Tumor Treating Fields (TTFields) combined with the drug repurposing approach CUSP9v3 induce metabolic reprogramming and synergistic anti-glioblastoma activity in vitro.

BACKGROUND: Glioblastoma represents a brain tumor with a notoriously poor prognosis. First-line therapy may include adjunctive Tumor Treating Fields (TTFields) which are electric fields that are continuously delivered to the brain through non-invasive arrays. On a different note, CUSP9v3 represents a drug repurposing strategy that includes 9 repurposed drugs plus metronomic temozolomide. Here, we examined whether TTFields enhance the antineoplastic activity of CUSP9v3 against this disease. METHODS: We performed preclinical testing of a multimodal approach of TTFields and CUSP9v3 in different glioblastoma models. RESULTS: TTFields had predominantly synergistic inhibitory effects on the cell viability of glioblastoma cells and non-directed movement was significantly impaired when combined with CUSP9v3. TTFields plus CUSP9v3 significantly enhanced apoptosis, which was associated with a decreased mitochondrial outer membrane potential (MOMP), enhanced cleavage of effector caspase 3 and reduced expression of Bcl-2 and Mcl-1. Moreover, oxidative phosphorylation and expression of respiratory chain complexes I, III and IV was markedly reduced. CONCLUSION: TTFields strongly enhance the CUSP9v3-mediated anti-glioblastoma activity. TTFields are currently widely used for the treatment of glioblastoma patients and CUSP9v3 was shown to have a favorable safety profile in a phase Ib/IIa trial (NCT02770378) which facilitates transition of this multimodal approach to the clinical setting.

Detecting Förster resonance energy transfer in living cells by conventional and spectral flow cytometry.

Assays based on Förster resonance energy transfer (FRET) can be used to study many processes in cell biology. Although this is most often done with microscopy for fluorescence detection, we report two ways to measure FRET in living cells by flow cytometry. Using a conventional flow cytometer and the "3-cube method" for intensity-based calculation of FRET efficiency, we measured the enzymatic activity of specific kinases in cells expressing a genetically-encoded reporter. For both AKT and protein kinase A, the method measured kinase activity in time-course, dose-response, and kinetic assays. Using the Cytek Aurora spectral flow cytometer, which applies linear unmixing to emission measured in multiple wavelength ranges, FRET from the same reporters was measured with greater single-cell precision, in real time and in the presence of other fluorophores. Results from gene-knockout studies suggested that spectral flow cytometry might enable the sorting of cells on the basis of FRET. The methods we present provide convenient and flexible options for using FRET with flow cytometry in studies of cell biology.

Metabolomic Signatures Discriminate Horses with Clinical Signs of Atypical Myopathy from Healthy Co-grazing Horses.

Atypical myopathy (AM) is a severe rhabdomyolysis syndrome that occurs in grazing horses. Despite the presence of toxins in their blood, all horses from the same pasture are not prone to display clinical signs of AM. The objective of this study was to compare the blood metabolomic profiles of horses with AM clinical signs with those of healthy co-grazing (Co-G) horses. To do so, plasma samples from 5 AM horses and 11 Co-G horses were investigated using untargeted metabolomics. Metabolomic data were evaluated using unsupervised, supervised, and pathway analyses. Unsupervised principal component analysis performed with all detected features separated AM and healthy Co-G horses. Supervised analyses had identified 1276 features showing differential expression between both groups. Among them, 46 metabolites, belonging predominantly to the fatty acid, fatty ester, and amino acid chemical classes, were identified by standard comparison. Fatty acids, unsaturated fatty acids, organic dicarboxylic acids, and fatty esters were detected with higher intensities in AM horses in link with the toxins' pathological mechanism. The main relevant pathways were lipid metabolism; valine, leucine, and isoleucine metabolism; and glycine metabolism. This study revealed characteristic metabolite changes in the plasma of clinically affected horses, which might ultimately help scientists and field veterinarians to detect and manage AM. The raw data of metabolomics are available in the MetaboLights database with the access number MTBLS2579.

Design and Synthesis of Cathepsin-K-Activated Osteoadsorptive Fluorogenic Sentinel (OFS) Probes for Detecting Early Osteoclastic Bone Resorption in a Multiple Myeloma Mouse Model.

