Simplified Protocol to Demonstrate Gene Expression in Nicotiana benthamiana Using an Agrobacterium-Mediated Transient Assay.

Agrobacterium-mediated transient gene expression in Nicotiana benthamiana is widely used to study gene function in plants. One dramatic phenotype that is frequently screened for is cell death. Here, we present a simplified protocol for Agrobacterium-mediated transient gene expression by infiltration. Compared with current methods, the novel protocol can be done without a centrifuge or spectrometer, thereby suitable for K-12 outreach programs as well as rapidly identifying genes that induce cell death. Key features • The protocol simplifies the widely used Agrobacterium-mediated transient gene expression assay [1] and can be completed within one week when plants are available. • Rice XB3 gene can induce a dramatic and easily identifiable cell death phenotype in Nicotiana benthamiana. • Allows identification of cell death-inducing genes and is suitable for teaching. • Compared to the currently used methods, our protocol omits the use of agroinfiltration buffer, pH meter, temperature-controlled growth chamber, centrifuge, and spectrophotometer. Graphical overview Agrobacterium infiltration (agroinfiltration) of Nicotiana benthamiana. The photo demonstrates the method of agroinfiltration into the abaxial side of leaves using a needleless syringe.

Intravenous antibiotics for urinary tract infections in children with neurologic impairment.

BACKGROUND: Children with high-intensity neurologic impairment (HINI) have an increased risk of urinary tract infection (UTI) and prolonged intravenous (IV) antibiotic exposure. OBJECTIVE: To determine the association between short (≤3 days) and long (>3 days) IV antibiotic courses and UTI treatment failure in hospitalized children with HINI. METHODS: We performed a retrospective cohort study examining UTI hospitalizations at 49 hospitals in the Pediatric Health Information System from 2016 to 2021 for children (1-18 years) with HINI. The primary outcome was UTI readmission within 30 days. Our secondary outcome was the association of hospital-level variation in short IV antibiotic course use with readmission. Readmission rates were compared between short and long courses using multivariable regression. RESULTS: Of 5612 hospitalizations, 3840 (68.4%) had short IV antibiotic courses. In our adjusted model, children with short IV courses were less likely than with long courses to have a 30-day UTI readmission (4.0%, 95% CI [3.6%, 4.5%] vs. 6.3%, 95% CI [5.1%, 7.8%]). Despite marked hospital-level variation in short IV course use (50.0%-87.5% of hospitalizations), there was no correlation with readmissions. CONCLUSIONS: Children with HINI hospitalized with UTI had low UTI readmission rates, but those who received long IV antibiotic courses were more likely to experience UTI readmission versus those receiving short courses. While residual confounding may influence our results, we did not find that short IV courses impacted readmission at the hospital level despite variation in use across institutions. Long IV antibiotic courses are associated with risks and may not confer benefit in this population.

Pharmacologic Restraint Use During Mental Health Admissions to Children's Hospitals.

OBJECTIVES: Primary mental health admissions are increasing across US children's hospitals. These patients may experience agitation requiring pharmacologic restraint. This study characterized pharmacologic restraint use in medical inpatient units by primary mental health diagnosis. METHODS: This retrospective, cross-sectional study used the Pediatric Health Information System database. The study included children aged 5 to 17 years admitted with a primary mental health diagnosis between 2016 and 2021. Rates of pharmacologic restraint use per 1000 patient days were determined for 13 mental health diagnoses and trended over time with Poisson regression. RESULTS: Of 91 898 hospitalizations across 43 hospitals, 3% of admissions and 1.3% of patient days involved pharmacologic restraint. Trends in the rate of pharmacologic restraint use remained stable (95% confidence interval [CI], 0.7-2.1), whereas the incidence increased by 141%. Diagnoses with the highest rates of pharmacologic restraint days per 1000 patient days included autism (79.4; 95% CI, 56.2-112.3), substance-related disorders (45.0; 95% CI, 35.9-56.4), and disruptive disorders (44.8; 95% CI, 25.1-79.8). The restraint rate significantly increased in disruptive disorders (rate ratio [RR], 1.4; 95% CI, 1.1-1.6), bipolar disorders (RR, 2.0; 95% CI, 1.4-3.0), eating disorders (RR, 2.4; 95% CI, 1.5-3.9), and somatic disorders (RR, 4.2; 95% CI, 1.9-9.1). The rate significantly decreased for autism (RR, 0.8; 95% CI, 0.6-1.0) and anxiety disorders (RR, 0.3; 95% CI, 0.2-0.6). CONCLUSIONS: Pharmacologic restraint use among children hospitalized with a primary mental health diagnosis increased in incidence and varied by diagnosis. Characterizing restraint rates and trends by diagnosis may help identify at-risk patients and guide targeted interventions to improve pharmacologic restraint utilization.

Disparities in Pharmacologic Restraint for Children Hospitalized in Mental Health Crisis.

