As a rapidly developing cell engineering technique, cell electrofusion has been increasingly applied in the field of hybridoma preparation in recent years. However, electrofusion is a certain degree of difficulty to completely replace the polyethylene glycol-mediated cell fusion. The key elements limiting electrofusion in the field of hybridoma preparation are practical complicated. This review summarizes the state of art of cell electrofusion in hybridoma preparation based on recent published literatures, mainly focusing on electrofusion instruments and their components, process control, cell treatment, and process characterization. The review provides new information and insightful commentary, critically important to the promotion of further electrofusion development in the field of hybridoma preparation.

A major challenge in neuroscience is to pinpoint neurobiological correlates of specific cognitive and neuropsychiatric traits. At the mesoscopic level, promising candidates for establishing such connections are brain oscillations that can be robustly recorded as local field potentials with varying frequencies in the hippocampus in vivo and in vitro. Inbred mouse strains show natural variation in hippocampal synaptic plasticity (e.g., long-term potentiation), a cellular correlate of learning and memory. However, their diversity in expression of different types of hippocampal network oscillations has not been fully explored. Here, we investigated hippocampal network oscillations in three widely used inbred mouse strains: C57BL/6J (B6J), C57BL/6NCrl (B6N) and 129S2/SvPasCrl (129) with the particular aim to identify common oscillatory characteristics in inbred mouse strains that show aberrant emotional/cognitive behaviour (B6N and 129) and compare them to “control” B6J strain. First, we detected higher gamma oscillation power in the hippocampal CA3 of both B6N and 129 strains. Second, an increased incidence of hippocampal sharp wave-ripple (SW-R) transients was evident in these strains. Third, we observed prominent differences in the densities of distinct interneuron types and CA3 associative network activity which are indispensable for sustainment of mesoscopic network oscillations. Together, these results supports the notion that in vitro hippocampal network oscillations, similar to classical plasticity read-outs measured in hippocampal slices, can be used as robust reductionist models to study electrophysiological correlates of emotional and cognitive phenotypes. Importantly, we add further evidence to profound physiological differences among inbred mouse strains commonly used in neuroscience research.

In the present paper we investigate the qualitative behaviour of a fractional SEIR model with general incidence rate function and time delay where the fractional derivative is defined in the Caputo sense. The basic reproduction number $\mathcal{R}_{0}$ is derived using the method of next generation matrix and we give a complete study of local stability of both free and endemic equilibrium. Using Liapunov method we prove the global stability of free and endemic equilibrium under some hypotheses on the parameters of the system. Finally to illustrate our results, we use the model to predict the first peak of the COVID-19 epidemic in Algeria.

Background: Acute asthma exacerbations are a common cause for emergency department (ED) visits and hospitalizations in children. Since the outbreak of COVID-19 and the education system closure/ total lockdown in Israel on March 2020, we have noticed a decrease in pediatric ED visits and an increase in hospitalizations of asthma exacerbations. Objective: to examine the patterns of ED visits for asthma exacerbations during COVID-19 outbreak, in comparison to the previous year. Methods: a retrospective study comparing asthma related ED visits and hospitalizations among children aged 2-18 years at a tertiary center in southern Israel. Three time periods were selected: 2020A (pre- lockdown, 1/2/20-14/3/20), 2020B (lockdown, 15/3/20-15/5/20) and 2020 C (post- lockdown, 16/5/20-30/6/20) and compared to the three parallel time periods in 2019. Data regarding demographics, number of ED visits and clinical severity parameters were collected and analyzed. Results: 512 children visited the ED for asthma exacerbation: 273 children during 2019 and 239 children during 2020, with significantly fewer ED visits per day during the lockdown period (1.8 vs 1.43, p<0.001), compared to the parallel period in 2019. Significantly higher hospitalization rate (47.1% vs 33.7%, p=0.05) and longer length of stay (LOS) (3.15 vs 1.9 days, p= 0.03) were observed during the lockdown. Conclusion: lockdown is associated with fewer ED visits for asthma exacerbation, probably due to; reduced exposure to viral infections and environmental allergens, decreased availability of primary physicians and families’ reluctance to arrive to the ED. ED visits during lockdown were characterized by higher hospitalization rate and longer LOS.

