Physic nut (Jatropha curcas L.) is an important biofuel crop worldwide. Although it has been reported to be resistant to pests and diseases (1), stem cankers have been observed on this plant at several locations in Peninsular Malaysia since early February 2008. Necrotic lesions on branches appear as scars with vascular discoloration in the tissue below the lesion. The affected area is brownish and sunken in appearance. Disease incidence of these symptomatic nonwoody plants can reach up to 80% in a plantation. Forty-eight samples of symptomatic branches collected from six locations (University Farm, Setiu, Gemenceh, Pulau Carey, Port Dickson, and Kuala Selangor) were surface sterilized in 10% bleach, rinsed twice with sterile distilled water, air dried on filter paper, and plated on water agar. After 4 days, fungal colonies on the agar were transferred to potato dextrose agar (PDA) and incubated at 25°C. Twenty-seven single-spore fungal cultures obtained from all locations produced white, aerial mycelium that became dull gray after a week in culture. Pycnidia from 30-day-old pure cultures produced dark brown, oval conidia that were two celled, thin walled, and oval shape with longitudinal striations. The average size of the conidia was 23.63 × 12.72 μm with a length/width ratio of 1.86. On the basis of conidial morphology, these cultures were identified as Lasiodiplodia theobromae. To confirm the identity of the isolates, the internal transcribed spacer (ITS) region was amplified with ITS1/ITS4 primers and sequenced. The sequences were deposited in GenBank (Accession Nos. HM466951, HM466953, HM466957, GU228527, HM466959, and GU219983). Sequences from the 27 isolates were 99 to 100% identical to two L. theobromae accessions in GenBank (Nos. HM008598 and HM999905). Hence, both morphological and molecular characteristics confirmed the isolates as L. theobromae. Pathogenicity tests were performed in the glasshouse with 2-month-old J. curcas seedlings. Each plant was wound inoculated by removing the bark on a branch to a depth of 2 mm with a 10-mm cork borer. Inoculation was conducted by inserting a 10-mm-diameter PDA plug of mycelium into the wound and wrapping the inoculation site with wetted, cotton wool and Parafilm. Control plants were treated with plugs of sterile PDA. Each isolate had four replicates and two controls. After 6 days of incubation, all inoculated plants produced sunken, necrotic lesions with vascular discoloration. Leaves were wilted and yellow above the point of inoculation on branches. The control plants remained symptomless. The pathogen was successfully reisolated from lesions on inoculated branches. L. theobromae has been reported to cause cankers and dieback in a wide range of hosts and is common in tropical and subtropical regions of the world (2,3). To our knowledge, this is the first report of stem canker associated with L. theobromae on J. curcas in Malaysia. References: (1) S. Chitra and S. K. Dhyani. Curr. Sci. 91:162, 2006. (2) S. Mohali et al. For. Pathol. 35:385, 2005. (3) E. Punithalingam. Page 519 in: CMI Descriptions of Pathogenic Fungi and Bacteria. Commonwealth Mycological Institute, Kew, Surrey, UK. 1976.
Symptoms of gray leaf spot were first observed in June 2011 on pepper (Capsicum annuum) plants cultivated in the Cameron Highlands and Johor State, the two main regions of pepper production in Malaysia (about 1,000 ha). Disease incidence exceeded 70% in severely infected fields and greenhouses. Symptoms initially appeared as tiny (average 1.3 mm in diameter), round, orange-brown spots on the leaves, with the center of each spot turning gray to white as the disease developed, and the margin of each spot remaining dark brown. A fungus was isolated consistently from the lesions using sections of symptomatic leaf tissue surface-sterilized in 1% NaOCl for 2 min, rinsed in sterile water, dried, and plated onto PDA and V8 agar media (3). After 7 days, the fungal colonies were gray, dematiaceous conidia had formed at the end of long conidiophores (19.2 to 33.6 × 12.0 to 21.6 μm), and the conidia typically had two to six transverse and one to four longitudinal septa. Fifteen isolates were identified as Stemphylium solani on the basis of morphological criteria described by Kim et al. (3). The universal primers ITS5 and ITS4 were used to amplify the internal transcribed spacer region (ITS1, 5.8, and ITS2) of ribosomal DNA (rDNA) of a representative isolate (2). A 570 bp fragment was amplified, purified, sequenced, and identified as S. solani using a BLAST search with 100% identity to the published ITS sequence of an S. solani isolate in GenBank (1). The sequence was deposited in GenBank (Accession No. JQ736024). Pathogenicity of the fungal isolate was tested by inoculating healthy pepper leaves of cv. 152177-A. A 20-μl drop of conidial suspension (105 spores/ml) was used to inoculate each of four detached, 45-day-old pepper leaves placed on moist filter papers in petri dishes (4). Four control leaves were inoculated similarly with sterilized, distilled water. The leaves were incubated at 25°C at 95% relative humidity for 7 days. Gray leaf spot symptoms similar to those observed on the original pepper plants began to develop on leaves inoculated with the fungus after 3 days, and S. solani was consistently reisolated from the leaves. Control leaves did not develop symptoms and the fungus was not reisolated from these leaves. Pathogenicity testing was repeated with the same results. To our knowledge, this is the first report of S. solani causing gray leaf spot on pepper in Malaysia. References: (1) S. F. Altschul et al. Nucleic Acids Res. 25:3389, 1997. (2) M. P. S. Camara et al. Mycologia 94:660, 2002. (3) B. S. Kim et al. Plant Pathol. J. 15:348, 1999. (4) B. M. Pryor and T. J. Michailides. Phytopathology 92:406, 2002.
A leaf spot on eggplant (Solanum melongena) was observed in major eggplant growing regions in Malaysia, including the Cameron Highlands and Johor State, during 2011. Disease incidence averaged approximately 30% in severely infected regions in about 150 ha of eggplant fields and greenhouses examined. Early symptoms consisted of small, circular, brown, necrotic spots uniformly distributed on leaves. The spots gradually enlarged and developed concentric rings. Eventually, the spots coalesced and caused extensive leaf senescence. A fungus was recovered consistently by plating surface-sterilized (1% NaOCl) sections of symptomatic leaf tissue onto potato dextrose agar (PDA). For conidial production, the fungus was grown on potato carrot agar (PCA) and V8 agar media under a 16-h/8-h dark/light photoperiod at 25°C (4). Fungal colonies were a dark olive color with loose, cottony mycelium. Simple conidiophores were ≤120 μm long and produced numerous conidia in long chains. Conidia averaged 20.0 × 7.5 μm and contained two to five transverse septa and the occasional longitudinal septum. Twelve isolates of the fungus were identified as Alternaria tenuissima on the basis of morphological characterization (4). Confirmation of the species identification was obtained by molecular characterization of the internal transcribed spacer (ITS) region of rDNA amplified from DNA extracted from a representative isolate using universal primers ITS4 and ITS5 (2). The 558 bp DNA band amplified was sent for direct sequencing. The sequence (GenBank Accession No. JQ736021) was subjected to BLAST analysis (1) and was 99% identical to published ITS rDNA sequences of isolates of A. tenuissima (GenBank Accession Nos. DQ323692 and AY154712). Pathogenicity tests were performed by inoculating four detached leaves from 45-day-old plants of the eggplant cv. 125066x with 20 μl drops (three drops/leaf) of a conidial suspension containing 105 conidia/ml in sterile distilled water. Four control leaves were inoculated with sterile water. Leaves inoculated with the fungus and those treated with sterile water were incubated in chambers at 25°C and 95% RH with a 12-h photoperiod/day (2). Leaf spot symptoms typical of those caused by A. tenuissima developed on leaves inoculated with the fungus 7 days after inoculation, and the fungus was consistently reisolated from these leaves. The control leaves remained asymptomatic and the pathogen was not reisolated from the leaves. The pathogenicity test was repeated with similar results. To our knowledge, this is the first report of A. tenuissima causing a leaf spot on eggplant in Malaysia. A. tenuissima has been reported to cause leaf spot and fruit rot on eggplant in India (3). References: (1) S. F. Altschul et al. Nucleic Acids Res. 25:3389, 1997. (2) B. M. Pryor and T. J. Michailides. Phytopathology 92:406, 2002. (3) P. Raja et al. New Disease Rep. 12:31, 2005. (4) E. G. Simmons. Page 1 in: Alternaria Biology, Plant Diseases and Metabolites. J. Chelchowski and A. Visconti, eds. Elsevier, Amsterdam, 1992.
