Chitin Concentration Research Articles

Natural Feed Supplements From Crustacean Processing Side Streams for Improved Growth of Finfishes and Crustaceans: A Review.

Natural feed additives of plant/animal/microbial origin are researched as supplements in aquaculture to improve the properties of feed, minimize the usage of chemical alternatives, reduce food safety risks and ensure sustainability to combat global food and nutritional security. Side streams generated during shellfish processing possess valuable ingredients: protein, lipids, carotenoids, minerals and chitins. Considering the current trend of organic farming and antibiotic-free fish and shellfish, crustacean processing side streams and their derivatives seem promising and emerging resources as natural additives/supplements for formulating high-quality feeds with superior benefits. Lower concentrations of chitin and chitosan in diets are reported to stimulate the growth of shellfish and finfish under controlled conditions. Oligomers of chitosan and nano-chitosan are also the other potential derivatives as natural supplements in feed for better growth performance of aquaculture varieties. This review focuses on the significance of crustacean processing side streams and their derivatives, especially shrimp head meal, chitin, chitosan and chitosan oligosaccharides as potential natural additives in aquafeeds for promoting the growth performance of cultured fin fishes and shell fishes. Utilization in aquafeeds and the development of natural value-added supplements from crustacean processing side streams, especially shrimp head and shell leftover, offer an answer to the negative environmental impact due to its dumping; reduce the dependency on food fish for fish meal production & fishmeal for aquafeeds; solution to maintain the economic viability of the fish farmers & industry as well as to ensure the supply of safer and healthy aquatic foods to meet the objectives of sustainable development goals.

Characterisation, antibacterial activity, and prebiotic potential of dried Hermetia illucens L. larvae and of their fractions

Abstract In this study, black soldier fly (BSF) dried larvae were fractionated into their defatted, lipid, protein, and chitin-rich fractions. The samples were then characterised in terms of proximate composition, analysis of amino acids, analysis of the molecular weight distribution of proteins via gel electrophoresis, and fatty acids analysis of the lipid fraction. The antibacterial activity of all fractions was determined against Escherichia coli F4 and Streptococcus suis, two common swine pathogens. Finally, the prebiotic activity of the whole larvae and of their protein isolate and chitin-rich pellet was determined for Limosilactobacillus reuteri, a naturally occurring species in the gastro-intestinal tract of the pig. The results on amino acid profile showed that the protein extraction method used was not fully effective in cleaving chitin-bound proteins, as expected, as the true protein content of the chitin-rich sample was 48.5%. This was in accordance with the SDS-PAGE analysis, where the molecular weight distribution of proteins in the protein isolate sample did not show band at higher molecular weight, which were instead present in the chitin-rich sample. The analysis of fatty acid (FA) profile showed that lauric acid was the most abundant FA, in accordance with the literature. The results on microbiological analyses highlighted how all nitrogen-containing samples showed antibacterial activity against E. coli, the highest being the defatted fraction at 1% with a decrease of −0.77 log CFU/mL, while only the lipid fraction against S. suis (−0.48 log CFU/mL at 1%). All tested samples showed prebiotic activity for L. reuteri, but it was noticed that the increase in chitin concentration resulted in the decrease of the prebiotic effect. By combining the results of both antibacterial and prebiotic tests, it could be inferred that chitin might be considered as an antibacterial compound.

VmSom1 is essential for growth, development, maintenance of cell wall integrity and virulence in Valsa mali

