First Report of Powdery Mildew Caused by Erysiphe diffusa, Oidium neolycopersici, and Podosphaera xanthii on Papaya in Taiwan.

Powdery mildew can be found in most papaya (Carica papaya L.) fields during the winter and spring seasons in Taiwan. It usually causes severe yellowing of the leaf lamina and petiole and serious defoliation. Three types of powdery mildew fungi were isolated from papaya leaves in Chiayi City (23.28°N, 120.28°E) at the beginning of 2008. Conidia of the first one were single, globose, hyaline, and 24 to 36 × 14 to 18 µm (average 30.2 × 15.6 µm) without fibrosin bodies and with straight or occasionally flexuous conidiophores at the base. The second one had short pseudo-chains of two to four conidia which were ellipsoidal to ovoid, hyaline, and 24 to 40 × 12 to 16 µm (average 29.7 × 13.4 µm) without fibrosin bodies. The third type had chains of ellipsoidal conidia that were hyaline, 24 to 28 × 12 to 16 µm (average 26.3 × 14.4 µm) and contained fibrosin bodies. To confirm the identity of the three fungi, the internal transcribed spacer (ITS) region of rDNA was amplified using the primer pairs G1 (5'-TCC GTA GGT GAA CCT GCG GAA GGA T-3')/Ed2 (5'-CGC GTA GAG CCC ACG TCG GA-3'), G1 (5'-TCC GTA GGT GAA CCT GCG GAA GGA T-3')/On2 (5'-TGT GAT CCA TGT GAC TGG AA-3'), and S1 (5'-GGA TCA TTA CTG AGC GCG AGG CCC CG-3')/S2 (5'-CGC CGC CCT GGC GCG AGA TAC A-3'). The alignment of obtained sequences (GenBank Accession Nos. GU358452, 507 bp; GU358451, 580 bp; and GU358450, 455 bp) showed a sequence identity of 100, 99, and 99% with the ITS sequences of Erysiphe diffusa, Oidium neolycopersici, and Podosphaera xanthii (GenBank Accession Nos. FJ378880, EU909694, and GQ927254), respectively. On the basis of morphological characteristics and ITS sequence similarities, these fungi were identified as E. diffusa (Cooke & Peck) U. Braun & S. Takam., O. neolycopersici L. Kiss, and P. xanthii (Castagne) U. Braun & S. Takam., respectively (1,3). Single colonies on papaya leaves infected with powdery mildew were identified in the laboratory and maintained on papaya leaves as inoculum. Pathogenicity was confirmed through inoculations by gently pressing a single colony of each fungus onto leaves of healthy papaya seedlings (cv. Horng-Fe). Five seedlings were inoculated for each fungus and then covered with plastic bags for 2 days. Five noninoculated seedlings served as control. After inoculation, treated plants were maintained separately from the control in different rooms of a greenhouse at 25°C under natural daylight conditions. Seven days after inoculation, typical symptoms of powdery mildew were observed on inoculated plants, but not on noninoculated plants. The same species from diseased lesions following artificial inoculation with each fungus were identified with light microscopy. Papaya was previously described as a host to O. caricae Noack in many tropical and subtropical areas of the world including Taiwan (2). However E. cruciferarum, Golovinomyces cichoracearum, Oidiopsis sicula, O. caricae, O. caricae-papayae, O. caricicola, O. indicum, O. papayae, Ovulariopsis papayae, P. caricae-papayae, P. macularis, P. xanthii, and Streptopodium caricae were reported to infect papaya (4). To our knowledge, this is the first report of papaya powdery mildew caused by E. diffusa and O. neolycopersici in the world and the first report of the three fungi found on papaya in Taiwan. References: (1) U. Braun and S. Takamatsu. Schlechtendalia 4:1, 2000. (2) H. S. Chien and H. L. Wang. J. Agric. Res. China 33:320, 1984. (3) L. Kiss et al. Mycol. Res. 105:684, 2001. (4) J. R. Liberato et al. Mycol. Res. 108:1185, 2004.

First Report of Anthracnose on Cucurbitaceous Crops Caused by Glomerella magna in Taiwan.

