Transiently restricting individual amino acids protects Drosophila melanogaster against multiple stressors.

Nutrition and resilience are linked, though it is not yet clear how diet confers stress resistance or the breadth of stressors that it can protect against. We have previously shown that transiently restricting an essential amino acid can protect Drosophila melanogaster against nicotine poisoning. Here, we sought to characterize the nature of this dietary-mediated protection and determine whether it was sex, amino acid and/or nicotine specific. When we compared between sexes, we found that isoleucine deprivation increases female, but not male, nicotine resistance. Surprisingly, we found that this protection afforded to females was not replicated by dietary protein restriction and was instead specific to individual amino acid restriction. To understand whether these beneficial effects of diet were specific to nicotine or were generalizable across stressors, we pre-treated flies with amino acid restriction diets and exposed them to other types of stress. We found that some of the diets that protected against nicotine also protected against oxidative and starvation stress, and improved survival following cold shock. Interestingly, we found that a diet lacking isoleucine was the only diet to protect against all these stressors. These data point to isoleucine as a critical determinant of robustness in the face of environmental challenges.

Impact of the COVID-19 Pandemic on Inpatient Pediatric Medical Student Education in Hawai'i.

The COVID-19 pandemic has had many effects on medical student education, ranging from safety measures limiting patient exposure to changes in patient diagnoses encountered by medical students in their clerkship experience. This study aimed to identify the impact of the pandemic on the inpatient experiences of third- and fourth-year medical students by assessing patient volumes and diagnoses seen by students. Frequency and types of notes written by medical students on hospital-based pediatric rotations at Kapi'olani Medical Center for Women and Children as well as patient diagnoses and ages were compared between 2 time periods: pre-pandemic (July 2018-February 2020) and pandemic (May 2020-September 2021). On average, the number of patients seen by medical students was significantly reduced in the pandemic period from 112 patients/month to 88 patients/month (P=.041). The proportion of patients with bronchiolitis or pneumonia were also significantly reduced in the pandemic period (P<.001). Bronchiolitis was diagnosed in 1.3% of patients seen by medical students during the pandemic period, compared with 5.9% of patients pre-pandemic. Pneumonia was diagnosed in 1.0% of patients seen by medical students in the pandemic period compared with 4.6% pre-pandemic. There was no significant difference in patient age between the 2 groups (P=.092). During the first 18 months of the COVID-19 pandemic, medical students in this institution had a remarkably different inpatient experience from that of their predecessors. They saw fewer patients, and those patients had fewer common pediatric respiratory diseases. These decreases suggest these students may require supplemental education to compensate for these gaps in direct pediatric clinical experience.

A multi-nutrient array protocol to study disease-diet interactions in Drosophila melanogaster.

The availability of a defined fully synthetic diet for the fruit fly Drosophila melanogaster allows for complete and precise manipulation of its nutritional environment. Here, we present a protocol for performing large-scale multivariate nutrient analysis via the traditional diet preparation approach, or by adding nutrient solutions to a baseline medium. We detail procedures from sample collection to data analysis. This protocol has applications for the study of nutrition-life trait interactions and nutrigenomics, to reveal interactions between genotype and diet composition. For complete details on the use and execution of this protocol, please refer to Martelli et al.1 and Martelli et al.2.

Landolt Ring-Shaped Epithelial Keratopathy: A Novel Corneal Disease in a Non-Japanese White Patient.

PURPOSE: Landolt ring-shaped epithelial keratopathy is a corneal disease that has only been reported in 11 Japanese patients in 2014.1 We describe the first case of Landolt ring-shaped epithelial keratopathy in the United States in a patient of European background. METHODS: This is a single case report with longitudinal care. RESULTS: A 35-year-old White patient presented with a history of ocular burning, photophobia, and decreased vision. Corneal examination showed bilateral and asymmetric microcystic lesions in a unique Landolt ring (or the letter "C") shape, distributed randomly in the epithelium. Confocal microscopy revealed cellular ballooning and hyperreflective opacities in the basal layer of the corneal epithelium. The patient has had multiple recurrences of her symptoms year-round, each lasting 4 to 8 days. Topical treatment with cyclosporine, steroids, and lubrication resolved her symptoms but without complete resolution of signs on examination. CONCLUSIONS: Our patient's clinical signs and symptoms are similar to those described previously in 11 Japanese patients. However, unlike those patients, our patient demonstrates symptomatic response to topical treatment, no seasonal association to her condition, and to date, incomplete resolution of her disease after more than 2 years. This case highlights that Landolt ring-shaped epithelial keratopathy, a novel corneal disease of unclear origin, has relevance outside of the Japanese population.

