Exposure to the leaf litter microbiome of healthy adults protects seedlings from pathogen damage

It is increasingly recognized that microbiota affect host health and physiology. However, it is unclear what factors shape microbiome community assembly in nature, and how microbiome assembly can be manipulated to improve host health. All plant leaves host foliar endophytic fungi, which make up a diverse, environmentally acquired fungal microbiota. Here, we experimentally manipulated assembly of the cacao tree (Theobroma cacao) fungal microbiome in nature and tested the effect of assembly outcome on host health. Using next-generation sequencing, as well as culture-based methods coupled with Sanger sequencing, we found that manipulating leaf litter exposure and location within the forest canopy significantly altered microbiome composition in cacao. Exposing cacao seedlings to leaf litter from healthy conspecific adults enriched the seedling microbiome with Colletotrichum tropicale, a fungal endophyte known to enhance pathogen resistance of cacao seedlings by upregulating host defensive pathways. As a result, seedlings exposed to healthy conspecific litter experienced reduced pathogen damage. Our results link processes that affect the assembly and composition of microbiome communities to their functional consequences for host success, and have broad implications for understanding plant–microbe interactions. Deliberate manipulation of the plant– fungal microbiome also has potentially important applications for cacao production and other agricultural systems in generalIt is increasingly recognized that microbiota affect host health and physiology. However, it is unclear what factors shape microbiome community assembly in nature, and how microbiome assembly can be manipulated to improve host health. All plant leaves host foliar endophytic fungi, which make up a diverse, environmentally acquired fungal microbiota. Here, we experimentally manipulated assembly of the cacao tree (Theobroma cacao) fungal microbiome in nature and tested the effect of assembly outcome on host health. Using next-generation sequencing, as well as culture-based methods coupled with Sanger sequencing, we found that manipulating leaf litter exposure and location within the forest canopy significantly altered microbiome composition in cacao. Exposing cacao seedlings to leaf litter from healthy conspecific adults enriched the seedling microbiome with Colletotrichum tropicale, a fungal endophyte known to enhance pathogen resistance of cacao seedlings by upregulating host defensive pathways. As a result, seedlings exposed to healthy conspecific litter experienced reduced pathogen damage. Our results link processes that affect the assembly and composition of microbiome communities to their functional consequences for host success, and have broad implications for understanding plant–microbe interactions. Deliberate manipulation of the plant– fungal microbiome also has potentially important applications for cacao production and other agricultural systems in genera

Host specificity in a diverse Neotropical tick community: an assessment using quantitative network analysis and host phylogeny

