Molecular characterization of a grapevine leafroll-associated virus 4 from Slovenian vineyards.

During a survey conducted in vineyards in Slovenia, variety of grapevine leafroll disease symptoms were observed. Mixed infection with grapevine leafroll-associated viruses 3 and 4 (GLRaV-3, -4) in two grapevines from a vineyard in south-western part of Slovenia was confirmed by DAS-ELISA in 2010. The 3'final 1769 nucleotides of the Slovenian GLRaV-4 isolate were assembled from amplicons obtained by IC RT-PCR. The complete coat protein (CP) and p23 gene sequences were compared with other GLRaV-4 sequences from GenBank. Results showed that CP and p23 amino acid sequences of Slovenian variant (055-SI) are 88% and 85%, respectively, identical to corresponding genes of reference sequence GLRaV-4 LR106 (GenBank Acc. No. FJ467503). Phylogenetic analyses show that Slovenian variant clusters together with other corresponding strains of GLRaV-4. The sequencing results show great variability of the N-terminal part of the CP sequence indicating that this part of the genome is not suitable for molecular detection of the virus. To our knowledge this is also the first report of GLRaV-4 in Slovenian vineyards.

First Report of Bursaphelenchus mucronatus kolymensis Associated with Pinus sylvestris in Serbia.

Bursaphelenchus mucronatus kolymensis (Korentchenko) Braasch et al. (1), also referred to as the European type of B. mucronatus, is morphologically very similar to the pine wood nematode (PWN) B. xylophilus (Steiner & Buhrer) Nickle, the causal agent of pine wilt disease. The main morphological difference between the two species is the shape of the female tail mucro (3); however, some populations of PWN vary in mucro shape, which can lead to misidentification. Since PWN was found and identified for the first time in Portugal in 1999 (4), concern about the spread of PWN to pines and other conifers all over Europe has increased. Therefore, the PWN survey in every European country is essential. In 2011, the Serbian Ministry of Agriculture, Forestry and Water Management implemented phytosanitary measures for detection of PWN in the territory of the Republic of Serbia. In September 2013, wood samples were collected from a 40-year-old wilted Pinus sylvestris located on Divcibare Mountain, western Serbia. Nematodes were extracted by a modified Baermann funnel method. Nematode specimens (10 females and 10 males) were characterized by an Aphelenchoid-type esophagus, female vulva flap, mucro on the end of female tail, and males with paired arcuate spicules. Morphological identification matched that of B. mucronatus kolymensis (1). The specific morphometric evidence for this European type subspecies of B. mucronatus was sub-cylindrical female tail with mucro 3 to 4 µm long, digitate, well offset from tail (5). Species identification was confirmed by PCR-RFLP of the rDNA ITS region using one female and one male nematode separately (2). The sizes of restricted DNA fragments as determined with agarose gel and capillary electrophoresis were in compliance with reported data in the literature (2,4) and support the identification of the nematode as B. mucronatus kolymensis. To our knowledge, this is the first report of any species belonging to Bursaphelenchus genus in Serbia. The presence of B. mucronatus kolymensis in Serbia indicates the possibility of establishment and spread of PWN. Therefore, the measures taken in case of PWN occurrence should be prepared and training foresters to identify pine wilt disease symptoms should be encouraged in Serbia. References: (1) H. Braasch et al. J. Nematode Morphol. Syst. 14:77, 2011. (2) W. Burgermeister et al. Nematology 11:649, 2009. (3) Y. Mamiya and N. Enda. Nematologica 25:353, 1979. (4) M. M. Mota et al. Nematology 1:727, 1999. (5) M. M. Mota and P. Vieira. Page 146 in: Pine Wilt Disease: A Worldwide Threat to Forest Ecosystems. Springer Science and Business Media, 2008.

First Report of the Root-Knot Nematode Meloidogyne ethiopica on Tomato and Cucumber in Turkey.

