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Berlin Brandenburg

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  • 1
    In: Molecular Microbiology, January 2010, Vol.75(2), pp.261-263
    Description: Integrative conjugative elements (ICEs) occur frequently in Gram‐positive and Gram‐negative bacteria. In contrast to plasmids, they are stably integrated in the bacterial genome, often inserted in a tRNA gene. They are excised from the host chromosome upon induction in order to be transferred to a recipient cell. When conjugative transfer is completed, they stably reintegrate in the chromosome. It is generally thought that ICEs are incapable of autonomous replication, instead relying on replication and segregation along with the host chromosome. In this issue of Lee and co‐workers demonstrate that ICE from is capable of autonomous plasmid‐like replication in its circular form after excision. The authors show that ICE replication is unidirectional; it initiates at and requires the ICE‐encoded conjugative relaxase NicK. Replication also requires the catalytic subunit of the host DNA polymerase PolC, the host processivity clamp DnaN and the host‐encoded alternative helicase PcrA. Autonomous replication of ICE appears to be important for its stable maintenance, but not for horizontal transfer of the element. Lee and co‐workers argue that plasmid‐like replication is likely a common property of ICEs, probably contributing to stability and maintenance of ICEs in bacterial populations. I discuss these findings in context with data on other ICEs from Gram‐positive and Gram‐negative bacteria and with respect to possible consequences of the findings for basic research on mobile genetic elements from Gram‐positive bacteria and their applications in biotechnology.
    Keywords: Transfer Rna ; Transposons;
    ISSN: 0950-382X
    E-ISSN: 1365-2958
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  • 2
    In: The Journal of Bacteriology, 2010, Vol. 192(1), p.23
    Description: Similarities of ICEBs1 with other ICEs are highlighted, the intriguing regulation of conjugative transfer of ICEBs1 as well as the role of putative key players in the process are discussed. The commentary ends with perspectives on potential applications of ICEBs1 in prokaryotic genetics/genetic engineering.
    Keywords: Biology;
    ISSN: 0021-9193
    ISSN: 00219193
    E-ISSN: 10985530
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  • 3
    Language: English
    In: Chemosphere, June 2011, Vol.84(1), pp.175-181
    Description: ► Bacteria were isolated from soil and identified by 16S rDNA sequencing. ► MICs of all the isolates were determined for pesticides and heavy metals. ► Bacterial isolates were tested for the presence of Inc group plasmids. ► DNA from bacteria and soil revealed the presence of IncP-specific sequences. ► IncP plasmids are responsible for transferring resistance genes among bacterial species. A total of 35 bacteria from contaminated soil (cultivated fields) near pesticide industry from Chinhat, Lucknow, (India) were isolated and tested for their tolerance/resistance to pesticides, heavy metals and antibiotics. Bacterial isolates were identified by 16S rDNA sequencing. Gas Chromatography analysis of the soil samples revealed the presence of lindane at a concentration of 547 ng g and α-endosulfan and β-endosulfan of 422 ng g and 421 ng g respectively. Atomic Absorption Spectrophotometry analysis of the test sample was done and Cr, Zn, Ni, Fe, Cu and Cd were detected at concentrations of 36.2, 42.5, 43.2, 241, 13.3 and 11.20 mg kg respectively. Minimum inhibitory concentrations of all the isolates were determined for pesticides and heavy metals. All the multi-resistant/tolerant bacterial isolates were also tested for the presence of incompatibility (Inc) group IncP, IncN, IncW, IncQ plasmids and for rolling circle plasmids of the pMV158-family by PCR. Total community DNA was extracted from pesticide contaminated soil. PCR amplification of the bacterial isolates and soil DNA revealed the presence of IncP-specific sequences ( and which was confirmed by dot blot hybridization with RP4-derived DIG-labelled probes. Plasmids belonging to IncN, IncW and IncQ group were neither detected in the bacterial isolates nor in total soil DNA. The presence of conjugative or mobilizable IncP plasmids in the isolates indicate that these bacteria have gene transfer capacity with implications for dissemination of heavy metal and antibiotic resistance genes. We propose that IncP plasmids are mainly responsible for the spread of multi-resistant bacteria in the contaminated soils.
