scholarly journals Phage ϕC2 Mediates Transduction of Tn6215, Encoding Erythromycin Resistance, between Clostridium difficile Strains

mBio ◽  
2013 ◽  
Vol 4 (6) ◽  
Author(s):  
Shan Goh ◽  
Haitham Hussain ◽  
Barbara J. Chang ◽  
Warren Emmett ◽  
Thomas V. Riley ◽  
...  
Keyword(s):  
Clostridium Difficile ◽  
Genetic Manipulation ◽  
Recipient Strain ◽  
Open Reading Frames ◽  
Human Pathogen ◽  
Erythromycin Resistance ◽  
Content Type ◽  
Filter Mating ◽  
First Time ◽  
Reading Frames

ABSTRACTIn this work, we show thatClostridium difficilephage ϕC2 transduceserm(B), which confers erythromycin resistance, from a donor to a recipient strain at a frequency of 10−6per PFU. The transductants were lysogenic for ϕC2 and contained theerm(B) gene in a novel transposon, Tn6215. This element is 13,008 bp in length and contains 17 putative open reading frames (ORFs). It could also be transferred at a lower frequency by filter mating.IMPORTANCEClostridium difficileis a major human pathogen that causes diarrhea that can be persistent and difficult to resolve using antibiotics.C. difficileis potentially zoonotic and has been detected in animals, food, and environmental samples.C. difficilegenomes contain large portions of horizontally acquired genetic elements. The conjugative elements have been reasonably well studied, but transduction has not yet been demonstrated. Here, we show for the first time transduction as a mechanism for the transfer of a novel genetic element inC. difficile. Transduction may also be a useful tool for the genetic manipulation ofC. difficile.

scholarly journals Bacteriophages LIMElight and LIMEzero of Pantoea agglomerans, Belonging to the “phiKMV-Like Viruses”

2011 ◽  
Vol 77 (10) ◽  
pp. 3443-3450 ◽  
Author(s):  
Evelien M. Adriaenssens ◽  
Pieter-Jan Ceyssens ◽  
Vincent Dunon ◽  
Hans-Wolfgang Ackermann ◽  
Johan Van Vaerenbergh ◽  
...  
Keyword(s):  
Soil Samples ◽  
Pantoea Agglomerans ◽  
Open Reading Frames ◽  
Human Pathogen ◽  
Content Type ◽  
Host Diversity ◽  
Double Stranded Dna ◽  
Opportunistic Human Pathogen ◽  
Reading Frames

ABSTRACTPantoea agglomeransis a common soil bacterium used in the biocontrol of fungi and bacteria but is also an opportunistic human pathogen. It has been described extensively in this context, but knowledge of bacteriophages infecting this species is limited. Bacteriophages LIMEzero and LIMElight ofP. agglomeransare lytic phages, isolated from soil samples, belonging to thePodoviridaeand are the firstPantoeaphages of this family to be described. The double-stranded DNA (dsDNA) genomes (43,032 bp and 44,546 bp, respectively) encode 57 and 55 open reading frames (ORFs). Based on the presence of an RNA polymerase in their genomes and their overall genome architecture, these phages should be classified in the subfamily of theAutographivirinae, within the genus of the “phiKMV-like viruses.” Phylogenetic analysis of all the sequenced members of theAutographivirinaesupports the classification of phages LIMElight and LIMEzero as members of the “phiKMV-like viruses” and corroborates the subdivision into the different genera. These data expand the knowledge ofPantoeaphages and illustrate the wide host diversity of phages within the “phiKMV-like viruses.”

scholarly journals SalmonellaGenomic Island 3 Is an Integrative and Conjugative Element and Contributes to Copper and Arsenic Tolerance ofSalmonella enterica

2019 ◽  
Vol 63 (9) ◽  
Author(s):  
Nobuo Arai ◽  
Tsuyoshi Sekizuka ◽  
Yukino Tamamura ◽  
Masahiro Kusumoto ◽  
Atsushi Hinenoya ◽  
...  
Keyword(s):  
Open Reading Frames ◽  
Trna Genes ◽  
Sequencing Analysis ◽  
Content Type ◽  
Arsenic Tolerance ◽  
Filter Mating ◽  
Serovar Typhimurium ◽  
Resistance Island ◽  
Reading Frames ◽  
Integrative And Conjugative Element

