scholarly journals Synthetic and Evolutionary Construction of a Chlorate-Reducing Shewanella oneidensis MR-1

mBio ◽  
2015 ◽  
Vol 6 (3) ◽  
Author(s):  
Iain C. Clark ◽  
Ryan A. Melnyk ◽  
Matthew D. Youngblut ◽  
Hans K. Carlson ◽  
Anthony T. Iavarone ◽  
...  
Keyword(s):  
Single Nucleotide Polymorphism ◽  
Copy Number ◽  
Shewanella Oneidensis ◽  
Respiratory Metabolism ◽  
Chromosome 11 ◽  
Nucleotide Polymorphism ◽  
Single Nucleotide ◽  
Single Strain ◽  
Content Type ◽  
Essential Components

ABSTRACTDespite evidence for the prevalence of horizontal gene transfer of respiratory genes, little is known about how pathways functionally integrate within new hosts. One example of a mobile respiratory metabolism is bacterial chlorate reduction, which is frequently encoded on composite transposons. This implies that the essential components of the metabolism are encoded on these mobile elements. To test this, we heterologously expressed genes for chlorate reduction fromShewanella algaeACDC in the non-chlorate-reducingShewanella oneidensisMR-1. The construct that ultimately endowed robust growth on chlorate includedcld, a cytochromecgene,clrABDC, and two genes of unknown function. Although strain MR-1 was unable to grow on chlorate after initial insertion of these genes into the chromosome, 11 derived strains capable of chlorate respiration were obtained through adaptive evolution. Genome resequencing indicated that all of the evolved chlorate-reducing strains replicated a large genomic region containing chlorate reduction genes. Contraction in copy number and loss of the ability to reduce chlorate were also observed, indicating that this phenomenon was extremely dynamic. Although most strains contained more than six copies of the replicated region, a single strain with less duplication also grew rapidly. This strain contained three additional mutations that we hypothesized compensated for the low copy number. We remade the mutations combinatorially in the unevolved strain and determined that a single nucleotide polymorphism (SNP) upstream ofcldenabled growth on chlorate and was epistatic to a second base pair change in the NarP binding sequence betweennarQPandnrfAthat enhanced growth.IMPORTANCEThe ability of chlorate reduction composite transposons to form functional metabolisms after transfer to a new host is an important part of their propagation. To study this phenomenon, we engineeredShewanella oneidensisMR-1 into a chlorate reducer. We defined a set of genes sufficient to endow growth on chlorate from a plasmid, but found that chromosomal insertion of these genes was nonfunctional. Evolution of this inoperative strain into a chlorate reducer showed that tandem duplication was a dominant mechanism of activation. While copy number changes are a relatively rapid way of increasing gene dosage, replicating almost 1 megabase of extra DNA is costly. Mutations that alleviate the need for high copy number are expected to arise and eventually predominate, and we identified a single nucleotide polymorphism (SNP) that relieved the copy number requirement. This study uses both rational and evolutionary approaches to gain insight into the evolution of a fascinating respiratory metabolism.

scholarly journals Whole-Genome Single-Nucleotide-Polymorphism Analysis for Discrimination of Clostridium botulinum Group I Strains

2014 ◽  
Vol 80 (7) ◽  
pp. 2125-2132 ◽  
Author(s):  
Narjol Gonzalez-Escalona ◽  
Ruth Timme ◽  
Brian H. Raphael ◽  
Donald Zink ◽  
Shashi K. Sharma
Keyword(s):  
Single Nucleotide Polymorphism ◽  
Clostridium Botulinum ◽  
Toxin Gene ◽  
Polymorphism Analysis ◽  
Snp Analysis ◽  
Whole Genome ◽  
Nucleotide Polymorphism ◽  
Single Nucleotide ◽  
Content Type ◽  
Group I

ABSTRACTClostridium botulinumis a genetically diverse Gram-positive bacterium producing extremely potent neurotoxins (botulinum neurotoxins A through G [BoNT/A-G]). The complete genome sequences of three strains harboring only the BoNT/A1 nucleotide sequence are publicly available. Although these strains contain a toxin cluster (HA+OrfX−) associated with hemagglutinin genes, little is known about the genomes of subtype A1 strains (termed HA−OrfX+) that lack hemagglutinin genes in the toxin gene cluster. We sequenced the genomes of three BoNT/A1-producingC. botulinumstrains: two strains with the HA+OrfX−cluster (69A and 32A) and one strain with the HA−OrfX+cluster (CDC297). Whole-genome phylogenic single-nucleotide-polymorphism (SNP) analysis of these strains along with other publicly availableC. botulinumgroup I strains revealed five distinct lineages. Strains 69A and 32A clustered with theC. botulinumtype A1 Hall group, and strain CDC297 clustered with theC. botulinumtype Ba4 strain 657. This study reports the use of whole-genome SNP sequence analysis for discrimination ofC. botulinumgroup I strains and demonstrates the utility of this analysis in quickly differentiatingC. botulinumstrains harboring identical toxin gene subtypes. This analysis further supports previous work showing that strains CDC297 and 657 likely evolved from a common ancestor and independently acquired separate BoNT/A1 toxin gene clusters at distinct genomic locations.

