Use of the enzyme preparation cellocandin from Geotrichum candidum 3C-106 for difibering of waste paper and dessication of cellulose suspension

The effect of cellocandin, an enzymatic preparation from Getrichum candidum 3C-106 with cellulolytic and xylanolytic activities, on defibring of waste paper and acceleration of desiccation of paper pulp was studied. It was shown that cellocandin accelerated defibring of waste paper to cellulose fibers and desiccation of cellulose suspension.     

Enzymatic treatment of primary municipal sludge with Trichoderma viride cellulase

Enzymatic hydrolysis of the cellulose in raw primary settled municipal sludge by Trichoderma viride cellulase achieved conversions of up to 75% of the cellulose, primarily to cellobiose, in 24 hr. Simultaneously the gel-like characteristic of raw primary sludge was changed to that of a slurry of fine particles in less than 2 hr, causing a radical change in the ability to ultrafilter the sludge. The use of raw primary sludge as a growth medium for T. viride cellulase production was also investigated. It was possible, with nitrogen supplements, to obtain an enzyme with a filter paper activity (FPA) of two compared to over four which is attainable on defined medium of similar strength. The potential use of cellulase treatment as a pretreatment or integral part of waste treatment processes is discussed and the alternatives evaluated.     

Isolation of cellulolytic mesophilic clostridia from a municipal solid waste digestor

Ten obligately anaerobic, cellulolytic mesophilic bacteria were isolated from a municipal solid waste digestor used for biogas production. The isolates were rod-shaped, spore-forming bacteria in anaerobic conditions, and stained Gram-positive in young cultures, and hence were identified asClostridium. Small regular translucent and unpigmented colonies were observed on cellulose plates. The strains were gelatinase-negative, hydrolyzed esculin and starch, and fermented xylose and arabinose. The lecithinase, lipase, and indole tests were negative. The major fermentation products from cellulose included ethanol and acetate. The morphological and other biochemical characteristics indicated that these clostridia did not correspond to any previously described species. All the strains produced high activities of extracellular cellulases in cellulose media and degraded paper. Offprint requests to: L. Benoit.     

Desiccation and egg viability of the ragwort flea beetle,Longitarsus flavicornis (Coleoptera: Chrysomelidae)

A study was conducted to investigate the effect of desiccation on the survival of eggs of Longitarsus flavicornis. Eclosion of L. flavicornis eggs in laboratory trials decreased with increasing desiccation time between 0 days (93% hatching) and 42 days (no egg hatching) at 50±2% relative humidity and 23±2°C. Probit analysis indicated that 25, 50 and 99% mortality of L. flavicornis eggs occurred after 5.7, 9.3 and 50.4 days desiccation, respectively. Egg development varied between a minimum of 8 days at 7 days desiccation to a maximum of 15 days at 28 days desiccation. Hatching span did not differ between treatments with all eggs hatching within 12 days of each other. A relative humidity of 88–100% was measured under ragwort rosettes in non-drought field conditions. This would be expected to facilitate successful egg eclosion. However, the occurrence of summer drought could be detrimental to egg survival.     

Freezing and desiccation injury resistance in the filamentous green alga Klebsormidium from the Antarctic, Arctic and Slovakia

