FIGURE 2 |.

In situ hybridization and immunohistochemistry staining demonstrate autophagy and ZIKV replication co-localize in human placentae of neonates affected by CZS. In situ hybridization with positive-strand ZIKV RNA probe set demonstrates ZIKV replication within villi from confirmed CZS cases but not in the placenta from the unaffected neonate (A: brown labeling, black arrowheads). Immunohistochemical staining for autophagic markers LC3B (B: dark red-brown labeling, black arrowheads) and SQSTM1/p62 (C: bright red labeling, black arrowheads) was noted in the same area of the villi where ZIKV replication was present. Probe-free controls confirm no non-specific binding in the absence of ZIKV, LC3B, SQSTM1/P62 probes, respectively. All images shown were obtained at 60X magnification. Micron bar scale in bottom right corner of images = 10μm. CV, chorionic villus; IVS, intervillous space; ST, syncytiotrophoblast.