Rod Botrytis i vrsta Botrytis cinerea: patogene, morfološke i epidemiološke karakteristike

Species of the genus Botrytis occur wherever their hosts are grown, ranging from cold areas of Alaska to warm and dry areas in Israel. They have a necrotrophic life style which is often associated with phenology of the host plant. The genus comprises 22 species, most of which have a narrow host range. Polifagous species Botritys cinerea, a causal agent of grey mould disease, is the most important and the most extensively studied representative of this genus. More than 350 papers related to all aspects of the research of this necrotrophic pathogen are published each year. In this paper up-to-date knowledge about pathogenic, morphological and epidemic characteristics of the genus Botrytis and, particularly, species B. cinerea are summarized. Symptoms caused by B. cinerea on various plant species and various plant parts are shown. Morphological and genetic variability of the species is described. The possible mechanisms of variability, as well as the attempts to divide the species into Group I (B. „pseudocinerea“) and Group II (B. cinerea „sensu-stricto“) are pointed out.Vrste roda Botrytis rasprostranjene su svuda gde se gaje ili su prisutne njihove biljke-domaćini – od hladnih zona Aljaske do toplih i suvih područja u Izraelu. Imaju nekrotrofni životni ciklus koji je najčešće tesno povezan sa fenologijom domaćina. Rod obuhvata 22 vrste od kojih većina ima uzak krug domaćina – parazitira biljke jednog roda ili jedne familije. Polifagna vrsta Botrytis cinerea, prouzrokovač sive truleži, najznačajniji je i najproučavaniji predstavnik ovog roda. Godišnje se objavi više od 350 radova koji se odnose na sve aspekte istraživanja ovog nekrotrofnog patogena. U ovom radu je dat pregled dosadašnjih saznanja o patogenim, morfološkim i epidemiološkim karakteristikama vrsta roda Botrytis, a posebno vrste B. cinerea. Detaljno su obrađeni simptomi oboljenja koje prouzrokuje B. cinerea na različitim biljnim vrstama i različitim delovima biljaka, morfološka i genetička varijabilnost vrste. Takođe, ukazano je i na moguće izvore varijabilnosti, kao i na pokušaje podele vrste na dve grupe – Grupu I (B. „pseudocinerea“) i Grupu II (B. cinerea „sensu-stricto“)