Background: Host specificity is a fundamental determinant of tick population and pathogen transmission dynamics, and therefore has important implications for human health. Tick host specificity is expected to be particularly high in the tropics, where communities of ticks, hosts and pathogens are most diverse. Yet the degree to which tropical tick species are host-specific remains poorly understood. Combining new field data with published records, we assessed the specificity of tick-host associations in Panama, a diverse Neotropical region. Methods: The resulting dataset includes 5,298 adult ticks belonging to 41 species of eight genera that were directly collected from 68 vertebrate host species of 17 orders. We considered three important aspects of tick host specificity: (i) the relative ecological importance of each host species (structural specificity); (ii) relatedness among host species (phylogenetic specificity); and (iii) spatial scale-dependence of tick-host relationships (geographical specificity). Applying quantitative network analyses and phylogenetic tools with null model comparisons, we assessed the structural and phylogenetic specificity across three spatial scales, ranging from central Panama to countrywide. Further, we tested whether species-rich tick genera parasitized a wider variety of hosts than species-poor genera, as expected when ticks specialize on different host species. Results: Most tick species showed high structural and/or phylogenetic specificity in the adult stage. However, after correcting for sampling effort, we found little support for geographical specificity. Across the three scales, adult ticks tended to be specific to a limited number of host species that were phylogenetically closely related. These host species in turn, were parasitized by tick species from distinct genera, suggesting switching among distantly related hosts is common at evolutionary timescales. Further, there was a strong positive relationship between the taxonomic richness of the tick genera and that of their hosts, consistent with distinct tick species being relatively specific to different host species. Conclusions: Our results indicate that in the adult stage, most ticks in the diverse Neotropical community studied are host specialists. This contrasts with earlier assessments, but agrees with findings from other host-parasite systems. High host specificity in adult ticks implies high susceptibility to local tick-host co-extirpation, limited ability to colonize new habitats and limited potential for interspecific pathogen transmission.Background: Host specificity is a fundamental determinant of tick population and pathogen transmission dynamics, and therefore has important implications for human health. Tick host specificity is expected to be particularly high in the tropics, where communities of ticks, hosts and pathogens are most diverse. Yet the degree to which tropical tick species are host-specific remains poorly understood. Combining new field data with published records, we assessed the specificity of tick-host associations in Panama, a diverse Neotropical region. Methods: The resulting dataset includes 5,298 adult ticks belonging to 41 species of eight genera that were directly collected from 68 vertebrate host species of 17 orders. We considered three important aspects of tick host specificity: (i) the relative ecological importance of each host species (structural specificity); (ii) relatedness among host species (phylogenetic specificity); and (iii) spatial scale-dependence of tick-host relationships (geographical specificity). Applying quantitative network analyses and phylogenetic tools with null model comparisons, we assessed the structural and phylogenetic specificity across three spatial scales, ranging from central Panama to countrywide. Further, we tested whether species-rich tick genera parasitized a wider variety of hosts than species-poor genera, as expected when ticks specialize on different host species. Results: Most tick species showed high structural and/or phylogenetic specificity in the adult stage. However, after correcting for sampling effort, we found little support for geographical specificity. Across the three scales, adult ticks tended to be specific to a limited number of host species that were phylogenetically closely related. These host species in turn, were parasitized by tick species from distinct genera, suggesting switching among distantly related hosts is common at evolutionary timescales. Further, there was a strong positive relationship between the taxonomic richness of the tick genera and that of their hosts, consistent with distinct tick species being relatively specific to different host species. Conclusions: Our results indicate that in the adult stage, most ticks in the diverse Neotropical community studied are host specialists. This contrasts with earlier assessments, but agrees with findings from other host-parasite systems. High host specificity in adult ticks implies high susceptibility to local tick-host co-extirpation, limited ability to colonize new habitats and limited potential for interspecific pathogen transmission

Erratum to: Theobroma cacao L. pathogenesis-related gene tandem array members show diverse expression dynamics in response to pathogen colonization

The original version of the manuscript [1] contained an incorrectly named Criollo gene ID on chromosome 1 in the first sentence, under the subheading “Organization of PR gene families into tandem arrays”. The second gene on chromosome 1, Tc##_g######, should therefore be Tc01_g000020.The original version of the manuscript [1] contained an incorrectly named Criollo gene ID on chromosome 1 in the first sentence, under the subheading “Organization of PR gene families into tandem arrays”. The second gene on chromosome 1, Tc##_g######, should therefore be Tc01_g000020

