The loss of unimproved grassland has led to species decline in a wide range of taxonomic groups. Agricultural intensification has resulted in fragmented patches of remnant grassland habitat both across Europe and internationally. The monitoring of remnant patches of this habitat is critically important, however, traditional surveying of large, remote landscapes is a notoriously costly and difficult task. The emergence of small-Unmanned Aircraft Systems (sUAS) equipped with low-cost multi-spectral cameras offer an alternative to traditional grassland survey methods, and have the potential to progress and innovate the monitoring and future conservation of this habitat globally. The aim of this article is to investigate the potential of sUAS for rapid detection of threatened unimproved grassland and to test the use of an Enhanced Normalized Difference Vegetation Index (ENDVI). A sUAS aerial survey is undertaken at a site nationally recognised as an important location for fragmented unimproved mesotrophic grassland, within the south east of England, UK. A multispectral camera is used to capture imagery in the visible and near-infrared spectrums, and the ENDVI calculated and its discrimination performance compared to a range of more traditional vegetation indices. In order to validate the results of analysis, ground quadrat surveys were carried out to determine the grassland communities present. Quadrat surveys identified three community types within the site; unimproved grassland, improved grassland and rush pasture. All six vegetation indices tested were able to distinguish between the broad habitat types of grassland and rush pasture; whilst only three could differentiate vegetation at a community level. The Enhanced Normalized Difference Vegetation Index (ENDVI) was the most effective index when differentiating grasslands at the community level. The mechanisms behind the improved performance of the ENDVI are discussed and recommendations are made for areas of future research and study.