Recoding anaerobic regulator fnr of Salmonella Typhimurium attenuates it's pathogenicity

C, Nikhil K.; Noatia, Laxmi; Priyadarshini, Swagatika; Pashupathi, M.; Gali, Jagan Mohanarao; Ali, M. Ayub; Behera, Sujit Kumar; Sharma, Bhaskar; Roychoudhury, P.; Kumar, Ajay; Behera, Parthasarathi

doi:10.1016/j.micpath.2022.105591

Cited by 5 publications

(4 citation statements)

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“…Referring to previous methods [ 27 , 28 ], 5 µL of overnight-cultured bacterial suspension was inoculated onto LB semi-solid agar plates (0.3% agar). The plates were then incubated at 37 °C for 5–6 h. The diameter of the swimming halo was measured, and the assay was repeated 3 times.…”

Section: Methodsmentioning

confidence: 99%

“…For the intracellular survival assay, as previously described [ 25 , 28 , 29 ], RAW264.7 cells were seeded at a density of 10 5 cells per well in 2–12 well plates. C50336, Δ cheV and Δ cheV + cheV infected the cells with an MOI of 100 and were incubated in a 37 °C, 5% CO 2 incubator for 2 h. The cells were then washed twice with PBS to remove non-adherent/invasive bacteria.…”

Section: Methodsmentioning

confidence: 99%

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CheV enhances the virulence of Salmonella Enteritidis, and the Chev-deleted Salmonella vaccine provides immunity in mice

Zhang,

Wu,

Wang

et al. 2024

BMC Vet Res

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Background Salmonella enteritidis (SE) is a major zoonotic pathogen and causes infections in a variety of hosts. The development of novel vaccines for SE is necessary to eradicate this pathogen. Genetically engineered attenuated live vaccines are more immunogenic and safer. Thus, to develop a live attenuated Salmonella vaccine, we constructed a cheV gene deletion strain of SE (named ΔcheV) and investigated the role of cheV in the virulence of SE. First, the ability to resist environmental stress in vitro, biofilm formation capacity, drug resistance and motility of ΔcheV were analyzed. Secondly, the bacterial adhesion, invasion, intracellular survival assays were performed by cell model. Using a mouse infection model, an in vivo virulence assessment was conducted. To further evaluate the mechanisms implicated by the reduced virulence, qPCR analysis was utilized to examine the expression of the strain’s major virulence genes. Finally, the immune protection rate of ΔcheV was evaluated using a mouse model. Results Compared to C50336, the ΔcheV had significantly reduced survival ability under acidic, alkaline and thermal stress conditions, but there was no significant difference in survival under oxidative stress conditions. There was also no significant change in biofilm formation ability, drug resistance and motility. It was found that the adhesion ability of ΔcheV to Caco-2 cells remained unchanged, but the invasion ability and survival rate in RAW264.7 cells were significantly reduced. The challenge assay results showed that the LD50 values of C50336 and ΔcheV were 6.3 × 105 CFU and 1.25 × 107 CFU, respectively. After the deletion of the cheV gene, the expression levels of fimD, flgG, csgA, csgD, hflK, lrp, sipA, sipB, pipB, invH, mgtC, sodC, rfbH, xthA and mrr1 genes were significantly reduced. The live attenuated ΔcheV provided 100% protection in mice against SE infection. Conclusion All the results confirmed that the deletion of the cheV gene reduces the virulence of SE and provides significant immune protection in mice, indicating that ΔcheV could be potential candidates to be explored as live-attenuated vaccines.

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Section: Methodsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

CheV enhances the virulence of Salmonella Enteritidis, and the Chev-deleted Salmonella vaccine provides immunity in mice

Zhang,

Wu,

Wang

et al. 2024

BMC Vet Res

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“…Referring to previous methods ( Nikhil et al, 2022 ), Five microliters of C50336, Δ pal , and Δ pal + pal bacterial cultures were each stabbed vertically into LB agar medium containing 0.3% agar and incubated at 37°C for 6 h. The motility diameters of C50336, Δ pal , and Δ pal + pal were measured.…”

Section: Methodsmentioning

confidence: 99%

The peptidoglycan-associated lipoprotein gene mutant elicits robust immunological defense in mice against Salmonella enteritidis

Zhao,

Duan,

Zhang

et al. 2024

Front. Microbiol.

