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Renato N. Inserra, Jason D. Stanley, Alberto Troccoli, John Chitambar and Sergei A. Subbotin

Hemicaloosia vagisclera n. sp. is described from Bermuda grass (Cynodon dactylon) in Florida. This new species is characterised by females with body slightly ventrally arcuate in death, face with a prominent elliptical oral disc, large amphids, slender stylet with mean length 64 μm, lateral field consisting of a single longitudinal line marked by continuous and discontinuous transverse striae, oval and full spermatheca, sclerotised vagina vera and tail annuli width greater than that of remaining body annuli. Diagnostic characters for the males are a C-shaped body, head with 4-5 faint lip annuli and pronounced oval oral disc, lateral field with two longitudinal lines intersected by transverse striae, tail digitate in the distal portion posterior to bursa, distinctly annulated and with a round terminus. Molecular characterisation of H. vagisclera n. sp. using the D2-D3 domain of 28S rRNA, partial 18S rRNA and ITS rRNA gene sequences is also provided. The phylogenetic relationships of this species with other representatives of the suborder Criconematina are presented and indicate that H. vagisclera n. sp. has sister relationships with Caloosia longicaudata supporting the classification of Caloosia together with Hemicaloosia as separate genera in the family Caloosiidae. A diagnostic PCR-ITS-RFLP profile for H. vagisclera n. sp. is also given together with an identification key for seven known species of Hemicaloosia.

Esther Van den Berg, Louwrens R. Tiedt, Jason D. Stanley, Renato N. Inserra and Sergei A. Subbotin

The genus Scutellonema contains more than 40 species of spiral nematodes with enlarged phasmids called scutella. In this study, we provide morphological and molecular characterisation of S. clavicaudatum sp. n., S. brachyurus, S. bradys, S. cavenessi, S. transvaalense, S. truncatum and Scutellonema sp. A. from North and Central America, and Africa. The new species, S. clavicaudatum sp. n., was found on sugarcane in South Africa and is characterised by a lack of lip annuli as in S. africanum, S. siamense and S. truncatum. The lip region, in both males and females, is conical and marked by six large rectangular blocks separated or fused with the submedian and lateral lip sectors, which surround a round and distinct labial disc. Females of this new species also have large vaginal glands, a functional spermatheca, the lateral field posterior to the scutellum ending in a bluntly pointed shape and a clavate tail. Morphological descriptions, measurements, light and scanning electron microscopic photos and drawings are also given for S. bradys, S. cavenessi, S. transvaalense and S. truncatum. The study of spiral nematode samples from Florida, USA, confirmed the presence of a morphologically and genetically atypical populations of S. bradys. The morphology of the S. bradys population from Bermuda grass in pasture land from central Florida fits that of type specimens of this species, but differs in having a truncate tail terminus rather than round and also a prominent spermatheca filled with flagellate spermatozoa. Other Florida Scutellonema samples analysed in this study belonged to S. cavenessi, a species native to West Africa. This is the first report of S. cavenessi in Florida, where it parasitises the ornamental plant Sansevieria trifasciata. Our study showed a high level of intraspecific variation for Scutellonema rRNA and mtDNA genes, which can reach 5.6% for the D2-D3 of 28S rRNA, 12.9% for the ITS rRNA genes and 14.4% for the COI gene. Phylogenetic relationships within Scutellonema are given as inferred from the analyses of the D2-D3 of 28S rRNA, ITS rRNA and the COI mtDNA gene sequences.

Alberto Troccoli, Sergei A. Subbotin, Jason D. Stanley, Brian Alford, Nicola Vovlas and Renato N. Inserra

Morphological and molecular analyses of three populations of Meloidoderita whittoni (Sledge & Christie, 1962) comb. n. (syn. Sphaeronema whittoni; Tumiota whittoni) collected in Florida from sweet gum (Liquidambar styraciflua L.) indicated that this species is a representative of Meloidoderita rather than Sphaeronema, where it was included in the original description, or Tumiota, to where it was subsequently transferred. However, this species can be considered an atypical representative of the genus, because it differs from the other species of this genus in having females retaining the eggs inside a thin-walled uterus, which remains encased inside their body. After egg deposition inside the uterus, the female dies and its body is transformed into a persistent tanned sac with a thick cuticle, devoid of ornamentations, which protects the eggs like a heteroderid cyst. The female secretes from the vulva a gelatinous matrix, which becomes hardened in time and encases its body for protection against predation and parasitism by biological antagonists as has been suggested for other tylenchuloid nematodes. No egg deposition outside the female body was observed. Second-stage juveniles of this species have a semi-endoparasitic, rather than endoparasitic, habit as in other known Meloidoderita species. This is the first report of a sphaeronematid having a cyst stage fitting the definition of a heteroderid cyst. Phylogenetic relationships between some species of Tylenchuloidea and Criconematina are analysed using the partial 18S rRNA, the D2-D3 of 28S rRNA and the ITS rRNA gene sequences.

