scholarly journals Benzoate- and Salicylate-Tolerant Strains of Escherichia coli K-12 Lose Antibiotic Resistance during Laboratory Evolution

2016 ◽  
Vol 83 (2) ◽  
Author(s):  
Kaitlin E. Creamer ◽  
Frederick S. Ditmars ◽  
Preston J. Basting ◽  
Karina S. Kunka ◽  
Issam N. Hamdallah ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Drug Resistance ◽  
Antibiotic Resistance ◽  
Multidrug Resistance ◽  
Gut Microbiome ◽  
Nucleotide Polymorphisms ◽  
Content Type ◽  
Large Deletions ◽  
Increased Sensitivity ◽  
K 12

ABSTRACT Escherichia coli K-12 W3110 grows in the presence of membrane-permeant organic acids that can depress cytoplasmic pH and accumulate in the cytoplasm. We conducted experimental evolution by daily diluting cultures in increasing concentrations of benzoic acid (up to 20 mM) buffered at external pH 6.5, a pH at which permeant acids concentrate in the cytoplasm. By 2,000 generations, clones isolated from evolving populations showed increasing tolerance to benzoate but were sensitive to chloramphenicol and tetracycline. Sixteen clones grew to stationary phase in 20 mM benzoate, whereas the ancestral strain W3110 peaked and declined. Similar growth occurred in 10 mM salicylate. Benzoate-evolved strains grew like W3110 in the absence of benzoate, in media buffered at pH 4.8, pH 7.0, or pH 9.0, or in 20 mM acetate or sorbate at pH 6.5. Genomes of 16 strains revealed over 100 mutations, including single-nucleotide polymorphisms (SNPs), large deletions, and insertion knockouts. Most strains acquired deletions in the benzoate-induced multiple antibiotic resistance (Mar) regulon or in associated regulators such as rob and cpxA, as well as the multidrug resistance (MDR) efflux pumps emrA, emrY, and mdtA. Strains also lost or downregulated the Gad acid fitness regulon. In 5 mM benzoate or in 2 mM salicylate (2-hydroxybenzoate), most strains showed increased sensitivity to the antibiotics chloramphenicol and tetracycline; some strains were more sensitive than a marA knockout strain. Thus, our benzoate-evolved strains may reveal additional unknown drug resistance components. Benzoate or salicylate selection pressure may cause general loss of MDR genes and regulators. IMPORTANCE Benzoate is a common food preservative, and salicylate is the primary active metabolite of aspirin. In the gut microbiome, genetic adaptation to salicylate may involve loss or downregulation of inducible multidrug resistance systems. This discovery implies that aspirin therapy may modulate the human gut microbiome to favor salicylate tolerance at the expense of drug resistance. Similar aspirin-associated loss of drug resistance might occur in bacterial pathogens found in arterial plaques.

scholarly journals Benzoate and Salicylate Tolerant Strains Lose Antibiotic Resistance during Laboratory Evolution ofEscherichia coliK-12

2016 ◽  
Author(s):  
Kaitlin E. Creamer ◽  
Frederick S. Ditmars ◽  
Preston J. Basting ◽  
Karina S. Kunka ◽  
Issam N. Hamdallah ◽  
...  
Keyword(s):  
Drug Resistance ◽  
Antibiotic Resistance ◽  
Gut Microbiome ◽  
Food Preservative ◽  
Large Deletions ◽  
Increased Sensitivity ◽  
K 12 ◽  
Similar Growth ◽  
External Ph ◽  
Mdr Genes

