Effect of soil tillage and crop rotation on the adult emergence of P. leporinus
Field trials were set up in different regions of Southwest Germany (Kirchardt, Neckarsulm, Ilvesheim, Dettenheim, Ladenburg, and Untereisesheim) and in the valley of the river Elbe (Arzberg) in autumn 2018 and 2019, following the sugar beet harvest. Trial sites were selected according to the following criteria: i) high presence of P. leporinus nymphs in the soil and strong and uniform SBR symptoms; and ii) uniform field topography. The adult emergence of P. leporinus was analysed after different soil tillage treatments (ploughing at depths of at least 20 cm and a cultivator at depths up to 20 cm) and the subsequent crops cultivated after the sugar beet harvest (barley or spring wheat in comparison to winter wheat in 2019 as well as maize (Zea mays) and bare soil in comparison to winter wheat in 2020) (Table 1). In Kirchardt 2019, winter durum (Triticum durum) was cultivated instead of winter wheat. Winter wheat after soil tillage with a cultivator was considered as the control treatment because it is the most common combination after sugar beet harvests in these regions. In our study, reduced tillage was carried out with a rigid tine cultivator instead of a disc harrow, as in the study by Bressan (2009), because this was more common in the regions analysed. In Ladenburg 2019 and Untereisesheim 2019, winter wheat was compared to spring barley; both after reduced tillage. However, in Ladenburg 2019, using a cultivator for soil tillage this was impossible due to the dry soil conditions, so a disc harrow was used instead. Agronomical measurements (time of soil tillage, tilling depths) and plant protection methods (use of herbicides and fungicides) were performed according to good agronomical practice in the wheat, barley and maize plots. Emerging weeds in bare soil treatment plots were controlled with the herbicides Adengo (Bayer CropScience Deutschland GmbH, Monheim, Germany), Glyphosate or with a combination of Laudis (Bayer CropScience Deutschland GmbH), Spectrum and Bo 235 (both BASF SE, Ludwigshafen, Germany) when necessary. Detailed information is given in Tables 1 and 2.
Table 1
Details of experimental field sites
Field trial site
|
Coordinates
|
Sugar beet harvest
|
Depths and dates of soil tillage
|
2018/19
|
Mouldboard plough
|
Rigid tine cultivator
|
Untereisesheim
|
49°12'54.1"N 9°11'27.0"E
|
18/9/18
|
|
20 cm, 15/10/18
|
Ladenburg
|
49°27'11.5"N 8°37'55.1"E
|
17/10/18
|
|
*10 cm, 17/10/18
|
Arzberg
|
51°30'31.3"N 13°09'46.0"E
|
2/11/18
|
22 cm, 8/11/18
|
18 cm, 8/11/18
|
Dettenheim
|
49°10'54.9"N 8°23'22.5"E
|
13/10/18
|
25 cm, 19/10/18
|
15 cm, 19/10/18
|
Kirchardt
|
49°12'33.0"N 8°58'20.1"E
|
1/10/18
|
28–30 cm, 7/10/18
|
10 cm, 5/10/18
|
2019/20
|
|
|
|
Arzberg
|
51°31'07.6"N 13°07'24.9"E
|
27/9/19
|
22 cm, 26/10/19
|
20 cm, 4/10/19
|
Neckarsulm
|
49°12'19.3"N 9°10'35.1"E
|
25/10/19
|
25 cm, 30/10/19
|
15–20 cm, 30/10/19
|
Ilvesheim
|
49°27'38.3"N 8°34'33.7"E
|
24/10/19
|
30 cm, 25/10/19
|
15 cm, 25/10/19
|
Dettenheim
|
49°10'51.3"N 8°23'31.1"E
|
25/10/19
|
20 cm, 26/10/19
|
10–12 cm, 26/10/19
|
*Disc harrow was used instead of the rigid tine cultivator in the Ladenburg testing site |
Table 2
