A new phoretic association: Bursaphelenchus minutus (Nematoda: Parasitaphelenchidae) and Orthotomicus erosus (Coleoptera: Scolitydae) recorded on Pinus pinea (L.)

A new phoretic association between Bursaphelenchus minutus and the bark beetle Orthotomicus erosus collected on Pinus pinea is described for the first time. Nematode identification was based on morphological observation and biometric measurements of males and females. Moreover, B. minutus was characterized by ITS sequences and RFLP profile.     

First report of Bursaphelenchus mucronatus kolymensis (Nematoda: Aphelenchoididae) on Monochamus sutor (Coleoptera: Cerambycidae) in Spain

This is the first report of Monochamus sutor as vector of B. mucronatus kolymensis, the European type of B. mucronatus (synonymous with B. kolymensis) in Spain. In 2013, a female of Monochamus sutor was caught in a Pinus sylvestris forest. Nematode species identification was based on morphological characteristics and on molecular and phylogenetic analysis of dauer juveniles and adults.     

First detection of Bursaphelenchus mucronatus (Nematoda: Aphelenchoididae) on Monochamus sutor (Coleoptera: Cerambycidae) in Romania

As a result of the detection of the pinewood nematode Bursaphelenchus xylophilus in Portugal, and its subsequent spread to Spain, intense surveys were conducted to screen for the presence of Bursaphelenchus species in Romania. Herein, we report recent surveys of insects potentially vectoring Bursaphelenchus species collected using trap trees or pheromone‐baited traps placed in the forest. Trap felled spruce trees (Picea abies) and pheromone‐baited traps were installed in six different counties in Romania (Bra?ov, Sibiu, Suceava, Hunedoara, Timi? and Dâmbovi?a). Ten different species of insects distributed among Curculionidae and Cerambycidae were obtained. Nematodes were extracted from insects and observed to validate the presence of Bursaphelenchus specimens. One female identified as Monochamus sutor was the only specimen carrying nematodes in the genus Bursaphelenchus. Nematodes were identified as B. mucronatus based on morphological and molecular features. This is the first detection and report of natural spread of B. mucronatus in Romania. The absence of B. xylophilus was confirmed in the areas of Romania surveyed in this work.     

The virulence of phytopathogenic fungi associated with the bark beetles Tomicus piniperda and Orthotomicus erosus in Tunisia

Phytopathogenic fungi associated with the bark beetles Tomicus piniperda and Orthotomicus erosus were isolated in various pine forests of Tunisia. Tomicus piniperda and its galleries yielded Leptographium wingfieldii, Ophiostoma minus, and Ophiostoma ips. Ophiostoma minus was the most frequent species associated with T. piniperda, in both the attacking and the emerging beetles. It was collected from most investigated forests, whereas O. ips and L. wingfieldii were obtained only from forests located in Central and Northern Tunisia. Frequencies of association with T. piniperda were always low, reaching 11.1% only once, for O. ips. Ophiostoma ips was the only blue stain fungus associated with O. erosus and its galleries. It was found in all the localities, but at a low and variable frequency, exceeding 15% very rarely. The virulence of 16 fungal isolates was tested by single inoculations into Pinus halepensis (Aleppo pine) at two localities. Differences were detected among species; L. wingfieldii was the most virulent and O. minus the least virulent species in terms of phloem reaction zone formation and fungal growth in the phloem. In a separate experiment, mass inoculations (400 and 800 inoculations per m² of bole on 1‐m high belts) were performed with two isolates of L. wingfieldii on Aleppo pine and on Pinus brutia (Brutia pine). Three months later, measurements of sapwood status (% of conductive transversal section) and of its specific hydraulic conductivity, as well as of the phloem reaction zone lengths, did not show any isolate or density effect. At these experimental sites, Brutia pines appeared significantly more susceptible than Aleppo pines.     

Bursaphelenchus fungivorus from Pinus pinaster bark in Portugal

Bursaphelenchus fungivorus is reported for the first time in Portugal, identified as associated with Pinus pinaster bark and characterized on the basis of morphological and morphometrical characters for this species. Species identification was confirmed using restriction fragment length polymorphism analysis and sequencing of the internal transcribed spacer (ITS) regions of ribosomal DNA. Intraisolate genetic variability was detected among ITS sequences of the Portuguese B. fungivorus isolate. Phylogenetic analysis, obtained from multiple sequence alignment between ITS sequences of Bursaphelenchus species, revealed that the Portuguese B. fungivorus isolate clusters with other B. fungivorus isolates, forming a separate group close to B. seani highlighting a molecular proximity of these two species.     

