Sedentary plant-parasitic cyst nematodes are biotrophs that cause significant losses in agriculture. Parasitism is based on modifications of host root cells that lead to the formation of a hypermetabolic feeding site (a syncytium) from which nematodes withdraw nutrients. The host cell cycle is activated in an initial cell selected by the nematode for feeding, followed by activation of neighboring cells and subsequent expansion of feeding site through fusion of hundreds of cells. It is generally assumed that nematodes manipulate production and signaling of the plant hormone cytokinin to activate cell division. In fact, nematodes have been shown to produce cytokinin in vitro; however, whether the hormone is secreted into host plants and plays a role in parasitism remained unknown. Here, we analyzed the spatiotemporal activation of cytokinin signaling during interaction between the cyst nematode, Heterodera schachtii, and Arabidopsis using cytokinin-responsive promoter:reporter lines. Our results showed that cytokinin signaling is activated not only in the syncytium but also in neighboring cells to be incorporated into syncytium. An analysis of nematode infection on mutants that are deficient in cytokinin or cytokinin signaling revealed a significant decrease in susceptibility of these plants to nematodes. Further, we identified a cytokinin-synthesizing isopentenyltransferase gene in H. schachtii and show that silencing of this gene in nematodes leads to a significant decrease in virulence due to a reduced expansion of feeding sites. Our findings demonstrate the ability of a plant-parasitic nematode to synthesize a functional plant hormone to manipulate the host system and establish a long-term parasitic interaction.Arabidopsis thaliana | cell cycle | cytokinin | cyst nematode | IPT P lant-parasitic nematodes are a significant threat to almost all economically important crops. International surveys revealed an average annual crop yield loss of more than 10% due to nematode infestation and up to 20% for certain crops, e.g., bananas (1). Most of this damage is caused by the sedentary rootknot (Meloidogyne spp.) and cyst nematodes (Globodera spp. and Heterodera spp.). Infective second-stage juveniles (J2) of both rootknot nematodes (RKNs) and cyst nematodes invade plant roots near the tip and move toward the vascular cylinder. On reaching the vascular cylinder, RKNs induce the formation of several giant cells, whereas cyst nematodes induce the formation of a syncytium. These feeding sites serve as the nematode's sole source of nutrients throughout its life cycle for several weeks. Cyst nematodes are dimorphic, but the mechanism of sex determination is not clearly understood. It has, nonetheless, been observed that the environment strongly influences the outcome of the sex ratio in cyst nematodes. Under favorable conditions with plenty of nutrients, the majority of juveniles develop into females. However, when the juveniles are exposed to adverse conditions, as seen in resistant plants, the percentage of males increase...

Beetles (Coleoptera) are the most diverse animal group on earth and interact with numerous symbiotic or pathogenic microbes in their environments. The red flour beetle Tribolium castaneum is a genetically tractable model beetle species and its whole genome sequence has recently been determined. To advance our understanding of the molecular basis of beetle immunity here we analyzed the whole transcriptome of T. castaneum by high-throughput next generation sequencing technology. Here, we demonstrate that the Illumina/Solexa sequencing approach of cDNA samples from T. castaneum including over 9.7 million reads with 72 base pairs (bp) length (approximately 700 million bp sequence information with about 30× transcriptome coverage) confirms the expression of most predicted genes and enabled subsequent qualitative and quantitative transcriptome analysis. This approach recapitulates our recent quantitative real-time PCR studies of immune-challenged and naïve T. castaneum beetles, validating our approach. Furthermore, this sequencing analysis resulted in the identification of 73 differentially expressed genes upon immune-challenge with statistical significance by comparing expression data to calculated values derived by fitting to generalized linear models. We identified up regulation of diverse immune-related genes (e.g. Toll receptor, serine proteinases, DOPA decarboxylase and thaumatin) and of numerous genes encoding proteins with yet unknown functions. Of note, septic-injury resulted also in the elevated expression of genes encoding heat-shock proteins or cytochrome P450s supporting the view that there is crosstalk between immune and stress responses in T. castaneum. The present study provides a first comprehensive overview of septic-injury responsive genes in T. castaneum beetles. Identified genes advance our understanding of T. castaneum specific gene expression alteration upon immune-challenge in particular and may help to understand beetle immunity in general.

Root-knot nematodes (genus Meloidogyne) are plant parasites causing huge economic loss in the agricultural industry and affecting severely numerous developing countries. Control methods against these plant pests are sparse, the preferred one being the deployment of plant cultivars bearing resistance genes against Meloidogyne species. However, M. enterolobii is not controlled by the resistance genes deployed in the crop plants cultivated in Europe. The recent identification of this species in Europe is thus a major concern. Here, we sequenced the genome of M. enterolobii using short and long-read technologies. The genome assembly spans 240 Mbp with contig N50 size of 143 kbp, enabling high-quality annotations of 59,773 coding genes, 4,068 non-coding genes, and 10,944 transposable elements (spanning 8.7% of the genome). We validated the genome size by flow cytometry and the structure, quality and completeness by bioinformatics metrics. This ensemble of resources will fuel future projects aiming at pinpointing the genome singularities, the origin, diversity, and adaptive potential of this emerging plant pest.

The beet cyst nematode Heterodera schachtii is able to infect Arabidopsis plants and induce feeding sites in the root. These syncytia are the only source of nutrients for the nematodes throughout their life and are a nutrient sink for the host plant. We have studied here the role of amino acid transporters for nematode development. Arabidopsis contains a large number of different amino acid transporters in several gene families but those of the AAP family were found to be especially expressed in syncytia. Arabidopsis contains 8 AAP genes and they were all strongly expressed in syncytia with the exception of AAP5 and AAP7, which were slightly downregulated. We used promoter::GUS lines and in situ RT-PCR to confirm the expression of several AAP genes and LHT1, a lysine- and histidine-specific amino acid transporter, in syncytia. The strong expression of AAP genes in syncytia indicated that these transporters are important for the transport of amino acids into syncytia and we used T-DNA mutants for several AAP genes to test for their influence on nematode development. We found that mutants of AAP1, AAP2, and AAP8 significantly reduced the number of female nematodes developing on these plants. Our study showed that amino acid transport into syncytia is important for the development of the nematodes.

The cyst nematode Heterodera filipjevi is a plant parasite causing substantial yield loss in wheat. Resistant cultivars are the preferred method of controlling cyst nematodes. Association mapping is a powerful approach to detect associations between phenotypic variation and genetic polymorphisms; in this way favorable traits such as resistance to pathogens can be located. Therefore, a genome-wide association study of 161 winter wheat accessions was performed with a 90K iSelect single nucleotide polymorphism (SNP) chip. Population structure analysis grouped into two major subgroups and first principal component accounted 6.16% for phenotypic diversity. The genome-wide linkage disequilibrium across wheat was 3 cM. Eleven quantitative trait loci (QTLs) on chromosomes 1AL, 2AS, 2BL, 3AL, 3BL, 4AS, 4AL, 5BL, and 7BL were identified using a mixed linear model false discovery rate of P < 0.01 that explained 43% of total genetic variation. This is the first report of QTLs conferring resistance to H. filipjevi in wheat. Eight QTLs on chromosomes 1AL, 2AS, 2BL, 3AL, 4AL, and 5BL were linked to putative genes known to be involved in plant-pathogen interactions. Two other QTLs on 3BL and one QTL on 7BL linked to putative genes known to be involved in abiotic stress.