ABSTRACTEscherichia coliK-12 W3110 grows in the presence of membrane-permeant organic acids that can depress cytoplasmic pH and accumulate in the cytoplasm. We conducted experimental evolution by daily diluting cultures in increasing concentrations of benzoic acid (up to 20 mM) buffered at external pH 6.5, a pH at which permeant acids concentrate in the cytoplasm. By 2,000 generations, clones isolated from evolving populations showed increasing tolerance to benzoate but were sensitive to chloramphenicol and tetracycline. Sixteen clones grew to stationary phase in 20 mM benzoate, whereas the ancestral strain W3110 peaked and declined. Similar growth occurred in 10 mM salicylate. Benzoate-evolved strains grew like W3110 in the absence of benzoate; in media buffered at pH 4.8, pH 7.0, or pH 9.0; or in 20 mM acetate or sorbate at pH 6.5. Genomes of 16 strains revealed over 100 mutations including SNPs, large deletions, and insertion knockouts. Most strains acquired deletions in the benzoate-induced multiple antibiotic resistance (Mar) regulon or in associated regulators such asrobandcpxA, as well as MDR efflux pumpsemrA,emrY, andmdtA. Strains also lost or down-regulated the Gad acid fitness regulon. In 5 mM benzoate, or in 2 mM salicylate (2-hydroxybenzoate), most strains showed increased sensitivity to the antibiotics chloramphenicol and tetracycline; some strains were more sensitive than amarAknockout. Thus, our benzoate-evolved strains may reveal additional unknown drug resistance components. Benzoate or salicylate selection pressure may cause general loss of MDR genes and regulators.IMPORTANCEBenzoate is a common food preservative, and salicylate is the primary active metabolite of aspirin. In the gut microbiome, genetic adaptation to salicylate may involve loss or downregulation of inducible multidrug resistance systems. This discovery implies that aspirin therapy may modulate the human gut microbiome to favor salicylate tolerance at the expense of drug resistance. Similar aspirin-associated loss of drug resistance might occur in bacterial pathogens found in arterial plaques.

scholarly journals Increased Survival of Antibiotic-Resistant Escherichia coli inside Macrophages

2012 ◽  
Vol 57 (1) ◽  
pp. 189-195 ◽  
Author(s):  
Migla Miskinyte ◽  
Isabel Gordo
Keyword(s):  
Escherichia Coli ◽  
Antibiotic Resistance ◽  
Bacterial Infections ◽  
Resistance Mutation ◽  
Fitness Cost ◽  
Resistance Mutations ◽  
Content Type ◽  
E Coli ◽  
Fitness Effects ◽  
K 12

ABSTRACTMutations causing antibiotic resistance usually incur a fitness cost in the absence of antibiotics. The magnitude of such costs is known to vary with the environment. Little is known about the fitness effects of antibiotic resistance mutations when bacteria confront the host's immune system. Here, we study the fitness effects of mutations in therpoB,rpsL, andgyrAgenes, which confer resistance to rifampin, streptomycin, and nalidixic acid, respectively. These antibiotics are frequently used in the treatment of bacterial infections. We measured two important fitness traits—growth rate and survival ability—of 12Escherichia coliK-12 strains, each carrying a single resistance mutation, in the presence of macrophages. Strikingly, we found that 67% of the mutants survived better than the susceptible bacteria in the intracellular niche of the phagocytic cells. In particular, allE. colistreptomycin-resistant mutants exhibited an intracellular advantage. On the other hand, 42% of the mutants incurred a high fitness cost when the bacteria were allowed to divide outside of macrophages. This study shows that single nonsynonymous changes affecting fundamental processes in the cell can contribute to prolonged survival ofE. coliin the context of an infection.

scholarly journals The Conjugation Window in an Escherichia coli K-12 Strain with an IncFII Plasmid

2020 ◽  
Vol 86 (17) ◽  
Author(s):  
Brendan Headd ◽  
Scott A. Bradford
Keyword(s):  
Escherichia Coli ◽  
Antibiotic Resistance ◽  
Antibiotic Resistance Genes ◽  
Time Frame ◽  
Environmental Aspect ◽  
Content Type ◽  
E Coli ◽  
Donor Cells ◽  
K 12 ◽  
Regulatory Controls

