A Polyuridine Insertion in the 3′ Untranslated Region of Classical Swine Fever Virus Activates Immunity and Reduces Viral Virulence in Piglets

Abstract

Low-virulence classical swine fever virus (CSFV) strains make CSF eradication particularly difficult. Few data are available on the molecular determinants of CSFV virulence. The aim of the present study was to assess a possible role for CSFV virulence of a unique, uninterrupted 36-uridine (poly-U) sequence found in the 3′ untranslated region (3′ UTR) of the low-virulence CSFV isolate Pinar de Rio (PdR). To this end, a pair of cDNA-derived viruses based on the PdR backbone were generated, one carrying the long poly-U insertion in the 3′ UTR (vPdR-36U) and the other harboring the standard 5 uridines at this position (vPdR-5U). Two groups of 20 5-day-old piglets were infected with vPdR-36U and vPdR-5U. Ten contact piglets were added to each group. Disease progression, virus replication, and immune responses were monitored for 5 weeks. The vPdR-5U virus was significantly more virulent than the vPdR-36U virus, with more severe disease, higher mortality, and significantly higher viral loads in serum and body secretions, despite similar replication characteristics in cell culture. The two viruses were transmitted to all contact piglets. Ninety percent of the piglets infected with vPdR-36U seroconverted, while only one vPdR-5U-infected piglet developed antibodies. The vPdR-5U-infected piglets showed only transient alpha interferon (IFN-α) responses in serum after 1 week of infection, while the vPdR-36U-infected piglets showed sustained IFN-α levels during the first 2 weeks. Taken together, these data show that the 3′ UTR poly-U insertion acquired by the PdR isolate reduces viral virulence and activates the innate and humoral immune responses without affecting viral transmission. IMPORTANCE Classical swine fever (CSF), a highly contagious viral disease of pigs, is still endemic in some countries of Asia and Central and South America. Considering that the 3′ untranslated region (3′ UTR) plays an important role in flavivirus replication, the present study showed for the first time that a long polyuridine sequence acquired in the 3′ UTR by an endemic CSFV isolate can activate immunity, control viral replication, and modulate disease in piglets. Our findings provide new avenues for the development of novel vaccines against infections with CSF virus and other flaviviruses. Knowledge of molecular virulence determinants is also relevant for future development of rapid and efficient diagnostic tools for the prediction of the virulence of field isolates and for efficient CSF control.

Foetal Immune Response Activation and High Replication Rate during Generation of Classical Swine Fever Congenital Infection

“…The capacity for trans-placental transmission and induction of foetal immune response was compared side by side between the high and low virulence strains (Figures 1 and 2, Tables 1 and 2). In accordance with previous data found in piglets, the highly virulent CSFV Margarita strain induced high serum IFN-α levels in sows over a short period of time [23]. By contrast, the IFN-α response induced by the low virulence PdR strain was lower, although it lasted one week longer.…”

“…The highest IFN-α response was found in the viraemic foetuses or in those that showed higher viral replication in organs from the group infected with the low virulence PdR strain. Notably, the capacity of the PdR strain for high and prolonged IFN-α activation in piglets has been associated with an uninterrupted 36-uridine sequence found in the 3 untranslated region of the CSFV genome [23]. Activation of IFN-α response in ruminant and human foetuses, following infection with bovine viral diarrhoea and Zika virus, respectively, has been described, and it may support the results obtained in this study [37,38].…”

“…Samples were considered positive when the Ct values were equal to or less than 42. In addition, using the Ct value, samples were determined to have either high (Ct value below 23), moderate (between 23 and 28), or low (Ct value above 28) CSFV RNA load, as previously described [23,50]. Samples in which fluorescence was undetectable (Undet) were considered negative.…”

High-Resolution Profiling of Innate Immune Responses by Porcine Dendritic Cell Subsets in vitro and in vivo

“…The observed differences might reflect exhaustion of pDC during infection with the highly virulent strain. On the other hand, it should be pointed out that the PdR strain has a unique poly-uridine sequence of the 3′UTR, which possibly promotes pDC activation ( 8 , 60 ).…”

Early and Solid Protection Afforded by the Thiverval Vaccine Provides Novel Vaccination Alternatives Against Classical Swine Fever Virus

“…The animals were monitored, and rectal temperature was measured daily by a trained veterinarian starting on the day of vaccination and until the end of the trial. A clinical score from 0 to 6 was assigned according to the clinical status of the animals, as previously described [ 20 , 21 ]: 0, no signs; 1, mild fever; 2, mild to moderate clinical signs; 3, moderate clinical signs; 4, moderate to severe clinical signs; 5, severe clinical signs; and 6, death. Serum and nasal and rectal swabs were collected on the day of vaccination for all the groups and at 4, 7, and 14 dpv for groups A and B.…”