Pleonal muscle development in the shrimp Penaeus (Litopenaeus) vannamei (Crustacea: Malacostraca: Decapoda: Dendrobranchiata)

Penaeoidean shrimp pleonal muscle is a valuable economic resource worldwide, but little is known of its development during larval stages. The development of pleonal muscle in Penaeus (Litopenaeus) vannamei was studied by rhodamine-phalloidin staining and laser-scanning confocal microscopy. Dorsal pleonal muscle was first evident at the protozoea I stage while ventral pleonal muscle was present by the protozoea II stage. Identifiable ventral pleonal muscles were evident by the protozoea III stage and all ventral muscle types were present in the mysis I. The tail flex response began at the mysis stage and growth of existing pleonal muscles continued. The pleopods formed during the mysis stages, with coxal and basis muscles developed by mysis III. The pleopods became functional beginning with the first post-larval stage. We conclude that the pleonal muscle pattern of P. vannamei larvae is similar to that of adult Penaeus setiferus, and that homologous muscles are present. The major formation of dorsal pleonal muscles occurs during the protozoea II stage, while significant development of ventral pleonal muscles occurs during the protozoea III stage.     

Cleavage and gastrulation in the shrimp Penaeus (Litopenaeus) vannamei (Malacostraca, Decapoda, Dendrobranchiata)

While most malacostracan crustaceans develop through superficial cleavage, in the Amphipoda, Euphausiacea, and Dendrobranchiata (Decapoda) cleavage is complete. Euphausiaceans and dendrobranchiate shrimp share a similar early cleavage pattern, early cleavage arrest and ingression of mesendoderm progenitor cells, a ring of crown cells (prospective naupliar mesoderm) around the blastopore, and hatching as a nauplius larva. Yet recent phylogenies do not support a close relationship between Euphausiacea and Decapoda. In addition, some variation is reported in the timing of mesendoderm cell arrest and number of crown cells for a number of dendrobranchiates. To determine the representative pattern of development in the Dendrobranchiata, embryos of the Pacific white shrimp Penaeus (Litopenaeus) vannamei were stained with Sytox Green to label chromosomes and nuclei and examined with confocal microscopy. The early cleavage pattern, mesendoblast arrest and subsequent ingression at the 32-cell stage, presence of 8 initial crown cells, and fates of the mesendoblasts are the same for P. vannamei (family Peneaeidae) and Sicyonia ingentis (family Sicyoniidae). The lineage of the primordial endoderm cells differs from that reported for P. kerathurus. These characters were discussed in the context of the evolution of development in the Dendrobranchiata and in comparison to the Euphausiacea.     

The basic isoelectric form of alpha-L-fucosidase from the hepatopancreas of the shrimp Penaeus monodon (Crustacea: Decapoda).

1. alpha-L-Fucosidase was purified ca 10,889-fold to homogeneity from Penaeus monodon, with a final spec. act. of 31,250 U/mg of protein. 2. By using SDS-polyacrylamide gel electrophoresis, the monomers of shrimp alpha-L-fucosidase were discovered to have mol. wts of 63,000 and those of human placental enzyme, 46,000 and 20,000. Since the active shrimp alpha-L-fucosidase was found to have a mol. wt of 233,000 by Superose 12 FPLC, it was concluded that the purified shrimp enzyme was tetrameric. 3. In contrast to the discovery of thermolability with human placental alpha-L-fucosidase, the shrimp enzyme was found to be stable to heating at 65 degrees C for 10 min. 4. The shrimp alpha-L-fucosidase has an isoelectric point (pI) of 8.5, but the human placental enzyme has a pI of 4.0. The shrimp enzyme was sialyated. 5. The shrimp alpha-L-fucosidase has a pH optimum at 5.5 and its Km was 22.2 microM with 4-methyl-umbelliferyl-alpha-L-fucopyranoside as substrate. The human enzyme has a broad pH optimum between 5.0 and 6.5.     

