140 »Neretva Science Week 2023« NATURA SLOVENIAE vol. 27, no. 3 (2025): 140–147 Assessing soil biodiversity in different land uses of the Nevesinje Field Vid NAGLIČ KRATKA ZNANSTVENA VEST / SHORT COMMUNICATION Agricultural institute of Slovenia, Hacquetova ulica 17, SI-1000 Ljubljana, Slovenia; E-mail: vid.naglic@kis.si Izdatajelj/Issued jointly by: Biotehniška fakulteta Univerze v Ljubljani in Nacionalni inštitut za biologijo / Biotechnical Faculty at University of Ljubljana and National Institute of Biology Založila/Published by: Založba Univerze v Ljubljani/University of Ljubljana Press ABSTRACT This study investigates soil biodiversity in agricultural landscapes and focus- es on the Nevesinje field in Bosnia and Herzegovina, which was sampled as part of the Neretva Science Week 2023. Soil microarthropods, including mites and springtails, play an important role in ecosystem health through nutrient cycling and organic matter decomposition. Using separate approaches — morphological techniques with the QBS-ar index and metabarcoding of DNA extracted from soil samples — we investigated the diversity and abundance of soil microarthropods at different sites. Our results showed that the arable field had the highest abundance of soil microarthropods, especially Acarina and Collembola, while the orchard had the lowest abundance but the high- est QBS-ar value, which reflects higher soil quality according to the QBS-ar index. Insects (Insecta) dominate with 68.6% of identified taxa, followed by arachnids (Arachnida) with 20.2%. The arable field next to the Zalomka River showed the highest taxonomic richness. The area faces imminent threats from the proposed dam construction, which could alter the hydrological dynamics and integrity of the habitat. Continued monitoring and further sampling are important to assess the ecological impact and develop conservation strate- gies to maintain soil biodiversity amid developmental pressures. IZVLEČEK Ocenjevanje biodiverzitete različnih tipov rabe tal na Nevesinjskem polju Ta študija raziskuje biotsko raznovrstnost tal v kmetijski krajini Nevesinjskega polja v Bosni in Hercegovini, ki je bilo vzorčeno v okviru Neretva Science Week 2023. Talni mikronevretenčarji, vključno s pršicami in skakači, imajo pomem- bno vlogo pri zdravju ekosistema z vplivom na kroženje hranil in razgradnjo organske snovi. Z dvema različnima pristopoma, morfološkimi tehnikami z indeksom QBS-ar in metabarkodiranjem DNK iz talnih vzorcev, smo raziskali raznolikost in številčnost talnih organizmov na različnih lokacijah. Naši rezul- tati so pokazali, da je imela njiva največjo številčnost talnih mikronevretenčar- jev, zlasti Acarina in Collembola, medtem ko je imel sadovnjak najmanjšo številčnost, vendar najvišjo vrednost QBS-ar, kar kaže na visoko kakovost tal glede na QBS. Taksonomska analiza DNK je pokazala prevlado žuželk (Insec- ta) s 68,6 %, z 20,2 % so pomembno prispevali še pajkovci (Arachnida). Njiva ob reki Zalomki je bila najbolj taksonomsko pestra od vseh zbranih vzorcev. To območje se sooča z neposrednimi grožnjami zaradi predlagane gradnje jezu, ki bi lahko spremenila hidrološko dinamiko in celovitost habitata. Stalno spremljanje in nadaljnje vzorčenje je pomembno za oceno ekološkega vpli- va in razvoj strategij za ohranjanje biotske raznovrstnosti tal v luči razvojnih pritiskov. Prejeto / Received: 24. 10. 2024 Sprejeto / Accepted: 30. 4. 2025 DOI: 10.14720/ns.20047 Objavljeno na spletu / Published online: 12. 11. 2025 NATURA SLOVENIAE vol. 27, no. 3 (2025) N AG LI Č: A ss es si ng s oi l b io di ve rs it y in d iff er en t l an d us es ... 141NATURA SLOVENIAE vol. 27, no. 3 (2025) NAGLIČ: Assessing soil biodiversity in different land uses... KRATKA ZNANSTVENA VEST / SHORT COMMUNICATION INTRODUCTION Soil microarthropods are crucial for maintaining soil health and play a key role in ecosystem stability through processes such as nutrient cycling and organic matter decomposition (Wall & Lynch 2000; Coleman et al. 2004). Despite their importance, these small soil organisms are often overlooked when assess- ing soil quality (Parisi et al. 2005). In agricultural landscapes, especially those exposed to anthropogenic pressures, the di- versity and abundance of soil microarthropods can serve as important indicators of ecosystem health and resilience (Men- ta 2012; Postma-Blaauw et