Molecular Characterization and Genomic Analysis of Novel Phage vB_ ShiP-A7 Infecting Multidrug-Resistant Shiglla Fexneri and Escherichia Coli

2020 ◽  
Author(s):  
Jing Xu ◽  
Yu Gu ◽  
Xinyan Yu ◽  
Ruiyang Zhang ◽  
Xuesen Zhang ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Case Reports ◽  
Burst Size ◽  
Shigella Flexneri ◽  
Genomic Analysis ◽  
Multidrug Resistant ◽  
Open Reading Frames ◽  
Mass Spectrometry Analysis ◽  
Spectrometry Analysis ◽  
Spherical Head

Abstract BackgroundPhage therapy has regained more attention due to the rise of multidrug-resistant (MDR) bacteria. Several case reports demonstrated clinical application of phage in resolving infections caused by MDR bacteria in recent years. ResultsWe isolated a new phage, vB_ShiP-A7, and then investigated its characteristics. Phage vB_ShiP-A7 is a member of Podoviridae that has an icosahedral spherical head and a short tail. vB_ShiP-A7 has large burst size and short replication time. vB_ShiP-A7’s genome is linear double stranded DNA composed of 40058 bp, encoding forty-three putative open reading frames. Comparative genome analysis demonstrated vB_ShiP-A7’s genome sequence is closely related to fifteen different phages (coverage 74-88%, identity 86-93%). Mass Spectrometry analysis revealed that twelve known proteins and six hypothetical proteins exist in particles of vB_ShiP-A7. Genome and proteome analyses confirmed the absence of lysogen-related proteins and toxic proteins in this phage. In addition, phage vB_ShiP-A7 can significantly reduce the growth of clinical MDR stains of Shigella flexneri and Escherichia coli in liquid culture. Furthermore, vB_ShiP-A7 can disrupt biofilms formed by Shigella flexneri or Escherichia coli in vitro. ConclusionPhage vB_ShiP-A7 is a stable novel phage, which has a strong application potential to inhibit MDR stains of Shigella flexneri and Escherichia coli. Comparing the genomes between vB_ShiP-A7 and other closely-related phages will help us better understand the evolutionary mechanism of phages.

scholarly journals Molecular Characteristics of Novel Phage vB_ShiP-A7 Infecting Multidrug-Resistant Shigella flexneri and Escherichia coli, and Its Bactericidal Effect in vitro and in vivo

2021 ◽  
Vol 12 ◽  
Author(s):  
Jing Xu ◽  
Ruiyang Zhang ◽  
Xinyan Yu ◽  
Xuesen Zhang ◽  
Genyan Liu ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Burst Size ◽  
Shigella Flexneri ◽  
Latency Period ◽  
Multidrug Resistant ◽  
Mass Spectrometry Analysis ◽  
Comparative Genomic ◽  
Molecular Characteristics ◽  
Infection Model

In recent years, increasing evidence has shown that bacteriophages (phages) can inhibit infection caused by multidrug-resistant (MDR) bacteria. Here, we isolated a new phage, named vB_ShiP-A7, using MDR Shigella flexneri as the host. vB_ShiP-A7 is a novel member of Podoviridae, with a latency period of approximately 35 min and a burst size of approximately 100 phage particles/cell. The adsorption rate constant of phage vB_ShiP-A7 to its host S. flexneri was 1.405 × 10–8 mL/min. The vB_ShiP-A7 genome is a linear double-stranded DNA composed of 40,058 bp with 177 bp terminal repeats, encoding 43 putative open reading frames. Comparative genomic analysis demonstrated that the genome sequence of vB_ShiP-A7 is closely related to 15 different phages, which can infect different strains. Mass spectrometry analysis revealed that 12 known proteins and 6 hypothetical proteins exist in the particles of phage vB_ShiP-A7. Our results confirmed that the genome of vB_ShiP-A7 is free of lysogen-related genes, bacterial virulence genes, and antibiotic resistance genes. vB_ShiP-A7 can significantly disrupt the growth of some MDR clinical strains of S. flexneri and Escherichia coli in liquid culture and biofilms in vitro. In addition, vB_ShiP-A7 can reduce the load of S. flexneri by approximately 3–10 folds in an infection model of mice. Therefore, vB_ShiP-A7 is a stable novel phage with the potential to treat infections caused by MDR strains of S. flexneri and E. coli.

scholarly journals T4-Like Phage Bp7, a Potential Antimicrobial Agent for Controlling Drug-Resistant Escherichia coli in Chickens

