First Report of the Dagger Nematode Xiphinema diversicaudatum in Citrus Orchards in Morocco

Abstract

Xiphinema species are migratory ectoparasitic nematodes that feed on an extensive range of hosts, and several species are vectors of nepoviruses. In May 2012, during a survey of the citrus-growing areas in the Gharb region of Morocco, several Xiphinema nematodes were detected in three locations: Kénitra (INRA, plot P1), Sidi Kacem, and Sidi Slimane. Samples were taken 30 cm deep at 50 cm distance from the tree trunks, in 40-year-old orange groves planted with the variety Maroc Late, grafted on rootstocks of Sour Orange and Citrange Carrizo. The trees showed yellowing of leaves, reduced tree vigor, and swellings at the tips of the roots. There were no weeds or grasses in the sampled area. Nematodes were extracted from soil using an automated centrifuge for extracting free-living nematode stages (2) and identified morphologically and by sequencing. All specimens were identified as Xiphinema diversicaudatum based on key morphological features from females and males. The average measurements of six females were: body length 4.1 mm, body width 60.4 μm, odontostylet 133.5 μm, odontophore 64.0 μm, spear 197.5 μm, tail length 45 μm, body width at anus 31 μm, and vulval position 48%. The females had two genital branches of similar length and structure, which contained a pseudo-Z differentiation. The average measurements of four males were: body length 4.3 mm, body width 51 μm, odontostylet 139 μm, odontophore 70 μm, spear 209 μm, tail length 45 μm, body width at anus 35 μm. To confirm the morphological identification, molecular observations were made. DNA was extracted from one nematode of each location. The D3 expansion region of the 26S rRNA gene was amplified using the primers D3A (5'-GACCCGTCTTGAAACACGGA-3') and D3B (5'-TCGGAAGGAACCAGCTACTA-3') (1). The PCR products were purified and sequenced (Macrogen, Inc., Seoul, Korea). All sequences obtained (GenBank Accession Nos. KF057879, KF057880, and KF057881) were compared with sequences available from the GenBank database including several species of Xiphinema. This comparison revealed a sequence similarity of 99 to 100% with X. diversicaudatum. Morphological and molecular identification demonstrated that the isolates of dagger nematodes from three citrus growing areas in Gharb belonged to X. diversicaudatum. An average of six X. diversicaudatum per 100 cm3 soil were found. This is the first report of this species in Morocco. X. diversicaudatum can transmit Arabis mosaic and Strawberry latent ringspot viruses (3). Arabis mosaic virus is of great economic importance in viticulture as it is associated with grapevine fanleaf degeneration disease, together with Grapevine fan leaf virus transmitted by X. index (4). As vineyards are planted amid citrus orchards in the Gharb region, particular attention should be given to this nematode, especially to the risk of its spread by soil. Our finding of X. diversicaudatum in a citrus orchard does not necessarily imply that X. diversicaudatum causes damage in citrus. However, its presence indicates that this nematode species can survive in this environment from where it could spread to other, more susceptible, crops.