Understanding the Representation of Asians and Asian Americans Within Academic Otolaryngology Leadership.

OBJECTIVE: Elucidate the representation of Asian and Asian Americans in academic otolaryngology and the influence of race on promotion and leadership opportunities. STUDY DESIGN: Retrospective analysis of the Association of American Medical Colleges Faculty Administrative Management Online User System. SETTING: Full-time otolaryngology faculty from all US medical schools from 2020 to 2023. METHODS: Faculty demographics, tenure, and rank were collected. Descriptive statistics, Fischer's exact test, Rank Equity Index (REI), and multivariable logistic and ordinal regressions were used to characterize our cohort and assess the impact of race on academic advancement and leadership, defined as promotion to tenure or full professorship. RESULTS: Asians comprised 20.53% of 9056 faculty over 4 years. Asians were most likely to hold tenure-eligible positions (n = 600, 30.74%) but were significantly less likely than non-Asians to be tenured (43.00% vs 48.65%, P = .015). Asians were slightly above parity in promotion from assistant to associate professor (REI = 1.09) but below parity in promotion from associate professor to professor (REI = 0.78). Relative to whites, Hispanics, and African Americans, Asians reported the lowest associate/professor and assistant/professor REIs. On multivariable regressions, Asian race was not associated with decreased odds of tenure-eligible positions but was associated with decreased odds of tenure (odds ratio [OR] = 0.77, 95% confidence interval [CI] = [0.64-0.93]) and rank promotion (OR = 0.82, 95% CI = [0.74-0.90]). CONCLUSION: Despite strong overall representation in otolaryngology, Asians are less likely to receive promotion, tenure, or full professorship relative to other racial groups. Future efforts should emphasize equitable advancement opportunities to ensure a diverse otolaryngology leadership.

Bilateral Vocal Fold Motion Impairment Associated With Diffuse Idiopathic Skeletal Hyperostosis.

Objective: To describe the clinical courses and interventions of symptomatic patients with bilateral vocal fold motion impairment (BVFMI) attributed to diffuse idiopathic skeletal hyperostosis (DISH). Study Design: Retrospective cohort study. Setting: Single Institution Academic Health Center. Methods: Retrospective chart review of patients ≥18 years old evaluated and treated for symptomatic BVFMI secondary to DISH between February 2021 and March 2023. A literature review was conducted. Results: A total of 4 cases were identified. All patients were male and had symptomatic BVFMI attributed to cervical spine DISH, as seen on imaging. Symptoms ranged from life-threatening dyspnea to breathy dysphonia in addition to dysphagia. Each patient was offered surgery for DISH. Two patients underwent osteophyte removal at the C5-C6 level with improved vocal fold (VF) mobility, breathing, and voice quality. Two patients elected serial observation as voice, swallow, and airway symptoms were manageable. The literature review showed a male-dominant (100%) presentation with an average of 70 years of age. Hypertension (45%) and diabetes mellitus (36%) were the most common comorbidities. Most patients were treated surgically (55%). Conclusion: Both surgical and conservative interventions may be considered for symptomatic relief and improvement in VF mobility on a patient-to-patient basis. Further study is warranted to investigate the etiology and treatment outcomes in these cases.

Factors associated with loss and recovery of smell and taste after COVID-19 infection.

Objective: To identify predictors associated with loss and recovery of smell and taste after COVID-19 infection. Methods: The Integrated Public Use Microdata Series (IPUMS) 2021 National Health Interview Series (NHIS) database was used to analyze factors associated with loss and recovery of smell and taste in respondents who had a previous COVID-19 infection. Significant variables from univariate analysis were included in a stepwise backward regression model to identify independent predictors. Results: Of the 3844 individuals who answered yes to having contracted COVID-19, 51.1% and 48.9% reported losing smell and taste, respectively. 95.7% recovered smell and 97% recovered taste. Predictors associated with higher odds of reporting a loss of smell included younger age (odds ratio [OR] 0.98; 95% confidence interval [CI] 0.98-0.99), female sex (OR 1.38; CI 1.17-1.63), use of e-cigarettes (OR 1.59; CI 1.25-2.02), and Mexican ethnicity (OR 1.61; CI 1.22-2.11). Predictors of taste loss were younger age (OR 0.98; CI 0.98-0.99), female sex (OR 1.31; CI 1.08-1.58), and higher BMI (OR 1.02; CI 1.00-1.04). Female sex was associated with decreased odds of reporting a recovery of smell (OR 0.74; CI 0.59-0.92) and taste (OR 0.54; CI 0.42-0.69). Black/African American race (OR 1.44; CI 1.03-2.03) and non-Mexican Hispanic ethnicities (OR 1.55; CI 1.02-2.34) were associated with an increased likelihood of reporting the recovery of smell. Conclusion: Various factors may be associated with the loss and recovery of smell and taste after COVID-19 infections. Clinicians may use this information to better counsel patients with these symptoms. Level of evidence: 4.

