scholarly journals Association of Genetically Related Escherichia coli Strains with Ulcerative Colitis - A Preliminary Study

2011 ◽  
Vol 2 (2) ◽  
pp. 93-96
Author(s):  
Ritu Agarwal ◽  
Chaman Deep ◽  
Saurabh K Patel ◽  
Ashok K Jain ◽  
Gopal Nath
Keyword(s):  
Escherichia Coli ◽  
Ulcerative Colitis ◽  
Genetic Relatedness ◽  
Bacterial Species ◽  
Rectal Mucosa ◽  
Whole Genome ◽  
Medical Sciences ◽  
Whole Genome Analysis ◽  
E Coli ◽  
Major Cluster

Objective: To explore the genetic relatedness among the Escherichia coli isolates recovered from rectal mucosa of patients with Ulcerative Colitis(UC) as well as from non specific diarrhoea patients by using ERIC PCR (whole genome analysis). Material & Methods: A total of 44 strains of E coli, each from patients suffering from UC with exacerbation while on maintenance therapy, were isolated to see if there is any association with specific genotype of E coli with the clini-cal condition. For comparison, 20 strains of E coli were also isolated from patients suffering from non specific diarrhoea. These isolates were subjected to ERIC PCR for analysing similarity/ dissimilarity with each other based on the distribution of ERIC sequences in the whole genome of the bacterial species. Results: The dendrogram prepared on the basis of banding pattern showed that majority of UC patients (39/44, 88.6%) grouped in to one major cluster while second major cluster comprised mostly strains isolated from patients with non specific diarrhoea i.e. controls (17/18, 94.4%). Moreover, in the cluster representing UC patients, a total of 11 strains were observed to be genotypically similar followed by 8 strains by ERIC PCR. Conclusion: Our results strongly indicate that specific Escherichia coli strains may be involved/ associated with UC and its relapse. Key Words: Ulcerative colitis; Escherichia coli; ERIC; PCR DOI: http://dx.doi.org/10.3126/ajms.v2i2.4769Asian Journal of Medical Sciences 2 (2011) 93-96

scholarly journals Comparative Whole Genome Analysis of Escherichia coli O157:H7 Isolates From Feedlot Cattle to Identify Genotypes Associated With the Presence and Absence of stx Genes

2021 ◽  
Vol 12 ◽  
Author(s):  
Mo Jia ◽  
Ifigenia Geornaras ◽  
Jennifer N. Martin ◽  
Keith E. Belk ◽  
Hua Yang
Keyword(s):  
Escherichia Coli ◽  
Metabolic Pathway ◽  
Genome Analysis ◽  
Feedlot Cattle ◽  
Phylogenomic Analysis ◽  
Whole Genome ◽  
Foodborne Outbreak ◽  
Whole Genome Analysis ◽  
E Coli ◽  
Red Recombination

A comparative whole genome analysis was performed on three newly sequenced Escherichia coli O157:H7 strains with different stx profiles, previously isolated from feedlot cattle [C1-010 (stx1−, stx2c+), C1-057 (stx−), and C1-067 (stx1+, stx2a+)], as well as five foodborne outbreak strains and six stx-negative strains from NCBI. Phylogenomic analysis demonstrated that the stx2c-carrying C1-010 and stx-negative C1-057 strains were grouped with the six NCBI stx-negative E. coli O157:H7 strains in Cluster 1, whereas the stx2a-carrying C1-067 and five foodborne outbreak strains were clustered together in Cluster 2. Based on different clusters, we selected the three newly sequenced strains, one stx2a-carrying strain, and the six NCBI stx-negative strains and identify their prophages at the stx insertion sites. All stx-carrying prophages contained both the three Red recombination genes (exo, bet, gam) and their repressor cI. On the other hand, the majority of the stx-negative prophages carried only the three Red recombination genes, but their repressor cI was absent. In the absence of the repressor cI, the consistent expression of the Red recombination genes in prophages might result in more frequent gene exchanges, potentially increasing the probability of the acquisition of stx genes. We further investigated each of the 10 selected E. coli O157:H7 strains for their respective unique metabolic pathway genes. Seven unique metabolic pathway genes in the two stx2a-carrying strains and one in the single stx2c-carrying and seven stx-negative strains were found to be associated with an upstream insertion sequence 629 within a conserved region among these strains. The presence of more unique metabolic pathway genes in stx2a-carrying E. coli O157:H7 strains may potentially increase their competitiveness in complex environments, such as feedlot cattle. For the stx2c-carrying and stx-negative E. coli O157:H7 strains, the fact that they were grouped into the same phylogenomic cluster and had the same unique metabolic pathway genes suggested that they may also share closely related evolutionary pathways. As a consequence, gene exchange between them is more likely to occur. Results from this study could potentially serve as a basis to help develop strategies to reduce the prevalence of pathogenic E. coli O157:H7 in livestock and downstream food production environments.

