The impact of UV-C radiation on the sugar metabolism of the red flour beetle Tribolium castaneum herbst (coleoptera; tenebrionidae).

PURPOSE: Ultraviolet-C (UV-C) is known to induce morphological abnormality in various parts of the red flour beetle, Tribolium castaneum, including its wings, antennae, eyes, legs, and reproductive organs. However, little is known about the effects of UV-C on T. castaneum's sugar content and enzyme activity. MATERIAL AND METHODS: We investigated the concentrations of glucose and trehalose as well as changes in trehalase activity in different developmental stages following UV-C radiation at different exposure periods (1, 2, 4, 8, 16, 32, and 64 min). In addition, the larval mortality and body weight were examined. RESULTS: A reduction in glucose content was recorded in 10-, 15- and 20-day-old larvae and trehalase enzyme activity was recorded in 5- and 10-day-old larvae, whereas an increase in trehalose content was found in adults irradiated with UV-C. In addition, UV-C radiation for 1-64 min caused larval mortality on the first and subsequent days post-irradiation. Moreover, UV-C irradiated larvae exhibited lower body weight, which aligned with the reduction of trehalase activity and glucose content from days 1-6 post-exposure, and the degree of these reductions corresponded to the exposure times. CONCLUSION: UV-C affected sugar content through the reduction of trehalase activity, and glucose declination may cause mortality in T. castaneum; however, further research is needed to provide a better understanding of the impact of UV-C on sugar metabolism.

UV-C radiation during the pupal stage affects morphological changes of wings in Tribolium castaneum (Col; Tenebrionidae).

Purpose: To reveal the effects of Ultraviolet-C (UV-C) on the elytra and hindwing morphology of Tribolium castaneum. Material and methods: Zero-day-old-pupae were irradiated with UV-C at a distance of 35 cm for 1, 2, 4, 8, 16, 32, or 64 min. Changes in wing morphologies were examined using light and scanning electron microscope. Results: UV-C radiation decreased the adult emergence rate and the insect body mass. Morphological changes of the elytra and hindwings in the adults were classified into nine grades. The treated insects had wrinkled and split elytra, and hindwings were not folded properly. Radiation altered the size of elytra, hindwings and wing shape. An analysis of the color intensity indicated that the irradiated beetles had darker elytra. The veins of hindwings became darker, while the membranous area had a lighter color than the control. UV-C radiation also affected the thickness of the elytra. Scanning electron microscopy revealed that UV-C caused deformity of elytra surface and decreased the number of hair sensilla. Conclusions: Results indicate that the elytra and hindwing morphology were altered by UV-C radiation. However, further analysis is required to evaluate the response of T. castaneum to UV-C radiation at the gene level.

Expression of Heat Shock Protein Genes in Different Developmental Stages and After Temperature Stress in the Maize Weevil (Coleoptera: Curculionidae).

The maize weevil, Sitophilus zeamais Motschulsky, is a major pest of rice and other postharvest grain stocks in tropical countries. Heating and cooling treatments have been adopted to control this pest. Because heat shock protein (hsp) genes respond to temperature stress, we examined the association of hsp genes with development and thermal stress in S. zeamais. The temperature response of the insect to heat and cold treatments was assessed at four developmental stages: egg, larva, pupa, and adult. LT50 values at high temperatures were similar among the four developmental stages, while adults were the most tolerant to low temperatures, and eggs, larvae, and pupae exhibited similar LT50 values. Expression levels of three hsps--Szhsp70, Szhsc70, and Szhsp90--fluctuated substantially throughout the four stages at a rearing temperature of 28°C. Heat shock and cold shock increased the expression of all three hsps, and the highest upregulation was observed at 40°C, although the intensity of upregulation varied among the three genes: strongly in Szhsp70, moderately in Szhsp90, and slightly in Szhsc70. Basal expression of the three hsps at 28°C and gene responses to heat and cold shock also varied significantly at the tissue level.

Trehalase activity in fungus-growing termite, Odontotermes feae (Isoptera: Termitideae) and inhibitory effect of validamycin.

