological preferences of phlebotomine sand flies and Leishmania spp. transmission. Sand fly entomological data in northern Kenya are scarce due to limited studies and neglect of leishmaniasis. The aim of this study was to investigate: (i) sand fly diversity and distribution; (ii) occurrence of Leishmania DNA within sand flies; and (iii) blood-meal sources of sand flies in Laisamis, northern Kenya. We conducted an entomological survey during February and March of 2021 in five areas of Laisamis sub-county using standard CDC light traps. A total of 1009 sand flies (394 male and 615 female) were morphologically identified, and representative samples verified by PCR amplification and sequencing of the cytochrome c oxidase subunit 1 (cox1) gene. Similarly, we identified blood-meal sources and Leishmania DNA in female sand flies by PCR amplicon sequencing of the vertebrate cytochrome b (cyt b) gene and internal transcribed spacer 1 (ITS1) of the 28S rRNA gene, respectively. Sergentomyia clydei (59.8%) was the most abundant sand fly species. Though collected mainly from one locality (Tirgamo), 14.8% of samples belonged to Phlebotomus (Artemievus) alexandri Sinton, 1928. We detected DNA of Leishmania major in 5.19% of Ph. alexandri, whereas Leishmania adleri DNA was detected in S. clydei (7.51%), Sergentomyia squamipleuris (8.00%), and Sergentomyia africanus (8.33%). Nine of 13 blood-fed sand flies had obtained blood from humans, of which 33.3% had L. major DNA. Both Ph. alexandri and S. clydei primarily fed on humans and could potentially be involved in the transmission of cutaneous leishmaniasis. The findings of this study contribute to the understanding of sand fly vector populations and their potential to transmit leishmaniasis in the area.

Extra-mandibular Osteoradionecrosis after the Treatment of Head and Neck Cancer.

AIMS: Osteoradionecrosis (ORN) is a serious toxicity of head and neck radiotherapy. It predominantly affects the mandible. Extra-mandibular ORN is rare. The aim of this study was to report the incidence and outcomes of extra-mandibular ORNs from a large institutional database. MATERIALS AND METHODS: In total, 2303 head and neck cancer patients were treated with radical or adjuvant radiotherapy. Of these, extra-mandibular ORN developed in 13 patients (0.5%). RESULTS: Maxillary ORNs (n = 8) were a consequence of the treatment of various primaries (oropharynx = 3, sinonasal = 2, maxilla = 2, parotid = 1). The median interval from the end of radiotherapy to the development of ORN was 7.5 months (range 3-42 months). The median radiotherapy dose in the centre of the ORN was 48.5 Gy (range 22-66.5 Gy). Four patients (50%) healed in 7, 14, 20 and 41 months. All temporal bone ORNs (n = 5) developed after treatment to the parotid gland (of a total of 115 patients who received radiotherapy for parotid gland malignancy). The median interval from the end of radiotherapy to the development of ORN was 41 months (range 20-68 months). The median total dose in the centre of the ORN was 63.5 Gy (range 60.2-65.3 Gy). ORN healed in only one patient after 32 months of treatment with repeated debridement and topical betamethasone cream. CONCLUSION: Extra-mandibular ORN is a rare late toxicity and this current study provides useful information on its incidence and outcome. The risk of temporal bone ORN should be considered in the treatment of parotid malignancies and patients should be counselled. More research is required to determine the optimal management of extra-mandibular ORN, particularly on the role of the PENTOCLO regimen.

The Extraordinary Case of a Woman with a 30-Year-Long Diffuse Leishmaniasis Cured with One Single Ampoule of Intranasal Pentavalent Antimoniate.

Infection with Leishmania amazonensis and L. mexicana may lead to diffuse cutaneous leishmaniasis. The cure is exceptional, especially for the strange case of this lady. Case report: The patient acquired the disease in childhood and remained with lesions for over 30 years, albeit several treatments. She worsened after a pregnancy, developing disseminated lesions. Miltefosine with amphotericin B and pentamidine resulted in remission. Lesions reappeared after one year, accompanied by intra-nasal infiltration of the disease. The nasal spraying of a single ampoule of pentavalent antimoniate resulted in the sustained disappearance of the nasal symptoms and all the cutaneous lesions. After over eight years, she remains disease-free, albeit under renal replacement therapy. The high nasal mucosal antimonial concentration may explain the long-lasting cure via new MHC class I epitope-specific CD8+ cell clones against L. amazonensis present in the nasal mucosa.

