单宁对苏云金芽孢杆菌库斯塔克变种杀棉铃虫效力的作用(英文)

本文报道棉铃虫、苏云金芽孢杆菌库斯塔克变种和单宁三者间的相互作用。苏云金芽孢杆菌芽孢晶体混合粉以6个不同的浓度（0％,0．005％,0．01％,0．015％,0．02％,0．025％湿重）分别加入含0．025％单宁和不含单宁的人工饲料中。初孵幼虫分别在上述饲料中饲养48h后,转入相应的不含苏云金芽孢杆菌的人工饲料上饲养,直到留存的幼虫化蛹。结果表明,单宁与苏云金芽孢杆菌混用可提高苏云金芽孢杆菌对棉铃虫的毒力,LD50降低45％,并可显著抑制该虫的生长发育,但二者在抑制生长发育方面无交互作用。选择取食试验显示,苏云金芽孢杆菌对五龄幼虫有显著的抑食性,但单宁无此作用,单宁与苏云金芽孢杆菌混用不会增强抑食性,但单宁能抑制苏云金芽孢杆菌菌落的生长,浓度高于15mg／100ml还能抑制芽孢的萌发。     

EFFECT OF TANNIC ACID ON THE EFFECTIVENESS OF BACILLUlS THURINGIENSIS VAR. KURSTAKI AGAINST HELICOVERPA ARMIGERA (HÜBNER)*

Abstract The role of tannic acid in increasing effectiveness of Bacillus thuringiensis var. kurstaki (HD-1) against Helicover pa armigera was examined in bioassays on semisynthetic diet. Concentrations of B. thuringiensis (0 %, 0.005 %, 0. 01 %, 0.015 %, 0.02 %, 0.025 % wet weight) were incorporated into the diet containing 0. 025% tannic acid and tannic acid-free diet. LD₅₀ of B. thuringiensis with tannic acid were 0.006% but that without tannic acid was 0.011%. Both B. thuringiensis and tannic acid retarded growth of H. armigera significantly, but there was no synergetic effect between them. Choice tests showed that B, thuringiensis deterred feeding of the fifth instar larvae of H. armigera but tannic acid had no such effect. Experiments on colony growth of B. thuringiensis on NBA media containing tannic acid (0, 1, 3, 6, 9, 12, 15, 18, 21 mg/100 ml) demonstrated that tannic acid reduced colony growth of B. thuringiensis, and inhibited sporulation above 15 mg/100 ml.     

Glass beads as a solid matrix in in vitro study of the role of polyamines in cold hardiness of white clover

Glass beads were used as an alternative to agar in the study of the role of polyamines in cold hardiness of white clover. Plantlet growth performance, in vitro hardening and cold stress tolerance were similar on both the agar solidified medium and the liquid medium in glass beads matrix. Glass beads allowed media exchange without plant transfer and an easy monitoring of the uptake of putrescine synthesis inhibitor, ¹⁴C difluoromethylornithine from the medium. The matrix is recommended in in vitro studies of whole plant physiology, screening procedures and bioassays where media exchange and/or uniform application of a selection pressure is required. There is also 60% saving on media components and the beads can be re-used after acid wash.Abbreviations DFMA difluoremethylarginine - DFMO difluoromethylornithine - MS Murashige and Skoog salts - Kcpm thousand counts per minute     

Extracellular enzymatic activity of aquatic and aero-aquatic conidial fungi

Nineteen species of aquatic and areo-aquatic conidial fungi were tested for their ability to produce extracellular enzymes which degrade cellulose, starch, lipids, proteins and tannic acid. The cellulolytic activity was determined by using both solid and liquid media. The activity of other enzymes was examined using solid media. Two-thirds of the species were able to hydrolyze soluble cellulose (CMC) incorporated in solid and liquid media with varying degrees of activity. Extracellular culture filtrates ofAegerita candida, Helicodendron giganteum andH. tubulosum contained a Cl-Cx enzyme complex that could degrade both soluble cellulose (CMC) and crystalline cellulose (filter paper). Lipase activity was demonstrated by 11 species. Fourteen of the species showed activity for amylase and protease, but only 11 of the 16 were capable of degrading tannic acid.     

