PROTECTING OUR CROPS - APPROACHES FOR PLANT PARASITIC NEMATODE CONTROL

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Gene expression changes in diapause or quiescent potato cyst nematode, Globodera pallida, eggs after hydration or exposure to tomato root diffusate

The authors thank the Education Spanish Ministry for the grant provided for the first author under the "Ayudas para la movilidad postdoctoral en centros extranjeros'' scheme. The James Hutton Institute receives funding from the Scottish Government.Plant-parasitic nematodes (PPN) need to be adapted to survive in the absence of a suitable host or in hostile environmental conditions. Various forms of developmental arrest including hatching inhibition and dauer stages are used by PPN in order to survive these conditions and spread to other areas. Potato cyst nematodes (PCN) (Globodera pallida and G. rostochiensis) are frequently in an anhydrobiotic state, with unhatched nematode persisting for extended periods of time inside the cyst in the absence of the host. This paper shows fundamental changes in the response of quiescent and diapaused eggs of G. pallida to hydration and following exposure to tomato root diffusate (RD) using microarray gene expression analysis encompassing a broad set of genes. For the quiescent eggs, 547 genes showed differential expression following hydration vs. hydratation and RD (H-RD) treatment whereas 708 genes showed differential regulation for the diapaused eggs following these treatments. The comparison between hydrated quiescent and diapaused eggs showed marked differences, with 2,380 genes that were differentially regulated compared with 987 genes following H-RD. Hydrated quiescent and diapaused eggs were markedly different indicating differences in adaptation for long-term survival. Transport activity is highly up-regulated following H-RD and few genes were coincident between both kinds of eggs. With the quiescent eggs, the majority of genes were related to ion transport (mainly sodium), while the diapaused eggs showed a major diversity of transporters (amino acid transport, ion transport, acetylcholine or other molecules).Publisher PDFPeer reviewe

‘Candidatus Xiphinematincola pachtaicus' gen. nov., sp. nov., an endosymbiotic bacterium associated with nematode species of the genus Xiphinema (Nematoda, Longidoridae)

An intracellular bacterium, strain IAST , was observed to infect several species of the plant-parasitic nematode genus Xiphinema (Xiphinema astaregiense, Xiphinema incertum, Xiphinema madeirense, Xiphinema pachtaicum, Xiphinema parapachydermum and Xiphinema vallense). The bacterium could not be recovered on axenic medium. The 16S rRNA gene sequence of IAST was found to be new, being related to the family Burkholderiaceae, class Betaproteobacteria. Fungal endosymbionts Mycoavidus cysteinexigens B1-EBT (92.9% sequence identity) and ‘Candidatus Glomeribacter gigasporarum’ BEG34 (89.8% identity) are the closest taxa and form a separate phylogenetic clade inside Burkholderiaceae. Other genes (atpD, lepA and recA) also separated this species from its closest relatives using a multilocus sequence analysis approach. These genes were obtained using a partial genome of this bacterium. The localization of the bacterium (via light and fluorescence in situ hybridization microscopy) is in the X. pachtaicum females clustered around the developing oocytes, primarily found embedded inside the epithelial wall cells of the ovaries, from where they are dispersed in the intestine. Transmission electron microscopy (TEM) observations supported the presence of bacteria inside the nematode body, where they occupy ovaries and occur inside the intestinal epithelium. Ultrastructural analysis of the bacterium showed cells that appear as mostly irregular, slightly curved rods with rounded ends, 0.8–1.2µm wide and 2.5–6.0µm long, possessing a typical Gram-negative cell wall. The peptidoglycan layer is, however, evident only occasionally and not detectable by TEM in most cells. Another irregularly occurring shell surrounding the endosymbiont cells or the cell clusters was also revealed, probably originating from the host cell membrane. Flagella or spore-like cells do not occur and the nucleoid is diffusely distributed throughout the cell. This endosymbiont is transmitted vertically through nematode generations. These results support the proposal of IAST as a new species, although its obligate intracellular and obligate endosymbiont nature prevented isolation of a definitive type strain. Strain IAST is therefore proposed as representing ‘Candidatus Xiphinematincola pachtaicus’ gen. nov., sp. nov

-Omics fields of study related to plant-parasitic nematodes

Plant-parasitic nematodes (PPN) cause significant losses and these pathogens must be addressed amid the growing demand for food, global warming, and the discarded use of inorganic pesticides. For these reasons, acquiring deeper knowledge about PPN and devising new management strategies are important in order to meet future food demand. This review focuses on PPN and their applicable and diverse –omics fields of study. While most efforts have been centered on transcriptomics, other –omics studies have recently begun to expand. The few genomes sequenced (Meloidogyne incognita, M. hapla, and Bursaphelenchus xylophilus) have shown high diversity in PPN. This review also discusses the future prospects and uses of –omics relative to PPN.

