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. 2019 Dec 13;9:19028. doi: 10.1038/s41598-019-54413-6

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© The Author(s) 2019

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.

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AeAmt1 abundance and immunolocalization in the alimentary canal, anal papillae, and carcass of wild-collected and laboratory A. aegypti larvae reared in freshwater (FW) and septic water (Septic). (a) AeAmt1 abundance and representative Western blot (right panel) in the carcass of wild A. aegypti larvae (n = 3). (b) AeAmt1 abundance and representative Western blot (right panel) in the Malpighian tubules (MT) and anal papillae (AP) of laboratory A. aegypti larvae (n = 3 FW, n = 4 Septic). The abundance of AeAmt1 protein was normalized to total protein (Coomassie protein stain, not shown), and Septic values are expressed relative to the control FW group (assigned a value of 1). Data shown as mean ± S.E.M. [Unpaired, two-tailed t-test; p < 0.05]. Representative transverse sections of anal papillae (AP) showing AeAmt1 (red) immunostaining from (c) wild FW-reared larvae, (d) wild Septic-reared larvae, (e) laboratory FW-reared larvae and (f) laboratory Septic-reared larvae. Nuclei are labelled by DAPI (blue) staining. Representative cross sections of the Malpighian tubules (MT) and rectum (RM) showing AeAmt1 (red) immunostaining from (g) wild FW-reared larvae, (h) wild Septic-reared larvae, (i) laboratory FW-reared larvae and (j) laboratory Septic-reared larvae. Nuclei are labelled by DAPI (blue) staining. Immunostaining of V₁ subunit of V-type H⁺-ATPase (VA⁾ is green (g–j). Co-localization of AeAmt1 with apical V₁ subunit of V-type H⁺-ATPase is indicated (dashed arrows) in the MT and RM (merge, yellow). Control sections (primary antibodies omitted, not shown) were devoid of red and green staining. Illustrations of the alimentary canal and anal papillae of A. aegypti larvae to the left of each immunofluorescence image indicates the region of the cross or transverse section (red rectangles). Lumen, (lm); anal papillae (AP); rectum (RM). Scale bars: 50 µm (c–j).