We describe the design and synthesis of OFS-1, an Osteoadsorptive Fluorogenic Sentinel imaging probe that is adsorbed by hydroxyapatite (HAp) and bone mineral surfaces, where it generates an external fluorescent signal in response to osteoclast-secreted cathepsin K (Ctsk). The probe consists of a bone-anchoring bisphosphonate moiety connected to a Förster resonance energy transfer (FRET) internally quenched fluorescent (IQF) dye pair, linked by a Ctsk peptide substrate, GHPGGPQG. Key structural features contributing to the effectiveness of OFS-1 were defined by structure-activity relationship (SAR) and modeling studies comparing OFS-1 with two cognates, OFS-2 and OFS-3. In solution or when preadsorbed on HAp, OFS-1 exhibited strong fluorescence when exposed to Ctsk (2.5-20 nM). Time-lapse photomicrographs obtained after seeding human osteoclasts onto HAp-coated well plates containing preadsorbed OFS-1 revealed bright fluorescence at the periphery of resorbing cells. OFS-1 administered systemically detected early osteolysis colocalized with orthotopic engraftment of RPMI-8226-Luc human multiple myeloma cells at a metastatic skeletal site in a humanized mouse model. OFS-1 is thus a promising new imaging tool for detecting abnormal bone resorption.

Dual metabolic reprogramming by ONC201/TIC10 and 2-Deoxyglucose induces energy depletion and synergistic anti-cancer activity in glioblastoma.

BACKGROUND: Dysregulation of the metabolome is a hallmark of primary brain malignancies. In this work we examined whether metabolic reprogramming through a multi-targeting approach causes enhanced anti-cancer activity in glioblastoma. METHODS: Preclinical testing of a combined treatment with ONC201/TIC10 and 2-Deoxyglucose was performed in established and primary-cultured glioblastoma cells. Extracellular flux analysis was used to determine real-time effects on OXPHOS and glycolysis. Respiratory chain complexes were analysed by western blotting. Biological effects on tumour formation were tested on the chorioallantoic membrane (CAM). RESULTS: ONC201/TIC10 impairs mitochondrial respiration accompanied by an increase of glycolysis. When combined with 2-Deoxyglucose, ONC201/TIC10 induces a state of energy depletion as outlined by a significant decrease in ATP levels and a hypo-phosphorylative state. As a result, synergistic anti-proliferative and anti-migratory effects were observed among a broad panel of different glioblastoma cells. In addition, this combinatorial approach significantly impaired tumour formation on the CAM. CONCLUSION: Treatment with ONC201/TIC10 and 2-Deoxyglucose results in a dual metabolic reprogramming of glioblastoma cells resulting in a synergistic anti-neoplastic activity. Given, that both agents penetrate the blood-brain barrier and have been used in clinical trials with a good safety profile warrants further clinical evaluation of this therapeutic strategy.

The combination of ibrutinib and rituximab demonstrates activity in first-line follicular lymphoma.

This phase 2 study evaluated the activity and safety of ibrutinib, a Bruton's tyrosine kinase inhibitor, plus rituximab in adults with previously untreated follicular lymphoma. Patients received once-daily ibrutinib 560 mg continuously plus once-weekly rituximab 375 mg/m2 for 4 weeks beginning Week 1 (Arm 1, n = 60) or Week 9 (following an 8-week ibrutinib lead-in) to explore biomarkers (Arm 2, n = 20). The primary endpoint was the best overall response rate (ORR). The median age was 58 years; most had an Eastern Cooperative Oncology Group Performance Status of 0 (74%) and Stage III/IV disease (84%). At a median study follow-up of 34 months in Arm 1 and 29 months in Arm 2, ORRs were 85% [95% confidence interval (CI) 73-93] and 75% (95% CI 51-91), respectively, with complete responses in 40% and 50%. The median duration of response was not reached in either arm; 30-month progression-free and overall survival rates were 67% and 97% (Arm 1) and 65% and 100% (Arm 2). The most common adverse events were fatigue, diarrhoea and nausea. Higher grade (Grade 3/4) haematological, haemorrhagic and cardiac events occurred infrequently. Ibrutinib plus rituximab was active and tolerable in first-line follicular lymphoma.

POLR1B and neural crest cell anomalies in Treacher Collins syndrome type 4.

