Structural studies of an exopolysaccharide produced by Streptococcus thermophilus THS

The structure of an extracellular polysaccharide (EPS) from Streptococcus thermophilus THS has been determined. A combination of component analysis, methylation analysis and NMR spectroscopy shows that the polysaccharide is composed of pentasaccharide repeating units. Sequential information was obtained by two-dimensional (1)H,(1)H-NOESY and (1)H,(13)C-HMBC NMR experiments. NMR data indicate different mobility within the EPS with a stiffer backbone and a more flexible side-chain.     

Carbohydrate distribution and cellular injuries in acid rain and cold-treated spruce needles

Summary The cellular structures of acid rain-irrigated needles of several provenances of Norway spruce (Picea abies L. Karst) seedlings were studied after winter experimental freezing. Frost injuries and recovery were characterized by visual damage scoring and classification of mesophyll cell alterations, also using histochemical methods for carbohydrate fluorescent staining. The treatment with-30° C during the late dormancy period was sufficient to cause significant injuries and intracellular degradation in the tissues of the green needles. The most affected seedlings in terms of visual injury scoring were found among those treated with clean water or at pH 3, while freezing injury, defined as an occlusion of phenolic substances in the central vacuole of the mesophyll cells, was most abundant in the needles from spruces irrigated either with clean water or at pH 4 or pH 3. Electron microscopy revealed the details of the injury, e. g. thinning out of the cytoplasm and chloroplast stroma, darkening of the chloroplasts and eventually swelling of the chloroplasts and protoplast. PAS and ConA reactions in the needle tissue revealed intense starch accumulation in the mesophyll and transfusion tissues as early as in March, with a tendency to increase, especially in the untreated needles during the recovery period. Plasma membrane disturbances were indicated by histochemical identification of callose deposits in the mesophyll cell walls, these being most abundant in the acid rain-treated needles. All these findings suggest that freezing at –30° C was more deleterious to the seedlings pretreated with acid or clean water than to those not given additional irrigation.     

Leukemia Prediction Using Sparse Logistic Regression

We describe a supervised prediction method for diagnosis of acute myeloid leukemia (AML) from patient samples based on flow cytometry measurements. We use a data driven approach with machine learning methods to train a computational model that takes in flow cytometry measurements from a single patient and gives a confidence score of the patient being AML-positive. Our solution is based on an regularized logistic regression model that aggregates AML test statistics calculated from individual test tubes with different cell populations and fluorescent markers. The model construction is entirely data driven and no prior biological knowledge is used. The described solution scored a 100% classification accuracy in the DREAM6/FlowCAP2 Molecular Classification of Acute Myeloid Leukaemia Challenge against a golden standard consisting of 20 AML-positive and 160 healthy patients. Here we perform a more extensive validation of the prediction model performance and further improve and simplify our original method showing that statistically equal results can be obtained by using simple average marker intensities as features in the logistic regression model. In addition to the logistic regression based model, we also present other classification models and compare their performance quantitatively. The key benefit in our prediction method compared to other solutions with similar performance is that our model only uses a small fraction of the flow cytometry measurements making our solution highly economical.     

Heat shock protein synthesis is induced by diethyl phthalate but not by di(2-ethylhexyl) phthalate in radish (Raphanus sativus)

The toxicity and effects on protein synthesis of the phthalate esters diethyl phthalate (DEP) and di(2-ethylhexyl) phthalate (DEHP) was studied in radish seedlings (Raphanus sativus cv. Kööpenhaminan tori). Phthalate esters are a class of commercially important compounds used mainly as plasticizers in high molecular-weight polymers such as many plastics. They can enter soil through various routes and can affect plant growth and development. First the effect of DEP and DEHP on the growth of radish seedlings was determined in an aqueous medium. It was found that DEP, but not DEHP, caused retardation of growth in radish. A further investigation on protein synthesis during DEP-stress was executed by in vivo protein labeling combined with two-dimensional gel electrophoresis (2D-PAGE). For comparisons with known stress-induced proteins a similar experiment was done with heat shock, and the induced heat shock proteins (HSPs) were compared with those of DEP-stress. The results showed that certain HSPs can be used as an indicator of DEP-stress, although the synthesis of most HSPs was not affected by DEP. DEP also elicited the synthesis of numerous proteins found only in DEP-treated roots. The toxic effect of phthalate esters and the roles of the induced proteins are discussed.     

Expression of a fungal hydrophobin in the Saccharomyces cerevisiae cell wall: effect on cell surface properties and immobilization

The aim of this work was to modify the cell surface properties of Saccharomyces cerevisiae by expression of the HFBI hydrophobin of the filamentous fungus Trichoderma reesei on the yeast cell surface. The second aim was to study the immobilization capacity of the modified cells. Fusion to the Flo1p flocculin was used to target the HFBI moiety to the cell wall. Determination of cell surface characteristics with contact angle and zeta potential measurements indicated that HFBI-producing cells are more apolar and slightly less negatively charged than the parent cells. Adsorption of the yeast cells to different commercial supports was studied. A twofold increase in the binding affinity of the hydrophobin-producing yeast to hydrophobic silicone-based materials was observed, while no improvement in the interaction with hydrophilic carriers could be seen compared to that of the parent cells. Hydrophobic interactions between the yeast cells and the support are suggested to play a major role in attachment. Also, a slight increase in the initial adsorption rate of the hydrophobin yeast was observed. Furthermore, due to the engineered cell surface, hydrophobin-producing yeast cells were efficiently separated in an aqueous two-phase system by using a nonionic polyoxyethylene detergent, C(12-18)EO(5).     

