Belonolaimus longicaudatus Host Status and Pathogenicity on Sweetpotato

These objectives were investigated in repeated greenhouse experiments conducted in Gainesville, FL. The experiment was arranged in a randomized complete block design with five replicates and two fully crossed factors: crop and sting nematode inoculation rate. Four host crops were used: (1) field corn, (2) sunn hemp, sweetpotato cultivars (3) ‘Beauregard’, and (4) ‘Covington’. For field corn, the cultivar NK 1573_3330 (Syngenta Crop Protection AG, Basel, Switzerland) was used. For sunn hemp, the cultivar Crescent Sun (Tropical Seeds, LLC, Miami, FL) was used. Field corn was selected as a known sting nematode host (Rhoades, 1978; Todd, 1989), and sunn hemp as a reported poor host (Braz et al., 2016). As previously described, ‘Covington’ and ‘Beauregard’ were selected for their varying susceptibilities to southern root-knot nematode. Sting nematode inoculation rates were uninoculated, medium, and high. In the first trial (Trial 1), the medium and high inoculation rates were 26 and 157 sting nematodes (mixture of males, females, and juveniles) per pot, respectively. In the second trial (Trial 2), the medium and high inoculation rates were 40 and 240 sting nematodes per pot, respectively. The primary purpose of varying the inoculation rates was to assess sting nematode damage to crops. The secondary purpose was to help ensure a representative assessment of the host status of the crops because if the crop growth is severely impaired from high sting nematode inoculation rates, it could result in misleading low final sting nematode populations.

Trials were established in a greenhouse at the University of Florida in Gainesville, FL, in summer 2021. Crops were planted on 28 May (Trial 1) and 1 June (Trial 2) into 15-cm-diameter clay pots filled with 1,000 cm³ autoclaved field soil. The field soil was a Chipley–Foxworth–Albany complex (91% sand, 6.8% silt, and 2.4% clay with 1.7% organic matter) obtained from the University of Florida North Florida Research and Education Center near Live Oak, FL. Field corn and sunn hemp were planted with three seeds and thinned to one seed per pot 7 days after planting (DAP). Sweetpotato was planted with one slip per pot. Slips are cuttings of plant stems grown from seed tubers and are standard commercial planting materials. Sweetpotato slips were obtained from the North Carolina State University breeding program fields, transported in cardboard boxes with adequate moisture, and stored under moist conditions until transplanting. The trial was established, and the slips were transplanted 1 day after arrival in Trial 1 and 4 days after arrival in Trial 2.

Sting nematodes were inoculated into pots at the previously indicated rates at 12 and 16 DAP in Trials 1 and 2, respectively. For inoculation, three holes of 2 cm depth were formed around each plant, and sting nematode inoculum was pipetted equally into each hole in a total solution of 12 mL per pot. Sting nematode inoculum was obtained from a culture maintained on St. Augustine grass ‘FX-313’ (Stenotaphrum secundatum (Walt.) Kuntze) and originally collected from ‘Tifway’ bermudagrass (Cynodon dactylon (L.) Pers.) in Sun City, FL. Sting nematodes for inoculation were extracted from the culture using the decanting and sieving method using a 400-mesh standard sieve (Cobb, 1918), followed by a modified Baermann method (McSorley and Frederick, 1991), with nematodes collected using a 500-mesh standard sieve, which was selected to maximize sting nematode viability. Nematode inoculum was quantified using a Zeiss (New York, USA) Primovert light microscope, stored at 5°C until inoculation, and was inoculated within 2 days of extraction. The plants were maintained for 70 days and 68 days after inoculation (DAI) in Trials 1 and 2, respectively, before the trials were terminated. The plants were watered daily by hand and maintained without supplemental light.

In each trial at 30 DAI, the plant height and number of leaves per plant were assessed. The plant height, leaves per plant, fresh shoot weight, and fresh root weight were assessed at trial termination. Soil nematode populations in each pot were quantified at the termination of each trial. To quantify soil sting nematode abundances, the soil was mixed manually, and nematodes were extracted from 100 cm³ soil using the sucrose centrifugation method (Jenkins, 1964). Sting nematodes were quantified morphologically by microscopy. In addition to soil abundances, at medium and high inoculation rates, reproductive factor was calculated. First, total sting nematode abundance per pot was calculated by multiplying the abundance in 100 cm³ soil by 10 since there was 1,000 cm³ soil in each pot. Then reproductive factor was calculated by dividing final soil nematode population per pot by the initial inoculum per pot. Typically, a crop that supports a reproductive factor >1 is considered a host, whereas a crop that supports a reproductive factor <1 is considered a poor or non-host (Timper and Hanna, 2005).

