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UV-Sensitivity of Shiga Toxin-Converting Bacteriophage Virions Φ24B, 933W, P22, P27 and P32.

Bloch S, Nejman-Faleńczyk B, Topka G, Dydecka A, Licznerska K, Narajczyk M, Necel A, Węgrzyn A, Węgrzyn G - Toxins (Basel) (2015)

Bottom Line: Under these conditions, a considerable release of phage DNA from virions was observed, and electron microscopy analyses indicated a large proportion of partially damaged virions.Infection of E. coli cells with UV-irradiated Stx phages resulted in significantly decreased levels of expression of N and cro genes, crucial for lytic development.We conclude that inactivation of Stx virions caused by relatively low dose of UV light is due to damage of capsids that prevents effective infection of the host cells.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular Biology, University of Gdańsk, Wita Stwosza 59, Gdańsk 80-308, Poland. sylwia.bloch@biol.ug.edu.pl.

ABSTRACT
Shiga toxin-converting bacteriophages (Stx phages) are present as prophages in Shiga toxin-producing Escherichia coli (STEC) strains. Theses phages can be transmitted to previously non-pathogenic E. coli cells making them potential producers of Shiga toxins, as they bear genes for these toxins in their genomes. Therefore, sensitivity of Stx phage virions to various conditions is important in both natural processes of spreading of these viruses and potential prophylactic control of appearance of novel pathogenic E. coli strains. In this report we provide evidence that virions of Stx phages are significantly more sensitive to UV irradiation than bacteriophage λ. Following UV irradiation of Stx virions at the dose of 50 J/m², their infectivity dropped by 1-3 log10, depending on the kind of phage. Under these conditions, a considerable release of phage DNA from virions was observed, and electron microscopy analyses indicated a large proportion of partially damaged virions. Infection of E. coli cells with UV-irradiated Stx phages resulted in significantly decreased levels of expression of N and cro genes, crucial for lytic development. We conclude that inactivation of Stx virions caused by relatively low dose of UV light is due to damage of capsids that prevents effective infection of the host cells.

No MeSH data available.


Related in: MedlinePlus

Expression of N and cro genes of bacteriophages λ (left panel) and Stx phage Φ24B (right panel) which were either non-irradiated (UV−) or irradiated with UV light at 50 J/m2 (UV+) prior to infection (at m.o.i. = 1). The mRNA levels were estimated in E. coli MG1655 host grown at 37 °C, at 7.5 min or 15 min after infection by λ or Φ24B, respectively. The presented results are mean values from three experiments with error bars indicating SD. Statistically significant differences between results obtained for non-irradiated and UV-irradiated phages (p < 0.001 in the t-test) are indicated by three asterisks.
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toxins-07-03727-f004: Expression of N and cro genes of bacteriophages λ (left panel) and Stx phage Φ24B (right panel) which were either non-irradiated (UV−) or irradiated with UV light at 50 J/m2 (UV+) prior to infection (at m.o.i. = 1). The mRNA levels were estimated in E. coli MG1655 host grown at 37 °C, at 7.5 min or 15 min after infection by λ or Φ24B, respectively. The presented results are mean values from three experiments with error bars indicating SD. Statistically significant differences between results obtained for non-irradiated and UV-irradiated phages (p < 0.001 in the t-test) are indicated by three asterisks.

Mentions: If our conclusion that decreased infectivity of UV-irradiated Stx phages, relative to untreated ones, results from a damage of capsids in a significant proportion of virions is true, one should expect that lower number of phages can introduce their DNA into host cells, and thus, level of expression of phage genes in a population of infected cells should be considerably lower than in E. coli infected with equal number of untreated phages. To test this hypothesis, a sample of the lysate of either λ or Φ24B virions was divided to two halves of equal titer (determined as pfu/mL). One half was UV irradiated at 50 J/m2 while the second was not, and then E. coli cells were infected with either treated or untreated phages. Following infection, total RNA was isolated, and expression of N and cro genes (transcribed from the pL and pR promoter, respectively), crucial for the lytic development [11,12,13,14], was estimated by reverse transcription real-time quantitative PCR (RT-qPCR). Analysis of phage gene expression (RNA level) instead of their copy numbers (DNA level) allowed to be devoid of signal becoming from a fraction of DNA released from capsids in response to UV exposure. In bacteriophage λ, UV irradiation of virions prior to infection did not result in any significant changes in expression of N and cro genes in the population of E. coli cells (Figure 4). On the contrary, in the E. coli population infected with irradiated Φ24B bacteriophage, the level of expression of both genes was significantly lower relative to bacteria infected with non-irradiated viruses (Figure 4).


