Elineide B. Silveira

Fungos endofíticos em Annona spp.: isolamento, caracterização enzimática e promoção do crescimento em mudas de pinha (Annona squamosa L.)

Endophytic isolates of fungi were obtained from leaves, stems and roots of 110 sweetsop and 90 soursop plants from Pernambuco. Twenty-nine isolates were analyzed for production of extracellular enzymes by qualitative assay in Petri dishes containing specific solid media, and for the capacity to promote growth of sweetsop seedlings. These isolates were identified as Acremonium (10.34%), Aspergillus (3.45%), Chaetomium (3.45%), Colletotrichum (10.34%), Cylindrocladium (13.8%), Fusarium (31.03%), Glomerella (3.45%), Nigrospora (6.9%), Penicillium (6.9%) and Phomopsis (10.34%). Nineteen isolates showed lypolytic activity while five showed proteolytic activity; cellulolytic and amylolytic activity were not detected. Eleven isolates of the genera Acremonium (GFR6 and GRR1), Colletotrichum (GFR4 and PFR4), Phomopsis (PFR3 and GCR4), Cylindrocladium (GRR4), Chaetomium (GRR7) and Fusarium (GRR5, PRR1 and PRR6) efficiently improved plant growth. Increase in shoot dry matter of sweetsop seedlings ranged from 23.2 to 32.7%; there was no increase in root dry matter. It is worthy of note that 20 isolates caused significant (P = 0.05) reduction in root dry matter of sweetsop seedlings. In apparently healthy tissues of sweetsop and soursop plants, some fungi promote shoot growth or reduce root growth, while others have no effect on growth of sweetsop seedlings.

Management of melon bacterial blotch by plant beneficial bacteria

Plant beneficial bacteria (PBB) have shown potential for disease control and are particularly important in the management of bacterial diseases, which are poorly controlled by conventional methods. In melon, bacterial fruit blotch caused by Acidovorax citrulli is a seedborne disease that is particularly destructive under certain conditions. PBB strains were screened for their ability to protect seeds and leaves from bacterial fruit blotch, and their antibiosis activity and plant colonization were studied. When Bacillus sp. RAB9 was applied to infected seeds, it reduced the area under the disease progress curve (AUDPC) by 47% and increased the incubation period (the time between inoculation and the first visible symptoms) by 35%. Three of the selected strains (JM339, MEN2 and PEP91) displayed antibiosis against A. citrulli. The RAB9Rif-Nal mutant colonized seeds epiphytically and roots and stems endophytically. Paenibacillus lentimorbus MEN2 sprayed on melon seedlings protected leaves, and when challenged with A. citrulli, it reduced the AUDPC (by 88%), disease index (by 81%) and incidence (by 77%). Given that the production of both melon seedlings and commercially grown greenhouse melons is increasing, biocontrol strategies may well be integrated into bacterial blotch management programs.

Bacterização de sementes e desenvolvimento de mudas de pepino

Epiphytic and endophytic bacteria were isolated from healthy cucumber plants, collected in several counties of Pernambuco State, Brazil, and evaluated for seedling growth promotion under greenhouse conditions. Three bioassays were performed. In the first, 93 strains were tested; in the second, 32, and, in the third, eight from cucumber plus 10 bacterial epiphytic strains from other crops were used. The cucumber seeds were bacterized by immersion in the bacterial suspension adjusted to A580 = 0.7, sown in polystyrene trays filled with organic substrate and analyzed ten days after sowing in relation to shoot (MSPA), root (MSR), and total (MST) dry matter. In the third bioassay the epiphytes PEP52, PEP8, PEP82, PEP91, and C22 were selected because they significantly (P=0.05) elevated MSR and MST in relation to the control reaching higher values than 70 and 40%, respectively. After compatibility tests the five isolates applied separately and in mixtures efficiently increased the index of MSPA, MSR, and MST in cucumber seedlings, without differing among them. It is worth noticing that PEP81 (Bacillus amyloliquefaciens) and PEP91 (Enterobacter cloacae) increased 55.5 and 34.5% (MSPA), 42.9 and 37.2% (MSR), and 41.6 and 34.0% (MST), respectively. The production of indolacetic acid, hydrogen cyanide, phosphate solubilization and changes in foliar levels of N, P, K, Ca, and Mg were evaluated as putative mechanisms of action for theses isolates, but they showed negative results. Seed bacterization with B. amyloliquefaciens PEP81 and E. cloacae PEP91 could improve quality of cucumber seedlings.