Soybean (Glycine max L.) is one of the most economically important crops in the world, and anthracnose is known to infect soybean in most countries. Colletotrichum truncatum is the common pathogen causing anthracnose of soybean. However, at least five species of Colletotrichum have been reported on soybean worldwide (2). In July 2010, anthracnose symptoms were observed on soybean in the experimental fields of the agriculture station in Ladang Dua, University Putra Malaysia located in Selangor state of Malaysia. Symptoms were initially observed on a few plants randomly within one field, but after 4 weeks, the disease was found in two additional fields scattered across an area of 1 km2. Pinkish-brown lesions were observed on the pods, and the formation of dark lesions on the leaves and stems was sometimes followed by stem girdling, dieback, and distorted growth. At later stages, numerous epidermal acervuli developed in the lesions, and mucilaginous conidial masses appeared during periods of high relative humidity. Conidia produced in acervuli were straight, cylindric, hyaline, and aseptate, with both ends rounded. Conidia measured (mean ± SD) 14.2 ± 0.6 × 3.6 ± 0.7 μm, and the L/W ratio was 3.95 μm. Six isolates of the fungus were obtained and identified as C. gloeosporioides on the basis of morphological characterization (3). The isolates were deposited in the University Putra of Malaysia Culture Collection (UPMCC). PDA cultures were white at first and subsequently became grayish to pink to reddish-brown. Amplification and sequence analysis of coding and none-coding regions of the ITS-rDNA (GenBank JX669450), actin (JX827430), β-tubulin (JX827454), histone (JX827448), chitin synthase (JX827436), and glyceraldehyde-3-phosphate dehydrogenase (JX827442) obtained from the representative isolate, CGM50, aligned with deposited sequences from GenBank and revealed 99 to 100% sequence identity with C. gloeosporioides strains (JX258757, JX009790, GQ849434, HM575301, JQ005413, and JX00948 from GenBank). One representative isolate, CGM50, was used for pathogenicity testing. Four non-infected detached leaves and pods of 24-day-old G. max var. Palmetto were surface-sterilized and inoculated by placing 10 μl of a conidial suspension (106 conidia ml-1) using either the wound/drop or non-wound/drop method (4), with 10 μl distilled water as a negative control. Leaves and pods were incubated at 25°C, 98% RH. The experiment was repeated twice. Five days after inoculation, the development of typical field symptoms, including acervuli formation, occurred on the leaves and pods of inoculated plants, but not on the negative controls. A fungus with the same colony and conidial morphology as CGM50 was recovered from the lesions on the inoculated leaves and pods. Anthracnose caused by C. gloeosporioides on soybean plants has been reported previously in different countries, but not in Malaysia (3). Geographically, the climate of Malaysia is highly conducive to maintain and cause outbreaks of anthracnose all year round; thus, the development of management recommendations will be inevitable for anthracnose control. To our knowledge, this is the first report of C. gloeosporioides causing anthracnose on soybean in Malaysia. References: (1) U. Damm et al. Fungal Diversity 39:45, 2009. (2) S. L. Chen et al. J. Phytopathol. 154:654, 2006. (3) B. C. Sutton. The Genus Glomerella and its Anamorph Colletotrichum. CAB International, Wallingford, UK, 1992. (4) P. P. Than et al. Plant Pathol. 57:562, 2008. ERRATUM: A correction was made to this Disease Note on May 19, 2014. The author N. Soleimani was added.
Banana is the second largest cultivated fruit crop in Malaysia, and is cultivated for both the domestic market and also for export. Anthranose is a well-known postharvest disease of banana and with high potential for damaging market value, as infection commonly occurs during storage. Anthracnose symptoms were observed on several varieties of banana such as mas, berangan, awak, nangka, and rastali in the states of Perak and Penang between August and October 2011. Approximately 80% of the fruits became infected with initial symptoms characterized as brown to black spots that later became sunken lesions with orange or salmon-colored conidial masses. Infected tissues (5 × 5 mm) were surface sterilized by dipping in 1% sodium hypochlorite (NaOCl) for 3 to 5 min, rinsed with sterile distilled water, and plated onto potato dextrose agar (PDA). Direct isolation was done by transferring the conidia from conidial masses using an inoculation loop and plating onto PDA. For both methods, the PDA plates were incubated at 27 ± 1°C with cycles of 12 h light and 12 h darkness. Visible growth of mycelium was observed after 4 to 5 days of incubation. Twenty isolates with conidial masses were recovered after 7 days of incubation. The isolates produced grayish white to grayish green and grey to moss dark green colony on PDA, pale orange conidial masses, and fusiform to cylindrical and hyaline conidia with an average size of 15 to 19 × 5 to 6 μm. Appresoria were ovate to obovate, dark brown, and 9 to 15 × 7 to 12 μm and setae were present, slightly swollen at the base, with a tapered apex, and brown. The cultural and morphological characteristics of the isolates were similar to those described for C. gleosporioides (1,2,3). All the C. gloeosporioides isolates were deposited in culture collection at Plant Pathology Lab, University Sains Malaysia. For confirmation of the identity of the isolates, ITS regions were sequenced using ITS4 and ITS5 primers. The isolates were deposited in GenBank with accessions JX163228, JX163231, JX163201, JX163230, JX163215, JX163223, JX163219, JX163202, JX163225, JX163222, JX163206, JX163218, JX163208, JX163209, JX163210, JX431560, JX163212, JX163213, JX431540, and JX431562. The resulting sequences showed 99% to 100% similarity with multiple C. gloeosporioides isolates in GenBank. Pathogenicity tests were conducted using mas, berangan, awak, nangka, and rastali bananas. Fruit surfaces were sterilized with 70% ethanol and wounded using a sterile scalpel. Two inoculation techniques were performed separately: mycelia plug and conidial suspension. Mycelial disc (5 mm) and a drop of 20 μl spore suspension (106 conidia/ml) were prepared from 7-day-old culture and placed on the fruit surface. The inoculated fruits were incubated at 27 ± 1°C for 10 days at 96.1% humidity. After 3 to 4 days of inoculation, brown to black spotted lesions were observed and coalesced to become black sunken lesions. Similar anthracnose symptoms were observed on all banana varieties tested. C. gloeosporioides was reisolated from the anthracnose lesions of all the inoculated fruit in which the cultural and morphological characteristics were the same as the original isolates. To our knowledge, this is the first report of C. gloeosporioides causing anthracnose of Musa spp. in Malaysia. References: (1) P. F. Cannon et al. Mycotaxon 104:189, 2008. (2) J. E. M. Mordue. Glomerella cingulata. CMI Description of Pathogenic Fungi and Bacteria, No. 315. CAB International,1971. (3) H. Prihastuti et al. Fungal Diversity 39:89, 2009.