Identifikacija Eutypa lata, parazita vinove loze

The phytopathogenic fungus Eutypa lata (Pers.: Fr.) Tul. and C. Tul., the causing agent of eutypa dieback, has been increasingly often identified in recent times as a cause of grapevine disease. It was first discovered and identified in Australian vineyards (Carter, 1973), where it represented one of the most dangerous fungus pathogens of this plant. A few years later it was discovered in European vineyards as well. This polyfagous fungus, known originally as E. armeniaca (Honsf. & Carter), was first discovered on apricot, on which it caused the ggummosis disease h. In Serbia, Eutypa lata has not been determined officially. However, bearing in mind the form of its spreading (anemochory), as well as the fact that our country is a major producer of grape and fruit, we need to pay special attention to this dangerous pathogen since there are indications that it is already present in our vineyards. During the period between 2003 and 2005, an inspection of a great number of vineyards in the areas of Vršac, Fruška Gora and Kruševac, was conducted. Many of them had grapevines with typical eutypa dieback symptoms. The aim of the inspection was to find grapevines with this disease, to mark them and take samples for laboratory analysis. Marking suspicious grapevines enabled us to monitor the volume of symptoms, as well as other changes on grapevines. Different colours were used for markings, according to the principle gsame colour . same year h The procedure revealed that the average period between early and mild disease symptoms and extreme changes, including withering of entire vines, was 2 to 3 years. The signs of eutypa dieback on diseased grapevines are manifested: on leaves in the form of chlorosis, twisting, necrosis of the edges, drying out and falling off; on shoots, where the shortening of internodia is noticable, as well as colour change and gzig-zag h distribution of internodes; on blossoms and clusters, where absence of flowering, partial bareness and irregular berry size are observed; and on trunks, where necrosis appears on the cross section in a typical gV h form. These studies also established that there is not a single grapevine plant with symptoms indicating eutypa dieback or a similar disease that is older than 8-10 years. The research results, based on examination and monitoring of disease development in the studied vineyards in previous years, indicate that the fungus Eutypa lata, the inducer of grapevine eutyposis, is most probably present in Serbia, and that our viticulture is facing yet another serious threat. In most European and other viticultural centers, highly specialized research teams are being organised to deal with the phenomenon of eutypa dieback, and its inducer Eutypa lata, which is more and more frequently classified as a top priority disease of wooden tissue, especially considering that the economic losses are very significant and occur in all vineyards, regardless of their location. Therefore, in order to prevent infection and fight the disease successfully, we need to take immediate measures of integral protection of grapevine without any delay.Među prouzrokovače bolesti vinove loze u novije vreme sve češće se ubraja i fitopatogena gljiva Eutypa lata (Pers.: Fr.) Tul. i C. Tul., uzročnik eutipoze. Prvi put je otkrivena i identifikovana u australijskom vinogorju (Carter, 1973), gde predstavlja jednu od najopasnijih mikoza ove biljke. Nekoliko godina kasnije otkrivena je i u vinogorjima Evrope. Ranije je ova polifagna gljiva, pošto je prvi put nađena na kajsiji, bila poznata pod imenom E. armeniaca (Honsf. & Carter), na kojoj prouzrokuje „gumoznu bolest”. U Srbiji Eutypa lata još nije zvanično konstatovana. Međutim, imajući u vidu način njenog širenja (anemohorija), kao i činjenicu da je naša zemlja značajan proizvođač grožđa i voća, potrebno je obratiti naročitu pažnju na ovog opasnog patogena, jer ima indicija da je prisutan i u našim vinogorjima. U periodu 2003-2005. godine obavljen je pregled većeg broja vinograda u vršačkom, fruškogorskom i kruševačkom vinogorju. U mnogima od njih nađeni su čokoti sa tipičnim simptomima eutipoze. Cilj ovih ispitivanja bio je pronalaženje ovako obolelih čokota, njihovo obeležavanje, kao i uzimanje uzoraka za dalje laboratorijske analize. Obeležavanje sumnjivih čokota omogućilo nam je da pratimo jačinu izraženosti simptoma, kao i sve ostale promene tih čokota. Za obeležavanje su korišćene različite boje, a primenjen je princip: jedna boja – jedna godina. Ovim postupkom je ustanovljeno da od pojave početnih, blažih simptoma oboljenja, pa do pojave izuzetno izraženih promena, uključujući i izumiranje celih čokota, treba da prođe 2 do 3 godine. Znaci eutipoze na napadnutim čokotima manifestuju se na lišću u vidu hloroze, kovrdžanja, nekroze oboda, sušenja i opadanja listova, na lastarima se uočava skraćivanje internodija, promena boje, „cik-cak” raspored internodusa, na cvastima i grozdovima primetno je izostajanje cvetanja, rehuljavost i neujednačenost veličine bobica, dok se na drvetu javlja nekroza na poprečnom preseku u obliku slova V, tipičnom za ovu bolest. U ovim ispitivanjima je takođe utvrđeno da nema zasada vinove loze starijeg 8-10 godina a da se ne nađu čokoti sa ovakvim simptomima koji upućuju na eutipozu ili slično oboljenje. Rezultati istraživanja dobijeni na osnovu obavljenih pregleda i praćenja razvoja simptoma oboljenja proteklih godina u proučavanim vinogorjima, upućuju na zaključak da je gljiva Eutypa lata, prouzrokovač eutipoze vinove loze, najverovatnije prisutna u Srbiji, i da je naše vinogradarstvo suočeno sa još jednom ozbiljnom opasnošću. U većini evropskih i ostalih vinogradarskih centara organizovani su timovi visokospecijalizovanih stručnjaka koji se bave istraživanjem fenomena eutipoze i njenog prouzrokovača - Eutypa lata, jer se ona sve češće ubraja u najznačajnije bolesti drvenastog tkiva, posebno kada se uzme u obzir da su ekonomski gubici veoma značajni i da se javljaju u svim vinogradima bez obzira na njihovu lokaciju. Zbog toga, a u cilju sprečavanja zaraze, kao i uspešne borbe protiv ove bolesti, neophodno je, bez odlaganja, primeniti mere integralne zaštite vinove loze

Da li je niska efikasnost fungicida uvek posledica razvoja rezistentnosti u populacijama patogena?

