The effect of accelerated seed ageing on cucumber germination following seed treatment with fungicides and microbial biocontrol agents for managing gummy stem blight by Didymella bryoniae

This study examined (1) the effect of the accelerated seed ageing on cucumber germination with treatments: Bacillus subtilis QST713 or Pseudomonas fluorescens CA in 1% methylcellulose and fungicides difenoconazole, carboxin or pyraclostrobin in 5% polyvinyl alcohol, and (2) the impact on disease severity of gummy stem blight (GSB) caused by Didymella bryoniae by the seed treatments and foliar spray application of methylcellulose-formulated B. subtilis or P. fluorescens. Difenoconazole, pyraclostrobin and microorganisms suppressed growth of D. bryoniae in a laboratory dual culture; carboxin had no effect on D. bryoniae growth. Germination of fungicide-treated seed was unaffected by accelerated seed ageing. Greenhouse: GSB disease severity with PVA and non-treated seed was 89% and 84%, respectively, whereas, difenoconazole, carboxin and pyraclostrobin, was significantly reduced, 56%, 53% and 40%, respectively. Germination of Bacillus-treated seed was unaffected by accelerated seed ageing, but was significantly reduced with Pseudomonas-treated seed. GSB disease severity with B. subtilis or P. fluorescens-treated seed was inconsistent; however, foliar spray application of B. subtilis significantly reduced GSB. Accelerated seed ageing exposed a significant negative impact on seed germination with P. fluorescens. Based on the accelerated ageing test, P. fluorescens-treated cucumber seed is detrimental to seed survival and therefore, is not a candidate for biocontrol activities for cucumber requiring seed storage.     

Growth promotion activity and biological control for the management of leaf blight incited by Alternaria helianthi

The Pseudomonas fluorescens (Pf1), Bacillus subtilis (Bs1) are the major potential biocontrol agents against foliar pathogens. MPf1 and MBs1 were found to be the most effective in inhibiting the mycelial growth of Alternaria helianthi. These biocontrol agents have the maximum capacity in controlling the spore germination, and data showing the growth-promoting effect of biocontrol agents and inhibition of seed-borne fungi are available. Seed-borne infections of A. helianthi are controlled by seed treatment with P. fluorescens, which showed least seed infection. The root length and shoot length has also been increased.     

Proline-based modulation of 2,4-diacetylphloroglucinol and viable cell yields in cultures of Pseudomonas fluorescens wild-type and over-producing strains

The antifungal compound 2,4-diacetylphloroglucinol (DAPG) is produced in the rhizosphere of wheat by pseudomonad populations responsible for the natural biological control phenomenon known as “take-all decline.” Studies were conducted to elucidate the impact of DAPG and its co-product 2,4,6-trihydroxyacetophenone (THA) on the production of Pseudomonas fluorescens for biological control. Increasing DAPG from 0.1 g/l to 0.5 g/l and THA from 0.05 g/l to 0.5 g/l significantly inhibited the growth and lowered the yield of viable bacteria in liquid cultures. On further examination of these metabolites applied in seed coatings, levels of DAPG and THA exceeding 0.05 mg/g seed significantly reduced wheat germination percentages. The three-way interaction of DAPG, THA, and culture medium ingredients was significant, and greatest seed germination loss (40–50%) was observed when 0.5 mg DAPG and 0.25 mg THA were combined in a coating of 0.5 ml culture medium per gram of seed. Based on the results of Biolog GN microplate, flask, and fermentor screens of C sources, proline was found to optimize the viable cell yields of the P. fluorescens strains tested. The combination of proline with glucose and urea as C and N sources in growth media could be optimized to minimize DAPG production and maximize the vitality of P. fluorescens Q8R1-96 and Q69c-80:miniTn5:phl20 (DAPG over-producer). In production cultures, the proline supply rate offers a potentially useful means to optimize the biological control agent yield and quality.     