Fecal microbiota of diarrheic calves: Before, during, and after recovering from disease.

BACKGROUND: It is unknown if gastrointestinal dysbiosis in diarrheic calves causes disease or is a consequence of the disease. OBJECTIVES: Describe the fecal microbiota of calves before, during, and after recovering from diarrhea. ANIMALS: Fifteen female Holstein calves of 0 to 21 days old from a single farm. Seven calves remained healthy throughout the study, and 8 developed diarrhea on Day 14. METHODS: Longitudinal cohort study. Microbiota composition was characterized by amplifying the V4 region of the 16S rRNA gene. RESULTS: Diversity (Shannon index) increased with age in healthy and diarrheic calves from Day 3 to 21, but diarrheic calves had a lower diversity on the day diarrhea was first observed (Day 14). By Day 21, diversity increased in calves that recovered from diarrhea and was not significantly different from that of their healthy counterparts (P > .05). Weighted UniFrac distance showed significant differences in the fecal microbiota between diarrheic and healthy calves at Day 14 of age (PERMANOVA, P < .05), but not before or after diarrhea (PERMANOVA, P > .05). Lactobacillus, Clostridium Sensu Stricto 1, and Collinsella were differentially abundant on Day 10 in calves that developed diarrhea on Day 14 (P < .05). CONCLUSION AND CLINICAL IMPORTANCE: The fecal microbiota of healthy and diarrheic calves evolved similarly during the first 10 days of age but differed significantly on the day of onset of diarrhea. Enriching Lactobacillus, Clostridium Sensu Stricto 1, and Collinsella before diarrhea onset could have been contributed to the development of diarrhea.

Endothelial glycocalyx degradation in critically ill foals.

BACKGROUND: Endothelial glycocalyx (EG) degradation occurs in septic humans and EG products can be used as biomarkers of endothelial injury. Information about EG biomarkers and their association with disease severity is lacking in hospitalized foals. OBJECTIVES: Measure serum syndecan-1 (SDC-1), heparan sulfate (HS), angiopoietin-2 (ANG-2), aldosterone (ALD), and plasma atrial natriuretic peptide (ANP) concentrations and to determine their association with disease severity and death in hospitalized foals. ANIMALS: Ninety foals ≤3 days old. METHODS: Prospective, multicenter, longitudinal study. Foals were categorized into hospitalized (n = 74; 55 septic; 19 sick nonseptic) and 16 healthy foals. Serum ([SDC-1], [HS], [ANG-2], [ALD]) and plasma (ANP) were measured over 72 hours using immunoassays. RESULTS: Serum ([SDC-1], [HS], [ANG-2], [ALD]) and plasma (ANP) were significantly higher in hospitalized and septic than healthy foals (P < .05). Serum (ANG-2) and plasma (ANP) were significantly higher in hospitalized nonsurvivors than in survivors (P < .05). On admission, hospitalized foals with serum (HS) > 58.7 ng/mL had higher odds of nonsurvival (odds ratio [OR] = 6.1; 95% confidence interval [CI] = 1.02-36.7). Plasma (ANP) >11.5 pg/mL was associated with the likelihood of nonsurvival in hospitalized foals (OR = 7.2; 95% CI = 1.4-37.4; P < .05). Septic foals with serum (ANG-2) >1018 pg/mL on admission had higher odds of nonsurvival (OR = 6.5; 95% CI =1.2-36.6; P < .05). CONCLUSION AND CLINICAL IMPORTANCE: Critical illness in newborn foals is associated with EG degradation and injury, and these biomarkers are related to the severity of disease on admission and the outcome of sick foals.