2013 ◽  
Vol 79 (18) ◽  
pp. 5559-5565 ◽  
Author(s):  
Can Zhang ◽  
Wenli Li ◽  
Wenhua Liu ◽  
Ling Zou ◽  
Chen Yan ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Burst Size ◽  
Multidrug Resistant ◽  
Open Reading Frames ◽  
Attractive Alternative ◽  
Drug Resistant ◽  
Animal Cells ◽  
Content Type ◽  
E Coli ◽  
Chicken Feces

ABSTRACTChicken-pathogenicEscherichia coliis severely endangering the poultry industry in China and worldwide, and antibiotic therapy is facing an increasing problem of antibiotic resistance. Bacteriophages can kill bacteria with no known activity in human or animal cells, making them an attractive alternative to antibiotics. In this study, we present the characteristics of a novel virulent bacteriophage, Bp7, specifically infecting pathogenic multidrug-resistantE. coli. Phage Bp7 was isolated from chicken feces. Bp7 belongs to the familyMyoviridae, possessing an elongated icosahedral head and contractile sheathed tail. It has a 168-kb double-stranded DNA genome. For larger yields, its optimal multiplicity of infection (MOI) to infectE. coliwas about 0.001. The latent period was 10 to 15 min, and the burst size was 90 PFU/infected cell. It was stable both at pH 5.0 to 10.0 and at 40°C or 50°C for at least 1 h. Bp7 could infect 46% of pathogenic clinicalE. colistrains. Bp7 harbored 791 open reading frames (ORFs) and 263 possible genes. Among the 263 genes, 199 possessed amino acid sequence identities with ORFs of phage T4, 62 had identities with other T4-like phages, and only one lacked any database match. The genome of Bp7 manifested obvious division and rearrangement compared to phages T4, JS98, and IME08. Bp7 is a new member of the “T4-like” genus, familyMyoviridae. Its wide host range, strong cell-killing activity, and high stability to pH make it an alternative to antimicrobials for controlling drug-resistantE. coliin chickens.

scholarly journals Morphological, biological, and genomic characterization of a newly isolated lytic phage Sfk20 infecting Shigella flexneri, Shigella sonnei, and Shigella dysenteriae1

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Bani Mallick ◽  
Payel Mondal ◽  
Moumita Dutta
Keyword(s):  
Burst Size ◽  
Shigella Flexneri ◽  
Alternative Therapy ◽  
Genomic Analysis ◽  
Multidrug Resistant ◽  
Lytic Activity ◽  
Lytic Phage ◽  
Tem Analysis ◽  
Host Interaction ◽  
Outbreak Area

AbstractShigellosis, caused by Shigella bacterial spp., is one of the leading causes of diarrheal morbidity and mortality. An increasing prevalence of multidrug-resistant Shigella species has revived the importance of bacteriophages as an alternative therapy to antibiotics. In this study, a novel bacteriophage, Sfk20, has been isolated from water bodies of a diarrheal outbreak area in Kolkata (India) with lytic activity against many Shigella spp. Phage Sfk20 showed a latent period of 20 min and a large burst size of 123 pfu per infected cell in a one-step growth analysis. Phage-host interaction and lytic activity confirmed by phage attachment, intracellular phage development, and bacterial cell burst using ultrathin sectioning and TEM analysis. The genomic analysis revealed that the double-stranded DNA genome of Sfk20 contains 164,878 bp with 35.62% G + C content and 241 ORFs. Results suggested phage Sfk20 to include as a member of the T4 myoviridae bacteriophage group. Phage Sfk20 has shown anti-biofilm potential against Shigella species. The results of this study imply that Sfk20 has good possibilities to be used as a biocontrol agent.

scholarly journals A Novel Polyvalent Bacteriophage vB_EcoM_swi3 Infects Pathogenic Escherichia coli and Salmonella enteritidis

2021 ◽  
Vol 12 ◽  
Author(s):  
Bingrui Sui ◽  
Lili Han ◽  
Huiying Ren ◽  
Wenhua Liu ◽  
Can Zhang
Keyword(s):  
Escherichia Coli ◽  
Salmonella Enteritidis ◽  
Burst Size ◽  
Large Subunit ◽  
Gc Content ◽  
Multidrug Resistant ◽  
Open Reading Frames ◽  
Morphological Observation ◽  
E Coli