scholarly journals Identification of Escherichia coli and Shigella Species from Whole-Genome Sequences

2016 ◽  
Vol 55 (2) ◽  
pp. 616-623 ◽  
Author(s):  
Marie A. Chattaway ◽  
Ulf Schaefer ◽  
Rediat Tewolde ◽  
Timothy J. Dallman ◽  
Claire Jenkins
Keyword(s):  
Escherichia Coli ◽  
Clonal Complex ◽  
Shigella Flexneri ◽  
Bacterial Species ◽  
Whole Genome ◽  
Content Type ◽  
E Coli ◽  
Shigella Species ◽  
Close Phylogenetic Relationship ◽  
Initial Identification

ABSTRACTEscherichia coliandShigellaspecies are closely related and genetically constitute the same species. Differentiating between these two pathogens and accurately identifying the four species ofShigellaare therefore challenging. The organism-specific bioinformatics whole-genome sequencing (WGS) typing pipelines at Public Health England are dependent on the initial identification of the bacterial species by use of a kmer-based approach. Of the 1,982Escherichia coliandShigellasp. isolates analyzed in this study, 1,957 (98.4%) had concordant results by both traditional biochemistry and serology (TB&S) and the kmer identification (ID) derived from the WGS data. Of the 25 mismatches identified, 10 were enteroinvasiveE. coliisolates that were misidentified asShigella flexneriorS. boydiiby the kmer ID, and 8 wereS. flexneriisolates misidentified by TB&S asS. boydiidue to nonfunctionalS. flexneriO antigen biosynthesis genes. Analysis of the population structure based on multilocus sequence typing (MLST) data derived from the WGS data showed that the remaining discrepant results belonged to clonal complex 288 (CC288), comprising bothS. boydiiandS. dysenteriaestrains. Mismatches between the TB&S and kmer ID results were explained by the close phylogenetic relationship between the two species and were resolved with reference to the MLST data.Shigellacan be differentiated fromE. coliand accurately identified to the species level by use of kmer comparisons and MLST. Analysis of the WGS data provided explanations for the discordant results between TB&S and WGS data, revealed the true phylogenetic relationships between different species ofShigella, and identified emerging pathoadapted lineages.

scholarly journals Genomic epidemiology and evolution of Escherichia coli in wild animals

2020 ◽  
Author(s):  
Robert Murphy ◽  
Martin Palm ◽  
Ville Mustonen ◽  
Jonas Warringer ◽  
Anne Farewell ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Population Structure ◽  
Host Species ◽  
Bacterial Species ◽  
Clinical Importance ◽  
Wild Animal ◽  
Whole Genome ◽  
Wild Animals ◽  
E Coli ◽  
Wild Hosts