A defined diet for pre-adult Drosophila melanogaster.

Drosophila melanogaster is unique among animal models because it has a fully defined synthetic diet available to study nutrient-gene interactions. However, use of this diet is limited to adult studies due to impaired larval development and survival. Here, we provide an adjusted formula that reduces the developmental period, restores fat levels, enhances body mass, and fully rescues survivorship without compromise to adult lifespan. To demonstrate an application of this formula, we explored pre-adult diet compositions of therapeutic potential in a model of an inherited metabolic disorder affecting the metabolism of branched-chain amino acids. We reveal rapid, specific, and predictable nutrient effects on the disease state consistent with observations from mouse and patient studies. Together, our diet provides a powerful means with which to examine the interplay between diet and metabolism across all life stages in an animal model.

Locality is the strongest predictor of expert performance in image-based differentiation of bacterial and fungal corneal ulcers from India.

PURPOSE: This study sought to identify the sources of differential performance and misclassification error among local (Indian) and external (non-Indian) corneal specialists in identifying bacterial and fungal keratitis based on corneal photography. METHODS: This study is a secondary analysis of survey data assessing the ability of corneal specialists to identify acute bacterial versus fungal keratitis by using corneal photography. One-hundred images of 100 eyes from 100 patients with acute bacterial or fungal keratitis in South India were previously presented to an international cohort of cornea specialists for interpretation over the span of April to July 2021. Each expert provided a predicted probability that the ulcer was either bacterial or fungal. Using these data, we performed multivariable linear regression to identify factors predictive of expert performance, accounting for primary practice location and surrogate measures to infer local fungal ulcer prevalence, including locality, latitude, and dew point. In addition, Brier score decomposition was used to determine experts' reliability ("calibration") and resolution ("boldness") and were compared between local (Indian) and external (non-Indian) experts. RESULTS: Sixty-six experts from 16 countries participated. Indian practice location was the only independently significant predictor of performance in multivariable linear regression. Resolution among Indian experts was significantly better (0.08) than among non-Indian experts (0.01; P < 0.001), indicating greater confidence in their predictions. There was no significant difference in reliability between the two groups ( P = 0.40). CONCLUSION: Local cornea experts outperformed their international counterparts independent of regional variability in tropical risk factors for fungal keratitis. This may be explained by regional characteristics of infectious ulcers with which local corneal specialists are familiar.

Identifying potential dietary treatments for inherited metabolic disorders using Drosophila nutrigenomics.

Inherited metabolic disorders are a group of genetic conditions that can cause severe neurological impairment and child mortality. Uniquely, these disorders respond to dietary treatment; however, this option remains largely unexplored because of low disorder prevalence and the lack of a suitable paradigm for testing diets. Here, we screened 35 Drosophila amino acid disorder models for disease-diet interactions and found 26 with diet-altered development and/or survival. Using a targeted multi-nutrient array, we examine the interaction in a model of isolated sulfite oxidase deficiency, an infant-lethal disorder. We show that dietary cysteine depletion normalizes their metabolic profile and rescues development, neurophysiology, behavior, and lifelong fly survival, thus providing a basis for further study into the pathogenic mechanisms involved in this disorder. Our work highlights the diet-sensitive nature of metabolic disorders and establishes Drosophila as a valuable tool for nutrigenomic studies for informing potential dietary therapies.

Early real-world experience monitoring circulating tumor DNA in resected early-stage non-small cell lung cancer.

OBJECTIVE: The study objective was to evaluate the impact of monitoring circulating tumor DNA on the detection and management of recurrence in patients with resected early-stage non-small cell lung cancer. METHODS: Between October 2021 and March 2023, postoperative circulating tumor DNA was monitored in patients with non-small cell lung cancer (N = 108). Longitudinal blood samples (n = 378 samples) were collected for prospective circulating tumor DNA analysis at 3-month intervals after curative-intent resection. A tumor-informed assay was used for the detection and quantification of circulating tumor DNA. The primary outcome measure was a circulating tumor DNA-positive result. The secondary outcome measure was changes in practice after a circulating tumor DNA-positive result. RESULTS: The mean age of the patients in this cohort was 68.1 years. Of the 108 patients, 12 (11.1%) were circulating tumor DNA positive at least at 1 timepoint postsurgery, of whom 8 (66.7%) had a clinically evident recurrence and the remaining 4 had limited clinical follow-up. Of the 10 patients with recurrent disease, 8 demonstrated circulating tumor DNA positivity and the remaining 2 patients had brain-only metastases. Postoperative clinical care was altered in 100% (12/12) of circulating tumor DNA-positive patients, with 58.3% (7/12) receiving an early computed tomography scan and 100% (12/12) receiving an early positron emission tomography computed tomography scan as part of their surveillance strategy. Among the patients who received an early positron emission tomography scan, 66.6% (8/12) were positive for malignant features. CONCLUSIONS: Routine monitoring of tumor-informed circulating tumor DNA after curative intent therapy improved patient risk stratification and prognostication.