Passiflora edulis Sims f. edulis, known as purple passion fruit, is a woody, perennial vine that is grown for its attractive two-part flower and its purple, edible fruit (4). In November 2009, passion fruit vines were collected during a regulatory nursery inspection in Santa Barbara County and submitted to the California Department of Food and Agriculture Plant Pest Diagnostics Laboratory. Nearly 100% of the plants inspected, all of which were approximately 1.25 m tall, appeared stunted, defoliated, and severely wilted. Dark brown vascular discoloration was present in the roots and lower stems of the plants. A pinkish violet Fusarium oxysporum colony containing chlamydospores, multiseptate macroconidia, and microconidia formed on monophialidic conidiophores was consistently isolated from roots and stems onto half-strength acidified potato dextrose agar (aPDA). All further experiments were done with an isolate obtained from a single conidium. A portion of the translation elongation factor gene (TEF-1α) was amplified and sequenced with primers ef1 and ef2 from our isolate (GenBank No. JF332039) (3). BLAST analysis of the 615-bp amplicon with the FUSARIUM-ID database showed 99% similarity with a F. oxysporum passion fruit isolate from Australia (NRRL 38273) (3). To confirm pathogenicity, washed roots of four-leaf stage seedlings approximately 10 cm tall were submerged in a conidial spore suspension (106 spores/ml) for 15 min. The conidial suspension was prepared by flooding 10-day-old cultures grown on aPDA medium with sterile distilled water. Seven seedlings were inoculated and planted in 10-cm2 pots and kept in a 25°C growth chamber with a 12-h photoperiod. Seven seedlings were mock inoculated with sterile water. After 3 weeks, four of the seven inoculated plants had leaves with yellow veins and discolored roots and had partially defoliated. Two of the four symptomatic plants also had brown stem cankers. F. oxysporum grew from the isolated roots and stems of all the inoculated plants. F. oxysporum did not grow from root and stem pieces from the water-dipped plants and the plants remained asymptomatic. Inoculations were repeated on plants approximately 15 cm tall with F. oxysporum growing from roots and stem pieces of all inoculated plants. Symptoms of yellow veins and root necrosis were not observed until 4 weeks after inoculation. Fusarium wilt caused by F. oxysporum f. sp. passiflorae is a significant disease of P. edulis f. edulis in Australia. The disease has also been reported in South Africa, Malaysia, Brazil, Panama, and Venezuela; but it is unclear as to whether the symptoms were caused by Fusarium wilt or Haematonectria canker (1). Banana poka (P. mollissima), P. ligularis, and P. foetida are also susceptible hosts (2). To our knowledge, this is the first report of Fusarium wilt caused by F. oxysporum f. sp. passiflorae on passion fruit in North America. Passion fruit is not commercially produced for consumption in California so the economic importance of this disease appears to be limited to nursery production and ornamental landscapes. The grower of the California nursery stated that the infected passion fruit plants had been propagated on site from seed. The source of inoculum at this nursery remains unknown. References: (1) I. H. Fischer and J. A. M. Rezende. Pest Tech. 2:1, 2008 (2) D. E. Garder. Plant. Dis. 73:476, 1989. (3) D. M. Geiser et al. Eur. J. Plant Pathol. 110:473, 2004. (4) F. W. Martin et al. Econ. Bot. 24:333, 1970.
Mango (Mangifera indica L.) is grown on approximately 20,000 ha in Taiwan. It is an economically important crop and the income of many fruit farmers comes primarily from mango production. During 2006 and 2007, a stem-end rot disease was observed 1 week after harvest on 28 to 36% of stored mangoes picked from six orchards in the Pingtung, Tainan, and Kaoshiung regions. Two popular mango cultivars, Keitt and Irwin, showed greater susceptibility to this disease, while 'Haden' was found to be moderately susceptible. In storage, symptoms initially appeared as light-to-dark brown lesions surrounding peduncles. Rot symptoms advanced slowly but eventually penetrated the mesocarp, which consequently reduced the commercial value of fruits. The fungus formed abundant pycnidia (0.1 to 0.6 mm in diameter) on infected fruits in advanced stages of symptom development. Pieces of symptomatic fruits plated on acidified potato dextrose agar (PDA) and incubated at 25 ± 1°C consistently yielded the same fungus. A single conidial isolate was cultured. Pycnidia developed on PDA after continuous exposure to light for 9 to 14 days. On the basis of morphological characteristics, the fungus was identified as Phomopsis mangiferae L. (2,3). Pycnidia released two types of conidia: α-conidia (5 to 10 × 2.3 to 4.0 μm) were hyaline and oval to fusoid; and β-conidia (15.0 to 37.5 × 1.3 to 2.5 μm) were hyaline and filiform with characteristic curves. Conidiophores were hyaline, filiform, simple or branched, septate, and 15 to 75 μm long. Cultures incubated under continuous fluorescent light (185 ± 35 μE·m-2·s-1) at 25°C for 3 days were used as inoculum for pathogenicity tests. Five fruits from 'Keitt' were wounded with a sterilized scalpel and each wound (2 × 2 × 2 mm) was inoculated with either a 5-mm mycelium agar plug or a 0.5-ml spore suspension (105 conidia per ml) of the fungus. Five wounded fruits inoculated with 5-mm PDA plugs or sterile water alone served as controls. Inoculated areas were covered with moist, sterile cotton. Fruits were enclosed in plastic bags and incubated at 24°C for 3 days. The test was performed three times. The same symptoms were observed on all inoculated fruits, whereas no decay was observed on control fruits. Reisolations from the inoculated fruits consistently yielded P. mangiferae, thus fulfilling Koch's postulates. This disease has previously been reported in Australia, Brazil, China, Cuba, India, Malaysia, and the United States (1). To our knowledge, this is the first report of P. mangiferae causing stem-end rot disease on mangoes in Taiwan. Our report necessitates taking preventive strategies in the field, prior to or after harvest, to contain postharvest losses in mangoes. References: (1) G. I. Johnson. Page 39 in: Compendium of Tropical Fruit Diseases. R. C. Ploetz et al., eds. The American Phytopathological Society. St. Paul, MN, 1994. (2) R. C. Ploetz, ed. Page 354 in: Diseases of Tropical Fruit Crops. CABI Publishing. Wallingford, UK, 2003. (3) E. Punithalingam. No. 1168 in: Descriptions of Pathogenic Fungi and Bacteria. CMI, Kew, Surrey, UK, 1993.
