Increased bioethanol production from commercial tobacco cultivars overexpressing thioredoxin f grown under field conditions

Bioethanol is mainly produced from food crops such as sugar cane and maize, and this has been held partly responsible for the rise of food commodity prices. Tobacco, integrated in biorefinery facilities for the extraction of different compounds, could become an alternative feedstock for biofuel production. When grown for energy production, using high plant densities and several mowings during the growing season, tobacco can produce large amounts of inexpensive green biomass. We have bred two commercial tobacco cultivars (Virginia Gold and Havana 503B) to increase the carbohydrate content by the overexpression of thioredoxin f in the chloroplast. Marker-free transplastomic plants were recovered and their agronomic performance under field conditions was evaluated. These plants were phenotypically equivalent to their wild types yet showed increased starch (up to 280 %) and soluble sugar (up to 74 %) contents in leaves relative to their control plants. Fermentable sugars released from the stalk were also higher (up to 24 %) for transplastomic plants. After heat pretreatment, enzymatic hydrolysis and yeast fermentation of leaf and stalk hydrolysates, an average of 20–40 % more ethanol was obtained from transplastomic plants than their wild-type controls. We propose an integral exploitation of the entire tobacco plant managed as a forage crop (harvesting sugar and starch-rich leaves and lignocellulosic stalks) that could considerably cheapen the entire production process.     

Associations of physical activity with gut microbiota in pre-adolescent children

[Purpose] To determine whether physical activity (PA), primarily the recommended 60 minutes of moderate-to-vigorous PA, is associated with gut bacterial microbiota in 10-year-old children.[Methods] The Block Physical Activity Screener, which provides minutes/day PA variables, was used to determine whether the child met the PA recommendations. 16S rRNA sequencing was performed on stool samples from the children to profile the composition of their gut bacterial microbiota. Differences in alpha diversity metrics (richness, Pielou’s evenness, and Faith’s phylogenetic diversity) by PA were determined using linear regression, whereas beta diversity (unweighted and weighted UniFrac) relationships were assessed using PERMANOVA. Taxon relative abundance differentials were determined using DESeq2.[Results] The analytic sample included 321 children with both PA and 16S rRNA sequencing data (mean age [SD] =10.2 [0.8] years; 54.2% male; 62.9% African American), where 189 (58.9%) met the PA recommendations. After adjusting for covariates, meeting the PA recommendations as well as minutes/day PA variables were not significantly associated with gut richness, evenness, or diversity (p ≥ 0.19). However, meeting the PA recommendations (weighted UniFrac R² = 0.014, p = 0.001) was significantly associated with distinct gut bacterial composition. These compositional differences were partly characterized by increased abundance of Megamonas and Anaerovorax as well as specific Christensenellaceae_R-7_group taxa in children with higher PA.[Conclusion] Children who met the recommendations of PA had altered gut microbiota compositions. Whether this translates to a reduced risk of obesity or associated metabolic diseases is still unclear.     

Recruitment of TBK1 to cytosol‐invading Salmonella induces WIPI2‐dependent antibacterial autophagy

Mammalian cells deploy autophagy to defend their cytosol against bacterial invaders. Anti‐bacterial autophagy relies on the core autophagy machinery, cargo receptors, and “eat‐me” signals such as galectin‐8 and ubiquitin that label bacteria as autophagy cargo. Anti‐bacterial autophagy also requires the kinase TBK1, whose role in autophagy has remained enigmatic. Here we show that recruitment of WIPI2, itself essential for anti‐bacterial autophagy, is dependent on the localization of catalytically active TBK1 to the vicinity of cytosolic bacteria. Experimental manipulation of TBK1 recruitment revealed that engagement of TBK1 with any of a variety of Salmonella‐associated “eat‐me” signals, including host‐derived glycans and K48‐ and K63‐linked ubiquitin chains, suffices to restrict bacterial proliferation. Promiscuity in recruiting TBK1 via independent signals may buffer TBK1 functionality from potential bacterial antagonism and thus be of evolutionary advantage to the host.     

Relationship between monokaryotic growth rate and mating type in the edible basidiomycete Pleurotus ostreatus

The edible fungus Pleurotus ostreatus (oyster mushroom) is an industrially produced heterothallic homobasidiomycete whose mating is controlled by a bifactorial tetrapolar genetic system. Two mating loci (matA and matB) control different steps of hyphal fusion, nuclear migration, and nuclear sorting during the onset and progress of the dikaryotic growth. Previous studies have shown that the segregation of the alleles present at the matB locus differs from that expected for a single locus because (i) new nonparental B alleles appeared in the progeny and (ii) there was a distortion in the segregation of the genomic regions close to this mating locus. In this study, we pursued these observations by using a genetic approach based on the identification of molecular markers linked to the matB locus that allowed us to dissect it into two genetically linked subunits (matBalpha and matBbeta) and to correlate the presence of specific matBalpha and matA alleles with differences in monokaryotic growth rate. The availability of these molecular markers and the mating type dependence of growth rate in monokaryons can be helpful for marker-assisted selection of fast-growing monokaryons to be used in the construction of dikaryons able to colonize the substrate faster than the competitors responsible for reductions in the industrial yield of this fungus.     

