Partitioning and diversity of nuclear and organelle markers in native and introduced populations of Epipactis helleborine (Orchidaceae)

Variability of allozymes (1170 individuals, 47 populations) and chloroplast DNA (692 individuals, 29 populations) was examined in native European and introduced North American populations of Epipactis helleborine (Orchidaceae). At the species level, the percentage of allozyme loci that were polymorphic (P(99)) was 67%, with a mean of 2.11 alleles (A) per locus, and an expected heterozygosity (H(exp)) of 0.294. At the population level, mean P(99) = 56%, mean A = 1.81, and mean H(exp) = 0.231. Although field observations suggest that self-pollination occurs frequently, populations had a genetic structure consistent with Hardy-Weinberg expectations and random mating (mean F(IS) = 0.002). There was significant deviation from panmixia associated with population differentiation (mean F(ST) = 0.206). The distribution of two chloroplast haplotypes showed that 15 of the 29 populations were polymorphic. Using both nuclear and organelle F(ST) estimates, a pollen to seed flow ratio of 1.43?:?1 was calculated. This is very low compared with published estimates for other plant groups, consistent with the high dispersability of orchid seeds. Finally, there was no evidence for a genetic bottleneck associated with the introduction of E. helleborine to North America.     

Variation in the Composition of Milk of Asian Elephants (Elephas maximus) Throughout Lactation

We investigated milk nutrient composition from three Asian elephant cows over the first 3 years of lactation, including two consecutive lactations in one cow. Body mass gain is presented for three calves during the first year. Milk samples (n = 74) were analyzed for dry matter (DM), fat, crude protein (CP), sugar, ash, calcium (Ca), phosphorus (P), and potassium (K); gross energy (GE) was calculated. Concentrations of most nutrients changed over lactation: DM, fat, CP, Ca, P, and GE were positively correlated to calf age; sugar was negatively correlated to calf age. GE doubled between birth (1 kcal/g) and 2 years of age (2 kcal/g). After accounting for calf age, GE, fat, Ca, and P concentrations differed among the cows. Milk composition also differed between two lactations from the same cow. When milk nutrients were expressed on a mg per kcal basis, the pattern changes: CP, Ca, and P remained relatively constant over lactation on a per energy basis. Calf mass quadrupled over the first year of life; mass gain was linear at 0.9 kg/day. Asian elephant milk composition is variable, both across lactations and between cows, complicating efforts to determine representative values for comparative studies and for the formulation of elephant milk formulas. The fact that CP, Ca, and P were all relatively constant when expressed on a per energy basis may be of biological significance. The increase in nutrient density over lactation undoubtedly limits maternal water loss, reducing the volume of milk necessary to support the calf. Zoo Biol 32:291–298, 2013. © 2012 Wiley Periodicals, Inc.     

Cloning and functional characterisation of polyunsaturated fatty acid elongases of marine and freshwater teleost fish

Enzymes that lengthen the carbon chain of polyunsaturated fatty acids are key to the biosynthesis of the highly unsaturated fatty acids, arachidonic, eicosapentaenoic and docosahexaenoic acids from linoleic and alpha-linolenic acids. A Mortierella alpina cDNA polyunsaturated fatty acid elongase sequence identified mammalian, amphibian, zebrafish and insect expressed sequence tags (ESTs) in GenBank. Consensus primers were designed in conserved motifs and used to isolate full length cDNA from livers of several fish species by Rapid Amplification of cDNA Ends (RACE). The amplified cDNAs encoded putative open reading frames (ORFs) of 288-294 amino acids that were highly conserved among the fish species. Heterologous expression in yeast, Saccharomyces cerevisiae, demonstrated that all of the ORFs encoded elongases with the ability to lengthen polyunsaturated fatty acid substrates with chain lengths from C18 to C22 and also monounsaturated fatty acids, but not saturated fatty acids. There were differences in the functional competence of the elongases from different fish species. Most of the fish elongases showed a pattern of activity towards different fatty acid substrates in the rank order C18>C20>C22, although the tilapia and turbot elongases had similar activity towards 18:4n-3 and 20:5n-3. The fish elongases generally showed greater activity or similar activities with n-3 than with n-6 homologues, with the exception of the cod enzyme which was more active towards n-6 fatty acids.     

Visualisation of mycorrhizal fungal structures and quantification of their surface area and volume using laser scanning confocal microscopy

 A method has been developed for the visualisation and three-dimensional (3D) measurement of mycorrhizal fungal structures inside plant roots. Sections of Allium porrum L. roots colonised by Glomus sp. 'City Beach' (WUM 16) and Lilium sp. roots colonised by Scutellospora calospora (Nicol. & Gerd.) Walker & Sanders (WUM 12(2)) were stained with acid fuchsin. This allowed fluorescence from the fungal structures to be observed under a laser scanning confocal microscope (LSCM) without interference from the plant cells. A series of horizontal optical sections were collected from a Glomus sp. arbuscule and from a hyphal coil of S. calospora. These data were used to produce extended focus images. Axial distortion in microscopic visualisation due to the refractive index mismatch between the immersion and mounting media was quantified using vertical scanning of the hyphae. A correction factor of 0.71 μm was used for the z-interval between the xy-slices. A series of binary xy-images from each structure was rendered into a 3D graphical model for viewing. The volume and surface area of the structures were estimated using computerised 3D measurement and also by stereological integration of binary xy-images. With both structures, the surface area estimates varied greatly between the two measuring systems, whereas differences in volume estimates were small. Computerised 3D measurement was considered more accurate than stereological integration of confocal binary images. Accepted: 25 August 1999     

