High Genetic Diversity in Predominantly Clonal Populations of the Powdery Mildew Fungus Podosphaera leucotricha from U.S. Apple Orchards.

Apple powdery mildew (APM), caused by Podosphaera leucotricha, is a constant threat to apple production worldwide. Very little is known about the biology and population structure of this pathogen in the United States and other growing regions, which affects APM management. A total of 253 P. leucotricha isolates, sampled from 10 apple orchards in Washington, New York, and Virginia, were genetically characterized with novel single sequence repeat and mating type markers. Eighty-three multilocus genotypes (MLGs) were identified, most of which were unique to a given orchard. Each isolate carried either a MAT1-1 or a MAT1-2 idiomorph at the mating type locus, indicating that P. leucotricha is heterothallic. Virulence tests on detached apple leaves showed that the 10 most frequent P. leucotricha MLGs were avirulent on a line containing a major resistance gene. Analysis of molecular variance showed significant differentiation (P < 0.001) among populations, a result supported by principal coordinate analysis revealing three genetic groups, each represented by nonoverlapping MLGs from Washington, New York, and Virginia. A Bayesian cluster analysis showed genetic heterogeneity between Washington populations, and a relative migration analysis indicated substantial gene flow among neighboring orchards. Random mating tests indicated that APM epidemics during the active cycle were dominated by clonal reproduction. However, the presence of sexual structures in orchards, the likelihood that five repeated MLGs resulted from sexual reproduction, and high genotypic diversity observed in some populations suggest that sexual spores play some role in APM epidemics. IMPORTANCE Understanding the population biology and epidemiology of plant pathogens is essential to develop effective strategies for controlling plant diseases. Herein, we gathered insights into the population biology of P. leucotricha populations from conventional and organic apple orchards in the United States. We showed genetic heterogeneity between P. leucotricha populations in Washington and structure between populations from different U.S. regions, suggesting that short-distance spore dispersal plays an important role in the disease's epidemiology. We presented evidence that P. leucotricha is heterothallic and that populations likely result from a mixed (i.e., sexual and asexual) reproductive system, revealing that the sexual stage contributes to apple powdery mildew epidemics. We showed that the major resistance gene Pl-1 is valuable for apple breeding because virulent isolates have most likely not emerged yet in U.S. commercial orchards. These results will be important to achieve sustainability of disease management strategies and maintenance of plant health in apple orchards.

Anatomical and physiological variation of the hyoid musculature during swallowing in infant pigs.

The function of a muscle is impacted by its line of action, activity timing and contractile characteristics when active, all of which have the potential to vary within a behavior. One function of the hyoid musculature is to move the hyoid bone during swallowing, yet we have little insight into how their lines of action and contractile characteristics might change during a swallow. We used an infant pig model to quantify the contractile characteristics of four hyoid muscles during a swallow using synchronized electromyography, fluoromicrometry and high-speed biplanar videofluoroscopy. We also estimated muscle line of action during a swallow using contrast-enhanced CT-scanned muscles animated to move with the hyoid bone and found that as the hyoid elevated, the line of action of the muscles attached to it became greater in depression. We also found that muscles acted eccentrically and concentrically, which was correlated with hyoid movement. This work contributes to our understanding of how the musculature powering feeding functions during swallowing.

No Evidence of Resistance to Trifloxystrobin, Triflumizole, and Boscalid in Podosphaera leucotricha Isolates From U.S. Commercial Apple Orchards.

