Thirty-five environmental isolates of Shiga toxin-producing Escherichia coli (STEC) were analyzed by MALDI-TOF-TOF mass spectrometry, top-down/middle-down proteomics and DNA sequencing. Clinically-relevant Shiga toxin 2 (Stx2) produced by these STEC strains were subtyped based on MS and MS/MS (tandem mass spectrometry) of the intact B-subunit (top-down) and A2 fragment (middle-down) of the A-subunit using antibiotic-induced protein expression. Antibiotic induction of Stx2 was found to be strain dependent. By proteomic analysis, seventeen strains were identified as Stx2a, six strains as Stx2c, four strains as either Stx2a or 2c and eight strains as either Stx2a, 2c or 2d. DNA sequencing indicated only stx_2a and stx_2c genes as being present in these strains. Weak induction of Stx2 for certain strains made it difficult to distinguish between clinical subtypes by proteomic analysis. Very weak toxin induction in eight strains was consistent with a ∼1300 bp transposon insertion in the stx_2c A-subunit gene identified by DNA sequencing. DNA sequencing also revealed the presence of two bacteriophage (BP) in three strains with a stx_2a gene in each BP genome. Middle-down proteomic analysis of the A2 fragment confirmed expression of two stx_2a genes present in one of these strains based on a slight difference in the amino acid sequence (D ↔ E substitution) in the two A2 fragments.

通过 MALDI-TOF-TOF 质谱、自上而下/中下蛋白质组学和 DNA 测序分析了35 种产志贺毒素大肠杆菌(STEC) 的环境分离物。由这些 STEC 菌株产生的临床相关志贺毒素 2 (Stx2) 基于完整 B 亚基（自上而下）和 A2 片段（中下）的 MS 和 MS/MS（串联质谱）进行亚型-亚基使用抗生素诱导的蛋白质表达。发现 Stx2 的抗生素诱导是菌株依赖性的。通过蛋白质组学分析，17 株被鉴定为 Stx2a，6 株为 Stx2c，4 株为 Stx2a 或 2c，8 株为 Stx2a、2c 或 2d。DNA 测序表明只有stx _2a和stx _2c基因存在于这些菌株中。某些菌株对 Stx2 的弱诱导使得难以通过蛋白质组学分析区分临床亚型。毒素诱导8株非常弱的是与在一个~1300 bp的转座子插入一致STX _2c中通过DNA测序来鉴定A-亚基基因。DNA 测序还揭示了在每个 BP 基因组中具有stx _2a基因的三个菌株中存在两种噬菌体 (BP) 。A2 片段的中下蛋白质组学分析证实了存在于这些菌株之一中的两个stx _2a基因的表达，这是基于两个 A2 片段中氨基酸序列的轻微差异（D ↔ E 取代）。