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635300.9828Diversity of silver resistance genes in IncH incompatibility group plasmids. Silver compounds are used as antimicrobial agents in medicine and bacteria that develop resistance to silver cations (Ag(+)) pose problems similar to those of antibiotic-resistant bacteria. The first set of Ag(+) resistance genes (sil) was from plasmid pMG101, now assigned to the IncHI incompatibility group. Questions of whether sil genes are unique to pMG101 or are more widely found, and whether they are associated with a specific incompatibility group or occur in many plasmid groups and on bacterial chromosomes were addressed. sil genes were identified in five IncH plasmids, but not in plasmids of the IncP incompatibility group. Three sil genes (silP, silR and silE) from these plasmids were PCR-amplified, cloned, sequenced and compared to those of pMG101. Differences of 0-50 nt per kb of sequence were found. Predicted gene products were 0-6% different in amino acid sequence, but the differences did not alter residues thought to be involved in protein function (see supplementary data at http://mic.sgmjournals.org or http://www.uic.edu/depts/mcmi/individual/gupta/index.htm). For representative IncH plasmid R476b and pMG101 the effects of Ag(+) exposure on resistance levels were measured by growth. The inducibility of silC, silR and silE gene expression after Ag(+) exposure was studied by reverse transcriptase (RT)-PCR. Silver resistance increased after Ag(+) exposure for strains carrying plasmid R476b. silC and silE expression from R476b was inducible after Ag(+) exposure and was constitutive and high from pMG101. The mRNA levels for the regulatory gene silR was constitutive for both pMG101 and R476b. Close homologues for silABC(ORF96)RS from pMG101 are clustered on the chromosomes of Escherichia coli strains K-12 and O157:H7, without contiguous silP and silE homologues. Insertion deletions of the E. coli K-12 chromosomal homologues for silA and silP gave Ag(+) hypersensitivity for growth. The silA homologue knockout was complemented back to wild-type resistance by the same gene cloned on a plasmid. Homologues of sil genes have also been identified on other enterobacterial genomes.200111739772
635010.9789Characterization and genomic analysis of chromate resistant and reducing Bacillus cereus strain SJ1. BACKGROUND: Chromium is a toxic heavy metal, which primarily exists in two inorganic forms, Cr(VI) and Cr(III). Chromate [Cr(VI)] is carcinogenic, mutational, and teratogenic due to its strong oxidizing nature. Biotransformation of Cr(VI) to less-toxic Cr(III) by chromate-resistant and reducing bacteria has offered an ecological and economical option for chromate detoxification and bioremediation. However, knowledge of the genetic determinants for chromate resistance and reduction has been limited so far. Our main aim was to investigate chromate resistance and reduction by Bacillus cereus SJ1, and to further study the underlying mechanisms at the molecular level using the obtained genome sequence. RESULTS: Bacillus cereus SJ1 isolated from chromium-contaminated wastewater of a metal electroplating factory displayed high Cr(VI) resistance with a minimal inhibitory concentration (MIC) of 30 mM when induced with Cr(VI). A complete bacterial reduction of 1 mM Cr(VI) was achieved within 57 h. By genome sequence analysis, a putative chromate transport operon, chrIA1, and two additional chrA genes encoding putative chromate transporters that likely confer chromate resistance were identified. Furthermore, we also found an azoreductase gene azoR and four nitroreductase genes nitR possibly involved in chromate reduction. Using reverse transcription PCR (RT-PCR) technology, it was shown that expression of adjacent genes chrA1 and chrI was induced in response to Cr(VI) but expression of the other two chromate transporter genes chrA2 and chrA3 was constitutive. In contrast, chromate reduction was constitutive in both phenotypic and gene expression analyses. The presence of a resolvase gene upstream of chrIA1, an arsenic resistance operon and a gene encoding Tn7-like transposition proteins ABBCCCD downstream of chrIA1 in B. cereus SJ1 implied the possibility of recent horizontal gene transfer. CONCLUSION: Our results indicate that expression of the chromate transporter gene chrA1 was inducible by Cr(VI) and most likely regulated by the putative transcriptional regulator ChrI. The bacterial Cr(VI)-resistant level was also inducible. The presence of an adjacent arsenic resistance gene cluster nearby the chrIA1 suggested that strong selective pressure by chromium and arsenic could cause bacterial horizontal gene transfer. Such events may favor the survival and increase the resistance level of B. cereus SJ1.201020723231
