Morphologic, Morphometric, and Molecular Characterization of Vietnamese Populations of Meloidogyne incognita.

Meloidogyne incognita is considered the most damaging and common root-knot nematode to numerous host plants worldwide. During a survey of nematodes in Vietnam, 1,106 samples from 22 different plant species were collected. M. incognita was recorded on 13 of the 22 host plants. Four populations of M. incognita from four host plants were chosen for comparison and confirmation of their morphologic, morphometric, and molecular characteristics. Genetically based phylogenetic trees were constructed to show relationships among root-knot nematodes. Molecular barcodes of four gene regions, ITS, D2-D3 of 28S rRNA, COI, and Nad5 mtDNA, integrated with morphologic and morphometric data were used as reliable references for molecular identification of M. incognita. Our analyses indicated that tropical root-knot nematodes are very similar in characterization of ITS, D2-D3 of 28S rRNA, and COI regions. However, these gene regions can be used to separate the tropical root-knot nematode group from other groups. On the other hand, the analysis of Nad5 mtDNA and multiplex-PCR with specific primers can be used to distinguish tropical species.

First report of Pratylenchus penetrans (Nematoda: Pratylenchidae) associated with artichokes in Vietnam.

Pratylenchus penetrans is one of the world's most common and destructive root-lesion nematodes and can parasitize more than 400 plant species. P. penetrans has been reported to cause serious damage to artichokes in several countries, such as Greece, Brazil, and France. Until now, there have been no reports of P. penetrans associated with artichokes in Vietnam. In this study, we recorded this species in artichoke fields in Lam Dong province, Vietnam with an average density of 50 nematodes/100g of soil (frequency of appearance at 64.7%). This nematode was associated with symptoms such as yellowing leaves, stunt, and root necrosis of artichokes in Vietnam, indicating its high damaging potential and a need for suitable control strategies. The identification of this species in our study was confirmed by morphology, morphometric data, and molecular characterization of 18S and 28S rRNA regions. Our study also provides the first molecular data of P. penetrans in Vietnam. The inclusion of molecular data for P. penetrans in Vietnam represents a significant contribution to the scientific community and a pivotal advancement in addressing nematode-related challenges in agriculture. This dataset serves as an invaluable reference for various molecular-focused endeavors, including but not limited to molecular identification, pathogenicity studies, and the development of effective management strategies.

First report and new molecular and morphological characterizations of root-knot nematode, Meloidogyne javanica, infecting ginger and long coriander in Vietnam.

Ginger (Zingiber officinale Roscoe) and long coriander (Eryngium foetidum L.) are commonly grown and used as important spices and medicinal plants in Vietnam. Our study recovered for the first time one of the most damaging tropical root-knot nematodes, Meloidogyne javanica, associated with these plants in the Western Highlands of Vietnam. In this study, M. javanica was characterized based on morphology and molecular characterization of D2-D3 fragment of 28S rRNA, ITS, and Nad5 mtDNA regions. The identification of this species was done based on a combination of morphology, multiplex-PCR with specific primer, network haplotype analysis, and PPNID program.

First report of Rotylenchulus reniformis infecting turmeric in Vietnam and consequent damage.

Turmeric (Curcuma longa L.) is one of the common medicinal crops of high economical value in Vietnam. A survey in the Central Highlands of Vietnam revealed a turmeric growing area showing serious disease symptoms, including stunting of the plant, yellowing or darkening of the leaf margins and tips, and underdeveloped dry and rotten rhizomes. An inspection for plant pathogens in soil samples from this area revealed a high density and frequency of Rotylenchus reniformis, with a significant relationship between the density of R. reniformis, rhizome weight, and level of plant damage. This study provides the first report of R. reniformis found in parasitic association with turmeric in Vietnam with the support of molecular data and examines its resulting damage.Turmeric (Curcuma longa L.) is one of the common medicinal crops of high economical value in Vietnam. A survey in the Central Highlands of Vietnam revealed a turmeric growing area showing serious disease symptoms, including stunting of the plant, yellowing or darkening of the leaf margins and tips, and underdeveloped dry and rotten rhizomes. An inspection for plant pathogens in soil samples from this area revealed a high density and frequency of Rotylenchus reniformis, with a significant relationship between the density of R. reniformis, rhizome weight, and level of plant damage. This study provides the first report of R. reniformis found in parasitic association with turmeric in Vietnam with the support of molecular data and examines its resulting damage.

First report of Hemicriconemoides litchi associated with Piper sarmentosum and revision of the genus Hemicriconemoides in Vietnam.

