Species Delimitation and Description of Mesocriconema nebraskense n. sp. (Nematoda: Criconematidae), a Morphologically Cryptic, Parthenogenetic Species from North American Grasslands

Abstract

Nematode surveys of North American grasslands conducted from 2010 to 2015 frequently recovered a species of criconematid nematode morphologically resembling Mesocriconema curvatum. These specimens were recovered from remnant native prairies in the central tallgrass ecoregion of North America, and not from surrounding agroecosystems. Historical records indicate that M. curvatum is a cosmopolitan species feeding on a wide range of agronomic and native plants. DNA barcoding indicates North American grasslands contain at least 10 phylogenetically distinct lineages of Mesocriconema that resemble, but are not, M. curvatum. Analysis of the two most common lineages reveals two distinctly different population structures. The variation in population structure suggests unique evolutionary histories associated with their diversification. These two major lineages share a sympatric distribution and their slight morphological differences contrast with a high level of genetic separation. Based on their genetic divergence, fixed diagnostic nucleotides, population structure, species delimitation metrics, and a sympatric distribution, we believe that one of these distinct lineages warrants formal nomenclatural recognition. Herein, we provide formal recognition for Mesocriconema nebraskense n. sp. and discuss its relationship to other Mesocriconema lineages discovered in native North American grasslands.

Description and Distribution of Three Criconematid Nematodes from Hangzhou, Zhejiang Province, China

“…Several sets of primers (synthesized by Invitrogen, Shanghai, China) were used in the PCR analyses to amplify the near full-length18S region of rDNA and COI region. Two sets of primers: the forward 18S39F (5′-AAA GAT TAA GCC ATG CAT G-3′) and the reverse 18S977R (5′-TTT ACG GTT AGA ACT AGG GCG G-3′), the forward 18S900F (5′-AAG ACG GAC TAC AGC GAA AG-3′) and the reverse 18S1713R (5′-TCA CCT ACA GCT ACC TTG TTA CG-3′) for amplification of the nearly full-length 18S rRNA (Olson et al, 2017). For the amplification of COI the primers used were COI-F5-(5′-AATWTWGGTGTTGGAACTTCTT -GAAC-3′) and COI-R9-(5′ CTTAAAACATAATGRAAAT-GWGCWACWACATAATAAGTATC-3) (Powers et al, 2014).…”

“…The 25-µl PCR was performed using 2x-TsingKe Master Mix DNA polymerase (Beijing TsingKe Biotech Co., Ltd) according to the manufacturer's protocol in a BIOER-XP thermocycler. The thermal cycler program for 18S and COI was as follows: denaturation at 95°C for 5 min, followed by 40 cycles (18S) or 50 cycles (COI) of denaturation at 94°C for 30 s, annealing at 50°C (18S) or 48°C (COI) for 30 s, and extension at 72°C for 90 s. A final extension was performed at 72°C for 5 min as described by Powers et al (2014) and Olson et al (2017). PCR products were separated and visualized on 1% agarose gels and stained with ethidium bromide.…”

DNA barcoding evidence for the North American presence of alfalfa cyst nematode, Heterodera medicaginis

“…Amplification conditions were as follows: denaturation at 94°C for 5 min, followed by 45 cycles of denaturation at 94°C for 30 s, annealing at 48.0°C or 50.0°C for 30 s, and extension at 72°C for 90 s with a 0.5°C per second ramp rate to 72°C. A final extension was performed at 72°C for 5 min as described by Powers et al (2014) and Olson et al (2017). PCR products were separated and visualized on 1% agarose using 0.5× TBE and stained with ethidium bromide.…”

Description of Gracilacus paralatescens n. sp. (Nematoda:Paratylenchinae) found from the rhizosphere of Bamboo in Zhejiang, China

“…Several sets of primers (synthesized by Invitrogen, Shanghai, China) were used in the PCR analyses to amplify the near-full-length 18S region, D2-D3 of 28S, and ITS region of rDNA. Primers for amplification of 18S were 18s39F-18s977R and 18s900-18s1713 (Olson et al, 2017). Primers for amplification of ITS were TW81-AB28 (Joyce et al, 1994).…”

“…The primers for amplification of D2-D3 of 28S were D2A and D3B (De Ley et al, 1999). PCR conditions were as described by Ye et al (2007) and Powers et al (2010). PCR products were evaluated on 1% agarose gels stained with ethidium bromide.…”