A new staining technique for dual identification of plankton and detritus in seawater

A new method is described that employs dual staining and epifluorescencemicroscopy for the identification of plankton and detritus inseawater. The staining technique uses the fluorochromes propidiumiodide (PI) and 4'.6-diamidino-2-phenylindole (DAPI) in conjunctionwith UV excitation. PI/DAPI staining identifies living matterdistinct from non-living matter by selectively staining detritusa deep orange color while cells are either blue-white or pink,depending upon stain and cell type. Comparison of this techniquewith existing epifluorescence techniques and staining reagentsindicates that PI/DAPI offers an improved approach for Stainingdetritus and plankton when evaluation of both is desired. Thisstep is a prerequisite for quantitation of the volumes of detritusand plankton, preliminary to independent estimates of theircarbon and nitrogen pools in aquatic ecosystems.     

On assessment of prey ingestion by copepods

The consumption of photosynthetic and heterotrophic cells byan abundant calanoid copepod species feeding on natural planktoncommunities was quantified with a state-of-the-art image-analysissystem. Late copepodid stages of Eucalanus pileatus did notingest bactena, small photosynthetic and heterotrophic nanoplankton,or the abundant Ceratium spp. in quantifiable amounts Althoughdiatoms were by far the most abundant cells (in terms of POC1^–1), the copepods ingested a higher percentage of ciliatesin relation to their abundance than of diatoms and small heterotrophicdinoflagellates in the first experiment, and ingested a higherpercentage of dinoflagel lates and ciliates compared to diatomsin the second experiment Heterotrophic cells sufficiently largeto be captured were repeatedly preferred by E.pileatus overautotrophs of similar or larger size. More over, among the cellswhich could be individually perceived by this calanoid, largerones were not pre ferred over smaller cells, implying that someaspect of food quality can be as significant as prey size. Theseresults support the notion (e.g. Kleppel, Mar. Ecol Prog. Ser.,99, s183–195, 1993) that feeding by copepods will be underestimatedif ingestion of heterotrophic food organisms is not quantified.While the proposed microscope-based method is comparativelyslow (     

Evaluation of the effects of various chemicals on discharge of and pain caused by jellyfish nematocysts

Jellyfish tentacles in contact with human skin can produce pain swelling and redness. The pain is due to discharge of jellyfish nematocysts and associated toxins and discharge can be caused by a variety of mechanical and chemical stimuli. A series of tests were carried out with chemicals traditionally used to treat jellyfish stings e.g. acetic acid ammonia meat tenderizer baking soda and urea to determine if these chemicals stimulated or inhibited nematocyst discharge and if they brought relief to testers who were exposed to jellyfish tentacles. Chrysaora quinquecirrha (sea nettle) Chiropsalmus quadrumanus (sea wasp) and Physalia physalis (Portuguese man-of-war) were used in the study. It was found that many of the chemicals traditionally used to treat jellyfish stings stimulated nematocyst discharge and did not relieve the pain. However there was immediate relief when a common anesthetic lidocaine was sprayed on the skin of testers in contact with jellyfish tentacles. Initial exposure of tentacle suspensions to lidocaine prevented the nematocyst discharge by subsequent exposure to acetic acid ethanol ammonia or bromelain. Thus lidocaine in addition to acting as an anesthetic on skin in contact with jellyfish tentacles inhibited nematocyst discharge possibly by blocking sodium and/or calcium channels of the nematocytes.     

Drought Leads to Collapse of Black-Tailed Prairie Dog Populations Reintroduced to the Chihuahuan Desert

ABSTRACT Recently, a conservation strategy developed to restore populations of black-tailed prairie dog (Cynomys ludovicianus) suggested reintroducing animals into the Chihuahuan Desert grasslands of the southwestern United States. Rainfall in desert habitats is lower and more variable compared to rainfall near the center of the prairie dog's range. Additionally, peak rainfall comes months after prairie dogs reproduce in these desert systems. Thus, southwestern populations may be less prolific and fluctuate more than those found in northerly climes. Using mark-recapture and mark-resight techniques, we estimated reproduction and monthly survival from 577 individuals inhabiting 6 reintroduced colonies from 2003 to 2005 in the northern Chihuahuan Desert. During 2003 precipitation was 64% of the long-term average, whereas both 2004 and 2005 had near-average precipitation. Probability that a female became pregnant, number of juvenile prairie dogs emerging from maternity burrows, and date of emergence were all correlated to adult female body mass. Adult monthly survival decreased from >0.95 during spring to 0.70 in summer 2003, following a rapid loss in adult body mass that coincided with low precipitation. In 2003 monthly juvenile survival was near zero on 2 of the 3 largest colonies and growth rates of juveniles were half that of subsequent years. Estimated population size declined by 68% (range = 18–91%) from 2003 to 2004, and 5 of 6 populations declined an average of 75% from their original introduction size. Prairie dog populations in desert environs may have a high risk of extirpation caused by weather patterns indicative of desert climates. Our results are important for those managers involved in the conservation of prairie dogs and we suggest that regional differences should be carefully considered prior to any reintroduction effort.     

