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1.
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2.
The occurrence and geographic distribution of longidorid nematode species inhabiting the rhizosphere of cultivated olive (cvs. Chemlali and Chétoui) in Tunisia were investigated. Morphological and morphometrical studies identified three Longidorus and six Xiphinema species, with frequencies of prevalence as following: Longidorus africanus (23.0 %), L. euonymus (4.5 %), L. glycines (13.7 %), Xiphinema conurum (13.7 %), X. italiae (36.4 %), X. meridianum (13.7 %), X. pachtaicum (18.2 %), X. robbinsi (9.1 %), and Xiphinema sp. (4.5 %). The three Longidorus species were reported for the first time in Tunisia, in addition to two species of Xiphinema (viz. X. meridianum and X. robbinsi). Molecular characterisation using D2-D3 expansion regions of 28S rRNA and ITS1-rRNA was carried out and Bayesian inference analysis was used to reconstruct phylogenetic relationships among these species and with other longidorids. Twenty-five new D2-D3 of 28S rRNA gene sequences were obtained in the present study, seven for Longidorus and 18 for Xiphinema spp., as well as 14 new ITS1 rRNA gene sequences (seven for Longidorus and seven for Xiphinema spp.).  相似文献   

3.
Plant parasitic nematodes of the family Trichodoridae cause substantial yield losses in many agricultural crops. Rapid and accurate identification of trichodorids to the species level is critical for selection of appropriate measures for control. This study analysed 99 sequences of the D2–D3 expansion segments of the 28S rRNA gene and 131 sequences of the 18S rRNA gene from the stubby nematodes belonging to the genera Nanidorus, Paratrichodorus and Trichodorus. Species delimiting was based on the integration of morphological identification, which is not provided in the present article, and molecular‐based phylogenetic inference and sequence analysis. Twenty‐two valid species and several species complexes were identified among nematodes included in the analysis. PCR‐RFLPs of the partial 18S rDNA and the D2–D3 expansion segments of the 28S rDNA were tested and proposed for identification of these nematodes. Gel PCR‐RFLP profiles and tables with restriction fragment lengths for several diagnostic enzymes are provided for identification. Some problems of taxonomy and phylogeny of nematodes of the family Trichodoridae are also discussed.  相似文献   

4.
The plant parasitic nematode Longidorus poessneckensis found in Austria, the Czech Republic, Germany, Poland and the Slovak Republic was molecularly characterized. Mitochondrial genes encoding cytochrome c oxidase subunit I (COI) and nicotinamide dehydrogenase subunit 4 (nad4), the D2 and D3 expansion segments of 28S rRNA and Internal transcribed spacer 1 (ITS1) rRNA were sequenced for 16 L. poessneckensis populations. Six haplotypes of COI and five haplotypes of nad4 were determined. Nucleotide intraspecific variation was up to 17.1% for the partial sequenced COI gene and up to 17.7% for the partial sequenced nad4 gene, the latter being the highest up to date known intraspecific variation in this genus. The analyses of multiple amino acid sequence alignments of mitochondrial genes revealed low variability (0–2.4%) for COI gene and high divergence (0–7.6%) for nad4 gene. Intraspecific sequence diversity for the D2-D3 of 28S rRNA gene was up to 1.2% and for ITS1 rRNA gene was up to 1.6%. It has been hypothesized, that during the Last Glacial Maximum, L. poessneckensis populations probably persisted in refuge areas in the Carpathian Mountains and subsequently expanding from these areas and colonizing other European regions.  相似文献   

5.
The plant–parasitic nematode Longidorus poessneckensis from the Czech Republic was morphologically and molecularly characterised. Molecular analyses were carried out using mitochondrial DNA (cytochrome c oxidase subunit 1—cox1) and ribosomal DNA (ITS2—second internal transcribed spacer, 18S gene and D2/D3 expansion segments of the 28S gene), which were amplified and sequenced. Phylogenetic relationship of L. poessneckensis with three morphologically closely related species, i.e. L. macrosoma, L. helveticus and L. uroshis, was inferred by using maximum likelihood and maximum parsimony methods, with a female of Xiphinema diversicaudatum and a bivulval female of X. vuittenezi as outgroups. All multiple alignments yielded similar basic trees supporting the uniqueness of L. poessneckensis and the validity of the four Longidorus species identified using morphological characters. Phylogenetic analyses revealed that L. poessneckensis is more closely related to L. macrosoma and L. helveticus than to L. uroshis. High inter-population diversity (19%) was observed across the cox1 gene between two populations of L. poessneckensis.  相似文献   

