Spot(ters) the difference: Bringing traditional anatomical examinations online.

The COVID-19 pandemic caused a shift in anatomy education forcing institutions to find innovative ways to teach and assess online. This study details the development of an online spotter across multiple modules that allowed students to sit the examination at home whilst still maintaining the integrity of the assessment. The online spotter consisted of individual, Zoom calls between students and examiners whereby slides with images and questions were screen shared. To examine the viability of this spotter in non-lockdown scenarios several parameters were considered. Mean marks were compared to traditional versions and Pearson's r correlation coefficients were calculated between online and traditional spotters and between online spotters and overall performance in anatomy modules. A survey was carried out to determine the students' view of the assessment. Pearson's r was between 0.33 and 0.49 when comparing online spotters to the traditional format, and between 0.65 and 0.75 (p < 0.01) when compared to a calculated anatomy score. The survey indicated overall student satisfaction as 82.5% reported that it was a fair way to test their knowledge and 55% reported the same or lower levels of anxiety when compared to traditional spotters. However, there was nothing to indicate that the students preferred this format over laboratory-based spotters. These results indicate that this new exam format would be useful for small cohorts who are undertaking online or hybrid courses, or in circumstances when running a full spotter is too costly, and represents a fair and robust way to assess practical anatomical knowledge online.

Cross-resistance of barnyardgrass [Echinochloa crus-galli (L.) P. Beauv.] to aryloxyphenoxypropionate herbicides.

Managing emerged weeds that have evolved resistance to acetyl CoA carboxylase (ACCase)-inhibiting herbicides is a challenging task. A dose-response experiment was conducted on barnyardgrass biotypes resistant (R) and susceptible (S) to three aryloxyphenoxypropionate herbicides cyhalofop-butyl (CyB), fenoxaprop-ethyl (FeE), and quizalofop-ethyl (QuE) along with investigations into the potential resistance mechanism of these biotypes. The tested R barnyardgrass biotypes had strong resistance to CyB and FeE (resistant/susceptible ratio: 7.9-14.4) but weak resistance to QuE (resistant/susceptible ratio: 2.4-3.1). Absorption, translocation, and total metabolism of CyB and QuE were not associated with differences among S and R barnyardgrass biotypes. However, differences between S and R barnyardgrass were observed in production of active acid forms of each herbicide (cyhalofop-acid and quizalofop-acid). Production of cyhalofop-acid was >1.6-fold less in R barnyardgrass (3-8%) for 24 h after herbicide application than in the S barnyardgrass (8-16%). Meanwhile, production of quizalofop-acid was less in R barnyardgrass (< 14%) throughout the study period than in the S barnyardgrass (< 22%). Sequencing results of ACCase gene showed no difference between S and R barnyardgrass. Overall results show that a non-target-site resistance mechanism altering metabolism of CyB and QuE likely contributes to resistance of the barnyardgrass biotypes to these herbicides.

Absorption, translocation, and metabolism of florpyrauxifen-benzyl and cyhalofop-butyl in cyhalofop-butyl-resistant barnyardgrass [Echinochloa crus-galli (L.) P. Beauv.].

Dose-response experiments were conducted to assess the sensitivity of one susceptible and three putative resistant (R1, R2, and R3) barnyardgrass [Echinochloa crus-galli (L.) P. Beauv.] biotypes to florpyrauxifen-benzyl and cyhalofop-butyl alone and as a formulated premix. Subsequently, potential resistance mechanisms of the barnyardgrass were evaluated. Based on biomass reduction results, resistant/susceptible ratios were calculated for R1 (7.0-50), R2 (7.0-150), and R3 (18-214) biotypes. Absorption and translocation of [14C]-florpyrauxifen-benzyl decreased in R1 and R3 biotypes, but not for [14C]-cyhalofop-butyl. The metabolism of [14C]-florpyrauxifen-benzyl to [14C]-florpyrauxifen-acid was >2-fold less in resistant biotypes (9-11%) than in the susceptible biotype (23%). Moreover, the production of [14C]-florpyrauxifen-acid in susceptible barnyardgrass (not in the R biotypes) increased 3-fold when florpyrauxifen-benzyl and cyhalofop-butyl were applied in mixture compared to florpyrauxifen-benzyl applied alone. The tested barnyardgrass biotypes had no mutation in the Transport Inhibitor Response1, auxin-signaling F-box, and acetyl coenzyme A carboxylase genes. Although further studies on cyhalofop-butyl resistance with respect to analysis of specific metabolites are needed, our findings in this study demonstrates that the evolution of florpyrauxifen-benzyl resistance in multiple resistant barnyardgrass can be related to non-target-site resistance mechanisms reducing absorption and translocation of the herbicide and causing reduced conversion or rapid degradation of florpyrauxifen-acid.