In March 2005, a fruit rot disease was found in several commercial strawberry (Fragaria × ananassa Duchesne) fields at Fongyuan, 24.25°N, 120.72°E, in Taichung County in central Taiwan. The disease was rare and was negligible in most cultivated areas. However, disease incidence has increased by 4 to 5% over the last 2 years and causes significant postharvest losses. In storage, symptoms on berries include light brown-to-black, sunken, irregularly shaped lesions. The lesions gradually enlarge and become firm with a dark green-to-black, velvety surface composed of mycelia, conidiophores, and conidia. Twelve single conidial isolates (AF-1 to AF-12) of a fungus were isolated by placing portions of symptomatic fruit from four locations onto acidified potato dextrose agar (PDA) and incubating at 24 ± 1°C. One isolate from each of the four locations, AF-2, 6, 9, and 12, was selected for identification and pathogenicity studies. The fungus was identified as an Alternaria sp. according to the morphological descriptions of A. tenuissima (2,3). Conidiophores were simple or branched, straight or flexuous, septate, pale to light brown, 3.0 to 5.0 μm in diameter, and bore two to six conidia in a chain. Conidia were dark brown, obclavate or oval, and multicellular with seven transverse (in most cases) and numerous longitudinal septa. Conidia were 15.5 to 56.5 μm (average 35.0 μm) long × 6.0 to 15.0 μm (average 11.0 μm) wide at the broadest point. The pathogen was consistently isolated from berries in the field or in storage. Pathogenicity tests were conducted by inoculating 12 surface-sterilized berries with each of the four isolates. Approximately 300 μl of a spore suspension (2 × 105 conidia per ml) was placed at two points on the uninjured surface of each fruit and allowed to dry for 5 min. Control fruits were treated with sterile water. The berries were then enclosed in a plastic bag and incubated at 24 ± 1°C for 2 days. Disease symptoms similar to those described above were observed on 95% of inoculated berries 3 days after inoculation, while no symptoms developed in control berries. Reisolation from the inoculated berries consistently yielded the Alternaria sp. described above. Pathogenicity tests were performed three times. Previously, strawberry fruit rot caused by A. tenuissima was reported from Florida (2) and Malaysia (1), however, to our knowledge, this is the first report of fruit rot of strawberry caused by a species of Alternaria in Taiwan. References: (1) W. D. Cho et al. List of Plant Diseases in Korea. Korean Society of Plant Pathology, 2004. (2) C. M. Howard and E. E. Albregts. Phytopathology 63:938, 1973. (3) R. D. Milholland. Phytopathology 63:1395, 1973.
Rambutan (Nephelium lappaceum Linn.) is a tropical, exotic fruit that has a rapidly expanding niche market in Hawaii. Diseased rambutan fruit was commonly observed in orchards in the Hilo and Kona districts of Hawaii Island during 2006. In surveys conducted in January, symptoms appeared as dark brown-to-black spots on mature fruit and blackened areas at the base of spinterns (hair-like projections) of mature and immature fruits. Pieces of infected fruit (cv. R167) were surface sterilized for 2 min in 0.5% NaOCl, plated on potato dextrose agar, and incubated at 24 ± 1°C for 7 days. The fungus growing on PDA was pale buff with sparse, aerial mycelium and acervuli containing black, slimy spore masses. All isolates had five-celled conidia. Apical and basal cells were hyaline, while the three median cells were olivaceous; the upper two were slightly darker than the lower one. Conidia (n = 40) were 20.3 ± 0.1 × 6.8 ± 0.1 μm. There were typically three apical appendages averaging 16.8 ± 0.2 μm long. The average basal appendage was 3.8 ± 0.1 μm long. The fungus was initially identified as Pestalotiopsis virgatula (Kleb.) Stey. on the basis of conidial and cultural characteristics (3). The identification was confirmed by molecular analysis of the 5.8S subunit and flanking internal transcribed spacers (ITS1 and ITS2) of rDNA amplified from DNA extracted from single-spore cultures with the ITS1/ITS4 primers (1,4) and sequenced (GenBank Accession No. EU047943). To confirm pathogenicity, agar pieces, 3 mm in diameter, from 7-day old cultures were used as inoculum. Five mature fruit from rambutan cv. R134 were rinsed with tap water, surface sterilized with 0.5% NaOCl for 2 min, wounded with a needle head, inoculated in the laboratory, and maintained in a moist chamber for 7 days. Lesions resembling symptoms that occurred in the field were observed on fruit after 7 days. No symptoms were observed on fruit inoculated with agar media. The fungus reisolated from diseased fruit was identical to the original isolates, confirming Koch's postulates. The disease appears to be widespread in Hawaii. Preharvest symptoms may have the potential to affect postharvest fruit quality if fruits are not stored at the proper conditions. Pestalotiopsis spp. have been reported on rambutan in Malaysia, Brunei, and Australia (2). To my knowledge, this is the first report of P. virgatula causing fruit spots on rambutan in Hawaii. References: (1) G. Caetano-Annolles et al. Curr. Genet. 39:346, 2001. (2) D. F. Farr et al. Fungal Databases. Systematic Botany and Mycology Laboratory. On-line publication. ARS, USDA, 2007. (3) E. F. Guba. Monograph of Pestalotia and Monochaetia. Harvard University Press, Cambridge, MA, 1961. (4) T. J. White et al. PCR Protocols: A Guide to Methods and Applications. Academic Press, San Diego, CA. 1990.
Red-fleshed dragon fruit (Hylocereus polyrhizus [Weber] Britton & Rose) is a newly introduced and potential crop in the Malaysian fruit industry. Besides its nutritious value, the fruit is being promoted as a health crop throughout Southeast Asia. In April of 2007, a new disease was observed in major plantations of H. polyrhizus throughout five states (Kelantan, Melaka, Negeri Sembilan, Penang, and Perak) in Malaysia with 41 and 25% disease incidence and severity, respectively. Stems of H. polyrhizus showed spots or small, circular, faint pink-to-beige necrotic lesions that generally coalesced as symptoms progressed. Symptom margins of diseased stem samples were surface sterilized with a 70% alcohol swab, cut into small blocks (1.5 × 1.5 × 1.5 cm), soaked in 1% sodium hypochlorite (NaOCI) for 3 min, and rinsed in several changes of sterile distilled water (each 1 min). The surface-sterilized tissues were placed onto potato dextrose agar (PDA) and incubated under alternating 12-h daylight and black light for 7 days. A fungus was consistently isolated from the stems of symptomatic H. polyrhizus and identified as Curvularia lunata (Wakker) Beodijn (1-3) that showed pale brown multicelled conidia (phragmoconidia; three to five celled) that formed apically through a pore (poroconidia) in sympodially, elongating, geniculated conidiophores. Conidia are relatively fusiform, cylindrical, or slightly curved, with one of the central cells being larger and darker (26.15 ± 0.05 μm). All 25 isolates of C. lunata obtained from diseased H. polyrhizus are deposited at the Culture Collection Unit, Universiti Sains Malaysia and available on request. Isolates were tested for pathogenicity by injecting conidial suspensions (1 × 106 conidia/ml) and pricking colonized toothpicks on 25 healthy H. polyrhizus stems. Controls were treated with sterile distilled water and noncolonized toothpicks. All inoculated plants and controls were placed in a greenhouse with day and night temperatures of 30 to 35°C and 23 to 30°C, respectively. Development of external symptoms on inoculated plants was observed continuously every 2 days for 2 weeks. Two weeks after inoculation, all plants inoculated with all isolates of C. lunata developed stem lesions similar to those observed in the field. No symptoms were observed on the control plants and all remained healthy. C. lunata was reisolated from 88% of the inoculated stems, completing Koch's postulates. The pathogenicity test was repeated with the same results. To our knowledge, this is the first report of C. lunata causing a disease on H. polyrhizus. References: (1) M. B. Ellis. Dematiaceous Hyphomycetes. Commonwealth Mycological Institute, Kew, Surrey, England, 1971. (2) R. R. Nelson and F. A. Hassis. Mycologia 56:316, 1964. (3) C. V. Subramanian. Fungi Imperfecti from Madras V. Curvularia. Proc. Indian Acad. Sci. 38:27, 1955.