Biological control of seedling blight of wheat caused by Fusarium graminearum with beneficial rhizosphere microorganisms

Fusarium graminearum is associated with the cereal damping-off complex which reduces germination, seedling stand and yield. Fifty-two bacterial strains and six Trichoderma spp. isolated from the wheat rhizosphere were evaluated for biocontrol of seedling blight of wheat caused by F. graminearum. Their potential as biocontrol agents was tested in vitro and in the greenhouse. Isolates varied in their ability to inhibit the mycelial growth of F. graminearum in agar plate bioassays by 0–79%. This parameter was not related with biocontrol efficacy of in vivo assays. In greenhouse trials, all isolates were initially evaluated for reducing disease on wheat cultivars Klein Centauro (moderately resistant to F. graminearum) and Pro INTA Oasis (susceptible) planted in sterilized soil artificially infested with the pathogen. Among the 25 bacteria and six fungal isolates that exhibited a pronounced suppressive effect, the most efficient 10 for both cultivars were further assayed on eight cultivars (Buck Candil, Buck Catriel, Buck Chambergo, Buck Poncho, Buck Topacio, Klein Cacique, Klein Centauro and Pro INTA Oasis) potted in cultivated–inoculated soil. Three weeks after sowing, plant stand, percentage of diseased emerging seedlings, plant height and dry weight were evaluated. Among the antagonists only Stenotrophomonas maltophilia was significantly better than the control for the average of the eight cultivars for plant stand, height and dry weight. Stenotrophomonas maltophilia also caused a non-significant decrease in the percentage of diseased plants. Three strains of Bacillus cereus and one isolate of Trichoderma harzianum gave also a good control in some cultivars. The ability of these isolates to affect the infection of wheat seedlings by F. graminearum may be of potential value in field trials.     

Methyl jasmonate stimulates polyamines biosynthesisand resistance against leaf rust in wheat plants

Abstract

Leaf rust of wheat (Triticum aestivum L.), incited by Puccinia recondita ex Desm. f. sp. tritici Eriks, is one of the most important wheat diseases in Egypt. Methyl jasmonate (MJ) is a potential plant elicitor which induces a wide range of chemical and anatomical defense reactions in conifers and might be used to increase systemic resistance against biotic damage. In the greenhouse, different concentrations of MJ (10, 20 and 30 mM) were applied as seed soaking plus foliar spray or only as foliar spray to control leaf rust and induction of secondary compound production in leaves of wheat plants. Foliar spray was applied after 30 and 50 days of sowing. Results indicated that all concentrations and treatments reduced the severity of rust disease caused by P. recondita f. sp. tritici in wheat leaves during 45 days of inoculations. Disease incidence was decreased significantly in MJ-treated plants as seed soaking plus foliar spray with 20 and 30 mM when compared to 10 mM MJ or control plants. The study revealed that, with increasing concentrations of MJ, the secondary metabolites were greatly increased. Endogenous levels of both free and conjugated putrescine, spermidine and spermine increased in response to the elicitor. Activities of polyamine biosynthetic enzymes of ornithine decarboxylase (ODC) and polyamine oxidase (PAO) displayed up to threefold increases relative to untreated control. Moreover, significant increases in activities of plant defense-related protein, enzymes as peroxidase and chitinase as well as free and conjugated phenols contents were recorded in treated plants compared with untreated and infected plants. Furthermore, MJ treatment increased the chlorophyll-a, chlorophyll-b and carotenoids pigments contents, the higher increase was obtained with combined treatment between seeds soaking plus foliar spray at 20 and 30 mM of MJ. Under field conditions, three concentrations of MJ, i.e. 10, 20 and 30 mM as combined treatment between seeds soaking plus foliar spray or only as foliar spray were applied to study their effect against rust disease. Foliar spray was applied after 30 and 80 days of sowing. Results showed that the high reduction in disease severity was obtained with combined treatments between seeds soaking plus foliar spray with MJ at 20 and 30 mM compared with other treatments and control. At the same time, all treatments increased the growth and grain yield of wheat plants. It could be suggested that combination treatment between seeds soaking plus foliar spray with methyl jasmonate might be used commercially for controlling rust disease of wheat plants under field conditions.     