al. 2010). The Nevesinje field in Bos- nia and Herzegovina is an agriculturally important region that mostly features traditional extensive type of agriculture. That area could soon experience significant environmental changes due to the planned construction of a dam (Gallop 2023). Such developments may lead to changes in land use and potentially affect soil biodiversity (Foley et al. 2005). In order to assess the potential ecological impact of such infrastructure projects, it is important to understand the current biodiversity of soil organ- isms in this region. The aim of our research was to assess the biodiversity of soil microarthropods in the Nevesinje field using two complementary approaches: morphological analysis with the QBS-ar index and DNA metabarcoding (defined here as DNA extracted from homogenized bulk soil samples without prior removal of soil organisms, also referred to as eDNA). The for- mer provides a qualitative assessment of soil biological quality based on the presence of soil-adapted microarthropods (Parisi et al. 2005), while the latter provides a detailed genetic insight into the soil community and identifies organisms that may be missed by conventional microscopy (Taberlet et al. 2012, 2018). Morphological analysis is highly time-consuming, requir- ing careful extraction and identification of microarthropods, which limited the number of samples that could be processed. Due to time constraints, we were only able to collect morpho- logical samples from three sites, while DNA metabarcoding was applied to samples from six sites. We hypothesized that the agricultural land of the Nevesinje field would harbor a rich and diverse soil fauna due to its habitat diversity (Wardle et al. 2004). By combining traditional morphological techniques with modern genetic tools, this study aims to provide a basic understanding of soil biodiversity in this little-studied region. MATERIAL AND METHODS STUDY AREA AND SAMPLING Soil samples were collected from six sites within the Nevesinje field, Bosnia and Herzegovina, representing different types of agricultural use. They included a meadow, an orchard, an ara- ble field, a riparian habitat, an arable field near Zalomka river, and an oak forest. These locations were chosen on the basis of land use and proximity to the proposed dam construction site. GPS coordinates were recorded for each sampling location: • Meadow: N 43.184816, E 18.160509 (QBS, DNA) • Orchard: N 43.179056, E 18.158028 (QBS, DNA) • Arable field: N 43.178778, E 18.157995 (QBS, DNA) • Riparian habitat near Zalomka river: N 43.210843, E 18.221617 (DNA) • Arable field near Zalomka river: N 43.211345, E 18.222782 (DNA) • Oak forest: N 43.185420, E 18.121981 (DNA) APSTRAKT Procjena biodiverziteta različitih tipova korištenja zemljišta u Nevesinjskom polju Ova studija istražuje biodiverzitet zemljišta u poljoprivrednom području Nevesinjskog polja u Bosni i Hercegovini, koji je proučavan u okviru Sedmice nauke Neretva 2023. Zemljišni mikroartropodi, uključujući grinje i skokune, ig- raju važnu ulogu u održanju zdravih ekosistema utičući na kruženje hranjivih materija i razgradnju organske materije. Koristeći dva različita pristupa, mor- fološke tehnike s QBS-ar indeksom i metabarkodiranje DNK iz uzoraka zem- ljišta, istražili smo raznolikost i brojnost organizama u zemljištu na različitim lokacijama. Naši rezultati su pokazali da je obradivo zemljište imalo najveću zastupljenost mikroinvertebrata, posebno Acarina i Collembola, dok je voćn- jak imao najmanju zastupljenost, ali najvišu QBS-ar vrijednost, što ukazuje na visok kvalitet zemljišta prema QBS indeksu. Insekti (Insecta) dominiraju sa 68,6 % identifikovanih taksona, a slijede ih paukoliki zglavkari (Arachnida) sa 20,2 %. Obradivo zemljište uz rijeku Zalomku bilo je taksonomski najraznovrsni- je od svih prikupljenih uzoraka. Ovo područje se suočava sa neposrednim pri- jetnjama zbog predložene izgradnje brane, koja bi mogla promijeniti hidrološku dinamiku i integritet staništa. Kontinuirano praćenje i dalje uzorkovanje su važni za procenu ekološkog uticaja i razvoj strategija zaštite kako bi se održao biodiver- zitet zemljišta usled razvojnih pritisaka. KEY WORDS: soil microarthropods, DNA metabarcoding, QBS-ar, mesofauna, Upper Horizons project KLJUČNE BESEDE: talni mikronevretenčarji, DNA metabarkodiranje, QBS-ar, mezofavna, projekt