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Variation of the C. unicolor and T. versicolor mono- and dual cultures laccase activity in dependence on the mandarin peels’ concentration. MP mandarin peels
Source publication
The present study aimed to identify a pair of fungal strains that promote laccase production in the co-cultivation of white-rot basidiomycetes and to determine the optimum conditions to enhance enzyme synthesis under co-fermentation of mandarin peels. Co-cultivation of Cerrena unicolor with Trametes versicolor, Lenzites betulina, and Panus lecomtei...
Citations
... Collectively, these studies underscore the significant influence of carbon sources on WRF laccase productivity. Despite ongoing efforts by scientists to optimize carbon sources to enhance WRF laccase productivity as a standardized process [28][29][30], the regulatory mechanism underlying these phenomena has received insufficient attention and remains unclear in the long term. In addition to conventional strategies, technological innovations have spurred advancements in genetic and protein engineering [20]. ...
Background
White-rot fungi are known to naturally produce high quantities of laccase, which exhibit commendable stability and catalytic efficiency. However, their laccase production does not meet the demands for industrial-scale applications. To address this limitation, it is crucial to optimize the conditions for laccase production. However, the regulatory mechanisms underlying different conditions remain unclear. This knowledge gap hinders the cost-effective application of laccases.
Results
In this study, we utilized transcriptomic and metabolomic data to investigate a promising laccase producer, Cerrena unicolor 87613, cultivated with fructose as the carbon source. Our comprehensive analysis of differentially expressed genes (DEGs) and differentially abundant metabolites (DAMs) aimed to identify changes in cellular processes that could affect laccase production. As a result, we discovered a complex metabolic network primarily involving carbon metabolism and amino acid metabolism, which exhibited contrasting changes between transcription and metabolic patterns. Within this network, we identified five biomarkers, including succinate, serine, methionine, glutamate and reduced glutathione, that played crucial roles in co-determining laccase production levels.
Conclusions
Our study proposed a complex metabolic network and identified key biomarkers that determine the production level of laccase in the commercially promising Cerrena unicolor 87613. These findings not only shed light on the regulatory mechanisms of carbon sources in laccase production, but also provide a theoretical foundation for enhancing laccase production through strategic reprogramming of metabolic pathways, especially related to the citrate cycle and specific amino acid metabolism.
... Maximal xylanase activity was measured on the 7th day of fermentation with wheat bran (34.63 U/mL) (Ramanjaneyulu et al. 2017;Marinho et al. 2023). Additionally, Kachlishvili et al. (2021) reported a high xylanase activity of 14 200 U/L with T. versicolor 159 cultivated on semi synthetic medium supplemented with mandarin peels. On the other hand, it was shown that the genome of C. hoffmannii does contain 9 genes of endo-1,4-xylanases (Mondo et al. 2019). ...
Nowadays, natural resources like lignocellulosic biomass are gaining more and more attention. This study was conducted to analyse chemical composition of dried and ground samples (500 μm) of various Algerian bioresources including alfa stems (AS), dry palms (DP), olive pomace (OP), pinecones (PC), and tomato waste (TW). AS exhibited the lowest lignin content (3.60 ± 0.60%), but the highest cellulose (58.30 ± 2.06%), and hemicellulose (20.00 ± 3.07%) levels. DP, OP, and PC had around 30% cellulose, and 10% hemicellulose. OP had the highest lignin content (29.00 ± 6.40%), while TW contained (15.70 ± 2.67% cellulose, 13.70 ± 0.002% hemicellulose, and 17.90 ± 4.00% lignin). Among 91 isolated microorganisms, nine were selected for cellulase, xylanase, and/or laccase production. The ability of Bacillus mojavensis to produce laccase and cellulase, as well as B. safensis to produce cellulase and xylanase, is being reported for the first time. In submerged conditions, TW was the most suitable substrate for enzyme production. In this conditions, T. versicolor K1 was the only strain able to produce laccase (4,170 ± 556 U/L). Additionally, Coniocheata hoffmannii P4 exhibited the highest cellulase activity (907.62 ± 26.22 U/L), and B. mojavensis Y3 the highest xylanase activity (612.73 ± 12.73 U/L). T. versicolor K1 culture showed reducing sugars accumulation of 18.87% compared to initial concentrations. Sucrose was the predominant sugar detected by HPLC analysis (13.44 ± 0.02 g/L). Our findings suggest that T. versicolor K1 holds promise for laccase production, while TW represents a suitable substrate for sucrose production.
