Baculoviruses establish systemic infections within susceptible insect hosts, even though host tissues are surrounded by basal lamlnae, extracellular matrices that exclude particles smaller than these viruses. Using a recombinant Autgrapha californwca M nuclear polyhedrosis virus containing a lacZ reporter gene under the control of a constitutive promoter, we followed the progression of infection in Tichoplusia ni larvae. We discovered that infection of the larval insect tracheal system (and not hemocytes, as thought previously) provides the major conduit for this virus to pass through basal laminae and to spread throughout the host. Tracheal epidermal cells, the only known cellular components of the tracheal system, share a common lymph system. Locally these cells contact one another by interdigitating cytoplasmic extensions called epidermal feet. These two features of the tracheal system are likely to facilitate the rapid systemic spread of the virus. The findings reported here have major implications for the fields of insect pathology and biological control and usher in an important consideration regarding host-range factors.Nuclear polyhedrosis viruses (NPV) (family Baculoviridae) are enveloped double-stranded DNA viruses that infect only arthropod hosts, primarily lepidopteran insect larvae. NPVs are unusual among viruses because they produce two phenotypes, one that transmits infection between hosts and one that spreads infection within the host. The first phenotype consists of one or more enveloped virions, each containing one or more nucleocapsids sequestered within a crystalline matrix ofprotein. Such viral occlusions are called polyhedra. As with the spore stages of many bacteria and fungi, polyhedra resist desiccation and allow for temporal escape from inhospitable environments. After ingestion by susceptible larval insect hosts, polyhedra rapidly dissociate in the alkaline gut juices and release occlusion-derived virus (ODV). Even though ODV lack surface spikes or peplomers, they enter midgut columnar epithelial cells by membrane fusion (1-3). These infected epithelial cells produce the second phenotype, budded virus (BV), which buds through the basal cell membrane at sites containing gp64, a viral-encoded glycoprotein that forms spikes on the surface of BV. These gp64-containing structures mediate entry of BV into target cells by promoting fusion of the BV envelope and endocytic vesicular membranes (4-6).Early investigations of baculovirus infections focused on host pathology and the potential use of baculoviruses for biological control purposes. Accurate descriptions of the progression of pathogenesis in virus-infected insects, however, were hampered by the limits of the light and electron microscopy techniques commonly used, both with and without accompanying immunohistochemistry. Rare events are difficult to capture by using these techniques because they require meticulous serial sectioning and the three-dimensional reconstruction of hundreds, if not thousands, of sections for each insect....

As an initial investigation into the long-term evolution of protostellar disks, we explore the conditions required to explain the large outbursts of disk accretion seen in some young stellar objects. We use onedimensional time-dependent disk models with a phenomenological treatment of the magnetorotational instability (MRI) and gravitational torques to follow disk evolution over long timescales. Comparison with our previous two-dimensional disk model calculations (Zhu et al. 2009b indicates that the neglect of radial effects and two-dimensional disk structure in the one-dimensional case makes only modest differences in the results; this allows us to use the simpler models to explore parameter space efficiently. We find that the mass infall rates typically estimated for low-mass protostars generally result in AU-scale disk accretion outbursts,as predicted by our previous analysis (Zhu et al. 2009a. We also confirm quasi-steady accretion behavior for high mass infall rates if the values of α-parameter for the magnetorotational instability is small, while at this high accretion rate convection from the thermal instability may lead to some variations. We further constrain the combinations of the α-parameter and the MRI critical temperature, which can reproduce observed outburst behavior. Our results suggest that dust sublimation may be connected with full activation of the MRI. This is consistent with the idea that small dust captures ions and electrons to suppress the MRI. In a later paper we will explore both long-term outburst and diskevolution with this model, allowing for infall from protostellar envelopes with differing angular momenta.

Larvae of lepidopteran insects commonly become increasingly resistant to baculovirus infections as they age. The mechanism responsible for this development resistance is not known, but the phenomenon does not occur if the viral inoculum is administered intrahemocoelically instead of orally, which is the natural route of infection. This observation indicates that the factors mediating developmental resistance are operative during infection of the primary target tissue, the larval midgut, and not during subsequent systemic infection. To learn more about the mechanism of developmental resistance, we orally inoculated four cohorts of fourth instar Trichoplusia ni larvae with a recombinant of Autographa californica M nuclear polyhedrosis virus expressing a reporter gene. While these cohorts differed only by a few hours in age, we found increasing resistance to infection in successively older cohorts. By assessing the presence and location of infected cells at intervals during the first 48 hr after inoculation, we identified two key factors relevant to the resistance pattern among the developmental cohorts. These factors were: (i) an age-dependent rate of establishing and/or sloughing infected midgut cells and (ii) the ability of fourth instar T. ni to completely clear infection of the midgut epithelium by ecdysis to the fifth instar.

The unintended consequences of gene targeting in mouse models have not been thoroughly studied and a more systematic analysis is needed to understand the frequency and characteristics of off-target effects. Using RNA-seq, we evaluated targeted and neighboring gene expression in tissues from 44 homozygous mutants compared with C57BL/6N control mice. Two allele types were evaluated: 15 targeted trap mutations (TRAP); and 29 deletion alleles (DEL), usually a deletion between the translational start and the 3’ UTR. Both targeting strategies insert a bacterial beta-galactosidase reporter (LacZ) and a neomycin resistance selection cassette. Evaluating transcription of genes in +/- 500 kb of flanking DNA around the targeted gene, we found up-regulated genes more frequently around DEL compared with TRAP alleles, however the frequency of alleles with local down-regulated genes flanking DEL and TRAP targets was similar. Down-regulated genes around both DEL and TRAP targets were found at a higher frequency than expected from a genome-wide survey. However, only around DEL targets were up-regulated genes found with a significantly higher frequency compared with genome-wide sampling. Transcriptome analysis confirms targeting in 97% of DEL alleles, but in only 47% of TRAP alleles probably due to non-functional splice variants, and some splicing around the gene trap. Local effects on gene expression are likely due to a number of factors including compensatory regulation, loss or disruption of intragenic regulatory elements, the exogenous promoter in the neo selection cassette, removal of insulating DNA in the DEL mutants, and local silencing due to disruption of normal chromatin organization or presence of exogenous DNA. An understanding of local position effects is important for understanding and interpreting any phenotype attributed to targeted gene mutations, or to spontaneous indels.

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