Short communication: Pepino mosaic virus, a new threat for Serbia’s tomatoes

Aim of study: To report the occurrence of Pepino mosaic virus (PepMV) on tomato in Serbia and to genetically characterize Serbian PepMV isolates.Area of study: Tomato samples showing virus-like symptoms were collected in the Bogojevce locality (Jablanica District, Serbia).Material and methods: Collected tomato samples were assayed by DAS-ELISA using antisera against eight economically important or quarantine tomato viruses. Three selected isolates of naturally infected tomato plants were mechanically transmitted to tomato ‘Novosadski jabučar’ seedlings. For confirmation of PepMV infection, RT-PCR was performed using specific primers PepMV TGB F/PepMV UTR R. Maximum-likelihood phylogenetic tree was constructed with 47 complete CP gene sequences of PepMV to determine the genetic relationship of Serbian PepMV isolates with those from other parts of the world.Main results: The results of DAS-ELISA indicated the presence of PepMV in all tested samples. Mechanically inoculated ‘Novosadski jabučar’ seedlings expressed yellow spots and light and dark green patches, bubbling, and curled leaves. All tested tomato plants were RT-PCR positive for the presence of PepMV. The CP sequence analysis revealed that the Serbian PepMV isolates were completely identical among themselves and shared the highest nucleotide identity of 95.1% (99.2% aa identity) with isolate from Spain (FJ263341). Phylogenetic analysis showed clustering of the Serbian PepMV isolates into CH2 strain, but they formed separate subgroup within CH2 strain.Research highlights: This is the first data of the presence of PepMV in protected tomato production in Serbia. Considering increased incidence and rapid spread in Europe, the presence of PepMV on tomato could therefore represent serious threat to this valuable crop in Serbia

Virusi jabučastih voćaka u Bosni i Hercegovini

Field surveys and laboratory tests were carried out to assess the sanitary status of pome fruit trees in Bosnia and Herzegovina. Field surveys were carried out in 10 orchards, two nurseries and one varietal collectione during 2005-2006. A total of 65 apple and 51 pear cultivars were tested for the presence of the four most important pome fruit viruses: Apple chlorotic leaf spot virus (ACLSV), Apple stem pitting virus (ASPV), Apple stem grooving virus (ASGV) and Apple mosaic virus (ApMV). The most frequent viruses of apple were ACLSV (72%) and ASPV (69%), whereas for pear those were ASGV (69%) and ACLSV (64%). Biological indexing was more reliable than ELISA for virus detection. Multiplex RT-PCR results of 20 randomly selected apple cultivars were in line with biological indexing. This is the first report of the presence of ACLSV, ASPV, ASGV and ApMV in Bosnia and Herzegovina in pome fruits.Pregled voćnjaka i laboratorijska testiranja vršena su u cilju utvrđivanja sanitarnog statusa jabučastih voćaka u Bosni i Hercegovini. Pregledano je 10 voćnjaka, dva rasadnika i jedan kolekcioni zasad tokom 2005. godine. Ukupno 65 sorti jabuke i 50 kruške testirano je na prisustvo četiri najznačajnija virusa jabučastih voćaka: virus hlorotične lisne pegavosti jabuke (Apple chlorotic leaf spot virus, ACLSV), virus jamičavosti stabla jabuke (Apple stem pitting virus, ASPV), virus brazdavosti stabla jabuke (Apple stem grooving virus, ASGV) i virus mozaika jabuke (Apple mosaic virus, ApMV). Na ispitivanim sortama jabuke, najzastupljeniji su bili ACLSV (72%) i ASPV (69%), dok je najznačajnije prisustvo, na određenim sortama kruške, utvrđeno za ASGV (69%) i ACLSV (64%). Biološko indeksiranje se pokazalo kao pouzdanija tehnika za detekciju virusa jabučastih voćaka od ELISA. Kod 20 slučajno odabranih sorti jabuke, rezultati dobijeni biološkim indeksiranjem su potvrđeni multiplex RT-PCR. Ovaj rad predstavlja prvo saopštenje o prisustvu virusa ACLSV, ASPV, ASGV i ApMV na jabučastim voćkama u Bosni i Hercegovini