Estimating Incidence of Attachment of Pasteuria penetrans Endospores to Meloidogyne spp. with Tally Thresholds

Pasteuria penetrans has .been identified as an important biological control agent of root-knot nematodes. In this study the use of tally thresholds was evaluated for estimating P. penetrans endospore attachment to second-stage juveniles (J2) of Meloidogyne spp. A tally threshold (T) is defined as the maximum number of individuals in a sample unit that may be treated as absent based on binomial sampling. Three different data sets that originated from centrifugal bioassay, incubation bioassay, and field experiments were investigated. The data sets each contained 70, 33, and 111 estimates of the mean number of endospores attached per J2 (m), respectively. Empirical relationships between m and proportions of J2 with ≤T endospores attached (P_T) were developed using parameters from the linear regression of ln(m) on P_T (0 < P_T < 1): ln(m) = a + b P_T, T was set to 0, 1, 2, 3, 4, 5, 8, and 10 endospores/J2. The results indicated that the variances of linear equations tended to decrease with increasing T values for all three data sets. T values of 0, 1, 8, and 10 endospores/J2 for centrifugal bioassay and incubation bioassay, and of 0, 1, 2, and 3 endospores/J2 for field experiments were associated with an r² of >= 0.8. These T values were robust for estimating m from P_T, reducing the variability as well as the time and effort spent in estimating the mean number of endospores attached per J2.     

Growth regulatory peptide production by human breast carcinoma cells

The mechanisms by which human breast cancers regulate their own growth have been studied by us in an in vitro model system. We showed that specific growth factors (IGF-I, TGF alpha, PDGF) are secreted by human breast cancer cells. A variety of experiments suggest that they are involved in tumor growth and progression. These activities are induced by estradiol in hormone-dependent breast cancer cells and secreted constitutively by estrogen-independent cells. Concentrates of conditioned medium derived from breast cancer cells can induce the growth of hormone-dependent cells in vivo in athymic nude mice. Hormone-dependent breast cancer cells also secrete TGF beta. TGF beta is growth inhibitory. Growth inhibitors such as antiestrogens or glucocorticoids increase TGF beta secretion. An antiestrogen-resistant mutant of MCF-7 cells does not secrete TGF beta when treated with antiestrogen, but is growth inhibited when treated with exogenous TGF beta. Thus, TGF beta functions as a negative autocrine growth regulator and is probably responsible for some of the growth inhibitory effects of antiestrogens.     

Combination treatment with cholestyramine and bezafibrate for heterozygous familial hypercholesterolaemia.

Cholestyramine and bezafibrate were compared individually and in combination in the treatment of 18 patients with heterozygous familial hypercholesterolaemia. The study used a double blind, placebo controlled block design with a placebo run in period of two months followed by three phases of active treatment, each of two months'' duration. Patients were randomly allocated to one of the six possible sequences of medication so that three patients would be treated with each sequence. Two patients withdrew from the study before completion. The median concentration of total cholesterol decreased from 9.65 mmol/l (interquartile range 8.62 to 8.72) to 7.24 mmol/l (6.70 to 7.52) with cholestyramine, to 8.09 mmol/l (7.18 to 8.68) with bezafibrate, and to 6.31 mmol/l (5.84 to 7.27) with the combination. This fall was due almost entirely to a decrease in the low density lipoprotein cholesterol concentration, and the combination was significantly more effective than either drug alone. The 98% confidence intervals for the median differences between the combination and cholestyramine and the combination and bezafibrate were 0.04 to 1.49 mmol/l and 0.51 to 2.18 mmol/l respectively. These results suggest that this combination is an effective and useful treatment in heterozygous familial hypercholesterolaemia.     

Trichinella spiralis: Proteins and antigens isolated from a large-particle fraction derived from the muscle larva

Proteins and antigens derived from a large-particle fraction of muscle larvae of Trichinella spiralis (i.e., the S₃ fraction) were characterized in terms of their molecular weights, isoelectric points, carbohydrate contents, electrophoretic mobilities, antigenicity, and their ability to induce protection in mice. Gel filtration on Sephacryl S-200 yielded 5 major peaks of material while electrophoresis in polyacrylamide gel with sodium dodecyl sulfate revealed a minimum of 28 proteins ranging in MW from 11,000 to 200,000. Analytical isoelectric focusing on acrylamide gel yielded 37 bands of protein, while the periodic acid-Schiff reaction performed on a similar gel revealed 22 glycoproteins. Most proteins were within a pI range of 4.0–7.0, while all of the glycoproteins had pI ranging from 4.0 to 6.5. Immunoelectrophoresis of the S₃ fraction using hyperimmune rabbit serum demonstrated a minimum of 19 precipitin arcs, while crossed immunoelectrophoresis yielded 16 peaks. These determinations were made on several batches of material isolated in the same fashion and gave the same results. Preparative isoelectric focusing yielded 30 fractions. These fractions were assayed for the presence of antigens, then pooled and tested for their ability to induce protection in mice against an oral challenge infection. Fused rocket immunoelectrophoresis of all 30 fractions revealed the presence of a minimum of 18 antigens with pI ranging from 4.0 to 9.0. The pooled fractions (i.e., 1–9; 10–20; 21–30) all protected mice against oral challenge infection, while fraction 5 (pI = 4.3) protected best.