Identification and real-time expression analysis of selected Toxoplasma gondii in-vivo induced antigens recognized by IgG and IgM in sera of acute toxoplasmosis patients

BMC Infectious Diseases

Rahmah Noordin²  Muhammad Hafiznur Yunus²  Sabariah Osman²  Izzati Zahidah Abdul Karim²  Majid Golkar¹  Ai Ying Teh²  Boon Yin Khoo²  Atefeh Amerizadeh² 

[1] Parasitology Department, Molecular Parasitology Laboratory, Pasteur Institute of Iran, Tehran, Iran;Institute for Research in Molecular Medicine (INFORMM), Universiti Sains Malaysia (USM), Penang 11800, Malaysia

关键词: Real-time polymerase chain reaction (PCR);    mRNA expression analysis;    Acute toxoplasmosis sera;    cDNA library immunoscreening;    In-vivo induced antigen technology (IVIAT);    Toxoplasma gondii;   

Others  :  1147517
DOI  :  10.1186/1471-2334-13-287

 received in 2013-03-20, accepted in 2013-06-13,  发布年份 2013

【 摘 要 】

Background

Toxoplasma gondii is an obligate intracellular zoonotic parasite of the phylum Apicomplexa which infects a wide range of warm-blooded animals, including humans. In this study in-vivo induced antigens of this parasite was investigated using in-vivo induced antigen technology (IVIAT) and pooled sera from patients with serological evidence of acute infection.

Methods

The pooled sera was first pre-absorbed against three different preparations of antigens from in-vitro-grown cells of each T. gondii and E. coli XL1-Blue MRF’, subsequently it was used to screen T. gondii cDNA phage expression library. Positive clones from each group were subjected to quantitative real-time PCR expression analysis on mRNA of in-vivo and in-vitro grown parasites.

Results

A total of 29 reactive clones from each IgM and IgG immunoscreenings were found to have high homology to T. gondii genes. Quantitative real-time PCR expression analysis showed that 20 IgM-detected genes and 11 IgG-detected genes were up-regulated in-vivo relative to their expression levels in-vitro. These included genes encoding micronemes, sterol-regulatory element binding protein site, SRS34A, MIC2-associated protein M2AP, nucleoredoxin, protein phosphatase 2C and several hypothetical proteins. A hypothetical protein (GenBank accession no. 7899266) detected by IgG had the highest in-vivo over in-vitro fold change of 499.86; while another up-regulated hypothetical protein (GenBank accession no. 7898829) recognized by IgM showed high sensitivity (90%) and moderate specificity (70%) in detecting T. gondii antibodies when tested with 20 individual serum samples.

Conclusion

The highly up-regulated genes and the corresponding proteins, in particular the hypothetical proteins, may be useful in further studies on understanding the disease pathogenesis and as potential vaccine candidates.

【 授权许可】

   
2013 Amerizadeh et al.; licensee BioMed Central Ltd.

【 图 表 】

Figure 1.

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