Tool for automatic macrozone characterization from EBSD data sets of titanium alloys.

Microtexture heterogeneities are commonly found in titanium forgings because of the thermomechanical processing. Also known as macrozones, these regions can reach millimetres in length, with grains sharing a similar crystallographic orientation leading to less resistance to crack propagation. Since the link between macrozones and the reduction of cold-dwell-fatigue performance on rotative components in gas turbine engines was established, efforts have been put into macrozone definition and characterization. The electron backscatter diffraction (EBSD) technique, widely used for texture analysis, allows for a qualitative macrozone characterization; however, further processing is required to define the boundaries and disorientation spread of each macrozone. Current approaches often use c-axis misorientation criteria, but this can sometimes lead to a large disorientation spread within a macrozone. This article describes the development and application of a computational tool implemented in MATLAB for automatic macrozone identification from EBSD data sets on the basis of a more conservative approach where both the c-axis tilting and rotation are considered. The tool allows for detection of macrozones according to the disorientation angle and density-fraction criteria. The clustering efficiency is validated by pole-figure plots, and the effects of the key parameters defining the macrozone clustering (disorientation and fraction) are discussed. In addition, this tool was successfully applied to both fully equiaxed and bimodal microstructures of titanium forgings.

What do we want? Training! When do we want it? Now? A training needs analysis for adaptive radiotherapy for therapeutic radiographers.

INTRODUCTION: Therapeutic radiographers (TRs) have adapted to the changing requirements and demands of the oncology service and in response to advanced techniques such as on-line adaptive MRI-guided radiotherapy (MRIgRT). The skills required for MRIgRT would benefit many TRs not just those involved in this technique. This study presents the results of a training needs analysis (TNA) for the required MRIgRT skills in readiness for training TRs for current and future practice. METHODS: A UK-based TNA was used to ask TRs about their knowledge and experience with essential skills required for MRIgRT based on previous investigations into the topic. A five-point Likert scale was used for each of the skills and the difference in values were used to calculate the training need for current and future practice. RESULTS: 261 responses were received (n = 261). The skill rated the most important to current practice was CBCT/CT matching and/or fusion. The current highest priority needs were radiotherapy planning and radiotherapy dosimetry. The skill rated the most important to future practice was CBCT/CT matching and/or fusion. The future highest priority needs were MRI acquisition and MRI Contouring. Over 50% of participants wanted training or additional training in all skills. There was an increase in all values for skills investigated from current to future roles. CONCLUSION: Although the examined skills were viewed as important to current roles, the future training needs, both overall and high priority, were different compared to current roles. As the 'future' of radiotherapy can arrive rapidly, it is essential that training is delivered appropriately and timely. Before this can occur, there must be investigations into the method and delivery of this training. IMPLICATIONS FOR PRACTICE: Role development. Education changes for therapeutic radiographers.

Role of Adjuvant Radiotherapy in Acinic Cell Carcinoma of the Salivary Glands: A Systematic Review.

A systematic review was carried out to evaluate if adjuvant radiotherapy for acinic cell carcinomas (ACCs) of salivary glands improves survival. Twelve retrospective studies published between 2000 and 2020 that analysed the effect of radiotherapy on salivary gland neoplasms and ACCs of salivary glands and met the inclusion criteria were included in the review. The overall quality of the studies was moderate to low. There was no high-quality evidence for improved survival with radiotherapy for ACCs of the salivary gland. Some evidence suggests that there may be an advantage for patients with high-grade tumours, but these data should be interpreted with caution due to the small number of patients and low-quality evidence. Good quality of evidence is lacking. Recommendation for adjuvant radiotherapy for tumours with poor prognostic factors will require discussion and shared decision-making with the patients.

Changes in Depression and Anxiety Among Children and Adolescents From Before to During the COVID-19 Pandemic: A Systematic Review and Meta-analysis.