First report on Bursaphelenchus sexdentati (Nematoda: Aphelenchoididae) in Israel

This is the first report and characterization of Bursaphelenchus sexdentati and the first official report of the presence of the genus Bursaphelenchus in Israel. This species was isolated from Orthotomicus erosus (Coleoptera: Scolytinae) found on Pinus halepensis in the Judean foothills in Israel. Nematodes collected from insect galleries were reared on fungal cultures and identified based on morphological diagnostic characters for the genus Bursaphelenchus. Sequencing analyses of the Internal Transcribed Spacer (ITS) region, small subunit (SSU) and large subunit (LSU) of ribosomal DNA confirmed the identification of this nematode species collected from wood and directly from the insect body.     

On the genus Bursaphelenchus Fuchs, 1937 (Nematoda: Parasitaphelenchinae) associated with wood and insects from declining forest trees in the Czech Republic

An overview of the genus Bursaphelenchus in the Czech Republic is presented, based on a recent survey for monitoring the presence of the pinewood nematode, Bursaphelenchus xylophilus, as well as on previous reports of this genus in the country. In addition, we provide a morphological and molecular characterization of four Bursaphelenchus species (B. eremus, B. pinophilus, B. vallesianus and B. borealis) found during the monitoring programme for forest pests, conducted during 2006–2010, within the Moravian and Bohemian regions. Nematodes were extracted from over 1917 insects and 1493 wood samples collected from deciduous and coniferous trees exhibiting wilting and declining symptoms. Bursaphelenchus species were found only in 0.73% of insects and 0.47% of the total number of wood samples. Bursaphelenchus borealis and B. pinophilus dauer juveniles were found associated with the insect vectors Dryocetes autographus and Pityogenes bidentatus, respectively. While a total of seven Bursaphelenchus species are now reported from the Czech Republic, the status of B. xylophilus remains as absent.     

On the track of Bursaphelenchus pinophilus Brzeski and Baujard, 1997 (Nematoda: Aphelenchoididae) in Portugal

This is the first report and characterization of Bursaphelenchus pinophilus in Portugal. This species was isolated from a young dying Pinus pinaster tree located in Valverde, in the Alentejo region. Nematodes were identified using several morphological diagnostic characters for this species (male spicule structure, number of lateral incisures, number and distribution of the male papillae, presence of female vulval flap and female tail shape) and confirmed using RFLP analysis of the internal transcribed spacer (ITS) regions of ribosomal DNA.     

松材线虫病死木皮下的南京隐翅虫(鞘翅目:隐翅虫科)

【目的】南京隐翅虫是在受松材线虫病危害的病死木皮下发现的隐翅虫新种,与小蠹、天牛等多种次期性害虫的时间和空间生态位跟随关系密切,具有作为媒介昆虫防治蛀干害虫的潜力。研究南京隐翅虫不同发育阶段个体的形态特征、成虫的食性及昼夜活动节律,能够为进一步的研究和应用提供参考。【方法】室内观察南京隐翅虫不同发育阶段个体的形态特征,通过人工饲养并结合口器解剖研究南京隐翅虫成虫的食性;应用"陷阱法"观察试虫的昼夜活动节律。【结果】南京隐翅虫幼虫为寡足型,口器咀嚼式,胸足发达,蛹为裸蛹,成虫体长1.5～2.0 mm,身体狭长平行,头、中后胸、腹部6～8节颜色较深,身体其他部分浅黄色。成虫喜食人为杀死的跳虫成虫、蝇科幼龄幼虫及天牛幼虫,对于小蠹科、象甲科、叩甲科、蚁科、蠼螋科的幼虫尸体表现出取食行为,试验过程中并未观察到捕食其他生物活体。室内南京隐翅虫成虫昼夜活动节律高峰期分别是8:00—11:00以及20:00—22:00。【结论】南京隐翅虫成虫为尸食性,其昼夜活动节律受温度影响较大。     

Bursaphelenchus tusciae in Tunisia associated with Hylurgus ligniperda

Bursaphelenchus tusciae is reported for the first time in Tunisia and North Africa, associated with the insect Hylurgus ligniperda Fabricius (Coleoptera: Curculionidae: Scolytinae). Nematode identification was based on restriction fragment length polymorphism analysis and sequencing of the internal transcribed spacer (ITS) regions of ribosomal DNA. Phylogenetic analysis revealed that Tunisian B. tusciae clusters together with two other B. tusciae isolates forming a separate group close to B. hildegardae and B. eggersi. As H. ligniperda is among maritime pine scolytids pests in Tunisia and is widely distributed in North Africa, this study is an important contribution to the knowledge of Bursaphelenchus species associated with bark beetles of pine forests in Tunisia and North Africa.     

