Development of Colletotrichum acutatum in the Foliar Tissue of Strawberry Plants

P oc. 6 h Con . EFPP 2002, P ague. Plan P o ec . Sci., 38 (Special Issue 2), 2002: 429–431 429 Plan P o ec ion Science – 2002 INTRODUCTION Colle o ichum spp. cause an h acnose disease o lea bligh on all signi ican ag icul u al c ops and o namen al plan s a ound he wo ld, being one o he mos des uc i e disease on s awbe y. In ac se e al species o Colle o ichum ha e been epo ed om majo a eas o s awbe ies p oduc ion c op in a la ge amoun o coun ies. Colle o ichum acu a um J. H. Simmonds a acks ae ial pa s o he plan inducing high losses o seedlings and ui s. Spain is he i s p oduce , o esh ma ke , a e Uni ed S a es. Huel a in Andalucia, Sou h-wes e n Spain, con ibu es wi h mo e han 90% o he na ional p oduc ion (DE LOS SANTOS & ROMERO MUÑOZ 1999). In ui ing ields in Huel a C. acu a um cause c own o and plan dea h, ui o , owe bligh and lesions on pe ioles, s olons and lea es, which a e ligh -b own wi h eddish colo in ma gins (DE LOS SANTOS & ROMERO MUÑOZ 1999) and a e di e en o black lea spo caused by C. aga iae and C. gloeospo ioides desc ibed by HOWARD and ALBREGTS (1983) in he summe nu se y. Lesions on olia issue a e no e y equen and se e al s udies ha e sugges ed ha can de elop quiescen in ec ions on s awbe y plan s ha ing a epidemiological ole p io o coloniza ion o ui s and ano he issues. The ini ial s ages o Colle o ichum in ec ion is well s udied and gene ally include conidia adhe e o, and ge mina i e on plan su aces, p oduce ge m- ubes which di e en ia e o o m melanized app esso ia (VAN DYKE & MIMS 1991; BYRNE e al. 1997). On symp omless s awbe y lea es has been epo ed p oduc ion o seconda y conidia o mic ocyclic conidia ion by C. acu a um and may be a signi ican sou ce o inoculum o in ec ions (LEANDRO e al. 2001). The in ec ion p ocess o Colle o ichum species causing la en in ec ion on cowpea lea es e eals ha ungus colonized he mesophyll by in e cellula hyphae, wi hou ini ially isible symp oms (LATUNDE-DADA e al. 1999). Assymp oma ic s ages in some in ec ed plan issues di icul s he knowledge o he p ocess o pa hogenesis. This wo k p e ends o con ibu e o he cla i ica ion o he la ency phase o C. acu a um in he olia issue o he s awbe y plan s h oughou he De elopmen o Colle o ichum acu a um in he Folia Tissue o S awbe y Plan s F. T. ARROYO1*, F. J. MORENO2, P. DAZA2, J. TORREBLANCA2, G. GARCÍA-HERDUGO2, B. DE LOS SANTOS1, C. BARRAU1, C. BLANCO1, M. PORRAS1 and F. ROMERO1 1C.I.F.A. Las To es- omejil, Alcalá del Río, 41200. Se illa, Spain; 2Depa amen o de Biología Celula , Uni e sidad de Se illa, 41012 Se illa, Spain *E-mail: [email p o ec ed] Abs ac S awbe y an h acnose, caused by Colle o ichum acu a um, is one o he mos des uc i e disease o his c op h oughou he wo ld. Assymp oma ic s ages in he plan ha e been he aim o his wo k. Inocula ed lea es samples we e aken a di e en imes and hey we e p ocessed o scanne elec on mic oscopy (SEM) and ansmission elec on mic oscopy (TEM). Conidial de elopmen on bo h su aces lea es was de e mined. The ul as uc u al s udy o ungus pene a ion in o plan cell was cha ac e ized by he o ma ion o esicles o e he ungus pe iphe y and is a mo phological pa am- e e o he in ense memb anes a ic, also could be a e idence o a ansc ip ional ac i i y and enzyma ic cell sec e ion. Di e ences o symp oms on bo h su aces o lea es we e obse ed. Keywo ds: an h acnose; Colle o ichum acu a um; la en in ec ion; ul as uc u e 430 431 Vol. 38, Special Issue 2: 429–431 Plan P o ec ion Science – 2002 Plan P o ec ion Science – 2002 Vol. 38, Special Issue 2: 429–431 s udy o he conidium de elopmen and ul as uc u al in he ela ionship hos -pa hogen. MATERIALS AND METHODS Suscep ible c . Cama osa lea es, we e inocula ed wi h 100 µl d ops o a conidial suspension (106 conidia pe ml dis illed s e il wa e ) o C. acu a um CECT 20240 (ARROYO e al. 2001). Plan s we e incuba ed a 25°C, 100% RH and 16 h ligh pho ope iod in a g ow h chambe . Fo he s udy o he conidium de elopmen on he olia su ace using scaning elec onic mic oscopy (SEM), 1 cm2 pieces o lea we e sampled a 4, 8, 12, 16, 20, 24, 36, 48, 72, 96, and 120 hou s a e -inocula ion (hpi), hen hey we e ixed in glu a aldehyde, dehyd a ed and d ied up by means o he c i ical poin d ying echnique. The s udy o he in ec ion p ocess a ul as uc u al le el using ansmission elec onic mic