Integrated bioinformatics, modelling, and gene expression analysis of the putative pentose transporter from Candida tropicalis during xylose fermentation with and without glucose addition.

The transport of substrates across the cell membrane plays an essential role in nutrient assimilation by yeasts. The establishment of an efficient microbial cell factory, based on the maximum use of available carbon sources, can generate new technologies that allow the full use of lignocellulosic constituents. These technologies are of interest because they could promote the formation of added-value products with economic feasibility. In silico analyses were performed to investigate gene sequences capable of encoding xylose transporter proteins in the Candida tropicalis genome. The current study identified 11 putative transport proteins that have not yet been functionally characterized. A phylogenetic tree highlighted the potential C. tropicalis xylose-transporter proteins CtXUT1, CtXUT4, CtSTL1, CtSTL2, and CtGXT2, which were homologous to previously characterized and reported xylose transporters. Their expression was quantified through real-time qPCR at defined times, determined through a kinetic analysis of the microbial growth curve in the absence/presence of glucose supplemented with xylose as the main carbon source. The results indicated different mRNA expression levels for each gene. CtXUT1 mRNA expression was only found in the absence of glucose in the medium. Maximum CtXUT1 expression was observed in intervals of the highest xylose consumption (21 to 36 h) that corresponded to consumption rates of 1.02 and 0.82 g/L/h in the formulated media, with xylose as the only carbon source and with glucose addition. These observations indicate that CtXUT1 is an important xylose transporter in C. tropicalis. KEY POINTS: • Putative xylose transporter proteins were identified in Candida tropicalis; • The glucose concentration in the cultivation medium plays a key role in xylose transporter regulation; • The transporter gene CtXUT1 has an important role in xylose consumption by Candida tropicalis.

The Secretome of Phanerochaete chrysosporium and Trametes versicolor Grown in Microcrystalline Cellulose and Use of the Enzymes for Hydrolysis of Lignocellulosic Materials.

The ability of white-rot fungi to degrade polysaccharides in lignified plant cell walls makes them a suitable reservoir for CAZyme prospects. However, to date, CAZymes from these species are barely studied, which limits their use in the set of choices for biomass conversion in modern biorefineries. The current work joined secretome studies of two representative white-rot fungi, Phanerochaete chrysosporium and Trametes versicolor, with expression analysis of cellobiohydrolase (CBH) genes, and use of the secretomes to evaluate enzymatic conversion of simple and complex sugarcane-derived substrates. Avicel was used to induce secretion of high levels of CBHs in the extracellular medium. A total of 56 and 58 proteins were identified in cultures of P. chrysosporium and T. versicolor, respectively, with 78-86% of these proteins corresponding to plant cell wall degrading enzymes (cellulolytic, hemicellulolytic, pectinolytic, esterase, and auxiliary activity). CBHI predominated among the plant cell wall degrading enzymes, corresponding to 47 and 34% of the detected proteins in P. chrysosporium and T. versicolor, respectively, which confirms that Avicel is an efficient CBH inducer in white-rot fungi. The induction by Avicel of genes encoding CBHs (cel) was supported by high expression levels of cel7D and cel7C in P. chrysosporium and T. versicolor, respectively. Both white-rot fungi secretomes enabled hydrolysis experiments at 10 FPU/g substrate, despite the varied proportions of CBHs and other enzymes present in each case. When low recalcitrance sugarcane pith was used as a substrate, P. chrysosporium and T. versicolor secretomes performed similarly to Cellic® CTec2. However, the white-rot fungi secretomes were less efficient than Cellic® CTec2 during hydrolysis of more recalcitrant substrates, such as acid or alkaline sulfite-pretreated sugarcane bagasse, likely because Cellic® CTec2 contains an excess of CBHs compared with the white-rot fungi secretomes. General comparison of the white-rot fungi secretomes highlighted T. versicolor enzymes for providing high glucan conversions, even at lower proportion of CBHs, probably because the other enzymes present in this secretome and CBHs lacking carbohydrate-binding modules compensate for problems associated with unproductive binding to lignin.