Virus bronzavosti paradajza – jedan od najdestruktivnijih biljnih virusa

Tomato spotted wilt virus (TSWV) has one of the largest host ranges among plant viruses and is widespread in all climates. TSWV is responsible for numerous epidemics in many parts of the world in different crops, mainly vegetables, tobacco and ornamentals. Its highly polyphagous nature, effectiveness of virus transmission by the thrips as its vectors, rapidity with which new variants arise, as well as difficulties in controlling the vectors make TSWV one of the most dangerous plant viruses. The ability of this virus to cause such severe losses on a broad range of crops, as well as its intriguing biological and molecular characteristics place TSWV amongst the most extensively studied plant viruses in the world at present. This paper provides a general overview of TSWV, encompassing all the major aspects of its biology and current knowledge on host range, symptomatology, molecular biology, vector relationship, control and diagnosis.Smatra se da virus bronzavosti paradajza (Tomato spotted wilt virus, TSWV) ima najširi krug domaćina i da je jedan od najrasprostranjenijih biljnih virusa. Ovaj virus je odgovoran za brojne epidemije na različitim usevima u mnogim delovima sveta, a najčešće na povrću, duvanu i ukrasnim biljkama. Visoko polifagna priroda virusa, efikasnost prenošenja vektorima – tripsima, brzina kojom se stvaraju nove varijante virusa, kao i teškoće u kontroli vektora, čine TSWV jednim od najopasnijih biljnih virusa. Zbog ekonomske važnosti na širokom krugu domaćina, kao i interesantnih bioloških i molekularnih karakteristika, ovo je danas jedan od najproučavanijih biljnih virusa. Ovaj pregledni rad o virusu bronazavosti paradajza bliže objašnjava sve aspekte njegove biologije i sadrži savremene podatke o nekim njegovim osobinama koje se odnose na niz domaćina, simptomatologiju, molekularnu biologiju, odnos sa vektorom, kontrolu i dijagnozu oboljenja koje prouzrokuje

Characterization of carbon fibrous material from platanus achenes as platinum catalysts support

Carbon materials with developed porosity are usually used as supports for platinum catalysts. Physico-chemical characteristics of the support influence the properties of platinum deposited and its catalytic activity. In our studies, we deposited platinum on carbon fibrous like materials obtained from platanus seeds - achenes. The precursor was chemically activated with different reagents: NaOH, pyrogallol, and H2O2, before the carbonization process. Platinum was deposited on all substrates to study the influence of the substrate properties on the activity of the catalyst. Carbon materials were characterized by nitrogen adsorption/desorption isotherms measurements, X-ray diffraction, and scanning electron microscopy. It was noticed that the adsorption characteristics of carbon support affected the structure of platinum deposits and thus their activity

Uticaj neravnomerne distribucije virusa bronzavosti paradajza na serološko dokazivanje virusa u paradajzu, paprici i ukrasnim biljkama

