Pantoea bathycoeliae sp. nov and Sodalis sp. are core gut microbiome symbionts of the two-spotted stink bug.

Stink bug species (Pentatomoidea superfamily) have developed an interdependence with obligate bacterial gut symbionts in specialized midgut crypts (M4 sub-region). Species of the Enterobacteriaceae family (predominantly Pantoea) are vertically transferred to their offspring and provide nutrients that cannot be obtained from plant sap food sources. However, the bacteria in the other gut compartments of stink bugs have rarely been investigated. The two-spotted stink bug, Bathycoelia distincta, is a serious pest of macadamias in South Africa. Nothing is currently known regarding its gut microbiome or how symbionts are transferred between insect generations. In this study, the consistency of B. distincta gut bacteria across geographic locations and life stages was determined with 16S rRNA metabarcoding, considering both the M4 and other gut compartments. A novel Pantoea species was found to be the primary M4 gut symbiont and is vertically transferred to the offspring. The other gut compartments had a low bacterial diversity and genera varied between stink bug populations but a Sodalis species was prominent in all populations. Sequence data of the M4 compartment were used to produce high-quality metagenome-assembled genomes (MAGs) for the Pantoea and Sodalis species. Functional analyses suggested a similar role in nutrient provision for the host, yet also unique metabolites produced by each species. The Sodalis sp. also had additional traits, such as secretion systems, that likely allowed it to establish itself in the host. The Pantoea species was described as Pantoea bathycoeliae sp. nov based on the rules of the SeqCode.

Growth rate is a dominant factor predicting the rhizosphere effect.

The root microbiome is shaped by plant root activity, which selects specific microbial taxa from the surrounding soil. This influence on the microorganisms and soil chemistry in the immediate vicinity of the roots has been referred to as the rhizosphere effect. Understanding the traits that make bacteria successful in the rhizosphere is critical for developing sustainable agriculture solutions. In this study, we compared the growth rate potential, a complex trait that can be predicted from bacterial genome sequences, to functional traits encoded by proteins. We analyzed 84 paired rhizosphere- and soil-derived 16S rRNA gene amplicon datasets from 18 different plants and soil types, performed differential abundance analysis, and estimated growth rates for each bacterial genus. We found that bacteria with higher growth rate potential consistently dominated the rhizosphere, and this trend was confirmed in different bacterial phyla using genome sequences of 3270 bacterial isolates and 6707 metagenome-assembled genomes (MAGs) from 1121 plant- and soil-associated metagenomes. We then identified which functional traits were enriched in MAGs according to their niche or growth rate status. We found that predicted growth rate potential was the main feature for differentiating rhizosphere and soil bacteria in machine learning models, and we then analyzed the features that were important for achieving faster growth rates, which makes bacteria more competitive in the rhizosphere. As growth rate potential can be predicted from genomic data, this work has implications for understanding bacterial community assembly in the rhizosphere, where many uncultivated bacteria reside.

A Detection Assay to Identify Alternative Food Sources of the Two-Spotted Stink Bug, Bathycoelia distincta (Hemiptera: Pentatomidae).

The two-spotted stink bug, Bathycoelia distincta Distant (Hemiptera: Pentatomidae), is a serious pest in South African macadamia orchards. This pest is predominantly controlled using insecticides, thus alternative control methods are essential. The stink bugs arrive as adults in the orchards, during the early nut set season, but little is known about their alternative plant hosts before their arrival. The aim of this study was to develop a PCR-based metabarcoding assay to identify plant material in the gut of B. distincta. Thereafter, the persistence of plant DNA in the gut, after switching food sources, was determined by rearing the stink bugs on Zea mays L. (Cyperales: Poaceae), transferring them to Macadamia sp. and then collecting insects at different time points. As a proof of concept, the assay was tested on insects collected from commercial macadamia orchards to determine if it can identify alternative food sources. The chloroplast gene markers, trnL and trnF, were most successful for plant DNA amplification. The time trial suggested that plant material can be detected 24 h after switching to the alternate food source and one of the samples still contained Z. mays DNA after five days. Various plant species were detected from the orchard collected samples, including known food sources of other stink bugs, such as tea plants (Camellia sinensis L. (Ericales:Theaceae)) and sunflowers (Helianthus annuus L. (Asterales: Asteraceae)). This study provides the first indication of potential alternative food sources of B. distincta. The assay developed in this study can now be implemented for large-scale field surveys to contribute to future integrated pest management strategies.