ABSTRACT Many studies have examined the role that conjugation plays in disseminating antibiotic resistance genes in bacteria. However, relatively little research has quantitively examined and modeled the dynamics of conjugation under growing and nongrowing conditions beyond a couple of hours. We therefore examined growing and nongrowing cultures of Escherichia coli over a 24-h period to understand the dynamics of bacterial conjugation in the presence and absence of antibiotics with pUUH239.2, an IncFII plasmid containing multiantibiotic- and metal-resistant genes. Our data indicate that conjugation occurs after E. coli cells divide and before they have transitioned to a nongrowing phase. The result is that there is only a small window of opportunity for E. coli to conjugate with pUUH239.2 under both growing and nongrowing conditions. Only a very small percentage of the donor cells likely are capable of even undergoing conjugation, and not all transconjugants can become donor cells due to molecular regulatory controls and not being in the correct growth phase. Once a growing culture enters stationary phase, the number of capable donor cells decreases rapidly and conjugation slows to produce a plateau. Published models did not provide accurate descriptions of conjugation under nongrowing conditions. We present here a modified modeling approach that accurately describes observed conjugation behavior under growing and nongrowing conditions. IMPORTANCE There has been growing interest in horizontal gene transfer of antibiotic resistance plasmids as the antibiotic resistance crisis has worsened over the years. Most studies examining conjugation of bacterial plasmids focus on growing cultures of bacteria for short periods, but in the environment, most bacteria grow episodically and at much lower rates than in the laboratory. We examined conjugation of an IncFII antibiotic resistance plasmid in E. coli under growing and nongrowing conditions to understand the dynamics of conjugation under which the plasmid is transferred. We found that conjugation occurs in a narrow time frame when E. coli is transitioning from a growing to nongrowing phase and that the conjugation plateau develops because of a lack of capable donor cells in growing cultures. From an environmental aspect, our results suggest that episodic growth in nutrient-depleted environments could result in more conjugation than sustained growth in a nutrient rich environment.

scholarly journals Contagious Antibiotic Resistance: Plasmid Transfer among Bacterial Residents of the Zebrafish Gut

2021 ◽  
Vol 87 (9) ◽  
Author(s):  
Wesley Loftie-Eaton ◽  
Angela Crabtree ◽  
David Perry ◽  
Jack Millstein ◽  
Justin Baytosh ◽  
...  
Keyword(s):  
Antibiotic Resistance ◽  
Multidrug Resistance ◽  
Gut Microbiome ◽  
Bacterial Communities ◽  
Plasmid Transfer ◽  
Culturable Bacteria ◽  
Content Type ◽  
Resistance Plasmid ◽  
Resistance Plasmids

ABSTRACTBy characterizing the trajectories of antibiotic resistance gene transfer in bacterial communities such as the gut microbiome, we will better understand the factors that influence this spread of resistance. Our aim was to investigate the host network of a multidrug resistance broad-host-range plasmid in the culturable gut microbiome of zebrafish. This was done throughin vitroandin vivoconjugation experiments withEscherichia colias the donor of the plasmid pB10::gfp. When this donor was mixed with the extracted gut microbiome, only transconjugants ofAeromonas veroniiwere detected. In separate matings between the same donor and four prominent isolates from the gut microbiome, the plasmid transferred to two of these four isolates,A. veroniiandPlesiomonas shigelloides, but not toShewanella putrefaciensandVibrio mimicus. When theseA. veroniiandP. shigelloidestransconjugants were the donors in matings with the same four isolates, the plasmid now also transferred fromA. veroniitoS. putrefaciens.P. shigelloideswas unable to donate the plasmid, andV. mimicuswas unable to acquire it. Finally, when theE. colidonor was addedin vivoto zebrafish through their food, plasmid transfer was observed in the gut, but only toAchromobacter, a rare member of the gut microbiome. This work shows that the success of plasmid-mediated antibiotic resistance spread in a gut microbiome depends on the donor-recipient species combinations and therefore their spatial arrangement. It also suggests that rare gut microbiome members should not be ignored as potential reservoirs of multidrug resistance plasmids from food.IMPORTANCETo understand how antibiotic resistance plasmids end up in human pathogens, it is crucial to learn how, where, and when they are transferred and maintained in members of bacterial communities such as the gut microbiome. To gain insight into the network of plasmid-mediated antibiotic resistance sharing in the gut microbiome, we investigated the transferability and maintenance of a multidrug resistance plasmid among the culturable bacteria of the zebrafish gut. We show that the success of plasmid-mediated antibiotic resistance spread in a gut microbiome can depend on which species are involved, as some are important nodes in the plasmid-host network and others are dead ends. Our findings also suggest that rare gut microbiome members should not be ignored as potential reservoirs of multidrug resistance plasmids from food.