Apple powdery mildew, caused by Podosphaera leucotricha, continues to be a challenge in commercial apple orchards in the U.S. Pacific Northwest and worldwide. In this study, P. leucotricha isolates were collected in 2018 and 2019 from two organic (baseline) and eight conventional (exposed) apple orchards in Washington, New York, and Virginia, and assessed for their sensitivity to trifloxystrobin (TRI, n = 232), triflumizole (TFZ, n = 217), and boscalid (BOS, n = 240) using a detached leaf assay. Effective concentrations inhibiting 50% growth (EC50) were not significantly different between baseline and exposed isolates, and ranged from 0.001 to 0.105, 0.09 to 6.31, and 0.05 to 2.18 µg/ml, for TRI, TFZ, and BOS, respectively. Reduction in sensitivity by factors of 105, 63, and 22 to TRI, TFZ, and BOS, respectively, were observed in some isolates, but all isolates were controlled by the commercial label rates of the three fungicides on detached leaves. Sequencing of the cytochrome b (cytb), cytochrome P450 sterol 14α-demethylase (CYP51), and the iron-sulfur protein subunit (SdhB) genes in isolates with high EC50 revealed no mutation previously reported to confer resistance to these fungicides in other fungi, and presence of a group I intron after codon 143 in the cytb gene. Significant (P < 0.001) moderate positive correlations (r = 0.38) observed between sensitivity to TRI and TFZ warrant continuous rotations of fungicides with different modes of action in conventional orchards. The established baseline sensitivities and the molecular markers will help in selecting discriminatory doses and bypassing the challenging in vivo testing for future sensitivity monitoring in P. leucotricha.

Fungal Evolution in Anthropogenic Environments: Botrytis cinerea Populations Infecting Small Fruit Hosts in the Pacific Northwest Rapidly Adapt to Human-Induced Selection Pressures.

Many fungal pathogens have short generation times, large population sizes, and mixed reproductive systems, providing high potential to adapt to heterogeneous environments of agroecosystems. Such adaptation complicates disease management and threatens food production. A better understanding of pathogen population biology in such environments is important to reveal key aspects of adaptive divergence processes to allow improved disease management. Here, we studied how evolutionary forces shape population structure of Botrytis cinerea, the causal agent of gray mold, in the Pacific Northwest agroecosystems. Populations of B. cinerea from adjacent fields of small fruit hosts were characterized by combining neutral markers (microsatellites) with markers that directly respond to human-induced selection pressures (fungicide resistance). Populations were diverse, without evidence for recombination and association of pathogen genotype with host. Populations were highly localized with limited migration even among adjacent fields within a farm. A fungicide resistance marker revealed strong selection on population structure due to fungicide use. We found no association of resistance allele with genetic background, suggesting de novo development of fungicide resistance and frequent extinction/recolonization events by different genotypes rather than the spread of resistance alleles among fields via migration of a dominant genotype. Overall our results showed that in agroecosystems, B. cinerea populations respond strongly to selection by fungicide use with greater effect on population structure compared to adaptation to host plant species. This knowledge will be used to improve disease management by developing strategies that limit pathogen local adaptation to fungicides and other human-induced selection pressures present in Pacific Northwest agroecosystems and elsewhere.IMPORTANCE Agroecosystems represent an efficient model for studying fungal adaptation and evolution in anthropogenic environments. In this work, we studied what evolutionary forces shape populations of one of the most important fungal plant pathogens, B. cinerea, in small fruit agroecosystems of the Pacific Northwest. We hypothesized that host, geographic, and anthropogenic factors of agroecosystems structure B. cinerea populations. By combining neutral markers with markers that directly respond to human-induced selection pressures, we show that pathogen populations are highly localized and that selection pressure caused by fungicide use can have a greater effect on population structure than adaptation to host. Our results give a better understanding of population biology and evolution of this important plant pathogen in heterogeneous environments but also provide a practical framework for the development of efficient management strategies by limiting pathogen adaptation to fungicides and other human-induced selection pressures present in agroecosystems of the Pacific Northwest and elsewhere.

A Genome Resource for the Apple Powdery Mildew Pathogen Podosphaera leucotricha.

