Specific tagging of the egress-related osmiophilic bodies in the gametocytes of Plasmodium falciparum

<p>Abstract</p> <p>Background</p> <p>Gametocytes, the blood stages responsible for <it>Plasmodium falciparum </it>transmission, contain electron dense organelles, traditionally named osmiophilic bodies, that are believed to be involved in gamete egress from the host cell. In order to provide novel tools in the cellular and molecular studies of osmiophilic body biology, a <it>P. falciparum </it>transgenic line in which these organelles are specifically marked by a reporter protein was produced and characterized.</p> <p>Methodology</p> <p>A <it>P. falciparum </it>transgenic line expressing an 80-residue N-terminal fragment of the osmiophilic body protein Pfg377 fused to the reporter protein DsRed, under the control of <it>pfg377 </it>upstream and downstream regulatory regions, was produced.</p> <p>Results</p> <p>The transgenic fusion protein is expressed at the appropriate time and stage of sexual differentiation and is trafficked to osmiophilic bodies as the endogenous Pfg377 protein. These results indicate that a relatively small N-terminal portion of Pfg377 is sufficient to target the DsRed reporter to the gametocyte osmiophilic bodies.</p> <p>Conclusions</p> <p>This is the first identification of a <it>P. falciparum </it>aminoacid sequence able to mediate trafficking to such organelles. To fluorescently tag such poorly characterized organelles opens novel avenues in cellular and imaging studies on their biogenesis and on their role in gamete egress.</p

Antiplasmodial Effects of a few Selected Natural Flavonoids and their Modulation of Artemisinin Activity

The direct antiplasmodial effects of five structurally-related flavonoids, namely quercetin, rutin, eriodictyol, eriodictyolchalcone and catechin, were analyzed in vitro on P. falciparum. Notably, all these flavonoids, with the only exception of rutin, caused relevant inhibition of P. falciparum growth when given at 1 mM concentration. In addition, they were found to affect greatly the potent antiplasmodial activity of artemisinin, leading to significant additive and even synergistic effects. In particular, quercetin induced a pronounced synergistic effect. The observed synergisms might be conveniently exploited to design new and/or more effective combination therapies

Multiple sclerosis: peripheral mononuclear cells inhibit <i>Plasmodium falciparum</i> growth and are activated by parasite antigens

The human genome has been subjected to selective pressures to resist to infectious agents in spite of a heavy segregational load. With this regard, thalassaemia and glucose-6-phosphate dehydrogenase deficiency have been considered an efficient genetic protection against P. falciparum malaria in Sardinia, insular Italy. In this island, some multiple sclerosis (MS)-associated HLA haplotypes have the highest odds ratios in the same highestrate malarious areas of the island. Moreover, tumor necrosis factor (TNF) polymorphisms epidemiologically associated with both MS and malaria are ten-fold more frequent amongst Sardinians compared to other populations worldwide4. A possible association between MS and malaria in this island was never analysed experimentally. We studied the immunological response of mononuclear cells to P. falciparum and the killing effect of macrophages on parasites in Sardinian MS patients and in matched healthy controls (HC)

Recent genetic exchanges and admixture shape the genome and population structure of the zoonotic pathogen Cryptosporidium parvum

Cryptosporidium parvum is a globally distributed zoonotic pathogen and a major cause of diarrhoeal disease in humans and ruminants. The parasite's life cycle comprises an obligatory sexual phase, during which genetic exchanges can occur between previously isolated lineages. Here, we compare 32 whole genome sequences from human- and ruminant-derived parasite isolates collected across Europe, Egypt and China. We identify three strongly supported clusters that comprise a mix of isolates from different host species, geographic origins, and subtypes. We show that: (1) recombination occurs between ruminant isolates into human isolates; (2) these recombinant regions can be passed on to other human subtypes through gene flow and population admixture; (3) there have been multiple genetic exchanges, and most are probably recent; (4) putative virulence genes are significantly enriched within these genetic exchanges, and (5) this results in an increase in their nucleotide diversity. We carefully dissect the phylogenetic sequence of two genetic exchanges, illustrating the long-term evolutionary consequences of these events. Our results suggest that increased globalization and close human-animal contacts increase the opportunity for genetic exchanges between previously isolated parasite lineages, resulting in spillover and spillback events. We discuss how this can provide a novel substrate for natural selection at genes involved in host–parasite interactions, thereby potentially altering the dynamic coevolutionary equilibrium in the Red Queens arms race

Comparative genomics revealed adaptive admixture in Cryptosporidium hominis in Africa