Tracking Molecular Interactions in Membranes by Simultaneous ATR-FTIR-AFM

In situ atomic force microscopy (AFM) is an exceedingly powerful and useful technique for characterizing the structure and assembly of proteins in real-time, in situ, and especially at model membrane interfaces, such as supported planar lipid bilayers. There remains, however, a fundamental challenge with AFM-based imaging. Conclusions are inferred based on morphological or topographical features. It is conventionally very difficult to use AFM to confirm specific molecular conformation, especially in the case of protein-membrane interactions. In this case, a protein may undergo subtle conformational changes upon insertion in the membrane that may be critical to its function. AFM lacks the ability to directly measure such conformational changes and can, arguably, only resolve features that are topographically distinct. To address these issues, we have developed a platform that integrates in situ AFM with attenuated total reflection-Fourier transform infrared (ATR-FTIR) spectroscopy. This combination of tools provides a unique means of tracking, simultaneously, conformational changes, not resolvable by in situ AFM, with topographical details that are not readily identified by conventional spectroscopy. Preliminary studies of thermal transitions in supported lipid bilayers and direct evidence of lipid-induced conformational changes in adsorbed proteins illustrates the potential of this coupled in situ functional imaging strategy.     

MIC-A allele profiles and HLA class I associations in Behçet's disease

Recently a new family of non-classical MHC molecules, the MHC class I chain-related protein (MIC), encoded by genes located in the major histocompatability complex have been identified. On the basis of the location of MIC genes and the structure and expression of MIC molecules it has been postulated that MIC may be a susceptibility factor in Behçet's disease (BD). We investigated the association of the 16 described external domain alleles and the transmembrane triplet repeats of MIC-A with BD in a Middle Eastern population. DNA from ninety-five patients and 102 age- and sex-matched controls were analyzed by polymerase chain reaction using allele specific primers. Our results show an increase of MIC-A*009 in the BD patient group 44/95 (46%) compared with controls 24/102 (24%) (χ²=11.3, OR=2.8, P=0.00078). MIC-A*009 was also found to be strongly associated with HLA-B51 in the patients 39/44 (88%) when compared with controls 10/24 (42%) (χ²=4, P=0.04). MIC-A*009 was also found in linkage disequilibrium with HLA-B52, but only in controls. The A6 form of a MIC-A transmembrane triplet repeat was found to be significantly raised in the patients (80/95; 84%;) compared with controls (58/102, 57%) (χ²=17.5, OR=4, P=0.000028). Although the MIC-A associations described are highly significant, the association with HLA-B51 independently remains the most significant factor (χ²=56.8, P<10^–6). The data suggests that as both MIC-A*009 and A6 are in strong linkage disequilibrium with HLA-B51, they are unlikely to be the susceptibility gene for BD but may be markers for additional risk factors.     

SplitsTree: analyzing and visualizing evolutionary data

MOTIVATION: Real evolutionary data often contain a number of different and sometimes conflicting phylogenetic signals, and thus do not always clearly support a unique tree. To address this problem, Bandelt and Dress (Adv. Math., 92, 47-05, 1992) developed the method of split decomposition. For ideal data, this method gives rise to a tree, whereas less ideal data are represented by a tree-like network that may indicate evidence for different and conflicting phylogenies. RESULTS: SplitsTree is an interactive program, for analyzing and visualizing evolutionary data, that implements this approach. It also supports a number of distances transformations, the computation of parsimony splits, spectral analysis and bootstrapping.      

Conditionally Immortalized Neural Cell Lines: Potential Models for the Study of Neural Cell Function

Studies on primary cell cultures have contributed significantly to our understanding of neural cell function. Nevertheless, for many studies the value of these primary cell cultures has been limited by the time the cultures survivein vitro,the quantity of cellular material available for analysis, and the need to prepare the cells on a regular basis from fresh tissue. Techniques for immortalizing cells have existed for some time, but the repertoire of immortalizing genes has grown significantly. This has expanded our ability to generate useful cell lines of specific neural types that are better models of thein vivophenotype than previously. The constitutive expression of oncogenes keeps cells in a proliferative state that could lead to the loss of differentiated gene expression and function. An appealing improvement of immortalization methodology is the use of temperature-sensitive oncogenes that generate cell lines that can proliferate at a permissive temperature and “differentiate” at a nonpermissive temperature. The proliferation of such conditionally immortalized cell lines can be suppressed simply by increasing the temperature. Cell lines maintained at the nonpermissive temperature can enter into a stage in which they express differentiated properties of the cell. The potential ability of conditionally immortalized neural cell lines to accurately reflect theirin vivofunction has now been demonstrated on several occasions through transplantation experiments. In this report, the generation of these cell lines is described along with a discussion of their potential applications in neurobiology.