Molting dynamics and juvenile hormone titer profiles in the nymphal stages of a lower termite, Cryptotermes secundus (Kalotermitidae) - signatures of developmental plasticity

Termites are social cockroaches and this sociality is founded on a high plasticity during development. Three molting types (progressive, stationary and regressive molts) are fundamental to achieve plasticity during alate/sexual development, and they make termites a major challenge to any model on endocrine regulation in insect development. As the endocrine signatures underpinning this plasticity are barely understood, we studied the developmental dynamics and their underlying juvenile hormone OH) titers in a wood-dwelling termite. Cryptotermes secundus, which is characterized by an ancestral life style of living in dead wood and individuals being totipotent in development. The following general pattern elements could be identified during winged sexual development (i) regressive molts were accompanied by longer intermolt periods than other molting types, (ii) JH titers decreased gradually during the developmental transition from larva (immatures without wing buds), to nymph (immatures with wing buds), to winged adult, (iii) in all nymphal stages, the JH titer rose before the next molt and dropped thereafter within the first week, (iv) considerable variation in JH titers occurred in the midphase of the molting cycle of the 2nd and 3rd nymphal instar, inferring that this variation may reflect the underlying endocrine signature of each of the three molting types, (v) the 4th nymphal instar, the shortest of all, seems to be a switch point in development, as nymphs in this stage mainly developed progressively. When comparing these patterns with endocrine signatures seen in cockroaches, the developmental program of Cryprotermes can be interpreted as a co-option and repetitive use of hormonal dynamics of the post dorsal-closure phase of cockroach embryonic development. (C) 2012 Elsevier Ltd. All tights reserved.

Cryptocyanin, a crustacean molting protein: Evolutionary link with arthropod hemocyanins and insect hexamerins

Cryptocyanin, a copper-free hexameric protein in crab (Cancer magister) hemolymph, has been characterized and the amino acid sequence has been deduced from its cDNA. It is markedly similar in sequence, size, and structure to hemocyanin, the copper-containing oxygen-transport protein found in many arthropods. Cryptocyanin does not bind oxygen, however, and lacks three of the six highly conserved copper-binding histidine residues of hemocyanin. Cryptocyanin has no phenoloxidase activity, although a phenoloxidase is present in the hemolymph. The concentration of cryptocyanin in the hemolymph is closely coordinated with the molt cycle and reaches levels higher than hemocyanin during premolt. Cryptocyanin resembles insect hexamerins in the lack of copper, molt cycle patterns of biosynthesis, and potential contributions to the new exoskeleton. Phylogenetic analysis of sequence similarities between cryptocyanin and other members of the hemocyanin gene family shows that cryptocyanin is closely associated with crustacean hemocyanins and suggests that cryptocyanin arose as a result of a hemocyanin gene duplication. The presence of both hemocyanin and cryptocyanin in one animal provides an example of how insect hexamerins might have evolved from hemocyanin. Our results suggest that multiple members of the hemocyanin gene family—hemocyanin, cryptocyanin, phenoloxidase, and hexamerins—may participate in two vital functions of molting animals, oxygen binding and molting. Cryptocyanin may provide important molecular data to further investigate evolutionary relationships among all molting animals.

Performance and physiological responses of combined t-bar and PIT tagged giant mud crabs (Scylla serrata)

Mud crabs (Scylla spp.) are intensively caught throughout South-East Asia and support a very substantial commercial, recreational fishing and aquaculture industry. Identification of individual animals is important to improve understanding and management of this species. However, tagging of crustaceans is difficult as they frequently molt and internal tags can pose a hazard to consumers. In this pilot study we tested a new method combining passive integrated transponder tags and t-bar tags externally. 45 giant mud crabs (Scylla serrata) were captured from the wild and kept in tanks for a maximum of 10 months. We inserted tags into the abdomen of 35 giant mud crabs and tested a modified method where the combined t-bar/PIT-tag was inserted into the muscle tissue of the rear leg between the dorsal carapace plate and the top of the abdominal flap. Tagged crabs with the modified method showed 85% tag retention for molting crabs. We tested the same method in the field where 852 individuals were tagged with combined t-bar/PIT-tags of which 82 were recaptured showing 100% tag retention but without any evidence of molting having occurred. The tested method of combined t-bar/PIT-tags in giant mud crabs can further improve monitoring for wild and aquaculture populations and can be deployed widely with low cost.

