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During the spring of 2003, gall symptoms on the roots and crowns of young raspberry plants cv. Vilamette were observed near Valjevo. Phytopathogenic bacteria were isolated from diseased plant samples. Based on the pathogenic, morphological, differential biochemical and physiological characteristics, the isolated strains were identified as tumorigenic Agrobacterium radiobacter (biovar 1 Agrobacterium). In order to confirm the identity of isolated strains by polymerase chain reaction (PCR) primers complementary to tms2 gene located on the Ti plasmid were used. In the first PCR protocol using a tms2F1 + tms2R2 primer pair, 617 bp products specific for tumorigenic Agrobacterium strains were amplified. The second PCR protocol, using a tms2F1 + tms2B primer pair, amplified the expected 458 bp products. On the basis of multiplex PCR with primers complementary to chromosomal gene coding for 23S rRNA, the isolated strains were classified as biovar 1 Agrobacterium (A. radiobacter). This is the first report of tumorigenic A. radiobacter on raspberry in Serbia. Tokom proleća 2003. godine, zapaženi su tumori na korenu i korenovom vratu mladih biljaka maline sorte Vilamette u okolini Valjeva. Iz obolelih uzoraka izolovane su fitopatogene bakterije. Proučavanjem patogenih, morfoloških i diferencijalnih biohemijsko-fizioloških karakteristika izolovanih sojeva, utvrđeno je da proučavani sojevi pripadaju bakteriji Agrobacterium radiobacter (biovar 1). U cilju potvrde identiteta izolovanih sojeva lančanom reakcijom polimeraze (PCR), korišćeni su prajmeri komplementarni tms2 genu lociranom na Ti plazmidu. U prvoj PCR reakciji izvedenoj korišćenjem tms2F1 + tms2R2 para prajmera, detektovani su fragmenti nukleinske kiseline veličine 617 baznih parova (bp), specifični za tumorogene sojeve A. tumefaciens. U drugoj PCR reakciji korišćenjem para prajmera tms2F1 i tms2B su amplifikovani PCR produkti očekivane veličine 458 bp, karakteristični za ovu bakteriju. Na osnovu multiplex PCR reakcije korišćenjem prajmera komplementarnih na hromozomalni gen koji kodira 23S rRNA sojevi izolovani iz tumora na korenu maline svrstani su u biovar 1 Agrobacterium (A. radiobacter). Ovo je prvi nalaz ove bakterije na malini u Srbiji.

The test results of Pseudomonas syringae strains, isolated by a pear trees are as given. The symptoms of disesease, caused by this bacterium, appeared in two types: a blossom blast, a trunk necrosis and a branch followed by canker formation. All strains are Gram negative, fluorescent on a King medium B and oxydative (O/F test). The tested strains are HR positive, producing a levan, but don't oxydase, arginindehydrolase and pectinase (LOPAT +---+). Strains originated from a pear trees caused necrosis on an artificial inoculated pear, cherry and lemon fruits, as well as a syringae leaves and bean pods. The results of differential tests for P. syringae pv. syringae and P. syringae pv. morsprunorum (GATT) revealed that the tested strains hydrolise gelatin and esculin, but the negative results are recorded in tyrosinase production and tartrate utilization tests. The PCR analysis, by using a BOX primer, shows the high level of symilarity among the Serbian P. syringae strains, isolated from a pear fruit trees, but also within a slice differencies, compared with a check strain CFBP 1582. These results confirm a previous data about genetic diversity among P. syringae strains originated from a different areas all around the world. Based on the obtained results, it's concluded that the tested strains belong to P. syringae pv. syringae. Further characterization of P. syringae