Showing papers on "Bacillus thuringiensis published in 2002"
TL;DR: The understanding of the biochemical and genetic basis of resistance to Bt can help design appropriate management tactics to delay or reduce the evolution of resistance in insect populations.
Abstract: Bacillus thuringiensis (Bt) is a valuable source of insecticidal proteins for use in conventional sprayable formulations and in transgenic crops, and it is the most promising alternative to synthetic insecticides. However, evolution of resistance in insect populations is a serious threat to this technology. So far, only one insect species has evolved significant levels of resistance in the field, but laboratory selection experiments have shown the high potential of other species to evolve resistance against Bt. We have reviewed the current knowledge on the biochemical mechanisms and genetics of resistance to Bt products and insecticidal crystal proteins. The understanding of the biochemical and genetic basis of resistance to Bt can help design appropriate management tactics to delay or reduce the evolution of resistance in insect populations.
907 citations
TL;DR: Transgenic plants expressing insecticidal proteins from the bacterium, Bacillus thuringiensis (Bt), are revolutionizing agriculture and have shown positive economic benefits to growers and reduced the use of other insecticides.
Abstract: ▪ Abstract Transgenic plants expressing insecticidal proteins from the bacterium, Bacillus thuringiensis (Bt), are revolutionizing agriculture. Bt, which had limited use as a foliar insecticide, has become a major insecticide because genes that produce Bt toxins have been engineered into major crops grown on 11.4 million ha worldwide in 2000. Based on the data collected to date, generally these crops have shown positive economic benefits to growers and reduced the use of other insecticides. The potential ecological and human health consequences of Bt plants, including effects on nontarget organisms, food safety, and the development of resistant insect populations, are being compared for Bt plants and alternative insect management strategies. Scientists do not have full knowledge of the risks and benefits of any insect management strategies. Bt plants were deployed with the expectation that the risks would be lower than current or alternative technologies and that the benefits would be greater. Based on th...
781 citations
TL;DR: Identification of more than 20 bacterial isolates capable of enzymatic inactivation of AHLs from different sources are reported, indicating that these isolates probably belong to the species Bacillus thuringiensis.
Abstract: A range of gram-negative bacterial species use N-acyl homoserine lactone (AHL) molecules as quorum-sensing signals to regulate different biological functions, including production of virulence factors. AHL is also known as an autoinducer. An autoinducer inactivation gene, aiiA, coding for an AHL lactonase, was cloned from a bacterial isolate, Bacillus sp. strain 240B1. Here we report identification of more than 20 bacterial isolates capable of enzymatic inactivation of AHLs from different sources. Eight isolates showing strong AHL-inactivating enzyme activity were selected for a preliminary taxonomic analysis. Morphological phenotypes and 16S ribosomal DNA sequence analysis indicated that these isolates probably belong to the species Bacillus thuringiensis. Enzymatic analysis with known Bacillus strains confirmed that all of the strains of B. thuringiensis and the closely related species B. cereus and B. mycoides tested produced AHL-inactivating enzymes but B. fusiformis and B. sphaericus strains did not. Nine genes coding for AHL inactivation were cloned either by functional cloning or by a PCR procedure from selected bacterial isolates and strains. Sequence comparison of the gene products and motif analysis showed that the gene products belong to the same family of AHL lactonases.
480 citations
Patent•
28 Feb 2002
TL;DR: In this paper, the Bacillus thuringiensis delta-endotoxin H04 was used to control insect pests and to confer insect resistance in trenasgenic plants.
Abstract: Synthetic nucleotide sequences optimized for expression in plants encode varying forms of the hybrid Bacillus thuringiensis delta-endotoxin H04, the toxin portion of which contains domains I and II of Cry1Ab and domain III of Cry1C. Compositions and formulations containing the insecticidal toxins are capable of controlling insect pests. The invention is further drawn to methods of making the hybrid toxins and to methods of using the nucleotide sequences, for example in microorganisms to control insect pests and in trenasgenic plants to confer insect resistance.
420 citations
TL;DR: Administration of slapn double-stranded RNA to S. litura larvae reduces its expression and this results directly implicate larval midgut aminopeptidase N as receptor for Bacillus thuringiensis insecticidal proteins.
333 citations
TL;DR: The entire 127,923-bp sequence of the toxin-encoding plasmid pBtoxis from Bacillus thuringiensis subsp.
Abstract: The entire 127,923-bp sequence of the toxin-encoding plasmid pBtoxis from Bacillus thuringiensis subsp. israelensis is presented and analyzed. In addition to the four known Cry and two known Cyt toxins, a third Cyt-type sequence was found with an additional C-terminal domain previously unseen in such proteins. Many plasmid-encoded genes could be involved in several functions other than toxin production. The most striking of these are several genes potentially affecting host sporulation and germination and a set of genes for the production and export of a peptide antibiotic.