Interventions for the prevention of weight gain during festive and holiday periods in children and adults: A systematic review.

Some periods during the year, such as festive and summer holiday periods, have been associated with weight gain. We aimed to assess the effect of interventions for the prevention of body weight gain during festive and holiday periods in children and adults. A systematic search was conducted in six databases and supplementary sources until January 4, 2023. We included randomized controlled trials (RCTs), cluster-RCTs, and non-RCTs. Our primary outcome measure was the change in body weight in adults or the change in BMI z-score or BMI percentile in children and adolescents. From 4216 records, 12 primary studies (from 22 reports) met the inclusion criteria-10 from the United States, one from the United Kingdom, and one from Chile. Two studies had a low risk of bias, two moderate, seven high, and one critical risk of bias. The meta-analysis in children included four of seven studies during the summer holidays (six interventions) and showed a mean difference in BMI z-score favoring the intervention group (-0.06 [95% CI -0.10, -0.01], p = 0.01, I2 = 0%, very low certainty evidence). The meta-analysis in adults included five studies during festive periods with a mean difference in weight favoring the intervention group (-0.99 kg [95% CI -2.15, 0.18], p = 0.10, I2 = 89%, very low certainty evidence). This review has highlighted potential interventions to prevent the increase in body weight during holiday periods. More work is needed to improve the quality of the evidence and to extend it to countries outside of the United States and United Kingdom and to the adolescent population.

New Highly Selective Antivirals for Chikungunya Virus identified from the Screening of a Drug-Like Compound Library.

Chikungunya fever is a mosquito-borne disease caused by Chikungunya virus (CHIKV). Treatment of CHIKV infections is currently supportive and does not limit viral replication or symptoms of persistent chronic arthritis. Although there are multiple compounds reported as antivirals active against CHIKV in vitro, there are still no effective and safe antivirals. Thus, active research aims at the identification of new chemical structures with antiviral activity. Here, we report the screen of the Pandemic Response Box library of small molecules against a fully infectious CHIKV reporter virus. Our screening approach successfully identified previously reported CHIKV antiviral compounds within this library and further expanded potentially active hits, supporting the use of reporter-virus-based assays in high-throughput screening format as a reliable tool for antiviral drug discovery. Four molecules were identified as potential drug candidates against CHIKV: MMV1634402 (Brilacidin) and MMV102270 (Diphyllin), which were previously shown to present broad-spectrum antiviral activities, in addition to MMV1578574 (Eravacycline), and the antifungal MMV689401 (Fluopicolide), for which their antiviral potential is uncovered here.

Early exposure to glyphosate during larval development induces late behavioural effects on adult honey bees.

As the most abundant pollinator insect in crops, Apis mellifera is a sentinel species of the pollinator communities. In these ecosystems, honey bees of different ages and developmental stages are exposed to diverse agrochemicals. However, most toxicological studies analyse the immediate effects during exposure. Late effects during adulthood after early exposure to pollutants during larval development are poorly studied in bees. The herbicide glyphosate (GLY) is the most applied pesticide worldwide. GLY has been detected in honey and beebread from hives near treated crops. Alterations in growth, morphogenesis or organogenesis during pre-imaginal development could induce late adverse effects after the emergence. Previous studies have demonstrated that GLY alters honey bee development, immediately affecting survival, growth and metabolism, followed by late teratogenic effects. The present study aims to determine the late impact on the behaviour and physiology of adult bees after pre-imaginal exposure to GLY. For that, we reared brood in vitro or in the hive with sub-chronic exposure to the herbicide with the average detected concentration in hives. Then, all newly emerged bees were reared in an incubator until maturity and tested when they became nurse-aged bees. Three behavioural responses were assessed as markers of cognitive and physiological impairment. Our results show i) decreased sensitivity to sucrose regardless of the rearing procedure, ii) increased choice latency and locomotor alterations during chemotaxis and iii) impaired associative learning. These late toxicity signs could indicate adverse effects on task performance and colony efficiency.

