Sensitive and specific detection of Clavibacter xyli subsp. xyli, causal agent of ratoon stunting disease of sugarcane, with a polymerase chain reaction-based assay

A sensitive, specific polymerase chain reaction-based assay was developed for the detection of the causal agent of ratoon stunting disease of sugarcane, Clavibacter xyli subsp. xyli. This assay uses oligonucleotide primers derived from the internal transcribed spacer region between the 16S and 23S rRNA genes of the bacterial rRNA operon. The assay is specific for C. xyli subsp. xyli and does not produce an amplification product from the template of the closely related bacterium C. xyli subsp. cynodontis, nor from other bacterial species. The assay was successfully applied to the detection of C. xyli subsp. xyli in fibrovascular fluid extracted from sugarcane and was sensitive to approximately 22 cells per PCR assay. A multiplex PCR test was also developed which identified and differentiated C. xyli subsp. xyli and C. xyli subsp. cynodontis in a single PCR assay.

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.

Bacterial dieback of pistachio in Australia

Xanthomonas bacteria have been found in association with dieback of pistachio trees in Australia. Identification of the bacterial species and/or pathovar and epidemiological studies are in progress.

Effect of a highly concentrated lipopeptide extract of Bacillus subtilis on fungal and bacterial cells

Lipopeptides produced by Bacillus subtilis are known for their high antifungal activity. The aim of this paper is to show that at high concentration they can damage the surface ultra-structure of bacterial cells. A lipopeptide extract containing iturin and surfactin (5 mg mL-1) was prepared after isolation from B. subtilis (strain OG) by solid phase extraction. Analysis by atomic force microscope (AFM) showed that upon evaporation, lipopeptides form large aggregates (0.1-0.2 mu m2) on the substrates silicon and mica. When the same solution is incubated with fungi and bacteria and the system is allowed to evaporate, dramatic changes are observed on the cells. AFM micrographs show disintegration of the hyphae of Phomopsis phaseoli and the cell walls of Xanthomonas campestris and X. axonopodis. Collapses to fungal and bacterial cells may be a result of formation of pores triggered by micelles and lamellar structures, which are formed above the critical micelar concentration of lipopeptides. As observed for P. phaseoli, the process involves binding, solubilization, and formation of novel structures in which cell wall components are solubilized within lipopeptide vesicles. This is the first report presenting evidences that vesicles of uncharged and negatively charged lipopeptides can alter the morphology of gram-negative bacteria.

Factors associated with mortality in patients with bloodstream infection and pneumonia due to Stenotrophomonas maltophilia

Severe infections caused by Stenotrophomonas maltophilia are associated with high mortality, and strategies to improve the clinical outcome for infected patients are needed. A retrospective cohort study of patients with bloodstream infection (BSIs) and pneumonia caused by S. maltophilia was conducted. Multivariate analysis was performed to access factors associated with 14-day mortality. A total of 60 infections were identified. Among these, eight (13%) were pneumonias and 52 were BSIs; 33.3% were primary, 13% were central venous catheter (CVC)-related and 40% were secondary BSIs. Fifty-seven (85%) patients had received previous antimicrobial therapy; 88% had CVC, 57% mechanical ventilation and 75% were in the intensive care unit at the onset of infection. Malignancy (45%) was the most frequent underlying disease. The mean of the Acute Physiology and Chronic Health Evaluation II (APACHE II) scores was 17 and for the Sepsis-related Organ Failure Assessment (SOFA) score, it was 7 points. The overall and 14-day mortality were, respectively, 75% and 48%. Forty-seven (78%) patients were treated and, of these, 74% received trimethoprim-sulfamethoxazole. Independent risk factors associated with mortality were SOFA index > 6 points (0.005) and septic shock (0.03). The Kaplan-Meier estimations curves showed that patients with APACHE II score > 20 and SOFA score > 10 had a survival chance of, respectively, less than 8% and less than 10% (P <= 0.001) at 21 days after the first positive S. maltophilia culture. Our results suggest that the independent factors associated with outcome in patients with infection caused by S. maltophilia are septic shock and higher SOFA index.

Producción de factores de virulencia por fitopatógenos bacterianos de soja. Utilización de productos naturales como biocontrol. Production of virulence factors by soybean bacterial phytopathogens. The use of natural products as biocontrolers.

