Dataset Information

The AS87_04050 gene is involved in bacterial lipopolysaccharide biosynthesis and pathogenicity of Riemerella anatipestifer.

ABSTRACT: Riemerella anatipestifer is reported worldwide as a cause of septicemic and exudative diseases of domestic ducks. In this study, we identified a mutant strain RA2640 by Tn4351 transposon mutagenesis, in which the AS87_04050 gene was inactivated by insertion of the transposon. Southern blot analysis indicated that only one insertion was found in the genome of the mutant strain RA2640. SDS-PAGE followed by silver staining showed that the lipopolysaccharide (LPS) pattern of mutant strain RA2640 was different from its wild-type strain Yb2, suggesting the LPS was defected. In addition, the phenotype of the mutant strain RA2640 was changed to rough-type, evident by altered colony morphology, autoaggregation ability and crystal violet staining characteristics. Bacterial LPS is a key factor in virulence as well as in both innate and acquired host responses to infection. The rough-type mutant strain RA2640 showed higher sensitivity to antibiotics, disinfectants and normal duck serum, and higher capability of adherence and invasion to Vero cells, compared to its wild-type strain Yb2. Moreover, the mutant strain RA2640 lost the agglutination ability of its wild-type strain Yb2 to R. anatipestifer serotype 2 positive sera, suggesting that the O-antigen is defected. Animal experiments indicated that the virulence of the mutant strain RA2640 was attenuated by more than 100,000-fold, compared to its wild-type strain Yb2. These results suggested that the AS87_04050 gene in R. anatipestifer is associated with the LPS biosynthesis and bacterial pathogenicity.

SUBMITTER: Wang X

PROVIDER: S-EPMC4193840 | biostudies-literature | 2014

REPOSITORIES: biostudies-literature

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The AS87_04050 gene is involved in bacterial lipopolysaccharide biosynthesis and pathogenicity of Riemerella anatipestifer.

Wang Xiaolan X Ding Chan C Wang Shaohui S Han Xiangan X Hou Wanwan W Yue Jiaping J Zou Jiechi J Yu Shengqing S

PloS one 20141010 10

Riemerella anatipestifer is reported worldwide as a cause of septicemic and exudative diseases of domestic ducks. In this study, we identified a mutant strain RA2640 by Tn4351 transposon mutagenesis, in which the AS87_04050 gene was inactivated by insertion of the transposon. Southern blot analysis indicated that only one insertion was found in the genome of the mutant strain RA2640. SDS-PAGE followed by silver staining showed that the lipopolysaccharide (LPS) pattern of mutant strain RA2640 was ...[more]

PMID: 25303276

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Project description:Duck infectious serositis, also known as Riemerella anatipestifer disease, infects domestic ducks, geese, and turkeys and wild birds. However, the regulatory mechanism of its pathogenicity remains unclear. The PhoPR two-component system (TCS) was first reported in Gram-negative bacteria in our previous research and was demonstrated to be involved in virulence and gene expression. Here, DNA affinity purification sequencing (DAP-seq) was applied to further explore the regulation of PhoPR in relation to pathogenicity in R. anatipestifer. A conserved motif was identified upstream of 583 candidate target genes which were directly regulated by PhoP. To further confirm the genes which are regulated by PhoR and PhoP, single-gene-deletion strains were constructed. The results of transcriptome analysis using next-generation RNA sequencing showed 136 differentially expressed genes (DEGs) between the ΔphoP strain and the wild type (WT) and 183 DEGs between the ΔphoR strain and the WT. The candidate target genes of PhoP were further identified by combining transcriptome analysis and DAP-seq, which revealed that the main direct regulons of PhoP are located on the membrane and PhoP is involved in regulating aerotolerance. Using the in vivo duck model, the pathogenicity of ΔphoP and ΔphoR mutants was found to be significantly lower than that of the WT. Together, our findings provide insight into the direct regulation of PhoP and suggest that phoPR is essential for the pathogenicity of R. anatipestifer. The gene deletion strains are expected to be candidate live vaccine strains of R. anatipestifer which can be used as ideal genetic engineering vector strains for the expression of foreign antigens. IMPORTANCE Riemerella anatipestifer is a significant pathogen with high mortality in the poultry industry that causes acute septicemia and infectious polyserositis in ducks, chickens, geese, and other avian species. Previously, we characterized the two-component system encoded by phoPR and found that R. anatipestifer almost completely lost its pathogenicity for ducklings when phoPR was deleted. However, the mechanism of PhoPR regulation of virulence in R. anatipestifer had not been deeply explored. In this study, we utilized DAP-seq to explore the DNA-binding sites of PhoP as a response regulator in the global genome. Furthermore, phoP and phoR were deleted separately, and transcriptomics analysis of the corresponding gene deletion strains was performed. We identified a series of directly regulated genes of the PhoPR two-component system. The duckling model showed that both PhoP and PhoR are essential virulence-related factors in R. anatipestifer.

Project description:The bacterium Riemerella anatipestifer requires iron for growth, but the mechanism of iron uptake is not fully understood. In this study, we disrupted the Feo system and characterized its function in iron import in R. anatipestifer ATCC 11845. Compared to the parent strain, the growth of the ΔfeoA, ΔfeoB, and ΔfeoAB strains was affected under Fe3+-limited conditions, since the absence of the feo system led to less intracellular iron than in the parent strain. In parallel, the ΔfeoAB strain was shown to be less sensitive to streptonigrin, an antibiotic that requires free iron to function. The sensitivity of the ΔfeoAB strain to hydrogen peroxide was also observed to be diminished compared with that of the parent strain, which could be related to the reduced intracellular iron content in the ΔfeoAB strain. Further research revealed that feoA and feoB were directly regulated by iron through the Fur regulator and that the transcript levels of feoA and feoB were significantly increased in medium supplemented with 1 mM MnCl2, 400 μM ZnSO4, and 200 μM CuCl2. Finally, it was shown that the ΔfeoAB strain of R. anatipestifer ATCC 11845 was significantly impaired in its ability to colonize the blood, liver, and brain of ducklings. Taken together, these results demonstrated that FeoAB supports ferrous iron acquisition in R. anatipestifer and plays an important role in R. anatipestifer colonization. IMPORTANCE In Gram-negative bacteria, the Feo system is an important ferrous iron transport system. R. anatipestifer encodes an Feo system, but its function unknown. As iron uptake may be required for oxidative stress protection and virulence, understanding the contribution of iron transporters to these processes is crucial. This study showed that the ΔfeoAB strain is debilitated in its ability to import iron and that its intracellular iron content was constitutively low, which enhanced the resistance of the deficient strain to H2O2. We were surprised to find that, in addition to responding to iron, the Feo system may play an important role in sensing manganese, zinc, and copper stress. The reduced colonization ability of the ΔfeoAB strain also sheds light on the role of iron transporters in host-pathogen interactions. This study is important for understanding the cross talk between iron and other metal transport pathways, as well as the pathogenic mechanism in R. anatipestifer.

Dataset Information

The AS87_04050 gene is involved in bacterial lipopolysaccharide biosynthesis and pathogenicity of Riemerella anatipestifer.

Publications

The AS87_04050 gene is involved in bacterial lipopolysaccharide biosynthesis and pathogenicity of Riemerella anatipestifer.

Similar Datasets

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