No adverse effects of transgenic maize on population dynamics of endophytic Bacillus subtilis strain B916-gfp.
ABSTRACT: Endophytic bacterial communities play a key role in promoting plant growth and combating plant diseases. However, little is known about their population dynamics in plant tissues and bulk soil, especially in transgenic crops. This study investigated the colonization of transgenic maize harboring the Bacillus thuringiensis (Bt) cry1Ah gene by Bacillus subtilis strain B916-gfp present in plant tissues and soil. Bt and nontransgenic maize were inoculated with B916-gfp by seed soaking, or root irrigation under both laboratory greenhouse and field conditions. During the growing season, B916-gfp colonized transgenic as well as nontransgenic plants by both inoculation methods. No differences were observed in B916-gfp population size between transgenic and nontransgenic plants, except at one or two time points in the roots and stems that did not persist over the examination period. Furthermore, planting transgenic maize did not affect the number of B916-gfp in bulk soil in either laboratory or field trials. These results indicate that transgenic modification of maize with the cry1Ah gene has no influence on colonization by the endophytic bacteria B916-gfp present in the plant and in bulk soil.
Project description:The aim of the experiment was to determine if temporal variations of belowground activity reflect the influence of the Cry1Ab protein from transgenic maize on soil bacteria and, hence, on a regulatory change of the microbial community (ability to metabolize sources belonging to different chemical guilds) and/or a change in numerical abundance of their cells. Litter placement is known for its strong influence on the soil decomposer communities. The effects of the addition of crop residues on respiration and catabolic activities of the bacterial community were examined in microcosm experiments. Four cultivars of Zea mays L. of two different isolines (each one including the conventional crop and its Bacillus thuringiensis cultivar) and one control of bulk soil were included in the experimental design. The growth models suggest a dichotomy between soils amended with either conventional or transgenic maize residues. The Cry1Ab protein appeared to influence the composition of the microbial community. The highly enhanced soil respiration observed during the first 72 h after the addition of Bt-maize residues can be interpreted as being related to the presence of the transgenic crop residues. This result was confirmed by agar plate counting, as the averages of the colony-forming units of soils in conventional treatments were about one-third of those treated with transgenic straw. Furthermore, the addition of Bt-maize appeared to induce increased microbial consumption of carbohydrates in BIOLOG EcoPlates. Three weeks after the addition of maize residues to the soils, no differences between the consumption rate of specific chemical guilds by bacteria in soils amended with transgenic maize and bacteria in soils amended with conventional maize were detectable. Reaped crop residues, comparable to post-harvest maize straw (a common practice in current agriculture), rapidly influence the soil bacterial cells at a functional level. Overall, these data support the existence of short Bt-induced ecological shifts in the microbial communities of croplands' soils.
Project description:Background: Asian corn borer (ACB), Ostrinia furnacalis can develop resistance to transgenic Bacillus thuringiensis (Bt) maize expressing Cry1Ah-toxin. However, the mechanisms that regulate the resistance of ACB to Cry1Ah-toxin are unknown. Objective: In order to understand the molecular basis of the Cry1Ah-toxin resistance in ACB, "omics" analyses were performed to examine the difference between Cry1Ah-resistant (ACB-AhR) and susceptible (ACB-BtS) strains of ACB at both transcriptional and translational levels. Results: A total of 7,007 differentially expressed genes (DEGs) and 182 differentially expressed proteins (DEPs) were identified between ACB-AhR and ACB-BtS and 90 genes had simultaneous transcription and translation profiles. Down-regulated genes associated with Cry1Ah resistance included aminopeptidase N, ABCC3, DIMBOA-induced cytochrome P450, alkaline phosphatase, glutathione S-transferase, cadherin-like protein, and V-ATPase. Whereas, anti-stress genes, such as heat shock protein 70 and carboxylesterase were up-regulated in ACB-AhR, displaying that a higher proportion of genes/proteins related to resistance was down-regulated compared to up-regulated. The Kyoto encyclopedia of genes and genomes (KEGG) analysis mapped 578 and 29 DEGs and DEPs, to 27 and 10 pathways, respectively (P < 0.05). Furthermore, real-time quantitative (qRT-PCR) results based on relative expression levels of randomly selected genes confirmed the "omics" response. Conclusion: Despite the previous studies, this is the first combination of a study using RNA-Seq and iTRAQ approaches on Cry1Ah-toxin binding, which led to the identification of longer length of unigenes in ACB. The DEGs and DEPs results are valuable for further clarifying Cry1Ah-mediated resistance.