During the summer and fall of 2006, leaf anthracnose samples were collected from fields of cucumber (Cucumis sativus L.), calabash gourd (Lagenaria siceraria (Molina) Standley), and luffa (Luffa cylindrica (L.) M. Roem.) in southern Taiwan. On cucumber leaves, spots start as water-soaked areas and expand into brown spots. Leaf lesions on calabash gourd and luffa begin as water soaked and then become light brown-to-reddish spots. Centers of lesions sometimes fall out; giving infected leaves a shot-hole appearance. Small pieces (approximately 2 × 2 mm) of diseased leaf tissue from margins of individual lesions were surface disinfected in 1% sodium hypochlorite solution for 1 min, rinsed in sterile water, plated on water agar, and incubated at 25°C. After 4 days, mycelium was isolated, transferred to potato dextrose agar (PDA), and then incubated at 25°C in a 12-h light/darkness regimen. Fast-growing colonies on PDA were white to orange or pink with abundant acervuli but no perithecium. One-celled conidia were ovoid to oblong and 12 to 20 × 4 to 6 (15.9 × 5.0) µm. The morphological traits were identical to those of Colletotrichum magna (teleomorph Glomerella magna Jenkins & Winstead) and clearly distinct from those of C. orbiculare (Berk. & Mont.) Arx (synonym C. lagenarium (Pass.) Ellis & Halst. (conidia were mostly oblong, measuring 7 to 11 × 2 to 6 [9.3 × 4.2] µm, with slow-growing gray colonies) (2,3). Koch's postulates were performed to verify that the isolates were capable of causing anthracnose on cucurbitaceous crops. Pathogenicity tests were conducted in the greenhouse at 25°C under natural daylight conditions. Isolate C0604 was grown on PDA for 14 days and a spore suspension was made (106 spores/ml). Three 14-day-old seedlings at the two- to three-leaf stage of muskmelon (Cucumis melo L. var. reticulatus Naud., cv. Sapphire), squash (Cucurbita moschata Duch., cv. Achen), calabash gourd (cv. Huapu), and luffa (cv. 623) were sprayed with the spore suspension and then covered with plastic bags. Control treatments were sprayed with sterile water. After 2 days, the bags were removed. Typical anthracnose symptoms developed on all inoculated seedlings 7 days after inoculation. G. magna was reisolated from inoculated leaves following the protocol used for the original isolation. Control seedlings developed no symptoms. To confirm the identity of the fungus, PCR amplification and DNA sequencing of the internal transcribed spacer 1 (ITS1)-5.8S-ITS2 of rRNA gene of the isolate C0604 was performed by using ITS1/ITS4 as the PCR and sequencing primers. Sequence analysis of the 558-bp PCR product (GenBank Accession No. GU358453) showed 100% identity to the rRNA sequence of G. magna (GenBank Accession No. DQ003103) (1). PCR amplification of the ITS region was also carried out using species-specific primer GmF (5'- GTG AAC ATA CCT CAA ACG TTG CC -3')/GmR (5'- GGA GGG TCC GCC ACT GTA TTT CG -3') designed in this study. A DNA fragment of approximately 378 bp was amplified from nine isolates of G. magna, whereas no amplification products were obtained from reference cultures of C. gloeosporioides (Penz.) Penz. & Sacc. and C. orbiculare. To our knowledge, this is the first report of G. magna causing anthracnose on cucurbitaceous crops in Taiwan. References: (1) M. Du et al. Mycologia 97:641, 2005. (2) S. F. Jenkins, Jr. and N. N. Winstead. Phytopathology 54:452, 1964. (3) T. A. Zitter et al., eds. Compendium of Cucurbit Diseases. The American Phytopathological Society, St. Paul, MN, 1996.

First Report of Papaya Scab Caused by Cladosporium cladosporioides in Taiwan.