Molecular testing for equine herpesviruses 1 (EHV-1) in healthy postpartum broodmares.

Objective: Our objective was to determine whether equine herpesviruses 1 (EHV-1) viral nucleic acids could be detected immediately after foaling from nasal and vaginal swabs, whole blood, and placental tissue of healthy mares. Animals procedure and results: Nasal and vaginal swabs, EDTA blood, and placental tissue (296 samples) were collected from 74 clinically healthy postpartum broodmares within 24 h after giving birth to live, clinically healthy foals. All samples were tested (PCR) for nucleic acids of neuropathogenic and non-neuropathogenic strains of EHV-1, and all were negative. Conclusion and clinical relevance: As EHV-1 was not detected in the immediate postpartum period in healthy mares with uncomplicated foaling, we inferred that EHV-1-positive samples from aborting mares and/or EHV-1 detection in fetal membranes indicate EHV-1-associated abortion.

Tests moléculaires pour l'herpèsvirus équin 1 (EHV-1) chez des juments poulinières post-partum en bonne santé. Objectif: Notre objectif était de déterminer si les acides nucléiques viraux de l'herpèsvirus équin 1 (EHV-1) pouvaient être détectés immédiatement après la mise bas à partir de prélèvements nasaux et vaginaux, de sang total et de tissus placentaires de juments saines. Animaux procédure et résultats: Des écouvillons nasaux et vaginaux, du sang EDTA et du tissu placentaire (296 échantillons) ont été prélevés sur 74 juments poulinières post-partum cliniquement saines dans les 24 heures suivant la naissance de poulains vivants et cliniquement sains. Tous les échantillons ont été testés (PCR) pour les acides nucléiques des souches neuropathogènes et non-neuropathogènes de l'EHV-1, et tous se sont révélés négatifs. Conclusion et pertinence clinique: Comme l'EHV-1 n'a pas été détecté dans la période post-partum immédiate chez des juments en bonne santé avec un poulinage sans complication, nous avons déduit que les échantillons positifs pour l'EHV-1 provenant de juments qui ont avorté et/ou la détection de l'EHV-1 dans les membranes foetales indiquent un avortement associé à l'EHV-1.(Traduit par Dr Serge Messier).

Integrating Dynamic Network Analysis with AI for Enhanced Epitope Prediction in PD-L1:Affibody Interactions.

Understanding binding epitopes involved in protein-protein interactions and accurately determining their structure are long-standing goals with broad applicability in industry and biomedicine. Although various experimental methods for binding epitope determination exist, these approaches are typically low throughput and cost-intensive. Computational methods have potential to accelerate epitope predictions; however, recently developed artificial intelligence (AI)-based methods frequently fail to predict epitopes of synthetic binding domains with few natural homologues. Here we have developed an integrated method employing generalized-correlation-based dynamic network analysis on multiple molecular dynamics (MD) trajectories, initiated from AlphaFold2Multimer structures, to unravel the structure and binding epitope of the therapeutic PD-L1:Affibody complex. Both AlphaFold2 and conventional molecular dynamics trajectory analysis were ineffective in distinguishing between two proposed binding models, parallel and perpendicular. However, our integrated approach, utilizing dynamic network analysis, demonstrated that the perpendicular mode was significantly more stable. These predictions were validated using a suite of experimental epitope mapping protocols, including cross-linking mass spectrometry and next-generation sequencing-based deep mutational scanning. Conversely, AlphaFold3 failed to predict a structure bound in the perpendicular pose, highlighting the necessity for exploratory research in the search for binding epitopes and challenging the notion that AI-generated protein structures can be accepted without scrutiny. Our research underscores the potential of employing dynamic network analysis to enhance AI-based structure predictions for more accurate identification of protein-protein interaction interfaces.

Kalanchoe tomentosa: Phytochemical Profiling, and Evaluation of Its Biological Activities In Vitro, In Vivo, and In Silico.

