Exclusive Decoration of Simian Immunodeficiency Virus Env with High-Mannose Type N-Glycans Is Not Compatible with Mucosal Transmission in Rhesus Macaques

Human immunodeficiency virus (HIV) and simian immunodeficiency virus (SIV) envelope (Env) proteins are extensively decorated with N-glycans, predominantly of the high-mannose type. However, it is unclear how high-mannose N-glycans on Env impact viral spread. We show that exclusive modification of SIV Env with these N-glycans reduces viral infectivity and abrogates mucosal transmission, despite increasing viral capture by immune cell lectins. Thus, high-mannose N-glycans have opposed effects on SIV infectivity and lectin reactivity, and a balance might be required for efficient mucosal transmission.     

Multiple blood feeding in mosquitoes shortens the Plasmodium falciparum incubation period and increases malaria transmission potential

Many mosquito species, including the major malaria vector Anopheles gambiae, naturally undergo multiple reproductive cycles of blood feeding, egg development and egg laying in their lifespan. Such complex mosquito behavior is regularly overlooked when mosquitoes are experimentally infected with malaria parasites, limiting our ability to accurately describe potential effects on transmission. Here, we examine how Plasmodium falciparum development and transmission potential is impacted when infected mosquitoes feed an additional time. We measured P. falciparum oocyst size and performed sporozoite time course analyses to determine the parasite’s extrinsic incubation period (EIP), i.e. the time required by parasites to reach infectious sporozoite stages, in An. gambiae females blood fed either once or twice. An additional blood feed at 3 days post infection drastically accelerates oocyst growth rates, causing earlier sporozoite accumulation in the salivary glands, thereby shortening the EIP (reduction of 2.3 ± 0.4 days). Moreover, parasite growth is further accelerated in transgenic mosquitoes with reduced reproductive capacity, which mimic genetic modifications currently proposed in population suppression gene drives. We incorporate our shortened EIP values into a measure of transmission potential, the basic reproduction number R₀, and find the average R₀ is higher (range: 10.1%–12.1% increase) across sub-Saharan Africa than when using traditional EIP measurements. These data suggest that malaria elimination may be substantially more challenging and that younger mosquitoes or those with reduced reproductive ability may provide a larger contribution to infection than currently believed. Our findings have profound implications for current and future mosquito control interventions.     

Populations of a widespread hexacoral have trophic plasticity and flexible syntrophic interactions across the Indo-Pacific Ocean

Coral Reefs - Benthic cnidarians are suspension feeders that ingest items ranging from particulate organic matter to macrozooplankton. Additionally, many species receive photosynthetic products...     

Endocrine modulating actions of a phytosterol mixture and its oxidation products in zebrafish (Danio rerio)

In this study, a phytosterol preparation ("ultrasitosterol"; 80% beta-sitosterol) and an oxidized ultrasitosterol preparation were evaluated for reproductive effects in zebrafish. Adults were exposed in a continuous flow to 10 microg/L and 100 microg/L ultrasitosterol and oxidized ultrasitosterol, and to 0.27 microg/L 17beta-estradiol and 0.28 microg/L testosterone for 3 weeks. Biomarkers analysed included plasma vitellogenin, testosterone, 11-ketotestosterone, 17beta-estradiol, and gonadal histopathology. Ovarian steroid production of testosterone and 17beta-estradiol was examined in isolated zebrafish follicles exposed in vitro to the compounds at the same concentrations as in vivo. Vtg was induced in males exposed to ultrasitosterol, and in males and females exposed to 17beta-estradiol. Males exposed to oxidized phytosterols showed increased levels of testosterone and 11-ketotestosterone, and accelerated spermatogenesis. Increased follicular atresia was observed in females exposed to oxidized phytosterols and 17beta-estradiol. Correlation analyses between biomarkers revealed more intercorrelated values for females than for males, and the strongest associations were found in females exposed to oxidized phytosterols. Testosterone production was significantly increased in follicles exposed to the oxidized phytosterol preparations. These findings indicate that the phytosterol mixture is weakly estrogenic in male fish, and that the oxidized phytosterol mixture contains substances that may interfere with spermatogenesis, oogenesis and gonadal steroidogenesis in zebrafish.     

Localization of Usher 1 proteins to the photoreceptor calyceal processes,which are absent from mice

The mechanisms underlying retinal dystrophy in Usher syndrome type I (USH1) remain unknown because mutant mice lacking any of the USH1 proteins—myosin VIIa, harmonin, cadherin-23, protocadherin-15, sans—do not display retinal degeneration. We found here that, in macaque photoreceptor cells, all USH1 proteins colocalized at membrane interfaces (i) between the inner and outer segments in rods and (ii) between the microvillus-like calyceal processes and the outer segment basolateral region in rods and cones. This pattern, conserved in humans and frogs, was mediated by the formation of an USH1 protein network, which was associated with the calyceal processes from the early embryonic stages of outer segment growth onwards. By contrast, mouse photoreceptors lacked calyceal processes and had no USH1 proteins at the inner–outer segment interface. We suggest that USH1 proteins form an adhesion belt around the basolateral region of the photoreceptor outer segment in humans, and that defects in this structure cause the retinal degeneration in USH1 patients.     

