First report of leaf blight disease of <Emphasis Type="Italic">Gloriosa superba</Emphasis> L. caused by <Emphasis Type="Italic">Alternaria alternata</Emphasis> (Fr.) Keissler in India

Leaf blight disease was found on Gloriosa superba L. (Liliaceae), an endangered, herbaceous, perennial, climbing lily that produces colchicine, in West Bengal, India in 2004. Small brownish spots on leaves developed into concentric rings, which eventually darkened and coalesced to blight the entire leaf. The causal fungus was morphologically identified as Alternaria alternata (Fr.) Keissler. This is the first record of A. alternata on G. superba.     

Specific inhibition of spore germination of <Emphasis Type="Italic">Alternaria brassicicola</Emphasis> by fistupyrone from <Emphasis Type="Italic">Streptomyces</Emphasis> sp. TP-A0569

Fistupyrone (FP), a metabolite from Streptomyces sp. TP-A0569, inhibited the in vivo infection of Chinese cabbage seedlings by Alternaria brassicicola. To detect the possible action sites of FP, the effect of FP on the infection behavior of A. brassicicola and A. alternata was investigated. When spores of A. brassicicola were suspended in FP solution and inoculated on host leaves, FP at 0.1ppm significantly inhibited spore germination, appressorial formation, and infection hypha formation of A. brassicicola. Host-specific AB-toxin production and lesion formation by A. brassicicola spores were also reduced significantly by treatment with FP 1ppm. The effect of FP seemed to be irreversible because significant washing of FP-treated spores with distilled water (DW) did not change the inhibitory effects. In contrast, A. alternata isolates such as Japanese pear pathotype, apple pathotype, and saprophyte behaved almost equally in both FP- and DW-treated spores. Mycelial dry weight in potato dextrose broth and mycelial diameters on potato dextrose agar, gelatin glucose agar, and Czapek solution agar of both A. brassicicola and A. alternata were not different with or without addition of FP. These results indicate that FP at low concentrations has a fungicidal effect on spores of A. brassicicola but not on spores of A. alternata; FP also does not affect the vegetative phase of these fungi.     

Alternaria Brown Spot of Minneola in Greece; Evaluation of Citrus Species Susceptibility

During the last three years, a new disease was observed in northwestern Greece on Minneola trees, hybrid of mandarin and grapefruit. On May small brown necrotic leaf spots surrounded by yellow halo areas of various sizes appeared and covered a major portion of the leaves with extension of necrosis into the veins. On young fruits small, slightly depressed black spots were the first symptoms, which later became 2–7 mm in diameter. Brown spots were observed on the leaves and fruits in several orchards in the same area, causing leaves and fruits to drop. In some orchards over 50% of the fruits were affected. From the fruit and leaf spots the typical small-spore species Alternaria alternata was isolated. Pathogenicity tests were performed by artificially inoculating fruits of Minneola, common mandarin and Clementine. The symptoms of the disease were reproduced only on fruits of Minneola hybrids by the specific strain of the fungus Alternaria alternata pv. citri. Different citrus susceptibility tests indicated that mandarins Minneola, Nova and Page were very susceptible to tested isolates while Clementine SRA and Poros Clementine were not. All lemons and lime Seedless were not susceptible. Grapefruit New Hall was not susceptible, while the Star Ruby was. Orange Lane Late, Navel Late, Oval Poros, Olinda, Navel Athos were not susceptible and only Moro showed reaction being slightly susceptible only to one isolate.     

Pathological evaluation of host-specific AAL-toxins and fumonisin mycotoxins produced by Alternaria and Fusarium species

Host-specific AAL-toxins and mycotoxin fumonisins are structurally related and were originally isolated from the tomato pathotype of Alternaria alternata and from Fusarium verticillioides, respectively. Previous reports on the production of fumonisin derivatives by the tomato pathotype suggested a possible involvement in the pathogenicity of the pathogen. Here, we have evaluated the role of fumonisin in A. alternata–tomato interactions. The results indicate that highly pathogenic isolates of A. alternata tomato pathotype produce AAL-toxin as the sole toxin, strongly implicating it as a pathogenicity factor. The related compound, fumonisin, is also toxigenic and has infection-inducing activity on susceptible tomato plants.     

