The Novel Macrolide-Lincosamide-Streptogramin B Resistance Gene
            <i>erm</i>
            (44) Is Associated with a Prophage in Staphylococcus xylosus

Wipf, Juliette Ramona Karin; Schwendener, Sybille; Perreten, Vincent

doi:10.1128/aac.02949-14

Cited by 37 publications

(20 citation statements)

References 47 publications

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“…This suggested the presence of a macrolide, lincosamide, and streptogramin B (MLS B ) resistance methylase (Erm) (7). However, none of the erm genes commonly occurring in staphylococci were detected by microarray analysis (8,9). We therefore examined S. fleurettii strain JW205 for a novel MLS B resistance mechanism by genome comparison with MLS Bsusceptible S. fleurettii strain JW404 (Table 1).…”

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confidence: 99%

The New Macrolide-Lincosamide-Streptogramin B Resistance Gene erm (45) Is Located within a Genomic Island in Staphylococcus fleurettii

Wipf

Schwendener

Nielsen

et al. 2015

Antimicrob Agents Chemother

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Genome alignment of a macrolide, lincosamide, and streptogramin B (MLS B )-resistant Staphylococcus fleurettii strain with an MLS B -susceptible S. fleurettii strain revealed a novel 11,513-bp genomic island carrying the new erythromycin resistance methylase gene erm(45). This gene was shown to confer inducible MLS B resistance when cloned into Staphylococcus aureus. The erm(45)-containing island was integrated into the housekeeping gene guaA in S. fleurettii and was able to form a circular intermediate but was not transmissible to S. aureus. Staphylococcus fleurettii is a commensal bacterium of various animal species and an occasional cause of bovine mastitis (1-3). It naturally contains the methicillin resistance gene mecA within its chromosome and is therefore suspected to have been the source of the mecA gene found in staphylococcal cassette chromosome mec (SCCmec) of methicillin-resistant staphylococci, including methicillin-resistant Staphylococcus aureus (MRSA) (4).Due to this intrinsic resistance to ␤-lactams, other antibiotic classes such as macrolides or lincosamides are being used for the treatment of mastitis caused by S. fleurettii (5, 6).S. fleurettii strain JW205, recently isolated from bovine milk in Switzerland, exhibited resistance to erythromycin and inducible resistance to clindamycin (3). This suggested the presence of a macrolide, lincosamide, and streptogramin B (MLS B ) resistance methylase (Erm) (7). However, none of the erm genes commonly occurring in staphylococci were detected by microarray analysis (8, 9). We therefore examined S. fleurettii strain JW205 for a novel MLS B resistance mechanism by genome comparison with MLS Bsusceptible S. fleurettii strain JW404 (Table 1).Detection and characterization of erm(45). Genomes of strain JW205 and JW404 were sequenced using Ion Torrent (Life Technologies, Grand Island, NY) at the UZH/ETH Functional Genomics Center (Zurich, Switzerland) and Illumina MiSeq (Illumina, San Diego, CA) at the Department of Clinical Microbiology at Hvidovre Hospital (Hvidovre, Denmark), respectively. Contigs of the sequenced strains were aligned using the progressive Mauve algorithm (10). This revealed an 11,513-bp integrated genomic island in JW205, which was absent in strain JW404. The island contained 18 open reading frames (ORFs) (Fig. 1), which were identified by the Prokaryotic Dynamic Programming Genefinding Algorithm (Prodigal) (11) and compared to protein sequences and conserved domains in the BLASTp program (http: //blast.ncbi.nlm.nih.gov/Blast.cgi) and the Swiss Institute of Bioinformatics PROSITE database (http://prosite.expasy.org/). The rightmost ORF of the island encoded a 245-amino-acid (aa) protein, which contained the rRNA adenine dimethylase PROSITE signature PS01131, which is present in nearly all Erm 23S rRNA methylases (12). Of all 36 currently described Erm determinants, this methylase exhibited the highest similarity to Erm(B), with 64% aa and 67% nucleotide (nt) identity (Fig. 2).The novel gene was assigned the name erm(45) according to th...

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The New Macrolide-Lincosamide-Streptogramin B Resistance Gene erm (45) Is Located within a Genomic Island in Staphylococcus fleurettii

Wipf

Schwendener

Nielsen

et al. 2015

Antimicrob Agents Chemother

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“…We recently cloned the new MLS B resistance gene erm (44) into vector pBUS1-P cap , generating plasmid pBJW13 (26). For that purpose, the open reading frame of erm (44) was inserted between the NdeI start codon and the TGA stop codon of the MCS and expressed from the strong promoter P cap (Fig.…”

Section: Resultsmentioning

confidence: 99%

“…The shuttle vectors were tested for plasmid stability and copy number in S. aureus. The macrolide-lincosamide-streptogramin B (MLS B ) resistance gene erm (44), cloned downstream of P cap (26), was used to demonstrate the feasibility of rgs-his 6 codon fusion and the production of Erm(44)-RGS-His 6 protein in both S. aureus and E. coli.…”

