Release of Angiostrongylus cantonensis larvae from live intermediate hosts under stress stimuli

The metastrongyloid nematode Angiostrongylus cantonensis is known to cause 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 mollusc or paratenic host (poikilotherm 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 stimuli. This study aimed to compare larval release from gastropods with and without stress stimuli. 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 the 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 stress and non-stress period. In contrast, individual larvae of A. cantonensis were found in mucus from Li. fulica after stress exposure which corresponds to an increased number of DNA-positive mucus samples after stress. Apparently, stress stimuli of intensity comparable to transport or snail handling stimulate larval release, especially from highly infected intermediate hosts. However, the small number of larvae released probably does not pose a significant risk of human infection.