The freezing and desiccation tolerance of 12 Klebsormidium strains, isolated from various habitats (aeroterrestrial, terrestrial, and hydro-terrestrial) from distinct geographical regions (Antarctic — South Shetlands, King George Island, Arctic — Ellesmere Island, Svalbard, Central Europe — Slovakia) were studied. Each strain was exposed to several freezing (−4°C, −40°C, −196°C) and desiccation (+4°C and + 20°C) regimes, simulating both natural and semi-natural freeze-thaw and desiccation cycles. The level of resistance (or the survival capacity) was evaluated by chlorophyll a content, viability, and chlorophyll fluorescence evaluations. No statistical differences (Kruskal-Wallis tests) between strains originating from different regions were observed. All strains tested were highly resistant to both freezing and desiccation injuries. Freezing down to −196°C was the most harmful regime for all studied strains. Freezing at −4°C did not influence the survival of studied strains. Further, freezing down to −40°C (at a speed of 4°C/min) was not fatal for most of the strains. RDA analysis showed that certain Antarctic and Arctic strains did not survive desiccation at +4°C; however, freezing at −40°C, as well as desiccation at +20°C was not fatal to them. On the other hand, other strains from the Antarctic, the Arctic, and Central Europe (Slovakia) survived desiccation at temperatures of +4°C, and freezing down to −40°C. It appears that species of Klebsormidium which occupy an environment where both seasonal and diurnal variations of water availability prevail, are well adapted to freezing and desiccation injuries. Freezing and desiccation tolerance is not species-specific nor is the resilience only found in polar strains as it is also a feature of temperate strains. Presented at the International Symposium Biology and Taxonomy of Green Algae V, Smolenice, June 26–29, 2007, Slovakia. This paper is dedicated to the memory of the late Dr. Bohuslav Fott (1908–1976), Professor of Botany at the Charles University in Prague, to mark the centenary of his birth.     

Effects of cellulose on growth,enzyme production,and ultrastructure of a Cellulomonas species

Cellulomonas sp. (NRCC 2406) was grown on complex medium (peptone-tryptone-yeast extract) alone, or with the addition of different celluloses (solka floc, avicel, CF 11 cellulose or Whatman No. 1 filter paper) and/or glucose. Cultures growing on the complex medium without cellulose produced low levels of endo- and exo-cellulases and very little -glucosidase. Adding cellulose stimulated growth, as measured by cellular protein or by viable counts, and also stimulated production of cellulases. Adding glucose in the prescene of cellulose inhibited growth and cellulose breakdown. Glucose also inhibited attachment of growing cells to cellulose fibres. Electron microscope studies showed that Cellulomonas sp. adhered to the cellulose fibers. In the presence of cellulose in the media, the cells developed a thicker outer layer which probably helps in the adhesion process.Abbreviations PTYE peptone, tryptone, yeast extract medium - DNS dinitrosalicylic acid - CMC carboxymethyl cellulose - cfu/ml colony-forming units per ml     

In vitro desiccation tolerance of the epiphytic Ghost Orchid,Dendrophylax lindenii (Lindl.) Benth x. Rolfe

The Ghost Orchid, Dendrophylax lindenii (Lindl.) Benth x. Rolfe, is a rare and endangered epiphytic orchid native to south Florida and Cuba. The orchid is considered difficult to propagate under greenhouse conditions, requiring high humidity and low air movement. In contrast, the orchid’s native habitat seasonally dries out with decreased precipitation and humidity. This suggests some level of desiccation tolerance. Ghost Orchid plants were assessed for potential desiccation tolerance and ability to recover from desiccation stress under in vitro conditions. In vitro-derived plants were placed into sterile baby food jars and transferred to chambers maintained at 10% relative humidity, which is extremely low compared to relative humidity levels (40–100%) recorded under natural field conditions. Plants were removed every week for 4 wk and recovered on P723 medium supplemented with banana powder for 4 wk. Data were collected at the initiation of the experiment, after the desiccation periods, and after 4 wk in vitro recovery. Ghost Orchid plants demonstrated extremely high desiccation tolerance. Even after 4 wk desiccation, plant survival was observed at 79.2% after recovery. Desiccated plants exhibited significant decreases in tissue water potential (− 18.44 MPa), fresh weight (65.5% loss), and water content (14.2%); however, high plant survival was still observed under these conditions similar to poikilohydric plants. Overall, the Ghost Orchid demonstrated high desiccation tolerance, which should be considered for future greenhouse culture and for its application in the direct field establishment of in vitro-derived plants without greenhouse acclimatization.