Identification of putative defense-related genes in Japanese pear against <Emphasis Type="Italic">Alternaria alternata</Emphasis> infection using suppression subtractive hybridization and expression analysis

The Japanese pear pathotype of Alternaria alternata, a toxin-dependent necrotrophic pathogen, causes black spot of Japanese pear by producing the host-specific AK-toxin. Pre-inoculation with nonpathogenic A. alternata or pretreatment with an elicitor prepared from A. alternata reduced disease symptoms caused by the pathogen. Salicylic acid- and jasmonic acid-dependent signaling pathways are not involved in the induced resistance to infection by the pathogen. The expression of multiple defense-related genes in Japanese pear leaves inoculated with nonpathogenic A. alternata was examined using suppression subtractive hybridization. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users. The nucleotide sequence data reported are available in the DDBJ/EMBL/GenBank database as accessions DC993229–DC993535.     

Detoxification of α-tomatine by tomato pathogens Alternaria alternata tomato pathotype and Corynespora cassiicola and its role in infection

In tomato plants, α-tomatine, a steroidal glycoalkaloid saponin, inhibits fungal growth. Tomato pathogens that produce host-specific toxins, Alternaria alternata tomato pathotype causing Alternaria stem canker and Corynespora cassiicola causing Corynespora target spot, were investigated for sensitivity to α-tomatine. Although spore germination of A. alternata pathogenic and nonpathogenic to tomato and of C. cassiicola pathogenic to tomato was not affected by 0.1 mM α-tomatine, spore germination of C. cassiicola nonpathogenic to tomato was significantly inhibited. This result showed that A. alternata, regardless of its pathogenicity, and only the C. cassiicola pathogenic to tomato are resistant to α-tomatine. Germinating spores of A. alternata and C. cassiicola resistant to α-tomatine detoxified α-tomatine by degrading it to a less polar product. After inoculation of tomato leaves, spores of A. alternata and C. cassiicola nonpathogenic to tomato germinated and formed appressoria, but did not form infection hyphae in host tissues. When a host-specific toxin (CCT-toxin) produced by C. cassiicola pathogenic to tomato was added to nonpathogenic spores, colonization within leaves was observed in A. alternata, but not in C. cassiicola. On the other hand, when spores of C. cassiicola nonpathogenic to tomato were suspended in spore germination fluid of nonpathogenic A. alternata with α-tomatine detoxification activity, the fungus could be induced to colonize leaves in the presence of CCT-toxin. These results indicate that A. alternata tomato pathotype and C. cassiicola pathogenic to tomato detoxify α-tomatine during infection and that this detoxification is essential for host colonization by pathogens that produce host-specific toxins.     

Frequency of Alternaria brassicicola in commercial cabbage seeds in Japan

Plug seedlings, widely used in cabbage cultivation in Japan, are often infected by seed-borne pathogens, especially the serious pathogen Alternaria brassicicola. Because information on seed infestation is scant in Japan, we investigated fungal infestation in commercial batches of cabbage seeds produced between 1984 and 2001. A total of 123 lots were divided into six groups by production period (1984–1989, 1994–1998, and 2001) and by use or nonuse of fungicide. One hundred seeds from each lot were incubated separately on agar at 25°C to isolate the predominant fungus. Alternaria brassicicola was isolated most frequently, 0%–94% of the seeds depending on seed lot or 6%–21% of the seeds grouped by production period and fungicide treatment. Thus, the pathogen was isolated even from seeds refrigerated for 17 years. Alternaria brassicicola accounted for 57%–95% of all isolated fungi by the group and was higher on older or fungicide-treated seeds. Seeds that were not treated with fungicide in lots grouped by production districts in western Japan were infested with A. brassicicola at a rate of over 12%, higher than that in the eastern region (<4%). Infestation was higher in the warmer areas of Japan. Eighty-five isolates, other than A. brassicicola, produced spots on cabbage cotyledons, although they were not isolated as frequently: less than 5% of seeds by group separated by production period and fungicide treatment. Most of these isolates were Alternaria alternata. This is the first report on the frequency of fungal infestation of commercial cabbage seeds in Japan.     

Variability of Cuban and International Populations of Alternaria solani from Different Hosts and Localities: AFLP Genetic Analysis

As causal agent of early blight disease in tomato and potato, Alternaria solani is an internationally important horticultural pathogen. Genetic variability was surveyed by amplified fragment length polymorphism analysis in a total of 112 isolates from potato and tomato, representing pathogen populations from different Cuban provinces together with isolates from the USA, Brazil, Turkey, Greece and China. Also included in the analysis were isolates from catenulated Alternaria spp. from Brazil, Canada, Greece and Russia, along with single isolates of Alternaria porri, Alternaria alternata and a Curvularia sp. UPGMA clustering revealed a differentiation between the isolates of A. solani and all other species with the exception of A. porri which could not be distinguished from A. solani. Among the isolates of A. solani, two distinct subclusters were formed, with high genetic significance revealed by bootstrapping, corresponding to a general subdivision based on the respective solanaceous host. The results are discussed with regard to potential host specificity of A. solani on tomato and potato, and in terms of the comparative contributions of regional and international genetic variability in populations of this ubiquitous plant pathogen.     