ABSTRACTSalmonellagenomic island 3 (SGI3) was first described as a chromosomal island inSalmonella4,[5],12:i:–, a monophasic variant ofSalmonella entericasubsp.entericaserovar Typhimurium. The SGI3 DNA sequence detected fromSalmonella4,[5],12:i:– isolated in Japan was identical to that of a previously reported one across entire length of 81 kb. SGI3 consists of 86 open reading frames, including a copper homeostasis and silver resistance island (CHASRI) and an arsenic tolerance operon, in addition to genes related to conjugative transfer and DNA replication or partitioning, suggesting that the island is a mobile genetic element. We successfully selected transconjugants that acquired SGI3 after filter-mating experiments using theS. entericaserovars Typhimurium, Heidelberg, Hadar, Newport, Cerro, and Thompson as recipients. Southern blot analysis using I-CeuI-digested genomic DNA demonstrated that SGI3 was integrated into a chromosomal fragment of the transconjugants. PCR and sequencing analysis demonstrated that SGI3 was inserted into the 3′ end of the tRNA genespheVorpheR. The length of the target site was 52 or 55 bp, and a 55-bpattIsequence indicating generation of the circular form of SGI3 was also detected. The transconjugants had a higher MIC against CuSO4compared to the recipient strains under anaerobic conditions. Tolerance was defined by thecusgene cluster in the CHASRI. The transconjugants also had distinctly higher MICs against Na2HAsO4compared to recipient strains under aerobic conditions. These findings clearly demonstrate that SGI3 is an integrative and conjugative element and contributes to the copper and arsenic tolerance ofS. enterica.

scholarly journals Lausannevirus Encodes a Functional Dihydrofolate Reductase Susceptible to Proguanil

2017 ◽  
Vol 61 (4) ◽  
Author(s):  
L. Mueller ◽  
P. M. Hauser ◽  
F. Gauye ◽  
G. Greub
Keyword(s):  
Dihydrofolate Reductase ◽  
Expression System ◽  
Open Reading Frames ◽  
Dna Viruses ◽  
Content Type ◽  
The Family ◽  
Nucleocytoplasmic Large Dna Viruses ◽  
First Time ◽  
Reading Frames

ABSTRACT Lausannevirus belongs to the family Marseilleviridae within the group of nucleocytoplasmic large DNA viruses (NCLDVs). These giant viruses exhibit unique features, including a large genome, ranging from 100 kb to 2.5 Mb and including from 150 to more than 2,500 genes, as well as the presence of genes coding for proteins involved in transcription and translation. The large majority of Lausannevirus open reading frames have unknown functions. Interestingly, a bifunctional dihydrofolate reductase-thymidylate synthase (DHFR-TS) is encoded in the Lausannevirus genome. The enzyme plays central roles in DNA precursor biosynthesis. DHFR is the pharmacological target of antifolates, such as trimethoprim, pyrimethamine, and proguanil. First, the functionality of Lausannevirus DHFR-TS was demonstrated by the successful complementation of a DHFR-deficient Saccharomyces cerevisiae strain with a plasmid expressing the heterologous gene. Additionally, using this heterologous expression system, we demonstrated the in vitro susceptibility of Lausannevirus DHFR-TS to proguanil and its resistance to pyrimethamine and trimethoprim. Proguanil may provide a unique and useful treatment if Lausannevirus proves to be a human pathogen. To our knowledge, this is the first time that a DHFR-TS has been described and characterized in an NCLDV.

scholarly journals First Report of Macrolide Resistance Geneerm(T) Harbored by a Novel Small Plasmid from Erysipelothrix rhusiopathiae

2015 ◽  
Vol 59 (4) ◽  
pp. 2462-2465 ◽  
Author(s):  
Chang-Wen Xu ◽  
An-Yun Zhang ◽  
Chun-Mei Yang ◽  
Yun Pan ◽  
Zhong-Bin Guan ◽  
...  
Keyword(s):  
Low Frequency ◽  
Macrolide Resistance ◽  
Open Reading Frames ◽  
The Novel ◽  
Erysipelothrix Rhusiopathiae ◽  
Small Plasmid ◽  
Content Type ◽  
Macrolide Resistance Gene ◽  
First Time ◽  
Reading Frames

ABSTRACTThe macrolide resistance geneerm(T) was identified for the first time in a porcineErysipelothrix rhusiopathiaeisolate from swine in China. The novel 3,749-bp small plasmid pER29, which carrieserm(T), had a G+C content of 31% and four distinct open reading frames. The presence of pER29 increased by at least 128-fold the MICs of clindamycin and erythromycin forE. rhusiopathiae. The fitness cost of pER29 could be responsible for the low frequency oferm(T) inE. rhusiopathiae.

scholarly journals Two Unrelated 8-Vinyl Reductases Ensure Production of Mature Chlorophylls in Acaryochloris marina

