Role of alarmin cytokines and microRNAs in the host-schistosome interaction [version 1; referees: 3 approved]

Schistosomiasis is a serious but neglected tropical infectious disease, afflicting more than 240 million people in 78 countries. Lack of an effective vaccine and obscuring disease mechanism could be the main hurdles to effectively control and eradicate this disease. A better understanding of the host–schistosome interaction is the key to clearing these hurdles. Recently, accumulating evidence shows that alarmin cytokines and microRNAs (miRNAs) are crucial regulators in the host–schistosome interaction. Alarmin cytokines are proven to be potent mechanisms driving type 2 immunity, which is the central disease mechanism of schistosomiasis. MiRNA deregulation is a hallmark of a variety of human diseases, including schistosomiasis. In this review, we summarize the research advances on the role of alarmin cytokines and miRNAs in the host–schistosome interaction

No miRNA were found in Plasmodium and the ones identified in erythrocytes could not be correlated with infection

<p>Abstract</p> <p>Background</p> <p>The transcriptional regulation of <it>Plasmodium </it>during its complex life cycle requires sequential activation and/or repression of different genetic programmes. MicroRNAs (miRNAs) are a highly conserved class of non-coding RNAs that are important in regulating diverse cellular functions by sequence-specific inhibition of gene expression. What is know about double-stranded RNA-mediated gene silencing (RNAi) and posttranscriptional gene silencing (PTGS) in <it>Plasmodium </it>parasites entice us to speculate whether miRNAs can also function in <it>Plasmodium</it>-infected RBCs.</p> <p>Results</p> <p>Of 132 small RNA sequences, no <it>Plasmodium</it>-specific miRNAs have been found. However, a human miRNA, miR-451, was highly expressed, comprising approximately one third of the total identified miRNAs. Further analysis of miR-451 expression and malaria infection showed no association between the accumulation of miR-451 in <it>Plasmodium falciparum</it>-iRBCs, the life cycle stage of <it>P. falciparum </it>in the erythrocyte, or of <it>P. berghei </it>in mice. Moreover, treatment with an antisense oligonucleotide to miR-451 had no significant effect on the growth of the erythrocytic-stage <it>P. falciparum</it>.</p> <p>Methods</p> <p>Short RNAs from a mixed-stage of <it>P. falciparum</it>-iRBC were separated in a denaturing polyacrylamide gel and cloned into T vectors to create a cDNA library. Individual clones were then sequenced and further analysed by bioinformatics prediction to discover probable miRNAs in <it>P. falciparum</it>-iRBC. The association between miR-451 expression and the parasite were analysed by Northern blotting and antisense oligonucleotide (ASO) of miR-451.</p> <p>Conclusion</p> <p>These results contribute to eliminate the probability of miRNAs in <it>P. falciparum</it>. The absence of miRNA in <it>P. falciparum </it>could be correlated with absence of argonaute/dicer genes. In addition, the miR-451 accumulation in <it>Plasmodium</it>-infected RBCs is independent of parasite infection. Its accumulation might be only the residual of erythroid differentiation or a component to maintain the normal function of mature RBCs.</p

Solution structure of a Plasmodium falciparum AMA-1/MSP 1 chimeric protein vaccine candidate (PfCP-2.9) for malaria

Background: The Plasmodium falciparum chimeric protein PfCP-2.9 is a promising asexual-stage malaria vaccine evaluated in clinical trials. This chimeric protein consists of two cysteine-rich domains: domain III of the apical membrane antigen 1 (AMA-1 [III]) and the C-terminal region of the merozoite surface protein 1 (MSP1-19). It has been reported that the fusion of these two antigens enhanced their immunogenicity and antibody-mediated inhibition of parasite growth in vitro. Methods: The N-15-labeled and C-13/N-15-labeled PfCP-2.9 was produced in Pichia pastoris for nuclear magnetic resonance (NMR) structure analysis. The chemical shift assignments of PfCP-2.9 were compared with those previously reported for the individual domains (i.e., PfAMA-1(III) or PfMSP 1-19). The two-dimensional spectra and transverse relaxation rates (R-2) of the PfMSP1-19 alone were compared with that of the PfCP-2.9. Results: Confident backbone assignments were obtained for 122 out of 241 residues of PfCP-2.9. The assigned residues in PfCP-2.9 were very similar to those previously reported for the individual domains. The conformation of the PfMSP1-19 in different constructs is essentially the same. Comparison of transverse relaxation rates (R-2) strongly suggests no weak interaction between the domains. Conclusions: These data indicate that the fusion of AMA-1(III) and MSP1-19 as chimeric protein did not change their structures, supporting the use of the chimeric protein as a potential malaria vaccine.Infectious DiseasesParasitologyTropical MedicineSCI(E)5ARTICLEnull

Effects of Polymorphisms in the SjSP-13 Gene of Schistosoma japonicum on Its Diagnostic Efficacy and Immunogenicity

