Complete DNA Sequence and Analysis of the Transferable Multiple-Drug Resistance Plasmids (R Plasmids) from Photobacterium damselae subsp. piscicida Isolates Collected in Japan and the United States

Abstract

Photobacterium damselae subsp. piscicida is a bacterial fish pathogen that causes a disease known as pasteurellosis. Two transferable multiple-drug resistance (R) plasmids, pP99-018 (carrying resistance to kanamycin, chloramphenicol, tetracycline, and sulfonamide) and pP91278 (carrying resistance to tetracycline, trimethoprim, and sulfonamide), isolated from P. damselae subsp. piscicida strains from Japan (P99-018) and the United States (P91278), respectively, were completely sequenced and analyzed, along with the multipledrug resistance regions of three other R plasmids also from P. damselae subsp. piscicida strains from Japan. The sequence structures of pP99-018 (150,057 bp) and pP91278 (131,520 bp) were highly conserved, with differences due to variation in the drug resistance and conjugative transfer regions. These plasmids, shown to be closely related to the IncJ element R391 (a conjugative, self-transmitting, integrating element, or constin), were divided into the conjugative transfer, replication, partition, and multiple-drug resistance regions. Each of the five multiple-drug resistance regions sequenced exhibited unique drug resistance marker composition and arrangement.Antimicrobial drugs have historically been used to control bacterial and mycotic infections in fish farming (16). This practice has helped the industry improve production but at the same time allowed for the emergence of drug-resistant pathogens, which is known to be made possible by transferable R plasmids present in fish-pathogenic gram-positive and gramnegative bacteria (17). The R plasmids of Photobacterium damselae subsp. piscicida, a gram-negative pathogen, carry several drug resistance genes, also called drug resistance markers, that include a class A ␤-lactamase gene for ampicillin resistance (Ap r ), catI and catII for chloramphenicol resistance (Cp r ), ppflo for florfenicol resistance (Ff r ), aphA7 for kanamycin resistance (Km r ), sul2 for sulfonamide resistance (Sa r ), and tet(A) for tetracycline resistance (Tc r ). Some of these drugs (ampicillin, florfenicol, sulfonamide, and tetracycline) are used in aquaculture in Japan, while others (chloramphenicol and kanamycin) are not (7).P. damselae subsp. piscicida is the causative agent of pasteurellosis, a disease responsible for serious economic losses in fish farms across the world (17). However, even though the drug resistance patterns and genes contained in R plasmids from P. damselae subsp. piscicida strains in Japan, the United States, and Europe have been identified, the complete sequences of these R plasmids from P. damselae subsp. piscicida and the molecular mechanism responsible for the formation of resistance determinants in this species have not been well characterized.In this study, we determined and analyzed the complete nucleotide sequences of R plasmids pP99-018 (Japan) and pP91278 (United States) from P. damselae subsp. piscicida, as well as the multiple-drug resistance regions of three other R plasmids (pSP98048, pSP98026, and pP9014) also collected in Japan. ...

The Importance of Mobile Genetic Elements in the Evolution of Salmonella: Pathogenesis, Antibiotic Resistance and Host Adaptation

“…Nevertheless, the nucleotide sequences of the regions shared with the bla CMY-2 -bearing IncA/C plasmids were identical, suggesting that the bla CMY-2 -bearing plasmids could be the result of the insertion of DNA modules into this smaller precursor plasmid Wiesner et al, 2011). The general agreement from the analysis of the genetic structure of the IncA/C group,is that plasmid evolution progresses faster through the insertion/deletion of DNA stretches rather than by point mutations (Welch et al, 2007;Kim et al, 2008;Fricke et al, 2009;Call et al, 2010;Wiesner et al, 2011).…”

“…They have attracted the attention of the research community due to their ability to acquire antimicrobial resistance traits and to mobilize across geographical and taxonomical borders (Fricke et al, 2009). Recent comparative studies have addressed the evolutionary relationships among the IncA/C plasmids from Salmonella enterica, Escherichia coli, Yersinia pestis, Yersinia ruckeri, Vibrio cholera, Photobacterium damselae and Aeromonas salmonicida (Welch et al, 2007;Kim et al, 2008;McIntosh et al, 2008;Pan et al, 2008;Fricke et al, 2009;Call et al, 2010). The genomic comparison of seven IncA/C plasmids showed that these plasmids share a common backbone, including the origin of replication and a conjugative plasmid transfer system (Welch et al, 2007;Fricke et al, 2009).…”

Similarity in the Structure of <i>tetD</i>-Carrying Mobile Genetic Elements in Bacterial Strains of Different Genera Isolated from Cultured Yellowtail

“…PCR with a set of primers TCr3-F1 5 -AGCCTGACCAATCTGAC-3 and TCr3-R5 5 -GTGAAGAAGTGGCAGATC-3 described in Fig.2, and TA3 as a template DNA indicated the presence of adhC and ORF2, and IS26-Right in opposite orientation to the IS26-Left Fig.2A . The flanking region by IS26-Left and -Right was identical to the structure found in Photobacterium damselae Kim et al, 2008;Kim and Aoki, 1994 except for the ISRSB101. The inserted ISRSB101 was 100% identical to that of plasmid pFBOAT6 of Aeromonas punctata Rhodes et al, 2004 , and was bordered by direct repeats of 9 nucleotide 5 -TGTTGCAAA-3 originating from IS26-Inner.…”

“…TA3 pTCKM3 . Solid bars indicate the regions which exhibit homology to the reported sequences; pP99-018 from Photobacterium damselae Kim et al, 2008;Kim and Aoki, 1994 , Salmonella serovar Ordnez Allard et al, 1993 , and pFBAOT6 from Aeromonas punctata Rhodes et al, 2004 . B The structure found in Salmonella sp.…”

“…TC67 pCM67-5 . Solid bars indicate the regions which exhibit homology to the reported sequences; pP99-018 from Photobacterium damselae Kim et al, 2008;Kim and Aoki, 1994 , class I integron from Aeromonas hydrophila Poole et al, 2006 , TnSF1 form Shigella flexneri accession no. AF188331 , pPCVM196330_110 from Salmonella serovar Schwarzenground accession no.…”