During March 2007, a fruit rot disease was observed in several loquat (Eriobotrya japonica (Thunberg) Lindley) fields located in Taichung, Nantou, and Miaoli counties. Loquat is a valuable fruit crop grown predominantly in central Taiwan, and hence, even a minor yield loss by this new disease is economically significant. Symptoms on fruits initially appeared as small lesions (<1 mm) that later developed into light-to-dark brown, circular, larger (7 mm), sunken lesions, indicating invasion of a pathogen into the fruit. Pieces of rotted fruit tissue (1 × 1 × 1 mm) were immersed for 1 min in 3% commercial bleach, followed by 70% ethanol, cultured on potato dextrose agar (PDA), and incubated under constant fluorescent light (185 ± 35 μE·m-2·s-1) at 24°C for 2 days. Three single conidial isolates (AS1 to AS3) were selected and used in morphological and pathogenicity studies. All three isolates were identified as an Alternaria sp. (1-3) and formed abundant, dark brown mycelium when cultured on PDA with light at 24°C. Conidiophores were 60 to 89 × 3 to 5 μm, densely fasciculate, cylindrical, simple or branched, and had distinct conidial scars. Conidia were 12 to 74 × 6 to 14 μm, golden brown, straight or curved, obclavate with beaks measuring half the length of the conidium, and observed in chains of 10 or more spores with four to seven transverse septa and several longitudinal septa. Pathogenicity tests were conducted twice by inoculating eight surface-sterilized wounded or unwounded fruits with each of the three isolates in each experiment. Two cuts (1 × 1 × 1 mm) were made on each fruit 3 cm apart with a sterile scalpel, and a 300-μl spore suspension (2 × 105 conidia per ml) was placed on each wound. Similarly, a 300-μl spore suspension was placed on unwounded fruits and air dried for 5 min. Control fruits were similarly treated with sterile water. Inoculated fruits were enclosed in a plastic bag and kept at 24 ± 1°C. Symptoms of soft rot were observed on 60% (unwounded) and 100% (wounded) of inoculated fruits 5 days after inoculation, while control fruits did not develop disease symptoms. Reisolation from the symptomatic fruits consistently yielded an Alternaria sp. This fungus previously has been reported as the causal agent of fruit rot or black spot of papaya, mango, kiwifruit, pear, and carambola from Australia, India, Malaysia, South Africa, and the United States (1-3). To our knowledge, this is the first report of fruit rot of loquat caused by an Alternaria sp. in Taiwan. To manage this disease, growers may resort to fungicidal sprays followed by bagging of fruits to reduce pre- and postharvest losses. References: (1) A. L. Jones and H. S. Aldwinckle. Compendium of Apple and Pear Diseases. The American Phytopathological Society. St. Paul, MN, 1990. (2) R. C. Ploetz. Diseases of Tropical Fruit Crops. CABI Publishing. Wallingford, Oxfordshire, UK, 2003. (3) R. C. Ploetz et al. Compendium of Tropical Fruit Diseases. The American Phytopathological Society. St. Paul, MN, 1994.
In December 2008, infected leaves of Trichosanthes cucumerina were observed on commercial cucurbit farms located in Pontian, Johor (south of West Malaysia). Bright yellow and small necrotic lesions were observed on the adaxial surface of the leaves, whereas sporangiophores were observed on pale yellowish brown-to-brown lesions on the abaxial surface. The length and width of the sporangia ranged from 19 to 36 μm (28.6) and 11 to 23 μm (17.6), respectively. The length of the sporangiophores ranged from 310 to 450 μm, with an average length of 380 μm. The pathogen was identified as Pseudoperonospora cubensis on the basis of the morphological criteria described by Palti and Cohen (2). To confirm the morphological findings, DNA was extracted from symptomatic tissue and the internal transcribed spacer (ITS) region was PCR amplified using primers ITS5-P2 and ITS4 (3). The appropriate-sized amplicon was gel excised and column purified and then submitted for direct sequencing. The resulting 802 bp amplified ITS region was 100% identical to published P. cubensis sequences (GenBank Accession Nos. EU876603, EU876584, and AY198306). This sequence was deposited with NCBI GenBank under the Accession No. GU233293. In this study, pathogenicity tests were conducted using detached leaf disc assays (1) and a P. cubensis isolate obtained from T. cucumerina. For this purpose, leaf discs were excised from 6- to 8-week-old leaves of T. cucumerina using a 20-mm cork borer. Five leaf discs were placed with their abaxial surface facing upward on moist filter paper in petri dishes. Each of four leaf discs was inoculated with four 10-μl droplets of a 1 × 105 per ml sporangial suspension, whereas the fifth disc was inoculated with water droplets and served as a control. Three replications were completed. The leaf discs were placed in darkness at 14 ± 2°C for 24 h and subsequently incubated with a 12-h photoperiod. After 10 days, sporulation was observed on the sporangia-inoculated leaf discs with similar morphological features to the initial field samples. To our knowledge, this is the first report of P. cubensis causing downy mildew of T. cucumerina in Malaysia. References: (1) A. Lebeda and M. P. Widrlechner. J. Plant Dis. Prot. 110:337, 2003. (2) J. Palti and Y. Cohen. Phytoparasitica 8:109, 1980. (3) H. Voglmayr and O. Constantinescu. Mycol. Res. 112:487, 2008.