Reliable detection of plant pathogens does not only mean the development of sufficiently sensitive laboratory techniques for their routine testing. Regardless of the sensitivity of applied methods, the proper selection of samples to be tested has crucial influence on method reliability. Due to uneven distribution of Tomato spotted wilt virus (TSWV) in naturally infected systemic host plants, the collection and sampling of material for assaying is a critical moment upon which the reliability of laboratory procedure depends. The effect of irregular virus distribution on its serological detection was examined in tomato, pepper and four ornamental species, as its most important host plants in our country. The reliability of virus detection, depending on its uneven distribution, was assessed by serological testing of tomato and pepper symptomatic leaves and fruits, and symptomatic and asymptomatic young and old leaves, as well as flower petals of ornamentals. Although TSWV was detected using ELISA in the majority of plants included in the experiment, the tests indicated an uneven distribution and unequal concentrations of TSWV in different parts of the plants. The virus could not be detected in a certain number of subsamples, prepared from infected tomato and pepper fruits and older ornamental leaves. The virus also could not be detected in some ornamentals and tomato plants with intensive symptoms. Conversely, the virus was detected in three ornamental plants without any symptoms. Examining the virus distribution in different plant parts indicated that the reliability of ELISA could be reached not only by sampling younger ornamental leaves, but also by preparing compound samples with as much leaves as possible, or by testing a greater number of subsamples of the tested plant. Considering a small possibility of TSWV detection in tomato and pepper fruits, the infection of these should be established by testing their leaves. Besides, the data show that negative test results for the known host plants with characteristic symptoms require tests to be repeated using more sensitive methods than ELISA. These results are important as guidelines for growers and professionals who submit samples, as well as for laboratories performing routine testing.Pouzdana detekcija fitopatogenih organizama ne podrazumeva samo razvijanje dovoljno osetljivih laboratorijskih tehnika za rutinsko testiranje. Bez obzira koliko primenjene metode bile osetljive, na njihovu pouzdanost presudno utiče pravilan izbor uzoraka koji će se testirati. Za virus bronzavosti paradajza (Tomato spotted wilt virus, TSWV), zbog neravnomerne distribucije u sistemično zaraženim biljkama domaćinima, prikupljanje i priprema uzoraka za testiranje predstavljaju kritični momenat od koga zavisi pouzdanost laboratorijske metode. Sprovedena su ispitivanja uticaja neravnomerne distribucije virusa u paradajzu, paprici i četiri vrste ukrasnih biljaka, kao najznačajnijim domaćinima ovog virusa u našoj zemlji. Pouzdanost detekcije virusa, u zavisnosti od njegove neravnomerne distribucije, utvrđivana je serološkim testiranjem lišća i plodova paradajza i paprike sa simptomima i simptomatičnog i asimptomatičnog mlađeg i starijeg lišća, kao i kruničnih listića ukrasnih biljaka. Iako je primenom ELISA metode TSWV dokazan u većini biljaka uključenih u eksperiment, obavljena testiranja ukazala su na neravnomernu distribuciju i različitu koncentraciju TSWV u različitim delovima biljaka. Prisustvo virusa nije moglo biti dokazano u određenom broju poduzoraka pripremljenih od zaraženih biljaka, plodova paradajza i paprike i starijeg lišća ukrasnih biljaka. Virus nije mogao biti detektovan ni u nekim ukrasnim biljkama i biljkama paradajza sa izraženim simptomima. Nasuprot tome, TSWV je dokazan u tri ukrasne biljke bez simptoma. Ispitivanja distribucije virusa u biljci ukazala su da se pouzdanost ELISA metode može postići uzorkovanjem ne samo mlađeg lišća ukrasnih biljaka, već pripremom zbirnog uzorka sa što više listova ili ispitivanjem većeg broja poduzoraka biljke koja se testira. S obzirom na malu mogućnost detekcije TSWV u plodovima paradajza i paprike, utvrđivanje zaraženosti ovih biljaka treba obaviti testiranjem lišća. Pored toga, dobijeni rezultati ukazuju da je u slučaju pojave negativnih rezultata testiranjem uzoraka sa karakterističnim simptomima na poznatim domaćinima, potrebno ponoviti testiranje osetljivijim metodama od ELISA. Dobijeni rezultati značajni su kao smernica za proizvođače i stručna lica koja dostavljaju uzorke i laboratorije zadužene za rutinska testiranja