Apple Valsa canker disease, caused by Valsa mali Miyabe et Yamada, severely endangers the healthy growth of apple trees. The Som1, located downstream of the cyclic AMP-dependent protein kinase A (cAMP-PKA) pathway, plays crucial roles in the growth, development, morphological differentiation, and virulence of filamentous fungi. In this study, we identify and functionally characterize VmSom1, a homolog of Som1, in Valsa mali. The VmSom1 gene is located on chromosome 12, encoding an 824 amino acid protein. Phylogenetic analysis reveals VmSom1 as a fungal Som1 homolog. The VmSom1 deletion mutants exhibit slower growth rates and fail to produce pycnidia. Additionally, their hyphal growth is significantly inhibited on media containing Calcofluor White, Congo Red, NaCl, and sorbitol. The growth rate of VmSom1 deletion mutants is reduced on maltose, lactose, sucrose and fructose media but increases on glucose medium. Moreover, the mycelial growth rate of the VmSom1 deletion mutant is significantly lower than that of the wild-type strain in peptone, NH4SO4, NaNO3, and no nitrogen. Notably, the distances between the septa increase, and chitin concentration shifts to the hyphal tip in the VmSom1 deletion mutant. Furthermore, compared with the wild-type strain, the VmSom1 deletion mutant exhibits fewer diseased spots on apple fruit and branches. Overall, our findings demonstrate that VmSom1 is involved in regulating the growth and development, colony surface hydrophobicity, osmotic stress, cell wall integrity maintenance, carbon and nitrogen source utilization, septa formation, and virulence of V. mali.

UPC2 mutations and development of azole resistance in Candida albicans hospital isolates from Lebanon

ObjectivesThis study evaluated the role of Upc2 in the development of azole resistance in Candida albicans isolates from Lebanese hospitalized patients and determined a correlation between resistance and virulence. MethodsThe UPC2 gene which codes for an ergosterol biosynthesis regulator was sequenced and analysed in two azole-resistant and one azole-susceptible C. albicans isolates. An amino acid substitution screening was carried out on Upc2 with a focus on its ligand binding domain (LBD) known to interact with ergosterol. Then, Upc2 protein secondary structure prediction and homology modelling were conducted, followed by total plasma membrane ergosterol and cell wall chitin quantifications. For virulence, mouse models of systemic infection were generated and an agar adhesion and invasion test was performed. ResultsAzole-resistant isolates harboured novel amino acid substitutions in the LBD of Upc2 and changes in protein secondary structures were observed. In addition, these isolates exhibited a significant increase in plasma membrane ergosterol content. Resistance and virulence were inversely correlated while increased cell wall chitin concentration does not seem to be linked to resistance since even though we observed an increase in chitin concentration, it was not statistically significant. ConclusionsThe azole-resistant C. albicans isolates harboured novel amino acid substitutions in the LBD of Upc2 which are speculated to induce an increase in plasma membrane ergosterol content, preventing the binding of azoles to their target, resulting in resistance.

Shifts in soil C stabilization mechanisms are linked to reindeer-induced changes in plant communities and associated fungi in subarctic tundra

Arctic tundra ecosystems store a significant proportion of the global soil organic carbon (C). However, warming-induced shrub encroachment and reindeer (Rangifer tarandus L.) grazing regimes promoting graminoid vegetation may strongly influence tundra soil C stability. Here, we studied how reindeer grazing intensity and experimental warming affect soil C stabilization in a tundra ecosystem. We hypothesized that under light grazing, persistent complexes formed by fungal necromass (FNM) and condensed tannins (CT) from shrub roots stabilize the soil C, whereas, under heavy grazing, the soil C stabilization is affected by glomalin-related soil proteins (GRSP) produced by arbuscular mycorrhizal fungi of graminoids. In addition, we expect warming to mediate grazing effects, diminishing the potential for C stabilization.Our results show no effect of grazing on stable C concentration, however, under light grazing the labile C concentration was higher. We found higher concentrations of chitin and tannins under light grazing, indicative of soil C stabilization potential through FNM-CT complexes. By contrast, we found more root ergosterol under heavy grazing, suggesting a high abundance of endophytes, usually melanized, and a slightly higher GRSP concentration. Warming did not cause changes in stable C concentration but was associated with changes in the soil chemical quality, pointing to a decrease of lignin, polypeptides, and polysaccharides.We conclude that different soil C stabilization mechanisms operate under light and heavy grazing pressures and that these mechanisms are closely linked to changes in the vegetation and the fungi typically associated with them.

