Physical aspects of artificial diets

The physical properties of artificial diets, texture, hardness, homogeneity, and water content are regulated by the addition of cellulose, agar, polysaccharide gums, and other large molecules. These physical aspects are important in promoting good growth and development of insects.The development of the boll weevil, Anthonomus grandis Boheman, was improved by adding more cellulose to the diet. Additional agar did not improve growth. Polysaccharide gums made the diet too viscous and were poor substitutes for agar. This insect developed satisfactorily on diets with moderate ranges of water and nutrient content.The preparation of the diets is also important in regulating physical properties. Heating stops enzyme action in plant products, ruptures cells, and affects solubility of ingredients. The addition of stabilizers protects nutrients and keeps them mixed homogeneously.

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Zusammenfassung Die physikalischen Eigenschaften künstlicher Diäten für Insekten umfassen Struktur, Härte, Homogenität und Wassergehalt sowie Faktoren, die diese beeinflussen. Diese Gründsätze gelten für feste Diäten bei Insekten mit beißenden Mundwerkzeugen.Strukturelle Eigenschaften werden gewöhnlich durch Hinzufügen von Zellulose hervorgerufen, welche die Konsistenz liefert und zum Fraß anregt. In Diäten mit hohem Wassergehalt wird die Konsistenz durch Zugabe von Agar reguliert. Natürliche polysaccharide Gummis verleihen den Diäten Andickung, Stabilisierung, Gelierung und strukturelle Eigenschaften. Diäten für den Baumwollkapselbohrer, Anthonomus grandis Boheman, werden abgewandelt durch Hinzufügen zusätzlicher Zellulosemengen, Agar und Stärke sowie ersatzweise durch Carrageenan, Heuschrecken- Bohnen- Gummi und Guar-Gummi für die Gesamtheit oder Teile des Agars. In den beschriebenen Versuchen verkürzte Zellulose die Entwicklungszeit. Agar und Stärke verursachten wenig Veränderung. Im allgemeinen bildeten die Gummis visköse Gemische und gelierten nicht. Daher ergab sich eine schlechte Entwicklung für den Baumwollkapselkäfer. Carrageenan konnte als teilweiser Ersatz für Agar verwendet werden. Große Moleküle, wie Zellulose, Stärke, Pektin, Phospholipide und Proteine, die in natürlichen Produkten vorkommen, beeinflussen die physikalischen Eigenschaften der Diäten ebenfalls. Diese Eigenschaft der Naturstoffe ist oft übersehen worden, weil die Hauptbedeutung immer in ihrem Nährwert gesehen wurde.Obwohl die Nahrung pflanzenfressender Insekten einen hohen Wassergehalt hat, können viele mit Nahrung sehr unterschiedlicher Wasserkonzentrationen auskommen und einige vermögen trockene Nahrung zu verzehren und Wasser zu trinken. Die Herstellung der Diäten kann ihre physikalischen Eigenschaften ebenfalls beeinflussen. Die Bestandteile der Diät können chemische und enzymatische Veränderungen erfahren, welche die Diät und damit das Insekt beeinflussen. Hitze-Behandlung stoppt die enzymatischen Reaktionen in den Geweben, zerreißt die Zellen und löst ihre Inhaltsstoffe. Polysaccharide Gummis, Phospholipide und gelierende Mittel werden benutzt, um die Bestandteile der Diät in einem beständigen, homogenen Gemisch zu erhalten. Diese Substanzen können auch in flüssigen Diäten verwendet werden, um die Nährstoffe in Suspension zu halten.

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in cooperation with Texas Agricultural Experiment Station Texas A & M University, U.S.A.     