Allatostatins of the tiger prawn,Penaeus monodon (Crustacea: Penaeidea)

More than 40 peptides belonging to the -Y/FXFGL-NH(2) allatostatin superfamily have been isolated and identified from the central nervous system (CNS) of the tiger prawn, Penaeus monodon (Crustacea: Penaeidea). The peptides can be arranged in seven sub-groups according to the variable post-tyrosyl residue represented by Ala, Gly, Ser, Thr, Asn, Asp, and Glu. Two of the residues (Thr and Glu) have not been observed in this position previously in either insects or crustaceans. Also reported for the first time for allatostatins, two of the peptides are N-terminally blocked by a pyroglutamic acid residue. The yields of certain peptides with similar amino acid sequences to each other were, in some instances, very different. As an example, the yield of ANQYTFGL-NH(2) was 2pmol, compared with ASQYTFGL-NH(2), with a yield of 156 pmol. There are several possibilities to account for this. If, as in all species so far investigated, there is a single allatostatin gene in P. monodon, then it would appear that different sub-populations have contributed mutant forms of particular peptides to the extract. Another, less likely possibility is that this species has more than one allatostatin gene, producing a variable array of peptides albeit in different molar ratios. Several peptides were present apparently as a result of the loss of one or more residues at the N-terminus of a larger form, either due to N-terminal degradation or specific post-translational processing. The number of peptides identified exceeds that for any other insect or crustacean species previously investigated. None is identical to any of the 60-70 insect allatostatins so far identified, and only three are common to other crustaceans. Immunohistochemical study of the CNS of P. monodon, with the same antisera as used to monitor the purification, confirms the widespread nature and complexity of allatostatinergic neural pathways in arthropods. Thus, all neuromeres of the brain, and all except one of the ventral cord ganglia, possess allatostatin neurons and extensive areas of allatostatin-innervated neuropile. In addition to the cytological evidence that the allatostatins act as neurotransmitters, associated with tissues as varied as eyes and legs, their presence in neurohemal areas such as the sinus gland and the perineural sheath of the thoracic ganglia suggests a neuroendocrine function. As well as posing a challenge to physiologists assigning specific functions to the allatostatins, their extensive intra-species multiplicity, linked to their inter-species variability, also presents a complex problem to geneticists and evolutionists.     

Cleavage and gastrulation of the euphausiacean<Emphasis Type="Italic"> Meganyctiphanes norvegica</Emphasis> (Crustacea,Malacostraca)

According to the Articulata hypothesis the cleavage of arthropods must be derived from spiral cleavage. However, arthropods show a great variety of cleavage modes with a widespread occurrence of superficial cleavage. In the Malacostraca, holoblastic cleavage occurs in some taxa such as Amphipoda, Euphausiacea and Dendrobranchiata. In particular, the cleavage of euphausiaceans has been proposed to be a modified spiral cleavage. The cell lineage of early stages up to blastoderm formation of the euphausiacean Meganyctiphanes norvegica is reconstructed using recent methods of fluorescent staining. Only the oblique angle of the mitotic spindles during the transition from the 2- to the 4-cell stage resembles the spiral cleavage mode. At the 8-cell stage, four cells each form a pattern of two interlocking bands which is preserved until the 122-cell stage. One blastomere is delayed in division and shows an oblique division from the fourth cleavage on. It is the precursor cell of two enlarged and cleavage-arrested cells at the 32-cell stage. At the 62-cell stage, these two cells are surrounded by eight cells following a specific cell division pattern during the subsequent division cycles. The cleavage pattern of M. norvegica occurs in two mirror images. A comparative approach reveals distinct similarities between the early cleavage patterns of Euphausiacea and Dendrobranchiata which are suggested to be homologous. Furthermore, the relationships to non-malacostracan cleavage patterns are discussed. It is shown that the early cleavage pattern of M. norvegica does not offer an example of a spiral cleavage within arthropods.     

Molecular detection methods developed for a systemic rickettsia-like bacterium (RLB) in Penaeus monodon (Decapoda: Crustacea)

Molecular detection methods were developed to aid in the diagnosis of a rickettsia-like bacterium (RLB) which caused severe mortalities of farm-raised Penaeus monodon in Madagascar. Using primers derived from the 16S rRNA gene of bacteria, a PCR assay was optimized to amplify this region of the genome of the RLB, using extracted DNA from infected P. monodon tissue as the template. The resulting amplified PCR product was sequenced and 2 novel primers were selected from the variable region of the gene. These primers amplified a 532 bp fragment of DNA originating from the rickettsia-infected samples. The PCR assay was optimized and tested on DNA extracted from specific pathogen-free (SPF) P. vannamei tissue and several other strains of bacteria. The PCR assay with the rickettsia-specific primers was specific for this RLB and did not amplify the other DNA samples tested. The 532 bp PCR-amplified fragment was labeled with digoxigenin (DIG) for in situ hybridization assays. This probe was tested on SPF, RLB and bacteria-infected shrimp specimens preserved in Davidson's fixative. The probe was specific for both natural and experimental rickettsial infections. Hybridization with this probe required a stringent temperature of 65 degrees C, otherwise cross-reactivity was observed with other types of bacteria.     