Utility of by‐products of black soldier fly larvae (Hermetia illucens) production as feed ingredients for Pacific Whiteleg shrimp (Litopenaeus vannamei)

AbstractProjected growth in insect production as alternative feedstuffs will yield novel by‐products that are potentially valuable for aquafeed applications. We analyzed the nutrient composition of three by‐products occurring from black soldier fly larvae production (exuvia, cocoon, imago) and the bioavailability of key nutrients for Pacific Whiteleg shrimp (Litopenaeus vannamei). Protein accounted for 317 g kg−1 in exuviae, 433 g kg−1 in cocoons, and up to 521 g kg−1 in adult flies (imagines). Considerable amounts of essential amino acids were detected in imago meal, which significantly matched the ideal dietary amino acid composition for penaeid shrimp (r2 = 0.66, p = 0.0076). Exuviae and cocoons contained moderate amounts of lipids (64–140 g kg−1), while imagines comprised 356 g kg−1 total lipid. Saturated fatty acids predominated in all insect materials (47%–83% of total fatty acids). Chitin concentration was highest in cocoons and exuviae (154 and 139 g kg−1) and low in imagines (51 g kg−1). A feeding trial with shrimp, L vannamei, revealed apparent digestibility coefficients of 20%–59% for protein, 24%–54% for energy, 25%–49% for carbon, and 27%–68% for copper. Defatting of imago meal increased the digestibility of protein, energy, and carbon by 77%, 64%, and 61%, respectively. Defatted imago meal can serve as a protein supplement for shrimp diets.

Eco-friendly and safe alternatives for the valorization of shrimp farming waste.

The seafood industry generates waste, including shells, bones, intestines, and wastewater. The discards are nutrient-rich, containing varying concentrations of carotenoids, proteins, chitin, and other minerals. Thus, it is imperative to subject seafood waste, including shrimp waste (SW), to secondary processing and valorization for demineralization and deproteination to retrieve industrially essential compounds. Although several chemical processes are available for SW processing, most of them are inherently ecotoxic. Bioconversion of SW is cost-effective, ecofriendly, and safe. Microbial fermentation and the action of exogenous enzymes are among the significant SW bioconversion processes that transform seafood waste into valuable products. SW is a potential raw material for agrochemicals, microbial culture media, adsorbents, therapeutics, nutraceuticals, and bio-nanomaterials. This review comprehensively elucidates the valorization approaches of SW, addressing the drawbacks of chemically mediated methods for SW treatments. It is a broad overview of the applications associated with nutrient-rich SW, besides highlighting the role of major shrimp-producing countries in exploring SW to achieve safe, ecofriendly, and efficient bio-products.

A novel lytic polysaccharide monooxygenase from enrichment microbiota and its application for shrimp shell powder biodegradation.

Lytic polysaccharide monooxygenases (LPMO) are expected to change the current status of chitin resource utilization. This study reports that targeted enrichment of the microbiota was performed with chitin by the selective gradient culture technique, and a novel LPMO (M2822) was identified from the enrichment microbiota metagenome. First, soil samples were screened based on soil bacterial species and chitinase biodiversity. Then gradient enrichment culture with different chitin concentrations was carried out. The efficiency of chitin powder degradation was increased by 10.67 times through enrichment, and chitin degradation species Chitiniphilus and Chitinolyticbacter were enriched significantly. A novel LPMO (M2822) was found in the metagenome of the enriched microbiota. Phylogenetic analysis showed that M2822 had a unique phylogenetic position in auxiliary activity (AA) 10 family. The analysis of enzymatic hydrolysate showed that M2822 had chitin activity. When M2822 synergized with commercial chitinase to degrade chitin, the yield of N-acetyl glycosamine was 83.6% higher than chitinase alone. The optimum temperature and pH for M2822 activity were 35°C and 6.0. The synergistic action of M2822 and chitin-degrading enzymes secreted by Chitiniphilus sp. LZ32 could efficiently hydrolyze shrimp shell powder. After 12 h of enzymatic hydrolysis, chitin oligosaccharides (COS) yield reached 4,724 μg/mL. To our knowledge, this work is the first study to mine chitin activity LPMO in the metagenome of enriched microbiota. The obtained M2822 showed application prospects in the efficient production of COS.