521720.9789UV Resistance of bacteria from the Kenyan Marine cyanobacterium Moorea producens. UV resistance of bacteria isolated from the marine cyanobacterium Moorea producens has not been observed previously, findings which highlight how unsafe germicidal UV irradiation for sterilization of air, food, and water could be. Further, UV resistance of Bacillus licheniformis is being observed for the first time. This study focused on bacteria isolated from the marine cyanobacterium M. producens collected off the Kenyan coast at Shimoni, Wasini, Kilifi, and Mida. UV irradiance of isolates (302 nm, 70 W/m(2) , 0-1 hr) established B. licheniformis as the most UV resistant strain, with the following order of taxon resistance: Bacilli> γ proteobacteria > Actinobacteria. UV resistance was independent of pigmentation. The maximum likelihood phylogenetic distance determined for both B. licheniformis and Bacillus aerius relative to M. producens CCAP 1446/4 was 2.0. Survival of B. licheniformis upon UV irradiance followed first-order kinetics (k = 0.035/min, R(2)  = 0.88). Addition of aqueous extracts (2, 10, 20 and 40 mg/ml) of this B. licheniformis strain on the less resistant Marinobacterium stanieri was not significant, however, the commercial sunscreen benzophenone-3 (BP-3) positive control and the time of irradiance were significant. Detection of bacteria on M. producens filaments stained with acridine orange confirmed its nonaxenic nature. Although the chemistry of UV resistance in cyanobacteria has been studied in depth revealing for example the role of mycosporine like amino acids (MAAs) in UV resistance less is known about how bacteria resist UV irradiation. This is of interest since cyanobacteria live in association with bacteria.201930123980
522130.9781Molecular cloning of the DNA gyrase genes from Methylovorus sp. strain SS1 and the mechanism of intrinsic quinolone resistance in methylotrophic bacteria. The genes encoding the DNA gyrase A (GyrA) and B subunits (GyrB) of Methylovorus sp. strain SS1 were cloned and sequenced. gyrA and gyrB coded for proteins of 846 and 799 amino acids with calculated molecular weights of 94,328 and 88,714, respectively, and complemented Escherichia coli gyrA and gyrB temperature sensitive (ts) mutants. To analyze the role of type II topoisomerases in the intrinsic quinolone resistance of methylotrophic bacteria, the sequences of the quinolone resistance-determining regions (QRDRs) in the A subunit of DNA gyrase and the C subunit (ParC) of topoisomerase IV (Topo IV) of Methylovorus sp. strain SS1, Methylobacterium extorquens AM1 NCIB 9133, Methylobacillus sp, strain SK1 DSM 8269, and Methylophilus methylotrophus NCIB 10515 were determined. The deduced amino acid sequences of the QRDRs of the ParCs in the four methylotrophic bacteria were identical to that of E. coli ParC. The sequences of the QRDR in GyrA were also identical to those in E. coli GyrA except for the amino acids at positions 83, 87, or 95. The Ser83 to Thr substitution in Methylovorus sp. strain SS1, and the Ser83 to Leu and Asp87 to Asn substitutions in the three other methylotrophs, agreed well with the minimal inhibitory concentrations of quinolones in the four bacteria, suggesting that these residues play a role in the intrinsic susceptibility of methylotrophic bacteria to quinolones.200516404155
