Imaging movement of malaria parasites during transmission by Anopheles mosquitoes

Malaria is contracted when Plasmodium sporozoites are inoculated into the vertebrate host during the blood meal of a mosquito. In infected mosquitoes, sporozoites are present in large numbers in the secretory cavities of the salivary glands at the most distal site of the salivary system. However, how sporozoites move through the salivary system of the mosquito, both in resting and feeding mosquitoes, is unknown. Here, we observed fluorescent Plasmodium berghei sporozoites within live Anopheles stephensi mosquitoes and their salivary glands and ducts. We show that sporozoites move in the mosquito by gliding, a type of motility associated with their capacity to invade host cells. Unlike in vitro, sporozoite gliding inside salivary cavities and ducts is modulated in speed and motion pattern. Imaging of sporozoite discharge through the proboscis of salivating mosquitoes indicates that sporozoites need to locomote from cavities into ducts to be ejected and that their progression inside ducts favours their early ejection. These observations suggest that sporozoite gliding allows not only for cell invasion but also for parasite locomotion in host tissues, and that it may control parasite transmission.     

Independent roles of apical membrane antigen 1 and rhoptry neck proteins during host cell invasion by apicomplexa

During invasion, apicomplexan parasites form an intimate circumferential contact with the host cell, the tight junction (TJ), through which they actively glide. The TJ, which links the parasite motor to the host cell cytoskeleton, is thought to be composed of interacting apical membrane antigen 1 (AMA1) and rhoptry neck (RON) proteins. Here we find that, in Plasmodium berghei, while both AMA1 and RON4 are important for merozoite invasion of erythrocytes, only RON4 is required for sporozoite invasion of hepatocytes, indicating that RON4 acts independently of AMA1 in the sporozoite. Further, in the Toxoplasma gondii tachyzoite, AMA1 is dispensable for normal RON4 ring and functional TJ assembly but enhances tachyzoite apposition to the cell and internalization frequency. We propose that while the RON proteins act at the TJ, AMA1 mainly functions on the zoite surface to permit correct attachment to the cell, which may facilitate invasion depending on the zoite-cell combination.     

Towards systematic identification of Plasmodium essential genes by transposon shuttle mutagenesis

After the deciphering of the genome sequences of several Plasmodium species, efforts must turn to elucidating gene function and identifying essential gene products. However, random approaches are lacking and gene targeting is inefficient in Plasmodium. Here, we established shuttle transposon mutagenesis in Plasmodium berghei. We constructed a mini-Tn5 derivative that can transpose into parasite genes cloned in Escherichia coli, providing an efficient means of generating knockout fragments. A 10⁴-fold increase in frequencies of double-crossover homologous recombination in the parasite using a new electroporation technology permits to reproducibly generate pools of distinct mutants after transfection with mini-Tn5-interrupted sequences. The procedure opens the way to the systematic identification of essential genes in Plasmodium.     

Deficiency in OGG1 protects against inflammation and mutagenic effects associated with H. pylori infection in mouse

BACKGROUND: Helicobacter pylori infection is associated with gastric cancer. Study with the Big Blue mouse model has reported a mutagenic effect associated with the H. pylori infection, as a result in part of oxidative DNA damage. The present work investigates the consequences of a deficiency in the OGG1 DNA glycosylase, responsible for the excision of 8-oxo guanine, on the inflammatory and genotoxic host response to the infection. MATERIALS AND METHODS: Big Blue Ogg1-/- C57BL/6 mice were orally inoculated with H. pylori strain SS1 or vehicle only, and sacrificed after 1, 3, or 6 months. The serologic response, histologic lesions, mutant frequency, and spectra of mutations were assessed in the stomach and compared to what observed in the wild-type (Wt) context. RESULTS: Inflammatory lesions induced in the gastric mucosa of H. pylori-infected mice, corresponding to a moderate gastritis, were less severe in Ogg1-/- than in Wt Big Blue mice. Analysis of antimicrobial humoral immunity exhibited a lower IgG2a serum level (Th1 response) after 6 months of infection in Ogg1-/- than in the Wt mice. In these conditions, the H. pylori-SS1 infection in the Ogg1-/- mice did not induce a mutagenic effect at the gastric epithelial cells level, either after 3 or 6 months. CONCLUSIONS: The inactivation of the OGG1 DNA glycosylase in mouse leads to less severe inflammatory lesions and abolished the mutagenic effect at the gastric epithelial cells level, induced by the H. pylori infection. These data suggest for the OGG1deficiency a protective role against inflammation and genotoxicity associated to the H. pylori infection.     

