Occurrence of Root-Lesion Nematode Pratylenchus coffeae on Corn in Xinjiang Uygur Autonomous Region, China.

Abstract

Pratylenchus coffeae Filipjev & Schuurmans Stekhoven, 1941, is one of the most important root-lesion nematodes (RLN) parasitizing many agronomic and industrial crops (Wang et al. 2021). Corn (Zea mays L.) is one economically important crop in China, with 35 million hectares cultivated annually (Li et al. 2019). In July 2019, a survey of RLN was carried out in corn field planting with cultivar Heyu 187 in Chuanba village in Qitai County, Xinjiang Uygur Autonomous Region, China. Five root/soil samples were collected from poor growing plants with distinct brown lesions. Nematodes were extracted from the collected root/soil samples with the modified Baermann funnel method (Hooper et al. 2005). The average of 157 RLN per 100 cm3 of soil and 43 RLN per gram of fresh root were extracted. The obtained RLN were sterilized with 0.3% streptomycin sulfate and cultured on carrot disks at 25°C. Twenty petri dishes with carrot disks, each inoculated with one female. The morphological and molecular characteristics of RLN cultured on carrot disks were examined for species identification. Morphological measurements of adult females (n=15) included body length (range = 529.0 to 658.0 μm, mean = 571.0 μm), head with two lip annuli, stylet (15.5 to 17.0 μm, 16.0 μm), tail length (27.5 to 32.5 μm, 30.5 μm), a (23.8 to 32.9, 28.5), b (5.8 to 7.1, 6.5), c (16.5 to 23.4, 18.9), and V (76.6 to 83.1%, 80.8%). Morphological measurements of adult males (n=15) were body length (range = 479.5 to 568.0 μm, mean = 516.0 μm), head with two lip annuli, stylet (14.5 to 15.5 μm, 15.0 μm), tail length (24.0 to 29.0 μm, 26.0 μm), spicule length (16.4 to 19.0 μm, 17.5 μm), gubernaculum length (4.4 to 5.3 μm, 4.9 μm), a (29.2 to 32.5, 31.0), b (5.7 to 6.9, 6.2), and c (18.2 to 22.6, 19.8). The morphological characters of this population are consistent with the description of P. coffeae (Castillo and Vovlas, 2007). Nematode DNA was extracted from an individual female. The primers of D2A/D3B (5'-ACAAGTACCGTGAGGGAAAGTTG-3'/5'-TCGGAAGGAACCAGCTACTA-3') (Subbotin et al. 2006) and 18S/26S (5'-TTGATTACGTCCCTGCCCTTT-3' / 5'-TTTCACTCGCCGTTACTAAGG-3') (Vrain et al. 1992) were used to amplify the D2/D3 expansion region of the 28S rRNA gene and the rDNA internal transcribed spacer (ITS) region, respectively. The PCR products were purified and transformed to E. coli strain DH5α, and then sequenced by Sangon Biotech Co. Ltd. (Shanghai, China). The obtained sequences of the D2/D3 region (793 bp) and the ITS region (1,242 bp) were submitted to GenBank, and the accession numbers for D2/D3 region were OK103614 and OK103619 which had 98.6% and 100% identity with the reported P. coffeae sequences (KC490925); the two obtained ITS sequences accession numbers OK103603 and OK103613) had more than 99% identity with published P. coffeae sequences from GenBank (e.g., LC030410, LC030395, MH134508 and LC030380). Hence, both morphological and molecular data demonstrated the presence of P. coffeae. To further confirm reproduction on corn, the obtained RLN population was used to inoculate corn plants in 2-liter pots containing 1.8-liter sterilized and mixed soil with 2 pastoral soil: 1 substrate in greenhouse at 27°C. About 15 days after sowing, each pot with one corn plant (cv. Heyu 187) with the same growth status was selected to inoculate P. coffeae. Five small holes near the roots were made using a glass rod. Approximately 1,000 mixed stage nematodes of P. coffeae were then pipetted into the holes of each plant. Eight replications were performed. Eight additional pots of uninoculated corn plants were used as control. After 2 months, corn roots were washed and brown lesions were observed on roots. The average number of RLN/pot was approximately 5,030 in soil and 2,870 in roots, and each pot had an average of 7.9 reproduction factors (final population/initial population), indicating that this nematode population infects and reproduces well on this corn cultivar. No nematodes and symptoms was detected in the control pot. The nematode of P. coffeae has only been reported on corn in Guangdong, Liaoning, Shangdong and Henan Provinces in China (Liu et al. 1996; Liu et al. 2001; Xia et al. 2021). To our knowledge, this is the first report of P. coffeae infecting corn in Xinjiang Uygur Autonomous Region of China. Since RLN can cause considerable damage to corn, one of the most important food crops produced in China, strategic measures should be taken to prevent the spread of P. coffeae to other regions.