Trehalase is the hydrolytic enzyme that catalyzed the hydrolysis of trehalose to glucose. In this study, trehalase activity in the fungus-growing termite, Odontotermes feae Wasmann had been examined. Trehalase activity in digestive tract and carcass of O. feae was higher than that in wood-feeding termite, Coptotermes gestroi Wasmann. The intestinal tract of worker caste of O. feae was the main source of trehalase compared with that in salivary, fat body, and carcass. In particular, the highest activity was found in the midgut and hindgut parts. More specifically, the contents of midgut and hindgut had higher enzyme activity compared with that trehalase prepared from their epithelial tissue. The enzyme activity of gut trehalase in three different termite castes, worker, soldier, and reproductive, had been determined. The result showed that female alate had the highest activity, followed by worker, male alate, and soldier castes. Trehalose concentration in the reproductive caste was at lowest level, while soldier and worker contained the high trehalose concentration. This study indicates that high trehalase activity locates in the midgut and hindgut contents and change in trehalase activity in fungus-growing termite is caste-specific. Validamycin inhibited trehalase activity of O. feae in vivo and caused high mortality, indicating that this trehalase inhibitor is valuable tools for termite control.

Regulation of soluble and membrane-bound trehalase activity and expression of the enzyme in the larval midgut of the bamboo borer Omphisa fuscidentalis.

Diapausing larvae of Omphisa fuscidentalis contain soluble and membrane-bound trehalase in the midgut. Soluble trehalase activity accounts for three-fourths of the total trehalase activity in midgut homogenates. The exposure of diapausing larvae to juvenile hormone analog (JHA) induced pupation, accompanied by an increase in soluble trehalase activity at the beginning of the prepupal period. Injection of 20-hydroxyecdysone (20E) increased the level of soluble trehalase activity 5 days postinjection in a dose-dependent manner. In contrast, no increase in membrane-bound trehalase activity was observed under the same conditions. We cloned the cDNAs that encode the soluble and membrane-bound forms of trehalase in O. fuscidentalis trehalase-1 (OfTreh-1) and trehalase-2 (OfTreh-2), respectively. Treh-1 encodes a 581-aa protein while Treh-2 encodes a 648-aa protein with one putative transmembrane domain near the C-terminus. The mRNA expression level of Treh-1 was 27-fold higher than that of Treh-2 in diapausing larval midgut. Following the exposure of diapausing larvae to JHA, Treh-1 mRNA expression increased gradually until the prepupal period whereupon it increased dramatically; in contrast, the mRNA expression of Treh-2 remained at its initial level. Similarly, 20E upregulated Treh-1 expression but had no effect on Treh-2 expression. Taken together, these results suggest that an increase in the soluble trehalase activity at pupation is caused by upregulation of Treh-1 gene. Moreover, membrane-bound trehalase does not appear to be involved in the dynamic changes in the hemolymph trehalose concentration that occur during the larval-pupal transformation.

Characteristic expression of three heat shock-responsive genes during larval diapause in the bamboo borer Omphisa fuscidentalis.

Three heat shock-responsive genes, Ofhsp70, Ofhsc70, and Ofhsp90, were identified in Omphisa fuscidentalis. These genes encode proteins with molecular weights of 70, 72, and 82 kDa, respectively, and were upregulated during heat shock. Ofhsp70 was expressed actively in the pre-diapause period, but its expression was low during diapause. While Ofhsc70 expression was low during pre-diapause and the first half of diapause, its expression increased greatly in the second half before diapause termination. In contrast, Ofhsp90 expression decreased by half during the transition to diapause and conspicuously decreased during late diapause. The results suggest that Ofhsp90 is associated with the maintenance of diapause and Ofhsc70 with the termination of diapause, whereas the expression of Ofhsp70 is unrelated to diapause. The insect molting hormone 20-hydroxyecdysone (20E) and juvenile hormone analog (JHA) caused increases in the mRNA expression of Ofhsc70, but not Ofhsp70 or Ofhsp90, in diapausing larvae. These transcriptional responses to 20E are consistent with the changes in ecdysteroid titer in the hemolymph relative to gene expression during diapause.

Hormonal mechanisms underlying termination of larval diapause by juvenile hormone in the bamboo borer, Omphisa fuscidentalis.

Topical application of methoprene, a juvenile hormone analogue (JHA), induces pupation by activating the prothoracic glands (PGs) in diapausing larvae of the bamboo borer, Omphisa fuscidentalis. To determine the minimum stimulation period for PG activation, we transplanted PGs of JHA-treated larvae (donors) into non-treated larvae (recipients) on successive days after JHA treatment and observed the recipients for pupation. JHA stimulation for 1 day was sufficient to induce pupation. In recipient larvae, the hemolymph ecdysteroid titer increased transiently on day 18 after transplantation and significantly on days 24-28, prior to pupation. Secretory activity of recipient PGs increased transiently on day 16 and days 22-28. Because the recipient PG activity was too low to account for an increased ecdysteroid titer, the JHA-stimulated donor PGs must produce the major part of hemolymph ecdysteroids. In addition, the ecdysteroid produced by the donor PGs might have stimulated the recipient PGs. We examined the possible involvement of two ecdysone receptor (EcR) isoforms, OfEcR-A and OfEcR-B1, in PG activation by JHA, and found that although both isoforms were up-regulated, accompanied by an increased ecdysteroid titer in the hemolymph, the isoform mRNA levels were not altered at all before the increase in PG secretory activity. Thus, EcR expression might not be involved in feedback activation of PGs.