Endomelanconiopsis, a new anamorph genus in the Botryosphaeriaceae

A new lineage is discovered within the Botryosphaeriaceae (Ascomycetes, Dothideomycetes, incertae sedis). Consistent with current practice of providing generic names for independent lineages, this lineage is described as Endomelanconiopsis gen. nov., with the anamorphic species E. endophytica sp. nov. and E. microspora comb. nov. (5 Endomelanconium microsporum). Endomelanconiopsis is characterized by eustromatic conidiomata and holoblastically produced, brown, nonapiculate, unicellular conidia, each with a longitudinal germ slit. Phylogenetic analysis of partial sequences of LSU, ITS and translation elongation factor 1 alpha (tef1) indicate that E. endophytica is sister of E. microspora and that they are nested within the Botryosphaeriaceae. However because there is no support for the ‘‘backbone’’ of the Botryosphaeriacae we are not able to see the interrelationships among the many genera in the family. Neither species is known to have a teleomorph. Endomelanconiopsis differs from Endomelanconium because conidia of the type species of Endomelanconium, E. pini, are papillate at the base, conidiogenous cells proliferate sympodially and the pycnidial wall is thinner; we postulate that the teleomorph of E. pini as yet unknown is an inoperculate discomycete. Endomelanconiopsis endophytica was isolated as an endophyte from healthy leaves of Theobroma cacao (cacao, Malvaceae) and Heisteria concinna (Erythroplaceae) in Panama. Endomelanconiopsis microspora was isolated from soil in EuropeA new lineage is discovered within the Botryosphaeriaceae (Ascomycetes, Dothideomycetes, incertae sedis). Consistent with current practice of providing generic names for independent lineages, this lineage is described as Endomelanconiopsis gen. nov., with the anamorphic species E. endophytica sp. nov. and E. microspora comb. nov. (5 Endomelanconium microsporum). Endomelanconiopsis is characterized by eustromatic conidiomata and holoblastically produced, brown, nonapiculate, unicellular conidia, each with a longitudinal germ slit. Phylogenetic analysis of partial sequences of LSU, ITS and translation elongation factor 1 alpha (tef1) indicate that E. endophytica is sister of E. microspora and that they are nested within the Botryosphaeriaceae. However because there is no support for the ‘‘backbone’’ of the Botryosphaeriacae we are not able to see the interrelationships among the many genera in the family. Neither species is known to have a teleomorph. Endomelanconiopsis differs from Endomelanconium because conidia of the type species of Endomelanconium, E. pini, are papillate at the base, conidiogenous cells proliferate sympodially and the pycnidial wall is thinner; we postulate that the teleomorph of E. pini as yet unknown is an inoperculate discomycete. Endomelanconiopsis endophytica was isolated as an endophyte from healthy leaves of Theobroma cacao (cacao, Malvaceae) and Heisteria concinna (Erythroplaceae) in Panama. Endomelanconiopsis microspora was isolated from soil in Europ

ECOLOGICAL IMPLICATIONS OF ANTI-PATHOGEN EFFECTS OF TROPICAL FUNGAL ENDOPHYTES AND MYCORRHIZAE

We discuss studies of foliar endophytic fungi (FEE) and arbuscular mycorrhizal fungi (AMF) associated with Theobroma cacao in Panama. Direct, experimentally controlled comparisons of endophyte free (E—) and endophyte containing (E+) plant tissues in T. cacao show that foliar endophytes (FEE) that commonly occur in healthy host leaves enhance host defenses against foliar damage due to the pathogen (Phytophthora palmivora). Similarly, root inoculations with commonly occurring AMF also reduce foliar damage due to the same pathogen. These results suggest that endophytic fungi can play a potentially important mutualistic role by augmenting host defensive responses against pathogens. There are two broad classes of potential mechanisms by which endophytes could contribute to host protection: (1) inducing or increasing the expression of intrinsic host defense mechanisms and (2) providing additional sources of defense, extrinsic to those of the host (e.g., endophytebased chemical antibiosis). The degree to which either of these mechanisms predominates holds distinct consequences for the evolutionary ecology of host-endophyte-pathogen relationships. More generally, the growing recognition that plants are composed of a mosaic of plant and fungal tissues holds a series of implications for the study of plant defense, physiology, and genetics.We discuss studies of foliar endophytic fungi (FEE) and arbuscular mycorrhizal fungi (AMF) associated with Theobroma cacao in Panama. Direct, experimentally controlled comparisons of endophyte free (E—) and endophyte containing (E+) plant tissues in T. cacao show that foliar endophytes (FEE) that commonly occur in healthy host leaves enhance host defenses against foliar damage due to the pathogen (Phytophthora palmivora). Similarly, root inoculations with commonly occurring AMF also reduce foliar damage due to the same pathogen. These results suggest that endophytic fungi can play a potentially important mutualistic role by augmenting host defensive responses against pathogens. There are two broad classes of potential mechanisms by which endophytes could contribute to host protection: (1) inducing or increasing the expression of intrinsic host defense mechanisms and (2) providing additional sources of defense, extrinsic to those of the host (e.g., endophytebased chemical antibiosis). The degree to which either of these mechanisms predominates holds distinct consequences for the evolutionary ecology of host-endophyte-pathogen relationships. More generally, the growing recognition that plants are composed of a mosaic of plant and fungal tissues holds a series of implications for the study of plant defense, physiology, and genetics

