Genomic overview of closely related fungi with different Protea host ranges

Genome comparisons of species with distinctive ecological traits can elucidate genetic divergence that influenced their differentiation. The interaction of a microorganism with its biotic environment is largely regulated by secreted compounds, and these can be predicted from genome sequences. In this study, we considered Knoxdaviesia capensis and Knoxdaviesia proteae, two closely related saprotrophic fungi found exclusively in Protea plants. We investigated their genome structure to compare their potential inter-specific interactions based on gene content. Their genomes displayed macrosynteny and were approximately 10 % repetitive. Both species had fewer secreted proteins than pathogens and other saprotrophs, reflecting their specialized habitat. The bulk of the predicted species-specific and secreted proteins coded for carbohydrate metabolism, with a slightly higher number of unique carbohydrate-degrading proteins in the broad host-range K. capensis. These fungi have few secondary metabolite gene clusters, suggesting minimal competition with other microbes and symbiosis with antibiotic-producing bacteria common in this niche. Secreted proteins associated with detoxification and iron sequestration likely enable these Knoxdaviesia species to tolerate antifungal compounds and compete for resources, facilitating their unusual dominance. This study confirms the genetic cohesion between Protea-associated Knoxdaviesia species and reveals aspects of their ecology that have likely evolved in response to their specialist niche.     

Early colonization of Protea flowers enable dominance of competitively weak saprobic fungi in seed cones,benefitting their hosts

Sporothrix and Knoxdaviesia fungi use pollinators to colonize Protea flowers at anthesis. These saprobes remain dominant in the nutrient-rich, fire-retardant Protea seed-cones (infructescences) for at least a year after flowering. We tested the hypothesis that they competitively exclude potentially detrimental fungi from infructescences during this time. We compared seed set and longevity of infructescences containing Sporothrix and Knoxdaviesia vs. those that contain ‘contaminant’ saprobes. Hereafter we evaluated their competitive abilities against the ‘contaminant’ saprobes. Infructescences devoid of Sporothrix and Knoxdaviesia were dominated by Penicillium cf. toxicarium, Cladosporium cf. cladosporoides and Fusarium cf. anthophilum. Sporothrix and Knoxdaviesia presence did not affect seed viability, but infructescences persisted longer than those colonised by ‘contaminant’ fungi. The ‘contaminant’ species were stronger competitors than Sporothrix and Knoxdaviesia. However, Sporothrix and Knoxdaviesia could defend captured space well against ‘contaminant’ species. This effect was enhanced when fungal taxa grew on media prepared from their usual Protea host species, clarifying their dominance and host consistency observed in the field. Sporothrix and Knoxdaviesia from Protea are therefore weak competitors against common saprobes, especially when growing on alternative hosts, and need to colonise flowers very early (before colonization by other fungi) to dominate in this environment. They may delay seed release from infructescences longer than if these are colonised by other saprobes, increasing chances of seed release to occur after fire, when conditions are more favourable for Protea recruitment.     

Convergent evolution unites the population genetics of Protea-associated ophiostomatoid fungi

Knoxdaviesia and Sporothrix species occupy the flower heads of some Protea plants in southern Africa. Knoxdaviesia species display exceptional genetic diversity within the Core Cape Subregion (CCR) and are readily dispersed across large distances. This study aimed to determine whether overlapping ecologies have led to a similar population genetic structure in Sporothrix splendens. Two DNA sequence markers, β-tubulin and a microsatellite region, were amplified in 97 S. splendens strains from eight populations that span its host distribution. Genetic diversity was low in a geographically isolated population, but high elsewhere. CCR populations were closely related, showing isolation by distance with populations at the eastern edge of the sampling range. Like Knoxdaviesia species, long-distance dispersal of S. splendens spores is prevalent, although likely affected by patchy host populations. This study is the first to consider populations of a non-clinical Sporothrix species, providing insights into the population attributes of a naturally distributed species.     

