Relating Mobility of dsRNA in Nanoporous Silica Particles to Loading and Release Behavior.

Nanoparticle delivery of polynucleic acids traditionally relies on the modulation of surface interactions to achieve loading and release. This work investigates the additional role of confinement in mobility of dsRNA (84 and 282 base pair (bp) sequences of Spodoptera frugiperda) as a function of silica nanopore size (nonporous, 3.9, 8.0, and 11.3 nm). Amine-functionalized nanoporous silica microspheres (NPSMs, ∼10 µm) are used to directly visualize the loading and exchange of fluorescently labeled dsRNA. Porous particles are fully accessible to both lengths of dsRNA by passive diffusion, except for 282 bp dsRNA in 3.9 nm pores. The stiffness of dsRNA suggests that encapsulation occurs by threading into nanopores, which is inhibited when the ratio of dsRNA length to pore size is large. The mobility of dsRNA at the surface and in the core of NPSMs, as measured by fluorescence recovery after photobleaching, is similar. The mobility increases with pore size (from 0.0002 to 0.001 µm2/s for 84 bp dsRNA in 3.9-11.3 nm pores) and decreases with the length of dsRNA. However, when the dsRNA is unable to load into the pores (on nonporous particles and for 282 bp dsRNA in 3.9 nm pores), surface mobility is not detectable. The pore structure appears to serve as a "source" to provide a mobile network of dsRNA at the particle surface. The importance of mobility is demonstrated by exchange experiments, where NPSMs saturated with mobile dsRNA can exchange dsRNA with the surrounding solution, while immobile dsRNA is not exchanged. These results indicate that nanoparticle synthesis techniques that provide pores large enough to take up polynucleic acids internally (and not simply on the external surface of the particle) can be harnessed to design polynucleic acid/nanoporous silica combinations for controlled mobility as a path forward toward effective nanocarriers.

Effect of Confinement in Nanopores on RNA Interactions with Functionalized Mesoporous Silica Nanoparticles.

Amine-functionalized mesoporous silica nanoparticles (MSNPAs) are ideal carriers for oligonucleotides for gene delivery and RNA interference. This investigation examines the thermodynamic driving force of interactions of double-stranded (ds) RNA with MSNPAs as a function of RNA length (84 and 282 base pair) and particle pore diameter (nonporous, 2.7, 4.3, and 8.1 nm) using isothermal titration calorimetry, extending knowledge of solution-based nucleic acid-polycation interactions to RNA confined in nanopores. Adsorption of RNA follows a two-step process: endothermic interactions driven by entropic contribution from counterion (and water) release and an exothermic regime dominated by short-range interactions within the pores. Evidence of hindered pore loading of the longer RNA and pore size-dependent confinement of RNA in the MSPAs is provided from the relative contributions of the endothermic and exothermic regimes. Reduction of endothermic and exothermic enthalpies in both regimes in the presence of salt for both lengths of RNA indicates the significant contribution of short-range electrostatic interactions, whereas ΔH and ΔG values are consistent with conformation changes and desolvation of nucleic acids upon binding with polycations. Knowledge of the interactions between RNA and functionalized porous nanoparticles will aid in porous nanocarrier design suitable for functional RNA delivery.

Genomic architecture of endogenous ichnoviruses reveals distinct evolutionary pathways leading to virus domestication in parasitic wasps.

