Noroviruses are major agents of viral gastroenteritis worldwide. The infectivity of Norwalk virus, the prototype norovirus, has been studied in susceptible human volunteers. A new variant of the hit theory model of microbial infection was developed to estimate the variation in Norwalk virus infectivity, as well as the degree of virus aggregation, consistent with independent (electron microscopic) observations. Explicit modeling of viral aggregation allows us to express virus infectivity per single infectious unit (particle). Comparison of a primary and a secondary inoculum showed that passage through a human host does not change Norwalk virus infectivity. We estimate the average probability of infection for a single Norwalk virus particle to be close to 0.5, exceeding that reported for any other virus studied to date. Infected subjects had a dose-dependent probability of becoming ill, ranging from 0.1 (at a dose of 10(3) NV genomes) to 0.7 (at 10(8) virus genomes). A norovirus dose response model is important for understanding its transmission and essential for development of a quantitative risk model. Norwalk virus is a valuable model system to study virulence because genetic factors are known for both complete and partial protection; the latter can be quantitatively described as heterogeneity in dose response models.

Accumulation of autophagosomes because of impaired autophagy during valosin-containing protein (VCP)–linked dementia is explained by the absence or reduced activity of VCP.

Enterotoxigenic Escherichia coli (ETEC) is a prevalent cause of traveler's diarrhea and infant mortality in thirdworld countries. Heat-labile enterotoxin (LT) is secreted from ETEC via vesicles composed of outer membrane and periplasm. We investigated the role of ETEC vesicles in pathogenesis by analyzing vesicle association and entry into eukaryotic cells. Fluorescently labeled vesicles from LT-producing and LT-nonproducing strains were compared in their ability to bind adrenal and intestinal epithelial cells. ETEC-derived vesicles, but not control nonpathogenderived vesicles, associated with cells in a time-, temperature-, and receptor-dependent manner. Vesicles were visualized on the cell surface at 41C and detected intracellularly at 371C. ETEC vesicle endocytosis depended on cholesterol-rich lipid rafts. Entering vesicles partially colocalized with caveolin, and the internalized vesicles accumulated in a nonacidified compartment. We conclude that ETEC vesicles serve as specifically targeted transport vehicles that mediate entry of active enterotoxin and other bacterial envelope components into host cells. These data demonstrate a role in virulence for ETEC vesicles.

TDP-43 is found in ubiquitinated inclusions (UBIs) in some frontotemporal dementias (FTD-U). One form of FTD-U, due to mutations in VCP, occurs with an inclusion body myopathy (IBMPFD). Since IBMPFD brain has TDP-43 in UBIs, we looked for TDP-43 inclusions in IBMPFD muscle. In normal muscle TDP-43 is present in nuclei. In IBMPFD muscle TDP-43 is additionally present as large inclusions within UBIs in muscle cytoplasm. TDP-43 inclusions were also found in 78% of sIBM muscles. In IBMPFD and sIBM muscle TDP-43 migrated with an additional band on immunoblot similar to that reported in FTD-U brains. This study adds sIBM and hereditary inclusion body myopathies to the growing list of TDP-43 positive inclusion diseases.

R eliable and quantitative assays to measure in vivo autophagy are essential. Currently, there are varied methods for monitoring autophagy; however, it is a challenge to measure "autophagic flux" in an in vivo model system. Conversion and subsequent degradation of the microtubule-associated protein 1 light chain 3 (MAP1-LC3/LC3) to the autophagosome associated LC3-II isoform can be evaluated by immunoblot. However, static levels of endogenous LC3-II protein may render possible misinterpretations since LC3-II levels can increase, decrease or remain unchanged in the setting of autophagic induction. Therefore, it is necessary to measure LC3-II protein levels in the presence and absence of lysomotropic agents that block the degradation of LC3-II, a technique aptly named the "autophagometer." In order to measure autophagic flux in mouse skeletal muscle, we treated animals with the microtubule depolarizing agent colchicine. Two days of 0.4 mg/kg/day intraperitoneal colchicine blocked autophagosome maturation to autolysosomes and increased LC3-II protein levels in mouse skeletal muscle by >100%. The addition of an autophagic stimulus such as dietary restriction or rapamycin led to an additional increase in LC3-II above that seen with colchicine alone. Moreover, this increase was not apparent in the absence of a "colchicine block." Using this assay, we evaluated the autophagic response in skeletal muscle upon denervation induced atrophy. Our studies highlight the feasibility of performing an "in vivo autophagometer" study using colchicine in skeletal muscle.

SUMMARY Electron microscopy, considered by some to be an old technique, is still on the forefront of both clinical viral diagnoses and viral ultrastructure and pathogenesis studies. In the diagnostic setting, it is particularly valuable in the surveillance of emerging diseases and potential bioterrorism viruses. In the research arena, modalities such as immunoelectron microscopy, cryo-electron microscopy, and electron tomography have demonstrated how viral structural components fit together, attach to cells, assimilate during replication, and associate with the cellular machinery during replication and egression. These studies provide information for treatment and vaccine strategies.

This is a case report of an immunocompromised individual who presented with progressive alopecia, friable follicular spinous processes, and erythematous, indurated papules. Examination of skin biopsies using light microscopy and immunohistochemistry revealed pathologic changes of the follicular inner root sheath epithelium with dystrophic trichohyaline granules. Electron microscopy of thin sections of tissue revealed intracellular viral particles with a size and appearance consistent with those in the Papovaviridae family. Electron microscopy of negatively stained extract from a homogenized lesion also demonstrated icosahedral viruses with papovavirus morphology. We believe this is a previously unreported folliculocentric viral infection in an immunosuppressed human host and have termed this entity "trichodysplasia spinulosa".

Adaptation to new environments often occurs in the face of gene flow. Under these conditions, gene flow and recombination can impede adaptation by breaking down linkage disequilibrium between locally adapted alleles. Theory predicts that this decay can be halted or slowed if adaptive alleles are tightly linked in regions of low recombination, potentially favouring divergence and adaptive evolution in these regions over others. Here, we compiled a global genomic data set of over 1,300 individual threespine stickleback from 52 populations and compared the tendency for adaptive alleles to occur in regions of low recombination between populations that diverged with or without gene flow. In support of theory, we found that putatively adaptive alleles (F and d outliers) tend to occur more often in regions of low recombination in populations where divergent selection and gene flow have jointly occurred. This result remained significant when we employed different genomic window sizes, controlled for the effects of mutation rate and gene density, controlled for overall genetic differentiation, varied the genetic map used to estimate recombination and used a continuous (rather than discrete) measure of geographic distance as proxy for gene flow/shared ancestry. We argue that our study provides the first statistical evidence that the interaction of gene flow and selection biases divergence toward regions of low recombination.