     

Cellulase production by six <Emphasis Type="Italic">Trichoderma</Emphasis> spp. fermented on medicinal plant processings

Capabilities of cellulase production, using delignified bioprocessings of medicinal and aromatic plants, viz. citronella (Cymbopogon winterianus) and Artemisia annua (known as marc of Artemisia) and garden waste (chiefly containing Cynodon dactylon), by the six species of Trichoderma were comparatively evaluated. Among the members of Trichoderma studied, T. citrinoviride was found to be the most efficient producer of cellulases along with a high level of β- glucosidase (produced 102.4 IU g⁻¹ on marc of Artemisia; 101.33 IU g⁻¹ on garden waste; 81.86 IU g⁻¹ on distillation waste of citronella and 94.77 IU g⁻¹ on pure cellulose). Although T. virens was noticed to be the minimal enzyme producer fungus, it interestingly could not produce complete cellulase enzyme complex on any test waste or pure cellulose, except on marc of Artemisia, where it produced all three enzymes of the complex. Immediate reduction in pH was also noticed during fermentation in the case of pure polymer (cellulose) by all tested fungi, while it was delayed with delignified agrowastes. The pH profile varied with the substrate used as well as with individual species of Trichoderma. On the other hand, no alteration in pH with any species of Trichoderma was noticed when grown on marc of A. annua, which might be due to the buffering capacity of this marc.     

Desiccation survival of the eggs of the rotifer Adleta vaga (Davis, 1873)

Age at the time of drying affects the desiccation survival of the embryos of the bdelloid rotifer Adineta vaga. Although the embryos younger than 24 hours do not survive desiccation, up to 71% and 6% of the embryos at least 45 hours old survive desiccation for 2 and 10 days, respectively.     

Production of cellulase by immobilized whole cells of Haloarcula

Halophilic Archaea are adapted to a life in the extreme conditions and some of them are capable of growth on cellulosic waste as carbon and energy source by producing cellulase enzyme. The production of cellulase using free and immobilized cells of halophilic archaeal strain Haloarcula 2TK2 isolated from Tuzkoy Salt Mine and capable of producing cellulose was studied. The cells were cultured in a liquid medium containing 2.5 M NaCl to obtain the maximum cellulase activity and immobilized on agarose or polyacrylamide or alginate. Optimal salt dependence of free and immobilized cells of Haloarcula 2TK2 was established and the effects of pH and temperature were investigated. Immobilization to Na-alginate enhanced the enzymatic activity of the Haloarchaeal cells when compared to free cells and other polymeric supports. From the results obtained it is reasonable to infer that decomposition of plant polymers into simpler end products does occur at high salinities and cellulase producing haloarchael cells may be potentially utilized for the treatment of hypersaline waste water to remove cellulose.     

Endoglucanase Activity of Compost-Dwelling Fungus Paecilomyces inflatus is Stimulated by Humic Acids and Other Low Molecular Mass Aromatics

Summary Paecilomyces inflatus isolated from municipal waste compost was found to have cellulolytic activity in several solid and liquid media. This study was done to reveal the multifarious effects of municipal waste compost on endoglucanase activity of P. inflatus. The highest enzyme activities under the conditions of solid-state fermentation were measured in authentic compost samples compared with wood, straw and bran substrates. In surface liquid cultures glucose, cellobiose, xylan, Avicel cellulose, carboxymethylcellulose (CM-cellulose), starch and citrus pectin were used as carbon sources. All carbon sources supported the growth of P. inflatus. However, only CM-cellulose, cellobiose and pectin noticeably stimulated endoglucanase (EG) activity. Further stimulation of EG activity was obtained in cultures containing 1% CM-cellulose as a carbon source by supplementation with low-molecular mass aromatic compounds vanillin, veratric acid and benzoic acid, and with soil humic acid (SHA). SHA and veratric acid were found to be the most efficient elicitors of the cellulolytic activity. P. inflatus was able to utilize nitrate and ammonium as pure nitrogen sources in media containing cellulose.     

Energies of freezing and frost desiccation

A stable cellulose paper system was studied to relate water distribution data, as obtained previously from plant tissues, to the analysis of freezing energy. Water distribution data for the cellulose system were obtained by several techniques and were coordinated with calorimetric data. The effect of the cellulose system on the latent heat of freezing was evaluated to estimate activation energies as functions of the amount of associated liquid water. Similar activation energies of water phase transitions in critical plant tissue systems may be heritable characteristics that affect freezing stress. Adhesion energy, that develops between ice and hydrophilic polymer systems as they compete for liquid water in a complex interface, was suggested as one possible source of freezing stress. This does not occur in frost desiccation.     