Evaluation of dicamba volatilization when mixed with glyphosate using imazethapyr as a tracer.

Expansion of dicamba-resistant crops increased the frequency of off-target movement issues, especially in the midsouthern United States. Six field trials were conducted over two growing seasons with the purpose to determine the contribution of volatilization and physical suspension of particles to the off-target movement of dicamba when applied with glyphosate and imazethapyr - a non-volatile herbicide used as a tracer for physical off-target movement. Applications included dicamba at 560 g ha-1, glyphosate at 1260 g ha-1, and imazethapyr at 105 g ha-1. Applicators include glyphosate with dicamba to increase the spectrum of weed control from these applications; however, this addition increases potential for dicamba volatilization. Following application of the mixture, air samplers were placed in the field to collect dicamba and imazethapyr. Results showed there was at least 50 times more dicamba than imazethapyr detected even though the dicamba:imazethapyr ratio applied was 5.3:1. Dicamba was detected in the treated area and the off-site locations and all intervals of air sampling, ranging from 126 to 5990 ng. No more than 37.5 ng of imazethapyr was detected during the first 24-h after application (HAA) inside the treated area. Imazethapyr was only detected in 9 of the 20 sampling combinations during these experiments, and most of these detections (6) occurred during the first 24 HAA and inside the treated area. While some movement from the suspension of particles occurred based on the detection of imazethapyr in air samples, results show that most dicamba detection was due to the volatilization of the herbicide.

Extracellular matrix protein anosmin-1 modulates olfactory ensheathing cell maturation in chick olfactory bulb development.

Olfactory ensheathing cells (OECs) are a specialized class of glia, wrapping around olfactory sensory axons that target the olfactory bulb (OB) and cross the peripheral nervous system/central nervous system boundary during development and continue to do so post-natally. OEC subpopulations perform distinct subtype-specific functions dependent on their maturity status. Disrupted OEC development is thought to be associated with abnormal OB morphogenesis, leading to anosmia, a defining characteristic of Kallmann syndrome. Hence, anosmin-1 encoded by Kallmann syndrome gene (KAL-1) might modulate OEC differentiation/maturation in the OB. We performed in ovo electroporation of shRNA in the olfactory placode to knock-down kal in chick embryos, resulting in abnormal OB morphogenesis and loss of olfactory sensory axonal innervation into OB. BLBP-expressing OECs appeared to form a thinner and poorly organized outmost OB layer where SOX10 expressing OECs were completely absent with emergence of GFAP-expressing OECs. Furthermore, in embryonic day 10 chick OB explant cultures, GFAP expression in OECs accumulating along the OB nerve layers was dramatically reduced by recombinant anosmin-1. We then purified immature OECs from embryonic day 10 chick OB. These cells express GFAP after 7 days in vitro, exhibiting a multipolar morphology. Overexpression of chick anosmin, exogenous anosmin-1 or FGF2 could inhibit GFAP expression with cells presenting elongated morphology, which was blocked by the FGF receptor inhibitor Su5402. These data demonstrate that anosmin-1 functions via FGF signalling in regulating OEC maturation, thereby providing a permissive glial environment for axonal innervation into the OB during development.

Development of the cerebellum: simple steps to make a 'little brain'.