Records of Bursaphelenchus spp. (Nematoda,Parasitaphelenchidae) in coniferous timber imported from the Asian part of Russia

Coniferous wood imported from the Asian part of Russia was surveyed in Germany (Mukran ferry terminal, highway and railway border station in Frankfurt/Oder) and Austria (railway in Marchegg, Retz and Wr. Neustadt, Lower Austria). The consignments consisted of mixed timber of Pinus/Picea or Pinus/Larix. Out of 625 samples investigated, 51 samples (8.5%) yielded eight Bursaphelenchus species. Bursaphelenchus mucronatus was found 42 times in Pinus, Picea and Larix wood, Bursaphelenchus hylobianum and Bursaphelenchus fraudulentus were detected twice, in Pinus/Larix and in Larix, respectively. The following species were each found once: Bursaphelenchus leoni in mixed timber of Pinus/Picea, Bursaphelenchus ‘borealis’ in Pinus, Bursaphelenchus hellenicus and Bursaphelenchus paracorneolus in Larix. One sample of Larix wood from Krasnoyarsk region contained a few specimens corresponding to Bursaphelenchus xylophilus. Bursaphelenchus mucronatus was present in about 30% of the samples showing signs of insect attack. A Monochamus species was found in a sample from Irkutsk. Most of the B. mucronatus isolates found belonged to the European genotype, whereas the East Asian genotype was found in three instances. This is the first report of B. fraudulentus, B. hellenicus, B. leoni, B. paracorneolus and the East Asian genotype of B. mucronatus in Russia. Bursaphelenchus hylobianum is the only species found in Russian wood and not in Europe so far. The species were identified morphologically and by internal transcribed spacer (ITS)‐restriction fragment length polymorphism (RFLP) technique. Species‐specific ITS‐RFLP patterns were established for B. hylobianum. In the case of the isolate morphologically corresponding to B. xylophilus, DNA extraction from the available low number of specimens failed to yield sufficient rDNA for ITS‐RFLP analysis.     

Occurrence of <Emphasis Type="Italic">Bursaphelenchus mucronatus</Emphasis> (Nematoda; Aphelenchoididae) in France and association with <Emphasis Type="Italic">Monochamus galloprovincialis</Emphasis> (Coleoptera: Cerambycidae)

As a consequence of the recent introduction of the pine wood nematode Bursaphelenchus xylophilus in Portugal, nematodes of the genus Bursaphelenchus were looked for in various French pine forests, in trees attacked by Monochamus galloprovincialis, the vector insect of B. xylophilus, and in the insects themselves. Trap trees were felled in 12 localities distributed all over the country. Nematodes were extracted from transversal stem discs; insects emerging from the trap trees were studied. B. hellenicus, B. leoni, B. mucronatus and B. sexdentati were isolated, but not B. xylophilus. The presence of B. mucronatus and the absence of B. xylophilus were confirmed by molecular markers. B. mucronatus was isolated from several regions with an average prevalence of 19%. The infestation of M. galloprovincialis by B. mucronatus reached 26.7%. The wide distribution of B. mucronatus in France could have an effect on the extension of B. xylophilus in a case of an introduction.     

New morphometric ranges for Bursaphelenchus willibaldi Schönfeld et al. 2006 (Nematoda: Parasitaphelenchidae) from Iran

Bursaphelenchus willibaldi, associated with trees in two separate locations in forests of northern Iran, are characterized and illustrated based on morphological, morphometric and molecular data. The Iranian population of B. willibaldi has a body length of 392–595 μm, stylet length of 12–14 μm, c‐index of 9.0–14.5 and PUS length of 51–82 μm. Males have spicules 15–18 μm long along the arch line. The Iranian population showed morphometric variations compared with the originally described samples. For example, it has a shorter body in females and males, greater c‐index, slightly greater range of V, slightly smaller range of PUS and tail length. Molecular phylogenetic analysis of the recovered populations revealed both sequenced isolates forming a clade with one European isolate of the species using Bayesian inference (BI) analysis with full Bayesian posterior probability (BPP).     

