Bulletin OEPP/EPPO Bulletin 23,461-412 (1993)

Chemical control of the olive rot caused by Glomerella cingulata'

by A. M. PENNISI, G. E. AGOSTEO and S. GRASS0 Istituto di Difesa delle Piante, Universitk di Reggio Calabria,

Piazza S. Francesco di Sales 2, 89061 Gallina, Reggio Calabria (Italy)

The effect of various fungicides on Glomerella cingulata, the causal agent of a serious rot of olive fruits, was evaluated both in uitro and in the field. The following fungicides were tested: copper oxychloride, Bordeaux mixture, ziram, myclobutanil, hexaconazole, metiram, bitertanol and benomyl. Field trials were carried out in Calabria (southern Italy) for 2 years on two olive cultivars, Ottobratica (susceptible) and Sinopolese (resistant). The minimum effective concentrations of bitertanol, hexaconazole, myclobutanil, benomyl, metiram, ziram, and copper oxychloride on mycelial growth in uitro were < 5, i 50, i 50, i 1000, < 1000, i 1000, i 1000 p g ml-', respectively. The ED90 values for conidial germination were 0.01, 0.02, 50, 50, 1000, 1000, and 1000 pg ml-' of benornyl, ziram, myclobutanil, copper oxychloride, bitertanol, hexaconazole and metiram. respectively. In field trials, Bordeaux mixture and copper oxychloride proved to be the most effective products; however zirarn, myclobutanil, bitertanol, metiram, and hexaconazole reduced the incidence of fruit rot significantly.

Introduction

Fruit rot caused by Glomerella cinguluta (Stoneman) Spaulding & Schrenk (anamorph: Colletotrichum gloeosporioides) is the most serious disease of olive in the humid areas of Calabria (southern Italy) characterized by environmental conditions favouring epidemic development (Ciccarone, 1950; Graniti, 1955; Martelli, 1959, 1960). The first infections on olive fruits appear in autumn and also cause the dieback of twigs and branches (Graniti & Laviola, 1981; Cirulli et al., 1981). Control of the disease in the field has traditionally been based on copper fungicide treatments, such as copper oxychloride or Bordeaux mixture. However olive growers have refrained from using these fungicides to spray the trees because they are phytotoxic.

The aim of the present work was to test the efficacy of new systemic fungicides as an alternative to copper compounds. Some of these were ergosterol-biosynthesis-inhibitor (EBI) fungicides known to be effective against Colletotrichum spp., G. cinguluta and other Ascomycetes (Kuck & Scheinpflug, 1986; Siddi, 1989; Maggioni et al., 1991).

Materials and methods

The following fungicides were assessed: Bordeaux mixture (Siaram, 13.5% Cu), copper oxychloride (Cupravit, 50% Cu), ziram (Fuclasin, 86.5% a.i.), metiram (Polyram, 7 I .2%1 a.i.), benomyl (Benlate, 50% a i ) , myclobutanil (Thiocur 12, 13.4% a i ) , hexaconazole (Anvil, 3% a.i.) and bitertanol (Baycor, 25% a.i.).

Laboratory tests

The isolate of G. cingulutu used in vitro tests was obtained from a rotted olive fruit. The

Paper presented at the Joint MPU/EPPO Conference on Olive Diseases, Sounion (GR), 1992-05-05/08.

467

468 A . M . Pennisi et al.

concentrations assessed to evaluate fungicide activity on mycelial growth and conidial germination were 0.0, 0.001, 0.01, 0.1, I , 5, 10, 50, 100, 1000 p g a.i. ml-I. Fungicides were suspended in sterile deionized water and added to media after autoclaving and cooling to 45°C. Potato dextrose agar (PDA Difco, 39 g per litre of distilled water) was used as medium and was poured (25 ml per plate) into 100-mm disposable Petri dishes. Plates were inoculated on the same day with 6-mm agar plugs collected from 7-day-old colonies of the test fungus. For each of the concentrations tested, five replicate plates of PDA were inoculated. Two orthogonal diameters of the colonies, developed after 7 days’ incubation at 24 f 1“C, were measured. To determine the inhibitory effect of the fiingicides on conidial germination, aliquots (80 pl) of suspensions at varying concentrations diluted in sterile distilled water were placed on sterile glass slides and dried at 45°C. Aliquots (80 pl) of a conidial suspension in Gorter solution (Gorter, 1961; Martelli, 1962) containing about 2.5 x lo6 conidia per ml were put on each glass slide. After 24-h incubation at 24f 1”C, the conidia were stained with lactophenol-cotton blue. For each of the concentrations tested, three replicate glass slides were prepared. The percentage of conidia showing normal germ tubes was assessed by microscopic observation of 100 conidia per replicate. Yo inhibition of both mycelial growth and germination for each fungicide tested was calculated with respect to the control.

