Since the time of European settlement, land-use history, management practices, the introduction of non-native invasive species (NNIS), and climate change have dramatically changed the successional pathways of Southern Illinois forests. Biodiversity is considered essential for ecosystem health and resiliency, so understanding the impact theses forest changes have on biodiversity is necessary to guide future management decisions. This study was conducted at Touch of Nature Environmental Center (TONEC) in Southern Illinois using the Shannon diversity index (H') to compare the levels of biodiversity in the overstory, shrub, seedling, and herbaceous layers across early, mid, and mature successional stages. Twenty plots were randomly placed within early, mid, and mature forest successional stages for a total of 60 plots. Four circular nested vegetation plots were recorded at each plot location. In the overstory plots (area 314.16 m2), woody stems above 6.5 cm diameter at breast height (dbh), age class, and crown were all recorded. In the shrub layer (area 28.27 m2), all woody stems between 2.5 and 6.5 cm dbh were recorded, while in the seedling layer (area 3.14 m2), all woody stems < 1 cm dbh were recorded. In the herbaceous layer, (area 1 m2) species were measured by percent cover. Shannon diversity index (H') was calculated for each plot. When vegetation layers were combined and mean H' within forest successional stages were compared, no differences were found. However, when successional stages were combined within each vegetation layer, the shrub layer had the least H' increasing to seedling, increasing again to overstory and herbaceous; these being equal. When breaking down the vegetation layers separately and comparing H' across successional stages within each, both the overstory and shrub layers had no difference. Early successional was the least diverse in seedling, rising to mid and mature, which were equal. The herbaceous layer showed an opposite trend with early having the highest H' decreasing to mid and decreasing again to mature. Finally looking at the differences across vegetation layers within early, mid, and mature successional stages. Early-successional had the most variability with the highest H' in herbaceous; this decreased progressively to overstory then to shrub and seedling, which were equal. In mid-successional plots, overstory, seedling, and herbaceous layers were equal with H' decreasing in shrub. In mature plots, all vegetation layers had the same H'. Across all vegetation layers, the highest percentage of NNIS was in early-successional, followed by mid, with notably less found in mature forest. Even in instances where H' values were the same, species composition across forest successional stages were quite different. While overall biodiversity is the same across successional stages, differences in H' can be seen when looking at the vegetation layers. Management should focus on removing NNIS in the early and mid-successional forests paying particular attention to the shrub layer, which has the overall lowest H' with the most dramatic differences in composition and the presence of NNIS across all successional stages. Also of concern is the low H' in the seedling layer in early-successional forest, which reduces the likelihood of successful regeneration of these hardwood stands in the future.

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