Performance and physiological responses of combined t-bar and PIT tagged giant mud crabs (Scylla serrata)

Mud crabs (Scylla spp.) are intensively caught throughout South-East Asia and support a very substantial commercial, recreational fishing and aquaculture industry. Identification of individual animals is important to improve understanding and management of this species. However, tagging of crustaceans is difficult as they frequently molt and internal tags can pose a hazard to consumers. In this pilot study we tested a new method combining passive integrated transponder tags and t-bar tags externally. 45 giant mud crabs (Scylla serrata) were captured from the wild and kept in tanks for a maximum of 10 months. We inserted tags into the abdomen of 35 giant mud crabs and tested a modified method where the combined t-bar/PIT-tag was inserted into the muscle tissue of the rear leg between the dorsal carapace plate and the top of the abdominal flap. Tagged crabs with the modified method showed 85% tag retention for molting crabs. We tested the same method in the field where 852 individuals were tagged with combined t-bar/PIT-tags of which 82 were recaptured showing 100% tag retention but without any evidence of molting having occurred. The tested method of combined t-bar/PIT-tags in giant mud crabs can further improve monitoring for wild and aquaculture populations and can be deployed widely with low cost.

Studies on "Sappe" disease of the silkworm, Bombyx mori : II. Effect of age of larvae on the manifestation of the disease

The effect of age of the larvae on the manifestation of the "Sappe" disease of the silkworm by oral inoculation of different pathogens, viz., Aerobacter cloacae, Pseudomonas boreopolis, Escherichia freundii, Achromobacter delmarvae, A. Superficialis, Pseudomonas ovalis, and Staphylococcus albus was tested. It was found that the reaction of the larva to the pathogen was influenced by its age. Some, e.g., Escherichia freundii, were more lethal when introduced at early stages whereas certain others, e.g., Aerobacter cloacae and Staphylococcus albus, caused maximum damage when invading older larvae. Irrespective of the age of infection, death of the worms mainly occurred during molting and before spinning. The studies also indicated that growth and mortality of the larvae were affected differentially by the pathogens.

Marine Ranching: Proceedings of the Eighteenth U.S.-Japan Meeting on Aquaculture Port Ludlow, Washington, 18-19 September 1989

Female reproduction in penaeid shrimp is carefully regulated by several different endocrine factors. Their precise modes of action have not yet been fully elucidated. Three endocrine factors, each representing a different chemical class of hormones, have been investigated in the penaeid shrimp Sicyonia ingentis in our laboratory: ecdysteroids, vitellogenesis-inhibiting hormone (VIH) , and methyl farnesoate (MF). Ecdysteroids (the steroid molting hormones of arthropods; predominantly 20-hydroxyecdysone), are initially present in low levels (<10 ng/mg) in shrimp embryos. As development of the embryos nears time of hatch, the ecdysteroid levels increase to approximately 150 ng/ mg, indicating that they may be of embryonic origin and involved in embryonic development. An assay was developed for shrimp VIH, which presumably is a protein. Delay of onset of the next reproductive cycle was observed following injection of sinus gland extracts into shrimp that had previously had their eyestalks removed. A photoaffinity analog was synthesized for the putative shrimp reproductive hormone MF-a terpenoid. This analog, farnesyl diazomethyl ketone (FDK) , was used to demonstrate the presence of specific binding proteins for MF in shrimp hemolymph. (PDF file contains 136 pages.)

The effect of timing of tagging on streamer-tag recapture rates for American lobster (Homarus americanus)