Inferring Processes of Coevolutionary Diversification in a Community of Panamanian Strangler Figs and Associated Pollinating Wasps

The fig and pollinator wasp obligate mutualism is diverse (~750 described species), ecologically important, and ancient (~80-90 Ma), providing model systems for generating and testing many questions in evolution and ecology. Once thought to be a prime example of strict one-to-one cospeciation, current thinking suggests that genera of pollinator wasps coevolve with corresponding subsections of figs, but the degree to which cospeciation or other processes contributes to the association at finer scales is unclear. Here we use genome-wide sequence data from a community of Panamanian strangler figs (Ficus subgenus Urostigma, section Americana) and associated fig wasp pollinators (Pegoscapus spp.) to infer the process of coevolutionary diversification in this obligate mutualism. Using a model-based approach adapted from the study of gene family evolution, our results indicate pervasive and ongoing host switching of pollinator wasps at this fine phylogenetic and regional scale. Although the model estimates a modest amount of cospeciation, simulations reveal this signal to be consistent with levels of co-association expected under a model of free host switching. Our findings provide an outline for testing how ecological and evolutionary processes can be modeled to evaluate the history of association of interacting lineages in a phylogenetic framework

Host body size and the diversity of tick assemblages on Neotropical vertebrates

AbstractIdentifying the factors that influence the species diversity and distribution of ticks (Acari: Ixodida) across vertebrate host taxa is of fundamental ecological and medical importance. Host body size is considered one of the most important determinants of tick abundance, with larger hosts having higher tick burdens. The species diversity of tick assemblages should also be greater on larger-bodied host species, but empirical studies testing this hypothesis are lacking. Here, we evaluate this relationship using a comparative dataset of feeding associations from Panama between 45 tick species and 171 host species that range in body size by three orders of magnitude. We found that tick species diversity increased with host body size for adult ticks but not for immature ticks. We also found that closely related host species tended to have similar tick species diversity, but correcting for host phylogeny did not alter the relationships between host body size and tick species diversity. The distribution of tick species was highly aggregated, with approximately 20% of the host species harboring 80% of all tick species, following the Pareto principle or 20/80 Rule. Thus, the aggregated pattern commonly observed for tick burdens and disease transmission also holds for patterns of tick species richness. Our finding that the adult ticks in this system preferentially parasitize large-bodied host species suggests that the ongoing anthropogenic loss of large-bodied vertebrates is likely to result in host-tick coextinction events, even when immature stages feed opportunistically. As parasites play critical roles in ecological and evolutionary processes, such losses may profoundly affect ecosystem functioning and services

The Roots of Diversity: Below Ground Species Richness and Rooting Distributions in a Tropical Forest Revealed by DNA Barcodes and Inverse Modeling