scholarly journals Complete Sequence of the FII Plasmid p42-2, CarryingblaCTX-M-55,oqxAB,fosA3, andfloRfrom Escherichia coli

2016 ◽  
Vol 60 (7) ◽  
pp. 4336-4338 ◽  
Author(s):  
Qiu E. Yang ◽  
Timothy Rutland Walsh ◽  
Bao Tao Liu ◽  
Meng Ting Zou ◽  
Hui Deng ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Antibiotic Resistance ◽  
Multidrug Resistance ◽  
Resistance Genes ◽  
Antibiotic Resistance Genes ◽  
Complete Sequence ◽  
Content Type ◽  
Resistance Determinants

ABSTRACTWe sequenced a novel conjugative multidrug resistance IncF plasmid, p42-2, isolated fromEscherichia colistrain 42-2, previously identified in China. p42-2 is 106,886 bp long, composed of a typical IncFII-type backbone (∼54 kb) and one distinct acquired DNA region spanning ∼53 kb, harboring 12 antibiotic resistance genes [blaCTX-M-55,oqxA,oqxB,fosA3,floR,tetA(A),tetA(R),strA,strB,sul2,aph(3′)-II, and ΔblaTEM-1]. The spread of these multidrug resistance determinants on the same plasmid is of great concern and, because of coresistance to antibiotics from different classes, is therapeutically challenging.

scholarly journals Genome-Based Comparison of Cyclic Di-GMP Signaling in Pathogenic and Commensal Escherichia coli Strains

2015 ◽  
Vol 198 (1) ◽  
pp. 111-126 ◽  
Author(s):  
Tatyana L. Povolotsky ◽  
Regine Hengge
Keyword(s):  
Escherichia Coli ◽  
Bacterial Species ◽  
Content Type ◽  
E Coli ◽  
Large Deletions ◽  
Diguanylate Cyclases ◽  
Core Set ◽  
Genes Encoding ◽  
K 12 ◽  
Encoding Genes

ABSTRACTThe ubiquitous bacterial second messenger cyclic di-GMP (c-di-GMP) has recently become prominent as a trigger for biofilm formation in many bacteria. It is generated by diguanylate cyclases (DGCs; with GGDEF domains) and degraded by specific phosphodiesterases (PDEs; containing either EAL or HD-GYP domains). Most bacterial species contain multiples of these proteins with some having specific functions that are based on direct molecular interactions in addition to their enzymatic activities.Escherichia coliK-12 laboratory strains feature 29 genes encoding GGDEF and/or EAL domains, resulting in a set of 12 DGCs, 13 PDEs, and four enzymatically inactive “degenerate” proteins that act by direct macromolecular interactions. We present here a comparative analysis of GGDEF/EAL domain-encoding genes in 61 genomes of pathogenic, commensal, and probioticE. colistrains (including enteric pathogens such as enteroaggregative, enterohemorrhagic, enteropathogenic, enterotoxigenic, and adherent and invasiveEscherichia coliand the 2011 German outbreak O104:H4 strain, as well as extraintestinal pathogenicE. coli, such as uropathogenic and meningitis-associatedE. coli). We describe additional genes for two membrane-associated DGCs (DgcX and DgcY) and four PDEs (the membrane-associated PdeT, as well as the EAL domain-only proteins PdeW, PdeX, and PdeY), thus showing the pangenome ofE. colito contain at least 35 GGDEF/EAL domain proteins. A core set of only eight proteins is absolutely conserved in all 61 strains: DgcC (YaiC), DgcI (YliF), PdeB (YlaB), PdeH (YhjH), PdeK (YhjK), PdeN (Rtn), and the degenerate proteins CsrD and CdgI (YeaI). In all other GGDEF/EAL domain genes, diverse point and frameshift mutations, as well as small or large deletions, were discovered in various strains.IMPORTANCEOur analysis reveals interesting trends in pathogenicEscherichia colithat could reflect different host cell adherence mechanisms. These may either benefit from or be counteracted by the c-di-GMP-stimulated production of amyloid curli fibers and cellulose. Thus, EAEC, which adhere in a “stacked brick” biofilm mode, have a potential for high c-di-GMP accumulation due to DgcX, a strongly expressed additional DGC. In contrast, EHEC and UPEC, which use alternative adherence mechanisms, tend to have extra PDEs, suggesting that low cellular c-di-GMP levels are crucial for these strains under specific conditions. Overall, our study also indicates that GGDEF/EAL domain proteins evolve rapidly and thereby contribute to adaptation to host-specific and environmental niches of various types ofE. coli.