Principal component analysis of interrelations in stored-wheat ecosystems infested with multiple species of insects

Principal component analysis (PCA) was used to determine the effects of infestation of bulk stored wheat by multiple species of insects at 30±2°C for 60 wk. Eight 204-liter drums containing wheat at 15.5% moisture content were used as three distinct man-made ecosystems: (a) Control system (2 drums), insect-free; (b) RST system (3 drums), artificially infested with Rhyzopertha dominica (F.),Sitophilus oryzae (L.), and Tribolium castaneum (Herbst ); and (c) COT system (3 drums), infested with Cryptolestes ferrugineus (Stephens ),Oryzaephilus surinamensis (L.), and Tribolium castaneum. The variables measured tri-weekly within each system included carbon dioxide, oxygen, temperature, grain moisture, seed damage, grain weight and volume, dust weight and volume, fat acidity values (FAV) of the wheat, seed germination, microflora including Alternaria alternata(Fr.) Keissler ,Aspergillus glaucus group, Aspergillus candidusLink , and bacteria, insects and the mite Tarsonemus granariusLindquist . PCA provided multivariate synopsis of the data quantifying several important relationships among the variables monitored. Tri-weekly and cumulative 60-wk analyses of each system showed that high bacterial counts were associated with high FAV levels;Tarsonemus numbers were positively related to Aspergillus; Alternaria and seed germination were negatively related to FAV, bacteria and grain damage; and that the number of insects was related to the presence of Aspergillus and negatively related to the presence of bacteria. Seed germination and Alternaria infection often decrease rapidly presumably because of infection by fungi of the Aspergillus glaucus group. The combined action of R. dominica and Aspergillus spp. enhanced seed damage and increased grain moisture content thus promoting bacterial growth which in turn inhibited insect and mold growth. Fat acidity values increased with time unless seed damage and bacterial infection were extensive as in the RST system.     

Bioagent based integrated management of Phytophthora blight of pigeonpea

Phytophthora blight (Phytophthora drechsleri f. sp. cajani) is one of the major yield limiting factors of short duration varieties of pigeonpea (Cajanus cajan). For eco-friendly and sustainable management of the disease, four antagonists, Pseudomonas fluorescens, Bacillus subtilis, Trichoderma viride and T. hamatum, were evaluated under in vitro and in vivo conditions. P. fluorescens produced maximum inhibition zone, while Trichoderma species overgrew on the host mycelium and caused lysis. The fungicides apron (metalaxyl), ridomil MZ (metalaxyl + mancozeb), capton (captaf), difolatan (captafol), thiram (tetramethylthiuram disulphide) and bavistin (carbendazim) at both 0.3 and 0.6% were found to be compatible with P. fluorescens while B. subtilis was compatible only with apron at 0.3%. T. viride grew and sporulated well at 0.3 and 0.6% apron and 0.2% ridomil MZ. The bioagents proved to have potential and those which were compatible under in vitro were evaluated under field conditions. The efficacy of fungicide in combination with bioagents was greater compared to fungicide alone. The integration of P. fluorescens with apron or ridomil MZ as seed treatment significantly reduced the Phytophthora blight incidence and enhanced seed germination and grain yield.     

Efficacy of Pseudomonas fluorescens and Paecilomyces lilacinus against Meloidogyne graminicola infecting rice under system of rice intensification

The talc-based formulations of the plant growth promoting rhizobacterium, Pseudomonas fluorescens and egg parasitic fungi, Paecilomyces lilacinus, were evaluated as seed treatment, soil application and combination of both for the management of M. graminicola in fields of rice grown under system of rice intensification. Both the bioformulations significantly reduced the root invasion and soil populations of M. graminicola but P. fluorescens was most effective when applied as seed cum soil application and seed treatment alone. Effect of these treatments was comparable with the standard chemical carbofuran application. The introduced P. fluorescens survived significantly in rice roots when applied as seed cum soil application and seed application alone than as soil application. There was significant increase in phenol, peroxidase and chitinase accumulation in plants treated with P. fluorescens. Application of bioagents had positive influence on growth parameters such as plant height, root length, shoot weight, root weight and number of tillers per hill. Application of P. fluorescens as seed cum soil treatment resulted in higher grain yield, which was 20.6%–26.9% increase over control followed by P. fluorescens as seed treatment alone that increased grain yield of rice by 10.7%–11.2% than control. However, economic returns per investment was higher when P. fluorescens was applied as seed treatment alone (1:8.8–1:12.0 incremental cost benefit ratio) followed by the P. fluorescens as seed cum soil treatment (1:6.2–1:9.7 incremental cost benefit ratio).     

