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  • 1
    Language: English
    In: Applied and environmental microbiology, February 2013, Vol.79(4), pp.1410-3
    Description: To study the role of broad-host-range IncP-1 plasmids in bacterial adaptability to irregular environmental challenges, a quantitative real-time PCR assay was developed that specifically detects the korB gene, which is conserved in all IncP-1 plasmids, in environmental samples. IncP-1 plasmid dynamics in a biopurification system for pesticide wastes were analyzed.
    Keywords: Environmental Microbiology ; Bacteroidetes -- Genetics ; Plasmids -- Analysis
    ISSN: 00992240
    E-ISSN: 1098-5336
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  • 2
    Language: English
    In: Applied Microbiology and Biotechnology, 2016, Vol.100(21), pp.9343-9353
    Description: Pig manures are frequently used as fertilizer or co-substrate in biogas plants (BGPs) and typically contain antibiotic residues (ARs), as well as bacteria carrying resistance genes (RGs) and mobile genetic elements (MGEs). A survey of manures from eight pig fattening and six pig breeding farms and digestates from eight BGPs in Lower Saxony, Germany was conducted to evaluate the link between antibiotic usage and ARs to RGs and MGEs present in organic fertilizers. In total, 11 different antibiotics belonging to six substance classes were applied in the farms investigated. Residue analysis revealed concentrations of tetracycline up to 300 mg kg −1 dry weight (DW) in manures and of doxycycline up to 10.1 mg kg −1 DW in digestates indicating incomplete removal during anaerobic digestion. RGs ( sul1 , sul2 , tet (A), tet (M), tet (X), qacE ∆ 1 ) were detected in total community DNA of all samples by PCR-Southern blot hybridization. Broad-host range plasmids (IncP-1, IncQ, IncN, and IncW) and integron integrase genes ( intI1 , intI2 ) were found in most manure samples with IncN and IncW plasmids being more abundant in manure from pig breeding compared to pig fattening farms. IntI1 , IncQ, and IncW plasmids were also detected in all digestates, while IncP-1, IncN, and LowGC plasmids were detected only sporadically. Our findings strongly reinforce the need for further research to identify mitigation strategies to reduce the level of contamination of organic fertilizers with ARs and transferable RGs that are applied to soil and that might influence the mobile resistome of the plant microbiome.
    Keywords: Antibiotic resistance genes ; Mobile genetic elements ; Antibiotics ; Pig husbandry ; Manures ; Digestates
    ISSN: 0175-7598
    E-ISSN: 1432-0614
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  • 3
    Language: English
    In: Applied Microbiology and Biotechnology, 2017, Vol.101(11), pp.4815-4825
    Description: On-farm biopurification systems (BPSs) represent an efficient technology for treating pesticide-contaminated wastewater. Biodegradation by genetically adapted bacteria has been suggested to perform a major contribution to the removal of pesticides in BPSs. Recently, several studies pointed to the role of IncP-1 plasmids in the degradation of pesticides in BPSs but this was never linked with catabolic markers. Therefore, a microcosm experiment was conducted in order to examine whether changes in mobile genetic element (MGE) abundances in response to the application of phenylurea herbicide linuron are linked with changes in catabolic genes. Denaturing gradient gel electrophoresis (DGGE) fingerprints of 16S ribosomal RNA gene fragments amplified from total community (TC)-DNA suggested significant shifts in the bacterial community composition. PCR-Southern blot-based detection of genes involved in linuron hydrolysis ( libA and hylA ) or degradation of its metabolite 3,4-dichloroaniline ( dcaQ I , dcaQ II , and ccdC ) in TC-DNA showed that the abundance of the hylA gene was increased faster and stronger in response to linuron application than that of the libA gene, and that the dcaQ II gene was more abundant than the isofunctional gene dcaQ I 20 and 60 days after linuron addition. Furthermore, a significant increase in the relative abundance of the IncP-1-specific korB gene in response to linuron was recorded. Our data suggest that different bacterial populations bearing isofunctional genes coding for enzymes degrading linuron seemed to be enriched in BPSs in response to linuron and that IncP-1 plasmids might be involved in their dissemination.
    Keywords: Total community DNA ; 16S rRNA genes ; Degradative genes ; PCR hybridization ; Plasmids
    ISSN: 0175-7598
    E-ISSN: 1432-0614
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  • 4
    Language: English
    In: Journal of Hazardous Materials, 2010, Vol.184(1), pp.523-532
    Description: Twenty-six different plant species were analyzed regarding their performance in soil contaminated with petroleum oil. Two well-performing species, Italian ryegrass ( var. Taurus), Birdsfoot trefoil ( var. Leo) and the combination of these two plants were selected to study the ecology of plant-associated, culturable alkane-degrading bacteria. Hydrocarbon degrading bacteria were isolated from the rhizosphere, root interior and shoot interior and subjected to the analysis of 16S rRNA gene, the 16S and 23S rRNA intergenic spacer region and alkane hydroxylase genes. Furthermore, we investigated whether alkane hydroxylase genes are plasmid located. Higher numbers of culturable, alkane-degrading bacteria were associated with Italian ryegrass, which were also characterized by a higher diversity, particularly in the plant interior. Only half of the isolated bacteria hosted known alkane hydroxylase genes ( and cytochrome P153-like). Degradation genes were found both on plasmids as well as in the chromosome. In regard to application of plants for rhizodegradation, where support of numerous degrading bacteria is essential for efficient break-down of pollutants, Italian ryegrass seems to be more appropriate than Birdsfoot trefoil.
