**1. Introduction**

Soybean (*Glycine max* L. Merril) is one of the oldest crops with high oil and protein content in the grain [1,2]. The protein and oil content depends on the variety and growing conditions and can vary between 35–50% protein and 18–24% soybean oil [3]. This oilseed is used in oil production, food production and animal nutrition. In the food industry, it is used in the form of soybean, oil, flour and milk, while grain, stalk or bread is used as livestock feed. However, the main reason for its cultivation is still livestock [3]. Besides its important role in human and livestock nutrition, it is also desirable in crop rotation. Through its symbiosis with nodule bacteria, it enriches the soil with nitrogen [4,5]. Soybean is a demanding crop that differs from other crops in complexity and cultivation requirements, especially in tillage and soil preparation for sowing. Basic ploughing is carried out to a depth of 30 cm, and in heavier soils levelling must be carried out in the autumn. In early spring, the soil must be closed as early as possible to retain all accumulated moisture [6].

Frequent and intensive tillage of any crop, including soybean cultivation, results in greater soil compaction or disruption of the continuity of larger pores and corridors of organisms in the soil. Such soil affects both the abundance and diversity of soil fauna as compaction creates unfavourable living conditions, especially anaerobic conditions [7,8]. One of the most important components of soil, apart from its chemical and physical

**Citation:** Lemic, D.; Pajaˇc Živkovi´c, I.; Posari´c, M.; Bažok, R. Influence of Pre-Sowing Operations on Soil-Dwelling Fauna in Soybean Cultivation. *Agriculture* **2021**, *11*, 474. https://doi.org/10.3390/ agriculture11060474

Academic Editor: Eric Blanchart

Received: 10 April 2021 Accepted: 18 May 2021 Published: 21 May 2021

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properties, is its biological component or soil organisms. The biological component or soil biodiversity is a very important but at the same time insufficiently known component of the soil ecosystem [9,10]. Biodiversity consists of soil organisms that spend all or part of their life cycle either in the soil or on its surface (including crop residues or mulch) and are responsible for processes that are very important for soil health and fertility [8]. Tillage is one of the most aggressive activities affecting soil biological balance. Biological balance refers to the interactions among organisms, including the structure of food webs and the ability of ecological systems to sustain themselves over time. In general, deeper and more frequent tillage increases negative impacts on soil organisms, while no-till, strip tillage and compatible tillage systems maintain biodiversity and soil organism richness in crop production. Improper and inappropriate tillage results in greater soil compaction or disruption of the continuity of larger soil pores as well as the corridors of soil organisms. This mainly affects the abundance, but also the diversity of the biological component of the soil [11], as greater soil compaction creates less favourable living and especially anaerobic conditions, which are only suitable for a smaller number of soil organisms [8].

The organisms in the soil are divided into three categories according to their influence on agriculture: Beneficial, Indifferent and Pests, and according to their size into four basic groups: Microfauna, Mesofauna, Macrofauna and Megafauna [12]. The abundance of beneficial organisms is extremely important as it is often used as an indicator to assess the viability of the agroecosystem. Higher numbers of beneficial soil organisms indicate better sustainability and positive impact on the crops grown [13]. Beneficial fauna has a positive impact on increasing soil fertility (decomposition and mineralisation of organic matter; mixing, transport and combination of organic and mineral soil components; transport of microorganisms...) and regulating the water–air ratio (creation and maintenance of soil pores) [14].

In the cultivation of soybeans, the occurrence of pests affects the quality and quantity of the grain. To prevent such damage, all available control measures are used, including chemical measures. Pesticides can be used in soybean production to control insects, mites, weeds and pathogens [15]. The use of pesticides has negative effects and destroys beneficial soil organisms [8,16,17]. Nietupski [18] states that of all pesticides used, only herbicides have negative effects on beneficial Carabidae.

The most numerous beneficial insects in soil fauna are species from the orders Collembola and Coleoptera, which are often referred to as bio-indicators [19]. These organisms have different feeding strategies and functional roles within soil processes. Collembola communities influence nutrient availability through their interactions with soil organisms [20], such as rates of bacterial and fungal consumption and spore transport. The relationships of soil collembolan fauna to their ecological niches and the stability of community composition at a given site provide good starting points for bioindication of changes in soil properties and impacts of human activities [21]. Carabids are often used as indicators of habitat change. They have been used in grasslands and boreal forests where species numbers and/or abundance have been found to change along a habitat disturbance gradient [22]. Their numbers are influenced by many factors, one of which is the pre-seeding procedure.

Glyphosate has been the subject of controversy for several years, ever since the World Health Organization (WHO) warned of possible carcinogenic and genotoxic effects on humans. Glyphosate is the active ingredient in many commercial herbicides, of which the best-known commercial product in the world is called Roundup, while in Croatia it is better known as Cidokor [23]. The use of glyphosate is extremely widespread in agriculture and horticulture [24]. Vandenberg et al. [25] noted that more than 1500 studies have been conducted on the safety of glyphosate in the last decade, potentially changing the regulatory view. More intensive research on the effects of glyphosate on beneficial (and harmful) soil fauna has not been conducted. Currently, there are no studies on the impact of pre-sowing intervention or glyphosate application on overall soil fauna and particularly on beneficial insects in soybean production.

Based on all the above, the hypothesis of this study is as follows: in soybean cultivation, more intensive tillage before sowing and glyphosate application have a negative impact on the whole soil fauna and especially on the members of the beneficial fauna. Based on the hypothesis, the objective of the study was to determine the total soybean soil fauna and the effects of different pre-sowing interventions on the abundance and composition of soil fauna in soybean cultivation.

#### **2. Materials and Methods**

#### *2.1. The Locality of the Experiment*

In 2019, a survey was conducted in six localities in the area of Šašinovec (45◦5100" N 16◦1001" E), a village near Zagreb in the central part of Croatia. Six soybean fields were sown in each of these six localities (36 soybean fields in total). In each field, different soil treatments were applied before sowing. Cover crops were sown in field 1. Field 2 was mulched, while field 3 was ploughed. Glyphosate was applied to field 4 for weed control. Field 5 was ploughed under, and field 6 had standard tillage (stubble ploughing at 10 cm, deep fall ploughing at 25 cm in 2018, and winter furrow closure and standard soil preparation for seeding in 2019). Mulching, cover plants and glyphosate applications do not involve soil activities, while ploughing, undermining and standard tillage represent interventions in shallower and/or deeper soil layers. More detailed data on tillage operations can be found in Table 1.


**Table 1.** Pre-sowing interventions and implementation dates.

\* Usual interventions of the field owner = conventional tillage in soybean cultivation.

#### *2.2. Soil Fauna Sampling*

Soil fauna sampling was conducted on three dates, April, June, and September, from the beginning to the end of soybean cultivation. Traps were active for two weeks in each specified sampling period. Soil fauna was collected using epigeic covered pitfall traps. Polythene pots (Ø = 12 cm, h = 18 cm) were incorporated 18 cm into the soil and covered with PVC roofs (Ø = 16 cm) approximately 4 cm above ground level. Each trap was half filled with salted water (20% solution) for captures conservation. Four pitfall traps were placed in each field, two at the edge and two in the middle of the plot. All collected samples were stored in plastic containers with appropriate labelling prior to determination.