The taxonomic status of Hemicriconemoides litchi, H. mangiferae, and H. strictathecatus has been in debates for many years. In this study, a population of H. litchi collected from the rhizosphere of Piper sarmentosum is characterized using a combination of morphology and molecular data. Our results are in agreement with other authors to maintain the validity of H. litchi and provide sequences of H. litchi with correct names on GenBank. A revision on the genus Hemicriconemoides in Vietnam is also discussed.The taxonomic status of Hemicriconemoides litchi, H. mangiferae, and H. strictathecatus has been in debates for many years. In this study, a population of H. litchi collected from the rhizosphere of Piper sarmentosum is characterized using a combination of morphology and molecular data. Our results are in agreement with other authors to maintain the validity of H. litchi and provide sequences of H. litchi with correct names on GenBank. A revision on the genus Hemicriconemoides in Vietnam is also discussed.

First report of Xiphinema hunaniense Wang & Wu, 1992 (Nematoda: Longidoridae) in Vietnam.

For the first time, a survey of plant-parasitic nematodes in the Central Highlands of Vietnam discovered a population of Xiphinema hunaniense Wang & Wu, 1992. The Vietnamese population of X. hunaniense is characterized by having an offset lip region, lack of anterior genital branch, vagina directed backward, and a digitate tail. Morphological features and morphometrics of this population are in agreement with the type population of X. hunaniense except for some variations. In addition, molecular characterization of this population and phylogenetic tree of 28S rDNA sequences of the genus are also provided.For the first time, a survey of plant-parasitic nematodes in the Central Highlands of Vietnam discovered a population of Xiphinema hunaniense Wang & Wu, 1992. The Vietnamese population of X. hunaniense is characterized by having an offset lip region, lack of anterior genital branch, vagina directed backward, and a digitate tail. Morphological features and morphometrics of this population are in agreement with the type population of X. hunaniense except for some variations. In addition, molecular characterization of this population and phylogenetic tree of 28S rDNA sequences of the genus are also provided.

First report of Paratylenchus lepidus Raski, 1975 associated with green tea (Camellia sinensis (L.) Kuntze) in Vietnam.

The pin nematodes, Paratylenchus spp., are relatively small nematodes that can feed on a wide range of host plants. The morphological identification of this nematode is greatly hampered by their small size and variable characters. This study provides the first report of Paratylenchus lepidus from Vietnam with a combination of morphological and molecular characterizations. The 28S rDNA phylogenetic tree of the genus and the first COI mtDNA barcode of this species are also provided.

First Report of Scutellonema brachyurus (Steiner, 1938) Andrassy, 1958 and Occurrence of Meloidogyne incognita (Kofoid & White, 1919) Chitwood, 1949 in Belgium.

A study of plant-parasitic nematodes in the Botanical garden at Ghent University in Belgium revealed the presence of two tropical nematode species, i.e. Scutellonema brachyurus and Meloidogyne incognita. Scutellonema brachyurus was recovered, only once, for the first time in Belgium from Musa basjoo and is morphologically characterized. M. incognita, forming galls on Hedychium greenii, was recovered in all seasons over three consecutive years and is morphologically and molecularly characterized. Although no unequivocal evidence was found to indicate that these nematodes pose a current threat in Belgium, in the light of climate change, it is crucial to improve our knowledge of potential tropical nematode activity in more Northern countries.

First report of Longidorus mindanaoensis Coomans, De Ley, Jimenez and De Ley, 2012 (Nematoda: Longidoridae) From a Mangrove Forest in Vietnam.

Longidorus mindanaoensis was recovered from a mangrove forest in Vietnam. The recovered population is in general morphological agreement with the type population, and the characters of pharyngeal bulb, i.e. the same unique pattern of pharyngeal glands nuclei as well as the lip region morphology, amphidial fovea shape and size and position of vulva corroborated its identity. Molecular studies of the recovered population using D2-D3 expansion segments of large subunit ribosomal DNA (LSU rDNA D2-D3) revealed the D2-D3 sequence of recovered population is 99.6% similar to the sequence of the type population. A new morphometric range for body size was recorded for the species based upon present Vietnamese population, and present study emphasized the diversity of Longidorus spp. in Vietnam could be higher than previously assumed.

Description of Rotylenchus rhomboides n. sp. and a Belgian population of Rotylenchus buxophilus (Tylenchomorpha: Hoplolaimidae).