Effects of electric field strengths on fusion and in vitro development of domestic cat embryos derived by somatic cell nuclear transfer

The present study was conducted to determine the effect of electric field strength on the rate of membrane fusion between the somatic cell and cytoplast and on subsequent in vitro development of reconstructed embryos. Additionally, the in vitro developmental competence of cat oocytes artificially activated after 44 h of maturation culture was examined. An efficient fusion rate (64.2%) was obtained by applying a single pulse of 1.5 kV/cm for 50 micros, and the fusion rate remained almost constant at the higher field intensity (59.8 and 54.9% at 1.7 and 2.0 kV/cm, respectively). Although the cleavage rate of fused embryos increased with an increase of the electric field strength, there were no differences among the groups with respect to the proportion of development to the morula and blastocyst stages. In the additional experiment, oocytes at the metaphase II stage after culture for 44 h were activated by the combination of calcium ionophore (CaI) with cycloheximide (CHX). Some (11.8%) of activated oocytes developed to the blastocyst stage. Results from this study indicated that electric field strength affects the rates of fusion and cleavage but has no significant effects on the development to the blastocyst stage of reconstructed embryos. Prolonged maturation culture of cat oocytes (up to 44 h) decreased their ability to develop to the blastocyst stage.     

Sentulic acid: a cytotoxic ring A-seco triterpenoid from Sandoricum koetjape Merr

A new ring A-seco triterpenoid, sentulic acid, along with a known oleanane-type triterpenoid, 3-oxoolean-12-en-27-oic acid, were isolated from the Indonesian plant Sandoricum koetjape Merr. Their chemical structures were elucidated by spectroscopic analysis. In addition, the cytotoxic effects of these compounds on human promyelocytic leukemia HL-60 cells were studied. The results of 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assays and trypan blue dye exclusion tests showed that sentulic acid and 3-oxoolean-12-en-27-oic acid were able to induce cytotoxicity in these cells. Furthermore, morphological examination and DNA fragmentation analysis indicated that these cytotoxic effects were mediated by apoptosis.     

Decreased expression of Apaf‐1 with progression of melanoma

Defects in apoptotic system may contribute in the pathogenesis and resistance of malignant melanoma cells to chemotherapy. Apoptotic protease‐activating factor‐1 (Apaf‐1) is a cell death effector that acts with cytochrome c and caspase‐9 to mediate apoptosis. Recently it was shown that metastatic melanomas often lose Apaf‐1 and are concomitantly resistant to apoptosis. It is not known, however, whether Apaf‐1 protein is lost during melanoma progression from localized to metastatic tumor. To this end, we evaluated Apaf‐1 protein expression by immunohistochemistry in 10 cases of human nevi, 11 melanomas in situ, 26 primary melanomas and 15 metastases. Significant decreases in Apaf‐1 expression was observed when comparing nevi and melanomas (chi‐square = 33.719; P < 0.0001). Moreover, primary melanomas with greater tumor thickness showed lesser expression of Apaf‐1 (chi‐square = 16.182; P < 0.003). Intriguingly, we were unable to detect Apaf‐1 expression in lesions of metastatic melanomas. These data demonstrated that there is an inverse correlation between Apaf‐1 expression and pathologic stage of melanoma. This suggests that the decreased expression of Apaf‐1 seen in correlation with melanoma progression renders melanoma more resistant to chemotherapy.     

Effects of oxygen tension on the development and quality of porcine in vitro fertilized embryos

The present study was conducted to examine the effect of oxygen tension during in vitro culture (IVC) of porcine oocytes/embryos on their development and quality using two different culture systems. Porcine cumulus oocyte complexes (COCs) were matured (IVM) and fertilized (IVF) in vitro, and subsequently cultured for 6 days in a simple and economical portable incubator or a standard CO(2) incubator. While the same temperature (38.5 degrees C) and CO(2) concentration (5%) were used in the both systems, the portable incubator was operated in a negative air pressure (- 300 mmHg) to create an O(2) level at 8-10% (low O(2) concentration), or in a positive air pressure (high O(2) concentration). To compare the two culture systems, IVM and IVF of COCs and subsequent IVC of in vitro produced (IVP) embryos were carried out in the portable incubator with a low O(2) concentration (Group I) or in the standard incubator with a high O(2) concentration (Group II). To assess the effect of O(2) concentration on IVC of IVP embryos, some oocytes that had been cultured in the standard incubator for IVM and IVF were subsequently cultured in the portable incubator with a low O(2) concentration (Group III) or a high O(2) concentration (Group IV). The occurrence of DNA fragmentation in the blastocysts produced under different culture conditions was examined by TUNEL staining to assess embryo quality. The rates of oocytes that reached MII and were penetrated by spermatozoa following IVF did not differ between the two incubation systems. In contrast, the proportions of development to blastocysts and the mean cell number of blastocysts in Group I were higher than those in Group II and Group IV. The index of DNA-fragmented nucleus in the blastocysts of Group I was significantly lower than that in the blastocysts of Group II. Therefore, low oxygen tension during IVM, IVF and IVC enhanced the subsequent development of IVP embryos to the blastocyst stage and improved their quality.     