    Keywords: Conjugative Plasmids ; Pesticide Tolerance ; Antibiotic Resistance ; Metal Resistance ; Alluvial Soil ; Chemistry ; Ecology
    ISSN: 0045-6535
    E-ISSN: 1879-1298
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  • 4
    Language: English
    In: Chemosphere, February 2017, Vol.168, pp.1637-1647
    Description: Poultry manure is a nitrogen rich fertilizer, which is usually recycled and spread on agricultural fields. Due to its high nutrient content, chicken manure is considered to be one of the most valuable animal wastes as organic fertilizer. However, when chicken litter is applied in its native form, concerns are raised as such fertilizers also include high amounts of antibiotic resistant pathogenic and heavy metals. We studied the impact of an anaerobic thermophilic digestion process on poultry manure. Particularly, microbial antibiotic resistance profiles, mobile genetic elements promoting the resistance dissemination in the environment as well as the presence of heavy metals were focused in this study. The initiated heat treatment fostered a community shift from pathogenic to less pathogenic bacterial groups. Phenotypic and molecular studies demonstrated a clear reduction of multiple resistant pathogens and self-transmissible plasmids in the heat treated manure. That treatment also induced a higher release of metals and macroelements. Especially, Zn and Cu exceeded toxic thresholds. Although the concentrations of a few metals reached toxic levels after the anaerobic thermophilic treatment, the quality of poultry manure as organic fertilizer may raise significantly due to the elimination of antibiotic resistance genes (ARG) and self-transmissible plasmids.
    Keywords: Anaerobic Digestion ; Poultry Manure ; Antibiotic Resistance Genes ; Metals ; Conjugative Plasmids ; Thermophilic Temperature ; Chemistry ; Ecology
    ISSN: 0045-6535
    E-ISSN: 1879-1298
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  • 5
    Language: English
    In: Applied and environmental microbiology, September 2014, Vol.80(17), pp.5282-91
    Description: Wastewater contains large amounts of pharmaceuticals, pathogens, and antimicrobial resistance determinants. Only a little is known about the dissemination of resistance determinants and changes in soil microbial communities affected by wastewater irrigation. Community DNAs from Mezquital Valley soils under irrigation with untreated wastewater for 0 to 100 years were analyzed by quantitative real-time PCR for the presence of sul genes, encoding resistance to sulfonamides. Amplicon sequencing of bacterial 16S rRNA genes from community DNAs from soils irrigated for 0, 8, 10, 85, and 100 years was performed and revealed a 14% increase of the relative abundance of Proteobacteria in rainy season soils and a 26.7% increase in dry season soils for soils irrigated for 100 years with wastewater. In particular, Gammaproteobacteria, including potential pathogens, such as Pseudomonas, Stenotrophomonas, and Acinetobacter spp., were found in wastewater-irrigated fields. 16S rRNA gene sequencing of 96 isolates from soils irrigated with wastewater for 100 years (48 from dry and 48 from rainy season soils) revealed that 46% were affiliated with the Gammaproteobacteria (mainly potentially pathogenic Stenotrophomonas strains) and 50% with the Bacilli, whereas all 96 isolates from rain-fed soils (48 from dry and 48 from rainy season soils) were affiliated with the Bacilli. Up to six types of antibiotic resistance were found in isolates from wastewater-irrigated soils; sulfamethoxazole resistance was the most abundant (33.3% of the isolates), followed by oxacillin resistance (21.9% of the isolates). In summary, we detected an increase of potentially harmful bacteria and a larger incidence of resistance determinants in wastewater-irrigated soils, which might result in health risks for farm workers and consumers of wastewater-irrigated crops.
    Keywords: Biota ; Soil Microbiology ; Waste Water ; Agricultural Irrigation -- Methods ; Bacteria -- Classification
    ISSN: 00992240
    E-ISSN: 1098-5336
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  • 6
    Language: English
    In: Applied and Environmental Microbiology, 2012, Vol. 78(3), p.895
    Description: On the basis of pIP501, a green fluorescent protein (GFP)-tagged monitoring tool was constructed for quantifying plasmid mobilization among Gram-positive bacteria and between Gram-positive Enterococcus faecalis and Gram-negative Escherichia coli. Furthermore, retromobilization of the GFP-tagged monitoring tool was shown from E. faecalis OG1X into the clinical isolate E. faecalis T9.
