Selection of antagonists of postharvest apple parasites: Penicillium expansum and Botrytis cinerea

The objectives of this study were to constitute a collection of pathogenic agents of economic importance which cause losses of apple fruits after harvest namely Botrytis cinerea and Penicillium expansum and to select in vivo efficient antagonistic strains able to protect fruits against both pathogens at 5 degrees C (P. expansum) and 25 degrees C (P. expansum & B. cinerea). Twenty strains of P. expansum and ten strains of B. cinerea have been isolated from infected apple fruits. Potential antagonistic micro-organisms (thirty three isolates) belonging to yeast, bacteria and fungi have been isolated from apple surface. Six of them (strains Ach1-1, Ach2-1, Ach2-2 belonging to Aureobasidium pullulans (De Bary) Arnaud, and strains 1112-3, 1113-10 and 1113-5 belonging to Aureobasidium pullulans (de Bary) Am. v. pullulans) showed a high level of protection (more than 80%) at 25 degrees C. once inoculated with P. expansum or B. cinerea for 5 days. The highest level of protection against P. expansum (96%) was observed with the application of Ach 2-1. Six days after inoculation of B. cinerea, strains Ach 2-2 and Ach 2-1 insured 100% and 96% of protection, respectively. At lower temperature (5 degrees C), first symptoms of P. expansum appeared 13 days after its inoculation. Percentages of protection observed after apple treatment with one of the six antagonistic strains were ranged from 78% to 94% 20 days after P. expansum inoculation. Strains labelled Ach showed a protective level higher than 90% against this pathogen, followed by strain 1113-10 (90%), strain 1113-5 (89%) and strain 1112-3 (82%). At 26 days post-inoculation, levels of protection decreased but remained higher than 60% (more than 80% with strain Ach2-2 and strain 1113-5, 75% with strain Ach2-1 and 1113-10, 72% with ach1-1, 61% for the other strains). Strain Ach2-2 and 1113-10 were retained as the best antagonists for the subsequent studies.     

Bacillus isolates from the spermosphere of peas and dwarf French beans with antifungal activity against Botrytis cinerea and Pythium species

A range of isolation procedures including washing, sonication and incubation in nutrient broth were used separately and in combination to obtain potential bacterial antagonists to Botrytis cinerea and Pythium mamillatum from the testae and cotyledons of peas and dwarf French beans. Heat treatment was also used to bias this selection towards spore-forming bacteria. Ninety-two bacterial isolates were obtained, 72 of which were provisionally characterized as species of Bacillus . Four of these Bacillus isolates (B3, C1, D4 and J7) displayed distinct antagonism in vitro against Botrytis cinerea and P. mamillatum when screened using dual culture analysis. Further characterization of these antagonists using API 50CHB biochemical profiling identified isolate D4 as Bacillus polymyxa and isolates B3, C1 and J7 as strains of B. subtilis . In vitro screening techniques, using cell-free and heat-killed extracts of liquid cultures against Botrytis cinerea , demonstrated the production of antifungal compounds by these four Bacillus antagonists. With each isolate the antifungal activity was found not to be either exclusively spore-bound nor released entirely into the medium but present in both fractions. The antifungal compounds produced by these isolates were shown to be heat-stable. Their identification, production and release require further study for exploitation as biocontrol systems.     

Evaluation of the effects of chemical versus biological control on Botrytis cinerea agent of gray mould disease of strawberry

