Longitudinal serological and virological study on porcine torovirus (PToV) in piglets from Spanish farms

Abstract

A study was performed to evaluate porcine torovirus (PToV) seroprevalence and infection in three multi-site farms from the North-eastern region of Spain. Serum samples from 120 piglets and faecal samples from 36 piglets were longitudinally collected at 1, 3, 7, 11 and 15 weeks of age. Serum samples from their dams (n=30) were also taken 1-week post-farrowing. PToV antibodies in serum were monitored by ELISA, while viral infection was assessed by real-time RT-PCR in faeces. A high seroprevalence (about 100%) was observed in animals older than 11 weeks and in adult sows. Moreover, all 1-week-old animals were seropositive, indicating maternal antibody transference through colostrum. The antibody titers declined to close to or below the ELISA cut-off value by the age of weaning (3 weeks of age). Development of a significant antibody response to PToV occurred before 7 weeks of age in about 50% of piglets, and the remaining animals developed the response by weeks 11 or 15. These results indicate that PToV infection occurred soon after weaning. Although the prevalence of infection in suckling piglets varied among the studied farms, PToV prevalences in 7 and 11-week-old pigs were between 50-67% and 58-75%, respectively, in all farms. Sequencing results indicated that more than one PToV strains were circulating in the studied farms. Present data suggest that PToV was endemic on the studied farms, and provide new insights on the epidemiology of PToV.

Fig. 1

Longitudinal analysis of IgG response to PToV in pigs from three Spanish herds. Serum samples from 120 piglets and 30 sows were collected from three farms. Samples collected at weeks 1, 3, 7, 11 and 15 of piglet age and samples from sows collected at 1 week post-farrowing were analyzed for the presence of IgG to PToV-N protein by ELISA. (A) Percentages of seropositive (black bars) and seronegative (grey bars) animals over the study period. (B) Mean IgG ELISA values of sows and piglets at each sampling time are represented by grey and black squares, respectively. IgG ELISA cut-off value is indicated (- - -).

Fig. 2

Analysis of the different patterns of IgG antibody development. The dynamics of IgG antibody rising determined by ELISA through weeks in each individual piglet were classified using MeV software using a K-means algorithm and four antibody profiles (1–4) were determined. Antibody profile for each individual piglet is represented by grey lines and the mean ELISA value between all piglets belonging to a profile is represented by a black line.

Fig. 3

Comparison between the levels of IgG and neutralizing antibodies to PToV in nine selected piglets from different antibody dynamics profiles. Serum samples from 9 piglets representative from the groups established based on IgG profile (profiles 1, 3 and 4) were selected (3 piglets per group) and analyzed by virus neutralization test. Neutralizing antibody titers (grey square) were determined in serum samples taken at weeks 1, 3, 7, 11 and 15 of piglet age. Mean IgG ELISA values from the selected piglets from each group are shown (black triangle). Mean sows neutralizing titers (empty square) and ELISA IgG values (empty triangle) for each group is also indicated.

Fig. 4

PToV detection by rt-RT-PCR in farms. Faecal samples were collected from 12 piglets from farms A (black square), B (grey triangle) and C (black dot) at weeks 1, 3, 7, 11 and 15. PToV was detected by rt-RT-PCR using primers rtNII5′ and rtNII3′ to amplify a region from N gene. Number of PToV positive piglets in each farm at different weeks is shown.

Fig. 5

Phylogenetic analysis from partial PToV-N gene sequences obtained from Spanish farms. Gene sequences were aligned using the ClustalW method and the phylogenetic tree was performed by the neighbour joining method. Statistical bootstrap value obtained after 1000 replicates is shown in each branch.

Fig. 6

Phylogenetic analysis from complete HE gene from samples 12.11, 13.11, 14.7, 52.7 and 52.11 from farm B. Gene sequences were aligned using the ClustalW method and the phylogenetic tree was performed by the neighbour joining method. Statistical bootstrap value obtained after 1000 replicates is shown in each branch.