The root-knot nematode Meloidogyne ethiopica Whitehead, mainly reported from African countries, was first described in 1968 in Tanzania (4). It was further detected in South America (Brazil, Chile, and Peru) (2). In 2004, M. ethiopica was recorded for the first time in Europe on tomato (3) and later in field soil samples from maize (Zea mays L.) and kiwi [Actinidia deliciosa (A. Chev.) C. F. Liang & A. R. Ferguson] collected in the area of Kavalla in North Greece (1). M. ethiopica was included on the EPPO alert list of harmful organisms in 2011. In summer 2009, severe stunting, leaf wilting, and extensive root galling of a presumed root-knot nematode (Meloidogyne sp.) were detected during a routine survey for root nematodes attacking tomato (Solanum lycopersicum L.) crops from two greenhouses in the campus of University of Ondokuz Mayis and attacking cucumber (Cucumis sativus L.) crops from commercial greenhouses in Çarsamba district of Samsun Province, Turkey. Perineal patterns of Meloidogyne sp. females collected from infested roots were variable, with moderately high to high dorsal arch, lateral line mostly indistinct and striae usually wavy, which is very similar to that seen in M. incognita Kofoid & White. Such variability among perineal patterns precluded its accurate identification. For further species identification, protein extracted from young egg-laying females were resolved in 3% stacking (pH 6.8) and 7% separating (pH 8.8) polyacrylamide gel with Tris-glycine buffer. The gels were stained with α-naphthyl acetate substrate for esterase activity (1). The esterase phenotypes exhibited a three banded pattern, E3, that was identical to M. ethiopica. Therefore, esterase studies were repeated including nematodes from M. ethiopica culture from Slovenia used as a reference. In addition, small subunit rDNA sequence analyses were performed to complete species identification. DNA sequences of a 1.6-kb rDNA fragment were generated using two sets of primers (1) and deposited in the NCBI GenBank with the accession number KC551945. The sequence was compared to the corresponding fragments of M. ethiopica and closely related species from the NCBI GenBank. DNA sequence of M. ethiopica from Turkey exhibited the highest identity of 99.8% to the sequence FJ559408 of M. ethiopica from Slovenia. Sequences KC551945, FJ559408, AY942630, and JQ768373 of M. ethiopica from Turkey, Slovenia, Brazil, and Greece, respectively, clustered together and formed a separate clade using phylogenetic analyses. This species may pose a threat for vegetable production in Turkey since it has a wide host range infesting numerous higher plants including monocotyledons, dicotyledons, herbaceous, and woody plants. Furthermore, M. ethiopica may have been present in Turkey for many years because correct identification based only on perineal pattern is difficult. Thus, misidentification might have been possible. The distribution of this nematode in more extensive vegetable fields should be determined. To our knowledge, this is the first report of M. ethiopica in Turkey and the third finding of this species in Europe. References: (1) I. L. Conceição et al. Eur. J. Plant Pathol. 134:451, 2012. (2) S. N. Murga-Gutierrez et al. Nematropica. 42:57, 2012. (3) S. Sirca et al. Plant Disease. 88:680, 2004. (4) A. G. Whitehead. Nematology. 15:315, 1969.

First Report of the Pale Potato Cyst Nematode Globodera pallida from Slovenia.