Genome comparisons suggest an association between Ceratocystis host adaptations and effector clusters in unique transposable element families.

Ceratocystis fimbriata is a host specific fungal pathogen of sweet potato (Ipomoea batatas). The closely related species, C. manginecans, is an important pathogen of trees (e.g. Acacia mangium and Mangifera indica) but has never been isolated from tuber crops. The genetic factors that determine the host range and host specificity of these species have not been determined. The aim of this study was to compare the genomes of C. fimbriata and C. manginecans in order to identify species-specific genetic differences that could be associated with host specificity. This included whole-genome alignments as well as comparisons of gene content and transposable elements (TEs). The genomes of the two species were found to be very similar, sharing similar catalogues of CAZymes, peptidases and lipases. However, the genomes of the two species also varied, harbouring species-specific genes (e.g. small secreted effectors, nutrient processing proteins and stress response proteins). A portion of the TEs identified (17%) had a unique distribution in each species. Transposable elements appeared to have played a prominent role in the divergence of the two species because they were strongly associated with chromosomal translocations and inversions as well as with unique genomic regions containing species-specific genes. Two large effector clusters, with unique TEs in each species, were identified. These effectors displayed non-synonymous mutations and deletions, conserved within a species, and could serve as mutational hot-spots for the development of host specificity in the two species.

QTL mapping of mycelial growth and aggressiveness to distinct hosts in Ceratocystis pathogens.

Some species of Ceratocystis display strong host specificity, such as C. fimbriata sensu stricto that is restricted to sweet potato (Ipomoea batatas) as host. In contrast, the closely related C. manginecans, infects Acacia mangium and Mangifera indica but is not pathogenic to I. batatas. Despite the economic importance of these fungi, knowledge regarding the genetic factors that influence their pathogenicity and host specificity is limited. A recent inheritance study, based on an interspecific cross between C. fimbriata and C. manginecans and the resultant 70 F1 progeny, confirmed that traits such as mycelial growth rate, spore production and aggressiveness on A. mangium and I. batatas are regulated by multiple genes. In the present study, a quantitative trait locus (QTL) analysis was performed to determine the genomic loci associated with these traits. All 70 progeny isolates were genotyped with SNP markers and a linkage map was constructed. The map contained 467 SNPs, distributed across nine linkage groups, with a total length of 1203 cm. Using the progeny genotypes and phenotypes, one QTL was identified on the linkage map for mycelial growth rate, one for aggressiveness to A. mangium and two for aggressiveness to I. batatas (P < 0.05). Two candidate genes, likely associated with mycelial growth rate, were identified in the QTL region. The three QTLs associated with aggressiveness to different hosts contained candidate genes involved in protein processing, detoxification and regions with effector genes and high transposable element density. The results provide a foundation for studies considering the function of genes regulating various quantitative traits in Ceratocystis.

Global population genomics of the forest pathogen Dothistroma septosporum reveal chromosome duplications in high dothistromin-producing strains.