Powdery mildew, caused by Podosphaera leucotricha, is an economically important disease of apple and pear trees. A single monoconidial strain (PuE-3) of this biotrophic fungus was used to extract DNA for Illumina sequencing. Data were assembled to form a draft genome of 43.8 Mb consisting of 8,921 contigs, 9,372 predicted genes, and 96.1% of complete benchmarking universal single copy orthologs (BUSCOs). This is the first reported genome sequence of P. leucotricha that will enable studies of the population biology, epidemiology, and fungicide resistance of this pathogen. Furthermore, this resource will be fundamental to uncover the genetic and molecular mechanisms of the apple-powdery mildew interaction, and support future pome fruit breeding efforts.

A new zygodactylid species indicates the persistence of stem passerines into the early Oligocene in North America.

BACKGROUND: The lake deposits of the informal Ruby Paper Shale unit, part of the Renova Formation of Montana, have yielded abundant plant fossils that document Late Eocene - Early Oligocene global cooling in western North America. A nearly complete small bird with feather impressions was recovered from this unit in in 1959, but has only been informally mentioned. RESULTS: Here we describe this fossil and identify it as a new species of Zygodactylus, a stem lineage passerine with a zygodactyl foot. The new taxon shows morphological traits that are convergent on crown Passeriformes, including an elongate hallux, reduced body size, and a comparative shortening of proximal limb elements. The fossil documents the persistence of this lineage into the earliest Oligocene (~ 33 Ma) in North America. It is the latest occurring North American species of a group that persists in Europe until the Miocene. CONCLUSIONS: Eocene-Oligocene global cooling is known to have significantly remodeled both Palearctic and Nearctic mammal faunas but its impact on related avifaunas has remained poorly understood. The geographic and temporal range expansion provided by the new taxon together with avian other taxa with limited fossil records suggests a similar pattern of retraction in North America followed by Europe.

My branch is your branch: Talar morphology correlates with relative substrate size in platyrrhines at Tiputini Biodiversity Station, Ecuador.

Given that most species of primates are predominantly arboreal, maintaining the ability to move among branches of varying sizes has presumably been a common selective force in primate evolution. However, empirical evaluations of the relationships between morphological variation and characteristics of substrate geometry, such as substrate diameter relative to an animal's body mass, have been limited by the lack of quantified substrate usage in the wild. Here we use recently published quantitative data to assess the relationships between relative substrate size and talar morphology in nine New World monkey species at the Tiputini Biodiversity Station, Ecuador. Within this sample, both fibular facet angle (the angle between the fibular facet and the trochlear rims) and body-mass-standardized area of the medial tibial facet decrease as average and maximum relative substrate size increases. Correlations between medial tibial facet area and relative substrate size are driven by the inclusion of callitrichids in this sample. Nevertheless, these findings strengthen the hypothesis that variation in fibular facet orientation and medial tibial facet area are functionally correlated with habitual degrees of pedal inversion. They also strengthen the notion that evolutionarily changing body mass could impact habitat geometry experienced by a lineage and thereby substantially impact major trends in primate morphological evolution. This study highlights the importance of empirical data on substrate use in living primates for inferring functional and evolutionary implications of morphological variation.

Palaeontological evidence for an Oligocene divergence between Old World monkeys and apes.

Apes and Old World monkeys are prominent components of modern African and Asian ecosystems, yet the earliest phases of their evolutionary history have remained largely undocumented. The absence of crown catarrhine fossils older than ∼20 million years (Myr) has stood in stark contrast to molecular divergence estimates of ∼25-30 Myr for the split between Cercopithecoidea (Old World monkeys) and Hominoidea (apes), implying long ghost lineages for both clades. Here we describe the oldest known fossil 'ape', represented by a partial mandible preserving dental features that place it with 'nyanzapithecine' stem hominoids. Additionally, we report the oldest stem member of the Old World monkey clade, represented by a lower third molar. Both specimens were recovered from a precisely dated 25.2-Myr-old stratum in the Rukwa Rift, a segment of the western branch of the East African Rift in Tanzania. These finds extend the fossil record of apes and Old World monkeys well into the Oligocene epoch of Africa, suggesting a possible link between diversification of crown catarrhines and changes in the African landscape brought about by previously unrecognized tectonic activity in the East African rift system.

