RyhB Paralogs Downregulate the Expressions of Multiple Survival-Associated Genes and Attenuate the Survival of Salmonella Enteritidis in the Chicken Macrophage HD11

Xia Meng; Mengping He; Binjie Chen; Pengpeng Xia; Jinqiu Wang; Chunhong Zhu; Heng Wang; Guoqiang Zhu

doi:10.3390/microorganisms11010214

RyhB Paralogs Downregulate the Expressions of Multiple Survival-Associated Genes and Attenuate the Survival of Salmonella Enteritidis in the Chicken Macrophage HD11

Microorganisms. 2023 Jan 15;11(1):214. doi: 10.3390/microorganisms11010214.

Authors

Xia Meng^{1

2

3}, Mengping He^{1

2

3}, Binjie Chen^{1

2}, Pengpeng Xia^{1

2

3}, Jinqiu Wang⁴, Chunhong Zhu⁵, Heng Wang^{1

2

3}, Guoqiang Zhu^{1

2

3}

Affiliations

¹ College of Veterinary Medicine, Yangzhou University, Yangzhou 225009, China.
² International Research Laboratory of Prevention and Control of Important Animal Infectious Diseases and Zoonotic Diseases of Jiangsu Higher Education Institutions, Yangzhou University, Yangzhou 225009, China.
³ Jiangsu Co-Innovation Center for Prevention and Control of Important Animal Infectious Diseases and Zoonoses, Yangzhou University, Yangzhou 225009, China.
⁴ Department of Animal Husbandry and Veterinary Medicine, Beijing Agricultural Vocational College, Beijing 102442, China.
⁵ Jiangsu Institute of Poultry Science, Yangzhou 225125, China.

Abstract

RyhB-1 and RyhB-2 are small non-coding RNAs in Salmonella that act as regulators of iron homeostasis by sensing the environmental iron concentration. Expressions of RyhB paralogs from Salmonella Typhimurium are increased within microphages. RyhB paralogs restrain the growth of S. Typhimurium in RAW264.7 macrophages by modulating the expression of Salmonella pathogenicity island 1 (SPI-1) genes sicA and rtsB. However, little is known about the regulatory role of RyhBs and their virulence-associated targets in Salmonella Enteritidis. We studied candidate targets of RyhB paralogs via RNA-Seq in conditions of iron limitation and hypoxia. RyhB paralogs were expressed when the S. Enteritidis strain CMCC(B)50336 (SE50336) interacted with the chicken macrophage line HD11. We analyzed gene expression associated with Salmonella survival and replication in macrophages in wild-type strain SE50336 and the RyhB deletion mutants after co-incubation with HD11 and screened out targets regulated by RyhBs. The expressions of both RyhB-1 and RyhB-2 were increased after co-incubation with HD11 for 8 h and several survival-associated genes within macrophages, such as ssaI, sseA, pagC, sodC, mgtC, yaeB, pocR, and hns, were upregulated in the ryhB-1 deletion mutant. Specifically, ssaI, the type-three secretion system 2 (T3SS-2) effector encoded by SPI-2, which promoted the survival of Salmonella in macrophages, was upregulated more than 3-fold in the ryhB-1 deletion mutant. We confirmed that both RyhB-1 and RyhB-2 downregulated the expression of ssaI to repress its mRNA translation by directly interacting with its coding sequence (CDS) region via an incomplete complementary base-pairing mechanism. The SPI-2 gene sseA was indirectly modulated by RyhB-1. The survival assays in macrophages showed that the ability of intracellular survival of ryhB-1 and/or ryhB-2 deletion mutants in HD11 was higher than that of the wild-type strain. These results indicate that RyhB paralogs downregulate survival-related virulence factors and attenuate the survival of S. Enteritidis inside chicken macrophage HD11.

Keywords: RyhB paralogs; Salmonella Enteritidis; macrophages; ssaI; survival.

Abstract

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