AbstractEscherichia coli is a common bacterial species in the gastrointestinal tracts of warm-blooded animals and humans. Pathogenic and antimicrobial resistance in E. coli may emerge via host switching from animal reservoirs. Despite its potential clinical importance, knowledge of the population structure of commensal E. coli within wild hosts and the epidemiological links between E. coli in non-human hosts and E. coli in humans is still scarce. In this study, we analysed the whole genome sequencing data of a collection of 119 commensal E. coli recovered from the guts of 68 mammal and bird species in Mexico and Venezuela in the 1990s. We observed low concordance between the population structures of E. coli colonizing wild animals and the phylogeny, taxonomy and ecological and physiological attributes of the host species, with distantly related E. coli often colonizing the same or similar host species and distantly related host species often hosting closely related E. coli. We found no evidence for recent transmission of E. coli genomes from wild animals to either domesticated animals or humans. However, multiple livestock- and human-related virulence factor genes were present in E. coli of wild animals, including virulence factors characteristic for Shiga toxin-producing E. coli (STEC) and atypical enteropathogenic E. coli (aEPEC), where several isolates from wild hosts harboured the locus of enterocyte effacement (LEE) pathogenicity island. Moreover, E. coli in wild animal hosts often harboured known antibiotic resistance determinants, including against ciprofloxacin, aminoglycosides, tetracyclines and beta-lactams, with some determinants present in multiple, distantly related E. coli lineages colonizing very different host animals. We conclude that although the genome pools of E. coli colonizing wild animal and human gut are well separated, they share virulence and antibiotic resistance genes and E. coli underscoring that wild animals could serve as reservoirs for E. coli pathogenicity in human and livestock infections.ImportanceEscherichia coli is a clinically importance bacterial species implicated in human and livestock associated infections worldwide. The bacterium is known to reside in the guts of humans, livestock and wild animals. Although wild animals are recognized to serve as potential reservoirs for pathogenic E. coli strains, the knowledge of the population structure of E. coli in wild hosts is still scarce. In this study we used the fine resolution of whole genome sequencing to provide novel insights into the evolution of E. coli genomes within a broad range of wild animal species (including mammals and birds), the co-evolution of E. coli strains with their hosts and the genetics of pathogenicity of E. coli strains in wild hosts. Our results provide evidence for the clinical importance of wild animals as reservoirs for pathogenic strains and necessitate the inclusion of non-human hosts in the surveillance programs for E. coli infections.

scholarly journals Whole-Genome Analysis of Antimicrobial-Resistant and Extraintestinal Pathogenic Escherichia coli in River Water

2016 ◽  
Vol 83 (5) ◽  
Author(s):  
Ryota Gomi ◽  
Tomonari Matsuda ◽  
Yasufumi Matsumura ◽  
Masaki Yamamoto ◽  
Michio Tanaka ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
River Water ◽  
Surface Waters ◽  
Pathogenic Bacteria ◽  
Resistant Bacteria ◽  
Whole Genome ◽  
Whole Genome Analysis ◽  
Content Type ◽  
E Coli ◽  
Resistance Determinants