Efficacy of four fungicides with different modes of action (vinclozolin, pyrimethanil, benomyl and fenhexamid) in control of B. cinerea in raspberry, was investigated in the paper. The trials were conducted at two localities in commercial raspberry plantations. In the case of unsatisfactory fungicide efficacy, qualitative and/or quantitative test of the susceptibility of the isolates to particular fungicide was performed, to determine whether the low efficacy is a consequence of resistance development in the pathogen population. At both localities, pyrimethanil and fenhexamid demonstrated the highest efficacy (73.2-89.6%), while the efficacy of vinclozolin was statistically significantly lower (48.7-63.4%) at both localities. However, qualitative and quantitative test of susceptibility to vinclozolin showed that all the isolates were susceptible to vinclozolin and that the reason for unsatisfactory efficacy should be primarily sought in inadequate fungicide application.U radu je ispitivana efikasnost četiri fungicida različitog mehanizma delovanja (vinklozolin, pirimetanil, benomil i fenheksamid) u zaštiti maline od B. cinerea. Ogledi su izvedeni na dva lokaliteta u komercijalnim zasadima maline. U slučajevima nezadovoljavajuće efikasnosti fungicida, kvalitativnim i/ili kvantitativnim testom osetljivosti izolata in vitro, utvrđeno je da li je niska efikasnost posledica razvoja rezistentnosti populacije patogena. Na oba lokaliteta, najveću efikasnost su ispoljili pirimetanil i fenheksamid (73,2-89,6%), dok je efikasnost vinklozolina bila statistički značajno niža (48,7-63,4%). Međutim, kvalitativni i kvantitativni test osetljivosti na vinklozolin je pokazao da su svi izolati u kategoriji osetljivih na vinklozolin, te da razlog za nezadovoljavajuću efikasnost ovog fungicida treba tražiti pre svega u neadekvatnoj primeni fungicida

Suzbijanje Botrytis cinerea i problem rezistentnosti na fungicide

Botrytis cinerea, the causal agent of grey mould, greatly affects fruit, grapevine, vegetable and ornamental crops production. It is a common causal agent of diseases in plants grown in protected areas, as well as fruit decay during storage and transport. The fungus invades almost all parts of the plant in all developmental stages, and the symptoms are usually described as grey mould, grey mildew, brown rot and seedling blight. The paper reviews the current knowledge on control possibilities of this necrotrophic pathogen. The attention is particularly paid to the mode of action of novel fungicides and to the problem of resistance. It is pointed out that by limiting the number of treatments in the growing season, avoiding the use of only one fungicide with a high risk for resistance development, appropriate application rate and timing, using mixtures of pesticides with different modes of action, as well as by alternative use of pesticides from different resistance groups, a longterm preservation of pesticide efficacy is provided.Botrytis cinerea, prouzrokovač sive truleži, značajno ugrožava proizvodnju voća, grožđa, povrća i ukrasnog bilja. Često izaziva bolesti biljaka gajenih u zaštićenom prostoru, kao i trulež plodova tokom skladištenja i transporta. Gljiva parazitira skoro sve delove biljke u svim fazama razvoja, a simptomi se najčešće opisuju kao siva trulež, siva plesan, palež, mrka trulež, a zapaženo je i poleganje klijanaca. U radu je dat pregled dosadašnjih saznanja o mogućnostima suzbijanja ovog nekrotrofnog patogena. Posebna pažnja je posvećena mehanizmu delovanja fungicida novije generacije kao i problemu rezistentnosti. Istaknuto je da se ograničenjem broja tretiranja u toku jedne vegetacije, izbegavanjem upotrebe samo jednog fungicida visokog rizika za razvoj rezistentnosti, odgovarajućom dozom i vremenom primene, upotrebom mešavina pesticida različitog mehanizma delovanja kao i alternativnom primenom pesticida iz različitih rezistentnih grupa omogućuje dugotrajno očuvanje efikasnosti pesticida