PURPOSE: Treacher Collins syndrome (TCS) is a rare autosomal dominant mandibulofacial dysostosis, with a prevalence of 0.2-1/10,000. Features include bilateral and symmetrical malar and mandibular hypoplasia and facial abnormalities due to abnormal neural crest cell (NCC) migration and differentiation. To date, three genes have been identified: TCOF1, POLR1C, and POLR1D. Despite a large number of patients with a molecular diagnosis, some remain without a known genetic anomaly. METHODS: We performed exome sequencing for four individuals with TCS but who were negative for pathogenic variants in the known causative genes. The effect of the pathogenic variants was investigated in zebrafish. RESULTS: We identified three novel pathogenic variants in POLR1B. Knockdown of polr1b in zebrafish induced an abnormal craniofacial phenotype mimicking TCS that was associated with altered ribosomal gene expression, massive p53-associated cellular apoptosis in the neuroepithelium, and reduced number of NCC derivatives. CONCLUSION: Pathogenic variants in the RNA polymerase I subunit POLR1B might induce massive p53-dependent apoptosis in a restricted neuroepithelium area, altering NCC migration and causing cranioskeletal malformations. We identify POLR1B as a new causative gene responsible for a novel TCS syndrome (TCS4) and establish a novel experimental model in zebrafish to study POLR1B-related TCS.

Changes in behavioural synchrony during dog-assisted therapy for children with autism spectrum disorder and children with Down syndrome.

BACKGROUND: Dog-assisted therapy (DAT) is hypothesized to help children with autism spectrum disorder (ASD) and Down syndrome (DS). METHODS: The present authors compared synchronous movement patterns of these children (n = 10) and their therapy dogs during the first and last session of a DAT programme, and their post-therapy changes in emotional and behavioural problems. RESULTS: The present authors found a significant increase in synchrony between child and therapy dog over time. Exploratory analyses suggest more synchrony between children with ASD and their therapy dogs, compared to the children with DS. CONCLUSIONS: This study is the first to test the synchrony hypothesis, shedding light upon a mechanism that may underlie the effect of DAT and how this may be different for children with ASD and DS.

The influence of hay steaming on clinical signs and airway immune response in severe asthmatic horses.

BACKGROUND: Avoidance of antigenic stimuli was found to significantly reverse airway obstruction of horses with severe equine asthma (sEA). To date, no published study investigated the influence of steaming hay on lower airway condition of sEA-affected horses. The objectives were to determine the clinical, cytological and cytokine respiratory responses of both sEA and control (CTL) horses experimentally exposed to steamed or dry hay. RESULTS: A cohort of 6 sEA horses and 6 CTL horses was involved in this field study. On day 0, both groups were fed with steamed hay for 5 consecutive days, followed by a wash-out period of 26 days prior to be fed with dry hay for 5 consecutive days. Investigations performed 2 days prior to and 5 days after each challenge included clinical score, tracheal mucus accumulation, and bronchoalveolar lavage fluid (BALF) cytology and cytokine mRNA expression. Feeding steamed hay significantly decreased its mould content (P < 0.001). Mucus score significantly increased when feeding dry hay (P = 0.01). No significant influence of challenge type was found on clinical score. Percentages of neutrophils (P < 0.001) as well as mRNA expression of IL-1ß (P = 0.024), IL-6R (P = 0.021), IL-18 (P = 0.009) and IL-23 (P = 0.036) in BALF of sEA affected horses were significantly increased after both (steamed and dry hay) challenges. Relative mRNA expression of IL-1ß, IL-6R and IL-23 in BALF were also significantly correlated to neutrophil percentages and both clinical and tracheal mucus score. CONCLUSIONS: Steaming significantly decreased mould content but inconsistently influenced the respiratory response of sEA affected horses when fed hay. Based on BALF cytology and cytokine profiles, its relevance might be controversial as a non-medicinal therapy for sEA-affected horses.

Multiple molecular detection of respiratory viruses and associated signs of airway inflammation in racehorses.