ABSTRACT Contamination of surface waters by antimicrobial-resistant bacteria and pathogenic bacteria is a great concern. In this study, 531 Escherichia coli isolates obtained from the Yamato River in Japan were evaluated phenotypically for resistance to 25 antimicrobials. Seventy-six isolates (14.3%) were multidrug resistant (MDR), 66 (12.4%) were nonsusceptible to one or two classes of agents, and 389 (73.3%) were susceptible. We performed whole-genome sequencing of selected strains by using Illumina technology. In total, the genome sequences of 155 strains were analyzed for antibiotic resistance determinants and phylogenetic characteristics. More than 50 different resistance determinants, including acquired resistance genes and chromosomal resistance mutations, were detected. Among the sequenced MDR strains (n = 66), sequence type 155 (ST155) complex (n = 9), ST10 complex (n = 9), and ST69 complex (n = 7) were prevalent. Among extraintestinal pathogenic E. coli (ExPEC) strains (n = 58), clinically important clonal groups, namely, ST95 complex (n = 18), ST127 complex (n = 8), ST12 complex (n = 6), ST14 complex (n = 6), and ST131 complex (n = 6), were prevalent, demonstrating the clonal distribution of environmental ExPEC strains. Typing of the fimH (type 1 fimbrial adhesin) gene revealed that ST131 complex strains carried fimH22 or fimH41, and no strains belonging to the fimH30 subgroup were detected. Fine-scale phylogenetic analysis and virulence gene content analysis of strains belonging to the ST95 complex (one of the major clonal ExPEC groups causing community-onset infections) revealed no significant differences between environmental and clinical strains. The results indicate contamination of surface waters by E. coli strains belonging to clinically important clonal groups. IMPORTANCE The prevalence of antimicrobial-resistant and pathogenic E. coli strains in surface waters is a concern because surface waters are used as sources for drinking water, irrigation, and recreational purposes. In this study, MDR and ExPEC strains in river water were characterized by genomic sequencing and analysis. We detected more than 50 resistance determinants and identified clonal groups specific to MDR and ExPEC strains. This study showed contamination of surface waters by E. coli strains belonging to clinically important clonal groups. Overall, this study advances our understanding of environmental MDR and ExPEC strains.

scholarly journals Phylogenetic Grouping of Human Ocular Escherichia coli Based on Whole-Genome Sequence Analysis

2020 ◽  
Vol 8 (3) ◽  
pp. 422
Author(s):  
Konduri Ranjith ◽  
Chinthala Reddy SaiAbhilash ◽  
Gumpili Sai Prashanthi ◽  
Shalem Raj Padakandla ◽  
Savitri Sharma ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Whole Genome Sequence ◽  
Whole Genome ◽  
Phylogenetic Groups ◽  
Whole Genome Analysis ◽  
E Coli ◽  
Phylogenetic Affiliation ◽  
Antimicrobial Resistance Genes ◽  
Phylogenetic Grouping ◽  
High Concordance

Escherichia coli is a predominant bacterium in the intestinal tracts of animals. Phylogenetically, strains have been classified into seven phylogroups, A, B1, B2, C, D, E, and F. Pathogenic strains have been categorized into several pathotypes such as Enteropathogenic (EPEC), Enterotoxigenic (ETEC), Enteroinvasive (EIEC), Enteroaggregative (EAEC), Diffusely adherent (DAEC), Uropathogenic (UPEC), Shiga-toxin producing (STEC) or Enterohemorrhagic (EHEC) and Extra-intestinal pathogenic E. coli (ExPEC). E. coli also survives as a commensal on the ocular surface. However, under conditions of trauma and immune-compromised states, E. coli causes conjunctivitis, keratitis, endopthalmitis, dacyrocystitis, etc. The phylogenetic affiliation and the pathotype status of these ocular E. coli strains is not known. For this purpose, the whole-genome sequencing of the 10 ocular E. coli strains was accomplished. Based on whole-genome SNP variation, the ocular E. coli strains were assigned to phylogenetic groups A (two isolates), B2 (seven isolates), and C (one isolate). Furthermore, results indicated that ocular E. coli originated either from feces (enteropathogenic and enterotoxigenic), urine (uropathogenic), or from extra-intestinal sources (extra-intestinal pathogenic). A high concordance was observed between the presence of AMR (Antimicrobial Resistance) genes and antibiotic resistance in the ocular E. coli strains. Furthermore, several virulent genes (fimB to fimI, papB to papX, etc.) and prophages (Enterobacteria phage HK97, Enterobacteria phage P1, Escherichia phage D108 etc.) were unique to ocular E. coli. This is the first report on a whole-genome analysis of ocular E. coli strains.

scholarly journals First Report of the Globally Disseminated IncX4 Plasmid Carrying themcr-1Gene in a Colistin-Resistant Escherichia coli Sequence Type 101 Isolate from a Human Infection in Brazil