This study investigates on effects of four fungicide and six isolate from Trichoderma and Gliocladium on Botrytis cinerea agent grey mold of strawberry under library and greenhouse condition. The effect of four fungicides i.e. benomyl, dichlofluanid, captan and triadimenol on B. cinerea was studied in the laboratory condition by method mixed poison to culture medium. It was shown that the fungicide including benomyl, triadimenol, dichlofluanid and captan were able to inhibit mycelial growth of B. cinerea on PDA plate with EC50 of 0.16, 1.42, 3.40 and 7.73 ppm respectively. These fungicides delayed myceliogenic germination of sclerotia at 1000 ppm, while exhibiting no fungicidal effect. Moreover, the antagonistic effects of six fungi including Trichoderma koningii (T21), T. viride (T4), T. harzionum (T5), T. viride (T2), G. virens (G2), G. virens (G8) on B. cinerea were assessed. This assessment was done under library condition and its results as follows: The antagonistic mechanism occurred through branching at the end of B. cinerea hyphae, hyphal contact, coiling, vacuolization and lyses. Volatile metabolites of T. koningii (T21) and non-volatile metabolites of G. virens (G2 and G8) and T. koningii (T21) caused maximum inhibition of the fungal growth. Trichoderma spp and G. virens were able to colonize and sporulate on sclerotia and caused their lysis within 7-21 days. In greenhouse, a completely randomized design with 11 treatments (4 chemical and 6 biological and one untreated control) each replicated five times were used for the comparison. Greenhouse studies revealed that application of fungicides i.e. captan, dichlofluanid, triadimenol and benomyl reduces disease severity by 42, 45, 48 and 52% respectively. The fungal antagonists reduce the grey mold disease severity between 5-42%. All treatments caused a decline in post harvest disease, as the most effective treatment of chemical control was benomyl with 68.33% and for the biological treatment this was T. koningii (T21) with 56%.     

Fungal ABC transporters and microbial interactions in natural environments

In natural environments, microorganisms are exposed to a wide variety of antibiotic compounds produced by competing organisms. Target organisms have evolved various mechanisms of natural resistance to these metabolites. In this study, the role of ATP-binding cassette (ABC) transporters in interactions between the plant-pathogenic fungus Botrytis cinerea and antibiotic-producing Pseudomonas bacteria was investigated in detail. We discovered that 2,4-diacetylphloroglucinol, phenazine-1-carboxylic acid and phenazine-1-carboxamide (PCN), broad-spectrum antibiotics produced by Pseudomonas spp., induced expression of several ABC transporter genes in B. cinerea. Phenazines strongly induced expression of BcatrB, and deltaBcatrB mutants were significantly more sensitive to these antibiotics than their parental strain. Treatment of B. cinerea germlings with PCN strongly affected the accumulation of [14C]fludioxonil, a phenylpyrrole fungicide known to be transported by BcatrB, indicating that phenazines also are transported by BcatrB. Pseudomonas strains producing phenazines displayed a stronger antagonistic activity in vitro toward ABcatrB mutants than to the parental B. cinerea strain. On tomato leaves, phenazine-producing Pseudomonas strains were significantly more effective in reducing gray mold symptoms incited by a ABcatrB mutant than by the parental strain. We conclude that the ABC transporter BcatrB provides protection to B. cinerea in phenazine-mediated interactions with Pseudomonas spp. Collectively, these results indicate that fungal ABC transporters can play an important role in antibiotic-mediated interactions between bacteria and fungi in plant-associated environments. The implications of these findings for the implementation and sustainability of crop protection by antagonistic microorganisms are discussed.     

Mannityl opine analogs allow isolation of catabolic pathway regulatory mutants

Five virulent Agrobacterium spp. strains that can catabolize the mannityl opines mannopine (MOP), mannopinic acid ( MOA ), and agropinic acid (AGA) were tested for their ability to grow on analogs of these compounds. Analogs containing alternative amino acids replacing glutamic acid or glutamine were generally refused by these bacteria, but mutants were obtained that catabolized the entire family of analogs. In the case of strain C58C1 (pRi 8196), we demonstrated that typical mutants were constitutive for MOP uptake, whereas the wild-type parent was inducible by MOP. Analogs of MOA prepared from a variety of sugars instead of mannose were generally refused, except for a strain carrying pTi B6-806, which grew well on all such analogs. The analogs allowed selection of mutants of all strains. Although most wild-type strains were inducible for AGA uptake, typical mutants selected from strain C58C1 (pRi 8196) were found to be constitutive for uptake of AGA, as was the wild-type strain carrying pTi B6-806. Such constitutive mutants grew on all sugar analogs of MOP, MOA , and AGA tested. The pTi B6-806-containing strain was tested for growth on a more extended series of analogs, including tetrose , triose, diose , and disaccharide analogs, all of which were accepted. Only ketose analogs were refused. Selection of promiscuous regulatory mutants by the two types of opine analogs suggests that the repressor proteins of MOP and AGA permease/ catabolase systems are chiefly responsible for the specificity of the pathways.     