The potato cyst nematodes (PCN) Globodera rostochiensis (Woll.) Behrens and G. pallida (Stone) Behrens are considered the most important nematode threat to potato production worldwide, and they are subject to strict quarantine regulations in many countries. The first report of the PCN in Slovenia dates back to 1971, when a single cyst of G. rostochiensis was detected (3). In the last decade, G. rostochiensis was detected several times, mainly in the central and northern parts of the country (2). Interceptions of G. pallida in imported consignments of ware potato were made several times, but had not been detected in soil in Slovenia. Therefore, the country was declared as a protected zone for G. pallida in the European Union by the national authorities in 2003. During the official PCN systematic survey in autumn 2011, the pale potato cyst nematode, G. pallida, was found in a soil sample. Two viable cysts were extracted and the nematode species was identified. The posterior part of the cysts containing eggs and juveniles were used for morphometrical analysis, while the anterior parts were used for DNA extraction and molecular analyses. The ribosomal internal transcribed spacer (ITS) region was amplified using ITS5 and PITSp4 primers and detected in real-time PCR using ABI7500 (Applied Biosystems, Life Technologies, Carlsbad, CA,) (1). Also, the ribosomal DNA region that extends from the 3' end of the 18S ribosomal subunit and includes all of ITS1, 5.8S, and ITS2, to the 5' end of the 28S ribosomal subunit, was used to generate a DNA sequence. The sequence obtained was compared with those from several Globodera species, revealing unequivocal similarity to G. pallida. The infested soil sample originated from a ware potato field near Ivancna Gorica, central Slovenia. Subsequently, an additional 69 samples were taken from the surroundings and viable G. pallida cysts were found in another five samples taken from two neighboring fields (one of grassland and the other of clover). Three fields, totaling 1.9 ha, were declared as G. pallida-infested. The eradication of the pest will take place by enforcing strict phytosanitary measures. Ware potatoes originating from areas where the pests occur is considered to be the most probable pathway for the introduction of G. pallida in Slovenia. A ware potato processing facility is situated in very close proximity to the infested fields. The waste waters from potato tuber washing were discharging onto the grassland, never used for potato or other field crop production in which the G. pallida infestation was found. The facility processes imported ware potato from several European and non-European countries. This case demonstrates that ware potato may pose a serious risk for the introduction of such pests, and should be therefore subjected to more intensive phytosanitary inspection. References: (1) J. Bacic et al. Russ. J. Nematol. 16:63, 2008. (2) S. Sirca et al. Phytopathol. Mediterr. 49:361, 2010. (3) G. Urek et al. Nematology 5:391, 2003.

Monitoring Diabrotica v. virgifera (Col.: Chrysomelidae) in southeastern Slovenia: increasing population trend and host spectrum expansion.

Ever since the western corn rootworm (WCR) (Diabrotica virgifera virgifera), an alien invasive species from North America, has been introduced into Europe on at least 3 separate occasions, it spread within 15 years over the entire area of south-eastern and central Europe (except Denmark). Until quite recently, Zea mays L. was the only known host plant whereas in North America WCR also attacks members of the plant family Cucurbitaceae. In August of 2006, we were able to validate these findings also in the Old World by observing WCR visiting blossoms of oil pumpkin (Cucurbita pepo L.). Beside this first report of WCR on this regionally and economically important crop, a population increase in Gaberje near Lendava, Eastern Slovenia, was observed. Some future consequences of multiple hosts for integrated pest management (IPM) of WCR are being discussed.

Xiphinema rivesi from Slovania Transmit Tobacco ringspot virus and Tomato ringspot virus to Cucumber Bait Plants.

The dagger nematode, Xiphinema rivesi Dalmasso, a member of the X. americanum group, was detected in 2002 for the first time in Slovenia and for the fourth time in Europe (4). X. rivesi is a vector of at least four North American nepoviruses including Cherry rasp leaf virus (CRLV), Tobacco ringspot virus (TRSV), Tomato ringspot virus (ToRSV), and Peach rosette mosaic virus (PRMV) (1,2). All of these viruses are included on the EPPO and EU lists of quarantine organisms, but none of the Xiphinema species found in Europe have been reported to transmit these nepoviruses. Three virus isolates, including TRSV (from Lobelia spp.; virus collection of the Plant Protection Service, Wageningen, The Netherlands), ToRSV (grapevine isolate PV-0381; DSMZ, Braunschweig, Germany), and Arabis mosaic virus (ArMV) (from Vinca spp.; virus collection of the Plant Protection Service), were used in transmission tests with a population of X. rivesi found in Slovenia. X. rivesi is not known to transmit ArMV and this virus was included as a check. The nematodes were extracted from peach orchard soil collected near the village of Dornberk, and transmission tests fulfilled the set of criteria proposed by Trudgill et al. (3). Cucumis sativus cv. Eva, grown in a growth chamber at 25°C, was used as acquisition hosts and transmission bait plants. The acquisition hosts were mechanically inoculated and showing systemic symptoms before the introduction of nematodes. Noninoculated acquisition plants were included as controls. After a 10-day acquisition feeding period, the nematodes were transferred to healthy bait plants and allowed a 14-day inoculation feeding period. X. rivesi transmitted TRSV and ToRSV but not ArMV. TRSV and ToRSV bait plants developed systemic symptoms 4 to 6 weeks after the nematodes were transferred. Transmission of TRSV and ToRSV was confirmed by testing leaf and root sap of bait plants in a double antibody sandwich (DAS)-ELISA. High virus concentrations were detected in the roots and leaves of TRSV and ToRSV symptomatic plants. DAS-ELISA on bait plants from nematodes that had been allowed to feed on ArMV-infected or the virus-free control acquisition plants gave negative results. No symptoms appeared on bait plants used for ArMV transmission or the control bait plants. To our knowledge, this is the first report of transmission of TRSV and ToRSV with a Xiphinema population from Europe. References: (1) D. J. F. Brown et al. Phytopathology 84:646, 1994. (2) L.W. Stobbs et al. Plant Dis. 80:105, 1996. (3) D. L. Trudgill et al. Rev. Nematol. 6:133, 1983. (4) G. Urek et al. Plant Dis. 87:100, 2002.

