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A study was conducted to examine the influence of CaCl2 on mushroom tyrosinase activity and membrane integrity on postharvest browning. Standardized bruising treatment were carried out on randomly selected mushrooms. Results suggested that increases in the calcium levels in mushrooms irrigated with CaCl2 may have minimized browning through increases in the integrity of the vacuolar membrane.

Introduction: Foot and ankle alignment plays a pivotal role in human gait and posture. Traditional assessment methods, relying on 2D standing radiographs, present limitations in capturing the dynamic 3D nature of foot alignment during weight-bearing and are prone to observer error. This study aims to integrate weight-bearing CT (WBCT) imaging and advanced deep learning (DL) techniques to automate and enhance quantification of the 3D foot and ankle alignment., Methods: Thirty-two patients who underwent a WBCT of the foot and ankle were retrospectively included. After training and validation of a 3D nnU-Net model on 45 cases to automate the segmentation into bony models, 35 clinically relevant 3D measurements were automatically computed using a custom-made tool. Automated measurements were assessed for accuracy against manual measurements, while the latter were analyzed for inter-observer reliability., Results: DL-segmentation results showed a mean dice coefficient of 0.95 and mean Hausdorff distance of 1.41 mm. A good to excellent reliability and mean prediction error of under 2 degrees was found for all angles except the talonavicular coverage angle and distal metatarsal articular angle., Conclusion: In summary, this study introduces a fully automated framework for quantifying foot and ankle alignment, showcasing reliability comparable to current clinical practice measurements. This operator-friendly and time-efficient tool holds promise for implementation in clinical settings, benefiting both radiologists and surgeons. Future studies are encouraged to assess the tool's impact on streamlining image assessment workflows in a clinical environment., Competing Interests: Declaration of Competing Interest Arne Burssens declares a previous consultancy for Curvebeam LLC. Other authors declare no competing interests, (Copyright © 2024 Elsevier B.V. All rights reserved.)

Traumatic injury is associated with several pulmonary complications, including pulmonary contusion, transfusion-related acute lung injury (TRALI), and the development of acute respiratory distress syndrome (ARDS). There is a lack of literature on these patients supported with veno-venous extracorporeal oxygenation (VV ECMO). Understanding the safety of using VV ECMO to support trauma patients and the ability to hold anticoagulation is important to broaden utilization. This is a single-center retrospective cohort study of adult trauma patients cannulated for VV ECMO during their initial admission over an 8 year period (2014-2021). We hypothesize that anticoagulation can be held in trauma patients on VV ECMO without increasing mortality or prothrombotic complications. We also describe the coagulopathy of traumatically injured patients on VV ECMO. Withholding anticoagulation was not associated with mortality in our study population, and there were no significant differences in bleeding or clotting complications between patients who did and did not receive systemic anticoagulation. Patients in the nonsurvivor group had increased coagulopathy both pre- and post-cannulation. Our study suggests anticoagulation can be safely withheld in traumatically injured VV ECMO patients without increasing mortality, complication rates, or transfusion requirements. Future, multicenter prospective studies with larger sample sizes are required to confirm our results., Competing Interests: Disclosure: The authors have no conflicts of interest to report., (Copyright © ASAIO 2024.)

Alterations in ankle's articular contact mechanics serve as one of the fundamental causes of significant pathology. Nevertheless, computationally intensive algorithms and lack of bilateral weightbearing imaging have rendered it difficult to investigate the normative articular contact stress and side-to-side differences. The aims of our study were two-fold: 1) to determine and quantify the presence of side-to-side contact differences in healthy ankles and 2) to establish normative ranges for articular ankle contact parameters. In this retrospective comparative study, 50 subjects with healthy ankles on bilateral weight-bearing CT were confirmed eligible. Segmentation into 3D bony models was performed semi-automatically, and individualized cartilage layers were modelled based on a previously validated methodology. Contact mechanics were evaluated by using the mean and maximum contact stress of the tibiotalar articulation. Absolute and percentage reference range values were determined for the side-to-side difference. Amongst a cohort of individuals devoid of ankle pathology, mean side-to-side variation in these measurements was < 12 %, while respective differences of > 17 % talar peak stress and > 31 % talar mean stress indicate abnormality. No significant differences were found between laterality in any of the evaluated contact parameters. Understanding these values may promote a more accurate assessment of ankle joint biomechanics when distinguishing acceptable versus pathological contact mechanics in clinical practice., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