296 citations
TL;DR: It was observed that recombinant Escherichia coli producing AiiA proteins also had AHL-degrading activity and could also attenuate the plant pathogenicity of Erwinia carotovora, indicating that insecticidal B. thuringiensis strains might have potential to compete with gram-negative bacteria in natural ecosystems by autoinducer-degRading activity.
Abstract: Gram-negative bacteria can communicate with each other by N-acyl homoserine lactones (AHLs), which are quorum-sensing autoinducers. Recently, the aiiA gene (encoding an enzyme catalyzing the degradation of AHL) has been cloned from Bacillus sp. strain 240B1. During investigations in the course of the ongoing Bacillus thuringiensis subsp. morrisoni genome project, an aiiA homologue gene in the genome sequence was found. These results led to consideration of the possibility of the widespread existence of the gene in B. thuringiensis. aiiA homologue genes were found in 16 subspecies of B. thuringiensis, and their sequences were determined. Comparison of the Bacillus sp. strain 240B1 aiiA gene with the B. thuringiensis aiiA homologue genes showed high homologies of 89 to 95% and 90 to 96% in the nucleotide sequence and deduced amino acid sequence, respectively. Among the subspecies of B. thuringiensis having an aiiA gene, the subspecies aizawai, galleriae, kurstaki, kyushuensis, ostriniae, and subtoxicus were shown to degrade AHL. It was observed that recombinant Escherichia coli producing AiiA proteins also had AHL-degrading activity and could also attenuate the plant pathogenicity of Erwinia carotovora. These results indicate that insecticidal B. thuringiensis strains might have potential to compete with gram-negative bacteria in natural ecosystems by autoinducer-degrading activity.
237 citations
TL;DR: The results indicated that the release of the Cry1Ab protein by roots is a common phenomenon with transgenic Btcorn and is not restricted to only the one Bt corn hybrid and tranformation event studied initially.
Abstract: The anti-lepidopteran toxin (Cry1Ab protein) encoded by truncated genes from Bacillus thuringiensis was released in the root exudates from all hybrids of Bt corn studied and which represented three transformation events (Bt11, MON810, and 176). In vitro and in situ studies indicated that the toxin released in root exudates accumulates in soil, as it adsorbs and binds rapidly on surface-active particles (e.g. clays and humic substances), and retains insecticidal activity for at least 180 d, the longest time studied. The results indicated that the release of the Cry1Ab protein by roots is a common phenomenon with transgenic Bt corn and is not restricted to only the one Bt corn hybrid (NK4640Bt) and tranformation event (Bt11) studied initially.
209 citations
TL;DR: Bioassays conducted with Cry1Ac and Cry2Aa showed that there was a small but significant negative interaction between these delta-endotoxins, and H. armigera was consistently more tolerant to B. thuringiensis insecticidal proteins than was H. punctigera, although both were susceptible to only a limited range of these proteins.
155 citations
TL;DR: Analysis of survival and growth of progeny from backcrosses suggest that resistance was controlled primarily by one or a few major loci, and the progression of resistance from 300- to 3,100-fold rules out the simplest model with one locus and two alleles.
Abstract: Laboratory selection with Cry1Ac, the Bacillus thuringiensis (Bt) toxin in transgenic cotton, initially produced 300-fold resistance in a field-derived strain of pink bollworm, Pectinophora gossypiella (Saunders), a major cotton pest. After additional selection increased resistance to 3,100-fold, we tested the offspring of various crosses to determine the mode of inheritance of resistance to Cry1Ac. The progeny of reciprocal F1 crosses (resistant male × susceptible female and vice versa) responded alike in bioassays, indicating autosomal inheritance. Consistent with earlier findings, resistance was recessive at a high concentration of Cry1Ac. However, the dominance of resistance increased as the concentration of Cry1Ac decreased. Analysis of survival and growth of progeny from backcrosses (F1 × resistant strain) suggest that resistance was controlled primarily by one or a few major loci. The progression of resistance from 300- to 3,100-fold rules out the simplest model with one locus and two alleles. Overall the patterns observed can be explained by either a single resistance gene with three or more alleles or by more than one resistance gene. The pink bollworm resistance to Cry1Ac described here fits “mode 1” resistance, the most common type of resistance to Cry1A toxins in Lepidoptera.
152 citations
TL;DR: These plants, in combination with the resistant DBM populations available, will allow greenhouse or field studies of resistance management strategies involving gene pyramiding.