2016 ◽  
Vol 60 (10) ◽  
pp. 6415-6417 ◽  
Author(s):  
Miriam R. Fernandes ◽  
John A. McCulloch ◽  
Marco A. Vianello ◽  
Quézia Moura ◽  
Paula J. Pérez-Chaparro ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Genome Analysis ◽  
Human Infection ◽  
Sequence Type ◽  
Whole Genome ◽  
Whole Genome Analysis ◽  
Content Type ◽  
Diabetic Foot Infection ◽  
E Coli ◽  
Food Animal

ABSTRACTA colistin-resistantEscherichia colistrain was recovered from a patient with a diabetic foot infection in Brazil. Whole-genome analysis revealed that theE. coliisolate belonged to the widespread sequence type (ST) 101 and harbored themcr-1gene on an IncX4 plasmid that was highly similar tomcr-1-bearing IncX4 plasmids that were recently identified inEnterobacteriaceaefrom food, animal, and human samples recovered on different continents. These results suggest that self-transmissible IncX4-type plasmids may represent promiscuous plasmids contributing to the intercontinental spread of themcr-1gene.

scholarly journals A Spontaneous rapZ Mutant Impairs Infectivity of Lytic Bacteriophage vB_EcoM_JS09 against Enterotoxigenic Escherichia coli

mSphere ◽  
2021 ◽  
Vol 6 (2) ◽  
Author(s):  
Yan Zhou ◽  
Hongduo Bao ◽  
Hui Zhang ◽  
Maoda Pang ◽  
Shujiao Zhu ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Genome Analysis ◽  
Resistant Bacteria ◽  
Whole Genome ◽  
Lytic Bacteriophage ◽  
Whole Genome Analysis ◽  
Content Type ◽  
E Coli ◽  
Stop Mutation ◽  
Phage Receptor

ABSTRACT Our understanding of the mechanisms underlying phage-bacterium interactions remains limited. In Escherichia coli, RapZ regulates glucosamine-6-phosphate (GlcN6P) metabolism, the formation of which initiates synthesis of the bacterial cell envelope, including lipopolysaccharides (LPS). However, the role of RapZ, if any, on phage infectivity remains to be investigated. Here, we isolated strains of enterotoxigenic E. coli (ETEC) resistant to its specific lytic bacteriophage vB_EcoM_JS09 (JS09) in a phage aerosol spray experiment. Whole-genome analysis of phage-resistant bacteria revealed the rapZ gene acquired a premature stop mutation at amino acid 227. Here, we report that the mutation in the rapZ gene confers resistance by inhibiting 93.5% phage adsorption. Furthermore, this mutation changes the morphology of phage plaques, reduces efficiency of plating and phage propagation efficiency, and impairs the infectivity of phage JS09 against ETEC. Using scanning electron microscopy assays, we attribute the inability of the phage to adsorb to the loss of receptors in strains with defective RapZ. Analysis of the LPS profile shows that strains with defective RapZ inhibit phage infection by changing the LPS profile in E. coli. Preincubation of phage JS09 with LPS extracted from a wild-type (WT) strain blocked infection, suggesting LPS is the host receptor for phage JS09 adsorption. Our data uncover the mechanism by which ETEC resists infection of phage JS09 by mutating the rapZ gene and then increasing the expression of glmS and changing the phage receptor-LPS profile. These findings provide insight into the function of the rapZ gene for efficient infection of phage JS09. IMPORTANCE The development of phage-resistant bacteria is a challenging problem for phage therapy. However, our knowledge of phage resistance mechanisms is still limited. RapZ is an RNase adaptor protein encoded by the rapZ gene and plays an important function in Gram-positive and Gram-negative bacteria. Here, we report the whole-genome analysis of a phage-resistant enterotoxigenic Escherichia coli (ETEC) strain, which revealed that the rapZ gene acquired a premature stop mutation (E227Stop). We show that the premature stop mutation of rapZ impairs the infectivity of phage JS09 in ETEC. Furthermore, our findings indicate that ETEC becomes resistant against the adsorption and infection of phage JS09 by mutating the rapZ gene, increasing the expression of glmS, and changing the phage receptor-LPS profile. It is also first reported here that RapZ is essential for efficient infection of phage JS09.

