1 Departamento de Microbiologia e Imunologia, Instituto de Biociências, Universidade Estadual Paulista "Júlio de Mesquita Filho" (UNESP), Botucatu, SP, Brazil.
2 Centro de Bacteriologia, Instituto Adolfo Lutz (IAL), São Paulo SP, Brazil.
J Med Microbiol. 2019 Jun;68(6):940-951. doi: 10.1099/jmm.0.000998. Epub 2019 May 20.
This study aimed to characterize 82 atypical enteropathogenic Escherichia coli (aEPEC) isolates, obtained from patients with diarrhea in Brazil, regarding their adherence patterns on HeLa cells and attaching and effacing (AE) lesion pathways.
The adherence and fluorescence-actin staining (FAS) assays were performed using HeLa cells. AE lesion pathways were determined through the detection of tyrosine residue 474 (Y474) phosphorylation in the Tir protein, after its translocation to host cells, and by PCR assays for tir genotyping and detection of Tir-cytoskeleton coupling protein (tccP) genes.
Regarding the adherence pattern, determined in the presence of d-mannose, 12 isolates (14.6 %) showed the localized adherence (LA)-like pattern, 3 (3.7 %) the aggregative adherence pattern and 4 (4.9 %) a hybrid LA/diffuse adherence pattern. In addition, 36 (43.9 %) isolates displayed an undefined adherence, and 26 (31.7 %) were non-adherent (NA), while one (1.2 %) caused cell detachment. Among the 26 NA aEPEC isolates, 11 showed a type 1 pilus-dependent adherence in assays performed without d-mannose, while 15 remained NA. Forty-eight (58.5 %) aEPEC were able to trigger F-actin accumulation underneath adherent bacteria (FAS-positive), which is an important feature of AE lesions. The majority (58.3 %) of these used the Tir-Nck pathway, while 39.6 % may use both Tir-Nck and Tir-TccP pathways to induce AE lesions.
Our results reveal the diversity of strategies used by aEPEC isolates to interact with and damage epithelial host cells, thereby causing diarrheal diseases.
本研究旨在对 82 株从巴西腹泻患者中分离出的非典型肠致病性大肠杆菌（aEPEC）菌株进行鉴定，评估它们在 HeLa 细胞上的黏附模式和附着与破坏（AE）病变途径。
采用 HeLa 细胞进行黏附实验和荧光肌动蛋白染色（FAS）实验。通过检测 Tir 蛋白易位到宿主细胞后酪氨酸残基 474（Y474）的磷酸化以及 Tir 基因分型和 Tir-细胞骨架偶联蛋白（tccP）基因的 PCR 检测，确定 AE 病变途径。
在存在 D-甘露糖的情况下，根据黏附模式的测定，有 12 株（14.6%）表现为局灶黏附（LA）样模式，3 株（3.7%）表现为聚集黏附模式，4 株（4.9%）表现为 LA/弥散黏附混合模式。此外，36 株（43.9%）表现为不明确的黏附，26 株（31.7%）为非黏附（NA），而 1 株（1.2%）导致细胞脱落。在 26 株非黏附性 aEPEC 菌株中，11 株在不使用 D-甘露糖的情况下，通过 1 型菌毛依赖的黏附实验显示出黏附性，而其余 15 株仍为非黏附性。48 株（58.5%）aEPEC 能够诱导黏附细菌下方肌动蛋白的积累（FAS 阳性），这是 AE 病变的一个重要特征。其中大多数（58.3%）菌株使用 Tir-Nck 途径，而 39.6%的菌株可能同时使用 Tir-Nck 和 Tir-TccP 途径来诱导 AE 病变。
我们的研究结果揭示了 aEPEC 菌株与上皮宿主细胞相互作用并造成损伤的多样性策略，从而导致腹泻疾病。