    Keywords: Conjugation, Genetic ; Gene Transfer, Horizontal ; Enterococcus Faecalis -- Genetics ; Escherichia Coli -- Genetics;
    ISSN: 1098-5336
    ISSN: 10985336
    ISSN: 00992240
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  • 7
    Language: English
    In: 2012, Vol.7(9), p.e45397
    Description: Irrigation with wastewater releases pharmaceuticals, pathogenic bacteria, and resistance genes, but little is known about the accumulation of these contaminants in the environment when wastewater is applied for decades. We sampled a chronosequence of soils that were variously irrigated with wastewater from zero up to 100 years in the Mezquital Valley, Mexico, and investigated the accumulation of ciprofloxacin, enrofloxacin, sulfamethoxazole, trimethoprim, clarithromycin, carbamazepine, bezafibrate, naproxen, diclofenac, as well as the occurrence of Enterococcus spp., and sul and qnr resistance genes. Total concentrations of ciprofloxacin, sulfamethoxazole, and carbamazepine increased with irrigation duration reaching 95% of their upper limit of 1.4 µg/kg (ciprofloxacin), 4.3 µg/kg (sulfamethoxazole), and 5.4 µg/kg (carbamazepine) in soils irrigated for 19–28 years. Accumulation was soil-type-specific, with largest accumulation rates in Leptosols and no time-trend in Vertisols. Acidic pharmaceuticals (diclofenac, naproxen, bezafibrate) were not retained and thus did not accumulate in soils. We did not detect qnrA genes, but qnrS and qnrB genes were found in two of the irrigated soils. Relative concentrations of sul1 genes in irrigated soils were two orders of magnitude larger (3.15×10 −3 ±0.22×10 −3 copies/16S rDNA) than in non-irrigated soils (4.35×10 −5 ±1.00×10 −5 copies/16S rDNA), while those of sul2 exceeded the ones in non-irrigated soils still by a factor of 22 (6.61×10 –4 ±0.59×10 −4 versus 2.99×10 −5 ±0.26×10 −5 copies/16S rDNA). Absolute numbers of sul genes continued to increase with prolonging irrigation together with Enterococcus spp. 23S rDNA and total 16S rDNA contents. Increasing total concentrations of antibiotics in soil are not accompanied by increasing relative abundances of resistance genes. Nevertheless, wastewater irrigation enlarges the absolute concentration of resistance genes in soils due to a long-term increase in total microbial biomass.
    Keywords: Research Article ; Agriculture ; Biology ; Chemistry ; Earth Sciences ; Engineering ; Medicine ; Chemistry ; Microbiology ; Ecology ; Pharmacology
    E-ISSN: 1932-6203
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  • 8
    In: PLoS ONE, 2014, Vol.9(10)
    Description: Background Enterococci are the third most common cause of healthcare-associated infections, which include urinary tract infections, bacteremia and endocarditis. Cell-surface structures such as lipoteichoic acid (LTA) have been poorly examined in E. faecalis , especially with respect to urinary tract infections (UTIs). The dlt operon is responsible for the D-alanylation of LTA and includes the gene dlt A, which encodes the D-alanyl carrier protein ligase (Dcl). The involvement of LTA in UTI infection by E. faecalis has not been studied so far. Here, we examined the role of teichoic acid alanylation in the adhesion of enterococci to uroepithelial cells. Results In a mouse model of urinary tract infection, we showed that E. faecalis 12030Δ dlt A mutant colonizes uroepithelial surfaces more efficiently than wild type bacteria. We also demonstrated that this mutant adhered four fold better to human bladder carcinoma cell line T24 compared to the wild type strain. Bacterial adherence could be significantly inhibited by purified lipoteichoic acid (LTA) and inhibition was specific. Conclusion In contrast to bacteraemia model and adherence to colon surfaces, E. faecalis 12030Δ dlt A mutant colonized uroepithelial surfaces more efficiently than wild-type bacteria. In the case of the uroepithelial surface the adherence to specific host cells could be prevented by purified LTA. Our results therefore suggest a novel function of alanylation of LTA in E. faecalis .