Biological control of Botrytis cinerea using the antagonistic and endophytic Burkholderia cepacia Cs5 for vine plantlet protection

Antifungal activity of the Burkholderia cepacia Cs5 was tested in vitro and in vivo for the control of Botrytis cinerea . Bacterial biomass was significantly improved by the amendment of ZnSO(4), Mo(7)(NH(4))(6)O(24), and mannitol to the NBY medium; consequently, the amount of the secreted fungicides was increased. The quantification of B. cinerea inhibition, in liquid and solid conditions, showed an important sensitivity of this fungus to the strain Cs5 fungicides. Microscopic monitoring impact of these fungicides on mycelium structure showed an important increase in their diameter and ramifications in the presence of 0.75% supernatant. For the in vivo application of the strain Cs5, Vitis vinifera plantlets were inoculated with a Cs5 bacterial suspension, then with B. cinerea spores. The plantlets protection was total and durable when these two inoculations were made 3 weeks apart, which is the time for the endophytic bacterium to colonize the plantlets up to the top leaves. This protection is due to Cs5 antagonism and the elicitation of the plantlets self-defense via the root overgrowth.     

Biodegradation of endosulfan and endosulfan sulfate by <Emphasis Type="Italic">Achromobacter xylosoxidans</Emphasis> strain C8B in broth medium

Endosulfan is one of the most widely used wide spectrum cyclodiene organochlorine insecticide. In environment, endosulfan can undergo either oxidation or hydrolysis reaction to form endosulfan sulfate and endosulfan diol respectively. Endosulfan sulfate is as toxic and as persistent as its parent isomers. In the present study, endosulfan degrading bacteria were isolated from soil through selective enrichment technique using sulfur free medium with endosulfan as sole sulfur source. Out of the 8 isolated bacterial strains, strain C8B was found to be the most efficient endosulfan degrader, degrading 94.12% α-endosulfan and 84.52% β-endosulfan. The bacterial strain was identified as Achromobacter xylosoxidans strain C8B on the basis of 16S rDNA sequence similarity. Achromobacter xylosoxidans strain C8B was also found to degrade 80.10% endosulfan sulfate using it as sulfur source. No known metabolites were found to be formed in the culture media during the entire course of degradation. Besides, the bacterial strain was found to degrade all the known endosulfan metabolites. There was marked increase in the quantity of released CO₂ from the culture media with endosulfan as sulfur source as compared to MgSO₄ suggesting that the bacterial strain, Achromobacter xylosoxidans strain C8B probably degraded endosulfan completely through the formation of endosulfan ether.     