In March of 2008, a leaf scab disease was observed in a papaya (Carica papaya L.) orchard at Guoshing, 24.03°N, 120.51°E, in Nantou County, Taiwan. Infected papayas developed symptoms of numerous, pale green, water-soaked areas, 0.5 to 1.5 mm. Infected leaves gradually turned white to gray on the upper surface and small, circular swellings were observed on the abaxial surface. Lesions may coalesce to cover more than 50% of the leaf, rendering them to fall prematurely. Lesions on the lower surface of the leaves were covered with olive-gray patches of mycelia and abundant conidia. Pieces (~2 × 2 mm) of diseased leaf tissue from margins of individual lesions were surface disinfected in 1% sodium hypochlorite solution for 1 min, rinsed in sterile water, plated on water agar, and incubated at 25°C. After 4 days, mycelium was isolated and transferred to potato dextrose agar (PDA). Five isolates (Cc-5 to Cc-9) were isolated and identified as Cladosporium cladosporioides (Fresen.) de Vries based on the velvety, olive-brown with almost black reverse colony color and dimensions and color of conidia and conidiophores (1). Conidia formed in long branched chains that readily disarticulate, single celled, elliptical to limoniform, 2 to 9 (4.6) × 2 to 3 (2.2) µm. Conidia were pale-to-olive brown and smooth to verruculose. Ramoconidia were 0 to 1 septate, 6 to 14 (9.4) × 2 to 4 (2.7) µm, smooth or sometimes minutely verruculose. Conidiophores were pale-to-olive brown, macro- and micronemateus, smooth or sometimes verruculose, 68 to 244 (141.7) × 3.2 to 4 (3.9) µm. To confirm the identity of the fungus, the internal transcribed spacer (ITS) 1 and 4 regions and mitochondrial small subunit (mtSSU) rDNA were sequenced (GenBank Accession Nos. EU935608 and FJ362555), which had 99% homology to the ITS and mtSSU rDNA of C. cladosporioides (GenBank Accession Nos. EU497957 and AY291273, respectively). Pathogenicity tests were conducted in the greenhouse at 25°C with natural daylight conditions. Fungal isolate Cc-6 was used; it was grown on PDA for 6 days and a spore suspension was made (106 spores/ml). Three papaya seedlings (cv. Horng-Fe) were sprayed with the spore suspension and covered with plastic bags. Control treatments were sprayed with sterile water. After 2 days, the bags were removed. Symptoms developed on all inoculated seedlings 4 days after inoculation. In all cases, the typical scab symptom, pale green, water-soaked areas on the lower leaf surface, were observed. C. cladosporioides was reisolated from inoculated leaves following the procedure used for the original isolation. Control seedlings developed no symptoms. The five isolates are being maintained at the DBST, NCYU, Taiwan. Previously, papaya scab reported in China was caused by C. cariciolum Corda (2), C. caricinum C. F. Zhang et P. K. Chi (3), and C. cladosporioides (4). To our knowledge, this is the first report of C. cladosporioides causing papaya scab in Taiwan. References: (1) M. B. Ellis. Dematiaceous Hyphomycetes. CMI, Kew, Surrey, England, 1971. (2) H.-H. Peng and Z.-Y. Zhang. J. Yunnan Agric. Univ. 12:23, 1997. (3) C.-F. Zhang. Ph.D. thesis. South China Agricultural University, Guangzhou, P.R.C., 1995. (4) Z. Y. Zhang et al. Flora Fungorum Sinicorum 14:1, 2003.

Comparison of the association of age with the infection of Salmonella and Salmonella enterica Serovar Typhimurium in Pekin ducks and Roman geese.