Streptococcus caprinus sp.nov., a tannin-resistant ruminal bacterium from feral goats

An undescribed bacterium capable of clearing tannic acid-protein complexes has been isolated from ruminal contents of feral goats browsing tannin-rich Acacia species. The bacterium is a Gram-positive facultative anaerobe, characterized as a Streptococcus , but DNA-DNA hybridization and 16S rDNA sequencing show that it is distinct from the common ruminal species Strep. bovis. We propose the name Streptococcus caprinus for this species. The type strain is Strep. caprinus 2.3, Australian Collection of Microorganisms (ACM) 3969. The bacterium grows in media containing at least 2.5% w/v tannic acid or condensed tannin and produces zones of clearing around colonies on nutrient agar plates with added tannic acid. Streptococcus caprinus is not a major inhabitant of domestic livestock, but is found in feral goats browsing tannin-rich Acacia species, at a population of up to 2 times 10⁶ cfu ml^-1 of rumen fluid.     

Nutritional Requirements of the Nematophagous Fungus Hirsutella rhossiliensis

Six natural media were examined for growth and sporulation of six isolates of the nematophagous fungus Hirsutella rhossiliensis , using solid and/or liquid culture. Twenty carbohydrates, 19 nitrogen (N) compounds, and nine vitamins were also tested for their effects on growth, sporulation, and spore germination of a further three isolates (ATCC46487, OWVT-1 and JA16-1). Variations in nutritional requirements existed among the fungal isolates. In general, V-8 juice agar (VA), cornmeal agar and potato dextrose agar were good media for growth, and malt extract agar, VA and yeast dextrose agar were good for sporulation of all six isolates. Glycogen was the best and sucrose, inulin, D- ( + ) - trehalose and soluble starch were also good carbon (C) sources for growth and spore germination of the three isolates ATCC46487, OWVT-1 and JA16-1 in both liquid and solid culture. None of the isolates utilized D- ( + )xylose as a C source. L- sorbose, D- ribose, citric acid and D- fructose were poor for growth of all isolates. The best C source for sporulation was D- ( + )-trehalose for ATCC46487, D- sorbitol for OWVT-1 and D- ( + )-cellobiose for JA16-1. Casein was the best N source for growth of ATCC46487 and OWVT-1, while peptone was best for JA16-1. L- asparagine, L- proline, and peptone were also good for growth of all three isolates. L - cystine was not utilized by H. rhossiliensis and DL- methionine inhibited growth of all isolates. Spore germination of all isolates was well supported by most N compounds examined but was inhibited by L- cystine. No significant difference in sporulation of ATCC46487 was observed among the N sources. DL- threonine was the best N source for spore production by OWVT-1 and L- phenylalanine was best for JA16-1. Vitamins generally enhanced fungal growth and sporulation, with thiamine having the greatest influence. Excluding some vitamins individually from the medium containing all other test vitamins sometimes increased growth and/or sporulation of certain isolates.     

A modified method for the detection of microbial proteases on agar plates using tannic acid

In routine assay for the screening of microbes producing proteases, 10% trichloroaceticacid (TCA) is flooded on the milk agar plates after inoculation and required incubation to precipitate the protein. However, the clarity of the hydrolyzed zone is not very sharp and distinct. We herein present an improved assay for detecting the presence of extracellular protease from microorganisms on agar plates. In this method 10% tannic acid is flooded on the milk agar plate (in place of, TCA) to observe the zone of hydrolysis. Tannic acid sharply increases the colour intensity of the plate, as it favours the precipitation of the unhydrolyzed protein in the plate, thereby improving the contrast between the intact zones and the enzymatic lyses zones of the substrate. Our results indicate that this method is useful to detect extracellular proteases produced by both fungi as well as bacteria. The method used in the present study is sensitive, and can be easily performed for screening of large number of microbial cultures. This is the first report on the use of tannic acid for the detection of microbial proteases.     

A modified method for the detection of microbial proteases on agar plates using tannic acid

In routine assay for the screening of microbes producing proteases, 10% trichloroaceticacid (TCA) is flooded on the milk agar plates after inoculation and required incubation to precipitate the protein. However, the clarity of the hydrolyzed zone is not very sharp and distinct. We herein present an improved assay for detecting the presence of extracellular protease from microorganisms on agar plates. In this method 10% tannic acid is flooded on the milk agar plate (in place of, TCA) to observe the zone of hydrolysis. Tannic acid sharply increases the colour intensity of the plate, as it favours the precipitation of the unhydrolyzed protein in the plate, thereby improving the contrast between the intact zones and the enzymatic lyses zones of the substrate. Our results indicate that this method is useful to detect extracellular proteases produced by both fungi as well as bacteria. The method used in the present study is sensitive, and can be easily performed for screening of large number of microbial cultures. This is the first report on the use of tannic acid for the detection of microbial proteases.     

