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1.
It was shown previously that spores and vegetative cells of Bacillus sphaericus (Bf) and Bacillus thuringiensis israelensis (Bti) are very sensitive to osmotic variations. Since spore osmotolerance has been associated with their SASP (small acid soluble spore proteins) content coded by ssp genes, hybridization assays were performed with sspE and sspA genes from B. subtilis as probes and showed that Bti and Bf strains could lack an sspE-like gene. The B. subtilis sspE gene was then introduced into Bti 4Q2 strain; spores were obtained and showed a 65 to 650 times higher level of osmotolerance to NaCl, without affecting other important properties: hypoosmotic resistance in vegetative cells, spore UV resistance, and larvicidal activity against diptera larvae. 相似文献
2.
Entomopathogenic bacilli B. thuringiensis israelensis (Bti) and B. sphaericus (Bf) exhibit low survival on field application. It was previously shown that their spores are very sensitive to different
stress effectors (heat, UV light) and especially to osmotic variations. Since SASP (Small, Acid-Soluble Spore Proteins), α/β
and γ type, are involved in spore tolerance to heat, UV light, peroxide, and salt, they were analyzed in Bti and Bf. The molecular
weight, migration pattern and amino acid composition of different SASP were determined and compared with other bacilli, in
particular to B. subtilis. A relation between spore osmotolerance, SASP content, and amino acid composition was shown. In addition, the absence of
γ SASP in Bti and Bf is discussed.
Received: 3 September 1997 / Accepted: 15 October 1997 相似文献
3.
The mosquitocidal bacterium Bacillus thuringiensis subsp. israelensis (Bti) produces four major endotoxin proteins, Cry4A, Cry4B, Cry11A, and Cyt1A, and has toxicity in the range of many synthetic chemical insecticides. Cry11B, which occurs naturally in B. thuringiensis subsp. jegathesan, is a close relative of Cry11A, but is approximately 10-fold as toxic to Culex quinquefasciatus. To determine whether the addition of Cry11B to Bti would improve its toxicity, we produced this protein in Bti. High levels of Cry11B synthesis were obtained by expression of the cry11B gene under the control of cyt1A promoters and the STAB-SD sequence. This construct was cloned into the shuttle vector pHT3101, yielding the derivative plasmid pPFT11Bs, which was then transformed by electroporation into acrystalliferous (4Q7) and crystalliferous (IPS-82) strains of Bti. Synthesis of Cry11B in Bti 4Q7 produced crystals approximately 50% larger than those produced with its natural promoters without STAB-SD. However, less Cry11B was produced per unit culture medium with this construct than with the wild-type construct, apparently because the latter construct produced more cells per unit medium. Nevertheless, the Bti IPS-82 strain that produced Cry11B with pPFT11Bs was twice as toxic as the parental IPS-82 strain (LC(50) = 1.4 ng/ml versus 3.3 ng/ml, respectively) to fourth instars of C. quinquefasciatus. Against fourth instars of Aedes aegypti, no statistically significant difference between parental Bti IPS-82 (LC(50) = 4.7 ng/ml) and the Bti IPS-82 recombinant producing Cry11B (LC(50) = 3.5 ng/ml) was found in toxicity. 相似文献
4.
Vector control can be an effective strategy to interrupt disease transmission and biolarvicides based on the entomopathogenic bacteria Bacillus sphaericus, and Bacillus thuringiensis serovar israelensis (Bti) have been successfully used to control species of public health relevance from the genera Aedes, Culex, Anopheles and Simulium. The most important feature of these agents is their ability to produce insecticidal proteins with selective action on the larval midgut. These protoxins are produced as crystals that, once ingested by larvae, are processed into active toxins, interact with receptors in the midgut epithelium and trigger cytopathological effects leading to larval death. B. sphaericus and Bti toxins share the initial steps of the mode of action; however, they interact with different midgut molecules. B. sphaericus presents a single larvicidal factor, the binary (Bin) toxin, whose action relies on the binding to one class of midgut receptors, while Bti crystals contain four protoxins (Cry4Aa, Cry4Ba, Cry11Aa and Cyt1Aa), which display interactions with multiple midgut receptors. The mode of action of B. sphaericus displays a greater potential for resistance selection, compared to Bti, and, to date, there is no record of insect resistance to the latter, contrarily to B. sphaericus. The set of mosquitocidal toxins and their interaction with midgut target sites are described in this review, as well as the implications for the potential to select resistance amongst exposed populations. These biolarvicides have specific mode of action that rely on unique interactions and make them the most selective agents to control Diptera insects actually available. 相似文献
5.