scholarly journals Copy-Number Variations Measured by Single-Nucleotide–Polymorphism Oligonucleotide Arrays in Patients with Mental Retardation

2007 ◽  
Vol 81 (4) ◽  
pp. 768-779 ◽  
Author(s):  
Janine Wagenstaller ◽  
Stephanie Spranger ◽  
Bettina Lorenz-Depiereux ◽  
Bernd Kazmierczak ◽  
Michaela Nathrath ◽  
...  
Keyword(s):  
Single Nucleotide Polymorphism ◽  
Mental Retardation ◽  
Copy Number ◽  
Copy Number Variations ◽  
Nucleotide Polymorphism ◽  
Single Nucleotide ◽  
Oligonucleotide Arrays

scholarly journals Genomic Investigation Reveals Contaminated Detergent as the Source of an Extended-Spectrum-β-Lactamase-Producing Klebsiella michiganensis Outbreak in a Neonatal Unit

2020 ◽  
Vol 58 (5) ◽  
Author(s):  
Paul Chapman ◽  
Brian M. Forde ◽  
Leah W. Roberts ◽  
Haakon Bergh ◽  
Debra Vesey ◽  
...  
Keyword(s):  
Single Nucleotide Polymorphism ◽  
Klebsiella Oxytoca ◽  
Multidrug Resistant ◽  
Hospital Environment ◽  
Nucleotide Polymorphism ◽  
Taxonomic Assignment ◽  
Single Nucleotide ◽  
Content Type ◽  
Extended Spectrum ◽  
Antimicrobial Resistance Genes

ABSTRACT Klebsiella species are problematic pathogens in neonatal units and may cause outbreaks, for which the sources of transmission may be challenging to elucidate. We describe the use of whole-genome sequencing (WGS) to investigate environmental sources of transmission during an outbreak of extended-spectrum-β-lactamase (ESBL)-producing Klebsiella michiganensis colonizing neonates. Ceftriaxone-resistant Klebsiella spp. isolated from neonates (or their mothers) and the hospital environment were included. Short-read sequencing (Illumina) and long-read sequencing (MinION; Oxford Nanopore Technologies) were used to confirm species taxonomy, to identify antimicrobial resistance genes, and to determine phylogenetic relationships using single-nucleotide polymorphism profiling. A total of 21 organisms (10 patient-derived isolates and 11 environmental isolates) were sequenced. Standard laboratory methods identified the outbreak strain as an ESBL-producing Klebsiella oxytoca, but taxonomic assignment from WGS data suggested closer identity to Klebsiella michiganensis. Strains isolated from multiple detergent-dispensing bottles were either identical or closely related by single-nucleotide polymorphism comparison. Detergent bottles contaminated by K. michiganensis had been used for washing milk expression equipment. No new cases were identified once the detergent bottles were removed. Environmental reservoirs may be an important source in outbreaks of multidrug-resistant organisms. WGS, in conjunction with traditional epidemiological investigation, can be instrumental in revealing routes of transmission and guiding infection control responses.

scholarly journals Genetic diversity of released Malaysian rice varieties based on single nucleotide polymorphism markers

2020 ◽  
Vol 56 (No. 2) ◽  
pp. 62-70 ◽  
Author(s):  
Shahril Ab Razak ◽  
Nor Helwa Ezzah Nor Azman ◽  
Rahiniza Kamaruzaman ◽  
Shamsul Amri Saidon ◽  
Muhammad Fairuz Mohd Yusof ◽  
...  
Keyword(s):  
Genetic Diversity ◽  
Single Nucleotide Polymorphism ◽  
Crop Improvement ◽  
Snp Markers ◽  
Group Method ◽  
Chromosome 11 ◽  
Nucleotide Polymorphism ◽  
Rice Varieties ◽  
Single Nucleotide ◽  
K Value

