Single-molecule analysis reveals that DNA replication dynamics vary across the course of schizogony in the malaria parasite Plasmodium falciparum.

The mechanics of DNA replication and cell cycling are well-characterized in model organisms, but less is known about these basic aspects of cell biology in early-diverging Apicomplexan parasites, which do not divide by canonical binary fission but undergo unconventional cycles. Schizogony in the malaria parasite, Plasmodium, generates ~16-24 new nuclei via independent, asynchronous rounds of genome replication prior to cytokinesis and little is known about the control of DNA replication that facilitates this. We have characterised replication dynamics in P. falciparum throughout schizogony, using DNA fibre labelling and combing to visualise replication forks at a single-molecule level. We show that origins are very closely spaced in Plasmodium compared to most model systems, and that replication dynamics vary across the course of schizogony, from faster synthesis rates and more widely-spaced origins through to slower synthesis rates and closer-spaced origins. This is the opposite of the pattern usually seen across S-phase in human cells, when a single genome is replicated. Replication forks also appear to stall at an unusually high rate throughout schizogony. Our work explores Plasmodium DNA replication in unprecedented detail and opens up tremendous scope for analysing cell cycle dynamics and developing interventions targetting this unique aspect of malaria biology.The work was supported by UK Research Councils [MR/L008823/1, BB/K009206/1 to CJM]; by the Agence Nationale de la Recherche within the frame of the “Investissements d’avenir” program [ANR11-LABX-0024-01 “PARAFRAP” to YS], the Centre National de la Recherche Scientifique (CNRS) and the French Ministry of Research and the Centre Hospitalier Universitaire of Montpellier [YS]; the “Fondation pour la Recherche Médicale” (FRM) and Agence Nationale de la Recherche (ANR) [SS

Using haematophagous fly blood meals to study the diversity of blood‐borne pathogens infecting wild mammals

Many emerging infectious diseases originate from wild animals, so there is a profound need for surveillance and monitoring of their pathogens. However, the practical difficulty of sample acquisition from wild animals tends to limit the feasibility and effectiveness of such surveys. Xenosurveillance, using blood-feeding invertebrates to obtain tissue samples from wild animals and then detect their pathogens, is a promising method to do so. Here, we describe the use of tsetse fly blood meals to determine (directly through molecular diagnostic and indirectly through serology), the diversity of circulating blood-borne pathogens (including bacteria, viruses and protozoa) in a natural mammalian community of Tanzania. Molecular analyses of captured tsetse flies (182 pools of flies totalizing 1728 flies) revealed that the blood meals obtained came from 18 different vertebrate species including 16 non-human mammals, representing approximately 25% of the large mammal species present in the study area. Molecular diagnostic demonstrated the presence of different protozoa parasites and bacteria of medical and/or veterinary interest. None of the six virus species searched for by molecular methods were detected but an ELISA test detected antibodies against African swine fever virus among warthogs, indicating that the virus had been circulating in the area. Sampling of blood-feeding insects represents an efficient and practical approach to tracking a diversity of pathogens from multiple mammalian species, directly through molecular diagnostic or indirectly through serology, which could readily expand and enhance our understanding of the ecology and evolution of infectious agents and their interactions with their hosts in wild animal communities

Malar J

BACKGROUND: Resistance to all available anti-malarial drugs has emerged and spread including artemisinin derivatives and their partner drugs. Several genes involved in artemisinin and partner drugs resistance, such as pfcrt, pfmdr1, pfK13 or pfpm2, have been identified. However, these genes do not properly explain anti-malarial drug resistance, and more particularly clinical failures observed in Africa. Mutations in genes encoding for Plasmodium falciparum proteins, such as P. falciparum Acetyl-CoA transporter (PfACT), P. falciparum UDP-galactose transporter (PfUGT) and P. falciparum cyclic amine resistance locus (PfCARL) have recently been associated to resistance to imidazolopiperazines and other unrelated drugs. METHODS: Mutations on pfugt, pfact and pfcarl were characterized on 86 isolates collected in Dakar, Senegal and 173 samples collected from patients hospitalized in France after a travel in African countries from 2015 and 2016 to assess their potential association with ex vivo susceptibility to chloroquine, quinine, lumefantrine, monodesethylamodiaquine, mefloquine, dihydroartemisinin, artesunate, doxycycline, pyronaridine and piperaquine. RESULTS: No mutations were found on the genes pfugt and pfact. None of the pfcarl described mutations were identified in these samples from Africa. The K784N mutation was found in one sample and the K734M mutation was identified on 7.9% of all samples for pfcarl. The only significant differences in ex vivo susceptibility according to the K734M mutation were observed for pyronaridine for African isolates from imported malaria and for doxycycline for Senegalese parasites. CONCLUSION: No evidence was found of involvement of these genes in reduced susceptibility to standard anti-malarial drugs in African P. falciparum isolates

