First Record of Aphelenchoides besseyi on Foxtail Millet (Setaria italica) in Henan Province of China

Abstract

Aphelenchoides besseyi is one of the important plant-parasitic nematodes on rice, reducing approximate 10-20% of the rice yield annually (Jones et al. 2013). Foxtail millet (Setaria italica) has been a major cereal crop in Northern China, especially in the semi-arid areas of this region, for thousands of years. In August of 2019 and 2020, a survey of nematodes on autumn grain crops was performed each year. One foxtail millet field (N34° 58′ 027″ and E113° 39′ 059″) in Yuanyang County of Henan Province caught our attention. Some upper leaves showed chlorosis without or with necrotic tips, and flag leaves presented crinkling and distortion, stalks were colored, earheads were vertical, glumes were brown or light black and open, and grains became thin. A total of ten samples were collected, and the nematodes were isolated from the spike pieces by shallow plate method and counted under a stereomicroscope. The average number of nematodes per earhead of foxtail millet counted up to 1738.75 ± 107.72. Morphologically, females were slender with a short stylet, an oval metacorpus with a distinct valve, a labial region slightly wider than the first body annulus and a conoid tail with a terminus bearing a star-shaped mucro with four pointed processes. The females were characterized as follows (mean ± SD; n=20): body length (L) = 668.92 ± 12.73 µm (647.38 to 689.70 µm); maximum body width (W) = 14.35 ± 1.11 µm (12.12 to 16.88 µm); L/W = 46.83 ± 2.94 (40.44 to 50.03); tail length = 38.93 ± 3.48 µm (33.41 to 45.92 µm); L/tail length = 17.31 ± 1.44 (14.47 to 19.62); and stylet length (ST) = 11.57 ± 0.57 µm (10.77 to 12.34 µm). The males had three pairs of ventrosubmedian papillae with the first one adanal, spicula curved with a slight basal process, terminus bearing four mucrones arranged variably, and the whole worm was in ‘J’ shape. The males could be described as follows (mean ± SD, n = 20): L = 606.66 ± 10.70 µm (586.49 to 626.37 µm); W = 13.95 ± 0.60 µm (12.71 to 14.94 µm); L/W = 43.55 ± 1.69 (40.73 to 46.43); tail length = 35.54 ± 1.93 µm (31.41 to 38.18 µm); L/tail length = 17.07 ± 0.79 (16.05 to 18.67); ST = 11.53 ± 0.56 µm (1061 to 12.76 µm). All the key morphometrics were consistent with those of A. besseyi reported from Brazil (Favoreto et al. 2018) and China (Lin et al. 2004; Ou et al. 2014). The amplifications of rDNA internal transcribed spacer (ITS) fragments generated a PCR fragment of 830 bp from a single nematode, using the primers set TW81 (5′-GTTTCCGTAGGTGAACCTGC-3′) and AB28 (5′-ATATGCTTAAGTTCAGCGGGT-3′) (Joyce et al. 1994). Five independent PCR experiments were conducted, and all the PCR products were purified and sequenced. Nucleotide sequence of ITS-rDNA was deposited in GenBank with Accession Number OK090549.1. The obtained ITS region sequence was more than 99% identical to those of A. besseyi reported from China (MW216945.1) and India (JF826518.1, JF826519.1 and JF826517.1). These ITS sequence results further supported that the isolated nematodes were A. besseyi. Subsequently, the species-specific primers of A. besseyi (BSF, 5′-TCGATGAAGAACGCAGTGAATT-3′ and BSR, 5′-AGATCAAAAGCCAATCGAATCAT-3′) were used for confirmation by PCR (Cui et al. 2010). An expected PCR fragment of 312 bp was obtained, which was consistent with those of A. besseyi reported previously. The pathogenicity of identified A. besseyi was confirmed by infection of foxtail millet (Setaria italica cv. ‘Yugu33’). Foxtail millet budding seeds were sown in the pots contained 150 mL of sterile soil mixture. In two weeks, 10 seedlings were inoculated with 100 A. besseyi each, and 4 plants were non-inoculated as the control. The foxtail millet seedlings were grown in a plant-growth chamber at 25/30°C under 12 h dark/12 h light. On the average, 73.3 and 138.2 of A. besseyi were isolated from each plant at 15 and 40 days post inoculation, respectively. Both the morphological and molecular characteristics were identical with those nematodes obtained from the original samples. All the upper leaves of the inoculated plants showed chlorosis and necrosis, symptoms that were similar to those observed in the field, and neither symptom developed on the non-inoculated control plants, nor were nematodes re-isolated from the control plants. To the best of our knowledge, this is the first record of A. besseyi on foxtail millet in Henan Province of North China. Henan is one of the most important grain-producing areas in China, and A. besseyi is an important domestic quarantine nematode, which may become a severe threat to cereal production in Henan Province. Our findings will be very beneficial for A. besseyi management and further research on foxtail millet in Henan Province of North China.

Seed borne nematode of small millets: A review

“…It can survive in stored seeds for several years under dry conditions. A. besseyi was reported first time on foxtail millet (Setaria italica) from Japan (Yoshi and Yamamota, 1950) [27] , India (Dave et al, 1979, Lal andMathur, 1988) [6,12] and China (Cui et al, 1989, Chen et al, 2022) [5,2] . Lal and Rajan (1995) [13] collected 1138 accessions of foxtail millet from diverse geographical regions of India and examined for seed borne nematodes by soaking the seeds in water for 24-48 h at 30 0 C. About 8% (91) accessions were found infected with A. besseyi with inoculum level varying from 2 to 36 per seed.…”

“…Foxtail millet is a major cereal crop in Northern China especially in the semi-arid areas and is an important domestic quarantine nematode in Henan province of China. The average number of nematode per earhead of foxtail millet was up to 1738.75±107.72 (Chen et al, 2022) [2] . The disease caused by A. besseyi in foxtail millet was referred as Ear blight of italian/foxtail millet (Yoshi and Yamamota, 1950) [27] and Light ear disease of foxtail millet (Dave et al, 1979, Shukla et al, 1997) [6,23] .…”

“…Sometimes completely sterile glumes were also observed. Chen et al (2022) [2] reported that some upper leaves showed chlorosis without or with necrotic tips. Flag leaves presented crinkling and distortion, coloured stalks, vertical ear heads, open brown to light black glumes and thin grains due to infestation of A. besseyi in foxtail millet.…”

“…Larger body length, greatest body width, oesophageal length, tail length of female and male, vulva of female, spear and spicules of male were greater in rice isolates than the foxtail millet and proso millet isolates. Chen et al (2022) [2] characterized the females isolated from foxtail millet were slender with a short stylet, an oval metacorpus with a distinct valve, a labial region slightly wider than the first body annules and a conoid tail with a terminus bearing a star shaped mucro with four pointed processes. The males had three pairs of ventro-submedian papillae with the first one adanal, spicula curved with a slight basal process, terminus bearing four mucrons arranged variably and the whole worm was in 'J' shape.…”