Background: Right ventricular failure (RVF) following Left Ventricular Assist Device (LVAD) implantation is associated with worse outcomes. Prediction and early identification of RVF with speckle-tracking echocardiography (STE) has been proposed. Methods: We queried multiple databases for articles reporting on pre-operative/intraoperative global longitudinal strain (GLS) and free-wall strain (FWS) in LVAD recipients. We performed a systematic review and meta-analysis of published literature. The standard mean difference (SMD) in GLS and FWS in patients with and without RVF postoperatively was pooled using random effects model. Results: Fifteen studies, with a total of 967 LVAD recipients were included. There was statistically significant difference in GLS among patients who did and did not develop RVF; SMD= -3.09 (95% CI: -4.62 to -1.57; p-value <0.0001). There was significant difference in FWS between two groups; SMD: -2.75 (95% CI: -3.72 to -1.79; p-value <0.0001). Upon subgroup analysis of imaging modality, transthoracic echocardiography (TTE)-derived GLS and FWS remained predictive for RVF with SMD of -3.97 (95% CI: -5.40 to -2.54; p-value <0.001) and -3.05 (95% CI: -4.11 to -1.99; p-value <0.001), respectively. However, there was no significant difference between RVF and non-RVF groups upon using transesophageal echocardiography (TEE) to assess GLS and FWS. Conclusion: GLS and FWS assessment of the RV by STE is a useful tool to predict postoperative RVF in LVAD recipients. While the predictive role of TTE was robust, the TEE-derived measures seemed to be less predictive. Future studies need to specify the strain cut-off value that can predict the adverse outcome of RVF

Xq28 (involving MECP2) duplication syndrome is a severe neurodevelopmental disorder in males, most females are asymptomatic carriers, but there are phenotypic heterogeneities in the females. Skewed X-chromosome inactivation (XCI) seems to prevent duplicated region activation in asymptomatic females, but it remains controversial. Herein we reported two asymptomatic females (daughter and mother) with interstitial Xq28 duplication. HUMARA and RP2 assays showed that both had complete skewed XCI, the Xq28 duplicated chromosome was inactivated in the daughter, but surprisingly, it was activated in her mother. Interestingly, by combining RNA sequencing and whole-exome sequencing, we confirmed that XIST only expressed in the Xq28 duplication chromosomes of the two females, indicating that the Xq28 duplication chromosomes were inactive. Meanwhile, MECP2 and most XCI genes in the duplicated X-chromosomes were not transcriptionally expressed or upregulated, precluding major clinical phenotypes in the two females, especially the mother. We showed that XCI status detected by RNA sequencing was more relevant for establishing the clinical phenotype of MECP2 duplication females. It suggested there were other factors maintaining the XCI status in addition to DNA methylation, a possible additional inhibition mechanism occured at the transcriptional level in the unmethylated X-chromosome, counter balancing the MECP2 duplication’s detrimental phenotype effects

The dynamic of fractional covid-19 epidemic model with a convex incidence rate is studied in this article. Under Caputo operator, existence and uniqueness for the solutions of the fractional covid-19 epidemic model have been ana- lyzed using xed point theorems. We study all the basic properties and results including local and global stability. We show the global stability of disease free equilibrium using the method of Lyapunov function theory while for disease endemic, we use the method of geometrical approach. Moreover, sensitivity analysis complemented by simulations are performed to determine how changes in parameters affect the dynamical behavior of the system.

Castleman disease is a rare, heterogeneous disorder that driven by proinflammatory responses. Human herpes virus-8 has a major role in pathogenesis of multicentric Castleman disease. There is also a subgroup of cases, human herpes virus-8 negative, idiopathic multicentric Castleman disease. The role of primary immunodeficiencies in idiopathic Castleman disease are poorly described. DOCK8 deficiency is a combined primary immunodeficiency. It has a broad clinic spectrum including atopy, autoimmunity and cancer. We present a 10-year-old, DOCK8 deficient patient. He had giant lobular capillary hemangiomas on his neck, iliac and gluteal regions and multiple lymphadenopathies. Abdominal lymph node pathology revealed hyaline vascular type Castleman disease and human herpes virus-8 staining was negative. His lesions were shown to be infected with orf virus. Our case is the first case to relate idiopathic multicentric Castleman disease and DOCK8 deficiency; also, very unusual presentation of orf virus infection in humans.

