Toxoplasmosis is an immunologically complex disease, particularly in immunocompromised patients. Although there are several therapeutic regimens for such disease, the majority of them have many drawbacks. Therefore, it is of utmost importance to improve the current regimen in an effort to achieve a well-tolerated therapy while also enhancing the host immune response. Famous for their immunomodulatory effect, Lactobacillus delbrueckii and Lactobacillus fermentum probiotics were chosen to be evaluated in this study as an adjuvant therapy against the virulent RH Toxoplasma gondii (T. gondii) strain. Experimental mice were divided into control and treated groups. The control group was further subdivided into two groups: group I: 10 uninfected mice and group II: 20 infected untreated mice. The treated experimental group was subdivided into three groups (20 mice each); group III: sulfamethoxazole-trimethoprim (SMZ-TMP) treated, group IV: probiotics treated, and group V: SMZ-TMP combined with probiotics. The results obtained revealed that combined therapy increased survival rate and time up to 95% and 16 days, respectively, with an 82% reduction of tachyzoites and marked distortion, as detected by the scanning electron microscope (SEM). Additionally, combined therapy alleviated the severity and the extent of the inflammatory cells’ infiltration, thereby reducing hepatocyte degeneration. Intriguingly, serum IF-γ level showed a significant increase to 155.92 ± 10.12 ng/L with combined therapy, reflecting the immunological role of the combined therapy. The current results revealed that probiotics have a high adjuvant potential in alleviating the impact of toxoplasmosis. Using probiotics as a synergistic treatment to modulate conventional therapy in systemic toxoplasmosis may gain popularity due to their low cost and current availability.

Background: Previous studies have reported involvement of Toxoplasma gondii (T. gondii) infections in the pathogenesis of some autoimmune diseases, such as polymyositis, rheumatoid arthritis, autoimmune thyroiditis, and Crohn’s disease. However, data on the association between T. gondii infections and Type 1 diabetes mellitus (T1DM) are still controversial. Therefore, in the present study, we aimed to investigate the pancreatic pathological changes in mouse models with acute and chronic toxoplasmosis and their association with T1DM. Materials and Methods: Three groups (10 mice each) of male Swiss Albino mice were used. One group of mice was left uninfected, whereas the second and third groups were infected with the acute virulent T. gondii RH strain and the chronic less virulent Me49 T. gondii strain, respectively. T. gondii-induced pancreatic pathological changes were evaluated by histopathological examination of pancreatic tissues. Moreover, the expression of insulin, levels of caspase-3, and the pancreatic infiltration of CD8⁺ T cells were evaluated using immunohistochemical staining. Results: Pancreatic tissues of T. gondii-infected animals showed significant pathological alterations and variable degrees of insulitis. Mice with acute toxoplasmosis exhibited marked enlargement and reduced numbers of islets of Langerhans. However, mice with chronic toxoplasmosis showed considerable reduction in size and number of islets of Langerhans. Moreover, insulin staining revealed significant reduction in β cell numbers, whereas caspase-3 staining showed induced apoptosis in islets of Langerhans of acute toxoplasmosis and chronic toxoplasmosis mice compared to uninfected mice. We detected infiltration of CD8⁺ T cells only in islets of Langerhans of mice with chronic toxoplasmosis. Conclusions: Acute and chronic toxoplasmosis mice displayed marked pancreatic pathological changes with reduced numbers of islets of Langerhans and insulin-producing-β cells. Since damage of β cells of islets of Langerhans is associated with the development of T1DM, our findings may support a link between T. gondii infections and the development of T1DM.

Globally, Toxoplasma gondii remains a major One Health challenge, with substantial public health and socioeconomic consequences. Despite its widespread threat, effective control measures are limited by gaps in vaccination strategies. In this study, we experimentally evaluated a Multi-Epitope Vaccine MEV-2A, an in silico-designed multi-epitope recombinant protein incorporating a built-in cholera toxin B subunit as an adjuvant, in addition to B- and T-cell epitopes from seven antigen families. The MEV-2A was expressed, purified, and administered to BALB/c mice (n = 15/group), either alone or with a second adjuvant, either Freund’s or a novel polyethyleneimine-based black seed oil nano-emulsion (PEI/BSO-NE). Specific humoral and cellular immune responses were evaluated, and their protective efficacy was assessed by reduction in parasite DNA burden in peritoneal fluid and survival rate analysis following challenge. The synthesized nano-emulsion (NE) was well dispersed, nano-sized, and had high encapsulation efficiency and pH-responsive release. All three MEV-2A-containing formulations elicited significant specific humoral and cellular immune responses relative to unvaccinated and adjuvant-only controls. They also induced a mixed Th1/ Th2 immune response. Furthermore, they resulted in a notable reduction in parasite DNA burden and prolonged the median survival time. The MEV-2A@Freund’s formulation elicited the highest antibody reactivity, IL-2, and IFN-γ levels, along with 71.38% reduction in parasite DNA burden and median survival of 11 days, whereas the MEV-2A@PEI/BSO-NE induced higher IL-4 and IL-10, 91.28% reduction in parasite DNA burden and median survival of 13 days. These findings identify MEV-2A as a promising vaccine candidate and PEI/BSO-NE as a formulation worth further evaluation. Future studies are warranted to evaluate the protective efficacy of MEV-2A against different genotypes and to comprehensively assess the safety and immunological properties of PEI/BSO-NE as a delivery method and adjuvant platform.