Goodbye PAM: Phage λ’s Red recombination system cripples PAMs and helps dodge CRISPR attacks

Ho, Khang; Harshey, Rasika M.

doi:10.1016/j.chom.2021.09.011

Cell Host & Microbe

2021

DOI: 10.1016/j.chom.2021.09.011

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Goodbye PAM: Phage λ’s Red recombination system cripples PAMs and helps dodge CRISPR attacks

Khang Ho

Rasika M. Harshey

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2022

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“…Generation of the steE-deficient S. Pullorum and its complementation. The steE deletion mutant of S. Pullorum was constructed by λ-Red recombination system as previously described (Ho et al, 2021). Briefly, the kanamycin resistance cassette (KanR) was amplified from pKD4 plasmid using the primers P1/P2.…”

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Construction and Characterization of Stee Deletion Mutant of Salmonella Pullorum

Liu¹,

Фотіна²,

Петров³

et al. 2022

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Salmonella Pullorum (S. Pullorum) is one of the host-restricted serotypes causing systemic infection in poultry. After S. Pullorum infection, chicks and turkeys usually have acute systemic infection. The main clinical symptoms are white dysentery and dyspnea, and the mortality can be as high as 100%. In adult chickens, local and chronic infections are the most common without obvious clinical symptoms, and can be transmitted vertically to offspring through ovary. Although the use of antibiotics reduces the death of sick chickens, it can not completely eliminate the pathogenic microorganisms in hosts, and is prone to public health problems such as drug resistance and drug residues. No study has ever reported the role of steE in HD-11 cells infected by S. Pullorum. The growth and biochemical characteristics of S. Pullorum ΔsteE were similar to that of S. Pullorum. Furthermore, we also observed the effects of steE on cell proliferation and apoptosis in S. Pulloruminfected HD-11 cells.In order to define the pathogenicity of steE gene of S. Pullorum, the steE deletion strain of S. Pullorum and its complemented strain were successfully constructed, and then its characterization were analyzed. S. Pullorum was preserved by the microbiology laboratory of the college of animal science and veterinary medicine, Henan Institute of Science and Technology. The pKD4, pKD46 and pCP20 or pBR322 plasmids were used for the λ-Red recombination system or complementary strain. The biological characteristics of S. Pullorum ΔsteE were consistent with those of its parent strain S. Pullorum and complementary strain S. Pullorum ΔsteE (pBR322-steE). Construction and confirmation of the ΔsteE strain. To identify the roles of steE in S. Pullorum, the steE deletion mutant of S. Pullorum was correctly constructed.The virulence test showed S. Pullorum ΔsteE decreased the proliferation and apoptosis of HD-11 cells compared to that of S. Pullorum and S. Pullorum ΔsteE (pBR322-steE). Taken together, our data demonstrate that the deletion of steE in S. Pullorum had no effect the growth and biochemical characteristics, but its proliferation ability decreased significantly in HD-11 cells, which decreased cell apoptosis, indicating that steE was closely related to virulence of S. Pullorum. Altogether, our research suggest that the steE gene was required for S. Pullorum virulence, which laid a foundation for further related research in S. Pullorum vaccine strains.

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confidence: 99%

Construction and Characterization of Stee Deletion Mutant of Salmonella Pullorum

Liu¹,

Фотіна²,

Петров³

et al. 2022

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Goodbye PAM: Phage λ’s Red recombination system cripples PAMs and helps dodge CRISPR attacks

Cited by 1 publication

References 10 publications

Construction and Characterization of Stee Deletion Mutant of Salmonella Pullorum

Construction and Characterization of Stee Deletion Mutant of Salmonella Pullorum

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