Life history strategy and adult and larval behavior of Macrodiplosis selenis (Diptera: Cecidomyiidae), a species that induces leaf‐margin fold galls on deciduous Quercus (Fagaceae)

Life historical, behavioral and ecological traits of Macrodiplosis selenis, which induces leaf‐margin fold galls on Quercus serrata, Q. mongolica and Q. dentata (Fagaceae) in Japan and South Korea, were studied. Daily activity and larval development indicate that M. selenis is a diurnal and univoltine gall midge. In April, females lay their eggs both on upper and under surfaces of fresh leaves. The duration of the egg stage varies from 5 to 9 days, depending on daily temperatures. Hatched larvae crawl to the upper surface of the leaf margin, where they start to induce galls. Larvae become full‐grown in October, drop to the ground in November and overwinter in cocoons on the ground, while larvae of congeners mature in May and drop to the ground in June. A relatively long period of the second larval stadium from July to October on the host trees seems to be effective for M. selenis in avoiding summer mortalities caused by predation and aridity on the ground and by ectoparasitoids that attack mature larvae or pupae on the host leaves. The spatial distribution pattern of M. selenis leaf galls is contagious and the mean gall density per leaf is significantly correlated with the mean crowding. This study adds new insights of life history strategy and adult and larval behavioral pattern to the ecological knowledge of gall midges, and these kinds of information are essential for further studies of M. selenis population dynamics and interactions with other Quercus‐associated herbivores.     

Nucleotide-induced conformational changes of PMP70, an ATP binding cassette transporter on rat liver peroxisomal membranes

Nucleotide-induced conformational changes of the 70-kDa peroxisomal membrane protein (PMP70) were investigated by means of limited-trypsin digestion. Rat liver peroxisomes preincubated with various nucleotides were subsequently digested by trypsin. The digestion products were subjected to immunoblot analysis with an anti-PMP70 antibody that recognizes the carboxyl-terminal 15 amino acids of the protein. PMP70 was initially cleaved in the boundary region between the transmembrane and nucleotide-binding domains and a carboxyl-terminal 30-kDa fragment resulted. The fragment in turn was progressively digested at the helical domain between the Walker A and B motifs. The fragment, however, could be stabilized with MgATP or MgADP. In contrast to MgATP, MgATP-gammaS protected whole PMP70 as well as the fragment. The 30-kDa fragment processed by trypsin was recovered in the post-peroxisomal fraction as a complex with a molecular mass of about 60 kDa irrespective of the presence of MgATP. These results suggest that PMP70 exists as a dimer on the peroxisomal membranes and the binding and hydrolysis of ATP induce conformational changes in PMP70 close to the boundary between the transmembrane and nucleotide binding domains and the helical domain between the Walker A and B motifs.     

Phagocytosis of different matrix components by different cell types at bone-forming sites in cultured mouse calvariae

Summary Sites of bone formation on fragments of parietal bone of fetal-mice cultured for 10 days were examined by electron microscopy after addition of either ruthenium red or ferrocyanide to the postfixation fluid. Osteoclasts, osteoblast-like cells, and macrophages were the principal active cells at these formation sites. The mononuclear cells (osteoblast-like cells and macrophages) in the osteoid tissue showed evidence of having incorporated elements of calcified tissue. Osteoblast-like cells had phagocytized collagen fibrils and calcified bone matrix. This occurred more frequently in the calcifying area. Mononuclear macrophages showed not only phagocytosis and digestion of cellular debris and bone spicules in the osteoid, but also active incorporation of calcified bone matrix that had been detached from its surroundings by its pseudopod-like projections from long cytoplasmic processes. Collagen fibrils were seldom observed within the macrophages. These observations suggest that in our culture system osteoblast-like cells and macrophages at bone formation sites have a phagocytic role in bone remodeling.This study was supported in part by a grant from the Ministry of Education. Science and Culture of Japan (No. 59771321)     

Structural Uniformity of Pullulan Produced by Several Strains of Pullularia pullulans

Extracellular polysaccharides produced by 3 strains of Pullularia pullulans were fractionated by treating with cetyl trimethyl ammonium hydroxide into soluble and insoluble fractions, and the structure of the former fraction, i.e., pullulan, was studied. The yield and the ratio of 2 fractions varied widely according to the strains. But the structure of pullulan was found to be uniform irrespective of the strains used. All 3 samples of pullulan gave only glucose on complete acid hydrolysis, and 93~95% maltotriose and 5~7% maltotetraose after isoamylase (pullulanase) action. The ratio of α-1,4- to α-1,6-glucosidic linkages calculated from periodate oxidation data coincided very well with the value expected from the ratio of maltotriose to maltotetraose units. An evidence for the complete absence of branch structure in pullulan was presented from the results of hydrolysis by pullulan 4-glucanohydrolase.     