Parasitoids associated with Monochamus galloprovincialis (Oliv.) (Coleoptera: Cerambycidae) within the pine wilt nematode-affected zone in Portugal

Until recently, the pine sawyer Monochamus galloprovincialis (Olivier) (Coleoptera; Cerambycidae) was considered a secondary forest pest in Portugal. Its pest status changed when it was found to be the main vector of the recently introduced pine wood nematode (PWN) Bursaphelenchus xylophilus (Steiner and Bührer) Nickle (Nematoda; Aphelenchoididae). A survey in three locations within the PWN-affected zone was conducted in order to identify parasitoids associated with immature stages of the beetle and to evaluate their importance. No parasitoid was found associated with the insect eggs whereas 10% of the larvae were found parasitised by three Braconidae. Cyanopterus flavator Fabricius was the most numerous species; Iphiaulax impostor (Scopoli) and Coeloides sordidator Ratzeburg were the other species present. A review of the parasitoids associated with Monochamus in Europe, eastern Asia and North America was made, and the possibility of using the local species in future biological control programmes in Portugal is briefly discussed.     

Blue stain fungi associated withIps cembrae (Coleoptera: Scolytidae) in Japanese larch in connection with their brood development

The association between blue stain fungi andIps cembrae Heer (Coleoptera: Scolytidae) was investigated in the Japanese larch,Larix leptolepis Gordon, in the Nagoya University Forest, Aichi Prefecture, central Japan.I. cembrae had one or two generation(s) in a year in this study area. Two blue stain fungi,Ophiostoma piceae andLeptographium sp., were isolated from the body surface of both male adults in mating chambers and female adults in parent galleries, suggesting that this beetle species was a vector of these fungi. Although no blue stain fungi were isolated from non-stained wood, both fungi were isolated from the mating chambers, the center and the uppermost end of the galleries throughout the season. The fact thatO. piceae was consistently isolated with high frequency from adults and from their galleries strongly suggested that this species would be the principal blue stain fungus infecting the beetle-attacked larch trees.     

New reports of Bursaphelenchus species associated with conifer trees in Romania

The 2013 annual monitoring programme for the pinewood nematode, Bursaphelenchus xylophilus, analysed a total of 267 wood samples collected from declining or symptomatic coniferous trees distributed among national forests, gardens, public parks, distribution centres, wood‐processing industries, as well as 104 samples collected from wood packing material originated from several other countries. From a total of eight species found, five have been previously reported from Romania, while three other species (Bursaphelenchus abietinus, Bursaphelenchus fraudulentus and Bursaphelenchus fuchsi) represent new findings for Romania. In addition, B. mucronatus was found in packing wood originated from Russia. Herein, we provide a morphological, morphometric and molecular characterization for the new species found for Romania. Bursaphelenchus xylophilus was not detected.     

Blue stain fungi associated withTomicus piniperda (Coleoptera: Scolytidae) on Japanese red pine

A number of various species of blue-stain fungi were isolated fromTomicus piniperda adults at various stages of development, as well as from the galleries, pupal chambers and sapwood underneath galleries on Japanese red pine. This study was an attempt to identify the species, composition of blue-stain fungi associated withT. piniperda, the frequency of occurrence of the fungi, and their role in the sapwood-staining of Japanese red pine in Tsukuba City, central Japan. Among the seven species of blue-stain fungi isolated, an undescribed species ofOphiostoma together withO. minus were the dominant species and closely associated withT. piniperda. These two species occurred on newly emerging adults more frequently than the overwintered adults.Hormonema dematioides was also associated with the beetle, however, its frequency of occurrence from the emerged new adults was very low. Although the two other species,O. ips andGraphium sp. were also isolated from emerged beetles, the frequency of these fungi from gallery systems suggested that they were accidentally carried byT. piniperda. Leptographium wingfieldii, known to be associated with the beetle in Europe, was also isolated at a very low frequency and the fungus seemed not to be closely associated with the beetle.Ophiostoma sp. andO. minus appear to be the most important causes of blue-stain of Japanese red pine sapwood after infestation byT. piniperda.     

松材线虫的低温冷冻保存

研究-80℃下松材线虫的冷冻保存方法,比较不同冷冻保护剂对松材线虫低温保存的保护效果,以及解冻后松材线虫的繁殖情况。结果表明:15%甘油对松材线虫保护效果最好,解冻后存活率达36.5%,其次为10%甘油、20%甘油、25%甘油,保护效果最差的是1%二甲基亚砜(DMSO);解冻后线虫能够正常生长繁殖,冻存3天和冻存30天的线虫存活率和繁殖无显著性差异。