The Incidence of Root-Knot Nematodes Meloidogyne arenaria, M. incognita, and M. javanica on Vegetables and Weeds in Montenegro.

Root-knot nematodes (Meloidogyne spp.) are common pathogens that parasitize vegetables and other crops and cause significant yield reductions worldwide. In the early spring of 2006, severe plant stunting, chlorosis, and extensive root galling were observed on cucumber plants grown in a greenhouse on Zeta plain, Zetska ravnica, Montenegro. In the summer and autumn of 2006, infected roots of different crops were collected from greenhouses and vegetable production fields of Zeta plain, which represents the largest area of Montenegro's vegetable production. Several vegetable crops were found to be infected with root-knot nematodes, including tomato (Lycopersicon esculentum Mill.), squash (Cucurbita pepo L.), cucumber (Cucumis sativus L.), pepper (Capsicum annuum L.), and lettuce (Lactuca sativa L.). Symptoms on these crops included root-galling, leaf chlorosis, and stunting. Heavily infected tomato plants growing in two greenhouses also displayed early flower and fruit drop. Nematode species were determined based on characterization of (i) female perineal patterns, (ii) male and second-stage juvenile morphology (2,3), and (iii) esterase and malate dehydrogenase phenotypes (PhastSystem; Amersham Biosciences, Piscataway, NJ) from young egg-laying females (1). The most prevalent species was M. incognita, which was isolated from the roots of tomatoes, peppers, cucumbers, and lettuce from nine locations. Meloidogyne arenaria was detected at three locations from the roots of tomatoes grown in a greenhouse and the weeds Convolvulus arvensis L. and Solanum nigrum L., which were growing in open fields in separate locations. M. javanica was found on tomato and squash in the same field where M. arenaria was also found on S. nigrum. M. javanica was isolated from tomato and squash. In this study, we found high incidence of Meloidogyne spp. in intensive vegetable production areas of Montenegro. The implementation of an effective integrated pest management program is essential for future use of infested areas. To our knowledge, this is the first report of M. arenaria, M. incognita, and M. javanica from Montenegro. References: (1) P. R. Esbenshade and A. C. Triantaphyllou. J. Nematol. 17:6, 1985. (2) S. B. Jepson. Identification of Root-Knot Nematodes. CAB International, Wallingford, UK, 1987. (3) G. Karssen. The Plant-Parasitic Nematode Genus Meloidogyne Göldi, 1892 (Tylenchida) in Europe. Koninklijke Brill NV, Leiden, the Netherlands, 2002.

Diabrotica virgifera virgifera in eastern Slovenia: increasing population trend 2005.

