Multi-copy alpha-amylase genes are crucial for Ditylenchus destructor to parasitize the plant host

Abstract

Ditylenchus destructor is a migratory plant-parasitic nematode that causes huge damage to global root and tuber production annually. The main plant hosts of D . destructor contain plenty of starch, which makes the parasitic environment of D . destructor to be different from those of most other plant-parasitic nematodes. It is speculated that D . destructor may harbor some unique pathogenesis-related genes to parasitize the starch-rich hosts. Herein, we focused on the multi-copy alpha-amylase genes in D . destructor , which encode a key starch-catalyzing enzyme. Our previously published D . destructor genome showed that it has three alpha-amylase encoding genes, Dd_02440 , Dd_11154 , and Dd_13225 . Comparative analysis of alpha-amylases from different species demonstrated that the other plant-parasitic nematodes, even Ditylenchus dipsaci in the same genus, harbor only one or no alpha-amylase gene, and the three genes from D . destructor were closely clustered in the phylogenetic tree, indicating that there was a unique expansion of the alpha-amylase gene in D . destructor . The enzymatic activity of the three alpha-amylase proteins was verified by an enzyme assay. Quantitative real-time PCR assay showed that the expression of the three alpha-amylase genes in the post-hatching stage of D . destructor was found to be significantly higher than that in eggs. In the in situ hybridization assay, the expression of the genes was localized to the intestine, implying the association of these genes with nematode digestion. An infection assay in sweet potato demonstrated that RNA interference of any one alpha-amylase gene had no influence on the infectivity of D . destructor . Using the multi-target dsRNA cocktail method, it was found that silencing of two of the three genes inhibited nematode infection, and the infectivity of worms treated with three dsRNA simultaneously changed the most, which decreased by 76.6%. Thus, the multi-copy alpha-amylase genes in D . destructor are compensatory and crucial for nematodes to parasitize the plant host.

The Minichromosome Maintenance Complex Component 2 (MjMCM2) of Meloidogyne javanica is a potential effector regulating the cell cycle in nematode-induced galls

“…Further mining of the M. javanica genome should be performed using the effectR package, designed to predict effector proteins, including other RxLR effector proteins. Use of this approach by Tabima and Grünwald 48 resulted in the discovery of several genes in the M. javanica genome, e.g., SOD and α-amylase, whose secretion and function have been studied during parasitism by other PPNs 90 , 91 . Similarly, other cell wall-associated proteins, such as GH which catalyzes the hydrolysis of the glycosidic linkage of glycosides 57 , the 5-L-arabinanase 1 gene that is required for polysaccharide degradation 56 and a laminin-like protein that interacts with receptors anchored in the plasma membrane of cells adjacent to basement membranes 92 , have all been shown to carry degenerate RxLR-like motifs.…”

Silencing Ditylenchus destructor cathepsin L-like cysteine protease has negative pleiotropic effect on nematode ontogenesis

“…The inoculation, cultivation and extraction of D. destructor Dd01 strain, as well as screening eggs out from the extracted mixed-stage nematodes, were performed as previously described 10 . One-cell stage eggs were collected from the screened eggs by observation under an IX71 inverted microscope (Olympus, Tokyo, Japan).…”

“…First-strand complementary DNA (cDNA) was synthesized individually using the PrimeScript™ RT reagent Kit with gDNA Eraser (Cat# RR047Q, Takara, Dalian, China). The alpha-tubulin ( tba-1 ) and actin ( act-1 ) genes from D. destructor were used as internal reference genes 10 . The primers are listed in Table 1 .…”

“…Disturbing fundamental life processes such as growth, development, and reproduction, by RNAi is considered as a valuable approach for pest control. To date, only few D. destructor genes, including the paramyosin gene 8 , venom allergen-like protein gene 9 , alpha-amylase gene 10 and voltage-gated calcium channel subunit gene 11 – 13 , have been studied and identified as parasitic or pathogenesis-related genes, which could serve as potential targets for the control of the nematode. Many other putative target genes of D. destructor still need to be explored.…”

Complete Genome Sequence of Brachybacterium sp. Strain NBEC-018, Isolated from Nematode-Infected Potatoes

“…Strain NBEC-018 was isolated from a sample of rotten potatoes ( Solanum tuberosum ) collected in a farm field in Wuhan, China (30.62N, 114.13E). The collected potatoes were infected with potato rot nematodes ( Ditylenchus destructor ), an important plant-parasitic nematode ( 4 ). After the nematode-infected wounds on the potatoes were washed with sterile water, a bacterial suspension was obtained.…”