Laboratory evaluation of grain protectant efficacy of different foliar extracts against maize weevil – Sitophilus zeamais (Motschulsky)

The present study was aimed to evaluate six indigenous plant extracts for their ability to protect maize from infestation by maize weevil Sitophilus zeamais (M). The plant extracts were tested for its toxicity by fumigation and grain surface coating methods. Further, the effects of botanical treatments on F1 progeny, seed emergence and antifungal activities were studied. The results indicated that the extracts of Eicchornia crassipes controlled the test insect in grain surface coating method (100%, P = 0.05). Further, the same plant extract had significantly reduced the development of F1 progeny (97 ± 0.7%). In germination experiments, T + UEx samples showed normal seed germination of maize. However, in T + Ex samples, the seed germination depended on the grain protectant efficacy of the tested botanical extracts. Always higher percantage seed germination was recorded in those treatments where there was less infestation. Extracts of Carica papaya had significantly reduced the incidence of seed-borne fungus Aspergillus flavus (4 ± 2.4%) and did not affect the seed germination when compared to other treatments and untreated control.     

小麦赤霉病拮抗菌JB7的拮抗活性及鉴定

由禾谷镰刀菌(Fusarium graminearum, Fg)引起的赤霉病是限制小麦生产的主要病害之一。生物防治是一种高效且可持续的防治方法。【目的】从小麦种子内筛选具有抑制禾谷镰刀菌的菌株并对其生防潜力进行评估,为小麦赤霉病生防制剂的开发与利用提供菌种资源及理论支撑。【方法】采用平板对峙、孢子萌发法和无菌上清液抑菌试验筛选小麦种子内对禾谷镰刀菌具有拮抗活性的内生菌株;利用扫描电镜(scanning electron microscope, SEM)和共聚焦扫描电镜(confocal laser scanning microscope, CLSM)观察并分析无菌上清液对Fg的分生孢子形态、膜完整性以及胞内活性氧的影响;通过盆栽试验验证内生菌对小麦赤霉病的生防效果;应用二代Illumina HiSeq测序平台进行全基因组测序。【结果】从小麦种子中分离出一株高效抑制Fg生长的内生菌株JB7,其衰亡期无菌上清液对Fg孢子萌发抑制率高达85.23%。菌株JB7的无菌上清液使Fg孢子表面凹陷,破坏其细胞膜,造成核酸和蛋白质的渗漏,诱导Fg菌丝活性氧的累积,引起Fg菌丝可溶性蛋白和丙二醛含量的显著升高。该菌株具有分泌蛋白酶、纤维素酶、葡聚糖酶和产铁载体的能力。盆栽试验表明菌株JB7能显著降低小麦赤霉病的病情指数(P<0.05)。经全基因组学鉴定为甲基营养型芽孢杆菌(Bacillus methylotrophicus) JB7,该菌株基因组中含有12个抑菌功能的次级代谢产物合成基因簇。【结论】菌株JB7能抑制禾谷镰刀菌的生长,对小麦赤霉病有较强的防效,可作为生物防治小麦赤霉病的候选菌株。     

Prevalence and control of wheat (Triticum aestivum L.) seed borne fungi in farmer-saved seeds