6.
Longidorus helveticus was found at two out of 285 sampling sites for the first time in the Czech Republic. Females, males and juvenile stages were analyzed morphologically and morphometrically. The morphological identification of samples was verified by polymerase chain reaction using a species specific primer. Four markers of ribosomal DNA (18S, ITS1, ITS2, D2-D3 expansion segments of 28S rRNA) and two markers of mitochondrial DNA (cox1 and nad4) were sequenced and analyzed and compared with published gene sequences of other populations of L. helveticus. The partial mitochondrial cytochrome c-oxidase subunit 1 gene and partial nicotinamide dehydrogenase subunits 4 gene showed relatively high genetic variation within the species compared with ribosomal DNA markers.  相似文献   

7.
The occurrence and geographic distribution of longidorid nematode species inhabiting the rhizosphere of grapevine plants in southern Spain were investigated. Nematode surveys were conducted on 77 vineyards during the spring seasons of 2006, 2007 and 2008 in the main Andalusian grapevine-growing areas, including the provinces of Cádiz, Córdoba, and Huelva. Morphological and morphometrical studies identified two Longidorus and nine Xiphinema species, viz.: Longidorus alvegus, L. magnus, Xiphinema adenohystherum, X. hispidum, X. index, X. italiae, X. lupini, X. nuragicum, X. pachtaicum, X. rivesi, and X. turcicum. Overall, frequencies of infestation were, in decreasing order: X. pachtaicum 90.8%, X. index 30.3%, X. italiae 13.2%, L. magnus 11.8%, X. hispidum 7.9%, X. lupini 3.9%, L. alvegus and X. rivesi 2.6%, and X. adenohystherum, X. nuragicum and X. turcicum 1.3%. Xiphinema hispidum, X. lupini, L. alvegus and L. magnus were compared with nematode type specimens and are reported for the first time in Spain. Furthermore, the male of L. alvegus is described for the first time in the literature. Molecular characterisation of these species using D2–D3 expansion regions of 28S rRNA, 18S rRNA and ITS1-rRNA was carried out and maximum likelihood and Bayesian inference analysis were used to reconstruct phylogenetic relationships among these species and with other longidorids. The monophily of the genera Xiphinema and Longidorus was accepted and the genera Paralongidorus and Xiphidorus were rejected by the Shimodaira-Hasegawa test based on tree topologies.  相似文献   

8.
The Japanese cyst nematode Heterodera elachista was detected parasitizing corn cv Rixxer in Bosco Mesola (Ferrara Province) in Northern Italy. The only previous report of this nematode was in Asia (Japan, China and Iran) attacking upland rice; being this work the first report of this cyst nematode in Europe, and confirmed corn as a new host plant for this species. Integrative morphological and molecular data for this species were obtained using D2-D3 expansion regions of 28S rDNA, ITS1-rDNA, the partial 18S rDNA, the protein-coding mitochondrial gene, cytochrome oxidase c subunit I (COI), and the heat-shock protein 90 (hsp90). Heterodera elachista identified in Northern Italy was morphologically and molecularly clearly separated from other cyst nematodes attacking corn (viz. H. avenae, H. filipjevi, H. delvii, H. oryzae, H. sacchari, H. sorghi, H. zeae, Punctodera chalcoensis, and Vittadera zeaphila) and rice (H. oryzae, H. sacchari). The phylogenetic relationships of H. elachista from Northern Italy with other cyst-nematodes using rDNA and mtDNA showed a separation of the genus Heterodera in various morphospecies groups based on vulval cone structures. The development and parasitic habit of H. elachista on naturally infected corn cv Rixxer confirmed a typical susceptible reaction, including multinucleate syncytial cells in parenchymatic cells. Under greenhouse conditions, H. elachista successfully reproduced on two crops widely used in Northern Italy, such as corn (cv PR 33) and rice (cv Baldo). Considering the limited host-range of this nematode, that include two of the three world’s most important crops, special attention is needed for avoiding the dispersal of this nematode into new areas, by movement of soil on equipment, water, and contaminated containers infested soil, or agricultural practices.  相似文献   