F. Andrew Jones is with the Smithsonian Tropical Research Institute, David L. Erickson is with the Smithsonian Institution, Moises A. Bernal is with the Smithsonian Tropical Research Institute and UT Austin, Eldredge Bermingham is with the Smithsonian Tropical Research Institute, W. John Kress is with the Smithsonian Institution, Edward Allen Herre is with the Smithsonian Tropical Research Institute, Helene C. Muller-Landau is with the Smithsonian Tropical Research Institute, Benjamin L. Turner is with the Smithsonian Tropical Research Institute.Background -- Plants interact with each other, nutrients, and microbial communities in soils through extensive root networks. Understanding these below ground interactions has been difficult in natural systems, particularly those with high plant species diversity where morphological identification of fine roots is difficult. We combine DNA-based root identification with a DNA barcode database and above ground stem locations in a floristically diverse lowland tropical wet forest on Barro Colorado Island, Panama, where all trees and lianas >1 cm diameter have been mapped to investigate richness patterns below ground and model rooting distributions. Methodology/Principal Findings -- DNA barcode loci, particularly the cpDNA locus trnH-psba, can be used to identify fine and small coarse roots to species. We recovered 33 species of roots from 117 fragments sequenced from 12 soil cores. Despite limited sampling, we recovered a high proportion of the known species in the focal hectare, representing approximately 14% of the measured woody plant richness. This high value is emphasized by the fact that we would need to sample on average 13 m2 at the seedling layer and 45 m2 for woody plants >1 cm diameter to obtain the same number of species above ground. Results from inverse models parameterized with the locations and sizes of adults and the species identifications of roots and sampling locations indicates a high potential for distal underground interactions among plants. Conclusions -- DNA barcoding techniques coupled with modeling approaches should be broadly applicable to studying root distributions in any mapped vegetation plot. We discuss the implications of our results and outline how second-generation sequencing technology and environmental sampling can be combined to increase our understanding of how root distributions influence the potential for plant interactions in natural ecosystems.FAJ acknowledges the support of a Tupper postdoctoral fellowship in tropical biology and the National Science Foundation (DEB 0453665). Funding was provided by the Smithsonian Institution Global Earth Observatory, the Smithsonian Tropical Research Institute/Center for Tropical Forest Sciences endowment fund, and the Smithsonian Tropical Research Institute/Frank Levinson fund. We would like to thank Autoridad Nacional del Ambiente and the Smithsonian Tropical Research Institute for processing research permits. We thank S. Hubbell and R. Condit for access to plot data, S. Schnitzer for liana census data (NSF DEB 0613666), and L. Comita and S. Hubbell for access to seedling data (NSF DEB 0075102 and DEB 0823728). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.Marine Scienc

Host affinity of endophytic fungi and the potential for reciprocal interactions involving host secondary chemistry

PREMISE: Interactions between fungal endophytes and their host plants present useful systems for identifying important factors affecting assembly of host-associated microbiomes. Here we investigated the role of secondary chemistry in mediating host affinity of asymptomatic foliar endophytic fungi using Psychotria spp. and Theobroma cacao (cacao) as hosts. METHODS: First, we surveyed endophytic communities in Psychotria species in a natural common garden using culture-based methods. Then we compared differences in endophytic community composition with differences in foliar secondary chemistry in the same host species, determined by liquid chromatography–tandem mass spectrometry. Finally, we tested how inoculation with live and heat-killed endophytes affected the cacao chemical profile. RESULTS: Despite sharing a common environment and source pool for endophyte spores, different Psychotria host species harbored strikingly different endophytic communities that reflected intrinsic differences in their leaf chemical profiles. In T. cacao, inoculation with live and heat-killed endophytes produced distinct cacao chemical profiles not found in uninoculated plants or pure fungal cultures, suggesting that endophytes, like pathogens, induce changes in secondary chemical profiles of their host plant. CONCLUSIONS: Collectively our results suggest at least two potential processes: (1) Plant secondary chemistry influences assembly and composition of fungal endophytic communities, and (2) host colonization by endophytes subsequently induces changes in the host chemical landscape. We propose a series of testable predictions based on the possibility that reciprocal chemical interactions are a general property of plant–endophyte interactionsPREMISE: Interactions between fungal endophytes and their host plants present useful systems for identifying important factors affecting assembly of host-associated microbiomes. Here we investigated the role of secondary chemistry in mediating host affinity of asymptomatic foliar endophytic fungi using Psychotria spp. and Theobroma cacao (cacao) as hosts. METHODS: First, we surveyed endophytic communities in Psychotria species in a natural common garden using culture-based methods. Then we compared differences in endophytic community composition with differences in foliar secondary chemistry in the same host species, determined by liquid chromatography–tandem mass spectrometry. Finally, we tested how inoculation with live and heat-killed endophytes affected the cacao chemical profile. RESULTS: Despite sharing a common environment and source pool for endophyte spores, different Psychotria host species harbored strikingly different endophytic communities that reflected intrinsic differences in their leaf chemical profiles. In T. cacao, inoculation with live and heat-killed endophytes produced distinct cacao chemical profiles not found in uninoculated plants or pure fungal cultures, suggesting that endophytes, like pathogens, induce changes in secondary chemical profiles of their host plant. CONCLUSIONS: Collectively our results suggest at least two potential processes: (1) Plant secondary chemistry influences assembly and composition of fungal endophytic communities, and (2) host colonization by endophytes subsequently induces changes in the host chemical landscape. We propose a series of testable predictions based on the possibility that reciprocal chemical interactions are a general property of plant–endophyte interaction

