scholarly journals Phenotypic Characterization and Whole-Genome Analysis of a Novel Bacteriophage HCF1 Infecting Citrobacter amalonaticus and C. freundii

2021 ◽  
Vol 12 ◽  
Author(s):  
Prince Kumar ◽  
Mukesh K. Meghvansi ◽  
Dev V. Kamboj
Keyword(s):  
Environmental Stress ◽  
Host Range ◽  
Growth Curve ◽  
Phenotypic Characterization ◽  
Query Coverage ◽  
One Step ◽  
Step Growth ◽  
Tract Infections ◽  
Citrobacter Amalonaticus

Citrobacter species often occur in sewage, food, soil, wastewater, and in the intestinal tract of animals and humans. Citrobacter spp. cause urinary tract infections (UTIs) and infantile meningitis in humans. Due to the presence of plasmid-encoded resistance genes, Citrobacter spp. are often resistant to many antibiotics. In this study, Citrobacter virus HCF1, a novel virulent bacteriophage capable of killing Citrobacter amalonaticus and Citrobacter freundii, was isolated from the sewage water. The isolated bacteriophage was characterized with respect to transmission electron microscopy, one-step growth curve, host range, in vitro efficacy, storage stability, and environmental stress tolerance. The one-step growth curve analysis revealed that the latent period of HCF1 was 30 min and the estimated burst size was 121 plaque-forming units (PFU) per bacterial cell. Host range testing indicated that the HCF1 was specific to the Citrobacter genus. In vitro efficacy assay in the effluent of an anaerobic biodigester showed that the HCF1 completely eliminated the host within 4 and 5 h at MOI:100 and MOI:10, respectively, thereby indicating its potential for combating C. amalonaticus infections. The isolated bacteriophage is considerably stable and tolerant to environmental stress. Furthermore, the complete genome of HCF1 was sequenced using Oxford Nanopore sequencing and the data were subjected to detailed bioinformatic analyses. NCBI-BLASTn analysis revealed that the HCF1 genome had a query coverage of 15–21% and a maximum similarity of 77.27–78.49% with 11 bacteriophages of the Drexlerviridae family. Detailed bioinformatic analysis of the genome profile suggests that HCF1 is a novel T1svirus belonging to the Tempevirinae subfamily of the Drexlerviridae family.

scholarly journals Random Transposon-Mediated Mutagenesis of the Essential Large Tegument Protein pUL36 of Pseudorabies Virus

2010 ◽  
Vol 84 (16) ◽  
pp. 8153-8162 ◽  
Author(s):  
Britta S. Möhl ◽  
Sindy Böttcher ◽  
Harald Granzow ◽  
Walter Fuchs ◽  
Barbara G. Klupp ◽  
...  
Keyword(s):  
Amino Acids ◽  
Pseudorabies Virus ◽  
Plaque Assay ◽  
Tegument Protein ◽  
Nuclear Pore ◽  
Viral Spread ◽  
One Step ◽  
Step Growth ◽  
Insertion Mutants

ABSTRACT Homologs of the pseudorabies virus (PrV) essential large tegument protein pUL36 are conserved throughout the Herpesviridae. pUL36 functions during transport of the nucleocapsid to and docking at the nuclear pore as well as during virion formation after nuclear egress in the cytoplasm. Deletion analyses revealed several nonessential regions within the 3,084-amino-acid PrV pUL36 (S. Böttcher, B. G. Klupp, H. Granzow, W. Fuchs, K. Michael, and T. C. Mettenleiter, J. Virol. 80:9910-9915, 2006; S. Böttcher, H. Granzow, C. Maresch, B. Möhl, B. G. Klupp, and T. C. Mettenleiter, J. Virol. 81:13403-13411, 2007), while the C-terminal 62 amino acids are essential for virus replication (K. Coller, J. Lee, A. Ueda, and G. Smith, J. Virol. 81:11790-11797, 2007). To identify additional functional domains, we performed random mutagenesis of PrV pUL36 by transposon-mediated insertion of a 15-bp linker. By this approach, 26 pUL36 insertion mutants were selected and tested in transient transfection assays for their ability to complement one-step growth and/or viral spread of a PrV UL36 null mutant. Ten insertion mutants in the N-terminal half and 10 in the C terminus complemented both, whereas six insertion mutants clustering in the center of the protein did not complement in either assay. Interestingly, several insertions within conserved parts yielded positive complementation, including those located within the essential C-terminal 62 amino acids. For 15 mutants that mediated productive replication, stable virus recombinants were isolated and further characterized by plaque assay, in vitro growth analysis, and electron microscopy. Except for three mutant viruses, most insertion mutants replicated like wild-type PrV. Two insertion mutants, at amino acids (aa) 597 and 689, were impaired in one-step growth and viral spread and exhibited a defect in virion maturation in the cytoplasm. In contrast, one functional insertion (aa 1800) in a region which otherwise yielded only nonfunctional insertion mutants was impaired in viral spread but not in one-step growth without a distinctive ultrastructural phenotype. In summary, these studies extend and refine previous analyses of PrV pUL36 and demonstrate the different sensitivities of different regions of the protein to functional loss by insertion.

scholarly journals Characterization of RNA synthesis during a one-step growth curve and of the replication mechanism of bovine viral diarrhoea virus

1996 ◽  
Vol 77 (11) ◽  
pp. 2729-2736 ◽  
Author(s):  
Y. Gong ◽  
R. Trowbridge ◽  
T. B. Macnaughton ◽  
E. G. Westaway ◽  
A. D. Shannon ◽  
...  
Keyword(s):  
Growth Curve ◽  
Rna Synthesis ◽  
Bovine Viral Diarrhoea Virus ◽  
Bovine Viral Diarrhoea ◽  
Replication Mechanism ◽  
Diarrhoea Virus ◽  
One Step ◽  
Step Growth

scholarly journals Study of the One-Step Growth Curve of Equine Infectious Anemia Virus by Immunofluorescence