The first invasion of the western corn rootworm (WCR), Diabrotica virgifera virgifera LeConte (Coleoptera: Chrysomelidae) into Slovenia was recorded in 2003. A few beetles were found in the western province of Slovenia near Italy. More beetles were detected in the eastern district of Prekmurje bordering Austria, Hungary and Croatia. While the western population remained the same in 2005, the eastern population increased considerably. By means of the very sensitive Metcalf sticky trap baited with the female sex pheromone, early infestations could be recorded. At corn fields, Zea mays L., near the villages of Dolina, Gornji Lakos and Pince, Diabrotica flights were observed in 2005 which had their maxima during the first week of August and yielded, respectively, 57, 112, and 45 beetles/trap per 3-4 day trapping interval. The increase from 2004 to 2005 in this eastern most district of Slovenia is noteworthy. Generally in the western and central part of Slovenia, most maize is grown in rotation, typically every second year. It is estimated that about 10 % of the surface of maize is grown as monoculture, typically on farms with intensive cattle production. In eastern Slovenia, a higher proportion of maize is grown in monoculture (about 30 % in the 3-year period from 2002-2004); this percentage decreased in 2005 due to official requirements by the Slovenian Phytosanitarian Administration.

The western corn rootworm diabrotica Virgifera virgifera en route to Germany.

The western corn rootworm Diabrotica virgifera virgifera LeConte (Col.:Chrysomelidae) (D.v.v.) is one of the most important maize pests in North America. Ever since its invasion into Europe and its detection near Belgrade airport by BACA in 1993 it quickly spread all over southeastern Europe and is now advancing towards central Europe. Up until summer 2004 considered free of D.v.v., Germany is, with the exception of its northern and northeastern borders, surrounded by countries with proven D.v.v. infestations. In addition to simultaneous spot introductions by airplanes, three main routes for terrestrial introduction into Germany are likely: 1. from south to north via Lombardy (Italy) through Switzerland to the State of Baden-Wuerttemberg in the southwest; 2. from south east to northwest via Croatia, Slovenia, Austria into the State of Bavaria; and 3. from Belgium and the Netherlands in southeasterly direction to the state of Northrhine-Westfalia. From these, progress of D.v.v. along route 1 is so far the most advanced. It follows the well established network of road and rail connections through Switzerland and underscores the active role mankind and its technology plays as an active distribution vector for D.v.v. Mandatory crop rotation in the Swiss Canton of Ticino did slow down but could not prevent the northbound advance of D.v.v. in 2004. Considering the recent discovery of D.v.vu near the South German border, its introduction into German territory is only a matter of time and may be ecologically unavoidable. In Slovenla, another relatively small southern transit state, the D.v.v. population density is still much lower than in Switzerland but with significantly increasing trend during 2004 and with special emphasis in its southeastern provinces. Considering its relatively short distance to southeastern Bavaria and the well developed transalpine rail, road and tunnel system, Slovenia as a transit state may provide another access route for D.v.v. of lesser but still significant importance to Germany.

Monitoring presence and advance of the alien invasive western corn rootworm beetle in eastern Slovenia with highly sensitive Metcalf traps.

The American Chrysomelid beetle Diabrotica virgifera virgifera LeConte (D.v.v.), also called the western corn rootworm, spread from the location of its original introduction into Europe, Belgrade airport (BACA 1993), in all directions. Within a decade it occupied almost all countries of South-eastern and Central Europe. However, it reached Slovenia as late as 2003: Only 19 specimen were found in maize fields of the eastern and also western provinces. Already in the summer of 2004, their number had risen to 386 which were mainly found in the eastern provinces near the borders to Croatia, Austria and Hungary. For their monitoring, a simple trap is being described which can be acquired in high numbers at a very low price and can guaranty a most sensitive detection of beetles. The Metcalf cup trap which in Slovenia so far has been unknown performed the task of monitoring quite well at five locations. It will also facilitate the future search for new and increasing infestations. Comparing 2004 with the year 2003, D.v.v. expanded its range in eastern Slovenia by about 15 km. These new infestations will include territories in which fields of the regionally Important oil seed pumpkin Cucurbita pepo are located. With important traffic connections between South-eastern and Central Europe, Slovenia will occupy a bridgehead function in the preventive protection of maize from D.v.v. spreading into more northerly European regions including western Austria and southern Germany.

First Report of the East-Asian Type of Bursaphelenchus mucronatus in Pinus sylvestris in Slovenia.