614940.9781Characterization and whole-genome sequencing of an extreme arsenic-tolerant Citrobacter freundii SRS1 strain isolated from Savar area in Bangladesh. Citrobacter freundii SRS1, gram-negative bacteria, were isolated from Savar, Bangladesh. The strain could tolerate up to 80 mmol L(-1) sodium arsenite, 400 mmol L(-1) sodium arsenate, 5 mmol L(-1) manganese sulfate, 3 mmol L(-1) lead nitrate, 2.5 mmol L(-1) cobalt chloride, 2.5  mmol L(-1) cadmium acetate, and 2.5 mmol L(-1) chromium chloride. The whole-genome sequencing revealed that the genome size of C. freundii SRS1 is estimated to be 5.4 Mb long, and the G + C content is 51.7%. The genome of C. freundii SRS1 contains arsA, arsB, arsC, arsD, arsH, arsR, and acr3 genes for arsenic resistance; czcA, czcD, cbiN, and cbiM genes for cobalt resistance; chrA and chrB genes for chromium resistance; mntH, sitA, sitB, sitC, and sitD genes for manganese resistance; and zntA gene for lead and cadmium resistance. This novel acr3 gene has never previously been reported in any C. freundii strain except SRS1. A set of 130 completely sequenced strains of C. freundii was selected for phylogenomic analysis. The phylogenetic tree showed that the SRS1 strain is closely related to the C. freundii 62 strain. Further analyses of the genes involved in metal and metalloid resistance might facilitate identifying the mechanisms and pathways involved in high metal resistance in the C. freundii SRS1 strain.202336332226
45550.9780An inducible tellurite-resistance operon in Proteus mirabilis. Tellurite resistance (Te(r)) is widespread in nature and it is shown here that the natural resistance of Proteus mirabilis to tellurite is due to a chromosomally located orthologue of plasmid-borne ter genes found in enteric bacteria. The P. mirabilis ter locus (terZABCDE) was identified in a screen of Tn5lacZ-generated mutants of which one contained an insertion in terC. The P. mirabilis terC mutant displayed increased susceptibility to tellurite (Te(s)) and complementation with terC carried on a multicopy plasmid restored high-level Te(r). Primer extension analysis revealed a single transcriptional start site upstream of terZ, but only with RNA harvested from bacteria grown in the presence of tellurite. Northern blotting and reverse transcriptase-PCR (RT-PCR) analyses confirmed that the ter operon was inducible by tellurite and to a lesser extent by oxidative stress inducers such as hydrogen peroxide and methyl viologen (paraquat). Direct and inverted repeat sequences were identified in the ter promoter region as well as motifs upstream of the -35 hexamer that resembled OxyR-binding sequences. Finally, the 390 bp intergenic promoter region located between orf3 and terZ showed no DNA sequence identity with any other published ter sequences, whereas terZABCDE genes exhibited 73-85 % DNA sequence identity. The ter operon was present in all clinical isolates of P. mirabilis and Proteus vulgaris tested and is inferred for Morganella and Providencia spp. based on screening for high level Te(r) and preliminary PCR analysis. Thus, a chromosomally located inducible tellurite resistance operon appears to be a common feature of the genus Proteus.200312724390
619560.9779Differential gene expression analysis shows that cephalosporin resistance is intrinsic to Clostridioides difficile strain 630. Clostridioides difficile infection (CDI) is the most common nosocomial infection in the US. CDI has become a growing concern due to C. difficile's resistance to several antibiotics, including cephalosporins. Furthermore, patients administered cephalosporins are at higher risk of contracting CDI. Cephalosporins are β-lactam antibiotics, which prevent bacterial cell wall synthesis by inhibiting penicillin-binding proteins (PBPs). β-lactam-resistant bacteria evade these antibiotics by producing β-lactamases or by harboring low-affinity PBPs. A genomic analysis of C. difficile strain 630 identified 31 putative β-lactam resistance genes. Upon cefoxitin exposure, few C. difficile strain 630 putative antibiotic-resistant genes were overexpressed. Most notably, the β-lactamase blaCDD gene was upregulated approximately 600-fold, as previously reported. Deletion of the blaCDD locus did not change in cephalosporin susceptibility. Deletion of the second most upregulated gene, the PBP vanY, was also ineffective at decreasing cephalosporin resistance. Cefoxitin exposure of the C. difficile strain 630ΔblaCDD mutant did not increase upregulation of other putative antibiotic resistance genes compared to wildtype C. difficile strain 630. Transcriptomic analyses of wildtype C. difficile strain 630 exposed to cephradine, cefoxitin, ceftazidime, or cefepime revealed the shared upregulation of a putative heterodimeric ABC transporter encoded by loci CD630_04590 (ABC transporter ATP-binding protein) and CD630_04600 (ABC transporter permease). These genes are genomically located directly downstream of blaCDD (CD630_04580). The deletion mutant CD630_04600 remained resistant to a number of antibiotics. Thus, even though blaCDD, CD630_04590, and CD630_04600 are all upregulated when exposed to cephalosporins, they do not seem to be involved in antibiotic resistance in C. difficile strain 630.202539672901
