Functional Characterization of FeoAB in Iron Acquisition and Pathogenicity in Riemerella anatipestifer

Abstract

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 Δ<i>feoA,</i> Δ<i>feoB,</i> and Δ<i>feoAB</i> strains was affected under Fe³⁺-limited conditions, since the absence of the <i>feo</i> system led to less intracellular iron than in the parent strain. In parallel, the Δ<i>feoAB</i> strain was shown to be less sensitive to streptonigrin, an antibiotic that requires free iron to function. The sensitivity of the Δ<i>feoAB</i> 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 Δ<i>feoAB</i> strain. Further research revealed that <i>feoA</i> and <i>feoB</i> were directly regulated by iron through the Fur regulator and that the transcript levels of <i>feoA</i> and <i>feoB</i> were significantly increased in medium supplemented with 1 mM MnCl₂, 400 μM ZnSO₄, and 200 μM CuCl₂. Finally, it was shown that the Δ<i>feoAB</i> 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. <b>IMPORTANCE</b> 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 Δ<i>feoAB</i> 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 Δ<i>feoAB</i> 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.