Botrytis cinerea na malini II: Brzina rasta i virulentnost izolata

Growth rate and virulence of 130 isolates of Botrytis cinerea, derived from raspberry fruit originating from six commercial fields in a raspberry growing region of Serbia and classified in two morphological and four genetic groups were studied. The results showed significant differences in mycelial growth rate among the isolates. The highest and lowest recorded growth rates were 24.5 mm/day and 8.4 mm/day, respectively, while the growth rate of most isolates ranged from 15.8 to 21.8 mm/day. The growth rate of isolates that belong to different morphological and genetic subgroups varied similarly. Furthermore, growth rate intervals of all subgroups overlapped, suggesting that the groups cannot be distinguished based on growth rates of the isolates contained. The studied B. cinerea isolates exibited different levels of virulence towards vine, sunflower and raspberry leaves, while an analysis of variance revealed that both the isolates and the inoculated host species were significant sources of variation (P<0.01). Sunflower and raspberry leaves were significantly more sensitive than vine leaves. However, correlation between isolate virulence and different hosts was not found.U radu su predstavljeni rezultati proučavanja brzine rasta i virulentnosti 130 izolata Botrytis cinerea, dobijenih iz obolelih plodova maline poreklom sa šest lokaliteta iz područja komercijalnog gajenja maline u Srbiji i razvrstanih u dve morfološke i četiri genetičke grupe. Rezultati su pokazali da je razlika između izolata u brzini rasta statistički značajna. Najveći zabeleženi porast bio je 24,5 mm/dan, dok je najmanji porast iznosio 8,4 mm/dan. Utvrđena su slična variranja u porastu izolata koji pripadaju različitim morfološkim ili genetičkim grupama. Drugim rečima, rasponi brzine rasta izolata iz različitih morfoloških i genetičkih grupa međusobno se preklapaju, što ukazuje da ovaj parametar nije pogodan za razvrstavanje izolata u grupe. Proučavani izolati su ispoljili različit nivo virulentnosti za listove vinove loze, suncokreta i maline, dok je analiza varijanse pokazala da su i izolati i domaćini statistički značajan izvor variranja (P<0,01). Listovi suncokreta i maline bili su značajno osetljiviji od listova vinove loze. Međutim, korelacija u virulentnosti izolata za različite domaćine nije ustanovljena

Botrytis cinerea na malini u Srbiji I: Morfološka i molekularna karakterizacija

Morphological and molecular characterisation of 130 isolates of Botrytis cinerea, derived from raspberry fruit originating from six commercial fields in a raspberry growing region of Serbia (locations: Požega, Prilike, Arilje, Ivanjica, Šabac and Valjevo) was performed. The results showed that all isolates formed white, uniform, aerial mycelia with entire margin on PDA medium. First morphological differences among the isolates appeared after six days of incubation. Three-week old isolates were grouped into eight distinct morphological types – four mycelial and four sclerotial. Mostly, they were of sclerotial type (81.5%) and the most frequently found was an S3 type, which formed large irregularly placed sclerotia. This type was dominant in five of six investigated locations and represented 45-65% of the isolates. The least frequent was the mycelial type M3 (0.7% of the isolates) characterized by mycelial masses. The presence of Boty and/or Flipper transposons was detected in isolates originating from all investigated locations. It was discovered that the B. cinerea population in raspberry in Serbia, besides the well-described genetically isolated sympatric species transposa (43.1%) and vacuma (10.8%), contains also another two, boty (44.6%) and flipper (1.5%) species with only one transposon (either Boty or Flipper) in the genome. In addition, it was revealed that all isolates from raspberry collected in Serbia, transposa, vacuma, boty or flipper, are sensitive or weakly resistant to fenhexamid and therefore belong to the B. cinerea genetical Group II.U radu su predstavljeni rezultati morfološke i molekularne karakterizacije 130 izolata Botrytis cinerea, dobijenih iz obolelih plodova maline poreklom sa šest lokaliteta iz područja komercijalnog gajenja maline u Srbiji (Požega, Prilike, Arilje, Ivanjica, Šabac i Valjevo). Utvrđeno je da u početnim fazama razvoja na KDA podlozi svi izolati B. cinerea formiraju belu, uniformnu, rastresitu, vazdušnu miceliju ravnog oboda. Razlike među izolatima počinju da se javljaju posle inkubacije od šest dana. Na osnovu izgleda kolonije tri nedelje od zasejavanja, izolati su razvrstani u osam morfoloških tipova – četiri micelijska i četiri sklerocijska. Većina izolata je formirala kolonije sklerocijskog tipa (81,5%), a najzastupljeniji je bio tip S3 sa krupnim, nepravilno raspoređenim sklerocijama, koji je dominirao u pet od šest proučavanih populacija patogena i predstavljao 45-65% izolata. Najređi je bio micelijski tip M3 (0,7% izolata) koji se odlikuje nakupinama vazdušne micelije. Prisustvo transpozona Boty i/ili Flipper otkriveno je u genomu izolata sa svih lokaliteta. Utvrđeno je da u populaciji patogena na malini u Srbiji, osim genetički izolovanih subpopulacija transposa (43,1%) i vacuma (10,8%), postoje još dve – boty (44,6%) i flipper (1,5%) sa izolatima koji sadrže samo jednu vrstu transpozona u genomu. Istraživanje je takođe pokazalo da su svi izolati B. cinerea na malini u Srbiji, bilo da su transposa, vacuma, boty ili flipper, osetljivi ili slabo rezistentni na fenheksamid i da, prema tome, pripadaju genetičkoj Grupi II B. cinerea