Inheritance of Type I Glandular Trichomes in Cucumis melo L.

Glandular trichomes Type I (Picture 1 and 2), firstly described in Cucumis melo by Gómez-Guillamón et al. (2006), are involved in the early rejection of melon plants as hosts by Aphis gossypii Glover, and, in high density, they could be considered as an additional factor to the aphid resistance conferred by Vat gene (Sarria et al., 2007). The knowledge of the inheritance mode of their density would enhance their utilization in melon breedingThis work has been financed by the CICYT Research Project: AGL2005-03850-C02-01Peer reviewe

Root-lesion nematodes of the genus Pratylenchus (Nematoda: Pratylenchidae) from Costa Rica with molecular identification of P. gutierrezi and P. panamaensis topotypes

Root-lesion nematodes of the genus Pratylenchus are among the most damaging nematodes for crop and ornamental plants worldwide. There are about 75 described species in this genus with a wide host range, but a difficult diagnosis because of morphological similarities and overlapping of morphometric characters among species. Four species of Pratylenchus were detected parasitizing cultivated and ornamental plants in Costa Rica: Pratylenchus bolivianus, P. gutierrezi, P. pseudocoffeae, and P. zeae; while P. panamaensis was detected in coffee in the type locality at Panama. The specimens were identified using morphological and molecular methods. Morphometrics and morphology using light and scanning electron microscopy are given for all of them. The presence of P. bolivianus and P. pseudocoffeae are reported for the first time in Costa Rica. The molecular and morphological variability associated with former populations of P. gutierrezi showed that we are in front of complex of species. This has been resolved with the study of topotype populations of P. gutierrezi and P. panamaensis combined with rDNA sequences (D2-D3 expansion segments of 28S and ITS). Our results, including SEM en face view data on topotypes of P. gutierrezi and P. panamaensis demonstrated that both species are almost undistinguishable morphologically, but clearly separated molecularly with D2-D3 expansion segments of 28S (89 % similar, 84 nucleotides and 19 indels) and ITS rDNA (only 36 % coverage with a 88 % similarity value). Consequently, P. gutierrezi and P. panamaensis need to be maintained as different valid taxa. Molecular data on D2-D3 expansion segments of 28S and ITS rDNA of topotypes of P. panamaensis were also coincident (99 % similarity) with root-lesion nematodes parasitizing coffee in Guatemala and Costa Rica, and banana in Madeira, and previously identified as P. gutierrezi (K1-AF170440, K2-AF170441, FR692277, respectively), which need to be considered conspecific with P. panamaensis. These results showed us the necessity of integrative taxonomy in the identification and characterization of Pratylenchus spp.Los nematodos lesionadores de raíces del género Pratylenchus se encuentran entre los nematodos más dañinos para cultivos y plantas ornamentales en todo el mundo. Hay alrededor de 75 especies descritas en este género con una amplia gama de hospedadores, pero un diagnóstico difícil debido a las similitudes morfológicas y la superposición de caracteres morfométricos entre las especies. Se detectaron cuatro especies de Pratylenchus parasitando plantas cultivadas y ornamentales en Costa Rica: Pratylenchus bolivianus, P. gutierrezi, P. pseudocoffeae y P. zeae; mientras que P. panamaensis se detectó en café en la localidad tipo de Panamá. Las muestras se identificaron mediante métodos morfológicos y moleculares. Para todos ellos se da morfometría y morfología mediante microscopía óptica y electrónica de barrido. Se reporta por primera vez en Costa Rica la presencia de P. bolivianus y P. pseudocoffeae. La variabilidad molecular y morfológica asociada a antiguas poblaciones de P. gutierrezi mostró que estamos frente a un complejo de especies. Esto se ha resuelto con el estudio de poblaciones de topotipos de P. gutierrezi y P. panamaensis combinadas con secuencias de ADNr (segmentos de expansión D2-D3 de 28S e ITS). Nuestros resultados, incluidos los datos de SEM en face view sobre topotipos de P. gutierrezi y P. panamaensis demostraron que ambas especies son casi indistinguibles morfológicamente, pero claramente separadas molecularmente con segmentos de expansión D2-D3 de 28S (89% similares, 84 nucleótidos y 19 indeles ) e ITS rDNA (solo cobertura del 36% con un valor de similitud del 88%). En consecuencia, P. gutierrezi y P. panamaensis deben mantenerse como taxones válidos diferentes. Los datos moleculares sobre los segmentos de expansión D2-D3 de 28S e ITS rDNA de topotipos de P. panamaensis también coincidieron (99% de similitud) con los nematodos lesionadores de raíces que parasitan el café en Guatemala y Costa Rica, y el banano en Madeira, y previamente identificados como P .gutierrezi (K1-AF170440, K2-AF170441, FR692277, respectivamente), que deben considerarse conespecíficas con P. panamaensis. Estos resultados nos mostraron la necesidad de una taxonomía integradora en la identificación y caracterización de Pratylenchus spp.Universidad Nacional, Costa RicaConsejo Superior de Investigaciones Científicas, EspañaUniversidad de Jaén, EspañaEscuela de Ciencias Agraria