Zahra Tanha Maafi, Majid Amani, Jason D. Stanley, Renato N. Inserra, Esther Van den Berg and Sergei A. Subbotin

During a survey conducted on banana plantations in Sistan and Blouchestan province, south-east Iran, a new species of Tylenchulus was extracted from the soil and roots of banana plants. This species, named Tylenchulus musicola sp. n., is characterised by mature females having a swollen, hook-shaped body with a conical and elongate post-vulval portion ending in a round terminus, males having a weak stylet and a cylindrical and thick tail ending in a bluntly rounded and smooth terminus, and by second-stage juveniles having a slender body and a posterior body portion ending in a finely pointed or mucronate terminus. The results of glasshouse host tests indicated that the new species does not parasitise sugarcane ratoons or sour orange seedlings. Tylenchulus musicola sp. n. is distinguished from other known Tylenchulus species by the sequences of D2-D3 expansion segments of 28S rRNA and ITS rRNA genes. Phylogenetic relationships within Tylenchulus were reconstructed based on rRNA gene sequences using Bayesian inference. Diagnostic PCR-ITS-RFLP profiles are presented for T. musicola sp. n., T. furcus, T. graminis, T. palustris, T. semipenetrans and Trophotylenchulus floridensis. PCR with species-specific primers and genus-specific primer are tested and developed for rapid identification of five Tylenchulus species. An identification key to Tylenchulus species is provided.

Esther Van den Berg, Louwrens R. Tiedt, Renato N. Inserra, Jason D. Stanley, Nicola Vovlas, Juan E. Palomares Rius, Pablo Castillo and Sergei A. Subbotin

Sheathoid nematodes of the genus Hemicriconemoides are migratory root-ectoparasites of many plants including various agricultural crops and fruit trees. They are generally found inhabiting warm areas of the world and presently consist of 52 valid species. In this study we provide morphological and molecular characterisation of 12 species of this genus viz.: H. alexis, H. brachyurus, H. californianus, H. chitwoodi, H. macrodorus, H. minutus, H. ortonwilliamsi, H. promissus, H. silvaticus, H. strictathecatus, H. wessoni and Hemicriconemoides sp. originating from China, Greece, Japan, Myanmar, Spain, South Africa and the USA. Morphological descriptions, measurements, light and scanning electron microscopic observations and drawings are given for several species. Phylogenetic relationships within Hemicriconemoides, as inferred from the analyses of the D2-D3 of 28S rRNA and ITS-rRNA gene sequences, resulted in trees with three major clades that corresponded with species groupings based on morphology of the lip pattern and vulval flap. PCR with species-specific primers were developed for H. californianus, H. chitwoodi and H. strictathecatus.

Sergei A. Subbotin, Jason D. Stanley, Antoon T. Ploeg, Zahra Tanha Maafi, Emmanuel A. Tzortzakakis, John J. Chitambar, Juan E. Palomares-Rius, Pablo Castillo and Renato N. Inserra

Needle nematode populations of Longidorus orientalis associated with date palm, Phoenix dactylifera, and detected during nematode surveys conducted in Arizona, California and Florida, USA, were characterised morphologically and molecularly. The nematode species most likely arrived in California a century ago with propagative date palms from the Middle East and eventually spread to Florida on ornamental date palms that were shipped from Arizona and California. This is the first validated continental record of this needle nematode species in the USA and the Americas. The USA populations of L. orientalis contained a small number of males that were not reported in the original description and are herein described. Longidorus orientalis was able to survive for at least 4 years at very low numbers in the warm and humid environment of Florida on date palms imported from California and Arizona. Association of L. orientalis with L. africanus was observed in all of the surveyed sites, indicating that date palm is a host of both nematodes. Phylogenetic relationships of L. orientalis with closely related Longidorus species, in addition to relationships between populations of L. orientalis from the USA, Greece, Iran and Spain, were inferred from the analyses of D2-D3 of 28S rRNA, ITS1 rRNA and partial coxI gene sequences. The PCR-D2-D3 expansion segments of 28S rDNA-RFLP diagnostic profile is provided. Longidorus orientalis populations display a high level of intraspecific variation (up to 15.5%) in coxI mtDNA sequences. Analysis of phylogenetic relationships of nematode populations revealed incongruence of the ITS1 rRNA and coxI mtDNA gene trees, which might be the result of selective introgression of mtDNA through gene flow between previously isolated populations introduced simultaneously into new geographical regions.