Laboratory studies were conducted to determine the sublethal effects of exposure to selected larvicides on the critical swimming speed (Ucrit) of crimson-spotted rainbowfish, Melanotaenia duboulayi (Castlenau). This native fish is common throughout southeastern Queensland, and it is increasingly being distributed as a biological control agent of mosquitoes. The selected larvicides included, two organophosphate (OP) compounds (temephos and pirimiphos-methyl), two microbial larvicides (Bacillus thuringiensis spp. israelensis [Bti] de Barjac and Bacillus sphaericus [Bs] Neide), and an insect growth regulator (IGR) (s-methoprene). Exposure to the OP temephos at 10 times the effective field concentration (EFC; 0.33 mg/liter), and OP pirimiphos-methyl at the EFC (0.50 mg/liter), resulted in a significant reduction in the Ucrit of M. duboulayi under controlled conditions. Conversely, exposure to the microbial (Bti and Bs) and IGR (s-methoprene) larvicides at 10 times the EFC had no effect on the Ucrit of M. duboulayi. Accordingly, these products are suitable for integrated pest management programs in Australia. 相似文献
6.
Aedes albopictus transmits several arboviral infections. In the absence of vaccines, control of mosquito populations is the only strategy to prevent vector-borne diseases. As part of the search for novel, biological and environmentally friendly strategies for vector control, the isolation of new bacterial species with mosquitocidal activity represents a promising approach. However, new bacterial isolates may be difficult to grow and genetically manipulate. To overcome these limits, here we set up a system allowing the expression of mosquitocidal bacterial toxins in the well-known genetic background of Bacillus subtilis. As a proof of this concept, the ability of B. subtilis to express individual or combinations of toxins of Bacillus thuringiensis israelensis (Bti) was studied. Different expression systems in which toxin gene expression was driven by IPTG-inducible, auto-inducible or toxin gene-specific promoters were developed. The larvicidal activity of the resulting B. subtilis strains against second-instar Ae. albopictus larvae allowed studying the activity of individual toxins or the synergistic interaction among Cry and Cyt toxins. The expression systems here presented lay the foundation for a better improved system to be used in the future to characterize the larvicidal activity of toxin genes from new environmental isolates. 相似文献
7.
The Upper Rhine Valley, a “hotspot of biodiversity” in Germany, has been treated with the biocide Bacillus thuringiensis var. israelensis (Bti) for mosquito control for decades. Previous studies discovered Bti nontarget effects in terms of severe chironomid abundance reductions. In this study, we investigated the impact of Bti on species level and addressed the community composition of the nontarget family Chironomidae by use of community metabarcoding. Chironomid emergence data were collected in three mosquito‐control relevant wetland types in the Upper Rhine Valley. For all three sites the chironomid species composition, based on operational taxonomic units (OTUs), was different to varying degrees in the Bti‐treated samples versus control samples, ranging from a significant 63% OTU reduction to an OTU replacement. We assumed that predatory chironomids are less prone to Bti than filter feeders, as the latter feed on floating particles leading to direct ingestion of Bti. However, a comparable percentage of predators and filter feeders (63% and 65%, respectively) was reduced in the Bti samples, suggesting that the feeding strategy is not the main driver for Bti sensitivity in chironomids. Finally, our data was compared to a three‐year‐old data set, indicating possible chironomid community recovery due to species recolonization a few years after the last Bti application. Considering the currently discussed worldwide insect decline we recommend a rethinking of the usage of the biocide Bti, and to prevent its ongoing application especially in nature protection reserves to enhance ecological resilience and to prevent boosting the current biodiversity loss. 相似文献
8.
