Continued emergence, re-emergence and spread of zoonotic diseases demonstrates the imperative need for multisectoral communication and joint coordination of disease detection and response. While there are existing international frameworks underpinning One Health capacity building for pandemic prevention and response, often guidance does not account for challenges faced by countries undergoing long-term conflict and sociopolitical instability. The purpose of this research was to identify Libya’s laboratory and surveillance networks and routes of inter- and multisectoral communication and coordination for priority zoonotic diseases. The One Health Systems Assessment for Priority Zoonoses (OH-SAPZ) tool is an established methodology that was adapted and applied to the Libyan context to support prioritization of zoonotic diseases, development of systems map schematics outlining networks of communication and coordination, and analysis of operations for targeted capacity building efforts. Five zoonotic diseases were selected to undergo assessment: highly pathogenic avian influenza, brucellosis, Rift Valley fever, leishmaniasis and rabies. Through decisive acknowledgement of Libya’s unique health setting, we mapped how patient and sample information is both communicated within and between the human, animal and environmental health sectors, spanning from local index case identification to international notification. Through our assessment we found strong communication within the public and animal health sectors, as well as existing multisectoral coordination on zoonotic disease response. However, local-level communication between the sectors is currently lacking. Due to the ongoing conflict, resources (financial and human) and access have been severely impacted, resulting in limited laboratory diagnostic capacity and discontinued disease prevention and control measures. We sought to identify opportunities to leverage existing operations for endemic diseases like brucellosis for emerging zoonotic threats, such as Rift Valley fever. Analysis of these operations and capabilities supports the development of targeted recommendations that address gaps and may be used as an implementation guide for future One Health capacity building efforts.