634870.9779Overexpression of cold shock protein A of Psychromonas arctica KOPRI 22215 confers cold-resistance. A polar bacterium was isolated from Arctic sea sediments and identified as Psychromonas artica, based on 16S rDNA sequence. Psychromonas artica KOPRI 22215 has an optimal growth temperature of 10 degrees C and a maximum growth temperature of 25 degrees C, suggesting this bacterium is a psychrophile. Cold shock proteins (Csps) are induced upon temperature downshift by more than 10 degrees C. Functional studies have researched mostly Csps of a mesophilic bacterium Escherichia coli, but not on those of psychrophilic bacteria. In an effort to understand the molecular mechanisms of psychrophilic bacteria that allow it withstand freezing environments, we cloned a gene encoding a cold shock protein from P. artica KOPRI 22215 (CspA(Pa)) using the conserved sequences in csp genes. The 204 bp-long ORF encoded a protein of 68 amino acids, sharing 56% homology to previously reported E. coli CspA protein. When CspA(Pa) was overexpressed in E. coli, it caused cell growth-retardation and morphological elongation. Interestingly, overexpression of CspA(Pa) drastically increased the host's cold-resistance by more than ten times, suggesting the protein aids survival in polar environments.201020169403
615180.9779Novel arsenic hyper-resistant bacteria from an extreme environment, Crven Dol mine, Allchar, North Macedonia. Novel hyper-resistant bacteria were isolated from the Crven Dol mine (Allchar, North Macedonia), arsenic-rich extreme environment. Bacteria were recovered from a secondary mineral mixture, an alteration of hydrothermal realgar rich in arsenates (pharmacolite, hornesite, and talmessite). The sample was recovered from the dark part of the mine at 28 m depth. Three bacterial strains and a bacterial consortium were isolated for their capacity to survive exposure to 32 g/L (209 mM) of arsenite, and 176 g/L (564 mM) of arsenate. The 16S rRNA gene analysis identified bacterial isolates as Stenotrophomonas sp. and two Microbacterium spp. This analysis also revealed that bacterial consortium comprise two Bacteriodetes exhibiting similarity to Olivibacter ginsengisoli and to uncultured bacterium, and one γ-proteobacteria with similarity to Luteimonas sp. Among all isolates Stenotrophomonas sp. exhibited the highest tolerance to As compound as well as the capacity to accumulate As inside the cells. Analysis of genes involved in As-resistance showed that recovered isolates possess the genes encoding the ArsB, Acr3(1) and Acr3(2) proteins, indicating that at least a part of their resistance could be ascribed to As-efflux systems described in isolates obtained from human-polluted environments.202132712355
18890.9779Resistance to ag(i) cations in bacteria: environments, genes and proteins. Bacterial resistance to Ag(I) has been reported periodically with isolates from many environments where toxic levels of silver might be expected to occur, but initial reports were limited to the occurrence of resistant bacteria. The availability of silver-resistance conferring DNA sequences now allow genetic and mechanistic studies that had basically been missing. The genes determining Ag(I) resistance were sequenced from a plasmid found in a burn ward isolate. The 14.2 kb determinant contains seven recognized genes, arranged in three mRNA transcriptional units. The silE gene determines an extracellular (periplasmic space) metal-binding protein of 123 amino acids, including ten histidine residues implicated in Ag(I) binding. SilE is homologous to PcoE, of copper resistance. The next two genes, silR and silS, determine a two protein, histidine-kinase membrane sensor and aspartyl phosphate transcriptional responder, similar to other two component systems such as CzcR and CzcS (for cadmium, zinc and cobalt resistance) and PcoR and PcoS (for copper resistance). The remaining four genes, silCBAP, are co-transcribed and appear to determine Ag(+) efflux, with SilCBA homologous to CzcCBA, a three component cation/proton antiporter, and SilP a novel P-type ATPase with a amino-terminal histidine-rich cation-specificity region. The effects of increasing Ag(+) concentrations and growth medium halides (Cl-, Br- and I-) have been characterized, with lower Cl- concentrations facilitating resistance and higher concentrations toxicity. The properties of this unique Ag(I)-binding SilE protein are being characterized. Sequences similar to the silver-resistance DNA are being characterized by Southern blot DNA/DNA hybridization, PCR in vitro DNA synthesis and DNA sequencing. More than 25 additional closely related sequences have been identified in bacteria from diverse sources. Initial DNA sequencing results shows approximately 5-20% differences in DNA sequences.199918475907