A novel virulent bacteriophage vB_EcoM_swi3 (swi3), isolated from swine feces, lyzed 9% (6/65) of Escherichia coli and isolates 54% (39/72) of Salmonella enteritidis isolates, which were all clinically pathogenic multidrug-resistant strains. Morphological observation showed that phage swi3 belonged to the Myoviridae family with an icosahedral head (80 nm in diameter) and a contractile sheathed tail (120 nm in length). At the optimal multiplicity of infection of 1, the one-step growth analysis of swi3 showed a 25-min latent period with a burst size of 25-plaque-forming units (PFU)/infected cell. Phage swi3 remained stable both at pH 6.0–8.0 and at less than 50°C for at least 1 h. Genomic sequencing and bioinformatics analysis based on genomic sequences and the terminase large subunit showed that phage swi3 was a novel member that was most closely related to Salmonella phages and belonged to the Rosemountvirus genus. Phage swi3 harbored a 52-kb double-stranded DNA genome with 46.02% GC content. Seventy-two potential open reading frames were identified and annotated, only 15 of which had been assigned to functional genes. No gene associated with pathogenicity and virulence was identified. The effects of phage swi3 in treating pathologic E. coli infections in vivo were evaluated using a mouse model. The administration of a single intraperitoneal injection of swi3 (106 PFU) at 2 h after challenge with the E. coli strain (serotype K88) (108 colony-forming units) sufficiently protected all mice without toxic side effects. This finding highlighted that phage swi3 might be used as an effective antibacterial agent to prevent E. coli and S. enteritidis infection.

scholarly journals Heterogeneous and Flexible Transmission ofmcr-1in Hospital-AssociatedEscherichia coli

mBio ◽  
2018 ◽  
Vol 9 (4) ◽  
Author(s):  
Yingbo Shen ◽  
Zuowei Wu ◽  
Yang Wang ◽  
Rong Zhang ◽  
Hong-Wei Zhou ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Genomic Analysis ◽  
Multidrug Resistant ◽  
Fluoroquinolone Resistance ◽  
Gram Negative Bacteria ◽  
Colistin Resistance ◽  
Gram Negative ◽  
Content Type ◽  
E Coli ◽  
Genes Encoding

ABSTRACTThe recent emergence of a transferable colistin resistance mechanism, MCR-1, has gained global attention because of its threat to clinical treatment of infections caused by multidrug-resistant Gram-negative bacteria. However, the possible transmission route ofmcr-1amongEnterobacteriaceaespecies in clinical settings is largely unknown. Here, we present a comprehensive genomic analysis ofEscherichia coliisolates collected in a hospital in Hangzhou, China. We found thatmcr-1-carrying isolates from clinical infections and feces of inpatients and healthy volunteers were genetically diverse and were not closely related phylogenetically, suggesting that clonal expansion is not involved in the spread ofmcr-1. Themcr-1gene was found on either chromosomes or plasmids, but in most of theE. coliisolates,mcr-1was carried on plasmids. The genetic context of the plasmids showed considerable diversity as evidenced by the different functional insertion sequence (IS) elements, toxin-antitoxin (TA) systems, heavy metal resistance determinants, and Rep proteins of broad-host-range plasmids. Additionally, the genomic analysis revealed nosocomial transmission ofmcr-1and the coexistence ofmcr-1with other genes encoding β-lactamases and fluoroquinolone resistance in theE. coliisolates. These findings indicate thatmcr-1is heterogeneously disseminated in both commensal and pathogenic strains ofE. coli, suggest the high flexibility of this gene in its association with diverse genetic backgrounds of the hosts, and provide new insights into the genome epidemiology ofmcr-1among hospital-associatedE. colistrains.IMPORTANCEColistin represents one of the very few available drugs for treating infections caused by extensively multidrug-resistant Gram-negative bacteria. The recently emergentmcr-1colistin resistance gene threatens the clinical utility of colistin and has gained global attention. Howmcr-1spreads in hospital settings remains unknown and was investigated by whole-genome sequencing ofmcr-1-carryingEscherichia coliin this study. The findings revealed extraordinary flexibility ofmcr-1in its spread among genetically diverseE. colihosts and plasmids, nosocomial transmission ofmcr-1-carryingE. coli, and the continuous emergence of novel Inc types of plasmids carryingmcr-1and newmcr-1variants. Additionally,mcr-1was found to be frequently associated with other genes encoding β-lactams and fluoroquinolone resistance. These findings provide important information on the transmission and epidemiology ofmcr-1and are of significant public health importance as the information is expected to facilitate the control of this significant antibiotic resistance threat.

scholarly journals Dissection of the Role of Pili and Type 2 and 3 Secretion Systems in Adherence and Biofilm Formation of an Atypical Enteropathogenic Escherichia coli Strain

