Soil texture, infective juvenile concentration, and soil organic matter influence the efficacy of Steinernema feltiae isolate Lican Ray

IJs of S. feltiae LR were cultured at 25 ± 0.1°C using last larval instars of G. mellonella (Kaya and Stock, 1997), and IJs were stored at 8°C in tissue culture flasks with distilled water for less than five days before their use.

Three different soils were used in the experiments. Sandy loam and clay soils were obtained from Pichidegua town, located in the Libertador Bernardo O´Higgins Region, Chile. Loam soil was obtained from Universidad de Chile’s experimental field located at La Pintana, in the Metropolitan Region, Santiago, Chile.

The clay soil was sieved, and all soils were sterilized in metallic cans using the water bath method at 90°C for 2 hr, and then physical and chemical parameters of the soil samples were determined in the Laboratory of Soil from Universidad de Chile (Table 1). The samples were moistened up to field capacity and stored in closed plastic containers at 25 ± 2°C for up to 48 hr before use.

OM in the form of Vermicompost was obtained from a vermiculture farm, which used rests of vegetables from the kitchen of a cafeteria. Three types of vermicompost were used according to their age and degree of decomposition: initial (less than one month), medium (three months) and advanced (six months). All the samples were sterilized, as described previously, and prior to use, the vermicompost was sieved (3 mm opening for initial; 2 mm for others), chemical parameters determined as above (Table 2), and stored at −80°C to stop the decomposition process. On the day of the assay, it was moistened up to field capacity.

Bioassays were conducted using a 75 mm diameter and 25 cm long t-shaped PVC tube, denominated matrix (Fig. 1A), filled with soil. Through the lateral opening (Fig. 1B), a modified 60 mm diameter plastic petri dish was introduced. This dish had both sides replaced by 1 mm stainless steel mesh, and contained soil corresponding to the treatment, and five last larval instars of G. mellonella. Removable lids closed the end matrix openings, and the lateral lid was modified with a metallic mesh for gas exchange (Fig. 1B). According to the treatments, matrices were filled with different soil type, previously sterilized and moistened.

Figure 1:

The S. feltiae isolate LR evaluated in this work displayed the expected dispersal behavior in the soil, according to S. feltiae foraging strategy (Alekseev et al., 2006; Campos-Herrera and Gutiérrez, 2014). The geographic diversity of Chile (from 18 °S latitude to 56 °S latitude) results in a myriad of soils under different management strategies, and biotic and abiotic characteristics. In this work, we verified the potential of this native isolate to be used under different conditions for the control of soils pests.

Although IJ concentration for the control of different pests have been widely studied (Gazit et al., 2000; Batista et al., 2014; Guo et al., 2017), the influence of soil texture at different IJ concentrations on pest control is poorly understood. For instance, Koppenhöfer and Fuzy (2006) found that the highest concentration of Steinernema scarabaei caused the highest mortality levels on Anomala orientalis (Coleoptera: Scarabeidae) in all substrates studied (loamy sand, sandy loam, loam, silt loam, acidic sand and potting mix). In contrast, in a study by Campos-Herrera and Gutiérrez (2009), the infectivity of S. feltiae isolate the Rioja against S. littoralis was affected by both soil type and IJ dosage, with higher mortality achieved with higher IJ doses in heavier soils. However, in these soils, the number of days to larval death was highest. Similarly, we observed that IJ dosage and soil texture affect IJ movement and its capacity to reach and kill the insect host. In our study, initially, the highest levels of mortality were achieved in loam soil, regardless of the IJ concentration, achieving the highest levels of mortality at six DAI. However, as time passed, both texture and IJ concentration influenced larval mortality, particularly in light-textured soil (sandy loam), where there was a clear tendency to higher mortality as the concentration increased.

These results suggest that in field conditions, higher IJ concentrations of S. feltiae should be utilized in applications for light soils. In heavy soils (clay), even though no significant differences were observed, the highest mortality was achieved with the highest IJ concentration, but it is likely that the percentage of clay in the soil will define the feasibility for EPN to disperse. According to Wallace (1968), the nematodes are confined to the existing soil geometry, and pore size correlates with movement; nematodes cannot pass through spaces much narrower than their body diameter. Therefore, the isolate selected should consider its body size and soil clay content.

Several studies have shown that soil texture affects the capacity of EPN to disperse and infect the host, such as Steinernema riobrave and H. bacteriophora on Diaprepes abbreviatus (Coleoptera: Curculionidae) (Shapiro et al., 2000), H. bacteriophora, H. megidis and S. feltiae on Diabrotica virgifera virgifera (Coleoptera: Chrysomelidae) (Toepfer et al., 2010), and S. carpocapsae and H. bacteriophora on potato tuber moth, P. operculella (Hassani-Kakhki et al., 2013). In some cases, light textures were the most beneficial for various EPNs species (Hassani-Kakhki et al., 2013).