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BackgroundSalmonella enteritidis (S. enteritidis), a zoonotic pathogen with a broad host range, presents a substantial threat to global public health safety. Vaccination stands as an effective strategy for the prevention and control of S. enteritidis infection, highlighting an immediate clinical need for the creation of safe and efficient attenuated live vaccines.MethodsIn this study, a S. enteritidis peptidoglycan-associated lipoprotein (pal) gene deletion strain (Δpal), was constructed. To assess its virulence, we conducted experiments on biofilm formation capability, motility, as well as cell and mouse infection. Subsequently, we evaluated the immune-protective effect of Δpal.ResultsIt was discovered that deletion of the pal gene reduced the biofilm formation capability and motility of S. enteritidis. Cell infection experiments revealed that the Δpal strain exhibited significantly decreased abilities in invasion, adhesion, and intracellular survival, with downregulation of virulence gene expression, including mgtC, invH, spvB, sipA, sipB, ssaV, csgA, and pipB. Mouse infection experiments showed that the LD50 of Δpal increased by 104 times, and its colonization ability in mouse tissue organs was significantly reduced. The results indicated that the pal gene severely affected the virulence of S. enteritidis. Further, immunogenicity evaluation of Δpal showed a significant enhancement in the lymphocyte transformation proliferation capability of immunized mice, producing high titers of specific IgG and IgA, suggesting that Δpal possesses good immunogenicity. Challenge protection tests demonstrated that the strain could provide 100% immune protection against wild-type strains in mice.DiscussionThis study proves that the pal gene influences the virulence of S. enteritidis, and Δpal could serve as a candidate strain for attenuated live vaccines, laying the foundation for the development of attenuated live vaccines against Salmonella.

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“…STM harbors a specific stress tolerance mechanism to survive inside the host cells with low partial pressure of oxygen by switching their metabolism from aerobic to anaerobic respiration [3]. The global anaerobic regulators like Fnr, ArcAB, SsrB, NarX/L, and so on, are responsible for the swift change from aerobic to anaerobic metabolism in bacteria, thereby either upregulating or downregulating the expression of a plethora of downstream genes required for their colonization in hypoxic target niches [4][5][6][7].…”

Section: Introductionmentioning

confidence: 99%

Role of narL gene in the pathogenesis of Salmonella Typhimurium

Mani,

Priyadarsini,

K. Channabasappa

et al. 2023

J Basic Microbiol

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Salmonella Typhimurium (STM) is a facultative anaerobe and one of the causative agents of nontyphoidal salmonellosis (NTS). Its anaerobic metabolism is enabled under the hypoxic environment that is encountered inside macrophages and the gut lumen of the host. In both of these niches, free radicals and oxidative intermediates are released by neutrophils as an inflammatory response. These chemical species further undergo reactions to produce nitrate, which is preferably taken up by STM as an electron acceptor in the absence of oxygen. NarL, the response regulator of the two‐component regulatory system NarX/L, and a transcription factor, gets activated under anaerobic nitrate‐rich conditions and upregulates the nitrate reduction during anaerobic respiration of STM. To understand the role of NarL in the pathogenesis of STM, we generated a narL‐knockout (STM:ΔnarL) as well as a narL‐complemented strain of STM. Anaerobically, the mutant displayed no growth defect but a significant attenuation in the swimming (26%) and swarming (61%) motility, and biofilm‐forming ability (73%) in vitro, while these morphotypes got rescued upon genetic complementation. We also observed a downregulation in the expression of genes associated with nitrate reduction (narG) and biofilm formation (csgA and csgD) in anaerobically grown STM:ΔnarL. As compared with wild STM, narL mutant exhibited a threefold reduction in the intracellular replication in both intestinal epithelial cells (INT‐ 407) and monocyte‐derived macrophages of poultry origin. Further, in vivo competitive assay in the liver and spleen of the murine model showed a competitive index of 0.48 ± 0.58 and 0.403668 ± 0.32, respectively, for STM:ΔnarL.

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Recoding anaerobic regulator fnr of Salmonella Typhimurium attenuates it's pathogenicity

Cited by 5 publications

References 58 publications

CheV enhances the virulence of Salmonella Enteritidis, and the Chev-deleted Salmonella vaccine provides immunity in mice

CheV enhances the virulence of Salmonella Enteritidis, and the Chev-deleted Salmonella vaccine provides immunity in mice

The peptidoglycan-associated lipoprotein gene mutant elicits robust immunological defense in mice against Salmonella enteritidis

Role of narL gene in the pathogenesis of Salmonella Typhimurium

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