scHilda: Hierarchical Integration of LLM with KG database for single cell type annotation.

Abstract

Cell type annotation in single-cell RNA sequencing is a critical bottleneck, with existing automated methods facing limitations in accuracy, interpretability, and generalization to novel cell types. Although Large Language Models (LLMs) have recently shown potential in single-cell annotation, they are prone to inherent "hallucinations". Furthermore, a critical challenge is utilizing imperfect and potentially noisy external knowledge bases in a principled and robust manner to effectively constrain and enhance the LLM's reasoning capabilities. To address this, we propose scHilda, a novel framework designed to tackle this challenge. It deeply integrates an external Knowledge Graph into the LLM's reasoning process and employs a hierarchical arbitration annotation strategy. This strategy first identifies major cell lineages with the support of a global knowledge base and then dynamically retrieves focused subgraph domain information related to that lineage to precisely resolve cell subtypes. This dynamic knowledge-enhanced reasoning mechanism effectively constrains the LLM's decision space, reduces the risk of hallucination, and mitigates potential misguidance from knowledge base deficiencies. Tests on multiple benchmark datasets show that scHilda outperforms existing methods, achieving state-of-the-art (SOTA) performance. Notably, scHilda demonstrates exceptional robustness when handling complex mixed samples and enables lower-cost lightweight LLMs to achieve annotation performance close to that of top-tier models. Furthermore, rigorous statistical evaluations, alongside detailed interpretability case studies and query complexity analyses, validate the framework's efficiency and transparent decision-making. By deeply integrating the reasoning power of LLMs with structured biological knowledge, scHilda not only improves the accuracy and interpretability of cell annotation but also provides a new paradigm for building the next generation of trustworthy biological AI systems.