Infectivity and Screening of Anti-piperaquine Genes in Mice Infected with Piperaquine-Sensitive and Piperaquine-Resistant Plasmodium berghei
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Piperaquine (PQ) is one of the major components of artemisinin-based combination therapy for malaria. However, the mechanism of PQ resistance has remained unclear.
In this study, we infected mice with PQ-resistant Plasmodium berghei ANKA strain line (PbPQR) or PQ-sensitive P. berghei ANKA strain line (PbPQS) and their survival rates, parasitemia, and spleen sizes were compared. In addition, we constructed genomic DNA subtractive library of spleens from the infected mice, and screened the potential PQ-resistant related genes from genomic DNA of PbPQR line using the representational difference analysis (RDA) method. Clones of the subtractive library were screened by PCR, and related genes were sequenced and analyzed using BLAST software of NCBI.
Compared to PbPQS-infected mice, PbPQR-infected mice survived significantly longer, and had significantly lowered parasitemia rate and significantly increased splenomegaly. Among the total of 502 clones picked, 494 were sequenced and 96 unique PCR fragments were obtained; in which 24 DNA fragments were homologous to chromosomes related to immune function of mice. ORF Finder blasting showed that at the protein level, 26 encoded proteins were homologous to 18 hypothetical PbANKA proteins and 13 encoded proteins were homologous to “ferlin-like protein” family of PbANKA. In addition, there were more immune-related DNA molecules, ubiquitous PbANKA homology at the ORF fragment level, and enriched ferlin-like protein families identified from PbPQR-infected mice than those from PbPQS-infected mice.
These findings suggest that PbPQR may induce stronger protective immune response than that of PbPQS in infected mice.
KeywordsPlasmodium berghei ANKA Piperaquine resistance Subtractive library Representational difference analysis Ferlin-like protein
Research reported in this publication was supported by the Natural Science Foundation of China (no. 81060140). The content is solely the responsibility of the authors
Compliance with Ethical Standards
Conflict of Interest
The authors declare that they have no competing interests.
All experiments were performed in compliance with the requirements of the Animal Ethics Committee at Sun Yat-sen University.
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