Method of determining the antimicrobial activity of alcohol extracts of propolis]

It is proposed to determine the antimicrobial activity of propolys alcohol extracts by the method of subsequent dilutions in solid nutrient media. Dilution of the extracts immediately in hot agar eliminated the inhibiting effect of the extragent on the microbial growth. Opalesence appearing in the agar did not prevent estimation of the results in contrast to the method of subsequent dilutions in liquid nutrient media.     

The effect of selected allelochemicals on germination of conidia and blastospores and mycelial growth of the entomopathogenic fungus,Paecilomyces fumosoroseus (Deuteromycotina: Hyphomycetes)

Selected allelochemicals that protect plants from invasion by plant pathogenic fungi were investigated for their activity against the entomopathogenic fungus, Paecilomyces fumosoroseus. The alkaloids tomatine, solanine, and camptothecin; the furanocoumarin, xanthotoxin; and the phenolic, tannic acid were tested for their effects on germination of conidia and blastospores and growth of mycelia. The LC50 values (corresponding to 50% inhibition of germination) for tomatine, solanine, camptothecin, xanthotoxin and tannic acid were 51.6, 95.9, 55.9, 83.0 and 72.8 mg/l respectively. When blastospores were placed on media containing a concentration of the individual allelochemicals that inhibit germination in approximately 50% of conidia, all but blastospores on tomatine had significantly less germination than did aerial conidia. Growth rates of mycelia were slowest in the camptothecin medium, followed by those of tomatine and xanthotoxin and were not significantly different from controls in the media containing solanine and tannic acid. A multitude of biotic and abiotic factors are responsible for specificity and degree of pathogenicity of entomopathogens. The effect of crop plant chemistry on the efficacy of entomopathogens should be quantified further in order to maximize their potential when used concomitantly with resistant plant varieties. This revised version was published online in June 2006 with corrections to the Cover Date.     

Discrepancies in the Enumeration of Escherichia coli

Stationary-phase cells of Escherichia coli were enumerated by the pour plate method on Trypticase soy agar containing 0.3% yeast extract (TSYA), violet red-bile agar, and desoxycholate-lactose agar, and by the most-probable-number method in Brilliant Green-bile broth and lauryl sulfate broth. Maximum counts were assumed to be those on TSYA. In general, numbers detected were lower with the selective solid media and higher with the selective liquid media. Inhibitory effects, especially on selective solid media varied with the strains of E. coli. The lower detection on selective solid media was partly due to the stress induced in some cells by the temperature of the melted media used in the pour plate method. These cells apparently failed to repair and form colonies in the selective media. Improved detection on the selective solid media was achieved by using 1% nonfat milk solids, 1% peptone, or 1% MgSO(4).7H(2)O in the dilution blanks. Higher detection on selective agar media was effected by surface plating or by surface-overlay plating of the cells. The surface-overlay method appeared to be superior for the direct enumeration of E. coli in foods.     

A xyloglucan from seeds of the native Brazilian species <Emphasis Type="Italic">Hymenaea courbaril</Emphasis> for micropropagation of Marubakaido and Jonagored apples

Xyloglucan was extracted from seeds of Hymenaea courbaril and mixed with agar to prepare a solid culture medium used for micropropagation of the Marubakaido apple rootstock (Malus prunifolia Borkh) and cv. Jonagored (Malus domestica). The performance on gels created from a blend of 0.4%agar and 0.2% xyloglucan (w/v) was compared with that on media gelled with a standard concentration 0.6% (w/v) of agar. The growth of shoots and the multiplication rate were higher on the modified culture medium than on the agar-gelled medium. The occurrence of hyperhydric shoots was lower on the modified medium. In the absence of auxin, shoot rooting reached 70% (Marubakaido) and 66% (Jonagored) on the agar-xyloglucan medium and 6.7% and 10.4%, respectively, on the agar medium. When 0.25 microM indole-3-butyric acid (IBA) was added to both media, the modified medium gave better results in terms of rooting percentage and quality of roots than the agar-gelled medium.     