Field trials

Field trials were carried out in the Gioia Tauro area in Calabria in 1989/1990 and 1991/1992 on the two olive cvs Ottobratica and Sinopolese. In these years, a split-split plot design was set up in different groves, and every treatment was replicated 3 times. The following treatments were scheduled: ( 1 ) one spray treatment a year, applied in September; (2) two spray treatments a year, applied in Septembcr and October; (3) three spray treatments a year, applied in September, October and November (only in the 1991 experiment). All applications were carried out in the last 10 days of the month by high-volume application from a tractor-mounted sprayer, using 25 litre per tree a t 30 atm (30.9 kg cm-2). The fungicides applied in the first year were Bordeaux mixture a t 135, ziram at 173, benomyl at 50 and myclobutanil at 6.7 g a.i. per 100 1. In the second

+ Zlram

-E Metlram

+ Benomyl

* Bltertanol

* Hexaconarole

0.001 0.01 0.1 1 10 100 1,000

Concentration (Irglml)

Fig. 1. Dose-response relationships showing the fungicide sensitivity of conidial germination in Glomerellu cingulutu. Relations dose-reponse rnettant en evidence la sensibilite aux fongicides de la germination des conidies de G. cingulcitu.

Chemical control of G. cingulata 469

year the olive trees were sprayed with copper oxychloride at 200, metiram at 142, myclobutanil a t 6.7, hexaconazole at 3 and bitertanol a t 25 g a.i. per 100 1. The effectiveness of the treatments was evaluated as O h infected fruits, by collecting from each tree one lot of 500 drupes in the first 10 days of December, January, and February. Three sample collections were made in accordance with the local custom of allowing the olives to fall naturally into nets left under the trees.

Results

In the in uitro test, copper oxychloride and ziram showed maximum activity on conidial germination at a concentration between 50 and 100 pg ml-I. All the other products completely inhibited conidial germination only at concentrations of 1000 p g ml-l or above (Fig. 1) . Bitertanol inhibited mycelial growth at the concentration of 5 p g ml-l, while the inhibitory concentration of both hexaconazole and myclobutanil was 50 p g ml-' (Fig. 2).

The two olive cultivars tested in the field showcd differing susceptibility to the disease. Cv. Ottobratica was more susceptible than cv. Sinopolese. Disease incidence was 30 and 44% of rotted olive fruits in 1990 and 1992 for non-treated cv. Ottobratica, and 18 and 10% respectively for cv. Sinopolese (Table 1). The period of picking also influenced the incidence of olive rot. Delay in the olive harvest significantly increased the incidence of fruit rot. This was especially true in the second year, since disease incidence in the control rose from 17% in December to 39 "Al in February (Table 2). Similarly the difference in olive rot susceptibility between cvs Sinopolese and Ottobratica was more pronounced with late harvesting (Table 2) due to the higher incidcnce of the disease in February.

Examination of the olive fruits harvested in the field showed that copper fungicide, ziram, and metiram were the most effective products tested. They significantly reduced disease incidence relative to the control (Table 3) and their activity increased with the number of treatments. The results in Table 4 show the effect of multiple sprays on %I drupes infected during the 1991/1992 field trials. Analysis of variance showed that the interactions between fungicides, olive cultivars, number of treatments, and picking period were significant.

. . . . . . . . . . . .

1 10 100

-Cu oxychloride

+ Zlram

* Metlram

+ Benomyl

* Bitertanol

+- Hexaconazole

* Myclobutgnll

Concentration @g/ml)

Measurements taken from 7day old colonies grown on PDA at optimum growth temperature (25'C)

Fig. 2. Dose-response relationships showing the fungicide sensitivity of mycelial growth in Glomerella cingulata. Relations dose-reponse mettant en evidence la sensibilite aux fongicides de la croissance mycelienne de G. cingulata.

A . M . Pennisi et al.

Table 1. Incidence of fruit rot due to Glonierellu cinguluta on olive cvs Ottobratica and Sinopolese in relation to harvest period",b Incidence de la pourriture des fruits due a G. cingulura chez les cultivars d'olivier Ottobratica et Sinopolese, en fonction de I'tpoque de la recolte

Year 1989/1990

Harvest period

Year 1991/1992

Total for Olive cvs December January February year

Ottobratica 24" 30" 34" 30" Sinopolese 1 Oh I 5b 28b I gh

Total 17 22 31 24

Total for December January February year

29" 41" 62" 44" 6' gb 17b 1 Ob

17 24 39 27

a Values are the percentage of infected olives from untreated control trees; letters do not differ significantly (P=0.05).

values followed by the same

Table 2: Effect of harvest time on Influence de l'epoque de la rttcolte sur le '% de pourriture des olives par G. cingulata dans les essais de plein champ de 1991/1992

olive rot due to Glomerellu cingulutu in 1991/1992 field trials"

Fungicides 199 1-1 2 1992-0 1 1992-02

Copper oxychloride 4 a 10 a 16 a Metiram l b I 1 a 16 a Myclo butanil 13 c 21 b 29 c Hex a c o n a z o 1 e 11 c 20 b 26 c Bitertanol 14 cd 19 b 21 b Control 17 d 24 c 39 d

a Values followed by the same letters do not differ significantly (P=0.05).