During a survey in the Botanical garden of Ghent University, a new species Rotylenchus rhomboides n. sp. and a population of Rotylenchus buxophilus were found. Rotylenchus rhomboides n. sp. is characterized by the presence of a rhomboid-like widening of the mid-ridge of lateral field at the level of vulva, a feature previously unknown within the genus. The population of the new species, composed only by females, has a rounded labial region with 4 to 5 annuli, robust stylet (31-37 µm long), short dorsal esophageal gland (9-19 µm) overlap of the intestine, vulva located slightly posterior to mid-body, and hemispherical or rounded tail shape with large phasmids located 3 to 5 annuli anterior to the level of anus. The hierarchical cluster analysis based on morphological features indicated that the new species closely resembles R. corsicus, R. gracilidens, and R. rugatocuticulatus. The DNA analyses of the D2-D3 of 28S rDNA, ITS rDNA, and COI mtDNA sequences of Rotylenchus rhomboides n. sp. show a close relationship with R. buxophilus, R. goodeyi, R. laurentinus, R. pumilus, and R. incultus, all of which can also be differentiated from the new species by morphological features. The combination of morphological, morphometric, and molecular characteristics confirmed the new species and the first report of R. buxophilus on yam (Dioscorea tokoro) in Belgium.During a survey in the Botanical garden of Ghent University, a new species Rotylenchus rhomboides n. sp. and a population of Rotylenchus buxophilus were found. Rotylenchus rhomboides n. sp. is characterized by the presence of a rhomboid-like widening of the mid-ridge of lateral field at the level of vulva, a feature previously unknown within the genus. The population of the new species, composed only by females, has a rounded labial region with 4 to 5 annuli, robust stylet (31­37 µm long), short dorsal esophageal gland (9­19 µm) overlap of the intestine, vulva located slightly posterior to mid-body, and hemispherical or rounded tail shape with large phasmids located 3 to 5 annuli anterior to the level of anus. The hierarchical cluster analysis based on morphological features indicated that the new species closely resembles R. corsicus, R. gracilidens, and R. rugatocuticulatus. The DNA analyses of the D2-D3 of 28S rDNA, ITS rDNA, and COI mtDNA sequences of Rotylenchus rhomboides n. sp. show a close relationship with R. buxophilus, R. goodeyi, R. laurentinus, R. pumilus, and R. incultus, all of which can also be differentiated from the new species by morphological features. The combination of morphological, morphometric, and molecular characteristics confirmed the new species and the first report of R. buxophilus on yam (Dioscorea tokoro) in Belgium.

First report of Mesocriconema sphaerocephalum (Taylor, 1936) Loof, 1989 associated with carrot (Daucus carota subsp. Stativus) in Vietnam.

Our study recorded the presence of Mesocriconema sphaerocephalum on carrot in Hanoi city and Hai Duong province in Vietnam. This species was identified by morphometric, morphological characterizations, and molecular characterization of D2D3 of 28S rDNA sequence. To our knowledge, this is the first report of M. sphaerocephalum on carrot in Vietnam.Our study recorded the presence of Mesocriconema sphaerocephalum on carrot in Hanoi city and Hai Duong province in Vietnam. This species was identified by morphometric, morphological characterizations, and molecular characterization of D2D3 of 28S rDNA sequence. To our knowledge, this is the first report of M. sphaerocephalum on carrot in Vietnam.

Description of Geocenamus vietnamensis sp. n. (Nematoda: Merliniidae) from Vietnam.

A new species of the genus Geocenamus was isolated from soil and root samples from the rhizosphere of Casuarina equisetifolia (Casuarinaceae) in Quang Nam province, Vietnam. This species is characterized by a round-to-hexagonal labial disc, the presence of a labial region, which is continuous or slightly offset from the body with six sectors, lateral sectors of first labial annulus being smaller than the submedian sectors, the presence of six to seven labial annules; the absence of deirids; stylet length 24 to 28 µâ€Šm long, body length 776 to 979 µâ€Šm long; lateral field with six to eight lateral lines, without areolation at mid-body and with areolation in outer bands at the tail region and a pointed tail terminus. Geocenamus vietnamensis n. sp. most closely resembles G. boghiae in having a non-sclerotized head framework and lacking a bursa in the males. It can be clearly distinguished from all other species of the genus Geocenamus by these characteristics. The combination of morphology, morphometric features, and phylogenetic trees, based on D2-D3 of 28S and ITS rDNA sequences, showed that this new species can be clearly separated from all other sequenced species. This record is the first for Geocenamus in Vietnam.A new species of the genus Geocenamus was isolated from soil and root samples from the rhizosphere of Casuarina equisetifolia (Casuarinaceae) in Quang Nam province, Vietnam. This species is characterized by a round-to-hexagonal labial disc, the presence of a labial region, which is continuous or slightly offset from the body with six sectors, lateral sectors of first labial annulus being smaller than the submedian sectors, the presence of six to seven labial annules; the absence of deirids; stylet length 24 to 28 µâ€Šm long, body length 776 to 979 µâ€Šm long; lateral field with six to eight lateral lines, without areolation at mid-body and with areolation in outer bands at the tail region and a pointed tail terminus. Geocenamus vietnamensis n. sp. most closely resembles G. boghiae in having a non-sclerotized head framework and lacking a bursa in the males. It can be clearly distinguished from all other species of the genus Geocenamus by these characteristics. The combination of morphology, morphometric features, and phylogenetic trees, based on D2­D3 of 28S and ITS rDNA sequences, showed that this new species can be clearly separated from all other sequenced species. This record is the first for Geocenamus in Vietnam.

