Pseudaphelenchus zhoushanensis n. sp. was isolated from a dead Pinus thunbergii at Changgang Mountain, Zhoushan Islands, Zhejiang Province, China. It is characterised by the small to medium length body, cuticle slightly annulated, presence of three lateral lines, stylet 9.0-10.7 μm with small but conspicuous basal knobs, excretory pore located from same level as the metacorpus to slightly anterior to metacorpus, true bursa surrounding entire tail but inconspicuous, male tail conical with a single mucron, spicule with distinct condylus and rostrum strongly arcuate to a pointed end, female tail conical with annulation, strongly ventrally bent in distal part of tail, with terminus bluntly pointed or finely mucronate. Phylogenetic analyses using sequences of the18S and 28S D2-D3 regions of rDNA confirmed the status of P. zhoushanensis n. sp. as a new species. Combining the molecular phylogenetic analyses, morphology and biology of P. zhoushanensis n. sp. and Tylaphelenchus jiaae indicates that T. jiaae is a member of Pseudaphelenchus to which it is herein transferred as P. jiaae n. comb. (= T. jiaae).
A short insert genomic library for Xiphinema index, the natural vector of
Grapevine Fanleaf Virus, was constructed from degenerate oligonucleotide
primed PCR (DOP-PCR) products. The genomic library was screened for (CA)n
microsatellites. Screening of 6200 colonies and comparison of the sequencing
results revealed seven (CA)n containing microsatellites, coded here as
XIMSL1, XIMSL2, XIMSL3, XIMSL4, XIMSL5, XIMSL6 and XIMSS1. XIMSL prefixed
microsatellites were followed by the motif of the same long interspersed
element. Microsatellite XIMSS1 has some similarity to the short interspersed
element. Except for XIMSL1, all microsatellites were proven to be effective
diagnostic tools at species level. Genetic diversity between and within
populations was also evaluated for each microsatellite.
Bursaphelenchus decraemerae n. sp., isolated in Ningbo, China, from packaging wood made of Pinus sp. imported from the USA, is described. It is characterised by the lateral field with three lines, a stylet 10-12 μm long with small basal swellings, excretory pore located 4-12 μm posterior to median bulb, spicule small, ventrally arcuate with lamina gradually tapering to distal tip, rostrum triangular pointed to conical, condylus elongated with bluntly rounded tip, and cucullus minute, barely visible. There are seven caudal papillae arranged as a single ventral precloacal papilla, one adcloacal pair and two postcloacal pairs. The bursal flap is narrow and oval. A small vulval flap is present in lateral view of the female and the tail is conical, strongly ventrally curved with the terminus finely pointed and sometimes appearing like a mucron. The new species is characterised by morphological and molecular analysis and is close to B. gerberae and B. sakishimanus.
Ruehmaphelenchus quercophilus n. sp., isolated from a dying oak, Quercus robur, in the State Forests, Łopuchówko District, Poland, is described and figured. It is characterised by three lateral lines, relatively stout body, excretory pore posterior to nerve ring, five caudal papillae, spicules relatively small (12-14 μm) with weakly developed condylus and rostrum, bursa absent, vulva positioned at ca 78% of body length, vulval flap absent, post-uterine branch extending for slightly over half of vulva to anus distance, and conoid tail with a bend at the distal end forming a terminal projection 10-14 μm long. The new species is very similar to four other described species, namely R. formosanus, R. juliae, R. thailandae, and R. sirisus. Detailed phylogenetic analysis based on 18S, 28S D2-D3 and ITS sequences has confirmed the status of this nematode as a new species.
Bursaphelenchus parapinasteri is redescribed morphologically and with new molecular characterisation. It was isolated from a dead Pinus thunbergii on Changgang Mountain, Zhoushan Islands, Zhejiang Province, China. Detailed morphology of the spicule, female gonad, hemizonid position, arrangement of male caudal papillae, and female tail terminus shape are documented. The ITS-RFLP patterns and the ITS1/2, partial 28S D2-D3 and partial 18S gene sequences were characterised. The phylogenetic analyses revealed that B. parapinasteri belongs to the hofmanni-group sensu Braasch and is close to B. anamurius, B. hofmanni, B. mazandaranense, B. paracorneolus, B. pinasteri and B. ulmophilus. A diagnostic key to species of the hofmanni-group is presented.
Two Sheraphelenchus populations were isolated from pine wood packaging from Italy and onion bulbs from South Korea at Ningbo Port, China, respectively. The former, on the basis of morphology and molecular sequencing, is described as S. parabrevigulonis n. sp., which is characterised by a stylet length of 11 μm, three lines in the lateral field, excretory pore at level with, or slightly posterior to, nerve ring, V = 93.1, and spicules with long condylus and membrane-like rostrum in a shallow triangular shape. The species from onion, also on the basis of morphology and sequence data, was identified as S. sucus, which is characterised by a stylet length of 13 μm (erroneously stated to be ca 6 μm in the type description), excretory pore at level of, or posterior to, median bulb, and V = 80.9. The phylogeny trees based on rDNA 18S, 28S D2-D3 and ITS1/2 sequences revealed that S. parabrevigulonis n. sp. and S. sucus clustered into a clear clade and grouped with Bursaphelenchus species. The study enriched the taxonomic information of Sheraphelenchus and confirmed its status as an independent genus.