Gornji Horizonti KLJUČNE RIJEČI: mikroartropodi u zemljištu, DNK metabarkodiranje, QBS-ar, mezofauna, projekat Gornji Horizonti Figure 1. Cumulative abundance of soil microarthropods for each site. Slika 1. Kumulativna številčnost talnih mikronevretenčarjev za vsako lokacijo. Slika 1. Kumulativna brojnost mikroartropoda u tlu za svaki lokalitet. Site QBS-ar Number of EMI groups Arable Field 102 14 Orchard 106 11 Meadow 86 12 Table 1. QBS-ar index and number of ecomorphological (EMI) groups for each site. Tabela 1. Indeks QBS-ar in število ekomorfoloških (EMI) skupin za vsako lokacijo. Tabela 1. QBS-ar indeks i broj ekomorfoloških (EMI) grupa za svaki lokalitet. 142NATURA SLOVENIAE vol. 27, no. 3 (2025) NAGLIČ: Assessing soil biodiversity in different land uses... KRATKA ZNANSTVENA VEST / SHORT COMMUNICATION Sampling was conducted over two days (5. and 6. 6. 2023), approximately at the same time of the day. Descriptions of sampling locations are included in the supplementary mate- rial. Samples for morphological analysis (QBS-ar) were taken at three sites (arable field, meadow, orchard). At each, three subsamples of soil were taken from the top 10 cm of the soil profile using a soil corer with a diameter of 11.3 cm. Samples for DNA analysis were collected at six sites. On each site, three subsamples for DNA analysis were collected, using the same sampling technique as for the QBS. Subsamples were joined and manually homogenized in the field. Both the QBS and DNA samples were stored in a cool box immediately after collection to preserve their integrity. Three days after sampling, the QBS samples were processed using a Kempson extractor with the organisms heat-extracted and stored in ethanol, while the DNA samples were frozen until subsequent analysis. MORPHOLOGICAL ANALYSIS Morphological analysis of soil samples was performed using the QBS-ar (Biological Soil Quality Index) method (Parisi et al. 2005). This index assesses soil biological quality by examining the presence of soil microarthropods with morphological ad- aptations to life in the soil. The organisms were identified to class or order level under a stereomicroscope, and a combined value of QBS-ar was calculated for each site based on three collected samples. The QBS-ar index is calculated as the sum of the Ecomorphological Index (EMI) values assigned to taxa identified in the sample. EMI values range from 1 to 20, with higher values assigned to taxa with greater adaptation to soil life. In addition, the abundance of animals was recorded in all subsamples. DNA EXTRACTIONS FROM SOIL, LIBRARY PREPARATIONS, SEQUENCING AND BIOINFORMATIC ANALYSIS Six composite samples underwent DNA analysis using the DNeasy PowerLyzer PowerSoil isolation kit (Qiagen, Denmark). Approximately 40 ml of soil per sample was used. Soil sam- ples were freeze-dried for 72 hours and homogenized with a Bead Ruptor Elite (Omni International) at 4 m s-1 for 30 seconds, the process was repeated three times. For the PowerSoil ex- tractions, 0.25 g of homogenized soil was used, following the manufacturer’s instructions. DNA concentrations were quan- tified using a Qubit 4.0 Fluorometer (Invitrogen, USA). The COI region was amplified with mlCOlintF/jgHCO2198 primers (Geller et al. 2013) using a two-step PCR and dual indexing for Illumina MiSeq sequencing. Final PCR products were cleaned with HighPrep™ magnetic beads (MagBio Genomics Inc.) and pooled for sequencing. Illumina reads were processed in QIIME2 (v2020.10.0) (Bolyen et al. 2019), where primer sequenc- es were trimmed, and reads truncated after 230 base pairs. Reads were filtered, denoised, merged, and dereplicated using DADA2 (Callahan et al. 2016), producing Amplicon Sequence Variants (ASVs). Details about the number of reads before and after processing are included in the supplementary material. Taxonomic classification was performed via BLAST against the NCBI database (Sayers et al. 2022). ASVs associated with the thirty-three taxonomic groups used for QBS-ar index identi- fication were retained, including soil microarthropods (e.g. Acari, Collembola), myriapods (Diplopoda, Chilopoda, Paurop- oda, Symphyla), isopods, insects (Coleoptera, Hymenoptera, Diptera), and other soil-dwelling arthropods. Data analysis and visualization were conducted in Python, version 3.13. RESULTS AND DISCUSSION We