The effects of culture media, solid substrates, and relative humidity on growth, sporulation and conidial discharge of Valdensinia heterodoxa

Valdensinia heterodoxa (Sclerotiniacae) is a potential fungal bioherbicide for control of salal (Gaultheria shallon). The effect of culture media, substrates and relative humidity (RH) on growth, sporulation and conidial discharge of V. heterodoxa was determined for two isolates PFC2761 and PFC3027 in vitro. Culture media significantly affected the growth, sporulation, and conidial discharge of V. heterodoxa. Of eight agar media used, colony radial growth was optimal on salal oatmeal agar and salal potato dextrose agar for isolates PFC2761 and PFC3027, respectively; whereas sporulation was at an optimum on salal oatmeal agar for both isolates. Of the eight liquid media tested, mycelial production was highest on wheat bran–salal–potato dextrose broth. Growth on solid substrates greatly stimulated sporulation and conidial discharge of V. heterodoxa. Of the 12 solid substrates used, the greatest numbers of discharged conidia were observed from wheat bran and wheat bran–salal within 14 d of sporulation. Sporulation on solid substrates continued for 42 d. RH significantly affected the sporulation and conidial discharge for both isolates across all solid substrates tested. No conidia were produced or discharged below 93 % RH on wheat bran–salal and millet. With an increase of the RH from 93 to 97 %, sporulation and the number of discharged conidia increased significantly for both isolates on wheat bran–salal, but not on millet.     

Characterization of Growth and Conidia Production of Exserohilum monoceras on Different Substrates

On agar media the maximum conidia production of Exserohilum monoceras occurred on V-8 juice agar (VA) or centrifuged V-8 juice agar, whereas the optimal radial mycelial growth occurred on Czapek-Dox agar. The optimal temperatures for radial mycelial growth and conidia production were 28 and 27°C respectively. Light prohibited E. monoceras conidia production. The best sporulation occurred under continuous dark conditions. Echinochloa leaf decoction significantly increased conidia production on potato dextrose agar (PDA) and VA, and significantly increased germ tube length on PDA, lima bean agar and VA, but did not affect conidia germination. No conidia were produced in liquid media. Of 22 agricultural-based products evaluated as solid substrates, the most abundant sporulation (1.8 × 10⁶ conidia g^-1 of dry weight) occurred on corn leaves. The conidia production of E. monoceras on corn leaves was affected by incubation period, moisture content and substrate quantity. There were no differences in germination rate, germ tube length and virulence of conidia produced on agar media or corn leaves.     

单宁酸对根田鼠断乳幼体生长和存活的影响

在食物中含10％和20％蛋白质条件下,测定了单宁酸对根田鼠幼体在断乳后20d内的生长和存活的影响。结果表明,食物中含蛋白质为10％的条件下,摄食含3％和6％单宁酸食物的试验个体生长速率分别为-0．135g/d和-0．25g/d,食物利用效率均显低于对照组,断乳后20d内平均存活天数较对照组分别下降26．23％和49．36％。在食物蛋白质为20％的条件下,摄食含3％和6％单宁酸食物的试验个体生长速率分别为0．134g/d和-0．116g／d,摄食6％单宁酸食物的试验个体食物利用效率显低于摄食3％单宁酸食物的试验个体和对照组个体,断乳后20d内的平均存活天数较对照组下降39．41％,摄食3％单宁酸食物的试验个体较对照组略有下降,但不显。上述结果验证了单宁酸能显影响植食性小哺乳动物生长和存活的假设。     

Selection of Media for the Isolation of Common Bacterial L-Phase Organisms from a Clinical Specimen

The L-phase of 13 bacteria commonly associated with disease were induced by penicillin and inoculated into various solid and broth media; their growth was recorded for a period of 14 days. Plates containing highly purified agar and sucrose as the stabilizing agent and those incubated under aerobic conditions gave the best results. Magnesium seems to be necessary for growth in broth media on primary isolation, although it may not be necessary on multiple transfers after a more stable state has been reached. Growth in broth media is much more difficult to achieve. Reversion is aided by using a higher concentration of agar in plates, by decreasing the sucrose concentration, and by omitting the antibiotics and horse serum. A procedure has been outlined for the routine culture and identification of L-phase organisms from a clinical specimen.     