BACKGROUND: The potential involvement of viruses in inflammatory airway disease (IAD) was previously investigated through either serology or PCR from nasopharyngeal swabs (NS). The aims of this study were to determine the prevalence and incidence of viral genome detection by qPCR in the equine airways, and their association with respiratory clinical signs. METHODS: Both NS and tracheal washes (TW) were collected monthly on 52 Standardbred racehorses at training, over 27 consecutive months (581 samples). Equid herpesviruses (EHV)-1, -4, -2 and -5, equine rhinitis virus-A and -B (ERBV), equine adenovirus-1 and -2, equine coronavirus and equine influenza virus were systematically investigated in both NS and TW. Nasal discharge, coughing, tracheal mucus score and TW neutrophil proportions were simultaneously recorded. RESULTS: Genome for 7/10 viruses were detected at least once throughout the study; up to 4 different viruses being also concomitantly detected. Monthly incidence in TW was respectively 27.9% (EHV-5), 24.8% (EHV-2), 7.1% (ERBV), 3.8% (EHV-4), 1.9% (EAdV1) and 0.2% (EHV-1; ERAV). Neither agreement nor correlation between NS and TW was found for respectively genome detection and viral loads. Detection of viral genome in NS was not associated with any clinical sign. Coughing was significantly associated with TW detection of EHV-2 DNA (OR 3.1; P = 0.01) and ERBV RNA (OR 5.3; P < 0.001). Detection of EHV-2 DNA in TW was also significantly associated with excess tracheal mucus (OR 2.1; P = 0.02). CONCLUSIONS: Detection and quantification of EHV-2 and ERBV by qPCR in TW, but not in NS, should be considered when investigating horses with IAD.

Connective tissue growth factor regulates adipocyte differentiation of mesenchymal stromal cells and facilitates leukemia bone marrow engraftment.

Mesenchymal stromal cells (MSCs) are a major component of the leukemia bone marrow (BM) microenvironment. Connective tissue growth factor (CTGF) is highly expressed in MSCs, but its role in the BM stroma is unknown. Therefore, we knocked down (KD) CTGF expression in human BM-derived MSCs by CTGF short hairpin RNA. CTGF KD MSCs exhibited fivefold lower proliferation compared with control MSCs and had markedly fewer S-phase cells. CTGF KD MSCs differentiated into adipocytes at a sixfold higher rate than controls in vitro and in vivo. To study the effect of CTGF on engraftment of leukemia cells into BM, an in vivo model of humanized extramedullary BM (EXM-BM) was developed in NOD/SCID/IL-2rg(null) mice. Transplanted Nalm-6 or Molm-13 human leukemia cells engrafted at a threefold higher rate in adipocyte-rich CTGF KD MSC-derived EXM-BM than in control EXM-BM. Leptin was found to be highly expressed in CTGF KD EXM-BM and in BM samples of patients with acute myeloid and acute lymphoblastic leukemia, whereas it was not expressed in normal controls. Given the established role of the leptin receptor in leukemia cells, the data suggest an important role of CTGF in MSC differentiation into adipocytes and of leptin in homing and progression of leukemia.

Olanzapine inhibits proliferation, migration and anchorage-independent growth in human glioblastoma cell lines and enhances temozolomide's antiproliferative effect.

The poor prognosis of patients with glioblastoma fuels the search for more effective therapeutic compounds. We previously hypothesised that the neuroleptic olanzapine may enhance antineoplastic effects of temozolomide the standard chemotherapeutic agent used in this disease. This study tested this hypothesis. The anti-proliferative effect of olanzapine was examined by MTT assays and cell count analysis. Soft-agar assays were performed to examine colony-forming ability. In addition, the inhibitory effect of olanzapine on the migratory capacity of U87MG and A172 cells was analyzed by Transwell(®) assays. Moreover, staining for annexin V/propidium iodide or carboxyfluorescein succinimidyl ester was performed prior to flow cytometric analysis in order to better understand the subjacent cellular mechanism. Our initial hypothesis that olanzapine may enhance temozolomide's anti-tumor activity could be confirmed in U87MG and A172 glioblastoma cell lines. Moreover, treatment with olanzapine alone resulted in a marked anti-proliferative effect on U87MG, A172 and two glioma stem-like cells with IC50 values ranging from 25 to 79.9 µM. In U87MG cells, anchorage-independent growth was dose-dependently inhibited. In A172 cells, migration was also shown to be inhibited in a dose-dependent manner. In addition, olanzapine was shown to exert a cell line-dependent pleomorphism with respect to the induction of apoptosis, necrosis and/or cytostasis. Our data show that the neuroleptic olanzapine enhances the anti-tumor activity of temozolomide against glioblastoma cell lines. Moreover, this is the first study to show that olanzapine provides on its own anti-cancer activity in glioblastoma and thus may have potential for repurposing.

