A safe and effective pan-filovirus vaccine is highly desirable since the filoviruses Ebola virus (EBOV) and Marburg virus (MARV) cause highly lethal disease typified by unimpeded viral replication and severe hemorrhagic fever. Previously, we showed that expression of the homologous glycoprotein (GP) and matrix protein VP40 from a single filovirus, either EBOV or MARV, resulted in formation of wild-type virus-like particles (VLPs) in mammalian cells. When used as a vaccine, the wild-type VLPs protected from homologous filovirus challenge. The aim of this work was to generate a multi-agent vaccine that would simultaneously protect against multiple and diverse members of the Filoviridae family. Our initial approach was to construct hybrid VLPs containing heterologous viral proteins, of EBOV and MARV, and test the efficacy of the hybrid VLPs in a guinea pig model. Our data indicate that vaccination with GP was required and sufficient to protect against a homologous filovirus challenge, as heterologous wild-type VLPs or hybrid VLPs that did not contain the homologous GP failed to protect. Alternately, we vaccinated guinea pigs with a mixture of wild-type Ebola and Marburg VLPs. Vaccination with a single dose of the multivalent VLP vaccine elicited strong immune responses to both viruses and protected animals against EBOV and MARV challenge. This work provides a critical foundation towards the development of a pan-filovirus vaccine that is safe and effective for use in primates and humans.