Gall-forming arthropods are highly specialized herbivores that, in combination with their hosts, produce extended phenotypes with unique morphologies [1]. Many are economically important, and others have improved our understanding of ecology and adaptive radiation [2]. However, the mechanisms that these arthropods use to induce plant galls are poorly understood. We sequenced the genome of the Hessian fly (Mayetiola destructor; Diptera: Cecidomyiidae), a plant parasitic gall midge and a pest of wheat (Triticum spp.), with the aim of identifying genic modifications that contribute to its plant-parasitic lifestyle. Among several adaptive modifications, we discovered an expansive reservoir of potential effector proteins. Nearly 5% of the 20,163 predicted gene models matched putative effector gene transcripts present in the M. destructor larval salivary gland. Another 466 putative effectors were discovered among the genes that have no sequence similarities in other organisms. The largest known arthropod gene family (family SSGP-71) was also discovered within the effector reservoir. SSGP-71 proteins lack sequence homologies to other proteins, but their structures resemble both ubiquitin E3 ligases in plants and E3-ligase-mimicking effectors in plant pathogenic bacteria. SSGP-71 proteins and wheat Skp proteins interact in vivo. Mutations in different SSGP-71 genes avoid the effector-triggered immunity that is directed by the wheat resistance genes H6 and H9. Results point to effectors as the agents responsible for arthropod-induced plant gall formation.

The genetic tractability of the Hessian fly (HF, Mayetiola destructor) provides an opportunity to investigate the mechanisms insects use to induce plant gall formation. Here we demonstrate that capacity using the newly sequenced HF genome by identifying the gene (vH24) that elicits effector-triggered immunity in wheat (Triticum spp.) seedlings carrying HF resistance gene H24. vH24 was mapped within a 230-kb genomic fragment near the telomere of HF chromosome X1. That fragment contains only 21 putative genes. The best candidate vH24 gene in this region encodes a protein containing a secretion signal and a type-2 serine/threonine protein phosphatase (PP2C) domain. This gene has an H24-virulence associated insertion in its promoter that appears to silence transcription of the gene in H24-virulent larvae. Candidate vH24 is a member of a small family of genes that encode secretion signals and PP2C domains. It belongs to the fraction of genes in the HF genome previously predicted to encode effector proteins. Because PP2C proteins are not normally secreted, our results suggest that these are PP2C effectors that HF larvae inject into wheat cells to redirect, or interfere, with wheat signal transduction pathways.

The Hessian fly (HF, Mayetiola destructor) is a plant-galling parasite of wheat (Triticum spp.). Seven percent of its genome is composed of highly diversified signal-peptideencoding genes that are transcribed in HF larval salivary glands. These observations suggest that they encode effector proteins that are injected into wheat cells to suppress basal wheat immunity and redirect wheat development towards gall formation. Genetic mapping has determined that mutations in four of these genes are associated with HF larval survival (virulence) on plants carrying four different resistance (R) genes. Here, this line of investigation was pursued further using bulked-segregant analysis combined with whole genome resequencing (BSA-seq). Virulence to wheat R genes H6, Hdic, and H5 was examined. Mutations associated with H6 virulence had been mapped previously. Therefore, we used H6 to test the capacity of BSA-seq to map virulence using a fieldderived HF population. This was the first time a non-structured HF population had been used to map HF virulence. Hdic virulence had not been mapped previously. Using a structured laboratory population, BSA-seq associated Hdic virulence with mutations in two candidate effector-encoding genes. Using a laboratory population, H5 virulence was previously positioned in a region spanning the centromere of HF autosome 2. BSA-seq resolved H5 virulence to a 1.3 Mb fragment on the same chromosome but failed to identify candidate mutations. Map-based candidate effectors were then delivered to Nicotiana plant cells via the type III secretion system of Burkholderia glumae bacteria. These experiments demonstrated that the genes associated with virulence to wheat R genes H6 and H13 are capable of suppressing plant immunity. Results are consistent with the hypothesis that effector proteins underlie the ability of HFs to survive on wheat.

The coffee berry borer (CBB) Hypothenemus hampei is the most limiting pest of coffee production worldwide. The CBB genome has been recently sequenced; however, information regarding the presence and characteristics of transposable elements (TEs) was not provided. Using systematic searching strategies based on both de novo and homology-based approaches, we present a library of TEs from the draft genome of CBB sequenced by the Colombian Coffee Growers Federation. The library consists of 880 sequences classified as 66% Class I (LTRs: 46%, non-LTRs: 20%) and 34% Class II (DNA transposons: 8%, Helitrons: 16% and MITEs: 10%) elements, including families of the three main LTR (Gypsy, Bel-Pao and Copia) and non-LTR (CR1, Daphne, I/Nimb, Jockey, Kiri, R1, R2 and R4) clades and DNA superfamilies (Tc1-mariner, hAT, Merlin, P, PIF-Harbinger, PiggyBac and Helitron). We propose the existence of novel families: Hypo, belonging to the LTR Gypsy superfamily; Hamp, belonging to non-LTRs; and rosa, belonging to Class II or DNA transposons. Although the rosa clade has been previously described, it was considered to be a basal subfamily of the mariner family. Based on our phylogenetic analysis, including Tc1, mariner, pogo, rosa and Lsra elements from other insects, we propose that rosa and Lsra elements are subfamilies of an independent family of Class II elements termed rosa. The annotations obtained indicate that a low percentage of the assembled CBB genome (approximately 8.2%) consists of TEs. Although these TEs display high diversity, most sequences are degenerate, with few full-length copies of LTR and DNA transposons and several complete and putatively active copies of non-LTR elements. MITEs constitute approximately 50% of the total TEs content, with a high proportion associated with DNA transposons in the Tc1-mariner superfamily.

The coffee berry borer (CBB); Hypothenemus hampei (Coleoptera: Curculionidae), is widely recognized as the major insect pest of coffee crops. Like many other arthropods, CBB harbors numerous bacteria species that may have important physiological roles in host nutrition, detoxification, immunity and protection. To date, the structure and dynamics of the gut-associated bacterial community across the CBB life cycle is not yet well understood. A better understanding of the complex relationship between CBB and its bacterial companions may provide new opportunities for insect control. In the current investigation, we analyzed the diversity and abundance of gut microbiota across the CBB developmental stages under field conditions by using high-throughput Illumina sequencing of the 16S ribosomal RNA gene. Overall, 15 bacterial phyla, 38 classes, 61 orders, 101 families and 177 genera were identified across all life stages, including egg, larva 1, larva 2, pupa, and adults (female and male). Proteobacteria and Firmicutes phyla dominated the microbiota along the entire insect life cycle. Among the 177 genera, the 10 most abundant were members of Ochrobactrum (15.1%), Pantoea (6.6%), Erwinia (5.7%), Lactobacillus (4.3%), Acinetobacter (3.4%), Stenotrophomonas (3.1%), Akkermansia (3.0%), Agrobacterium (2.9%), Curtobacterium (2.7%), and Clostridium (2.7%). We found that the overall bacterial composition is diverse, variable within each life stage and appears to vary across development. About 20% of the identified OTUs were shared across all life stages, from which 28 OTUs were consistently found in all life stage replicates. Among these OTUs there are members of genera Pantoea, Erwinia, Agrobacterium, Ochrobactrum, Pseudomonas, Acinetobacter, Brachybacterium, Sphingomonas and Methylobacterium, which can be considered as the gut-associated core microbiota of H. hampei. Our findings bring additional data to enrich the understanding of gut microbiota in CBB and its possible use for development of insect control strategies.