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PubMed2001

Production of the biocontrol agent Pantoea agglomerans strain CPA-2 using commercial products and by-products.

The aim of this paper was to find the nitrogen and carbon sources that provide maximum biomass production of strain CPA-2 of the biocontrol agent Pantoea agglomerans and minimum cost of media, whilst maintaining biocontrol efficacy. To reduce the cost of media, commercial products and by-products were tested. P. agglomerans can be produced using a combination of nitrogen sources such as yeast extract (5 g l(-1)) and dry beer yeast (10 g l(-1)) with inexpensive carbohydrates such as sucrose (10 g l(-1)) and molasses (20 g l(-1)), respectively, maintaining the efficacy of the biocontrol agent against Penicillium digitatum and P. italicum on oranges. The results obtained in this study could be used to provide a reliable basis for a scale-up of this fermentation process to an industrial level.

Biomass↗
PubMed1996

Sarcocystis singaporensis: studies on host specificity, pathogenicity, and potential use as a biocontrol agent of wild rats.

Host specificity and pathogenicity of Sarcocystis singaporensis were investigated as a prerequisite to a subsequent application of the parasite as a biocontrol agent of wild rats in Egypt. After inoculation of 7 snake species comprising the families Elapidae, Viperidae, Colubridae, and Boidae with sarcocysts, sporocyst development was only observed in a reticulated python. Among amphibians, reptiles, and rodents that orally received various sporocyst doses in the laboratory, 2 x 10(4) sporocysts or more were lethal to roof rats Rattus rattus frugivorous, brown rats Rattus norvegicus, and bandicoot rats Nesokia indica. Sarcocysts developed in Rattus spp. and Nile grass rats Arvicanthis niloticus. Subsequently, the pathogenicity of S. singaporensis was tested under natural control situations offering bait pellets containing high amounts of sporocysts to a free-living population of roof rats, which was monitored by indirect census baiting commonly used in rodenticide evaluation. Ten days after consumption of the bait pellets, the infected population collapsed, leading to a control success of 73%. A negative control population, which received a placebo, remained stable. These data demonstrate for the first time that S. singaporensis can be used as a biocontrol agent of wild rats. However, an immunization experiment with roof rats in the laboratory showed that these are capable of mounting a rapid specific immune response resulting in survival of acute sarcocystosis.

Amphibians↗
PubMed2007

Expressed sequence tags-based identification of genes in the biocontrol agent Chaetomium cupreum.

Chaetomium cupreum has a potential as biocontrol agent against a range of plant pathogens on the basis of production of antifungal metabolites, mycoparasitism, competition for space and nutrients, or various combinations of these. To explore genes expressed in C. cupreum, a cDNA library was constructed from mycelium and 3,066 expressed sequence tags (ESTs) were generated. Clusters analysis enabled the identification of 1,471 unigenes with 392 contigs and 1,079 singleton sequences. Putative functions were assigned to 874 unigenes that exhibited strong similarity to genes/ESTs in public databases putatively containing genes involved in cellular component, molecular function, and biological process. Other 597 ESTs representing novel genes showed no significant similarity to public database resource of NCBI. A proportion of genes was identified related to degradation of pathogen cell wall, antifungal metabolite production, as was estimated in the biocontrol fungus. The paper described is a first step towards the knowledge of the C. cupreum genome. The results present the useful application of EST analysis on C. cupreum and provide a preliminary indication of gene expression putatively involved in biocontrol.

Chaetomium↗
PubMed1995

Liquid-culture pH, temperature, and carbon (not nitrogen) source regulate phenazine productivity of the take-all biocontrol agent Pseudomonas fluorescens 2-79.