Table 3. Effect of fungicide applications on %I infection of olives by Clumerellu cingulutu" Influence des traitements fongicides sur le % d'infection des olives par G. cingulutu

Fungicides Year 1989/1990 Year 1991/1992

Bordeaux mixture 18 a Copper oxychloride 10 a Ziram 19 ab Metiram 11 ab Benomyl 21 b Myclobutanil 20 ab 21 c Hexaconazole 19 c Bitertanol 18 c Control 24 c 27 d

a Values followed by the same letters do not differ significantly (P=0.05)

Chemical control of G. cingulata 47 1

Table 4. Effect of number of fungicide treatments on YO infection of olives by Glomerellu cinguluta in 1991/ 1992 field trialsa Influence du nombre de traitements fongicides sur le % d’infection des olives par G . cingulutu dans les essais de plein champ de 1991/1992

Number of sprays

Fungicides One Two Three

Copper oxychloride 15 a 10 a 5 a Metiram 17 a 10 a 6 a Myclobutanil 24 bc 20 bc 12 b Hexaconazole 22 b 22 c 13 b Bitertanol 23 bc 18 b 14 b Control 35 d 26 d 19 c

a Values followed by the same letters do not differ significantly (P=0.05).

Discussion The different susceptibility to olive rot of cvs Ottobratica and Sinopolese is in accordance with previous observations of other authors (Martelli, 1959; Martelli & Piglionica, 1961). Copper fungicides, ziram, and metiram were most effective in reducing the incidence of olive rot in the field, but no fungicide tested alone gave satisfactory control of the disease. On the basis of the laboratory results, it could be supposed that a mixture of copper or non-copper fungicides with systemic substances (bitertanol, hexaconazole and myclobutanil) could give better results. In in vitro tests, the former were more effective on conidial germination, the latter on mycelial growth.

Multiple applications of fungicides from September to November were more effective than a single treatment in September in reducing the severity of disease. However the values for the fungicide treatments did not differ significantly from the control, when they were applied on cv. Sinopolese or the incidence of disease was very low. Thus, the efficacy and convenience of the chemical treatments depends on the susceptibility of the olive cultivar and on the climatic conditions favouring disease spread. Early harvesting can result in a reduction of disease severity, so choice of harvesting time could also be regarded as part of the management strategy for olive rot.

Lutte chimique contre Glomerella cingulata sur olivier

L’action de plusieurs fongicides sur Gloinerella cingulata, agent d’une grave pourriture des fruits de l’olivier, a ete evalute in vitro et au champ. Les fongicides suivants ont etk testes: oxychlorure de cuivre, bouillie bordelaise, zirame, myclobutanil, hexaconazole, metirame, bitertanol et bknomyl. Des essais en plein champ ont etC rCalises dans des oliveraies dans la region de Calabria (sud de l’Italie), pendant deux anntes consecutives et sur deux cultivars: Ottobratica (sensible) et Sinopolese (resistant). La concentration minimale ayant un effet sur la croissance mycelienne in oitro pour le bitertanol, I’hexaconazole, le myclobutanil, le benomyl, le metirame, le zirame et l’oxychlorure de cuivre, etait respectivement de: < 5 , < 50, < 50, > 1000, > 1000, > 1000, > 1000 pg ml-I. Pour la germination des conidies, les DEgo etaient respectivement de 0,01,0,02, 50,50,1000,1000 et 1000 pg ml-I pour le btnomyl, le zirame, le myclobutanil, I’oxychlorure de cuivre, le bitertanol, l’hexaconazole et le metirame. Dans des essais de plein champ, la bouillie bordelaise et l’oxychlorure de cuivre se sont averes les plus efficaces; cependant le zirame, le myclobutanil, le bitertanol, le mCtirame et I’hexaconazole ont permis de rtduire significative- ment les pourritures sur fruits.

472 A. M . Pennisi et al.

References

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SIDDI, G. (1989) Myclobutanil: a systemic fungicide. Informatore Fitoputologico 39 (lo), 19-28.