Possible factors responsible for the toxicity of Cochlodinium polykrikoides,a red tide phytoplankton

Cochlodinium polykrikoides, a harmful red tide dinoflagellate, is highly toxic to fish, but the toxic mechanism is still unknown. Recent study has suggested that C. polykrikoides generates reactive oxygen species (ROS) such as superoxide anion (O(2)(-)) and hydrogen peroxide (H(2)O(2)), and the ROS-mediated ichthyotoxicity has been proposed. In this study, we found that the levels of O(2)(-) and H(2)O(2) detected in C. polykrikoides were trace levels as compared with those of Chattonella marina which is well-known to produce ROS. Furthermore, no significant increase in O(2)(-) generation by C. polykrikoides was observed in the presence of lectins such as concanavalin A (Con A) and wheat germ agglutinin (WGA) or fish mucus prepared from skin and gill of yellowtail, whereas C. marina generated increased level of O(2)(-) responding to these stimuli. Interestingly, the cell-free aqueous extract prepared from C. polykrikoides showed toxic effect on the HeLa cells, but the extract of C. marina had no significant effect. Furthermore, gradual accumulation of polysaccharides in the medium was observed during the growth of C. polykrikoides, and the medium gradually became viscous, but no such changes were observed in the medium of C. marina. These results suggest that multiple factors may be responsible for the toxic mechanism of C. polykrikoides.     

Molecular characteristics and transcription of the gene encoding a multifunctional alcohol dehydrogenase in relation to the deactivation of pyruvate formate-lyase in the ruminal bacterium<Emphasis Type="Italic"> Streptococcus bovis</Emphasis>

To clarify the deactivation mechanism of pyruvate formate-lyase (PFL) and its role in the regulation of fermentation in Streptococcus bovis, the molecular properties and genetic expression of multifunctional alcohol dehydrogenase (ADHE) were investigated. S. bovis was found to have ADHE, which was deduced to consist of 872 amino acids with a molecular mass of 97.4 kDa. The ADHE was shown to harbor three enzyme activities: (1) alcohol dehydrogenase, (2) coenzyme-A-linked acetaldehyde dehydrogenase that catalyzes the conversion of acetyl-CoA to ethanol, and (3) PFL deactivase. Similar to Escherichia coli ADHE, S. bovis ADHE required Fe2+ for its activity. The gene encoding ADHE ( adhE) was shown to be monocistronic. The level of adhE mRNA changed in parallel with the mRNA levels of the genes encoding PFL (pfl) and PFL-activating enzyme (act) as the growth conditions changed, although these genes are independently transcribed. Synthesis of ADHE, PFL-activating enzyme, and PFL appears to be regulated concomitantly. Overexpression of ADHE did not cause a change in the formate-to-lactate ratio. It is conceivable that ADHE is not significantly involved in the reversible inactivation of active PFL under anoxic conditions. Partition of the flow from pyruvate appears to be mainly regulated by the activities of lactate dehydrogenase and PFL.     

Differential staining of insect neurons with nickel and cobalt

Pairs of neurons were differentially stained by intracellular injection of the divalent cations, nickel and cobalt. Pairs of neuronal tracts could be similarly labelled following immersion of their axon bundles in the markers described. Use of nickel and a 4 : 6 mixture of cobalt and nickel according to the protocols presented here revealed the morphology of cricket neuron processes as thin as 1 μm in diameter and yielded a favourable contrast between the cells marked, as well as with respect to the background of unstained tissue.     

Genetic variation of the cytochrome b gene in the rosy bitterling, Rhodeus ocellatus (Cyprinidae) in Japan

According to conventional views, the rosy bitterling, Rhodeus ocellatus, comprises two subspecies, R. ocellatus kurumeus and R. ocellatus ocellatus, the former being native to Japan whereas the latter was introduced into Japan from China during World War II. To examine the genetic structure of Japanese R. ocellatus, part of the mitochondrial cytochrome b gene from 48 individuals collected from various locations in Japan was sequenced. Three major mitochondrial lineages were found. Based on historical evidence, two of these represent R. ocellatus ocellatus and the third R. ocellatus kurumeus. The existence of two distinct lineages of R. ocellatus ocellatus in Japan suggests at least two colonizations. Some local populations comprised purely R. ocellatus kurumeus, but those from Kashima and Ogori included both subspecies. Because the proportion of R. ocellatus ocellatus in Kashima increased from 1994 to 1995, invasion by R. ocellatus ocellatus into R. ocellatus kurumeus habitats is apparently in progress. Received: April 30, 1999 / Revised: March 22, 2000 / Accepted: December 20, 2000     

Effects of temperature, salinity and irradiance on the growth of the red tide dinoflagellate Gyrodinium instriatum Freudenthal et Lee

The effects of temperature, salinity and irradiance on the growth of the red tide dinoflagellate Gyrodinium instriatum Freudenthal et Lee were examined in the laboratory. Exposed to 45 different combinations of temperature (10–30 °C) and salinity (0–40) under saturating irradiance, G. instriatum exhibited its maximum growth rate of 0.7 divisions/day at a combination of 25 °C and a salinity of 30. Optimum growth rates (>0.5 divisions/day) were observed at temperatures ranging from 20 to 30 °C and at salinities from 10 to 35. The organism could not grow at ≤10 °C. In addition, G. instriatum burst at a salinity of 0 at all temperatures, but grew at a salinity of 5 at temperatures between 20 and 25 °C. It is noteworthy that G. instriatum is a euryhaline organism that can live under extremely low salinity. Factorial analysis revealed that the contributions of temperature and salinity to its growth of the organism were almost equal. The irradiance at the light compensation point (I₀) was 10.6 μmol/(m² s) and the saturated irradiance for growth (I_s) was 70 μmol/(m² s), which was lower than I_s for several other harmful dinoflagellates (90–110 μmol/(m² s)).