In this work, the leaves of K. tomentosa were macerated with hexane, chloroform, and methanol, respectively. The phytochemical profiles of hexane and chloroform extracts were unveiled using GC/MS, whereas the chemical composition of the methanol extract was analyzed using UPLC/MS/MS. The antibacterial activity of extracts was determined against gram-positive and gram-negative strains through the minimal inhibitory concentration assay, and in silico studies were implemented to analyze the interaction of phytoconstituents with bacterial peptides. The antioxidant property of extracts was assessed by evaluating their capacity to scavenge DPPH, ABTS, and H2O2 radicals. The toxicity of the extracts was recorded against Artemia salina nauplii and Caenorhabditis elegans nematodes. Results demonstrate that the hexane and chloroform extracts contain phytosterols, triterpenes, and fatty acids, whereas the methanol extract possesses glycosidic derivatives of quercetin and kaempferol together with sesquiterpene lactones. The antibacterial performance of extracts against the cultured strains was appraised as weak due to their MIC90 values (>500 µg/mL). As antioxidants, treatment with extracts executed high and moderate antioxidant activities within the range of 50-300 µg/mL. Extracts did not decrease the viability of A. salina, but they exerted a high toxic effect against C. elegans during exposure to treatment. Through in silico modeling, it was recorded that the flavonoids contained in the methanol extract can hamper the interaction of the NAM/NAG peptide, which is of great interest since it determines the formation of the peptide wall of gram-positive bacteria. This study reports for the first time the biological activities and phytochemical content of extracts from K. tomentosa and proposes a possible antibacterial mechanism of glycosidic derivatives of flavonoids against gram-positive bacteria.

Molecular basis of RNA recombination in the 3'UTR of chikungunya virus genome.

Chikungunya virus (CHIKV) is a rapidly spreading re-emergent virus transmitted from mosquitoes to humans. The emergence of epidemic variants has been associated with changes in the viral genome, such as the duplication of repeated sequences in the 3' untranslated region (UTR). Indeed, blocks of repeated sequences seemingly favor RNA recombination, providing the virus with a unique ability to continuously change the 3'UTR architecture during host switching. In this work, we provide experimental data on the molecular mechanism of RNA recombination and describe specific sequence and structural elements in the viral 3'UTR that favor template switching of the viral RNA-dependent RNA polymerase on the 3'UTR. Furthermore, we found that a 3'UTR deletion mutant that exhibits markedly delayed replication in mosquito cells and impaired transmission in vivo, recombines in reference laboratory strains of mosquitoes. Altogether, our data provide novel experimental evidence indicating that RNA recombination can act as a nucleic acid repair mechanism to add repeated sequences that are associated to high viral fitness in mosquito during chikungunya virus replication.

Infection and chronic disease activate a systemic brain-muscle signaling axis.

Infections and neurodegenerative diseases induce neuroinflammation, but affected individuals often show nonneural symptoms including muscle pain and muscle fatigue. The molecular pathways by which neuroinflammation causes pathologies outside the central nervous system (CNS) are poorly understood. We developed multiple models to investigate the impact of CNS stressors on motor function and found that Escherichia coli infections and SARS-CoV-2 protein expression caused reactive oxygen species (ROS) to accumulate in the brain. ROS induced expression of the cytokine Unpaired 3 (Upd3) in Drosophila and its ortholog, IL-6, in mice. CNS-derived Upd3/IL-6 activated the JAK-STAT pathway in skeletal muscle, which caused muscle mitochondrial dysfunction and impaired motor function. We observed similar phenotypes after expressing toxic amyloid-ß (Aß42) in the CNS. Infection and chronic disease therefore activate a systemic brain-muscle signaling axis in which CNS-derived cytokines bypass the connectome and directly regulate muscle physiology, highlighting IL-6 as a therapeutic target to treat disease-associated muscle dysfunction.

Dengue virus preferentially uses human and mosquito non-optimal codons.