5770100.9778Prevalence of silver resistance genes in bacteria isolated from human and horse wounds. The aim of this study was to investigate the prevalence of silver resistance genes in 172 bacterial strains which had been isolated from both human and equine wounds. PCR screening for 8 currently named genes in 3 silver resistance transcriptional units, silE, silRS and silP, silCBA and silF was performed on total DNA extracted from all clinical isolates. Plasmids were isolated from sil-positive strains to determine if the genes were present on the chromosome. MICs and zone of inhibition assays were utilised to examine phenotypic resistance to silver nitrate and ionic silver. Evidence of silver resistance genes was demonstrated in six strains of Enterobacter cloacae, an organism rarely implicated as a primary pathogen in chronic wounds. MIC data showed that all strains were inhibited at silver nitrate concentrations > or =5mg/L. When tested against a silver-containing absorbent wound dressing all strains showed inhibition of growth after 24h. In MIC and zone of inhibition studies, inhibition was evident but reduced in strains which contained sil genes. Although sil genes were found in six of the wound isolates studied, the genes were consistently associated with a non-pathogenic bacterium. Furthermore, investigation of phenotypic resistance in sil-positive isolates showed that silver continued to be effective.200919362435
5218110.9777Expression of a Shiga-Like Toxin during Plastic Colonization by Two Multidrug-Resistant Bacteria, Aeromonas hydrophila RIT668 and Citrobacter freundii RIT669, Isolated from Endangered Turtles (Clemmys guttata). Aeromonas hydrophila RIT668 and Citrobacter freundii RIT669 were isolated from endangered spotted turtles (Clemmys guttata). Whole-genome sequencing, annotation and phylogenetic analyses of the genomes revealed that the closest relative of RIT668 is A. hydrophila ATCC 7966 and Citrobacter portucalensis A60 for RIT669. Resistome analysis showed that A. hydrophila and C. freundii harbor six and 19 different antibiotic resistance genes, respectively. Both bacteria colonize polyethylene and polypropylene, which are common plastics, found in the environment and are used to fabricate medical devices. The expression of six biofilm-related genes-biofilm peroxide resistance protein (bsmA), biofilm formation regulatory protein subunit R (bssR), biofilm formation regulatory protein subunit S (bssS), biofilm formation regulator (hmsP), toxin-antitoxin biofilm protein (tabA) and transcriptional activator of curli operon (csgD)-and two virulence factors-Vi antigen-related gene (viaB) and Shiga-like toxin (slt-II)-was investigated by RT-PCR. A. hydrophila displayed a >2-fold increase in slt-II expression in cells adhering to both polymers, C. freundii adhering on polyethylene displayed a >2-fold, and on polypropylene a >6-fold upregulation of slt-II. Thus, the two new isolates are potential pathogens owing to their drug resistance, surface colonization and upregulation of a slt-II-type diarrheal toxin on polymer surfaces.202032752245
405120.9776Characterization of a small plasmid (pMBCP) from bovine Pseudomonas pickettii that confers cadmium resistance. This is the first report of isolation of Pseudomonas pickettii from a normal adult bovine duodenum. This organism was one of several bacteria isolated as part of a study to examine cadmium resistance genes (cad(r)) for use in generating transgenic plants to reclaim cadmium-contaminated soils in Kansas. P. pickettii containing a plasmid of 2.2kb (designated pMBCP) grew in Luria-Bertani broth and agar containing up to 800 microM of cadmium chloride and was resistant to 16 antibiotics. Curing the organism of plasmid revealed that antibiotic resistances were not