BACKGROUND: Polydnaviruses (PDVs) are mutualistic endogenous viruses inoculated by some lineages of parasitoid wasps into their hosts, where they facilitate successful wasp development. PDVs include the ichnoviruses and bracoviruses that originate from independent viral acquisitions in ichneumonid and braconid wasps respectively. PDV genomes are fully incorporated into the wasp genomes and consist of (1) genes involved in viral particle production, which derive from the viral ancestor and are not encapsidated, and (2) proviral segments harboring virulence genes, which are packaged into the viral particle. To help elucidating the mechanisms that have facilitated viral domestication in ichneumonid wasps, we analyzed the structure of the viral insertions by sequencing the whole genome of two ichnovirus-carrying wasp species, Hyposoter didymator and Campoletis sonorensis. RESULTS: Assemblies with long scaffold sizes allowed us to unravel the organization of the endogenous ichnovirus and revealed considerable dispersion of the viral loci within the wasp genomes. Proviral segments contained species-specific sets of genes and occupied distinct genomic locations in the two ichneumonid wasps. In contrast, viral machinery genes were organized in clusters showing highly conserved gene content and order, with some loci located in collinear wasp genomic regions. This genomic architecture clearly differs from the organization of PDVs in braconid wasps, in which proviral segments are clustered and viral machinery elements are more dispersed. CONCLUSIONS: The contrasting structures of the two types of ichnovirus genomic elements are consistent with their different functions: proviral segments are vehicles for virulence proteins expected to adapt according to different host defense systems, whereas the genes involved in virus particle production in the wasp are likely more stable and may reflect ancestral viral architecture. The distinct genomic architectures seen in ichnoviruses versus bracoviruses reveal different evolutionary trajectories that have led to virus domestication in the two wasp lineages.

Improving the baculovirus expression vector system with vankyrin-enhanced technology.

The baculovirus expression vector system (BEVS) is a widely used platform for the production of recombinant eukaryotic proteins. However, the BEVS has limitations in comparison to other higher eukaryotic expression systems. First, the insect cell lines used in the BEVS cannot produce glycoproteins with complex-type N-glycosylation patterns. Second, protein production is limited as cells die and lyse in response to baculovirus infection. To delay cell death and lysis, we transformed several insect cell lines with an expression plasmid harboring a vankyrin gene (P-vank-1), which encodes an anti-apoptotic protein. Specifically, we transformed Sf9 cells, Trichoplusia ni High FiveTM cells, and SfSWT-4 cells, which can produce glycoproteins with complex-type N-glycosylation patterns. The latter was included with the aim to increase production of glycoproteins with complex N-glycans, thereby overcoming the two aforementioned limitations of the BEVS. To further increase vankyrin expression levels and further delay cell death, we also modified baculovirus vectors with the P-vank-1 gene. We found that cell lysis was delayed and recombinant glycoprotein yield increased when SfSWT-4 cells were infected with a vankyrin-encoding baculovirus. A synergistic effect in elevated levels of recombinant protein production was observed when vankyrin-expressing cells were combined with a vankyrin-encoding baculovirus. These effects were observed with various model proteins including medically relevant therapeutic proteins. In summary, we found that cell lysis could be delayed and recombinant protein yields could be increased by using cell lines constitutively expressing vankyrin or vankyrin-encoding baculovirus vectors. © 2017 American Institute of Chemical Engineers Biotechnol. Prog., 33:1496-1507, 2017.

De novo sequencing and transcriptome analysis of female venom glands of ectoparasitoid Bracon hebetor (Say.) (Hymenoptera: Braconidae).

Venom is a key-factor in the regulation of host physiology by parasitic Hymenoptera and a potentially rich source of novel bioactive substances for biotechnological applications. The limited study of venom from the ectoparasitoid Bracon hebetor, a tiny wasp that attacks larval pest insects of field and stored products and is thus a potential insect control agent, has not described the full complement and composition of these biomolecules. To have a comprehensive picture of genes expressed in the venom glands of B. hebetor, a venom gland transcriptome was assembled by using next generation sequencing technologies followed by de novo assemblies of the 10.81 M sequence reads yielded 22,425 contigs, of which 10,581 had significant BLASTx hits to know genes. The majority of hits were to Diachasma alloeum, an ectoparasitoid from same taxonomic family, as well as other wasps. Gene ontology grouped the sequences into molecular functions in which catalytic activity with 42.2% was maximum, cellular components in which cells with 33.8% and biological processes among which metabolic process with 30% had the most representatives. In this study, we highlight the most abundant sequences, and those that are likely to be functional components of the venom for parasitization. Full length ORFs of Calreticulin, Venom Acid Phosphatase Acph-1 like protein and arginine kinase proteins were isolated and their tissue specific expression was studied by RT-PCR. Our report is the first to characterize components of the B. hebetor venom glands that may be useful for developing control tools for insect pests and other applications.