Novi pravci primene otpornosti tikava u kontroli virusnih oboljenja

As there is a growing frequency of viral plant diseases in epidemic proportions, the possibilities for successful control are constantly being explored. Despite the fact that integral and simultaneous employment of numerous control measures may contribute to the decreasing amount of yield losses, especially concerning non-persistently aphid-transmitted viruses, these measures are often not efficient enough. Research into the basis of resistance to viral infection and principles of its inheritance, introduction of sources of resistance in susceptible genotypes, by conventional or genetic manipulations, are very intensive for cucurbit crops, especially pumpkins. Pumpkin crops are being endangered by a great number of different viruses, among which the Zucchini yellow mosaic virus, (ZYMV), Watermelon mosaic virus (WMV) and Cucumber mosaic virus (CMV) are present every year in Serbia, frequently causing epidemics. The majority of pumpkin cultivars are not resistant or tolerant to viral infections, but sources of resistance have been identified in various related species. So far, the identified sources of resistance to the ZYMV are found in Cucurbita moschata and Citrullus lanatus var. lanatus genotypes and consist of one or several major dominant genes of resistance. It is a similar case with WMV, although the sources of dominant major genes are identified in C. lanatus and C. colocynthis. The sources of resistance to CMV in the form of one dominant gene have been identified in the genotype C. moschata, although the introduction of this gene by conventional means proved to be very difficult. Besides the aforementioned, substantial efforts are being made in developing genotypes with multiple resistance against several viruses and even other pathogens, as well as genotypes with resistance to the most significant plant aphid species, through mechanisms of antixenosis or antibiosis. The other way of obtaining resistant genotypes includes genetic manipulation. Genetically modified resistant pumpkins have been among the first successfully developed crops. Genotypes with pathogen derived resistance can already be found in commercially grown pumpkins in some parts of the world, and they have been developed by introducing the coat protein gene of one, two or all three viruses which are the most frequent, ZYMV, WMV and CMV. Yet, this approach to the control of pumpkin viral diseases is related to possible negative consequences, mostly through the already detected gene transfer to wild plants and development of resistant transgenic weeds of unpredictable impact on the environment. Improved host plant genetic resistance to viral infections or biological vectors, developed by conventional or genetic engineering methods, represents the most dynamic and prominent field of research. It is economically and ecologically the most justified approach to the control of pumpkin and other plant diseases caused by viruses non-persistently transmitted by aphids.Kako je pojava brojnih viroza gajenih biljaka u epidemijskim razmerama sve češća, mogućnosti za uspešnu kontrolu stalno se ispituju. Integralna i istovremena primena brojnih pojedinačnih mera kontrole, naročito u slučaju virusa koji se prenose biljnim vašima na neperzistentan način, mogu da doprinesu smanjenju šteta, ali često nisu dovoljno efikasne. Proučavanja osnova otpornosti i načina nasleđivanja, unošenje izvora otpornosti u osetljive genotipove, konvencionalnim ili genetičkim manipulacijama, vrlo su intenzivna kod vrežastih kultura, posebno tikve. Tikve ugrožava veliki broj raznorodnih virusa od kojih se u Srbiji svake godine javljaju virus žutog mozaika cukinija (Zucchini yellow mosaic virus, ZYMV), virus mozaika lubenice (Watermelon mosaic virus, WMV) i virus mozaika krastavca (Cucumber mosaic virus, CMV), a pojedinih godina izazivaju epidemije. Većina sorti gajenih tikava ne poseduje otpornost ili tolerantnost na virusne zaraze, ali izvori otpornosti identifikovani su u različitim srodnim vrstama. Do sada identifikovani izvori otpornosti na ZYMV nalaze se u genotipovima Cucurbita moschata i Citrullus lanatus var. lanatus i obuhvataju jedan ili nekoliko major gena rezistentnosti koji se nasleđuju dominantno. Slična situacija je i sa WMV, mada su izvori dominantnih major gena identifikovani u C. lanatus i C. colocynthis. Izvor otpornosti na CMV u vidu jednog dominantnog gena identifikovan je u genotipu C. moschata, mada je unošenje ovog gena konvencionalnim putem bilo veoma teško. Pored toga, veliki napori ulažu se u dobijenje genotipova sa istovremenom otpornošću na više virusa čak i drugih patogena, kao i genotipova koji ispoljavaju otpornost na najznačajnije vrste biljnih vaši, kroz mehanizme antiksenoze ili antibioze. Drugi pravac dobijanja otpornih genotipova podrazumeva genetičke manipulacije. Genetski modifikovane otporne tikve su među prvima, od svih gajenih biljaka, uspešno razvijene. Genotipovi sa PD (pathogen derived) otpornošću već se nalaze u komercijalnoj proizvodnji tikava u nekim delovima sveta i dobijeni su unošenjem gena za CP (coat protein) jednog, dva ili sva tri u svetu najraširenija virusa, ZYMV, WMW i CMV. Ipak, ovaj prilaz kontroli virusnih oboljenja tikava vezan je za uočene moguće negativne posledice, pre svega kroz već registrovan transfer gena u biljke spontane flore i nastanak otpornih transgenih korova nepredvidljivog ponašanja i značaja u prirodi. Poboljšana genetička otpornost biljke domaćina bilo na infekciju virusima, bilo na biološke vektore, dobijena konvencionalnim ili metodama genetičkog inženjeringa predstavlja najdinamičnije i najperspektivnije polje istraživanja kao ekonomski i ekološki najopravdaniji pristup kontroli oboljenja tikava i drugih biljaka koje izazivaju virusi koji se neperzistetno prenose biljnim vašima

Prisustvo i rasprostranjenost virusa uljane tikve i molekularna detekcija virusa žutog mozaika cukinija