En Argentina el cultivo de soja ocupa el primer lugar en superficie sembrada. El 90% de la producción se obtiene en la zona central del pais. La siembra directa favorece la multiplicación y supervivencia de fitopatógenos causantes de tizón y pústula bacterianos. El tizón es producido por Pseudomonas syringae pv. glycinea observándose manchas marrones en las hojas. Produce gran variedad de toxinas: coronatina, faseolotoxina, siringomicina, tabtoxina, proteínas “nucleation ice”, entre otras, las cuales contribuyen a la clorosis y necrosis. En la infección, además, están involucrados exopolisacáridos (levano y alginato). La celulosa ha sido relacionada en la adhesión bacteriana y en la formación de biofilm. La pústula es causada por Xanthomonas axonopodis pv. glycines. Produce manchas pequeñas con una pequeña pústula de color claro. Libera enzimas como α-amilasa, proteasa, endo β-mannanasa, actividad peptolítica, que degradan componentes vegetales. Xantan, producido por X. axonopodis es uno de los componentes necesarios para la formación de biofilm. Este último es considerado un importante factor de virulencia porque proporciona una estrategia de colonización que otorga mayor resistencia a ambientes desfavorables, tolerancia a antimicrobianos, producción de metabolitos y exoenzimas, etc. Actualmente el control de bacterias fitopatógenas se realiza mediante pesticidas con alta toxicidad para los consumidores y el ambiente. Para evitar las bacteriosis en la práctica se sugiere la rotación de cultivos y utilizar semillas certificadas. Se están probando compuestos naturales derivados de plantas medicinales como pesticidas; estos se pueden dividir en varias categorías fitoquímicas. Varios estudios confirman la actividad antibacteriana, antifúngica y antiviral de estos productos. Extractos vegetales con alto contenido de flavonoides y aceite esenciales poseen una importante actividad antibacteriana. Además, algunos aceites esenciales podrían estar incidiendo en la liberación y/o producción de biofilm, exopolisacáridos y exoproteínas. La gran incidencia de las infecciones por fitopatógenos y las pérdidas económicas que estas acarrean hacen que su control presente grandes dificultades para la agricultura sustentable en soja de nuestro país. En este trabajo se propone estudiar los diferentes factores de virulencia de cepas bacterianas fitopatógenas y evaluar el rol que cumplen en el proceso de la enfermedad en cultivos de soja y desarrollar estrategias para el control de bacteriosis vegetales aplicando productos naturales aislados de plantas aromáticas. La correcta utilización de productos antimicrobianos de origen natural aplicados sobre el cultivo y/o sobre las semillas evitaría la dispersión de la enfermedad y la eliminación al medio ambiente de productos contaminantes no deseados. In Argentina, soybean cultivation occupies the first place; 90% of this cereal is produced in the central region of the country. Intensive tillage practices favour multiplication and survival of bacterial phytopathogens causing blight and pustule diseases. Pseudomonas syringae pv. glycinea produce several toxins like coronatine, faseolotoxine, siringomicine, tabtoxine and proteins of nucleation ice that contribute to the develop of chlorosis and necrosis, characteristic of bacterial blight. It also produces levan and alginate, cellulose and biofilm. Pustule disease is caused by Xanthomonas axonopodis pv glycines, which produce enzymes like α-amilase, protease, endo β-mannanase, peptolitic activity, xanthan and biofilm. Nowadays the control of phytopathogenic bacteria consists in the application of pesticides that are toxic for the environment and man. Natural products from medicinal plants are a new alternative for the treatment of phytopathogens. Researches made with phytochemical compounds (flavonoids, phenols, quinones, cummarines, essential oils, terpenes) support the antimicrobial activity of these natural products. What is more, these substances could suppress the biofilm, exoproteins and exopolisaccharides formation and release of them. The infections caused by phytopathogens provoke economical loses and its control presents big difficulties in our country. The proposal of this work is the characterization of phytopatoghenic strains, its virulence factors and the role they play in the disease process. The development of a new alternative for the control of vegetable bacteriosis using natural products obtained from aromatic plants and the correct application of them on sown fields or on seeds is also an objective in this work.

Nematódeos e bactérias em fôlhas de alcachôfra

Diseased plants of artichoke (Cynara scolimus L.) from the Instituto Agronômico, Campinas, Brazil, harboured nematodes in the leaves. The nematodes proved to belong to the species Protorhabditis oxyuris (Claus, 1862) Dougherty, 1955. As it could not be charged as the primary agent of the disease, a study of the bacteria found in the material was made. This disclosed that 4 different bacterial forms were present, namely Bacillus subtilis, Bacterium udum, Xanthomonas beticola var. cynarae n. var., and Cory neb acterium fascians var. cynarae n. var. The typical forms of X. beticola and of C. fascians are known as pathogenic to a number of plants: but, in the present case, unfortunately no inoculation trial could be made as an attempt to clear up the possible pathogenicity of any of them also to Cynara scolimus. The nematode P. oxyuris is the secondary agent, which however possibly play an important part in the distribution of the bacteria, as it has been repeatedly pointed out by previous writers with reference to closely related species. The symptoms of the disease are described. The main injuries found on leaves were necrotic areas progressing from the distal part of the limb, resulting in a great reduction of the leaf superficies.