Dates and depths of seedbed preparation in the different field trial sites. Seedbed preparation was carried out with a rotary harrow at each testing site
Field trial site
|
Depths and dates of seedbed preparation
Depths and dates of sowing
|
2018/19
|
Winter wheat
|
Spring wheat
|
Barley
|
Untereisesheim
|
4–5 cm, 16/10/18 2–3 cm, 16/10/18
|
|
4–5 cm, 28/2/19
2–3 cm, 28/2/19
|
Ladenburg
|
3–5 cm, 2/11/18
5 cm, 2/11/18
|
|
3–5 cm, 23/2/19
5 cm, 23/2/19
|
Arzberg
|
4 cm, 8/11/18
2 cm, 8/11/18
|
4 cm, 4/3/19
2 cm, 4/3/19
|
|
Dettenheim
|
3–5 cm, 4/11/18
2 cm, 4/11/18
|
8 cm, 26/2/19
2 cm, 26/2/19
|
|
Kirchardt
|
5 cm, 8/10/18
2–3 cm, 8/10/18
|
5 cm, 13/3/19
2–3 cm, 13/3/19
|
|
2019/20
|
Winter wheat
|
Maize
|
Bare soil
|
Arzberg
|
4 cm, 26/10/19
2 cm, 26/10/19
|
8 cm, 14/4/20
8 cm, 15/4/20
|
|
Neckarsulm
|
4–5 cm, 31/10/19
3 cm, 31/10/19
|
10 cm, 11/4/20
8 cm, 14/4/20
|
8 cm, 11/4/20
|
Ilvesheim
|
8 cm, 26/10/19
3 cm, 26/10/19
|
8 cm: 1/4/20, 8/4/20
8 cm, 11/4/20
|
8 cm: 1/4/20, 8/4/20
|
Dettenheim
|
3–5 cm, 27/10/19
2 cm, 27/10/19
|
8–10 cm, 9/4/20
8 cm, 11/4/20
|
-
|
The field trial layout used a completely randomized block design. A plan of the layout is provided in Fig. S1. In 2019 the trials were designed with three blocks. This was increased to four blocks in 2020, due to the nonhomogeneous nature of P. leporinus adult emergence observed in 2019. Plot width was 6 m except at the sites Untereisesheim 2019 (3 m), Ladenburg 2019 (3 m) and Arzberg 2020 (12 m).
Quantification of field-emerging P. leporinus adults
Three cages (base area approx. 3.6 m² with a 1.5 m maximum height, Fig. S2) were set up in each plot to catch P. leporinus adults emerging from the soil. The cages were placed 10 m apart. An additional cage per plot was erected in Arzberg in 2020. All cages were covered with a 1.35 mm mesh gauze (FA.BIO 01 Kulturschutznetz Rettichnetz, Hartmann-Brockhaus, Pfaffenhofen-Wagenhofen, Germany) with gauze ends buried in the ground. Inside each cage, a 10 x 25 cm yellow sticky trap (Gelbe Insekten-Leimtafeln, Aeroxon Insect Control GmbH, Waiblingen, Germany) was attached at the top to catch emerging adults. Each sticky trap was replaced once per week. Counting of adult P. leporinus started on 13/5/19 in Arzberg and on 14/5/19 in all other field trial sites in 2019, and on 29/4/20 in Arzberg and on 22/4/20 in all other field trial sites in 2020. On 29/6/20 in Neckarsulm, Ilvesheim and Dettenheim and on 30/6/20 in Arzberg, maize plants were cut to a maximum height of 1 m, to avoid that the plants reached the top of the cages and to ensure that emerging adult P. leporinus were still attracted by the sticky traps while the maize plants stayed alive.
P. leporinus identification
All trapped planthopper adults were identified to genus level according to their scutellum, vertex, pronotum and hind tarsus, using a stereomicroscope. From each field trial site, a representative sample of 25 male Pentastiridius sp. were then identified to species level according to their genital morphology. The key of Biedermann and Niedringhaus (2004) was used. All analysed insects in the genus Pentastiridius sp. were identified as P. leporinus, so, therefore, we counted all Pentastiridius sp. as P. leporinus.