    Keywords: Research Article ; Biology And Life Sciences ; Medicine And Health Sciences
    E-ISSN: 1932-6203
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  • 9
    Language: English
    In: The Journal of biological chemistry, 18 January 2013, Vol.288(3), pp.2018-28
    Description: Conjugative plasmid transfer is the most important means of spreading antibiotic resistance and virulence genes among bacteria and therefore presents a serious threat to human health. The process requires direct cell-cell contact made possible by a multiprotein complex that spans cellular membranes and serves as a channel for macromolecular secretion. Thus far, well studied conjugative type IV secretion systems (T4SS) are of Gram-negative (G-) origin. Although many medically relevant pathogens (e.g., enterococci, staphylococci, and streptococci) are Gram-positive (G+), their conjugation systems have received little attention. This study provides structural information for the transfer protein TraM of the G+ broad host range Enterococcus conjugative plasmid pIP501. Immunolocalization demonstrated that the protein localizes to the cell wall. We then used opsonophagocytosis as a novel tool to verify that TraM was exposed on the cell surface. In these assays, antibodies generated to TraM recruited macrophages and enabled killing of pIP501 harboring Enteroccocus faecalis cells. The crystal structure of the C-terminal, surface-exposed domain of TraM was determined to 2.5 Å resolution. The structure, molecular dynamics, and cross-linking studies indicated that a TraM trimer acts as the biological unit. Despite the absence of sequence-based similarity, TraM unexpectedly displayed a fold similar to the T4SS VirB8 proteins from Agrobacterium tumefaciens and Brucella suis (G-) and to the transfer protein TcpC from Clostridium perfringens plasmid pCW3 (G+). Based on the alignments of secondary structure elements of VirB8-like proteins from mobile genetic elements and chromosomally encoded T4SS from G+ and G- bacteria, we propose a new classification scheme of VirB8-like proteins.
    Keywords: Conjugation, Genetic ; Bacterial Proteins -- Chemistry ; Cell Wall -- Genetics ; Enterococcus Faecalis -- Genetics ; Plasmids -- Genetics ; Virulence Factors -- Chemistry
    ISSN: 00219258
    E-ISSN: 1083-351X
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  • 10
    Language: English
    In: Journal of bacteriology, October 2013, Vol.195(19), pp.4436-44
    Description: pIP501 is a conjugative broad-host-range plasmid frequently present in nosocomial Enterococcus faecalis and Enterococcus faecium isolates. We focus here on the functional analysis of the type IV secretion gene traG, which was found to be essential for pIP501 conjugative transfer between Gram-positive bacteria. The TraG protein, which localizes to the cell envelope of E. faecalis harboring pIP501, was expressed and purified without its N-terminal transmembrane helix (TraGΔTMH) and shown to possess peptidoglycan-degrading activity. TraGΔTMH was inhibited by specific lytic transglycosylase inhibitors hexa-N-acetylchitohexaose and bulgecin A. Analysis of the TraG sequence suggested the presence of two domains which both could contribute to the observed cell wall-degrading activity: an N-terminal soluble lytic transglycosylase domain (SLT) and a C-terminal cysteine-, histidine-dependent amidohydrolases/peptidases (CHAP) domain. The protein domains were expressed separately, and both degraded peptidoglycan. A change of the conserved glutamate residue in the putative catalytic center of the SLT domain (E87) to glycine resulted in almost complete inactivity, which is consistent with this part of TraG being a predicted lytic transglycosylase. Based on our findings, we propose that TraG locally opens the peptidoglycan to facilitate insertion of the Gram-positive bacterial type IV secretion machinery into the cell envelope.
    Keywords: Bacterial Proteins -- Metabolism ; Enterococcus Faecalis -- Enzymology ; Enterococcus Faecium -- Enzymology ; Gene Expression Regulation, Bacterial -- Physiology ; Gene Expression Regulation, Enzymologic -- Physiology ; Peptidoglycan -- Metabolism
    ISSN: 00219193
    E-ISSN: 1098-5530
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