UV-Sensitivity of Shiga Toxin-Converting Bacteriophage Virions Φ24B, 933W, P22, P27 and P32.

Bloch S, Nejman-Faleńczyk B, Topka G, Dydecka A, Licznerska K, Narajczyk M, Necel A, Węgrzyn A, Węgrzyn G - Toxins (Basel) (2015)

Expression of N and cro genes of bacteriophages λ (left panel) and Stx phage Φ24B (right panel) which were either non-irradiated (UV−) or irradiated with UV light at 50 J/m2 (UV+) prior to infection (at m.o.i. = 1). The mRNA levels were estimated in E. coli MG1655 host grown at 37 °C, at 7.5 min or 15 min after infection by λ or Φ24B, respectively. The presented results are mean values from three experiments with error bars indicating SD. Statistically significant differences between results obtained for non-irradiated and UV-irradiated phages (p < 0.001 in the t-test) are indicated by three asterisks.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4591643&req=5

toxins-07-03727-f004: Expression of N and cro genes of bacteriophages λ (left panel) and Stx phage Φ24B (right panel) which were either non-irradiated (UV−) or irradiated with UV light at 50 J/m2 (UV+) prior to infection (at m.o.i. = 1). The mRNA levels were estimated in E. coli MG1655 host grown at 37 °C, at 7.5 min or 15 min after infection by λ or Φ24B, respectively. The presented results are mean values from three experiments with error bars indicating SD. Statistically significant differences between results obtained for non-irradiated and UV-irradiated phages (p < 0.001 in the t-test) are indicated by three asterisks.
Mentions: If our conclusion that decreased infectivity of UV-irradiated Stx phages, relative to untreated ones, results from a damage of capsids in a significant proportion of virions is true, one should expect that lower number of phages can introduce their DNA into host cells, and thus, level of expression of phage genes in a population of infected cells should be considerably lower than in E. coli infected with equal number of untreated phages. To test this hypothesis, a sample of the lysate of either λ or Φ24B virions was divided to two halves of equal titer (determined as pfu/mL). One half was UV irradiated at 50 J/m2 while the second was not, and then E. coli cells were infected with either treated or untreated phages. Following infection, total RNA was isolated, and expression of N and cro genes (transcribed from the pL and pR promoter, respectively), crucial for the lytic development [11,12,13,14], was estimated by reverse transcription real-time quantitative PCR (RT-qPCR). Analysis of phage gene expression (RNA level) instead of their copy numbers (DNA level) allowed to be devoid of signal becoming from a fraction of DNA released from capsids in response to UV exposure. In bacteriophage λ, UV irradiation of virions prior to infection did not result in any significant changes in expression of N and cro genes in the population of E. coli cells (Figure 4). On the contrary, in the E. coli population infected with irradiated Φ24B bacteriophage, the level of expression of both genes was significantly lower relative to bacteria infected with non-irradiated viruses (Figure 4).

Bottom Line: Under these conditions, a considerable release of phage DNA from virions was observed, and electron microscopy analyses indicated a large proportion of partially damaged virions.Infection of E. coli cells with UV-irradiated Stx phages resulted in significantly decreased levels of expression of N and cro genes, crucial for lytic development.We conclude that inactivation of Stx virions caused by relatively low dose of UV light is due to damage of capsids that prevents effective infection of the host cells.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular Biology, University of Gdańsk, Wita Stwosza 59, Gdańsk 80-308, Poland. sylwia.bloch@biol.ug.edu.pl.

ABSTRACT
Shiga toxin-converting bacteriophages (Stx phages) are present as prophages in Shiga toxin-producing Escherichia coli (STEC) strains. Theses phages can be transmitted to previously non-pathogenic E. coli cells making them potential producers of Shiga toxins, as they bear genes for these toxins in their genomes. Therefore, sensitivity of Stx phage virions to various conditions is important in both natural processes of spreading of these viruses and potential prophylactic control of appearance of novel pathogenic E. coli strains. In this report we provide evidence that virions of Stx phages are significantly more sensitive to UV irradiation than bacteriophage λ. Following UV irradiation of Stx virions at the dose of 50 J/m², their infectivity dropped by 1-3 log10, depending on the kind of phage. Under these conditions, a considerable release of phage DNA from virions was observed, and electron microscopy analyses indicated a large proportion of partially damaged virions. Infection of E. coli cells with UV-irradiated Stx phages resulted in significantly decreased levels of expression of N and cro genes, crucial for lytic development. We conclude that inactivation of Stx virions caused by relatively low dose of UV light is due to damage of capsids that prevents effective infection of the host cells.

No MeSH data available.


Related in: MedlinePlus