Porcine Circovirus 3 Detection in Aborted Fetuses and Stillborn Piglets from Swine Reproductive Failure Cases

Abstract

Porcine circovirus 3 (PCV-3) has been widely detected in healthy and diseased pigs; among different pathologic conditions, the strongest evidence of association comes from reproductive disease cases. However, simple viral detection does not imply the causality of the clinical conditions. Detection of PCV-3 within lesions may provide stronger evidence of causality. Thus, this study aimed to assess the frequency of PCV-3 detection in tissues from fetuses/stillborn piglets in cases of reproductive problems in domestic swine, as well as the histopathologic assessment of fetal tissues. Fetuses or stillborn piglets from 53 cases of reproductive failure were collected and analyzed by PCV-3 qPCR. The presence of porcine reproductive and respiratory syndrome virus (PRRSV), porcine circovirus 2 (PCV-2), and porcine parvovirus 1 (PPV1) was also checked. PCV-3 qPCR positive samples with a high viral load were tested by PCV-3 in situ hybridization (ISH), sequenced, and phylogenetically analyzed. PCV-3 DNA was detected in 18/53 (33.9%) reproductive failure cases and in 16 of them PCV-3 was the only pathogen found. PCV-2 DNA was found in 5/53 (9.4%), PRRSV RNA in 4/53 (7.5%) and PPV1 was not detected. Four out of the six PCV-3 qPCR-positive cases with Ct value <30 were positive when tested by ISH. In these samples, PCV-3 was detected within mild histopathologic lesions, such as arteritis and periarteritis in multiple tissues. The present work emphasizes the need to include PCV-3 as a potential causative agent of reproductive failure in swine.

Keywords: aborted fetuses; histopathology; in situ hybridization; porcine circovirus 3 (PCV-3); quantitative PCR; reproductive failure; stillborn.

Figure 1

Histology (H&E stain, a,c,e,g) and PCV-3 ISH (hematoxylin counterstain, b,d,f,h) results. (a) Fetal lung from case No. 3 with no apparent histopathologic lesions. (b) PCV-3 genome in macrophage-like cells and in a perivascular area in lung fetus from case No. 3. (c) Moderate tunica media swelling in a splenic artery with a mild lymphocytic infiltration at perivascular and arteriolar locations (arrow) of a stillborn piglet from case No. 52. (d) PCV-3 detection in smooth muscle of spleen artery and perivascular inflammation in the same fetus. (e) Kidney–pelvis artery of a stillborn piglet from case No. 52. (f) PCV-3 detection in smooth muscle and in lamina propria of a kidney artery of the same fetus. (g) Cerebellum from a stillborn piglet with no apparent histopathologic lesions of fetus from case No. 52. (h) High amount of PCV-3 nucleic acid in cerebellum white matter and mild-to-moderate in grey matter of the same animal.

Figure 2

Phylogenetic analysis of PCV-3 complete genome sequences from this study, and the PCV-3 reference sequences from Franzo et al. [31]. The maximum likelihood tree was constructed with the Tamura–Nei model with Gamma plus I distribution (1000 replicates). The width of the branches is proportional to bootstrap values, and the scale bar indicates nucleotide substitutions per site. Sequences obtained in the present study are colored in red; sequences classified as PCV-3a are in the blue shadow, and the sequences of a tentative PCV-3b in orange shadow.