Dual control of midgut trehalase activity by 20-hydroxyecdysone and an inhibitory factor in the bamboo borer Omphisa fuscidentalis Hampson.

During larval diapause lasting 9 months from September to May, trehalase activity in the midgut of the bamboo borer Omphisa fuscidentalis Hampson (Lepidoptera: Crambidae) was low from December to April, followed by a fourfold increase in May that remained high during the pupal stage in July. An application of juvenile hormone analog (JHA) produced increases in the ecdysteroid titer, while trehalase activity was increased by both JHA and 20-hydroxyecdysone (20E) injection. The trehalase activity in the midgut of diapausing larvae was doubled by incubating the midgut with 20E for 48h. During diapause as well as after JHA application, expression of two ecdysone receptor isoform genes (EcR-A and EcR-B1) in the midgut increased simultaneously with the increase in hemolymph ecdysteroid titer, followed by an increase in trehalase activity. The hemolymph of diapausing larvae contained a trehalase inhibitor and inhibitory activity was high during diapause. After 20E injection, trehalase inhibition decreased as midgut trehalase activity increased. Taken together, at least two factors may participate in the change in midgut trehalase activity: increase in trehalase activity and decrease in trehalase inhibitor activity, both of which may be induced by 20E.

Identification, characterization, and developmental regulation of two storage proteins in the bamboo borer Omphisa fuscidentalis.

Two insect storage proteins, OfSP1 (75 kDa) and OfSP2 (72 kDa), were purified using three different chromatographies from the hemolymph of Omphisa fuscidentalis larvae during diapause, and their genes were cloned. OfSP1 and OfSP2 concentrations in the hemolymph were high during diapause. During pupation, OfSP1 levels decreased in the male hemolymph and disappeared from the female hemolymph. OfSP1 and OfSP2 mRNA levels in the fat bodies were low during the third instar, but increased greatly during the fourth and fifth larval instars. During diapause, mRNA expression continued at a lower level than during the feeding period. The injection of 20-hydroxyecdysone (20E) into diapausing larvae caused an increase in OfSP1 and OfSP2 mRNA levels 2-3 days post-injection, followed by a decrease in expression until pupation, which occurred 2-4 days thereafter. When larvae were treated with juvenile-hormone analog (JHA), OfSP1 and OfSP2 mRNA levels gradually decreased until the onset of pupation. In Omphisa, OfSP1 and OfSP2 proteins are produced and released by the larval fat bodies in the fourth and fifth-instar larvae, and the proteins accumulate in the hemolymph until the insects enter diapause. OfSP1 may be reabsorbed by the fat bodies at the end of diapause for subsequent re-use during pupation.

Correlation of oxygen consumption, cytochrome c oxidase, and cytochrome c oxidase subunit I gene expression in the termination of larval diapause in the bamboo borer, Omphisa fuscidentalis.

The moth Omphisa fuscidentalis (Lepidoptera, Pyralidae) is a univoltine insect with a larval diapause period lasting up to 9 months. We studied changes in O(2) consumption in conjunction with cytochrome c oxidase activity and cytochrome c oxidase subunit I (cox1) gene expression. O(2) consumption changed within a day, showing a supradian rhythm with a ca.12-h cycle at 25 degrees C. During the first two-thirds of the diapause period, from October to March, O(2) consumption was constant until January and then increased by March. Topical application of methoprene, a juvenile hormone analog (JHA), to diapausing larvae terminated the diapause and was associated with an increase in O(2) consumption rate at diapause termination. In JHA-treated larvae, cytochrome c oxidase activity in fat bodies was high at the beginning of the prepupal period and highest at pupation. cox1 expression in fat bodies displayed a transient peak 8 days after JHA application and peaked in the prepupal period. Taken together, our results show that the break of diapause by JHA is associated with the activation of cox1, bringing about an increase in cytochrome c oxidase activity, followed by an increase in O(2) consumption rate.