Characterization of Hoplolaimus seinhorsti and Hoplolaimus pararobustus (Tylenchina: Hoplolaimidae) from banana, with phylogeny and species delineation in the genus Hoplolaimus.

The morphological and molecular characterisations of two lance nematode species isolated from the rhizosphere of banana, Hoplolaimus seinhorsti and H. pararobustus, are provided based on an integrative study that includes light and scanning electron microscopy, phylogenetic analysis and two tree-based molecular species delimitation methods (GMYC and bPTP). Nineteen new sequences were obtained, including 5 partial 18S rRNA, 6 D2-D3 of 28S rRNA, 1 ITS rRNA and 7 COI mtDNA (the first COI sequences of H. seinhorsti and H. pararobustus), and an updated morphological character comparison of 37 Hoplolaimus species is presented. The tree-based molecular species-delimitation approaches employed gave markedly differing results, and also showed remarkable discrepancies among the investigated genes, although the bPTP output was found to agree well with established morphological species delimitations. Both species-delimitation approaches did, however, provide the same output for the COI mtDNA sequences, and the COI mtDNA gene sequence was also found to correspond better to established morphological species. It is therefore recommended by this paper as representing the most suitable barcode marker for Hoplolaimus species identification. This integrative study also resulted in the corrective reassignment of 17 gene sequences that were previously unidentified or incorrectly classified, as well as concluding that H. pararobustus consists of two cryptic species.

Morphological and molecular characterization of Pratylenchus species from Yam (Dioscorea spp.) in West Africa.

The root-lesion nematodes (RLN), Pratylenchus spp., are among the major plant-parasitic nematodes affecting yam (Dioscorea spp.) production in West Africa. The distribution and diversity of RLN species associated with yam was investigated through a soil and tuber survey of the main producing areas in Nigeria and Ghana. Pratylenchus spp. were detected in the yam rhizosphere in 59% of 81 soil samples from Ghana and 39% of 114 soil samples from Nigeria. Pratylenchus spp. were detected in 24 of 400 tubers examined, in combination with root-knot nematodes (Meloidogyne spp.) and their associated damage of galls and crazy roots (79%), and with yam nematode (Scutellonema bradys) and their associated damage of dry rot (17%), although no specific additional symptoms were observed for Pratylenchus spp. Species of Pratylenchus were identified by their morphological features and by sequences of the D2-D3 region of the 28 S rDNA gene and the mitochondrial cytochrome oxidase I gene (COI). Pratylenchus brachyurus was the most frequent RLN species in both the rhizosphere and tubers of yam. Pratylenchus hexincisus was recovered from one tuber collected in Nigeria. While further investigations are required to establish the host status of yam for this nematode, this appears to be the first record of P. hexincisus on yam. The present taxonomical status of P. scribneri and P. hexincisus is discussed.

Diversity of Root-knot Nematodes Associated with Tubers of Yam (Dioscorea spp.) Established Using Isozyme Analysis and Mitochondrial DNA-based Identification.

The root-knot nematodes (RKN), Meloidogyne spp., represent an important threat to yam (Dioscorea spp.) production in West Africa. With the aim to establish the diversity of RKN species affecting yam tubers, for control and resistance screening purposes, surveys were conducted in the main yam producing areas of Nigeria. Galled tubers (N = 48) were collected from farmers' stores and markets in nine states in Nigeria and in one district in Ghana. RKN isolated from yam tubers were identified using enzyme phenotyping (esterase and malate dehydrogenase) and mitochondrial DNA (mtDNA) NADH dehydrogenase subunit 5 (Nad5) barcoding. Examination of 48 populations revealed that yam tubers were infested by Meloidogyne incognita (69%), followed by M. javanica (13%), M. enterolobii (2%), and M. arenaria (2%). Most of the tubers sampled (86%) were infected by a single species, and multiple species of RKN were detected in 14% of the samples. Results of both identification methods revealed the same species, confirming their accuracy for the identification of these tropical RKN species. In addition to M. incognita, M. javanica, and M. enterolobii, we report for the first time M. arenaria infecting yam tubers in Nigeria. This finding extends the list of yam pests and calls for caution when developing practices for yam pest management.

An Improved Phenotyping Method for Evaluation of Yam (Dioscorea spp.) Resistance to Nematodes Belonging to the Genera Meloidogyne and Scutellonema.

Phenotyping yam (Dioscorea spp.) germplasm for resistance to parasitic nematodes is hampered by the lack of an efficient screening method. In this study, we developed a new method using rooted yam vine cuttings and yam plantlets generated from semi-autotrophic hydroponics (SAHs) propagation for phenotyping yam genotypes for nematode resistance. The method was evaluated using 26 genotypes of D. rotundata for their reaction to Scutellonema bradys and four root-knot nematode species, Meloidogyne arenaria, M. enterolobii, M. incognita, and M. javanica. Yam plantlets established in nursery bags filled with steam-sterilized soil were used for screening against single nematode species. Plants were inoculated four weeks after planting and assessed for nematode damage eight weeks later. A severity rating scale was used to classify genotypes as resistant, tolerant, or susceptible determine based on the nematode feeding damage on tubers and the rate of nematode multiplication in the roots of inoculated plants. The results demonstrated putative resistance and tolerance against S. bradys in 58% of the genotypes and 88%, 65%, 65%, and 58% against M. arenaria, M. javanica, M. incognita, and M. enterolobii, respectively. The method is rapid, flexible, and seasonally independent, permitting year-round screening under controlled conditions. This method increases the throughput and speed of phenotyping and improves the selection process.

Morphological and molecular characterisation of Scutellonema species from yam (Dioscorea spp.) and a key to the species of the genus.

The yam nematode, Scutellonema bradys, is a major threat to yam (Dioscorea spp.) production across yam-growing regions. In West Africa, this species cohabits with many morphologically similar congeners and, consequently, its accurate diagnosis is essential for control and for monitoring its movement. In the present study, 46 Scutellonema populations collected from yam rhizosphere and yam tubers in different agro-ecological zones in Ghana and Nigeria were characterised by their morphological features and by sequencing of the D2-D3 region of the 28S rDNA gene and the mitochondrial COI genes. Molecular phylogeny, molecular species delimitation and morphology revealed S. bradys, S. cavenessi, S. clathricaudatum and three undescribed species from yam rhizosphere. Only S. bradys was identified from yam tuber tissue, however. For barcoding and identifying Scutellonema spp., the most suitable marker used was the COI gene. Additionally, 99 new Scutellonema sequences were generated using populations obtained also from banana, carrot, maize and tomato, including the first for S. paralabiatum and S. clathricaudatum, enabling the development of a dichotomous key for identification of Scutellonema spp. The implications of these results are discussed.