Kyosuke Ikuta

The mitochondrial genome of a filamentous brown alga Pylaiella littoralis (strain CCMP 1907) has been reported to contain four group IIB introns in the LSU rRNA gene and three group IIA introns in the cox1 gene. We found extreme variability in the number of group II introns for these two genes by analyzing eight P. littoralis specimens collected at worldwide habitats. The first intron of the LSU rRNA gene from a specimen collected in France and the fourth intron from a specimen harvested in Japan exhibited an exceptionally long evolutionary distance when compared with the cognate introns found in P. littoralis specimens. Moreover, these introns harbored an intact or nearly intact tripartite ORF, suggesting they are the result of a recent invasion of cognate introns. Based on the fact that many of the target sites were intronless, we propose that opportunity of intron infection is the bottleneck step of the group II intron cycle which consists of invasion, degeneration, and complete loss from the target site.

The adult female of the freshwater ostracod Cyprinotus uenoi Brehm, 1936 (Podocopida: Cypridoidea) has a pair of long, sac-like ovaries separately lying in the posterior part of the left and the right carapace valves. Oogonia and very early previtellogenic oocytes are located in the terminal germarium of each ovary. In the germarium, the oogonia occur in the most terminal region, and the very early previtellogenic oocytes are located in the remainder, arranged in order of size, the larger ones nearer the ovarian lumen. Most of the growing oocytes, previtellogenic and vitellogenic, are found in the ovarian lumen, the larger ones farther from the germarium. In the germarium, a cytoplasmic bridge connects a pair of adjoining germ cells, resulting from an incomplete cytokinesis of oogonial division. Among the previtellogenic and early vitellogenic oocytes in the ovarian lumen, "nurse cells" are found as small, spherical cells in mostly the same number as these oocytes. A cytoplasmic bridge connects each "nurse cell" to an adjoining oocyte. Based on the manner of connection and some morphological features, we consider that each "nurse cell" originates from one of each pair of adjoining germ cells connected by a cytoplasmic bridge in the germarium, as in the true nurse cells of several branchiopod crustaceans and insects with meroistic ovarioles.

Three types of respiratory deficient mitochondrial strains have been reported in Chlamydomonas reinhardtii: a deficiency due to (i) two base substitutions causing an amino acid change in the apocytochrome b (COB) gene (i.e., strain named dum-15), (ii) one base deletion in the COXI gene (dum-19), or (iii) a large deletion extending from the left terminus of the genome to somewhere in the COB gene (dum-1, -14, and -16). We found that these respiratory deficient strains of C. reinhardtii can be divided into two groups: strains that are constantly transformable and those could not be transformed in our experiments. All transformable mitochondrial strains were limited to the type that has a large deletion in the left arm of the genome. For these mitochondria, transformation was successful not only with purified intact mitochondrial genomes but also with DNA-constructs containing the compensating regions. In comparison, mitochondria of all the non-transformable strains have both of their genome termini intact, leading us to speculate that mitochondria lacking their left genome terminus have unstable genomes and might have a higher potential for recombination. Analysis of mitochondrial gene organization in the resulting respiratory active transformants was performed by DNA sequencing and restriction enzyme digestion. Such analysis showed that homologous recombination occurred at various regions between the mitochondrial genome and the artificial DNA-constructs. Further analysis by Southern hybridization showed that the wild-type genome rapidly replaces the respiratory deficient monomer and dimer mitochondrial genomes, while the E. coli vector region of the artificial DNA-construct likely does not remain in the mitochondria.

In adult females of the halocyprid ostracod Conchoecia imbricata, paired saclike ovaries lie separately at the both sides of the alimentary canal in the posterior region of the trunk. The ovarian wall consists of a thin layer of the ovarian epithelium, making itself into many folds. Oogonia, very early previtellogenic oocytes, and young somatic interstitial cells are gathered in a nipple-shaped germarium in each terminal of the ovaries. In the germarium, oogonia are concentrated in the most terminal region, and very early previtellogenic oocytes; are located in the remainder, arranged in order of their sizes, the larger ones nearer to the ovarian lumen. These oocytes grow and leave the germarium, not to enter the ovarian lumen, but to ride on the outer surface of the ovarian wall, sandwiched between the ovarian epithelium and its basement membrane. They grow further to mature, supported by a few tall ovarian epithelial cells and raising the basement membrane of the ovarian epithelium. The larger oocytes lie farther from the germarium on the ovarian wall. Both the basic ovarian structure and the arrangement of the early female germ cells in the germarium of C. imbricata are common to those of the podocopid ostracods, many other crustaceans, and other mandibulate arthropods, but the further growth of the oocytes on the outer surface of the ovarian wall is similar to the pattern seen in the myodocopid ostracods, the branchiurans, the pentastomids, and the chelicerate arthropods. Such an intermediate oogenetic mode in C. imbricata is unique, not only among the ostracods and other crustaceans, but also among other mandibulates and the chelicerates.

