Antibody and cytokine response to Cystoisospora suis infections in immune-competent young pigs

Abstract

Background: To date, investigations on the immune response to Cystoisospora suis infections focused on suckling piglets, the age group clinically most affected. Actively immunizing piglets is unfeasible due to their immature immune system and the typically early infection in the first days after birth. Therefore, understanding and possibly enhancing the immune response of immune-competent animals is the prerequisite to develop a passive immunization strategy for piglets which currently rely on very limited treatment options.

Methods: To investigate antibody and cytokine responses of immune-competent animals and the impact of the oral immunization protocol on their immune response, growers with unknown previous exposure to C. suis (10-11 weeks-old) were infected one or three times with different doses (600 and 6000 or 200 and 2000, respectively) of C. suis oocysts, and compared to uninfected controls. Oocyst excretion was evaluated, and blood and intestinal mucus antibody titers were determined by IFAT. Systemic production of Th1, Th2, inflammatory and regulatory cytokines was determined in different immune compartments at mRNA and (after stimulation with a recombinant merozoite-protein) at protein level by PCR and multiplex fluorescent immunoassay, respectively.

Results: Infection generated significantly increased serum IgA and IgG levels against C. suis sporozoites and merozoites, irrespective of infection mode, with IgG against merozoites showing the strongest increase. No clinical signs and only occasional excretion were observed. The systemic cytokine response to C. suis was only weak. Nonetheless, in white blood cells, IL-4, IL-6 and IL-10 mRNA-levels significantly increased after infection, whereas IFN-ɣ, IL-2 and TGF-β expression tended to decrease. In mesenteric lymph nodes (MLN), IL-10 and TNF-α levels were elevated while splenic cytokine expression was unaltered upon infection. Stimulated MLN-derived lymphocytes from infected pigs produced slightly more IL-12 and less IFN-α than controls.

Conclusions: An infection and a subsequent systemic immune response can be induced in immune-competent animals by all evaluated infection models and growers can be used as models to mimic sow immunizations. The immune response to C. suis, although mild and with considerable variation in cytokine expression, was characterized by a Th2-associated and regulatory cytokine profile and antibody production. However, none of the parameters clearly stood out as a potential marker associated with protection. Antibody titers were significantly positively related with oocyst excretion and might thus serve as correlates for parasite replication or severity of infection.

Keywords: Antibody; Coccidiosis; Cytokine; Immunity; Immunization; Pig; Stimulation; T helper 2.

Fig. 1

Development of mean blood serum antibody titers (IgG, IgA) against C. suis sporozoites and merozoites after infection. Titers were converted to numerics, starting with 1 representing a titer of 1:20, followed by 2 representing 1:40, and so on. Means and standard deviations are given for each group on each study day. Study day 1: day of (first) infection. Abbreviations: SL, Single Low; SH, Single High; TL, Trickle Low; TH, Trickle High; C, Control

Fig. 2

Cytokine mRNA expression of white blood cells after infection with C. suis and sham-treatment, respectively. Each sample from study day 8, 15 and 29 was normalized to its corresponding sample from study day 1 [i.e. before (first) infection]. Thus, fold expression values are given relative to values of this time point. Length of boxes indicates the interquartile range; the embedded line represents the median. Whiskers extend to the largest and smallest values still within 1.5 times the length of the interquartile range from the upper and lower quartiles, respectively. Data lying outside this range are plotted individually as dots. Abbreviations: SL, Single Low; SH, Single High; TL, Trickle Low; TH, Trickle High; C, Control

Fig. 3

Mean IFN-α (a) and IL-12 (b) concentrations in supernatants of stimulated lymphocytes. MLN-derived lymphocytes (extracted on SD 29) were stimulated with rCSUI_005805 [44]. Adjusted values (cytokine concentrations of unstimulated controls were subtracted from those of stimulated cells) are given. Length of boxes: interquartile range; embedded line: median. Whiskers extend to the largest and smallest values within 1.5 times the length of the interquartile range from the upper and lower quartiles, respectively. Data lying outside this range are plotted as dots