Report on Second Annual Balkan Week of Plant Health /  Abstracts of Presentation 
The Meeting  lasted from 31st of May until 1st of June and was attended by 87 participants from 7 different countries, who presented 15 oral contributions and participated in three discussion rounds.

Abstracts

1. V. Todorov
The Director General of the Bulgarian National Service for Plant Protection Dr. Ventsislav Todorov opened the Second Balkan Week of Plant Health with a presentation regarding the challenges facing Plant Protection in Bulgaria as a full member of the EU.  In the introduction Dr. Todorov elucidated the structure and the activities of NSPP. A major accent was put on the work that was done for full harmonization of the Bulgarian legislation with the one of the Community and the development of sufficient administrative capacity for its appropriate implementation. He emphasized the major aspects that the activity of NSPP encompasses – phytosanitary control, biological testing and registration of plant protection products, operative plant protection, and control of the application of plant protection products.
Main aspects in the activity of Plant Protection Institute were pointed out – development of technologies for biological and integrated plant protection and elements of organic agriculture, Risk assessment of contamination of plant production with pesticide residues and mycotoxins, development of flexible and effective expert systems for diagnostics and identification of pests by application of fast and reliable technologies at the level of European standards, pest risk assessment for emergence and spread of quarantine pests.
The audience was also introduced to the activities of the Central Laboratory for Chemical Testing and Control and the Central Laboratory of Plant Quarantine.
As a conclusion Dr. Todorov said that NSPP has a sufficient administrative and scientific capacity to perform all its duties and responsibilities set in the contract of accession of Bulgaria in the EU.


2. Tospovirus problems fort he Balkan: New developments and risks.
Günter Adam

The list of approved and and assigned new Tospovirus species is presented including their new clustering according to serology. Also the phylogenetic clustering according to the amino acid sequence of the N protein is presented which reveals beside the TSWV- and WMSMV-cluster a new cluster which contains the Near East species. These have, different from the “New World”- and “Asian”- cluster, the same vector species that are also endemic in the Balkan countries. Therefore it is recommended that the plant inspection have an increased interest for the detection of these new species, as possible new Tospoviruses to come.
Finally some new research from the laboratory of J. Moyer will be presented where via pseudorecombinant generation the reason for loss of insect transmission was pinpointed towards the Gn protein with probably a single point mutation. This is the first experimental evidence for a function of Gn for thrips transmission besides the fact that the complete loss of G proteins causes loss of particle formation and transmission.



3. Identification and Bioinformation – two important steps in modern pest thrips control.
Gerald Moritz

Situation: In the last decades the importance of pest thrips has increased considerably worldwide. These species are usually highly adaptable, with a well developed resistance against certain insecticides and with a tendency to be vectors of fungi, bacteria and viruses. Some of them are invasive, and are notorious for causing extensive crop damage. Their identification is important to understand their biology and to empower integrated pest management strategies. However, it is questionable how effective current taxonomic methods and research are to adequately serve these problems and future scenarios.
Classical Identification: The development of dichotomous keys is time consuming, expensive to print, of most use to professional taxonomists with good reference collections and libraries, and mostly focus on specific thrips taxa of local interest; their effective market niche is thus limited. In contrast, computing power is now inexpensive and available globally, and we should not ignore this availability.
Digital Identification: Several computerized identification and information systems, especially for pest thrips, are available. These systems provide fully illustrated and user-friendly means of recognising most of the thrips species that have been recorded as pests in various parts of the world. These multivariate computer keys have important advantages and features and include a range of information on the host plants, distributions and body structure of these thrips. Furthermore, you can use expert routes combined with original computerized colour pictures, movie clips and the possibility to compare the identified species with other species of the key.
Molecular Identification: The latest thrips key combines an illustrated visual key with molecular methods based on the ITS-RFLP technique and allows the identification of all ontogenetic Stages including eggs. In 2008 computerized key of Bulgarian pest thrips will be available.