Unseen fungal biodiversity and complex inter-organismal interactions in Protea flower heads

A unique microbiome occurs within the flower heads of various Protea species endemic to Africa. These include two lineages of ophiostomatoid fungi, Knoxdaviesia (Microascales) and Sporothrix (Ophiostomatales), that have members occurring exclusively in this environment and that rely on mites as their primary mode of spore dissemination. The mites, in turn, attach to the bodies of Protea-pollinating beetles and the beaks and bodies of birds for long-distance movement, establishing a hierarchical dispersal network for the ophiostomatoid fungi. This inter-organismal network is highly successful, achieving fungal dispersal over vast distances. Multiple species of fungi, mites and bacteria have been described from this unique niche over the past four decades. The intricacies of their symbiotic interactions continue to be unravelled. This review covers all current knowledge of the “distinctly African” Protea-ophiostomatoid fungus environment and illustrates the depth of a fascinating unseen fungal biodiversity niche.     

Electroantennogram (EAG) responses of Microplitis croceipes and Cotesia marginiventris and their lepidopteran hosts to a wide array of odor stimuli: Correlation between EAG response and degree of host specificity?

In order to test whether the electroantennogram (EAG) response spectrum of an insect correlates to its degree of host specificity, we recorded EAG responses of two parasitoid species with different degrees of host specificity, Microplitis croceipes (specialist) and Cotesia marginiventris (generalist), to a wide array of odor stimuli including compounds representing green leaf volatiles (GLVs), herbivore-induced plant volatiles (HIPV), ecologically irrelevant (not used by the parasitoid species and their hosts for host location) plant volatiles, and host-specific odor stimuli (host sex pheromones, and extracts of host caterpillar body and frass). We also tested the EAG responses of female moths of the caterpillar hosts of the parasitoids, Heliothis virescens and Spodoptera exigua, to some of the odor stimuli. We hypothesized that the specialist parasitoid will have a narrower EAG response spectrum than the generalist, and that the two lepidopteran species, which are similar in their host plant use, will show similar EAG response spectra to plant volatiles. As predicted, the specialist parasitoid showed greater EAG responses than the generalist to host-specific odor and one HIPV (cis-3-hexenyl butyrate), whereas the generalist showed relatively greater EAG responses to the GLVs and unrelated plant volatiles. We detected no differences in the EAG responses of H. virescens and S. exigua to any of the tested odor.     

Understanding How the Complex Molecular Architecture of Mannan-degrading Hydrolases Contributes to Plant Cell Wall Degradation

Microbial degradation of plant cell walls is a central component of the carbon cycle and is of increasing importance in environmentally significant industries. Plant cell wall-degrading enzymes have a complex molecular architecture consisting of catalytic modules and, frequently, multiple non-catalytic carbohydrate binding modules (CBMs). It is currently unclear whether the specificities of the CBMs or the topology of the catalytic modules are the primary drivers for the specificity of these enzymes against plant cell walls. Here, we have evaluated the relationship between CBM specificity and their capacity to enhance the activity of GH5 and GH26 mannanases and CE2 esterases against intact plant cell walls. The data show that cellulose and mannan binding CBMs have the greatest impact on the removal of mannan from tobacco and Physcomitrella cell walls, respectively. Although the action of the GH5 mannanase was independent of the context of mannan in tobacco cell walls, a significant proportion of the polysaccharide was inaccessible to the GH26 enzyme. The recalcitrant mannan, however, was fully accessible to the GH26 mannanase appended to a cellulose binding CBM. Although CE2 esterases display similar specificities against acetylated substrates in vitro, only CjCE2C was active against acetylated mannan in Physcomitrella. Appending a mannan binding CBM27 to CjCE2C potentiated its activity against Physcomitrella walls, whereas a xylan binding CBM reduced the capacity of esterases to deacetylate xylan in tobacco walls. This work provides insight into the biological significance for the complex array of hydrolytic enzymes expressed by plant cell wall-degrading microorganisms.     