Aims: To investigate mechanisms of osmotic tolerance in Burkholderia cenocepacia, a member of the B. cepacia complex (Bcc) of closely related strains, which is of clinical as well as environmental importance. Methods and Results: We employed NMR‐based metabolic profiling (metabolomics) to elucidate the metabolic consequences of high osmotic stress for five isolates of B. cenocepacia. The strains differed significantly in their levels of osmotic stress tolerance, and we identified three different sets of metabolic responses with the strains least impacted by osmotic stress exhibiting higher levels of the osmo‐protective metabolites glycine‐betaine and/or trehalose. Strains either increased concentrations or had constitutively high levels of these metabolites. Conclusions: Even within the small set of B. cenocepacia isolates, there was a surprising degree of variability in the metabolic responses to osmotic stress. Significance and impact of the study: The metabolic responses, and hence osmotic stress tolerance, vary between different B. cenocepacia isolates. This study provides a first look into the potentially highly diverse physiology of closely related strains of one species of the Bcc and illustrates that physiological or clinically relevant phenotypes are unlikely to be inferable from genetic relatedness within this species group. 相似文献
9.
Two insecticidal bacteria are used as larvicides to control larvae of nuisance and vector mosquitoes in many countries, Bacillus thuringiensis ssp. israelensis and B. sphaericus. Field studies show both are effective, but serious resistance, as high as 50 000‐fold, has evolved where B. sphaericus is used against Culex mosquitoes. To improve efficacy and deal with even greater potential problems of resistance, we previously developed several recombinant larvicidal bacteria that combine the best mosquitocidal proteins of these bacteria. In the present study, we report laboratory selection studies using our best recombinant strain against larvae of Culex quinquefasciatus. This recombinant, Bti/BsBin, is a strain of B. thuringiensis ssp. israelensis engineered to produce a large amount of the B. sphaericus binary (Bin) toxin, which makes it more than 10‐fold as mosquitocidal as the its parental strains. Here we show that larvae exposed to Bti/BsBin failed to develop significant resistance after 30 successive generations of heavy selection pressure. The highest level of resistance obtained at the LC 95 level was 5.2‐fold, but declined to less than two‐fold at the 35th generation. Testing the selected populations against B. sphaericus alone showed resistance to Bin evolved, but was masked by combination with B. thuringiensis ssp. israelensis. These results suggest that recombinant bacterial strains have improved mosquito and vector management properties compared with the wild‐type strains used in current commercial formulations, and should prove useful in controlling important human diseases such as malaria and filariasis on a long‐term basis, even when used intensively under field conditions. 相似文献
10.
Bacillus thuringiensis (Bt) Berliner is a promising agent for microbial control of agriculturally and medically important insects. This study aimed
at searching for Bt strains encoding Cry proteins that act more efficiently against fall armyworm. Thirty Bt strains were
isolated from soil samples in Pernambuco State and evaluated through bioassays. Among these, strain I4A7 was the most efficient
against the fall armyworm, Spodoptera frugiperda (J. E. Smith, 1797) (Lepidoptera: Noctuidae), and thus it was characterized by biochemical sodium dodecyl sulfate–polyacrylamide
gel electrophoresis (SDS–PAGE) and molecular (polymerase chain reaction (PCR) and sequencing reaction) methods. The protein
pattern of this strain on a SDS–PAGE was similar to that of B. thuringiensis
israelensis (Bti). Moreover, I4A7 cry DNA sequence showed high identity (99–100%) to genes cry4Aa, 4Ba, 10Aa, 11Aa, cyt1Aa and cyt2B from Bti. The toxicity of the newly isolated Bti-like strain upon S. frugiperda should be considered as this strain might be used in combination with other Bt strains, such as B. thuringiensis var. kurstaki (Btk).