9.
A new species in the genus Ditylenchus, D. stenurus n. sp. collected from western Iran, is described and illustrated herein based on morphological and molecular studies. The new species is characterised by a body length of 772 (663–863) μm, delicate stylet 6 (5–7) μm long, six lines in the lateral field. Median bulb of pharynx well-developed, muscular with crescentic valve. Post-vulval uterine sac well-developed, 35 (30–45) μm long, female tail elongate-conoid, becoming narrow suddenly with finely rounded terminus. The new species comes close in morphology and morphometrics to five known species of the genus, namely D. arachis, D. caudatus, D. clarus, D. myceliophagus, and D. nanus. DNA sequencing data was obtained on the partial 18S, D2/D3 expansion segments of the 28S rRNA gene and internal transcribed spacer (ITS). The phylogenetic relationships of this species with other Ditylenchus spp. using partial 18S–rDNA and D2/D3 indicated that D. stenurus n. sp. clustered together with several species belongs to the D. triformis-group i. e. D. africanus, D. destructor and D. halictus: all sharing a rounded tail terminus and six lines in lateral fields.  相似文献   

10.
A new mono-sexual root-lesion nematode species, Pratylenchus oleae n. sp., parasitizing roots of olive plants cv. Koroneiki in commercial fields at Ouled Chamekh (central Tunisia), and wild and cultivated olive (cv. Picual) plants in Agua Amarga (southern Spain) is described. The new species is characterised by the female having a lip region slightly offset and bearing three annuli, stylet 16.5 (14.5-17.0) μm long, with prominent rounded knobs, pharyngeal overlapping rather long (22–36) μm, lateral fields areolated and with four incisures and diagonal lines in middle band, spermatheca rounded but non-functional, tail short, conoid-rounded to subcylindrical, usually annulated terminus, males unknown, and a specific D2-D3, ITS1, 18S-rRNA, hsp90 and COI sequences. Morphologically this species is related to P. cruciferus, P. delattrei, and P. kumamotoensis. The results of the phylogenetic analysis based on sequences of the D2-D3 expansion regions of 28S, partial 18S and ITS rRNA genes confirmed the close relationship of P. oleae n. sp. with P. dunensis, P. penetrans, P. pinguicaudatus, from which was clearly separated. A PCR-based diagnostic assay was also developed for identification of P. oleae n. sp. using the species-specific primers Poleae_fw1_4 and Poleae_rv1 that amplify a 547-bp fragment in the internal transcribed spacer (ITS1) region of ribosomal DNA, which clearly separate from other root-lesion nematodes damaging olive such as P. penetrans and P. vulnus.  相似文献   

11.
四种短体线虫的形态和分子生物学鉴定   总被引：1，自引：0，他引：1  
 采用形态学和分子生物学方法对来自荷兰的藏橐吾和铃兰、泰国的高山榕、缅甸的香蕉等种苗(球)分别鉴定出玻利维亚短体线虫Pratylenchus bolivianus、铃兰短体线虫P. convallariae、咖啡短体线虫P. coffeae和斯佩奇短体线虫 P. speijeri。对这4种线虫的形态特征进行较为详细的描述后,认为头环、口针、侧区、生殖系统和尾形等形态特征是种类鉴定的重要依据;进一步利用引物28S-D2A/28S-D3Br扩增测序得到上述4种线虫的28S rRNA基因D2/D3区序列,序列分析发现玻利维亚短体线虫从欧洲到美洲的不同种群之间的遗传距离变异较小,仅为0~0.007;铃兰短体线虫同一种群不同个体之间的遗传变异较大,为0.006~0.029;咖啡短体线虫不同种群之间的平均遗传距离为0.013;斯佩奇短体线虫不同种群之间的平均遗传距离为0.010;咖啡短体线虫P. coffeae和斯佩奇短体线虫P. speijeri亲缘关系很近,二者种间平均遗传距离仅为0.026。本研究再次证明线虫28S rRNA基因D2/D3区基因序列可作为短体线虫种间鉴定的依据。  相似文献   