High incidence of diseases associated with the use of seeds saved from previous harvests as desire of maintaining local varieties with special attributes is of increased concern in wheat industry worldwide. Prevalent of seed-borne fungi in farmer-saved seeds and seed dressing fungicides to prevent infection from seeds to seedlings was studied in Northern Tanzania. One hundred and thirty five untreated farmer-saved seed lots were collected randomly from farmers. Alternaria alternata, Bipolaris sorokiniana, Drechslera tritici, Fusarium graminearum, Fusarium moniliforme, Aspergillus flavus, Cladosporium sphaerospermum, Epicoccum purpurascens, Pyricularia oryzae and Penicillium corylophilum were fungi isolated in farmer-saved seeds. Mean seed infection was 29% causing average grain yield loss of 1.2 mt/ha^?1. Seed dressing with Metalaxy plus (Methyl carboxaitide), Mancozeb (Manganase-zinc salt) and Baytan (Chlorophenoxy ethanol) increased seed germination by 14, 13 and 17%, respectively, and grain yield by 28, 20 and 18%, respectively. Farmer-saved seeds were heavily infected by fungi with low grain yield performance.     

Bacterial wilt of tomato in Karnataka and its management by <Emphasis Type="Italic">Pseudomonas fluorescens</Emphasis>

Field surveys undertaken in major tomato growing districts of the Karnataka state, located in southern part of India, revealed a high incidence of bacterial wilt caused by Ralstonia solanacearum and it is one of the most destructive bacterial diseases of economically important crops. Across all the tomato cultivars under evaluation, the disease incidence in plants ranged from 9% to 39% whereas the incidence in seeds ranged from 4% to 18%. The effects of tomato seed treatments with Pseudomonas fluorescens in the control of bacterial wilt under greenhouse conditions revealed that the treatments protected plants against soil-borne infections of the bacterial wilt organism. Seed treatment with antagonistic P. fluorescens strain significantly improved the quality of seed germination and seedling vigour. The disease incidence was significantly reduced in plants raised from P. fluorescens treated seeds followed by challenge inoculation with R. solanacearum. Periodic field surveys for the incidence of bacterial wilt of tomato could be recommended to monitor the populations of the bacterial wilt pathogen. Workable measures are presented that could lead to the reduction of the prevalence of this serious disease in affected fields of the small farm-holders.     

凋落物物理阻隔对格氏栲种子萌发及胚根生长的影响

为了探讨凋落物物理阻隔对格氏栲天然林自然更新状况的影响,通过模拟野外凋落物覆盖,设置格氏栲种子上层覆盖厚0 cm(CK)、2 cm(D2)、4 cm(D4)、6 cm(D6)、8 cm(D8)及种子下层铺垫厚2 cm(U2)、4 cm(U4)凋落物等7个处理,分析凋落物覆盖方式及厚度对格氏栲种子萌发及胚根生长的影响。结果表明:(1)凋落物覆盖方式及厚度对种子萌发进程存在显著影响。CK萌发持续时间最长,上层覆盖处理(D)次之,下层铺垫处理(U)的种子起始萌发时间显著滞后。(2)CK种子萌发率最高,其次D6处理发芽速度较快,发芽整齐;U处理较D处理的发芽率及发芽势均显著降低,且萌发抑制率显著增加。(3)D处理的种子胚根生长速度快,胚根长度大于CK;U处理的种子胚根生长速度呈先慢后快趋势。可见,凋落物是影响格氏栲种子萌发及胚根生长的重要因素,主要通过阻碍种子与土壤接触而抑制萌发,影响格氏栲林更新。     

Effect of Pseudomonas fluorescens RB4 and Bacillus subtilis 189 on the phytoremediation potential of Catharanthus roseus (L.) in Cu and Pb-contaminated soils