De novo sequencing and transcriptome analysis of venom glands of endoparasitoid Aenasius arizonensis (Girault) (=Aenasius bambawalei Hayat) (Hymenoptera, Encyrtidae).

Aenasius bambawalei Hayat (Encyrtidae: Hymenoptera) has been synonymized with Aenasius arizonensis (Girault) is a small, newly discovered endoparasitoid of the cotton mealybug Phenacoccuss solenopsis Tinsley (Pseudococcidae: Hemiptera), which completes its life cycle inside the body of its host and it is a potential insect control tool. Despite the acquired knowledge regarding host-parasitoid interaction, little information is available on the factors of parasitoid origin able to modulate mealybug physiology. The components of A. arizonensis venom have not been well studied but venom from other parasitoids and wasps contain biologically active proteins that have potential applications in pest management or may be of medicinal importance. To provide an insight into the transcripts expressed in the venom gland of A. arizonensis, a transcriptomic database was developed utilizing high throughput RNA sequencing approaches to analyze the genes expressed in venom glands of this endoparasitic wasp. The resulting A. arizonensis RNA sequences were assembled de-novo with contigs then blasted against the NCBI non-redundant sequence database. Contigs which matched database sequences were mostly homologous to genes from hymenopteran parasitoids such as Nasonia vitripennis, Copidosoma floridanum, Fopius arsenus and Pteromalas puparium. Further analysis of the A. arizonensis database was then performed which focused on selected genes encoding proteins potentially involved in host developmental arrest, disrupting the host immune system, host paralysis, and transcripts that support these functions. Sequenced mRNAS predicted to encode full length ORFs of Calreticulin, Serine Protease Precursor and Arginine kinase proteins were identified and the tissue specific expression of these putative venom genes was analyzed by RT-PCR. In addition, results also demonstrate that de novo transcriptome assembly allows useful venom gene expression analysis in a species lacking a genome sequence database and may provide useful information for devising control tools for insect pests and other applications.

Nancy E. Beckage (1950-2012): pioneer in insect host-parasite interactions.

Nancy E. Beckage is widely recognized for her pioneering work in the field of insect host-parasitoid interactions beginning with endocrine influences of the tobacco hornworm, Manduca sexta, host and its parasitoid wasp Apanteles congregatus (now Cotesia congregata) on each other's development. Moreover, her studies show that the polydnavirus carried by the parasitoid wasp not only protects the parasitoid from the host's immune defenses, but also is responsible for some of the developmental effects of parasitism. Nancy was a highly regarded mentor of both undergraduate and graduate students and more widely of women students and colleagues in entomology. Her service both to her particular area and to entomology in general through participation on federal grant review panels and in the governance of the Entomological Society of America, organization of symposia at both national and international meetings, and editorship of several different journal issues and of several books is legendary. She has left behind a lasting legacy of increased understanding of multilevel endocrine and physiological interactions among insects and other organisms and a strong network of interacting scientists and colleagues in her area of entomology.

Specificities of ichnoviruses associated with campoplegine wasps: genome, genes and role in host-parasitoid interaction.

Ichnoviruses (IVs), unique symbiotic viruses carried by ichneumonid campoplegine wasps, derive from integration of a paleo-ichnovirus into an ancestral wasp genome. The modern 'genome' is composed of both regions that are amplified, circularized and encapsidated into viral particles and non-encapsidated viral genomic regions involved in particle morphogenesis. Packaged genomes include multiple circular dsDNAs encoding many genes mostly organized in gene families. Virus particles are assembled in specialized ovarian cells from which they exit into the oviduct lumen; mature virions are injected during oviposition into the insect host. Expression of viral proteins in infected cells correlates with physiological alterations of the host enabling success of parasitism.