morphologically identified individuals in the soil samples to the level of morphospecies and counted cumulative abun- dance of important classes, number of EMI groups and QBS-ar index, for three sites. The arable field and orchard had QBS-ar values higher than 93.7 (Tab. 1), the tentative threshold value to distinguish high-quality soils (Menta et al. 2018), while the meadow had a lower QBS-ar value (Tab. 1). Based on morpho- logical identification, the arable field had the most EMI groups, followed by the meadow and the orchard (Tab. 1). The arable field had the highest cumulative abundance (absolute counts of individuals) of soil microarthropods, especially mites (Aca- rina) and springtails (Collembola) (Fig. 1), while meadow and orchard had lower cumulative abundances. Figure 2. Taxon richness across various habitats. The numbers above each bar represent the total taxon richness (calculated as the number of unique ASVs identified) for each habitat. The bar colors indicate the proportional contributions of Acari (light gray), Collembola (dark gray), and other taxa (black) to the total richness. Slika 2. Taksonomska pestrost v različnih habitatih. Številke nad posameznimi stolpci predstavljajo skupno taksonomsko pestrost (izraženo kot število unikatnih ASV-jev, ki so bili identificirani) za vsak habitat. Barve stolpcev označujejo proporcionalni prispevek pršic (Acari) (svetlo siva), skakačev (Collembola) (temno siva) in drugih taksonov (črna) k skupni pestrosti. Slika 2. Taksonomsko bogastvo vrstama u različitim staništima. Brojevi iznad svake kolone predstavljaju ukupnu taksonomsko bogastvo (iz- raženu kao broj jedinstvenih ASV-ova koji su identifikovani) za svako stanište. Boje u stupcima označavaju proporcionalni doprinos grinja (Acari) (svijetlosiva), skokuna (Collembola) (tamnosiva) i ostalih taksona (crna) ukupnoj raznolikosti. 143NATURA SLOVENIAE vol. 27, no. 3 (2025) NAGLIČ: Assessing soil biodiversity in different land uses... KRATKA ZNANSTVENA VEST / SHORT COMMUNICATION We acknowledge that statistical comparison between differ- ent localities cannot be made, since we do not have biological replicates. However, the first results are in congruence with our expectations for all analysed types of soil. For example, low QBS-ar value of the meadow can be explained with lower plant diversity and soil compaction due to grazing or use of machinery, which can have a negative effect on the microar- thropod communities in the soil, resulting in lower diversity and abundance of soil microarthropods (Wardle et al. 2004; Beylich et al. 2010). The reason could also be the low organic matter content in the soil, which may limit the availability of food resources necessary to sustain a diverse and abundant microarthropod community, although this was not directly tested as no physico-chemical analysis of the soil was conduct- ed. Overall, first insights based on QBS-ar index point to the fact that agricultural land of the Nevesinje field indeed harbors a rich and diverse soil fauna. Next, we employed the DNA metabarcoding approach to analyse the taxonomic composition of soil microarthropods at six different sites. We filtered out ASVs associated with fau- na groups used in QBS-ar index and analysed taxon richness (Fig. 2) and relative abundance of most represented classes and orders (Fig. 3). Taxon richness for each site was calculated as the number of unique ASVs identified and assigned to analysed taxonomic groups, for each site (Fig. 2). The agricultural lands had higher taxon richness of soil related taxa than natural habitats. These results are preliminary and based on a single sampling event, yet they align with several studies, that showed that agricul- tural practices, particularly in arable fields, may promote soil biodiversity by increasing organic matter and creating diverse microhabitats, which are known to increase the diversity of soil microarthropods (Mäder et al. 2002; Wardle et al. 2004). In contrast, the riparian habitat and the meadow had lower taxon richness, probably due to specific habitat characteristics that limit the occurrence of certain soil taxa. Riparian zones, although dynamic, can also have fluctuating moisture levels that may limit the diversity of certain soil fauna (Naiman & Décamps 1997). Meadows, on the other hand, may have lower taxon richness due to factors