4. Economically important thrips species in greenhouses in Bulgaria
Olia Karadjova, Vladimir Krumov

Agriculture is a traditional branch in our country’s economy. There has been a crisis in agriculture resulting from the transition period in Bulgaria in the early 90-s of the past century. In the course of the last 10 years there has been a steady trend of stabilization and increase in crop output. There is a tendency of establishment of new perennial crops – vineyards and orchards. Before the transition there were numerous greenhouses that were using ineffective methods for maintenance. Recently new and modern greenhouses were established.
A higher quality of plant production is being sought using organic agriculture. This brings out a need for more effective methods for pest control and, in particular, research directed towards new approaches in control of insects vectoring plant viruses.
With highest economic importance in Bulgarian greenhouses is Frankliniella occidentalis. Since the Western Flower Thrips was first reported in Bulgaria in 1991, it has become one of the most important greenhouse crop pests. Mass infestations of WFT transmitting TSWV, cause severe damage on crops, sometimes reaching 96-100% and leading to total loss of the yield.
INSV was detected several times during the monitoring program in Impatiens spp. seedlings imported mainly from Israel, but the infected plants were destroyed. Since then the virus has not been reported.
Thrips tabaci is second by its economic importance in Bulgaria. Its populations are vectors of TSWV on tobacco and tomatoes on the field and sometimes on vegetables in greenhouses. In the past, there were many outbreaks on tobacco, the most serious of which were in 1969 with 57% reduction in yield and in 1983 - 53%. The second virus that is transmitted by Th. tabaci - IYSV is not reported yet in Bulgaria.
E. americanus was introduced in Europe in 1993. The species was first established in Bulgaria in 2001. In the year of 2002 the species was found on different floricultural greenhouses and markets in Sofia, Plovdiv and Burgas. This thrips is not a vector of plant viruses. E. americanus is susceptible to most insecticides.


5. Diagnosis of the main Tospoviruses in greenhouses in Bulgaria
Dimitrinka Hristova, PPI, Bulgaria

The task is focused on Tomato spotted wilt virus (TSWV) and the main vector of the virus Thrips tabaci. TSWV was reported for first time in Bulgaria in 1952 in the tobacco growing areas in the south and two years later in tomato fields in the north part of Bulgaria. Since then TSWV epidemics have frequently occurred in major tobacco areas and neighboring tomato fields. In 1993 TSWV epidemics have frequently occurred in glasshouses all over the country, caused by the rapid expansion of the new vector species Francliniella occidentalis.
A survey of tospoviruses presented in Bulgaria was conducted during three vegetation periods as 258 different isolates from tobacco, tomato, pepper, weeds and ornamentals in the open field and from glasshouses were sampled in Bulgaria. The results obtained for 14 representative viral samples are analyzed and prove that only TSWV isolates were present.
TSWV were with variable symptom phenotype and among the isolates analyzed two of them-DH37 and GD98, were obtained that could overcome the resistance of some N- transgenic tobacco lines produced in Bulgaria. The absence of INSV ,indicates that until now the tospovirus population in Bulgaria is endogeneous and probably not contaminated with strains from Western Europe.
The complete S- RNA sequences of four Bulgarian isolates -10HK96,BS97,DH37 and GD98 and one German isolate of TSWV were determined in Institute of Applied Botany, Hamburg University. All isolates show a high conservation in their N proteins, while the NSs proteins and the intergenic regions (IGR) were more variable.
The Bulgarian isolate 10HK96 has the largest S-RNA (3364 nucleotides) among tomato spotted wilt viruses reported so far. The enlargement is based on an insertion of 365 nts in the IGR that may have resulted from stuttering of the viral polymerase or non-homologous recombination.
All these results confirm the need of more detailed investigation of diversity of Tospoviruses not only in Bulgaria but in the whole Balkan region. There is a need to continue efforts of identification of new Tospoviruses in Bulgaria and in the Balkan region.