12.
四种短体线虫的形态和分子生物学鉴定   总被引：1，自引：0，他引：1  
 采用形态学和分子生物学方法对来自荷兰的藏橐吾和铃兰、泰国的高山榕、缅甸的香蕉等种苗(球)分别鉴定出玻利维亚短体线虫Pratylenchus bolivianus、铃兰短体线虫P. convallariae、咖啡短体线虫P. coffeae和斯佩奇短体线虫 P. speijeri。对这4种线虫的形态特征进行较为详细的描述后,认为头环、口针、侧区、生殖系统和尾形等形态特征是种类鉴定的重要依据;进一步利用引物28S-D2A/28S-D3Br扩增测序得到上述4种线虫的28S rRNA基因D2/D3区序列,序列分析发现玻利维亚短体线虫从欧洲到美洲的不同种群之间的遗传距离变异较小,仅为0~0.007;铃兰短体线虫同一种群不同个体之间的遗传变异较大,为0.006~0.029;咖啡短体线虫不同种群之间的平均遗传距离为0.013;斯佩奇短体线虫不同种群之间的平均遗传距离为0.010;咖啡短体线虫P. coffeae和斯佩奇短体线虫P. speijeri亲缘关系很近,二者种间平均遗传距离仅为0.026。本研究再次证明线虫28S rRNA基因D2/D3区基因序列可作为短体线虫种间鉴定的依据。  相似文献   

13.
During nematode surveys in southern Spain and Italy 14 populations of Xiphinema species tentatively identified as Xiphinema americanum-group were detected. Morphological and morphometrical studies identified three new species and six known Xiphinema americanum-group species, viz.: Xiphinema parabrevicolle n. sp., Xiphinema parapachydermum n. sp., Xiphinema paratenuicutis n. sp., Xiphinema duriense, Xiphinema incertum, Xiphinema opisthohysterum, Xiphinema pachtaicum, Xiphinema rivesi, and Xiphinema santos. The Xiphinema americanum-group is the most difficult Xiphinema species group for diagnosis since the morphology is very conservative and morphometric characters often overlap. This group includes vectors of several important plant pathogenic viruses that cause significant damage to a wide range of agricultural crops. Molecular characterisation of these species using D2-D3 expansion regions of 28S rRNA, 18S rRNA, ITS1-rRNA and the protein-coding mitochondrial gene, cytochrome oxidase c subunit 1 was carried out and maximum likelihood and Bayesian inference analysis were used to reconstruct phylogenetic relationships among these species and with other Xiphinema americanum-group species.  相似文献   

14.
The occurrence and geographic distribution of longidorid nematode species inhabiting the rhizosphere of cultivated and wild olive and grapevine in Crete Island were investigated. Morphological and morphometrical studies identified five Longidorus and six Xiphinema species, with frequencies of prevalence (for wild and cultivated olives and grapevines, respectively) as follows: Longidorus closelongatus (2.0–13.3 %), L. cretensis (1.0–6.7 %), L. moesicus (13.3 % only in grapevines), L. orientalis (3.3 % only in grapevines), L. pseudoelongatus (7.0 % only in olives), Xiphinema cretense n. sp. (3.0 % only in olives), X. index (3.0–23.3 %), X. israeliae (6.3 % only in olives), X. italiae (3.3–10.0 %), X. pachtaicum (26.7–42 %) and X. simile (3.3 % only in grapevines). Xiphinema cretense n. sp. is characterized by a body size 3,872–6,135 μm long, lip region anteriorly rounded, separated from the rest of the body by a depression, odontostyle and odontophore 140.6 and 80.3 μm long respectively, vulva position at 46.0–50.5 %, female tail 31.0–38.0 μm long, nearly hemispherical with curvature essentially dorsal and with a tip completely rounded or presenting a very short bulge, c ratio (119.1–186.9), c’ ratio (0.7–0.8). Molecular characterisation using D2-D3 expansion regions of 28S rRNA, 18S rRNA and ITS1-rRNA was carried out and maximum likelihood and Bayesian inference analysis were used to reconstruct phylogenetic relationships among these species and with other longidorids.  相似文献   