Strain 2-79 is a biocontrol agent against take-all, an important disease of wheat caused by Gaeumannomyces graminis var. tritici. In the rhizosphere, it produces the antibiotic phenazine 1-carboxylic acid (PCA) as the primary means of disease suppression. One barrier to commercial use of phenazine-producing pseudomonads, like strain 2-79, is the lack of liquid-culture technology for mass production. For instance, there is little published research concerning the impact of liquid-culture secondary metabolism on the biocontrol qualities of the cell harvest, i.e., efficacy, phytotoxicity, and storage survival. Yet it is important to know whether the fermentation process should be designed to enhance or eliminate secondary metabolite accumulation. To enable future exploration of this issue, we identified liquid-culture parameters that could be manipulated to control the phenazine productivity of strain 2-79. Our results indicated that PCA accumulation was very sensitive to the culture pH and temperature. It was possible to produce large cell populations with either high or low phenazine productivity by choosing to control culture pH at 7 and 8 respectively. Although high cell accumulations were achieved over the broad 25-34 degrees C range studied, high, moderate, or low PCA productivities were observed at 25-27 degrees C, 29-32.5 degrees C, or 34 degrees C respectively. When pH was controlled at 7, specific PCA productions at 25 degrees C could be modulated by the choice of carbon source supplied. PCA accumulation per unit biomass reached 0.31 g/g on glucose, 0.16 g/g on glycerol and xylose, and only 0.09 g/g on fructose.(ABSTRACT TRUNCATED AT 250 WORDS)

Carbon↗
PubMed2002

Water activity, temperature, and pH effects on growth of the biocontrol agent Pantoea agglomerans CPA-2.

The growth response of the biocontrol agent Pantoea agglomerans to changes in water activity (a(w)), temperature, and pH was determined in vitro in nutrient yeast extract-sucrose medium. The minimum temperature at which P. agglomerans was able to grow was 267-272 kelvins (-6 to -1 degrees C), and growth of P. agglomerans did not change at varying pH levels (4.5-8.6). The minimum a(w) for growth was 0.96 in media modified with glycerol and 0.95 in media modified with NaCl or glucose. Solute used to reduce water activity had a great influence on bacterial growth, especially at unfavourable conditions (e.g., low pH or temperature). NaCl stimulated bacterial growth under optimum temperatures but inhibited it under unfavourable pH conditions (4.5 or 8.6). In contrast, the presence of glucose in the medium allowed P. agglomerans to grow over a broad range of temperature (3-42 degrees C) or pH (5-8.6) regimes. This study has defined the range of environmental conditions (a(w), pH, and temperature) over which the bacteria may be developed for biological control of postharvest diseases.

Biomass↗
PubMed2002

Antibiotic production by bacterial biocontrol agents.

Interest in biological control of plant pathogens has been stimulated in recent years by trends in agriculture towards greater sustainability and public concern about the use of hazardous pesticides. There is now unequivocal evidence that antibiotics play a key role in the suppression of various soilborne plant pathogens by antagonistic microorganisms. The significance of antibiotics in biocontrol, and more generally in microbial interactions, often has been questioned because of the indirect nature of the supporting evidence and the perceived constraints to antibiotic production in rhizosphere environments. Reporter gene systems and bio-analytical techniques have clearly demonstrated that antibiotics are produced in the spermosphere and rhizosphere of a variety of host plants. Several abiotic factors such as oxygen, temperature, specific carbon and nitrogen sources, and microelements have been identified to influence antibiotic production by bacteria biocontrol agents. Among the biotic factors that may play a determinative role in antibiotic production are the plant host, the pathogen, the indigenous microflora, and the cell density of the producing strain. This review presents recent advances in our understanding of antibiotic production by bacterial biocontrol agents and their role in microbial interactions.

Anti-Bacterial Agents↗
PubMed2005

Survival of the biocontrol agents Brevibacillus brevis ZJY-1 and Bacillus subtilis ZJY-116 on the spikes of barley in the field.

Fusarium head blight (FHB) caused by Fusarium graminearum is a devastating disease that results in extensive yield losses to wheat and barley. A green fluorescent protein (GFP) expressing plasmid pRP22-GFP was constructed for monitoring the colonization of two biocontrol agents, Brevibacillus brevis ZJY-1 and Bacillus subtilis ZJY-116, on the spikes of barley and their effect on suppression of FHB. Survival and colonization of the Brevibacillus brevis ZJY-1 and Bacillus subtilis ZJY-116 strains on spikes of barley were observed by tracking the bacterial transformants with GFP expression. Our field study revealed that plasmid pRP22-GFP was stably maintained in the bacterial strains without selective pressure. The retrieved GFP-tagged strains showed that the bacterial population fluctuation accorded with that of the rain events. Furthermore, both biocontrol strains gave significant protection against FHB on spikes of barley in fields. The greater suppression of barley FHB disease was resulted from the treatment of barley spikes with biocontrol agents before inoculation with F. graminearum.

Bacillus↗
PubMed2005

Effect of temperature and host plant leaf morphology on the efficacy of two entomopathogenic biocontrol agents of Thrips palmi (Thysanoptera: Thripidae).