A supracellular system of actin-lined canals controls biogenesis and release of virulence factors in parasitoid venom glands.

Parasitoid wasps produce virulence factors that bear significant resemblance to viruses and have the ability to block host defense responses. The function of these virulence factors, produced predominantly in wasp venom glands, and the ways in which they interfere with host development and physiology remain mysterious. Here, we report the discovery of a specialized system of canals in venom glands of five parasitoid wasps that differ in their infection strategies. This supracellular canal system is made up of individual secretory units, one per secretory cell. Individual units merge into the canal lumen. The membrane surface of the proximal end of each canal within the secretory cell assumes brush border morphology, lined with bundles of F-actin. Systemic administration of cytochalasin D compromises the integrity of the secretory unit. We show a dynamic and continuous association of p40, a protein of virus-like particles from a Drosophila parasitoid, L. heterotoma, with the canal and venom gland lumen. Similar structures in three Leptopilina species and Ganaspis xanthopoda, parasitoids of Drosophila spp., and Campoletis sonorenesis, a parasitoid of Heliothis virescens, suggest that this novel supracellular canal system is likely to be a common trait of parasitoid venom glands that is essential for efficient biogenesis and delivery of virulence factors.

Genomic and morphological features of a banchine polydnavirus: comparison with bracoviruses and ichnoviruses.

Many ichneumonid and braconid endoparasitoids inject a polydnavirus (PDV) into their caterpillar hosts during oviposition. The viral entities carried by wasps of these families are referred to as "ichnoviruses" (IVs) and "bracoviruses" (BVs), respectively. All IV genomes characterized to date are found in wasps of the subfamily Campopleginae; consequently, little is known about PDVs found in wasps of the subfamily Banchinae, the only other ichneumonid taxon thus far shown to carry these viruses. Here we report on the genome sequence and virion morphology of a PDV carried by the banchine parasitoid Glypta fumiferanae. With an aggregate genome size of approximately 290 kb and 105 genome segments, this virus displays a degree of genome segmentation far greater than that reported for BVs or IVs. The size range of its genome segments is also lower than those in the latter two groups. As reported for other PDVs, the predicted open reading frames of this virus cluster into gene families, including the protein tyrosine phosphatase (PTP) and viral ankyrin (ank) families, but phylogenetic analysis indicates that ank genes of the G. fumiferanae virus are not embedded within the IV lineage, while its PTPs and those of BVs form distinct clusters. The banchine PDV genome also encodes a novel family of NTPase-like proteins displaying a pox-D5 domain. The unique genomic features of the first banchine virus examined, along with the morphological singularities of its virions (IV-like nucleocapsids, but enveloped in groups like some of the BVs), suggest that they could have an origin distinct from those of IVs and BVs.

Shared and species-specific features among ichnovirus genomes.

During egg-laying, some endoparasitic wasps transmit a polydnavirus to their caterpillar host, causing physiological disturbances that benefit the wasp larva. Members of the two recognized polydnavirus taxa, ichnovirus (IV) and bracovirus (BV), have large, segmented, dsDNA genomes containing virulence genes expanded into families. A recent comparison of IV and BV genomes revealed taxon-specific features, but the IV database consisted primarily of the genome sequence of a single species, the Campoletis sonorensis IV (CsIV). Here we describe analyses of two additional IV genomes, the Hyposoter fugitivus IV (HfIV) and the Tranosema rostrale IV (TrIV), which we compare to the sequence previously reported for CsIV. The three IV genomes share several features including a low coding density, a strong A+T bias, similar estimated aggregate genome sizes ( approximately 250 kb) and the presence of nested genome segments. In addition, all three IV genomes contain members of six conserved gene families: repeat element, cysteine motif, viral innexin, viral ankyrin, N-family, and a newly defined putative family, the polar-residue-rich proteins. The three genomes, however, differ in their degree of segmentation, in within-family gene frequency and in the presence, in TrIV, of a unique gene family (TrV). These interspecific variations may reflect differences in parasite/host biology, including virus-induced pathologies in the latter.