Background and Objective: With improved survival in neonates with meconium aspiration syndrome (MAS), the focus is currently on mitigating the morbidities. The objective of this study was to predict factors determining prolonged hospital stay in neonates with MAS. Materials and methods: It was a retrospective cohort from five centres of south India between 2018 and 2020. Neonates ≥35 weeks of gestation admitted to NICU with the diagnosis of MAS and requiring oxygen beyond 24 hours of life were included in the study. The morbidities in the neonates with stay ≤7 days (short stay) were compared with >7 days (prolonged stay). Logistic regression by the backward stepwise method was used for predictive score creation. Results: Out of 347 neonates with MAS discharged home, 103 (29%) had a short stay and 244 (71%) had prolonged stay. The primary support beyond O2 (CPAP/MV) (42% vs 83%, p<0.001), FiO2 at 1hr>30% (45% vs 87%, p<0.001), HIE stage 2 or 3 (2% vs 27%, p<0.001), moderate-severe PPHN (3% vs 31%, p<0.001) were independent factors associated with prolonged stay on logistic regression. A prediction model was devised using weighted scores of these four associated morbidities. The clinical score thus developed had 83% sensitivity, 68% specificity for the prediction of prolonged stay [AUC- 82, 95% CI (78-87), p<0.001]. Conclusion: More than two-thirds of neonates with MAS had prolonged stay. The primary support beyond oxygen, Fio2 requirement >30%, Moderate to severe PPHN, HIE stage 2 or 3 were predictive of prolonged stay in neonates with MAS.

Quantum chemical calculations were carried out to establish the half-sandwich structural behaviour between heavier group-14 elements (Si-Pb) and neutral Be3 ring. The proposed complexes are found to be global minima on the potential energy surface. Quantum chemical investigation revealed that the complexes found possess high bond dissociation energy and also favorable thermodynamics of their formation. The complexes were also found to possess significant aromatic behaviour. In addition, the half-sandwich complexes were found to possess promising chemical properties to be useful for potential H2 storage material under reversible conditions.

Landscape anthropization has been identified as one of the main drivers of pathogen emergence worldwide, facilitating pathogen spillover between domestic species and wildlife. The present study investigated Carnivore protoparvovirus-1 (CPPV) infection using molecular methods in 98 free-ranging wild guignas (Leopardus guigna) and 262 co-occurring owned, free-roaming rural domestic cats. We also assessed landscape anthropization variables as potential drivers of infection. CPPV DNA was detected in guignas across their entire distribution range, with observed prevalence of 13.3% (real-time PCR) and 9% (conventional PCR) in guignas, and 6.1% (conventional PCR) in cats. Prevalence in guigna did not vary depending on age, sex, study area or landscape variables. Prevalence was higher in juvenile cats (16.7%) than in adults (4.4%). Molecular characterization of the virus by amplification and sequencing of almost the entire vp2 gene (1746 bp) from one guigna and five domestic cats was achieved, showing genetic similarities to canine parvovirus 2c (CPV-2c) (one guigna and one cat), feline panleukopenia virus (FPV) (one cat), CPV-2 (no subtype identified) (two cats), CPV-2a (one cat). The CVP-2c-like sequence found in a guigna clustered together with domestic cat and dog CPV-2c sequences from South America, suggesting possible spillover from a domestic to a wild species as the origin of infection in guigna. No clinical signs of disease were found in PCR-positive animals except for the CPV-2c-infected guigna, which had hemorrhagic diarrhea and died a few days after arrival at a wildlife rescue center. Our findings reveal widespread presence of Carnivore protoparvovirus-1 across the guigna distribution in Chile and suggest that virus transmission potentially occurs from domestic to wild carnivores, causing severe disease and death in susceptible wild guignas.

During the COVID-19 pandemic, multiple skin manifestations have been described. These include an urticarial rash, morbilliform rash, maculopapular rash, vascular lesions, and varicella-like eruptions. A 30-year-old woman presented with a mild cough, then hives and pruritic rash for 3 days, followed by fever, dyspepsia, and throat pain for one day.

Classical absent pulmonary valve syndrome (APVS) with tetralogy of Fallot (TOF) is a rare congenital cardiac anomaly commonly associated with absent of patent ductus arteriosus (PDA), which is mostly diagnosed after 20 weeks of gestation by fetal echocardiography. This case of APVS with TOF was suspected at 13 weeks of gestation and diagnosed at 14 weeks of gestation with an obvious PDA. The pulmonary arterial trunk and the branches did not dilated obviously. 15 days later, the PDA narrowed down with the discovery of pulmonary artery ectasia at the same time. This progress indicated that the absent of PDA is not necessary for the survival of APVS with TOF in utero, in contrast, the absent or restriction of PDA may be nothing less than adaptation to the disease. Fetal autopsy confirmed the accuracy of fetal echocardiography. Chromosome microarray analysis (CMA) showed 20p12 deletion in this fetus, which is rare among TOF cases.