Cosmos caudatus Kunth. (Asteraceae), commonly known as ulam raja, is widely grown as an herbal aromatic shrub. In Malaysia, its young leaves are popularly eaten raw as salad with other greens and have been reported to possess extremely high antioxidant properties, which may be partly responsible for some of its believed medicinal functions. In early 2010, a suspected powdery mildew was observed on ulam raja plants at the Agricultural Park of Universiti Putra Malaysia. Initially, individual, white, superficial colonies were small and almost circular. Later, they enlarged and coalesced to cover the whole abaxial leaf surface. With development of the disease, all green parts (leaves, stems, and petioles) became covered with a continuous mat of mildew, giving a dusty appearance. Newly emerged leaves rapidly became infected. Diseased leaves ultimately senesced and dried up, making them aesthetically unattractive and unmarketable. The pathogen produced conidia in short chains (four to six conidia) on erect conidiophores. Conidiophores were unbranched, cylindrical, 125 to 240 μm long, with a slightly swollen foot cell. Individual conidia were hyaline, ellipsoid, and 25 to 30 (27.5) × 15 to 20 (17.5) μm with fibrosin inclusions. Morphological descriptions were consistent with those described for Sphaerotheca fuliginea or S. fusca, which has lately been reclassified as Podosphaera fusca (1). From extracted genomic DNA of P. fusca UPM UR1, the internal transcribed spacer (ITS) region was amplified with ITS1 (5'-TCCGTAGGTGAACCTGCGG-3') and ITS4 (5'-TCCTCCGCTTATTGATATGC-3'). A BLAST search of GenBank with an ITS rDNA sequence of this fungus (GenBank Accession No. HQ589357) showed a maximum identity of 98% to the sequences of two P. fusca isolates (GenBank Accession Nos. AB525915.1 and AB525914.1). To satisfy Koch's postulates, the pathogenicity of fungal strain UPM UR1 was verified on 4-week-old plants. Inoculation was carried out by gently rubbing infected leaves onto healthy plants of C. caudatus. Ten pots of inoculated plants were kept under a plastic humid chamber and 10 pots of noninoculated plants, placed under another chamber, served as controls. After 48 h, the plants were then placed under natural conditions (25 to 28°C). Powdery mildew symptoms, similar to those on diseased field plants, appeared after 7 days on all inoculated plants. The white, superficial colonies enlarged and merged to cover large areas within 2 weeks. The infected leaf tissues became necrotic 6 to 8 days after the appearance of the first symptoms. Sporulation of P. fusca was observed on all infected leaves and stems. No symptoms were seen on the control plants. To our knowledge, this is the first report of P. fusca causing powdery mildew on C. caudatus in Malaysia. This pathogen has also been reported previously to be economically important on a number of other hosts. With ulam raja plants, more attention should be given to prevention and control measures to help manage this disease. Reference: (1) U. Braun and S. Takamatsu. Schlechtendalia 4:1, 2000.
On May 9, 2013, symptoms reminiscent of bacterial leaf streak (BLS) caused by Xanthomonas oryzae pv. oryzicola were observed on rice plants at the panicle emergence stage at Musenyi, Gihanga, and Rugombo fields in Burundi. Affected leaves showed water-soaked translucent lesions and yellow-brown to black streaks, sometimes with visible exudates on leaf surfaces. Symptomatic leaves were ground in sterile water and the suspensions obtained were subjected to a multiplex PCR assay diagnostic for X. oryzae pathovars (3). Three DNA fragments (331, 691, and 945 bp) corresponding to X. oryzae pv. oryzicola were observed after agarose gel electrophoresis. Single bacterial colonies were then isolated from surface-sterilized, infected leaves after grinding in sterile water and plating of 10-fold dilutions of the cell suspension on semi-selective PSA medium (4). After incubation at 28°C for 5 days, each of four independent cultures yielded single yellow, mucoid Xanthomonas-like colonies (named Bur_1, Bur_2, Bur_6, and Bur_7) that resembled the positive control strain MAI10 (1). These strains originated from Musenyi (Bur_1), Gihanga (Bur_2), and Rugumbo (Bur_6 and Bur_7). Multiplex PCR assays on the four putative X. oryzae pv. oryzicola strains yielded the three diagnostic DNA fragments mentioned above. All strains were further analyzed by sequence analysis of portions of the gyrB gene using the universal primers gyrB1-F and gyrB1-R for PCR amplification (5). The 762-bp DNA fragment was identical to gyrB sequences from the Asian X. oryzae pv. oryzicola strains BLS256 (Philippines), ICMP 12013 (China), LMG 797 and NCPPB 2921 (both Malaysia), and from the African strain MAI3 (Mali) (2). The partial nucleotide sequence of the gyrB gene of Bur_1 was submitted to GenBank (Accession No. KJ801400). Pathogenicity tests were performed on greenhouse-grown 4-week-old rice plants of the cvs. Nipponbare, Azucena, IRBB 1, IRBB 2, IRBB 3, IRBB 7, FKR 14, PNA64F4-56, TCS 10, Gigante, and Adny 11. Bacterial cultures were grown overnight in PSA medium and re-suspended in sterile water (1 × 108 CFU/ml). Plants were inoculated with bacterial suspensions either by spraying or by leaf infiltration (1). For spray inoculation, four plants per accession and strain were used while three leaves per plant and four plants per accession and strain were inoculated by tissue infiltration. After 15 days of incubation in a BSL-3 containment facility (27 ± 1°C with a 12-h photoperiod), the spray-inoculated plants showed water-soaked lesions with yellow exudates identical to those seen in the field. For syringe-infiltrated leaves, the same symptoms were observed at the infiltrated leaf area. Re-isolation of bacteria from symptomatic leaves yielded colonies with the typical Xanthomonas morphology that were confirmed by multiplex PCR to be X. oryzae pv. oryzicola, thus fulfilling Koch's postulates. Bur_1 has been deposited in the Collection Française de Bactéries Phytopathogènes as strain CFBP 8170 ( http://www.angers-nantes.inra.fr/cfbp/ ). To our knowledge, this is the first report of X. oryzae pv. oryzicola causing bacterial leaf streak on rice in Burundi. Further surveys will help to assess its importance in the country. References: (1) C. Gonzalez et al., Mol. Plant Microbe Interact. 20:534, 2007. (2) A. Hajri et al. Mol. Plant Pathol. 13:288, 2012. (3) J. M. Lang et al. Plant Dis. 94:311, 2010. (4) L. Poulin et al. Plant Dis. 98:1423, 2014. (5) J. M. Young et al. Syst. Appl. Microbiol. 31:366, 2008.