Effects of substrate and phylogeny on quadrupedal gait in free-ranging platyrrhines.

OBJECTIVES: Primate diagonal sequence (DS) gaits are often argued to be an adaptation for moving and foraging in the fine-branch niche; however, existing data have come predominantly from laboratory studies that are limited in taxonomic breadth and fail to account for the structural and ecological variation of natural substrates. We test the extent to which substrate diameter and orientation influence gait sequence type and limb phase in free-ranging primates, as well as how phylogenetic relatedness might condition response patterns. MATERIALS AND METHODS: We filmed quadrupedal locomotion in 11 platyrrhine species at field sites in Ecuador and Costa Rica and measured the diameter and orientation of locomotor substrates using remote sensors. We quantified limb phase values and classified strides by gait sequence type (N = 988 strides). RESULTS: Our results show that most of the species in our sample consistently used DS gaits, regardless of substrate diameter or orientation; however, all taxa also used asymmetrical and/or lateral sequence gaits. By incorporating phylogenetic eigenvectors into our models, we found significant differences in gait sequence patterns and limb phase values among the major platyrrhine clades, suggesting that phylogeny may be a better predictor of gait than substrate diameter or orientation. DISCUSSION: Our field data generally corroborate locomotor patterns from laboratory studies but capture additional aspects of gait variability and flexibility in response to the complexity of natural environments. Overall, our results suggest that DS gaits are not exclusively tailored to narrow or oblique substrates but are used on arboreal substrates in general.

Production of the antibiotic secondary metabolite solanapyrone A by the fungal plant pathogen Ascochyta rabiei during fruiting body formation in saprobic growth.

Fungi are noted producers of a diverse array of secondary metabolites, many of which are of pharmacological importance. However, the biological roles of the vast majority of these molecules during the fungal life cycle in nature remain elusive. Solanapyrones are polyketide-derived secondary metabolites produced by diverse fungal species including the plant pathogen Ascochyta rabiei. This molecule was originally thought to function as a phytotoxin facilitating pathogenesis of A. rabiei. Chemical profiling and gene expression studies showed that solanapyrone A was specifically produced during saprobic, but not parasitic growth of A. rabiei. Expression of the gene encoding the final enzymatic step in solanapyrone biosynthesis was specifically associated with development of the asexual fruiting bodies of the fungus on certain substrates. In confrontation assays with saprobic fungi that were commonly found in chickpea debris in fields, A. rabiei effectively suppressed the growth of all competing fungi, such as Alternaria, Epicoccum and Ulocladium species. Solanapyrone A was directly detected in the inhibitory zone using a MALDI-imaging mass spectrometry, and the purified compound showed significant antifungal activities against the potential saprobic competitors. These results suggest that solanapyrone A plays an important role for competition and presumably the survival of the fungus.

Evolutionary aspects of the development of teeth and baleen in the bowhead whale.

In utero, baleen whales initiate the development of several dozens of teeth in upper and lower jaws. These tooth germs reach the bell stage and are sometimes mineralized, but toward the end of prenatal life they are resorbed and no trace remains after birth. Around the time that the germs disappear, the keratinous baleen plates start to form in the upper jaw, and these form the food-collecting mechanism. Baleen whale ancestors had two generations of teeth and never developed baleen, and the prenatal teeth of modern fetuses are usually interpreted as an evolutionary leftover. We investigated the development of teeth and baleen in bowhead whale fetuses using histological and immunohistochemical evidence. We found that upper and lower dentition initially follow similar developmental pathways. As development proceeds, upper and lower tooth germs diverge developmentally. Lower tooth germs differ along the length of the jaw, reminiscent of a heterodont dentition of cetacean ancestors, and lingual processes of the dental lamina represent initiation of tooth bud formation of replacement teeth. Upper tooth germs remain homodont and there is no evidence of a secondary dentition. After these germs disappear, the oral epithelium thickens to form the baleen plates, and the protein FGF-4 displays a signaling pattern reminiscent of baleen plates. In laboratory mammals, FGF-4 is not involved in the formation of hair or palatal rugae, but it is involved in tooth development. This leads us to propose that the signaling cascade that forms teeth in most mammals has been exapted to be involved in baleen plate ontogeny in mysticetes.