scholarly journals Selective Conditions for a Multidrug Resistance Plasmid Depend on the Sociality of Antibiotic Resistance

2016 ◽  
Vol 60 (4) ◽  
pp. 2524-2527 ◽  
Author(s):  
Michael J. Bottery ◽  
A. Jamie Wood ◽  
Michael A. Brockhurst
Keyword(s):  
Drug Resistance ◽  
Antibiotic Resistance ◽  
Multidrug Resistance ◽  
Efflux Pump ◽  
Antibiotic Resistance Genes ◽  
Mechanisms Of Resistance ◽  
Content Type ◽  
Drug Concentrations ◽  
Selection For ◽  
Free Strain

ABSTRACTMultidrug resistance (MDR) plasmids frequently carry antibiotic resistance genes conferring qualitatively different mechanisms of resistance. We show here that the antibiotic concentrations selecting for the RK2 plasmid inEscherichia colidepend upon the sociality of the drug resistance: the selection for selfish drug resistance (efflux pump) occurred at very low drug concentrations, just 1.3% of the MIC of the plasmid-free antibiotic-sensitive strain, whereas selection for cooperative drug resistance (modifying enzyme) occurred at drug concentrations exceeding the MIC of the plasmid-free strain.

scholarly journals Limited and Strain-Specific Transcriptional and Growth Responses to Acquisition of a Multidrug Resistance Plasmid in Genetically Diverse Escherichia coli Lineages

mSystems ◽  
2021 ◽  
Vol 6 (2) ◽  
Author(s):  
Steven Dunn ◽  
Laura Carrilero ◽  
Michael Brockhurst ◽  
Alan McNally
Keyword(s):  
Escherichia Coli ◽  
Antibiotic Resistance ◽  
Multidrug Resistance ◽  
Resistance Genes ◽  
Antibiotic Resistance Genes ◽  
Growth Responses ◽  
Fitness Costs ◽  
Transcriptional Responses ◽  
Content Type ◽  
E Coli

