Site-specific integration and excision of pMEA100 in Nocardia mediterranei

Summary Nocardia mediterranei strain LBG A3136 contains the 23.7 kb element pMEA100 in a chromosomally integrated form as well as in the free state (Moretti et al. 1985). The integrated form of this element can be excised precisely from the Nocardia chromosome without any accompanying rearrangements in flanking chromosomal DNA. After transfer into plasmid-free mutant strains, pMEA100 reintegrates site specifically into its original chromosomal locus. The exact mapping of the pMEA100 integration site was accomplished by restriction analysis and DNA sequencing. The attachment site of pMEA100, the junctions of its integrated form and plasmid-free chromosomal DNA of N. mediterranei contain an identical 47 bp long sequence which is probably required for site-specific recombination connected with integration and excision of pMEA100. Only one such sequence was found in the chromosome of pMEA100-free N. mediterranei derivatives as suggested by the single integration locus.     

Seston quality controls zebra mussel (Dreissena polymorpha ) energetics in turbid rivers

Feeding processes and energetic balance of zebra mussels were both related to the quantity and quality of natural seston. Filtration rate and pseudofeces production increased while clearance rate remained constant with increasing seston concentration. Ingestion rate, assimilation efficiency, and assimilation rate all increased with increasing food quality, measured as the ratio of organic to inorganic material in the seston. Respiration rate did not change with either food quantity or quality. As a result, scope for growth declined with decreasing food quality, and fell below 0 cal mg⁻¹ h⁻¹ at an organic:inorganic ratio of 0.5. The association between feeding processes and food quality appears related to a breakdown in the ability of zebra mussels to selectively ingest high-quality organic particles when the organic content of the seston is low. Ingestion, assimilation efficiency, assimilation rate and scope for growth were all higher when seston was amended with an addition of a natural assemblage of algae. Food quality may be a better indicator of environmental conditions suitable for growth than food quantity. These results suggest that the conditions of high suspended inorganic sediment concentrations in large turbid rivers represent a difficult growth environment for the zebra mussel. Received: 12 May 1997 / Accepted: 7 July 1998     

Identification and characterization of glucose transport proteins in plasma membrane- and Golgi vesicle-enriched fractions prepared from lactating rat mammary gland.

Previous studies have indicated that turkey erythrocyte and rat liver membranes contain endogenous alpha beta heterodimeric insulin receptors in addition to the disulphide-linked alpha 2 beta 2 heterotetrameric complexes characteristic of most cell types. We utilized 125I-insulin affinity cross-linking to examine the structural properties of insulin receptors from rat liver and turkey erythrocyte membranes prepared in the absence and presence of sulphydryl alkylating agents. Rat liver membranes prepared in the absence of sulphydryl alkylating agents displayed specific labelling of Mr 400,000 and 200,000 bands, corresponding to the alpha 2 beta 2 heterotetrameric and alpha beta heterodimeric insulin receptor complexes respectively. In contrast, affinity cross-linking of membranes prepared with iodoacetamide (IAN) or N-ethylmaleimide identified predominantly the alpha 2 beta 2 heterotetrameric insulin receptor complex. Similarly, affinity cross-linking and solubilization of intact turkey erythrocytes in the presence of IAN resulted in exclusive labelling of the alpha 2 beta 2 heterotetrameric insulin receptor complex, whereas in the absence of IAN both alpha 2 beta 2 and alpha beta species were observed. Turkey erythrocyte alpha 2 beta 2 heterotetrameric insulin receptors from IAN-protected membranes displayed a 3-4-fold stimulation of beta subunit autophosphorylation and substrate phosphorylation by insulin, equivalent to that observed in intact human placenta insulin receptors. Turkey erythrocyte alpha beta heterodimeric insulin receptors, prepared by defined pH/dithiothreitol treatment of IAN-protected membranes, were also fully competent in insulin-stimulated protein kinase activity compared with alpha beta heterodimeric human placenta receptors. In contrast, endogenous turkey erythrocyte alpha beta heterodimeric insulin receptors displayed basal protein kinase activity which was insulin-insensitive. These data indicate that native turkey erythrocyte and rat liver insulin receptors are structurally and functionally similar to alpha 2 beta 2 heterotetrameric human placenta insulin receptors. The alpha beta heterodimeric insulin receptors previously identified in these tissues most likely resulted from disulphide bond reduction and denaturation of the alpha 2 beta 2 holoreceptor complexes during membrane preparation.     

Abscisic acid and salinity stress induced somaclonal variation and increased histone deacetylase (HDAC) activity in <Emphasis Type="Italic">Ananas comosus</Emphasis> var. MD2

Somaclonal and phenotypic variation caused by genetic and/or epigenetic modifications, are a valuable source of genetic variation to improve desirable polygenetic traits in crops. In this study, we induced somaclonal variation in vitro pineapple (Ananas comosus var. MD2) through hormonal induction, NaCl, and abscisic acid (ABA) supplementation. Our results showed that supplementation of high concentration of 6-benzylaminopurine (4.0 mg/L BAP) alone or combined with indole-butyric acid (IBA) produced the highest percentage of dwarf variants (100%). Murashige and Skoog (MS) media containing 4.0 mg/L BAP plus 2.0 mg/L IBA produced the shortest plantlets (1.9?±?0.1 cm). In comparison, MS media containing 1.0% NaCl induced formation of dwarf plantlets with a mean plantlet height of 1.4?±?0.3 cm, whereas 1.0 mg/L ABA generated plantlets with a mean plantlet height of 1.7?±?0.1 cm. We then analyzed the histone deacetylase (HDAC) enzyme activity for dwarf and non-dwarf plantlets. In general, dwarf plantlets exhibited higher HDAC activity than non-dwarf plantlets. The highest HDAC activity (109, 333.33?±?4.40 ng/min/mg) was recorded for dwarf plantlets grown on media supplemented with 1.0 mg/L ABA. The dwarf variants also underwent phenotypic recovery to normal phenotype within 8 months after transferred to MS basal media. No ploidy alteration was detected in these dwarf plantlets after analyzed by flow cytometry. Taken together, although the generated dwarf plantlets showed higher HDAC activity compared to non-dwarf plantlets, their capability of reverting to non-dwarf phenotype suggested that it might be due to epigenetic modulation.     