Predicting Ascospore Release of Monilinia vaccinii-corymbosi of Blueberry with Machine Learning.

Mummy berry, caused by Monilinia vaccinii-corymbosi, causes economic losses of highbush blueberry in the U.S. Pacific Northwest (PNW). Apothecia develop from mummified berries overwintering on soil surfaces and produce ascospores that infect tissue emerging from floral and vegetative buds. Disease control currently relies on fungicides applied on a calendar basis rather than inoculum availability. To establish a prediction model for ascospore release, apothecial development was tracked in three fields, one in western Oregon and two in northwestern Washington in 2015 and 2016. Air and soil temperature, precipitation, soil moisture, leaf wetness, relative humidity and solar radiation were monitored using in-field weather stations and Washington State University's AgWeatherNet stations. Four modeling approaches were compared: logistic regression, multivariate adaptive regression splines, artificial neural networks, and random forest. A supervised learning approach was used to train the models on two data sets: training (70%) and testing (30%). The importance of environmental factors was calculated for each model separately. Soil temperature, soil moisture, and solar radiation were identified as the most important factors influencing ascospore release. Random forest models, with 78% accuracy, showed the best performance compared with the other models. Results of this research helps PNW blueberry growers to optimize fungicide use and reduce production costs.

Flight feather attachment in rock pigeons (Columba livia): covert feathers and smooth muscle coordinate a morphing wing.

Mechanisms for passively coordinating forelimb movements and flight feather abduction and adduction have been described separately from both in vivo and ex vivo studies. Skeletal coordination has been identified as a way for birds to simplify the neuromotor task of controlling flight stroke, but an understanding of the relationship between skeletal coordination and the coordination of the aerodynamic control surface (the flight feathers) has been slow to materialize. This break between the biomechanical and aerodynamic approaches - between skeletal kinematics and airfoil shape - has hindered the study of dynamic flight behaviors. Here I use dissection and histology to identify previously overlooked interconnections between musculoskeletal elements and flight feathers. Many of these structures are well-placed to directly link elements of the passive musculoskeletal coordination system with flight feather movements. Small bundles of smooth muscle form prominent connections between upper forearm coverts (deck feathers) and the ulna, as well as the majority of interconnections between major flight feathers of the hand. Abundant smooth muscle may play a role in efficient maintenance of folded wing posture, and may also provide an autonomically regulated means of tuning wing shape and aeroelastic behavior in flight. The pattern of muscular and ligamentous linkages of flight feathers to underlying muscle and bone may provide predictable passive guidance for the shape of the airfoil during flight stroke. The structures described here provide an anatomical touchstone for in vivo experimental tests of wing surface coordination in an extensively researched avian model species.

The evolution of mammal-like crocodyliforms in the Cretaceous Period of Gondwana.

Fossil crocodyliforms discovered in recent years have revealed a level of morphological and ecological diversity not exhibited by extant members of the group. This diversity is particularly notable among taxa of the Cretaceous Period (144-65 million years ago) recovered from former Gondwanan landmasses. Here we report the discovery of a new species of Cretaceous notosuchian crocodyliform from the Rukwa Rift Basin of southwestern Tanzania. This small-bodied form deviates significantly from more typical crocodyliform craniodental morphologies, having a short, broad skull, robust lower jaw, and a dentition with relatively few teeth that nonetheless show marked heterodonty. The presence of morphologically complex, complementary upper and lower molariform teeth suggests a degree of crown-crown contact during jaw adduction that is unmatched among known crocodyliforms, paralleling the level of occlusal complexity seen in mammals and their extinct relatives. The presence of another small-bodied mammal-like crocodyliform in the Cretaceous of Gondwana indicates that notosuchians probably filled niches and inhabited ecomorphospace that were otherwise occupied by mammals on northern continents.