Abstract: This study was undertaken to determine the effects of pyramiding two Bacillus thuringiensis (Bt) genes in the same plant on the production of Bt proteins and the control of diamondback moths (DBM, Plutella xylostella) resistant to one or the other protein. Broccoli lines carrying both cry1Ac and cry1C Bt genes were produced by sexual crosses of cry1Ac- and cry1C-transgenic plants. Plants containing both genes were selected by tests for resistance to kanamycin and hygromycin, and confirmed by PCR analysis for the Bt genes. Both cry1Ac and cry1C mRNAs were detected in the hybrid lines, and Cry1Ac and Cry1C proteins were stably produced at levels comparable to the parental plants. Plants producing both Cry1Ac and Cry1C proteins caused rapid and complete mortality of DBM larvae resistant to Cry1A or Cry1C, and suffered little or no leaf damage. These plants, in combination with the resistant DBM populations available, will allow greenhouse or field studies of resistance management strategies involving gene pyramiding.
TL;DR: A new family of insecticidal crystal proteins was discovered by screening sporulated Bacillus thuringiensis cultures for oral activity against western corn rootworm larvae by finding the 44-kDa polypeptide and the 41.9- and 51.4-k da binary dipteran insecticidal proteins from Bacillus sphaericus are evolutionarily related.
Abstract: A new family of insecticidal crystal proteins was discovered by screening sporulated Bacillus thuringiensis cultures for oral activity against western corn rootworm (WCR) larvae. B. thuringiensis isolates PS80JJ1, PS149B1, and PS167H2 have WCR insecticidal activity attributable to parasporal inclusion bodies containing proteins with molecular masses of ca. 14 and 44 kDa. The genes encoding these polypeptides reside in apparent operons, and the 14-kDa protein open reading frame (ORF) precedes the 44-kDa protein ORF. Mutagenesis of either gene in the apparent operons dramatically reduced insecticidal activity of the corresponding recombinant B. thuringiensis strain. Bioassays performed with separately expressed, biochemically purified 14- and 44-kDa polypeptides also demonstrated that both proteins are required for WCR mortality. Sequence comparisons with other known B. thuringiensis insecticidal proteins failed to reveal homology with previously described Cry, Cyt, or Vip proteins. However, there is evidence that the 44-kDa polypeptide and the 41.9- and 51.4-kDa binary dipteran insecticidal proteins from Bacillus sphaericus are evolutionarily related. The 14- and 44-kDa polypeptides from isolates PS80JJ1, PS149B1, and PS167H2 have been designated Cry34Aa1, Cry34Ab1, and Cry34Ac1, respectively, and the 44-kDa polypeptides from these isolates have been designated Cry35Aa1, Cry35Ab1, and Cry35Ac1, respectively.
TL;DR: The amount of Cry1Ac protein accumulated as a result of continuous use of transgenic Bt cotton, and subsequent incorporation of plant residues into the soil by postharvest tillage, is extremely low and does not result in detectable biological activity.
Abstract: Soil samples were collected from within and outside six fields where insect-resistant transgenic cotton (Bollgard) encoding the Bacillus thuringiensis Berliner (Bt) subsp. kurstaki cry1Ac gene had been grown and subsequently incorporated into soil by postharvest tillage for 3–6 consecutive years. The level of Cry1Ac protein in these samples (collected 3 mo after the last season’s tillage) was evaluated using both enzyme-linked immunosorbent assays (ELISA) and bioassays with a susceptible insect species, Heliothis virescens (F.), the tobacco budworm. Both methods revealed that no detectable Cry1Ac protein was present in any of the soil samples collected from within or outside the Bollgard fields. Based on the results from reference standards, the limit of detection for the ELISA was 3.68 ng of extractable protein per gram of soil, and that of the bioassay (measured by EC50) was 8 ng of biologically active protein per gram of soil. Together, these findings demonstrate that the amount of Cry1Ac protein accumulated as a result of continuous use of transgenic Bt cotton, and subsequent incorporation of plant residues into the soil by postharvest tillage, is extremely low and does not result in detectable biological activity.
TL;DR: Recruitment of BT-R(1) by B. thuringiensis indicates that the bacterium interacts with a specific cell adhesion molecule during its pathogenesis, which apparently indicates that Cry toxins, like other bacterial toxins, attack epithelial barriers by targeting cellAdhesion molecules within susceptible insect hosts.
TL;DR: The isotope data on moths collected in Texas early in the season indicate that the majority of overwintering H. zea do not originate from cotton-feeding larvae and may be migrants from Mexico, and support other research indicating that many late-season H.Zea moths captured in Louisiana and Texas are migrants whose larvae developed on corn in more northern locations.
