Gene silencing in root lesion nematodes (Pratylenchus spp.) significantly reduces reproduction in a plant host

Abstract

Accepted ManuscriptThis is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review of the resulting proof before it is published in its final form. Please note that during the production process errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain. hours in soaking solutions containing 10-50 mM octopamine, 0.1-1.0 mg/mL FITC, and 0.5-6 mM spermidine did not affect vitality. Spermidine phosphate salt hexahydrate rather than spermidine or spermidine trihydrochloride increased uptake of FITC by nematodes, and this resulted in more effective gene silencing. Silencing pat-10 and unc-87 genes of P. thornei and P. zeae resulted in paralysis and uncoordinated movements in both species, although to a higher degree in P. thornei.There was also a greater reduction in transcript of both genes in P. thornei indicating that it may be more susceptible to RNAi. For P. thornei treated with dsRNA of pat-10 and unc-87 there was a significant reduction (77-81%) in nematode reproduction on carrot mini discs over a 5 week period. The results show that RLNs are clearly amenable to gene silencing, and that in planta delivery of dsRNA to target genes in root lesion nematodes should confer host resistance.Moreover, for the two genes, dsRNA derived from either nematode species silenced the corresponding gene in both species. This implies cross-species control of nematodes via RNAi is possible.

De Novo Transcriptome Sequencing and Analysis of the Cereal Cyst Nematode, Heterodera avenae

“…An example of such phenotype in PPNs is the silencing of two genes involved in movement, pat-10 and unc-87 , in J2s of P. thornei which reduced reproduction by 77–81% on carrot mini discs. The phenotypic effect after feeding dsRNA to these nematodes was abnormal behaviour including twitching, slow movement, repeated banging of the head against the body and loss of orientation [72].…”

Advances in Understanding the Molecular Mechanisms of Root Lesion Nematode Host Interactions

“…When treated nematodes were transferred to carrot minidiscs for 5 weeks, there was a significant reduction in reproduction of 77-81%: Moreover, for these two genes, dsRNA derived from either nematode species silenced the corresponding gene in both species. This observation suggests that cross-species control of nematodes via RNAi is possible (106). RNAi has also been demonstrated for a transcript with a SPRY domain and calreticulin of P. zeae (32) and endoglucanases of P. vulnus (26).…”

“…The first papers that demonstrated that RNAi could also be applied to RLNs were the effective silencing of two movement-related genes, calponin (unc-87) and troponin C ( pat-10) of P. coffeae (102) and P. thornei (106): RNAi results have since been substantiated for P. vulnus (26), P. zeae (32), and P. penetrans (112). When mixed stages of P. thornei or P. zeae were soaked for 16 h in dsRNA to unc-87 or pat-10, there was a significant reduction in expression of these target genes, and the treatment resulted in paralysis and uncoordinated movements in both species.…”

“…Further functional studies are needed to provide firm evidence of how and where candidate effectors function in RLN-host interactions. A caveat to using RNAi as a functional tool is that the extent of silencing depends on several factors, including the amount of dsRNA delivered, the sequence chosen, the natural level of target gene expression, whether the target is a member of a multigene family, and whether there is apparent recovery from RNAi phenotypes-these parameters vary with target gene and species (26,102,106). However, the recent demonstration that hairy roots engineered to produce nematode dsRNA can silence the nematode target genes of P. thornei, P. vulnus, and P. penetrans feeding on them is a positive demonstration that host-induced gene silencing could provide an effective alternative to current chemical and conventional control strategies for RLNs (29,106,112,114).…”

Genomics Approaches Used to Control Plant Parasitic Nematodes - A Review

“…Besides, assisting in the identification of the key genes involved in parasitism, the genomic and EST sequences so generated could also provide clue to researchers to pinpoint appropriate target genes for RNAi experiments. Recently a report has been published by Tan et al, (2013) where they described the dsRNA mediated gene silencing in two important RLN species P. thornei and P.zeae. Artificial feeding silenced two important genes, calponin and troponin C, essential for nematode structural integrity and proper muscle contraction.…”