Background: Talar displacement is considered the main predictive factor for poor outcomes and the development of post-traumatic osteoarthritis after ankle fractures. Isolated lateral talar translation, as previously studied by Ramsey and Hamilton using carbon powder imprinting, does not fully replicate the multidirectional joint subluxations seen in ankle fractures. The purpose of this study was to analyze the influence of multiple uniplanar talar displacements on tibiotalar contact mechanics utilizing weightbearing computed tomography (WBCT) and finite element analysis (FEA)., Methods: Nineteen subjects (mean age = 37.6 years) with no history of ankle surgery or injury having undergone WBCT arthrogram (n = 1) and WBCT without arthrogram (n = 18) were included. Segmentation of the WBCT images into 3D simulated models of bone and cartilage was performed. Three-dimensional (3D) multiple uniplanar talar displacements were simulated to investigate the respective influence of various uniaxial displacements (including lateral translation, anteroposterior translation, varus-valgus angulation, and external rotation) on the tibiotalar contact mechanics using FEA. Tibiotalar peak contact stress and contact area were modeled for each displacement and its gradations., Results: Our modeling demonstrated that peak contact stress of the talus and tibia increased, whereas contact area decreased, with incremental displacement in all tested directions. Contact stress maps of the talus and tibia were computed for each displacement demonstrating unique patterns of pressure derangement. One millimeter of lateral translation resulted in 14% increase of peak talar contact pressure and a 3% decrease in contact area., Conclusion: Our model predicted that with lateral talar translation, there is less noticeable change in tibiotalar contact area compared with prior studies whereas external rotation greater than 12 degrees had the largest effect on peak contact stress predictions., Level of Evidence: Level V, computational simulation study., Competing Interests: Declaration of Conflicting InterestsThe author(s) declared no potential conflicts of interest with respect to the research, authorship, and/or publication of this article. Disclosure forms for all authors are available online.

Background: Minimally invasive surgical (MIS) osteotomies are increasing as a surgical option for treating midfoot and forefoot conditions. This study aimed to evaluate the impact of each burr pass on the degree of correction, gap size, and alignment in MIS Akin and first metatarsal dorsiflexion osteotomies (DFO)., Methods: MIS Akin and first metatarsal DFO were performed on ten cadaveric specimens. Fluoroscopic measurements included the metatarsal dorsiflexion angle (MDA), dorsal cortical length (MDCL), first phalangeal medial cortical length (PCML) and proximal to distal phalangeal articular angle (PDPAA)., Results: The average decrease in PCML with each burr pass was as follows: 1.53, 1.33, 1.27, 1.23 and 1.13 mm at the 1st to 5th pass, respectively. The MDCL sequentially decreased by 1.80, 1.59, 1.35, 0.75, and 0.60 mm. The MDA consistently decreased, and the PDPAA incrementally became more valgus oriented., Conclusion: On average, a first metatarsal dorsal wedge resection of 4.7 mm and first phalangeal medial wedge resection of 2.9 mm was achieved after 3 and 2 burr passes, respectively. This data may aid surgeons determine the optimal number of burr passes required to achieve the desired patient-specific surgical correction., (Copyright © 2024 European Foot and Ankle Society. Published by Elsevier Ltd. All rights reserved.)

Background: The debridement and Achilles tendon reinsertion (DATR) have been the most common surgical approach for the treatment of Insertional Achilles Tendinopathy (IAT), while dorsal closing wedge calcaneal osteotomy (DCWCO) has recently gained popularity as an alternative surgical option. This study aimed to systematically review the published literature on both surgical techniques and compare their clinical outcomes and complication rates., Methods: A systematic review was performed according to the PRISMA guidelines using Medline, Embase, and Scopus databases. The inclusion criteria encompassed clinical studies reporting functional outcomes and complications, with a minimum of 10 patients and at least 12 months of follow-up., Results: Seven studies (n = 169) were included for the analysis of DATR, and eight studies (n = 227) were included for the analysis of open DCWCO. Both groups showed a similar improvement in AOFAS score. The overall complication rates were 16.6% in the DATR group and 9.2% in the DCWCO group, but the difference was not statistically significant. However, there was a significantly higher incidence of wound complications in the DATR group (10.1%, 95% C.I.: 4.7-15.6) compared to the DCWCO group (2.5%, 95% C.I.: 0.6-4.4) as the confidence intervals did not overlap., Conclusions: Clinical outcomes and overall complication rates of both techniques were comparable, although DCWCO had a lower incidence of wound complications. Further research should be focused on prospective studies comparing the two techniques to corroborate the current findings., Level of Evidence: Level IV; meta-analysis., Competing Interests: Declaration of Competing Interest The authors declare no conflicts of interest., (Copyright © 2023 European Foot and Ankle Society. Published by Elsevier Ltd. All rights reserved.)