already described in the first part of the results chapter: we obtained similar results by means of morphological identification with the meadow having the low- est QBS-ar score out of the three sampled sites. These patterns we observe may also reflect the fact that certain habitats (i.e. those used for QBS assessment) inherently contain fewer taxa that were included in our analysis. Additionally, limitations in Figure 3. Relative abundance of taxonomic classes (A) and orders (B) of soil microarthropods across all sampling sites. Slika 3. Relativna številčnost taksonomskih razredov (A) in redov (B) talnih mikronevretenčarjev na vseh vzorčnih mestih. Slika 3. Relativna brojnost taksonomskih klasa (A) i redova (B) mikroartropoda u zemljištu na svim mjestima uzorkovanja. 144NATURA SLOVENIAE vol. 27, no. 3 (2025) NAGLIČ: Assessing soil biodiversity in different land uses... KRATKA ZNANSTVENA VEST / SHORT COMMUNICATION taxonomic databases, particularly for some microarthropod groups, or the relatively low number of our samples, combined with the rapid, non-replicated sampling approach, could have influenced the observed richness patterns. With the given time constraints, this was the most comprehensive assessment we could achieve. The taxonomic composition of soil invertebrates derived from the DNA metabarcoding analysis for different habitats is shown in the pie charts in Fig. 3. The left-hand diagram (Fig. 3A) shows the distribution of ASVs across the five most represented class- es. Insects (Insecta) account for 68.6% of the ASVs identified, followed by arachnids (Arachnida) with 20.2%. Other class- es, such as Malacostraca, Collembola, and Diplopoda, were represented at lower percentages. The diagram on the right (Fig. 3B) shows the distribution of ASVs assigned to the ten most represented orders in our dataset. Hymenoptera ac- counted for 22.9% of the identified ASVs, followed by Coleop- tera with 16.9%. It is important to note that these percentages represent the proportion of ASVs classified within each order, not the actual abundance of organisms in the soil community. The observed percentages reflect the taxonomic resolution achieved through DNA metabarcoding, which depends on ref- erence databases and the biases associated with PCR ampli- fication. When comparing these results with morphological identifica- tion, it is important to note that the two different approaches – morphology-based abundance and DNA-based diversity of soil microarthropods – do not necessarily correlate. Nevertheless, we find that two orders of mites (Sarcoptiformes and Trombid- iformes) are among the most represented orders in our DNA analysis, and they were also abundant in morphological anal- ysis. In contrast, Collembola were among the most abundant taxa in the morphological results but was only represented at the class level in the DNA data. Different Collembola orders were included in the »Other« category in Fig. 3B because their relative representation among the identified ASVs was insuf- ficient to rank them among the top ten orders. We also found that many Collembola sequences could not be identified at a higher taxonomic resolution, possibly due to the limited capa- bilities of the reference databases for Collembola in the soil. This highlights the challenges in metabarcoding studies asso- ciated with incomplete reference libraries, which can hinder accurate taxonomic assignment (Porco et al. 2013). Improving these databases is crucial to increase the resolution and reli- ability of metabarcoding in soil biodiversity assessment. The results should be interpreted as the first qualitative obser- vations in the area rather than definitive conclusions about soil diversity. This is a snapshot study based on a single sampling event, which does not account for biological replicates, tempo- ral variability or the full extent of soil biodiversity. However, it offers the first insight into soil diversity of this remote, under- studied and critically endangered area. The results highlight the ecological significance of the study area, suggesting that the relatively high soil quality in agricultural lands may be at- tributed to a combination of extensive farming and traditional agricultural practices. This biodiversity is threatened by the planned dam construction, which could disrupt hydrological patterns, destroy