Dothistroma needle blight is one of the most devastating pine tree diseases worldwide. New and emerging epidemics have been frequent over the last 25 years, particularly in the Northern Hemisphere, where they are in part associated with changing weather patterns. One of the main Dothistroma needle blight pathogens, Dothistroma septosporum, has a global distribution but most molecular plant pathology research has been confined to Southern Hemisphere populations that have limited genetic diversity. Extensive genomic and transcriptomic data are available for a D. septosporum reference strain from New Zealand, where an introduced clonal population of the pathogen predominates. Due to the global importance of this pathogen, we determined whether the genome of this reference strain is representative of the species worldwide by sequencing the genomes of 18 strains sampled globally from different pine hosts. Genomic polymorphism shows substantial variation within the species, clustered into two distinct groups of strains with centres of diversity in Central and South America. A reciprocal chromosome translocation uniquely identifies the New Zealand strains. Globally, strains differ in their production of the virulence factor dothistromin, with extremely high production levels in strain ALP3 from Germany. Comparisons with the New Zealand reference revealed that several strains are aneuploids; for example, ALP3 has duplications of three chromosomes. Increased gene copy numbers therefore appear to contribute to increased production of dothistromin, emphasizing that studies of population structure are a necessary adjunct to functional analyses of genetic polymorphisms to identify the molecular basis of virulence in this important forest pathogen.

IMA Genome-F 11: Draft genome sequences of Fusarium xylarioides, Teratosphaeria gauchensis and T. zuluensis and genome annotation for Ceratocystis fimbriata.

Draft genomes of the fungal species Fusarium xylarioides, Teratosphaeria gauchensis and T. zuluensis are presented. In addition an annotation of the genome of Ceratocystis fimbriata is presented. Overall these genomes provide a valuable resource for understanding the molecular processes underlying pathogenicity and potential management strategies of these economically important fungi.

Inheritance of phenotypic traits in the progeny of a Ceratocystis interspecific cross.

Ceratocystis fimbriata is a fungal plant pathogen that causes black rot on Ipomoea batatas. Based on inoculation studies on numerous tree species, the pathogen is known to be host specific. The closely related species, Ceratocystis manginecans, causes severe wilt on a broad range of tree hosts, including Mangifera indica, Acacia mangium and other leguminous tree species. The genetic factors underlying the pathogenicity and host specificity of Ceratocystis species have rarely been investigated. In this study, an F1 population of 70 recombinant progeny from a cross between C. fimbriata and C. manginecans was generated and the inheritance of various phenotypic traits was investigated. Results showed that colony colour, growth rate, asexual spore production and aggressiveness to I. batatas and A. mangium are all quantitative traits with high levels of heritability. However, conidia production and aggressiveness appeared to be regulated by a small number of genes. No correlation could be found between aggressiveness and other phenotypic traits, suggesting that these are inherited independently. This is the first study to consider genetic inheritance of pathogenicity and host specificity in Ceratocystis species and the results will contribute, in future, to the identification of quantitative trait loci and candidate genes associated with the traits investigated.

A possible centre of diversity in South East Asia for the tree pathogen, Ceratocystis manginecans.

The fungal pathogen, Ceratocystis manginecans, has caused serious canker and wilt disease on Mangifera indica (mango), on legume tree species in Oman and Pakistan and on Acacia spp. in Indonesia. A Ceratocystis species, with similar morphology to C. manginecans, has recently been reported in Vietnam, causing severe disease of Acacia trees. Previous population genetic studies on isolates from M. indica in Oman and Pakistan have shown that the pathogen represents a single clonal haplotype, indicative of an introduced pathogen. The aim of this study was to investigate the genetic diversity and population structure of 160 C. manginecans isolates, from four host-associated populations from Oman, Pakistan, Indonesia and Vietnam. This was done by applying a combination of 14 previously developed microsatellite markers and a new set, designed in this study from two different C. manginecans genomes. Sequence data confirmed that the isolates in Vietnam are the same species as those in Indonesia and were thus identified as C. manginecans. Unlike the populations in Oman and Pakistan, relatively high levels of genetic variation were found for the isolates from Indonesia and Vietnam. The Vietnam population was significantly differentiated from the other populations and isolates from this area had the highest level of genetic diversity thus far encountered for the pathogen.