Burrowing shrimp of the infraorders Gebiidea and Axiidea (Crustacea: Decapoda)

This review is devoted to the different aspects of biology of burrowing shrimp of the infraorders Gebiidea and Axiidea (Crustacea: Decapoda). Information on their taxonomy, morphology and physiology, distribution, burrow architecture, and trophic mode is reviewed. The peculiarities of breeding and development, life history, and the role in the ecosystem of these infaunal species are analyzed. Special attention is given to the species that inhabit Peter the Great Bay of the Sea of Japan.     

Ontogeny of gut morphology in the white shrimp Penaeus setiferus (Decapoda,Penaeidae)

Ontogeny of the gut in Penaeus setiferus was investigated by reconstruction of serial sections examined by light microscopy. Development of the gut into the adult form is protracted over several weeks beyond metamorphosis in steps that may be directly related to the unique postlarval life history of Penaeus. The gastric mill is lacking in larval stages of P. setiferus. In protozoeal stages Z₁-Z₃, the pyloric ampullae are blind sacs that do not communicate with the midgut. The gland filter first appears in mysis stage M₂. The gastric mill in early postlarval (PL) stages consists of poorly chitinized lobes with flexible setae. By PL₂₁ the ossicles of the gastric mill are rigid and setae are replaced by spine-like denticles, but even by PL₃₅ the gastric mill is neither as massive nor heavily chitinized as in adults. During the mysis stages and early PL stages, the hepatopancreas communicates freely with both the foregut and the midgut trunk. By PL₃₅ the hepatopancreatic ducts are essentially isolated from the remainder of the midgut by foregut ossicles. The midgut in Z₁ consists of two pairs of simple caeca and the midgut trunk. During larval growth, each of the lateral midgut caeca develops into a number of lobes. After metamorphosis these lobes begin to ramify into small-diameter tubules, and by PL₃₅ have completely ramified into the hepatopancreas of adults. From M₁ to PL₄, the anterior midgut caeca decrease in absolute size and become a single anterior diverticulum. The posterior midgut diverticulum first appears in PL₂₁ as a simple sac and thereafter increases in size and complexity.     

Identification of abundant and informative microsatellites from shrimp (Penaeus monodon) genome

Microsatellites were isolated from P. monodon genomic libraries by direct sequencing of recombinant clones without probe screening. Forty-nine out of 83 clones sequenced contained 99 microsatellite arrays of three or more repeats. When five or more and ten or more repeats were considered, 28 and 14 microsatellites were detected, respectively. The 99 microsatellites were classified as perfect (75%), imperfect (6%), compound perfect (3%) and compound imperfect (16%). The abundance of di-, tri-, tetra- and hexanucleotide repeats were 67%, 20%, 9% and 3%, respectively. The dinucleotide repeats included 36 (CT)n, 31 (GT)n, 17(AT)n and 3 (CG)n. One octanucleotide repeat (ATTTATTC)5 was found within a large repeat sequence. Optimal annealing temperatures were determined for PCR using 11 primer sets encompassing 15 microsatellites. Ten primer sets provided successful amplifications with allele sizes generally ranging from 139 to 410 bp. All these primers amplified polymorphic loci with PIC values ranging from 0.63 to 0.96. Two primer sets amplified additional bands which can easily be distinguished from the bands of the main locus. Three out of 10 P. monodon microsatellites also amplified alleles in P. vannamei. The abundance and informative nature of P. monodon microsatellites and their potential for cross-species amplification make them useful for genetic studies.     

Early embryonic development of the freshwater shrimp Caridina multidentata (Crustacea,Decapoda, Atyidae)

Decapod crustaceans show a great diversity of developmental modes at all levels. In particular, early cleavage varies from total via mixed to superficial modes and from determinate cleavage with a stereotyped pattern to indeterminate cleavage. However, the ground pattern of early decapod development is not clear. To address this problem, we studied the early embryonic development of the caridean shrimp Caridina multidentata with a combination of confocal laser scanning microscopy, scanning electron microscopy, 4D microscopy and 3-D reconstruction software. Despite a yolk-rich egg, the cleavage is holoblastic and shows a distinct pattern of blastomere arrangement, characterized by two interlocking cell bands. This resembles the conditions in dendrobranchiate shrimps, which most likely are the sister group to Pleocyemata to which C. multidentata belongs. Hence, our results offer the possibility to assume total cleavage with blastomeres arranged in two interlocking cell bands as ancestral cleavage mode for Decapoda.     

