Tomoyasu Toshifumi, Yamasaki Takahiro, Chiba Shinya, Kusaka Shingo, Tabata Atsushi, Whiley Robert A, Nagamune Hideaki
Department of Bioscience and Bioindustry, Graduate School of Bioscience and Bioindustry, Tokushima University Graduate School, Tokushima, Japan.
Department of Resource Circulation Engineering, Center for Frontier Research of Engineering, Tokushima University Graduate School, Tokushima, Japan.
Infect Immun. 2017 Aug 18;85(9). doi: 10.1128/IAI.00379-17. Print 2017 Sep.
is an opportunistic bacterial pathogen secreting a human-specific cytolysin called intermedilysin (ILY) as a major pathogenic factor. This bacterium can degrade glycans into monosaccharides using two glycosidases, multisubstrate glycosidase A (MsgA) and neuraminidase (NanA). Here, we detected a stronger hemolytic activity mediated by ILY when PC574 was cultured in fetal bovine serum (FBS) than when it was grown in the standard culture medium. FBS-cultured cells also showed higher MsgA and NanA activity, although overproduction of ILY in FBS was undetectable in mutants -null and -null. Addition of purified MsgA and NanA to the FBS resulted in a release of 2.8 mM galactose and 4.3 mM -acetylneuraminic acid; these sugar concentrations were sufficient to upregulate the expression of ILY, MsgA, and NanA. Conversely, when strain PC574 was cultured in human plasma, no similar increase in hemolytic activity was observed. Moreover, addition of human plasma to the culture in FBS appeared to inhibit the stimulatory effect of FBS on ILY, MsgA, and NanA, although there were individual differences among the plasma samples. We confirmed that human plasma contains immunoglobulins that can neutralize ILY, MsgA, and NanA activities. In addition, human plasma had a neutralizing effect on cytotoxicity of toward HepG2 cells in FBS, and a higher concentration of human plasma was necessary to reduce the cytotoxicity of an ILY-high-producing strain than an ILY-low-producing strain. Overall, our data show that blood contains factors that stimulate and inhibit ILY expression and activity, which may affect pathogenicity of .
是一种机会性细菌病原体，分泌一种名为中间溶血素（ILY）的人类特异性细胞溶素作为主要致病因素。这种细菌可以使用两种糖苷酶，即多底物糖苷酶A（MsgA）和神经氨酸酶（NanA），将聚糖降解为单糖。在这里，我们检测到当PC574在胎牛血清（FBS）中培养时，ILY介导的溶血活性比在标准培养基中培养时更强。FBS培养的细胞也显示出更高的MsgA和NanA活性，尽管在null和null突变体中未检测到FBS中ILY的过量产生。向FBS中添加纯化的MsgA和NanA导致释放2.8 mM半乳糖和4.3 mM N-乙酰神经氨酸；这些糖浓度足以上调ILY、MsgA和NanA的表达。相反，当PC574菌株在人血浆中培养时，未观察到溶血活性有类似增加。此外，向FBS中的培养物中添加人血浆似乎抑制了FBS对ILY、MsgA和NanA的刺激作用，尽管血浆样本之间存在个体差异。我们证实人血浆中含有可以中和ILY、MsgA和NanA活性的免疫球蛋白。此外，人血浆对FBS中对HepG2细胞的细胞毒性有中和作用，并且需要更高浓度的人血浆来降低高ILY产生菌株的细胞毒性，而不是低ILY产生菌株。总体而言，我们的数据表明血液中含有刺激和抑制ILY表达及活性的因子，这可能会影响的致病性。
