Seeds are notoriously high in polyphenols and polysaccharides, which reduce RNA quality and yield, and interfere with downstream applications. We present simple modifications to a rapid RNA extraction protocol for use with seeds. The inclusion of polyethylene glycol in place of polyvinylpyrrolidone reduced polyphenol and polysaccharide contamination. In addition, replacing NaCl with KCl improved the RNA yield from Arabidopsis seeds still bound by mucilage. On extraction of Arabidopsis seed recovered from field soils clean RNA pellets with no accompanying gelatinous matrix (polysaccharide) were seen, with A260/230 ratios greater than 1.8 confirming the lack of polysaccharide carry-over. When Brassica oleracea and Sinapis arvensis seeds were extracted, 260/230 ratios greater than 1.8 were seen. RNA yields in excess of 10 µg per 100 mg seed suitable for RT-QPCR were obtained.

Aggregation of colloidal clay particles (Na-montmorillonite) by CaCl2 and anionic polysaccharide (succinoglycan) in turbulent conditions was investigated using time-resolved size measurements by laser diffraction on diluted (50 mg l –1) and stirred suspensions. Excess of Ca2+ ions promotes coagulation of the clay, reducing interparticle repulsions, and allows adsorption of succinoglycan, inducing bridging flocculation. Growth/breakage cycles, characteristic of the turbulent conditions, cause the macromolecules to be incorporated in the innermost of the flocs, where the morphological units are shown by confocal microscopy to be the micrometric Ca-clay particles. Such incorporation results in an increased floc tensile strength, depending on the amount of macromolecules adsorbed, with a maximum at polysaccharide concentrations of 2 wt.% with respect to clay mass.

The potential of various quantitative lateral flow (LF) based assays utilizing up-converting phosphor (UCP) reporters for the diagnosis of schistosomiasis is reviewed including recent developments. Active infections are demonstrated by screening for the presence of regurgitated worm antigens (genus specific polysaccharides), whereas anti-Schistosoma antibodies may indicate ongoing as well as past infections. The circulating anodic antigen (CAA) in serum or urine (and potentially also saliva) is identified as the marker that may allow detection of single-worm infections. Quantitation of antigen levels is a reliable method to study effects of drug administration, worm burden and anti-fecundity mechanisms. Moreover, the ratio of CAA and circulating cathodic antigen (CCA) is postulated to facilitate identification of either Schistosoma mansoni or Schistosoma haematobium infections. The UCP-LF assays allow simultaneous detection of multiple targets on a single strip, a valuable feature for antibody detection assays. Although antibody detection in endemic regions is not a useful tool to diagnose active infections, it gains potential when the ratio of different classes of antibody specific for the parasite/disease can be determined. The UCP-LF antibody assay format allows this type of multiplexing, including testing a linear array of up to 20 different targets. Multiple test spots would allow detection of specific antibodies, e.g. against different Schistosoma species or other pathogens as soil-transmitted helminths. Concluding, the different UCP-LF based assays for diagnosis of schistosomiasis provide a collection of tests with relatively low complexity and high sensitivity, covering the full range of diagnostics needed in control programmes for mapping, screening and monitoring.

Chitin is a natural polysaccharide found in many diverse phyla and almost always occurs in association with protein. In this study the chitin polymer is characterized by X-ray diffraction from its naturally occurring unprocessed state through various steps used in the purification procedure. In addition, the effect of different treatments on the final product is examined. These studies show that native chitin has a characteristic diffraction pattern that is not altered by the mild treatments used to isolate relatively pure chitin. Chitins prepared from different sources exhibit the same characteristic diffraction pattern. In addition, chitin films prepared using non-degrading solvents retain most of the characteristic patterns. De-acylation of chitin to produce chitosan results in large changes to the diffraction pattern. To a very limited extent features present in the diffraction pattern of native chitin can be recovered by re-acylation of chitosan.

Colonial Microcystis aeruginosa were obtained when the unicellular algae were exposed to flagellate Ochromonas sp. filtrate. To investigate the benefit of this morphological change, flagellates were added into cultures of unicellular and colonial M. aeruginosa, respectively. The clearance rates of flagellates on algae were markedly decreased when they were cultivated with induced colonial M. aeruginosa. This result indicated that colony formation in M. aeruginosa was a predator-induced defense, which could reduce predation risk from flagellate. The increased content of soluble extracellular polysaccharide (sEPS) and bound extracellular polysaccharide (bEPS) may play an important role in adhering M. aeruginosa cells together to form colonies. The decrease of ΦPS II and the increase of sinking rates of induced colonial M. aeruginosa showed that the costs of grazed-induced colony formation in M. aeruginosa may reflect in the photosystem II efficiency, and in the sinking rates.

The production and composition of extracellular polymeric substances (EPS) in axenic batch cultures of the benthic marine epipelic diatoms Navicula salinarum and Cylindrotheca closterium were investigated. EPS was secreted into the medium and the bulk was loosely associated with the cells. Neither N. salinarum nor C. closterium formed a well-defined polysaccharide capsule. EPS of both N. salinarum and C. closterium consisted predominantly of polysaccharide but small quantities of protein were present as well. EPS also contained uronic acids and SO42− groups. Analysis of monosaccharides using gas chromatography showed that for both species glucose and xylose were the main constituents, but several other monosaccharides were present in smaller quantities. Two fractions of EPS were distinguished: a small amount was secreted into the medium and a second fraction was extracted in water at 30°C. For both species the two fractions differed somewhat in composition, indicating that they represented two different types of EPS. The EPS produced by N. salinarum and by C. closterium differed in their composition. The rate of EPS production in batch culture was highest during the transition from exponential growth to stationary growth. Negatively charged groups such as uronic acids and sulphated sugars determine the adhesion capacity of EPS and probably play an important role in the stabilization of intertidal sediments on which these diatoms grow and produce biofilms.