A meta-analytic review of the relations between anxiety and empathy.

Although theory suggests that empathy may signal a risk for anxiety (Tone & Tully, 2014), the relation between these constructs remains unclear due to the lack of a quantitative synthesis of empirical findings. We addressed this question by conducting three meta-analyses assessing anxiety and general, cognitive, and affective empathy (k's = 70-102 samples; N's = 19,410-25,102 participants). Results suggest that anxiety has a small and significant association with general empathy (r = .08). The relation of clinical anxiety with cognitive empathy was significant but very weak (r = -.03), and small for affective empathy (r = .16). Geographic region and the type of cognitive (e.g., perspective taking, fantasy) and affective empathy (e.g., affective resonance, empathic concern) emerged as moderators. Results suggest that anxiety has a weaker association with general empathy but a stronger association with affective empathy in participants from predominantly collectivistic geographic regions. Further, greater anxiety was weakly associated with less perspective-taking and greater fantasy, and anxiety had a more modest association with empathic concern than other types of affective empathy. Targeting affective empathy (e.g., promoting coping strategies when faced with others' distress) in interventions for anxiety may be beneficial.

Validation of the PyloPlus Urea Breath Test System in pediatric patients.

IMPORTANCE: For the diagnosis and post-treatment monitoring of H. pylori infection, non-invasive testing methodologies improve patient comfort, particularly for children. Previously, only the BreathTek UBT had FDA approval for use in pediatric patients and required an adjustment calculation based on age, height, and weight of the patient. The purpose of this study was to evaluate the performance of the PyloPlus UBT assay in a pediatric population.

Parthenolide As a Therapeutic for Disseminated Canine Neoplasms.

This study provides a unique translational research opportunity to help both humans and dogs diagnosed with diseases that carry dismal prognoses in both species: histiocytic sarcoma (HS), hemangiosarcoma (HSA), and disseminated mastocytosis/mast cell tumor (MCT). Although exceedingly rare in humans, these so called "orphan diseases" are relatively more common in dogs. For these and other more commonplace cancers like lymphoma (Lym), dogs are an excellent translational model for human disease due to remarkably similar disease biology. In this study, assays were performed to assess the therapeutic potential of parthenolide (PTL), a known canonical nuclear factor kappa B (NF-κB) signaling inhibitor with additional mechanisms of antineoplastic activity, including alteration of cellular reduction-oxidation balance. Canine cell lines and primary cells are sensitive to PTL and undergo dose-dependent apoptosis after exposure to drug. PTL exposure also leads to glutathione depletion, reactive oxygen species generation, and NF-κB inhibition in canine cells. Standard-of-care therapeutics broadly synergize with PTL. In two canine HS cell lines, expression of NF-κB pathway signaling partners is downregulated with PTL therapy. Preliminary data suggest that PTL inhibits NF-κB activity of cells and extends survival time in a mouse model of disseminated canine HS. These data support further investigation of compounds that can antagonize canonical NF-κB pathway signaling in these cancers and pave the way for clinical trials of PTL in affected dogs. As dogs are an excellent natural disease model for these cancers, these data will ultimately improve our understanding of their human disease counterparts and hopefully improve care for both species. SIGNIFICANCE STATEMENT: Disseminated neoplasms in human and canine cancers are challenging to treat, and novel therapeutic approaches are needed to improve outcomes. Parthenolide is a promising treatment for histiocytic sarcoma, hemangiosarcoma, and mast cell neoplasia.

A Simple Method for Quantifying Larval Locomotion in Drosophila melanogaster.