oscopy (TEM) was ca ied ou in s ips o issue ob ained a 24, 48 and 120 hpi. Samples we e ixed in glu a aldehyde and pos ixed in OsO4 , dehyd a ed wi h ace one and embedded in Epon 812 esin. RESULTS Conidia o C. acu a um we e le adhe ed on o- lia su ace h ough ma ix ex acellula sec e ed by hemsel es. Ge mina ion on bo h, uppe and down olia su aces began wi hin 4 hpi. A 8 hpi some app esso ia we e obse ed in bo h shee s o he lea su ace ha g ew om ge mina i e ubes, gene ally sho ubes o sessile on he uppe lea su ace and longe on he lowe one (Figu es 1A and 1B, espec i ely). A his ime i was also app e- cia ed o ma ion o seconda y conidia (mic ocyclic conidia ion) o igina ed om conidiogenes s uc u es ep esen ed by hypha and conidia phialide (Figu es 1C and 1D). Anas omoses o bo h hypha and p ima y conidia we e de ec ed and hey became o make accumula ions un il 100 µm. F om his s uc u es we e also obse ed p ocesses o conidiogenesis (Figu e 1E). A 72 hpi was pe cei ed on he uppe lea su ace a collapse and lysis o ungi s uc u es ha i was inc eased a 120 hpi (Figu e 1F). Pene a ion o cu icle om app esso ium was ob- se ed a 24 hpi, no being a synch onous p ocess (Figu e 2A). In ec ion in acellula was also mani- es ed in he lowe lea su ace a 24 hpi whe eas in he uppe one i could only be de ec ed hypha inside cell wall (Figu es 2B and 2C, espec i ely). Mac oscopics symp oms on lea es e eals a di e - en ial answe be ween bo h olia su aces, nec o ic spo s o se e al mm in he lowe lea su ace and symp omless in he uppe one. Obse a ion o hese small spo s in he lowe lea su ace seems o be co - Figu es 1. Conidial de elopmen on he lea su ace. A and B, app esso ia on he uppe and lowe lea su aces esp. a 8 hpi. C and D, hypha and conidia phialide a 72 hpi. E, anas omoses on he lowe one a 120 hpi. F, ungi collapse on uppe one a 120 hpi 430 431 Vol. 38, Special Issue 2: 429–431 Plan P o ec ion Science – 2002 Plan P o ec ion Science – 2002 Vol. 38, Special Issue 2: 429–431 ela ed wi h a educed numbe o cells died by hypha in acellula , mainly in eins bu also in he shee (Figu es 3A and 3B). In he uppe lea su ace his pa hogen appea s o su i e inside epide mic cell wall whe e is conside ed an in ense ac i i y o enzyma ic sec e ion, mani es ed by a la ge amoun o endo- and exocy osis esicles and he deg ada ion o wall in his a ea, can exis . The eco e y o he pa hogen is 100% in bo h lea su aces a 120 hpi. Conclusions The in ec ion and de elopmen o pa hogen in he olia issue o s awbe y plan s shows he capaci y o i o be loca ed and o emain asymp oma ic, being able o con ibu e o he disease dispe sion h ough c op ields. Re e ences ARROYO F.T., MORENO F.J., GARCÍA-HERDUGO G., ROMERO F. (2001): Respues a di e encial de Colle o- ichum acu a um y del ejido olia en plan as de e- sa. In: IV Sem. Cien í ico In . P o ección de Plan as, 41ª Reunión Anual de la Sociedad de Fi opa ología de Amé ica-Di isión Ca ibe (APS-CD). Va ade o, Cuba. BYRNE J.M., HAUSBECK M.K., HAMMERSCHMIDT R. (1997): Conidial ge mina ion and app esso ium o ma- ion o Colle o ichum coccodes on oma o oliage. Plan Dis., 81: 715–718. DE LOS SANTOS B., ROMERO MUÑOZ F. (1999): Occu - ence o Colle o ichum acu a um, causal o ganism o s awbe y an h acnose in sou hwes e n Spain. Plan Dis., 83: 301. HOWARD C.M., ALBREGTS E.E. (1983): Black lea spo phase o s awbe y an h acnose caused by C. gloeo- spo ioides. Plan Dis., 67: 1144–1146. LATUNDE-DADA A.O., O’CONNELL R.J., NASH C., LUCAS J.A. (1999): S oma al pene a ion o cowpea (Vigna unguicula a) lea es by a Colle o ichum species causing la en an h acnose. Plan Pa hol., 48: 777–785. LEANDRO L.F.S., GLEASON M.L., NUTTER F.W., WEGULO S.N., DIXON P.M. (2001): Ge mina ion and spo ula ion o Colle o ichum acu a um on symp omless s awbe y lea es. Phy opa hology, 91: 659–664. VAN DYKE C.G., MIMS C.W. (1991): Ul as uc u e o conidia, conidium ge mina ion, and app esso ium de e- lopmen in he plan pa hogenic ungus Colle o ichum unca um. Can. J. Bo ., 69: 2455–2467. Figu es 2. T ansmission elec on mic og aphs o in e ac ion be ween C. acu aum and s awbe y lea es. A, app esso ium on he uppe lea su ace a 24 hpi, which is o ming a pene a ion peg. B, in acellula hypha in he lowe one a 24 hpi. C, ungi inside epide mic cell wall wi h shows a la ge amoun o esicles, i is obse ed deg ada ion o cell wall Figu es 3. A and B, mac o- scopic symp oms on he lowe lea su ace