Function and diversity of P0 proteins among cotton leafroll dwarf virus isolates.

BACKGROUND: The RNA silencing pathway is an important anti-viral defense mechanism in plants. As a counter defense, some members of the viral family Luteoviridae are able to evade host immunity by encoding the P0 RNA silencing suppressor protein. Here we explored the functional diversity of P0 proteins among eight cotton leafroll dwarf virus (CLRDV) isolates, a virus associated with a worldwide cotton disease known as cotton blue disease (CBD). METHODS: CLRDV-infected cotton plants of different varieties were collected from five growing fields in Brazil and their P0 sequences compared to three previously obtained isolates. P0's silencing suppression activities were scored based on transient expression experiments in Nicotiana benthamiana leaves. RESULTS: High sequence diversity was observed among CLRDV P0 proteins, indicating that some isolates found in cotton varieties formerly resistant to CLRDV should be regarded as new genotypes within the species. All tested proteins were able to suppress local and systemic silencing, but with significantly variable degrees. All P0 proteins were able to mediate the decay of ARGONAUTE proteins, a key component of the RNA silencing machinery. CONCLUSIONS: The sequence diversity observed in CLRDV P0s is also reflected in their silencing suppression capabilities. However, the strength of local and systemic silencing suppression was not correlated for some proteins.

Postharvest conservation of the tuberous roots of Pachyrhizus Ahipa (Wedd) Parodi.

This paper aimed to evaluate the effects of storage periods on the conservation of Pachyrhizus ahipa roots at different temperatures and packaging materials. The roots were harvested, washed, packed in PVC, plastic bags, without wrappings (control) and stored in polystyrene trays in refrigerators, or cold chambers, or at room temperature. Total titratable acidity (TTA), total soluble solids (TSS), pH, as well as their ash, lipid, total carbohydrate and protein (dry basis) contents were analyzed. The lowest loss of root fresh weight was observed in the cold chamber and plastic bags. The TTA remained higher among roots stored in the cold chamber and in PVC packaging. The lowest TSS contents were observed for roots stored in the cold chamber, and these did not vary among the packing materials. The average carbohydrate content percentage for all treatments was 84.9%. The percentage of lipids was highest in roots stored at room temperature while protein and ash contents were highest in roots under refrigeration. The best storage conditions for roots are plastic bags packaging in a cold chamber, with the roots retaining appropriate quality for commercialization for up to 30 days.

Degradation of caffeine by photo-Fenton process: optimization of treatment conditions using experimental design.

The degradation of caffeine in different kind of effluents, via photo-Fenton process, was investigated in lab-scale and in a solar pilot plant. The treatment conditions (caffeine, Fe(2+) and H(2)O(2) concentrations) were defined by experimental design. The optimized conditions for each variable, obtained using the response factor (% mineralization), were: 52.0 mg L(-1)caffeine, 10.0 mg L(-1)Fe(2+) and 42.0 mg L(-1)H(2)O(2) (replaced in kinetic experiments). Under these conditions, in ultrapure water (UW), the caffeine concentration reached the quantitation limit (0.76 mg L(-1)) after 20 min, and 78% of mineralization was obtained respectively after 120 min of reaction. Using the same conditions, the matrix influence (surface water - SW and sewage treatment plant effluent - STP) on caffeine degradation was also evaluated. The total removal of caffeine in SW was reached at the same time in UW (after 20 min), while 40 min were necessary in STP. Although lower mineralization rates were verified for high organic load, under the same operational conditions, less H(2)O(2) was necessary to mineralize the dissolved organic carbon as the initial organic load increases. A high efficiency of the photo-Fenton process was also observed in caffeine degradation by solar photocatalysis using a CPC reactor, as well as intermediates of low toxicity, demonstrating that photo-Fenton process can be a viable alternative for caffeine removal in wastewater.

Global alteration of microRNAs and transposon-derived small RNAs in cotton (Gossypium hirsutum) during Cotton leafroll dwarf polerovirus (CLRDV) infection.