Codon optimality refers to the effect that codon composition has on messenger RNA (mRNA) stability and translation level and implies that synonymous codons are not silent from a regulatory point of view. Here, we investigated the adaptation of virus genomes to the host optimality code using mosquito-borne dengue virus (DENV) as a model. We demonstrated that codon optimality exists in mosquito cells and showed that DENV preferentially uses nonoptimal (destabilizing) codons and avoids codons that are defined as optimal (stabilizing) in either human or mosquito cells. Human genes enriched in the codons preferentially and frequently used by DENV are upregulated during infection, and so is the tRNA decoding the nonoptimal and DENV preferentially used codon for arginine. We found that adaptation during single-host passaging in human or mosquito cells results in the selection of synonymous mutations towards DENV's preferred nonoptimal codons that increase virus fitness. Finally, our analyses revealed that hundreds of viruses preferentially use nonoptimal codons, with those infecting a single host displaying an even stronger bias, suggesting that host-pathogen interaction shapes virus-synonymous codon choice.

Reproductive Tract Microbiota of Mares.

The female reproductive tract microbiota is a complex community of microorganisms that might be crucial in maintaining a healthy reproductive environment. Imbalances in the bacterial community (dysbiosis) and the reduction of beneficial organisms and pathogen proliferation are associated with disease. Endometritis is a common cause of fertility problems in mares, and it is still challenging to diagnose and treat based on routine culture results of certain microorganisms. Although high-throughput sequencing studies provide helpful information regarding the composition of the reproductive tract microbiota in mares, there are still challenges in defining a "normal" microbiota. The primary objective of this literature review is to summarize the current knowledge regarding the microbiota present in the reproductive tract of mares, including the vagina, cervix, and uterus. The second objective is to describe the relevant factors that can impact the reproductive microbiota of mares, including the estrous cycle stage, the type of species (genera) investigated, season, and geographic location. The rationality of identifying the normal microbiota in the reproductive tract of a mare will likely aid in understanding the impact of the microbiota on the host's reproductive health and contribute to the treatment and prevention of equine sub and infertility issues.

Vaginal and Uterine Microbiota of Healthy Maiden Mares during Estrus.

This descriptive cross-sectional study compared the microbiota of the uterus, vagina, clitoral fossa (CF), and perineal skin in healthy maiden mares during estrus. Twelve synchronized, healthy maiden mares (3-4 years old) from one single recipient mare herd were included. Microbial communities were characterized by amplifying the V3-V4 region of the 16S rRNA gene using the Illumina MiSeq platform. The uterine and vaginal microbiota had significantly lower richness (Chao-1) than the skin (p < 0.05). The uterine and vagina bacterial composition was similar in presence and abundance and could be differentiated from that of the CF and perineal skin. The microbial composition (Jaccard and Bray-Curtis distances) significantly differed across body-site locations (p < 0.05), which explained approximately 14% and 19% of the variation in microbial composition for Jaccard and Bray-Curtis distances, respectively. Firmicutes, Actinobacteria, Proteobacteria, and Bacteroidetes were the dominant taxa in the uterus and vagina, with higher proportions of Proteobacteria in the vaginal samples compared to the uterine samples. Streptococcaceae and Staphylococcaceae were present in high abundance in the uterine and vaginal samples, while Lactobacillaceae were not (<10%). We demonstrate that the uterine and vaginal microbiota of healthy maiden mares during estrus is similar but both distinct from that of the CF and perineal skin.

Allogeneic B cell immunomodulatory therapy in amyotrophic lateral sclerosis.

Amyotrophic lateral sclerosis (ALS) is an orphan neurodegenerative disease. Immune system dysregulation plays an essential role in ALS onset and progression. Our preclinical studies have shown that the administration of exogenous allogeneic B cells improves outcomes in murine models of skin and brain injury through a process termed pligodraxis, in which B cells adopt an immunoregulatory and neuroprotective phenotype in an injured environment. Here, we investigated the effects of B-cell therapy in the SOD1G93A mouse preclinical model of ALS and in a person living with ALS. Purified splenic mature naïve B cells from haploidentical donor mice were administered intravenously in SOD1G93A mice for a total of 10 weekly doses. For the clinical study in a person with advanced ALS, IgA gammopathy of unclear significance, and B lymphopenia, CD19+ B cells were positively selected from a healthy haploidentical donor and infused intravenously twice, at a 60-day interval. Repeated intravenous B-cell administration was safe and significantly delayed disease onset, extended survival, reduced cellular apoptosis, and decreased astrogliosis in SOD1G93A mice. Repeated B-cell infusion in a person with ALS was safe and did not appear to generate a clinically evident inflammatory response. An improvement of 5 points on the ALSFRS-R scale was observed after the first infusion. Levels of inflammatory markers showed persistent reduction post-infusion. This represents a first demonstration of the efficacy of haploidentical B-cell infusion in the SOD1G93A mouse and the safety and feasibility of using purified haploidentical B lymphocytes as a cell-based therapeutic strategy for a person with ALS.