Comportamento ''in vitro" de uma bactéria fitopatogênica frente a diversos antibióticos

Foi estudada a resistência da bactéria fitopatogênica Xanthomonas campestris com relação a cinco antibióticos. Quando os antibióticos foram adicionados ao meio sólido no qual a bactéria iria se desenvolver, colônias resistentes à estreptomicina e éritromicina, apareceram até nas mais altas concentrações usadas. No caso da penicilina, aureomicina e cloranfenicol, em concentrações superiores a 200 mcg/ml, 0,3 mcg/ml e 128 mcg/ml respectivamente, não apareceram colônias resistentes. Os resultados mostraram que a penicilina foi o antibiótico menos efetivo e a aureomicina a mais efetiva em inibir o crescimento de X. campestris. Os resultados sugerem também que o modêlo de resistência à estreptomicina e éritromicina é o de um só passo enquanto que o modêlo de resistência aos três outros antibióticos ensaiados, é o de múltiplos passos.

Trichurid nematodes in ring-necked pheasants from backyard flocks of the State of Rio de Janeiro, Brazil: frequency and pathology

The present investigation is related to the frequency of infection and to the gross and microscopic lesions associated to the presence of trichurid worms in 50 ring-necked pheasants (Phasianus colchicus) from backyard flocks in the state of Rio de Janeiro, Brazil. In the investigated birds, the overall infection rate was of 74%, with the presence of Eucoleus perforans with 72% of prevalence and 21.2 of mean intensity, in the esophageal and crop mucosa and rarely in the junction of the proventriculus and esophagus, E. annulatus with 2% and 3 in the crop mucosa, Capillaria phasianina, with 12% and 4.3 in the cecum and small intestine and Baruscapillaria obsignata, for the first time referred in this host, with 2% and 1 in the small intestine. Clinical signs were absent. The gross lesions observed in the crop and esophagus of 14 (38.9%) pheasants parasitized with E. perforans were thickening, small nodules, congestion, and petechial haemorrhages in the mucosa. These birds presented a mean infection of 37.5 and a range of infection of 10-82. The microscopic lesions revealed chronic esophagitis with diffuse inflammatory process in the lamina propria characterized mostly by a mononuclear cell infiltrate and also with the presence of granulocytes. In the case of the parasitism of pheasants with C. phasianina, the gross lesions were absent; microscopic lesions were characterized by chronic typhlitis with mononuclear infiltrate. Gross and microscopic lesions were absent in the pheasants parasitized with E. annulatus and B. obsignata.

Simptomatologia i microorganismes associats a la necrosi bruna apical de la noguera (Juglans regia)

La Necrosi Apical Bruna (BAN, brown apical necrosis, segons les sigles en anglès)Es va detectar per primer cop l’any 1997 a Extremadura degut a la severa caiguda defruits. Avui dia la malaltia és present a quasi totes les zones productores de lamediterrània. Els símptomes difereixen dels provocats per Xanthomonas arboricola pv.juglandis i Gnomonia leptsostyla.. S’observa que els fruits afectats presenten una tacabruna a la zona apical i necrosi dels teixits interiors. El grup de Patologia Vegetal de laUniversitat de Girona participa i dirigeix la tasca sobre l’etiologia de la BAN dins laxarxa europea d’investigació en bacteris patògens de fruiters ,COST873. Hi ha una certacontrovèrsia en la definició dels símptomes i agents causals. Tots els grups coincideixenen afirmar que es tracta d’una malaltia complexa amb diferents organismes implicats

Lactic acid bacteria from fresh fruit and vegetables as biocontrol agents of phytopathogenic bacteria and fungi

This study evaluated the efficacy of lactic acid bacteria (LAB) isolated from fresh fruits and vegetables as biocontrol agents against the phytopathogenic and spoilage bacteria and fungi, Xanthomonas campestris, Erwinia carotovora, Penicillium expansum, Monilinia laxa, and Botrytis cinerea. The antagonistic activity of 496 LAB strains was tested in vitro and all tested microorganisms except P. expansum were inhibited by at least one isolate. The 496 isolates were also analyzed for the inhibition of P. expansum infection in wounds of Golden Delicious apples. Four strains (TC97, AC318, TM319, and FF441) reduced the fungal rot diameter of the apples by 20%; only Weissella cibaria strain TM128 decreased infection levels by 50%. Cell-free supernatants of selected antagonistic bacteria were studied to determine the nature of the antimicrobial compounds produced. Organic acids were the preferred mediators of inhibition but hydrogen peroxide was also detected when strains BC48, TM128, PM141 and FF441 were tested against E. carotovora. While previous reports of antifungal activity by LAB are scarce, our results support the potential of LAB as biocontrol agents against postharvest rot. [Int Microbiol 2008; 11(4):231-236]