All variables were analyzed separately for each trial because there were significant (ANOVA, P < 0.05) treatment-by-trial interactions in a preliminary analysis of sting nematode abundances. For each trial, sting nematode abundance and reproductive factor were analyzed by two-way ANOVA, with crop and inoculation rate as factors. If there were significant crop–inoculation rate interactions, crop effects on sting nematode parameters were analyzed separately with each inoculation rate. If crop effects were significant (P < 0.05), crop treatments were separated by Fisher’s protected LSD (a = 0.05). For crop growth parameters (number leaves, plant height, shoot weight, and root weight), the inoculation rate effect was analyzed for each crop using one-way ANOVA since direct growth comparisons among crops were not of scientific interest for this study. If inoculation rate effects on crop growth parameters were significant (P < 0.05), inoculation rate treatments were separated by Fisher’s protected LSD (a = 0.05). Before completing ANOVA, response variables were transformed, if needed, to meet assumptions of homogeneity of variance using Levene’s test (Levene, 1960) and normality of residuals based on graphing (Cook and Weisburg, 1999). In Trial 1, soil sting nematode abundances were transformed by square root to meet model assumptions for the full two-way ANOVA model only. For all other variables and models, transformation was not necessary. Analyses were conducted in R statistical software (version 3.4.4, The R Foundation for Statistical Computing, Vienna, Austria).

For both trials, there were significant crop– inoculation rate effects on sting nematode parameters (data not shown), so the parameters were analyzed separately by inoculation rate. For both trials, soil sting nematode abundances were significantly affected by crop at the medium and high inoculation rates, but no nematodes were detected in the uninoculated treatment (Fig. 1). At both medium and high inoculation rates, soil sting nematode abundances were generally significantly greater for corn than for any other crop in both trials (Fig. 1). The only exception was that in Trial 1, at the medium inoculation rate, soil sting nematode abundances for ‘Covington’ were not significantly different from those for corn or the other crops. Sting nematode reproductive factor followed the same trends as soil sting nematode abundances (Fig. 2). In both trials and inoculation rates, sting nematode reproductive factor was >1 for corn, indicating a susceptible host, although values were numerically greater in Trial 2 than in Trial 1 (Fig. 2). Sunn hemp and ‘Beauregard’ had reproductive factors <1 in each trial, as well as each inoculation rate. ‘Covington’ had a reproductive factor <1, except in Trial 2 at the medium inoculation rate, when reproductive factor was at 1 (Fig. 2). Across both trials and inoculation rates, sting nematode reproductive factors were 2.5, 0.6, 0.2, and 0.2 for field corn, ‘Covington’, ‘Beauregard’, and sunn hemp, respectively.

Figure 1

Figure 2

Sting nematode had relatively little impact on crop growth. Sting nematode inoculation rate did not significantly affect ‘Beauregard’ or ‘Covington’ plant height at any time (Table 1). At 30 DAI in Trial 2, the corn plant height was greater at the medium rate than at the uninoculated or high inoculation rate but was not affected at any other time (Table 1). In Trial 1, the sunn hemp plant height was significantly greater at the medium rate than at the uninoculated or high rate at both 30 DAI and termination but was not significantly affected in Trial 2. Sting nematode inoculation rate did not have a consistent effect on leaves per plant at 30 or termination DAI in either trial (Table 2). In Trial 2 at 30 DAI, there were significantly more corn leaves per plant at the medium rate than at the uninoculated or high inoculation rate. In Trial 1 at 30 DAI, there were significantly more ‘Beauregard’ leaves per plant at the high than at the uninoculated rate. There were no other significant effects on corn or ‘Beauregard’ leaf counts and no effects on ‘Covington’ or sunn hemp leaf counts.

There were no significant inoculation rate effects on corn, sunn hemp, or ‘Covington’ final shoot weight in either trial (Fig. 3). For ‘Beauregard’, the final shoot weight was significantly greater at the high rate than at the medium or uninoculated rate in Trial 1 but was not affected in Trial 2. There were no significant inoculation rate effects on the final root weight for any crop in either trial (Fig. 4).

Figure 3

Figure 4

Qualitatively, minor indications consistent with sting nematode damage (Grabau and Vann, 2021), including root stunting, pruned lateral roots, and infrequent root tip necrosis, were observed on some corn plants. No obvious indications of nematode damage were observed on sunn hemp or sweetpotato roots. No sweetpotato tubers had formed by the termination of either trial, so there was no observation of potential symptoms on tubers. No formal damage ratings were made because indications were minor and relatively infrequent.