15.
The reniform nematodes of the genus Rotylenchulus are semi-endoparasites of numerous herbaceous and woody plant roots and distributed in regions with Mediterranean, subtropical and tropical climates. In this study, we provide morphological and molecular characterisation of three out of 11 valid species of the genus Rotylenchulus: R. macrodoratus, R. macrosoma, and R. reniformis from Greece (Crete), Italy and Spain. The overall prevalence of reniform nematodes in wild and cultivated olives in Greece, Italy, and Spain was 11.5%, 19.0% and 0.6%, respectively. In Greece, R. macrodoratus and R. macrosoma were detected in cultivated olive with a prevalence of 8.2% and 6.2%, respectively, but none of them were found in wild olive. This is the first report of R. macrosoma in Greece. Only one reniform nematode species was detected in olive from Italy and Spain, viz. R. macrodoratus and R. macrosoma, respectively. The parasitism of R. macrosoma on hazelnut in northern Spain was also confirmed for the first time. This study demonstrates that R. macrodoratus and R. macrosoma have two distinct rRNA gene types in their genomes, specifically the two types of D2-D3 for R. macrosoma and R. macrodoratus, the two types of ITS for R. macrodoratus and the testing of the ITS variability in other R. macrosoma populations in different countries. Rotylenchulus macrosoma from Greece and Spain showed differences in nucleotide sequences in the ITS region and D2-D3 of 28S rRNA gene.  相似文献   

16.
A nematode survey conducted in 2013 in Algeria, revealed that potato cyst nematodes (PCN) and cereal cyst nematodes (CCN) are widely distributed in several potato and cereal growing regions of the country. Sixteen PCN populations from five localities and five CCN populations from four of these localities were collected and characterized at the morphological and molecular levels. The PCN populations were identified as Globodera rostochiensis and G. pallida occurring separately or in mixed populations. Two species of CCN were detected. Heterodera avenae was found in four localities, whereas H. hordecalis only in one locality in association with H. avenae. The morphological and morphometric identification of PCN and CCN was confirmed by diagnostic ITS-RFLP profiles and sequencing. Phylogenetic analysis of the ITS, D2-D3 expansion domains of the 28S rRNA gene and 18S rRNA gene was made for PCN and CCN populations. Globodera pallida and G. rostochiensis from Algeria show great similarity with European and South American populations. Because of the high divergence among Algerian populations of G. pallida and G. rostochiensis it can be assumed that they were multi-introduced in Algeria. The most divergent population of G. pallida, that formed a well-separated group with some populations from Chile and Peru, suggests a later or independent introduction of this population into Algeria. Heterodera avenae and H. hordecalis formed a well-supported cluster with the corresponding populations.  相似文献   

17.
The Bursaphelenchus genus (Nematoda: Parasitaphelenchidae) comprises mostly wood‐inhabiting nematodes that feed on various tree‐colonizing fungi. One species of the genus, B. xylophilus, has been proven as an agent causing pine wilt disease (PWD). However, involvement of other Bursaphelenchus species in the PWD remains enigmatic. In the current paper, comparative molecular analysis is performed based on nuclear ribosomal DNA (rDNA) of B. vallesianus, a species that was recently isolated from pine trees (Pinus sylvestris) exhibiting wilting and declining symptoms in the Czech Republic. Sequencing of the nuclear‐encoded ITS1–5·8S–ITS2 rDNA region confirmed previous taxonomic conclusions based on morphology. Evolutionary reconstructions resulted in a phylogenetic tree, where the Czech isolate of B. vallesianus occupied a common clade together with other species belonging to the so‐called B. sexdentati group. Unexpectedly, comprehensive analysis of the sequence data revealed a genetic variation distinguishing the Czech isolate of B. vallesianus from all other species of the B. sexdentati group. This dissimilarity consists of the presence of a four nucleotide exchange found in the 5·8S rRNA‐coding gene. The newly identified genetic variation appears to affect the 5·8S rRNA folding, as deduced from secondary structure models. Additionally, it is shown that for the first time, to the authors’ knowledge, both bursaphelenchid internal transcribed spacers (ITS1 and ITS2) fold into the multibranched closed loops. While the ITS2 closed loop is formed with help of canonical 5·8S‐28S rRNA pairing, the ITS1 forms the thermodynamically stable closed loop with no support of flanking rRNA sequences. The current information on bursaphelenchid ITS rDNA sequence diversity and structure is further discussed.  相似文献   

