R. Sales

First Report of Alternaria Brown Spot of Citrus in Spain

In 1998, a new disease of Fortune mandarin trees was detected in orchards in the eastern province of Valencia. This is one of the most important late-maturing cultivars grown in Spain. Symptoms were typical of Alternaria brown spot of citrus (2). Young leaves showed brown necrotic and irregular blighted areas with characteristic yellow halos. The necrosis had a tendency to follow the veins. On fruits, symptoms included light brown, slightly depressed spots to circular and dark brown areas on the external surface. Infected young fruits and leaves often fell and the mature fruits were unmarketable due to lesions, resulting in important economic losses. Isolations on potato dextrose agar supplemented with 0.5 mg/ml of streptomycin sulfate (PDAS) from affected leaves and fruits consistently yielded Alternaria alternata (Fr.:Fr.) Keissl., which was identified based on conidial morphological characteristics. Pathogenicity tests were conducted using 15 isolates from fruit and leaves by inoculating detached immature Fortune leaves with a sterile water suspension of 5 × 105 conidia per ml. Drops of this suspension (40 μl each) were placed on the lower surfaces of each leaflet using four leaves per isolate. Leaves were incubated in a moist chamber in the dark at 27°C (1). After 48 h, most of these isolates caused necrotic lesions on the leaves similar to those observed in the field, and the fungus was reisolated, confirming Kochs postulates. In 1999, the fungus spread to other citrus-growing areas, and to date the disease has been detected affecting Fortune and Nova mandarins and Minneola tangelo. This is the first report of Alternaria brown spot of citrus in Spain. References: (1) K. Kohmoto et al. Phytopathology 81:719, 1991. (2) J. O. Whiteside. Plant Dis. Rep. 60:326, 1976.

First report of Monosporascus cannonballus on watermelon in Brazil.

In 2008 and 2009, vine decline symptoms were observed in three watermelon (Citrullus lanatus (Thunb.) Matsum. & Nakai) fields located in the municipalities of Mossoró (Rio Grande do Norte State) and Quixeré (Ceará State) in northeastern Brazil. Symptoms included yellowing of crown leaves just prior to harvest and collapse of many of the vines. Mean maximum daily temperatures for the first and second half of the season were 28.6 and 25.1°C, respectively. Affected plants exhibited necrotic root systems and lacked most of the secondary and tertiary feeder roots. Numerous perithecia on the roots contained asci and ascospores characteristics of Monosporascus cannonballus Pollack & Uecker (1,2). Small pieces of primary and secondary roots were surface disinfected and plated onto potato dextrose agar (PDA) medium with 0.5 g liter-1 of streptomycin sulfate and incubated for 7 days at 25°C in the dark. Hyphal tips from all colonies were transferred to PDA and further incubated for 30 to 40 days at 25°C in the dark for subsequent growth and sporulation. Isolations consistently yielded colonies of white mycelium, which became dark grayish after 10 to 15 days, and perithecia with one-spored asci. The internal transcribed spacer regions of ribosomal DNA of isolates 18-5 and 19-1 were sequenced (GenBank Accession Nos. GQ891544 and GQ891545). These sequences were identical to sequences of M. cannonballus (GenBank Accession Nos. AM167936 and AM167937). Pathogenicity of these two isolates was confirmed on watermelon cv. Crimson Sweet in a greenhouse maintained at 25 to 30°C. Inoculum was produced in a sand-oat hulls (Avena sativa) medium (0.5 liter of sand, 46 g of ground oat hulls, and 37.5 ml of distilled water) and incubated at 25°C for 1 month. CFU were quantified by serial dilution using 1% hydroxyethyl cellulose. A sterilized mixture of equal portions (vol/vol) of sand and peat moss was used to fill 17-cm-diameter plastic pots and inoculum was added to produce an inoculum concentration of 20 CFU g-1. Five watermelon seeds planted in each pot were later thinned to one seedling per pot. There were five replicated pots for each treatment with an equal number of noninfested pots. Plants were evaluated for disease 45 days after sowing. All isolates of M. cannonballus were highly aggressive and caused severe root necrosis compared with the noninoculated controls. M. cannonballus was reisolated from symptomatic plants, confirming Kochs postulates. In 2004, M. cannonballus was reported in the same Brazilian cucurbit-growing areas causing root rot and vine decline of muskmelon (Cucumis melo L.) (3), but to our knowledge, this is the first report of M. cannonballus on watermelon in Brazil. References: (1) R. D. Martyn and M. E. Miller. Plant Dis. 80:716, 1996. (2) F. G. Pollack and F. A. Uecker. Mycologia 66:346, 1974. (3) R. Sales Jr. et al. Plant Dis. 88:84, 2004.

Variabilidade de isolados de Myrothecium roridum provenientes de meloeiro cultivado no Estado do Rio Grande do Norte

The variability of 53 isolates of Myrothecium roridum obtained from melon (Cucumis melo) fields in the State of Rio Grande do Norte (Brazil) was estimated based on: severity of Myrothecium stem canker in the cultivars AF-682 and Orange Flesh, the pathogens physiology (mycelium growth rate - MGR - and sporulation - SP) and its sensitivity to the fungicide fluazinam. All isolates were pathogenic to AF-682 and Orange Flesh; however, better discrimination of pathogen variability was achieved in the latter. Variation among isolates was high when assessing all variables, except for MGR and sensitivity to fungicides (all sensitive). There was no correlation between SEV and any other variables. The variation among isolates within municipalities (60%) was higher than the variation among municipalities. There is evidence of high genetic variability in the populations of M. roridum, but no association between the assessed variables and locations was detected.