Esther Van den Berg, Louwrens R. Tiedt, Renato N. Inserra, Jason D. Stanley, Nicola Vovlas, Juan E. Palomares-Rius, Pablo Castillo and Sergei A. Subbotin

The results of morphological and molecular analyses of a Florida topotype and other populations of Hemicriconemoides strictathecatus showed that this sheathoid nematode consists of two morphotypes, both with an average stylet length of more than 70 μm, but having different tail termini, bluntly pointed or rounded. These findings confirmed the morphological similarity of H. strictathecatus with H. mangiferae, which was considered a junior synonym of this species as previously proposed by Decraemer & Geraert (1992, 1996). Populations of a sheathoid nematode with a stylet length ranging from 62.5 to 72.0 μm from Taiwan, China, South Africa and Venezuela and identified in previous studies as H. strictathecatus were found to be morphologically and molecularly different from this species and are now considered as representatives of H. litchi. Another sheathoid nematode population from Florida, considered to be H. mangiferae by McSorley et al. (1980), was also found to be morphologically and molecularly congruous with H. litchi. During nematological surveys in Florida, a new sheathoid nematode was detected on date palms imported from California into Florida and is described herein as H. phoenicis sp. n. This new species is related morphologically to the H. strictathecatus morphotype with pointed tail terminus. Both have a stylet longer than 70 μm. The new species is phylogenetically related to H. strictathecatus and H. litchi. It differs morphologically from other Hemicriconemoides species by the cuticular ornamentation of the annuli, which are marked by coarse longitudinal ridges, grooves and thick margins. Morphological and molecular characterisations of H. cocophillus from Mozambique and Florida, USA are also elucidated in this study. New phylogenies of the genus Hemicriconemoides as inferred from the analyses of the ITS rRNA, D2-D3 of 28S rRNA and partial coxI gene sequences are provided.

Esther Van den Berg, Louwrens R. Tiedt, Gracia Liébanas, John J. Chitambar, Jason D. Stanley, Renato N. Inserra, Pablo Castillo and Sergei A. Subbotin


Hemicycliophora presently contains 132 valid species of sheath nematodes. Within several samples obtained from surveys in Canada, South Africa, Spain and the USA, we distinguished six valid and six putative unidentified species by integrating the results of morphological and molecular analyses. Valid species included: H. californica, H. gracilis, H. parvana, H. poranga, H. raskii, and H. signata. The putative unidentified species were indicated as Hemicycliophora sp. 10, sp. 12, sp. 15, sp. 16, sp. 17, and sp. 18. Two new species of sheath nematodes from Spain and the USA were described and named as H. onubensis sp. n. and H. robbinsi sp. n., respectively. Hemicycliophora wyei is proposed as a junior synonym of H. parvana and H. ripa is proposed as a junior synonym of H. poranga. Eighteen valid and 13 unidentified species of sheath nematodes were characterised using the partial COI mtDNA gene. A total of 94 new sequences of which 77 were for the COI mtDNA gene were obtained in this study. Phylogenetic relationships within Hemicycliophora, using the D2-D3 expansion segments of 28S rDNA, ITS rRNA and COI gene sequences, are presented as inferred from Bayesian analysis.

Alberto Troccoli, Sergei A. Subbotin, John J. Chitambar, Toon Janssen, Lieven Waeyenberge, Jason D. Stanley, Larry W. Duncan, Paula Agudelo, Gladis E. Múnera Uribe, Javier Franco and Renato N. Inserra

Amphimictic populations of root-lesion nematodes with numerous males and females having three lip annuli, a functional spermatheca and non-areolated lateral field occur on sword fern (Nephrolepis exaltata) in Florida. Identified for decades as Pratylenchus penetrans, they appeared to be a morphologically separated species on the basis of a longer stylet (17.8-18.3 μm) than P. penetrans (15-17 μm) and different lip pattern in en face view (rectangular vs dumb-bell in P. penetrans). Morphologically similar amphimictic root-lesion nematodes have also been detected on flax lily in Costa Rica. Subsequent morphological observations indicated that these amphimictic root-lesion nematodes from fern and flax lily are closely related to the parthenogenetic species P. bolivianus, which has areolated lateral fields. In spite of the reproductive and morphological dissimilarities between these populations, their separation into different species was not supported by the results of molecular analyses of their DNA sequences. The populations used in these analyses included those that are amphimictic from Florida and Costa Rica and others that are parthenogenetic from the type locality in Bolivia, and geographically distant localities in Chile, China, Colombia and Europe. Phylogenetic analyses of the ITS and D2-D3 expansion segments of the 28S rRNA gene indicated that they belong to the same species, P. bolivianus, which consists of two morphotypes, P. bolivianus (am) amphimictic and P. bolivianus (pm) parthenogenetic, herein described and illustrated. Contradictory results were obtained by the analyses using a portion of the hsp90 gene. The phylogenetic study, which included sequences of other root-lesion nematodes, a topotype and geographical distant populations of P. zeae, revealed that P. bolivianus and P. zeae formed highly supported clades in the majority consensus trees. PCR with species-specific primers for rapid diagnostics of P. bolivianus and P. zeae were developed and tested.