A sialidase from the hepatopancreas of the shrimp Penaeus japonicus (Crustacea: Decapoda): reversible binding with the acidic beta-galactosidase

1. The sialidase purified from the hepatopancreas of Penaeus japonicus is able to bind the acidic beta-galactosidase in vitro. No protective protein, Mr 32,000, was detected in either purified enzyme preparation. 2. The specific activity of the isolated sialidase is 55.0 mU/mg of protein. After polyacrylamide gel electrophoresis under denaturing conditions, the purified shrimp enzyme was found to consist of monomers of Mr 32,000. 3. The sialidase from shrimp has an isoelectric point (pI) of 4.6 +/- 0.1. 4. The shrimp enzyme has the pH optimum at 5.0 and its Km was 5.5 microM with 2'-(4-methylumbelliferyl)-alpha-D-N-acetylneuraminic acid as substrate. The enzyme activity was inhibited by either Hg2+ or Cu2+ ions.     

Introducing foreign DNA into tiger shrimp (Penaeus monodon) by electroporation

Electroporation was used to introduce pFLAG-CMV-1-BAP, a DNA fragment that includes a bacterial alkaline phosphatase gene driven by a human cytomegalovirus (CMV) promoter, into Penaeus monodon zygotes. The transgenic tiger shrimp was achievedby using 10kV, 28 pulses, 120 g sec pulse time, 10 cycles, and a DNA concentration of 37.5 microg/mL. The hatching rate of electroporated zygotes (46%) was significantly lower than that of zygotes in the untreated group (89%). The survival rate of postlarvae in the electroporated group using a DNA concentration of 37.5 microg/mL decreased from 0.6% for postlarva 45 to 0.4% for postlarva 120. Based on dot blot analysis, the rate of gene transfer was 37% in mysis-stage, 23% postlarva 15(PL15), 19% postlarva 45(PL45), and 21% 4-month-old (about PL120). Genomic Southern blotting demonstrated that DNA from transgenic tiger shrimp contained fragments of exogenous DNA that were smaller, larger and of the same molecular size as pFLAG-CMV-1-BAP. Transferred DNA fragments were integrated into the genomes of 31% of the transgenic tiger shrimp. The exogenous DNA was mosaically distributed in a wide variety of tissues. Immunohistochemical staining revealed that the FLAG-BAP fused-protein encoded by pFLAG-CMV-1-BAP was present in the ovaries of some transgenic tiger shrimp.     

Phylogenetic analysis of the Malacostraca (Crustacea)

The Malacostraca comprises about 28 000 species with a broad disparity in morphology, anatomy, embryology, behaviour and ecology. The phylogenetic relationships of the major taxa are still under debate. Is the Leptostraca the sister group of the remaining Malacostraca, or is this taxon more closely related to other Crustacea? Does the Stomatopoda or the Bathynellacea represent the most basal taxon within the remaining taxa? Is the Peracarida monophyletic or are some peracarid taxa more closely related to other ‘caridoid’ taxa? Is the Thermosbaenacea part of the Peracarida or its sister group, and how much support is there for a taxon Amphipoda + Isopoda? To answer these questions a phylogenetic analysis of the Malacostraca combining different phylogenetic approaches was undertaken. In a first step, the monophyly of the Malacostraca including the Leptostraca is shown using the ‘Hennigian approach’. A computer cladistic analysis of the Malacostraca was carried out with NONA and PEE ‐WEE , based on 93 characters from morphology, anatomy and embryology. Nineteen higher malacostracan taxa are included in our analysis. Taxa whose representatives are exclusively fossils were not included. The Leptostraca was used as an operational out‐group. The present analysis supports the basal position of the Stomatopoda. Syncarida and Peracarida (including Thermosbaenacea) are supported as monophyletic, the Eucarida is not. Instead a sister‐group relationship is suggested between Euphausiacea and Peracarida (including Thermosbaenacea), with the Syncarida as the sister group to both taxa. Certain embryonic characters are interpreted as support for the monophyly of the Peracarida (without Thermosbaenacea) because convergences or reversals of these characters seem implausible. Within the Peracarida, the Mysidacea (Lophogastrida + Mysida) represents the sister group to the remaining taxa. A sister‐group relationship between Amphipoda and Isopoda is not supported.     