Streamer tags are commonly used to study the ecology and population biology of the American lobster (Homarus americanus). Aquarium observations suggest that streamer tag loss, either through tag-induced mortality or tag shedding, is related to the molt stage of the lobster at the time of tagging, and the molting event itself. Tag-induced mortality, where lobsters did not molt, occurred within eleven and sixteen days following tagging for lobsters tagged in postmolt (4%) and late premolt (10%) stages, respectively; whereas no lobsters tagged in early premolt or intermolt stages died. Taginduced mortality at time of molting was observed for lobsters tagged in late premolt stage (11%), and tag shedding was observed for lobsters tagged both in early (25%) and late premolt (11%) stages, but was significantly higher (P=0.014) for lobsters tagged in early premolt stages. Autopsies revealed that lobsters died mainly of organ perforations (hepato-pancreas and pericardial sac) following the tagging process, and rupture of the dorsal thoraco-abdominal membrane during the molting process. The total tag loss was estimated at 4% for lobsters tagged after molting, and 27% and 31% for lobsters tagged in early and late premolt stages, respectively. There was no tag loss for lobsters tagged in the intermolt stage during four months of laboratory observations (July−October). To minimize streamer tag loss, lobsters should be tagged during the intermolt or postmolt stage. Based on field studies, recapture rates for lobsters tagged in premolt stage are always lower than those of lobsters tagged in postmolt stage. Furthermore, recapture rates during the second year, for lobsters that molt in the year following tagging, were drastically reduced, and no lobster was recaptured after four years at large. Finally, to account for tag loss during the first year at large, a minimal adjustment of 24.9% (SD 2.9%) and 4.4% (SD 1.6%) for the recapture rate of lobsters tagged immediately before and after the molting season, respectively, is recommended. Adjustments beyond one year at large are not recommended for the American lobster at this time.

Larval rearing of a freshwater prawn Macrobrachium gangeticum

Observations were made on the larval development of a freshwater prawn, Macrobrachium gangeticum, which revealed that hatching occurred in freshwater but the larvae failed to survive after 2nd molting. Salinity was necessary for survival of the larvae after 2nd molting. The complete larval development involved nine larval stages and the 10th stage was considered to be the post-larva which measured between 4.5 and 5.0 mm in length. All the larval stages were completed within 26 days of hatching. Specimens from each larval stage were taken out and examined under a microscope.

A simple method of tagging prawns

The recognition of individual animals is crucial to many aspects of research. Prawns (Penaeus monodon) present unique difficulties in this respect since they molt regularly. Thus, almost all tagging and marking methods developed for prawns so far have proven inadequate. Some tags or marks are lost during molting; others cause injury to the prawns. A new and efficient method has been developed at the Igang Seafarming Station of the Aquaculture Department. Rectangular brass tags measuring 5 mm by 20 mm and numbered consecutively are used. The prawn is held gently but firmly at the base of the carapace with the left hand while the right hand slips the brass tag around the stalk of the unablated eye and presses the tag gently. All tagging must be made under water to avoid stress. From May 29 to September 7 1977 to a total of 348 unilaterally-ablated adult female prawns were tagged on the unablated eyestalk in 5 batches to enable individual observations on gonadal maturation, molting, and growth. Periodic examinations were made four times a month to coincide with the different phases of the lunar cycle. On each examination, survival and recovery rates were recorded. The data included death due to immediate mortality during ablation and loss to cannibalism for the duration of the experiments. In all five tagging experiments, most of the prawns recovered had their tags intact. These included even dead and molting animals. The eyestalk tagging method is suitable for prawns because the tags can be attached without causing injury and has no effect on the rate of growth, maturity, molting and behavior of the animal. The tags are identifiable and permanent; they remain attached to the animal even after death.

Tolerance of Penaeus monodon larvae to cupric sulfate added in bath

Copper is used to deter the growth of bacterial, fungal and protozoan disease organism in fishes. Zoeae (Z SUB-1 ), myses (M SUB-1 ) and postlarvae (P SUB-1 ) were exposed to copper sulfate at concentrations of 0 . 025, 0 . 05, 0 . 75, 0 . 1 and 0 . 2 ppm from 24 to 96 hours. The number of surviving larvae were counted at the end of each 24-hour period and the percentage of survival is determined for each dose level. The LC SUB-50 for each of the larval stages was interpolated from the data whenever possible. Three trials with 2 replicates per trial were conducted. The physico-chemical characteristics of the bath taken before and at the end of the experimental period show insignificant differences between initial and final values in each trial. Results indicate that mortality rates of all larval stages increased with exposure time and that mortality rates of the experimental group is higher than the control. Interpolation of the LC SUB-50 is possible only for the 48-h and 72-h exposure times for both zoeae and myses and for the 48-h exposure time for the postlarvae. This is due to the high survival percentage of the 24-h group and the low survival percentage (below 50%) of the larvae exposed for 96 hours. The 48-hour LC SUB-50 for Z SUB-1 , M SUB-1 and P SUB-1 are 0 . 225, 0 . 350 and 0 . 125 ppm respectively. Postlarvae seem to be more sensitive than either of the 2 larval stages having a lower 48-h LC SUB-50 and a low survival rate after 72 hours. The larvae were observed to lose their balance and were lethargic, producing few swimming movements so that they were mostly confined to the bottom of the aquaria. Moribund larvae observed under the microscope had a faster but weak heartbeat compared to healthy larvae. Slight or complete loss of feeding ability indicated by empty guts and delayed molting of Z SUB-1 to Z SUB-2 were also noted.