plasmid-mediated. Low-level cadmium resistance was conferred by the plasmid because uncured organism grew significantly better (P<0.05) at 55 microM compared to cured organism. Both plasmid and chromosomal DNA were probed by DNA-DNA hybridization for the presence of known cadmium resistance genes (cadA, cadC, and cadD from Gram-positive (Staphylococcus aureus), but none were detected. The plasmid had one restriction site each for BamHI, PstI, SmaI, and XhoI; two sites each for HincII, SacI, and SphI; and multiple sites for AluI and XcmI. DNA sequence analyses of the cloned and original plasmids showed a GC content of greater than 60% and no homology to any published sequences in the GenBank, European Bioinformatics Institute, or Japanese Genome Net databases. The DNA sequence is contained in GenBank accession number AF144733. Thus, pMBCP offers low-level cadmium resistance to P. picketttii.200312651180
494130.9775The mercury resistance operon of the IncJ plasmid pMERPH exhibits structural and regulatory divergence from other Gram-negative mer operons. The bacterial mercury resistance determinant carried on the IncJ plasmid pMERPH has been characterized further by DNA sequence analysis. From the sequence of a 4097 bp Bg/II fragment which confers mercury resistance, it is predicted that the determinant consists of the genes merT, merP, merC and merA. The level of DNA sequence similarity between these genes and those of the mer determinant of Tn21 was between 56 center dot 4 and 62 center dot 4%. A neighbour-joining phylogenetic tree of merA gene sequences was constructed which suggested that pMERPH bears the most divergent Gram-negative mer determinant characterized to date. Although the determinant from pMERPH has been shown to be inducible, no regulatory genes have been found within the Bg/II fragment and it is suggested that a regulatory gene may be located elsewhere on the plasmid. The cloned determinant has been shown to express mercury resistance constitutively. Analysis of the pMERPH mer operator/promoter (O/P) region in vivo has shown constitutive expression from the mer PTCPA promoter, which could be partially repressed by the presence of a trans-acting MerR protein from a Tn21-like mer determinant. This incomplete repression of mer PTCPA promoter activity may be due to the presence of an extra base between the -35 and -10 sequences of the promoter and/or to variation in the MerR binding sites in the O/P region. Expression from the partially repressed mer PTCPA promoter could be restored by the addition of inducing levels of Hg2+ ions. Using the polymerase chain reaction with primers designed to amplify regions in the merP and merA genes, 1 center dot 37 kb pMERPH-like sequences have been amplified from the IncJ plasmid R391, the environmental isolate SE2 and from DNA isolated directly from non-cultivated bacteria in River Mersey sediment. This suggests that pMERPH-like sequences, although rare, are nevertheless persistent in natural environments.19968932707
481140.9775Characterization and structure prediction of partial length protein sequences of pcoA, pcoR and chrB genes from heavy metal resistant bacteria from the Klip River, South Africa. The Klip River has suffered from severe anthropogenic effects from industrial activities such as mining. Long-term exposure to heavy metal pollution has led to the development of heavy metal resistant strains of Pseudomonas sp. KR23, Lysinibacillus sp. KR25, and E. coli KR29. The objectives of this study were to characterize the genetics of copper and chromate resistance of the isolates. Copper and chromate resistance determinants were cloned and sequenced. Open reading frames (ORFs) related to the genes CopA and CopR were identified in E. coli KR29, PcoA in Lysinibacillus sp. KR25 and none related to chromate resistance were detected. The 3D-models predicted by I-TASSER disclose that the PcoA proteins consist of β-sheets, which form a part of the cupredoxin domain of the CopA copper resistance family of genes. The model for PcoR_29 revealed the presence of a helix turn helix; this forms part of a DNA binding protein, which is part of a heavy metal transcriptional regulator. The bacterial strains were cured using ethidium bromide. The genes encoding for heavy metal resistance and antibiotic resistance were found to be located on the chromosome for both Pseudomonas sp. (KR23) and E. coli (KR29). For Lysinibacillus (KR25) the heavy metal resistance determinants are suspected to be located on a mobile genetic element, which was not detected using gel electrophoresis.201525837632