Multiple copies of <Emphasis Type="Italic">AMT2</Emphasis> are prerequisite for the apple pathotype of <Emphasis Type="Italic">Alternaria alternata</Emphasis> to produce enough AM-toxin for expressing pathogenicity

The apple pathotype of Alternaria alternata produces the cyclic depsipeptide AM-toxin and causes Alternaria blotch of apple. Previously, we cloned AMT2 from the apple pathotype as an orthologue of AFTS1, which is required for biosynthesis of the decatrienoic acid ester AF-toxin I of the strawberry pathotype. These genes were predicted to encode aldo-keto reductases involved in biosynthesis of a common precursor, 2-hydroxy-isovaleric acid, of AF-toxin I and AM-toxin. In this study, we analyzed the function of AMT2 in AM-toxin biosynthesis in the apple pathotype. DNA gel blot analysis of the apple pathotype strain IFO8984 with five restriction enzymes suggested that this strain has a single copy of AMT2 in the genome. However, gene disruption experiments showed that IFO8984 probably has three copies of AMT2. We made mutants having one or two copies of AMT2 disrupted. The single-copy mutants produced less AM-toxin than did the wild type and were still as pathogenic as the wild type. The two-copy mutants produced trace or undetectable amounts of AM-toxin and were markedly reduced in pathogenicity. Thus, AMT2 was verified to be required for AM-toxin biosynthesis and hence pathogenicity. The fact that the two-copy mutants have a remaining copy of AMT2 suggests that multiple copies of AMT2 are prerequisite for the pathogen to produce enough AM-toxin for full pathogenicity.     

<Emphasis Type="Italic">Alternaria alternata</Emphasis> toxin detection by fluorescence derivatization and separation by high performance liquid chromatography

An improved high performance liquid chromatographic (HPLC) method was developed for the detection and quantitation of host-specific AA-toxin produced byAlternaria alternata, a water hyacinth pathogen. Precolumn derivatization of the toxin witho-phthalaldehyde (OPA) plus mercaptoethanol yielded highly fluorescent products showing well resolved peaks on reverse-phase HPLC. The minimum amounts of AA-toxins detectable by this method were approximately 1 ng. This method seems to offer advantages over conventional techniques, because prechromatographic derivatization of the toxin with OPA is rapid and easy, and fluorescent derivatives permit excellent detection sensitivity. Toxin production in culture filtrates and spore-germinated fluids of the pathogen were analyzed quantitatively using this procedure. http://www.phytoparasitica.org posting Dec. 16, 2002.     

Black Spot of Peach Caused by <Emphasis Type="Italic">Alternaria alternata</Emphasis> (Fr.) Keissler

A new disease of peach (Prunus persica Batsch var. vulgaris Maxim.), causing brown, sunken lesions and brownish to blackish brown spots with cracks on peach fruits, was found in Okayama prefecture, Japan, in 1995. The disease was observed not only on peach fruits but also on twigs and leaves. An Alternaria sp. was consistently isolated from these diseased fruits, twigs and leaves. The isolates were pathogenic to peach fruits and leaves. Based on the morphological characteristics, the causal fungus was identified as Alternaria alternata (Fr.) Keissler. After cross-inoculation with isolates from peach, Japanese pear and apple, the isolates were found to be pathogenic only to their original host. This is the first report on a peach disease caused by a host-specific A. alternata; therefore, the common name of black spot (`Kokuhanbyo' in Japanese) was proposed. Received 25 June 1999/ Accepted in revised form 12 October 1999     

北京市生菜链格孢根腐病病原菌鉴定

为明确北京市生菜链格孢根腐病的病原菌种类,采用常规组织分离法分离获得病原菌,依据柯赫氏法则对病原菌进行致病力检测,并利用分子生物学技术结合形态学鉴定确定病原菌分类地位。结果显示,从生菜病样组织中分离到2种病原菌共18株,形态学鉴定结果为芸薹链格孢Alternaria brassicae和万寿菊链格孢A. tagetica,分离比例分别为55.6%和44.4%,且二者均能单独侵染生菜根部,前者致病力较后者强,亦能复合侵染。对致病菌株进行GAPDH基因的PCR扩增和测序,并建立了基于GAPDH基因序列的系统发育树,聚类分析结果与形态学鉴定结果一致,因此证实北京市生菜链格孢根腐病是由芸薹链格孢和万寿菊链格孢复合侵染所致。     

