Coccolithophores: functional biodiversity, enzymes and bioprospecting

Emiliania huxleyi is a single celled, marine phytoplankton with global distribution. As a key species for global biogeochemical cycling, a variety of strains have been amassed in various culture collections. Using a library consisting of 52 strains of E. huxleyi and an 'in house' enzyme screening program, we have assessed the functional biodiversity within this species of fundamental importance to global biogeochemical cycling, whilst at the same time determining their potential for exploitation in biocatalytic applications. Here, we describe the screening of E. huxleyi strains, as well as a coccolithovirus infected strain, for commercially relevant biocatalytic enzymes such as acid/alkali phosphodiesterase, acid/alkali phosphomonoesterase, EC1.1.1-type dehydrogenase, EC1.3.1-type dehydrogenase and carboxylesterase.     

Nutritional quality of the protein in quinoa (Chenopodium quinoa,Willd) seeds

The nutritional quality of protein in quinoa seeds has been determined by amino acid assay and by animal feeding experiments. The amino acid composition of the protein in raw quinoa and washed quinoa show similar pattern. The first limiting amino acids were the aromatic amino acids thyrosine + phenylalanine giving a chemical score of 86 for protein in raw quinoa and 85 for protein in washed quinoa. Threonine was the next limiting amino acid followed by lysine. The amount of lysine and sulfur amino acids (methionine + cystine) was relatively high. In general, the content of essential amino acids in quinoa is higher than in common cereals. The animal experiments showed NPU values of 75.7, BV of 82.6 and TD value of 91.7 for the protein in raw quinoa. Results of the in-vitro enzymatic methods showed that the digestibility of the protein in quinoa is comparable to that of other high quality food proteins. The corresponding experiments carried out with samples of guinoa seeds, which have been processed to remove the saponins, showed that, the saponins do not exert any negative effect on the nutritive quality of the protein.     

Preparation of indigestible pyrodextrins from different starch sources

Starch-modifying processes, such as pyrodextrinization, are potential ways to alter the nutritional features of this polysaccharide. A widely used method for pyrodextrinizing maize starch was also applied to lentil, sorghum, cocoyam, sagu, and cassava starches, and the in vitro digestibility of the products was evaluated. Pyrodextrins were produced by heating starch at 140 degrees C for 3 h, with catalytic amounts of HCl. The enzymatically available starch content of all preparations decreased by 55-65% after modification. Thus, pyrodextrinization seems to be an effective way to produce indigestible glucans from different starches. Pyrodextrins obtained were complex mixtures of starch derivatives with a wide range of molecular weight as estimated by gel filtration chromatography. Both their molecular weight profiles and contents of indigestible fractions varied with starch source. Experiments with lentil and cassava starches showed that changing dextrinization conditions also affects the susceptibility to enzymatic hydrolysis of the product.     

Real-time monitoring of thermal flavor generation in skim milk powder using atmospheric pressure chemical ionization mass spectrometry

The feasibility of monitoring volatile flavor compounds formed by thermal treatment of skimmed milk powder in real time by atmospheric pressure chemical ionization mass spectrometry (APCIMS) was established. Skim milk powder samples were heated isothermally (70 to 120 degrees C) at different moisture contents (2.2 and 12.7 g water/100 g dry solids). Headspace was sampled and analyzed continuously in full scan mode (30-180 amu) by APCIMS. The identity of the volatile compounds monitored by APCIMS was confirmed by coupled GC-EI-APCIMS. The concentration measured by the APCIMS was the net effect of three processes, namely formation of the compound, partition from the skim milk powder into the gas phase, and dilution due to the headspace sampling method used. Preliminary experiments established that the technique could follow the effects of heating temperature and moisture content on the formation of selected compounds from skim milk powder.     

Effect of Inoculation with Plant Growth-Promoting Bacteria on Growth and Copper Uptake by Sunflowers

The effect of plant growth-promoting bacteria inoculation on Helianthus annuus growth and copper (Cu) uptake was investigated. For this, the strains CC22, CC24, CC30, and CC33 previously isolated from heavy metal- and hydrocarbon-polluted soil were selected for study. These strains were characterized on the basis of their 16S rDNA sequences and identified as Pseudomonas putida CC22, Enterobacter sakazakii CC24, Acinetobacter sp. CC30, and Acinetobacter sp. CC33. Strains were able to synthesize indole, solubilize phosphorus, and produce siderophores in vitro, which are proper characteristics of plant growth-promoting (PGP) bacteria. Bacteria were also able to bioaccumulate Cu(II), and most of them could use aromatic hydrocarbons as a sole carbon source. Furthermore, Acinetobacter sp. CC33 exhibited the greatest extent of Cu(II) accumulation, and CC30 the widest range for degrading hydrocarbons. Acinetobacter sp. CC30 was selected for pot experiments on the basis of its plant growth-promoting properties. Inoculation with CC30 significantly increased the plant biomass (dry weight and length of root and shoot) and improved the photosynthetic pigment content in non- and Cu-contaminated soil (p < 0.05). Additionally, plant Cu uptake was improved by CC30 inoculation showing a significantly enhanced root Cu content (p < 0.05). Our findings evidenced that the strain CC30 protected the plant against the deleterious effect of Cu contamination and improved the Cu extraction by plant, hence concluding that its inoculation represents an alternative to improve phytoremediation process of heavy metals, particularly Cu, in contaminated environments.     