Rubber tree (Hevea brasiliensis) is an important crop in tropical regions of China. In October 2013, a new stem rot disease was found on cv. Yunyan77-4 at a rubber tree plantation in Hekou, Yunnan Province. There were about 100 plants, and diseased rubber trees accounted for 30% or less. Initially, brown-punctuate secretion appeared on the stem, which was 5 to 6 cm above the ground. Eventually, the secretion became black and no latex produced from the rubber tree bark. After removing the secretion, the diseased bark was brown putrescence, but the circumambient bark was normal. Upon peeling the surface bark, the inner bark and xylem had brown rot and was musty. The junction between health and disease was undulate. On the two most serious plants, parts of leaves on the crown were yellow, and the root near the diseased stem was dry and puce. The pathogen was isolated and designated HbFO01; the pathogenicity was established by following Koch's postulates. The pathogen was cultivated on a potato dextrose agar (PDA) plate at 28°C for 4 days. Ten plants of rubber tree cv. Yunyan77-4 were selected from a disease-free plantation in Haikou, Hainan Province, and the stem diameter was about 7 cm. The bark of five plants was peeled, and one mycelium disk with a diameter of 1 cm was inserted into the cut and covered again with the bark. The other five plants were treated with agar disks as controls. The inoculation site was kept moist for 2 days, and then the mycelium and agar disk were removed. On eighth day, symptoms similar to the original stem lesions were observed on stems of inoculated plants, while only scars formed on stems of control plants. The pathogen was re-isolated from the lesions of inoculated plants. On PDA plates, the pathogen colony was circular and white with tidy edges and rich aerial hyphae. Microscopic examination showed microconidia and chlamydospores were produced abundantly on PDA medium. The falciform macroconidia were only produced on lesions and were slightly curved, with a curved apical cell and foot shaped to pointed basal cell, usually 3-septate, 16.2 to 24.2 × 3.2 to 4.0 μm. Microconidia were produced in false heads, oval, 0-septate, 6.2 to 8.2 × 3.3 to 3.8 μm, and the phialide was cylindrical. Chlamydospores were oval, 6.4 to 7.2 × 3.1 to 3.8 μm, alone produced in hypha. Morphological characteristics of the specimen were similar to the descriptions for Fusarium oxysporum (2). Genomic DNA of this isolate was extracted with a CTAB protocol (4) from mycelium and used as a template for amplification of the internal transcribed spacer (ITS) region of rDNA with primer pair ITS1/ITS4 (1). The full length of this sequence is 503 nt (GenBank Accession No. KJ009335), which exactly matched several sequences (e.g., JF807394.1, JX897002.1, and HQ451888.1) of F. oxysporum. Williams and Liu had listed F. oxysporum as the economically important pathogen of Hevea in Asia (3), while this is, to our knowledge, the first report of stem rot caused by F. oxysporum on rubber tree in China. References: (1) D. E. L. Cooke et al. Fungal Genet. Biol. 30:17, 2000. (2) J. F. Leslie and B. A. Summerell. The Fusarium Laboratory Manual, 2006. (3) T. H. Williams and P. S. W. Liu. A host list of plant diseases in Sabah, Malaysia, 1976. (4) J. R. Xu et al. Genetics 143:175, 1996.
Hylocereus undatus widely grows in southern China. Some varieties are planted for their fruits, known as dragon fruits or Pitaya, while some varieties for their flowers known as Bawanghua. Fresh or dried flowers of Bawanghua are used as routine Chinese medicinal food. Since 2008, a serious anthracnose disease has led to great losses on Bawanghua flower production farms in the Baiyun district of Guangzhou city in China. Anthracnose symptoms on young stems of Bawanghua are reddish-brown, sunken lesions with pink masses of spores in the center. The lesions expand rapidly in the field or in storage, and may coalesce in the warm and wet environment in spring and summer in Guangzhou. Fewer flowers develop on infected stems than on healthy ones. The fungus overwinters in infected debris in the soil. The disease caused a loss of up to 50% on Bawanghua. Putative pathogenic fungi with whitish-orange colonies were isolated from a small piece of tissue (3 × 3 mm) cut from a lesion margin and cultured on potato dextrose agar in a growth chamber at 25°C, 80% RH. Dark colonies with acervuli bearing pinkish conidial masses formed 14 days later. Single celled conidia were 11 to 18 × 4 to 6 μm. Based on these morphological characteristics, the fungi were identified as Colletotrichum gloeosporioides (Penz.) Penz. & Sacc (2). To confirm this, DNA was extracted from isolate BWH1 and multilocus analyses were completed with DNA sequence data generated from partial ITS region of nrDNA, actin (ACT) and glutamine synthetase (GS) nucleotide sequences by PCR, with C. gloeosporioides specific primers as ITS4 (5'-TCCTCCGCTTATTGATATGC-3') / CgInt (5'-GGCCTCCCGCCTCCGGGCGG-3'), GS-F (5'-ATGGCCGAGTACATCTGG-3') / GS-R (5'-GAACCGTCGAAGTTCCAC-3') and actin-R (5'-ATGTGCAAGGCCGGTTTCGC-3') / actin-F (5'-TACGAGTCCTTCTGGCCCAT-3'). The sequence alignment results indicated that the obtained partial ITS sequence of 468 bp (GenBank Accession No. KF051997), actin sequence of 282 bp (KF712382), and GS sequence of 1,021 bp (KF719176) are 99%, 96%, and 95% identical to JQ676185.1 for partial ITS, FJ907430 for ACT, and FJ972589 for GS of C. gloeosporioides previously deposited, respectively. For testing its pathogenicity, 20 μl of conidia suspension (1 × 106 conidia/ml) using sterile distilled water (SDW) was inoculated into artificial wounds on six healthy young stems of Bawanghua using sterile fine-syringe needle. Meanwhile, 20 μl of SDW was inoculated on six healthy stems as a control. The inoculated stems were kept at 25°C, about 90% relative humidity. Three independent experiments were carried out. Reddish-brown lesions formed after 10 days, on 100% stems (18 in total) inoculated by C. gloeosporioides, while no lesion formed on any control. The pathogen was successfully re-isolated from the inoculated stem lesions on Bawanghua. Thus, Koch's postulates were fulfilled. Colletotrichum anthracnose has been reported on Pitaya in Japan (3), Malaysia (1) and in Brazil (4). To our knowledge, this is the first report of anthracnose disease caused by C. gloeosporioides on young stems of Bawanghua (H. undatus) in China. References: (1) M. Masyahit et al. Am. J. Appl. Sci. 6:902, 2009. (2) B. C. Sutton. Page 402 in: Colletotrichum Biology, Pathology and Control. J. A. Bailey and M. J. Jeger, eds. CAB International, Wallingford, UK, 1992. (3) S. Taba et al. Jpn. J. Phytopathol. 72:25, 2006. (4) L. M. Takahashi et al. Australas. Plant Dis. Notes 3:96, 2008.
The fight against cancer has been a never-ending battle. Limitations of conventional therapies include lack of selectivity, poor penetration and highly toxic to the host. Using genetically modified bacteria as a tumour therapy agent has gained the interest of scientist from the past few decades. Low virulence and highly tolerability of Salmonella spp. in animals and humans make it as the most studied pathogen with regards to anti-tumour therapy. The present study aims to construct a genetically modified S. Agona auxotroph as an anti-tumour agent. LeuB and ArgD metabolic genes in ΔSopBΔSopD double knockout S. Agona were successfully knocked out using a Targetron gene knockout system. The knockout was confirmed by colony PCR and the strains were characterized in vitro and in vivo. The knockout of metabolic genes causes significant growth defect in M9 minimal media. Quadruple knockout ΔSopBΔSopDΔLeuBΔArgD (BDLA) exhibited lowest virulence among all of the strains in all parameters including bacterial load, immunity profile and histopathology studies. In vivo anti-tumour study on colorectal tumour bearing-BALB/c mice revealed that all strains of S. Agona were able to suppress the growth of the large solid tumour as compared with negative control and ΔLeuBΔArgD (LA) and BDLA auxotroph showed better efficacy. Interestingly, higher level of tumour growth suppression was noticed in large tumour. However, multiple administration of bacteria dosage did not increase the tumour suppression efficacy. In this study, the virulence of BDLA knockout strain was slightly reduced and tumour growth suppression efficacy was successfully enhanced, which provide a valuable starting point for the development of S. Agona as anti-tumour agent.