Vrste roda Monilinia na jabučastim i koštičavim vrstama voćaka

Different species of the genus Monilinia are common plant pathogens that endanger pome and stone fruit production worldwide. In Serbia, two species of this genus are widely distributed – M. laxa and M. fructigena, while M. fructicola, which is officially on the A2 EPPO List of quarantine pest organisms in Europe and on the 1A part I List of quarantine pest organisms in Serbia, has so far been detected only on stored apple and nectarine fruits. The most important control measures against these pathogens include chemical control in combination with adequate cultural practices, particularly under favourable conditions for disease development. Concerning that species of this genus can cause significant economic losses, knowledge of the pathogen biology, disease epidemiology and pathogen-host interactions is a necessary prerequisite for stable and profitable production of pome and stone fruits.Različite vrste roda Monilinia dobro su poznati biljni patogeni koji ugrožavaju proizvodnju jabučastog i koštičavog voća širom sveta. U našoj zemlji široko su rasprostranjene dve vrste ovog roda – M. laxa i M. fructigena, dok je M. fructicola koja se zvanično nalazi na A2 EPPO listi karantinskih štetnih organizama u Evropi i na A1 listi karantinskih štetnih organizama u našoj zemlji, do sada detektovana samo na uskladištenim plodovima jabuke i plodovima nektarine. Najznačajnija mera borbe protiv ovih patogena je hemijsko suzbijanje uz primenu adekvatnih agrotehničkih mera, naročito u godinama povoljnim za razvoj oboljenja. S obzirom da vrste ovog roda mogu prouzrokovati velike ekonomske gubitke, stepen poznavanja biologije patogena, epidemiologije bolesti i odnosa parazit-domaćin predstavljaju neophodan preduslov stabilne i profitabilne proizvodnje jabučastog i koštičavog voća

Razvoj formulacije na bazi etarskog ulja timijana i njen uticaj na Monilinia fructigena

Antifungal activity of thyme essential oil against Monilinia fructigena and development of an effective and stable oil formulation for agricultural use were studied in this paper. Bioactivity of the oil itself and its developed formulation, an emulsifiable concentrate (EC), was tested in vitro and in vivo. In vitro experiments, using a slightly modified agar overlay technique, showed that the initially emulsified thyme essential oil, as well as the developed formulation, significantly inhibited mycelial growth of M. fructigena in vitro. Experiments in vivo, performed on inoculated apple fruits, revealed that the formulation successfully decreased oil evaporation from the treated area and provided a significant level of M. fructigena suppression, 64.7-72.1% compared to the control. To our knowledge, an EC formulation of thyme essential oil for agricultural uses had never been developed before. The presented results are initial findings and evaluation of product activity should be continued in the field to determine its efficacy and activity spectrum, and to estimate the economic aspect of its use.U radu je proučavana antifungalna aktivnost etarskog ulja timijana na Monilinia fructigena, kao i mogućnost razvoja efikasne i stabilne formulacije ulja za upotrebu u poljoprivredi. Bioaktivnost etarskog ulja kao i razvijene formulacije – koncentrata za emulziju (EC) testirana je u ogledima in vitro i in vivo. Ogledi in vitro, izvedeni po delimično modifikovanoj metodi testiranja antimikrobne aktivnosti na površini agara, pokazali su da i inicijalno emulgovano ulje i razvijena formulacija značajno inhibiraju porast micelije izolata M. fructigena. U ogledima in vivo na inokulisanim plodovima jabuke pokazano je da je procesom formulisanja značajno smanjena isparljivost ulja sa tretirane površine i postignuta inhibicija razvoja mrke truleži ploda 64,7-72,1% u poređenju sa kontrolom. Koliko nam je poznato, u ovom radu je po prvi put razvijena EC formulacija etarskog ulja timijana za upotrebu u poljoprivredi. Dobijeni rezultati predstavljaju početak istraživanja koje treba dopuniti rezultatima ispitivanja efekata razvijene formulacije u uslovima praktične primene, proučavanja spektra njenog delovanja, kao i ekonomske isplativosti njene primene