Knoxdaviesia capensis: dispersal ecology and population genetics of a flower-associated fungus

Protea-associated fungi are dispersed between flower heads by mites, beetles and possibly birds. For the ophiostomatoid fungus, Knoxdaviesia proteae, these vectors offer regular dispersal between distant floral hosts. Unlike K. proteae, Knoxdaviesia capensis occupies multiple Protea host species. In this study, we aimed to determine whether the generalist K. capensis shares the long-distance dispersal pattern with specialist K. proteae and whether it moves freely between different host species. We evaluated the genetic structure of K. capensis from five populations of a wide-spread host and between sympatric hosts. Twelve K. capensis-specific microsatellite markers were developed and applied to 90 individuals. K. capensis showed high genetic diversity and almost maximal genotypic diversity. All populations were poorly differentiated, indicating the presence of long-distance dispersal. No differentiation could be detected between sympatric host populations, suggesting free dispersal between different hosts. This implies that the beetle and bird vectors that pollinate Protea species show the same non-specific movement.     

Metarhizium robertsii Produces an Extracellular Invertase (MrINV) That Plays a Pivotal Role in Rhizospheric Interactions and Root Colonization

As well as killing pest insects, the rhizosphere competent insect-pathogenic fungus Metarhizium robertsii also boosts plant growth by providing nitrogenous nutrients and increasing resistance to plant pathogens. Plant roots secrete abundant nutrients but little is known about their utilization by Metarhizium spp. and the mechanistic basis of Metarhizium-plant associations. We report here that M. robertsii produces an extracellular invertase (MrInv) on plant roots. Deletion of MrInv (⊿MrInv) reduced M. robertsii growth on sucrose and rhizospheric exudates but increased colonization of Panicum virgatum and Arabidopsis thaliana roots. This could be accounted for by a reduction in carbon catabolite repression in ⊿MrInv increasing production of plant cell wall-degrading depolymerases. A non-rhizosphere competent scarab beetle specialist Metarhizium majus lacks invertase which suggests that rhizospheric competence may be related to the sugar metabolism of different Metarhizium species.     

Ecophysiology of six selected shrub species in different plant communities at the periphery of the Atlantic Forest of SE-Brazil

The Atlantic rain forest of Brazil is one of the 25 biodiversity hotspots of the world. It is surrounded by distinct marginal plant communities which are floristically related to the rain forest. We performed an in situ ecophysiological study of six shrub and tree species which are abundant in one or more of these marginal ecosystems. They formed three pairs of taxonomically related species, which, however, differed in geographic distribution and/or habitat preference. We aimed to answer the following questions: (1) Do habitat generalists and specialists differ in ecophysiological behaviour? (2) Are there general trends responsive to site characteristics, irrespective of taxonomic affiliation? (3) In which variables between-site intraspecific variation is more often detected? Plants chosen were the Clusiaceae Calophyllum brasiliense Cambess (swamp forest specialist) and Rheedia brasiliensis (Mart.) Planch &; Triana (a generalist plant, occurring in swamp forests and dry, sandy coastal vegetation); the Myrsinaceae Myrsine parvifolia A.DC. (widespread in sandy plains) and Myrsine gardneriana A.DC. (widespread in high-altitude habitats); and the Euphorbiaceae Styllingia dichotoma Muell. Arg. (a coastal inselberg specialist) and Croton compressus Lam. (a generalist found in coastal inselbergs and dry forests). We compared measurements of photosynthesis (chlorophyll fluorescence parameters), stable isotope (δ¹³C, δ¹⁵N) and metabolic carbon and nitrogen compounds. Generalists and specialists did not always differ in ecophysiological behaviour: for instance, while widespread generalists such as M. parvifolia and C. compressus were ecophysiologically versatile, R. brasiliense appeared to perform below optimum in all sampled habitats. Light and water were abiotic factors which apparently explained most of the variation found. Overall, ecophysiological responses were often more clearly related to individual species rather than to taxonomic affinities in higher hierarchies (family, genus) or habitat. These results are discussed as a further indication of high biodiversity even in habitats marginal to the rain forest hotspot.     