2016 ◽  
Vol 198 (9) ◽  
pp. 1393-1400 ◽  
Author(s):  
Guangyu E. Chen ◽  
Andrew Hitchcock ◽  
Philip J. Jackson ◽  
Roy R. Chaudhuri ◽  
Mark J. Dickman ◽  
...  
Keyword(s):  
Transcriptional Control ◽  
Sequence Similarity ◽  
Open Reading Frames ◽  
High Sequence Similarity ◽  
Content Type ◽  
Ethyl Group ◽  
Acaryochloris Marina ◽  
Vinyl Group ◽  
Vinyl Reductase ◽  
Reading Frames

ABSTRACTThe major photopigment of the cyanobacteriumAcaryochloris marinais chlorophylld, while its direct biosynthetic precursor, chlorophylla, is also present in the cell. These pigments, along with the majority of chlorophylls utilized by oxygenic phototrophs, carry an ethyl group at the C-8 position of the molecule, having undergone reduction of a vinyl group during biosynthesis. Two unrelated classes of 8-vinyl reductase involved in the biosynthesis of chlorophylls are known to exist, BciA and BciB. The genome ofAcaryochloris marinacontains open reading frames (ORFs) encoding proteins displaying high sequence similarity to BciA or BciB, although they are annotated as genes involved in transcriptional control (nmrA) and methanogenesis (frhB), respectively. These genes were introduced into an 8-vinyl chlorophylla-producing ΔbciBstrain ofSynechocystissp. strain PCC 6803, and both were shown to restore synthesis of the pigment with an ethyl group at C-8, demonstrating their activities as 8-vinyl reductases. We propose thatnmrAandfrhBbe reassigned asbciAandbciB, respectively; transcript and proteomic analysis ofAcaryochloris marinareveal that bothbciAandbciBare expressed and their encoded proteins are present in the cell, possibly in order to ensure that all synthesized chlorophyll pigment carries an ethyl group at C-8. Potential reasons for the presence of two 8-vinyl reductases in this strain, which is unique for cyanobacteria, are discussed.IMPORTANCEThe cyanobacteriumAcaryochloris marinais the best-studied phototrophic organism that uses chlorophylldfor photosynthesis. Unique among cyanobacteria sequenced to date, its genome contains ORFs encoding two unrelated enzymes that catalyze the reduction of the C-8 vinyl group of a precursor molecule to an ethyl group. Carrying a reduced C-8 group may be of particular importance to organisms containing chlorophylld. Plant genomes also contain orthologs of both of these genes; thus, the bacterial progenitor of the chloroplast may also have contained bothbciAandbciB.

scholarly journals First Staphylococcal Cassette ChromosomemecContaining amecB-Carrying Gene Complex Independent of Transposon Tn6045in a Macrococcus caseolyticus Isolate from a Canine Infection

2015 ◽  
Vol 59 (8) ◽  
pp. 4577-4583 ◽  
Author(s):  
Elena Gómez-Sanz ◽  
Sybille Schwendener ◽  
Andreas Thomann ◽  
Stefanie Gobeli Brawand ◽  
Vincent Perreten
Keyword(s):  
Integration Site ◽  
Direct Repeat ◽  
Open Reading Frames ◽  
Gene Complex ◽  
Direct Repeats ◽  
Content Type ◽  
Canine Infection ◽  
The Right ◽  
Lactamase Gene ◽  
Reading Frames

ABSTRACTA methicillin-resistantmecB-positiveMacrococcus caseolyticus(strain KM45013) was isolated from the nares of a dog with rhinitis. It contained a novel 39-kb transposon-defective completemecB-carrying staphylococcal cassette chromosomemecelement (SCCmecKM45013). SCCmecKM45013contained 49 coding sequences (CDSs), was integrated at the 3′ end of the chromosomalorfXgene, and was delimited at both ends by imperfect direct repeats functioning as integration site sequences (ISSs). SCCmecKM45013presented two discontinuous regions of homology (SCCmeccoverage of 35%) to the chromosomal and transposon Tn6045-associated SCCmec-like element ofM. caseolyticusJCSC7096: (i) themecgene complex (98.8% identity) and (ii) theccr-carrying segment (91.8% identity). Themecgene complex, located at the right junction of the cassette, also carried the β-lactamase geneblaZm(mecRm-mecIm-mecB-blaZm). SCCmecKM45013contained two cassette chromosome recombinase genes,ccrAm2andccrBm2, which shared 94.3% and 96.6% DNA identity with those of the SCCmec-like element of JCSC7096 but shared less than 52% DNA identity with the staphylococcalccrABandccrCgenes. Three distinct extrachromosomal circularized elements (the entire SCCmecKM45013, ΨSCCmecKM45013lacking theccrgenes, and SCCKM45013lackingmecB) flanked by one ISS copy, as well as the chromosomal regions remaining after excision, were detected. An unconventional circularized structure carrying themecBgene complex was associated with two extensive direct repeat regions, which enclosed two open reading frames (ORFs) (ORF46 and ORF51) flanking the chromosomalmecB-carrying gene complex. This study revealedM. caseolyticusas a potential disease-associated bacterium in dogs and also unveiled an SCCmecelement carryingmecBnot associated with Tn6045in the genusMacrococcus.