Método generalizado de análise de dialelos desbalanceados

Apresenta-se um procedimento generalizado de estimação das capacidades geral e específica de combinação em cruzamentos dialélicos, com número desigual de repetições para tratamentos (genitores e combinações híbridas F1), avaliados em um delineamento com restrições na casualização. Neste caso, as médias ajustadas estimadas são interdependentes e heterocedásticas, devendo-se, para estimar os parâmetros desejados, utilizar o modelo linear generalizado de GaussMarkov. O objetivo deste trabalho foi a dedução teórica do método e sua aplicação a um exemplo prático. Foram analisados os dados obtidos de um dialelo completo, sem os recíprocos, envolvendo cinco genitores: CB 511687-1, CB 733753, Diamante Negro, Rosinha G2 e Compuesto Chimaltenango 2. Os genitores CB 733753, CB 511687-1 e Diamante Negro contribuem geneticamente para a resistência, enquanto Rosinha G2 e Compuesto Chimaltenango 2 contribuem para a suscetibilidade do feijoeiro ao crestamento-bacteriano comum. Na maioria dos cruzamentos analisados constatou-se a dominância parcial da suscetibilidade do feijoeiro a Xanthomonas axonopodis pv. phaseoli.

BRS 137: cultivar de soja para cultivo no Sul do Brasil

A liberação de cultivares de elevado potencial produtivo e com resistência a doenças é fundamental para continuar agregando rendimento na produção brasileira de soja. A cultivar BRS 137 é resultado do programa de melhoramento de soja desenvolvido pela Embrapa. A cultivar apresentou rendimento médio de grãos 2% superior ao da cultivar IAS 5, em 24 ambientes, no Rio Grande do Sul. É resistente à pústula-bacteriana (Xanthomonas axonopodis pv. glycines), ao cancro-da-haste (Diaporthe phaseolorum f. sp. meridionalis), à podridão-parda-da-haste (Phialophora gregata), à mancha-olho-de-rã (Cercospora sojina) e ao oídio (Microsphaera diffusa). É indicada para cultivo no Rio Grande do Sul, em semeaduras realizadas a partir de meados de outubro até fim de novembro, com população máxima de 300.000 plantas/ha.

Cultivar de soja BRS 205

A BRS 205, cultivar de soja de ciclo semiprecoce, obtida do cruzamento [BR-16(2) x Ocepar 8] x Tracy-M, é indicada para o Rio Grande do Sul, em semeaduras de novembro. Possui crescimento determinado, plantas de flor branca, pubescência marrom, porte baixo e grãos de hilo preto. Tem resistência ao cancro-da-haste (Diaporthe phaseolorum f. sp. meridionalis), à podridão-parda-da-haste (Phialophora gregata), à mancha-olho-de-rã (Cercospora sojina) e à pústula bacteriana (Xanthomonas axonopodis pv. glycines). É moderadamente resistente ao oídio (Microsphaera diffusa). O rendimento médio de grãos foi 6% e 11% superior ao das cultivares RS 7-Jacuí e BR-16, respectivamente.

Identificação e controle com antibióticos de bactérias endofíticas contaminantes em explantes de batata micropropagados

Este trabalho teve por objetivos isolar, caracterizar e identificar bactérias endofíticas contaminantes encontradas em tecidos de batata durante a micropropagação e selecionar antibióticos para o controle in vitro desses microrganismos por meio da determinação da concentração bactericida mínima inibitória. Brotações de batata apresentando contaminação bacteriana durante a etapa de multiplicação in vitro, foram superficialmente esterilizadas e os internódios transferidos para placas de Petri com ágar nutriente, onde permaneceram incubadas a 28°C por até cinco dias. Após purificação, as bactérias foram caracterizadas e identificadas por testes taxonômicos. Um total de oito estirpes bacterianas foram isoladas e identificadas como pertencentes às famílias Acetobacteriaceae (1) e Enterobacteriaceae (2) e aos gêneros Corynebacterium (3), Pseudomonas (1) e Xanthomonas (1). Os melhores resultados para a inibição do crescimento bacteriano foram obtidos com os antibióticos ampicilina, cloranfenicol, estreptomicina e tetraciclina em concentrações que variaram de 32 a 256 mg L-1.