Pervasive effects of a dominant foliar endophytic fungus on host genetic and phenotypic expression in a tropical tree

It is increasingly recognized that macro-organisms (corals, insects, plants, vertebrates) consist of both host tissues and multiple microbial symbionts that play essential roles in their host’s ecological and evolutionary success. Consequently, identifying benefits and costs of symbioses, as well as mechanisms underlying them are research priorities. All plants surveyed under natural conditions harbor foliar endophytic fungi (FEF) in their leaf tissues, often at high densities. Despite producing no visible effects on their hosts, experiments have nonetheless shown that FEF reduce pathogen and herbivore damage. Here, combining results from three genomic, and two physiological experiments, we demonstrate pervasive genetic and phenotypic effects of the apparently asymptomatic endophytes on their hosts. Specifically, inoculation of endophyte-free (E−) Theobroma cacao leaves with Colletotrichum tropicale (E+), the dominant FEF species in healthy T. cacao, induces consistent changes in the expression of hundreds of host genes, including many with known defensive functions. Further, E+ plants exhibited increased lignin and cellulose content, reduced maximum rates of photosynthesis (Amax), and enrichment of nitrogen-15 and carbon-13 isotopes. These phenotypic changes observed in E+ plants correspond to changes in expression of specific functional genes in related pathways. Moreover, a cacao gene (Tc00g04254) highly up-regulated by C. tropicale also confers resistance to pathogen damage in the absence of endophytes or their products in host tissues. Thus, the benefits of increased pathogen resistance in E+ plants are derived in part from up-regulation of intrinsic host defense responses, and appear to be offset by potential costs including reduced photosynthesis, altered host nitrogen metabolism, and endophyte heterotrophy of host tissues. Similar effects are likely in most plant-endophyte interactions, and should be recognized in the design and interpretation of genetic and phenotypic studies of plantsIt is increasingly recognized that macro-organisms (corals, insects, plants, vertebrates) consist of both host tissues and multiple microbial symbionts that play essential roles in their host’s ecological and evolutionary success. Consequently, identifying benefits and costs of symbioses, as well as mechanisms underlying them are research priorities. All plants surveyed under natural conditions harbor foliar endophytic fungi (FEF) in their leaf tissues, often at high densities. Despite producing no visible effects on their hosts, experiments have nonetheless shown that FEF reduce pathogen and herbivore damage. Here, combining results from three genomic, and two physiological experiments, we demonstrate pervasive genetic and phenotypic effects of the apparently asymptomatic endophytes on their hosts. Specifically, inoculation of endophyte-free (E−) Theobroma cacao leaves with Colletotrichum tropicale (E+), the dominant FEF species in healthy T. cacao, induces consistent changes in the expression of hundreds of host genes, including many with known defensive functions. Further, E+ plants exhibited increased lignin and cellulose content, reduced maximum rates of photosynthesis (Amax), and enrichment of nitrogen-15 and carbon-13 isotopes. These phenotypic changes observed in E+ plants correspond to changes in expression of specific functional genes in related pathways. Moreover, a cacao gene (Tc00g04254) highly up-regulated by C. tropicale also confers resistance to pathogen damage in the absence of endophytes or their products in host tissues. Thus, the benefits of increased pathogen resistance in E+ plants are derived in part from up-regulation of intrinsic host defense responses, and appear to be offset by potential costs including reduced photosynthesis, altered host nitrogen metabolism, and endophyte heterotrophy of host tissues. Similar effects are likely in most plant-endophyte interactions, and should be recognized in the design and interpretation of genetic and phenotypic studies of plant