Toxoplasma gondii Tetravalent Chimeric Proteins as Novel Antigens for Detection of Specific Immunoglobulin G in Sera of Small Ruminants

Abstract

The detection of Toxoplasma gondii infection in small ruminants has important significance for public health and veterinary medicine. This study, for the first time, describes the reactivity of four tetravalent chimeric proteins (AMA1N-SAG2-GRA1-ROP1, AMA1C-SAG2-GRA1-ROP1, AMA1-SAG2-GRA1-ROP1, and SAG2-GRA1-ROP1-GRA2) containing immunodominant regions from the AMA1 (apical membrane antigen 1), SAG2 (surface antigen 2), GRA1 (dense granule antigen 1), GRA2 (dense granule antigen 2), and ROP1 (rhoptry antigen 1) with specific IgG antibodies from the sera of small ruminants with the use of an indirect enzyme-linked immunosorbent assay (ELISA). The reactivity of individual chimeric antigens was analyzed in relation to the results obtained in IgG ELISA based on a Toxoplasma lysate antigen (TLA). All chimeric proteins were characterized by high specificity (between 96.39% to 100%), whereas the sensitivity of the IgG ELISAs was variable (between 78.49% and 96.77%). The highest sensitivity was observed in the IgG ELISA test based on the AMA1-SAG2-GRA1-ROP1. These data demonstrate that this chimeric protein can be a promising serodiagnostic tool for T. gondii infection in small ruminants.

Descriptive Comparison of ELISAs for the Detection of Toxoplasma gondii Antibodies in Animals: A Systematic Review

“…Out of 57 studies describing the evaluation of diagnostic performance of different ELISAs for the detection of T. gondii antibodies, the majority originated from Europe ( n = 21), followed by Asia ( n = 16), South America ( n = 9), North America ( n = 6), Africa ( n = 3) and Australia ( n = 2) ( Figure 2 ). Out of the 20 animal species covered, most of the studies (91%) focused on domesticated animals [ 33 , 37 , 46 , 47 , 48 , 49 , 50 ], but 7% were on zoo and wild animals [ 28 , 44 , 51 , 52 ], and one study targeted both domestic and wild animals [ 43 ]. Serum was the most widely used (89% of studies) sample for the detection of T. gondii -specific antibodies in animals, followed by meat juice/tissue fluid (7%) [ 37 , 45 , 53 , 54 ] and milk (4%) [ 55 , 56 ].…”

“…Three main types of antigen, including native (66% of the 50 studies) [ 28 , 35 , 43 , 51 , 61 ], recombinant (30%) [ 24 , 44 , 50 , 57 , 62 ], and chimeric proteins (4%) [ 7 , 48 ], were used for the detection of T. gondii -specific antibodies in animals. Seven studies did not clearly indicate the type of antigen used, and hence were not included in the timeline below ( Figure 3 ).…”

“…However, recombinant and chimeric antigens possess other advantages over native antigens, including the ease of production, reduced cost, and less exposure to biohazardous procedures [ 88 ]. Moreover, assay optimisation and standardisation are easier and researchers have the freedom to precisely construct the required antigen composition of interest [ 48 , 88 ]. However, there are some disadvantages, including inefficient expression and misfolding of proteins during the production process inside the prokaryotic systems, which could affect the affinity of the assay [ 89 ].…”

“…Infected hosts mount varying humoral immune responses depending on the infective stage of the parasite; thus, a single recombinant antigen is probably not capable of binding with all stage-specific antibodies [ 70 ]. In contrast, a combination of different recombinant antigens may allow T. gondii -specific antibodies to recognise multiple epitopes from different parasitic stages [ 48 , 70 ]. Based on the diagnostic performance reported, the combination of SAG and GRA antigens, including SAG1, SAG2, and GRA7, may provide better diagnostic performance.…”

Toxoplasma gondii Recombinant Antigens in the Serodiagnosis of Toxoplasmosis in Domestic and Farm Animals

“…Only a few publications raised the issue of cross-reactions between T. gondii and N. caninum , which are the most antigenically similar among Apicomplexa parasites. In even fewer works, N. caninum seropositive sera were used to determine cut-off values or as controls [ 80 , 84 , 85 , 89 , 91 ]. Apicomplexa-type parasites are antigenically similar and additional tests on sera from animals infected with other parasites must be performed to fully assess the diagnostic potential of recombinant antigens, in particular, the species Neospora sp.…”