Mechanisms of biocontrol by Pantoea dispersa of sugar cane leaf scald disease caused by Xanthomonas albilineans

Albicidins, a family of phytotoxins and antibiotics produced by Xanthomonas albilineans, are important in sugar cane leaf scald disease development. The albicidin detoxifying bacterium Pantoea dispersa (syn. Erwinia herbicola) SB1403 provides very effective biocontrol against leaf scald disease in highly susceptible sugar cane cultivars. The P. dispersa gene (albD) for enzymatic detoxification of albicidin has recently been cloned and sequenced. To test the role of albicidin detoxification in biocontrol of leaf scald disease, albD was inactivated in P. dispersa by site-directed mutagenesis. The mutants, which were unable to detoxify albicidin, were less resistant to the toxin and less effective in biocontrol of leaf scald disease than their parent strain. This indicates that albicidin detoxification contributes to the biocontrol capacity of P. dispersa against X. albilineans. Rapid growth and ability to acidify media are other characteristics likely to contribute to the competitiveness of P. dispersa against X. albilineans at wound sites used to invade sugar cane.

Effects of Black Leaf Streak Disease and Sigatoka Disease on fruit quality and maturation process of bananas produced in the subtropical conditions of southern Brazil

Banana fruits are harvested at the green-mature stage (pre-climacteric) in order to allow sufficient time for transport and marketing. The time between the harvest and the initiation of the natural ripening process is called green life (GL), which is closely correlated to physiological age. Sigatoka Disease (SD: also called yellow Sigatoka) and Black Leaf Streak Disease (BLSD; also called black Sigatoka) are the main foliar diseases affecting banana production. The aim of this work was to investigate the influence of these diseases on banana GL and postharvest behavior in subtropical conditions (southeastern Brazil). The results showed that both diseases shortened the banana's GL when compared to control bananas of the same physiological age. Moreover, fruits from infested plots showed higher values of CO2 (+100% for SD and +300% for BLSD) and C2H4 production (+30% for SD and +60% for BLSD) at the climacteric peak. BLSD caused 40% reduction in fruit weight. Fruits from plants with a high degree of SD or BLSD undergo an altered maturation process. (C) 2011 Elsevier Ltd. All rights reserved.

A new Alternaria leaf-spot disease of cherries...

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Genetic uniformity of international isolates of Leifsonia xyli subsp xyli, causal agent of ratoon stunting disease of sugarcane

An international collection of the sugarcane ratoon stunting disease pathogen, Leifsonia xyli subsp. xyli, was analysed to assess genetic diversity. DNA fingerprinting using BOX primers was performed on 105 isolates, comprising 65 Australian isolates and an additional 40 isolates from Indonesia (n = 8), Japan (n = 1), USA (n = 3), Brazil (n = 2), Mali (n = 2), Zimbabwe (n = 13), South Africa (n = 9) and Reunion (n = 2). Sixty-two of these isolates were also screened using ERIC primers. No variation was found among any of the isolates. The intergenic spacer (IGS) region of the ribosomal RNA genes from 54 isolates was screened for sequence variation using single-stranded conformational polymorphism (SSCP), but none was observed. Direct sequencing of the IGS from a subset of nine isolates, representing all of the countries sampled in this study, confirmed the results of the SSCP analysis. Likewise, no sequence variation was found in the 16S ribosomal RNA genes of the same subset. Four Colombian isolates from sugarcane, morphologically similar to L. xyli subsp. xyli, were putatively shown to be an undescribed Agrococcus species of unknown pathogenicity. The lack of genetic variation among L. xyli subsp. xyli isolates, independent of time of sampling, cultivar of isolation, or country of origin, suggests the worldwide spread of a single pathogenic clone, and further suggests that sugarcane cultivars resistant to ratoon stunting disease in one area should retain this property in other regions.

In vivo antifungal activity of neem oil and aqueous extracts against leaf spot disease caused by Cercospora abelmoschii on okra

The cercospora leaf spot, caused by Cercospora abelmoschi Ellis and Everhart, is quite common in okra culture. Therefore, this study aimed to evaluate the efficiency of aqueous extracts of neem ( Azadirachta indica A. Juss), citronella ( Cymbopogon nardus (L.) Rendle), eucalyptus ( Eucalyptus grandis L.), ecolife®, A. indica oil and fungicide cercobin 700 PM® in control of cercospora leaf spot on okra in greenhouse. The extracts and neem oil were tested in concentration 10%, the fungicide cercobin 700PM® in dose 2.5 g.l-1, applied 10 days after pathogen inoculation by leaf spray and the citric biomass extract ecolife® in concentration 5.0 ml.l-1, applied 10 days before pathogen inoculation. All treatments, except ecolife®, were effective in controlling cercospora leaf spot and may be recommended as alternatives in agroecological systems.