1972 ◽  
Vol 5 (6) ◽  
pp. 890-895 ◽  
Author(s):  
Chuzo Ushimi ◽  
James B. Henson ◽  
John R. Gorham
Keyword(s):  
Growth Curve ◽  
Equine Infectious Anemia Virus ◽  
Equine Infectious Anemia ◽  
One Step ◽  
Step Growth ◽  
The One ◽  
Anemia Virus

A BACTERIOPHAGE THAT ATTACKS NUMEROUS PHYTOPATHOGENIC XANTHOMONAS SPECIES

1958 ◽  
Vol 4 (5) ◽  
pp. 493-497 ◽  
Author(s):  
M. D. Sutton ◽  
H. Katznelson ◽  
C. Quadling
Keyword(s):  
Burst Size ◽  
Lytic Phage ◽  
Plant Materials ◽  
Single Burst ◽  
Xanthomonas Species ◽  
One Step ◽  
Average Burst Size ◽  
Step Growth ◽  
Rot Disease

This paper reports the isolation of a lytic phage that attacks in vitro numerous phytopathogenic Xanthomonas species, including X. campestris (Pammel) Dowson, the cause of black rot disease of crucifers. Although 'one-step' growth experiments suggested an average burst size of ca. four for this phage-host system, 'single burst' experiments indicated a burst size of ca. one hundred phage particles per bacterium. The particles have typical phage morphology, as determined by electron microscopy. This phage gave satisfactory results when used in the rapid plaque count test for detection of phage-sensitive bacteria in plant materials.

scholarly journals ONE-STEP GROWTH CURVE OF WESTERN EQUINE ENCEPHALOMYELITIS VIRUS ON CHICKEN EMBRYO CELLS GROWN IN VITRO AND ANALYSIS OF VIRUS YIELDS FROM SINGLE CELLS

1954 ◽  
Vol 99 (2) ◽  
pp. 183-199 ◽  
Author(s):  
R. Dulbecco ◽  
Marguerite Vogt
Keyword(s):  
Chicken Embryo ◽  
Cell Layer ◽  
Growth Curves ◽  
Virus Growth ◽  
Embryo Cells ◽  
Cell Layers ◽  
A Cell ◽  
One Step ◽  
Step Growth

The rate of adsorption of WEE virus onto chicken embryo cells in vitro was determined both on a cell layer and on a cell suspension. One-step growth curves were determined in cell suspensions and on cell layers. The latent period varied between 2 and 3½ hours; it was shorter on cell layers and decreased with higher multiplicity of infection. The shortest period is probably the real latent period. The growth curves of the virus showed an initial exponential rise and reached a maximal constant value after 6 to 8 hours. The maximum virus yield per cell varied between 200 and 1000 on the cell layer, and between 100 and 200 in suspended cells. The yield of single infected cells was determined. An analysis of the distributions of the individual yields obtained after various periods of virus growth led to two main conclusions: (1) that virus is released from the same cell over a long period of time; (2) that one phase of the intracellular virus growth is exponential.

VIRAL REPLICATION: ONE-STEP GROWTH CURVE

Virology ◽  
1992 ◽  
pp. 100-106
Author(s):  
FLORENCE G. BURLESON ◽  
THOMAS M. CHAMBERS ◽  
DANNY L. WIEDBRAUK
Keyword(s):  
Viral Replication ◽  
Growth Curve ◽  
One Step ◽  
Step Growth

scholarly journals Pathogenicity-Associated Islands in Extraintestinal Pathogenic Escherichia coli Are Fitness Elements Involved in Intestinal Colonization

2010 ◽  
Vol 192 (19) ◽  
pp. 4885-4893 ◽  
Author(s):  
Médéric Diard ◽  
Louis Garry ◽  
Marjorie Selva ◽  
Thomas Mosser ◽  
Erick Denamur ◽  
...  
Keyword(s):  
Genomic Islands ◽  
Phenotypic Characterization ◽  
Intestinal Colonization ◽  
Human Pathogens ◽  
Biologically Relevant ◽  
E Coli ◽  
Pathogenic Escherichia Coli ◽  
Tract Infections

ABSTRACT The virulence of many human pathogens does not seem to be an evolutionarily selected trait, but an accidental by-product of the selection that operates in another ecological context. We investigated the possibility that virulence of the ex traintestinal p athogenic E scherichia c oli (ExPEC) strains, which frequently cause disease in the host in which they asymptomatically colonize the intestine, is the consequence of commensalism. Most of the ExPEC virulence factors are clustered on genomic islands called p athogenicity- a ssociated i slands (PAIs). We constructed and characterized several mutants of the ExPEC 536 strain with either (i) deletions of each single PAI or (ii) a complete deletion of all seven PAIs. In vitro phenotypic characterization of 536 mutants showed that the seven PAIs were dispensable for growth in the absence of external stress, as well as under a range of biologically relevant stressors, i.e., serum, bile, and oxidative, nitrosative, hyperosmotic, and acidic stress. However, challenge against the wild-type (WT) strain in a murine model shows that the deletion of all seven PAIs drastically reduces the fitness of 536 during persistent intestinal colonization. This defect seems to be linked to the hypermotility observed for mutants devoid of all seven PAIs. In addition, we show that PAIs diminish fitness of their carrier during growth in urine, suggesting that urinary tract infections are unlikely to provide selective pressure for the maintenance of ExPEC PAIs. Our results are in accordance with the coincidental-evolution hypothesis postulating that extraintestinal E. coli virulence is a by-product of commensalism.

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