番茄灰霉病拮抗内生细菌的筛选、鉴定及其活性

对安徽省淮北市番茄植株根、茎、叶中内生细菌及其数量进行了调查和筛选,并测定了其抑菌活性。结果表明,番茄根、茎和叶中的内生细菌的数量分别为5.69×105、5.16×105和2.83×105CFU.g-1鲜重。根据分离部位和表型特征,从健康番茄植株体内分离到267株内生细菌,通过对峙实验,筛选到11株对番茄灰霉病菌有拮抗作用的菌株,占所分离内生细菌总数的4.12%。来自茎组织中的菌株XF136的抑菌效果最佳,抑菌带宽度达32.2mm。根据形态特征、生理生化特性、(G+C)mol%和16SrDNA序列分析,将菌株XF136鉴定为解淀粉芽孢杆菌(Bacillus amyloliquefaciens)。室内测定菌株XF136发酵滤液对灰霉病菌菌丝生长及分生孢子萌发的抑制作用,结果表明,菌株XF136发酵滤液可以抑制灰霉病菌菌丝生长和分生孢子萌发,且浓度越高,抑制能力越强;当发酵滤液浓度为20%时则完全抑制灰霉病菌菌丝生长和分生孢子萌发。盆栽防效试验结果表明,10%菌株XF136发酵滤液对番茄灰霉病防治效果与50%多菌灵600倍液相当,20%发酵滤液对番茄灰霉病的防治效果高于50%多菌灵600倍液。本研究表明,菌株XF136是防治番茄灰霉病潜在的优良生防菌株,具有良好的开发应用价值。     

Survey of taurine uptake and metabolism in Staphylococcus aureus

Taurine has been reported to be a component of the capsular polysaccharide of the encapsulated M strain of Staphylococcus aureus. This led to a study of the uptake and metabolism of [1,2-14C]taurine in a variety of encapsulated and unencapsulated S. aureus strains. Taurine was taken up by all strains studied. A discrepancy between uptake measured as depletion of radioactivity from growth medium and as cell-associated radioactivity suggested that taurine may be catabolized to CO2 in some strains. In most strains, cell-associated radioactivity was located mainly in cold TCA-soluble (pool metabolites) fractions. About 90% of the cell-associated radioactivity was present in the pool metabolites fraction in the M strain, and about 10% in hot TCA-soluble (nucleic acid-teichoic acid-capsular polysaccharide) fraction. Radioactivity in spent medium and the capsular polysaccharide-containing fraction appeared to be present as taurine in this strain. Radioactivity in the pool metabolites fraction of three of the strains examined did not chromatograph as taurine, indicating that taurine was converted into other cell metabolites. One strain incorporated radioactivity from taurine into cellular macromolecules, thus revealing a heterogeneity of staphylococcal taurine metabolism.     

Degradation of 4-aminobenzenesulfonate by a two-species bacterial coculture

The mutualistic interactions in a 4-aminobenzenesulfonate (sulfanilate) degrading mixed bacterial culture were studied. This coculture consisted of Hydrogenophaga palleronii strain S1 and Agrobacterium radiobacter strain S2. In this coculture only strain S1 desaminated sulfanilate to catechol-4-sulfonate, which did not accumulate in the medium but served as growth substrate for strain S2. During growth in batch culture with sulfanilate as sole source of carbon, energy, nitrogen and sulfur, the relative cell numbers (colony forming units) of both strains were almost constant. None of the strains reached a cell number which was more than threefold higher than the cell number of the second strain. A mineral medium with sulfanilate was inoculated with different relative cell numbers of both strains (relative number of colony forming units S1:S2 2200:1 to 1:500). In all cases, growth was found and the proportion of both strains moved towards an about equal value of about 3:1 (strain S1:strain S2). In contrast to the coculture, strain S1 did not grow in a mineral medium in axenic culture with 4-aminobenzenesulfonate or any other simple organic compound tested. A sterile culture supernatant from strain S2 enabled strain S1 to grow with 4-aminobenzenesulfonate. The same growth promoting effect was found after the addition of a combination of 4-aminobenzoate, biotin and vitamin B₁₂. Strain S1 grew with 4-aminobenzenesulfonate plus the three vitamins with about the same growth rate as the mixed culture in a mineral medium. When (resting) cells of strain S1 were incubated in a pure mineral medium with sulfanilate, up to 30% of the oxidized sulfanilate accumulated as catechol-4-sulfonate in the culture medium. In contrast, only minor amounts of catechol-4-sulfonate accumulated when strain S1 was grown with 4ABS in the presence of the vitamins.Abbreviations 4ABS 4-aminobenzenesulfonate - CFU colony forming units - 4CS catechol-4-sulfonate - 4HB 4-hydroxybenzoate     