Performance of nymphs on different host plants
The survival and development of P. leporinus nymphs on different plant species was analysed under controlled environmental conditions (20.9 ± 1°C, 45 ± 11.6% relative humidity, 24 h darkness). Plants at the seedling stage (4–10 days old) were then offered to nymphs, as this is the stage under field conditions when they have their first contact. The parental adult generation was caught between 19/6 and 20/6/2020 in a sugar beet field (49°12'17.2"N 9°10'48.8"E). Oviposition was performed on sugar beet plants under controlled environmental conditions and egg batches were kept on Petri dishes until the nymphs hatched. Directly after hatching, the first instar nymphs were individually transferred with a small paint brush to wheat (cv. Dekan, KWS SAAT SE & Co. KGaA, Einbeck, Germany), and barley (cv. Orbit, KWS SAAT SE & Co. KGaA) or maize (cv. Ronaldinio, KWS SAAT SE & Co. KGaA) plants. Seeds were not treated with insecticides or fungicides. Plants were grown in conical 600 mL plastic pots (11 cm upper diameter; 9 cm height) that were filled with sand (diameter 0–2 mm) under controlled environmental conditions (as described above, with a 16:8 h light/dark photoperiod and 80 µmoL (s m²)−1 light intensity). Light was provided by full-spectrum LED lights (‘Bioledex GoLeaf E2 LED Pflanzenleuchte Vollspektrum 120cm 50W IP44’, DEL-KO GmbH, Germany). After 4–10 days, the seedlings were carefully removed from the sand and cleaned with tap water. A total of 24 nymphs were analysed per plant species (four nymphs each from six different parental females) to ensure genetic variation of specimens.
The experiment was carried out in 11 x 8 x 5 cm plastic containers with 25 g of substrate [(3:1 parts mixture of Fruhstorfer Erde Typ P 25 (HAWITA Gruppe GmbH, Vechta, Germany) and sand (diameter 0–2 mm)]. All boxes were closed with lids, which had tiny holes inserted to allow gas exchange. Seedling roots were placed on top of the substrate to allow the nymphs direct feeding access. Boxes received two seedlings of the wheat and barley treatments and one seedling of the maize treatment. Each box was inoculated with a single nymph. Seedlings were replaced when the first signs of deterioration were observed or after a maximum of eight days. The substrate was kept moist and was replaced after four weeks. The experiment was carried out under controlled environmental conditions. During the first 160 days of the trial, the survival and development of the nymphs were evaluated every second day using a stereomicroscope. Due to slow development and low mortality in the higher nymphal instars, the evaluation was carried out twice per week, starting after 160 days. Nymphal development was evaluated by observation of moulting and measurement of head capsule width according to Pfitzer et al. (2022).
Statistical analysis
Statistical analysis was carried out using SAS 9.4 (SAS Institute Inc., Cary, USA). The total number of fieldemerged P. leporinus adults per m² was analysed. The mean values of the emerged adults were calculated from three or four cages within each plot. Those means were then used for a two-way analysis of variance (ANOVA) and tested for their homogeneities of variance and normal distribution. Least square means were determined with LSMEANS in the PROC GLIMMIX procedure separately for each testing site to analyse for significant differences at the p < 0.05 significance level. The assumption of the homogeneity of the variances was not fulfilled in the field trials, where the role of soil tillage and cultivation of winter wheat was compared to spring wheat or to maize and bare soil. To fulfil the assumption of homogeneity of variances, the data were square root transformed to analyse for significant differences. The data were then back transformed to calculate the estimated mean values and standard errors in each treatment and for each testing site. For the results of the field trials, where the role of cultivation of winter wheat compared to barley was tested, the values were not transformed as the assumptions of their homogeneities of variance and normal distribution were fulfilled (SAS Institute Inc. 2022).
Cox proportional hazard models and PROC PHREG were used in SAS 9.4 for survival analysis of the no-choice oviposition experiment. Hazard ratios (HR) > 1 or < 1 describe the higher or lower probabilities of mortality compared to the referred treatment. To obtain a letter code in this experiment, survival was also analysed using PROC LIFEREG in SAS 9.4. The Weibull, exponential, and logistic distributions of the data were tested. The Akaike information criterion (AIC) was lowest for the Weibull distribution and assumed for this analysis. Least square means were determined with LSMEANS, to identify significant differences between the treatments at p < 0.05 (SAS Institute Inc. 2022).