Immiscible solvents enabled nanostructure formation for efficient polymer photovoltaic cells.

Organic photovoltaics (OPVs) fabricated via solution processing are an attractive way to realize low cost solar energy harvesting. Bulk heterojunction (BHJ) devices are the most successful design, but their morphology is less controllable. In this manuscript, we describe a simple approach to realize 'ordered' BHJ morphology using two immiscible solvents with different boiling point and a quasi-bilayer approach. Tunable fine structures were demonstrated in poly(3-hexylthiophene) (P3HT) and [6,6]-Phenyl C61 butyric acid methyl ester (PCBM) model systems, and the devices with optimized fine structure showed a 33% efficiency enhancement compared to those with a planar bilayer structure.

Microarray molecular mapping of horses with severe asthma.

BACKGROUND: Severe asthma (SA) in horses, resembling human asthma, is a prevalent, debilitating allergic respiratory condition marked by elevated allergen-specific immunoglobulin E (IgE) against environmental proteins; however, research exploring the exposome's influence on IgE profiles is currently limited but holds paramount significance for diagnostic and therapeutic developments. ANIMALS: Thirty-five sports horses were analyzed, consisting of environmentally matched samples from France (5 SA; 6 control), the United States (6 SA; 6 control), and Canada (6 SEA; 6 control). METHODS: This intentional cross-sectional study investigated the sensitization profiles of SA-affected and healthy horses via serological antigen microarray profiling. Partial least square-discriminant analysis (PLS-DA) was used to identify and rank the importance of allergens for class separation (ie, affected/non-affected) as variable influence of projection (VIP), and allergen with commonality internationally established via frequency analysis. RESULTS: PLS-DA models showed high discriminatory power in predicting SA in horses from Canada (area under the curve [AUC] 0.995) and France (AUC 0.867) but poor discriminatory power in horses from the United States (AUC 0.38). Hev b 5.0101, Cyn D, Der p 2, and Rum cr were the only shared allergens across all geographical groups. CONCLUSIONS AND CLINICAL IMPORTANCE: Microarray profiling can identify specific allergenic components associated with SA in horses, while mathematical modeling of this data can be used for disease classification, highlighting the variability of sensitization profiles between geographical locations and emphasizing the importance of local exposure to the prevalence of different allergens. Frequency scoring analysis can identify important variables that contribute to the classification of SA across different geographical regions.

Cytological diagnosis of exercise-induced pulmonary haemorrhage: Comparison of tracheal wash and bronchoalveolar lavage in standardbred racehorses.

BACKGROUND: Cytology of airway samples is sensitive for diagnosis of exercise-induced pulmonary haemorrhage (EIPH), but the association between tracheal wash (TW) and bronchoalveolar lavage fluid (BALF) is unknown. The objective of this study was to determine whether diagnosis of EIPH, using haemosiderophages/macrophages (H/M) ratio, differs when based on TW or BALF. METHODS: A prospective cross-sectional study was conducted on 102 standardbred horses in training. TW and BALF were collected concomitantly from all horses at rest (at least 24 hours after their last training or race), and their H/M ratios were calculated. Spearman's correlation, Cohen's kappa and Gwet's coefficient tests were performed to evaluate the association between TW and BALF samples. RESULTS: With BALF, 21 horses met the cytological inclusion criteria for an EIPH diagnosis from individual and/or pooled samples. With TW, 20 horses had occasional (H/M < 10%) haemosiderophages, and nine, one and three horses had small (10%-25%), moderate (25%-50%) and large (>50%) proportions, respectively. Poor correlations and inconsistent concordances between TW and BALF were found for H/M ratio. LIMITATIONS: Limitations include the use of a single staining method and the absence of a total haemosiderin score. CONCLUSION: No association between TW and BALF was found for the cytological diagnosis of EIPH. Based on H/M ratio, BALF remains the sample type of choice for cytological diagnosis of EIPH.

Globus Lucidus: A porcine study of an intracranial implant designed to deliver closed, repetitive photodynamic and photochemical therapy in glioblastoma.