Real-time RT-PCR expression analysis of chitinase and endoglucanase genes in the three-way interaction between the biocontrol strain Clonostachys rosea IK726, Botrytis cinerea and strawberry

Clonostachys rosea is a well-known biocontrol agent against Botrytis cinerea, the causal agent of gray mold in strawberry. The activity of cell wall-degrading enzymes might play a significant role for successful biocontrol by C. rosea. The expression pattern of four chitinases, and two endoglucanase genes from C. rosea strain IK726 was analyzed using real-time RT-PCR in vitro and in strawberry leaves during interaction with B. cinerea. Specific primers were designed for beta-tubulin genes from C. rosea and B. cinerea, respectively, and a gene encoding a DNA-binding protein (DBP) from strawberry, allowing in situ activity assessment of each fungus in vitro and during their interaction on strawberry leaves. Growth of B. cinerea was inhibited in all pathogen-antagonist interactions while the activity of IK726 was slightly increased. In all in vitro interactions, four of the six genes were upregulated while no change in expression of two endochitinases was measured. In strawberry leaves, the chitinase genes were upregulated 2-12-fold, except one of the endochitinases, whereas no change in expression of the two endoglucanases was measured. The results suggest that three out of four chitinase genes of IK726 are involved in biocontrol on leaves. This is the first example of monitoring of expression of chitinolytic genes in interactions between biocontrol agents and pathogens in plant material.     

Investigations on the destruxin production of the entomopathogenic fungus Metarhizium anisopliae

The dynamics of cyclic peptide destruxins (dtxs) produced by Metarhizium anisopliae strains V245 and V275 were monitored both on solid and in liquid media. The results showed that both strains did not produce dtxs in large-scale fermenter cultures or solid Czapek Dox (CD) agar. Production of the major dtxs A and B could be determined in both strains when grown on rice for up to 10-30 days. The main dtxs A, B, E, and E diol were detected in CD liquid culture filtrate from both strains after three days post-inoculation on. Parallel decrease of dtx E and increase of E diol in the culture medium were found, indicating that the latter is the hydrolytic product from the former. Production of dtxs A and B was significantly positively correlated. A negative correlation was observed between the production of the metabolites and pH value of the medium. The influence of different nutrient sources on dtx production was evaluated by using media with different carbon and nitrogen ratios as well as with different insect homogenates. The findings showed that the amount of dtxs A, B, and E increased with the increasing content of peptone in the medium. When insect homogenate was used as single nutrient source or added to CD medium, no toxins were detected in the culture filtrate. The potential risk posed by the toxic metabolites during mass production is discussed.     

哈尔滨地区假丝酵母菌DNA异质性及药物敏感性分析

研究假丝酵母菌的DNA异质性及药物敏感性,为预防和监控院内假丝酵母菌感染奠定基础。将临床分离的假丝酵母菌菌株,用科玛嘉显色培养基鉴定菌种,经纸片扩散法进行药敏试验,应用随机扩增多态性DNA（RAPD）技术对这些菌株进行基因分型。结果显示：93株假丝酵母菌中白假丝酵母菌68株,非白假丝酵母菌25株,所有菌株对制霉菌素,两性霉素B两种药物的敏感率最高（100％）,酮康唑其次（70．9％）,氟康唑的敏感率最低（50．5％）,引物1和引物2将来源不同的68株白假丝酵母菌分别分成4型（A1、B1、C1、D1）和6型（A2、B2、C2、D2、E2、F2）。哈尔滨地区的假丝酵母菌感染以白假丝酵母菌为主,且主要为A1、B1型（引物1）或A2、B2型（引物2）;基因型与药敏谱无明显相关性。     