Handling Editor: Helen Roy. 相似文献
11.
Novel strains of rhizobacteria, Pseudomonas fluorescens Pf 9A‐14, Pseudomonas sp. Psp. 8D‐45 and Bacillus subtilis Bs 8B‐1, showed broad‐spectrum antagonistic activity and provided suppression of Pythium damping‐off and root rot of cucumber. Their biocontrol potential was further investigated for suppression of additional seedling diseases of cucumber ( Phytophthora capsici) and radish ( Rhizoctonia solani). Bacterial strains were also characterised for production of antibiotics, metabolites, volatiles, phytohormones and lytic enzymes. Seed and pre‐plant applications of all three antagonistic bacteria as cell suspension and talc or irradiated peat formulations to the infested potting mix provided overall high level of suppression of Phytophthora damping‐off and root rot of cucumber (66–85% healthy seedlings) and relatively low level of suppression of Rhizoctonia damping‐off of radish (18–38% healthy seedlings). Bacterial treatments also resulted in higher plant fresh masses. Seed coating with irradiated peat formulation of a mixture of three bacteria resulted in superior control of Phytophthora damping‐off and root rot of cucumber and much higher plant fresh masses. The presence of genes for biosynthesis of phenazine‐1‐carboxylic acid, 2,4‐diacetylphloroglucinol, pyrrolnitrin and pyoluteorin was confirmed in Pseudomonas strains, and that of fengycin, bacillomycin, bacilysin, surfactin and iturin A in B. subtilis Bs 8B‐1. All three strains produced HCN, salicylic acid, indole‐3‐acetic acid, protease and β‐1,3‐glucanase. Both Pseudomonas strains produced siderophores and only P. fluorescens Pf 9A‐14 showed phosphate solubilisation and chitinase activity. All three strains inhibited pathogen growth by producing volatiles, and gas chromatography–mass spectrometry analysis revealed eight compounds in Pf 9A‐14, 10 in Bs 8B‐1 and 4 in Psp 8D‐45, some with known antifungal activity. The antagonistic and plant‐growth promotion activities of these strains might be due to production of antibiotics, metabolites, lytic enzymes or phytohormones. 相似文献
13.
Fusarium oxysporum Schlecht. (Hypocreales: Nectriaceae) is one of the most devastating plant pathogens worldwide, causing vascular wilt in several crops. Management of this disease primarily relies on chemical fungicides and resistant cultivars in high value crops. However, due to the limited efficacy of these methods, alternative control methods are needed. Biological control is a sustainable, safe, and effective alternative, but the use of a single biological control agent (BCA) usually has inconsistent results. The consistency of biocontrol could be enhanced using microbial consortia. In this context, the aim of this work was to select an effective microbial consortium against vascular wilt in cape gooseberry (Physalis peruviana L.) caused by Fusarium oxysporum f. sp. physali, from a mixture of four strains of Trichoderma spp. Pers. (Hypocreales: Hypocraceae) and Bacillus velezensis (Bacillales: Bacillaceae) Bs006. The calculated synergy factor was used as a selection criterion. Then the selected consortium was evaluated in the field and compared to carbendazim. The Trichoderma virens Gl006 and B. velezensis Bs006 consortium showed synergistic activity against vascular wilt under greenhouse and field conditions and efficacy similar to chemical control. These results suggest that Gl006 and Bs006 have a higher potential in controlling Fusarium wilt in cape gooseberry when applied as a consortium compared to separate, single strains. 相似文献
14.
