Stage-specific expression of NcSAG4 as a marker of chronic Neospora caninum infection in a mouse model

Abstract

Neospora caninum infection persists throughout the life of its intermediate host due to the conversion of tachyzoites to slowly dividing bradyzoites that encyst in the brain. This event results in persistent N. caninum infection in bovine herds and partially explains the poor efficacy of many chemotherapeutic agents and vaccine formulations. Thus, there is a need for greater understanding of the tachyzoite-to-bradyzoite conversion mechanisms. Here we studied for the first time the transcription kinetics of the N. caninum bradyzoite-specific gene NcSAG4 in brain samples from chronically infected mice by means of real-time RT-PCR. NcSAG4-messenger RNA (mRNA) levels increased significantly during the chronic phase but followed 2 different expression patterns depending on the isolate used for murine inoculation. NcSAG4-mRNA levels in brains from Nc-1-inoculated mice peaked during late chronic infection (on day 64 post-infection, p.i.), whereas those from Nc-Liv-inoculated mice peaked earlier during the chronic infection (on day 32 p.i.). This difference could be a reflection of the different abilities of these isolates to replicate and form cysts in parasitized brains. These results are consistent with our observations of anti-rNcSAG4 antibody production; low levels were present at seroconversion and slowly increased during the chronic phase. In contrast, NcSAG1 transcription levels, which mark the tachyzoite stage, were maintained without variation in both groups of mice. This suggests the presence of a significant amount of tachyzoites or intermediate zoites expressing NcSAG1 in the brain, even during the late chronic infection.

Clinical outcome and vertical transmission variability among canine Neospora caninum isolates in a pregnant mouse model of infection

“…This is supported by the increased levels of specific antibody responses against the recombinant proteins rNcGRA7 and rNcSAG4 in the dams. It has been postulated that, for highly virulent isolates, a higher parasite burden or a more rapid conversion to the bradyzoite stage increases the exposure of mice to these proteins and, consequently, results in strong anti-rNcGRA7 and anti-rNcSAG4 responses (Aguado-Martinez et al 2009;Jimenez-Ruiz et al 2013). The most virulent N. caninum isolates in this study (Nc-Bahia and Nc-Liv) induced the highest antibody levels against rNcGRA7 and rNcSAG4.…”

Detecção de ácidos nucleicos em tecidos de gerbils submetidos à infecção aguda por Neospora caninum

“…Avaliação clínica Os animais foram avaliados diariamente quanto à presença de sinais clínicos como: a inclinação lateral da cabeça, pêlos arrepiados, anorexia, aumento de volume abdominal, prostração, incoordenação motora, paralisia de membros e movimentos circulares (Neosporose Cerebral) (AGUADO-MARTÍNEZ, et al, 2009). …”

“…As consequências clínicas destas duas fases diferem significativamente (AGUADO-MARTÍNEZ et al, 2009). Por essa razão, a determinação de regiões com maior probabilidade de receberem cistos teciduais contendo bradizoítos do protozoário, em tecidos bovinos infectados, torna-se uma ferramenta de fundamental importância para o diagnóstico e compreensão da patogênese da neosporose.…”

Characterisation of NcGRA7 and NcSAG4 proteins: Immunolocalisation and their role in the host cell invasion by Neospora caninum tachyzoites

“…Until now, two bradyzoite-specific proteins of N. caninum have been identified and cloned: NcSAG4 (Fernández-García et al 2006) and NcBSR4 (Risco-Castillo et al 2007). NcSAG4 is expressed early during the process of stage conversion (Fernández-García et al 2006) and has been shown to be a reliable marker for chronic infection (Aguado-Martínez et al 2008, 2009b. Moreover, antibodies against the T. gondii bradyzoite antigen 1 (TgBAG1), specifically expressed in this stage (Bohne et al 1995), cross-react with N. caninum tissue cysts and bradyzoites obtained in vivo and in vitro, respectively (Weiss et al 1999) suggesting that an orthologous protein exists in this parasite.…”