Improved control of moldy-core decay (<Emphasis Type="Italic">Alternaria alternata</Emphasis>) in Red Delicious apple fruit by mixtures of DMI fungicides and captan

The sterol biosynthesis inhibitors bromuconazole and difenoconazole and tank mixes of each fungicide with captan were applied to apples and evaluated as controls for moldy-core and fruit decay caused by Alternaria alternata. Effectiveness of a mixture of bromuconazole and captan in controlling colonization by the fungus was also evaluated. Decay formation by A. alternata on mature detached fruits was partially inhibited by bromuconazole at 0.5 μg ml⁻¹ and was completely inhibited at 50 μg ml⁻¹; it was significantly affected by either bromoconazole at 5 μg ml⁻¹ or captan at 1,250 μg ml⁻¹, and was completely inhibited by their mixture. In general, three foliar applications of bromuconazole or difenoconazole in the field, during the bloom period, reduced the numbers of infected fruits by 40–60% compared with untreated control trees. However, tank mixes of either fungicide with captan improved control of moldy-core in fruits at harvest. Tank mixtures of bromuconazole and captan also significantly reduced the percentage of fruits colonized by A. alternata when sampled at various days after full bloom. Artificial inoculations in the orchard at full bloom did not change the inhibitory effects of the tank mixtures. Large-scale demonstration trials in commercial orchards supported these findings. The inhibitory effects of tank mixes on decay development in detached fruits, and on moldy-core in the field indicate that a control programme based on mixtures of either bromuconazole or difenoconazole with captan during the bloom period can effectively reduce moldy-core on Red Delicious apples.     

Detection of fungi producing infection-inhibiting metabolites against <Emphasis Type="Italic">Alternaria alternata</Emphasis> Japanese pear pathotype from fungi inhabiting internal tissues of Japanese pear shoots

Fungi inhabiting Japanese pear were isolated from internal tissues of cv. Nijisseiki, and culture filtrates (CFs) of 100 isolates were evaluated for their inhibitory activity against infection by Alternaria alternata Japanese pear pathotype. CFs of 11 isolates inhibited lesion formation on the pear by the pathogen. Among these isolates, CFs of five isolates inhibited spore germination. CFs of the six other isolates inhibited appressorial formation, infection hypha formation, AK-toxin production, or a combination of these actions. Analysis of sequence homology in the rDNA ITS1 regions of these isolates showed that most isolates had high homology with some fungal endophytes.     

烟草赤星病菌对苯醚甲环唑和氟环唑敏感性测定及田间防效试验

为明确山东省主要烟草产区烟草赤星病菌Alternaria alternata对苯醚甲环唑和氟环唑的敏感性,采用菌丝生长速率法检测苯醚甲环唑和氟环唑对86株烟草赤星病菌菌株的EC_(50),用Spearman’s秩相关系数法分析供试菌株对2种杀菌剂敏感性的相关性,并评价其对烟草赤星病的田间防效。结果表明,苯醚甲环唑对烟草赤星病菌菌丝生长EC_(50)值介于5.3883～8.0316 mg/L之间,平均值为6.9594 mg/L,敏感性频率分布呈连续性偏正态分布,田间烟草赤星病菌对苯醚甲环唑仍然较为敏感,可建立田间烟草赤星病菌对苯醚甲环唑的敏感基线。氟环唑对烟草赤星病菌菌丝生长的EC_(50)值介于0.1253～0.2517 mg/L,平均值为0.1773 mg/L,敏感性频率分布呈连续性偏正态分布,可建立田间烟草赤星病菌对氟环唑的敏感基线。相关性分析结果表明,烟草赤星病菌对苯醚甲环唑和氟环唑敏感性之间不存在明显的相关性。田间试验结果表明,30%苯醚甲环唑悬浮剂对烟草赤星病具有较好的防效,其有效成分150 g/hm~2处理的防效最高,为73.99%;12.5%氟环唑悬浮剂对烟草赤星病的最高防效为70.57%。表明2种三唑类杀菌剂对烟草赤星病防效较好。     

Induced resistance in tomato plants to the toxin-dependent necrotrophic pathogen Alternaria alternata

A necrotrophic pathogen, the tomato pathotype of Alternaria alternata (Aa) causes Alternaria stem canker on tomato. Its pathogenicity depends on the production of host-specific AAL-toxin. Pre-inoculation with nonpathogenic Aa or pretreatment an elicitor prepared from Aa reduced disease symptoms by the pathogen. Salicylic acid (SA)- and jasmonic acid (JA)-dependent defense responses in tomato are not involved in the resistance to the pathogen induced by nonpathogenic Aa. The results suggest that an alternative and unknown signaling pathway independent of SA- and JA-signaling might modulate the induced resistance by activating the expression of the multiple defense genes.     