6. Bemisia tabaci – Biology, vector potential and control in greenhouses
Piero Caciagli.

A brief description of biology, developmental stages and life cycle of Bemisia tabaci was made. Tools for pest identification and some references of use were mentioned. Some main differences between Trialeurodes vaporariorum and Bemisia tabaci were pointed out. Twenty biotypes of tobacco whitefly were presented, determined with the use of DNA-based techniques like esterase banding pattern, amplified fragment length polymorphism and others.
Some data on Bemisia tabaci, biotype B were given – squash silver leaf transmition, biological traits, with special emphasis on temperatures and the possibilities for survival in open areas of Italy. Mean annual temperatures of thirteen European countries were compared in order to assess the risk of pest establishment in the open field. Conclusions based on climatic conditions in the central Balkans estimate the risk of Bemisia tabaci colonizing the fields as low to moderate and colonizing greenhouses as moderate. Plant viruses, transmitted by tobacco whitefly were also mentioned with emphasis on viruses present in Europe.
Some information on control methods – mechanical, conventional, biological, and plant resistance was also presented.


7. Virus diseases of the grapevine and the Directive 2005/43/EC.
Giovanni Martelli.

The taxonomy of currently known grapevine viruses was presented briefly. Production damage and economic losses were emphasized.
Effect of nepoviruses, presence in Europe and their vectors; leafrol viruses, crop losses and vectors; rugose wood complex and transmission of rugose-wood associated viruses; fleck complex; graft incompatibility and agents were mentioned.
The approach of EEC on the sanitary status of grapevine nursery productions was summarized since the original directive in 1968 till now with some of the amendments, proposals and recommendations.
For direct detection of FD and BN phytoplasmas bi-specific PCR procedure (multiplex nested-PCR assay) was used.
The stolbur phytoplasmas of grapevine have already been reported in neighboring country as Greece (Davis et al.,1977), Macedonia (Segura et al.,2003) and Serbia (Deduk et al.2003) but this is the first report of the phytoplasma presence on grapevine in Bulgaria. Greece and in Macedonia.
On the basis of these results the NPPS has taken measures for control against spreading the disease (including inspection of mother plants during the growing season), eradication of contaminated canes ( if it is necessary) and also prophylactic treatments against spreading of the leafhopper Scaphoideus titanus, Hyalesthes obsoletus, Repthalus spp., Zygina spp., Empoasca vitis and etc. Another alternative for prognosis and integrated management is the chemical control.


8. Fruit tree indexing.
Wilhelm Jelkmann

The way fruit trees are indexed in Europe are describe by EPPO in three reports, one for Malus, Pyrus and Cydonia, one for almond, apricot, peach and plum and the last for cherry. These reports contain lists of the relevant pathogens, indexing varieties and indexing parameters for field and greenhouse testing, but also other suitable testing procedures if known. For many examples the symptoms on indicator plants will be presented. The advantages and disadvantages of the indexing procedure in comparison to other tests will be discussed and explained. It will also be touched briefly, how pathogenfree fruit trees can be generated.


9. Citrus Black Spot (Guiguardia citricarpa) – epidemiology and detection.
Irene Vloutoglou

The taxonomical position of Citrus black spot and the susceptible and tolerant hosts were mentioned. Different types of symptoms are hard spots and pycnidia, freckle spots, virulent spots, cracked spots.
The life cycle and sources of inoculum – leaf litter (perithecia - ascospores), attached leaves, fruits twigs (pycnidia-pycnidiospores), citrus plant organs (latent mycelium) were presented.
The favorable environmental conditions (temperature 24 – 25 °C, high relative humidity) and world distribution of Guiguardia citricarpa were mentioned. The pest is registered only in the Southest parts of the world – Argentina, South Africa, Australia and some parts of Asia. The pest is in A1 list of EPPO and regulations of import were emphasized.
Some methods of detection and identification like visual inspection of symptoms, microscopic examination of pycnidia, and two methods for detection when pycnidia are absent with their advantages and disadvantages were presented.
10. Monitoring Program for Quarantine Pests on grapevine in Bulgaria
Zhelju Avramov