Over the past decade, intensive spread of virus infections of oilseed pumpkin has resulted in significant economic losses in pumpkin crop production, which is currently expanding in our country. In 2007 and 2008, a survey for the presence and distribution of oilseed pumpkin viruses was carried out in order to identify viruses responsible for epidemics and incidences of very destructive symptoms on cucurbit leaves and fruits. Monitoring and collecting samples of oil pumpkin, as well as other species such as winter and butternut squash and buffalo and bottle gourd with viral infection symptoms, was conducted in several localities of Vojvodina Province. The collected plant samples were tested by DAS-ELISA using polyclonal antisera specific for the detection of six most economically harmful pumpkin viruses: Cucumber mosaic virus (CMV), Zucchini yellow mosaic virus (ZYMV), Watermelon mosaic virus (WMW), Squash mosaic virus (SqMV), Papaya ringspot virus (PRSV) and Tobacco ringspot virus (TRSV) that are included in A1 quarantine list of harmful organisms in Serbia. Identification of viruses in the collected samples indicated the presence of three viruses, ZYMV, WMV and CMV, in individual and mixed infections. Frequency of the identified viruses varied depending on locality and year of investigations. In 2007, WMV was the most frequent virus (94.2%), while ZYMV was prevalent (98.04%) in 2008. High frequency of ZYMV determined in both years of investigation indicated the need for its rapid and reliable molecular detection. During this investigation, a protocol for ZYMV detection was developed and optimized using specific primers CPfwd/Cprev and commercial kits for total RNA extraction, as well as for RT-PCR. In RT-PCR reaction using these primers, a DNA fragment of approximately 1100 bp, which included coat protein gene, was amplified in the samples of infected pumkin leaves. Although serological methods are still useful for large-scale testing of a great number of samples, this protocol, due to its high sensitivity and specificity, is an important improvement in rapid diagnosis of diseases caused by this virus. In addition, the protocol provides a basis for further characterization of ZYMV isolates originating from Serbia.Intenzivno širenje virusa infektivnih za uljanu tikvu (Cucurbita pepo), poslednjih deset godina imalo je za posledicu značajne ekonomske gubitke u proizvodnji ove kulture koja se gaji na sve većim površinama u našoj zemlji. Kako bi se identifikovali virusi, odgovorni za epidemijsku pojavu i ispoljavanje veoma destruktivnih simptoma na lišću i plodovima uljane tikve, tokom 2007. i 2008. sprovedeno je ispitivanje njihove pojave i rasprostranjenosti. Pregled i sakupljanje uzoraka uljane tikve, kao i nekih drugih vrsta tikava, kao što su bundeva, muskatna i bizonska tikva i vrg sa simptomima virusnih zaraza, obavljeno je na više različitih lokaliteta gajenja uljane tikve u Vojvodini. Sakupljeni uzorci testirani su DAS-ELISA metodom primenom poliklonalnih antiseruma specifičnih za detekciju u svetu šest ekonomski najznačajnijih virusa tikava: virusa mozaika krastavca (Cucumber mosaic virus, CMV), virusa žutog mozaika cukinija (Zucchini yellow mosaic virus, ZYMV), virusa mozaika lubenice (Watermelon mosaic virus, WMW), virusa mozaika bundeve (Squash mosaic virus, SqMV), virusa prstenaste pegavosti papaje (Papaya ringspot virus, PRSV) i virusa prstenaste pegavosti duvana (Tobacco ringspot virus, TRSV), koji se nalazi na A1 karantinskoj listi štetnih organizama u Srbiji. Identifikacija virusa u sakupljenim uzorcima ukazala je na prisustvo tri virusa, ZYMV, WMV i CMV, koji su se javili u pojedinačnim ili mešanim infekcijama. Njihova učestalost je po pojedinim godinama i lokalitetima bila različita. Tokom 2007. najčešće je dokazan WMV (94,2%), dok je 2008. godine prevalentan virus bio ZYMV (98,04%). Velika učestalost ZYMV u obe godine ispitivanja ukazala je na potrebu za brzom i pouzdanom molekularnom detekcijom ovog virusa. U toku ovih istraživanja razvijen je i optimizovan protokol za detekciju ZYMV primenom specifičnih prajmera CPfwd/CPrev i komercijalnih kitova za ekstrakciju ukupne RNA i RT-PCR. Korišćenjem ovih prajmera, kojim se amplifikuje deo genoma ZYMV kojim je obuhvaćen i gen za proteinski omotač, umnožen je DNK fragment dužine oko 1100 bp iz lišća zaraženih biljaka. Mada serološke metode i dalje imaju veliku prednost u primeni za masovna testiranja velikog broja uzoraka, razvijeni protokol molekularne detekcije, zbog visoke osetljivosti i specifičnosti predstavlja značajno poboljšanje brze dijagnoze oboljenja koja ovaj virus izaziva. Osim toga, ovaj protokol pruža osnovu za dalju karakterizaciju ZYMV izolata poreklom iz Srbije