Molekularna detekcija Monilinia fructigena prouzrokovača truleži ploda dunje

Species of the genus Monilinia are important causal agents of fruit rot on pome and stone fruits in Serbia. The pathogen is very harmful, especially on small properties and cottage plantations where intensive control measures are not applied. Quince is important host for the pathogens of this genus. During spring 2010, intensive occurrence of mummified fruits overwintering on branches of the quince was observed. The pathogen was isolated using standard phytopathological methods. Pathogenicity of eight obtained isolates was tested by artificial inoculation of injured apple fruits. Identification was performed according to pathogenic, morphological and ecological properties, and was confirmed by Multiplex polimeraze chain reaction, PCR. All the isolates studied caused brown rot on inoculated apple fruits. The isolates form light yellow colonies with lobate margins, with single-celled, transparent, elliptical or oval conidia in chains, regardless temperature or light presence. Sclerotia are observed in 14 days old cultures. The highest growth rate of most of the isolates is at 27°C and in dark. Based on studied pathogenic, morphological and ecological characteristics, it was found that the Monilinia fructigena is causal agent of brown rot of quince. Using specific primers (MO368-5, MO368-8R, MO368-10R, Laxa-R2) for detection of Monilinia species in Multiplex PCR reaction, the expected fragment 402 bp in size was amplified, which confirmed that the studied isolates belonged to the species M. fructigena.Vrste roda Monilinia su značajni prouzrokovači truleži ploda jabučastog i koštičavog voća u našoj zemlji. Parazit nanosi velike štete, posebno na malim posedima i u vikend zasadima u kojima se ne primenjuje redovna zaštita. Dunja je značajan domaćin patogena ovog roda. Tokom proleća 2010. godine ustanovljena je intenzivnija pojava mumificiranih plodova prezimelih na granama. Cilj ovog rada bio je da se identifikuje prouzrokovač mumifikacije plodova dunje, prikupljenih sa različitih lokaliteta. Patogen je izolovan primenom standardnih fitopatoloških metoda. Patogenost osam dobijenih izolata proverena je veštačkom inokulacijom povređenih plodova jabuke. Identifikacija je obavljena na osnovu patogenih, morfoloških i ekoloških osobina, a potvrđena je primenom Multiplex PCR. Svi proučavani izolati su na inokulisanim plodovima jabuke prouzrokovali trulež smeđe boje. Proučavani izolati na KDA podlozi formiraju svetlo-žute kolonije režnjevitog oboda. U kulturi gljiva, nezavisno od uticaja temperature i prisustva ili odsustva svetlosti, formira jednoćelijske, providne, eliptične ili ovalne konidije u nizovima. Sklerocije se uočavaju u kulturama starosti 14 dana. Većina izolata ostvaruje najbolji porast na temperaturi od 27°C i u odsustvu svetlosti. Na osnovu proučenih patogenih, morfoloških i odgajivačkih odlika, utvrđeno je da izolati pripadaju vrsti Monilinia fructigena. Korišćenjem specifičnih prajmera (MO368-5, MO368-8R, MO368-10R, Laxa-R2) za detekciju vrsta roda Monilinia u Multiplex PCR reakciji, amplifikovan je očekivani fragment veličine oko 402 bp čime je potvrđeno da proučavani izolati pripadaju vrsti M. fructigena