Cryptosporidiosis is a major cause of diarrhoeal illness among African children, and is associated with childhood mortality, malnutrition, cognitive development and growth retardation. Cryptosporidium hominis is the dominant pathogen in Africa, and genotyping at the glycoprotein 60 (gp60) gene has revealed a complex distribution of different subtypes across this continent. However, a comprehensive exploration of the metapopulation structure and evolution based on whole-genome data has yet to be performed. Here, we sequenced and analysed the genomes of 26 C. hominis isolates, representing different gp60 subtypes, collected at rural sites in Gabon, Ghana, Madagascar and Tanzania. Phylogenetic and cluster analyses based on single-nucleotide polymorphisms showed that isolates predominantly clustered by their country of origin, irrespective of their gp60 subtype. We found a significant isolation-by-distance signature that shows the importance of local transmission, but we also detected evidence of hybridization between isolates of different geographical regions. We identified 37 outlier genes with exceptionally high nucleotide diversity, and this group is significantly enriched for genes encoding extracellular proteins and signal peptides. Furthermore, these genes are found more often than expected in recombinant regions, and they show a distinct signature of positive or balancing selection. We conclude that: (1) the metapopulation structure of C. hominis can only be accurately captured by whole-genome analyses; (2) local anthroponotic transmission underpins the spread of this pathogen in Africa; (3) hybridization occurs between distinct geographical lineages; and (4) genetic introgression provides novel substrate for positive or balancing selection in genes involved in host–parasite coevolution

A retrospective molecular study of Cryptosporidium species and genotypes in HIV-infected patients from Thailand

Abstract Background Opportunistic infections represent a serious health problem for HIV-infected people. Among enteric infections, cryptosporidiosis, a severe and life-threatening diarrheal disease, is of particular importance in low economic settings where access to anti-retroviral therapy is limited. Understanding transmission routes is crucial in establishing preventive measures, and requires the use of informative genotyping methods. In this study, we performed a retrospective analysis of Cryptosporidium species in 166 stool samples collected from 155 HIV-infected patients during 1999–2004 at the Siriraj Hospital in Bangkok, Thailand. Results Microscopic examination of stools identified 104 of the 155 patients as positive for Cryptosporidium. Other common pathogens identified were microsporidia, Isospora, Giardia, Strongyloides and Opisthorchis. All samples were tested by amplification of a fragment of the 18S rDNA locus, and sequencing showed the presence of Cryptosporidium hominis (n = 42), C. meleagridis (n = 20), C. canis (n = 12), C. felis (n = 7), C. suis (n = 6) and C. parvum (n = 5). Genotyping at the glycoprotein 60 (gp60) locus revealed substantial variability in isolates of C. hominis and C. meleagridis. Among C. hominis isolates, subtype IeA11G3T3 was the most prevalent, but allelic family Id was the more diverse with four subtypes described, two of which were identified for the first time. Among C. meleagridis isolates, seven subtypes, two of which were new, were found in the allelic family IIIb, along with new subtypes in allelic families IIIe and IIIg. In the four C. parvum isolates, subtype IIoA16G1, a rare subtype previously reported in a Swedish patient who had traveled to Thailand, was identified. Conclusions This study confirms the high susceptibility of HIV-infected individuals to infection with different Cryptosporidium species and subtypes, and further stresses the importance of surveillance for opportunistic intestinal protozoans

First molecular investigation of Cryptosporidium spp. in young calves in Algeria

To date, no information is available on the prevalence and genetic identity of Cryptosporidium spp. in cattle in Algeria. In this study, 17 dairy farms in the province of Batna, located in the northeast of the country, were visited to collect 132 fecal samples from young calves (< 8 weeks old). Samples were examined microscopically using the modified Ziehl-Neelsen acid-fast staining method, and at least one sample per farm was submitted for molecular analysis. Amplification of a fragment of the small subunit ribosomal RNA gene was positive for 24 of the 61 samples (40%), and sequence analysis identified three species, namely Cryptosporidium bovis (n = 14), C. ryanae (n = 6), and C. parvum (n = 4). The C. parvum IIaA13G2R1 subtype, an uncommon zoonotic subtype, was identified in two isolates from a single farm by sequencing a fragment of the GP60 gene. This is the first report about genotyping and subtyping of Cryptosporidium in calves in Algeria

First molecular investigation of

To date, no information is available on the prevalence and genetic identity of Cryptosporidium spp. in cattle in Algeria. In this study, 17 dairy farms in the province of Batna, located in the northeast of the country, were visited to collect 132 fecal samples from young calves (< 8 weeks old). Samples were examined microscopically using the modified Ziehl-Neelsen acid-fast staining method, and at least one sample per farm was submitted for molecular analysis. Amplification of a fragment of the small subunit ribosomal RNA gene was positive for 24 of the 61 samples (40%), and sequence analysis identified three species, namely Cryptosporidium bovis (n = 14), C. ryanae (n = 6), and C. parvum (n = 4). The C. parvum IIaA13G2R1 subtype, an uncommon zoonotic subtype, was identified in two isolates from a single farm by sequencing a fragment of the GP60 gene. This is the first report about genotyping and subtyping of Cryptosporidium in calves in Algeria