Schistosomiasis japonica is one of the most prevalent parasitic diseases in China. The scarcity of effective diagnostic tools is a major factor that contributes to the high prevalence of schistosomiasis japonica. SjSP-13 is a promising serological diagnostic biomarker of the disease. However, it is unclear whether polymorphisms in SjSP-13 affect its diagnostic efficacy and immunogenicity. Here, we found the SjSP-13 gene was highly polymorphic, and all the alleles of the gene were clustered into two clades, clade A and B. SjSP-13.6 and SjSP-13.25, the representative alleles of clade A and B, were produced in Escherichia coli. The diagnostic value of SjSP-13.6 (AUC = 0.983 ± 0.006), was found to be similar to the SjSP-13.25 (AUC = 0.973 ± 0.009) by receiver operating characteristic (ROC) analysis. SjSP-13.6 and SjSP-13.25 have the same specificity (96.7%), while the sensitivity of SjSP-13.6 (90.4%) is slightly but not significantly higher than SjSP-13.25 (85.2%). The combination use of the two alleles (SjSP-13.6/25) didn’t increase the diagnostic performance of SjSP-13 as the AUC value of SjSP-13.6/25 is 0.977 ± 0.009, lower than individual SjSP-13.6 (AUC = 0.983 ± 0.006). In addition, we found the immunogenicity of clade A alleles is significantly higher than clade B in Schistosoma japonicum naturally infected animals and patients, as the mean antibody levels of SjSP-13.6 was significantly higher than SjSP-13.25. We conclude that polymorphisms of the SjSP-13 gene should not affect its diagnostic efficacy, and it is not necessary to combine the alleles of the two clades for diagnosis of schistosomiasis

High-throughput sequencing of RNAs isolated by cross-linking immunoprecipitation (HITS-CLIP) reveals Argonaute-associated microRNAs and targets in Schistosoma japonicum

Sequences of SjAgo-associated novel miRNAs by the HITS-CLIP assay. (XLSX 16 kb

Epitope mapping of PfCP-2.9, an asexual blood-stage vaccine candidate of Plasmodium falciparum

<p>Abstract</p> <p>Background</p> <p>Apical membrane antigen 1 (AMA-1) and merozoite surface protein 1 (MSP1) of <it>Plasmodium falciparum </it>are two leading blood-stage malaria vaccine candidates. A <it>P. falciparum </it>chimeric protein 2.9 (PfCP-2.9) has been constructed as a vaccine candidate, by fusing AMA-1 domain III (AMA-1 (III)) with a C-terminal 19 kDa fragment of MSP1 (MSP1-19) via a 28-mer peptide hinge. PfCP-2.9 was highly immunogenic in animal studies, and antibodies elicited by the PfCP-2.9 highly inhibited parasite growth <it>in vitro</it>. This study focused on locating the distribution of epitopes on PfCP-2.9.</p> <p>Methods</p> <p>A panel of anti-PfCP-2.9 monoclonal antibodies (mAbs) were produced and their properties were examined by Western blot as well as <it>in vitro </it>growth inhibition assay (GIA). In addition, a series of PfCP-2.9 mutants containing single amino acid substitution were produced in <it>Pichia pastoris</it>. Interaction of the mAbs with the PfCP-2.9 mutants was measured by both Western blot and enzyme-linked immunosorbent assay (ELISA).</p> <p>Results</p> <p>Twelve mAbs recognizing PfCP-2.9 chimeric protein were produced. Of them, eight mAbs recognized conformational epitopes and six mAbs showed various levels of inhibitory activities on parasite growth <it>in vitro</it>. In addition, seventeen PfCP-2.9 mutants with single amino acid substitution were produced in <it>Pichia pastoris </it>for interaction with mAbs. Reduced binding of an inhibitory mAb (mAb7G), was observed in three mutants including M62 (Phe⁴⁹¹→Ala), M82 (Glu⁵¹¹→Gln) and M84 (Arg⁵¹³→Lys), suggesting that these amino acid substitutions are critical to the epitope corresponding to mAb7G. The binding of two non-inhibitory mAbs (mAbG11.12 and mAbW9.10) was also reduced in the mutants of either M62 or M82. The substitution of Leu³¹to Arg resulted in completely abolishing the binding of mAb1E1 (a blocking antibody) to M176 mutant, suggesting that the Leu residue at this position plays a crucial role in the formation of the epitope. In addition, the Asn¹⁵residue may also play an important role in the global folding of PfCP-2.9, as its substitution by Arg lead to reduced binding of most mAbs and abolishing the binding of mAb6G and mAbP5-W12.</p> <p>Conclusions</p> <p>This study provided valuable information on epitopes of PfCP-2.9 vaccine candidate through generation of a panel of mAbs and a series of PfCP-2.9 mutants. The information may prove to be useful for designing more effective malaria vaccines against blood-stage parasites.</p