and fragment habitats and ultimately reduce the richness and abundance of the soil microarthropods. Fast changes in water flow and habitat structure could lead to the loss of important microhabitats. Given the potential impact on this fragile ecosystem, continuous sampling is essential to monitor these changes and develop strategies to protect soil biodiversity in the face of development pressure. This study represents the first attempt to assess the quality of soil, using a combination of traditional and DNA techniques in this area. We hope that it will shed some light on this hidden yet important ecosystem, which is tightly connected with local traditions and human use of the landscape. 145NATURA SLOVENIAE vol. 27, no. 3 (2025) NAGLIČ: Assessing soil biodiversity in different land uses... KRATKA ZNANSTVENA VEST / SHORT COMMUNICATION POVZETEK V raziskavi smo preučevali biotsko raznovrstnost talnih or- ganizmov na območju Nevesinjskega polja v Bosni in Her- cegovini, kjer načrtovana gradnja jezu predstavlja poten- cialno grožnjo za tamkajšnji ekosistem. Zbrali smo vzorce tal na šestih različnih lokacijah, ki vključujejo dve njivi, sa- dovnjak, travnik, obrežni habitat ob reki Zalomki in hrastov gozd. Raziskava je temeljila na dveh pristopih: morfološki analizi preko QBS-ar indeksa ter DNA metabarkodiranju, kar nam je omogočilo vpogled v genetsko pestrost talnih orga- nizmov. Morfološki rezultati kažejo, da je bila največja šte- vilčnost talnih mikronevretenčarjev, predvsem pršic (Acari- na) in skakačev (Collembola), zaznana na njivi, medtem ko je imel sadovnjak najmanjšo številčnost, a najvišji QBS-ar indeks (106), ki nakazuje na visoko biološko kakovost tal. Nasprotno je bil QBS-ar indeks na njivi 102, na travniku pa 86. Z metabarkodiranjem smo največjo taksonomsko pes- trost mikronevretenčarjev zaznali na njivi ob reki Zalomki, kar kaže na bogato biotsko raznovrstnost tega območja, medtem ko sta travnik in obrežni habitat pokazala nižjo pestrost. Taksonomska analiza DNA je razkrila prevlado žuželk (Insecta), ki so predstavljale 68,6 % vseh prepoznanih taksonov, z znatno prisotnostjo pajkovcev (Arachnida) (20,2 %). Med najbolj pogostimi redovi so bili kožekrilci (Hymenoptera) in hrošči (Coleoptera). Red Collembola je bil vključen v analizo, vendar njihova relativna številčnost tak- sonomskih skupin ni dosegla prve deseterice kljub temu, da so bile glede na morfološke rezultate vsaj v nekaterih vzorcih najštevilčnejše, kar nakazuje na potrebo po izboljšanih podatkovnih bazah za njihovo natančnejšo identifikacijo. Ti rezultati kažejo na pomembno biotsko raznovrstnost Nevesinjskega polja, ki pa je zaradi načrtovane gradnje jezu resno ogrožena. Nadaljnje spremljanje stanja in dodat- no vzorčenje sta ključnega pomena za oceno dolgoročnih ekoloških posledic in za razvoj strategij, ki bodo zagotovile ohranitev talne biotske raznovrstnosti v tem občutljivem okolju. SAŽETAK Tokom istraživanja smo proučavali biodiverzitet organi- zama u zemljištu na području Nevesinjskog polja u Bosni i Hercegovini, gdje planirana izgradnja brane predstavlja potencijalnu prijetnju lokalnim ekosistemima. Uzorke zem- ljišta prikupili smo na šest različitih lokacija, uključujući dva obradiva polja, voćnjak, livadu, priobalno stanište uz rijeku Zalomku i hrastovu šumu. Istraživanje je zasnovano na dva pristupa: morfološkoj analizi putem QBS-ar indeksa i DNK metabarkodiranju, što nam je omogućilo da steknemo uvid u genetsku raznolikost organizama u zemljištu. Morfološki rezultati pokazuju da je najveća brojnost mikroartropoda u zemljištu, uglavnom grinja (Acarina) i skokuna (Collem- bola), otkrivena na polju, dok je voćnjak imao najmanju brojnost, ali najviši QBS-ar indeks (106), što ukazuje na visok biološki kvalitet tla. Nasuprot tome, QBS-ar indeks na polju je bio 102, a na livadi 86. Korištenjem metabarko- diranja, detektovali smo najveću taksonomsku raznolikost mikroinvertebrata na polju uz rijeku Zalomku, što ukazuje na bogatu biodiverzitet ovog područja, dok su livadsko i priobalno stanište pokazali nižu raznolikost. Taksonomska analiza DNK otkrila je dominaciju insekata (Insecta), koji su predstavljali 68,6% svih prepoznatih taksona, sa značajnim prisustvom paučnjaka (Arachnida) (20,2%). Među najčešćim redovima bili su Hymenoptera i Coleoptera. Red Collembo- la je uključen u analizu, ali njihova relativna brojnost