Release of Angiostrongylus cantonensis larvae from live intermediate hosts under stress

The metastrongyloid nematode Angiostrongylus cantonensis causes eosinophilic meningitis in a variety of homeothermic hosts including humans. Third-stage infectious larvae develop in gastropods as intermediate hosts. Humans are usually infected by intentional or incidental ingestion of an infected mollusk or paratenic host (poikilothermic vertebrates and invertebrates). The infection may also hypothetically occur through ingestion of food or water contaminated by third-stage larvae spontaneously released from gastropods. Larvae are thought to be released in greater numbers from the intermediate host exposed to stress. This study aimed to compare larval release from stressed with unstressed gastropods. Experimentally infected Limax maximus and Lissachatina fulica were exposed to a stress stimulus (shaking on an orbital shaker). The mucus was collected before and after the stress and examined microscopically and by qPCR for the presence of A. cantonensis larvae and their DNA. In the case of L. maximus, no larvae were detected microscopically in the mucus, but qPCR analysis confirmed the presence of A. cantonensis DNA in all experimental replicates, without clear differences between stressed and non-stressed individuals. In contrast, individual larvae of A. cantonensis were found in mucus from Li. fulica after stress exposure, which also reflects an increased number of DNA-positive mucus samples after stress. Stress stimuli of intensity similar to the transport or handling of mollusks can stimulate the release of larvae from highly infected intermediate hosts. However, these larvae are released in small numbers. The exact number of larvae required to trigger neuroangiostrongyliasis is unknown. Therefore, caution is essential when interacting with potential intermediate hosts in regions where A. cantonensis is endemic.