Nontyphoid Salmonella have a broad host range in poultry and mammals, and serovar Typhimurium is a threat to public health. In this study, normal and sick ducks and geese were collected from 12 farms in Taiwan to investigate the age-associated infection of Salmonella and Salmonella Typhimurium in Roman geese (Anser anser domesticus) and Pekin ducks (Anas platyrhynchos domesticus). In normal birds, the prevalence of Salmonella differed between species, and with age [e.g., 1-wk group, 37.5% (30/80) for ducks and 5.2% (6/116) for goslings (P < 0.05) vs. 4-wk group, 1% (1/96) for ducks and 12.1% (21/174) for geese]. Salmonella Typhimurium was identified from the visceral organs of moribund young geese suffering with colibacillosis and riemerellosis isolated from 2 goose farms (farm A and B, respectively). At farm B, 22.9% (27/118) of 4-wk geese with diarrhea were Salmonella Typhimurium-positive compared with 4.6% (8/174) of 4-wk normal geese. All Salmonella Typhimurium strains except one harbored a 94.7-kb virulence plasmid. Subcutaneous injection of Salmonella Typhimurium isolate 91NGL1 resulted in different clinical signs and pathogenesis between ducks and geese. In addition, the mean infectivity dose ratios of ducks to geese were 3.2 and 85.0 for 4- and 12-d birds, respectively, suggesting that goslings were more susceptible to Salmonella Typhimurium and resistance to Salmonella Typhimurium increased with age, especially for ducks. Therefore, Salmonella Typhimurium infection should be more common in goose farms than in duck farms, especially in the younger birds.

First Report of Simplicillium lanosoniveum Causing Brown Spot on Salvinia auriculata and S. molesta in Taiwan.

Salvinia spp. are small, floating ferns that grow in long chains of two oval leaves and a root-like third leaf. S. natans (L.) All., a native floating fern distributed in paddy fields, ponds, and ditches in Taiwan, has become critically endangered. Another two exotic species, S. auriculata Aublet (eared salvinia) and S. molesta Mitchell (giant salvinia), are sold in increasing frequency at local flower markets and aquarium shops and pose a serious threat when they find their way into the natural environment. Brown spot of S. auriculata was found in a home aquarium in December 2006 in Chiayi, Taiwan. Symptoms of the disease included many, irregular, dark brown spots on both upper and lower leaf surfaces. Lesions on the upper surface of the leaves were covered with white patches of mycelia and abundant conidia. Small pieces (approximately 2 × 2 mm) of diseased leaf tissue from the margin of individual lesions were surface disinfected in 1% sodium hypochlorite solution for 1 min, rinsed in sterile water, plated on water agar, and incubated at 25°C. Six isolates of the fungus were then isolated and transferred to potato dextrose agar (PDA). Isolate Cs0701 was identified morphologically as Simplicillium lanosoniveum (van Beyma) Zare & W. Gams on the basis of morphology of asexual reproduction structures and rDNA sequence analysis (1). In culture, this fungus formed whitish-to-whitish yellow, pulvinate colonies with matted surfaces. The reverse side of cultures was yellow to light brown. Small, ovate to spherical, hyaline conidia, 2.2 to 3.0 × 1.6 to 2.0 µm (average 2.4 × 1.9 µm) were formed. To confirm the identity of the fungus, PCR amplification and DNA sequencing of the internal transcribed spacer (ITS1-5.8S-ITS2 rRNA gene cluster) was conducted on isolates Cs0701 and Cs0702. The sequence of the PCR product was compared with sequences of closely related species listed in the GenBank database. Except for a single nucleotide, the ITS sequence of both isolates (480 bp; GenBank Accession No. EU939525) was identical to the rRNA of Simplicillium lanosoniveum (GenBank Accession No. AJ292396). Koch's postulates were performed to confirm the pathogenicity of the fungus on S. auriculata and S. molesta. After 14 days of growth on PDA, a spore suspension of isolate Cs0701 (106 spores per ml) was sprayed onto approximately 5 and 10 g of healthy S. auriculata and S. molesta plants, respectively, floated in 500-ml beakers filled with 300 ml of tap water. All treatments, including controls misted with sterile water, were replicated three times. The beakers were covered with plastic bags and placed in a growth chamber maintained at 25°C with 12-h fluorescent light cycles. After 2 days, the bags were removed. Symptoms developed on all inoculated plants 4 days after inoculation. In all cases, typical brown spots were observed. Simplicillium lanosoniveum was reisolated from all surface-disinfested infected tissues. Control plants developed no symptoms. Six isolates of the fungus are being maintained at the Department of Microbiology and Immunology, National Chiayi University, Taiwan. To our knowledge, this is the first report of Simplicillium lanosoniveum causing brown spot of S. auriculata and S. molesta in Taiwan. Reference: (1) R. Zare and W. Gams. Nova Hedwigia 73:1, 2001.