Experiments employing both broth and soil cultures demonstrated the capacity for bidirectional genetic exchange between the eubacterial species Bacillus subtilis and Bacillus licheniformis. The process was studied using standard laboratory strains and wild isolates of these species. The genetic exchange in soil occurs spontaneously. The interspecific recombination involved markers for antibiotic resistance and for the use of specific carbon sources (API characters). Hybrids frequently had unstable phenotypes, i.e., lacked a consistent expression of foreign genes over repeated transfer and growth. This instability often involved a “correction” back toward the phenotype of one or the other of the parental species for many differentiating characters; the final phenotype was always that of the more probable or actually known recipient species. This “correction” process is reminiscent of phenomena associated with the instability of artificial fusion protoplasts or noncomplementing diploids of B. subtilis, as well as the merodiploids formed by intergeneric crosses with enteric bacteria. The hybrids observed here must also be diploid, in some manner, because they sequentially express traits of both parental species at rates well above the frequency of mutation. Among the unstable changes in hybrids of the wild strains there was a 3:1 bias in favor of “correction.” The dynamics of the hybridization process in soil are described. It appears that the hybrids are formed most rapidly following outgrowth from spores and during the early growth of parental vegetative cell populations. Later on, the hybrids are much less frequent in the soil cultures, suggesting that they are competitively inferior to the parental species. It is argued that the capacity for recombination found between B. subtilis and B. licheniformis could locally erase their distinctness, even though they possess only about 15% DNA sequence homology. Yet they remain distinct in the wild. The methods and results of these experiments prepare the way for detailed studies of the nature of species and species boundaries throughout the genus Bacillus. 相似文献
15.
田间试验结果表明 ,阿维菌素对美洲斑潜Liriomyzasativae蝇幼虫防效较好 ,优于伊维菌素对美洲斑潜蝇的防效 ,可较好地控制美洲斑潜蝇幼虫的危害 ,而Btg,Bti,Bs不宜单独用于防治美洲斑潜蝇幼虫。阿维菌素用量为 0 2 7,0 3 6,0 45g (a .i.) 667m2 时 ,对美洲斑潜蝇幼虫防效第 7d校正防效为 65 %左右 ,施药后第 1 1d校正防效为 85 48%~ 99 0 5 % ,施药后第 1 5d校正防效为 90 94%~ 99 89%。伊维菌素 0 5g (a .i.) 667m2 施药后第 3 ,7,1 1d校正防效分别为 61 67% ,90 5 3 % ,90 93 % ,伊维菌素 0 2 5g(a i ) 667m2 相应防效为 5 7.71 % ,84 68% ,85 83 %。Btg ,Bti,Bs施药后第 3 ,7,1 1d校正防效为 3 3 88%~ 5 5 5 4%。 相似文献
16.
A chromosomal DNA fragment containing the Bacillus macquariensis (Bm) ATP-dependent phosphofructokinase-encoding gene (pfk) was cloned from a subgenomic library in pUC19 using a PCR-derived probe. The region containing pfk, including flanking sequences, was sequenced and the deduced amino acid sequence (aa) was found to be homologous to other PFK, but it contained two single-aa changes conserved in a range of other organisms from pro- and eukaryotic origins. Enzymatic studies with PFK purified from overproducing Escherichia coli (Ec) host cells showed that the Bm enzyme is similar to B. stearothermophilus (Bs) PFK in many respects and that it is relatively cold stable. 相似文献
17.
