Thomas F. McCutchan’s research while affiliated with Second Military Medical University, Shanghai and other places

Presently the arsenal of anti-malarial drugs is limited and needs to be replenished. We evaluated the potential anti-malarial activity of two water soluble derivatives of nocathiacin (BMS461996 and BMS411886) against asexual blood stages of Plasmodium falciparum. Nocathiacins are a thiazolyl peptide group of antibiotics, structurally related to thiostrepton, have potent activity against a wide spectrum of multi drug resistant gram positive bacteria, and inhibit protein synthesis. We evaluated the potential anti-malarial activity of the derivatives of nocathiacin (BMS461996 and BMS411886) against asexual blood stages of Plasmodium falciparum. The in vitro growth inhibition assay was done using three laboratory strains of P. falciparum displaying varying levels of chloroquine (CQ) susceptibility. Our results indicates that, BMS461996 has potent anti-malarial activity and inhibits parasite growth with mean IC50 of 51.55 nM for P. falciparum 3D7 (CQ susceptible), 85.67 nM for P. falciparum Dd2 (accelerated resistance to multiple drugs, ARMD) and 99.44 nM for P.falciparum K1 strain (resistant to CQ, pyrimethamine and sulfadoxine). Similar results at approximately 7-fold higher IC50 values were obtained with BMS411886 in comparison to BMS461996. We also tested the effect of BMS491996 on gametocytes, our results show that at 20 fold excess of mean IC50 concentration, gametocytes were deformed with a pyknotic nucleus and growth of stage I-IV gametocytes was arrested. This preliminary study shows significant potential for nocathiacin analogues to be developed as anti-malarial drug candidates and warrants further investigation. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

One unique feature of malaria parasites is the differential transcription of structurally distinct rRNA (rRNA) genes at different developmental stages: the A-type genes are transcribed mainly in asexual stages, whereas the S-type genes are expressed mostly in sexual or mosquito stages. Conclusive functional evidence of different rRNAs in regulating stage-specific parasite development, however, is still absent. Here we performed genetic crosses of Plasmodium yoelii parasites with one parent having an oocyst development defect (ODD) phenotype and another producing normal oocysts to identify the gene(s) contributing to the ODD. The parent with ODD—characterized as having small oocysts and lacking infective sporozoites—was obtained after introduction of a plasmid with a green fluorescent protein gene into the parasite genome and subsequent passages in mice. Quantitative trait locus analysis of genome-wide microsatellite genotypes of 48 progeny from the crosses linked an ~200-kb segment on chromosome 6 containing one of the S-type genes (D-type small subunit rRNA gene [D-ssu]) to the ODD. Fine mapping of the plasmid integration site, gene expression pattern, and gene knockout experiments demonstrated that disruption of the D-ssu gene caused the ODD phenotype. Interestingly, introduction of the D-ssu gene into the same parasite strain (self), but not into a different subspecies, significantly affected or completely ablated oocyst development, suggesting a stage- and subspecies (strain)-specific regulation of oocyst development by D-ssu. This study demonstrates that P. yoelii D-ssu is essential for normal oocyst and sporozoite development and that variation in the D-ssu sequence can have dramatic effects on parasite development.

Abstract Plasmodium parasites have recently been documented in the endangered Galapagos penguin (Spheniscus mendiculus). Because avian malaria (caused by Plasmodium spp.) causes high mortality in other avian species after initial exposure, there is great concern for the conservation of the endemic Galapagos penguin. Using a Plasmodium spp. circumsporozoite protein antigen, we have standardized an enzyme-linked immunosorbent assay (ELISA) to test the level of exposure to the parasite in this small population, as indicated by seroprevalence. Sera from adult and juvenile Galapagos penguins collected between 2004 and 2009 on the Galapagos archipelago was tested for the presence of anti - Plasmodium spp. antibodies. Penguins were also tested for prevalence of avian malaria parasite DNA, determined by polymerase chain reaction (PCR) screening. Total seroprevalence of malarial antibodies in this sample group was 97.2%, while total prevalence of Plasmodium parasite DNA by PCR screening was 9.2%. This large discrepancy suggests high exposure to the parasite and low Plasmodium - induced mortality, at least under normal environmental conditions. The results of this study also suggest that parasite prevalence may be under-detected through PCR screening and multiple detection methods are necessary to better understand the extent of Plasmodium exposure on the archipelago.

Can we predict the rise and spread of resistance to multi-drug therapy in a more predictable manner? We raise this question after analyzing over 500 Plasmodium vivax isolates collected from different, geographically isolated regions of China for sequence variation in and around the dhfr and dhps genes. We find: that resistance lineages have arisen at least once in each region; that there appears to have been little movement of parasite populations between these areas; and that highly resistant parasites contain dhfr and dhps alleles that are in linkage disequilibrium. We show a direct relationship between this linkage disequilibrium and a parasite's fitness in the absence of drug pressure. Such fitness would increase the spread of drug resistant phenotypes and is thus a selectable trait. These conclusions raise questions about the appropriate use of some other drug combinations to prevent and treat infection.

Supporting information

Background: γ-irradiation is commonly used to create attenuation in Plasmodium parasites. However, there are no systematic studies on the survival, reversion of virulence, and molecular basis for γ-radiation-induced cell death in malaria parasites. Methods: The effect of γ-irradiation on the growth of asexual Plasmodium falciparum was studied in erythrocyte cultures. Cellular and ultrastructural changes within the parasite were studied by fluorescence and electron microscopy, and genome-wide transcriptional profiling was performed to identify parasite biomarkers of attenuation and cell death. Results: γ-radiation induced the death of P. falciparum in a dose-dependent manner. These parasites had defective mitosis, sparse cytoplasm, fewer ribosomes, disorganized and clumped organelles, and large vacuoles-observations consistent with "distressed" or dying parasites. A total of 185 parasite genes were transcriptionally altered in response to γ-irradiation (45.9% upregulated, 54.1% downregulated). Loss of parasite survival was correlated with the downregulation of genes encoding translation factors and with upregulation of genes associated with messenger RNA-sequestering stress granules. Genes pertaining to cell-surface interactions, host-cell remodeling, and secreted proteins were also altered. Conclusions: These studies provide a framework to assess the safety of γ-irradiation attenuation and promising targets for genetic deletion to produce whole parasite-based attenuated vaccines.

Supplementary Info

Schistosomes, unlike malaria parasites, are in their diploid stage when targeted by the human immune system. Diploids can be either homozygous or heterozygous. The difference has profound significance for developing immunity and yet has not previously been addressed. We examined the implications of zygosity on immunity to a diploid pathogen, Schistosoma japonicum and showed that the diploid state, and its associated heterozygous advantage, significantly affects the outcome of attack by the immune system and the accumulation of antigenic diversity in the parasite population. We demonstrate here that diploidy provides a novel means of immune evasion for diploid pathogens.

Although clinically benign, malaria fever is thought to have significant relevance in terms of parasite growth and survival and its virulence which in turn may alter the clinical course of illness. In this article, the historical literature is reviewed, providing some evolutionary perspective on the genesis and biological relevance of malaria fever, and the available molecular data on the febrile-temperature-inducible parasite factors that may contribute towards the regulation of parasite density and alteration of virulence in the host is also discussed. The potential molecular mechanisms that could be responsible for the induction and regulation of cyclical malaria fevers caused by different species of Plasmodium are also discussed.