Small RNAs (sRNAs) are a class of non-coding RNAs ranging from 20- to 40-nucleotides (nts) that are present in most eukaryotic organisms. In plants, sRNAs are involved in the regulation of development, the maintenance of genome stability and the antiviral response. Viruses, however, can interfere with and exploit the silencing-based regulatory networks, causing the deregulation of sRNAs, including small interfering RNAs (siRNAs) and microRNAs (miRNAs). To understand the impact of viral infection on the plant sRNA pathway, we deep sequenced the sRNAs in cotton leaves infected with Cotton leafroll dwarf virus (CLRDV), which is a member of the economically important virus family Luteoviridae. A total of 60 putative conserved cotton miRNAs were identified, including 19 new miRNA families that had not been previously described in cotton. Some of these miRNAs were clearly misregulated during viral infection, and their possible role in symptom development and disease progression is discussed. Furthermore, we found that the 24-nt heterochromatin-associated siRNAs were quantitatively and qualitatively altered in the infected plant, leading to the reactivation of at least one cotton transposable element. This is the first study to explore the global alterations of sRNAs in virus-infected cotton plants. Our results indicate that some CLRDV-induced symptoms may be correlated with the deregulation of miRNA and/or epigenetic networks.

Detection of paramyxoviruses in Magellanic penguins (Spheniscus magellanicus) on the Brazilian tropical coast.

Aquatic migratory birds are a major vectors by which influenza viruses and paramyxoviruses are spread in nature. Magellanic penguins (Spheniscus magellanicus) are usually present on the southern shores of South America and can swim as far as the southern coast of Brazil in winter. In 2008, however, several Magellanic penguins were observed on the northeastern coast of Brazil. Paramyxoviruses were isolated from Magellanic penguins on the Espírito Santo state coast, approximately 4000 km from their breeding colonies, although influenza viruses were not detected. Among the paramyxoviruses, five Avulavirus isolates belonging to serotype APMV-2 and the serotype APMV-10, which was proposed by Miller et al. (2010), were identified. These results highlight the risks associated with the spread of paramyxoviruses between natural to non-natural habitats by birds exhibiting unusual migration patterns, and they document for the first time the presence of the APMV-2 and APMV-10 serotypes on penguins in Brazil. The local avifauna may become infected with these viruses through close contact between migratory and resident birds. Continued surveillance of virus incidence in these migratory populations of penguins is necessary to detect and prevent the potential risks associated with these unusual migration patterns.

Profile of small interfering RNAs from cotton plants infected with the polerovirus Cotton leafroll dwarf virus.

BACKGROUND: In response to infection, viral genomes are processed by Dicer-like (DCL) ribonuclease proteins into viral small RNAs (vsRNAs) of discrete sizes. vsRNAs are then used as guides for silencing the viral genome. The profile of vsRNAs produced during the infection process has been extensively studied for some groups of viruses. However, nothing is known about the vsRNAs produced during infections of members of the economically important family Luteoviridae, a group of phloem-restricted viruses. Here, we report the characterization of a population of vsRNAs from cotton plants infected with Cotton leafroll dwarf virus (CLRDV), a member of the genus Polerovirus, family Luteoviridae. RESULTS: Deep sequencing of small RNAs (sRNAs) from leaves of CLRDV-infected cotton plants revealed that the vsRNAs were 21- to 24-nucleotides (nt) long and that their sequences matched the viral genome, with higher frequencies of matches in the 3- region. There were equivalent amounts of sense and antisense vsRNAs, and the 22-nt class of small RNAs was predominant. During infection, cotton Dcl transcripts appeared to be up-regulated, while Dcl2 appeared to be down-regulated. CONCLUSIONS: This is the first report on the profile of sRNAs in a plant infected with a virus from the family Luteoviridae. Our sequence data strongly suggest that virus-derived double-stranded RNA functions as one of the main precursors of vsRNAs. Judging by the profiled size classes, all cotton DCLs might be working to silence the virus. The possible causes for the unexpectedly high accumulation of 22-nt vsRNAs are discussed. CLRDV is the causal agent of Cotton blue disease, which occurs worldwide. Our results are an important contribution for understanding the molecular mechanisms involved in this and related diseases.