Morphological and Molecular Characteristics of Pratylenchus haiduongensis sp. n., a New Species of Root-Lesion Nematodes Associated with Carrot in Vietnam.

Pratylenchus haiduongensis sp. n. is described as associated with carrot (Daucus carota subsp. sativus (Hoffm.) Schübl. & G. Martens) in Hai Duong Province, Vietnam. P. haiduongensis sp. n. is characterized by the lip region with three annuli and slightly separated from the body. Stylet knobs are rounded (never indented anteriorly). The lateral field includes four incisures, bearing areolation at the pharynx region and tail region and occasionally appears in the vulval region. Sometimes the appearances of oblique broken striaes divide the lateral field into five or six incisures. The ovary is distinct with one row of oocytes. Spermatheca is oval in shape with round central cavity, without sperm or reduced in some specimens. The postvuval uterine sac is long surpassing the vulva body diameter by 2 to 2.5 times (PUS = 31 to 65 µm). High vulva position with V = 66 to 75%. The tail shape can be subhemispherical with a smooth, slightly indented, broadly smooth, or cleft terminus observed in some specimens. The matrix code of P. haiduongensis sp. n. is: A2, B1, C4, D(1,3), E1, F(5,6), G(1,2), H(1,4); I(1,2,3,4), J1, K(1,2) according to Castillo and Vovlas (2007). The LSU-D2D3 segment and the ITS-rDNA region of this species were amplified and sequenced. The morphological characters and molecular phylogenetic analyses confirmed that this is a new species of the genus Pratylenchus in Vietnam.

Remarks on phylogeny and molecular variations of criconematid species (Nematoda: Criconematidae) with case studies from Vietnam.

The family Criconematidae is a remarkable group of nematodes, containing roughly 600 nominal root-ectoparasitic species, of which many species are known to be significant agricultural pests. Strikingly, our phylogenetic analyses based on 18S, D2-D3 of 28S rRNA, and COI mtDNA sequences of criconematid species, supported by tree topology tests (SH and AU tests), revealed that almost all studied genera, including Criconema, Ogma, Crossonema, Discocriconema, Hemicriconemoides, Criconemoides, Mesocriconema, and Lobocriconema, are not monophyletic groups, a finding that is partly contrary to those of previous studies on these groups. Our results suggest that key morphological characters used in the classification of Criconematidae are the consequence of convergent evolution. It is clear from our studies that the species status of at least 40 sequences of criconematid species from GenBank must be either revised or reconsidered, with analyses based on a polyphasic approach that includes different tree- and distance-based molecular species-delimitation methods (bPTP, GMYC, ABGD1, and ABGD2). Our studies found the ABGD2 output of the automatic barcode method to agree remarkably well with established species delimitations, while in general, the four species-delimitation results corresponding to three barcode regions forwarded significantly more putative species compared to those originally considered. This study also characterised for the first time the populations of Criconemoides myungsugae and Discocriconemella hensungica associated with Vietnamese ginseng, one of the most precious and rare ginseng varieties in the world. Although these populations are morphologically in agreement with the original descriptions of C. myungsugae and D. hengsungica, their molecular data display notable variations compared to the sequences deposited in GenBank. These species demonstrate clearly the immense molecular variations that can be observed in several species of the family Criconematidae.

Integrative taxonomy of Mesocriconema onoense (Tylenchida: Criconematidae) from Vietnam highly suggests the synonymization of Mesocriconema brevistylus and related species.

The genus Mesocriconema is one of the most diverse genera within the family Criconematidae, known as ring nematodes, with more than 90 species. Although species in this genus usually show distinct morphological characterizations, the identification based only on morphology can lead to misidentification in many studies resulted in a number of synonymizations in the genus over time. In this study, an integrated approach has been applied in characterizing Mesocriconema onoense from Vietnam. The molecular data of 28S rRNA, ITS, 18S rRNA regions were analyzed and discussed to confirm the correct names on GenBank. Besides, phylogenetic analyses of 28S rRNA, ITS, and 18S rRNA regions of Mesocriconema species revealed that Mesocriconema brevistylus should be considered as a junior synonym of M. onoense. Consequently, M. helicus, M. onostris, and M. paronostris should also be considered as the synonyms of M. onoense.