Multi-locus nuclear barcode assessment of genetic diversity in Swiss Orthoptera unveils conservation status limitations

With accelerating biodiversity loss, tracking both inter- and intraspecific diversity is critical, as within-species variation underpins population viability and adaptive potential. Traditional single-locus barcoding, targeting short mitochondrial or plastid fragments, generally provides reliable species-level resolution but poorly captures intraspecific variation. To address this, we developed an Orthoptera-specific marker panel targeting 398 nuclear loci plus the complete mitogenome and ribosomal DNA, sequencing these regions in 645 specimens representing every Swiss Orthoptera species sampled during the nationwide Red List update. This multilocus dataset produced a well-supported phylogeny and resolved several taxonomic ambiguities with direct conservation relevance. Genetic variation was further used to infer population structure and to test whether intraspecific diversity correlates with ecological traits and Red List status. While species’ IUCN threat levels were not correlated with genetic diversity, we found a significant negative association between genetic diversity and dependence on riparian habitats: river-bank specialists showed lower diversity, likely reflecting the severe fragmentation and alteration of these ecosystems. Our results demonstrate that genetic diversity measures from multilocus markers provide valuable, complementary insights into conservation status. However, the lack of correlation with IUCN threat levels highlights a challenge for integrating genetic data into Red List assessments: while low diversity often flags advanced decline and urgent action, high diversity does not necessarily signal low extinction risk, as it can persist for some time after population decline. Genomic data should therefore be incorporated cautiously alongside traditional criteria. These findings are being integrated into the upcoming Swiss Orthoptera Red List update. Metadata.xlsx Species, sampling locality and date, altitude, biogeographical region, legacy donor as well as extraction and library concentration per sample. Coordinates are provided in LV03+. Orthology_inference_results.xlsx Location of identified Ultra-Conserved Elements (UCEs) and Unique Single-Copy Orthologs (USCOs) in reference genomes. Loci_results_summary.xlsx Summary statistics of global multiple sequence alignments of COI barcodes, mitochondrial genes, ribosomal DNA, Ultra-Conserved Elements (UCEs), and Unique Single-Copy Orthologs (USCOs) per family. Sample_results_summary.xlsx Number of raw, clean, and mapped reads per sample, as well as the number and length of Ultra-Conserved Elements (UCEs) and Unique Single-Copy Orthologs (USCOs). The file also provides genetic diversity values, the number and length of mitochondrial genes, and the lengths of COI sequences and ribosomal DNA. Data_genetic_diversity_estimates. Percentage of polymorphic sites per species, calculated using UCEs and USCOs, ribosomal DNA, the entire mitogenome or the COI barcode. The dataset includes associated ecological and conservation factors for each species: IUCN Red List national threat status, Conservation priority designation at the Swiss level, Degree of habitat specificity, Indicator status for riparian habitats, Indicator status for dry meadow and pasture habitats, Estimated species mobility, Preference for warm habitats, Elevational range or preference. Folder MSA_Phylogenies Family-level phylogenetic tree inferred with IQ-TREE v2.0.5 using COI barcodes, mitochondrial genes, ribosomal DNA, Ultra-Conserved Elements (UCEs), and Unique Single-Copy Orthologs (USCOs).随着生物多样性丧失速率不断加快，同时追踪种间与种内多样性至关重要，因为种内变异是种群存续能力与适应潜力的基础。传统单基因条形码技术靶向线粒体或质体的短片段，通常可提供可靠的物种级分辨率，但难以捕捉种内变异。为解决这一局限，我们开发了一套直翅目（Orthoptera）专属标记面板，靶向398个核基因座以及完整线粒体基因组与核糖体DNA，并对645份标本的上述区域进行测序——这些标本涵盖了全国性红色名录更新期间采样的所有瑞士直翅目物种。 该多基因座数据集构建了支持度良好的系统发育树，并解决了若干与保护直接相关的分类学歧义。研究进一步利用遗传变异推断种群结构，并检验种内多样性是否与生态性状及红色名录等级存在关联。尽管物种的世界自然保护联盟（IUCN）威胁等级与遗传多样性并无关联，但我们发现遗传多样性与河岸生境依赖程度存在显著负相关：河岸生境专性物种的遗传多样性更低，这或许反映了此类生态系统的严重破碎化与改造。 我们的研究结果表明，基于多基因座标记的遗传多样性评估可为保护现状评估提供极具价值的补充性见解。然而，遗传多样性与IUCN威胁等级并无关联这一现象，也为将遗传数据整合进红色名录评估带来了挑战：尽管低遗传多样性往往预示着种群已严重衰退、亟待采取保护行动，但高遗传多样性并不必然代表低灭绝风险——因为种群衰退后，高遗传多样性仍可能持续一段时间。因此，基因组数据应与传统评估标准谨慎结合使用。 上述研究发现已被整合进即将更新的瑞士直翅目红色名录。 Metadata.xlsx：包含每份标本的物种信息、采样地点与日期、海拔、生物地理区域、既往捐赠方，以及每份样本的提取浓度与文库浓度。坐标采用LV03+坐标系标注。 Orthology_inference_results.xlsx：包含参考基因组中已鉴定的超保守元件（Ultra-Conserved Elements, UCEs）与单拷贝独特同源基因（Unique Single-Copy Orthologs, USCOs）的位置信息。 Loci_results_summary.xlsx：包含各科COI条形码、线粒体基因、核糖体DNA、超保守元件（UCEs）以及单拷贝独特同源基因（USCOs）的全局多序列联配统计汇总。 Sample_results_summary.xlsx：包含每份样本的原始读段、清洁读段与比对读段数量，以及超保守元件（UCEs）与单拷贝独特同源基因（USCOs）的数量与长度。该文件还提供了遗传多样性数值、线粒体基因的数量与长度，以及COI序列与核糖体DNA的长度。 Data_genetic_diversity_estimates.xlsx：包含基于超保守元件（UCEs）、单拷贝独特同源基因（USCOs）、核糖体DNA、完整线粒体基因组或COI条形码计算得到的各物种种多态位点占比。数据集还包含各物种的相关生态与保护因子信息：瑞士国家级IUCN红色名录威胁等级、瑞士层面的保护优先级划分、生境专化程度、河岸生境指示物种地位、干燥草甸与牧场生境指示物种地位、估计的物种移动性、暖生境偏好、海拔范围或海拔偏好。 MSA_Phylogenies文件夹：包含使用IQ-TREE v2.0.5基于COI条形码、线粒体基因、核糖体DNA、超保守元件（UCEs）以及单拷贝独特同源基因（USCOs）构建的科水平系统发育树。