Cytoplasmic utilization of human immunodeficiency virus type 1 genomic RNA is not dependent on a nuclear interaction with gag

In some retroviruses, such as Rous sarcoma virus and prototype foamy virus, Gag proteins are known to shuttle between the nucleus and the cytoplasm and are implicated in nuclear export of the viral genomic unspliced RNA (gRNA) for subsequent encapsidation. A similar function has been proposed for human immunodeficiency virus type 1 (HIV-1) Gag based on the identification of nuclear localization and export signals. However, the ability of HIV-1 Gag to transit through the nucleus has never been confirmed. In addition, the lentiviral Rev protein promotes efficient nuclear gRNA export, and previous reports indicate a cytoplasmic interaction between Gag and gRNA. Therefore, functional effects of HIV-1 Gag on gRNA and its usage were explored. Expression of gag in the absence of Rev was not able to increase cytoplasmic gRNA levels of subgenomic, proviral, or lentiviral vector constructs, and gene expression from genomic reporter plasmids could not be induced by Gag provided in trans. Furthermore, Gag lacking the reported nuclear localization and export signals was still able to mediate an efficient packaging process. Although small amounts of Gag were detectable in the nuclei of transfected cells, a Crm1-dependent nuclear export signal in Gag could not be confirmed. Thus, our study does not provide any evidence for a nuclear function of HIV-1 Gag. The encapsidation process of HIV-1 therefore clearly differs from that of Rous sarcoma virus and prototype foamy virus.     

Signal perception by the secretion stress-responsive CssRS two-component system in Bacillus subtilis

The CssRS two-component system responds to heat and secretion stresses in Bacillus subtilis by controlling expression of HtrA and HtrB chaperone-type proteases and positively autoregulating its own expression. Here we report on the features of the CssS extracellular loop domain that are involved in signal perception and on CssS subcellular localization. Individual regions of the CssS extracellular loop domain contribute differently to signal perception and activation. The conserved hydrophilic 26-amino-acid segment juxtaposed to transmembrane helix 1 is involved in the switch between the deactivated and activated states, while the conserved 19-amino-acid hydrophobic segment juxtaposed to transmembrane 2 is required for signal perception and/or transduction. Perturbing the size of the extracellular loop domain increases CssS kinase activity and makes it unresponsive to secretion stress. CssS is localized primarily at the septum but is also found in a punctate pattern with lower intensity throughout the cell cylinder. Moreover, the CssRS-controlled HtrA and HtrB proteases are randomly distributed in foci throughout the cell surface, with more HtrB than HtrA foci in unstressed cells.     

A Basic Approach Towards the Development of Bioelectric Bacterial Biosensors for the Detection of Plant Viruses

There is a growing need for virus sensors with improved sensitivity and dynamic range for disease diagnosis, pharmaceutical research, agriculture and homeland security. Membrane‐engineered animal cells bearing antibodies against viral antigens have been previously used for biorecognition biosensors for the ultrarapid (3 min), sensitive (1 ng/ml) detection of plant viruses, such as the cucumber mosaic virus. We here report a new approach for the construction of cell‐based sensors for virus detection, based on membrane (antibody)‐engineered bacteria. The novel method was applied for the detection of tobacco mosaic virus (TMV) and cherry leaf roll virus (CLRV) using sensors containing modified Escherichia coli XL‐1Blue MRF’ bacteria. E. coli membranes have been engineered with electro‐inserted, virus‐homologous antibodies. The detection principle was based on the measurement of changes in the bacterial membrane potential as a result of virus–antibody binding. After optimization of the membrane‐engineering process, the virus detection limit for TMV and CLRV with the bacteria‐based biosensor system was 1 pg/ml, representing a 1000‐fold improvement over currently available methods. Although the novel biosensor is still in its proof‐of‐concept stage of development, its sensitivity and speed (assay time: 60–100 s) could make it a very promising tool for high throughput, field‐based virus screening.     

Proteomic methods applied to the analysis of immobilized biocatalysts

Methods adapted from proteomics can directly characterize proteins present in immobilized biocatalysts. Complete hydrolysis followed by HPLC analysis of Tyr and Phe estimates total protein bound, and is preferable to conventional difference methods, as tested with subtilisin Carlsberg on silica. This new method shows that various treatments give quantitative desorption of proteins immobilized by adsorption. Intact desorbed proteins may be analyzed by electrospray mass spectrometry. The Candida antarctica lipase B from Novozyme 435 was shown to be heavily glycosylated, while the lipase from Lipozyme RM IM was a mixture of four N-terminally truncated forms. Peptides from selective cleavage were analyzed by tandem mass spectrometry, leading to automatic identification of proteins present. A second major protein present in Lipozyme RM IM was thus found to be alpha-amylase from Aspergillus oryzae. These methods should be valuable complements to activity measurements in understanding immobilized enzyme activity and stability.