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M Oliva, R Monno, P Addabbo, G Pesole, M Scrascia, C Calia, A M Dionisi, M Chiara, D S Horner, C Manzari, C Pazzani
In the present study we report the identification of a sul3-associated class 1 integron containing the dfrA12-orfF-aadA2-cmlA1-aadA1-qacH array embedded in a Tn21-derived element that is part of a conjugative FII plasmid named pST1007-1A. The plasmid was identified in the Salmonella Typhimurium strain ST1007, a member of a clinically relevant clonal MDR lineage diffuse in Italy. ST1007 exhibited resistance to ampicillin, chloramphenicol, streptomycin, sulphamethoxazole, tetracycline and trimethoprim encoded by blaTEM-1 , cmlA1, (aadA1, aadA2, strAB), (sul2, sul3), tet(B) and dfrA12 genes, respectively...
October 16, 2018: Plasmid
Ganesh K Maurya, Hari S Misra
The gene knockouts are mostly created using homologous recombination-based replacement of target gene(s) with the expressing cassette of selection marker gene(s). Here, we constructed a series of plasmids bearing the expressing cassettes of genes encoding different antibiotics markers like nptII (KanR ), aadA (SpecR ), cat (CmR ) and aac(3) (GenR ). D. radiodurans is a radioresistant Gram positive bacterium that does not support the independent maintenance of colE1 origin-based plasmids. Using these constructs, the disruption mutants of both single and multiple genes involved in segregation of secondary genome elements have been generated in this bacterium...
September 24, 2018: Plasmid
Ewa Sadowy
Linezolid is considered a last resort drug in treatment of severe infections caused by Gram-positive pathogens, resistant to other antibiotics, such as vancomycin-resistant enterococci (VRE), methicillin-resistant staphylococci and multidrug resistant pneumococci. Although the vast majority of Gram-positive pathogenic bacteria remain susceptible to linezolid, resistant isolates of enterococci, staphylococci and streptococci have been reported worldwide. In these bacteria, apart from mutations, affecting mostly the 23S rRNA genes, acquisition of such genes as cfr, cfr(B), optrA and poxtA, often associated with mobile genetic elements (MGE), plays an important role for resistance...
September 22, 2018: Plasmid
Xiu-Zhong Zhang, Chang-Wei Lei, Jin-Xin Zeng, Yan-Peng Chen, Zhuang-Zhuang Kang, Yu-Long Wang, Xiao-Lan Ye, Xi-Wen Zhai, Hong-Ning Wang
We have identified an IncX1 plasmid named pQJDSal1 from Salmonella enterica subsp. enterica serovar Pullorum (S. Pullorum). The plasmid is 67,685 bp in size and has 72 putative genes. pQJDSal1 harbors a conserved IncX1-type backbone with predicted regions for conjugation, replication and partitioning, as well as a toxin/antitoxin plasmid addiction system. Two regions (A and B) that have not been previously reported in IncX1 plasmids are inserted into the backbone. Region A (10.7 kb), inserted between parA and taxD, consists of a new Tn6168-like transposon containing an arsenic resistant operon arsB2CHR and sulfonamide resistance gene sul2...
September 21, 2018: Plasmid
Monika Dolejska, Costas C Papagiannitsis
Multidrug resistance (MDR) Gram-negative bacteria have been increasingly reported in humans, companion animals and farm animals. The growing trend of plasmid-mediated resistance to antimicrobial classes of critical importance is attributed to the emergence of epidemic plasmids, rapidly disseminating resistance genes among the members of Enterobacteriaceae family. The use of antibiotics to treat humans and animals has had a significant impact on the environment and on wild animals living and feeding in human-influenced habitats...
September 20, 2018: Plasmid
João Alves Gama, Rita Zilhão, Francisco Dionisio
Naturally occurring plasmids have medical importance given that they frequently code for virulence or antibiotic resistance. In many cases, plasmids impose a fitness cost to their hosts, meaning that the growth rate of plasmid-bearing cells is lower than that of plasmid-free cells. However, this does not fit with the fact that plasmids are ubiquitous in nature nor that plasmids and their hosts adapt to each other very fast - as has been shown in laboratory evolutionary assays. Even when plasmids are costly, they seem to largely interact in such a way that the cost of two plasmids is lower than the cost of one of them alone...
September 19, 2018: Plasmid
Alma K Tamez-Castrellón, Ricardo Romo-Lucio, Iván Martínez-Duncker, Héctor M Mora-Montes
Some members of the Sporothrix genus can cause sporotrichosis, a worldwide distributed mycosis that affects several mammalian species, including human beings. Sporothrix schenckii and Sporothrix brasiliensis are the fungal species frequently associated with this disease, and the latter has gained significant interest because of the increased number of cases associated with transmission by cats. Despite the relevance of these organisms in the medical field, limited strategies for their genetic manipulation have been explored...