Resuscitation of viable but non‐culturable bacteria to enhance the cellulose‐degrading capability of bacterial community in composting

Nowadays, much of what we know regarding the isolated cellulolytic bacteria comes from the conventional plate separation techniques. However, the culturability of many bacterial species is controlled by resuscitation‐promoting factors (Rpfs) due to entering a viable but non‐culturable (VBNC) state. Therefore, in this study, Rpf from Micrococcus luteus was added in the culture medium to evaluate its role in bacterial isolation and enhanced effects on cellulose‐degrading capability of bacterial community in the compost. It was found that Proteobacteria and Actinobacteria were two main phyla in the compost sample. The introduction of Rpf could isolate some unique bacterial species. The cellulase activity of enrichment cultures with and without Rpf treatment revealed that Rpf treatment significantly enhanced cellulase activity. Ten isolates unique in Rpf addition displayed carboxymethyl‐cellulase (CMCase) activity, while six isolates possessed filter paper cellulase (FPCase) activity. This study provides new insights into broader cellulose degraders, which could be utilized for enhancing cellulosic waste treatment.     

Dopamine release by Ulvaria obscura (Chlorophyta): environmental triggers and impacts on photosynthesis,growth, and survival of the releaser

In the NE Pacific, Ulvaria obscura is a common component of “green tide” blooms. It is also the only alga known to produce dopamine, which is released into seawater on sunny days when Ulvaria is emersed and then rehydrated. To better understand the mechanisms associated with dopamine release, we experimentally determined whether light quantity and quality, desiccation, temperature, exudates from conspecifics, and dissolved dopamine caused dopamine release. We also examined the effects of desiccation on Ulvaria's ability to photosynthesize, grow, and survive. Desiccation was the only factor that caused significant amounts of dopamine to be lost from U. obscura tissues. The loss of water from Ulvaria tissues was strongly and positively correlated with the loss of dopamine after rehydration. Only 56% of desiccated algae survived for 1 week, compared to 100% of undesiccated control algae. Desiccated algae lost 77% of their pigmented surface area and grew only 15% as much as undesiccated algae, which remained fully pigmented. The oxygen saturation of water containing Ulvaria that was desiccated and then rehydrated was significantly lower than that of seawater containing undesiccated algae. Thus, desiccation, which is coupled with dopamine release, is associated with the deterioration and death of some, but not all, tissues in Ulvaria. Although dopamine released into seawater can reduce the survival or growth of potential competitors, its release is associated with significant physiological stress and tissue mortality. However, the survival and continued growth of some Ulvaria tissues indicates that a net fitness benefit to release dopamine following desiccation cannot be ruled out.     

Effect of various carbon and nitrogen sources on cellulose synthesis by Acetobacter xylinum

The effect of various carbon and nitrogen sources on cellulose membrane production by Acetobacter xylinum was evaluated. Among the carbon sources, sucrose, glucose and mannitol were found to be suitable for optimum levels of cellulose production. The strain was able to utilize a wide range of protein and nitrogen sources such as peptone, soybean meal, glycine, casein hydrolysate, and glutamic acid for cellulose synthesis. Sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDS-PAGE) analysis of pellicle proteins (PP) revealed electrophoretic bands of molecular masses in the range of 116–20 kDa. Furthermore, the strain can be useful for the removal of various nitrogenous and carbon substrates present in waste waters.     

