Chilo iridescent virus (CIV; IIV-6) is the type member of the genus Iridovirus (family Iridoviridae, large icosahedral cytoplasmic DNA viruses). CIV induces death and deformity in the cotton boll weevil, Anthonomus grandis, replicates productively in larvae of the cotton boll weevil, and significantly reduces laboratory populations of the cotton aphid, Aphis gossypii. CIV virion protein extract (CVPE) shuts down host protein synthesis in several insect cell lines and induces mortality in neonate boll weevil larvae. We report here that CVPE induces apoptosis in spruce budworm and boll weevil cell lines, as detected by blebbing, DNA fragmentation, and TUNEL assay. Tissue culture toxicity dose assays (TCTD(50)) showed that spruce budworm cells were eight times more sensitive to CVPE than boll weevil cells. Pancaspase inhibitor suppressed apoptosis but had marginal effect on inhibition of host protein synthesis. Moreover, the CVPE dose for apoptosis was 1000-fold lower than the dose for shutdown of host synthesis. We also detected protein kinase activity in CVPE. Heating CVPE at 60 degrees C for 30 min destroyed all three activities. Our results suggest that one or more polypeptides in CIV induce apoptosis. This is the first study demonstrating apoptosis induction by a member of the genus Iridovirus and by virion extracts of a member of the family Iridoviridae.

The boll weevil, Anthonomus grandis Boheman, is a devastating pest of cotton. Chemical pesticides are problematic due to relative lack of target specificity and resistance. Microbial pesticides may provide viable alternatives because of their narrow host range. Chilo iridescent virus (CIV) is the type species for genus Iridovirus, family Iridoviridae: large, icosahedral cytoplasmic viruses containing a double-stranded DNA genome. Earlier work suggested that CIV replicated in the boll weevil; however, efficiency or production of infectious virus was not established. We showed that CIV undergoes a productive cycle in A. grandis. CIV DNA levels in boll weevil pupae increased significantly from 0 to 3 days post infection. Moreover, virogenic stromata and complete virus particles were observed in the cytoplasm by 7 days. An endpoint dilution assay using viral DNA replication as indicator suggested a 10(5)-fold increase in infectious virus titer over 7 days. This is the first such demonstration in larval infections with genus Iridovirus. Our study establishes that CIV undergoes a productive cycle in the boll weevil and provides an important and useful model system for replication at the organismal level. These results have important implications for the potential of CIV and its components in boll weevil control.

Apoptosis and inhibition of host gene expression are often associated with virus infections. Many viral polypeptides modulate apoptosis by direct interaction with highly conserved apoptotic pathways. Some viruses induce apoptosis during late stages of the infection cycle, while others inhibit apoptosis to facilitate replication or maintain persistent infection. In previous work, we showed that Chilo iridescent virus (CIV) or CIV virion protein extract induces apoptosis in spruce budworm and cotton boll weevil cell cultures. Here, we characterize the product of a CIV gene (iridovirus serine/threonine kinase; istk) with signature sequences for S/T kinase and ATP binding. ISTK appears to belong to the superfamily, vaccinia-related kinases (VRKs). The istk gene was expressed in Pichia pastoris vectors. Purified ISTK (48 kDa) exhibited S/T kinase activity. Treatment with ISTK induced apoptosis in budworm cells. A 35-kDa cleavage product of ISTK retaining key signature sequences was identified during purification. Pichia-expressed 35-kDa polypeptide, designated iridoptin, induced apoptosis and inhibition of host protein synthesis in budworm and boll weevil cells. A mutation in the ATP-binding site eliminated both kinase and apoptosis activity of iridoptin, suggesting that kinase activity is essential for induction of apoptosis. Analysis with custom antibody confirmed that ISTK is a structural component of CIV particles. This is the first demonstration of a viral kinase inducing apoptosis in any virus-host system and the first identification of a factor inducing apoptosis or host protein shutoff for the family Iridoviridae.

Trypsin inhibitor, lectin, phytate, and oligosaccharide levels were measured in the defatted decorticated Cucúrbita seed meals. Species surveyed included potential domesticates Cucúrbita foetidissima and Cucúrbita digitata (xerophytic gourds) as well as Cucúrbita maxima, Cucúrbita moschata, and Cucúrbita pepo (common squashes). Trypsin inhibitor activities in C. foetidissima and C. digitata samples were 5 times greater than in other cucurbits but were only 17% and 24% of that in soybean, the reference material. Heat treatment reduced the activity in all samples to negligible levels. Lectin activity was greatest in soybean, intermediate in C. digitata, and lowest in C. foetidissima and the domesticated cucurbits. Heat treatment substantially reduced the activity of lectins in all samples except those in C. digitata and soybeans. Levels of phytate and sugars were found to be similar among samples.Cultivated species of Cucúrbita have been associated with civilization for centuries (Whitaker and Davies, 1962). Squash, melon, cucumber and other cucurbits are presently supplying variety to human diets. Certain xerophytic Cucúrbita have been identified as possible food crops for arid and semiarid regions of the world. Buffalo gourd, Cucúrbita foetidissima, appears to be the most promising and has received considerable attention with respect to domestication and utilization (Bemis et al., 1978;Hogan

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