Divergences in protein activity and cellular localization within the Campoletis sonorensis Ichnovirus Vankyrin family.

Ichnoviruses (IVs) occur in obligate symbiotic associations with endoparasitic ichneumonid wasps. IVs are injected with eggs during parasitization, where viral infection and gene expression alter host physiology to ensure endoparasitoid survival. The seven Campoletis sonorensis IV (CsIV) vankyrin genes encode proteins that possess ankyrin repeat domains resembling the inhibitory domains of NF-kappaB transcription factor inhibitors (IkappaBs). The CsIV vankyrins are divided into two subclasses: those expressed primarily in the host fat body (three genes) and those expressed in host hemocytes (four genes). CsIV vankyrin proteins showed limited antigenic similarity when analyzed by Western blotting. Cellular localization and expression patterns of recombinant vankyrin proteins in High Five and Sf9 insect cells differed within and between the subclasses and in cells exposed to lipopolysaccharide, laminarin, or viral immune challenge. In unstimulated Sf9 cells, five vankyrins were detected in cell nuclei. The remaining two proteins localized predominantly to cytoplasmic granules. Immune stimulation of cells resulted in a nuclear-to-cytoplasmic shift of three vankyrins but did not affect localization of other variants. When expressed from recombinant Autographa californica multiple nucleopolyhedroviruses (AcMNPVs), all vankyrins showed a nuclear localization during early stages of infection with patterns resembling those of immune-challenged cells as the infection progressed. Two fat body vankyrins also produced unique biological effects when expressed from recombinant AcMNPV. Insect cells infected with these viruses exhibited enhanced longevity compared to those infected with viruses expressing other vankyrins. Together, these data suggest that vankyrin proteins in CsIV have divergent physiological functions.

Polydnavirus genes that enhance the baculovirus expression vector system.

The baculovirus expression vector system (BEVS) is a powerful and versatile system for protein expression, which has many advantages. However, a limitation of any lytic viral expression system, including BEVS, is that death and lysis of infected insect cells terminates protein production. This results in interruption of protein production and higher production costs due to the need to set up new infections, maintain uninfected cells, and produce pure viral stocks. Genetic methods to slow or prevent cell death while maintaining high-level, virus-driven protein production could dramatically increase protein yields. Several approaches have been used to improve the BEVS and increase the synthesis of functional proteins. Successful enhancement of the BEVS was obtained when various gene elements were added to the virus, secretion and posttranslational processing were modified, or protein integrity was improved. A gene family from the insect virus Campoletis sonorensis ichnovirus (CsIV) was discovered that delays lysis of baculovirus-infected cells, thereby significantly enhancing recombinant protein production in the BEVS system. By using the CsIV vankyrin gene family, protein production in the vankyrin-enhanced BEVS (VE-BEVS) was increased by a factor of 4- to 15-fold by either coexpressing the vankyrin protein from a dual BEVS or by providing its activity in trans by expressing the vankyrin protein from a stably transformed cell line. In sum, VE-BEVS is an enhancement of the existing BEVS technology that markedly improves protein expression levels while reducing the cost of labor and materials.

Potential uses of Cys-motif and other polydnavirus genes in biotechnology.

Exploiting the ability of insect pathogens, parasites, and predators to control natural and damaging insect populations is a cornerstone of biological control. Here we focus on an unusual group of viruses, the polydnaviruses (PDV), which are obligate symbionts of some hymenopteran insect parasitoids. PDVs have a variety of important pathogenic effects on their parasitized hosts. The genes controlling some of these pathogenic effects, such as inhibition of host development, induction of precocious metamorphosis, slowed or reduced feeding, and immune suppression, may have use for biotechnological applications. In this chapter, we consider the physiological functions of both wasp and viral genes with emphasis on the Cys-motif gene family and their potential use for insect pest control.

Response of immunocompetent and immunosuppressed Spodoptera littoralis larvae to baculovirus infection.