Abstract: Data have been lacking on the proportion of Helicovera zea larvae that develop on noncotton host plants that can serve as a refuge from selection pressure for adaptation to transgenic cotton varieties that produce a toxin from the bacterium Bacillus thuringiensis. We found that individual H. zea moths that develop as larvae on cotton and other plants with C3 physiology have a different ratio of 13C to 12C than moths that develop on plants with C4 physiology, such as corn. We used this finding in determining the minimum percentage of moths that developed on noncotton hosts in two cotton-growing areas. Our results indicate that local corn can serve as a refuge for H. zea in midsummer. Our results contrast dramatically with the prevailing hypothesis that the large majority of late-season moths are produced from larvae feeding on cotton, soybean, and other C3 plants. Typically, <50% of moths captured in August through October have isotope ratios indicative of larval feeding on C3 plants. In one October sample, 100% of the moths originated from C4 hosts even though C4 crops were harvested at least 1 mo earlier, and no common wild C4 hosts were available. These findings support other research indicating that many late-season H. zea moths captured in Louisiana and Texas are migrants whose larvae developed on corn in more northern locations. Our isotope data on moths collected in Texas early in the season indicate that the majority of overwintering H. zea do not originate from cotton-feeding larvae and may be migrants from Mexico. Non-Bt corn in Mexico and the U.S. corn belt appears to serve as an important refuge for H. zea.
TL;DR: The characterization of a mini-Tn10 mutant of B. thuringiensis that is defective in flagellum filament assembly and in swimming and swarming motility as well as in the production of hemolysin BL and phosphatidylcholine-preferring phospholipase C is described.
Abstract: Bacillus thuringiensis is being used worldwide as a biopesticide, although increasing evidence suggests that it is emerging as an opportunistic human pathogen. While phospholipases, hemolysins, and enterotoxins are claimed to be responsible for B. thuringiensis virulence, there is no direct evidence to indicate that the flagellum-driven motility plays a role in parasite-host interactions. This report describes the characterization of a mini-Tn10 mutant of B. thuringiensis that is defective in flagellum filament assembly and in swimming and swarming motility as well as in the production of hemolysin BL and phosphatidylcholine-preferring phospholipase C. The mutant strain was determined to carry the transposon insertion in flhA, a flagellar class II gene encoding a protein of the flagellar type III export apparatus. Interestingly, the flhA mutant of B. thuringiensis synthesized flagellin but was impaired in flagellin export. Moreover, a protein similar to the anti-sigma factor FlgM that acts in regulating flagellar class III gene transcription was not detectable in B. thuringiensis, thus suggesting that the flagellar gene expression hierarchy of B. thuringiensis differs from that described for Bacillus subtilis. The flhA mutant of B. thuringiensis was also defective in the secretion of hemolysin BL and phosphatidylcholine-preferring phospholipase C, although both of these virulence factors were synthesized by the mutant. Since complementation of the mutant with a plasmid harboring the flhA gene restored swimming and swarming motility as well as secretion of toxins, the overall results indicate that motility and virulence in B. thuringiensis may be coordinately regulated by flhA, which appears to play a crucial role in the export of flagellar as well as nonflagellar proteins.
TL;DR: If plants with Cry1Ac plus Cry2Ab are deployed while genes that confer resistance to each of these toxins are rare, and if the inheritance of resistance to both toxins is recessive, the efficacy of transgenic cotton might be greatly extended.
Abstract: Crops genetically engineered to produce Bacillus thuringiensis toxins for insect control can reduce use of conventional insecticides, but insect resistance could limit the success of this technology. The first generation of transgenic cotton with B. thuringiensis produces a single toxin, Cry1Ac, that is highly effective against susceptible larvae of pink bollworm (Pectinophora gossypiella), a major cotton pest. To counter potential problems with resistance, second-generation transgenic cotton that produces B. thuringiensis toxin Cry2Ab alone or in combination with Cry1Ac has been developed. In greenhouse bioassays, a pink bollworm strain selected in the laboratory for resistance to Cry1Ac survived equally well on transgenic cotton with Cry1Ac and on cotton without Cry1Ac. In contrast, Cry1Ac-resistant pink bollworm had little or no survival on secondgeneration transgenic cotton with Cry2Ab alone or with Cry1Ac plus Cry2Ab. Artificial diet bioassays showed that resistance to Cry1Ac did not confer strong cross-resistance to Cry2Aa. Strains with >90% larval survival on diet with 10 g of Cry1Ac per ml showed 0% survival on diet with 3.2 or 10 g of Cry2Aa per ml. However, the average survival of larvae fed a diet with 1 g of Cry2Aa per ml was higher for Cry1Ac-resistant strains (2 to 10%) than for susceptible strains (0%). If plants with Cry1Ac plus Cry2Ab are deployed while genes that confer resistance to each of these toxins are rare, and if the inheritance of resistance to both toxins is recessive, the efficacy of transgenic cotton might be greatly extended.
TL;DR: The results of this study confirm the presence of enterotoxin genes in natural isolates of Bacillus spp.