Development of interspecies cloned embryos in yak and dog

Interspecies nuclear transfer (NT) could be an alternative to replicate animals when supply of recipient oocytes is limited or in vitro embryo production systems are incomplete. In the present study, embryonic development was assessed following interspecies NT of donor cumulus cells derived from yak and dog into the recipient ooplasm of domestic cow. The percentages of fusion and subsequent embryo development to the eight-cell stage of interspecies NT embryos were comparable to those of intraspecies NT embryos (cow-cow NT embryos). The percentage of development to blastocysts was significantly lower (p < 0.05) in yak-cow NT embryos than that in cow-cow NT embryos (10.9% vs. 39.8%). In dog-cow NT embryos, only one embryo (0.4%) developed to the blastocyst stage. These results indicate that interspecies NT embryos possess equally developmental competence to the eight-cell stage as intraspecies NT embryos, but the development to blastocysts is very low when dog somatic cells are used as the donor nuclei.     

Tumor biomarker glycoproteins in the seminal plasma of healthy human males are endogenous ligands for DC-SIGN

DC-SIGN is an immune C-type lectin that is expressed on both immature and mature dendritic cells associated with peripheral and lymphoid tissues in humans. It is a pattern recognition receptor that binds to several pathogens including HIV-1, Ebola virus, Mycobacterium tuberculosis, Candida albicans, Helicobacter pylori, and Schistosoma mansoni. Evidence is now mounting that DC-SIGN also recognizes endogenous glycoproteins, and that such interactions play a major role in maintaining immune homeostasis in humans and mice. Autoantigens (neoantigens) are produced for the first time in the human testes and other organs of the male urogenital tract under androgenic stimulus during puberty. Such antigens trigger autoimmune orchitis if the immune response is not tightly regulated within this system. Endogenous ligands for DC-SIGN could play a role in modulating such responses. Human seminal plasma glycoproteins express a high level of terminal Lewis(x) and Lewis(y) carbohydrate antigens. These epitopes react specifically with the lectin domains of DC-SIGN. However, because the expression of these sequences is necessary but not sufficient for interaction with DC-SIGN, this study was undertaken to determine if any seminal plasma glycoproteins are also endogenous ligands for DC-SIGN. Glycoproteins bearing terminal Lewis(x) and Lewis(y) sequences were initially isolated by lectin affinity chromatography. Protein sequencing established that three tumor biomarker glycoproteins (clusterin, galectin-3 binding glycoprotein, prostatic acid phosphatase) and protein C inhibitor were purified by using this affinity method. The binding of DC-SIGN to these seminal plasma glycoproteins was demonstrated in both Western blot and immunoprecipitation studies. These findings have confirmed that human seminal plasma contains endogenous glycoprotein ligands for DC-SIGN that could play a role in maintaining immune homeostasis both in the male urogenital tract and the vagina after coitus.     