Mapping of Genomic Regions (Quantitative Trait Loci) Controlling Production and Quality in Industrial Cultures of the Edible Basidiomycete Pleurotus ostreatus

Industrial production of the edible basidiomycete Pleurotus ostreatus (oyster mushroom) is based on a solid fermentation process in which a limited number of selected strains are used. Optimization of industrial mushroom production depends on improving the culture process and breeding new strains with higher yields and productivities. Traditionally, fungal breeding has been carried out by an empirical trial and error process. In this study, we used a different approach by mapping quantitative trait loci (QTLs) controlling culture production and quality within the framework of the genetic linkage map of P. ostreatus. Ten production traits and four quality traits were studied and mapped. The production QTLs identified explain nearly one-half of the production variation. More interestingly, a single QTL mapping to the highly polymorphic chromosome VII appears to be involved in control of all the productivity traits studied. Quality QTLs appear to be scattered across the genome and to have less effect on the variation of the corresponding traits. Moreover, some of the new hybrid strains constructed in the course of our experiments had production or quality values higher than those of the parents or other commercial strains. This approach opens the possibility of marker-assisted selection and breeding of new industrial strains of this fungus.     

A radioimmunoprecipitation method in the serodiagnosis of HIV infection: the development of the optimal conditions for its performance and the evaluation of the possibilities for its use

The optimum conditions for using the method of radioimmunoprecipitation (RIP) for the detection of human immunodeficiency virus (HIV) in serum samples have been established. Out of several available cell lines persistently infected with HIV, specially selected line 17 has been chosen. The characteristic feature of this is the high and stable (under the conditions of prolonged cultivation) accumulation of virus-specific proteins in infected cells. The optimum conditions for making the test and its evaluation have also been established. The data of literature on the advantages of the method of RIP over such traditional methods as the enzyme immunoassay and immunoblotting have been confirmed. Thus, the presence of specific antibodies in several serum samples registered as false negative has been established. The intertypical reactivity of two serotypes of the virus, HIV-1 and HIV-2, has been studied. Cross reactivity of antibodies with respect to the HIV gene gag, but not with respect to viral glycoproteids, has been established. Ideas on the expediency and prospects of using RIP for the serological control of HIV infection are presented.     

Differentially Regulated, Vegetative-Mycelium-Specific Hydrophobins of the Edible Basidiomycete Pleurotus ostreatus

Three different hydrophobins (Vmh1, Vmh2, and Vmh3) were isolated from monokaryotic and dikaryotic vegetative cultures of the edible fungus Pleurotus ostreatus. Their corresponding genes have a number of introns different from those of other P. ostreatus hydrophobins previously described. Two genes (vmh1 and vmh2) were expressed only at the vegetative stage, whereas vmh3 expression was also found in the fruit bodies. Furthermore, the expression of the three hydrophobins varied significantly with culture time and nutritional conditions. The three genes were mapped in the genomic linkage map of P. ostreatus, and evidence is presented for the allelic nature of vmh2 and POH3 and for the different locations of the genes coding for the glycosylated hydrophobins Vmh3 and POH2. The glycosylated nature of Vmh3 and its expression during vegetative growth and in fruit bodies suggest that it should play a role in development similar to that proposed for SC3 in Schizophyllum commune.     

Molecular karyotype of the white rot fungus Pleurotus ostreatus.

The white rot fungus Pleurotus ostreatus is an edible basidiomycete with increasing agricultural and biotechnological importance. Genetic manipulation and breeding of this organism are restricted because of the lack of knowledge about its genomic structure. In this study, we analyzed the genomic constitution of P. ostreatus by using pulsed-field gel electrophoresis optimized for the separation of its chromosomes. We have determined that it contains 11 pairs of chromosomes with sizes ranging from 1.4 to 4.7 Mbp. In addition to chromosome separation, the use of single-copy DNA probes allowed us to resolve the ambiguities caused by chromosome comigration. When the two nuclei present in the dikaryon were separated by protoplasting, analysis of their karyotypes revealed length polymorphisms affecting various chromosomes. This is, to our knowledge, the clearest chromosome separation available for this species.