The fruit fly Drosophila melanogaster is a powerful genetic model that has been used for many decades to study nervous system function, development, and behavior. There are a large number of developmental and behavioral traits that can be measured to provide a broad readout of neurological function. These include patterned motor behaviors, such as larval locomotion, which can be used to assess whether genetic or environmental factors affect nervous system function to provide an entry point for deeper mechanistic studies. Here, we describe a protocol for quantifying larval locomotion using a simple camera setup and a freely available image analysis software. This protocol can be readily applied to human disease models or in toxicology studies, for example, to broadly assess the impact of treatments on neurological function.

Immunohistochemical Analysis of the Drosophila Larval Neuromuscular Junction.

Synapses are specialized junctions between cells that mediate neurotransmission to modify brain activity and body function. Studies on synapse structure and function play an important role in understanding how neurons communicate and the consequences of their dysfunction in neurological disorders. The Drosophila larval neuromuscular junction is an excellent model for dissecting the cellular and molecular mechanisms of the synapse, with its large size, accessibility, and well-characterized genetics. This protocol describes the steps required for morphological and immunohistochemical analysis of the Drosophila larval neuromuscular junction including its dissection and multiplex labeling of synaptic proteins. This technique can be used to assess the impact of genetic manipulations on synaptic development, integrity, and plasticity, thus providing a valuable tool for probing complex neurological processes in a whole animal system.

Assessing the Impact of Image Quality on Deep Learning Classification of Infectious Keratitis.

Objective: To investigate the impact of corneal photograph quality on convolutional neural network (CNN) predictions. Design: A CNN trained to classify bacterial and fungal keratitis was evaluated using photographs of ulcers labeled according to 5 corneal image quality parameters: eccentric gaze direction, abnormal eyelid position, over/under-exposure, inadequate focus, and malpositioned light reflection. Participants: All eligible subjects with culture and stain-proven bacterial and/or fungal ulcers presenting to Aravind Eye Hospital in Madurai, India, between January 1, 2021 and December 31, 2021. Methods: Convolutional neural network classification performance was compared for each quality parameter, and gradient class activation heatmaps were generated to visualize regions of highest influence on CNN predictions. Main Outcome Measures: Area under the receiver operating characteristic and precision recall curves were calculated to quantify model performance. Bootstrapped confidence intervals were used for statistical comparisons. Logistic loss was calculated to measure individual prediction accuracy. Results: Individual presence of either light reflection or eyelids obscuring the corneal surface was associated with significantly higher CNN performance. No other quality parameter significantly influenced CNN performance. Qualitative review of gradient class activation heatmaps generally revealed the infiltrate as having the highest diagnostic relevance. Conclusions: The CNN demonstrated expert-level performance regardless of image quality. Future studies may investigate use of smartphone cameras and image sets with greater variance in image quality to further explore the influence of these parameters on model performance. Financial Disclosures: Proprietary or commercial disclosure may be found after the references.

GCN2 mediates access to stored amino acids for somatic maintenance during Drosophila ageing.

Many mechanistic theories of ageing argue that a progressive failure of somatic maintenance, the use of energy and resources to prevent and repair damage to the cell, underpins ageing. To sustain somatic maintenance an organism must acquire dozens of essential nutrients from the diet, including essential amino acids (EAAs), which are physiologically limiting for many animals. In Drosophila, adulthood deprivation of each individual EAA yields vastly different lifespan trajectories, and adulthood deprivation of one EAA, phenylalanine (Phe), has no associated lifespan cost; this is despite each EAA being strictly required for growth and reproduction. Moreover, survival under any EAA deprivation depends entirely on the conserved AA sensor GCN2, a component of the integrated stress response (ISR), suggesting that a novel ISR-mediated mechanism sustains lifelong somatic maintenance during EAA deprivation. Here we investigated this mechanism, finding that flies chronically deprived of dietary Phe continue to incorporate Phe into new proteins, and that challenging flies to increase the somatic requirement for Phe shortens lifespan under Phe deprivation. Further, we show that autophagy is required for full lifespan under Phe deprivation, and that activation of the ISR can partially rescue the shortened lifespan of GCN2-nulls under Phe deprivation. We therefore propose a mechanism by which GCN2, via the ISR, activates autophagy during EAA deprivation, breaking down a larvally-acquired store of EAAs to support somatic maintenance. These data refine our understanding of the strategies by which flies sustain lifelong somatic maintenance, which determines length of life in response to changes in the nutritional environment.

Neuroligin 2 governs synaptic morphology and function through RACK1-cofilin signaling in Drosophila.