Effects of beta-mercaptoethanol and cycloheximide on survival and DNA damage of bovine embryos stored at 4 degrees C for 72 h

The objectives of this study were to determine the effects of cycloheximide (CHX) and beta-mercaptoethanol (beta-ME) during storage of in vitro-produced (IVP) bovine blastocysts for 72 h at 4 degrees C on their survival, hatching capacity and DNA damage. In Experiment 1, when blastocysts were stored in a medium supplemented with 25, 50 or 100 microg/mL of CHX, or 25, 50 or 100 microM of beta-ME, the blastocysts stored with 25 microg/mL of CHX had a significantly higher survival rate than that of the blastocysts stored without CHX (79.5% versus 54.2%). In contrast, beta-ME had no apparent effects on the survival and hatching capacity of stored embryos. In Experiment 2, to investigate synergistic effects of CHX and beta-ME during storage of blastocysts on their developmental parameters and DNA damage, they were stored in the medium with CHX (25 microg/mL) and beta-ME (50 microM). The combination of CHX and beta-ME had no significant effects on the survival of blastocysts. The proportion (6.8%) of DNA-fragmented cells in the blastocysts stored with CHX was similar to that (5.4%) in the non-stored blastocysts (positive control) and significantly lower than that (9.7%) in the blastocysts stored without CHX and beta-ME (negative control). However, there were no significant differences among the proportions of dead cells of blastocysts in the storage groups. Therefore, the supplementation of CHX in the storage medium had a beneficial effect on the proportions of survival and DNA-fragmented cells in the stored embryos, whereas the beta-ME alone or in combination with CHX had no positive effects on either of these proportions.     

Population dynamics of the whitefly,Bemisia tabaci (Gennadius) (Homoptera: Aleyrodidae), in Java,Indonesia, with special reference to spatio-temporal changes in the quantity of food resources

Six monitoring plots were established in the northern part of West Java, Indonesia, to clarify the factors that influence population fluctuations of the whitefly,Bemisia tabaci. Yellow sticky traps were used to monitor adult populations. To examine the relationship between the population fluctuations and the quantity of food resources, the quantity of food resources ofB. tabaci (i.e. soybean and mung bean) was also investigated routinely in each subdistrict where the experimental plot was located. The maximum number of adults per trap in each sowing season showed a similar noticeable seasonal trend in all experimental plots. Climatic factors did not have a major role to play in population fluctuations. The number of adults per trap in each experimental plot tended to increase when the quantity of food resources in each subdistrict increased. The changes of the quantity of food resources seemed to influence the population fluctuations. It is suggested that the operation of regulatory processes in population density is influenced largely by both the distance between habitat patches and the amplitude of temporal fluctuations of the quantity of food resources. An explanation for why the time lag between fluctuations of the quantity of food resources and the number of individuals ofB. tabaci occurred is discussed. Based on their flight ability and intrinsic rate of natural increase,B. tabaci is considered not to be a serious pest in the environment where the host plants are grown discontinuously in time and space.     

Regulation of Starch Biosynthesis in Plant Leaves: Activation and Inhibition of ADPglucose Pyrophosphorylase

The ADPglucose pyrophosphorylases of 7 plant-leaf tissues were partially purified and characterized. In all cases the enzymes showed stability to heat treatment at 65 degrees for 5 minutes in the presence of 0.02 m phosphate buffer, pH 7.0. The leaf ADPglucose pyrophosphorylases were activated 5 to 15-fold by 3-phosphoglycerate. Fructose-6-phosphate and fructose 1, 6-diphosphate stimulated ADPglucose pyrophosphorylase to lesser extents. The A(0.5) (conc of activator required to give 50% of the observed maximal activation) of 3-phosphoglycerate for the barley enzyme was 7 x 10(-6)m while for the sorghum enzyme it was 3.7 x 10(-4)m. Inorganic phosphate proved to be an effective inhibitor of ADPglucose synthesis. The I(0.5) (conc of inhibitor that gave 50% inhibition of activity for the various leaf enzymes varied from 2 x 10(-5)m (barley) to 1.9 x 10(-4)m (sorghum). This inhibition was reversed or antagonized by the activator 3-phosphoglycerate. These results form the basis for an hypothesis of the regulation of leaf starch biosynthesis.