A Novel Type Pathway-Specific Regulator and Dynamic Genome Environments of a Solanapyrone Biosynthesis Gene Cluster in the Fungus Ascochyta rabiei.

Secondary metabolite genes are often clustered together and situated in particular genomic regions, like the subtelomere, that can facilitate niche adaptation in fungi. Solanapyrones are toxic secondary metabolites produced by fungi occupying different ecological niches. Full-genome sequencing of the ascomycete Ascochyta rabiei revealed a solanapyrone biosynthesis gene cluster embedded in an AT-rich region proximal to a telomere end and surrounded by Tc1/Mariner-type transposable elements. The highly AT-rich environment of the solanapyrone cluster is likely the product of repeat-induced point mutations. Several secondary metabolism-related genes were found in the flanking regions of the solanapyrone cluster. Although the solanapyrone cluster appears to be resistant to repeat-induced point mutations, a P450 monooxygenase gene adjacent to the cluster has been degraded by such mutations. Among the six solanapyrone cluster genes (sol1 to sol6), sol4 encodes a novel type of Zn(II)2Cys6 zinc cluster transcription factor. Deletion of sol4 resulted in the complete loss of solanapyrone production but did not compromise growth, sporulation, or virulence. Gene expression studies with the sol4 deletion and sol4-overexpressing mutants delimited the boundaries of the solanapyrone gene cluster and revealed that sol4 is likely a specific regulator of solanapyrone biosynthesis and appears to be necessary and sufficient for induction of the solanapyrone cluster genes. Despite the dynamic surrounding genomic regions, the solanapyrone gene cluster has maintained its integrity, suggesting important roles of solanapyrones in fungal biology.

Identification and Enumeration of Small-Spored Alternaria Species Associated with Potato in the U.S. Northwest.

A number of Alternaria spp. have been isolated from potato worldwide but only Alternaria solani and A. alternata have been described as pathogenic to this host in the United States. These taxa are easily differentiated based on conidial morphology but species delimitation among the small-spored Alternaria spp. associated with potato are much more challenging. Accurate identification methods for small-spored Alternaria spp. are necessary so that a more thorough understanding of Alternaria epidemiology can be obtained. Isolations of Alternaria fungi from lesions on potato leaves collected in the U.S. Northwest were made between 2008 and 2011. Large-spored taxa (putatively A. solani), were isolated less frequently than small-spored taxa (putatively A. alternata sensu lato), except in 2010. Colletotrichum coccodes was isolated from necrotic lesions in 2008 to 2010 but not in 2011. Frequency of isolation ranged from 0.05 (5%) to 0.11 (11%) during the 3 years the fungus was detected. Anonymous genomic region OPA1-3, previously used for Alternaria systematics, allowed for the discrimination of phylogenetic lineages among 210 small-spored isolates. When OPA1-3 was restricted using enzyme ApaI, 65 isolates (31%) displayed a restriction banding pattern consistent with previously characterized morphospecies A. alternata and A. tenuissima and 145 (69%) displayed a restriction banding pattern consistent with the previously characterized morphospecies A. arborescens. Morphological characterization of a subsample of 59 small-spored Alternaria isolates randomly selected with each restriction pattern was compared with phylogenetic lineage. In all, 54 (92%) isolates were consistently assigned to the same group by both methods. Three isolates exhibited conidial morphologies that were inconsistent with any described morphospecies. A small number of isolates were identified as A. arbusti (infectoria group) via sequencing of the glyceraldehyde-3-phosphate-dehydrogenase locus and BLAST searches.

Macroevolutionary developmental biology: Embryos, fossils, and phylogenies.