芽孢杆菌和欧文氏菌的原生质体融合的研究

采用原生质体融合技术对带有链霉素标记的芽孢杆菌B12 13 2 和带有红霉素标记的欧文氏菌E2 6 2 3 进行原生质体融合。采用选择性及非选择性培养基对融合子进行 10次重复性传代试验后 ,获得稳定的融合子 2 84株 ,经重复性苎麻脱胶试验 ,从中获得 6株脱胶效果较好的融合子。     

Differential Behaviour of Mycelial Growth of Several Botrytis cinerea Strains on either Patchoulol- or Globulol-amended Media

Botrydial and dihidrobotrydial are two characteristic metabolites of the phytopathogenic fungus Botrytis cinerea , which are involved in the development of necrotic lesions on grapevine and tobacco. Patchoulol and globulol, two natural products which are analogues to precursors of botrydial and dihidrobotrydial, were tested on 10 B. cinerea strains which were isolated from different hosts and varied in aggressiveness on grapevine leaves. Mycelial growth of all strains was prevented when they were grown on either patchoulol- or globulol-amended malt agar media (200  μ g/ μ l). Each strain displayed a specific response pattern to those products, according to the high variability previously described for this species. Furthermore, strains were different from one another with regard to their level of aggressiveness against leaves detached from sherry grapevine vineyards.     

Extracellular glycosyl hydrolase activity of the clostridia producing acetone, butanol, and ethanol

Production of acetone, butanol, ethanol, acetic acid, and butyric acid by three strains of anaerobic bacteria, which we identified as Clostridium acetobutylicum, was studied. The yield of acetone and alcohols in 6% flour medium amounted to 12.7-15 g/l with butanol constituting 51.0-55.6%. Activities of these strains towards xylan, beta-glucan, carboxymethylcellulose, and crystalline and amorphous celluloses were studied. C. acertobutylicum 6, C. acetoburylicum 7, and C. acertobutylicum VKPM B-4786 produced larger amounts of acetone and alcohols and displayed higher cellulase and hemicellulase activities than the type strain C. acetobutylicum ATCC 824. It was demonstrated that starch in the medium could be partially substituted with plant biomass.     

Activity of Fluazinam against Strains of Botrytis cinerea Resistant to Benzimidazoles and/or Dicarboximides and to a Benzimidazole-phenylcarbamate Mixture

Fluazinam is a new active ingredient for the control of grey mould, belonging to the novel broad spectum phenylpyridinamine fungicides. The effect of fluazinam was studied on one wild type and four strains of Botrytis cinerea , which had been isolated from vegetable crops in Greece, and were resistant to benzimidazoles and/or dicarboximides and to the mixture of benzimidazoles (carbendazim) + phenylcarbamates (diethofencarb). In vitro fluazinam was found to be highly active against strains of B. cinerea which were sensitive or resistant to benzimidazoles or exhibited multiple resistance to benzimidazoles, dicarboximides and to the mixture carbendazim + diethofencarb [EC₅₀ and EC₉₅ values (concentration of active ingredient that suppresses mycelial growth to 50 and 95%, respectively, of that of the fungus on fungicide-free agar medium) calculated with probit analysis, ranged from 0.044 to 0.069 and 0.58 to 1.6 μg/ml, respectively]. No cross-resistance was observed between fluazinam and the market products benomyl, iprodione or carbendazim + diethofencarb. Preventive applications of fluazinam in vivo completely inhibited infections of cucumber seedlings by all the above-mentioned resistant strains of B. cinerea . Benomyl and iprodione did not effectively control the benzimidazole- and dicarboximide-resistant strains. The mixture of carbendazim + diethofencarb insufficiently controlled the strain of B. cinerea with moderate resistance to benzimidazoles. The results of this investigation indicate that it should be possible to use fluazinam as an alternative in resistance management programmes against grey mould.     