白术黑斑病菌对甲基硫菌灵和腐霉利的抗性检测及对咪鲜胺敏感性基线的建立

黑斑病是影响白术种植的主要病害之一,2015—2017年从浙江省磐安、天台、新昌和嵊州4地采集、分离得到137株白术黑斑病菌Alternaria alternata,采用菌丝生长速率法检测其对甲基硫菌灵和腐霉利的抗性。结果表明:供试的黑斑病菌群体 (n = 137) 对甲基硫菌灵的高水平抗性频率为77.4%,中等水平抗性频率为22.6%,未检测到敏感菌株;对腐霉利的抗性频率为20.4%,含21株低水平抗性菌株和7株中等水平抗性菌株。7种杀菌剂的室内毒力测定结果表明:咪鲜胺对白术黑斑病菌的活性最高,EC₅₀值为0.15 mg/L。供试的137株白术黑斑病菌对咪鲜胺的EC₅₀值分布在0.05~0.87 mg/L之间,平均EC₅₀值为 (0.29 ± 0.11) mg/L,可作为敏感性基线。     

DNA transposon fossils present on the conditionally dispensable chromosome controlling AF-toxin biosynthesis and pathogenicity of Alternaria alternata

The strawberry pathotype of Alternaria alternata produces host-specific AF-toxin and causes Alternaria black spot of strawberry. Previously, we isolated cosmid clones pcAFT-1 and pcAFT-2 from strain NAF8 of the strawberry pathotype that contain AF-toxin biosynthetic genes, named AFT genes. In a molecular characterization here of pcAFT-1 and pcAFT-2, 11 AFT genes and five transposon-like sequences, named TLS-S1 to TLS-S5, were detected. The nucleotide sequences of TLS-S1 and TLS-S4 share high homology, and their putative products have similarity to transposases of the hAT family transposons. Thus, TLS-S1 and TLS-S4 were renamed TLS-S1-1 and TLS-S1-2, respectively. Amino acid sequences deduced from TLS-S2, TLS-S3, and TLS-S5 have similarity to transposases of the Fot1/Pogo family transposons, but they are significantly different. All five sequences have incomplete open reading frames (ORFs) for transposases owing to deletions, termination codons, and/or frameshifts, indicating that they are inactivated elements. Analysis of genomic distribution of these sequences revealed that they are specifically distributed on a 1.05-Mb chromosome of NAF8, which has been identified as a conditionally dispensable (CD) chromosome encoding AFT genes. The presence of three, four, and three copies of TLS-S1, TLS-S2, and TLS-S3, respectively, and a single copy of TLS-S5 on the CD chromosome were estimated by DNA gel blot analysis. The remaining copy of TLS-S1 and the three copies of TLS-S2 were isolated and identified to also encode incomplete ORFs. Thus, it appears that all copies of the transposon-like sequences identified are inactivated elements (fossils) unique to the CD chromosome in the genome of the strawberry pathotype. The nucleotide sequence data reported are available in the DDBJ/EMBL/GenBank databases under the accession numbers AB236733 (TLS-S1-1), AB236734 (TLS-S1-2), AB236735 (TLS-S1-3), AB236736 (TLS-S2-1), AB236737 (TLS-S2-2), AB236738 (TLS-S3), and AB236739 (TLS-S5)     

Host specificity of a brazilian isolate of<Emphasis Type="Italic">Alternaria cassiae</Emphasis> (Cenargen CG593) under greenhouse conditions

SeveralAlternaria cassiae isolates were recovered from diseased sicklepod plants (Senna obtusifolia) in the southern regions of Brazil. A representative isolate (Cenargen CG593) was tested for its host range under greenhouse conditions. The fungus promoted symptoms in sicklepod, cassava (Manihot dulce), tomato (Lycopersicon esculentum) and eggplant (Solanum melongena) when tested at a spore concentration of 10⁶ spores ml⁻¹. When the plants were inoculated with a suspension of 10⁵ spores ml⁻¹ and held at a dew period of 12 h (cassava) or 18 h (tomato and eggplant), the plants showed symptoms of the disease, but they recovered and continued their normal vegetative growth. These results show that the fungusA. cassiae is safe to use for the control ofS. obtusifolia under Brazilian conditions, because it did not cause excessive damage in the three plants tested. http://www.phytoparasitica.org posting Jan. 14, 2007.