The remediation of heavy metal-contaminated soils has become a critical issue due to toxic effects of these metals on living organisms. The current research was conducted to study the effect of Pseudomonas fluorescens RB4 and Bacillus subtilis 189 on the growth and phytoremediation potential of Catharanthus roseus in Cu- and Pb-contaminated soils. The bacterial strains exhibited significantly higher level of water-extractable Pb and Cu in Pb, Cu, and Cu+Pb-contaminated. The P. fluorescens RB4 inoculated plants, produced 102%, 48%, and 45% higher fresh weight (FW) in soils contaminated with Cu, Pb, and both elements, respectively, as compared to un-inoculated control plants. Similarly, B. subtilis 189 inoculated plants produced 108%, 43%, and 114% more FW in the presence of Cu, Pb, and both elements. The plants co-cultivated with both bacteria exhibited 121%, 102%, and 177% higher FW, in Cu, Pb, and both elements contaminated soils, as compared to respective un-inoculated control. Co-cultivation of P. fluorescens RB4, B. subtilis 189, and P. fluorescens RB4 + B. subtilis 189 resulted in higher accumulation of Cu and Pb in shoots of the C. roseus grown in contaminated soils as compared to un-inoculated control. Bacterial treatments also improved the translocation and metal bioconcentration factors. The growth and phytoextraction capability of C. roseus was improved by inoculation of P. fluorescens RB4 and B. subtilis 189.     

Biotypes of the seed beetle Callosobruchus maculatus have differing effects on the germination and growth of their legume hosts

• 1 Populations of the seed beetle Callosobruchus maculatus (F.) exhibit considerable differences in body size and larval behaviour. We examined whether such variation modifies the relationship between beetle infestation and host plant performance.
• 2 Larvae from African and Asian biotypes were reared in seeds of four hosts that represented an almost four‐fold variation in seed mass. We estimated mass lost to larval consumption, and compared germination rates and seedling growth between infested and control seeds.
• 3 In seeds bearing a single larva, the larger‐bodied, contest‐competing larvae of the Asian biotype caused a 38–47% greater reduction in seed mass compared with the smaller‐bodied, scramble‐competing larvae of the African biotype. The amount of seed mass lost per larva remained similar in seeds with one or two scramble‐competing larvae but decreased significantly in seeds bearing two contest‐competing Asian larvae.
• 4 Differences in larval consumption and behaviour produced striking differences in the frequency of germination. Germination of singly‐infested mung bean (i.e. the smallest host) was 71% for African‐infested seeds versus 11% for Asian‐infested seeds. In cowpea (i.e. the largest host), 76% of Asian‐infested seeds germinated, whereas the germination rate of African‐infested cowpeas (92%) was similar to that of uninfested seeds.
• 5 Effects of beetle origin persisted after germination. Seedlings derived from Asian‐infested seeds had greater cotyledon damage 7 days after germination, and displayed lower height and less biomass 15 days after germination. Cotyledon damage was a good predictor of seedling performance (i.e. better than seed mass consumed) 15 days after germination.
• 6 Previous studies have suggested that population differences in larval size and burrowing behaviour (‘centripetal tendency’) reflect adaptation to different‐sized seeds. The present study demonstrates that these differences in turn influence the impact of larval feeding on host viability. Strong biotypic variation makes it difficult to generalize about pest impacts at the level of pest species.

     

Effect of electrostatic spray parameters on the viability of two bacterial biocontrol agents and their deposition on blueberry flower stigmas

The fungus Monilinia vaccinii-corymbosi infects blueberry flowers via the stigma-style ovary pathway to cause mummy berry disease. Previous laboratory experiments documented considerable activity of stigma-applied biofungicides containing the bacteria Bacillus subtilis and, to a lesser extent, Pseudomonas fluorescens against flower infection by the pathogen. However, adequate and targeted delivery of the biocontrol agents to the stigmatic surfaces of open flowers in the field has remained problematic. Here we consider the application of the biofungicides Serenade AS (containing B. subtilis QST713) and BlightBan A506 (containing P. fluorescens A506) to blueberry flowers by air-assisted electrostatic spraying. In laboratory experiments with typical field-use rates, viability of B. subtilis and P. fluorescens was unaffected by different levels of induction-charging voltage (0–1.2 kV) and atomizing pressure (138–276 kPa) applied to an electrostatic spray-charging nozzle, showing that the bacteria in both formulations readily survived exposure to the intense electrical fields and near-sonic atomizing air shear encountered during electrostatic spraying. Electrostatically charged application significantly (P<0.0001) increased deposition of B. subtilis on the stigmatic surfaces of detached blueberry flower clusters by a factor of 4.5 compared with conventional hydraulic spraying; a similar comparison showed that population densities of P. fluorescens on the stigma were increased by a factor of 2.9, but this effect was not statistically significant (P=0.1487). For Serenade, the increased coverage and/or retention on the flower stigma, along with the excellent bacterial survival, portend well for electrostatic application for mummy berry disease control in the field.     