strains, isolated from pear in Serbia use the ERIC and REP PCR and it's still underway. U radu su prikazani rezultati proučavanja fitopatogene bakterije Pseudomonas syringae, kao patogena kruške u Srbiji. Bolest se ispoljava u dva vida simptoma: palež cvasti i nekroza grana i debla mladih stabala kruške praćena obrazovanjem rak rana. Izolovani sojevi bakterije su Gramnegativni, fluorescentni, glukozu metabolišu isključivo u aerobnim uslovima (oksidativno). Stvaraju levan i prouzrokuju HR duvana ali ne stvaraju oksidazu, arginin dehidrolazu i pektinazu (LOPAT + - - - +). Prouzrokuju nekrozu inokulisanih plodova kruške, trešnje, limuna, listova jorgovana i mahuna boranije. Proučavani izolati hidrolizuju želatin i eskulin ali negativno reaguju pri testovima stvaranja tirozinaze i metabolizma tartarata (GATT). Na osnovu patogenih i biohemijskih odlika zaključeno je da proučavani izolati ispoljavaju izrazitu sličnost sa Psudomonas syringae pv. syringae, široko rasprostranjenim i ekonomski štetnim patogenom kruške. Primenom PCR analize korišćenjem BOX prajmera potvrđena je izrazita homogenost sojeva poreklom sa kruške, izolovanih u Srbiji, ali izvesne razlike u odnosu na kontrolne sojeve, što potvrđuje ranije dokazanu raznolikost sojeva ove bakterije u zavisnosti od lokaliteta i područja iz kog su izolovani. Na osnovu rezultata istraživanja dat je i predlog za uspostavljanje standardne procedure na bazi patogenih, biohemijskih i molekularnih metoda (primenom PCR) u cilju što brže i pouzdanije detekcije ove bakterije u biljnom materijalu, što je od velikog značaja pri proizvodnji sadnog materijala, proučavanja epidemiologije patogena i razrade mera njegovog suzbijanja.

Firelight symptoms were observed on ornamental plants Sorbs domestic (region of Niš, 2005) and S. terminals (Požega, 2007). Symptoms appeared as leaves and shoots blight followed with cancer forming on branches of S. domestic and shoot blight on S. terminals. Levant type single colony were collected after three days growth on nutrient agar enrich net with sucrose (SNA). Investigated strays produced HR in tobacco and necrosis of artificially inoculated unripe pear fruits followed by oozing of bacterial exudates. Symptoms similar to those registered in natural infections, were recorded on artificial inoculated S. domestic leaves and shoots. Based on the results of pathogen city tests, biochemical characteristics, ELISA test and PCR analysis it was confirmed that the investigated strains belonged to E. amylovora, widespread pathogen of poem fruit trees and ornamental plants in Serbia. This is the first report of E. amylovora as pathogen of Sorbs plant in Serbia. U radu su prikazani rezultati proučavanja izolovanih sojeva bakterije iz plamenjačom zahvaćenih mladara ukrasnih biljka Sorbus domestica i Sorbus torminalis. Proučavani sojevi su Gram negativni, ne stvaraju fluorescentni pigment na King-ovoj podlozi B, a glukozu metabolišu i oksidativno i fermentativno. Prouzrokuju HR duvana i nekrozu nesazrelih, inokulisanih plodova kruške (viljamova) praćenu obilnom produkcijom bakterijskog eksudata. Prvi simptomi na inokulisanim mladarima biljaka domaćina u očavaju se posle 2-3 dana, ispoljavaju se u vidu vlažnih pega koje se vremenom šire zahvatajući ga u celosti. Na osnovu patogenih, biohemijskih i seroloških odlika (ELISA) proučavanih izolata zaključeno je da patološke promene na ukrasnim biljkama Sorbus domestica i Sorbus torminalis prouzrokuje Erwinia amylovora. To je potvrđeno i primenom molekularne metode (PCR) korišćenjem specifičnog prajmera za bakteriju E. amylovora. Ovo je prvi nalaz E. amylovora na ukrsnim biljkama roda Sorbus u Srbiji.