Isolamento, seleção de bactérias e efeito de Bacillus spp. na produção de mudas orgânicas de alface

ABSTRACT Isolation, selection of bacteria, and effect of Bacillus spp. inthe production of organic lettuce seedlings Epiphytic and endophytic bacterial strains isolated from healthylettuce plants were evaluated for growth promotion of seedlings andplants respectively under greenhouse and field conditions of organicproduction of lettuce, in Brazil. The cultivar Veronica was utilized inthe greenhouse experiments and cvs. Veronica and Verdinha wereevaluated in the field. The strains were applied by simultaneousbacterization of seed and substrate. In the field the most efficient strainsC25 ( Bacillus thuringiensis subvar. kenyae ) and C116 ( Bacilluspumilus ) were utilized separated and in mixture after the compatibilityassay. In greenhouse root fresh weight (MFR), shoot fresh weight(MFPA) and total fresh weight (MFT) were evaluated 21 days afterbacterization. In field the total fresh weight of commercial plants wasdetermined 21 and 28 days after transplant, respectively for cvs.Verdinha and Veronica. The mechanisms of BPCP studied wereproduction of indolacetic acid, cyanidric acid, phosphate solubilizationand alterations of N, P, K, Ca and Mg foliar levels. In the greenhouse,seedlings treated with C116 showed significant increase in relation tocontrols for MFR, MFPA and MFT as well as those treated with C25for MFR and MFT. In the field cvs. Verdinha and Veronica treatedwith C25, C116 or mixture did not significantly differ from control.None of the analyzed mechanisms were positive but strain C25significantly increased the level of foliar N.

Severidade da mancha-aquosa em meloeiro sob diferentes condições de molhamento foliar e concentração de inóculo de Acidovorax avenae subsp. citrulli

The severity of melon (Cucumis melo) fruit blotch was evaluated at different intervals of the leaf wetness period (0, 6, 12, 24 and 48 h) from the time the leaf wetness period began (0, 6, 12, 24 and 48 h after inoculation), and at different inoculum concentrations of Acidovorax avenae subsp. citrulli (3.4 x 101 to 3.4 x 107 CFU.ml-1). Three pathogen strains and the yellow hybrids AF-646 and AF-682 were utilized. Leaves of 20-day old plants were atomized with bacterial suspension and placed in the greenhouse for determining incubation period, disease progress rate, disease index and area under the disease progress curve. The regression equations for the analyzed variables were better adjusted by the quadratic or logarithmic patterns. The incubation period ranged from 1.3 to 2.7 days and was greater in plants without leaf wetness. The disease index and area under the disease progress curve increased as the leaf wetness period elevated. Even in the absence of leaf wetness, disease symptoms were observed rating respectively 43.4% and 8.9 for disease index and area under the disease progress curve. The beginning of the leaf wetness period at 48 h significantly elevated (P<0,05) the incubation period and disease progress rate in relation to the other periods. The disease progress rate, disease index and area under the disease progress curve increased as the inoculum concentration increased reaching maximum values of 4.4 infection units/day, 74% and 19, respectively at 3.4 x 107 CFU ml-1.

Uso de antibióticos e leveduras para controle da podridão-mole em couve-chinesa

The soft rot caused by Pectobacterium carotovorum subsp. carotovorum (Pcc) may occur in Chinese cabbage (Brassica pekinensis) plantations presenting till 67% of incidence. In this research we evaluated the Pcc sensibility to bactericides in vitro, the effect of Mycoshield® at 3.0 and 1.5 g L-1, and yeasts at 108 cel/mL-1 to control the disease in greenhouse and field. Plants were sprayed with Mycoshield® (oxitetracycline 20%) and yeasts (Rh1 and Rh2 (Rhodotorula spp.) and Sc1 (Saccharomyces cerevisae)) seven days after transplant and inoculated with the isolate Pcc120 seven days and 12 h after treatment, respectively. In all experiments disease epidemiological components were evaluated. In vitro 40 Pcc isolates were resistant to copper sulfate and sensitive to oxitetracycline, streptomycin, oxitetracycline + streptomycin and oxitetracycline + copper sulfate all at 0.2 g L-1. Six Pcc isolates were more inhibited by Mycoshield® than by Agri-Micina® (oxitetracycline 1.5% + streptomycin 15%) both at 3.0 g L-1, but there was no inhibition by Kasumin® (Kasugamicin 2%) (2.0 mL L-1). In greenhouse Mycoshield® at 3.0 g L-1 reduced disease severity and disease index up to 47.4 and 19.0%. The yeast Sc1 reduced disease severity and area under the disease progress curve (AUDPC) up to 27.6 and 39.3% respectively, while Rh1 reduced AUDPC up to 33.5%. In field Mycoshield® reduced the disease index (14.4%) severity (15.5%) and AUDPC (28.9%), while Rh1 reduced disease index by 8.8% and Sc1 reduced the AUDPC by 15.7%. In conclusion Mycoshield® and yeasts showed low efficiency for controlling soft-rot of Chinese cabbage in field.