Exotic Lonicera species both escape and resist specialist and generalist herbivores in the introduced range in North America

The enemy release hypothesis predicts that invasive plant species may benefit from a lack of top-down control by co-evolved herbivores, particularly specialists, in their new range. However, to benefit from enemy escape, invasive plants must also escape or resist specialist or generalist herbivores that attack related species in the introduced range. We compared insect herbivore damage on the exotic shrub, Lonicera maackii, the native congener Lonicera reticulata, and the native confamilial Viburnum prunifolium in North America. We also compared the laboratory preference and performance of a North American honeysuckle specialist sawfly (Zaraea inflata) and the performance of a widespread generalist caterpillar (Spodoptera frugiperda) on cut foliage from native and exotic Lonicera species. L. maackii received significantly lower amounts of foliar herbivory than L. reticulata across three seasons, while damage levels observed on V. prunifolium for two seasons was generally intermediate between L. reticulata and L. maackii. The specialist sawfly damaged L. reticulata heavily, but was not detected on L. maackii in the field. There were few statistical differences in the performance of sawfly larvae on L. reticulata and L. maackii, but the sawfly achieved higher pupal masses on L. reticulata than on L. maackii, and they strongly preferred L. reticulata over L. maackii when given a choice. The sawfly was unable to complete development on native L. sempervirens and non-native L. japonica. In contrast, the generalist caterpillar performed similarly on all Lonicera species. While L. maackii experienced little herbivory in the field compared to native relatives in the same habitat, laboratory assays indicate L. maackii appears to be a suitable host that escapes selection by the specialist, but L. japonica and L. sempervirens are highly resistant to it. These findings indicate that both enemy escape and resistance (to a specialist, but not a generalist herbivore) may contribute to the success of exotic Lonicera species.     

Flower handling by bumblebees: a comparison of specialists and generalists

The hypothesis that specialists have evolved behavioural adaptations to handle their preferred food types more efficiently than related generalist species was tested. Naive workers of the specialist bumblebee, Bombus consobrinus, were more efficient than those of generalist species in acquiring flower-handling skills on their specialty plant, Aconitum (monkshood). With no previous foraging experience, B. consobrinus workers began probing in the vicinity of the nectary and quickly located the nectar. Generalists, on the other hand, showed no predisposition toward correct foraging on Aconitum. They probed in the wrong places, and many gave up before finding the nectar.     

No evidence for an ‘evolution of increased competitive ability’ for the invasive Lepidium draba

The ‘evolution of increased competitive ability’ (EICA) hypothesis states that reduced herbivory in the introduced range causes an evolutionary shift in resource allocation from herbivore defense to growth. Therefore, according to EICA, introduced genotypes are expected to grow more vigorously than conspecific native genotypes when cultivated under common standardized conditions. The EICA hypothesis also assumes that herbivores will perform better on introduced genotypes compared to native genotypes, because they are less well defended. However, selection for either defense or growth will depend on the type of defense (quantitative or qualitative) employed by the plant, and whether the plant is released from generalist or specialist herbivores. The predictions of the EICA hypothesis might be reversed if a plant experiences increased generalist herbivore pressure in the introduced range, and therefore invests more in qualitative defense. We examined this idea with the invasive perennial mustard, Lepidium draba. We grew a total of 16 populations of L. draba from both its native European and introduced western US ranges under common conditions in a greenhouse. We also tested for differences in plant resistance to the specialist herbivore, Psylliodes wrasei, by conducting a leaf disc feeding bioassay with native and introduced L. draba genotypes. Furthermore, we quantified the generalist herbivore load on L. draba in both ranges in order to assess the selection pressure for increased qualitative defense. Contrary to the original EICA prediction, all plant traits (biomass, number of shoots, length and diameter of longest leaf) tended to be greater for the native, rather than introduced L. draba genotypes. There was no significant difference in the proportion of leaf area consumed by the specialist herbivore between native and introduced genotypes. The generalist herbivore load on L. draba was significantly greater in the introduced range. Our data suggest that the EICA hypothesis does not explain the invasion success of L. draba in the US. Instead, we propose that the reduced vigor of introduced genotypes may be due to selection for increased defense against generalist herbivores in the introduced range.     