The cerebellum is a pre-eminent model for the study of neurogenesis and circuit assembly. Increasing interest in the cerebellum as a participant in higher cognitive processes and as a locus for a range of disorders and diseases make this simple yet elusive structure an important model in a number of fields. In recent years, our understanding of some of the more familiar aspects of cerebellar growth, such as its territorial allocation and the origin of its various cell types, has undergone major recalibration. Furthermore, owing to its stereotyped circuitry across a range of species, insights from a variety of species have contributed to an increasingly rich picture of how this system develops. Here, we review these recent advances and explore three distinct aspects of cerebellar development - allocation of the cerebellar anlage, the significance of transit amplification and the generation of neuronal diversity - each defined by distinct regulatory mechanisms and each with special significance for health and disease.

Transit amplification in the amniote cerebellum evolved via a heterochronic shift in NeuroD1 expression.

The cerebellum has evolved elaborate foliation in the amniote lineage as a consequence of extensive Atoh1-mediated transit amplification in an external germinal layer (EGL) comprising granule cell precursors. To explore the evolutionary origin of this layer, we have examined the molecular geography of cerebellar development throughout the life cycle of Xenopus laevis. At metamorphic stages Xenopus displays a superficial granule cell layer that is not proliferative and expresses both Atoh1 and NeuroD1, a marker of postmitotic cerebellar granule cells. Premature misexpression of NeuroD1 in chick partially recapitulates the amphibian condition by suppressing transit amplification. However, unlike in the amphibian, granule cells fail to enter the EGL. Furthermore, misexpression of NeuroD1 once the EGL is established both triggers radial migration and downregulates Atoh1. These results show that the evolution of transit amplification in the EGL required adaptation of NeuroD1, both in the timing of its expression and in its regulatory function, with respect to Atoh1.

The roof plate boundary is a bi-directional organiser of dorsal neural tube and choroid plexus development.

The roof plate is a signalling centre positioned at the dorsal midline of the central nervous system and generates dorsalising morphogenic signals along the length of the neuraxis. Within cranial ventricles, the roof plate gives rise to choroid plexus, which regulates the internal environment of the developing and adult brain and spinal cord via the secretion of cerebrospinal fluid. Using the fourth ventricle as our model, we show that the organiser properties of the roof plate are determined by its boundaries with the adjacent neuroepithelium. Through a combination of in ovo transplantation, co-culture and electroporation techniques in chick embryos between embryonic days 3 and 6, we demonstrate that organiser properties are maintained by interactions between the non-neural roof plate and the neural rhombic lip. At the molecular level, this interaction is mediated by Delta-Notch signalling and upregulation of the chick homologue of Hes1: chairy2. Gain- and loss-of-function approaches reveal that cdelta1 is both necessary and sufficient for organiser function. Our results also demonstrate that while chairy2 is specifically required for the maintenance of the organiser, its ectopic expression is not sufficient to recapitulate organiser properties. Expression of atonal1 in the rhombic lip adjacent at the roof plate boundary is acutely dependent on both boundary cell interactions and Delta-Notch signalling. Correspondingly, the roof plate boundary organiser also signals to the roof plate itself to specify the expression of early choroid plexus markers. Thus, the roof plate boundary organiser signals bi-directionally to acutely coordinate the development of adjacent neural and non-neural tissues.

Making teeth to order: conserved genes reveal an ancient molecular pattern in paddlefish (Actinopterygii).

Ray-finned fishes (Actinopterygii) are the dominant vertebrate group today (+30 000 species, predominantly teleosts), with great morphological diversity, including their dentitions. How dental morphological variation evolved is best addressed by considering a range of taxa across actinopterygian phylogeny; here we examine the dentition of Polyodon spathula (American paddlefish), assigned to the basal group Acipenseriformes. Although teeth are present and functional in young individuals of Polyodon, they are completely absent in adults. Our current understanding of developmental genes operating in the dentition is primarily restricted to teleosts; we show that shh and bmp4, as highly conserved epithelial and mesenchymal genes for gnathostome tooth development, are similarly expressed at Polyodon tooth loci, thus extending this conserved developmental pattern within the Actinopterygii. These genes map spatio-temporal tooth initiation in Polyodon larvae and provide new data in both oral and pharyngeal tooth sites. Variation in cellular intensity of shh maps timing of tooth morphogenesis, revealing a second odontogenic wave as alternate sites within tooth rows, a dental pattern also present in more derived actinopterygians. Developmental timing for each tooth field in Polyodon follows a gradient, from rostral to caudal and ventral to dorsal, repeated during subsequent loss of teeth. The transitory Polyodon dentition is modified by cessation of tooth addition and loss. As such, Polyodon represents a basal actinopterygian model for the evolution of developmental novelty: initial conservation, followed by tooth loss, accommodating the adult trophic modification to filter-feeding.