The Mediterranean lepidopteran pest Spodoptera littoralis is highly resistant to infection with the Autographa californica multiple nucleopolyhedrovirus (AcMNPV) via the oral route, but highly sensitive to infection with budded virus (BV) via the intrahaemocoelic route. To study the fate of AcMNPV infection in S. littoralis, vHSGFP, an AcMNPV recombinant that expresses the reporter green fluorescent protein gene under the control of the Drosophila heat-shock promoter, and high-resolution fluorescence microscopy were utilized. S. littoralis fourth-instar larvae infected orally with vHSGFP showed melanization and encapsulation of virus-infected tracheoblast cells serving the midgut columnar cells. At 72 h post-infection, the viral foci were removed during the moult clearing the infection. Thus, oral infection was restricted by immune responses to the midgut and midgut-associated tracheal cells. By contrast, injection of BV into the haemocoel resulted in successful infection of tracheoblasts, followed by spread of the virus through the tracheal epidermis to other tissues. However, in contrast to fully permissive infections where tracheoblasts and haemocytes are equally susceptible to infection, a severe limitation to vHSGFP infection of haemocytes was observed. To investigate the resistance of S. littoralis haemocytes to BV infection with AcMNPV, the larval immune system was suppressed with the Chelonus inanitus polydnavirus or a putatively immunosuppressive polydnavirus gene, P-vank-1. Both treatments increased the susceptibility of S. littoralis larvae to AcMNPV. It is concluded that the resistance of S. littoralis to AcMNPV infection involves both humoral and cellular immune responses that act at the gut and haemocyte levels. The results also support the hypothesis that tracheolar cells mediate establishment of systemic baculovirus infections in lepidopteran larvae. The finding that polydnaviruses and their encoded genes synergize baculovirus infection also provides an approach to dissecting the responses of the lepidopteran immune system to viruses by using specific polydnavirus immunosuppressive genes.

Effect of Campoletis sonorensis ichnovirus cys-motif proteins on Heliothis virescens larval development.

Polydnaviruses are obligate symbionts of some parasitic hymenopteran wasps responsible for modifying the physiology of their host lepidopteran larvae to benefit the endoparasite. Injection of Campoletis sonorensis ichnovirus (CsIV) into Heliothis virescens larvae alters larval growth, development and immunity but genes responsible for alterations of host physiology are not well described. Recent studies of polydnavirus genomes establish that these genomes encode families of related genes expressed in parasitized larvae. Here we evaluate five members of the CsIV cys-motif gene family for their ability to inhibit growth and development of lepidopteran larvae. To study the function of cys-motif proteins, recombinant proteins were produced from baculovirus expression vectors and injected or fed to H. virescens larvae in diet. rVHv1.1 was identified as the most potent protein tested causing a significant reduction in growth of H. virescens and Spodoptera exigua larvae. H. virescens larvae ingesting this protein also exhibited delayed development, reductions in pupation and increased mortality. Increased mortality was associated with chronic sub-lethal baculovirus infections. Taken together, these data indicate that the cys-motif proteins have pleiotropic effects on lepidopteran physiology affecting both development and immunity.

Polydnavirus genomes reflect their dual roles as mutualists and pathogens.

Symbionts often exhibit significant reductions in genome complexity while pathogens often exhibit increased complexity through acquisition and diversification of virulence determinants. A few organisms have evolved complex life cycles in which they interact as symbionts with one host and pathogens with another. How the predicted and opposing influences of symbiosis and pathogenesis affect genome evolution in such instances, however, is unclear. The Polydnaviridae is a family of double-stranded (ds) DNA viruses associated with parasitoid wasps that parasitize other insects. Polydnaviruses (PDVs) only replicate in wasps but infect and cause severe disease in parasitized hosts. This disease is essential for survival of the parasitoid's offspring. Thus, a true mutualism exists between PDVs and wasps as viral transmission depends on parasitoid survival and parasitoid survival depends on viral infection of the wasp's host. To investigate how life cycle and ancestry affect PDVs, we compared the genomes of Campoletis sonorensis ichnovirus (CsIV) and Microplitis demolitor bracovirus (MdBV). CsIV and MdBV have no direct common ancestor, yet their encapsidated genomes share several features including segmentation, diversification of virulence genes into families, and the absence of genes required for replication. In contrast, CsIV and MdBV share few genes expressed in parasitized hosts. We conclude that the similar organizational features of PDV genomes reflect their shared life cycle but that PDVs associated with ichneumonid and braconid wasps have likely evolved different strategies to cause disease in the wasp's host and promote parasitoid survival.