Grapevine yellows (GY) are associated with several different phytoplasmas worldwide but cannot be identified on the basis of symptoms.
Flavescence doree (FD) is the most dangerous GY, it dramatically affects large vinegrowing areas in France, Italy, Spain, Slovenia, Croatia and Serbia. The FD phytoplasma is a quarantine pest in Bulgaria and it is transmitted by a vine-feeding leafhopper Scaphoideus titanus Ball.
Bois noir (BN) which is associated with phytoplasma stolbur (STOL) or 16SrXII is the second economically important GY in Europe and is transmitted by the planthopper Hyalestes obsoletus Sing.
During the monitoring program for quarantine pests of the grapevine from 2003 to 2006 about 2500 samples were tested as all regions in Bulgaria are included. Number of analysis of phytoplasma for 2005 were 352 and for 2006 – 457.
The planthopper species Hyalesthes obsoletus Signoret known as both a vector of Stolbur phytoplasma and the vector of Bois noir (BN) disease of grapevine was identified in the regions of Bourgas, Varna, Vratsa, Rousse, Veliko Tarnovo, Stara Zagora and Pleven. For the first two years of the Monitoring programme Scaphoideus titanus was not detected but it was identified in the summer of 2006.
Species which were found on the territory of Bulgaria are: Zygina rhamni (F), Eupterix aurata (L), Empoasca vitis (Goethe), Neoaliturus fenestratus (Her.Schäffer), Cicadella viridis, (Linnaeus), Zyginidia sp., Philaenus sp., Reptalus sp.
Typical symptoms of GY were observed on red cultivar Merlot from the region of Sliven in 2005. The samples were analyzed by PRC using P1/P7 and R16F2n/R16R2 primers and AluII/RSA restriction enzymes.
For direct detection of FD and BN phytoplasmas bi-specific PCR procedure (multiplex nested-PCR assay) was used.
The stolbur phytoplasmas of grapevine have already been reported in neighboring country as Greece (Davis et al.,1977), Macedonia (Segura et al.,2003) and Serbia (Deduk et al.2003) but this is the first report of the phytoplasma presence on grapevine in Bulgaria. Greece and in Macedonia.
On the basis of these results the NPPS has taken measures for control against spreading the disease (including inspection of mother plants during the growing season), eradication of contaminated canes ( if it is necessary) and also prophylactic treatments against spreading of the leafhopper Scaphoideus titanus, Hyalesthes obsoletus, Repthalus spp., Zygina spp., Empoasca vitis and etc. Another alternative for prognosis and integrated management is the chemical control.

11. Phytosanitary threats and inspection requirements at import of plants and plant products.
Harry Arijs

The Council Directive 2000/29/EC consists of 29 articles with regulative protective  measures against the introduction of harmful organisms to plants or plant products into the Community, as well as spread therein. The attached five Annexes describe the organisms or commodities of specific plant and organisms which are subject of the regulations. In some examples the procedures of inspections for complex commodity casses are described, showing the measures which the Community takes to adopt its inspection to certain risks without interfering too much with international trade. As another example into this direction the ways to adjust the frequency of inspections will be explained, depending on associated risks or pending emergencies.
Finally the statistics for interceptions from year 2006 will be presented and explained in detail.


12. Problems faced in the field of phytosanitary control after accession of Bulgaria to the EC
Lubomir Kirilov,Nikolai Rosnev