scholarly journals Genomic Epidemiology and Evolution of Escherichia coli in Wild Animals in Mexico

mSphere ◽  
2021 ◽  
Vol 6 (1) ◽  
Author(s):  
Robert Murphy ◽  
Martin Palm ◽  
Ville Mustonen ◽  
Jonas Warringer ◽  
Anne Farewell ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Host Species ◽  
Bacterial Species ◽  
Clinical Importance ◽  
Wild Animal ◽  
Whole Genome ◽  
Wild Animals ◽  
Content Type ◽  
E Coli ◽  
Wild Hosts

ABSTRACT Escherichia coli is a common bacterial species in the gastrointestinal tracts of warm-blooded animals and humans. Pathogenicity and antimicrobial resistance in E. coli may emerge via host switching from animal reservoirs. Despite its potential clinical importance, knowledge of the population structure of commensal E. coli within wild hosts and the epidemiological links between E. coli in nonhuman hosts and E. coli in humans is still scarce. In this study, we analyzed the whole-genome sequencing data of a collection of 119 commensal E. coli strains recovered from the guts of 55 mammal and bird species in Mexico and Venezuela in the 1990s. We observed low concordance between the population structures of E. coli isolates colonizing wild animals and the phylogeny, taxonomy, and ecological and physiological attributes of the host species, with distantly related E. coli strains often colonizing the same or similar host species and distantly related host species often hosting closely related E. coli strains. We found no evidence for recent transmission of E. coli genomes from wild animals to either domesticated animals or humans. However, multiple livestock- and human-related virulence factor genes were present in E. coli of wild animals, including virulence factors characteristic of Shiga toxin-producing E. coli (STEC) and atypical enteropathogenic E. coli (aEPEC), where several isolates from wild hosts harbored the locus of enterocyte effacement (LEE) pathogenicity island. Moreover, E. coli isolates from wild animal hosts often harbored known antibiotic resistance determinants, including those against ciprofloxacin, aminoglycosides, tetracyclines, and beta-lactams, with some determinants present in multiple, distantly related E. coli lineages colonizing very different host animals. We conclude that genome pools of E. coli colonizing the guts of wild animals and humans share virulence and antibiotic resistance genes, underscoring the idea that wild animals could serve as reservoirs for E. coli pathogenicity in human and livestock infections. IMPORTANCE Escherichia coli is a clinically important bacterial species implicated in human- and livestock-associated infections worldwide. The bacterium is known to reside in the guts of humans, livestock, and wild animals. Although wild animals are recognized as potential reservoirs for pathogenic E. coli strains, the knowledge of the population structure of E. coli in wild hosts is still scarce. In this study, we used fine resolution of whole-genome sequencing to provide novel insights into the evolution of E. coli genomes from a small yet diverse collection of strains recovered within a broad range of wild animal species (including mammals and birds), the coevolution of E. coli strains with their hosts, and the genetics of pathogenicity of E. coli strains in wild hosts in Mexico. Our results provide evidence for the clinical importance of wild animals as reservoirs for pathogenic strains and highlight the need to include nonhuman hosts in the surveillance programs for E. coli infections.

scholarly journals Presence of β-Lactamase-producing Enterobacterales and Salmonella Isolates in Marine Mammals

2021 ◽  
Vol 22 (11) ◽  
pp. 5905
Author(s):  
Olivia M. Grünzweil ◽  
Lauren Palmer ◽  
Adriana Cabal ◽  
Michael P. Szostak ◽  
Werner Ruppitsch ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Marine Mammals ◽  
Virulence Genes ◽  
Multidrug Resistant ◽  
Healthcare Sector ◽  
Whole Genome ◽  
E Coli ◽  
High Prevalence ◽  
Lactamase Gene ◽  
Sequence Types