However, in our study, the native EPN, S. feltiae LR, was capable to disperse more in intermediate (loam), both vertically and horizontally. This isolate moved through the soil and reached the host at a distance of 30 cm horizontally in all soil types evaluated, except in loam, where they were able to move up to 40 cm. Furthermore, they achieved vertical distances of 70, 50 and 30 cm in loam, sandy loam and clay soil, respectively. Other studies found similar results with vertical movement of S. feltiae on the host G. mellonella (Neumann and Shields, 2006), also of S. carpocapsae on Anastrepha obliqua (Diptera: Tephritidae) (Toledo et al., 2009), and vertical and horizontal of Heterorhabdits amazonensis and Steinernema arenarium on Spodoptera frugiperda (Lepidopera: Noctuidae) (Andaló et al., 2012). In addition, in our study, mortality of G. mellonella decreased with the extreme soil textures evaluated. Horizontally at 30 cm, while in loam soil mortality reached 100% at the end of the experimental period, in sandy and clay soil it reached 80 and 45%, respectively. Similarly, at 40 cm, while in loam soil mortality reached 50%, in sandy and clay soil it reached only 20 and 0%, respectively.

However, the results have been diverse regarding which soil texture benefits EPNs activity, as the soil pore size can also affect IJ activity in the soil, because energy reserves are depleted with more movement. For example, smaller pores result in higher opposition to IJ movement and/or less oxygen available and consequently, IJs may lose efficacy finding, reaching and infecting their hosts (Wallace, 1968). Native isolates, even from the same species, may show better performance in specific soil textures. For example, in a study by Toepfer et al. (2010), the efficacy of three EPNs species, including S. feltiae, on D. v. virgifera larvae was higher in maize fields with heavy clay or silty clay soils than in sandy soils. In addition, S. carpocapsae and S. riobrave moved more easily in sandy loam and marine sand soils (Alekseev et al., 2006), and S. riobrave and H. bacteriophora dispersed more easily in sandy soils (Shapiro et al., 2000). Our study showed greater infective capacity efficacy of IJ in soils with medium pore size, however, other species, such as S. riobrave and H. bacteriophora, showed greater infection capacity on D. abbreviatus and longer persistence in Marl soil (80% silt and 15% clay) compared to sandy soil (Shapiro et al., 2000). Likewise, S. carpocapsae showed the highest percentage of infection on A. obliqua larvae in sand-clay mixture (60–82% depending on depth), compared to 45–64% in sand, and 23–30% in loamy-sand soil (Toledo et al., 2009).

Therefore, light soils (containing more sand and less clay) do not necessarily guarantee IJs efficacy, because there may be a strong relationship between soil pores and IJ size, which can benefit IJ dispersal. The dispersal rate in our study was higher in loam soil compared to the other textures, where IJs of S. feltiae LR reached G. mellonella (a non-mobile host), located 30 and 50 cm horizontally and vertically, respectively, in only three days. Similar studies were conducted by Bal and Grewal (2015) where S. carpocapsae and H. bacteriophora, moved horizontally approximately 15 and 10 cm in the same period, reaching a non-mobile host and 24 and 15 cm with a mobile host, respectively, in silt-loamy soil. However, further studies by Bal et al. (2017) showed that the same species moved an estimate of 150 cm in 3 days, under field conditions. Therefore, it is crucial to define the adequate EPN isolate for each soil type, which optimizes dispersal and time to reach the host, factors that should be considered together with IJ concentration in further field studies.

Although soil structure may be improved with the addition of OM, in this work, structure was not considered, because it was altered in the process of mixing the soil with OM and filling the tubes. Only the content of OM and the decomposition degree were considered. While EPN movement in organic substrate has received little attention, Griffin (2015), Ansari and Butt (2011) and Kapranas et al. (2017) point out that EPN behavior is influenced by the species and soil quality. For instance, both compaction and increasing peat content in sand, decreased IJ success in S. carpocapsae, Heterorhabditis downesi and S. feltiae (Kapranas et al., 2017). Also, according to Williams et al. (2013), it is likely that organic soils facilitate the active or passive movement of nematodes through the medium or promote their post-application survival. For example, Kruitbos et al. (2010) observed that while S. carpocapsae moved significantly less in sand than in peat, H. megidis dispersed well in both media, but only showed taxis toward hosts in sand. In addition, MacMillan et al. (2009) observed a more active dispersal of two Phasmarhabditis hermaphrodita strains in organic media (bark chips and leaf litter, and to a lesser extent peat) than in mineral soils. Similarly, our study evidenced, first, that the OM content influences IJ movement of S. feltiae LR, especially those treatments with OM content different from zero. Even though no significant differences in G. mellonella mortality were observed in the treatments that contained OM (2–8%), our results suggest that 4% would be the optimal content for this EPN and values above or below this, affect its capacity to reach and kill the host. Second, we observed that the OM decomposition degree (particle size), does not influence the capacity of S. feltiae LR to reach and kill the host. Regarding survival of the EPN, Herren et al. (2018) observed higher survival of S. feltiae in mature compost that in soil with no compost added; however, this was also affected by the application mode of the EPN. In their work, the survival ranged from only 3.2%, when EPNs were applied in aqueous suspension to soils with no compost added, to 62% when they were applied in clay-coated G. mellonella cadavers on soil with mature compost added.

Even though S. feltiae is a well-studied species, and several companies around the world have used it to formulate products commercially available, EPN intra-species variability concerning phenotypical and biological behavior has been demonstrated (Gaugler et al., 1989; Campos-Herrera and Gutiérrez, 2014). Therefore, when developing a product based on native EPN, it is important to study specific characteristics of these ecotypes to optimize its use under local conditions. Further studies under field conditions, using a wider range of soil OM content and including those textures that affected negatively the movement of S. feltiae LR (extreme textures), which could be improved adding OM, should be also considered.