Understanding genetic diversity is a main key for crop improvement and genetic resource management. In this study, we aim to evaluate the genetic diversity of the released Malaysian rice varieties using single nucleotide polymorphism (SNP) markers. A total of 46 released Malaysian rice varieties were genotyped using 1536 SNP markers to evaluate their diversity. Out of 1536 SNPs, only 932 SNPs (60.7%) represented high quality alleles, whereas the remainder either failed to amplify or had low call rates across the samples. Analysis of the 932 SNPs revealed that a total of 16 SNPs were monomorphic. The analysis of the SNPs per chromosome revealed that the average of the polymorphic information content (PIC) value ranged from 0.173 for chromosome 12 to 0.259 for chromosome 11, with an average of 0.213 per locus. The genetic analysis of the 46 released Malaysian rice varieties using an unweighted pair group method with arithmetic mean (UPGMA) dendrogram revealed the presence of two major groups. The analysis was supported by the findings from the STRUCTURE analysis which indicated the ∆K value to be at the highest peak at K = 2, followed by K = 4. The pairwise genetic distance of the shared alleles showed that the value ranged from 0.000 (MR159–MR167) to 0.723 (MRIA–Setanjung), which suggested that MR159 and MR167 were identical, and that the highest dissimilarity was detected between MRIA 1 and Setanjung. The results of the study will be very useful for the variety identification, the proper management and conservation of the genetic resources, and the exploitation and utilisation in future breeding programmes.

scholarly journals Quantitative Analysis of Single-Nucleotide Polymorphism for Rapid Detection of TR34/L98H- and TR46/Y121F/T289A-Positive Aspergillus fumigatus Isolates Obtained from Patients in Iran from 2010 to 2014

2015 ◽  
Vol 60 (1) ◽  
pp. 387-392 ◽  
Author(s):  
Faezeh Mohammadi ◽  
Seyed Jamal Hashemi ◽  
Jan Zoll ◽  
Willem J. G. Melchers ◽  
Haleh Rafati ◽  
...  
Keyword(s):  
Single Nucleotide Polymorphism ◽  
Quantitative Analysis ◽  
Single Nucleotide Polymorphisms ◽  
Aspergillus Fumigatus ◽  
Tandem Repeat ◽  
Promoter Region ◽  
Nucleotide Polymorphisms ◽  
Nucleotide Polymorphism ◽  
Single Nucleotide ◽  
Content Type

ABSTRACTWe employed an endpoint genotyping method to update the prevalence rate of positivity for the TR34/L98H mutation (a 34-bp tandem repeat mutation in the promoter region of thecyp51Agene in combination with a substitution at codon L98) and the TR46/Y121F/T289A mutation (a 46-bp tandem repeat mutation in the promoter region of thecyp51Agene in combination with substitutions at codons Y121 and T289) among clinicalAspergillus fumigatusisolates obtained from different regions of Iran over a recent 5-year period (2010 to 2014). The antifungal activities of itraconazole, voriconazole, and posaconazole against 172 clinicalA. fumigatusisolates were investigated using the European Committee on Antimicrobial Susceptibility Testing (EUCAST) broth microdilution method. For the isolates with an azole resistance phenotype, thecyp51Agene and its promoter were amplified and sequenced. In addition, using a LightCycler 480 real-time PCR system, a novel endpoint genotyping analysis method targeting single-nucleotide polymorphisms was evaluated to detect the L98H and Y121F mutations in thecyp51Agene of all isolates. Of the 172A. fumigatusisolates tested, the MIC values of itraconazole (≥16 mg/liter) and voriconazole (>4 mg/liter) were high for 6 (3.5%). Quantitative analysis of single-nucleotide polymorphisms showed the TR34/L98H mutation in thecyp51Agenes of six isolates. No isolates harboring the TR46/Y121F/T289A mutation were detected. DNA sequencing of thecyp51Agene confirmed the results of the novel endpoint genotyping method. By microsatellite typing, all of the azole-resistant isolates had genotypes different from those previously recovered from Iran and from the Dutch TR34/L98H controls. In conclusion, there was not a significant increase in the prevalence of azole-resistantA. fumigatusisolates harboring the TR34/L98H resistance mechanism among isolates recovered over a recent 5-year period (2010 to 2014) in Iran. A quantitative assay detecting a single-nucleotide polymorphism in thecyp51Agene ofA. fumigatusis a reliable tool for the rapid screening and monitoring of TR34/L98H- and TR46/Y121F/T289A-positive isolates and can easily be incorporated into clinical mycology algorithms.

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