The propagation of the mud crab Scylla serrata (F.) de Haan

The mud crab Scylla serrata is an important commercial species found in many brackish areas in the Philippines. During spawning and hatching, the berried females migrate to the sea. Seeds for pond stocking are obtained from the wild. Because of the unpredictability of seed supply, there is a need to propagate the species artificially. Thus, spawning, larval rearing, maturation, and rematuration of the species are being studied. The first attempts at hatching S. serrata were successful with rates varying between 75% and 90%. Two out of three trials on larval rearing yielded a few megalops. The first zoeal stages were fed diatoms, rotifers, Artemia salina, and bread yeast. Overfeeding programs were implemented during the critical premolting periods to prevent weakening of the larvae and lessen cannibalism. Larval weakening during the premolt makes them susceptible to attacks by fungi like Lagenidium and ciliates like Vorticella. S. serrata larvae survived salinity levels as low as 15 ppt until the 14th day of rearing. Other larvae were able to survive in salinities of 30-32 ppt for 8 to 13 days. Zoeal molting was hastened by lowering the salinity to 25-27 ppt. Artificial broodstocking of juveniles and adult crabs has been made possible using a simple refuge system made of three-compartmented hollow blocks. This system has been helpful in minimizing fighting among crabs. Remarkable growth rates have been observed with feeds like mussel meat and trash fish. Average growth increments of 11 mm carapace length and 20 . 35 g body weight have been observed every fortnight. A newly spent spawner could gain additional weight of 22 . 5 g in only 6 days. Feeding rates of juveniles and adult crabs have been established based on the average body weight from an experiment using mussel meat. Crabs feed more at night. In another experiment, eyestalk ablation was found to be effective in inducing growth and mating. Aside from hastening the molting process, copulation is induced even among the small crabs (average carapace length = 55 mm). Natural mating lasts about 26 hr. A copulation which lasted for seven days with a break in between was observed.

Notes on the induced maturation and spawning in four-month-old Penaeus monodon Fabricius by eyestalk ablation

The ablation technique consisted of making an incision across the eyeball to allow free flow of fluids while holding the prawn under water, squeezing the eyeball contents outwards, and pinching hard the eyestalk tissue. The cut area heals completely in about a week; no application of antibiotics is necessary. Spent spawners were tagged with thin brass rings (Rodriguez, 1976) around the unablated eyestalk for a separate experiment on rematuration. Two spawning yielding approximately 277,000 eggs were obtained three weeks after ablation, followed four days later by two more spawnings with 160,000 eggs; all four spawners weighed more than 100 g. With a hatching rate of 98% and 78% for the first and second batch, respectively, the spawnings produced viable nauplii. Water temperatures as low as 23 degree C due to a delayed cold spell in March depressed molting; weakened larvae had to be discharged at the mysis stage. Although ovarian development continued, no further spawnings were obtained due mainly to the onset of bacterial and fungal disease. Infection is initiated in injured portions of the exoskeleton, sometimes penetrating right through the muscles to the ovarian tissues. The non-flowthrough conditions and mussel meat feeding led to fouling of the culture water resulting in consecutive mortalities caused by disease. Female P.monodon held in maturation pens were ablated at the age of 15 months (Santiago, et al., 1976); they averaged only 16 g body weight after four months growth in ponds. In another experiment, pond-reared P.monodon females ranging from 50 to 80 g were ablated at approximately seven months (Aquacop, 1977). The present results show a minimum age of four months from postlarve that P.monodon is capable of ovarian development and spawning upon ablation. However, maturation is probably affected by size as well as age - the four-month old females weighed an average of 100 g in contrast to the smaller animals in the earlier experiments.