Toxigenicity and pathogenicity of <Emphasis Type="Italic">Fusarium poae</Emphasis> and <Emphasis Type="Italic">Fusarium avenaceum</Emphasis> on wheat

In a field experiment between 2004 and 2006, 14 winter wheat varieties were inoculated with either a mixture of three isolates of F. poae or a mixture of three isolates of F. avenaceum. In a subsequent climate chamber experiment, the wheat variety Apogee was inoculated with individual single conidium isolates derived from the original poly conidium isolates used in the field. Disease symptoms on wheat heads were visually assessed, and the yield as well as the fungal incidence on harvested grains (field only) was determined. Furthermore, grains were analysed using LC-MS/MS to determine the content of Fusarium mycotoxins. In samples from field and climate chamber experiments, 60 to 4,860 μg kg⁻¹ nivalenol and 2,400 to 17,000 μg kg⁻¹ moniliformin were detected in grains infected with F. poae and F. avenaceum, respectively. Overall, isolate mixtures and individual isolates of F. avenaceum proved to be more pathogenic than those of F. poae, leading to a higher disease level, yield reductions up to 25%, and greater toxin contamination. For F. poae, all variables except for yield were strongly influenced by variety (field) and by isolate (climate chamber). For F. avenaceum, variety had a strong effect on all variables, but isolate effects on visual disease were not reflected in toxin production. Correlations between visual symptoms, fungal incidence, and toxin accumulation in grains are discussed.     

Exploiting generic platform technologies for the detection and identification of plant pathogens

The detection and identification of plant pathogens currently relies upon a very diverse range of techniques and skills, from traditional culturing and taxonomic skills to modern molecular-based methods. The wide range of methods employed reflects the great diversity of plant pathogens and the hosts they infect. The well-documented decline in taxonomic expertise, along with the need to develop ever more rapid and sensitive diagnostic methods has provided an impetus to develop technologies that are both generic and able to complement traditional skills and techniques. Real-time polymerase chain reaction (PCR) is emerging as one such generic platform technology and one that is well suited to high-throughput detection of a limited number of known target pathogens. Real-time PCR is now exploited as a front line diagnostic screening tool in human health, animal health, homeland security, biosecurity as well as plant health. Progress with developing generic techniques for plant pathogen identification, particularly of unknown samples, has been less rapid. Diagnostic microarrays and direct nucleic acid sequencing (de novo sequencing) both have potential as generic methods for the identification of unknown plant pathogens but are unlikely to be suitable as high-throughput detection techniques. This paper will review the application of generic technologies in the routine laboratory as well as highlighting some new techniques and the trend towards multi-disciplinary studies.     

Characterisation of novel <Emphasis Type="Italic">Fusarium graminearum</Emphasis> microsatellite markers in different <Emphasis Type="Italic">Fusarium</Emphasis> species from various countries

Fifteen novel microsatellite markers were isolated from Fusarium graminearum. The level of polymorphism at these novel and 13 previously published microsatellite markers was analysed in 33 F. graminearum strains from Europe, North America, and Nepal. The number of alleles for each of the novel markers ranged from 4 to 20 and gene diversity from 0.417 to 0.962. In comparison with the previously published markers, the resolution for distinguishing among different strains was slightly increased. Twenty-seven markers were also detectable in three F. culmorum strains and one F. crookwellense strain. None of the markers was detected in three F. avenaceum and four F. poae strains, underlining the potential use of these microsatellite markers for species differentiation.     

Evaluation of a serological test (indirect ELISA) for the diagnosis of sarcoptic mange in red foxes (Vulpes vulpes)

Sarcoptic mange occurs in many parts of the world and is common in populations of domestic and wild canids, including red foxes (Vulpes vulpes). In recent years, an indirect antibody enzyme-linked immunosorbent assay (ELISA), with higher sensitivity and specificity than traditional diagnostic methods, has been successfully applied in the diagnosis of sarcoptic mange in dogs. The same ELISA has also demonstrated specific antibodies to Sarcoptes scabiei in experimentally infected red foxes. The aim of this study was to evaluate the indirect ELISA when used to detect antibodies to S. scabiei in field sera from Swedish red foxes. One cohort of both infected and non-infected red foxes (cohort 1; n = 88), and one cohort of apparently non-infected foxes (cohort 2; n = 67) were examined for skin lesions and presence of S. scabiei by thorough visual examination at autopsy and skin scrapings. Samples of blood-tinted body liquid from the abdomen or thorax cavity were collected and analysed by the indirect ELISA. The relative sensitivity and specificity of the ELISA at different cut-offs (OD values) were estimated by comparing the test results to the infection status as determined by examination and skin scrapings. The highest combination of relative sensitivity and specificity, calculated based on cohort 1, was 95.4 and 100.0%, respectively. These estimates were constant for cut-offs 0.150-0.225, which included the cut-off based on the mean plus three standard deviations of test results from cohort 2 (0.165). It is concluded that this test can be useful in diagnosis and epidemiological studies of S. scabiei infection in red foxes.     

Ascorbic Acid Effectively Improved Lutein Extraction Yield from Australian Sweet Lupin Flour

Plant Foods for Human Nutrition - Lutein is a xanthophyll, a&nbsp;bioactive phytochemical that presents itself as colourful pigments in plants. Australian sweet lupin flour has been...