Bursaphelenchus mucronatus Mamiya & Enda, a nematode associated with aboveground tissues of pine trees, is very similar morphologically to the pine wood nematode (PWN), B. xylophilus, a devastating pest that has caused widespread losses to pine woods in some parts of Asia since the early 1900s. During 1999, B. xylophilus was identified for the first time in Europe in Portugal (3), leading to concern that pines and other conifers all over Europe might be endangered. To protect forests from PWN and other pests, the European Union and other countries now regulate the importation of all coniferous chips, sawn wood, and logs and also have organized surveys to determine the extend of PWN infestation. Slovenia has approximately 1,227,832 ha of forest that makes up more than 60% of the surface of the country. The percentage of conifers within forests is 47.9% and comprised mainly of Picea, Pinus, Abies, and Larix species (2). During the summers of 2002-2004, approximately 100 ha of conifer forests in Slovenia were surveyed for the presence of PWN. In total, 170 wood samples from dead and dying pine (Pinus sylvestris L., P. nigra Arnold), spruce (Picea abies L.), and fir (Abies alba Miller) were analyzed. B. xylophilus (Steiner & Buhrer) Nickle was not found in the area, but a closely related nematode, B. mucronatus, previously shown to be pathogenic on conifer seedlings (4), was isolated from samples of P. sylvestris. Of 34 P. sylvestris samples analyzed, B. mucronatus was found in three samples. Samples were taken from a conifer forest located in the central part of Slovenia near the international airport of Ljubljana. The species was identified morphologically and also characterized by polymerase chain reaction restriction (PCR)-fragment length polymorphism (RFLP) analysis on the basis of ITS-rDNA fragment patterns. According to RFLP patterns (1), the analyzed species was recognized as the East-Asian type of B. mucronatus. Beside these findings, the specimens of B. homanni (Braasch) and Bursaphelenchus sp. have also been found in Slovenia. The results of the survey show that PWN was not found in Slovenia, but three different species of the Bursaphelenchus genus were present. References: (1) H. Braasch et al. Nachrbl. Dtsch. Pflanzenschutzd. 51:312, 1999. (2) M. Jurc et al. Zb. Gozdarstva in Lesarstva 72:121, 2004. (3) M. M. Mota et al. Nematology 1:727, 1999. (4) J. R. Sutherland. Pine wilt nematode. Pages 19-20 in: Compendium of Conifer Diseases. E. M. Hansen and K. J. Lewis, eds. The American Phytopathological Society, St. Paul, MN. 1997.

First Report of the Root-Knot Nematode Meloidogyne ethiopica on Tomato in Slovenia.

The root-knot nematode Meloidogyne ethiopica Whitehead originally described from Tanzania is also distributed in South Africa, Zimbabwe, and Ethiopia (3). Although this species is a relatively unknown root-knot nematode, M. ethiopica parasitizes several economical important crops, such as tomato, cowpea, bean, cabbage, pepper, pumpkin, tobacco, lettuce, and soybean (4). This species can be morphologically confused with M. incognita (Kofoid & White), and therefore, is probably easily overlooked (1). Recently, characteristic esterase isozyme patterns were described for this species, which provide a more reliable identification (2). In October 2003, Lycopersicum esculentum cv. Belle plants with large root-galls were observed in a greenhouse at Dornberk, Slovenia, which were identified as an unknown root-knot nematode within the M. incognita group. Subsequent sampling revealed infestation of all tomato plants within this greenhouse with obvious aboveground symptoms of stunting and wilting. Additional morphological and isozyme studies identified this root-knot nematode as M. ethiopica. The identification was based on female, male, and second-stage juvenile morphology and malate dehydrogenase and esterase isozyme phenotypes from young egg-laying females isolated from 20 tomato plants and compared with a culture of M. ethiopica from Africa. To our knowledge, this is not only the first report of M. ethiopica in Slovenia, but also the first report of this species in Europe. References: (1) A. M. Golden. Fundam. Appl. Nematol. 15:189, 1992. (2) W. Mandefro and K. Dagne. Afr. J. Plant Prot. 10:39, 2000. (3) A. G. Whitehead. Trans. Zool. Soc. Lond. 31:263, 1968. (4) A. G. Whitehead. Nematologica 15:315, 1969.