... Previous studies had indicated excellent laccase secretion ability of Cerrena unicolor (Jamroz et al. 2004;Rola et al. 2013;Kachlishvili et al. 2014Kachlishvili et al. , 2021Pawlik et al. 2021;Zhang et al. 2021). There are also many studies on the effect of lignocellulose materials on laccase activity from C. unicolor. ...
Coriolopsis trogii and Cerrena unicolor were investigated for laccase production in submerged fermentation with different nutrient medium containing metal ions and lignocellulosic materials. The maximum laccase activity of C. trogii Han751 was 8584.44 ± 98.45 U/L and was obtained from nutrient medium 7. However, the maximum laccase activity of C. unicolor Han 849 was 16144.26 ± 635.30 U/L from nutrient medium 9. Thus, the capacity of secreting laccase of C. unicolor Han 849 was superior to that of C. trogii Han751. Different fungal species have different medium components suitable for laccase production. The content of CuSO4·5H2O and MnSO4 in nutrient medium with the concentration of 0.25 g/L and 0.151 g/L, respectively, was more beneficial to C. trogii Han751 secreting laccase. However, the vital components of nutrient medium that contribute to the laccase activity of C. unicolor Han 849 were corncob, glucose, and CuSO4·5H2O, with the corresponding concentrations of 1 g/flask, 5 g/L, and 0.25 g/L, respectively. The results will contribute to the development of new methods to produce low-cost laccase.
... Cerrena unicolor (Cun) is a white-rot basidiomycete and a known producer of several bioactive compounds [17,18]. The fungus is also well-known as a source of miscellaneous enzymes, such as laccases, xylanases, cellulases, oxidases, and, as was recently reported, terpene cyclases [19][20][21][22][23]. In 2018, Cun was discovered to be a producer of volatile aromaactive compounds [24]. ...
... Grape pomace is also rich in oils and phenolic compounds [44] and can be used as a nitrogen source [45]. Citrus peel is rich in flavonoids, d-limonene, and sugars and was already successfully used as a medium supplement [21,46]. For all cultures of Cun supplemented with different side-streams, two general statements can be made. ...
... Grape pomace is also rich in oils and phenolic compounds [44] and can be used as a nitrogen source [45]. Citrus peel is rich in flavonoids, D-limonene, and sugars and was already successfully used as a medium supplement [21,46]. ...
The quality and harvest of essential oils depend on a large number of factors, most of which are hard to control in an open-field environment. Therefore, Basidiomycota have gained attention as a source for biotechnologically produced terpenoids. The basidiomycete Cerrena unicolor (Cun) was cultivated in submerged culture, and the production of sesquiterpenoids was analyzed via stir bar sorptive extraction (SBSE), followed by thermo-desorption gas chromatography coupled with mass spectrometry (TDS-GC-MS). Identification of aroma-active sesquiterpenoids was supported by GC, coupled with an olfactory detection port (ODP). Following the ideal of a circular bioeconomy, Cun was submerged (up-scalable) cultivated, and supplemented with a variety of food industrial side-streams. The effects of the different supplementations and of pure fatty acids were evaluated by liquid extraction and analysis of the terpenoids via GC-MS. As sesquiterpenoid production was enhanced by the most by lipid-rich side-streams, a cultivation with 13C-labeled acetate was conducted. Data confirmed that lipid-rich side-streams enhanced the sesquiterpene production through an increased acetyl-CoA pool.
... A fourfold turnover in laccase production was observed for MP, when compared with glycerol, a threecarbon substrate. Kachlishvili et al. (2021) reported the enhanced production of laccase in response to increasing MP concentrations; whereas a shortened time course for laccase was observed using the same substrate, compared to other carbon sources (Elisashvili et al. 2018). However, the aforemen- Fig. 1a Effect of nitrogen sources on laccase production. ...
The quest for circular bioeconomy has been on the rise in recent years, and it is anticipated to fulfil the environmental sustainability aspect of the sustainable development goals (SDG 2030). In this regard, our investigation attempted the biotechnological appraisal of an enzymatic derivative of bacterial (Pseudomonas sp. HRJ16) mandarin peelings (MP) fermentation as a vehicle for an environmentally sustainable textile bioscouring. Production of the bacterial exudate (HRJ16 laccase) was optimized by response surface methodology (RSM), using the common low-cost agroindustrial waste (MP). HRJ16 laccase was further assessed for its advantageous biochemical and catalytic properties, and then applied in synthetic dye decolorization and denim bleaching. Results emphasized the extremotolerance of the exudate to temperature, pH, salts, cations and surfactants, when at least ca. 80% residual activity was recollected after exposure to the different extreme operating conditions. The interesting capabilities of the HRJ16 in this study culminated in its successful bioscouring of denim fabric over 6 h and the spontaneous decolorization of the resultant effluent. This constitutive properties of HRJ16 might make it a crucial catalyst for achieving a circular bioeconomy in the textile industry.