Neuroligins are transmembrane cell adhesion proteins well-known for their genetic links to autism spectrum disorders. Neuroligins can function by regulating the actin cytoskeleton, however the factors and mechanisms involved are still largely unknown. Here, using the Drosophila neuromuscular junction as a model, we reveal that F-Actin assembly at the Drosophila NMJ is controlled through Cofilin signaling mediated by an interaction between DNlg2 and RACK1, factors not previously known to work together. The deletion of DNlg2 displays disrupted RACK1-Cofilin signaling pathway with diminished actin cytoskeleton proteo-stasis at the terminal of the NMJ, aberrant NMJ structure, reduced synaptic transmission, and abnormal locomotion at the third-instar larval stage. Overexpression of wildtype and activated Cofilin in muscles are sufficient to rescue the morphological and physiological defects in dnlg2 mutants, while inactivated Cofilin is not. Since the DNlg2 paralog DNlg1 is known to regulate F-actin assembly mainly via a specific interaction with WAVE complex, our present work suggests that the orchestration of F-actin by Neuroligins is a diverse and complex process critical for neural connectivity.

Conjunctival Lesions Secondary to Systemic Varicella Zoster Virus Infection.

Purpose: To report and present images of a case in which discrete conjunctival lesions developed in the setting of primary varicella zoster virus infection (ie, chickenpox). Methods: Case report and literature review. Results: This report describes a young, unvaccinated male who developed an acutely painful, red eye in the setting of disseminated primary varicella zoster infection. The cutaneous rash was widespread and included lesions on both eyelids. The patient was found to have multiple discrete de-epithelialized lesions involving the palpebral and bulbar conjunctiva. Throughout the disease course, good visual function was maintained and there was no evidence of intraocular involvement. The ocular surface lesions resolved without sequelae after 1 week of treatment with topical antibiotic ointment. Conclusions: Primary varicella zoster infection is an increasingly rare phenomenon in the setting of widespread vaccination. However, unvaccinated or undervaccinated individuals and other at-risk populations remain susceptible to developing severe infections. This case of chickenpox involved discrete conjunctival lesions that resolved without sequelae after conservative treatment with topical antibiotic ointment. While serious ophthalmic complications are uncommon in primary varicella infection, clinicians should be aware of the potential for ocular morbidity in this increasingly rare condition.

Review: The PI3K-AKT-mTOR signal transduction pathway in canine cancer.

Tumors in dogs and humans share many similar molecular and genetic features, incentivizing a better understanding of canine neoplasms not only for the purpose of treating companion animals, but also to facilitate research of spontaneously developing tumors with similar biologic behavior and treatment approaches in an immunologically competent animal model. Multiple tumor types of both species have similar dysregulation of signal transduction through phosphatidylinositol 3-kinase (PI3K), protein kinase B (PKB; AKT), and mechanistic target of rapamycin (mTOR), collectively known as the PI3K-AKT-mTOR pathway. This review aims to delineate the pertinent aspects of the PI3K-AKT-mTOR signaling pathway in health and in tumor development. It will then present a synopsis of current understanding of PI3K-AKT-mTOR signaling in important canine cancers and advancements in targeted inhibitors of this pathway.

Single-Cell RNA Sequencing Reveals That Adaptation of Human Aortic Endothelial Cells to Antiproliferative Therapies Is Modulated by Flow-Induced Shear Stress.

BACKGROUND: Endothelial cells (ECs) are capable of quickly responding in a coordinated manner to a wide array of stresses to maintain vascular homeostasis. Loss of EC cellular adaptation may be a potential marker for cardiovascular disease and a predictor of poor response to endovascular pharmacological interventions such as drug-eluting stents. Here, we report single-cell transcriptional profiling of ECs exposed to multiple stimulus classes to evaluate EC adaptation. METHODS: Human aortic ECs were costimulated with both pathophysiological flows mimicking shear stress levels found in the human aorta (laminar and turbulent, ranging from 2.5 to 30 dynes/cm2) and clinically relevant antiproliferative drugs, namely paclitaxel and rapamycin. EC state in response to these stimuli was defined using single-cell RNA sequencing. RESULTS: We identified differentially expressed genes and inferred the TF (transcription factor) landscape modulated by flow shear stress using single-cell RNA sequencing. These flow-sensitive markers differentiated previously identified spatially distinct subpopulations of ECs in the murine aorta. Moreover, distinct transcriptional modules defined flow- and drug-responsive EC adaptation singly and in combination. Flow shear stress was the dominant driver of EC state, altering their response to pharmacological therapies. CONCLUSIONS: We showed that flow shear stress modulates the cellular capacity of ECs to respond to paclitaxel and rapamycin administration, suggesting that while responding to different flow patterns, ECs experience an impairment in their transcriptional adaptation to other stimuli.