Integrating spatial and temporal variability into the analysis of fish food web linkages in Tijuana Estuary

Our understanding of fish feeding interactions at Tijuana Estuary was improved by incorporating estimates of spatial and temporal variability into diet analyses. We examined the stomach contents of 7 dominant species (n=579 total fish) collected between 1994 and 1999. General feeding patterns pooled over time produced a basic food web consisting of 3 major trophic levels: (1) primary consumers (Atherinops affinis, Mugil cephalus) that ingested substantial amounts of plant material and detritus; (2) benthic carnivores (Clevelandia ios, Hypsopsetta guttulata, Gillichthys mirabilis, and Fundulus parvipinnis) that ingested high numbers of calanoid copepods and exotic amphipods (Grandidierella japonica); and (3) piscivores (Paralichthys californicus and Leptocottus armatus) that often preyed on smaller gobiids. Similarity-based groupings of individual species' diets were identified using nonmetric multidimensional scaling to characterize their variability within and between species, and in space and time. This allowed us identify major dietary shifts and recognize events (i.e., modified prey abundance during 1997–98 El Ni no floods) that likely caused these shifts.     

Transience in the humpback whale population of New Caledonia and implications for abundance estimation

A phenomenon of transience in the humpback whale population breeding in New Caledonia has been highlighted in recent analyses. We used these data to illustrate the risk of flawed inference when transience is not properly accounted for in abundance estimation of resident populations. Transients are commonly defined as individuals that pass through the sampling area once, i.e., have a null probability of being caught again, and therefore induce heterogeneity in the detection process. The presence of transients can lead to severe bias in the estimation of abundance and we demonstrate how to correct for this feature when estimating abundance of resident populations. In New Caledonia, very different conclusions about the number of resident whales in the southern lagoon between 1999 and 2005 are obtained when the abundance estimate accounts for the transient whales. Without correction, the estimates of the abundance were up to twice as high across all years compared to the estimates of the resident population when a correction for transients had been incorporated. Having reliable population estimates when assessing the status of endangered species is essential in documenting recovery and monitoring of population trends. Therefore, we encourage researchers to account for transients when reporting abundances of resident populations.     

Interactions between osteosarcoma cell lines and dendritic cells immune function: An in vitro study

Dendritic cells (DCs) might be partly responsible for the defective immune response in tumor bearing hosts, but no study in osteosarcoma patients is still available. Therefore, we investigated in vitro whether human osteosarcoma cell lines have an inhibitor effect on different types of DCs: CD14+DCs, DC1 and DC2. DCs derived from healthy donors were cultured with osteosarcoma cell lines and appropriate cytokine cocktails and analysed for the expression of co-stimulatory molecules (CD40, CD80, CD83, CD86, HLA-DR). Each interaction resulted in a lower phenotypic expression of the DCs maturation markers, especially on DC2. Moreover, the addition of various cytokines and compounds (rhIL-12, CD40L, Indometacin) induced the DC1 and DC2 subsets towards the Th1 pattern as shown by ELISA. Osteosarcoma highly interferes with an in vitro DCs immune function as antigen presenting cells. The understanding of tumor biology underlines the need for a specific osteosarcoma immunotherapy able to reverse this immune-surveillance inhibition.     

Fish community responses to ecosystem stressors in coastal estuarine wetlands: a functional basis for wetlands management and restoration

Functional responses of estuarine fish species to environmental perturbations such as wetland impoundment, changes in water quality, and sediment accretion are investigated. The study focuses on the feeding, growth and habitat use by California killifish (Fundulus parvipinnis), topsmelt (Antherinops affinis), and juvenile California halibut (Paralichthys californicus) in impacted coastal wetlands to provide an ecological basis for guidance on the management and restoration of these ecosystems. The ecology of California killifish, Fundulus parvipinnis, is closely tied with the marsh surface, which they access at high tide to feed and grow. Field estimates of food consumption show that killifish can increase their food intake by two-fold to five-fold by adding marsh surface foods to their diet. Bioenergetics modeling predicts that killifish can grow over an order of magnitude faster if they add intertidal marsh surfaces to their subtidal feeding areas. Tidal inlet closures and increased marsh surface elevations due to sediment accretion can restrict killifish access to the marsh surface, affecting its growth and fitness. An open tidal inlet and tidal creek networks that allow killifish to access the marsh at high tide must be incorporated into the restoration design. Topsmelt and California halibut are also adversely affected by tidal inlet closures. Food consumption rates of topsmelt are 50% lower when the tidal inlet is closed, compared to when the estuary is tidally-flushed. Tidal inlet closures inadvertently induce variations in water temperature and salinity and negatively affect growth of juvenile California halibut. Tidal creek networks which consist of channels and creeks of various orders are also important to halibut. Large halibut (>200 mm TL) inhabit deeper, high order channels for thermal refuge, while small halibut (<120 mm TL) are abundant in lower order channels where they can feed on small-sized prey which are typically less abundant in high order channels. Maintaining an open tidal inlet, implementing sediment management programs and designing coastal wetlands with tidal creek networks adjacent to intertidal salt marsh habitat (for fish access) are key elements that need to be considered during the planning and implementation of coastal wetland restoration projects.