    Keywords: Alkane Hydroxylase ; 16s Rrna Gene ; Rhizosphere ; Endophytes ; Italian Ryegrass ; Birdsfoot Trefoil ; Engineering ; Law
    ISSN: 0304-3894
    E-ISSN: 1873-3336
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  • 5
    Language: English
    In: Applied and environmental microbiology, July 2012, Vol.78(14), pp.4933-41
    Description: Bananas are among the most widely consumed foods in the world. In Uganda, the country with the second largest banana production in the world, bananas are the most important staple food. The objective of this study was to analyze banana-associated microorganisms and to select efficient antagonists against fungal pathogens which are responsible for substantial yield losses. We studied the structure and function of microbial communities (endosphere, rhizosphere, and soil) obtained from three different traditional farms in Uganda by cultivation-independent (PCR-SSCP fingerprints of 16S rRNA/ITS genes, pyrosequencing of enterobacterial 16S rRNA gene fragments, quantitative PCR, fluorescence in situ hybridization coupled with confocal laser scanning microscopy, and PCR-based detection of broad-host-range plasmids and sulfonamide resistance genes) and cultivation-dependent methods. The results showed microhabitat-specific microbial communities that were significant across sites and treatments. Furthermore, all microhabitats contained a high number and broad spectrum of indigenous antagonists toward identified fungal pathogens. While bacterial antagonists were found to be enriched in banana plants, fungal antagonists were less abundant and mainly found in soil. The banana stem endosphere was the habitat with the highest bacterial counts (up to 10(9) gene copy numbers g(-1)). Here, enterics were found to be enhanced in abundance and diversity; they provided one-third of the bacteria and were identified by pyrosequencing with 14 genera, including not only potential human (Escherichia, Klebsiella, Salmonella, and Yersinia spp.) and plant (Pectobacterium spp.) pathogens but also disease-suppressive bacteria (Serratia spp.). The dominant role of enterics can be explained by the permanent nature and vegetative propagation of banana and the amendments of human, as well as animal, manure in these traditional cultivations.
    Keywords: Rhizosphere ; Soil Microbiology ; Crops, Agricultural -- Microbiology ; Enterobacteriaceae -- Isolation & Purification ; Fungi -- Isolation & Purification ; Musa -- Microbiology
    ISSN: 00992240
    E-ISSN: 1098-5336
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  • 6
    Language: English
    In: Applied and environmental microbiology, December 2012, Vol.78(23), pp.8492-7
    Description: Leaf lesions of Mandevilla sanderi were shown to be caused by Pseudomonas savastanoi. While BOX fingerprints were similar for P. savastanoi isolates from different host plants, plasmid restriction patterns and sequencing of plasmid-located pathogenicity determinants revealed that Mandevilla isolates contained similar plasmids distinct from those of other isolates. A repA-based detection method was established.
    Keywords: Apocynaceae -- Microbiology ; Plant Diseases -- Microbiology ; Pseudomonas -- Isolation & Purification
    ISSN: 00992240
    E-ISSN: 1098-5336
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  • 7
    Language: English
    In: Applied and environmental microbiology, August 2012, Vol.78(16), pp.5520-8
    Description: Mangroves are complex ecosystems that regulate nutrient and sediment fluxes to the open sea. The importance of bacteria and fungi in regulating nutrient cycles has led to an interest in their diversity and composition in mangroves. However, very few studies have assessed Archaea in mangroves, and virtually nothing is known about whether mangrove rhizospheres affect archaeal diversity and composition. Here, we studied the diversity and composition of Archaea in mangrove bulk sediment and the rhizospheres of two mangrove trees, Rhizophora mangle and Laguncularia racemosa, using denaturing gradient gel electrophoresis (DGGE) and pyrosequencing of archaeal 16S rRNA genes with a nested-amplification approach. DGGE profiles revealed significant structural differences between bulk sediment and rhizosphere samples, suggesting that roots of both mangrove species influence the sediment archaeal community. Nearly all of the detected sequences obtained with pyrosequencing were identified as Archaea, but most were unclassified at the level of phylum or below. Archaeal richness was, furthermore, the highest in the L. racemosa rhizosphere, intermediate in bulk sediment, and the lowest in the R. mangle rhizosphere. This study shows that rhizosphere microhabitats of R. mangle and L. racemosa, common plants in subtropical mangroves located in Rio de Janeiro, Brazil, hosted distinct archaeal assemblages.