Antimicrobial solid media for screening non-sterile Arabidopsis thaliana seeds

Stable genetic transformation of plants is a low-efficiency process, and identification of positive transformants usually relies on screening for expression of a co-transformed marker gene. Often this involves germinating seeds on solid media containing a selection reagent. Germination on solid media requires surface sterilization of seeds and careful aseptic technique to prevent microbial contamination, but surface sterilization techniques are time consuming and can cause seed mortality if not performed carefully. We developed an antimicrobial cocktail that can be added to solid media to inhibit bacterial and fungal growth without impairing germination, allowing us to bypass the surface sterilization step. Adding a combination of terbinafine (1 μM) and timentin (200 mg l⁻¹) to Murashige and Skoog agar delayed the onset of observable microbial growth and did not affect germination of non-sterile seeds from 10 different wild-type and mutant Arabidopsis thaliana accessions. We named this antimicrobial solid medium “MSTT agar”. Seedlings sown in non-sterile conditions could be maintained on MSTT agar for up to a week without observable contamination. This medium was compatible with rapid screening methods for hygromycin B, phosphinothricin (BASTA) and nourseothricin resistance genes, meaning that positive transformants can be identified from non-sterile seeds in as little as 4 days after stratification, and transferred to soil before the onset of visible microbial contamination. By using MSTT agar we were able to select genetic transformants on solid media without seed surface sterilization, eliminating a tedious and time-consuming step.     

Intraspecific variability in growth response to cadmium of the wood-rotting fungus Piptoporus betulinus

The intraspecific variability in growth response to cadmium (Cd) on agar media and in liquid culture was studied among fourteen strains of a wood-rotting fungus Piptoporus betulinus. The variability of Cd tolerance was found to be very high. The ED(50) ranged from 6.8 μM Cd in the most sensitive strain, up to 255.1 μM in the most resistant one. On agar media the addition of Cd to nutrient media resulted in reduction of relative growth rate and increased lag time. While the reduction of growth rate was already apparent at 10 μM Cd, the lag time was significantly increased in higher Cd concentrations. Five strains of P. betulinus failed to grow at 250 μM Cd and none grew at 500 μM metal. Biomass production in liquid culture was less sensitive to addition of Cd than the growth rate on solid media. At 100 μM Cd the radial growth rate of the mycelium was reduced to 27%, whereas the dry mass of mycelium was 77% of the respective control value. A group of four Cd-sensitive strains was found, showing low metal tolerance both on solid media and in liquid cultures. Although the isolates originated from sites with different Cd-pollution level, no correlation between level of Cd-pollution and resistance (ED(50)) was found. The growth rate of fourteen tested strains displayed lower variability than biomass production, showing that radial growth rate is more species-specific and therefore more valuable for interspecific comparisons of growth response.     

lbtA and lbtB are required for production of the Legionella pneumophila siderophore legiobactin

Under iron stress, Legionella pneumophila secretes legiobactin, a nonclassical siderophore that is reactive in the chrome azurol S (CAS) assay. Here, we have optimized conditions for legiobactin expression, shown its biological activity, and identified two genes, lbtA and lbtB, which are involved in legiobactin production. lbtA appears to be iron repressed and encodes a protein that has significant homology with siderophore synthetases, and FrgA, a previously described iron-regulated protein of L. pneumophila. lbtB encodes a protein homologous with members of the major facilitator superfamily of multidrug efflux pumps. Mutants lacking lbtA or lbtB were defective for legiobactin, producing 40 to 70% less CAS reactivity in deferrated chemically defined medium (CDM). In bioassays, mutant CDM culture supernatants, unlike those of the wild type, did not support growth of iron-limited wild-type bacteria in 2',2'-dipyridyl-containing buffered charcoal yeast extract (BCYE) agar and a ferrous iron transport mutant on BCYE agar without added iron. The lbtA mutant was modestly defective for growth in deferrated CDM containing the iron chelator citrate, indicating that legiobactin is required in conditions of severe iron limitation. Complementation of the lbt mutants restored both siderophore expression, as measured by the CAS assay and bioassays, and bacterial growth in deferrated, citrate-containing media. The lbtA mutant replicated as the wild type did in macrophages, amoebae, and the lungs of mice. However, L. pneumophila expresses lbtA in the macrophage, suggesting that legiobactin, though not required, may play a dispensable role in intracellular growth. The discovery of lbtAB represents the first identification of genes required for L. pneumophila siderophore expression.     

Improved Solid Medium for the Detection and Enumeration of Pectolytic Bacteria

An improved solid agar medium (MP medium) has been developed which allows detection of pectolytic activity in bacteria. Organisms tested exhibited a variety of regulatory controls governing pectate lyase synthesis. The medium contains mineral salts, pectin, and yeast extract. After growth of the organisms, the agar plate is flooded with a polysaccharide precipitant, and pectolytic activity is shown by clear zones around active colonies. High concentrations of phosphate are shown to be necessary for pectic enzyme formation on solid media. The medium has successfully been used to detect pectolytic organisms in soil, forest litter, and rotting vegetable samples.