Green Synthesis of Silver Nanoparticles with Extracts from Kalanchoe fedtschenkoi: Characterization and Bioactivities.

In this work, the hexane, chloroform, and methanol extracts from Kalanchoe fedtschenkoi were utilized to green-synthesize silver nanoparticles (Kf1-, Kf2-, and Kf3-AgNPs). The Kf1-, Kf2-, and Kf3-AgNPs were characterized by spectroscopy and microscopy techniques. The antibacterial activity of AgNPs was studied against bacteria strains, utilizing the microdilution assay. The DPPH and H2O2 assays were considered to assess the antioxidant activity of AgNPs. The results revealed that Kf1-, Kf2-, and Kf3-AgNPs exhibit an average diameter of 39.9, 111, and 42 nm, respectively. The calculated ζ-potential of Kf1-, Kf2-, and Kf3-AgNPs were -20.5, -10.6, and -7.9 mV, respectively. The UV-vis analysis of the three samples demonstrated characteristic absorption bands within the range of 350-450 nm, which confirmed the formation of AgNPs. The FTIR analysis of AgNPs exhibited a series of bands from 3500 to 750 cm-1, related to the presence of extracts on their surfaces. SEM observations unveiled that Kf1- and Kf2-AgNPs adopted structural arrangements related to nano-popcorns and nanoflowers, whereas Kf3-AgNPs were spherical in shape. It was determined that treatment with Kf1-, Kf2-, and Kf3-AgNPs was demonstrated to inhibit the growth of E. coli, S. aureus, and P. aeruginosa in a dose-dependent manner (50-300 µg/mL). Within the same range, treatment with Kf1-, Kf2-, and Kf3-AgNPs decreased the generation of DPPH (IC50 57.02-2.09 µg/mL) and H2O2 (IC50 3.15-3.45 µg/mL) radicals. This study highlights the importance of using inorganic nanomaterials to improve the biological performance of plant extracts as an efficient nanotechnological approach.

First report of two Pantoea species causing bacterial leaf blight on wheat in the United States.

