Radopholus similis
is an important migratory endoparasitic nematode, severely harms banana, citrus and many other commercial crops. Little is known about the molecular mechanism of infection and pathogenesis of
R. similis
. In this study, 64761 unigenes were generated from eggs, juveniles, females and males of
R. similis
. 11443 unigenes showed significant expression difference among these four life stages. Genes involved in host parasitism, anti-host defense and other biological processes were predicted. There were 86 and 102 putative genes coding for cell wall degrading enzymes and antioxidase respectively. The amount and type of putative parasitic-related genes reported in sedentary endoparasitic plant nematodes are variable from those of migratory parasitic nematodes on plant aerial portion. There were no sequences annotated to effectors in
R. similis
, involved in feeding site formation of sedentary endoparasites nematodes. This transcriptome data provides a new insight into the parasitic and pathogenic molecular mechanisms of the migratory endoparasitic nematodes. It also provides a broad idea for further research on
R. similis
.
Root-knot nematodes are one of the most harmful plant-parasitic nematodes (PPNs). In this paper, the predation of Stratiolaelaps scimitus against Meloidogyne incognita was tested in an individual arena, and the control efficiency of the mite on the nematode in the water spinach (Ipomoea aquatica) rhizosphere was studied with a pot experiment. The results showed that S. scimitus could develop normally and complete its life cycle by feeding on second-stage juveniles of M. incognita (Mi-J2). The consumption rate of a 24 h starving female mite on Mi-J2 increased with the increase of prey density at 25 °C. Among the starvation treatments, the nematode consumption rate of a female mite starved for 96 h at 25 °C was highest; and among temperature treatments, the maximum consumption rate of a 24 h starving female mite on Mi-J2 was at 28 °C. The number of M. incognita in the spinach rhizosphere could be reduced effectively by releasing S. scimitus into rhizosphere soil, and 400 mites per pot was the optimum releasing density in which the numbers of root knots and egg masses decreased by 50.9% and 62.8%, respectively. Though we have gained a greater understanding of S. scimitus as a predator of M. incognita, the biocontrol of M. incognita using S. scimitus under field conditions remains unknown and requires further study.
Radopholus similis is a migratory endoparasitic nematode that is extremely harmful to host plants. Venom allergen-like proteins (VAPs) are members of the cysteine-rich secretory protein family that are widely present in plants and animals. In this study, we cloned a VAP gene from R. similis, designated as RsVAP. RsVAP contains an open reading frame of 1089 bp encoding 362 amino acids. RsVAP is specifically expressed in the esophageal gland, and the expression levels of RsVAP are significantly higher in juveniles than in other life stages of R. similis. This expression pattern of RsVAP was consistent with the biological characteristics of juveniles of R. similis, which have the ability of infection and are the main infection stages of R. similis. The pathogenicity and reproduction rate of R. similis in tomato was significantly attenuated after RsVAP was silenced. In tobacco leaves transiently expressing RsVAP, the pathogen-associated molecular pattern-triggered immunity (PTI) induced by a bacterial flagellin fragment (flg22) was inhibited, while the cell death induced by two sets of immune elicitors (BAX and Gpa2/RBP-1) was repressed. The RsVAP-interacting, ras-related protein RABA1d (LeRabA1d) was identified in tomato hosts by yeast two-hybrid and co-immunoprecipitation assays. RsVAP may interact with LeRabA1d to affect the host defense response, which in turn facilitates nematode infection. This study provides the first evidence for the inhibition of plant defense response by a VAP from migratory plant-parasitic nematodes, and, for the first time, the target protein of R. similis in its host was identified.
Six candidate reference genes were chosen from the transcriptome database of Radopholus similis using the bioinformatics method, including four conventional reference genes (actin, Eukaryotic translation initiation factor 5A (eIF5A), Tubulin alpha (a-tubulin), ubiquitin (UBI)) and two new candidate reference genes (Ribosomal protein S21 (Rps21) and Serine/threonine protein phosphatase PP1-β catalytic subunit (β-PP1)). In addition, a traditional reference gene 18S ribosomal RNA (18S rRNA) obtained from NCBI databases was also added to the analysis. Real-time PCR was used to detect the expression of seven candidate reference genes in six populations of R. similis and four developmental stages (female, male, larva and egg) of a population. The stability of the expression of candidate genes was evaluated by three software programs, BestKeeper, geNorm and NormFinder. The results showed that eIF5A is the most suitable reference gene for gene functional research of different populations, while both Rps21 and eIF5A are the most suitable reference genes for different developmental stages of a population. Therefore, eIF5A is the best reference gene for studying R. similis. However, one defect of this study is that only seven candidate reference genes were analyzed; ideally, more genes should be tested.
Plant kinases containing the LysM domain play important roles in pathogen recognition and self-defense reactions. And it could recognize microbe-associated molecules including chitin and other polypeptides. The white tip nematode Aphelenchoides besseyi is a migratory parasitic nematode that infects plant shoots. It is distributed over almost all rice-producing areas and causes up to 50% economic losses. The rice OsRLK3 gene was a defense-related LysM kinase gene of rice. This study showed that the rice LysM kinase OsRLK3 could be induced by flg22, jasmonic acid, salicylic acid, and chitin. An interaction gene, Ab-atps from A. besseyi, was identified by screening the interaction between the rice gene OsRLK3 and an A. besseyi cDNA library using yeast two-hybrid screening. Ab-atps is a novel ATP synthase gene with a full length of 1341 bp, coding for 183 amino acids. The mRNA of Ab-atps was located in the esophagus and reproductive system of A. besseyi. The expression of Ab-atps was assessed at different developmental stages of the nematode and found to be the highest in the juvenile, followed by the egg, female, and male. Reproduction was significantly decreased in nematodes treated with Ab-atps double-stranded RNA (dsRNA) (p < 0.05). Transient expression experiments showed that Ab-ATPS-GFP was distributed in the nucleus, cytoplasm, and cell membrane, and Ab-ATPS-GFP triggered plant cell death. OsRLK3 was expressed significantly higher at 0.5 day and 1 day (p < 0.05) in rice plants inoculated with nematodes treated with Ab-atps dsRNA and gfp dsRNA for 0.5–7 days, respectively. Further, OsRLK3 expression under Ab-atps dsRNA treatment was significantly lower than with gfp dsRNA treatment at 0.5 day (p < 0.05) and significantly higher than with gfp dsRNA treatment at 1 day (p < 0.05). These results suggest that rice OsRLK3 could interact with A. besseyi Ab-atps, which plays an important role in growth, reproduction, and infection of the nematode. Our findings provide a theoretical basis to further understand the parasitic strategy of A. besseyi and its interaction mechanism with host plants, suggesting new ideas and targets for controlling A. besseyi.
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