DNA Barcoding and Morphometry Reveal Further Cryptic Bio-Diversity within the Pin Nematode Genus <i>Paratylenchus</i> (Nematoda: Tylenchulidae)

Paratylenchus species are obligate ectoparasitic nematodes on cultivated and wild herbaceous and woody plants occupying numerous soil categories. Several species may cause damage to several crops (viz. P. dianthus, P. enigmaticus, P. microdorus, P. hamatus and P. epacris on carnation, lettuce, rose and walnut, respectively). This investigation proves and emphasizes the relevance of applying integrative taxonomy for the accurate detection of Paratylenchus species in mountainous wild environments in the Malaga province, Southern Spain. This research analyzed 45 soil samples of maritimus pine and one of green heather in southern Spain and identified fourteen Paratylenchus species, two of them are described herein as new species (P. paraaonli sp. nov., P. plesiostraeleni sp. nov.), six of them were first reports for Spain (P. canchicus, P. nainianus, P. neonanus, P. salubris, Paratylenchus sp. 2 SAS, and P. wuae), and six species (P. caravaquenus, P. microdorus, P. nanus, P. neoamblycephalus, P. sheri, and P. variabilis) have been already reported in Spain. Accordingly, these data increase the biodiversity of pin nematodes in Spain comprising a total of 47 species (33.1% out of 142 total species of this genus). Phylogenetic analyses based on ribosomal and mitochondrial markers (D2-D3, ITS, and partial COI) resulted in a consistent position for the newly described Paratylenchus species in this study (P. plesiostraeleni sp. nov., P. paraaonli sp. nov.). Paratylenchus plesiostraeleni sp. nov. grouped in a separated subclade as unequivocal species from the P. straeleni-complex species (including P. straeleni and P. parastraeleni), and P. paraaonli sp. nov. clustered with P. vitecus, but clearly separate from this species. This study indicates that Paratylenchus species diversity in natural environments may be higher than expected, and this study may help in accurate identifications

Assessment of Helminth Biodiversity in Wild Rats Using 18S rDNA Based Metagenomics

<div><p>Parasite diversity has important implications in several research fields including ecology, evolutionary biology and epidemiology. Wide-ranging analysis has been restricted because of the difficult, highly specialised and time-consuming processes involved in parasite identification. In this study, we assessed parasite diversity in wild rats using 18S rDNA-based metagenomics. 18S rDNA PCR products were sequenced using an Illumina MiSeq sequencer and the analysis of the sequences using the QIIME software successfully classified them into several parasite groups. The comparison of the results with those obtained using standard methods including microscopic observation of helminth parasites in the rat intestines and PCR amplification/sequencing of 18S rDNA from isolated single worms suggests that this new technique is reliable and useful to investigate parasite diversity.</p></div

Species identification of isolated helminths based on 18S rDNA sequencing.

<p>Species identification of isolated helminths based on 18S rDNA sequencing.</p