Phytophthora Root and Crown Rot on Apples in Bulgaria
Phytophthora, prouzrokovač plamenjače korena i krošnje jabuke u Bugarskoj
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Author
Nakova, Mariana
Keywords
Phytophthora root and crown rotApples
Phytophthora sp.
Plamenjaèa korena i krošnje prouzrokovana rodom Phytophthora
jabuka
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Phytophthora is a genus of Oomycota responsible for some of the most serious diseases
with great economic impact (Judelson and Blanco, 2005). While 54 species were found in
the 20th century (Erwin and Ribeiro, 1996) another 51-54 new species have been identified
(Brasier, 2008) since the year 2000. They are spread worldwide and have broad range of host
plants – fruit trees, citrus, forest and park species. Phytophthora can cause serious damages
in orchards and nurseries of apples, cherries, etc. In Bulgaria they have been found first on
young apples and cherries (1998-1999) in Plovdiv region (Nakova, 2003).
Surveys have been done for discovering disease symptoms in Plovdiv and Kjustendil
regions. Isolates have been obtained from infected plant material (roots and stem
bases) applying baiting bioassay (green apples, variety Granny Smith) and/or PARP 10 selective
media. Phytophthora strains were identified based on standard morphology methods –
types of colonies on PDA, CMA, V 8, type and size of sporangia, oogonia and antheridia, and
oospores. Cardial temperatures for their growth were tested on CMA and PDA.
For molecular studies, DNA was extracted from mycelium using the DNA extraction kit.
DNA was amplified using universal primers ITS 6 and ITS 4. Amplification products concentrations
were estimated by comparison with the standard DNA. Sequencing was done at
the Scottish Crop Research Institute (SCRI, Dundee, Scotland).
Phytophthora root and crown rot symptoms first appear in early spring. Infected trees
show bud break delay, have small chlorotic leaves, and branches die all of a sudden. Later
symptoms are found in August-September. Leaves of the infected trees show reddish discoloration
and drop down. Both symptoms are connected with lesions (wet, necrotic in
appearance) at stem bases of the trees.
Disease spread was 2-3% in most gardens, only in an apple orchard in Bjaga (Plovdiv
region) it was up to 8-10%.
Morphologically, the isolates acquired from the apple trees were identified as
Phytophthora cactorum, P. citrophthora and P. cryptogea. Cardial temperatures for their
growth were tested on CMA and PDA.
PCR tests with ITS primers 4 and 6 generated a band at about 800 bP. Consequent
sequencing showed that 2 strains, Bg 1/1 and Bg ½, belong to Phytophthora cryptogea. Phytophthora je rod Oomycota odgovoran za neke od najozbiljnijih bolesti sa velikim
ekonomskim posledicama (Judelson i Blanco, 2005). Tokom 20. veka identifikovane su 54
vrste (Erwin i Ribeiro, 1996), a posle 2000. godine još 51-54 (Brasier, 2008). Ove vrste su raširene
po celom svetu na velikom broju biljaka domaćina – stablima različitih voćaka, citrusnih,
šumskih i parkovskih vrsta. Phytophthora može da prouzrokuje ozbiljnu štetu u voćnjacima
i rasadnicima jabuke, trešnje i drugih voćaka. U Bugarskoj je bolest najpre pronađena
na mladicama jabuke i trešnje (1998-1999) u regionu Plovdiva (Nakova, 2003).
Sprovedena su posmatranja kako bi se otkrili simptomi bolesti u području Plovdiva i
Kjustendila. Izolati su dobijeni sa zaraženog biljnog materijala (korena i korenovog vrata)
„metodom zamke“ u biotestu (zelena jabuka, sorta greni smit) i/ili na selektivnom medijumu
PARP 10. Sojevi Phytophthora su identifikovani standardnim morfološkim metodama
na osnovu: vrsta kolonija na PDA, CMA i V 8 medijumima, tipa i veličine sporangija, oogonija,
anteridija, kao i oospora. Kardijalna temperatura rasta ispitivana je na CMA i PDA
medijumima.
U molekularnim ispitivanjima DNK je ekstrahovana iz micelija pomoću DNK ekstrakcionog
kita. DNK je umnožena pomoću univerzalnih prajmera ITS 6 i ITS 4. Koncentracije produkata
amplifikacije ocenjivane su poređenjem sa standardnom DNK. Sekvencioniranje je
urađeno u Scottish Crop Research Institute (SCRI, Dundee, ŠkotskaPrvi simptomi plamenjače
korena i krošnje uzrokovani rodom Phytophthora pojavljuju se u rano proleće. Kod zaraženih
stabala javlja se okasnelo pucanje pupoljaka i mali hlorotični listovi, a grane iznenada
odumiru. Kasniji simptomi javljaju se u avgustu i septembru. Listovi zaraženog drveća imaju
crvenkastu boju i opadaju. Oba simptoma su u vezi sa lezijama (vlažnog, nekrotičnog izgleda)
u vratu korena drveća.
U većini voćnjaka, rasprostranjenost bolesti je bila 2-3%, osim u jednom voćnjaku u
Bjagi (Plovdiv) gde je raširenost bila 8-10%.
Izolati dobijeni sa stabala jabuke morfološki su identifikovani kao Phytophthora cactorum,
P. citrophthora i P. cryptogea. Kardijalne temperature za njihov rast ispitivane su na CMA
i PDA medijumima.
PCR testovi sa ITS prajmerima 4 i 6 dali su traku od oko 800 bP. Sekvencioniranje je pokazalo
da dva soja, Bg 1/1 i Bg 1/2, pripadaju vrsti Phytophthora cryptogea.