First Report of the Dagger Nematode Xiphinema rivesi, a Member of the X. americanum Group, from Slovenia.

The dagger nematode, Xiphinema rivesi Dalmasso, is a virus-transmitting plant-parasitic nematode described from France (2) and also reported from Germany, Portugal, and Spain. While this species is only known from a few local sites within Europe, it is widespread in North America and also present in Peru and Pakistan (3). X. rivesi is able to transmit at least four North American nepoviruses, including Cherry rasp leaf virus, Tobacco ringspot virus, Tomato ringspot virus, and Peach rosette mosaic virus (1,4). In early spring 2002, nematodes of the X. americanum group were detected in soil samples taken near the roots of peach plants from an orchard in Bilje near Nova Gorica, Slovenia. Resampling in June and September at the same site and at an adjacent, approximately 30-year-old peach orchard confirmed the presence of the nematode in relatively high numbers (5 per 100 ml of soil). Adult females were isolated and sent for further identification to the Plant Protection Service in Wageningen, the Netherlands. The nematode was morphologically identified as X. rivesi and confirmed at the rDNA-level with internal transcribed spacer restriction fragment length polymorphism analysis. Because both sampling sites where the nematode was found are close to the Vipava River in an area of extensive stone and pome fruit production, it is likely that the nematode is also present in other orchards in this region. The origin of this quarantine nematode is unknown as there is no direct link with Slovenian orchards and import of plant material from abroad in the last decades. To our knowledge, this is the first report of X. rivesi in Slovenia and the fourth report of this species in Europe. References: (1) D. J. F. Brown et al. Phytopathology 84:646, 1994. (2) A. Dalmasso. Mem. Mus. Hist. Nat. Nouv. Série A., Zool. 61:33, 1969. (3) Data Sheets on Quarantine Pests: Xiphinema americanum sensu lato. Page 629 in: Quarantine Pest for Europe, CABI, Wallingford, UK, 1997. (4) L. W. Stobbs et al. Plant Dis. 80:105, 1996.

Occurrence of the Root-Knot Nematodes Meloidogyne incognita and M. hapla in Slovenia.

The root-knot nematode Meloidogyne incognita (Kofoid & White) Chitwood is one of the most harmful species within the genus Meloidogyne. It has been detected regularly in greenhouses throughout Europe, while in the field it is restricted to the southern parts of Europe (4). M. incognita is able to parasitize more than 700 plant species and varieties, including the majority of our economically important crops (1). It has been recorded in most parts of the world, including former Yugoslavia; however, there is no data about its presence in Slovenia. In late summer of 2002, large galls were observed on roots of pepper, Capsicum annuum L., grown in a greenhouse located in Portoroz near the Adriatic Coast of Slovenia. Different root-knot nematode stages, i.e., males, females, and second-stage juveniles, were isolated and morphologically identified as M. incognita (2). Additional isozyme gel electrophoresis confirmed this (3). To our knowledge, this is the first report of M. incognita from Slovenia. In addition, M. hapla Chitwood was found for the first time in a field in Slovenia. Previously, it had been detected on different host plants, all restricted to greenhouses. In October 2002, M. hapla was isolated and identified from relatively small galls of sweet pepper, C. annuum L., grown in a field near Ljubljana, located in central Slovenia. Additional resampling of the soil comfirmed the presence of M. hapla in this field. References: (1) J. D. Eisenback. Root-Knot Nematode Taxonomic Database. CD-ROM, CABI, Wallingford, UK, 1998. (2) J. D. Eisenback and H. Hirschmann Triantaphyllou. Root-knot nematodes: Meloidogyne species and races. Pages 191-274 in: Manual of Agricultural Nematology. W. R. Nickle, ed. Marcel Dekkers, Inc., New York, 1991. (3) P. R. Esbenshade and A. C. Triantaphyllou. J. Nematol. 17:6, 1985. (4) G. Karssen. Revision of the European root-knot nematodes III on mono- and dicotyledons. Pages 119-121 in: The Plant-Parasitic Nematode Genus Meloidogyne Göldi, 1892 (Tylenchida) in Europe. Koninklijke Brill NV, Leiden, the Netherlands, 2002.