The evolution of the vertebrate cerebellum: absence of a proliferative external granule layer in a non-teleost ray-finned fish.

The cerebellum represents one of the most morphologically variable structures in the vertebrate brain. To shed light on its evolutionary history, we have examined the molecular anatomy and proliferation of the developing cerebellum of the North American paddlefish, Polyodon spathula. Absence of an external proliferative cerebellar layer and the restriction of Atonal1 expression to the rhombic lip and valvular primordium demonstrate that transit amplification in a cerebellar external germinal layer, a prominent feature of amniote cerebellum development, is absent in paddlefish. Furthermore, expression of Sonic hedgehog, which drives secondary proliferation in the mouse cerebellum, is absent from the paddlefish cerebellum. These data are consistent with what has been observed in zebrafish and suggest that the transit amplification seen in the amniote cerebellum was either lost very early in the ray-finned fish lineage or evolved in the lobe-finned fish lineage. We also suggest that the Atoh1-positive proliferative valvular primordium may represent a synapomorphy (shared derived character) of ray-finned fishes. The topology of valvular primordium development in paddlefish differs significantly from that of zebrafish and correlates with the adult cerebellar form. The distribution of proliferative granule cell precursors in different vertebrate taxa is thus the likely determining factor in cerebellar morphological diversity.

The amphioxus genome and the evolution of the chordate karyotype.

Lancelets ('amphioxus') are the modern survivors of an ancient chordate lineage, with a fossil record dating back to the Cambrian period. Here we describe the structure and gene content of the highly polymorphic approximately 520-megabase genome of the Florida lancelet Branchiostoma floridae, and analyse it in the context of chordate evolution. Whole-genome comparisons illuminate the murky relationships among the three chordate groups (tunicates, lancelets and vertebrates), and allow not only reconstruction of the gene complement of the last common chordate ancestor but also partial reconstruction of its genomic organization, as well as a description of two genome-wide duplications and subsequent reorganizations in the vertebrate lineage. These genome-scale events shaped the vertebrate genome and provided additional genetic variation for exploitation during vertebrate evolution.

Impact of volatility reduction agents on dicamba and glyphosate spray solution pH, droplet dynamics, and weed control.

BACKGROUND: Regulations in 2021 required the addition of a volatility reduction agent (VRA) to dicamba spray mixtures for postemergence weed control. Understanding the impact of VRAs on weed control, droplet dynamics, and spray pH is essential. RESULTS: Adding glyphosate to dicamba decreased the solution pH by 0.63 to 1.85 units. Across locations, potassium carbonate increased the tank-mixture pH by 0.85 to 1.65 units while potassium acetate raised the pH by 0.46 to 0.53 units. Glyphosate and dicamba in tank-mixture reduced Palmer amaranth control by 14 percentage points compared to dicamba alone and decreased barnyardgrass control by 12 percentage points compared to glyphosate alone 4 weeks after application (WAA). VRAs resulted in a 5-percentage point reduction in barnyardgrass control 4 WAA. Common ragweed, common lambsquarters, and giant ragweed control were unaffected by herbicide solution 4 WAA. Dicamba alone produced a larger average droplet size and had the fewest driftable fines (% volume < 200 µm). Potassium acetate produced a larger droplet size than potassium carbonate for Dv0.1 and Dv0.5 . The addition of glyphosate to dicamba decreased droplet size from the entire spray droplet spectrum (Dv0.1 , Dv0.5 , Dv0.9 ). CONCLUSION: A reduction in spray pH, droplet size, and weed control was observed from mixing dicamba and glyphosate. It may be advisable to avoid tank-mixtures of these herbicides and instead, apply them sequentially to maximize effectiveness. VRAs differed in their impacts on spray solution pH and droplet dynamics, but resulted in a minimal negative to no impact on weed control. © 2022 The Authors. Pest Management Science published by John Wiley & Sons Ltd on behalf of Society of Chemical Industry.