Reply to Morais-Almeida.To the Editor,We appreciate Dr. Morais-Almeida’s comments 1 about our Letter to the Editor, presenting additional literature about asthma prevalence in severe COVID-19 patients and highlighting data that contrasts our hypothesis that asthma, particularly type 2 asthma, may be protective against severe disease.The data that protection may be dependent on type 2 immunity is derived from the higher percentage of asthmatics being atopic2, also reflected in the series of ~2,500 patients regularly followed up in our Allergy Unit. Yu et al. 3 provided preliminary evidence about this in a single-center retrospective study, where COVID-19 atopic patients had less severe infections, milder lung damage compared to age- and gender- matched COVID-19 controls.ACE-2, the SARS-CoV-2 receptor, is linked to type 1 and 2 interferon signatures, and found to be overexpressed in type 2-low asthmatics4. Nevertheless, different outcomes in distinct asthma phenotypes still need to be addressed in COVID-19 studies.Besides Italy and China, reports from Russia 5 on ~1,300 intensive care unit patients with SARS-CoV-2 infection confirm the observation of a low prevalence of chronic lung diseases (i.e. asthma as well as COPD).Although preliminary data on the first COVID-19 cases in the US6 seem to contrast these observations, the higher prevalence of asthma in US COVID-19 hospitalised patients should be considered alongside a higher overall prevalence in these countries compared to Europe and China, as well as on the influence of other comorbidities (i.e. obesity) and host factors (i.e. age, race: 33% were non-Hispanic black patients in the study by Garg et al.) impacting COVID-19 outcomes. Another report from Sweden 7highlights the association between severe asthma and severe COVID-19.The severe asthma phenotype is often characterized by mixed granulocytic populations (neutrophilic and eosinophilic), prevalent type 1 inflammation, increased IFN-γ levels in the airways and ineffectiveness of ICS. This severe phenotype by itself, although accounting for less than 5% of asthmatic patients, would justify the CDC (and other institutions) including asthma as a risk factor for COVID-19. Data from the UK 8, apart from confirming the role of additional comorbidities, draw attention to the recent use of oral steroids, which, indeed, may be a clue for uncontrolled and/or severe asthma.Uncontrolled asthma is a risk factor for viral exacerbations and hospitalizations and we embrace the opportunity to stress the importance of optimal adherence to asthma controlling medications, regular follow-up and specialist-assessment of disease activity. Moreover, treatable comorbidities, which may impair asthma control, should always be managed. Promoting vaccination for preventable respiratory infections (i.e. Influenza and Pneumococcal pneumonia) is also advisable. Future studies may help better distinguishing the impact of different asthma phenotypes and comorbidities on COVID-19 outcome.Carli G.1, Cecchi L.1, Stebbing J.2, Parronchi P.3, Farsi A.11 SOS Allergy and Clinical Immunology, USL Toscana Centro, Prato Italy2 Department of Surgery and Cancer, Imperial College London, London SW7 2AZ, UK3 Department of Experimental and Clinical Medicine, University of Florence, Florence, Italy