A Tuber Rot of Cyperus esculentus Caused by Rosellinia necatrix

Tiger nut (Cyperus esculentus L.) is cultivated in Spain for the production of tiger nut milk. Over the past 5 years, important economic losses resulting from a new tuber rot have been observed near Valencia in eastern Spain. Affected tubers were covered by a white mycelium that turned black as the disease advanced, leading to a general rotting of tubers. Aboveground plant parts showed some early decay and under high-moisture environmental conditions the white mycelium was present on the soil surface. This mycelium showed pyriform swellings characteristic of Rosellinia necatrix Prill., and coremia were occasionally observed on external surfaces of tubers after incubation in a moist chamber for 1 to 2 months. Coremia produced small ellipsoid or obovoid single-celled conidia 3.7 to 5.0 × 2.0 to 2.2 μm. The teleomorph has not been observed. Pathogenicity tests were conducted with inoculum produced on wheat (Triticum aestivum L.) seeds that were soaked for 12 h in flasks filled with distilled water. Each flask contained 300 ml of seeds that were subsequently autoclaved after excess water was drained. Two fungal disks of a 2-week-old culture of R. necatrix grown on potato dextrose agar were placed aseptically in each flask. The flasks were incubated at 25°C for 4 weeks, and shaken once a week to avoid clustering of inoculum. Two plastic pots (35 cm in diameter) per isolate were filled with a sterilized mixture of equal portions (vol/vol) of soil, sand, and peat moss, and inoculum was added at a concentration of 30 g of infected wheat seeds per 1,200 g of substrate (1). Healthy tubers were surface disinfested in 1.5% (vol/vol) sodium hypochlorite for 1 min, washed twice in sterile water, sown, and subsequently thinned to one per pot after emergence. Plants were grown under field conditions. Six months after inoculation, symptomatic tubers appeared similar to those originally observed in the field. The fungus was reisolated from affected tubers, confirming Kochs postulates. Pathogenicity tests were also conducted on avocado (cv. Reed) and almond (cv. Garriges) seedlings, and apple rootstock (MM-106) as very susceptible host plants for R. necatrix (1). Inoculated plants showed symptoms of wilting and death 4 weeks after inoculation. The fungus was reisolated from affected plants. This is the first report of C. esculentus as a host of R. necatrix. Reference: (1) A. Sztejnberg and Z. Madar. Plant Dis. 64: 662, 1980.

Fitness components of Monosporascus cannonballus isolates from northeastern Brazilian melon fields

Monosporascus root rot and vine decline caused by Monosporascus cannonballus is one of the most important melon yield-limiting diseases in northeastern Brazil. This study investigated the fitness components of 57 isolates of M. cannonballus obtained from Brazilian melon fields by evaluating: i) their mycelial growth rate (MGR), and perithecia and ascospore production (PP and AP) on potato dextrose agar (PDA); ii) their sensitivity to the fungicide fluazinam; and iii) their virulence to melon seedlings. All M. cannonballus isolates showed variability in their MGR, PP and AP values. They were sensitive to the fungicide fluazinam, showing some degree of mycelial growth inhibition (MGI), and were pathogenic to melon seedlings, with a mean disease severity index (DSI) of 62.1%. By univariate analysis, the formation of groups of similarity amongst the isolates of M. cannonballus within each variable was not limited by the area of origin of each isolate, given that in most situations, different isolates of the same area were distributed into distinct groups of similarity. A multivariate cluster analysis allowed the separation of the 57 M. cannonballus isolates in 18 groups of similarity. The fitness variability among M. cannonballus isolates found in this study should be considered when possible sources of resistance are evaluated in melon breeding programs.

First Report of Basal Rot of Leek Caused by Fusarium culmorum in Spain

A severe basal rot of leek (Allium porrum L.) was first observed in 1996 in Sueca (Valencia) eastern Spain, and again from 1998 to 2000 in several fields in Haro (La Rioja) northern Spain, where the disease caused significant economic losses. Leeks were stunted, wilted, and had water-soaked tissues with a characteristic reddish purple discoloration at the base of the stem. Affected plants could be pulled up easily because of rotting of the basal plate and root system. Similar symptoms were also observed on seedlings 2 to 3 weeks after transplanting. Isolations from symptomatic stems and roots onto potato dextrose agar (PDA) supplemented with 0.5 mg/ml of streptomycin sulfate (PDAS) consistently yielded a Fusarium sp. Isolates were transferred to potato sucrose agar (PSA) and synthetic low nutrient agar (SNA) and incubated at 25°C for 10 days with a 12-h photoperiod. The isolates were identified as Fusarium culmorum (Wm. G. Sm.) Sacc. based on colony morphology on PSA and phialide and conidial morphology on SNA. This disease has been described on garlic (1), but there are few reports about its incidence on leek (2). Pathogenicity tests were conducted on leek cvs. Alora, Axel, Casado, Goliath, and Varea using five isolates of F. culmorum from different locations. Ninety-day-old seedlings were inoculated by introducing a spore suspension to obtain a soil infested with the fungus at 104 colony-forming units (CFU)/g of potting mix (a sterilized mixture of equal portions [v/v] of soil, sand, and peat moss). Plants were maintained in a greenhouse at 20 to 30°C. Within 45 to 70 days after inoculation, symptoms developed that were similar to those observed in the field. The fungus was reisolated from affected plants, completing Kochs postulates. All isolates were pathogenic, and all leek cvs. were highly susceptible to the pathogen. To our knowledge, this is the first report of F. culmorum affecting leek in Spain. References: (1) F. J. Crowe et al. Phytopathology 76:1094, 1986. (2) G. Tamietti and A. Garibaldi. Riv. di Patol. Veg. IV. 13:69, 1977.