Background: This retrospective cohort study sought to describe the ability of high protein regimens to achieve nitrogen equilibrium in patients receiving venovenous extracorporeal membrane oxygenation (VV ECMO)., Methods: Patients aged ≥18 years with a documented nitrogen balance study (NB) on VV ECMO between February 2018 and December 2021 were included. Studies with incomplete 24-h urine collections or changes in blood urea nitrogen ≥10 mg/dl were excluded. Data were summarized, correlation between first NB and potentially contributing variables was assessed with Kendall tau. Subanalysis described findings after stratifying for weight class (obese vs nonobese) and duration of VV ECMO at the time of NB., Results: A total of 68 NBs in 30 patients were included; 47% of the cohort had obesity. The number of NBs per patient was 2.2 ± 1.1, which were completed on a median of 31.5 (interquartile range: 16, 53.8) days receiving ECMO. Nitrogen equilibrium or positive balance was achieved in 72% of studies despite elevated nitrogen excretion. Patients received 87.9 ± 16.8% of prescribed protein on NB days for average intakes of 2.4 ± 0.4 g/kg of actual weight per day and 2.4 ± 0.5 g/kg of ideal weight per day in patients without and with obesity. Median NB in patients without obesity was -1.46 (-8.96, 2.98) g/day and -0.21 (-10.58, 4.04) g/day in patients with obesity. A difference in median NB after stratification for timing was observed (P = 0.029)., Conclusion: Nitrogen equilibrium can be achieved with high protein intake in adults receiving VV ECMO. NB monitoring is one tool to individualize protein prescriptions throughout the course of VV ECMO., (© 2023 American Society for Parenteral and Enteral Nutrition.)

Purpose: This study aimed to evaluate the impact of each burr pass on degree of correction, gap size and calcaneal morphology in MIS Zadek osteotomy., Methods: MIS Zadek osteotomy was performed on ten cadaveric specimens using a 3.1 mm Shannon burr. After each burr pass, the osteotomy gap was manually closed, and the subsequent burr passes were carried out with the foot held in dorsiflexion, which was repeated five times. Lateral X-rays were taken before and after each burr pass. Two independent reviewers measured the dorsal calcaneal length after each burr passage, as well as changes in several calcaneal parameters including X/Y ratio, Fowler Philip angle, and Böhler angle., Results: The average decrease in dorsal calcaneal cortical length with each burr pass was as follows: 2.6 ± 0.9 mm at the 1st pass, 2.4 ± 1 mm at the 2nd pass, 2 ± 1 mm at the 3rd pass, 1.6 ± 1 mm at the 4th pass, and 1.4 ± 0.7 mm at the 5th pass. The Fowler Philip and Böhler angles consistently decreased while the X/Y ratio consistently increased following each consecutive burr pass. Interobserver reliability analysis demonstrated good agreement for all parameters., Conclusion: The results revealed the trends of length and anatomical changes in the calcaneus with each burr pass. On average, a dorsal wedge resection of 10 mm was achieved after 5 burr passes. This data can aid surgeons in determining the optimal number of burr passes required for a particular amount of resection, ensuring the attainment of the desired patient-specific surgical outcome., Competing Interests: Conflict of interest The authors declare no conflicts of interest., (Copyright © 2023 European Foot and Ankle Society. Published by Elsevier Ltd. All rights reserved.)

Objectives: To evaluate the utility and diagnostic performance of portable handheld ultrasound for evaluating fibular rotation at the distal tibiofibular articulation after syndesmotic disruption., Methods: Four above-the-knee cadaveric specimens were included. Syndesmotic disruption was precipitated by transecting the Anterior Inferior Tibiofibular Ligament, Interosseous Ligament, and Posterior Inferior Tibiofibular Ligament. Thereafter, a proximal fibular osteotomy was performed, and three conditions were modeled at the distal syndesmosis: 1) reduced, 2) 5 degree internal rotation malreduction, and 3) 5 degree external rotation malreduction. Two blinded observers performed separate ultrasonographic examinations for each condition at the level of both the anterior and posterior distal tibiofibular articular surfaces. Syndesmotic gap penetrance, defined as the ability of the P-US to generate signal between the distal fibula and tibia at the level of the incisura, was graded positive if the sonographic waves penetrated between the distal tibiofibular joint and negative if no penetrating waves were detected. The accuracy measures of the anterior and posterior gap penetrance were evaluated individually., Results: Our preliminary results showed that posterior gap penetrance showed good performance when detecting either internal or external rotational malreduction of the fibula with very good specificity (87.5%) and PPV (90.0%). On the other hand, the anterior gap penetrance showed limited performance when detecting either form of rotational malreduction., Conclusion: We introduced a novel sign, the "gap penetrance sign", best measured from the posterior ankle, which can accurately detect syndesmotic malreduction using P-US in a manner that does not require specific quantitative measurements and is readily accessible to early P-US users., Competing Interests: None, (2024 © BY THE ARCHIVES OF BONE AND JOINT SURGERY.)