Fusarium species is one of the common pathogens of post-harvest disease to cause rot on tomato and other perishable vegetable fruits. The objectives of this study were to determine the diversity of Fusarium isolated species from post-harvest diseases of tomato fruit, to identify the causal organisms by using phenotype characteristics and to verify the pathogens of Fusarium fruit of tomato based on pathogenicity test. Carnation leaf-piece agar (CLA) and potato dextrose agar (PDA) media were used for phenotype-based identification of the Fusarium isolates with emphasis for characterizations of the shapes and sizes of the macroconidia and microconidia, colony features, growth rates, conidiogenous cells and chlamydospores. A total of 180 Fusarium isolates were obtained from 13 locations throughout Selangor. Fusarium solani was most abundantly isolated (34%) followed by F. semitectum (31%) and F. oxysporum (31%), F. subglutinans (3%) while the least was F. equiseti (1%). Twenty seven isolates were tested for pathogenicity test by injecting 1 mL of the conidial suspension onto healthy tomatoes. All the tested Fusarium isolates were pathogenic on tomato with different severity levels. The non-inoculated controls showed no symptoms of fruit rot. The most virulent was F. oxysporum isolate B711T with DSI 93.75%, while the least were isolates of F. solani (B647T) and F. oxysporum (B727T) with DSI 37.5%. Majority of the isolated Fusarium species can potentially produce mycotoxins as their secondary metabolites. The potential production of mycotoxins by pathogenic isolates of Fusarium species in contaminated tomato fruits could pose health hazards when consumed.
Fusarium nygamai Burgess & Trimboli was first described in 1986 in Australia (1) and subsequently reported in Africa, China, Malaysia, Thailand, Puerto Rico, and the United States. F. nygamai has been reported on sorghum, millet, bean, cotton, and in soil where it exists as a colonizer of living plants or plant debris. F. nygamai was also reported as a pathogen of the witch-weed Striga hermonthica (Del.) Benth. To our knowledge, no reports are available on its pathogenicity on crops of economic importance. In a survey of species of Fusarium causing seedling blight and foot rot of rice (Oryza sativa L.) carried out in Sardinia (Oristano, S. Lucia), F. nygamai was isolated in association with other Fusarium species-F. moniliforme, F. proliferatum, F. oxysporum, F. solani, F. compactum, and F. equiseti. Infected seedlings exhibited a reddish brown cortical discoloration, which was more intense in older plants. The identification of F. nygamai was based on monoconidial cultures grown on carnation leaf-piece agar (CLA) (2). The shape of macroconidia, the formation of microconidia in short chains and false heads, and the presence of chlamydospores were used as the criteria for identification. Two pathogenicity tests comparing one isolate of F. nygamai with one isolate of F. moniliforme were conducted on rice cv. Arborio sown in artificially infested soil in a greenhouse at 22 to 25°C. The inoculum was prepared by growing both Fusarium species in cornmeal sand (1:30 wt/wt) at 25°C for 3 weeks. This inoculum was added to soil at 20 g per 500 ml of soil. Pre- and post-emergence damping-off was assessed. Both F. nygamai and F. moniliforme reduced the emergence of seedlings (33 to 59% and 25 to 50%, respectively, compared to uninoculated control). After 25 days, the seedlings in infested soil exhibited a browning of the basal leaf sheaths, which progressed to a leaf and stem necrosis. Foot rot symptoms caused by F. nygamai and F. moniliforme were similar, but seedlings infected by F. nygamai exhibited a more intense browning on the stem base and a significant reduction of plant height at the end of the experiment. Either F. nygamai or F. moniliforme were consistently isolated from symptomatic tissue from the respective treatments. References: (1) L. W. Burgess and D. Trimboli. Mycologia 78:223,1986. (2) N. L. Fisher et al. Phytopathology 72:151,1982.
Mango (Mangifera indica L.; family Anacardiaceae) is one of the world's most important fruit crops and is widely grown in tropical and subtropical regions. Since 2001, a leaf spot disease was found in mango orchards of Taiwan. Now, the disease was observed throughout (approximately 21,000 ha) Taiwan in moderate to severe form, thus affecting the general health of mango trees and orchards. Initial symptoms were small, yellow-to-brown spots on leaves. Later, the irregularly shaped spots, ranging from a few millimeters to a few centimeters in diameter, turned white to gray and coalesced to form larger gray patches. Lesions had slightly raised dark margins. On mature lesions, numerous black acervuli, measuring 290 to 328 μm in diameter, developed on the gray necrotic areas. Single conidial isolates of the fungus were identified morphologically as Pestalotiopsis mangiferae (Henn.) Steyaert (2,3) and were consistently isolated from the diseased mango leaves on acidified (0.06% lactic acid) potato dextrose agar (PDA) medium incubated at 25 ± 1°C. Initially, the fungus grew (3 mm per day) on PDA as a white, chalky colony that subsequently turned gray after 2 weeks. Acervuli developed in culture after continuous exposure to light for 9 to 12 days at 20 to 30°C. Abundant conidia oozed from the acervulus as a creamy mass. The conidia (17.6 to 25.4 μm long and 4.8 to 7.1 μm wide) were fusiform and usually straight to slightly curved with four septa. Three median cells were olivaceous and larger than the hyaline apical and basal cells. The apical cells bore three (rarely four) cylindrical appendages. Pathogenicity tests were conducted with either 3-day-old mycelial discs or conidial suspension (105 conidia per ml) obtained from 8- to 10-day-old cultures. Four leaves on each of 10 trees were inoculated. Before inoculation, the leaves were washed with a mild detergent, rinsed with tap water, and then surface sterilized with 70% ethanol. Leaves were wounded with a needle and exposed to either a 5-mm mycelial disc or 0.2 ml of the spore suspension. The inoculated areas were wrapped with cotton pads saturated with sterile water and the leaves were covered with polyethylene bags for 3 days to maintain high relative humidity. Wounded leaves inoculated with PDA discs alone served as controls. The symptoms described above were observed on all inoculated leaves, whereas uninoculated leaves remained completely free from symptoms. Reisolation from the inoculated leaves consistently yielded P. mangiferae, thus fulfilling Koch's postulates. Gray leaf spot is a common disease of mangos in the tropics and is widely distributed in Africa and Asia (1-3); however, to our knowledge, this is the first report of gray leaf spot disease affecting mango in Taiwan. References: (1) T. K. Lim and K. C. Khoo. Diseases and Disorders of Mango in Malaysia. Tropical Press. Malaysia, 1985. (2) J. E. M. Mordue. No. 676 in: CMI Descriptions of Pathogenic Fungi and Bacteria. Surrey, England, 1980. (3) R. C. Ploetz et al. Compendium of Tropical Fruit Diseases. The American Phytopathological Society. St. Paul, MN, 1994.