The efficacy of two entomopathogenic biocontrol agents, Steinernema feltiae (Filipjev) and Verticillium lecanii (Zimmerman) Viégas (reclassified now as Lecanicillium muscarium (Petch) Zare & Gams), against Thrips palmi Karny was investigated. Assessments of the effect of temperature on the efficacy of S. feltiae indicated that higher mortality of T. palmi was recorded at 20 degrees C compared to either 15 or 25 degrees C, whereas significantly higher T. palmi mortality followed application of L. muscarium at 25 degrees C. Testing the control agents efficacy on three host plants; chrysanthemum, sweet pepper and cucumber, under constant temperature and high humidity conditions produced no significant difference in the level of T. palmi larval mortality on each host plant. Incorporating the chemical insecticide imidacloprid with both biological agents in a combined control strategy increased T. palmi juvenile mortality. The potential role of S. feltiae and L. muscarium within integrated pest management programmes for the control of T. palmi is discussed.

Animals↗
PubMed2005

Accumulation of the compatible solutes, glycine-betaine and ectoine, in osmotic stress adaptation and heat shock cross-protection in the biocontrol agent Pantoea agglomerans CPA-2.

AIMS: The effect of modifying the water activity (a(w)) of Pantoea agglomerans growth medium with the ionic solute NaCl on water stress resistance, heat-shock survival and intracellular accumulation of the compatible solutes glycine-betaine and ectoine were determined. METHODS AND RESULTS: The bacterium was cultured in an unmodified liquid medium or that modified with NaCl to 0.98 and 0.97 a(w), and viability of cells evaluated on a 0.96 a(w)-modified solid media to check water stress tolerance. Cells grown under ionic stress had better water stress tolerance than control cells. These cells also had cross-protection to heat stress (30 min, 45 degrees C). The modified cells accumulated substantial amounts of the compatible solutes glycine-betaine and ectoine in contrast to the control cells, which contained little or none of these two compounds. CONCLUSIONS: Improvement in osmotic and thermal tolerance of cells of the biocontrol agent P. agglomerans by modifying growth media with the ionic solute NaCl was achieved. The compatible solutes glycine-betaine and ectoine play a critical role in environmental stress tolerance improvement. SIGNIFICANCE AND IMPACT OF THE STUDY: This approach provides a method for improving the physiological quality of inocula and could have implications for formulation and shelf-life of biocontrol agents.

Adaptation, Physiological↗
PubMed2003

Seasonal variation in the effectiveness of the leaf-feeding beetle Zygogramma bicolorata (Coleoptera: Chrysomelidae) and stem-galling moth Epiblema strenuana (Lepidoptera: Tortricidae) as biocontrol agents on the weed Parthenium hysterophorus (Asteraceae).

Variation in the effectiveness of biocontrol agents on the weed Parthenium hysterophorus L. was evaluated at two properties (Mount Panorama and Plain Creek) in Queensland, Australia for four years (1996-2000) using a pesticide exclusion experiment. At Mount Panorama, higher levels of defoliation by the leaf-feeding beetle Zygogramma bicolorata Pallister and galling by the moth Epiblema strenuana Walker in 1996-97 coincided with an above average summer rainfall, but in the following three years with below average summer rainfall the defoliation and galling levels were significantly lower. Biocontrol had significant negative impact on the weed only in 1996-97 with no major impact in the following three years. At Plain Creek, galling by E. strenuana was evident in all the four years, but varied significantly between years due to non-synchrony between P. hysterophorus germination and E. strenuana emergence. At Plain Creek biocontrol had limited impact on the weed in 1996-97 and 1997-98, with no significant impact in the following two years. Over the 4-year period, defoliation and galling resulted in 70% reduction in the soil seed bank at Mount Panorama, but the reduction in the soil seed bank at Plain Creek due to galling was not significant. Effectiveness of Z. bicolorata and E. strenuana was dependent on weather conditions and as a result had only limited impact on the weed in three out of four years.

Animals↗
PubMed2006

Toxicity assessment of metabolites of fungal biocontrol agents using two different (Artemia salina and Daphnia magna) invertebrate bioassays.