Background: A medializing calcaneal osteotomy (MCO) is considered as one of the key inframalleolar osteotomies to correct progressive collapsing foot deformity (PCFD). While many studies were able to determine the post-operative hind- and midfoot alignment, alternations of the subtalar joint alignment remained obscured by superposition on plain radiography. Therefore, we aimed to assess the hind-, midfoot- and subtalar joint alignment pre- compared to post-operatively using 3D weightbearing CT (WBCT) imaging., Methods: Seventeen patients with a mean age of 42 ± 17 years were retrospectively analyzed. Inclusion criteria consisted of PCFD deformity corrected by a medializing calcaneal osteotomy (MCO) as main procedure and imaged by WBCT before and after surgery. Exclusion criteria were patients who had concomitant calcaneal lengthening osteotomies, mid-/hindfoot fusions, hindfoot coalitions, and supramalleolar procedures. Image data were used to generate 3D models and compute the hindfoot (HA), midfoot (MA) - and subtalar joint (STJ) alignment in the coronal, sagittal and axial plane, as well as distance maps., Results: Pre-operative measurements of the HA and MA improved significantly relative to their post-operative equivalents p < 0.05). The post-operative STJ alignment showed significant inversion (2.8° ± 1.7), abduction (1.5° ± 1.8), and dorsiflexion (2.3° ± 1.7) of the talus relative to the calcaneus (p < 0.05) compared to the pre-operative alignment. The displacement between the talus and calcaneus relative to the sinus tarsi increased significantly (0.6 mm±0.5; p < 0.05)., Conclusion: This study detected significant changes in the sagittal, coronal, and axial plane alignment of the subtalar joint, which corresponded to a decompression of the sinus tarsi. These findings contribute to our clinical practice by demonstrating the magnitude of alteration in the subtalar joint alignment that can be expected after PCFD correction with MCO as main procedure., (Copyright © 2023. Published by Elsevier Ltd.)

Background: Foot and ankle weightbearing CT (WBCT) imaging has emerged over the past decade. However, a systematic review of diagnostic applications has not been conducted so far., Method: A systematic literature search was performed according to the Preferred Reporting Items for Systematic Reviews and Meta-Analysis (PRISMA) guidelines after Prospective Register of Systematic Reviews (PROSPERO) registration. Studies analyzing diagnostic applications of WBCT were included. Main exclusion criteria were: cadaveric specimens and simulated WBCT. The Methodological Index for Non-Randomized Studies (MINORS) was used for quality assessment., Results: A total of 78 studies were eligible for review. Diagnostic applications were identified in following anatomical area's: ankle (n = 14); hindfoot (n = 41); midfoot (n = 4); forefoot (n = 19). Diagnostic applications that could not be used on weightbearing radiographs (WBRX) were reported in 56/78 studies. The mean MINORS was 9.8/24 (range: 8-12)., Conclusion: Diagnostic applications of WBCT were most frequent in the hindfoot, but other areas are on the rise. Post-processing of images was the main benefit compared to WBRX based on a moderate quality of the identified studies., (Copyright © 2023 European Foot and Ankle Society. Published by Elsevier Ltd. All rights reserved.)

Competing Interests: The author certifies that there are no funding or commercial associations (consultancies, stock ownership, equity interest, patent/licensing arrangements, etc.) that might pose a conflict of interest in connection with the submitted article related to the author or any immediate family members. All ICMJE Conflict of Interest Forms for authors and Clinical Orthopaedics and Related Research® editors and board members are on file with the publication and can be viewed on request.

Caterpillar salivary glucose oxidase (GOX) can function as both an elicitor or as an effector of plant defense responses depending upon the system. Treatment with GOX reduces the stomatal aperture of tomato and soybean leaves, thereby reducing the emission of volatile organic compounds (VOCs), that are important indirect defense responses of plants by attracting natural enemies of the caterpillars. Here we examined the effect of fungal GOX (fungal glucose oxidases have been used to determine specificity in defense response elicitation) on stomatal closure of maize leaves and on the volatile emission pattern whole maize plants. We also used salivary gland homogenate from wild-type and CRISPR-Cas9 Helicoverpa zea mutants deficient in GOX activity to determine the effect caterpillar saliva with and without GOX had on maize volatile emission. Collecting volatiles at 2-hour intervals allowed us to examine the changes in emission over time. Fungal GOX reduced the stomatal aperture in maize leaves, which may have influenced the observed significant reduction in total green leaf volatile (GLV) emission. Furthermore, fungal GOX significantly increased the emission of several key terpenes: linalool, DMNT, and Z-β-farnesene from maize, while salivary gland homogenate from wild type (WT; GOX+) H. zea increased the emission of α-pinene, β-pinene, and ocimene compared to H. zea unable to synthesize GOX. This study addressed a significant knowledge gap about the effect of GOX on maize volatiles and provides a baseline for further research on the effect of GOX on the regulation of terpene synthase genes and their relation to terpene volatile emission., (© 2023. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