ABSTRACT Multidrug-resistant (MDR) Escherichia coli strains are a major global threat to human health, wherein multidrug resistance is primarily spread by MDR plasmid acquisition. MDR plasmids are not widely distributed across the entire E. coli species, but instead are concentrated in a small number of clones. Here, we test if diverse E. coli strains vary in their ability to acquire and maintain MDR plasmids and if this relates to their transcriptional response following plasmid acquisition. We used strains from across the diversity of E. coli strains, including the common MDR lineage sequence type 131 (ST131) and the IncF plasmid pLL35, carrying multiple antibiotic resistance genes. Strains varied in their ability to acquire pLL35 by conjugation, but all were able to stably maintain the plasmid. The effects of pLL35 acquisition on cefotaxime resistance and growth also varied among strains, with growth responses ranging from a small decrease to a small increase in growth of the plasmid carrier relative to the parental strain. Transcriptional responses to pLL35 acquisition were limited in scale and highly strain specific. We observed transcriptional responses at the operon or regulon level—possibly due to stress responses or interactions with resident mobile genetic elements (MGEs). Subtle transcriptional responses consistent across all strains were observed affecting functions, such as anaerobic metabolism, previously shown to be under negative frequency-dependent selection in MDR E. coli. Overall, there was no correlation between the magnitudes of the transcriptional and growth responses across strains. Together, these data suggest that fitness costs arising from transcriptional disruption are unlikely to act as a barrier to dissemination of this MDR plasmid in E. coli. IMPORTANCE Plasmids play a key role in bacterial evolution by transferring adaptive functions between lineages that often enable invasion of new niches, including driving the spread of antibiotic resistance genes. Fitness costs of plasmid acquisition arising from the disruption of cellular processes could limit the spread of multidrug resistance plasmids. However, the impacts of plasmid acquisition are typically measured in lab-adapted strains rather than natural isolates, which act as reservoirs for the maintenance and transmission of plasmids to clinically relevant strains. Using a clinical multidrug resistance plasmid and a diverse collection of E. coli strains isolated from clinical infections and natural environments, we show that plasmid acquisition had only limited and highly strain-specific effects on bacterial growth and transcription under laboratory conditions. These findings suggest that fitness costs arising from transcriptional disruption are unlikely to act as a barrier to transmission of this plasmid in natural populations of E. coli.

scholarly journals Horsing Around: Escherichia coli ST1250 of Equine Origin Harboring Epidemic IncHI1/ST9 Plasmid with blaCTX-M-1 and an Operon for Short-Chain Fructooligosaccharide Metabolism

2021 ◽  
Vol 65 (5) ◽  
Author(s):  
Adam Valcek ◽  
Petra Sismova ◽  
Kristina Nesporova ◽  
Søren Overballe-Petersen ◽  
Ibrahim Bitar ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Multidrug Resistance ◽  
Integration Site ◽  
Draft Genome ◽  
Short Chain ◽  
Nucleotide Polymorphisms ◽  
Content Type ◽  
Plasmid Content ◽  
E Coli ◽  
Long Read

ABSTRACT The relatedness of the equine-associated Escherichia coli strain ST1250 and its single- and double-locus variants (ST1250-SLV/DLV), obtained from horses in Europe, was studied by comparative genome analysis. A total of 54 isolates of E. coli ST1250 and ST1250-SLV/DLV from healthy and hospitalized horses across Europe (Czech Republic [n = 23], The Netherlands [n = 18], Germany [n = 9], Denmark [n = 3], and France [n = 1]) from 2008 to 2017 were subjected to whole-genome sequencing. An additional 25 draft genome assemblies of E. coli ST1250 and ST1250-SLV/DLV were obtained from the public databases. The isolates were compared for genomic features, virulence genes, clade structure, and plasmid content. The complete nucleotide sequences of eight IncHI1/ST9 plasmids and one IncHI1/ST2 plasmid were obtained using long-read sequencing by PacBio or MinION. In the collection of 79 isolates, only 10 were phylogenetically close (<8 single nucleotide polymorphisms [SNP]). The majority of isolates belonged to phylogroup B1 (73/79 [92.4%]) and carried blaCTX-M-1 (58/79 [73.4%]). The plasmid content of the isolates was dominated by IncHI1 of ST9 (56/62 [90.3%]) and ST2 (6/62 [9.7%]), while 84.5% (49/58) of the blaCTX-M-1 genes were associated with the presence of the IncHI1 replicon of ST9 and 6.9% (4/58) with the IncHI1 replicon of ST2 within the corresponding isolates. The operon for the utilization of short-chain fructooligosaccharides (the fos operon) was present in 55 of 79 (69.6%) isolates, and all of these carried IncHI1/ST9 plasmids. The eight complete IncHI1/ST9 plasmid sequences showed the presence of blaCTX-M-1 and the fos operon within the same molecule. Sequences of IncHI1/ST9 plasmids were highly conserved (>98% similarity) regardless of country of origin and differed only in the structure and integration site of the multidrug resistance (MDR) region. E. coli ST1250 and ST1250-SLV/DLV are phylogenetically diverse strains associated with horses. A strong linkage of E. coli ST1250 with the epidemic multidrug resistance plasmid lineage IncHI1/ST9 carrying blaCTX-M-1 and the fos operon was identified.