BACKGROUND AND OBJECTIVES: Children hospitalized with a mental health crisis often receive pharmacologic restraint for management of acute agitation. We examined associations between pharmacologic restraint use and race and ethnicity among children admitted for mental health conditions to acute care nonpsychiatric children's hospitals. METHODS: We performed a retrospective cohort study of children (aged 5-≤18 years) admitted for a primary mental health condition from 2018 to 2022 at 41 US children's hospitals. Pharmacologic restraint use was defined as parenteral administration of medications for acute agitation. The association of race and ethnicity and pharmacologic restraint was assessed using generalized linear multivariable mixed models adjusted for clinical and demographic factors. Stratified analyses were performed based on significant interaction analyses between covariates and race and ethnicity. RESULTS: The cohort included 61 503 hospitalizations. Compared with non-Hispanic Black children, children of non-Hispanic White (adjusted odds ratio [aOR], 0.81; 95% confidence interval [CI], 0.72-0.92), Asian (aOR, 0.82; 95% CI, 0.68-0.99), or other race and ethnicity (aOR, 0.68; 95% CI, 0.57-0.82) were less likely to receive pharmacologic restraint. There was no significant difference with Hispanic children. When stratified by sex, racial/ethnic differences were magnified in males (aORs, 0.49-0.68), except for Hispanic males, and not found in females (aORs, 0.83-0.93). Sensitivity analysis revealed amplified disparities for all racial/ethnic groups, including Hispanic youth (aOR, 0.65; 95% CI, 0.47-0.91). CONCLUSIONS: Non-Hispanic Black children were significantly more likely to receive pharmacologic restraint. More research is needed to understand reasons for these disparities, which may be secondary to implicit bias and systemic and interpersonal racism.

Pharmacologic restraint use for children experiencing mental health crises in pediatric hospitals.

BACKGROUND: Children in mental health crises are increasingly admitted to children's hospitals awaiting inpatient psychiatric placement. During hospitalization, patients may exhibit acute agitation prompting pharmacologic restraint use. OBJECTIVE: To determine hospital-level incidence and variation of pharmacologic restraint use among children admitted for mental health conditions in children's hospitals. DESIGN, SETTING, AND PARTICIPANTS: We examined data for children (5 to ≤18 years) admitted to children's hospitals with a primary mental health condition from 2018 to 2020 using the Pediatric Health Information System database. Hospital rates of parenteral pharmacologic restraint use per 1000 mental health bed days were determined and compared after adjusting for patient-level and demographic factors. Cluster analysis (k-means) was used to group hospitals based on overall restraint use (rate quartiles) and drug class. Hospital-level factors for pharmacologic restraint use were compared. RESULTS: Of 29,834 included encounters, 3747 (12.6%) had pharmacologic restraint use. Adjusted hospital rates ranged from 35 to 389 pharmacologic restraint use days per 1000 mental health bed days with a mean of 175 (standard deviation: 72). Cluster analysis revealed three hospitals were high utilizers of all drug classes. No significant differences in pharmacologic restraint use were found in the hospital-level analysis. CONCLUSIONS: Children's hospitals demonstrate wide variation in pharmacologic restraint rates for mental health hospitalizations, with a 10-fold difference in adjusted rates between highest and lowest utilizers, and high overall utilizers order medications across all drug classes.

Improving Adherence to AAP Acute Otitis Media Guidelines in an Academic Pediatrics Practice through a Quality Improvement Project.

Introduction: Acute otitis media (AOM) is the most common reason for antibiotic use in children. The American Academy of Pediatrics (AAP) published its latest AOM guidelines in 2013. A safety-net antibiotic prescription (SNAP) is recommended for some patients based on age, severity, and duration of symptoms. At baseline, 78% of patients diagnosed with AOM in our general pediatrics practice met AAP guidelines, and 20% of eligible patients received a SNAP according to guidelines. We aimed to increase adherence to AAP AOM guidelines in an academic general pediatrics clinic from 78% to 90% by January 2020. Methods: A quality improvement team determined key drivers and developed interventions. Patients included were 6 months to 12 years old with AOM. Encounters were reviewed for adherence to AAP AOM guidelines. During the project, interventions included an ear pain note template, which generated guideline-based recommendations, note template education in clinic orientation sessions, a didactic session on AOM management, and reminders on workstations. Data were analyzed using P-charts. Results: Percent of AOM encounters (n = 1266) adhering to AAP AOM guidelines increased from 78% to 92%. We also reviewed two process measures. First, the use of the ear pain note template increased from 0% to 44%. Second, the percent of AOM encounters where an eligible patient received a SNAP increased from 21% to 78% (encounters n = 421). Conclusion: We demonstrate increased adherence to AAP AOM guidelines, including improved use of SNAPs after introducing a note template with clinical decision support and provider educational sessions.