Marine mammals have been described as sentinels of the health of marine ecosystems. Therefore, the aim of this study was to investigate (i) the presence of extended-spectrum β-lactamase (ESBL)- and AmpC-producing Enterobacterales, which comprise several bacterial families important to the healthcare sector, as well as (ii) the presence of Salmonella in these coastal animals. The antimicrobial resistance pheno- and genotypes, as well as biocide susceptibility of Enterobacterales isolated from stranded marine mammals, were determined prior to their rehabilitation. All E. coli isolates (n = 27) were screened for virulence genes via DNA-based microarray, and twelve selected E. coli isolates were analyzed by whole-genome sequencing. Seventy-one percent of the Enterobacterales isolates exhibited a multidrug-resistant (MDR) pheno- and genotype. The gene blaCMY (n = 51) was the predominant β-lactamase gene. In addition, blaTEM-1 (n = 38), blaSHV-33 (n = 8), blaCTX-M-15 (n = 7), blaOXA-1 (n = 7), blaSHV-11 (n = 3), and blaDHA-1 (n = 2) were detected. The most prevalent non-β-lactamase genes were sul2 (n = 38), strA (n = 34), strB (n = 34), and tet(A) (n = 34). Escherichia coli isolates belonging to the pandemic sequence types (STs) ST38, ST167, and ST648 were identified. Among Salmonella isolates (n = 18), S. Havana was the most prevalent serotype. The present study revealed a high prevalence of MDR bacteria and the presence of pandemic high-risk clones, both of which are indicators of anthropogenic antimicrobial pollution, in marine mammals.

scholarly journals L-Alanine Prototrophic Suppressors Emerge from L-Alanine Auxotroph through Stress-Induced Mutagenesis in Escherichia coli

2021 ◽  
Vol 9 (3) ◽  
pp. 472
Author(s):  
Harutaka Mishima ◽  
Hirokazu Watanabe ◽  
Kei Uchigasaki ◽  
So Shimoda ◽  
Shota Seki ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Pyruvate Dehydrogenase ◽  
Pyruvate Dehydrogenase Complex ◽  
Spontaneous Mutation ◽  
Point Mutations ◽  
Whole Genome Analysis ◽  
E Coli ◽  
Clone Pair ◽  
Induced Mutagenesis ◽  
Isogenic Mutants

In Escherichia coli, L-alanine is synthesized by three isozymes: YfbQ, YfdZ, and AvtA. When an E. coli L-alanine auxotrophic isogenic mutant lacking the three isozymes was grown on L-alanine-deficient minimal agar medium, L-alanine prototrophic mutants emerged considerably more frequently than by spontaneous mutation; the emergence frequency increased over time, and, in an L-alanine-supplemented minimal medium, correlated inversely with L-alanine concentration, indicating that the mutants were derived through stress-induced mutagenesis. Whole-genome analysis of 40 independent L-alanine prototrophic mutants identified 16 and 18 clones harboring point mutation(s) in pyruvate dehydrogenase complex and phosphotransacetylase-acetate kinase pathway, which respectively produce acetyl coenzyme A and acetate from pyruvate. When two point mutations identified in L-alanine prototrophic mutants, in pta (D656A) and aceE (G147D), were individually introduced into the original L-alanine auxotroph, the isogenic mutants exhibited almost identical growth recovery as the respective cognate mutants. Each original- and isogenic-clone pair carrying the pta or aceE mutation showed extremely low phosphotransacetylase or pyruvate dehydrogenase activity, respectively. Lastly, extracellularly-added pyruvate, which dose-dependently supported L-alanine auxotroph growth, relieved the L-alanine starvation stress, preventing the emergence of L-alanine prototrophic mutants. Thus, L-alanine starvation-provoked stress-induced mutagenesis in the L-alanine auxotroph could lead to intracellular pyruvate increase, which eventually induces L-alanine prototrophy.

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