scholarly journals Antibiotic Resistance in Vibrio cholerae: Mechanistic Insights from IncC Plasmid-Mediated Dissemination of a Novel Family of Genomic Islands Inserted at trmE

mSphere ◽  
2020 ◽  
Vol 5 (4) ◽  
Author(s):  
Nicolas Rivard ◽  
Rita R. Colwell ◽  
Vincent Burrus
Keyword(s):  
Drug Resistance ◽  
Antibiotic Resistance ◽  
Multidrug Resistance ◽  
Vibrio Cholerae ◽  
Resistance Genes ◽  
Mobile Genetic Elements ◽  
Genomic Islands ◽  
Phage Resistance ◽  
Cholera Outbreak ◽  
Content Type

ABSTRACT Cholera remains a formidable disease, and reports of multidrug-resistant strains of the causative agent Vibrio cholerae have become common during the last 3 decades. The pervasiveness of resistance determinants has largely been ascribed to mobile genetic elements, including SXT/R391 integrative conjugative elements, IncC plasmids, and genomic islands (GIs). Conjugative transfer of IncC plasmids is activated by the master activator AcaCD whose regulatory network extends to chromosomally integrated GIs. MGIVchHai6 is a multidrug resistance GI integrated at the 3′ end of trmE (mnmE or thdF) in chromosome 1 of non-O1/non-O139 V. cholerae clinical isolates from the 2010 Haitian cholera outbreak. In the presence of an IncC plasmid expressing AcaCD, MGIVchHai6 excises from the chromosome and transfers at high frequency. Herein, the mechanism of mobilization of MGIVchHai6 GIs by IncC plasmids was dissected. Our results show that AcaCD drives expression of GI-borne genes, including xis and mobIM, involved in excision and mobilization. A 49-bp fragment upstream of mobIM was found to serve as the minimal origin of transfer (oriT) of MGIVchHai6. The direction of transfer initiated at oriT was determined using IncC plasmid-driven mobilization of chromosomal markers via MGIVchHai6. In addition, IncC plasmid-encoded factors, including the relaxase TraI, were found to be required for GI transfer. Finally, in silico exploration of Gammaproteobacteria genomes identified 47 novel related and potentially AcaCD-responsive GIs in 13 different genera. Despite sharing conserved features, these GIs integrate at trmE, yicC, or dusA and carry a diverse cargo of genes involved in phage resistance. IMPORTANCE The increasing association of the etiological agent of cholera, Vibrio cholerae serogroup O1 and O139, with multiple antibiotic resistance threatens to deprive health practitioners of this effective tool. Drug resistance in cholera results mainly from acquisition of mobile genetic elements. Genomic islands conferring multidrug resistance and mobilizable by IncC conjugative plasmids were reported to circulate in non-O1/non-O139 V. cholerae clinical strains isolated from the 2010 Haitian cholera outbreak. As these genomic islands can be transmitted to pandemic V. cholerae serogroups, their mechanism of transmission needed to be investigated. Our research revealed plasmid- and genomic island-encoded factors required for the resistance island excision, mobilization, and integration, as well as regulation of these functions. The discovery of related genomic islands carrying diverse phage resistance genes but lacking antibiotic resistance-conferring genes in a wide range of marine dwelling bacteria suggests that these elements are ancient and recently acquired drug resistance genes.

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