Hydrogen Peroxide Production by Streptococcus pneumoniae Results in Alpha-hemolysis by Oxidation of Oxy-hemoglobin to Met-hemoglobin.
Hydrogen Peroxide Production by Streptococcus pneumoniae Results in Alpha-hemolysis by Oxidation of Oxy-hemoglobin to Met-hemoglobin.
复制标题
肺炎链球菌产生的过氧化氢通过氧血红蛋白氧化为血红蛋白导致α -溶血。
DOI：
10.1128/msphere.01117-20
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发表时间：
2020-12-09
期刊：
影响因子：
4.8
通讯作者：
Vidal JE
中科院分区：
文献类型：
--
作者：
McDevitt E;Khan F;Scasny A;Thompson CD;Eichenbaum Z;McDaniel LS;Vidal JE
有一种误解，认为在肺炎链球菌和其他α溶血性链球菌的血琼脂平板培养物上观察到的α溶血是由溶血素或过氧化氢引起的红细胞裂解产生的。我们在研究过程中注意到，野生型肺炎链球菌菌株和溶血素（例如肺炎球菌溶血素）敲除突变体在血琼脂平板上产生了α溶血晕。肺炎链球菌和其他链球菌在血琼脂平板上产生绿色光晕，称为α溶血。临床微生物学实验室利用这种表型来报告α-溶血性链球菌（包括肺炎链球菌）和其他细菌的培养结果。血琼脂平板上的 α 溶血晕与肺炎球菌肺炎球菌溶血素 (Ply) 的溶血活性有关，或者在较小程度上与肺炎链球菌产生的过氧化氢对红细胞的裂解有关。我们研究了肺炎链球菌产生的α溶血晕的分子基础。野生型菌株 TIGR4、D39、R6 和 EF3030 以及同基因衍生物 Δply 突变体在血琼脂平板上产生类似的 α 溶血晕，而过氧化氢敲除的 ΔspxB ΔlctO 突变体的培养物缺乏这种特征性晕。此外，在存在过氧化氢酶的情况下，野生型（wt）和Δply突变菌株的培养物中不存在α溶血晕。光谱研究表明，TIGR4 的培养上清液在孵育 30 分钟内从红细胞中释放出与血红蛋白结合的血红蛋白（血红蛋白），并将氧合血红蛋白氧化为高铁血红蛋白。正如预期的那样，鉴于 Ply 溶血活性以及过氧化氢有助于 Ply 的释放，TIGR4Δply 和 ΔspxB ΔlctO 同基因突变体显着减少了红细胞中血红素-血红蛋白的释放。然而，产生过氧化氢的TIGR4Δply将氧合血红蛋白氧化为高铁血红蛋白，而TIGR4ΔspxBΔlctO未能产生氧合血红蛋白的氧化。对所有其他野生型菌株和同基因突变体进行的研究得出了类似的结果。我们证明，所谓的α溶血晕是由肺炎链球菌产生的过氧化氢将氧合血红蛋白（Fe+2）氧化为非氧结合高铁血红蛋白（Fe+3）引起的。重要性 有一种误解，认为在肺炎链球菌和其他 α 溶血性链球菌的血琼脂平板培养物上观察到的 α 溶血是由溶血素或过氧化氢引起的红细胞裂解产生的。我们在研究过程中注意到，野生型肺炎链球菌菌株和溶血素（例如肺炎球菌溶血素）敲除突变体在血琼脂平板上产生了α溶血晕。相比之下，在四种不同菌株中制备的过氧化氢缺陷突变体缺乏特征性的α溶血晕。我们还证明野生型菌株和肺炎球菌溶血素突变体将氧合血红蛋白氧化为高铁血红蛋白。然而，过氧化氢敲除突变体无法氧化氧合血红蛋白。因此，肺炎链球菌和其他所谓的α溶血性链球菌培养物上形成的绿色晕圈是由过氧化氢产生的氧合血红蛋白氧化引起的。肺炎球菌肺炎期间，肺部可能会发生氧合血红蛋白氧化为非结合氧形式、高铁血红蛋白的情况。
There is a misconception that alpha-hemolysis observed on blood agar plate cultures of Streptococcus pneumoniae and other alpha-hemolytic streptococci is produced by a hemolysin or, alternatively, by lysis of erythrocytes caused by hydrogen peroxide. We noticed in the course of our investigations that wild-type S. pneumoniae strains and hemolysin (e.g., pneumolysin) knockout mutants produced the alpha-hemolytic halo on blood agar plates. Streptococcus pneumoniae and other streptococci produce a greenish halo on blood agar plates referred to as alpha-hemolysis. This phenotype is utilized by clinical microbiology laboratories to report culture findings of alpha-hemolytic streptococci, including S. pneumoniae, and other bacteria. The alpha-hemolysis halo on blood agar plates has been related to the hemolytic activity of pneumococcal pneumolysin (Ply) or, to a lesser extent, to lysis of erythrocytes by S. pneumoniae-produced hydrogen peroxide. We investigated the molecular basis of the alpha-hemolysis halo produced by S. pneumoniae. Wild-type strains TIGR4, D39, R6, and EF3030 and isogenic derivative Δply mutants produced similar alpha-hemolytic halos on blood agar plates, while cultures of hydrogen peroxide knockout ΔspxB ΔlctO mutants lacked this characteristic halo. Moreover, in the presence of catalase, the alpha-hemolysis halo was absent in cultures of the wild-type (wt) and Δply mutant strains. Spectroscopic studies demonstrated that culture supernatants of TIGR4 released hemoglobin-bound heme (heme-hemoglobin) from erythrocytes and oxidized oxy-hemoglobin to met-hemoglobin within 30 min of incubation. As expected, given Ply hemolytic activity and that hydrogen peroxide contributes to the release of Ply, TIGR4Δply and ΔspxB ΔlctO isogenic mutants had significantly decreased release of heme-hemoglobin from erythrocytes. However, TIGR4Δply that produces hydrogen peroxide oxidized oxy-hemoglobin to met-hemoglobin, whereas TIGR4ΔspxB ΔlctO failed to produce oxidation of oxy-hemoglobin. Studies conducted with all other wt strains and isogenic mutants resulted in similar findings. We demonstrated that the so-called alpha-hemolysis halo is caused by the oxidation of oxy-hemoglobin (Fe+2) to a non-oxygen-binding met-hemoglobin (Fe+3) by S. pneumoniae-produced hydrogen peroxide. IMPORTANCE There is a misconception that alpha-hemolysis observed on blood agar plate cultures of Streptococcus pneumoniae and other alpha-hemolytic streptococci is produced by a hemolysin or, alternatively, by lysis of erythrocytes caused by hydrogen peroxide. We noticed in the course of our investigations that wild-type S. pneumoniae strains and hemolysin (e.g., pneumolysin) knockout mutants produced the alpha-hemolytic halo on blood agar plates. In contrast, hydrogen peroxide-defective mutants prepared in four different strains lacked the characteristic alpha-hemolysis halo. We also demonstrated that wild-type strains and pneumolysin mutants oxidized oxy-hemoglobin to met-hemoglobin. Hydrogen peroxide knockout mutants, however, failed to oxidize oxy-hemoglobin. Therefore, the greenish halo formed on cultures of S. pneumoniae and other so-called alpha-hemolytic streptococci is caused by the oxidation of oxy-hemoglobin produced by hydrogen peroxide. Oxidation of oxy-hemoglobin to the nonbinding oxygen form, met-hemoglobin, might occur in the lungs during pneumococcal pneumonia.