Fungal biocontrol agents (BCAs) have been marketed for control of crop pests, weeds, and diseases. However, BCAs may produce toxic metabolites, whose presence in the formulated products, in the crops and in the environment should be considered along with the associated risk. Two invertebrate models, viz. Artemia salina and Daphnia magna were used to assess the acute toxicity of seven BCA metabolites, characterized by different chemical nature and mode of action, namely alamethicin (ALA), paracelsin (PCS), antiamoebin (AAM), gliotoxin (GTX), destruxin A (DA), oosporein (OOS), and elsinochrome A (EA). The two invertebrates were very sensitive to all the metabolites examined, except OOS. The LC50s after 24 and 36 h exposures showed the following toxicity ranks: A. salina, DA > ALA > EA > GTX > AAM > PCS (LC50s ranging from 9.78 to 40.84 microg/ml at 24 h and from 2.92 to 18.56 microg/ml at 36 h); D. magna, DA > GTX = EA > ALA > PCS > AAM (LC50s ranging from 0.20 to 24.41 microg/ml at 24h and from 0.16 to 11.98 microg/ml at 36 h). LC50 of OOS to D. magna increased dramatically in 36 h exposure, compared to 24 h exposures (5.84 and 68.40 microg/ml, respectively). A. salina and D. magna proved to be suitable models for rapid and inexpensive screening of toxicity of BCAs at an early stage of product development.

Animals↗
PubMed2006

The efficacy of two entomopathogenic biocontrol agents against adult Thrips palmi (Thysanoptera:Thripidae).

The efficacy of the two entomopathogenic biocontrol agents, Steinernema feltiae (Filipjev) and Lecanicillium muscarium (Petch), against juvenile Thrips palmi Karny in the laboratory situation is well documented. This study investigated the extrapolation of this knowledge to the control of adult stages. S. feltiae caused significantly higher mortality to juvenile T. palmi than to adult stages, whereas L. muscarium had a more significant impact on adult stages. The potential to develop an IPM strategy to control T. palmi utilising the two entomopathogens is discussed.

Age Factors↗
PubMed2006

[Compatibility of two fungal biocontrol agents conidia with commercial chemical acaricides].

In this paper, the biological compatibility of fungal biocontrol agents Beauveria bassiana and Paecilomyces fumosoroseus conidia with 10 commercial chemical acaricides were assayed, based on the conidial germination rates in nutritional liquid and on SDAY plate. The results showed that nutritional liquid was more available than SDAY plate in the assay. So far as the 24 h conidial viability concerned, there were significant differences among the test acaricides with the concentrations of recommended for field spray and 5 and 10 fold dilutions, as well as between the two fungal agents. Since acaricides azocyclotin, liuyangmycin, dicofol and avermectin had strong inhibitory effects on the 24 h germination rate of both fungal agents conidia, their combined application with fungal agents was unsuitable for mite control. However, the combined application of pyridaben, propargite, chlorpyrifos, hexythiazox or amitraz with either B. bassiana or P. fumosoroseus was practical, because of their short-term compatibility. When the mixtures of oil-based B. bassiana formulation with the three concentrations of pyridaben, propargite and chlorpyrifos were stored at 4 degrees C or at ambient temperature for 12 months, none of the three chemicals was considered to be good enough for a combined formulation due to the great variability in long-term compatibility. Nevertheless, chlorpyrifos exhibited an encouraging long-term compatibility with B. bassiana, because its low concentration in the fungal formulation did not affect the conidial viability during a 6.5-month period of storage at ambient temperature. When stored at 4 degrees C in dark, the B. bassiana formulation containing low or medium concentration of chlorpyrifos retained the conidial viability of > 90% for up to 12 months.

Animals↗
PubMed2006

GFP technology for the study of biocontrol agents in tritrophic interactions: a case study with Pseudozyma flocculosa.

GFP technology was applied to the biocontrol agent (BCA) Pseudozyma flocculosa to study its development and interactions at the tritrophic level plant-powdery mildew-BCA. Transformation experiments with GFP led to the production of a strongly fluorescent strain, Act-4, that displayed biocontrol traits typical of P. flocculosa WT. Following inundative applications, growth of P. flocculosa Act-4 was closely and almost exclusively associated with the colonies of the pathogen regardless of the powdery mildew species or the host plant tested. Development of P. flocculosa Act-4 on control leaves alone was extremely limited 24 h after its application and was typical of the epiphytic growth characterizing this type of yeast-like fungus. Based on the strong correlation between the colonization pattern of the different powdery mildew species tested and the presence of P. flocculosa Act-4, as determined by its fluorescence, it seems that growth of the BCA is dependant on the presence of powdery mildews. These results demonstrate that the GFP technology can be used to study plant-pathogen-BCA interactions and fulfill a wide array of purposes ranging from fundamental observations of the biocontrol behavior of a BCA to very applied ones serving some of the requirements for the registration of BCA's such as defining their environmental fate.