Abstract: Thirty-nine Bacillus strains obtained from a variety of environmental and food sources were screened by PCR for the presence of five gene targets (hblC, hblD, hblA, nheA, and nheB) in two enterotoxin operons (HBL and NHE) traditionally harbored by Bacillus cereus. Seven isolates exhibited a positive signal for at least three of the five possible targets, including Bacillus amyloliquefaciens, B. cereus, Bacillus circulans, Bacillus lentimorbis, Bacillus pasteurii, and Bacillus thuringiensis subsp. kurstaki. PCR amplicons were confirmed by restriction enzyme digest patterns compared to a positive control strain. Enterotoxin gene expression of each strain grown in a model food system (skim milk) was monitored by gene-specific reverse transcription-PCR and confirmed with the Oxoid RPLA and Tecra BDE commercial kits. Lecithinase production was noted on egg yolk-polymyxin B agar for all strains except B. lentimorbis, whereas discontinuous beta hemolysis was exhibited by all seven isolates grown on 5% sheep blood agar plates. The results of this study confirm the presence of enterotoxin genes in natural isolates of Bacillus spp. outside the B. cereus group and the ability of these strains to produce toxins in a model food system under aerated conditions at 32 degrees C.
TL;DR: It is shown that APN can function as a receptor for Cry1Ac1 in vivo, and the GAL4 enhancer trap technique is utilized to drive the expression of M. sexta APN in both midgut and mesodermal tissues of Cry1 Ac1 insensitive Drosophila larvae.
Abstract: The bacterium Bacillus thuringiensis synthesizes toxins (delta-endotoxins) that are highly specific for insects Once ingested, the activated form of the toxin binds to a specific receptor(s) located on the midgut epithelial cells, inserts into the membrane causing the formation of leakage pores and eventual death of the susceptible insect larvae Manduca sexta larvae are highly susceptible to Cry1Ac1, a toxin that is believed to bind M sexta Aminopeptidase N, a glycoprotein located on the apical membrane However, the binding data obtained to date only support the interaction of Cry1Ac1 with APN in vitro To explore the in vivo role of APN, we have utilized the GAL4 enhancer trap technique to drive the expression of M sexta APN in both midgut and mesodermal tissues of Cry1Ac1 insensitive Drosophila larvae Transgenic Drosophila fed the toxin were now killed, demonstrating that APN can function as a receptor for Cry1Ac1 in vivo
TL;DR: Results indicate that the B. thuringiensis InhA2 zinc metalloprotease has a vital role in virulence when the host is infected via the oral route.
Abstract: The entomopathogenic bacterium Bacillus thuringiensis is known to secrete a zinc metalloprotease (InhA) that specifically cleaves antibacterial peptides produced by insect hosts. We identified a second copy of the inhA gene, named inhA2, in B. thuringiensis strain 407 Cry−. The inhA2 gene encodes a putative polypeptide showing 66.2% overall identity with the InhA protein and harboring the zinc-binding domain (HEXXH), which is characteristic of the zinc-requiring metalloproteases. We used a transcriptional inhA2′-lacZ fusion to show that inhA2 expression is induced at the onset of the stationary phase and is overexpressed in a Spo0A minus background. The presence of a reverse Spo0A box in the promoter region of inhA2 suggests that Spo0A directly regulates the transcription of inhA2. To determine the role of the InhA and InhA2 metalloproteases in pathogenesis, we used allelic exchange to isolate single and double mutant strains for the two genes. Spores and vegetative cells of the mutant strains were as virulent as those of the parental strain in immunized Bombyx mori larvae infected by the intrahemocoelic route. Exponential phase cells of all the strains displayed the same in vitro potential for colonizing the vaccinated hemocoel. We investigated the synergistic effect of the mutant strain spores on the toxicity of Cry1C proteins against Galleria mellonella larvae infected via the oral pathway. The spores of ΔinhA2 mutant strain were ineffective in providing synergism whereas those of the ΔinhA mutant strain were not. These results indicate that the B. thuringiensis InhA2 zinc metalloprotease has a vital role in virulence when the host is infected via the oral route.
TL;DR: Overall, the data indicate that it is not important for risk assessment purposes to determine whether B. cereus and B. thuringiensis belong to a single or two species, and that the rate of gene flow was higher between strains of the same species, but that exchanges between B. Cereus andB.