The field of evolutionary developmental biology is broadly focused on identifying the genetic and developmental mechanisms underlying morphological diversity. Connecting the genotype with the phenotype means that evo-devo research often considers a wide range of evidence, from genetics and morphology to fossils. In this commentary, we provide an overview and framework for integrating fossil ontogenetic data with developmental data using phylogenetic comparative methods to test macroevolutionary hypotheses. We survey the vertebrate fossil record of preserved embryos and discuss how phylogenetic comparative methods can integrate data from developmental genetics and paleontology. Fossil embryos provide limited, yet critical, developmental data from deep time. They help constrain when developmental innovations first appeared during the history of life and also reveal the order in which related morphologies evolved. Phylogenetic comparative methods provide a powerful statistical approach that allows evo-devo researchers to infer the presence of nonpreserved developmental traits in fossil species and to detect discordant evolutionary patterns and processes across levels of biological organization.

Qualitative skeletal correlates of wing shape in extant birds (Aves: Neoaves).

BACKGROUND: Among living fliers (birds, bats, and insects), birds display relatively high aspect ratios, a dimensionless shape variable that distinguishes long and narrow vs. short and broad wings. Increasing aspect ratio results in a functional tradeoff between low induced drag (efficient cruise) and increased wing inertia (difficult takeoff). Given the wide scope of its functional effects, the pattern of aspect ratio evolution is an important factor that contributes to the substantial ecological and phylogenetic diversity of living birds. However, because the feathers that define the wingtip (and hence wingspan and aspect ratio) often do not fossilize, resolution in the pattern of avian wing shape evolution is obscured by missing information. Here I use a comparative approach to investigate the relationship between skeletal proxies of flight feather attachment and wing shape. RESULTS: An accessory lobe of the internal index process of digit II-1, a bony correlate of distal primary attachment, shows weak but statistically significant relationships to aspect ratio and mass independent of other skeletal morphology. The dorsal phalangeal fossae of digit II-1, which house distal primaries VIII and IX, also show a trend of increased prominence with higher aspect ratio. Quill knobs on the ulna are examined concurrently, but do not show consistent signal with respect to wing shape. CONCLUSIONS: Although quill knobs are cited as skeletal correlates of flight performance in birds, their relationship to wing shape is inconsistent among extant taxa, and may reflect diverging selection pressures acting on a conserved architecture. In contrast, correlates of distal primary feather attachment on the major digit show convergent responses to increasing aspect ratio. In light of the diversity of musculoskeletal and integumentary mophology that underlies wing shape in different avian clades, it is unlikely that a single skeletal feature will show consistent predictive power across Neoaves. Confident inference of wing shape in basal ornithurine birds will require multiple lines of evidence, together with an understanding of clade-specific evolutionary trends within the crown.

Functional Analyses of the Diels-Alderase Gene sol5 of Ascochyta rabiei and Alternaria solani Indicate that the Solanapyrone Phytotoxins Are Not Required for Pathogenicity.

Ascochyta rabiei and Alternaria solani, the causal agents of Ascochyta blight of chickpea (Cicer arietinum) and early blight of potato (Solanum tuberosum), respectively, produce a set of phytotoxic compounds including solanapyrones A, B, and C. Although both the phytotoxicity of solanapyrones and their universal production among field isolates have been documented, the role of solanapyrones in pathogenicity is not well understood. Here, we report the functional characterization of the sol5 gene, which encodes a Diels-Alderase that catalyzes the final step of solanapyrone biosynthesis. Deletion of sol5 in both Ascochyta rabiei and Alternaria solani completely prevented production of solanapyrones and led to accumulation of the immediate precursor compound, prosolanapyrone II-diol, which is not toxic to plants. Deletion of sol5 did not negatively affect growth rate or spore production in vitro, and led to overexpression of the other solanapyrone biosynthesis genes, suggesting a possible feedback regulation mechanism. Phytotoxicity tests showed that solanapyrone A is highly toxic to several legume species and Arabidopsis thaliana. Despite the apparent phytotoxicity of solanapyrone A, pathogenicity tests showed that solanapyrone-minus mutants of Ascochyta rabiei and Alternaria solani were equally virulent as their corresponding wild-type progenitors, suggesting that solanapyrones are not required for pathogenicity.