Increased Germination Rates of Baldcypress and Pondcypress Seed Following Treatments Affecting the Seed Coat

The effects of seed-coat removal, warm-water soaking, cold stratification, dry-cold storage and acid treatment on the germination of baldcypress (Taxodium distichum (L.) Rich.) and pondcypress (Taxodium distichum var. nutans (Ait.) Sweet) seed were studied. Removal of the hard seed coat resulted in a prompt and high percent germination, indicating that there was no physiological embryo dormancy. Cold stratification, warm-water soaking and acid treatment did not increase percent germination over the control after a 60-day germination period. They did promote the rate of germination however, having a significantly higher germination at 30 days. Dry-cold storage promoted the initial rate of germination, but resulted in a decreased percent germination below the control after 60 days. The above results suggest that moisture, and not cold, was the crucial component of stratification. The improved germination rate following acid treatment further suggests that any treatment which rendered the seed coat more permeable or softened it was more effective in increasing the rate of germination of baldcypress and pondcypress seed.     

Evaluation of Some Fungi and Bacteria for Potential Control of Safflower Rust

Some fungal and bacterial isolates applied as soil and seed treatments in greenhouse trials gave a good protection against Puccinia carthami Cda. on safflower, significantly reducing the infection from rust-infested seeds. Trichoderma viride and T. harzianum added alone and in combination as air-dried inoculum to the soil were the most effective fungal isolates. However T. viride at two higher rates reduced the rate of emergence of safflower seedlings. Among bacteria, Bacillus subtilis, B. cereus, isolates of Pseudomonas fluorescens and B. thuringiensis reduced rust infection when added both as soil drench and as seed treatment. Good results in the biocontrol of P. carthami were also obtained with some combination antagonist treatments such as T. viride + B. cereus, T. viride + P. fluorescens (16), T. viride + T. harzianum + B. cereus and T. viride +, T. harzianum + P. fluorescens (16). Fungal isolates added as soil treatment increased seedling length, whereas no similar effects were observed when these isolates were applied as seed treatment.     

Monitoring Population Size,Activity, and Distribution of gfp-luxAB-Tagged Pseudomonas fluorescens SBW25 during Colonization of Wheat

Increasingly, focus has been directed towards the use of microorganisms as biological control agents to combat fungal disease, as an alternative to chemical fungicides. Pseudomonas fluorescens SBW25 is one bacterial strain that has been demonstrated to promote plant growth by biocontrol of pathogenic fungi. To understand the mode of action of this bacterium, information regarding its localization and metabolic activity on plants is important. In this study, a gfp/luxAB-tagged derivative of P. fluorescens SBW25, expressing the green fluorescent protein (GFP) and bacterial luciferase, was monitored during colonization of wheat starting from seed inoculation. Since bacterial luciferase is dependent on cellular energy reserves for phenotypic expression, metabolically active cells were detected using this marker. In contrast, the stable GFP fluorescence phenotype was used to detect the cells independently of their metabolic status. The combination of these two markers enabled P. fluorescens SBW25 cells to be monitored on wheat plants to determine their specific location and metabolic activity. Studies on homogenized wheat plant parts demonstrated that the seed was the preferred location of P. fluorescens SBW25 during the 65-day time period studied, but the leaves and roots were also colonized. Interestingly, the bacteria were also found to be metabolically active on all plant parts examined. In situ localization of P. fluorescens SBW25 using a combination of different microscopic techniques confirmed the preference for the cells to colonize specific regions of the seed. We speculate that the colonization pattern of P. fluorescens SBW25 can be linked to the mechanism of protection of plants from fungal infection.