Differential hepatic gene expression of a dietary specialist (Neotoma stephensi) and generalist (Neotoma albigula) in response to juniper (Juniperus monosperma) ingestion

Dietary specialization is thought to be rare in mammalian herbivores because of limitations of their detoxification system in processing large doses of a single type of plant secondary compound (PSC). Therefore, in order to specialize on a single species of plant, mammalian herbivores must have a highly efficient detoxification system for the particular types of PSCs they ingest. Using microarray technology, we looked at the expression of hepatic genes of a dietary specialist, Neotoma stephensi, and a sympatric generalist, Neotoma albigula, in response to diets containing different levels of one-seeded juniper (Juniperus monosperma). We found large between species differences in gene expression, as well as large within species differences when specialists fed a low juniper diet (25% juniper) were compared to specialists fed their ecologically relevant level of juniper (70% juniper). We also tested the hypothesis that the specialist relies on less costly phase I detoxification enzymes more than phase II compared to the generalist. Although we found that the specialist had higher cumulative as well as average expression of phase I versus phase II enzymes, the generalist had a similar pattern of expression for phase I versus phase II enzymes.     

Response of pest control by generalist predators to local‐scale plant diversity: a meta‐analysis

Disentangling the effects of plant diversity on the control of herbivores is important for understanding agricultural sustainability. Recent studies have investigated the relationships between plant diversity and arthropod communities at the landscape scale, but few have done so at the local scale. We conducted a meta‐analysis of 32 papers containing 175 independent measures of the relationship between plant diversity and arthropod communities. We found that generalist predators had a strong positive response to plant diversity, that is, their abundance increased as plant diversity increased. Herbivores, in contrast, had an overall weak and negative response to plant diversity. However, specialist and generalist herbivores differed in their response to plant diversity, that is, the response was negative for specialists and not significant for generalists. While the effects of scale remain unclear, the response to plant diversity tended to increase for specialist herbivores, but decrease for generalist herbivores as the scale increased. There was no clear effect of scale on the response of generalist predators to plant diversity. Our results suggest that the response of herbivores to plant diversity at the local scale is a balance between habitat and trophic effects that vary according to arthropod specialization and habitat type. Synthesis and applications. Positive effects of plant diversity on generalist predators confirm that, at the local scale, plant diversification of agroecosystems is a credible and promising option for increasing pest regulation. Results from our meta‐analysis suggest that natural control in plant‐diversified systems is more likely to occur for specialist than for generalist herbivores. In terms of pest management, our results indicate that small‐scale plant diversification (via the planting of cover crops or intercrops and reduced weed management) is likely to increase the control of specialist herbivores by generalist predators.     

Generalized host‐plant feeding can hide sterol‐specialized foraging behaviors in bee–plant interactions

Host‐plant selection is a key factor driving the ecology and evolution of insects. While the majority of phytophagous insects is highly host specific, generalist behavior is quite widespread among bees and presumably involves physiological adaptations that remain largely unexplored. However, floral visitation patterns suggest that generalist bees do not forage randomly on all available resources. While resource availability and accessibility as well as nectar composition have been widely explored, pollen chemistry could also have an impact on the range of suitable host‐plants. This study focuses on particular pollen nutrients that cannot be synthesized de novo by insects but are key compounds of cell membranes and the precursor for molting process: the sterols. We compared the sterol composition of pollen from the main host‐plants of three generalist bees: Anthophora plumipes, Colletes cunicularius, and Osmia cornuta, as well as one specialist bee Andrena vaga. We also analyzed the sterols of their brood cell provisions, the tissues of larvae and nonemerged females to determine which sterols are used by the different species. Our results show that sterols are not used accordingly to foraging strategy: Both the specialist species A. vaga and the generalist species C. cunicularius might metabolize a rare C₂₇ sterol, while the two generalist species A. plumipes and O. cornuta might rather use a very common C₂₈ sterol. Our results suggest that shared sterolic compounds among plant species could facilitate the exploitation of multiple host‐plants by A. plumipes and O. cornuta whereas the generalist C. cunicularius might be more constrained due to its physiological requirements of a more uncommon dietary sterol. Our findings suggest that a bee displaying a generalist foraging behavior may sometimes hide a sterol‐specialized species. This evidence challenges the hypothesis that all generalist free‐living bee species are all able to develop on a wide range of different pollen types.     