Metabolic Exploration for Cyhalofop-Butyl Antagonism in Barnyardgrass [Echinochloa crus-galli (L.) P. Beauv.] Following Pretreatment of Malathion.

The present study investigated the effects of broad-spectrum metabolic inhibitors malathion (cytochrome P450 inhibitor) and/or 4-chloro-7-nitrobenzofurazan (NBD-Cl; glutathione S-transferase inhibitor) on the metabolism of cyhalofop-butyl (CyB) in barnyardgrass [Echinochloa crus-galli (L.) P. Beauv.] biotypes confirmed previously with multiple resistance to two herbicides CyB and florpyrauxifen-benzyl. The metabolic inhibitors were not effective at recovering the sensitivity of resistant barnyardgrass biotypes to CyB treated at the labeled rate (313 g ai ha-1). Rather, treatment with malathion followed by CyB caused antagonism, reducing the efficacy of CyB and promoting the growth of resistant biotypes. Pretreatment with malathion did not influence absorption/translocation of the applied form CyB and its conversion to the active herbicide form cyhalofop-acid (CyA), in both susceptible and resistant biotypes. In contrast, metabolism of the applied form (CyB) decreased 1.5 to 10.5 times by the malathion pretreatment. Taken together, the maintained CyA production against the reduced CyB metabolism could be the mechanism to account for the cause of CyB antagonism observed in barnyardgrass following malathion pretreatment. Additionally, the evolution of CyB resistance in barnyardgrass might be associated with reduced production of CyA in resistant biotypes, independent of activities of cytochrome P450 or GST enzymes.

Exploratory Analysis on Herbicide Metabolism and Very-Long-Chain Fatty Acid Production in Metolachlor-Resistant Palmer Amaranth (Amaranthus palmeri S. Wats.).

A Palmer amaranth (Amaranthus palmeri S. Wats.) biotype resistant to S-metolachlor was confirmed from crop fields in Arkansas, USA. This study investigated the metabolic effects of malathion (cytochrome P450 inhibitor) and 4-chloro-7-nitrobenzofurazan [NBD-Cl; glutathione S-transferase inhibitor] on the S-metolachlor-resistant A. palmeri biotype. Root elongation of the resistant biotype was 20% more inhibited by treatment of NBD-Cl (50 nM) and S-metolachlor (2 µM) in mixture than by treatment of S-metolachlor alone. Metabolites of S-metolachlor were 1.4-12.1 times greater produced in the resistant biotype for 7 d than in the susceptible standard. Production of cerotic acid, one of the very-long-chain fatty acids containing 26 carbons, was more reduced in the susceptible standard (3.8-fold) than in the resistant biotype (1.8-fold) by S-metolachlor treatment. Conclusively, evolution of S-metolachlor resistance observed in this study was likely associated with improved activity of glutathione S-transferases. Further studies are needed to genetically evaluate plant endogenous enzymes involving cerotic acid production.

Utility of roller wiper applications of dicamba for Palmer amaranth control in soybean.

BACKGROUND: The commercialization of dicamba-resistant soybean has resulted in increased concern for off-target movement of dicamba onto sensitive vegetation. To mitigate the off-target movement through physical drift, one might consider use of rope wicks and other wiper applicators. Although wiper-type application methods have been efficacious in pasture settings, the utility of dicamba using wiper applicators in agronomic crops is not available in scientific literature. To determine the utility of roller wipers for dicamba applications in dicamba-resistant soybean, two separate experiments were conducted in the summer of 2020 and replicated in both Keiser and Fayetteville, AR, USA. RESULTS: Utilizing opposing application directions and a 2:1:1 ratio of water: formulated glyphosate: formulated dicamba were the most efficacious practices for controlling Palmer amaranth. The high herbicide concentrations and wiping in opposing directions increased dicamba-resistant soybean injury when the wiper contacted the crop, but no yield loss was observed because of this injury. Broadcast applications resulted in greater Palmer amaranth mortality than roller wiper applications, and the most effective roller wiper treatments were when two sequential applications were made inside the crop canopy. CONCLUSIONS: Dicamba applications require adequate coverage for optimum weed control. While efforts can be made to increase roller wiper efficacy by optimizing coverage and timing of applications, broadcast applications are superior to roller wiper applicators for weed control. Roller wiper applications did not reduce soybean yield, thus wiper-type applications may be safely used in dicamba-resistant soybean, albeit the likelihood for off-target damage caused by volatilization of these treatments would need to be investigated. © 2022 The Authors. Pest Management Science published by John Wiley & Sons Ltd on behalf of Society of Chemical Industry.

