Dataset Information

Rapid detection of pecan root-knot nematode, Meloidogyne partityla, in laboratory and field conditions using loop-mediated isothermal amplification.

ABSTRACT: Meloidogyne partityla is the dominant root-knot nematode (RKN) species parasitizing pecan in Georgia. This species is known to cause a reduction in root growth and a decline in the yields of mature pecan trees. Rapid and accurate diagnosis of this RKN is required to control this nematode disease and reduce losses in pecan production. In this study, a loop-mediated isothermal amplification (LAMP) method was developed for simple, rapid, and on-site detection of M. partityla in infested plant roots and validated to detect the nematode in laboratory and field conditions. Specific primers were designed based on the sequence distinction of the internal transcribed spacer (ITS)-18S/5.8S ribosomal RNA gene between M. partityla and other Meloidogyne spp. The LAMP detection technique could detect the presence of M. partityla genomic DNA at a concentration as low as 1 pg, and no cross reactivity was found with DNA from other major RKN species such as M. javanica, M. incognita and M. arenaria, and M. hapla. We also conducted a traditional morphology-based diagnostic assay and conventional polymerase chain reaction (PCR) assay to determine which of these techniques was less time consuming, more sensitive, and convenient to use in the field. The LAMP assay provided more rapid results, amplifying the target nematode species in less than 60 min at 70°C, with results 100 times more sensitive than conventional PCR (~2-3 hrs). Morphology-based, traditional diagnosis was highly time-consuming (2 days) and more laborious than conventional PCR and LAMP assays. These features greatly simplified the operating procedure and made the assay a powerful tool for rapid, on-site detection of pecan RKN, M. partityla. The developed LAMP assay will facilitate accurate pecan nematode diagnosis in the field and contribute to the management of the pathogen.

SUBMITTER: Waliullah S

PROVIDER: S-EPMC7302683 | biostudies-literature | 2020

REPOSITORIES: biostudies-literature

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Publications

Rapid detection of pecan root-knot nematode, Meloidogyne partityla, in laboratory and field conditions using loop-mediated isothermal amplification.

Waliullah Sumyya S Bell Jessica J Jagdale Ganpati G Stackhouse Tammy T Hajihassani Abolfazl A Brenneman Timothy T Ali Md Emran ME Ali Md Emran ME

PloS one 20200618 6

Meloidogyne partityla is the dominant root-knot nematode (RKN) species parasitizing pecan in Georgia. This species is known to cause a reduction in root growth and a decline in the yields of mature pecan trees. Rapid and accurate diagnosis of this RKN is required to control this nematode disease and reduce losses in pecan production. In this study, a loop-mediated isothermal amplification (LAMP) method was developed for simple, rapid, and on-site detection of M. partityla in infested plant roots ...[more]

PMID: 32555580

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Project description:BackgroundSolanum torvum Sw is worldwide employed as rootstock for eggplant cultivation because of its vigour and resistance/tolerance to the most serious soil-borne diseases as bacterial, fungal wilts and root-knot nematodes. The little information on Solanum torvum (hereafter Torvum) resistance mechanisms, is mostly attributable to the lack of genomic tools (e.g. dedicated microarray) as well as to the paucity of database information limiting high-throughput expression studies in Torvum.ResultsAs a first step towards transcriptome profiling of Torvum inoculated with the nematode M. incognita, we built a Torvum 3' transcript catalogue. One-quarter of a 454 full run resulted in 205,591 quality-filtered reads. De novo assembly yielded 24,922 contigs and 11,875 singletons. Similarity searches of the S. torvum transcript tags catalogue produced 12,344 annotations. A 30,0000 features custom combimatrix chip was then designed and microarray hybridizations were conducted for both control and 14 dpi (day post inoculation) with Meloidogyne incognita-infected roots samples resulting in 390 differentially expressed genes (DEG). We also tested the chip with samples from the phylogenetically-related nematode-susceptible eggplant species Solanum melongena. An in-silico validation strategy was developed based on assessment of sequence similarity among Torvum probes and eggplant expressed sequences available in public repositories. GO term enrichment analyses with the 390 Torvum DEG revealed enhancement of several processes as chitin catabolism and sesquiterpenoids biosynthesis, while no GO term enrichment was found with eggplant DEG.The genes identified from S. torvum catalogue, bearing high similarity to known nematode resistance genes, were further investigated in view of their potential role in the nematode resistance mechanism.ConclusionsBy combining 454 pyrosequencing and microarray technology we were able to conduct a cost-effective global transcriptome profiling in a non-model species. In addition, the development of an in silico validation strategy allowed to further extend the use of the custom chip to a related species and to assess by comparison the expression of selected genes without major concerns of artifacts. The expression profiling of S. torvum responses to nematode infection points to sesquiterpenoids and chitinases as major effectors of nematode resistance. The availability of the long sequence tags in S. torvum catalogue will allow precise identification of active nematocide/nematostatic compounds and associated enzymes posing the basis for exploitation of these resistance mechanisms in other species.