Gill-cleaning mechanisms of a dendrobranchiate shrimp, Rimapenaeus similis (Decapoda, penaeidae): description and experimental testing of function.

Observations on functional morphology and results from experiments demonstrate that setiferous epipods compose the major gill-cleaning mechanism in a penaeoid shrimp, Rimapenaeus similis. Epipods on the second maxillipeds and on pereopods 1-3 are equipped with long setae bearing an array of digitate scale setules. These multidenticulate setae reach to most gills and are jostled among them during limb movements. Experiments were performed in which epipods were removed from the gill chamber on one side (experimental) but not the other (control); treated animals were exposed to fouling in a recirculating water system for 2 weeks. Particulate fouling, measured by reduction in relative gill transparency, was significantly greater on experimental than control gills. The pereopodal exopods, not previously implicated in gill cleaning in any decapod, were similarly identified as important gill-cleaning structures. Equipped with long multidenticulate setae like those on the epipods, exopods sweep back and forth over the gill filaments just under the gill cover, areas not reached by the epipods. Exopod-ablation experiments were conducted that showed that exopods prevent particulate fouling on gill surfaces over which they sweep. The similarity in action of the passive gill-cleaning system of R. similis to that of crayfish (Bauer [1998] Invert Biol 117:29-143) suggests the hypothesis that the epipodal and exopodal cleaning setae of R. similis are ineffective against epibionts. The reduction in epipodal and exopodal cleaning systems that occurs in the Penaeoidea is hypothesized to be compensated for by increased development of gill-cleaning setae on the branchiostegite, scaphognathite, or other structures.     

The complete mitochondrial genome of the Japanese mud shrimp Upogebia major (Crustacea, Decapoda)

We determined a full-length sequence of mitochondrial (mt) genome from Upogebia major. This is the first complete mt genome report for infraorder Thalassinidea in Decapoda, Crustacea. Our result showed that U. major generally followed a typical pancrustacean gene order but some tRNA genes showed a very unique gene arrangement such as duplication or translocation. Since none of the complete mt genome sequences in the infraorder Thalassinidea are available yet, this report will provide additional information in relation to mt genome diversity and evolution of the decapods.     

Diseases of the eye of farmed shrimp Penaeus monodon

Lesions were found in the eyes of cultured shrimp Penaeus monodon that displayed non-specific signs of disease, including lethargy, dark pigmentation, brown gills, empty midgut, anorexia, white tail muscle, necrosis of uropods and fouled cuticle. Eye lesions were associated with sexual development in moribund shrimp in at least 1 disease event. Suppurative inflammation, granuloma and malacia were observed in histological examination of the eye and the causative agents of lesions appear to be Vibrio spp. and a rod-shaped virus (similar to Lymphoid Organ Virus, Gill-Associated Virus [GAV] and Yellow-Head Virus). Suppurative inflammation was characterised by edema, infiltration of haemocytes and local sites of abscesses. Eyes with granuloma usually appeared white in pond-side examinations, and histology showed that fibrous tissue replaced ommatidia, ganglia and internal structures of the eye. Malacia of the eye was characterised by necrosis of nervous tissue, vacuolation and vascular proliferation in the medulla ganglia. Levels of presumptive Vibrionaceace were high in moribund specimens and Gram-negative rods were observed in some specimens as free particles in the interstitial fluid and haemolymph in the eye. Transmission electron microscopy showed that nerve cells in the fasciculated zone (near the basement membrane) contained cytoplasmic vesicles (1 to 3 microm in diameter) with particles (15 to 26 nm in diameter) and rod-shaped nucleocapsids. The rods were similar to those of GAV and were 130 to 260 nm long, 10 to 16 nm in diameter and had helical symmetry with a screw-like thread (2.4 to 3.5 nm pitch). Also, unidentified enveloped virions, averaging 74 nm in diameter, were observed in cytoplasmic vesicles in the fasciculated zone. In conclusion, it is suggested that bacterial and viral infections of the eye could result in impaired neuroendocrine functions, which may cause a range of clinical signs of disease.