Use of novel pollen species by specialist and generalist solitary bees (Hymenoptera: Megachilidae)

If trade-offs between flexibility to use a range of host species and efficiency on a limited set underlie the evolution of diet breadth, one resulting prediction is that specialists ought to be more restricted than generalists in their ability to use novel resource species. I used foraging tests and feeding trials to compare the ability of a generalist and a specialist solitary mason bee species to collect and develop on two pollen species that are not normally used in natural populations (novel pollens). Osmia lignaria (Hymenoptera: Megachilidae) is a generalist pollen feeder; O. californica, is more specialized. Adults of the specialist were more limited in use of novel hosts, but only in some contexts. Both bee species refused to collect one novel pollen. The specialist accepted a second novel pollen only when it was presented along with its normal pollen, whereas the generalist collected novel pollen whether presented alone or with normal pollen. Surprisingly, larvae of the specialist were more flexible than were generalists. The specialist grew well on mixtures of normal and novel pollen species, in some cases better than on its normal host alone. Larvae of the generalist grew more poorly on all diets containing novel pollens than on their normal host. Data on these two species of bees suggest that specialization by itself need not reduce flexibility on novel hosts. The findings also provide information about mechanisms of specialization in bees. Similar to some folivores, specific cues of the pollen host and the bee's interpretation of these contribute, along with foraging economics, to pollen choice by adults. The ability of the larvae to cope with specific components of one pollen species need not interfere with its ability to use others.     

Larval feeding induced defensive responses in tobacco: comparison of two sibling species of <Emphasis Type="Italic">Helicoverpa</Emphasis> with different diet breadths

Plants respond differently to damage by different herbivorous insects. We speculated that sibling herbivorous species with different host ranges might also influence plant responses differently. Such differences may be associated with the diet breadth (specialization) of herbivores within a feeding guild, and the specialist may cause less intensive plant responses than the generalist. The tobacco Nicotinana tabacum L. is the common host plant of a generalist Helicoverpa armigera (Hübner) and a specialist H. assulta Guenée (Lepidoptera, Noctuidae). The induced responses of tobacco to feeding of these two noctuid herbivores and mechanical wounding were compared. The results showed that the feeding of the specialist H. assulta and the generalist H. armigera resulted in the same inducible defensive system, but response intensity of plants was different to these two species. Inductions of jasmonic acid (JA), lipoxygenase (LOX), and proteinase inhibitors (PIs) were not significantly different concerning these two species, but H. assulta caused the less intensive foliar polyphenol oxidase (PPO) increase, more intensive nicotine and peroxidase (POD) increases in tobacco than H. armigera. The defensive response of plant to herbivores with different diet breadth seems to be more complicated than we expected, and the specialist does not necessarily cause less intensive plant responses than the generalist.     

Host specialization is ancestral in Torymus (Hymenoptera,Chalcidoidea) cynipid gall parasitoids

The host range of parasitoids varies greatly among species: Some only parasitize one to a few hosts (specialists), while others parasitize multiple species or a variety of host types (generalists). The direction of most host range shifts in parasitoid groups, that is from generalist to specialist or, alternatively, from specialist to generalist, is unknown. To explore the origin of host range shifts, we studied a clade within the genus Torymus (Hymenoptera, Chalcidoidea) that includes both generalist and specialist parasitoids of Cynipidae (Hymenoptera). We analysed the phylogenetic relationships of the species of Torymus on the basis of two gene fragments (cox1 and ITS2) of 246 specimens and performed an ancestral state reconstruction of the specialist/generalist trait. Our results revealed the following: (a) The ancestral state of this group of Torymus is specialist, with the generalist state evolving through a loss of specialization. (b) The species Torymus cyaneus and Torymus flavipes both have a strong genetic structure, suggesting the existence of different biological identities. (c) There has been a host plant shift in the lineage(s) leading to Torymus rubi and Torymus bedeguaris from galls on Quercus to those on Rosaceae. (d) The alien species Torymus sinensis and the native European species Torymus notatus are phylogenetically closely related. (e) Speciation within Torymus was likely associated with the diversification of their cynipid hosts, which itself was driven by the dramatic changes in climate and vegetation that occurred during the Miocene.