Detection of syndesmotic ankle instability remains challenging in clinical practice due to the limitations of two-dimensional (2D) measurements. The transition to automated three-dimensional (3D) measurement techniques is on the verge of a breakthrough but normative and side-to-side comparative data are missing. Therefore, our study aim was two-fold: (1) to establish 3D anatomical reference values of the ankle syndesmosis based on automated measurements and (2) to determine to what extent the ankle syndesmosis is symmetric across all 3D measurements. Patients without syndesmotic pathology with a non-weight-bearing CT scan (NWBCT; N = 38; Age = 51.6 ± 17.43 years) and weight-bearing CT scan (WBCT; N = 43; Age = 48.9 ± 14.3 years) were retrospectively included. After training and validation of a neural network to automate the segmentation of 3D ankle models, an iterative closest point registration was performed to superimpose the left on the right ankle. Subsequently, 3D measurements were manually and automatically computed using a custom-made algorithm and side-to-side comparison of these landmarks allowed one to investigate symmetry. Intra-observer analysis showed excellent agreements for all manual measurements (ICC range 0.85-0.99) and good (i.e. < 2.7° for the angles and < 0.5 mm for the distances) accuracy was found between the automated and manual measurements. A mean Dice coefficient of 0.99 was found for the automated segmentation framework. The established mean, standard deviation and range were provided for each 3D measurement. From these data, reference values were derived to differ physiological from pathological syndesmotic alignment. Furthermore, side-to-side symmetry was revealed when comparing left to right measurements (P > 0.05). In clinical practice, our novel algorithm could surmount the current limitations of manual 2D measurements and distinguish patients with a syndesmotic ankle lesion from normal variance., (© 2023. Springer Nature Limited.)

Insect herbivores face multiple challenges to their ability to grow and reproduce. Plants can produce a series of defenses that disrupt and damage the herbivore digestive system, which are heightened upon injury by insect feeding. Additionally, insects face threats from virulent microorganisms that can incur their own set of potential costs to hosts. Microorganisms that invade through the digestive system may function in concert with defenses generated by plants, creating combined assailments on host insects. In our study, we evaluated how tomato defenses interact with an enteric bacterial isolate, Serratia marcescens, in the corn earworm (Helicoverpa zea). We performed bioassays using different tomato cultivars that were induced by methyl jasmonate and larvae orally inoculated with a S. marcescens isolate. Untreated corn earworm larval mortality was low on constitutive tomato, while larvae inoculated with S. marcescens exhibited > 50% mortality within 5 days. Induction treatments elevated both control mortality (~ 45%) and in combination with S. marcescens (> 95%). Larvae also died faster when encountering induced defenses and Serratia. Using a tomato mutant, foliar polyphenol oxidase activity likely had stronger impacts on S. marcescens-mediated larval mortality. Induction treatments also elevated the number of bacterial colony-forming units in the hemolymph of larvae inoculated with Serratia. Larval mortality by S. marcescens was low (< 10%) on artificial diets. Our results demonstrate that plant chemical defenses enhance larval mortality from an opportunistic gut microbe. We propose that the combined damage from both the plant and microbial agent overwhelm the herbivore to increase mortality rates and expedite host death., (© 2023. This is a U.S. Government work and not under copyright protection in the US; foreign copyright protection may apply.)

Forced external rotation is hypothesized as the key mechanism of syndesmotic ankle injuries, inducing a three-dimensional deviation from the normal distal tibiofibular joint (DTFJ) alignment. However, current diagnostic imaging modalities are impeded by a two-dimensional assessment, without considering ligamentous stabilizers. Therefore, our aim is threefold: (1) to construct an articulated statistical shape model of the normal DTFJ with the inclusion of ligamentous morphometry, (2) to investigate the effect of weight-bearing on the DTFJ alignment, and (3) to detect differences in predicted syndesmotic ligament length of patients with syndesmotic lesions with respect to normative data. Training data comprised non-weight-bearing CT scans from asymptomatic controls (N = 76), weight-bearing CT scans from patients with syndesmotic ankle injury (N = 13), and their weight-bearing healthy contralateral side (N = 13). Path and length of the syndesmotic ligaments were predicted using a discrete element model, wrapped around bony contours. Statistical shape model evaluation was based on accuracy, generalization, and compactness. The predicted ligament length in patients with syndesmotic lesions was compared with healthy controls. With respect to the first aim, our presented skeletal shape model described the training data with an accuracy of 0.23 ± 0.028 mm. Mean prediction accuracy of ligament insertions was 0.53 ± 0.12 mm. In accordance with the second aim, our results showed an increased tibiofibular diastasis in healthy ankles after weight-bearing. Concerning our third aim, a statistically significant difference in anterior syndesmotic ligament length was found between ankles with syndesmotic lesions and healthy controls (p = 0.017). There was a significant correlation between the presence of syndesmotic injury and the positional alignment between the distal tibia and fibula (r = 0.873, p < 0,001). Clinical Significance: Statistical shape modeling combined with patient-specific ligament wrapping techniques can facilitate the diagnostic workup of syndesmosic ankle lesions under weight-bearing conditions. In doing so, an increased anterior tibiofibular distance was detected, corresponding to an "anterior open-book injury" of the ankle syndesmosis as a result of anterior inferior tibiofibular ligament elongation/rupture., (© 2022 Orthopaedic Research Society. Published by Wiley Periodicals LLC.)