scholarly journals Characterization of Two Virulent Phages of Lactobacillus plantarum

2012 ◽  
Vol 78 (24) ◽  
pp. 8719-8734 ◽  
Author(s):  
Mariángeles Briggiler Marcó ◽  
Josiane E. Garneau ◽  
Denise Tremblay ◽  
Andrea Quiberoni ◽  
Sylvain Moineau
Keyword(s):  
Lactobacillus Plantarum ◽  
Corn Silage ◽  
Open Reading Frames ◽  
High Identity ◽  
Content Type ◽  
Defense Systems ◽  
Double Stranded Dna ◽  
Type Phage ◽  
Reading Frames

ABSTRACTWe characterized twoLactobacillus plantarumvirulent siphophages, ATCC 8014-B1 (B1) and ATCC 8014-B2 (B2), previously isolated from corn silage and anaerobic sewage sludge, respectively. Phage B2 infected two of the eightL. plantarumstrains tested, while phage B1 infected three. Phage adsorption was highly variable depending on the strain used. Phage defense systems were found in at least twoL. plantarumstrains, LMG9211 and WCSF1. The linear double-stranded DNA genome of thepac-type phage B1 had 38,002 bp, a G+C content of 47.6%, and 60 open reading frames (ORFs). Surprisingly, the phage B1 genome has 97% identity with that ofPediococcus damnosusphage clP1 and 77% identity with that ofL. plantarumphage JL-1; these phages were isolated from sewage and cucumber fermentation, respectively. The double-stranded DNA (dsDNA) genome of thecos-type phage B2 had 80,618 bp, a G+C content of 36.9%, and 127 ORFs with similarities to those ofBacillusandLactobacillusstrains as well as phages. Some phage B2 genes were similar to ORFs fromL. plantarumphage LP65 of theMyoviridaefamily. Additionally, 6 tRNAs were found in the phage B2 genome. Protein analysis revealed 13 (phage B1) and 9 (phage B2) structural proteins. To our knowledge, this is the first report describing such high identity between phage genomes infecting different genera of lactic acid bacteria.

scholarly journals Function of the CRISPR-Cas System of the Human Pathogen Clostridium difficile

mBio ◽  
2015 ◽  
Vol 6 (5) ◽  
Author(s):  
Pierre Boudry ◽  
Ekaterina Semenova ◽  
Marc Monot ◽  
Kirill A. Datsenko ◽  
Anna Lopatina ◽  
...  
Keyword(s):  
Clostridium Difficile ◽  
Type I ◽  
Human Pathogen ◽  
Range Analysis ◽  
Data Set ◽  
Content Type ◽  
Crispr System ◽  
Nosocomial Diarrhea ◽  
Active Expression ◽  
Foreign Dna

ABSTRACT Clostridium difficile is the cause of most frequently occurring nosocomial diarrhea worldwide. As an enteropathogen, C. difficile must be exposed to multiple exogenous genetic elements in bacteriophage-rich gut communities. CRISPR (clustered regularly interspaced short palindromic repeats)-Cas (CRISPR-associated) systems allow bacteria to adapt to foreign genetic invaders. Our recent data revealed active expression and processing of CRISPR RNAs from multiple type I-B CRISPR arrays in C. difficile reference strain 630. Here, we demonstrate active expression of CRISPR arrays in strain R20291, an epidemic C. difficile strain. Through genome sequencing and host range analysis of several new C. difficile phages and plasmid conjugation experiments, we provide evidence of defensive function of the CRISPR-Cas system in both C. difficile strains. We further demonstrate that C. difficile Cas proteins are capable of interference in a heterologous host, Escherichia coli. These data set the stage for mechanistic and physiological analyses of CRISPR-Cas-mediated interactions of important global human pathogen with its genetic parasites. IMPORTANCE Clostridium difficile is the major cause of nosocomial infections associated with antibiotic therapy worldwide. To survive in bacteriophage-rich gut communities, enteropathogens must develop efficient systems for defense against foreign DNA elements. CRISPR-Cas systems have recently taken center stage among various anti-invader bacterial defense systems. We provide experimental evidence for the function of the C. difficile CRISPR system against plasmid DNA and bacteriophages. These data demonstrate the original features of active C. difficile CRISPR system and bring important insights into the interactions of this major enteropathogen with foreign DNA invaders during its infection cycle.