September 17, 2018: Plasmid
Jean-Yves Madec, Marisa Haenni
Antimicrobial resistance (AMR) plasmids have been recognized as important vectors for efficient spread of AMR phenotypes. The food reservoir includes both food-producing animals and food products, and a huge diversity of AMR plasmids have been reported in this sector. Based on molecular typing methods and/or whole genome sequencing approaches, certain AMR genes/plasmids combinations were found more frequently in food compared to other settings. However, the food source of a definite AMR plasmid is highly complex to confirm due to cross-sectorial transfers and international spread of AMR plasmids...
September 6, 2018: Plasmid
Andrew G Wiggins, Stephen P LaVoie, Joy Wireman, Anne O Summers
Multi-antibiotic resistant (MAR) bacteria cost billions in medical care and tens of thousands of lives annually but perennial calls to limit agricultural and other misuse of antibiotics and to fund antibiotic discovery have not slowed this MAR deluge. Since mobile genetic elements (MGEs) stitch single antibiotic resistance genes into clinically significant MAR arrays, it is high time to focus on how MGEs generate MAR and how disabling them could ameliorate the MAR problem. However, to consider only antibiotics as the drivers of MAR is to miss the significant impact of exposure to non-antibiotic toxic chemicals, specifically metals, on the persistence and spread of MAR...
September 4, 2018: Plasmid
Rafael Pinilla-Redondo, Valentine Cyriaque, Samuel Jacquiod, Søren J Sørensen, Leise Riber
The emergence of antimicrobial resistant bacteria constitutes an increasing global health concern. Although it is well recognized that the cornerstone underlying this phenomenon is the dissemination of antimicrobial resistance via plasmids and other mobile genetic elements, the antimicrobial resistance transfer routes remain largely uncharted. In this review, we describe different methods for assessing the transfer frequency and host ranges of plasmids within complex microbiomes. The discussion is centered around the critical evaluation of recent advances for monitoring the fate of fluorescently tagged plasmids in bacterial communities through the coupling of fluorescence activated cell sorting and next generation sequencing techniques...
August 4, 2018: Plasmid
Stephanie J Ambrose, Christopher J Harmer, Ruth M Hall
The large, broad host range IncC plasmids are important contributors to the spread of key antibiotic resistance genes and over 200 complete sequences of IncC plasmids have been reported. To track the spread of these plasmids accurate typing to identify the closest relatives is needed. However, typing can be complicated by the high variability in resistance gene content and various typing methods that rely on features of the conserved backbone have been developed. Plasmids can be broadly typed into two groups, type 1 and type 2, using four features that differentiate the otherwise closely related backbones...
August 3, 2018: Plasmid
Vicki Adams, Xiaoyan Han, Dena Lyras, Julian I Rood
Clostridium perfringens is an anaerobic bacterium that is a major human and animal pathogen. The key features of C. perfringens-mediated infections are that disease pathogenesis involves the production of protein toxins and that disease epidemiology generally involves the production of environmentally resistant endospores. Many of the toxins involved in these diseases are encoded on conjugative plasmids that are closely related to the paradigm tetracycline resistance plasmid pCW3. This plasmid encodes the Tet(P) tetracycline resistance determinant, and the tcp locus, which mediates conjugative transfer and is also present on the toxin plasmids...
July 26, 2018: Plasmid
Verena Kohler, Ankita Vaishampayan, Elisabeth Grohmann
Conjugative plasmid transfer is one of the major mechanisms responsible for the spread of antibiotic resistance and virulence genes. The incompatibility (Inc) 18 group of plasmids is a family of plasmids replicating by the theta-mechanism, whose members have been detected frequently in enterococci and streptococci. Inc18 plasmids encode a variety of antibiotic resistances, including resistance to vancomycin, chloramphenicol and the macrolide-lincosamide-streptogramine (MLS) group of antibiotics. These plasmids comprising insertions of Tn1546 were demonstrated to be responsible for the transfer of vancomycin resistance encoded by the vanA gene from vancomycin resistant enterococci (VRE) to methicillin resistant Staphylococcus aureus (MRSA)...
June 19, 2018: Plasmid
Andrea T Feßler, Yang Wang, Congming Wu, Stefan Schwarz
Lincosamide resistance in staphylococci is based on the expression of a number of genes which specify three major resistance mechanisms: (i) enzymatic inactivation by lincosamide nucleotidyltransferases, (ii) ribosome protection by ABC-F proteins, and (iii) methylation of the ribosomal target sites in the 23S rRNA by Cfr or Erm methylases. So far, only two lnu genes, lnu(A) and lnu(B), which code for different types of lincosamide nucleotidyltransferases, have been found in staphylococci. The ABC-F proteins are encoded by genes of the vga, lsa and sal classes...
