New evidence of nematode-endosymbiont bacteria coevolution based on one new and one known dagger nematode species of Xiphinema americanum-group (Nematoda, Longidoridae).

Three populations of Xiphinema primum n. sp. and two populations of X. pachtaicum were recovered from natural forests and cultural regions of northern Iran. Both species belong to the X. americanum-group and were characterized by their morphological, morphometric and molecular data. The new species, which was recovered in three locations, belongs to the X. brevicolle-complex and is characterized by 2124-2981 μm long females with a widely rounded lip region separated from the rest of the body by a depression, 103-125 μm long odontostyle, two equally developed genital branches with endosymbiont bacteria inside the ovary, which are visible under light microscope (LM), vulva located at 51.8-58.0%, the tail is 26-37 μm long with a bluntly rounded end and four juvenile developmental stages. It was morphologically compared with nine similar species viz. X. brevicolle, X. diffusum, X. incognitum, X. himalayense, X. luci, X. parabrevicolle, X. paramonovi, X. parataylori and X. taylori. The second species, X. pachtaicum, was recovered in two geographically distant points close to city of Amol. Molecular phylogenetic studies of the new species were performed using partial sequences of the D2-D3 expansion segments of the large subunit ribosomal RNA gene (LSU rDNA D2-D3), the internal-transcribed spacer rDNA (ITS = ITS1+5.8S+ITS2), and the mitochondrial cytochrome c oxidase I gene (COI mtDNA) regions. The Iranian population of X. pachtaicum was also phylogenetically studied based upon its LSU rDNA D2-D3 sequences. Both species were also inspected for their putative endosymbiont bacteria. Candidatus Xiphinematobacter sp. was detected from two examined populations of the new species, whereas the second endosymbiont bacterium, detected from three examined isolates of X. pachtaicum, was related to the plant and fungal endosymbionts of the family Burkholderiaceae. The phylogenetic analyses of the two endosymbiont bacteria were performed using partial sequences of 16S rDNA. In cophylogenetic analyses, significant levels of cophylogenetic signal were observed using both LSU rDNA D2-D3 and COI mtDNA markers of the host nematodes and 16S rDNA marker of the endosymbiont bacteria

Occurrence of Xiphinema santos Lamberti, Lemos, Agostinelli & D’Addabo 1993 (Nematoda: Longidoridae), a X. americanum-group member in Iran

A population of Xiphinema americanum-group was recovered in association with stone fruit trees in Isfahan province, center of Iran. A reverse taxonomic approach based upon the large subunit ribosomal DNA (LSU rDNA D2-D3) and the mitochondrial cytochrome c oxidase subunit I (COI mtDNA) gene sequences in integration with morphological studies, revealed that the recovered population belongs to Xiphinema santos. The Iranian population was mainly characterized by 1240–1868 μm long females with 60–84 μm long odontostyle, a = 37.2–51.9 and c = 42.8–54.6. It is further characterized by a lip region having a depression in junction with the body, presence of visible endosymbiont bacteria in ovaries under light microscope, dorsally convex and ventrally slightly concave conical tail with a blunt tip and three juvenile developmental stages. This population was similar to the type population in its morphology; however overlapped and extended morphometric data ranges, as well as differences in some indexes were observed. Compared to a Spanish population of this species, the Iranian population had a close morphology, similar morphometric data ranges and identical LSU and COI sequences. In LSU phylogeny, the relationship between the present and some previously sequenced isolates of the species and some isolates of three species X. georgianum, X. laevistriatum and X. citricolum was not resolved. In COI phylogeny, the clade of the Iranian and Spanish populations appeared as an independent lineage inside an unsupported clade including several species. The comparison with other populations of the species was reported and discussed. A second species, X. primum, that is native to Iran, was recovered from a new locality and characterized molecularly.The financial support of Tarbiat Modares University is appreciated. The authors acknowledge the critically reading of the MS by Dr. Mohammad Reza Atighi. The presented findings in this paper are part of the results of the project number 0-16-16-94176 by the first author entitled “Determination of infestation of pomegranate orchards and nurseries to plant parasitic nematodes with an emphasis on root knot nematodes”