... P. lecomtei BRM 044603 stood out when grown on substrates based on grain, moringa and pigeon pea, ST-12, OT-12 and CT-12, with laccase enzymatic activity of 416.8 ± 20.28, 382.6 ± 25.37 and 227.6 ± 14.12 U/g, respectively (Figures 1-3). When cultivated in submerged fermentation, P. lecomtei BRM 044603 also showed great potential in terms of laccase activity, reaching 799.8 ± 42.0 U/mL when using oil palm decanter cake as the carbon source [38]; however, when growing on mandarin peels, laccase activity of P. lecomtei BCC 903 decreased dramatically to 6.4 ± 1.0 U/mL [39]. These contrasting results corroborate that the production of enzymes by the fungus is strain and carbon source dependent. ...
There is an urgent need for improvements in animal production, particularly for ruminants, such that more sustainable and efficient processes are developed for obtaining more nutritious and efficient feeds. Filamentous fungi can add value to residual plant biomass, and may also have the potential to produce metabolites and enrich plant biomasses used in animal nutrition, converting them into nutraceutical sources. Thus, in this work, filamentous fungal fermentation of ruminant feed biomasses commonly used in Brazil was performed, and the enrichment for bioactive metabolites was tested. For this, Fistulina hepatica, Ganoderma lucidum, Pleurotus pulmonarius, Panus lecomtei, and Aspergillus terreus were grown for 28 days on different substrates: starchy grains- (sorghum, oat, and corn), fibrous substrates (coast-cross, rice husk, and moringa plant) and protein-rich substrates (cottonseed cake and pigeon pea plant). Fermented substrates were evaluated for laccase activity, crude protein, β-glucan, and lovastatin content. The highest growth rate was observed for G. lucidum in oat substrate (OT-01) (0.708 ± 0.035 cm/day) and F. hepatica in oat + coast-cross + pigeon pea treatment (OT-10) (0.607 ± 0.012 cm/day). High laccase activity was observed for P. lecomtei grown in starchy grain + moringa + pigeon pea substrate, reaching an activity of 416.8 ± 20.28 U/g. A. terreus growth in ST-09 (sorghum + pigeon pea) showed higher protein (15.3 ± 0.46%), β-glucan (503.56 ± 8.6 mg/g) and lovastatin (1.10 ± 0.17 mg/g) content compared to untreated substrates. These results demonstrate that filamentous fungi are an alternative for nutraceutical enrichment of ruminant feed biomasses. To the best of our knowledge, this is the first report in which P. lecomtei and F. hepatica are evaluated for their ability to be cultivated in ruminant feed substrates from Brazil.
... For example, laccase activity increased significantly in the interaction zone in the dual cultures of T. versicolor vs S. gausapatum and T. versicolor vs B. adusta on a 2% MA medium, in which S. gausapatum was replaced by T. versicolor, and B. adusta remained locked with T. versicolor. Dual cultures of Cerrena unicolor with T. versicolor led to up-regulation of laccase activity, and two new laccase isoenzymes were produced in interspecific interaction (Kachlishvili et al. 2021). The natural capacity of these proteins in communications has not known yet, upregulated laccases might be associated with melanin produced at the cooperation zone, which can fill in as a defensive hindrance between interfacing organisms (Presley et al. 2020). ...