Penetração e Colonização de Acidovorax avenae subsp. citrulli em Folhas, Frutos e Sementes de Melão Amarelo

Leaves, fruits and seeds of melon (Cucumis melo) type Yellow were inoculated with Acidovorax avenae subsp. citrulli (Aac1) in order to study the penetration and colonization of this pathogen using scanning electron microscopy. Leaves sampled at 24, 48, 72, 96 and 120 h after inoculation showed clusters of bacterial cells interconnected by fibrils, and very rarely isolated. Bacterial cells were located mainly on the cuticular flange of the abaxial epidermis, on the base of tector trichomes, near the diacytic stomata, and inside the stomat pores. Twenty-four hours after inoculation bacteria were observed mainly on the surface of 37 day-old fruits concentrated around stomata and lenticels; after 48 h bacteria were detected in the fruit pulp as well. In the other samples (72 and 96 h) few or no bacteria were found on fruit surfaces. Fruits 51 days old showed expressive colonization of the surface, stomata and lenticels until 72 h after inoculation. The pathogen was only found in the pulp of the 96-hour samples. These results suggest that A. avenae subsp. citrulli penetrated the fruits through the stomata and lenticels. In seeds, the bacteria colonized the internal and external teguments, embryo and endosperm.

Sobrevivência de Acidovorax avenae subsp. citrulli em meloeiro

The ability of Acidovorax avenae subsp. citrulli to survive epiphytically and endophytically on leaves, roots and rhizosphere of melon plants was determined by using a mutant resistant to rifampicin (Aac1Rif). Leaves of 18-day-old melon plants grown in greenhouse and field were sprayed with mutant suspensions at concentrations of 3.4 x 102, 3.4 x 103 and 3.4 x 104 cfu.ml-1. To determine survival on roots and rhizosphere, seeds of Yellow melon were sown in soil infested with suspensions of Aac1Rif at 3.4 x 105, 3.4 x 106 and 3.4 x 107 cfu.ml-1. At 6-day intervals samples of leaves, roots and rhizosphere soil were collected and processed for isolation on NYDA medium amended with rifampicin. Bacterial populations were determined as cfu.g-1 of sample and the data were transformed to log10 for regression analysis. On melon leaves, in greenhouse and field Aac1Rif survived epiphytically for 54 days. These epiphytic bacterial populations increased initially and decreased after a certain period of time. Both final populations were similar under the two conditions and ranged from 103 to 104 cfu.g-1 of leaf, without correlation with the inoculum concentration used. In greenhouse, bacterial populations on the roots and rhizosphere decreased with time, and 60 days after soil infestation they ranged from 102 to 103 cfu.g-1 of roots and 101 cfu.g-1 of soil. Aac1Rif was not detected as an endophyte in leaves or roots of melon plants.

Tratamento pós-colheita com cálcio e microrganismos para controle da podridão-mole em tomate

The effect of three calcium sources [CaCl2, Ca (CO3)2 and Ca (SO4)2] at 1; 2; 4; 6 and 8% (w/v) were evaluated on control of soft rot caused by Pectobacterium carotovorum subsp. carotovorum (Pcc) in tomato fruits. The calcium sources were applied using vacuum infiltration. Fruits were inoculated with pathogen 24 h later by deposition of 10 ml of a bacterial suspension containing 108 CFU/ml, on artificial wounds. Fruits were incubated in a moist chamber at 92±4% relative humidity during 48 h and evaluated by measuring the lesion diameter in two opposite sides. The disease severity reduction (DSR) was calculated in relation to the control with calcium application. The best calcium source and concentration was CaCl2 8% which reduced the disease severity by 69.5%. The antagonists Rhodotorula sp. (LD-19) and a fluorescent Pseudomonas sp. (P-2) were tested separately and along with CaCl2 8%. The antagonist was applied by pouring 10 ml of suspensions (DO580=0.7) on the wounds 24 h after calcium treatment and 2 h before Pcc inoculation. The difference was significant among treatments and, the combination CaCl2 8% + LD19 showed 90% DSR. Isolates P-2 and LD-19 applied separated showed lower percentage of DSR.

Identificação de progênies de tomateiro resistentes à murcha-bacteriana

Six hundred and sixty F6 plants obtained from the crossing of cultivars CL5915-93 (moderately resistant) and IPA-6 (susceptible) were screened for resistance to bacterial wilt (Ralstonia solanacearum) under field conditions (28 ± 4oC and RH 70 ± 5,5%) in March 1996 at UFRPE. Twenty-day-old seedlings were inoculated with biovar III of the pathogen by pouring 5 ml of a suspension (5x108 UFC/ml) at the base of each seedling, two hours prior to transplanting. Evaluations were performed at weekly intervals until 70 days after inoculation. At the end of the crop cycle, 151 symptomless plants were selected and from this material one progeny was backcrossed with the cultivar IPA-6. 40 progenies (F2) from this backcross were screened for resistance under greenhouse conditions (35 ± 5,5oC and RH 77 ± 2,5%) in September 1997, using the inoculation method previously described. Evaluations of disease incidence and severity were made four, seven, ten, thirteen and 16 days after inoculation. The resistance reaction was also evaluated through incubation and latency periods. Twenty-eight days after inoculation the existence of latent infection in the resistant progenies was evaluated through vascular discoloration and pathogen isolation from processing material. Disease incidence was observed in 95% of the progenies. Eight progenies were ranked as moderately resistant and eight as resistant. The average incubation and latency periods were short (4.5 at 4.8 days) for control susceptible progenies and long (11.6 at 12.9 days) for resistant progenies. Latent infection was detected in 45% of resistant progenies.