*4.3. Histological Study*

Two weeks after inoculation, fresh root segments were fixed in 50 mM phosphate buffer (pH 6.8) containing 2.5% glutaraldehyde and 1.6% paraformaldehyde for 4 h at room temperature. After fixation, the samples were rinsed three times with phosphoric acid buffer (pH 6.8, 0.1 M) for about 15 min each time and then dehydrated with a graded ethanol series (15% ethanol for 30 min; 30% ethanol for 30 min; 50% ethanol for 30 min; 70% ethanol for 1 h; 85% ethanol for 1 h; 95% ethanol for 1 h; absolute ethanol for 1 h). After dehydration, the samples were embedded in LRwhite gradient mixture (25% LRwhite for 24 h; 50% LRwhite for 24 h; 75% LRwhite for 24 h; 100% LRwhite for 24 h) and polymerized for 48 h at 60 ◦C. Next, three-μm-thick sections were cut using glass knives with a rotary microtome (Autocut 2040; Reichert-Jung; Germany). The sections were collected on slides and stained with 0.05% (*w*/*v*) toluidine blue O in benzoate buffer for general histology examinations. The

sections were examined, and images were captured digitally using a digital camera attached to the microscope (Axio Imager A1; Carl Zeiss, Oberkochen, Germany).
