Studies on Serological Cross-Reaction of Neospora caninum with Toxoplasma gondii and Hammondia heydorni.

Abstract

ABSTRACT. The enzyme-linked immunosorbent assay (ELISA) was used to examine cross-reactivity of Neospora caninum with Toxoplasma gondii and Hammondia heydorni. Anti-T. gondii mouse and cat sera cross-reacted with N. caninum soluble antigen (NLA), but not with the recombinant surface antigen (NcSRS2). Anti-H. heydorni dog sera showed no cross-reactivity with either the NLA antigen or the NcSRS2. Lack of cross-reactivity between anti-H. heydorni sera and N. caninum antigens, and the cross-reactivity of anti-T. gondii sera with the NLA suggest that N. caninum has common antigens to T. gondii except for NcSRS2 based on serology. In light of several studies suggesting a closer relationship between N. caninum and H. heydorni than with T. gondii, examination of serological cross-reactivity with N. caninum may be necessary to further classify the parasites in addition to molecular and morphological studies and clarification of the life cycle. KEY WORDS: ELISA, Neospora caninum.J. Vet. Med. Sci. 64(2): 161-164, 2002 Neospora caninum is a protozoan parasite of livestock and companion animals that causes abortion, neuromuscular paralysis and death [1,2]. Several studies have reported similarities and differences in the cyst wall morphology, tachyzoites ultrastructure, oocysts features, and life cycle of N. caninum, Toxoplasma gondii, Hammondia heydorni and Hammondia hammondi. The cyst wall of N. caninum bears resemblance to that of T. gondii and H. hammondi [12,13,17,18] [10,13]. In this paper, we investigate whether cross-reactivity occurs between N. caninum, T. gondii and H. heydorni using the surface protein NcSRS2 [5,7] and the soluble antigen (NLA) of N. caninum by using enzyme-linked immunosorbent assay (ELISA).Full-length NcSRS2 genes were cloned into baculovirus using the primer sets: 5'-ACG AAT TCA TGG CGA CGC ATG CTT-3', and 5'-GCG TCG ACT CAG TAC GCA AAG ATT-3'. The cDNA of N. caninum tachyzoites was used as the template in the polymerase chain reaction (PCR). PCR products were blunted by Klenow fragments, and ligated to a previously-digested Sma I baculovirus transfer vector pBacPAK8 (Clontech, Palo Alto, CA). Sf9 cells were co-transfected with a transfer vector and BacuoGold(r) Baculovirus DNA (PharMingen, San Diego, CA) using lipofectin TM reagent (Gibco BRL, Grand Island, NY). After 4 days of incubation at 27°C, the culture supernatant containing recombinant viruses was harvested and subjected to plaque purification. After three cycles of purification, recombinant viruses-expressing NcSRS2 were obtained. Autographa californica nuclear polyhedrosis virus and its recombinant products were grown in Spodoptera frugiperda (Sf9) cells in a TC-100 insect medium (Gibco BRL) supplemented with 10% FBS and 0.26% bacto tryptose broth (Difco, Detroit, MI).ELISA plates coated with the NLA antigen were prepared as described previously by Nishikawa et al. [15]. Monolayers of Sf9 cells that were grown in a 75 cm 2 flask and infected with 10 ml of recombinant baculovirus at 5 PFU/ cell in Sf-900II SFM medium (Gibco BRL). A...

Development of Rapid Immunochromatographic Test with Recombinant NcSAG1 for Detection of Antibodies toNeospora caninumin Cattle

“…2). These results suggest that the ICT with GST-NcSAG1t not only could detect the specific antibodies to N. caninum but also could discriminate between neosporosis and toxoplasmosis, which has been thought to be important because some animals, such as dogs, cattle, sheep, and horses, can be naturally infected with both N. caninum and T. gondii (5,10,11,12,13).…”

Protection against LethalNeospora caninumInfection in Mice Induced by Heterologous Vaccination with amic1 mic3KnockoutToxoplasma gondiiStrain

“…In this study, we have demonstrated that heterologous immunization with an attenuated T. gondii strain induced protec- Previous studies have provided evidence of humoral and cellular cross-reactivity between T. gondii and N. caninum in different hosts (16,19,21,32), but the extent to which T. gondii immunization could protect against subsequent N. caninum infection has never before been investigated using the mouse model. Whereas in a pregnant-ruminant model of N. caninum infection, a T. gondii vaccinal strain failed to induce protection against N. caninum-induced abortion (16), mice infected with N. caninum were protected against lethal toxoplasmosis (19).…”

“…Previous clinical and diagnostic studies have shown that specific antibodies directed against N. caninum or T. gondii crossreact in serological and immunohistochemical tests, suggesting a possible convergence of immune responses during infections with T. gondii and N. caninum (32,38). It has recently been shown that antibodies directed against N. caninum antigens inhibit host cell invasion by both these parasites (22,43).…”

Immunological comparison of 124 isolates of Toxoplasma gondii

“…Serologic cross reactions between T. gondii and Hammondia hammondi and Neospora caninum have been observed in intermediate hosts, but only at less than 1:100 in the dye test (Frenkel and Dubey 1975). Nishikawa et al (2002) found low titer cross-reactions with T. gondii in the ELISA test. Low-titer cross-reactions between N. caninum and T. gondii have also been observed (Dubey et al 1996).…”