Temperature effects at the NADP+ and NAD+ levels in the mycelium of P. nigricans Thom. strains grown on different carbon sources]

The effect of elevated temperature of the medium on the levels of NADP+ and NAD+ in the mycelium of highly productive strain 117 and low productive strain B of P. nigricans Thom was studied at various developmental periods on the mineral medium in the presence of glucose, succinate or acetate. It was found that in the presence of glucose the elevated temperature markedly stimulated the initial growth of the culture, decreased the level of NADP+ in strain 117 by the 48th hour and increased the level of NAD+ in both the strains. At later periods it decreased the rate of the dinucleotide accumulation during the culture development. With the use of succinate under these conditions the levels of NADP+ in both the strains during the growth phase (the 2nd and 5th days) and by the 9th day markedly increased, while the NAD+ concentration increased only by the 2nd day in both the strains and by the 9th day in strain B. When the strains were grown in the presence of acetate, the elevated temperature especially affected the level of NAD+.     

Selective enrichment media bias the types of Salmonella enterica strains isolated from mixed strain cultures and complex enrichment broths

For foodborne outbreak investigations it can be difficult to isolate the relevant strain from food and/or environmental sources. If the sample is contaminated by more than one strain of the pathogen the relevant strain might be missed. In this study mixed cultures of Salmonella enterica were grown in one set of standard enrichment media to see if culture bias patterns emerged. Nineteen strains representing four serogroups and ten serotypes were compared in four-strain mixtures in Salmonella-only and in cattle fecal culture enrichment backgrounds using Salmonella enrichment media. One or more strain(s) emerged as dominant in each mixture. No serotype was most fit, but strains of serogroups C2 and E were more likely to dominate enrichment culture mixtures than strains of serogroups B or C1. Different versions of Rappaport-Vassiliadis (RV) medium gave different patterns of strain dominance in both Salmonella-only and fecal enrichment culture backgrounds. The fittest strains belonged to serogroups C1, C2, and E, and included strains of S. Infantis, S. Thompson S. Newport, S. 6,8:d:-, and S. Give. Strains of serogroup B, which included serotypes often seen in outbreaks such as S. Typhimurium, S. Saintpaul, and S. Schwarzengrund were less likely to emerge as dominant strains in the mixtures when using standard RV as part of the enrichment. Using a more nutrient-rich version of RV as part of the protocol led to a different pattern of strains emerging, however some were still present in very low numbers in the resulting population. These results indicate that outbreak investigations of food and/or other environmental samples should include multiple enrichment protocols to ensure isolation of target strains of Salmonella.     

番茄灰霉病拮抗内生放线菌的筛选、鉴定及其活性评价

对安徽省淮北市番茄植株根、茎、叶中内生放线菌进行了分离、筛选,并测定了其抑菌活性。结果表明:番茄根、茎和叶中的内生放线菌的数量分别为5.66×104、0.67×104和0.39×104CFU.g-1鲜重。根据分离部位和表型特征,从健康番茄植株体内分离到93株内生放线菌,通过对峙实验,筛选到7株对番茄灰霉病菌有拮抗作用的菌株,占所分离内生放线菌总数的7.5%。来自根组织中的菌株HNU-EA27的抑菌效果最佳,抑菌圈直径达28.3mm。根据形态特征、培养特征、生理生化特性、细胞壁组分和16SrDNA序列分析,将菌株HNU-EA27鉴定为毒三素链霉菌(Streptomyces toxytricini)。室内测定菌株HNU-EA27发酵滤液对灰霉病菌菌丝生长及分生孢子萌发的抑制作用,结果表明:菌株HNU-EA27发酵滤液可以抑制灰霉病菌菌丝生长和分生孢子萌发,且浓度越高,抑制能力越强;当发酵滤液浓度为30%时则完全抑制灰霉病菌菌丝生长和分生孢子萌发。盆栽防效试验结果表明:30%菌株HNU-EA27发酵滤液对番茄灰霉病的预防与治疗效果分别为80.6%和73.8%,均高于50%多菌灵可湿性粉剂600倍液。本研究表明,菌株HNU-EA27是防治番茄灰霉病潜在的优良生防菌株,具有良好的开发应用价值。