Wheat (Triticum aestivum) is an important crop worldwide, contributing to about one third of the global caloric intake. In June 2021, leaves with bacterial blight symptoms, including yellow and necrotic lesions running parallel to veins, were found in several fields across five counties in eastern Colorado (Weld, Morgan, Sedgwick, Baca, and Kit Carson). Plants exhibiting these symptoms were scattered throughout fields, but symptoms appeared consistent across counties. To determine the causal agent and complete Koch's postulates, a 1 cm2 symptomatic leaf area was excised and macerated in 0.5 mL of sterilized water from four field samples. The lysate was spread on yeast extract dextrose calcium carbonate medium (YDC agar, 1% yeast extract, 2% dextrose, 2% calcium carbonate, 1.5% agar) to isolate bacteria. Single colonies of yellow, mucoid morphology were selected and streaked on new YDC plates. Isolate genomic DNA was extracted (Zymo Research Quick-DNA Fungal/Bacterial Miniprep Kit, #D6005), and ~30 ng of gDNA was used to amplify the 16S rRNA, gyrB, and rpoB genes of all four isolates (Barret et al., 2015; Delétoile et al., 2009; Krawczyk et al., 2020; Ogier et al., 2019). Amplified PCR products were cleaned (Zymo DNA Clean & Concentrator kit, #D4033) and Sanger sequenced, and all sequences have been deposited in NCBI (16S rRNA: OR707336, OR707337, OR707338, OR707339), (gyrB: PP407951, PP407952, PP407953, PP407954), (rpoB: PP407955, PP407956, PP407957, PP407958). A BLAST search against whole genomes identified one isolate from Kit Carson county (CO314) and two isolates from Baca county (CO316 and CO317) as Pantoea agglomerans with 100% identity for the 16S rRNA, gyrB, and rpoB genes, and one isolate from Weld county (CO315) was 100% identical to Pantoea allii for all three genes. To complete Koch's postulates and confirm Pantoea sp. as the causal disease agents, isolates were grown as lawns on DifcoTM Nutrient Agar (NA) medium (48h, 28℃), suspended in 10 mM MgCl2 using a final optical density of 0.1 (~109 colony forming units per milliliter (CFU/mL)), and syringe-infiltrated into the entire leaf area of 10-day-old wheat seedling leaves (var. Hatcher). Treatments of 10mM MgCl2 and a field isolate that does not cause symptoms, identified as Pseudomonas synxantha by 16S rRNA and gyrB sequencing, were negative controls. Inoculated wheat plants were transferred to a growth chamber (22℃, 90% relative humidity). Symptoms developed 14 days post inoculation (dpi), with the most severe appearing 21 dpi. Each of the four Pantoea isolates were re-isolated from symptomatic leaves by grinding them in a Tissue Lyser II (Qiagen) with two metal beads and diluting with 0.4 mL of sterile water. A 20 µL sample of each isolate was plated on NA (24h, 28℃). The colonies appeared phenotypically identical to the original isolates, and Sanger sequencing confirmed the identities of the isolates. To our knowledge, this is the first report of P. agglomerans causing disease in wheat in the United States, and the first report of P. allii as a wheat disease-causing agent. This report is consistent with previous communications showing P. agglomerans causing wheat disease in China (Gao et al., 2023), and P. ananatis in Poland (Krawczyk et al., 2020). The growing numbers of reports of Pantoea spp. as pathogens in recent years suggests increasing novel disease emergence on cereals worldwide.

Changes in the chikungunya virus E1 glycoprotein domain II and hinge influence E2 conformation, infectivity, and virus-receptor interactions.

In a previous study to understand how the chikungunya virus (CHIKV) E1 glycoprotein ß-strand c functions, we identified several attenuating variants at E1 residue V80 and the emergence of second-site mutations in the fusion loop (E1-M88L) and hinge region (E1-N20Y) with the V80 variants in vivo. The emergence of these mutations led us to question how changes in E1 may contribute to CHIKV infection at the molecular level. Here, we use molecular dynamics to understand how changes in the E1 glycoprotein may influence the CHIKV glycoprotein E1-E2 complex. We found that E1 domain II variants lead to E2 conformational changes, allowing us to hypothesize that emerging variants E1-M88L and E1-N20Y could also change E2 conformation and function. We characterized CHIKV E1-M88L and E1-N20Y in vitro and in vivo to understand how these regions of the E1 glycoprotein contribute to host-specific infection. We found that CHIKV E1-N20Y enhanced infectivity in mosquito cells, while the CHIKV E1-M88L variant enhanced infectivity in both BHK-21 and C6/36 cells and led to changes in viral cholesterol-dependence. Moreover, we found that E1-M88L and E1-N20Y changed E2 conformation, heparin binding, and interactions with the receptor Mxra8. Interestingly, the CHIKV E1-M88L variant increased replication in Mxra8-deficient mice compared to WT CHIKV, yet was attenuated in mouse fibroblasts, suggesting that residue E1-M88 may function in a cell-type-dependent entry. Taken together, these studies show that key residues in the CHIKV E1 domain II and hinge region function through changes in E1-E2 dynamics to facilitate cell- and host-dependent entry.IMPORTANCEArboviruses are significant global public health threats, and their continued emergence around the world highlights the need to understand how these viruses replicate at the molecular level. The alphavirus glycoproteins are critical for virus entry in mosquitoes and mammals, yet how these proteins function is not completely understood. Therefore, it is critical to dissect how distinct glycoprotein domains function in vitro and in vivo to address these gaps in our knowledge. Here, we show that changes in the CHIKV E1 domain II and hinge alter E2 conformations leading to changes in virus-receptor and -glycosaminoglycan interactions and cell-specific infection. These results highlight that adaptive changes in E1 can have a major effect on virus attachment and entry, furthering our knowledge of how alphaviruses infect mammals and insects.