Roadways vary in structural, geotechnical, locational, and operational properties, and synergies among these factors may present overwhelming challenges to understanding their full effects on the habitat quality (HQ). To explore the impact of dense roadway networks on an ecologically fragile region in the northwest of China, this study applied the Integrated Valuation of Ecosystem Services and Trade-offs (InVEST) to evaluate the HQ spatiotemporal distribution of the study area. Then, Generalised Estimating Equations (GEE) were formulated to examine the cumulative impact due to the operation of an increasing amount of roadways over the past two decades. According to the results, the influence of different road types on the HQ was apparent within the road-effect zone, and road grading reduction, road length and operation duration increase can harm the HQ within the road-effect zone. Overall, this study generates knowledge concerning the design and operation of environmentally-friendly roadways in ecologically fragile areas.