tak- sonomskih grupa nije dostigla prvih deset uprkos tome što su bili najbrojniji prema morfološkim rezultatima u barem nekim uzorcima, što ukazuje na potrebu za poboljšanim bazama podataka za njihovu precizniju identifikaciju. Ovi rezultati ukazuju na važan biodiverzitet Nevesinjskog pol- ja, koji je ozbiljno ugrožen planiranom izgradnjom brane. Daljnje praćenje i dodatno uzorkovanje ključni su za proc- jenu dugoročnih ekoloških posljedica i razvoj strategija koje će osigurati očuvanje biodiverziteta tla u ovom osjetljivom okruženju. ACKNOWLEDGEMENTS I would like to thank Špela Di Batista Borko for inviting me to participate in the Neretva Science Week, Marija Kravanja for her invaluable assistance during the fieldwork in Bosnia, Irena Bertoncelj for the introduction to the field of soil biodiversity and the Agricultural Institute of Slovenia for funding this re- search. I also extend my gratitude to the Environmental Micro- biology section at the Department of Environmental Science, Aarhus University, for their support and collaboration. I would also like to acknowledge the Research Infrastructure ELIXIR-SI (https://elixir-slovenia.org), funded by the European Regional Development Fund, the Ministry of Science, Education and Sports, and the Slovenian Research and Innovation Agency, which provided me with the computing resources needed for the bioinformatics analysis. 146NATURA SLOVENIAE vol. 27, no. 3 (2025) NAGLIČ: Assessing soil biodiversity in different land uses... KRATKA ZNANSTVENA VEST / SHORT COMMUNICATION REFERENCES Beylich A, Oberholzer H-R, Schrader S, Höper H, Wilke B-M. 2010. Evaluation of soil compaction effects on soil biota and soil biological processes in soils. Soil Tillage Research. 109(2): 133-143. https://doi.org/10.1016/j.still.2010.05.010 Bolyen E, Rideout JR, Dillon MR, Bokulich NA, Abnet CC, Al-Ghalith GA, Alexander H, Alm EJ, Arumugam M, Asnicar F, et al. 2019. Reproducible, interactive, scalable and extensible microbiome data science us- ing QIIME 2. 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Science. 304(5677): 1629-1633. https://doi.org/10.1126/science.1094875 Site Input Filtered % Input Passed Filter Denoised Merged % Input Merged Non- chimeric % Input Non- chimeric Oak forest 52599 47691 90.67 44200 36983 70.31 35578 67.64 Meadow 90556 82478 91.08 79486 70565 77.92 60280 66.57 Arable field near Zalomka river 68107 58259 85.54 55795 49413 72.41 47350 69.52 Riparian habitat near Zalomka river 36063 30273 83.94 28275 25423 70.5 24761 68.66 Arable field 76867 65772 85.57 62432 56006 72.86 54729 71.2 Orchard 41986 35864 85.42 33882 30662 73.03 30132 71.77 147NATURA SLOVENIAE vol. 27, no. 3 (2025) NAGLIČ: Assessing soil biodiversity in different land uses... KRATKA ZNANSTVENA VEST / SHORT COMMUNICATION Vsebina je dostopna pod pogoji licence Creative Commons Priznanje avtorstva 4.0 Mednarodna (CC BY 4.0), ki omogoča neomejeno uporabo, razširjanje in reproduciranje v kateremkoli mediju ali obliki, pod pogojem, da sta ustrezno navedena izvirni avtor in vir. The content is distributed under the terms of the Creative Commons Attribution 4.0 International License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. SUPPLEMENTARY MATERIAL DESCRIPTION OF LOCATIONS The meadow was a hay meadow with extensive management, showing no visible signs of recent fertilization or heavy grazing. The arable field had been recently ploughed, with no crops present at the time of sampling. The orchard consisted of Prunus trees, with a high undergrowth of various grasses and other vegetation, suggesting a less intensively managed system. The riparian habitat near the Zalomka River had a rocky soil structure with scarce vegetation, likely due to occasional flooding that limits plant establishment. The arable field near the Zalomka River was similarly ploughed at the time of sampling, with no visible crops. The oak forest consisted primarily of deciduous oak trees, with a well-developed litter layer and relatively sparse undergrowth, indicating a stable, less-disturbed environment. Table S1. Sequence processing summary for DNA metabarcoding analysis. Details of the sequence processing pipeline, including the number of raw sequences, sequences retained after quality filtering, denoising, merging, and chimera checking, as well as their respective percentages, for each sample.