Ikappabeta-related vankyrin genes in the Campoletis sonorensis ichnovirus: temporal and tissue-specific patterns of expression in parasitized Heliothis virescens lepidopteran hosts.

Polydnaviruses (PDVs) are unusual insect viruses that occur in obligate symbiotic associations with parasitic ichneumonid (ichnoviruses, or IVs) and braconid (bracoviruses, or BVs) wasps. PDVs are injected with eggs, ovarian proteins, and venom during parasitization. Following infection of cells in host tissues, viral genes are expressed and their products function to alter lepidopteran host physiology, enabling endoparasitoid development. Here we describe the Campoletis sonorensis IV viral ankyrin (vankyrin) gene family and its transcription. The seven members of this gene family possess ankyrin repeat domains that resemble the inhibitory domains of the Drosophila melanogaster NF-kappabeta transcription factor inhibitor (Ikappabeta) cactus. vankyrin gene expression is detected within 2 to 4 h postparasitization (p.p.) in Heliothis virescens hosts and reaches peak levels by 3 days p.p. Our data indicate that vankyrin genes from the C. sonorensis IV genome are differentially expressed in the tissues of parasitized hosts and can be divided into two subclasses: those that target the host fat body and those that target host hemocytes. Polyclonal antibodies raised against a fat-body targeting vankyrin detected a 19-kDa protein in crude extracts prepared from the 3 days p.p. fat body. Vankyrin-specific Abs localized to 3-day p.p. fat-body and hemocyte nuclei, suggesting a role for vankyrin proteins in the nuclei of C. sonorensis IV-infected cells. These data are evidence for divergent tissue specificities and targeting of multigene families in IVs. We hypothesize that PDV vankyrin genes may suppress NF-kappabeta activity during immune responses and developmental cascades in parasitized lepidopteran hosts of C. sonorensis.

Evaluation of early fetal loss induced by gavage with eastern tent caterpillars in pregnant mares.

OBJECTIVE: To determine whether gavage of pregnant mares (housed without access to pasture) with starved eastern tent caterpillars (ETCs) or their excreta is associated with early fetal loss (EFL), panophthalmitis, or pericarditis. DESIGN: Randomized clinical trial. ANIMALS: 15 mares. PROCEDURE: 15 mares with fetuses from 40 to 80 days of gestation (dGa) were randomly assigned to 1 of 3 groups and received 2.5 g of ETC excreta, 50 g of starved ETCs, or 500 mL of water, respectively, once daily for 10 days. Mares were housed in box stalls, walked twice daily, and not allowed access to pasture for 12 days before or during the 21-day trial. RESULTS: 4 of 5 mares gavaged with starved ETCs (group 2) aborted on trial days 8 (2 mares), 10, and 13. No control mares or mares that received excreta aborted. Differences between the ETC group and other groups were significant. Abortion occurred on 49, 64, 70, and 96 dGa. Allantoic fluids became hyperechoic the day before or the day of fetal death. Alpha streptococci were recovered from 1 fetus and Serratia marcescens from 3 fetuses. Neither panophthalmitis nor pericarditis was seen. The abortifacient component of the ETCs was not elucidated. CONCLUSIONS AND CLINICAL RELEVANCE: These findings suggest that mares with fetuses from 40 to 120 days of gestation should not be exposed to ETCs because they may induce abortion.