Importance: There is a growing body of high-quality cohort-based research that has examined changes in child and adolescent mental health during the COVID-19 pandemic vs before the pandemic. Some studies have found that child and adolescent depression and anxiety symptoms have increased, while others have found these symptoms to have remained stable or decreased. Objective: To synthesize the available longitudinal cohort-based research evidence to estimate the direction and magnitude of changes in depression and anxiety symptoms in children and adolescents assessed before and during the pandemic. Data Sources: Medline, Embase, and PsycInfo were searched for studies published between January 1, 2020, and May 17, 2022. Study Selection: Included studies reported on depression and/or anxiety symptoms, had cohort data comparing prepandemic to pandemic estimates, included a sample of children and/or adolescents younger than 19 years, and were published in English in a peer-reviewed journal. Data Extraction and Synthesis: In total, 53 longitudinal cohort studies from 12 countries with 87 study estimates representing 40 807 children and adolescents were included. Main Outcomes and Measures: Standardized mean changes (SMC) in depression and anxiety symptoms from before to during the pandemic. Results: The analysis included 40 807 children and adolescents represented in pre-COVID-19 studies and 33 682 represented in during-COVID-19 studies. There was good evidence of an increase in depression symptoms (SMC, 0.26; 95% CI, 0.19 to 0.33). Changes in depression symptoms were most conclusive for study estimates among female individuals (SMC, 0.32; 95% CI, 0.21 to 0.42), study estimates with mid to high income (SMC, 0.35; 95% CI, 0.07 to 0.63), and study estimates sourced from North America (SMC, 0.25; 95% CI, 0.15 to 0.36) and Europe (SMC, 0.35; 95% CI, 0.17 to 0.53). There was strong evidence that anxiety symptoms increased slightly during the pandemic (SMC, 0.10; 95% CI, 0.04 to 0.16), and there was some evidence of an increase in study estimates with mid to high income. Conclusions: This systematic review and meta-analysis of longitudinal studies including children and adolescents found an increase in depression symptoms during the COVID-19 pandemic, particularly among female individuals and those from relatively higher-income backgrounds.

Isoliensinine from Cissampelos pariera rhizomes exhibits potential gametocytocidal and anti-malarial activities against Plasmodium falciparum clinical isolates.

BACKGROUND: The unmet demand for effective malaria transmission-blocking agents targeting the transmissible stages of Plasmodium necessitates intensive discovery efforts. In this study, a bioactive bisbenzylisoquinoline (BBIQ), isoliensinine, from Cissampelos pariera (Menispermaceae) rhizomes was identified and characterized for its anti-malarial activity. METHODS: Malaria SYBR Green I fluorescence assay was performed to evaluate the in vitro antimalarial activity against D6, Dd2, and F32-ART5 clones, and immediate ex vivo (IEV) susceptibility for 10 freshly collected P. falciparum isolates. To determine the speed- and stage-of-action of isoliensinine, an IC50 speed assay and morphological analyses were performed using synchronized Dd2 asexuals. Gametocytocidal activity against two culture-adapted gametocyte-producing clinical isolates was determined using microscopy readouts, with possible molecular targets and their binding affinities deduced in silico. RESULTS: Isoliensinine displayed a potent in vitro gametocytocidal activity at mean IC50gam values ranging between 0.41 and 0.69 µM for Plasmodium falciparum clinical isolates. The BBIQ compound also inhibited asexual replication at mean IC50Asexual of 2.17 µM, 2.22 µM, and 2.39 µM for D6, Dd2 and F32-ART5 respectively, targeting the late-trophozoite to schizont transition. Further characterization demonstrated a considerable immediate ex vivo potency against human clinical isolates at a geometric mean IC50IEV = 1.433 µM (95% CI 0.917-2.242). In silico analyses postulated a probable anti-malarial mechanism of action by high binding affinities for four mitotic division protein kinases; Pfnek1, Pfmap2, Pfclk1, and Pfclk4. Additionally, isoliensinine was predicted to possess an optimal pharmacokinetics profile and drug-likeness properties. CONCLUSION: These findings highlight considerable grounds for further exploration of isoliensinine as an amenable scaffold for malaria transmission-blocking chemistry and target validation.