Project description:The experiment was to determine the chronic effects of two transgenic maize lines that contained the mCry1Ac gene from the Bacillus thuringiensis strain (BT) and the maroACC gene from Agrobacterium tumefaciens strain (CC), respectively, on ileal microbiota of laying hens. Seventy-two laying hens were randomly assigned to one of the three dietary treatments for 12 weeks, as follows: (1) nontransgenic near-isoline maize-based diet (CT diet), (2) BT maize-based diet (BT diet), and (3) CC maize-based diet (CC diet). Ileum histological examination did not indicate a chronic effect of two transgenic maize diets. Few differences were observed in any bacterial taxa among the treatments that used high-throughput 16S rRNA gene sequencing. The only differences that were observed for bacterial genera were that Bifidobacterium belong within the Bifidobacteriaceae family tended to be greater (p = 0.114) abundant in hens fed the transgenic maize-based diet than in hens fed the CT diet. Birds that consumed the CC maize diet tended to have less abundance (p = 0.135) of Enterobacteriaceae family in the ileum than those that consumed the CT maize diet. These results indicate the lack of adverse effects of the BT maize and the CC maize lines on the ileal microbiota of hens for long term and provide important data regarding biosafety assessment of the transgenic maize lines.
Project description:BACKGROUND: Despite all the benefits assigned to the genetically modified plants, there are still no sufficient data available in literature concerning the possible effects on the microbial communities associated with these plants. Therefore, this study was aimed at examining the effects of the genetic modifications of two transgenic maize genotypes (MON810--expressing the insecticidal Bt-toxin and TC1507--expressing the insecticidal Bt-toxin and the herbicide resistance PAT [phosphinothricin-N-acetyltransferase]) on their endophytic microbial communities, in comparison to the microbial community found in the near-isogenic non-transgenic maize (control). RESULTS: The structure of the endophytic communities (Bacteria, Archaea and fungi) and their composition (Bacteria) were evaluated by denaturing gradient gel electrophoresis (DGGE) and the construction of clone libraries, respectively. DGGE analysis and the clone libraries of the bacterial community showed that genotype TC1507 slightly differed from the other two genotypes. Genotype TC1507 showed a higher diversity within its endophytic bacterial community when compared to the other genotypes. Although some bacterial genera were found in all genotypes, such as the genera Burkholderia, Achromobacer and Stenotrophomonas, some were unique to genotype TC1507. Moreover, OTUs associated with Enterobacter predominated only in TC1507 clone libraries. CONCLUSION: The endophytic bacterial community of the maize genotype TC1507 differed from the communities of the maize genotype MON810 and of their near-isogenic parental genotypes (non-Bt or control). The differences observed among the maize genotypes studied may be associated with insertion of the gene coding for the protein PAT present only in the transgenic genotype TC1507.
