In this study, 58 Rhizoctonia isolates were obtained from plants exhibiting symptoms of root and stem rot in kiwifruit orchards in six different provinces in the Black Sea region of Turkey. The isolates were analysed for their cultural characteristics, anastomosis groups (AGs) and subgroups according to nucleotide sequence variations in the internal transcribed spacer (ITS) regions including the 5.8S gene of nuclear ribosomal DNA (rDNA) and pathogenicity. Based on ITS1‐5.8S‐ITS2 rDNA sequencing, 52 binucleate (BN) Rhizoctonia isolates were identified as belonging to AG‐O (17), AG‐L (1), AG‐G (7), AG‐I (1), AG‐A (9), AG‐Fa (9), AG‐Fb (1), AG‐R (3) and AG‐P (4). Of the remaining 6 isolates, three belonged to multinucleate (MN) R. solani AG 4 HG‐I, one belonged to AG 4 HG‐II, one belonged to AG 5 and one belonged to AG 1‐IB. The sequences of these isolates were aligned and compared with other Rhizoctonia sequences retrieved from GenBank (NCBI), and phylogenetic analysis was used to determine evolutionary relationships among them. All isolates used in the temperature experiments grew between 15–35°C, and their growth was completely inhibited at 5 and 40°C, with a few exceptions. In the pathogenicity tests carried out using 58.6% of the isolates on kiwifruit seedlings, it was determined that there were statistically significant virulence differences among BN and MN Rhizoctonia isolates (p < 0.05). The virulence of BN Rhizoctonia ranged from 0.2 to 4.0, whereas that of MN varied from 2.4 to 4.0. Unlike some isolates of AG‐R, AG 4 HG‐II and AG 1‐IB, which had no significant difference between them in virulence, AG‐P and AG 4 HG‐I isolates significantly reduced plant height, shoot and root dry weights and root length when compared to control plants. To our knowledge, it is the first detailed report of AGs of BN and MN Rhizoctonia, the causal agent of root and stem rot on kiwifruit in Turkey and in the world.