Anatomy of leaf apical hydathodes in four monocotyledon plants of economic and academic relevance.

Hydathode is a plant organ responsible for guttation in vascular plants, i.e. the release of droplets at leaf margin or surface. Because this organ connects the plant vasculature to the external environment, it is also a known entry site for several vascular pathogens. In this study, we present a detailed microscopic examination of leaf apical hydathodes in monocots for three crops (maize, rice and sugarcane) and the model plant Brachypodium distachyon. Our study highlights both similarities and specificities of those epithemal hydathodes. These observations will serve as a foundation for future studies on the physiology and the immunity of hydathodes in monocots.

Drought-induced alterations in photosynthetic, ultrastructural and biochemical traits of contrasting sugarcane genotypes.

Drought is an important factor which limits growth of sugarcane. To elucidate the physiological and biochemical mechanisms of tolerance, a pot experiment was conducted at Sugarcane Research Institute, Kaiyuan, China. Two genotypes (Yuetang 93-159-sensitive and Yunzhe 05-51-tolerant), were subjected to three treatments; 70±5% (control), 50±5% (moderate drought) and 30±5% (severe drought) of soil field capacity. The results demonstrated that drought induced considerable decline in morpho-physiological, biochemical and anatomical parameters of both genotypes, with more pronounced detrimental effects on Yuetang 93-159 than on Yunzhe 05-51. Yunzhe 05-51 exhibited more tolerance by showing higher dry biomass, photosynthesis and antioxidant enzyme activities. Compared with Yuetang 93-159, Yunzhe 05-51 exhibited higher soluble sugar, soluble protein and proline contents under stress. Yunzhe 05-51 illustrated comparatively well-composed chloroplast structure under drought stress. It is concluded that the tolerance of Yunzhe 05-51 was attributed to improved antioxidant activities, osmolyte accumulation and enhanced photosynthesis. These findings may provide valuable information for future studies on molecular mechanism of tolerance.

Methyltrioctylammonium chloride mediated removal of lignin from sugarcane bagasse for themostable cellulase production.

The present study was aimed to evaluate the Methyltrioctylammonium Chloride (IL) and Sodium hydroxide effect on sugarcane bagasse (SB) structure and its subsequent utilization to produce cellulase from a thermophilic bacterium Bacillus aestuarii UE25. The strain was isolated from a crocodile pond of Manghopir, Karachi. Ten different factors affecting IL pretreatment of SB and cellulase production by UE25 were evaluated by Plackett-Burman design and three significant factors were optimized by employing Box Behnken design. Under optimum conditions, the strain produced 118.4 IU mL-1 of EG and 75.01 IU mL-1 of BGL that corroborated well with the predicted values by the model. Scanning electron microscopy, gravimetric analysis, Fourier transform infrared spectroscopy and NMR of SB revealed removal of lignin, decrease in cellulose content and structural changes in the SB after pretreatment and fermentation. The data provide prospects of utilizing this IL in comparison to imidazolium based IL for pretreatment of biomass.

Crosstalk between sugarcane and a plant-growth promoting Burkholderia species.

Bacterial species in the plant-beneficial-environmental clade of Burkholderia represent a substantial component of rhizosphere microbes in many plant species. To better understand the molecular mechanisms of the interaction, we combined functional studies with high-resolution dual transcriptome analysis of sugarcane and root-associated diazotrophic Burkholderia strain Q208. We show that Burkholderia Q208 forms a biofilm at the root surface and suppresses the virulence factors that typically trigger immune response in plants. Up-regulation of bd-type cytochromes in Burkholderia Q208 suggests an increased energy production and creates the microaerobic conditions suitable for BNF. In this environment, a series of metabolic pathways are activated in Burkholderia Q208 implicated in oxalotrophy, microaerobic respiration, and formation of PHB granules, enabling energy production under microaerobic conditions. In the plant, genes involved in hypoxia survival are up-regulated and through increased ethylene production, larger aerenchyma is produced in roots which in turn facilitates diffusion of oxygen within the cortex. The detected changes in gene expression, physiology and morphology in the partnership are evidence of a sophisticated interplay between sugarcane and a plant-growth promoting Burkholderia species that advance our understanding of the mutually beneficial processes occurring in the rhizosphere.

Structural characterisation of pretreated solids from flow-through liquid hot water treatment of sugarcane bagasse in a fixed-bed reactor.