Sensibilité à la caspofungine des souches de Candida (expérience au CHRU de Montpellier)

MONTPELLIER-BU Médecine UPM (341722108) / SudocMONTPELLIER-BU Médecine (341722104) / SudocSudocFranceF

Molécules de surface de l' hématie parasitée par Plasmodium falciparum et implications dans la physiopathologie des accès palustres graves

PARIS7-Bibliothèque centrale (751132105) / SudocSudocFranceF

LYMPHOME MALIN NON HODGKINIEN ET VIRUS DE L'HEPATITE C

PARIS-BIUM (751062103) / SudocCentre Technique Livre Ens. Sup. (774682301) / SudocSudocFranceF

Diagnostic moléculaire de la toxoplasmose (étude comparative d'un kit commercial et de deux méthodes "maison")

MONTPELLIER-BU Pharmacie (341722105) / SudocSudocFranceF

Molecular diagnosis of visceral leishmaniasis: a French study comparing a reference PCR method targeting kinetoplast DNA and a commercial kit targeting ribosomal DNA

International audienceVisceral leishmaniasis is a life-threatening disease caused by the Leishmania donovani species complex. For direct diagnosis, molecular diagnosis on blood and bone marrow is an increasingly used technique. For Leishmania , most PCR assays are laboratory-developed, but marketed PCR assays are now available and should be evaluated independently of manufacturers. The “quanty Leishmaniae, Clonit” kit was compared to a laboratory-developed method widely used in France and considered here as a reference method. Performances were evaluated on serial dilution assays, on 5 external quality controls and on 35 clinical samples. The reference method performed better than the Clonit kit with higher “performance scores”: 20 of 28 (71%) and 4 of 12 (33%) vs 17 of 28 (61%) and 1 of 12 (8%) for the reference method and the Clonit kit, respectively. On clinical samples, six false negative results out of 27 positive samples (22%) were observed with the quanty Leishmaniae, Clonit method. These results are most likely due to the difference in the number of repeats of the PCR targets. The quanty Leishmaniae, Clonit, like most of marketed methods, targets the ribosomal DNA that has a lower number of copies than the kinetoplast DNA targeted by the reference PCR. This study confirms that the choice of target is crucial and should be taken into account in the development of new highly sensitive PCR methods. IMPORTANCE PCR revolutionized the direct diagnosis of infectious diseases, especially protozooses, where the infectious load is usually low. Commercial PCR methods are available and offer many advantages, including convenience and batch tracking as part of a quality system. For most parameters, the performance of commercial methods is at least as good as that of finely optimized methods developed in expert laboratories. This comparison work has not been done for the molecular diagnosis of visceral leishmaniasis. Leishmania sp. has a unique organelle, the kinetoplast, which corresponds to the mitochondrial DNA. It is organized into a large number of minicircles, which has made it a target for the development of diagnostic PCR. The quanty Leishmaniae, Clonit kit targeting ribosomal DNA was compared to a widely used laboratory-developed method based on kinetoplast DNA. This reference method gave significantly better results, probably due to the difference in the number of repeats of the PCR targets

Evaluation of Two Commercial Kits on the Automated ELITe InGenius PCR Platform for Molecular Diagnosis of Toxoplasmosis

International audienceMolecular diagnosis of toxoplasmosis is essential for establishing the diagnosis of congenital contaminations and for primary infection or reactivation of immunocompromised patients. An integrated extraction and real-time PCR-based system is of particular interest in this context. Commercial kits for automated extraction and amplification steps are now available. Herein, we assessed two commercial PCR assays for this diagnosis, those of Bio-Evolution and Elitech, on the ELITe InGenius platform. The Bio-Evolution assay showed a specificity and a sensitivity of 100% on clinical samples, but a lower analytical detection threshold than the Elitech assay. The latter showed a specificity of 100% and a sensitivity of 96%. The SP1000 cartridges, which allow DNA extraction from 1 mL of template, showed interesting performances on amniotic fluid samples. Overall, the two kits had good performances on the InGenius platform, which offers a turn-key solution suitable for the molecular diagnosis of toxoplasmosis