OBJECTIVE: Herein we describe initial results in a porcine model of a fully implantable device designed to allow closed, repetitive photodynamic treatment of glioblastoma (GBM). METHODS: This implant, Globus Lucidus, is a transparent quartz glass sphere with light-emitting diodes releasing wavelengths of 630 nm (19.5 mW/cm2), 405 nm (5.0 mW/cm2) or 275 nm (0.9 mW/cm2). 5-aminolevulinic acid was the photosensitizing prodrug chosen for use with Globus Lucidus, hence the implants illuminated at 630 nm or 405 nm. An additional 275 nm wavelength-emittance was included to explore the effects of photochemical therapy (PCT) by ultraviolet (UV) light. Twenty healthy domestic pigs underwent right-frontal craniotomies. The Globus Lucidus device was inserted into a surgically created right-frontal lobe cavity. After postoperative recovery, irradiation for up to 30 min daily for up to 14 d, or continuous irradiation for up to 14.6 h was conducted. RESULTS: Surgery, implants, and repeated irradiations using the different wavelengths were generally well tolerated. Social behavior, wound healing, body weight, and temperature remained unaffected. Histopathological analyses revealed consistent leukocyte infiltration around the intracerebral implant sites with no significant differences between experimental and control groups. CONCLUSION: This Globus Lucidus porcine study prepares the groundwork for adjuvant, long-term, repeated PDT of the GBM infiltration zone. This is the first report of a fully implantable PDT/PCT device for the potential treatment of GBM. A preclinical effectivity study of Globus Lucidus PDT/PCT is warranted and in advanced stages of planning.

Clade-defining mutations in human H1N1 hemagglutinin protein from 2021-2023 have opposing effects on in vitro fitness and antigenic drift.

Seasonal influenza viruses frequently acquire mutations that have the potential to alter both virus replication and antigenic profile. Recent seasonal H1N1 viruses have acquired mutations to their hemagglutinin (HA) protein receptor binding site (RBS) and antigenic sites, and have branched into the clades 5a.2a and 5a.2a.1. Both clades demonstrated improved in vitro fitness compared with the parental 5a.2 clade as measured through plaque formation, infectious virus production in human nasal epithelial cells, and receptor binding diversity. Both clades also showed reduced neutralization by serum from healthcare workers vaccinated in the 2022-23 Northern Hemisphere influenza season compared to the vaccine strain. To investigate the phenotypic impact of individual clade-defining mutations, recombinant viruses containing single HA mutations were generated on a 5a.2 genetic background. The 5a.2a mutation Q189E improved plaque formation and virus replication, but was more efficiently neutralized by serum from individuals vaccinated in 2022-23. In contrast, the 5a.2a mutation E224A and both 5a.2a.1 mutations P137S and K142R impaired aspects of in vitro fitness but contributed significantly to antigenic drift. Surprisingly, the E224A mutation and not Q189E caused broader receptor binding diversity seen in clinical isolates of 5a.2a and 5a.2a.1, suggesting that receptor binding diversity alone may not be responsible for the phenotypic effects of the Q189E mutation. These data document an evolutionary trade-off between mutations that improve viral fitness and those that allow for the evasion of existing host immunity.

The backbone model of the Arabis mosaic virus reveals new insights into functional domains of Nepovirus capsid.

Arabis mosaic virus (ArMV) and Grapevine fanleaf virus (GFLV) are two picorna-like viruses from the genus Nepovirus, consisting in a bipartite RNA genome encapsidated into a 30 nm icosahedral viral particle formed by 60 copies of a single capsid protein (CP). They are responsible for a severe degeneration of grapevines that occurs in most vineyards worldwide. Although sharing a high level of sequence identity between their CP, ArMV is transmitted exclusively by the ectoparasitic nematode Xiphinema diversicaudatum whereas GFLV is specifically transmitted by the nematode X. index. The structural determinants involved in the transmission specificity of both viruses map solely to their respective CP. Recently, reverse genetic and crystallographic studies on GFLV revealed that a positively charged pocket in the CP B domain located at the virus surface may be responsible for vector specificity. To go further into delineating the coat protein determinants involved in transmission specificity, we determined the 6.5 Å resolution cryo-electron microscopy structure of ArMV and used homology modeling and flexible fitting approaches to build its pseudo-atomic structure. This study allowed us to resolve ArMV CP architecture and delineate connections between ArMV capsid shell and its RNA. Comparison of ArMV and GFLV CPs reveals structural differences in the B domain pocket, thus strengthening the hypothesis of a key role of this region in the viral transmission specificity and identifies new potential functional domains of Nepovirus capsid.