Bakteriozni rak smatra se jednim od najznačajnijih i najrasprostranjenijih bakterioznih oboljenja vinove loze u svetu. Glavni prouzrokovač ovog oboljenja je vrsta Agrobacterium vitis. Tokom poslednjih nekoliko godina primećena je izražena pojava bolesti u većini vinogradarskih rejona u Srbiji. Stoga je u ovom radu proučena etiologija bakterioznog raka vinove loze u našoj zemlji. Takođe, izvršena je karakterizacija i proučen genetički diverzitet prouzrokovača bolesti. Primenjene su kako klasične bakteriološke, tako i molekularne metode. U radu su takođe korišćeni i brojni referentni sojevi A. vitis poreklom iz međunarodnih kolekcija. Simptomatični uzorci vinove loze prikupljeni su u 22 lokaliteta, raspoređena u gotovo svim vinogradarskim rejonima u našoj zemlji. Izolacija bakterija vršena je na neselektivne podloge sa manitolom i kvaščevim ekstraktom (YMA). Preliminarna identifikacija izolovanih sojeva, izvedena je primenom PCR metode, korišćenjem prajmera specifičnih za plazmidne (virC, virF, virD2, ipt, tms2) i hromozomske (pehA, 23S rRNK) gene. Među izolovanim sojevima identifikovan je tumorogeni A. vitis, a za dalji rad izdvojeno je ukupno 36 reprezentativnih sojeva. Rezultati dobijeni primenom PCR metode potkrepljeni su filogenetskom analizom zasnovanoj na parcijalnoj sekvenci 16S rRNK gena. Taksonomska pripadnost proučavanih sojeva dalje je potvrđena primenom klasičnih bakterioloških metoda. Svi proučavani sojevi ispoljili su morfološke, odgajivačke i biohemijsko-fiziološke odlike karakteristične za vrstu A. vitis. Jedini izuzetak bio je soj KFB 243, koji je i pri ponavljanju testa stvaranja kiseline iz eritritola kao izvora ugljenika ispoljio odlike svojstvene vrsti A. rhizogenes/biovar 2. U cilju provere patogenosti proučavanih sojeva, vršena je inokulacija biljaka vinove loze, suncokreta i paradajza. Od ukupno 36 proučavanih sojeva, 34 su prouzrokovala razvoj tipičnih tumora na inokulisanim biljkama vinove loze. U testu patogenosti na biljkama suncokreta i paradajza, zabeležene su značajne varijacije u intenzitetu simptoma, kako unutar pojedinačnog ogleda, tako i među različitim ponavljanjima... Crown gall is one of the most important and widespread bacterial diseases of grapevine throughout the world. It is predominantly caused by Agrobacterium vitis. During the last few years, serious outbreaks of the disease were observed in major viticultural regions in Serbia. Therefore, the objective of this research was to study etiology of the grapevine crown gall in this country. Characterization and study of genetic diversity of the pathogen were also performed, using conventional bacteriological and molecular methods. Various reference strains of A. vitis originating from international collections were also included. Symptomatic samples of grapevine were collected from 22 localities distributed in almost all viticultural regions in the country. Isolation of bacteria was performed on non-selective media with mannitol and yeast extract (YMA ). Preliminary identification of the isolated strains was performed using PCR method with primers specific for the plasmid (virC, virF, virD2, ipt, tms2) and chromosomal (pehA, 23S rRNA) genes. Among isolated strains, 36 representative tumorigenic A. vitis strains were selected for further study. Results obtained by PCR method were supported by phylogenetic analysis based on partial sequence of 16S rRNA gene. Taxonomic position of studied strains was further confirmed using conventional bacteriological methods. All the strains showed morphological, biochemical and physiological characteristics of A. vitis. The only exception was strain KFB 243, which showed characteristic of A. rhizogenes/biovar 2 in test of acid production from erythritol. Pathogenicity of the strains was studied by inoculation of grapevine, sunflower, and tomato plants. Out of 36 strains studied, 34 caused development of typical tumors on inoculated grapevine. In pathogenicity assay on tomato and sunflower plants, considerable variations in the intensity of symptoms were recorded, both within a single experiment, and among different repetitions. Therefore, use of tomato and sunflower as test plants, despite the rapid symptom development, cannot be recommended as a reliable test...