2446150.9774Low selection of topoisomerase mutants from strains of Escherichia coli harbouring plasmid-borne qnr genes. OBJECTIVES: To investigate mutations in the type II topoisomerase genes in quinolone-resistant mutants selected from bacteria harbouring plasmid-borne qnr genes. METHODS: Mutants were selected by nalidixic acid, ciprofloxacin and moxifloxacin from two Escherichia coli reference strains and corresponding transconjugants harbouring qnrA1, qnrA3, qnrB2 or qnrS1 genes. RESULTS: The proportion of resistant mutants selected by the three quinolones was, respectively, in the same range for qnr-positive transconjugants and reference strains. Only 20% (65/329) of the mutants selected from the transconjugants showed a gyrase mutation, whereas 79% (94/119) of those from the reference strains without a qnr gene did (P < 0.0001). At four times the MIC of the selector quinolone, gyrA mutants represented 49% and 95% of the mutants selected with nalidixic acid, 4% and 94% with ciprofloxacin and 0% and 54% with moxifloxacin for qnr-positive transconjugants and reference strains, respectively. Mutations within gyrA were distributed at codon 87 (D87G, H, N or Y) and at codon 83 (S83L) with three novel mutations (gyrA Ser83stop, gyrA Asp82Asn and gyrB insertion of Glu at 465) and three rare mutations (gyrA Gly81Asp, gyrA Asp82Gly and gyrA Ser431Pro), mainly obtained from reference strains after moxifloxacin selection. Strikingly, none of the mutants selected by moxifloxacin from qnr-positive transconjugants harboured a mutation in the topoisomerase genes. CONCLUSIONS: Topoisomerase mutants are rarely selected by ciprofloxacin and moxifloxacin from strains harbouring qnr. This suggests that the quinolone resistance-determining region domains are protected from quinolones by the Qnr protein and consequently other mechanisms are developed to acquire a further step of fluoroquinolone resistance.200818325893
6160160.9774Comparative transcriptional profiling of tildipirosin-resistant and sensitive Haemophilus parasuis. Numerous studies have been conducted to examine the molecular mechanism of Haemophilus parasuis resistance to antibiotic, but rarely to tildipirosin. In the current study, transcriptional profiling was applied to analyse the variation in gene expression of JS0135 and tildipirosin-resistant JS32. The growth curves showed that JS32 had a higher growth rate but fewer bacteria than JS0135. The cell membranes of JS32 and a resistant clinical isolate (HB32) were observed to be smoother than those of JS0135. From the comparative gene expression profile 349 up- and 113 downregulated genes were observed, covering 37 GO and 63 KEGG pathways which are involved in biological processes (11), cellular components (17), molecular function (9), cellular processes (1), environmental information processing (4), genetic information processing (9) and metabolism (49) affected in JS32. In addition, the relative overexpression of genes of the metabolism pathway (HAPS_RS09315, HAPS_RS09320), ribosomes (HAPS_RS07815) and ABC transporters (HAPS_RS10945) was detected, particularly the metabolism pathway, and verified with RT-qPCR. Collectively, the gene expression profile in connection with tildipirosin resistance factors revealed unique and highly resistant determinants of H. parasuis to macrolides that warrant further attention due to the significant threat of bacterial resistance.201728790420
433170.9773Expression of the strA-strB streptomycin resistance genes in Pseudomonas syringae and Xanthomonas campestris and characterization of IS6100 in X. campestris. Expression of the strA-strB streptomycin resistance (SMr) genes was examined in Pseudomonas syringae pv. syringae and Xanthomonas campestris pv. vesicatoria. The strA-strB genes in P. syringae and X. campestris were encoded on elements closely related to Tn5393 from Erwinia amylovora and designated Tn5393a and Tn5393b, respectively. The putative recombination site (res) and resolvase-repressor (tnpR) genes of Tn5393 from E. amylovora, P syringae, and X. campestris were identical; however, IS6100 mapped within tnpR in X. campestris, and IS1133 was previously located downstream of tnpR in E. amylovora (C.