Non-target-site resistance mechanism of barnyardgrass [Echinochloa crus-galli (L.) P. Beauv.] to florpyrauxifen-benzyl.

BACKGROUND: Florpyrauxifen-benzyl (FPB) is an arylpicolinate herbicide (Group IV) for barnyardgrass control in rice. One susceptible (Sus) and three putative FPB-resistant (R1, R2, and R3) barnyardgrass biotypes were selected based on resistant/susceptible (R/S) ratios obtained from dose-response tests and used to investigate the potential resistance mechanisms. RESULTS: Based on visual control results, the R/S ratios of barnyardgrass biotypes R1, R2, and R3 were 60-, 33-, and 16-fold greater than the Sus standard, respectively. Sequencing results of TIR1 and AFB genes in the tested barnyardgrass revealed no difference between Sus and R barnyardgrass biotypes. Absorption of [14 C]-FPB in Sus barnyardgrass increased over time and reached 90%, which was >10 percentage points greater than that in R biotypes. The [14 C]-FPB absorption in all R barnyardgrass equilibrated after 48 h. For both Sus and R barnyardgrass, most [14 C]-FPB absorbed was present in the treated leaf (79.8-88.8%), followed by untreated aboveground (9.5-18.6%) and belowground tissues (1.3-2.2%). No differences in translocation were observed. Differences between Sus and R barnyardgrass biotypes were found for FPB metabolism. Production of the active metabolite, florpyrauxifen-acid, was greater in Sus barnyardgrass (21.5-52.1%) than in R barnyardgrass (5.5-34.9%). CONCLUSION: In conclusion, reductions in FPB absorption and florpyrauxifen-acid production may contribute to the inability to control barnyardgrass with FPB. © 2021 Society of Chemical Industry.

Large-scale evaluation of physical drift and volatility of 2,4-D choline in cotton: a four-year field study.

BACKGROUND: Cotton with resistance to 2,4-d choline, glufosinate and glyphosate allows over-the-top use of these herbicides for postemergence weed control. Nontransgenic cotton is highly responsive to low rates of 2,4-d, causing concern among farmers when the herbicide is applied near the crop. Injury to nearby cotton following application of 2,4-d choline is sometimes blamed on volatilization of the herbicide. RESULTS: A large-scale experiment was conducted in 2018-2021 to better understand causes of damage to nearby sensitive cotton following an application of 2,4-d choline plus glufosinate-ammonium. The herbicides were applied to 0.4 ha in the center of a 4-ha non-Enlist cotton field. At 30 min after application, air samplers were established in the treated center and outside the treated area in cardinal directions. The 72-h cumulative air concentration of 2,4-d in the swath ranged from 3.3 to 7.1 ng m-3 , and most volatile residues (5.0-25.5 ng m-3 ) were detected in samplers established in the downwind direction directly adjacent to the treated field. Cotton plants in three downwind transects that were covered for 30 min after application were not damaged by 2,4-d, whereas noncovered plants along the downwind transects were injured. No cotton injury occurred outside the treated area, except in the downwind direction during application even though wind direction changed after application. CONCLUSION: 2,4-d choline volatilizes, but findings show that the volatilization is not sufficient to damage cotton in the neighboring area following applications, pointing to the importance of applicators understanding wind direction/shift during the application along with proximity of sensitive crops in the downwind direction. © 2022 Society of Chemical Industry.