Integument of the tapeworm scolex. 1. Freeze-fracture of the syncytial layer, microvilli and discoid bodies

The tegument of cestodes is the most important and structurally complex metabolic interface between these parasites and the hostile environment in which they reside. In spite of the complex metabolic, regulatory and immunological properties of this layer of syncytial cytoplasm, which are relatively well known, the detailed fine structural anatomy of the cestode tegument remains equivocal. The present study therefore reports the freeze-fracture morphology of the tapeworm (Hymenolepis diminuta) tegument. The most important features revealed by analysis of platinum replicas of freeze-fractured tapeworm scolex-neck tegument include: (a) presence of highly ordered linear and/or circumferentially-orientated rows of intramembrane particles situated on the PF fracture face of microvillar plasma membrane, which may participate in movements of the microvilli, (b) presence of apparent 'pores' (11 nm in diameter) at the tips of the tegumentary microvilli, which could serve as regulated gates through which extramicrovillar surface coating materials can be extruded, and (c) the alignment of cytoplasmic discoid bodies into positions at the bases of the surface microvilli such that they could move into the core of each microvillus and thereby release their contents for extrusion (via the pores) onto the outer surface of the microvilli. Concomitantly, the limiting membrane of the discoid bodies could be added to the tegument plasma membrane and thereby contribute to the rapid turnover of the tegumentary surface. This study provides the first detailed account of the ultrastructural anatomy of the tapeworm tegument and is intended to serve as a point of reference for future investigations of tapeworm tegumentary functions.     

A freeze-fracture study of the body wall of adult, in utero larvae and infective-stage larvae of Trichinella (Nematoda)

The surface layers of the cuticle, the hypodermal membranes and the muscle membranes of the adult, the in utero larvae and the infective-stage larvae of the nematode Trichinella spiralis have been studied by means of the freeze-fracturing technique. The surface of the cuticle of both adults and larvae fractures in ways different from membranes of internal cells. The surface coat on top of the epicuticle is probably the layer that changes antigenically. Reticulate ridges, with associated particles, on the E face of the outer hypodermal membrane of the adult are probably sites of attachment of the hypodermis to the cuticle. Longitudinally arranged ridges, with associated particles, of the outer hypodermal membrane are probably points of attachment to the cuticle in the in utero and infective larvae. Rectilinear arrays of particles are present on the P face of the inner hypodermal membrane and the P face of the muscle membrane adjacent to the hypodermis of adults and larvae and probably play a role in adhesion of the muscle membrane to the hypodermis. Particle-free areas of membrane lie external to the Z bundles of the muscle cell and are similar to the sites of attachment of Z lines in insect muscles.     

Determination of Sequences Required for Human Endogenous Retrovirus K Transduction and Its Recognition by Foreign Retroviral Virions

Sequences necessary for transduction of human endogenous retrovirus (HERV)-K_con, a consensus of the HERV-K(HML-2) family, were analyzed and found to reside in the leader/gag region. They act in an orientation-dependent way and consist of at least two sites working together. Having defined these sequences, we exploited this information to produce a simple system to investigate to what extent virions of HERV-K_con, murine leukemia virus, and HIV-1 have the ability to transduce each other''s genomes, leading to potential contamination of gene therapy vectors.     

EARLY LIFE HISTORIES,OCEAN CURRENTS,AND THE POPULATION GENETICS OF CARIBBEAN REEF FISHES

Tropical reef fishes, along with many benthic invertebrates, have a life cycle that includes a sedentary, bottom-dwelling reproductive phase and a planktonic stage that occurs early in development. The adult benthic populations occupy disjunct, patchy habitats; the extent of gene flow due to dispersal of the planktonic life stage is generally unknown.     

RGD-directed attachment of isolated rat osteoclasts to osteopontin, bone sialoprotein, and fibronectin

Osteoclasts isolated from the long bones of 5-day-old rats were seeded onto glass surfaces coated with osteopontin, bone sialoprotein, or fibronectin. Cell binding was promoted by all three proteins and inhibited in a dose-dependent manner by an RGD-containing peptide, while an RGE-containing peptide was ineffective. Immunocytochemistry of bone tissue showed enhanced concentration of osteopontin in bone opposite the clear zone of the osteoclasts, whereas immunolocalization of bone sialoprotein and fibronectin showed no accumulation on bone surfaces facing cells. The observations corroborate previous findings that the osteoclast is attached via an integrin to osteopontin on the bone surface. Although bone sialoprotein and fibronectin can mediate osteoclast binding in vitro, such a role in vivo is not supported by the immunocytochemical observations.     

REDD1 Is Essential for Optimal T Cell Proliferation and Survival

REDD1 is a highly conserved stress response protein that is upregulated following many types of cellular stress, including hypoxia, DNA damage, energy stress, ER stress, and nutrient deprivation. Recently, REDD1 was shown to be involved in dexamethasone induced autophagy in murine thymocytes. However, we know little of REDD1’s function in mature T cells. Here we show for the first time that REDD1 is upregulated following T cell stimulation with PHA or CD3/CD28 beads. REDD1 knockout T cells exhibit a defect in proliferation and cell survival, although markers of activation appear normal. These findings demonstrate a previously unappreciated role for REDD1 in T cell function.     

