Cubozoans (box jellyfish) are gonochoristic cnidarians with distinct reproductive strategies. This comparative histological study examines the gonad organization of Alatina alata and Copula sivickisi, two box jellyfish species that exhibit different modes of internal fertilization. A. alata reproduces via spermcasting aggregations while C. sivickisi reproduces via copulation; in both cases, internal fertilization occurs in the gastrovascular cavity. Herein, we provide the first histological description of subgastric sacs-structures unique to C. sivickisi. Although previously thought to function as sperm storage sacs, our findings reveal that subgastric sacs are nematocyst nests lacking sperm entirely. Conversely, we discovered that velarial spots in C. sivickisi females correspond to actual sperm storage structures. Histological examination of cubozoan sperm packages revealed that while sperm packages from both species have motile flagella, A. alata males produce nonencapsulated sperm bundles (i.e., "spermatozeugmata"), and C. sivickisi males produce encapsulated packages (i.e., "spermatophores"). Our findings corroborate the presence of several types of nematocysts in C. sivickisi embryo strands and spermatophores, and indicate their provenance to be both female and male gonads respectively, as well as subgastric sacs (i.e., nematocyst nests). In contrast to our findings of velarial spots as sperm storages structures in C. sivickisi females, and of nematocysts in the gonads of both sexes, we report that A. alata medusae lack both sperm storage structures and gonadal nematocysts. Finally, we discuss our findings on reproductive morphology of C. sivickisi and A. alata in light of the respective reproductive behavior of these two cubozoan species.

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The box jellyfish Chiropsalmus quadrumanus (Chirodropida: Cubozoa: Cnidaria) is common in warm waters. Although it is assumed that external fertilization is a characteristic of Chirodropida, the life history of C. quadrumanus is not yet known since its reproductive behavior has never been described, nor has the polyp has been found in nature. As a result, in the absence of documentation of reproductive behavior, we sought to test the hypothesis of external fertilization through a histological analysis of the female gonads. Herein, we analyze ten females collected in São Paulo and Rio de Janeiro (Brazil) and describe the gonadal organization and pattern of oocyte development. The discovery of four distinct stages of oocyte differentiation augments the scant existing reports of the structural and functional maturation of sex cells in Cubozoa species. Furthermore, the gonads of mature females comprise both mature (average diameter of 122 µm) and immature oocytes, suggesting that C. quadrumanus is iteroparous and exhibits multiple reproductive cycles during its life. Medusa bell size was not found to correlate with maturity state as even small females possessed a high percentage of oocytes in late vitellogenesis, suggesting that sexual maturation occurs rapidly in C. quadrumanus females.