Insect guts often harbor an abundance of bacteria. Many of these members are commensal, but some may emerge as opportunistic pathogens when the host is under stress. In this study, we evaluated how dietary nutritional concentration mediates a shift from commensal to pathogenic, and if host species influences those interactions. We used the lepidopterans (Noctuidae) fall armyworm (Spodoptera frugiperda), beet armyworm (Spodoptera exigua), and corn earworm (Helicoverpa zea) as hosts and a Serratia strain initially isolated from healthy fall armyworm. Diet concentration was altered by bulk reduction in nutritional content with dilution using cellulose. Our experiments revealed that low nutrient diet increased mortality from Serratia for beet armyworm and corn earworm. However, for fall armyworm, little mortality was observed in any of the diet combinations. Dietary nutrition and oral inoculation with Serratia did not change the expression of two antimicrobial peptides in fall and beet armyworm, suggesting that other mechanisms that mediate mortality were involved. Our results have implications for how pathogens may persist as commensals in the digestive tract of insects. These findings also suggest that diet plays a very important role in the switch from commensal to pathogen. Finally, our data indicate that the host response to changing conditions is critical in determining if a pathogen may overtake its host and that these three lepidopteran species have different responses to opportunistic enteric pathogens., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Published by Elsevier Inc.)

Gut microbiota can have diverse impacts on hosts, the nature of which often depend on the circumstances. For insect gut microbes, the quality and nature of host diets can be a significant force in swinging the pendulum from inconsequential to functionally important. In our study, we addressed whether beneficial microbes in one species impart similar functions to related species under identical conditions. Using fall armyworm (Spodoptera frugiperda), beet armyworm (Spodoptera exigua), and other noctuid hosts, we implemented an axenic rearing strategy and manipulated gut bacterial populations and dietary conditions. Our results revealed that some gut Enterococcus and Enterobacter isolates can facilitate utilization of a poor diet substrate by fall armyworm, but this was not the case for other more optimized diets. While Enterococcus provided benefits to fall armyworm, it was decidedly antagonistic to beet armyworm (Spodoptera exigua) under identical conditions. Unique isolates and bacterial introductions at early growth stages were critical to how both larval hosts performed. Our results provide robust evidence of the roles in which bacteria support lepidopteran larval growth, but also indicate that the directionality of these relationships can differ among congener hosts. IMPORTANCE Insects have intimate relationships with gut microbiota, where bacteria can contribute important functions to their invertebrate hosts. Lepidopterans are important insect pests, but how they engage with their gut bacteria and how that translates to impacts on the host are lacking. Here we demonstrate the facultative nature of gut microbiota in lepidopteran larvae and the importance of diet in driving mutualistic or antagonistic relationships. Using multiple lepidopteran species, we uncover that the same bacteria that can facilitate exploitation of a challenging diet in one host severely diminishes larval performance of another larval species. Additionally, we demonstrate the beneficial functions of gut microbiota on the hosts are not limited to one lineage, but rather multiple isolates can facilitate the exploitation of a suboptimal diet. Our results illuminate the context-dependent nature of the gut microbiomes in invertebrates, and how host-specific microbial engagement can produce dramatically different interactions.

Aspergillus fumigatus is one of the deadliest fungal species, causing hundreds of thousands of deaths each year. Because azoles provide the preferred first-line option for treatment of aspergillosis, the increase in rates of resistance and the poor therapeutic outcomes for patients infected with a resistant isolate constitute a serious global health threat. Azole resistance is frequently associated with specific tandem repeat duplications of a promoter element upstream of cyp51A , the gene that encodes the target for this drug class in A. fumigatus. This promoter element is recognized by the activating transcription factors SrbA and AtrR. This region also provides a docking platform for the CCAAT-binding complex (CBC) and HapX, which cooperate in the regulation of genes involved in iron-consuming pathways, including cyp51A . Here, we studied the regulatory contributions of SrbA, AtrR, CBC, and HapX binding sites to cyp51A expression and azole resistance under different iron availability employing promoter mutational analysis and protein-DNA interaction analysis. This strategy revealed iron status-dependent and -independent roles of these regulatory elements. We show that promoter occupation by both AtrR and SrbA is required for iron-independent steady-state transcriptional activation of cyp51A and its induction during short-term iron exposure relies on HapX binding. We further reveal the HapX binding site as a repressor element, disruption of which increases cyp51A expression and azole resistance regardless of iron availability. IMPORTANCE First-line treatment of aspergillosis typically involves the use of azole antifungals. Worryingly, their future clinical use is challenged by an alarming increase in resistance. Therapeutic outcomes for such patients are poor due to delays in switching to alternative treatments and reduced efficacy of salvage therapeutics. Our lack of understanding of the molecular mechanisms that underpin resistance hampers our ability to develop novel therapeutic interventions. In this work, we dissect the regulatory motifs associated with azole resistance in the promoter of the gene that encodes the azole drug target Cyp51A. These motifs include binding platforms for SrbA and AtrR, as well as the CCAAT-binding complex and HapX. Employing mutational analyses, we uncovered crucial cyp51A -activating and -repressing functions of the binding sites. Remarkably, disrupting binding of the iron regulator HapX increased cyp51A expression and azole resistance in an iron-independent manner.