Most of the Dominant Members of Amphibian Skin Bacterial Communities Can Be Readily Cultured

Currently, it is estimated that only 0.001% to 15% of bacteria in any given system can be cultured by use of commonly used techniques and media, yet culturing is critically important for investigations of bacterial function. Despite this situation, few studies have attempted to link culture-dependent and culture-independent data for a single system to better understand which members of the microbial community are readily cultured. In amphibians, some cutaneous bacterial symbionts can inhibit establishment and growth of the fungal pathogen Batrachochytrium dendrobatidis, and thus there is great interest in using these symbionts as probiotics for the conservation of amphibians threatened by B. dendrobatidis. The present study examined the portion of the culture-independent bacterial community (based on Illumina amplicon sequencing of the 16S rRNA gene) that was cultured with R2A low-nutrient agar and whether the cultured bacteria represented rare or dominant members of the community in the following four amphibian species: bullfrogs (Lithobates catesbeianus), eastern newts (Notophthalmus viridescens), spring peepers (Pseudacris crucifer), and American toads (Anaxyrus americanus). To determine which percentage of the community was cultured, we clustered Illumina sequences at 97% similarity, using the culture sequences as a reference database. For each amphibian species, we cultured, on average, 0.59% to 1.12% of each individual''s bacterial community. However, the average percentage of bacteria that were culturable for each amphibian species was higher, with averages ranging from 2.81% to 7.47%. Furthermore, most of the dominant operational taxonomic units (OTUs), families, and phyla were represented in our cultures. These results open up new research avenues for understanding the functional roles of these dominant bacteria in host health.     

Nef decreases HIV-1 sensitivity to neutralizing antibodies that target the membrane-proximal external region of TMgp41

Primate lentivirus nef is required for sustained virus replication in vivo and accelerated progression to AIDS. While exploring the mechanism by which Nef increases the infectivity of cell-free virions, we investigated a functional link between Nef and Env. Since we failed to detect an effect of Nef on the quantity of virion-associated Env, we searched for qualitative changes by examining whether Nef alters HIV-1 sensitivity to agents that target distinct features of Env. Nef conferred as much as 50-fold resistance to 2F5 and 4E10, two potent neutralizing monoclonal antibodies (nAbs) that target the membrane proximal external region (MPER) of TMgp41. In contrast, Nef had no effect on HIV-1 neutralization by MPER-specific nAb Z13e1, by the peptide inhibitor T20, nor by a panel of nAbs and other reagents targeting gp120. Resistance to neutralization by 2F5 and 4E10 was observed with Nef from a diverse range of HIV-1 and SIV isolates, as well as with HIV-1 virions bearing Env from CCR5- and CXCR4-tropic viruses, clade B and C viruses, or primary isolates. Functional analysis of a panel of Nef mutants revealed that this activity requires Nef myristoylation but that it is genetically separable from other Nef functions such as the ability to enhance virus infectivity and to downregulate CD4. Glycosylated-Gag from MoMLV substituted for Nef in conferring resistance to 2F5 and 4E10, indicating that this activity is conserved in a retrovirus that does not encode Nef. Given the reported membrane-dependence of MPER-recognition by 2F5 and 4E10, in contrast to the membrane-independence of Z13e1, the data here is consistent with a model in which Nef alters MPER recognition in the context of the virion membrane. Indeed, Nef and Glycosylated-Gag decreased the efficiency of virion capture by 2F5 and 4E10, but not by other nAbs. These studies demonstrate that Nef protects lentiviruses from one of the most broadly-acting classes of neutralizing antibodies. This newly discovered activity for Nef has important implications for anti-HIV-1 immunity and AIDS pathogenesis.     

Plasma HIV Viral Rebound following Protocol-Indicated Cessation of ART Commenced in Primary and Chronic HIV Infection

Objectives

The magnitude of HIV viral rebound following ART cessation has consequences for clinical outcome and onward transmission. We compared plasma viral load (pVL) rebound after stopping ART initiated in primary (PHI) and chronic HIV infection (CHI).

Design

Two populations with protocol-indicated ART cessation from SPARTAC (PHI, n = 182) and SMART (CHI, n = 1450) trials.

Methods

Time for pVL to reach pre-ART levels after stopping ART was assessed in PHI using survival analysis. Differences in pVL between PHI and CHI populations 4 weeks after stopping ART were examined using linear and logistic regression. Differences in pVL slopes up to 48 weeks were examined using linear mixed models and viral burden was estimated through a time-averaged area-under-pVL curve. CHI participants were categorised by nadir CD4 at ART stop.

Results

Of 171 PHI participants, 71 (41.5%) rebounded to pre-ART pVL levels, at a median of 50 (95% CI 48–51) weeks after stopping ART. Four weeks after stopping treatment, although the proportion with pVL≥400 copies/ml was similar (78% PHI versus 79% CHI), levels were 0.45 (95% CI 0.26–0.64) log₁₀ copies/ml lower for PHI versus CHI, and remained lower up to 48 weeks. Lower CD4 nadir in CHI was associated with higher pVL after ART stop. Rebound for CHI participants with CD4 nadir >500 cells/mm³ was comparable to that experienced by PHI participants.

Conclusions

Stopping ART initiated in PHI and CHI was associated with viral rebound to levels conferring increased transmission risk, although the level of rebound was significantly lower and sustained in PHI compared to CHI.