Nickel binding and immunological properties of the C-terminal domain of the Helicobacter pylori GroES homologue (HspA)

Helicobacter pylori synthesizes a heat-shock protein of the GroES class. The gene encoding this protein (heat-shock protein A, HspA) was recently cloned and it was shown to be unique in structure. H. pylori HspA consists of two domains: the N-terminal domain (domain A) homologous with other GroES proteins, and a C-terminal domain (domain B) corresponding to 27 additional residues resembling a metal-binding domain. Various recombinant proteins consisting of the entire HspA polypeptide, the A domain, or the B domain were produced independently as proteins fused to maltose-binding protein (MBP). Comparison of the divalent cation binding properties of the various MBP and MBP-fused proteins allowed us to conclude that HspA binds nickel ions by means of its C-terminal domain. HspA exhibited a high and specific affinity for nickel ions in comparison with its affinity for other divalent cations (copper, zinc, cobalt). Equilibrium dialysis experiments revealed that MBP–HspA binds nickel ions with an apparent dissociation constant (K_d) of 1.8 μM and a stoichiometry of 1.9 ions per molecule. The analysis of the deduced HspA amino acid sequences encoded by 35 independent clinical isolates demonstrated the existence of two molecular variants of HspA, i.e. a major and a minor variant present in 89% and 11% of strains, respectively. The two variants differed from each other by the simultaneous substitution of seven amino acids within the B domain, whilst the A domain was highly conserved amongst all the HspA proteins (99–100% identity). On the basis of serological studies, the highly conserved A domain of HspA was found to be the immunodominant domain. Functional complementation experiments were performed to test the properties of the two HspA variants. When co-expressed together with the H. pylori urease gene cluster in Escherichia coli cells, the two HspA variant-encoding genes led to a fourfold increase in urease activity, demonstrating that HspA in H. pylori has a specialized function with regard to the nickel metalloenzyme urease.     

Activation of hypothalamic RIP‐Cre neurons promotes beiging of WAT via sympathetic nervous system

Activation of brown adipose tissue (BAT) and beige fat by cold increases energy expenditure. Although their activation is known to be differentially regulated in part by hypothalamus, the underlying neural pathways and populations remain poorly characterized. Here, we show that activation of rat‐insulin‐promoter‐Cre (RIP‐Cre) neurons in ventromedial hypothalamus (VMH) preferentially promotes recruitment of beige fat via a selective control of sympathetic nervous system (SNS) outflow to subcutaneous white adipose tissue (sWAT), but has no effect on BAT. Genetic ablation of APPL2 in RIP‐Cre neurons diminishes beiging in sWAT without affecting BAT, leading to cold intolerance and obesity in mice. Such defects are reversed by activation of RIP‐Cre neurons, inactivation of VMH AMPK, or treatment with a β3‐adrenergic receptor agonist. Hypothalamic APPL2 enhances neuronal activation in VMH RIP‐Cre neurons and raphe pallidus, thereby eliciting SNS outflow to sWAT and subsequent beiging. These data suggest that beige fat can be selectively activated by VMH RIP‐Cre neurons, in which the APPL2–AMPK signaling axis is crucial for this defending mechanism to cold and obesity.     

Proteomic analysis during larval development and metamorphosis of the spionid polychaete Pseudopolydora vexillosa

Background  

While the larval-juvenile transition (metamorphosis) in the spionid polychaete Pseudopolydora vexillosa involves gradual morphological changes and does not require substantial development of juvenile organs, the opposite occurs in the barnacle Balanus amphitrite. We hypothesized that the proteome changes during metamorphosis in the spionids are less drastic than that in the barnacles. To test this, proteomes of pre-competent larvae, competent larvae (ready to metamorphose), and juveniles of P. vexillosa were compared using 2-dimensional gel electrophoresis (2-DE), and they were then compared to those of the barnacle.     

A Preliminary Study of Viral Metagenomics of French Bat Species in Contact with Humans: Identification of New Mammalian Viruses

The prediction of viral zoonosis epidemics has become a major public health issue. A profound understanding of the viral population in key animal species acting as reservoirs represents an important step towards this goal. Bats harbor diverse viruses, some of which are of particular interest because they cause severe human diseases. However, little is known about the diversity of the global population of viruses found in bats (virome). We determined the viral diversity of five different French insectivorous bat species (nine specimens in total) in close contact with humans. Sequence-independent amplification, high-throughput sequencing with Illumina technology and a dedicated bioinformatics analysis pipeline were used on pooled tissues (brain, liver and lungs). Comparisons of the sequences of contigs and unassembled reads provided a global taxonomic distribution of virus-related sequences for each sample, highlighting differences both within and between bat species. Many viral families were present in these viromes, including viruses known to infect bacteria, plants/fungi, insects or vertebrates, the most relevant being those infecting mammals (Retroviridae, Herpesviridae, Bunyaviridae, Poxviridae, Flaviviridae, Reoviridae, Bornaviridae, Picobirnaviridae). In particular, we detected several new mammalian viruses, including rotaviruses, gammaretroviruses, bornaviruses and bunyaviruses with the identification of the first bat nairovirus. These observations demonstrate that bats naturally harbor viruses from many different families, most of which infect mammals. They may therefore constitute a major reservoir of viral diversity that should be analyzed carefully, to determine the role played by bats in the spread of zoonotic viral infections.     

Co-Circulation of Dengue Virus Type 3 Genotypes in Vientiane Capital,Lao PDR

During the 2012 epidemic of dengue in Vientiane capital, Lao PDR, a major serotype switch from dengue 1 to 3 was observed. A molecular epidemiology study demonstrated that dengue 3 remained the predominant serotype in 2013, but also revealed the co-circulation of two genotypes, supporting the hypothesis of multiple geographic origins of dengue 3 strains circulating in Vientiane capital.