First Report of the Stubby Root Nematode Paratrichodorus allius on Sugar Beet in Minnesota

Stubby root nematodes (Paratrichodorus and Trichodorus) are migratory ectoparasites that feed on roots, transmit tobraviruses, and cause significant crop loss. In June 2015, three soil samples from a sugar beet field near Felton (Clay County), MN were submitted to the Nematology Laboratory at North Dakota State University for nematode assay. The soil texture was sandy, the current sugar beet cultivar was BTS 8337, and the field was previously planted with wheat in 2014. Nematodes were extracted from 100 cm3 soil using the sugar centrifugal flotation method. Plant-parasitic nematodes were identified and counted based on morphological features to genus. One of the samples was found to contain stubby root nematodes (60 per 100 cm3 soil) with several other nematodes. In August and September 2015, six soil samples were collected from the same field; five of them from the area with small and stunted plants, and one from the area with healthy plants. Nematodes were extracted, revealing all the five samples contained stubby root nematodes ranging from 40 to 200 (average 95) per 100 cm3 soil and the one from healthy plants had no stubby root nematodes. Other nematode genera recovered from these samples included Pratylenchus, Paratylenchus, Helicotylenchus, and Tylenchorhynchus. Individual stubby root nematodes were hand-picked and examined morphologically and molecularly for species identification. The specimens were identified as Paratrichodorus allius according to morphological and morphometric characteristics. Morphological measurements of adult females (n = 7) included body length (range = 532.0 to 785.0 μm, mean = 695.6 μm), onchiostyle (42.0 to 45.0, 43.6), body width (40.0 to 52.0, 45.4), anterior end to basal bulb (115.0 to 155.0, 130.0), a (12.6 to 17.4, 15.4), b (4.1 to 6.0, 5.2), and V (48.4 to 58.4%, 53.0%). The anus and caudal pores are subterminal. DNA was extracted from a single nematode (n = 6) in 20 µl of extraction buffer (1×PCR buffer, 60 µg/ml Proteinase K). The D2/D3 region of 28S rRNA, two segments of 18S rRNA, and ITS1 rDNA were amplified with primer pairs D2A/D3B, SSUF07/SSUR26, 18S965+18S18P, and BL18/5818, respectively. PCR products were purified and sequenced, and the sequences were deposited in GenBank. The two segments of 18S rRNA sequences (GenBank Accession No. KT892733, 846bp, and KT892734, 751 bp) were 100% identical with one population of P. allius (AJ439572) from Washington, and had 99% or less than 99% similarity with other