Structural studies of the exopolysaccharide produced by Lactobacillus rhamnosus strain GG (ATCC 53103)

The structure of the extracellular polysaccharide (EPS) from Lactobacillus rhamnosus strain GG has been investigated. In combination with component analysis, NMR spectroscopy shows that the polysaccharide is composed of hexasaccharide repeating units. Sequential information was obtained by two-dimensional (1)H,(1)H-NOESY, and (1)H,(13)C-HMBC NMR techniques. The structure of the repeating unit of the EPS from Lactobacillus rhamnosus strain GG was determined as: [carbohydrate structure: see text]     

Characterization of a highly evolved vaccine-derived poliovirus type 3 isolated from sewage in Estonia

Two types of vaccine-derived polioviruses have been recently designated to emphasize the different origins of the evolved viruses: circulating vaccine-derived polioviruses (cVDPV) associated with outbreaks of paralytic disease and strains isolated from chronically infected immunodeficient individuals (iVDPV). We describe here a type 3 VDPV (PV3/EST/02/E252; later E252) isolated from sewage collected in Tallinn, Estonia, in October 2002. Due to aberrant properties in subtyping, the virus was subjected to detailed characterization. Partial genomic sequencing suggested that the closest relative was the oral vaccine strain PV3/Sabin, but the two virus strains shared only 86.7% of the 900 nucleotides (nt) coding for the capsid protein VP1. Phylogenetic analysis of the nearly complete genome [nt 19 to poly(A)] revealed multiple nucleotide substitutions throughout the genome and a possible Sabin 3/Sabin 1-recombination junction site in the 2C coding region. A calculation based on the estimated mutation frequency of the P1 region of polioviruses suggested that the E252 virus might have replicated in one or more individuals for approximately 10 years. No persons chronically excreting poliovirus are known in Estonia. Amino acid substitutions were seen in all known antigenic sites, which was consistent with the observed aberrant antigenic properties of the virus demonstrated by both monoclonal antibodies and human sera from vaccinated children. In spite of the apparent transmission potential, no evidence was obtained for circulation of the virus in the Estonian population.     

Isolation and characterization of microsatellites in Drosophila virilis and their cross species amplification in members of the D. virilis group

We report the isolation and cross species amplification of 42 Drosophila virilis microsatellite loci. Nine loci were isolated from mapped P1 bacteriophage clones and 33 were obtained from genomic DNA or GenBank searches. Cross species amplification was tested for all members of the D. virilis group. The amplification success was high (varying from 45% to 100%) and most of the loci were polymorphic. This set of loci can be applied for several genetic studies such as mapping behavioural quantitative trait loci (QTL) and for studying population structure in a phylogeographical framework in D. virilis group species.     

Evaluation of chemical pretreatment of contaminated soil for improved PAH bioremediation

The efficiency of several chemical treatments as potential enhancers of the biodegradation of polycyclic aromatic hydrocarbons (PAHs) in contaminated soil was evaluated by analyzing the mineralization of ¹⁴C-labeled phenanthrene, pyrene, and benzo(a)pyrene. The effect of nonionic surfactants with Fenton oxidation and combinations of surfactants with the Fenton oxidation was evaluated in a microtiter plate assay. The surfactants selected for the study were Tween 80, Brij 35, Tergitol NP-10, and Triton X-100. The addition of Fentons reagent significantly enhanced the mineralization of pyrene at the two concentrations studied: 2.8 M H₂O₂ with 0.1 M FeSO₄ and 0.7 M H₂O₂ with 0.025 M FeSO₄. Phenanthrene mineralization was also positively induced by the Fenton treatments. However, none of the treatments had a significant effect on benzo(a)pyrene mineralization. Surfactant additions at concentrations of 20% and 80% of the aqueous critical micelle concentration did not significantly affect the mineralization rates. When surfactant addition was combined with the Fenton oxidation, reduced mineralization rates were obtained when compared with mineralization after Fentons treatment alone. The results indicate that the addition of Fentons reagent may enhance the mineralization of PAHs in contaminated soil, whereas the addition of surfactants has no significant beneficial effect. The efficiency of the Fenton oxidation may decrease when surfactants are added simultaneously with Fentons reagent to contaminated soil.     

Characterization of a shiitake mutant strain producing ballshaped fruiting bodies

Growth characteristics of a spontaneous mutant of shiitake Lentinula edodes (Berk.) Pegler were studied. The mutant was first detected as a result of changes in the growth habit of the normal strain in the liquid medium. Abundant formation of aerial hyphae was distinctive. In sawdust logs the mutant strain produced abnormal basidiocarps, lacking stipe, gill and spore formation.
Growth rates of the normal and the mutant strain were compared in two liquid media: malt-yeast extract and Leatham's medium. The increase in dry weight of the mutant's mycelium was much higher than that of the wild type in both media, which indicated better adaptation to liquid culture. In the sawdust, however, growth of the mutant was slower than that of the normal strain. The mutant's intracellular protein content was lower than that of the normal strain. The pH of the liquid cultures differed: the wild type decreased the pH during growth, while the mutant increased the pH. Comparison of the protein and esterase isoenzyme profiles of the vegetative hyphae of both strains indicated profound differences. One protein (pI 6.5, 39 kDa), which in earlier studies has been found to be typical of L. edodes species, was absent from the mutant's profile. Differences in the esterase profile were also clear.