Abstract: Little is known about genetic exchanges in natural populations of bacteria of the spore-forming Bacillus cereus group, because no population genetics studies have been performed with local sympatric populations. We isolated strains of Bacillus thuringiensis and B. cereus from small samples of soil collected at the same time from two separate geographical sites, one within the forest and the other at the edge of the forest. A total of 100 B. cereus and 98 B. thuringiensis strains were isolated and characterized by electrophoresis to determine allelic composition at nine enzymatic loci. We observed genetic differentiation between populations of B. cereus and B. thuringiensis. Populations of a given Bacillus species--B. thuringiensis or B. cereus--were genetically more similar to each other than to populations of the other Bacillus species. Hemolytic activity provided further evidence of this genetic divergence, which remained evident even if putative clones were removed from the data set. Our results suggest that the rate of gene flow was higher between strains of the same species, but that exchanges between B. cereus and B. thuringiensis were nonetheless possible. Linkage disequilibrium analysis revealed sufficient recombination for B. cereus populations to be considered panmictic units. In B. thuringiensis, the balance between clonal proliferation and recombination seemed to depend on location. Overall, our data indicate that it is not important for risk assessment purposes to determine whether B. cereus and B. thuringiensis belong to a single or two species. Assessment of the biosafety of pest control based on B. thuringiensis requires evaluation of the extent of genetic exchange between strains in realistic natural conditions.
TL;DR: The results indicate that N. lugens and its natural enemies will be exposed to Bt toxins from rice lines transformed with some Bt gene constructs, but that this exposure might not affect N. lividipennis fitness.
Abstract: Five transgenic rice lines, each containing an insecticidal toxin gene from Bacillus thuringiensis (Bt) under control of a different promoter, were tested for effects on two non-target insects: the brown planthopper, Nilaparvata lugens (Stal) (Homoptera: Delphacidae), and its predator Cyrtorhinus lividipennis (Hemiptera: Miridae). Bt toxin was detected by ELISA in the honeydew of N. lugens that fed on rice lines with the CaMV 35S and actin promoters. Nilaparvata lugens produced greater volumes of acidic honeydew (derived from xylem feeding) on all five Bt rice lines than on non-transgenic control lines. The amount of honeydew derived from phloem feeding did not differ between Bt and control lines. There were no differences between N. lugens reared on Bt and control lines in any of the five fitness parameters measured (survival to the adult stage, male and female weight, and male and female developmental time). There were no differences between C. lividipennis reared on N. lugens nymphs from Bt and control lines, in any of the three fitness parameters examined (survival to the adult stage and male and female developmental time). Our results indicate that N. lugens and its natural enemies will be exposed to Bt toxins from rice lines transformed with some Bt gene constructs, but that this exposure might not affect N. lugens and C. lividipennis fitness.
TL;DR: A significant variation in parasitism by the tachinids Lydella thompsoni (Herting) and Pseudoperichaeta nigrolineata (Walker) among sites, and more parasitism in non-Bt than in Bt fields is found, and implications for pest resistance management, population dynamics and risk assessment are discussed.
Abstract: In this study, we evaluated in field trials the effects on non-target species, of transgenic corn producing the Cry1Ab toxin of Bacillus thuringiensis (Bt). In 1998, we collected Ostrinia nubilalis (Hubner) larvae from transgenic Bt corn (Novartis Hybrid 176) and non-Bt corn at four geographical sites. We found a significant variation in parasitism by the tachinids Lydella thompsoni (Herting) and Pseudoperichaeta nigrolineata (Walker) among sites, and more parasitism in non-Bt than in Bt fields. The Bt effect did not vary significantly among fields. In 1999, we performed a field experiment at two sites, comparing the temporal abundance of non-target arthropods in Bt corn (Monsanto Hybrid MON810) and non-Bt corn. The non-target insects studied included the aphids Metopolophium dirhodum (Walker), Rhopalosiphum padi (L.) and Sitobion avenae (F.), the bug Orius insidiosus (Say), the syrphid Syrphus corollae (Meigen), the ladybird Coccinella septempunctata (L.), the lacewing Chrysoperla carnea (Stephens), thrips and hymenopteran parasitoids. For all species but one, the number of individuals varied greatly over the season but did not differ between the types of corn. The only exception was thrips which, at one site, was significantly more abundant in Bt corn than in non-Bt corn. However this difference did not remain significant when we took the multiple tests into account. Implications for pest resistance management, population dynamics and risk assessment are discussed.
TL;DR: To evaluate the possibility for discrimination of Bacillus cereus and B. thuringiensis using 16S rRNA and gyrB gene based PCR methods, and to obtain the sequences of the primer annealing sites so that the PCR results may be explained.
Abstract: Aims: To evaluate the possibility for discrimination of Bacillus cereus and B. thuringiensis using 16S rRNA and gyrB gene based PCR methods, and to obtain the sequences of the primer annealing sites so that the PCR results may be explained.