Bakteriozna plamenjača leske, čiji je prouzrokovač fitopatogena bakterija Xanthomonas arboricola pv. corylina (Xac), najznačajnije je bakteriozno oboljenje ove jezgraste voćke u svetu. U našoj zemlji bolest je prvi put uočena 50-ih godina prošlog veka, nakon čega nije bilo daljih proučavanja ovog patogena. U periodu 2008 - 2011. godine, u nekoliko rejona gajenja leske u Srbiji uočena je pojava simptoma pegavosti lišća, nekroze i uvelosti pupoljaka i letorasta, koji su ukazivali na moguću bakterioznu prirodu oboljenja. Simptomatični uzorci obolelih biljaka prikupljeni su sa različitih lokaliteta u našoj zemlji sa ciljem utvrđivanja etiologije oboljenja. Iz zaraženog biljnog tkiva izolovani su Gram i oksidaza negativni, katalazapozitivni aerobni sojevi bakterija žute boje kolnija na HA podlozi. Za dalji rad odabrano je ukupno 47 reprezentativnih sojeva, koji su proučeni primenom standardnih i savremenih metoda. U istraživanja su uključeni i sojevi poreklom iz Poljske, kao i referentni sojevi Xac poreklom iz međunarodnih kolekcija. Detaljno su proučene morfološke, patogene, biohemijsko-fiziološke, serološke i genetičke odlike sojeva, na osnovu kojih su svi proučavani sojevi identifikovani kao Xanthomonas arboricola pv. corylina. Na podlozi od hranljivog agara (HA) formirane su sitne i sjajne kolonije, dok su one na podlozi obogaćenoj glukozom (GYCA) bile krupne, sluzaste i ispupčene. Svi proučavani sojevi hidrolizuju skrob, želatin i eskulin, koriste glukozu, maltozu, saharozu kao izvor ugljenika, ali ne i sorbitol, metabolišu kvinate, razvijaju se pri 35°C i u prisustvu 2% NaCl. Sojevi su prouzrokovali hipersenzitivnu reakciju na listovima duvana i paradajza i mahunama boranije. Patogenost sojeva potvrđena je testom patogenosti na domaćinu i reprodukcijom simptoma prirodne infekcije. Na osnovu Biolog testa utvrđena je raznovrsnost proučavanih sojeva čiji su metabolički profili bili specifični za rod Xanthomonas... Bacterial blight of hazelnut, caused by Xanthomonas arboricola pv. corylina (Xac), is the most important bacterial disease of this nut crop worldwide. In our country, after the disease was initially described in the 1950s, there were no further data about the research of this pathogen. During the growing season of 2008-2011, symptoms of leaf spots, bud and twig necrosis and dieback, suggesting bacterial infection, were observed on hazelnut plants in several localities in Serbia. In order to study etiology of disease, symptomatic samples of diseased hazelnut plants were collected from different hazelnut-growing regions in the country. Isolations from the diseased tissue resulted in Gram and oxidase negative, catalase-positive, aerobic yellow bacterial colonies. Total of 47 representative strains were selected for further study. Strains isolated from hazelnut in Poland and various reference strains of Xac originating from international collections were also included in research. Based on their morphological, pathogenic, biochemical, physiological, serological and genetic characteristics tested strains were identified as Xanthomonas arboricola pv. corylina. Bacterial colonies formed on NA medium were small and glistening and on glucose amended GYCA medium large, mucous and domed. All strains hydrolyzed starch, gelatin and esculin, used glucose, maltose, sucrose as a carbon source, but not sorbitol, metabolized quinates, developed at 35 °C in the presence of 2% NaCl. The strains induced hypersensitive reaction of tobacco, tomato leaves and bean pods. Pathogenic nature of the strains was confirmed in pathogenicity test on host plants by reproduction of symptoms similar to natural infection. Based on carbon source utilization, metabolic diversity of investigated strains was confirmed. Strain's metabolic profiles were specific for Xanthomonas genus. Serological methods (ELISA i IF test) indicated similarity in serological characteristics among the studied strains, as well as the control Xac strain...