Project description:Possible non-target effect of transgenic cry1Ie maize exerts on natural enemy community biodiversity in the field is unresolved. In the present study, a 2-yr comparison of transgenic cry1Ie maize (Event IE09S034, Bt maize) and its near isoline (Zong 31, non-Bt maize) on natural enemy community biodiversity were compared with whole plant inspections, pitfall traps and suction sampler. Natural enemy diversity indices (Shannon-Wiener', Simpson's and Pielou's index) and abundance suggested there were no significant differences between the two types of maize. The only exceptions were the Pielou's index for whole plant inspections in 2013 and abundance for pitfall traps in 2012, which were significantly higher in Bt maize than those of non-Bt maize. The main species of natural enemies were identical in Bt and non-Bt maize plots for each method and the three methods combined. For whole plant inspections, Bt maize had no time-dependent effect on the entire arthropod natural enemy community, and also no effect on community dissimilarities between Bt and non-Bt maize plots. These results suggested that despite the presence of a relatively minor difference in natural enemy communities between Bt and non-Bt maize, transgenic cry1Ie maize had little, if any, effect on natural enemy community biodiversity.
Project description:BACKGROUND: Bt-maize is a transgenic variety of maize expressing the Cry toxin from Bacillus turingiensis. The potential accumulation of the relative effect of the transgenic modification and the cry toxin on the rhizobacterial communities of Bt-maize has been monitored over a period of four years. METHODOLOGY/PRINCIPAL FINDINGS: The accumulative effects of the cultivation of this transgenic plant have been monitored by means of high throughput DNA pyrosequencing of the bacterial DNA coding for the 16S rRNA hypervariable V6 region from rhizobacterial communities. The obtained sequences were subjected to taxonomic, phylogenetic and taxonomic-independent diversity studies. The results obtained were consistent, indicating that variations detected in the rhizobacterial community structure were possibly due to climatic factors rather than to the presence of the Bt-gene. No variations were observed in the diversity estimates between non-Bt and Bt-maize. CONCLUSIONS/SIGNIFICANCE: The cultivation of Bt-maize during the four-year period did not change the maize rhizobacterial communities when compared to those of the non-Bt maize. This is the first study to be conducted with Bt-maize during such a long cultivation period and the first evaluation of rhizobacterial communities to be performed in this transgenic plant using Next Generation Sequencing.
Project description:Agricultural practices are known to alter bulk soil microbial communities, but little is known about the effect of such practices on the plant endophytic community. We assessed the influence of long-term applications (20 years) of herbicides and different fertilizer types on the endophytic community of maize plants grown in different field experiments. Nested PCR-denaturing gradient gel electrophoresis (DGGE) analyses targeting general bacteria, type I or II methanotrophs, actinomycetes, and general fungi were used to fingerprint the endophytic community in the roots of Zea mays L. Low intraplant variability (reproducible DGGE patterns) was observed for the bacterial, type I methanotroph, and fungal communities, whereas the patterns for endophytic actinomycetes exhibited high intraplant variability. No endophytic amplification product was obtained for type II methanotrophs. Cluster and stability analysis of the endophytic type I methanotroph patterns differentiated maize plants cultivated by using mineral fertilizer from plants cultivated by using organic fertilizer with a 100% success rate. In addition, lower methanotroph richness was observed for mineral-fertilized plants than for organically fertilized plants. The use of herbicides could not be traced by fingerprinting the endophytic type I methanotrophs or by evaluating any other endophytic microbial group. Our results indicate that the effect of agrochemicals is not limited to the bulk microbial community but also includes the root endophytic community. It is not clear if this effect is due to a direct effect on the root endophytic community or is due to changes in the bulk community, which are then reflected in the root endophytic community.
Project description:Higher yields and reduced pesticide impacts are needed to mitigate the effects of agricultural intensification. A 2-year farm-scale evaluation of 81 commercial fields in Arizona show that use of transgenic Bacillus thuringiensis (Bt) cotton reduced insecticide use, whereas transgenic cotton with Bt protein and herbicide resistance (BtHr) did not affect herbicide use. Transgenic cotton had higher yield than nontransgenic cotton for any given number of insecticide applications. However, nontransgenic, Bt and BtHr cotton had similar yields overall, largely because higher insecticide use with nontransgenic cotton improved control of key pests. Unlike Bt and BtHr cotton, insecticides reduced the diversity of nontarget insects. Several other agronomic and ecological factors also affected biodiversity. Nevertheless, pairwise comparisons of diversity of nontarget insects in cotton fields with diversity in adjacent noncultivated sites revealed similar effects of cultivation of transgenic and nontransgenic cotton on biodiversity. The results indicate that impacts of agricultural intensification can be reduced when replacement of broad-spectrum insecticides by narrow-spectrum Bt crops does not reduce control of pests not affected by Bt crops.
