CryoGaussian: A Unified Gaussian Splatting Method for Homogeneous and Heterogeneous Cryo-EM Reconstruction

Single-particle cryo-electron microscopy (cryo-EM) resolves biomolecular structures at near-atomic resolution without crystallization. Reconstructing a biomolecule's 3D density from noisy 2D particle images presents two core challenges: (1) recovering as much reliable structural detail as the data support in both homogeneous and heterogeneous reconstruction; and (2) identifying rare and transient compositional states and tracing subtle, continuous conformational changes. To address both challenges, we propose CryoGaussian, a cryo-EM-native Gaussian splatting method that unifies homogeneous and heterogeneous reconstruction for high-fidelity structural recovery. Starting from randomly initialized Gaussians, CryoGaussian fits an explicit 3D density representation directly to particle images through orthographic line-integral projection modulated by the contrast transfer function. To model heterogeneity, CryoGaussian learns particle-image and per-Gaussian latents that jointly condition Gaussian attributes to capture compositional and conformational variation. The inferred states then guide optimization of the Gaussian representation for detailed per-state reconstruction. We validate CryoGaussian: (1) on all five synthetic CryoBench datasets, it achieves competitive performance in heterogeneity estimation and per-state reconstruction; (2) on homogeneous experimental data, it resolves structural detail in the RAG1-RAG2 complex, the SARS-CoV-2 spike on intact virions, and the 80S ribosome; and (3) on heterogeneous experimental data, it reveals previously validated structural features through unsupervised heterogeneity estimation, including a 50S assembly intermediate with rRNA helix 68 density but no central protuberance and continuous conformational variation in the SF3b and helicase regions of the pre-catalytic spliceosome. Sample maps and links are provided in the supplementary material; code available upon publication.