Infection by plant-parasitic nematodes (PPN) reduces global agricultural output by 10-15%. For rice alone this loss represents a commercial value of $35bn. All crops are affected to some degree, and the impact on subsistence farmers can be especially dire. The most pernicious PPN are highly adapted for life within plant roots. Some, such as the cyst (Globodera and Heterodera spp.) and root-knot nematodes (RKN: Meloidogyne spp.) establish a permanent feeding site by usurping host developmental circuitry. Others, including the lesion nematode (Pratylenchus spp.) function essentially as migratory herbivores. Understanding the molecular basis underpinning these distinct plant-nematode interactions is a key objective of the PPN research community.
The development of genetic tools for helminths has proven a challenge, largely because of the obligate, parasitic life stages within the host. For ethical and technical reasons, such barriers are much lower for PPN. Thus, we exploited M. hapla’s small diploid genome, plus its ability to both self- and out-cross, to develop necessary genetic resources. Parental lines (VW9 and LM) were established from field isolates that exhibited broad phenotypic diversity, and have been inbred and crossed. The resultant mapping populations (~100 RILs) segregate genotypic diversity, which we are correlating with nematode and plant phenotypes. One strategy has been a cross-species, eQTL analysis that samples each nematode gene to ask how it influences the expression of each plant gene. In parallel, each of the parents has been sequenced, and the VW9 transcriptome is now defined by more than a billion mapped Illumina reads derived from the RILs. The scaffolds from genome sequencing have been anchored to the 16 linkage groups of the map, and ~28,000 SNPs identified between the parents of our mapping populations. As more PPN genomes are obtained, it is clear that genome size and gene content can vary greatly within and between genera. For example, the M. hapla genome spans 54 Mb, whereas the genome of M. incognita is expanded to ~150 Mb. Recent analysis points to this species being a recent, interspecific hybrid, with a decaying, allo-hexaploid genome. At the other end of the spectrum, the 19.6 Mb Pratylenchus coffeae genome encodes a mere 6,700 genes, compared to ~14,500 for M. hapla. KEGG analysis revealed no obvious loss of biochemical ability as a result of being an obligate parasite. Broad comparisons, such as between P. coffeae, M. hapla, and C. elegans reveal patterns of gene family expansion/contraction, whereas closer comparisons (e.g., M. incognita vs M. hapla) provide a platform to understand evolution of host range, virulence and asexual reproduction.
However, despite the “taming” of M. hapla, several crucial deficiencies remain: 1) Seemingly trivial, cryopreservation is unreliable and largely ineffective. Currently, worm stocks must be continuously maintained on live plants; 2) Transformation remains elusive. In particular, gonadal microinjection is yet to be successful for RKN; 3) Mutagenesis is not practical. Unlike C. elegans, which can reproduce with huge loss of functionality, obligate parasites must complete complex interactions with their host in order to reproduce. Hence, in an obligate parasite, most genes are essential; 4) There is no lineage information for any PPN, limiting the scope of ablation experiments. Similarly, the anatomy is not well documented at cellular resolution. For example, it remains a debated point as to whether adult RKN retain a gut. Similarly, the mechanism of presumed muscle “degeneration” associated with the nematodes becoming sedentary is undocumented, as is the re-establishment of locomotion in males. Understanding how much anatomical and regulatory “equivalence” exists between C. elegans and PPN might be a useful guide. For example, is the logic of transcriptional regulation common to C. elegans and PPN (e.g., will M. hapla promoters drive GFP in an anticipated spatio-temporal pattern in C. elegans)? Can the 5 pharyngeal glands in C. elegans inform about the 3 such glands in PPN?