-S Chiou and A. L. Jones, J. Bacteriol. 175:732-740, 1993). Transcriptional fusions (strA-strB::uidA) indicated that a strong promoter sequence was located within res in Tn5393a. Expression from this promoter sequence was reduced when the tnpR gene was present in cis position relative to the promoter. In X. campestris pv. vesicatoria, analysis of promoter activity with transcriptional fusions indicated that IS6100 increased the expression of strA-strB. Analysis of codon usage patterns and percent G+C in the third codon position indicated that IS6100 could have originated in a gram-negative bacterium. The data obtained in the present study help explain differences observed in the levels of SMr expressed by three genera which share common genes for resistance. Furthermore, the widespread dissemination of Tn5393 and derivatives in phytopathogenic prokaryotes confirms the importance of these bacteria as reservoirs of antibiotic resistance in the environment.19957487022
6085180.9773Heavy metal resistance and genotypic analysis of metal resistance genes in gram-positive and gram-negative bacteria present in Ni-rich serpentine soil and in the rhizosphere of Alyssum murale. Forty-six bacterial cultures, including one culture collection strain, thirty from the rhizosphere of Alyssum murale and fifteen from Ni-rich soil, were tested for their ability to tolerate arsenate, cadmium, chromium, zinc, mercury, lead, cobalt, copper, and nickel in their growth medium. The resistance patterns, expressed as minimum inhibitory concentrations, for all cultures to the nine different metal ions were surveyed by using the agar dilution method. A large number of the cultures were resistant to Ni (100%), Pb (100%), Zn (100%), Cu (98%), and Co (93%). However, 82, 71, 58 and 47% were sensitive to As, Hg, Cd and Cr(VI), respectively. All cultures had multiple metal-resistant, with heptametal resistance as the major pattern (28.8%). Five of the cultures (about of 11.2% of the total), specifically Arthrobacter rhombi AY509239, Clavibacter xyli AY509235, Microbacterium arabinogalactanolyticum AY509226, Rhizobium mongolense AY509209 and Variovorax paradoxus AY512828 were tolerant to nine different metals. The polymerase chain reaction in combination with DNA sequence analysis was used to investigate the genetic mechanism responsible for the metal resistance in some of these gram-positive and gram-negative bacteria that were, highly resistant to Hg, Zn, Cr and Ni. The czc, chr, ncc and mer genes that are responsible for resistance to Zn, Cr, Ni and Hg, respectively, were shown to be present in these bacteria by using PCR. In the case of, M. arabinogalactanolyticum AY509226 these genes were shown to have high homology to the czcD, chrB, nccA, and mer genes of Ralstonia metallidurans CH34. Therefore, Hg, Zn, Cr and Ni resistance genes are widely distributed in both gram-positive and gram-negative isolates obtained from A. murale rhizosphere and Ni-rich soils.200717276484
399190.9773Identification of genes conferring arsenic resistance to Escherichia coli from an effluent treatment plant sludge metagenomic library. The majority of bacteria elude culture in the laboratory. A metagenomic approach provides culture-independent access to the gene pool of the whole bacterial community. A metagenomic library was constructed from an industrial effluent treatment plant sludge containing about 1.25 Gb of microbial community DNA. Two arsenic-resistant clones were selected from the metagenomic library. Clones MT3 and MT6 had eight- and 18-fold higher resistance to sodium arsenate in comparison with the parent strain, respectively. The clones also showed increased resistance to arsenite but not to antimony. Sequence analysis of the clones revealed genes encoding for putative arsenate reductases and arsenite efflux pumps. A novel arsenate resistance gene (arsN) encoding a protein with similarity to acetyltransferases was identified from clone MT6. ArsN homologues were found to be closely associated with arsenic resistance genes in many bacterial genomes. ArsN homologues were found fused to putative arsenate reductases in Methylibium petroleiphilum PM1 and Anaeromyxobacter dehalogenans 2CP-C and with a putative arsenite chaperone in Burkholderia vietnamiensis G4. ArsN alone resulted in an approximately sixfold higher resistance to sodium arsenate in wild-type Escherichia coli W3110.200919016868