Director General of NSPP is directly subordinate to the Minister of Agriculture and Forest. The position all structures of NSPP including PPI is presented.
Exchange of information is carried out between the NSPP, the main Directorate of Plant Protection, Phytosanitary Control Department, Central Laboratory for Plant Quarantine, 14 Regional Services for Plant Protection, Phytosanitary Control Department, territory survey and 12 Borders Control Points with lots connections in between.
Totally 541 people are working in the system of NSPP. Main directorate “Plant Protection” include Phytosanitary Control Department (with 225 people including headquarters and CLPQ), 4 NSPP headquarters, 24 people in Central Laboratory for Plant Quarantine, 73 in BIPs and 125 in RSPP.
Main objectives of the plant health regulatory system (phytosanitary control) are to prevent the introduction of harmful organisms (quarantine pests) and control their spread on the state territory and EC. Achieving the objects through implementation and enforcement of harmonized legislation, harmonized legislation in all Member States, Standing Committee of Plant Health (amendments and decisions).
Within 4 months Bulgarian legislation in the field of phytosanitary control was fully harmonized with EC Law.
Ordinances and directives in connection of phytosanitary control are listed.
Programme 1 is to increase BIPs up to 30 until the end of 2006.
Main problem which comes into being is that in 2006 on BIPs was carried out phytosanitary control on 33265 lots of plant origin. During the first 3 months of 2007 – on BIP Kapitan Andreevo was carried out phytosanitary control on 7000 lots with destination Bulgaria and other MS as the expected lots are 25000.
Weak points and irregularities are also tourist traffic (vacation, souvenirs), parcels per post, internet-parcels and to control these close cooperation with other services is needed.
Programme 2 is for control on the territory of the country (system of monitoring programs and national strategy), involvement of science potential in the plant health domain as CLPQ and Plant Protection Institute (ensuring export of the country as in 2006 Bulgaria was not notified for intercepted quarantine pests).
Programme 3 is for lab diagnostic network as the problems here are the recourses for maintenance, supplies, spare parts, modern equipment.
As a conclusion we could say that the main objective of plant health is quarantine and prevention spreading of harmful organisms.


13. Control of plants and plant products during their movement within the EU: control of potatoes and fruit tree planting material.
Harry Arijs

Almost similar regulations exist for the movement of material inside the EU to prevent the further spread of harmful organisms, already present in the Community. This is for a few exceptions regulated via plant passports issued for moving plants and some plant products. Exceptions are possible for distance and amount of material movved which is regulated by the different Member States individually. The term Protected Zone which defines areas that have special protection against one of the 26 harmful organisms in the EU is explained, as well as the emergency measures that can be demanded in case of especially harmful organisms like Phythophthora ramorum, Diabrotica virgifera or Bursaphelenchus xylophyllus will be explained. Finally as special cases for plant health measurements the conditions for the movement of potatoes and malus plants in the Community as well as into protected zones will be explained in detail.


14. Was cancelled


15. Globodera pallida – biology, identification methods and control.
Kenneth Evans

The potato cysts nematode occurs as two very closely related strains which are now two different species, the white cyst nematode, Globodera pallida and the golden cyst nematode, Globodera rostochiensis. They can be differentiated morphologically but with great difficulty or by protein patterns after isoelectric focussing with less problems. Differentiation is necessary to chose the right strategies for defence and prevention of building up large populations in potato fields, because G. pallida is more persistent and also more resistant to available nematicides. Different defence strategies will be presented, among which precision farming in combination with a population modelling computer program seems to offer greatest success. The program with its different parameters for predicting the success of several integrated treatments will be presented.


16. Details on the situation of Clavibacter michigahensis ssp. sepedonicus in Bulgaria
Mariana Laginova, Elka Dimitrova

Clavibacter michiganensis spp. sepedonicus (CMS) is a quarantine pathogen for Bulgaria according to the Plant Protection Law harmonized with Direct.  200/29 EC and 2006/56 EC. It was the aim of the described research to show that Bulgaria is free of CMS as demande by the free zone regulations, since Bulgaria was recognized as QMS free in the Decision 2005/870/EC.
In order to fulfill the requirements 310,55 ha seed potato area and 3098,2 ware potato producing are was tested. The seed and ware potato testing procedures and resulting statistics are presented for the las 4 years. In additions the intecept statistics for Imports of seed and ware potatos are presented.
According to the test results import potatos were found in three cases infected with CMS. Among the Bulgarian production sites from 2 and 14 areas of seed and ware potatos, respectively, CMS infected material was found. The imposed quarantine measures are described.