18.
Cyst nematodes obtained from commercial carrot fields in Ontario (Canada) and northern and southern Italy were subjected to morphological and molecular examination. Morphology of cyst cone tops, males and second-stage juveniles (J2) indicated the nematode species was the Carrot Cyst Nematode (CaCN), Heterodera carotae. The sequence of the Internal Transcribed Spacer (ITS), D2-D3 region of the 28S gene of ribosomal RNA, cytochrome oxidase I of mitochondrial DNA (coxI), and a heat shock protein gene (hsp90), from single cysts were also examined. Sequences of ITS and D2-D3 placed all the nematodes with Heterodera carotae and other Heterodera spp. belonging to the Goettingiana group in the same clade. The novel nine coxI sequences obtained also clustered in a well-supported phylogenetic clade for H. carotae. Similarly, the six new hsp90 sequences of H. carotae generated in this study were placed in a well-supported clade (PP = 1.00) together with other two sequences of H. carotae from Greece. Restriction Fragment Length Polymorphism (RFLP) of ITS-PCR products gave a restriction pattern for RsaI different than H. carotae but the other 6 restriction patterns were similar as described in former research. A diagnostic conventional PCR method was developed based on a primer set to be specific for H. carotae using coxI sequence. These primers were also used in real time PCR to generate a melt curve specific to H. carotae. Limit of detection for CaCN in conventional PCR reaction was a single J2.  相似文献   

19.
Several species of trichodorid nematodes cause economically important disease on plants. Trichodorus comprises most of the trichodorid species. Trichodorus golestanensis n. sp. is described from a forest park in northern Iran. The new species belongs to the T. lusitanicus morpho-species group and is characterized in male by three ventromedian cervical papillae of which two at level of onchiostyle region, three ventromedian precloacal supplements, the posterior one just anterior to retracted spicules and ventrally curved spicules with mid-blade indentation provided with a few bristles. Females are distinguished by well developed triangular vaginal sclerotized pieces in lateral optical view, a rhomboid-shaped vagina and a transverse slit-like vulva in ventral view. Molecular analysis using the sequence of D2-D3 expansion segment of 28S rDNA differentiated the new species and confirmed the relationships of T. golestanensis n. sp. with T. andalusicus and T. asturanus. The sequences of D2-D3 regions were also provided for previous trichodorid records from Iran T. gilanensis, P. teres and N. minor together with discussion of their relationship.  相似文献   

20.
A new longidorid nematode, Longidorus persicus n. sp., is described and illustrated from a population extracted from soil associated with wild rose (Rosa sp.) naturally growing in the mountains close to the village of Cheshmeh-e-Nezamei near the city of Gilan-e-Gharb, Kermanshah province, western Iran. The new needle nematode is characterised by a large body size (6550–7763 μm), an anteriorly flattened lip region, ca 13 μm wide, distinctly set off from body contour by a depression, amphidial fovea large, pocket shaped, slightly asymmetrically bilobed, a moderately long and flexible odontostyle ca 86 μm long, stylet guiding ring located at ca 25 μm from anterior end, posterior arrangement of the pharyngeal gland nuclei, vulva almost equatorial (49–54 %), tail short, about 3/4 of its width, dorsally convex-conoid, with rounded terminus, with a c’ ratio ca 1.3, bearing three pairs of caudal pores and male rare (ratio 1:10 females) with spicules ca 46 μm long. Integrative diagnosis was completed with molecular data obtained using D2-D3 expansion segments of 28S rDNA, ITS1-rDNA, and partial 18S-rDNA. The phylogenetic relationships of this species with other Longidorus spp. using D2-D3 expansion segments and partial 18S-rDNA indicated that L. persicus n. sp. clustered together with L. perangustus and L. euonymus: both of them sharing an anteriorly flattened lip region and distinctly set off from body contour by a depression.  相似文献   

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