    Keywords: Biodiversity ; DNA Barcoding, Taxonomic ; Denaturing Gradient Gel Electrophoresis ; Archaea -- Classification ; Combretaceae -- Microbiology ; Plant Roots -- Microbiology ; Rhizophoraceae -- Microbiology
    ISSN: 00992240
    E-ISSN: 1098-5336
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  • 8
    Language: English
    In: Applied and environmental microbiology, April 2011, Vol.77(7), pp.2527-30
    Description: Two soils were amended three times with pig manure. The abundance of sulfonamide resistance genes was determined by quantitative PCR 2 months after each application. In both soils treated with sulfadiazine-containing manure, the numbers of copies of sul1 and sul2 significantly increased compared to numbers after treatments with antibiotic-free manure or a control and accumulated with repeated applications.
    Keywords: Drug Resistance, Bacterial ; Genes, Bacterial ; Manure ; Metagenome ; Soil Microbiology ; Anti-Bacterial Agents -- Pharmacology ; Sulfadiazine -- Pharmacology
    ISSN: 00992240
    E-ISSN: 1098-5336
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  • 9
    Language: English
    In: Applied and environmental microbiology, March 2013, Vol.79(5), pp.1704-11
    Description: Spreading manure containing antibiotics in agriculture is assumed to stimulate the dissemination of antibiotic resistance in soil bacterial populations. Plant roots influencing the soil environment and its microflora by exudation of growth substrates might considerably increase this effect. In this study, the effects of manure from pigs treated with sulfadiazine (SDZ), here called SDZ manure, on the abundance and transferability of sulfonamide resistance genes sul1 and sul2 in the rhizosphere of maize and grass were compared to the effects in bulk soil in a field experiment. In plots that repeatedly received SDZ manure, a significantly higher abundance of both sul genes was detected compared to that in plots where manure from untreated pigs was applied. Significantly lower abundances of sul genes relative to bacterial ribosomal genes were encountered in the rhizosphere than in bulk soil. However, in contrast to results for bulk soil, the sul gene abundance in the SDZ manure-treated rhizosphere constantly deviated from control treatments over a period of 6 weeks after manuring, suggesting ongoing antibiotic selection over this period. Transferability of sulfonamide resistance was analyzed by capturing resistance plasmids from soil communities into Escherichia coli. Increased rates of plasmid capture were observed in samples from SDZ manure-treated bulk soil and the rhizosphere of maize and grass. More than 97% of the captured plasmids belonged to the LowGC type (having low G+C content), giving further evidence for their important contribution to the environmental spread of antibiotic resistance. In conclusion, differences between bulk soil and rhizosphere need to be considered when assessing the risks associated with the spreading of antibiotic resistance.
    Keywords: Drug Resistance, Bacterial ; Gene Transfer, Horizontal ; Manure ; Soil Microbiology ; Anti-Bacterial Agents -- Therapeutic Use ; Sulfadiazine -- Therapeutic Use
    ISSN: 00992240
    E-ISSN: 1098-5336
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  • 10
    In: Applied and Environmental Microbiology, 2010, Vol. 76(14), p.4765
    Description: A novel PCR primer system that targets a wide range of polycyclic aromatic hydrocarbon ring-hydroxylating dioxygenase (PAH-RHD(alpha)) genes of both Gram-positive and Gram-negative bacteria was developed and used to study their abundance and diversity in two different soils in response to phenanthrene spiking. The specificities and target ranges of the primers predicted in silico were confirmed experimentally by cloning and sequencing of PAH-RHD(alpha) gene amplicons from soil DNA. Cloning and sequencing showed the dominance of phnAc genes in the contaminated Luvisol. In contrast, high diversity of PAH-RHD(alpha) genes of Gram-positive and Gram-negative bacteria was observed in the phenanthrene-spiked Cambisol. Quantitative real-time PCR based on the same primers revealed that 63 days after phenanthrene spiking, PAH-RHD(alpha) genes were 1 order of magnitude more abundant in the Luvisol than in the Cambisol, while they were not detected in both control soils. In conclusion, sequence analysis of the amplicons obtained confirmed the specificity of the novel primer system and revealed a soil type-dependent response of PAH-RHD(alpha) gene-carrying soil bacteria to phenanthrene spiking.
    Keywords: Biodiversity ; Metagenome ; Soil Microbiology ; Bacteria -- Classification ; Bacterial Proteins -- Genetics ; Dioxygenases -- Genetics ; Phenanthrenes -- Metabolism;
    ISSN: 0099-2240
    ISSN: 00992240
    E-ISSN: 10985336
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