Microencephaly in fetal piglets following in utero inoculation of Zika virus

Figures & data

Fig. 1 Illustration of the experimental setup.

Sows 1543 and 1544 were subjected to laparotomy, after which all fetuses in the left uterine horn were inoculated, via IP route, with 10^5.6 TCID₅₀ of ZIKV strain PRVABC59. Sows 1545 and 1546 were inoculated via IV route with 10^6.4 TCID₅₀ of the same ZIKV strain and sows 1547 and 1548 were injected with culture medium, also via IV route

Fig. 2 Measurements of sows and fetuses subjected to ZIKV infection.

a Averages and standard deviations (SDs) of daily recorded sow body temperatures. b Numbers of live and dead (mummified) fetuses per sow and per uterine horn (left or right) at the time of necropsy. c ZIKV-specific VNT titers in sow serum at four weeks post challenge. Averages and SDs are presented. The limit of detection was set to 40. d–g Averages and SDs of crown-rump lengths d, head girths e, total weights f, and brain weights g of the live fetuses per experimental (sub)group. h, i Relative head size h and relative brain weight i of all individual fetuses. L left uterine horn,R right uterine horn. An asterisk indicates a statistical difference

Fig. 3 Macroscopic evidence of microencephaly in two fetal piglets following IP inoculation of ZIKV.

Top view of heads and brains of fetuses presenting stunted brain growth; 1543_FL3 a and 1544_FL7 b. Note the smaller gyri of the cerebral cortex of all lobes in fetus 1543_FL3 and of the parietal, temporal, and occipital lobes in fetus 1544_FL7 (white arrows)

Fig. 4 Microscopic evidence of microencephaly in 1543_FL3 and 1544_FL7.

Nissl-stained brain cross-sections of 1543_FL3 a, c and 1544_FL7 b, d. Cross-sections were made of a, b the telencephalon at the striatum (blue line), c, d diencephalon at the thalamus (red line), at the transition between diencephalon and mesencephalon (green line), and at the cerebellum and medulla oblongata (black line). Arrows in b and d indicate specific underdeveloped gyri in 1544_FL7. Fetuses and brain cross-sections from the negative control group are displayed for reference

Fig. 5 ZIKV is able to persist in pig placenta following in utero inoculation.

a ZIKV-specific RT-qPCR analysis of fetal brain and placenta samples. The cut-off for the PCR was 1.14 TCID₅₀ equivalent/g tissue. Infectious virus was recovered from the placenta sample from fetus 1543_FL6 (red circle). b IPMA of Vero cells inoculated with placenta sample 1543_FL6. c Representative panel of HE-stained placenta samples derived from the IU-inoculated sows

Fig. 6 Neurodepletion in the parietal lobe following IU inoculation of ZIKV.

a Overview of the localization of the parietal lobe and its cytoarchitecture in the normal fetal pig brain. The different cortical layers (I–VI) are indicated. b Images of Nissl and neurofilament (NF)-stained parietal lobe cross-sections of a negative control fetus and a higher magnification of the pyramidal neurons of cortical layer V. c A fetus presenting moderate neurodepletion in cortical layer V (1544_FL6) and d a fetus presenting severe neurodepletion in cortical layer V with prominent lobe hypoplasia (1543_FL3)

Fig. 7 Neurodepletion in all other lobes following IU inoculation of ZIKV.

Nissl and neurofilament (NF)-stained cross-sections of the cerebral cortical lobes at the striatum a and thalamus b of other inoculated fetuses presenting neurodepletion. The various cerebral lobes are indicated. For reference, images are included which are derived from similar cross-sections and from the same cerebral lobes of negative control fetuses. NF neurofilament