A fructan polymerase activity was partially purified and concentrated by sequential acid and salt precipitation from extracts of excised, illuminated leaves of timothy grass (Phleum pratense). The polymerase catalysed the de novo synthesis of oligo- and polyfructan from sucrose as sole substrate at near-physiological rates (0.5–0.9 mg g−1 fresh mass h−1; 0.9–1.5 nkat g−1). Rates of in vitro polymerisation were high, at up to 4.1 mg cm−3 h−1 (7.1 nkat cm−3) of total products of degree of polymerization greater than 2 (DP>2). The trisaccharides 1-kestose and 6-kestose together with oligosaccharides of up to DP[les ]c. 10 were synthesized in under 2 h at 30°C. In longer incubations, ethanol-precipitable polymers of DP=c. 10–35 (1.6–5.7 kDa) were detected by anion-exchange chromatography and pulsed amperometry. When this polymeric product was used as a primer and re-incubated with fresh enzyme and sucrose, abundant polymers of up to DP=50 (8.1 kDa) were formed. The structure of the polymeric enzyme product was compared with native fructan from timothy leaves and with standard inulin, using glycosyl-linkage analysis followed by identification of partially methylated alditol acetate derivatives by GC–MS. The deduced structure was a linear (unbranched) 2,6-linked fructose chain terminated with glucose and fructose. The linkage structures of the native and enzyme-generated polymers were identical, increasing confidence in the physiological relevance of the activity. After ultracentrifugation of tissue homogenates at 265 000 gav, the polymerase remained in the supernatant, demonstrating no tight association with particulate components. The polymerizing reaction was dependent on enzyme concentration, requiring at least 3 g fresh mass equivalent cm−3 (c. 2.7 nkat cm−3) for the efficient in vitro generation of fructans of DP>3. In common with other trisaccharide-synthesizing and oligofructan-glycosylating enzymes from grasses, the polymerase reaction exhibited both a maximal velocity at pH 5.0–5.5 and a low affinity for sucrose. The polymerization reaction did not saturate fully even at 1.5 M sucrose, and the concentration causing half maximal velocity (apparent Km) was c. 560 mM. The preparation contained substantial invertase activity (1.8 mg sucrose g−1 fresh mass h−1=1.5 nkat g−1 fresh mass) with a Km for sucrose hydrolysis of 5 mM. A single peak of polymerase activity with an Mr of 51 kDa was recovered from size-exclusion chromatography (SEC). Invertases of Mr 51 and 110 kDa were identified in the preparation. The 110-kDa invertase isoform exhibited no polymerase activity, but synthesized trisaccharide (mainly 1-kestose) from sucrose. The 51-kDa isoform co-eluted with the polymerase. The trisaccharide fraction produced by this isoform contained abundant 1- and 6-kestose. After SEC, the purification of the polymerase was 41-fold relative to the original tissue homogenate. The properties of enzymatic polymerization of fructan are discussed with respect to the physiology of accumulation in grass leaves and other systems.

The effect of copper on carbohydrate production in the diatom Cylindrotheca fusiformis and in the dinoflagellate Gymnodinium sp. was examined in batch cultures. Copper was shown to inhibit the growth of the two species to different extents: in particular the dinoflagellate was highly sensitive to the metal when metal chelators were absent. Analysis of the carbohydrate content showed that C. fusiformis produced a much higher total amount than Gymnodinium sp. and cells of the diatom which were exposed to toxic copper concentrations displayed a higher carbohydrate production which paralleled the increase in copper concentrations. The increased production could be attributed to extracellular rather than to intracellular carbohydrate and it occurred after 12 and 16 days of growth, respectively, for cells exposed to 0·2 and 0·5 mg l−1 Cu. The response of carbohydrate production by Gymnodinium sp. to the presence of copper was much smaller and occurred only in late stationary phase of growth (21st day). An analysis of the copper content showed that the amount of metal accumulated by Gymnodinium sp. was similar to that absorbed by C. fusiformis, but it was a larger fraction of the quantity supplied. The results presented indicate that carbohydrate extrusion is one of the mechanisms which may prevent toxic effects due to the presence of copper in the culture medium.

Xanthan gum (15 g/d) was given for 10 d to eighteen normal volunteers. In vivo measurements of stool output, transit time, frequency of defaecation and flatulence were compared with a preceding control period of 10 d. At the end of the control and test periods fresh faecal homogenate from each subject was anaerobically incubated with xanthan gum and control solutions to assess the ability of the bacteria to break down the gum. Xanthan gum was found to be a highly efficient laxative agent causing significant increases in stool output (P < 0.01), frequency of defaecation (P < 0.05) and flatulence (P < 0.01) whilst having variable effects on transit time. Before feeding xanthan gum, faecal samples from twelve of the eighteen subjects could reduce the viscosity of the gum in vitro. This rose to sixteen of the eighteen with significantly greater amounts (P < 0.05) of hydrogen and short-chain fatty acids also being produced, indicating bacterial adaptation in the presence of the substrate. Correlations between the in vivo and in vitro findings did not substantiate claims that the in vivo effect of a given polysaccharide can be predicted from its fermentation characteristics in vitro.