Methods and Results: Based on the sequence difference in the variable region (V1) of 16S rRNA and in the gyrB gene between B. cereus and B. thuringiensis, PCR primers specific to these Bacillus spp. were designed. When these primers were used to discriminate B. cereus and B. thuringiensis, six of 82 B. cereus strains were identified as B. thuringiensis while 67 of 73 B. thuringiensis strains were identified as B. cereus. Sequence analysis of the primer annealing sites showed that there is no clear-cut difference in the V1 region of 16S rRNA, and in the gyrB gene, between B. cereus and B. thuringiensis strains.
Conclusions: Although 16S rDNA based probes and gyrB gene based PCR primers have been suggested for the discrimination of B. cereus and B. thuringiensis strains, when a large number of Bacillus strains was tested, results showed that discrimination between B. cereus and B. thuringiensis is difficult. Therefore, to distinguish B. thuringiensis from B. cereus, a single feature, such as the presence of a parasporal crystal protein or cry gene, may sometimes be reliable.
Significance and Impact of the Study: Discrimination between B. cereus and B. thuringiensis is a challenging debate to which this paper makes a contribution.
TL;DR: Bacillus thuringiensis is distinguished from the very closely related Bacillus cereus and Bacillus anthracis by the presence of several plasmid-encoded δ-endotoxin genes, which are produced in large quantities during sporulation and are packaged into intracellular inclusions.
Abstract: Bacillus thuringiensis is distinguished from the very closely related Bacillus cereus and Bacillus anthracis by the presence of several plasmid-encoded δ-endotoxin genes. These δ-endotoxins, synthesized as protoxins, are produced in large quantities during sporulation and are packaged into intracellular inclusions. Ingestion of the inclusions by insect larvae leads to protoxin solubilization and conversion to toxins each specific for one of several orders of insects. The toxins form cation-selective channels in the membrane of cells lining the larval midgut with subsequent lethality. In most cases, δ-endotoxin synthesis and sporulation are closely coupled. The latter process in B. thuringiensis is probably virtually identical to that in Bacillus subtilis with the additional use of mother cell sporulation forms of RNA polymerase for the synthesis of the δ-endotoxins. There are other more subtle plasmid-encoded functions or plasmid interactions related to regulating protoxin synthesis. Consideration of both plasmid and chromosomal genes is thus critical for defining this organism.
09 Aug 2002
TL;DR: Longstanding Taxonomic Enigmas Within The a Bacillus Cereus Groupa Are On the Verge of Being Resolved By Far--Reaching Molecular Developments: Forecasts On the Possible Outcome By An Ad Hoc Team is Forecast.
Abstract: List of Contributors. Foreword. 1. Whither Bacillus? (Berkeley). 2. From Phylogeny To Systematics: Dissection Of The Genus Bacillus (Stackebrandt & Swiderski). 3. Longstanding Taxonomic Enigmas Within The a Bacillus Cereus Groupa Are On The Verge Of Being Resolved By Far--Reaching Molecular Developments: Forecasts On The Possible Outcome By An Ad Hoc Team. (Turnbull, Jackson, Hill, Keim, Kolsto & Beecher). 4. Bacillus Cereus And Food Poisoning (Granum). 5. Thermophilic Bacillus Isolates From Antarctic Environments (Nicolaus, Lama & Gambacorta). 6. Bacilli Associated With Spoilage In Dairy Products And Other Foods (Heyndrickx & Scheldeman). 7. Moderately Halophilic And Halotolerant Species Of Bacillus And Related Genera (Arahal & Ventosa). 8. Bacillus Identification -- Traditional Approaches (Fritze). 9. Modern Methods For Identification (Logan). 10. Nucleic Acid Analysis And SDS--PAGE Of Whole--Cell Proteins In Bacillus Taxonomy (De Vos). 11. Bacillus Thuringiensis Insecticides (Bishop). 12. Bt Crops: A Novel Insect Control Tool (Van Rie). 13. Bacillus Sphaericus And Its Insecticidal Toxins (Priest). 14. The Importance Of Bacillus Species In The Production Of Industrial Enzymes (Outtrop & Jorgensen). 15. Plant Growth Promotion By Bacillus And Relatives (Chanway). 16. Insertion Sequence Elements And Transposons In Bacillus (Mahillon). 17. Fingerprint Spectrometry Methods In Bacillus Systematics (Magee & Goodacre). 18. Whole--Cell Fatty Acid Analysis In The Systematics Of Bacillus And Related Genera (Kampfer). 19. Some Concluding Observations (Norris). Index
TL;DR: It was demonstrated that chitinase was widely produced in B. thuringiensis strains and some of the strains could enhance the toxicity of active B.Thuringienis strain against Spodoptera exigua larvae, and the bioassay results indicated that the chit inase‐producing B. Thuringiens is might play a role in the activity of the biopesticide.