Bacterial gut symbionts of insect herbivores can impact their host through different mechanisms. However, in most lepidopteran systems we lack experimental examples to explain how specific members of the gut bacterial community influence their host. We used fall armyworm (Spodoptera frugiperda) as a model system to address this objective. We implemented axenic and gnotobiotic techniques using two semi-artificial diets with pinto bean and wheat germ-based components. Following an initial screen of bacterial isolates representing different genera, larvae inoculated with Enterococcus FAW 2-1 exhibited increased body mass on the pinto bean diet, but not on the wheat germ diet. We conducted a systematic bioassay screening of Enterococcus isolated from fall armyworm, revealing they had divergent effects on the hosts' usage pinto bean diet, even among phylogenetically similar isolates. Dilution of the pinto bean diet revealed that larvae performed better on less-concentrated diets, suggesting the presence of a potential toxin. Collectively, these results demonstrate that some gut microorganisms of lepidopterans can benefit the host, but the dietary context is key towards understanding the direction of the response and magnitude of the effect. We provide evidence that gut microorganisms may play a wider role in mediating feeding breadth in lepidopteran pests, but overall impacts could be related to the environmental stress and the metabolic potentials of the microorganisms inhabiting the gut., (Published by Elsevier Ltd.)

Insects frequently confront different microbial assemblages. Bacteria inhabiting an insect gut are often commensal, but some can become pathogenic when the insect is compromised from different stressors. Herbivores are often confronted by various forms of plant resistance, but how defenses generate opportunistic microbial infections from residents in the gut are not well understood. In this study, we evaluated the pathogenic tendencies of Serratia isolated from the digestive system of healthy fall armyworm larvae (Spodoptera frugiperda) and how it interfaces with plant defenses. We initially selected Serratia strains that varied in their direct expression of virulence factors. Inoculation of the different isolates into the fall armyworm body cavity indicated differing levels of pathogenicity, with some strains exhibiting no effects while others causing mortality 24 h after injection. Oral inoculations of pathogens on larvae provided artificial diets caused marginal (< 7%) mortality. However, when insects were provided different maize genotypes, mortality from Serratia increased and was higher on plants exhibiting elevated levels of herbivore resistance (< 50% mortality). Maize defenses facilitated an initial invasion of pathogenic Serratia into the larval hemocoel¸ which was capable of overcoming insect antimicrobial defenses. Tomato and soybean further indicated elevated mortality due to Serratia compared to artificial diets and differences between plant genotypes. Our results indicate plants can facilitate the incipient emergence of pathobionts within gut of fall armyworm. The ability of resident gut bacteria to switch from a commensal to pathogenic lifestyle has significant ramifications for the host and is likely a broader phenomenon in multitrophic interactions facilitated by plant defenses., (© 2021. This is a U.S. government work and not under copyright protection in the U.S.; foreign copyright protection may apply.)

Koinobiont endoparasitoid wasps whose larvae develop inside a host insect alter several important facets of host physiology, potentially causing cascading effects across multiple trophic levels. For instance, the hijacking of the host immune responses may have effects on how insects interact with host plants and microbial associates. However, the parasitoid regulation of insect-plant-microbiome interactions is still understudied. In this study, we used the fall armyworm (FAW), Spodoptera frugiperda , and the braconid parasitoid Cotesia marginiventris to evaluate impacts of parasitism on the gut microbiome of FAW larvae, and respective maize plant defense responses. The level of reactive oxygen species and the microbial community in larval gut underwent significant changes in response to parasitism, leading to a significant reduction of Enterococcus , while elevating the relative abundance of Pseudomonas . FAW with parasitism had lower glucose oxidase (GOX) activity in salivary glands and triggered lower defense responses in maize plants. These changes corresponded to effects on plants, as Pseudomonas inoculated larvae had lower activity of salivary GOX and triggered lower defense responses in maize plants. Our results demonstrated that parasitism had cascading effects on microbial associates across trophic levels and also highlighted that insect gut bacteria may contribute to complex interrelationships among parasitoids, herbivores, and plants., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Wang, Mason, Ju, Xue, Tong, Peiffer, Song, Zeng and Felton.)