June 19, 2018: Plasmid
Lyle Harris, Leonardo J van Zyl, Bronwyn M Kirby-McCullough, Leonard H Damelin, Caroline T Tiemessen, Marla Trindade
The vaginal mucosa is dominated by Gram positive, rod shaped lactobacilli which serve as a natural barrier against infection. In both healthy- and bacterial vaginosis (BV)-infected women Lactobacillus crispatus and Lactobacillus jensenii have been found to be the predominant Lactobacillus species. Many studies have been conducted to assess factors influencing lactobacilli dominance in the vaginal microbiome. In the present study two plasmids, pLc4 and pLc17, isolated from vaginal Lactobacillus strains of both healthy and BV-infected women were characterized...
June 2018: Plasmid
Jack Schembri, Hayley D Gillis, Amanda L S Lang, Michelle Warhuus, Irene Martin, Walter Demczuk, May ElSherif, Shelly A McNeil, Jason J LeBlanc
BACKGROUND: Serotyping of Streptococcus pneumoniae is an integral part of disease surveillance, with over 92 serotypes characterized to date using traditional serotyping. To identify the most predominant disease causing serotypes, molecular serotyping methods are now increasingly being used, like conventional and real-time multiplex PCR (cmPCR and rmPCR, respectively). Given that cmPCR consists of eight reactions spanning 41 targets, and rmPCR consists of seven triplex reactions, standardizing positive controls for these assays is challenging...
June 2018: Plasmid
Zhongcheng Liu, Lifang Hao, Nanan Wang, Su Zhang, Nan Zhang, Zhenzhen Xu, Yanlei Yang, Yanfen Zhang
IgE/FcεRI signal pathway plays a crucial role in triggering allergic reactions, and there is no cross-recognition between IgE and FcεRI in human and rats. In order to obtain the hFcεRIα/ RBL-2H3 cell line, total RNA was extracted from U937 cells, and the human FcεRIα gene was obtained by RT-PCR technology. Then the amplified product was digested and inserted into the pIRES2 -EGFP vector. After the plasmid was transfected into the RBL-2H3 cells using lipofectamine, and the RBL-2H3 cell lines of stable expression were screened by G418...
June 2018: Plasmid
Deshui Yu, Libing Liao, Yi Zhang, Kedong Xu, Ju Zhang, Kun Liu, Xiaoli Li, Guangxuan Tan, Jurui Zheng, Yong He, Changling Xu, Jinjin Zhao, Beibei Fu, Jiaxing Xie, Jie Mao, Chengwei Li
RNA interference (RNAi), based on hairpin RNA (hpRNA) expression, plays an important role in functional analysis of plant genes. Traditional methods for making RNAi constructs usually involve multiple time-consuming cloning steps. We have developed a Gateway-compatible binary vector for RNAi-mediated gene knockdown in plants from pCAMBIA2301 and pHANNIBAL vectors. The new plant RNAi binary vector, named pCAMBIA2301-GW-RNAi, has two inverted repeated Gateway cassettes driven by the cauliflower mosaic virus 35S (CaMV 35S) promoter...
June 2018: Plasmid
Akira Nagasaki, Yoshio Kato, Keiichi Meguro, Ayana Yamagishi, Chikashi Nakamura, Taro Q P Uyeda
The CRISPR/Cas9 system is a powerful genome editing tool for disrupting the expression of specific genes in a variety of cells. However, the genome editing procedure using currently available vectors is laborious, and there is room for improvement to obtain knockout cells more efficiently. Therefore, we constructed a novel vector for high efficiency genome editing, named pGedit, which contains EGFP-Bsr as a selection marker, expression units of Cas9, and sgRNA without a terminator sequence of the U6 promoter...
June 2018: Plasmid
Fara Amelia Primelles Eguia, Henrique Roman Ramos, Stefanie Kraschowetz, Daniel Omote, Celso Raul Romero Ramos, Paulo Lee Ho, Eneas Carvalho, Viviane Maimoni Gonçalves
Expression vectors for industrial production should be stable and allow tight control of protein synthesis. This is necessary to ensure plasmid transmission to daughter cells in order to achieve a stable population capable of synthesizing high amounts of the target protein. A high-copy-number plasmid, pAE, was previously used for laboratory-scale production of recombinant human granulocyte colony-stimulating factor (rhG-CSF) and the Schistosoma mansoni fatty acid binding protein (rSm14), but it was unstable for large-scale production...
June 2018: Plasmid
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