Multifunctions of the upper lip in a bioluminescent myodocopid Vargula hilgendorfii were studied by video observation and histological method. The localization of luciferin and luciferase gland cells within the upper lip was partly successful. Two long protrusions of the upper lip, both of V. hilgendorfii and a non-luminescent species of the same family, immediately anterior to the mouth, were found to show very flexible movement especially while eating, as if smearing on the food surface a secretion from the protrusions (glands), which may support the hypothesized secretion of digestive enzymes from the upper lip. This hypothesis is further supported by the new finding of a pair of ducts which connect the basal part of the upper lip with the posterior digestive duct (stomach). Comparative studies of V. hilgendorfii with several sympatric non-luminescent species of the same family have also revealed that it has a characteristic reflecting organ immediately posterior to the anus. It is a conical small protrusion, as if dangling from the ventral edge of the abdomen at the apex of the cone. It is observable only in live specimens, when the furca, which is located outwardly to the organ, is sufficiently transparent. When illuminated, the reflecting organ reflects the distinct light. The diameter of the mirror (chemical composition provisionally analyzed) is about 6-8% of the carapace length. The organ develops from the very first stage of its ontogeny without reference to sex, which suggests that the function may be related to intraspecific signaling or predatory deterrence.

A pair of laterally flattened saclike ovaries separately lie on both sides of the alimentary canal in the posterior region of the trunk of the myodocopid ostracod Vargula hilgendorfii. Neither the oogonia nor the germarium as a young germ-cell cluster are found in the adult ovary. Several tens of synchronously growing oocytes, previtellogenic or vitellogenic, are found in the ovary, not in the ovarian lumen but on the outer surface of the ovarian wall facing the alimentary canal. Each of these oocytes is protruded outward from the ovarian wall, supported by a short stalk, a short protrusion of the ovarian epithelium composing the ovarian wall. These stalked oocytes are tightly covered by the extremely extended basement membrane of the ovarian epithelium. In the ovary of the fifth instar juveniles of V. hilgendorfii, the oogonia and very early previtellogenic oocytes are scattered among the ovarian epithelial cells of the ovarian wall facing the alimentary canal. Several tens of larger stalked previtellogenic oocytes of mostly similar size are protruded toward the alimentary canal. The oogenetic mode of V. hilgendorfii, in particular the growth of the oocytes on the outer surface of the ovarian wall, seems similar to those in branchiurans, pentastomids, and chelicerate arthropods, rather than to those in many other crustaceans. However. the ovary of V. hilgendorfii has a basic paired saclike structure, common to that in many other crustaceans, but different from unpaired tubular ovaries in chelicerates.