Novel Strategy for Electroanalytical Detection of Antipsychotic Drugs Chlorpromazine and Thioridazine; Possibilities for Simultaneous Determination

A simple and fast method for determination of two phenothiazine drugs, chlorpromazine (CPZ) and thioridazine (TDZ), at the boron-doped diamond electrode (BDDE) was proposed. Oxidation peaks on potentials higher than 1 V were used for quantitation of investigated phenothiazines due to better selectivity over common interfering compounds in urine. Differential pulse voltammetry was applied for trace determination of CPZ and TDZ, in Britton-Robinson buffer solution at optimal pH. Under optimized DPV conditions a linear analytical curve was obtained from 1.0x10(-7) to 4.0x10(-5) M with a detection limit of 0.3x10(-7) M (for CPZ) and in the concentration range of 2x10(-7) M to 4.0x10(-5) M, with a detection limit of 1.2x10(-7)M (for TDZ). The applicability of the method was proved by determination of CPZ and TDZ by proposed procedures in spiked urine samples. The electrochemical behavior of those two important tranquilizers in different solvents was described, exploring the possibility of simultaneous determination

Prisustvo i molekularna karakterizacija virusa mozaika lucerke u usevu duvana u Srbiji

Three-year investigation of the presence and distribution of tobacco viruses in Serbia revealed that Alfalfa mosaic virus (AMV) appeared every year with different frequency in tobacco crops. During 2008, the presence of AMV was detected in most of the tested samples (58.82%) and it was the second most common compared to all other viruses which presence was confirmed in Serbia. In 2006 and 2007, AMV was detected in a significantly lower percentage (2.80% and 13.64%, respectively). This study showed that Alfalfa mosaic virus was more commonly found in multiple infections with two, three or even four detected viruses. Single infections were detected only in 2006, in one tobacco field in the locality of Futog. During this investigation, a rapid and simple protocol was optimized and developed for molecular detection of AMV in tobacco leaves, using primers CPAMV1/CPAMV2 and commercially available kits for total RNA extraction as well as for RT-PCR (reverse transcription - polymerase chain reaction). Using RT-PCR and these primers that flank the AMV coat protein gene, a DNA fragment of 751 bp was amplified, sequenced, and compared with the sequences available in GenBank database. The sequence of isolate 196-08 (GenBank Acc. No. FJ527749) proved to be identical at the nucleotide level of 99 to 93% with those from other parts of the world. Phylogenetic analysis of 27 isolates based on 528 bp sequences of the coat protein gene did not show correlation of the isolates with their geographic origin or plant host and showed that these isolates fall into four molecular groups of strains. Serbian AMV isolate from tobacco belongs to group IV, the group that includes most of the isolates selected for phylogenetic analysis.Trogodišnjim proučavanjima pojave i rasprostranjenosti virusa u usevu duvana u Srbiji utvrđeno je da se virus mozaika lucerke (Alfalfa mosaic virus, AMV) javlja svake godine sa različitom učestalošću. Prisustvo AMV ustanovljeno je u najvećem broju testiranih uzoraka (58,82%) tokom 2008. godine kada je bio drugi po zastupljenosti u odnosu na sve ostale viruse čije je prisustvo utvrđeno u Srbiji. AMV je detektovan u znatno nižem procentu 2006. (2,80%) i 2007. godine (13,64%). Tokom ovih istraživanja virus mozaika lucerke češće je bio prisutan u mešanim infekcijama sa dva, tri ili čak četiri detektovana virusa. Prisustvo pojedinačne zaraze utvrđeno je samo 2006. godine u jednom usevu duvana na lokalitetu Futog. U toku ovih istraživanja razvijen je i optimiziran brz i precizan protokol za molekularnu detekciju AMV u lišću duvana primenom specifičnih prajmera CPAMV1/CPAMV2 i komercijalnih kitova za ekstrakciju ukupnih RNK i RT-PCR (reverzna transkripcija praćena lančanom reakcijom polimeraze). Korišćenjem ovih prajmera, koji omogućavaju umnožavanje gena za protein omotača, umnožen je fragment dužine 751 bp, sekvencioniran i upoređen sa sekvencama dostupnim u GenBank bazi podataka. Dobijena sekvenca izolata 196-08 (GenBank Acc. No. FJ527749) pokazala je 99-93% nukleotidne identičnosti sa izolatima AMV iz drugih delova sveta. Filogenetske analize 27 izolata, na osnovu 528 bp sekvence gena za protein omotača, nisu ukazale na korelaciju sa geografskim poreklom izolata ili biljkom domaćinom, a pokazale su grupisanje izolata u četiri molekularne grupe sojeva. Izolat AMV iz duvana iz Srbije pripada grupi IV, koja obuhvata većinu izolata odabranih za filogenetsku analizu