Identification of common bean resistant sources to angular leaf spot disease in a brazilian germplasm collection.

The aim of this study was to identify sources of resistance in the germplasm collection providing information of potential sources of resistance to introduce in breeding programs.

Physico-chemical characterization of banana varieties resistant to black leaf streak disease for industrial purposes.

A cultura da banana tem baixa diversidade genética, tornando a espécie susceptível a doenças dizimadoras como a Sigatoka negra. No entanto, a adoção de novas variedades necessita de avaliações agronômicas e físico-químicas. Neste estudo, as variedades de banana, resistentes à Sigatoka negra, foram caracterizadas e comparadas com a variedade tradicional (Grand Naine). Cada variedade foi avaliada considerando-se critérios relevantes para a agroindústria, como pH, sólidos solúveis totais, acidez total titulável, relação SST/ATT, açúcares totais, açúcares redutores e não redutores, umidade, sólidos totais e rendimento no processamento. A variedade Thap Maeo apresentou-se como a variedade mais potencial para substituição da Gran Naine na indústria, com altos teores de sólidos solúveis totais, açúcares redutores, açúcares totais e umidade. As variedades Caipira e FHIA 2 também podem substituir a Grand Naine. Na análise de agrupamentos, verificou-se que a variedade Grand Naine esteve muito próxima das variedades do subgrupo Gros Michel (Bucaneiro, Ambroisa e Calipso) e também da variedade Caipira, apresentando no seu genoma o grupo AAA. Conclui-se que há opções de variedades resistentes para substituição da variedade tradicional, nas regiões afetadas pela Sigatoka-negra.

High affinity binding of albicidin phytotoxins by the AlbA protein from Klebsiella oxytoca

Albicidins are a family of phytotoxins and antibiotics which play an important role in the pathogenesis of sugarcane leaf scald disease. The albA gene from Klebsiella oxytoca encodes a protein which inactivates albicidin by heat-reversible binding. Albicidin ligand binding to a recombinant AlbA protein, purified by means of a glutathione S-transferase gene fusion system, is an almost instant and saturable reaction. Kinetic and stoichiometric analysis of the binding reaction indicated the presence of a single high affinity binding site with a dissociation constant of 6.4 x 10(-8) M. The AlbA-albicidin complex is stable from 4 to 40 degrees C, from ph 5 to 9 and in high salt solutions. Treatment with protein denaturants released all bound albicidin. These properties indicate that AlbA may be a useful affinity matrix for selective purification of albicidin antibiotics. AlbA does not bind to p-nitrophenyl butyrate or alpha-naphthyl butyrate, the substrates of the albicidin detoxification enzyme AlbD from Pantoea dispersa. The potential exists to pyramid genes for different mechanisms in transgenic plants to protect plastid DNA replication from inhibition by albicidins.

Factors affecting biosynthesis by Xanthomonas albilineans of albicidin antibiotics and phytotoxins

Albicidins are important factors in systemic pathogenesis by Xanthomonas albilineans, which causes the devastating leaf scald disease of sugar cane. They ale also of substantial interest as antibiotics that selectively block prokaryote DNA replication. Albicidin biosynthesis is highly sensitive to medium composition. An optimized, chemically defined medium (SMG3) yielded 30-fold more albicidin from half the accumulated biomass, relative to sucrose peptone (SP) medium. Phosphate starvation stimulated albicidin production in SMG3 and SP media. Addition of other amino acids, ammonium ions or peptones to the defined medium increased the growth rate of X albilineans XA3, but differentially inhibited albicidin biosynthesis. Knowledge of these factors indicates new approaches to understanding mechanisms of pathogenesis and resistance to sugar cane leaf scald disease, and to strain improvement for production of albicidin antibiotics.