Functional Family Therapy for families of youth (age 11-18) with behaviour problems: A systematic review and meta-analysis.

Background: Functional Family Therapy (FFT) is a short-term family-based intervention for youth with behaviour problems. FFT has been widely implemented in the USA and other high-income countries. It is often described as an evidence-based program with consistent, positive effects. Objectives: We aimed to synthesise the best available data to assess the effectiveness of FFT for families of youth with behaviour problems. Search Methods: Searches were performed in 2013-2014 and August 2020. We searched 22 bibliographic databases (including PsycINFO, ERIC, MEDLINE, Science Direct, Sociological Abstracts, Social Services Abstracts, World CAT dissertations and theses, and the Web of Science Core Collection), as well as government policy databanks and professional websites. Reference lists of articles were examined, and experts were contacted to search for missing information. Selection Criteria: We included randomised controlled trials (RCTs) and quasi-experimental designs (QEDs) with parallel cohorts and statistical controls for between-group differences at baseline. Participants were families of young people aged 11-18 with behaviour problems. FFT programmes were compared with usual services, alternative treatment, and no treatment. There were no publication, geographic, or language restrictions. Data Collection and Analysis: Two reviewers independently screened 1039 titles and abstracts, read all available study reports, assessed study eligibility, and extracted data onto structured electronic forms. We assessed risks of bias (ROB) using modified versions of the Cochrane ROB tool and the What Works Clearinghouse standards. Where possible, we used random effects models with inverse variance weights to pool results across studies. We used odds ratios for dichotomous outcomes and standardised mean differences for continuous outcomes. We used Hedges g to adjust for small sample sizes. We assessed the heterogeneity of effects with χ 2 and I 2. We produced separate forest plots for conceptually distinct outcomes and for different endpoints (<9, 9-14, 15-23, and 24-42 months after referral). We grouped studies by study design (RCT or QED), and then assessed differences between these two subgroups of studies with χ 2 tests. We generated robust variance estimates, using correlated effects (CE) models with small sample corrections to synthesise all available outcome data. Exploratory CE analyses assessed potential moderators of effects within these domains. We used GRADE guidelines to assess the certainty of evidence on six primary outcomes at 1 year after referral. Main Results: Twenty studies (14 RCTs and 6 QEDs) met our inclusion criteria. Fifteen of these studies provided some valid data for meta-analysis; these studies included 10,980 families in relevant FFT and comparison groups. All included studies had high risks of bias on at least one indicator. Half of the studies had high risks of bias on baseline equivalence, support for intent-to-treat analysis, selective reporting, and conflicts of interest. Fifteen studies had incomplete reporting of outcomes and endpoints. Using the GRADE rubric, we found that the certainty of evidence for FFT was very low for all of our primary outcomes. Using pairwise meta-analysis, we found no evidence of effects of FFT compared with other active treatments on any primary or secondary outcomes. Primary outcomes were: recidivism, out-of-home placement, internalising behaviour problems, external behaviour problems, self-reported delinquency, and drug or alcohol use. Secondary outcomes were: peer relations and prosocial behaviour, youth self esteem, parent symptoms and behaviour, family functioning, school attendance, and school performance. There were few studies in the pairwise meta-analysis (k < 7) and little heterogeneity of effects across studies in most of these analyses. There were few differences between effect estimates obtained in RCTs versus QEDs. More comprehensive CE models showed positive results of FFT in some domains and negative results in others, but these effects were small (standardised mean difference [SMD] <|0.20|) and not significantly different from no effect with one exception: Two studies found positive effects of FFT on youth substance abuse and two studies found null results in this domain, and the overall effect estimate for this outcome was statistically different from zero. Over all outcomes (15 studies and 293 effect sizes), small positive effects were detected (SMD = 0.19, SE = 0.09), but these were not significantly different from zero effect. Prediction intervals showed that future FFT evaluations are likely to produce a wide range of results, including moderate negative effects and strong positive results (-0.37 to 0.75). Authors' Conclusions: Results of 10 RCTs and five QEDs show that FFT does not produce consistent benefits or harms for youth with behavioural problems and their families. The positive or negative direction of results is inconsistent within and across studies. Most outcomes are not fully reported, the quality of available evidence is suboptimal, and the certainty of this evidence is very low. Overall estimates of effects of FFT may be inflated, due to selective reporting and publication biases.