The biodegradation of lignocellulosic materials in nature is a complex process. To investigate the fungal competition patterns and degradation on Moso bamboo (Phyllostachys edulis), four wood decay fungi, including white-rot and brown-rot fungi, were selected as the test fungi in this research. Bamboo blocks were pre-colonized by different fungi then paired to form dual cultures. The results showed different competition outcomes among these fungi on potato dextrose agar, forest soil, and bamboo. Whit-rot fungus Trametes versicolor was highly competitive on bamboo where it could replace brown-rot fungi, and the biodegradation of bamboo was more severe in dual cultures, especially in the combination of Trametes versicolor and Gloeophyllum trabeum. The degradation of the bamboo components also differed with the monocultures and dual cultures. In monocultures of brown-rot fungi, cellulose was the most severely degraded component among the three primary components; while in monocultures of white-rot fungi, lignin was the most severely degraded component instead. The degradation of hemicellulose and lignin was more significant than that of cellulose in the dual cultures containing white-rot fungi. These results suggest that dual culturing of decay fungi has the potential to improve biomass utilization of bamboo and reduce the chemical treatment cost of raw materials. ARTICLE HISTORY
En el presente trabajo fue evaluado el efecto de los sobrenadantes de Trametes maxima CU1 sobre los parámetros físicos del pan. Para ello, se recuperaron los sobrenadantes de los cultivos de 18 días en un medio mineral suplementado con CuSO4, cáscara de naranja o cacahuete en combinación con paja de trigo, además de la presencia de un co-cultivo con Pycnoporus sanguineus CS2. Los títulos de lacasa, amilasas, celulasas y xilanasas se cuantificaron antes de su adición a la masa de pan. Una vez obtenidos los panes, se determinaron la altura, la dureza, la pérdida de peso (%), los poros por mm2 y el análisis del color. Los resultados de los títulos enzimáticos mostraron una diferencia estadísticamente significativa (p ≤ 0,05) entre los tratamientos, destacando en el medio con 350 mM CuSO4 la producción de lacasa, mientras que el medio suplementado con un 5% de cáscara de naranja presentó los cuatro tipos de actividades. En general, los panes presentaron diferencias (p ≤ 0.05) en los parámetros evaluados. Cabe destacar que el pan tratado con el sobrenadante con cáscara de naranja tuvo la mayor altura y color, al igual que el control, además de presentar la menor dureza entre los tratamientos enzimáticos. Por lo tanto, estos resultados demuestran el efecto del medio de cultivo sobre los perfiles enzimáticos del mismo hongo y su potencial aplicación en la industria panadera
The white rot fungus Cerrena unicolor 87613 has been previously shown to be a promising resource in laccase production, an enzyme with significant biotechnological applications. Conventional methods face technical challenges in improving laccase activity. Attempts are still being made to develop novel approaches for further enhancing laccase activity. This study aimed to understand the regulation of laccase activity in C. unicolor 87613 for a better exploration of the novel approach. Transcriptomic and metabolomic analyses were performed to identify key genes and metabolites involved in extracellular laccase activity. The findings indicated a strong correlation between the glutathione metabolism pathway and laccase activity. Subsequently, experimental verifications were conducted by manipulating the pathway using chemical approaches. The additive reduced glutathione (GSH) dose-dependently repressed laccase activity, while the GSH inhibitors (APR-246) and reactive oxygen species (ROS) inducer (H 2 O 2 ) enhanced laccase activity. Changes in GSH levels could determine the intracellular redox homeostasis in interaction with ROS and partially affect the expression level of laccase genes in C. unicolor 87613 in turn. In addition, GSH synthetase was found to mediate GSH abundance in a feedback loop. This study suggests that laccase activity is negatively influenced by GSH metabolism and provides a theoretical basis for a novel strategy to enhance laccase activity by reprogramming glutathione metabolism at a specific cultivation stage.
IMPORTANCE
The production of laccase activity is limited by various conventional approaches, such as heterologous expression, strain screening, and optimization of incubation conditions. There is an urgent need for a new strategy to meet industrial requirements more effectively. In this study, we conducted a comprehensive analysis of the transcriptome and metabolome of Cerrena unicolor 87613. For the first time, we discovered a negative role played by reduced glutathione (GSH) and its metabolic pathway in influencing extracellular laccase activity. Furthermore, we identified a feedback loop involving GSH, GSH synthetase gene, and GSH synthetase within this metabolic pathway. These deductions were confirmed through experimental investigations. These findings not only advanced our understanding of laccase activity regulation in its natural producer but also provide a theoretical foundation for a strategy to enhance laccase activity by reprogramming glutathione metabolism at a specific cultivation stage.
Endophytes are micro-organisms residing inside plants and are mostly transmitted by seeds. In this study, we used ITS meta-genomic sequencing analysis to investigate the fungal endophyte profile of seeds harvested from three different soybean genotypes (OT 13-08, AC Proteus and AC Harmony) grown in Manitoba, Canada. Protein and oil contents of these genotypes were different from each other. Out of the three soybean genotypes tested, AC Harmony showed the highest level (71.85%) of predicted fungal endophyte taxonomic units classified up to the genus level. At the species level analysis, 20 predicted micro-organisms were common in all genotypes. In addition, AC Harmony included the most diverse potential number (90) of endophyte species. The potential roles of the identified endophytes were further studied and the differential roles were observed based on previous evidence. This study will give insight into fungal endophytes in Canadian soybean genotypes for potential applications in agriculture.