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“…These vectors were derived from pBUS1 (24), which has frequently been used to clone genes in S. aureus (25)(26)(27)(28)(29)(30)(31)(32), and comprise the Gram-positive replicon pAM␣1 or pT181, the pBluescript II SK multiple cloning site (MCS) without a promoter, or a novel MCS with joinable rgs-his 6 codons downstream of the strong S. aureus type 1 capsule gene 1A promoter (P cap ) (33). The shuttle vectors were tested for plasmid stability and copy number in S. aureus.…”

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New Shuttle Vector-Based Expression System To Generate Polyhistidine-Tagged Fusion Proteins in Staphylococcus aureus and Escherichia coli

Schwendener

Perreten

2015

Appl Environ Microbiol

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Four Staphylococcus aureus-Escherichia coli shuttle vectors were constructed for gene expression and production of tagged fusion proteins. Vectors pBUS1-HC and pTSSCm have no promoter upstream of the multiple cloning site (MCS), and this allows study of genes under the control of their native promoters, and pBUS1-P cap -HC and pTSSCm-P cap contain the strong constitutive promoter of S. aureus type 1 capsule gene 1A (P cap ) upstream of a novel MCS harboring codons for the peptide tag Arg-GlySer-hexa-His (rgs-his 6 ). All plasmids contained the backbone derived from pBUS1, including the E. coli origin ColE1, five copies of terminator rrnB T 1 , and tetracycline resistance marker tet(L) for S. aureus and E. coli. The minimum pAM␣1 replicon from pBUS1 was improved through either complementation with the single-strand origin oriL from pUB110 (pBUS1-HC and pBUS1-P cap -HC) or substitution with a pT181-family replicon (pTSSCm and pTSSCm-P cap ). The new constructs displayed increased plasmid yield and segregational stability in S. aureus. Furthermore, pBUS1-P cap -HC and pTSSCm-P cap offer the potential to generate C-terminal RGS-His 6 translational fusions of cloned genes using simple molecular manipulation. BcgI-induced DNA excision followed by religation converts the TGA stop codon of the MCS into a TGC codon and links the rgs-his 6 codons to the 3= end of the target gene. The generation of the rgs-his 6 codon-fusion, gene expression, and protein purification were demonstrated in both S. aureus and E. coli using the macrolide-lincosamide-streptogramin B resistance gene erm(44) inserted downstream of P cap . The new His tag expression system represents a helpful tool for the direct analysis of target gene function in staphylococcal cells. Staphylococci are commensals of human and animal skin. In some species, particularly in Staphylococcus aureus, community-and hospital-associated clones have acquired a multitude of virulence and antibiotic resistance mechanisms, representing a serious public health risk (1, 2). An increased understanding of the resistance and pathogenicity mechanisms of staphylococci is needed. Therefore, molecular tools for genetic analysis, including gene disruption and ectopic expression, have been developed (3). S. aureus-Escherichia coli shuttle vectors have been constructed using restriction fragments derived from small, natural highcopy-number S. aureus plasmids, such as pUB110, pT181, pE194, and pC194 (4-7). These plasmids propagate through rolling-circle replication (RCR) in staphylococci, requiring the replication initiator protein (Rep) and the double-strand origin (dso) to initiate leading-strand synthesis as well as the single-strand origin (sso) for subsequent efficient lagging-strand synthesis (8). The absence of a functional sso sequence in RCR plasmids has been associated with single-stranded DNA (ssDNA) accumulation, low copy numbers, and plasmid instability (9-11). While some sso sequences support replication only in the native host, other sequences are functional in a number of ...

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“…Along with the erm(44) and erm(45) genes, which were also recently identified in staphylococci from bovine mastitis milk (9,22,23), the discovery of erm(48) highlights the role of dairy cows as a source of new MLS B resistance genes. Accession number(s).…”

Section: Figmentioning

confidence: 99%

New Macrolide-Lincosamide-Streptogramin B Resistance Gene erm (48) on the Novel Plasmid pJW2311 in Staphylococcus xylosus

Wipf

Riley

Kania

et al. 2017

Antimicrob Agents Chemother

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The Novel Macrolide-Lincosamide-Streptogramin B Resistance Gene erm (44) Is Associated with a Prophage in Staphylococcus xylosus

Cited by 37 publications

References 47 publications

The New Macrolide-Lincosamide-Streptogramin B Resistance Gene erm (45) Is Located within a Genomic Island in Staphylococcus fleurettii

The New Macrolide-Lincosamide-Streptogramin B Resistance Gene erm (45) Is Located within a Genomic Island in Staphylococcus fleurettii

New Shuttle Vector-Based Expression System To Generate Polyhistidine-Tagged Fusion Proteins in Staphylococcus aureus and Escherichia coli

New Macrolide-Lincosamide-Streptogramin B Resistance Gene erm (48) on the Novel Plasmid pJW2311 in Staphylococcus xylosus

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