Biološka varijabilnost virusa žutog mozaika cukinija u Srbiji

Zucchini yellow mosaic virus (ZYMV) has become an important pathogen of cucurbits over the past 20 years. In our country, this virus also causes significant losses, especially in oil pumpkin production. Although ZYMV variability has been extensively studied worldwide by biological and molecular characterization, such information is still limited for the European region. Herein, we present the first analysis of phenotypic variations of ZYMV isolates sampled from oil pumpkin cv. Olinka from different locations in Serbia during 2008. Twelve isolates were obtained by mechanical inoculation on the C. pepo hybrid Ezra F1, and four of them were chosen for further detailed biological study. These investigated isolates exhibited different phenotypical features and thus represented different variants, concerning their host range, and type and severity of symptoms. A previously developed primer pair, ZYMV CPfwd/CPrev, was tested to determine its suitability for detection of Serbian ZYMV isolates. Amplification of Serbian ZYMV isolates of different origin, in single and mixed infection, resulted in one distinctive band. The used primer pair proved to be useful not only for routine molecular detection, but also for further molecular characterization of Serbian ZYMV isolates because it amplifies the hypervariable genomic region of ZYMV. The result obtained in this study is a contribution to investigation of genetic diversity of Serbian ZYMV isolates and their molecular relationship with ZYMV isolates from other parts of the world.Poslednjih 20 godina virus žutog mozaika cukinija (Zucchini yellow mosaic virus, ZYMV) se javlja kao značajan patogen biljaka iz familije tikava. Kod nas ovaj virus, takođe, izaziva značajne gubitke, naročito u proizvodnji uljane tikve. Iako je biološka i molekularna varijabilnost ZYMV dosta ispitivana, još uvek nema dovoljno informacija o varijabilnosti evropskih izolata ovog virusa. Ovaj rad predstavlja prvu analizu fenotipske varijabilnosti izolata ZYMV poreklom iz Srbije, koji su tokom 2008. godine sakupljeni iz različitih lokaliteta gajenja uljane tikve sorte Olinka. Mehaničkim inokulacijama C. pepo hibrid Ezra F1, dobijeno je 12 izolata, od kojih su četiri odabrana za dalju biološku karakterizaciju. Na osnovu različitog kruga domaćina, tipa i jačine simptoma na test-biljkama, utvrđena je fenotipska varijabilnost ispitivanih izolata. Par prajmera ZYMV CPfwd/Cprev testiran je radi utvrđivanja pogodnosti za rutinsku detekciju izolata ZYMV poreklom iz Srbije. Na osnovu pojave traka očekivane veličine u gelu, ZYMV je detektovan i u pojedinačnim i mešanim infekcijama u biljkama sa različitih lokaliteta, što dokazuje pogodnost ovih prajmera. Kako ovi prajmeri amplifikuju hipervarijabilni deo genoma virusa, pogodni su ne samo za rutinsku molekularnu detekciju, već i za molekularnu karakterizaciju izolata ZYMV iz Srbije. Rezultati dobijeni u ovom radu omogućiće dalja proučavanja genetičke varijabilnosti izolata ZYMV iz Srbije, kao i njihove evolutivne povezanosti sa izolatima iz drugih delova sveta