Abstract: M . L I U , Q . X . C A I , H . Z . L I U , B . H . Z H A N G , J . P . Y A N A N D Z . M . Y U A N . 2002. Aims: To investigate the distribution of chitinase in Bacillus thuringiensis strains, and the enhancing effects of the chitinase-producing B. thuringiensis strains on insecticidal toxicity of active B. thuringiensis strain against Spodoptera exigua larvae. Methods and Results: The chitinolytic activities of B.thuringiensis strains representing the 70 serotypes were investigated by the whitish opaque halo and the colorimetric method. Thirtyeight strains produced different levels of chitinase at pH 7AE0, and so did 17 strains at pH 10AE0. The strain T04A001 exhibited the highest production, reaching a specific activity of 355 U ml )1 in liquid medium. SDS-PAGE and Western blotting showed that the chitinase produced by some B. thuringiensis strains had a molecular weight of about 61 kDa. The bioassay results indicated that the chitinase-producing B. thuringiensis strains could enhance the insecticidal activity of B. thuringiensis strain DL5789 against S. exigua larvae, with an enhancing ratio of 2AE35-fold. Conclusion: This study demonstrated that chitinase was widely produced in B. thuringiensis strains and some of the strains could enhance the toxicity of active B. thuringiensis strain. Significance and Impact of the Study: This is the first investigation devoted exclusively to analyse the distribution of chitinase in B. thuringiensis. It infers that the chitinase produced by B. thuringiensis might play a role in the activity of the biopesticide.
15 Oct 2002
TL;DR: The case of Bacillus thuringiensis (Bt) and its use in agriculture is discussed in this article, where the authors compare genetic modification of crops to alternatives and address the current controversy, positive outcomes, and potential risks associated with transgenic plants.
Abstract: Presents the case of Bacillus thuringiensis (Bt) and its use in agriculture. Compares genetic modification of crops to alternatives and addresses the current controversy, positive outcomes, and potential risks associated with transgenic plants. Makes specific recommendations for future research, evaluation and environmental monitoring, scientific coordination, and public education.
TL;DR: Field-release experiments indicated that the cry1Ab gene was stably transmitted in an intact manner via successive sexual generations, and the concentration of the Cry1Ab protein was kept quantitatively stable up to the R6 generation.
Abstract: The inheritance and expression patterns of the cry1Ab gene were studied in the progenies derived from different Bt ( Bacillus thuringiensis) transgenic japonica rice lines under field conditions. Both Mendelian and distorted segregation ratios were observed in some selfed and crossed F(2) populations. Crosses between japonica intra-subspecies had no significant effect on the segregation ratios of the cry1Ab gene, but crossing between japonica and indicainter-subspecies led to distorted segregation of the cry1Ab gene in the F(2)population. Field-release experiments indicated that the cry1Ab gene was stably transmitted in an intact manner via successive sexual generations, and the concentration of the Cry1Ab protein was kept quantitatively stable up to the R(6)generation. The cry1Ab gene, driven by the maize ubiquitinpromoter, displayed certain kinds of spatial and temporal expression patterns under field conditions. The content of the Cry1Ab protein varied in different tissues of the main stems, the primary tillers and the secondary tillers. Higher levels of the Cry1Ab protein were found in the stems, leaves and leaf sheaths than in the roots, while the lowest level was detected in grains at the maturation stage. The content of the Cry1Ab protein in the leaves peaked at the booting stage and was lowest at the heading stage. Furthermore, the Cry1Ab content of cry1Ab expression in different tissues of transgenic rice varied individually with temperature.
TL;DR: It may be necessary to compensate the original mixed cropping patterns in different areas for delaying resistance development of H. armigera to Bt cotton, and function of the refuge would closely depend on the proportion of Btotton.
Abstract: The density of Helicoverpa armigera (Hubner) populations on Bacillus thuringiensis Berliner (Bt) transgenic cotton, corn, peanut, and soybean; differences in its development on Bt cotton and common (nontransgenic) cotton; and the potential for mating among populations from Bt cotton fields and other crop fields were investigated in the suburbs of Xinxiang City (Henan Province) and Langfang City (Hebei Province) in the southern and northern parts of north China, respectively. Although development of H. armigera on Bt cotton was much slower than on common cotton, there was a still high probability of mating between populations from Bt cotton and other sources due to the scattered emergence pattern of H. armigera adults, and overlap of the second and third generations. In a cotton and corn growing region, early and late planted corn provided suitable refugia for the third and fourth generations of H. armigera, but not for the second generation. In a cotton and soybean/ peanut mix system, noncotton crops provided a natural refugia from the second- to fourth-generation H. armigera, but function of the refuge would closely depend on the proportion of Bt cotton. Consequently, it may be necessary to compensate the original mixed cropping patterns in different areas for delaying resistance development of H. armigera to Bt cotton.