A serine protease gene Evsp was cloned from the nematophagous fungus Esteya vermicola with strong virulence against Bursaphelenchus xylophilus. The full-length cDNA of Evsp contains 2280 nucleotides with a 1656 bp ORF encoding a protein with 551 amino acids. The genomic Evsp includes two exons (396 bp and 1260 bp) separated by an intron (207 bp). There is only one copy of Evsp gene in the fungal genome. The deduced amino acids sequences of Evsp showed highly homology with the catalytic domains in subtilisin serine proteases. Phylogenetic analyses based on the protein sequences revealed that E. vermicola is separated from nematode-trapping fungi but close to other nematophagous and entomopathogenic fungi. The recombinant serine protease rEvsp was induced in Escherichia coli with expression vector pET28a(+). The tests of protease and nematicidal activities for the purified and refolded rEvsp indicated it is possibly involved in the fungal infection process against B. xylophilus.
Bursaphelenchus parantoniae n. sp. isolated in Ningbo, China, from packaging wood made of Pinus sp. imported from Belgium, is described. It is characterised by a slim body (a = 32.6-51), lateral field with two lines, excretory pore located ca one metacorpal length posterior to median bulb, spicules stout and arcuate with triangular rostrum and flattened cucullus, and seven caudal papillae arranged as a single midventral precloacal papilla, one adcloacal pair and two postcloacal pairs. The vagina is inclined anteriorly, a vulval flap is present, the post-uterine branch extends for more than half of the vulva-anus distance, and the female tail is conical with a hyaline region and bluntly rounded terminus. Phylogenetic analyses showed that B. parantoniae n. sp. clusters with the hylobianum-group and appears close to B. antoniae, B. chengi, B. hylobianum and B. niphades. The new species is distinguished from these species by morphology, ITS-RFLP patterns and molecular characters.
Bursaphelenchus chengi sp. n. is described and illustrated. Dauer juveniles
were isolated from imported wood packaging materials from Taiwan to Nanjing
Port, China. Bursaphelenchus chengi sp. n. was reared and maintained on
Petri dish cultures of the fungus Botrytis cinerea. The new species is
characterised by the medium body size in both sexes, the presence of only
two incisures in the lateral field and the robust and strongly curved
spicules. The spicule lamina is angular distally, the rostrum digitate and
the condylus rounded. The tail is arcuate with a pointed terminus. The bursa
is usually truncate with the posterior margin indented in some specimens or
rounded with a fine axial point. Females have a small vulval flap formed by
a short extension of the cuticle of the anterior lip, and a conical tail
that gradually tapers to an almost straight or slightly recurved, pointed or
rounded terminus. Because of the presence of two lateral lines, similar
spicule shape, tapering female tail and the presence of a small vulval flap,
B. chengi sp. n. should be grouped in the abietinus-group sensu Braasch.
together with B. abietinus, B. antoniae, B. hellenicus, B. hylobianum and B.
rainulfi. ITS-RFLP profiles support the proposal of the new species, and
phylogenetic analysis of the 28S rDNA D2/D3 domain sequence places it close
to B. antoniae and other species of the abietinus-group.
Bursaphelenchus nematodes, extracted from 12 samples taken from imported
packaging wood originating from seven countries and regions at Nanjing,
China, during 2003 and 2004, were identified using morphology, morphometric
data and ITS-RFLP patterns. Two samples were identified as B. xylophilus:
one, from Canada, was the M-form and another, from Japan, the R-form. Five
samples were identified as B. mucronatus, three belonging to the European
genotype and two to the East-Asian genotype. Two samples (from Germany and
South Korea) were recognised as B. rainulfi. Two samples (South Korea and
Japan) were identified as B. doui and a sample from Hong Kong was identified
as B. thailandae. Additional morphometric data are provided for the
identified species, especially with respect to spicule morphometrics. The
CDA of morphometric data discriminated the five species by nine male and
eight female characters. The characters used for this analysis corresponded
partly to the main taxonomic-informative characters for the genus
Bursaphelenchus, viz., body length, stylet length, spicule length, a, c and
c′ values. The phylogenetic trees constructed from the D2D3 sequence data
supported the division of East Asian and European genotype groups of B.
mucronatus. Bursaphelenchus doui grouped with B. xylophilus and B.
mucronatus, all members of the xylophilus-group, which was separated from B.
rainulfi and B. thailandae.