Background Understanding the cellular host factors that promote and inhibit viral entry is important for identifying viral countermeasures. CRISPR whole genome screens can be used to rapidly discover host factors that contribute to or impair viral entry. However, when using the live viruses and cellular lethality for selection, these screens can identify an overwhelming number of genes without specificity for the stage of the viral infection cycle. New screening methods are needed to identify host machinery contributing to specific steps of viral infection. Here, we developed a CRISPR whole genome screen and counter screen strategy based on a pseudoviral platform that allowed identification of genes specific to SARS-CoV-2 spike and vesicular stomatitis virus glycoprotein VSV-G mediated entry. Methods To focus the screen onto the entry step, we used non-lytic fluorescent reporters in combination with a comparative counter screen strategy to distinguish host genes affecting the pseudoviral reporter from those unique to envelope-mediated entry. Screening of SARS-CoV-2 spike and VSV-G on the same lentiviral pseudovirus allowed identification of entry-specific genes relative to genes associated with retro-transcription, integration, and reporter expression from the lentiviral pseudovirus. Second, a Cre-Gag fusion protein in the pseudovirus was used to bypass retro-transcription and integration by directly activating a floxed GFP reporter upon entry to reduce the number of gene hits and increase specificity for viral entry. Results Our approach correctly identified SARS-CoV-2 and VSV-G receptors ACE2 and LDLR, respectively and distinguished genes associated with retroviral reporter expression from envelope-mediated entry. Moreover, the CRE-Gag fusion/flox reporter increased the screen specificity for viral entry associated genes. Validation of a few hits demonstrates that this approach distinguishes envelope-specific host factors from genes affecting reporter expression. Conclusion Overall, this approach provides a new strategy for identifying host genes influencing viral entry without the confounding complexity of live-viral screens which produce long gene lists associated with all aspects of viral pathogenesis and replication. This approach provides a pathway for increasing the specificity of CRISPR whole genome screens for identifying host genes contributing to specific steps in viral infection.

High dimensional models typically require a large computational overhead for multiphysics applications, which hamper their use for broad-sweeping domain interrogation. Herein, we develop a modeling framework to capture the through-plane fluid dynamic response of electrodes and flow fields in a redox flow cell, generating a computationally inexpensive two-dimensional (2D) model. We leverage a depth averaging approach that also accounts for variations in out-of-plane fluid motion and departures from Darcy’s law that arise from averaging across three-dimensions (3D). Our Resulting depth-averaged 2D model successfully predict the fluid dynamic response of arbitrary in-plane flow field geometries, with discrepancies of < 5% for both maximum velocity and pressure drop. This corresponds to reduced computational expense, as compared to 3D representations (< 1% of duration and 10% of RAM usage), providing a platform to screen and optimize a diverse set of cell geometries.

Spearing mantis shrimps are aggressive crustaceans using specialized appendages with sharp spikes to capture fishes with a fast movement. Each spike is a biological tool that have to combine high toughness, as required by the initial impact with the victim, with high stiffness and strength, to ensure sufficient penetration while avoid breaking. We performed a multimodal analysis to uncover the design strategies of this harpoon based on chitin. We found that the spike is a slightly hooked hollow beam with the outer surface decorated by serrations and grooves to enhance cutting and interlocking. The cuticle of the spike resembles a multilayer composite: an outer heavily mineralized, stiff and hard region (with average indentation modulus and hardness of 68 and 3 GPa), providing high resistance to contact stresses, is combined with a less mineralized region, which occupies a large fraction of the cuticle (up to 50%) and features parallel fibers oriented longitudinally, enhancing stiffness and strength. A central finding of our work is the presence of a tiny interphase (less than 10 μm in width) based on helical fibers and showing a spatial modulation in mechanical properties, which has the critical task to integrate the stiff but brittle outer layer with the more compliant highly anisotropic parallel fiber region. We highlighted the remarkable ability of this helicoidal region to stop nanoindentation-induced cracks. Using three-dimensional multimaterial printing to prototype spike-inspired composites, we showed how the observed construction principles can not only hamper damage propagation between highly dissimilar layers (resulting in composites with the helical interphase absorbing 50% more energy than without it) but can also enhance resistance to puncture (25% increase in the force required to penetrate the composites with a blunt tool). Such findings may provide guidelines to design lightweight harpoons relying on environmentally friendly and recyclable building blocks.

The rapid evolution of protein structure prediction tools has significantly broadened access to protein structural data. Although predicted structure models have the potential to accelerate and impact fundamental and translational research significantly, it is essential to note that they are not validated and cannot be considered the ground truth. Thus, challenges persist, particularly in capturing protein dynamics, predicting multi-chain structures, interpreting protein function, and assessing model quality. Interdisciplinary collaborations are crucial to overcoming these obstacles. Databases like the AlphaFold Protein Structure Database, the ESM Metagenomic Atlas, and initiatives like the 3D-Beacons Network provide FAIR access to these data, enabling their interpretation and application across a broader scientific community. Whilst substantial advancements have been made in protein structure prediction, further progress is required to address the remaining challenges. Developing training materials, nurturing collaborations, and ensuring open data sharing will be paramount in this pursuit. The continued evolution of these tools and methodologies will deepen our understanding of protein function and accelerate disease pathogenesis and drug development discoveries.