Physiological ecology of photosynthesis in Prasiola stipitata (Trebouxiophyceae) from the Bay of Fundy,Canada

The physiological ecology of Prasiola stipitata was examined in situ from two supralittoral sites in the Bay of Fundy (Nova Scotian, Canada) during November 2011, when the population was undergoing major expansion. Photosynthetic parameters (effective quantum yield, Φ_PSII, maximum quantum yield, F_v/F_m, and relative electron transport rate, rETR) were evaluated using chlorophyll fluorescence of PSII. A largely shaded and continuously moist population showed no change in Φ_PSII from one hour after sunrise to sunset in which natural irradiance varied between 3 and 300 μmol photons m^?2 s^?1. High irradiance (up to 1800 μmol photons m^?2 s^?1) had no apparent negative impacts on either quantum yield or rETR, but high desiccation in the field reduced quantum yield to almost zero. When thalli were brought into the laboratory, no change in F_v/F_m was observed up to 60% dehydration; however, there was a steep decline in F_v/F_m between 60% and 85% dehydration. Thalli showed complete recovery of F_v/F_m within one hour of reimmersion in seawater after 2 days of desiccation. After 15 days of desiccation full recovery required 24 h and after 30 days of desiccation thalli showed only partial recovery. These observations confirm the adaptation to photosynthesis in high irradiances and the rapid recovery following extreme desiccation observed in other Prasiola species.     

Seaweeds early development: detrimental effects of desiccation and attenuation by algal extracts

The effects of desiccation on the early development stages of Mazzaella laminarioides, Scytosiphon lomentaria and Lessonia nigrescens, algal species with different patterns of distribution across the intertidal zone, were examined in the laboratory. In addition, the protective effect against desiccation was evaluated using algal extracts, including those from Porphyra columbina, a macroalga tolerant to desiccation that lives in the uppermost part of the intertidal zone. Our results showed that M. laminarioides displayed the highest resistance to daily desiccation, followed by S. lomentaria, whereas L. nigrescens was the most susceptible. Spores from L. nigrescens exposed to desiccation, although being able to germinate, ceased further post-germination development. In addition, our results showed that all species exposed to extracts from desiccated P. columbina successfully completed their development and strongly suggest the occurrence of compounds with protective properties that help in attenuating the stress caused by desiccation. Finally, our results indicate that the magnitude of the effects generated by desiccation on the early algal development is related to the position of the species in the intertidal zone, and that the protective effects of P. columbina extracts reveal an exceptional metabolism of this species under desiccation stress.     

Isolation of a microbial consortium from activated sludge for the biological treatment of food waste

The bacterial community in the activated sludge of a local wastewater treatment plant was studied in an effort to understand and exploit the metabolic versatility of microorganisms for the efficient biological treatment of food waste. Microorganisms capable of and efficient in degrading domestic food waste were screened based on their ability to produce areas of clearing on selective media containing protein, fat, cellulose and starch. Nine microbial species belonging to the genera Flavobacterium, Pseudomonas, Micrococcus, Aeromonas, Xanthomonas, Vibrio and Sphingomonas were found to degrade all components of food waste. These bacteria were added to domestic wastewater and shown to cause a 60% reduction in the biochemical oxygen demand (BOD) level of wastewater compared to a control in which no microorganisms were added. The ability of the microbial consortium to degrade domestic wastewater as evidenced by the decrease in BOD levels suggests its potential for use in the biological treatment of food waste.     

Acid hydrolysis of sunflower seed husks for production of single cell protein

Summary Sunflower seed husks were chosen as a typical lignocellulosic waste product of low value. This model substrate was hydrolyzed with sulphuric acid at 120°C. The hydrolysis was carried out in two steps: hydrolysis of the pentosan fraction and subsequent hydrolysis of the cellulose fraction. The pentosan fraction was nearly quantitatively hydrolyzed. For the cellulose hydrolysis the yield was 79% of the theoretical yield. The hydrolyzates were neutralized to pH 5 with solid calcium hydroxide and used for preparation of growth media forCandida yeasts andPaecilomyces variotii. For the pentosan hydrolyzates the yields of yeast biomass were 35–36 g per 100 g available reducing sugars (supplied to the medium). In cellulose hydrolyzates the corresponding yields were 45–48 g withCandida utilis andC. tropicalis and about 30 g withC. pseudotropicalis. P. variotii was noticeably superior to the yeasts. In pentosan hydrolyzates it produced 63 g dry mycelium from 100 g reducing sugars supplied; in cellulose hydrolyzates, 94 g. This suggests that it must be an effective utilizer of a wide range of compounds, for example, organic acids in the medium.