Engineered detoxification confers resistance against a pathogenic bacterium

We generated transgenic sugarcane plants that express an albicidin detoxifying gene (albD), which was cloned from a bacterium that provides biocontrol against leaf scald disease. Plants with albicidin detoxification capacity equivalent to 1-10 ng of AlbD enzyme per mg of leaf protein did not develop chlorotic disease symptoms in inoculated leaves, whereas all untransformed control plants developed severe symptoms. Transgenic lines with high AlbD activity in young stems were also protected against systemic multiplication of the pathogen, which is the precursor to economic disease. We have shown that genetic modification to express a toxin-resistance gene can confer resistance to both disease symptoms and multiplication of a toxigenic pathogen in its host.

Analysis of the genes flanking xabB: a methyltransferase gene is involved in albicidin biosynthesis in Xanthomonas albilineans

Transposon mutagenesis and complementation studies previously identified a gene (xabB) for a large (526 kDa) polyketide-peptide synthase required for biosynthesis of albicidin antibiotics and phytotoxins in the sugarcane leaf scald pathogen Xanthomonas albilineans. A cistron immediately downstream from xabB encodes a polypeptide of 343 aa containing three conserved motifs characteristic of a family of S-adenosyl-L-methionine (SAM)-dependent O-methyltransferases. Insertional mutagenesis and complementation indicate that the product of this cistron (designated xabC) is essential for albicidin production, and that there is no other required downstream cistron. The xab promoter region is bidirectional, and insertional mutagenesis of the first open reading frame (ORF) in the divergent gene also blocks albicidin biosynthesis. This divergent ORF (designated thp) encodes a protein of 239 aa displaying high similarity to several IS21-like transposition helper proteins. The thp cistron is not located in a recognizable transposon, and is probably a remnant from a past transposition event that may have contributed to the development of the albicidin biosynthetic gene cluster. Failure of 'in trans' complementation of rhp indicates that a downstream cistron transcribed with thp is required for albicidin biosynthesis. (C) 2000 Elsevier Science B.V. All rights reserved.

Genes for albicidin biosynthesis and resistance span at least 69 kb in the genome of Xanthomonas albilineans

All Tn5 insertion mutants of Xanthomonas albilineans, the cause of leaf scald disease of sugar cane, which failed to produce albicidin antibiotics failed to cause chlorosis in inoculated sugar cane but- remained resistant to albicidin. Southern analysis revealed that mutants deficient in albicidin production carried the transposon on different chromosomal restriction fragments spanning at least: 50 kb in the X. albilineans genome, which is larger than any reported cluster of genes involved in the production of a bacterial phytotoxin. Albicidin-resistant cosmid clones from a Tox(-) Tn5 insertion mutant did not carry the transposon, and the subcloned albicidin resistance gene did not hybridize to any of the restriction fragments carrying Tn5 in the Tox(-) mutants, indicating that the albicidin biosynthesis and resistance genes are not closely linked in X. albilineans.

The gene for albicidin detoxification from Pantoea dispersa encodes an esterase and attenuates pathogenicity of Xanthomonas albilineans to sugarcane

Albicidin phytotoxins are pathogenicity factors in a devastating disease of sugarcane known as leaf scald, caused by Xanthomonas albilineans. A gene (albD) from Pantoea dispersa has been cloned and sequenced and been shown to code for a peptide of 235 amino acids that detoxifies albicidin, The gene shows no significant homology at the DNA or protein level to any known sequence, but the gene product contains a GxSxG motif that is conserved in serine hydrolases, The AlbD protein, purified to homogeneity by means of a glutathione S-transferase gene fusion system, showed strong esterase activity on p-nitrophenyl butyrate and released hydrophilic products during detoxification of albicidins. AlbD hydrolysis of p-nitrophenyl butyrate and detoxification of albicidins required no complex cofactors, Both processes were strongly inhibited by phenylmethylsulfonyl fluoride, a serine enzyme inhibitor, These data strongly suggest that AlbD is an albicidin hydrolase, The enzyme detoxifies albicidins efficiently over a pH range from 5.8 to 8.0, with a broad temperature optimum from 15 to 35 degrees C, Expression of albD in transformed X. albilineans strains abolished the capacity to release albicidin toxins and to incite disease symptoms in sugarcane, The gene is a promising candidate for transfer into sugarcane to confer a form of disease resistance.

Genes diferencialmente expressos em cana-de-açúcar inoculada com Xanthomonas albilineans, o agente causal da escaldadura da folha

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Perfil de expressão gênica de cana-de-açúcar submetida a estresse biótico com Xanthomonas albilineans

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)