Untreated sugarcane bagasse and sugarcane bagasse pretreated with flow-through liquid hot water (LHW) treatment (170-207°C and 204-250 ml/min) in a fixed-bed reactor have been structurally characterised. Field emission gun scanning electron microscopy (FEG-SEM) and transmission electron microscopy (TEM) were used to investigate changes in the residues, in particular due to the fate of lignin. FEG-SEM results show that the LHW treatment modified the surface morphology of the pretreated bagasse with lignin droplets being observed on the fibre surface. TEM showed an increase in the plant cell wall porosity and lignin migration across the plant cell wall. Increases in pretreatment temperature were observed to increase the average size and density of lignin droplets on the fibre surface. The results provide evidence that for LHW flow-through treatment, just as for batch treatment, lignin repolymerisation and deposition on the surface of pretreated sugarcane bagasse is an important consideration.

Carbonization on combustion and biodegradation of agricultural waste as a possible source of silica.

Agricultural waste being the major solid waste in the environment, the study has explored and identified the presence of minerals especially silica in the agricultural waste like sugarcane bagasse ash and rice husk ash by carbonization on combustion at different thermal conditions and biodegradation. Presence of silica in the ash samples has been well characterized by the XRD, FT-IR, EDX, SEM and N2 sorption techniques. Presence of crystal phases of silica like quartz and cristoballite is well indexed by the X-ray diffraction peaks that appeared at 2θ = 27, 40 and 60 which is further confirmed by the peaks at 1100, 820 and 620 cm(-1) of FT-IR. The elemental composition of the silica in ash is determined by EDX analysis. The exothermic reaction and the mass loss observed in the TG/DTG at the transient temperature of 840-850 °C has confirmed the presence of the α-quartz. SEM micrograph has also supported the presence of silica and has revealed the various crystal shapes that were present in the sugarcane and husk ash. The study has clearly revealed that the silica content has increased with the increase in temperature and refinement of the combustion condition to a maximum of 18.7-52 % and on biodegradation to about 48.3-92.4 %.

A novel surfactant-assisted ionic liquid pretreatment of sugarcane bagasse for enhanced enzymatic hydrolysis.

This study investigated a novel pretreatment method, as an essential step, for production of second generation bioethanol from sugarcane bagasse (SCB). Effect of tween 80 (TW) and polyethylene glycol 4000 (PEG) on SCB pretreatment was assessed using 1-butyl-3-methyl imidazolium chloride ([BMIM]Cl) as an ionic liquid (IL). Different concentrations of TW and PEG were used to determine the optimum concentration of surfactant for the highest percentage of cellulose conversion. TW and PEG increased lignin removal by 12.5% over the IL-only pretreated sample. The 3% (w/w) PEG showed a significant increase in enzymatic digestibility with an efficiency of 96.2% after 12h of hydrolysis; this was 23% higher than the efficiency of SCB pretreated with IL. The increase in digestibility of surfactant assisted IL pretreatment method can be attributed to the decrease in cellulose crystallinity, changes in the cellulose lattice, and delignification; which was confirmed by FT-IR, XRD and FE-SEM analysis.

Simultaneous production of cellulase and reducing sugar through modification of compositional and structural characteristic of sugarcane bagasse.

This study examined the potential of untreated and alkali-pretreated sugarcane bagasse (SCB) in cellulase, reducing sugar (RS) and fungal biomass production via solid state fermentation (SSF) using Pycnoporus sanguineus. The impact of the composition, structure and cellulase adsorption ability of SCB on the production of cellulase, RS and fungal biomass was investigated. From the morphological and compositional analyses, untreated SCB has relatively more structural changes with a higher percentage of depolymerisation on the cellulose, hemicellulose and lignin content compared to alkali-pretreated SCB. Thus, untreated SCB favoured the production of cellulase and fungal biomass whereas alkali-pretreated SCB yielded a higher amount of RS. The composition and morphology of untreated SCB did not encourage RS production and this suggested that RS produced during SSF might be consumed in a faster rate by the more abundantly grown fungus. Besides that, alkali-pretreated SCB with higher cellulase adsorption ability could have adsorbed the cellulase produced and resulted in a lower cellulase titre. In short, the production of specific bioproducts via SSF is dependent on the structure and composition of the substrate applied.

Tissue-specific cell wall hydration in sugarcane stalks.