2013 ◽  
Vol 81 (10) ◽  
pp. 3793-3802 ◽  
Author(s):  
Rodrigo T. Hernandes ◽  
Miguel A. De la Cruz ◽  
Denise Yamamoto ◽  
Jorge A. Girón ◽  
Tânia A. T. Gomes
Keyword(s):  
Escherichia Coli ◽  
Biofilm Formation ◽  
Coli Strain ◽  
Mass Spectrometry Analysis ◽  
Secretion Systems ◽  
Content Type ◽  
Spectrometry Analysis ◽  
Rigid Rod

ABSTRACTAtypical enteropathogenicEscherichia coli(aEPEC) strains are diarrheal pathogens that lack bundle-forming pilus production but possess the virulence-associated locus of enterocyte effacement. aEPEC strain 1551-2 produces localized adherence (LA) on HeLa cells; however, its isogenic intimin (eae) mutant produces a diffuse-adherence (DA) pattern. In this study, we aimed to identify the DA-associated adhesin of the 1551-2eaemutant. Electron microscopy of 1551-2 identified rigid rod-like pili composed of an 18-kDa protein, which was identified as the major pilin subunit of type 1 pilus (T1P) by mass spectrometry analysis. Deletion offimAin 1551-2 affected biofilm formation but had no effect on adherence properties. Analysis of secreted proteins in supernatants of this strain identified a 150-kDa protein corresponding to SslE, a type 2 secreted protein that was recently reported to be involved in biofilm formation of rabbit and human EPEC strains. However, neither adherence nor biofilm formation was affected in a 1551-2sslEmutant. We then investigated the role of the EspA filament associated with the type 3 secretion system (T3SS) in DA by generating a doubleeae espAmutant. This strain was no longer adherent, strongly suggesting that the T3SS translocon is the DA adhesin. In agreement with these results, specific anti-EspA antibodies blocked adherence of the 1551-2eaemutant. Our data support a role for intimin in LA, for the T3SS translocon in DA, and for T1P in biofilm formation, all of which may act in concert to facilitate host intestinal colonization by aEPEC strains.

scholarly journals Specific Roles of the iroBCDEN Genes in Virulence of an Avian Pathogenic Escherichia coli O78 Strain and in Production of Salmochelins

2008 ◽  
Vol 76 (8) ◽  
pp. 3539-3549 ◽  
Author(s):  
Mélissa Caza ◽  
François Lépine ◽  
Sylvain Milot ◽  
Charles M. Dozois
Keyword(s):  
Escherichia Coli ◽  
Respiratory Infections ◽  
Mass Spectrometry Analysis ◽  
Liquid Chromatography Mass Spectrometry ◽  
Avian Pathogenic Escherichia Coli ◽  
Prevalence Studies ◽  
Spectrometry Analysis ◽  
E Coli ◽  
Pathogenic Escherichia Coli ◽  
Negative Mutant

ABSTRACT Avian pathogenic Escherichia coli (APEC) strains are a subset of extraintestinal pathogenic E. coli (ExPEC) strains associated with respiratory infections and septicemia in poultry. The iroBCDEN genes encode the salmochelin siderophore system present in Salmonella enterica and some ExPEC strains. Roles of the iro genes for virulence in chickens and production of salmochelins were assessed by introducing plasmids carrying different combinations of iro genes into an attenuated salmochelin- and aerobactin-negative mutant of O78 strain χ7122. Complementation with the iroBCDEN genes resulted in a regaining of virulence, whereas the absence of iroC, iroDE, or iroN abrogated restoration of virulence. The iroE gene was not required for virulence, since introduction of iroBCDN restored the capacity to cause lesions and colonize extraintestinal tissues. Prevalence studies indicated that iro sequences were associated with virulent APEC strains. Liquid chromatography-mass spectrometry analysis of supernatants of APEC χ7122 and the complemented mutants indicated that (i) for χ7122, salmochelins comprised 14 to 27% of the siderophores present in iron-limited medium or infected tissues; (ii) complementation of the mutant with the iro locus increased levels of glucosylated dimers (S1 and S5) and monomer (SX) compared to APEC strain χ7122; (iii) the iroDE genes were important for generation of S1, S5, and SX; (iv) iroC was required for export of salmochelin trimers and dimers; and (v) iroB was required for generation of salmochelins. Overall, efficient glucosylation (IroB), transport (IroC and IroN), and processing (IroD and IroE) of salmochelins are required for APEC virulence, although IroE appears to serve an ancillary role.

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