Ascomycota↗
PubMed2005

Purification and characterization of an extracellular chitinase from the antifungal biocontrol agent Streptomyces halstedii.

An extracellular chitinase from Streptomyces halstedii AJ-7, a broad spectrum antifungal biocontrol agent, was characterized and purified. The apparent molecular weight of the purified protein was 55 kDa, Km value and Vmax of the protein for colloidal chitin were 3.2 mg ml(-1) and 118 micromol h(-1), respectively. The growth and chitinase activity of S. halstedii AJ-7 were enhanced by adding of 0.1% killed mycelium of Fusarium oxysporium in a medium containing 0.2% colloidal chitin.

Antifungal Agents↗
PubMed2002

Cuticular hydrocarbons of the flea beetles, Aphthona lacertosa and Aphthona nigriscutis, biocontrol agents for leafy spurge (Euphorbia esula).

The adult beetles Aphthona lacertosa and Aphthona nigriscutis, used as biocontrol agents for leafy spurge, had a complex mixture of hydrocarbons on their cuticular surface consisting of alkanes, methylalkanes, alkenes and alkadienes as determined by gas chromatography-mass spectrometry. A trace amount of wax esters were present. In both species, the hydrocarbons were the major cuticular lipid class and the gas chromatographic profiles of the total hydrocarbons were similar. However, the profiles for the saturated hydrocarbon fraction were distinct for each species. Alkanes (n-alkanes and methyl-branched alkanes), alkenes and alkadienes comprised 26, 44 and 30%, respectively, for A. lacertosa, and 48, 26 and 26%, respectively, for A. nigriscutis, of the total hydrocarbons. The major methyl-branched hydrocarbons were 2-methylalkanes: 2-methyloctacosane and 2-methyltriacontane. The major monoene was hentriacontene and the major diene was tritriacontadiene. The species were unique in that a number of di- and trimethyl-branched alkanes were present in minor quantities in which the first methyl branch was on carbon 2 or 3. Examples of structures were 2,10-, 2,12-, 2,6-, 2,4- and 3,7-dimethylalkanes. 2,10,12-Trimethylalkanes and a 2,10,12,24-tetramethylalkane with one methylene between adjacent methyl branch points also were identified. The adjacent methyl branch points of the 2,4- and 2,10,12- and 2,10,12,24-methyl-branched alkanes appeared to cause additional fragmentations in the mass spectra. Dimethylalkanes with an odd number of carbons in the backbone of the molecule were identified as 2,23-dimethylnonacosane and 2,25-dimethylhentriacontane; their mass spectra also corresponded to mass spectra expected for a 2,6 branching sequence. However, a 2,6 branching sequence is not biosynthetically feasible because such a structure has a straight-chain tail with an odd number of carbon atoms beyond the last methyl branch point. The 2,23 and 2,25 branching sequences could be synthesized starting with a primer derived from the amino acid leucine which would account for both the even number of carbons between the branch points and an even number of carbons beyond the last methyl branch point.

Alkanes↗
PubMed2002

Cloning and analysis of CoEXG1, a secreted 1,3-beta-glucanase of the yeast biocontrol agent Candida oleophila.

Lytic enzymes may have a role in the biological control of fungi. The yeast biocontrol agent, Candida oleophila, is an excellent subject to research this matter. In the present study, CoEXG1, which encodes for a secreted 1,3-beta-glucanase, is the first gene to be cloned from C. oleophila. It was isolated from a partial genomic library and analysed. Its open reading frame and putative promoter were expressed in baker's yeast, Saccharomyces cerevisiae. The reading frame, expressed under the inducible GAL1 promoter, caused an increased secretion of beta-glucanase, and the putative promoter region activated the lacZ reporter gene, to which it was fused. Sequencing analysis revealed that CoEXG1 carries the signature pattern of the 5 glycohydrolases family and has a putative secretion leader, as well as a high degree of identity to yeast 1,3-beta-glucanases. The GenBank Accession No. of CoEXG1 is AF393806.

Amino Acid Sequence↗