Aims: To determine the wet and dry density of spores of Bacillus anthracis and compare these values with the densities of other Bacillus species grown and sporulated under similar conditions. Methods and Results: We prepared and studied spores from several Bacillus species, including four virulent and three attenuated strains of B. anthracis, two Bacillus species commonly used to simulate B. anthracis ( Bacillus atrophaeus and Bacillus subtilis) and four close neighbours ( Bacillus cereus, Bacillus megaterium, Bacillus thuringiensis and Bacillus stearothermophilus), using identical media, protocols and instruments. We determined the wet densities of all spores by measuring their buoyant density in gradients of Percoll and their dry density in gradients of two organic solvents, one of high and the other of low chemical density. The wet density of different strains of B. anthracis fell into two different groups. One group comprised strains of B. anthracis producing spores with densities between 1·162 and 1·165 g ml ?1 and the other group included strains whose spores showed higher density values between 1·174 and 1·186 g ml ?1. Both Bacillus atrophaeus and B. subtilis were denser than all the B. anthracis spores studied. Interestingly and in spite of the significant differences in wet density, the dry densities of all spore species and strains were similar. In addition, we correlated the spore density with spore volume derived from measurements made by electron microscopy analysis. There was a strong correlation ( R2 = 0·95) between density and volume for the spores of all strains and species studied. Conclusions: The data presented here indicate that the two commonly used simulants of B. anthracis, B. atrophaeus and B. subtilis were considerably denser and smaller than all B. anthracis spores studied and hence, these simulants could behave aerodynamically different than B. anthracis. Bacillus thuringiensis had spore density and volume within the range observed for the various strains of B. anthracis. The clear correlation between wet density and volume of the B. anthracis spores suggest that mass differences among spore strains may be because of different amounts of water contained within wet dormant spores. Significance and Impact of the Study: Spores of nonvirulent Bacillus species are often used as simulants in the development and testing of countermeasures for biodefense against B. anthracis. The similarities and difference in density and volume that we found should assist in the selection of simulants that better resemble properties of B. anthracis and, thus more accurately represent the performance of countermeasures against this threat agent where spore density, size, volume, mass or related properties are relevant. 相似文献
18.
A loop-mediated isothermal amplification (LAMP) assay system was employed for detecting Bacillus anthracis spores in pure cultures as well as in various simulated powder samples. The specificity of the designed LAMP primer sets
was validated by assaying 13 B. anthracis strains and 33 non- B. anthracis species. The detection limits of the LAMP assay were 10 spores/tube for pure cultures and 100 spores/2 mg powder for simulated
powder samples. The results show that the LAMP protocol is a promising method for detecting B. anthracis. 相似文献
19.
Coinoculation with plant growth–promoting rhizobacteria (PGPR) and arbuscular mycorrhizal fungi (AMF) has been proposed as
an efficient method to increase plant growth. In this article we investigate how the interaction between three different AMF
isolates ( Glomus constrictum autochthonous, GcA; G. constrictum from collection, GcC; and commercial Glomus intraradices, Gi) and a Bacillus megaterium strain isolated from a Mediterranean calcareous soil affects Lactuca sativa L. plant growth. Inoculation with B. megaterium increased plant growth when in combination with two of the AMF isolates (GcA and Gi), but decreased it when in combination
with GcC. At the same time, plants inoculated with the GcC fungus alone or in combination with B. megaterium (GcC+Bm) showed leaf symptoms of stress injury by accumulating proline and reducing the amount of photosynthetic pigments,
whereas the opposite occurred in plants coinoculated with Gi fungus and B. megaterium (Gi+Bm). GcC+Bm leaves also presented the highest glucose-6-phosphate dehydrogenase (G6PDH) and the lowest glutamine synthetase
(GS) enzymatic activities, whereas Gi+Bm leaves showed the highest GS activity. Results on these enzymatic activities are
further discussed in relation to plant growth and performance. 相似文献
20.
Summary Three related strains of the genus Bacillus, viz. B. licheniformis, B. subtilis and Bacillus Q were all found to contain a minor species of 5 S RNA in an amount indicating that it is transcribed from only one of the multiple 5 S RNA cistrons known to be present in these strains. The major and minor types of 5 S RNA from each of the three strains could be separated from each other by polyacrylamide gel electrophoresis in the presence of urea. The complete nucleotide sequences of the minor B. subtilis and Bacillus Q 5 S RNAs have been determined. Comparison of these sequences to the previously determined sequence of the minor 5 S RNA from B. licheniformis (Raué et al., 1976) showed the three minor types of 5 S RNA to be identical except for the residues at positions 79, 85 and 95. Moreover, seven out of the eight sequence differences between the major and the minor 5 S RNA are the same in all three strains. Thus, the single cistron coding for minor 5 S RNA has been strongly conserved in all three strains, which may indicate a biological significance for the minor 5 S RNA species. 相似文献
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