Molecular characterization and effect of RNA interference of retinoid X receptor (RXR) on E75 and chitinase gene expression in Chinese shrimp Fenneropenaeus chinensis

Retinoid X receptor (RXR)/ultraspiracle (USP) is the heterodimeric partner of ecdysteroid receptor and is required for the molting process of arthropods. To better understand the molecular aspects governing the process of molting in shrimp, the full-length cDNA of two RXRs, named as FcRXR-1 and FcRXR-2 were obtained from Chinese shrimp Fenneropenaeus chinensis which were of 1715 and 1700 bp long, revealed a 1315 and 1300 bp open reading frame (ORF) respectively. Quantitative Real time PCR analysis showed a marked tissue-specific difference in the expression of FcRXR transcript, which revealed that the expression of FcRXR Could be regulated in a tissue-specific manner. Moreover, high expression of FcRXR mRNAs was observed in late pre-molt period (D3) and post molt stages (A-B) of shrimp. Among the two isoforms, FcRXR-2 appeared in a considerably high level in all the stages compared to the FcRXR-1. In addition, we examined the temporal expression of two chitinase genes: FcChitinase (FcChi) and FcChitinase-1 (FcChi-1) during the molt cycle of F chinensis. Both the FcChi and FcChi-1 transcripts were detected in all stages of molting, although considerable fluctuations observed through the molt cycle. Injection of double stranded RXR (dsRXR) into juvenile shrimp resulted in a maximum silencing effect at 48 h post injection. We analyzed the expression levels of FcChi, FcChi-1 and the ecdysone inducible gene E75 (FcE75) in samples of dsRXR injected shrimp. Significant reduction in levels of both FcE75, FcChi and FcChi-1 transcripts Occurred in the silenced shrimp. This correlation suggested that RXR might involve in the downstream regulation of E75 and chitinase gene transcription in the ecdysone signaling pathway of decapod crustaceans. (C) 2009 Published by Elsevier Inc.

Molecular characteristics and expression analysis of calreticulin in Chinese shrimp Fenneropenaeus chinensis

Calreticulin (CRT), as an endoplasmic reticulum luminal resident protein, plays important roles in Ca2+ homeostasis and molecular chaperoning. CRT on the surface of the cell can modulate cell adhesion, phagocytosis and integrin-dependent Ca2+ signaling. The full length cDNA of calreticulin (FcCRT) was cloned from Chinese shrimp Fenneropenaeus chinensis. It consists of 1672 by with an open reading frame of 1221 bp, encoding 406 amino acids. This is the first reported cDNA sequence of calreticulin in Crustacea. The deduced amino acid sequence of FcCRT showed high identity with those of Bombyx mori (88%), Drosophila melanogaster (83%), Mus musculus (82%) and Homo sapiens (82%). Highest expression of FcCRT was detected in ovary by Northern blot and in situ hybridization. Different mRNA levels of FcCRT were detected at various molting stages. Expression of FcCRT was induced significantly after 3 h of heat shock treatment, reached the maximum at 4 h and dropped after that. Differential expression profiles of FcCRT were observed in hepatopancreas and haemocytes when shrimp were challenged by white spot syndrome virus (WSSV). From the above results, we inferred that FcCRT might play important roles in Ca2+ homeostasis, chaperoning and immune function in shrimp. (c) 2007 Elsevier Inc. All rights reserved.

Effects of azadirachtin on six inorganic cation distributions in Ostrinia furnacalis (G.)

Azadirachtin (Az), as a botanical insecticide, is relatively safe and biodegradable. It affects a wide vaariety of biological processes, including the reduction of feeding, suspension of molting, death of larvae and pupae, and sterility of emerged adults in a dose-dependent manner. However, the mode of action of this toxin remains obscure. By using ion chromatography, we analyzed changes in six inorganic cation (Li+, Na+, NH4+, K+, Mg2+, and Ca2+) distributions of the whole body and hemolymph in Ostrinia furnacalis (G.) after exposure to sublethal doses of Az. The results showed that Az dramatically interfered with Na+, NH4+, K+, Mg2+, and Ca2+ distributions in hemolymph of O. furnacalis (G.) and concentrations of these five cations dramatically increased. However, in the whole body, the levels of K+, Mg2+, and Ca2+ significantly, decreased after exposure to Az, except that Na+ and NH4+ remained constant. Li+ was undetected in both the control and treated groups in the whole body and hemolymph. It is suggested that Az exerts its insecticidal effects on O. furnacalis (G.) by interfering with the inorganic cation distributions related to ion channels.