scholarly journals Characterization and Genome Sequence of Mycobacterium Phage XianYue

2020 ◽  
Vol 9 (25) ◽  
Author(s):  
Wesley D. Rhinehart ◽  
Amanda J. Laidlaw ◽  
Alexis M. O’Neal ◽  
Jessica A. Toller ◽  
Miriam Segura-Totten ◽  
...  
Keyword(s):  
Genome Sequence ◽  
Gc Content ◽  
Open Reading Frames ◽  
Nucleotide Identity ◽  
Content Type ◽  
Reading Frames

ABSTRACT Novel mycobacteriophage XianYue was isolated in Northeast Georgia and infects Mycobacteria smegmatis mc2155. Actinobacteriophages which share at least 50% nucleotide identity are grouped into clusters, with XianYue in cluster A2. Its genome is 52,907 bp with 91 open reading frames (ORFs) and 62.9% GC content, and it shares 86.51% nucleotide identity with mycobacteriophage Trixie.

scholarly journals A Role for Tetracycline Selection in Recent Evolution of Agriculture-Associated Clostridium difficile PCR Ribotype 078

mBio ◽  
2019 ◽  
Vol 10 (2) ◽  
Author(s):  
Kate E. Dingle ◽  
Xavier Didelot ◽  
T. Phuong Quan ◽  
David W. Eyre ◽  
Nicole Stoesser ◽  
...  
Keyword(s):  
Clostridium Difficile ◽  
Evolutionary History ◽  
Selective Pressure ◽  
Tetracycline Resistance ◽  
Clonal Expansion ◽  
Human Pathogen ◽  
Content Type ◽  
Resistance Determinants ◽  
Ribotype 078 ◽  
Human Infections

ABSTRACT The increasing clinical importance of human infections (frequently severe) caused by Clostridium difficile PCR ribotype 078 (RT078) was first reported in 2008. The severity of symptoms (mortality of ≤30%) and the higher proportion of infections among community and younger patients raised concerns. Farm animals, especially pigs, have been identified as RT078 reservoirs. We aimed to understand the recent changes in RT078 epidemiology by investigating a possible role for antimicrobial selection in its recent evolutionary history. Phylogenetic analysis of international RT078 genomes (isolates from 2006 to 2014, n = 400), using time-scaled, recombination-corrected, maximum likelihood phylogenies, revealed several recent clonal expansions. A common ancestor of each expansion had independently acquired a different allele of the tetracycline resistance gene tetM. Consequently, an unusually high proportion (76.5%) of RT078 genomes were tetM positive. Multiple additional tetracycline resistance determinants were also identified (including efflux pump tet40), frequently sharing a high level of nucleotide sequence identity (up to 100%) with sequences found in the pig pathogen Streptococcus suis and in other zoonotic pathogens such as Campylobacter jejuni and Campylobacter coli. Each RT078 tetM clonal expansion lacked geographic structure, indicating rapid, recent international spread. Resistance determinants for C. difficile infection-triggering antimicrobials, including fluoroquinolones and clindamycin, were comparatively rare in RT078. Tetracyclines are used intensively in agriculture; this selective pressure, plus rapid, international spread via the food chain, may explain the increased RT078 prevalence in humans. Our work indicates that the use of antimicrobials outside the health care environment has selected for resistant organisms, and in the case of RT078, has contributed to the emergence of a human pathogen. IMPORTANCE Clostridium difficile PCR ribotype 078 (RT078) has multiple reservoirs; many are agricultural. Since 2005, this genotype has been increasingly associated with human infections in both clinical settings and the community. Investigations of RT078 whole-genome sequences revealed that tetracycline resistance had been acquired on multiple independent occasions. Phylogenetic analysis revealed a rapid, recent increase in numbers of closely related tetracycline-resistant RT078 (clonal expansions), suggesting that tetracycline selection has strongly influenced its recent evolutionary history. We demonstrate recent international spread of emergent, tetracycline-resistant RT078. A similar tetracycline-positive clonal expansion was also identified in unrelated nontoxigenic C. difficile, suggesting that this process may be widespread and may be independent of disease-causing ability. Resistance to typical C. difficile infection-associated antimicrobials (e.g., fluoroquinolones, clindamycin) occurred only sporadically within RT078. Selective pressure from tetracycline appears to be a key factor in the emergence of this human pathogen and the rapid international dissemination that followed, plausibly via the food chain.

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