Project description:Meloidogyne polycephannulata n. sp. is described from specimens collected from an area cultivated with carrot cv. Brasilia, in the city of Rio Paranaíba, in the region of Alto Paranaíba, Minas Gerais State, Brazil. The perineal pattern of the female is circular to ovoid with a high dorsal arch that has widely spaced, coarse annulations. The lateral field may have a deep furrow separating the dorsal and ventral arches. The medial lips are short and wide, whereas the lateral lips are large and triangular. The female stylet is 15-16 ?m long with wide knobs, distinctly divided by an indentation in the center. Its tip is slightly curved dorsally. The excretory pore opens 34-65 ?m from the anterior end. Females retain eggs and second-stage juveniles in their body cavity, similar to that of the cyst-forming nematodes. Males are 1.3-1.7 mm long and have a high head cap that is rounded and slopes posteriorly. The labial disc is fused to the medial lips. The head region has several irregular annulations that are similar in appearance to the first or second body annules that are likewise irregular, making the head region appear to be extremely large. The stylet of the male is 21-24 ?m long; it is slender, and has small, rounded knobs, that are distinctly indented medially and appear heart-shaped. The shaft has several tiny projections throughout its length. Mean second-stage juvenile length is 411.7 ?m. The juvenile head cap is elevated, the medial lips are small, and the lateral lips are elongate to triangular-shaped. The head region has several short, incomplete and irregular transverse annulations. The juvenile stylet is 14-23 ?m long with small, rounded, and sloping knobs. The thin tail ends with a short hyaline portion that is variable in size (16-26 ?m) and with a small, rounded tip. Isozyme profiles of esterases from Meloidogyne javanica show 3 strong bands (SB) at Rm 46, 59, and 66; profiles of M. polycephannulata n. sp. show a SB at Rm 47 and a weak band (WB) at Rm 52; M. petuniae has two SB at Rm 44 and 53; M. phaseoli has a SB at 53, 58, and 64 Rm; M. brasilensis has three SB at Rm 40, 58, and 66 and a WB at Rm 71; M. pisi has a SB at Rm 40, 60, and 64 and two WB at 46 and 50 Rm. Data from sequencing the 18S rDNA region of M. polycephannulata n. sp. confirms that it is different from M. arabicida, M. arenaria, M. ethiopica, M. incognita, M. javanica, M. paranaensis, and M. thailandica. Sequence identity among these eight species ranged between 85 to 93.4%. Meloidogyne polycephannulata n. sp. reproduces very well on carrot and tomato; poorly on pepper; and not at all on cotton, peanut, tobacco, watermelon, and sweet corn.

Dataset Information

Rapid detection of pecan root-knot nematode, Meloidogyne partityla, in laboratory and field conditions using loop-mediated isothermal amplification.

Publications

Rapid detection of pecan root-knot nematode, Meloidogyne partityla, in laboratory and field conditions using loop-mediated isothermal amplification.

Similar Datasets

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