Plant cell walls contain water, especially under biological and wet processing conditions. The present work characterizes this water in tissues of sugarcane stalks. Environmental scanning electron microscopy shows tissue deformation upon drying. Dynamic vapor sorption determines the equilibrium and kinetics of moisture uptake. Thermoporometry by differential scanning calorimetry quantifies water in nanoscale pores. Results show that cell walls from top internodes of stalks are more deformable, slightly more sorptive to moisture, and substantially more porous. These differences of top internode are attributed to less lignified walls, which is confirmed by lower infrared spectral signal from aromatics. Furthermore, cell wall nanoscale porosity, an architectural and not directly compositional characteristic, is shown to be tissue-specific. Nanoscale porosities are ranked as follows: pith parenchyma > pith vascular bundles > rind. This ranking coincides with wall reactivity and digestibility in grasses, suggesting that nanoscale porosity is a major determinant of wall recalcitrance.

Sugarcane cell wall structure and lignin distribution investigated by confocal and electron microscopy.

Lignocellulosic plant cell wall is considered a potential source for second generation biofuels. The plant cell wall is a highly complex structure mainly composed of cellulose, hemicelluloses, and lignin that form a network of crosslinked fibers. The structural organization of the sugarcane cell wall has not been previously analyzed in detail, and this analysis is a prerequisite for further studies on the recalcitrance and deconstruction of its biomass. In this work, cellulose and lignin localization were investigated by confocal laser scanning microscopy. In addition, the internode sugarcane cell wall structural organization was analyzed by electron microscopy. Internode stem anatomy showed a typical monocot structure consisting of epidermis, hypoderm, and vascular bundles scattered throughout ground parenchyma tissue and surrounded by sclerenchyma fibers. Confocal images of safranin labeled sugarcane showed that lignin distribution was predominant in the vessel elements, cell wall corners (CC), and middle lamella (ML), while cellulose-rich cell walls were randomly distributed in the ML and organized in the other cell wall layers. KMnO4 cytochemistry revealed that lignin was predominantly distributed in secondary cell walls, ML and CC. Cell wall sublayers (S1, S2, and S3) were identified and measured by transmission electron microscopy. Our results provide insights that may help further understanding of sugarcane cell wall organization, which is crucial for the research and technology of plant-based biofuel production.

Association of wet disk milling and ozonolysis as pretreatment for enzymatic saccharification of sugarcane bagasse and straw.

Ozonolysis was studied separately and in combination with wet disk milling (WDM) for the pretreatment of sugarcane bagasse and straw, with the aim of improving their enzymatic saccharification. The glucose yields for ozonolysis followed by WDM were 89.7% for bagasse and 63.1% for straw, whereas the use of WDM followed by ozonolysis resulted in glucose yields of 81.1% for bagasse and 92.4% for straw, with shorter WDM time. This last procedure allowed a substantial decrease in energy consumption in comparison to the use of WDM alone or of ozonolysis followed by WDM. Higher overall saccharification yields with shorter milling times were observed when ozonolysis was carried out before WDM. This effect might be related to the higher specific surface area. Additionally, a finer morphology was observed by the association of the two treatments in comparison to the sole use of ozonolysis or WDM.

Bioconversion of novel substrate Saccharum spontaneum, a weedy material, into ethanol by Pichia stipitis NCIM3498.

Rising fuel prices and environmental issues have paved the way for the exploration of cellulosic ethanol. However, challenges involving substrate hydrolysis and cost-effectiveness still limit the efficient bioconversion and utilization of cellulosic ethanol. We aimed to evaluate a cheaper and abundantly available wild sugarcane variety, Saccharum spontaneum, as the raw substrate for bioconversion of ethanol by Pichia stipitis NCIM3498. Three different strategies for substrate hydrolysis using acid (dilute sulfuric acid) and alkali (dilute sodium hydroxide) and aqueous ammonia (AA) treatment followed by enzymatic hydrolysis were studied. A maximum of 631.5±3.25 mg/g sugars with 89.38% hydrolytic efficiency (HE) could be achieved after enzymatic hydrolysis of AA-pretreated S. spontaneum. All the substrate hydrolysates were evaluated for ethanol conversion in batches by P. stipitis. The microbial fermentation of released sugars into ethanol showed (g/g) 0.36±0.011, 0.384±0.022, 0.391±0.02, and 0.40±0.01 yield from detoxified acid hydrolysate and acid-, NaOH- and AA-pretreated substrate S. spontaneum enzymatic hydrolysates, respectively.