High-resolution videos, scanning electron microscopy and histology were used to study the feeding mechanism of myodocopid ostracods from the Pacific Coast of Japan, as exemplified by Vargula hilgendorfii (Muller, 1890) and a few other cypridinid species. Ostracods observed in the laboratory were attracted to a wide spectrum of natural food sources, behaving as predators of living prey (e.g. polychaete annelids), as opportunistic scavengers on dead animals (e.g. annelids, fishes, squid), and also consuming artificial food. Food sources may be detected by chemoreception. The fourth limb (endopodites with strong sclerotized setae) and the furcal lamellae (claws with teeth) act in coordination to abrade and eventually tear open the protective integument of living/dead prey such as annelids. The mandibular palps are used mainly to hold the food. Food sections and soft-body contents are transferred to the mouth by the fourth limb (endopodial "rake") and fifth limb (exopodite with pectinate setae) and are passed to the oesophagus by the endites (mandibles, fourth and fifth limbs). Food is subsequently pumped up to the stomach by peristaltic contractions of the oesophagus (ring muscles) and stored in the stomach pouch. The upper lip of bioluminescent (V. hilgendorfii) and non-bioluminescent species of Cypridinidae often contact food, suggesting that some of the glands housed in this organ may emit digestive enzymes prior to ingestion. Ostracods are able to ingest massive quantities of food within a few minutes and to survive starvation for several weeks. In V. hilgendorfii, the midgut is a huge sac-like organ with no partition and is lined with a single layer of columnar epithelial cells. No differentiated hepatopancreas is present. The cypridinid produces a single faecal pellet wrapped in a thin reticulated, peritrophic membrane. Myodocopid ostracods exhibit a wide range of feeding strategies (detritus-feeding, comb-feeding, scavenging, predation, ectoparasitism) in both benthic and pelagic niches, and constitute a substantial source of food for many zooplankters. Adaptation of cypridinids to scavenging/predation is reflected in the morphology of their furcae, mandibles, fourth and fifth limbs, and their digestive system. Palaeontological data suggest that early Triassic cypridinids and possible late Ordovician myodocopids may have been carnivorous scavengers feeding on carcasses of cephalopods (ammonoids or orthoconic nautiloids), thus playing the same role of "recyclers" as modern representatives of the group.

Eggshell ultrastructure is described in a branchiuran, Argulus japonicus. Growing oocytes of various sizes are found on the outer surface of the ovary. They are protruded outward, covered by a thin basal lamina of the ovarian epithelium which connects them on the ovarian surface. Many microvilli develop on the surface of oocytes from the late previtellogenic stage. These microvilli elongate to raise the basal lamina from the oocyte surfaces. In the early vitellogenic oocytes, large and small electron-dense granules appear and increase in number in the outer and inner regions, respectively, of the perivitelline space between the basal lamina and the oocyte surfaces. In the late vitellogenic oocytes, other large electron-lucent granules appear and increase in number in the innermost region of the perivitelline space. Mature eggs in the oviduct have a two-layered eggshell: the outer layer composed of large electron-dense granules and the inner layer of small electron-dense granules and large electron-lucent granules. Some unique features on the origin and formation of the eggshell in A. japonicus are discussed.

The adult ovary of the branchiuran Argulus japonicus is a single, median, long sac-like organ located in the thorax above the alimentary canal. A long germarium, including oogonia, very early previtellogenic oocytes, and young somatic cells (interstitial cells), is embedded in the dorsal ovarian wall along the median longitudinal line of the ovary. The ovarian wall, consisting of a layer of the ovarian epithelial cells, is folded repeatedly and distinctively in the lateral and ventral portions of the ovary. Growing oocytes, previtellogenic and vitellogenic, occur on the outer surface of the ovarian wall, not in the ovarian lumen. The smaller oocytes are located nearer to the germarium and the larger ones on the more ventral surface of the ovary. These structural features of the branchiuran ovary are compared with those of other crustaceans and the pentastomids to consider their phylogenetic implications. (C) 1997 Wiley-Liss, Inc.

The gills of the ostracode Leuroleberis surugaensis Hiruta, 1982, a nektobenthic cylindroleberidid myodocopid from the Pacific Coast of central Japan are described. They consist of 7 pairs of integumental laminae attached to the thoracic wall. Each individual lamina has numerous lacunae (diameter up to 60 mu m), smaller subcuticular spaces, an (efferent) epibranchial and a (afferent) hypobranchial canal, pillarlike cells, and cells bearing a large nucleus (5-10 mu m in diameter), comparable to the nephrocytes of decapods. The lacunae form an extensive anastomosing network of arcuate sinuses containing hemolymph and amoeboid free-cells (hemocytes). The epibranchial canals open dorsally into the pericardial cavity suggesting that hemolymph contained in gills Rows back to the heart through a dorsal route. The 7 pairs of gill laminae of Leuroleberis are interpreted as the major respiratory surfaces of the animal and may also be involved in the degradation of metabolic waste products. Ventilation over the respiratory surfaces is provided by the rhythmic movement of epipodial plates (fifth limb). The book gills of Leuroleberis and other cylindroleberidids are interpreted as possible epipodial remnants of lost limbs posterior to the seventh limb.