Project description:Diazotrophic bacteria can reduce N2 into plant-available ammonium (NH4 +), promoting plant growth and reducing nitrogen (N) fertilizer requirements. However, there are few systematic studies on the effects of diazotrophic bacteria on biological N2 fixation (BNF) contribution rate and host plant N uptake and metabolism. In this study, the interactions of the diazotrophic Paenibacillus beijingensis BJ-18 with wheat, maize, and cucumber were investigated when it was inoculated to these plant seedlings grown in both low N and high N soils, with un-inoculated plants as controls. This study showed that GFP-tagged P. beijingensis BJ-18 colonized inside and outside seedlings, forming rhizospheric and endophytic colonies in roots, stems, and leaves. The numbers of this bacterium in the inoculated plants depended on soil N levels. Under low N, inoculation significantly increased shoot dry weight (wheat 86.1%, maize 46.6%, and cucumber 103.6%) and root dry weight (wheat 46.0%, maize 47.5%, and cucumber 20.3%). The 15N-isotope-enrichment experiment indicated that plant seedlings derived 12.9-36.4% N from BNF. The transcript levels of nifH in the inoculated plants were 0.75-1.61 folds higher in low N soil than those in high N soil. Inoculation enhanced NH4 + and nitrate (NO3 -) uptake from soil especially under low N. The total N in the inoculated plants were increased by 49.1-92.3% under low N and by 13-15.5% under high N. Inoculation enhanced activities of glutamine synthetase (GS) and nitrate reductase (NR) in plants, especially under low N. The expression levels of N uptake and N metabolism genes: AMT (ammonium transporter), NRT (nitrate transporter), NiR (nitrite reductase), NR, GS and GOGAT (glutamate synthase) in the inoculated plants grown under low N were up-regulated 1.5-91.9 folds, but they were not obviously changed under high N. Taken together, P. beijingensis BJ-18 was an effective, endophytic and diazotrophic bacterium. This bacterium contributed to plants with fixed N2, promoted plant growth and N uptake, and enhanced gene expression and enzyme activities involved in N uptake and assimilation in plants. However, these positive effects on plants were regulated by soil N status. This study might provide insight into the interactions of plants with beneficial associative and endophytic diazotrophic bacteria.
Project description:A polyphasic approach has been developed to gain knowledge of suitable key indicators for the evaluation of environmental impact of genetically modified Bt 11 and Bt 176 corn lines on soil ecosystems. We assessed the effects of Bt corn (which constitutively expresses the insecticidal toxin from Bacillus thuringiensis, encoded by the truncated Cry1Ab gene) and non-Bt corn plants and their residues on rhizospheric and bulk soil eubacterial communities by means of denaturing gradient gel electrophoresis analyses of 16S rRNA genes, on the nontarget mycorrhizal symbiont Glomus mosseae, and on soil respiration. Microcosm experiments showed differences in rhizospheric eubacterial communities associated with the three corn lines and a significantly lower level of mycorrhizal colonization in Bt 176 corn roots. In greenhouse experiments, differences between Bt and non-Bt corn plants were detected in rhizospheric eubacterial communities (both total and active), in culturable rhizospheric heterotrophic bacteria, and in mycorrhizal colonization. Plant residues of transgenic plants, plowed under at harvest and kept mixed with soil for up to 4 months, affected soil respiration, bacterial communities, and mycorrhizal establishment by indigenous endophytes. The multimodal approach utilized in our work may be applied in long-term field studies aimed at monitoring the real hazard of genetically modified crops and their residues on nontarget soil microbial communities.