Abstract
The accessory nidamental gland (ANG) is a female reproductive organ found in most squid and cuttlefish that contains a consortium of bacteria. These symbiotic bacteria are transmitted from the marine environment and selected by the host through an unknown mechanism. In animals, a common antimicrobial mechanism of innate immunity is iron sequestration, which is based on the development of transferrin (TF)-like proteins. To understand this mechanism of host-microbe interaction, we attempted to characterize the role of transferrin in bigfin reef squid (Sepioteuthis lessoniana) during bacterial transmission. qPCR analysis showed that Tf was exclusively expressed in the outer layer of ANG,and this was confirmed by in situ hybridization, which showed that Tf was localized in the outer epithelial cell layer of the ANG. Western blot analysis indicated that TF is a soluble glycoprotein. Immunohistochemical staining also showed that TF is localized in the outer epithelial cell layer of the ANG and that it is mainly expressed in the outer layer during ANG growth. These results suggest that robust Tf mRNA and TF protein expression in the outer layer of the ANG plays an important role in microbe selection by the host during bacterial transmission.
Highlights
Birds[13,18] and reptiles[19]
Because bacteria were observed on the surface of the accessory nidamental gland (ANG) at stage 1 (Figs. 2 and 3), but later appeared within the invaginating primordial and secondary tubules at stage 2 (Figs. 2 and 3), our data support the hypothesis that the ANG houses a bacterial consortium that is horizontally transmitted from the environmental bacterial community
Fluorescence in situ hybridization (FISH) analyses showed that the tubules of the ANG are dominated by single taxonomic groups[29,30]
Summary
Birds[13,18] and reptiles[19]. LTF has similar functions to ovotransferrin and is found in most milk and tear secretions in mammals, where its role is to inhibit the proliferation of invading microorganisms[20]. TF may play an important antimicrobial role through iron sequestration in teleost and invertebrates. To elucidate the potential role of TF in the ANG of cephalopods during bacterial transmission, we cloned a Tf gene from the bigfin reef squid ( called oval squid, Sepioteuthis lessoniana). To visulize the localization of TF in the ANG of bigfin reef squid, we made a TF-specific antibody and demonstrated the presence of Tf-expressing cells and TF itself in the outer epithelial cell layer of the ANG. We found that TF is mainly distributed in the outer epithelial cell layer of the ANG during the developmental stage in which bacterial colonization occurs. TF may, play an important role in the primary phase of bacterial selection in the ANG of bigfin reef squid
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