Objective: The role of the syndesmotic ankle ligaments as extrinsic stabilizers of the distal tibiofibular joint (DTFJ) has been studied extensively in patients with high ankle sprains (HAS). However, research concerning the fibular incisura as intrinsic stabilizer of the DTFJ has been obscured by a two-dimensional assessment of a three-dimensional structure. Therefore, we aimed to compare the morphometry of the incisura fibularis between patients with HAS and a control group using three-dimensional radiographic techniques., Materials and Methods: Fifteen patients with a mean age of 44 years (SD = 15.2) diagnosed with an unstable HAS and twenty-five control subjects with a mean age of 47.4 years (SD = 6.5) were analyzed in this retrospective comparative study. The obtained CT images were converted to three-dimensional models, and the following radiographic parameters of the incisura fibularis were determined using three-dimensional measurements: incisura width, incisura depth, incisura height, incisura angle, incisura width-depth ratio, and incisura-tibia ratio., Results: The mean incisura depth (M = 4.7 mm, SD = 1.1 mm), incisura height (M = 36.1 mm, SD = 5.3 mm), and incisura angle of the control group (M = 137.2°, SD = 7.9°) differed significantly from patients with a HAS (resp., M = 3.8 mm, SD = 1.1 mm; M = 31.9 mm, SD = 3.2 mm; M = 143.2°, SD = 8.3°) (P < 0.05). The incisura width, incisura width-depth ratio, and incisura-tibia ratio demonstrated no significant difference (P > 0.05)., Conclusion: Our three-dimensional comparative analysis has detected a shallower and shorter fibular incisura in patients with HAS. This distinct morphology could have repercussion on the intrinsic or osseous stability of the DTFJ. Future prospective radiographic assessment could determine to what extend the fibular incisura morphology contributes to syndesmotic ankle injuries caused by high ankle sprains.

Herbivore-induced plant volatiles (HIPVs) are widely recognized as an ecologically important defensive response of plants against herbivory. Although the induction of this 'cry for help' has been well documented, only a few studies have investigated the inhibition of HIPVs by herbivores and little is known about whether herbivores have evolved mechanisms to inhibit the release of HIPVs. To examine the role of herbivore effectors in modulating HIPVs and stomatal dynamics, we conducted series of experiments combining pharmacological, surgical, genetic (CRISPR-Cas9) and chemical (GC-MS analysis) approaches. We show that the salivary enzyme, glucose oxidase (GOX), secreted by the caterpillar Helicoverpa zea on leaves, causes stomatal closure in tomato (Solanum lycopersicum) within 5 min, and in both tomato and soybean (Glycine max) for at least 48 h. GOX also inhibits the emission of several HIPVs during feeding by H. zea, including (Z)-3-hexenol, (Z)-jasmone and (Z)-3-hexenyl acetate, which are important airborne signals in plant defenses. Our findings highlight a potential adaptive strategy where an insect herbivore inhibits plant airborne defenses during feeding by exploiting the association between stomatal dynamics and HIPV emission., (© 2021 The Authors. New Phytologist © 2021 New Phytologist Foundation.)

Silicon (Si) is a beneficial mineral that enhances plant protection against abiotic and biotic stresses, including insect herbivores. Si increases mechanical and biochemical defenses in a variety of plant species. However, the use of Si in agriculture remains poorly adopted despite its widely documented benefits in plant health. In this study, we tested the effect of Si supplementation on the induction of plant resistance against a chewing herbivore in crops with differential ability to accumulate this element. Our model system comprised the generalist herbivore fall armyworm (FAW) Spodoptera frugiperda and three economically important plant species with differential ability to uptake silicon: tomato (non-Si accumulator), soybean, and maize (Si-accumulators). We investigated the effects of Si supply and insect herbivory on the induction of physical and biochemical plant defenses, and herbivore growth using potted plants in greenhouse conditions. Herbivory and Si supply increased peroxidase (POX) activity and trichome density in tomato, and the concentration of phenolics in soybean. Si supplementation increased leaf Si concentration in all plants. Previous herbivory affected FAW larval weight gain in all plants tested, and the Si treatment further reduced weight gain of larvae fed on Si accumulator plants. Notably, our results strongly suggest that non-glandular trichomes are important reservoirs of Si in maize and may increase plant resistance to chewing herbivores. We conclude that Si offers transient resistance to FAW in soybean, and a more lasting resistance in maize. Si supply is a promising strategy in management programs of chewing herbivores in Si-accumulator plants., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Acevedo, Peiffer, Ray, Tan and Felton.)