Understanding the impact of main cell wall polysaccharides on the decomposition of ectomycorrhizal fungal necromass

AbstractThe extramatrical mycelium of ectomycorrhizal fungi (EMF) is an important source of soil carbon and nitrogen. While the importance of recalcitrant compounds in the fungal cell wall has been explored earlier, the contribution of highly abundant but labile components, like glucans, and the role of their temporal dynamics during decomposition remains unknown. For the first time, we examined how the concentration of three main fungal cell wall components (chitin, melanin, glucans) in EMF necromass are related to necromass decomposition, over a period of 6 weeks. Although the initial concentrations of the three components were not good predictors of necromass loss, we found species–specific trends of chitin and glucans loss over time. The chitin concentration during decomposition was tightly linked to the weekly necromass degradation, with trends of chitin loss being dissimilar across fungal species. Chitin concentration was positively correlated with the mass loss in the first week, but in the remaining 5 weeks, it was found to be weakly negatively correlated with mass loss. The similarity in susceptibility to the decomposition of glucans and chitin likely compensates for the impacts of interspecific differences in their initial concentration, leading to overall similar decomposition patterns. Alternatively, other, non‐measured, components (e.g., glycoproteins, N content) may contribute to explaining similar decomposition patterns. Our results indicate that ectomycorrhizal necromass decomposition processes differ from those of plant litter decomposition with, unlike in plants, differences in initial concentrations of major structural carbohydrates (e.g., glucans) being unrelated to differences in decomposition rates. These findings indicate that the decomposition of fungal material cannot be inferred from assumptions based on data provided by studies of plant decomposition.Highlights Necromass of different ectomycorrhizal species decomposes similarly Main cell wall components show different decomposition patterns The relationship between cell wall compounds and mass loss changes over time Mycorrhizal necromass decomposition differs substantially from that of plant litter.

Impact of elicitors on essential oil compositions and phytochemical constituents in Lavandula stoechas L.

Lavandula stoechas L. (LS) is an aromatic evergreen herb used broadly in the food, pharmaceutical, and perfume industries. However, the inducer effect of elicitors on secondary metabolites (SMs) biosynthesis in LS is nebulous. In addition, the precise mechanism of elicitors in cells remains unexplored. Hence, the primary objective of this study was to analyze the changes in phytochemical compositions of LSs treated with a biotic elicitor (chitin) and an abiotic one (copper nanoparticle) under in vitro and greenhouse conditions, with the aim of choosing an appropriate elicitor type, concentration, and exposure time for this species. In this study, the biochemical function of four chitin concentrations (i.e. 0, 50, 100 & 200 mg L−1) and four copper oxide nanoparticle concentrations (i.e. 0, 25, 50, & 100 mg L−1) at two exposure times (i.e. 5 & 10 days) was investigated in LS so as to compare with the un-elicited explants and the original plant materials. The analysis showed that the highest contents of the total phenolic (4.68 mg g−1 FW), flavonoids (0.68 mg g−1 FW), anthocyanins (36.51 mg g−1 FW), and flavonols (0.29 mg g−1 FW) compounds were observed in the elicited LSs. Besides, the role of elicitors in augmenting the percentage of SMs was intelligible, especially ‘lavandulol’, ‘1,8-cineole’, ‘germacrene D’, and ‘(E)-nerolidol’, which increased by 21.68%, 17.21%, 9.33%, and 8.11%, respectively. In conclusion, these findings indicate that utilizing elicitors at optimal concentrations and with timely durations of exposure can largely assist in improving the biotechnological production of SMs in LS, so that their potential for industrial use can be actualized.

"PREPARATION AND CHARACTERIZATION OF CHITIN HYDROGEL COMPOSITED WITH HALLOYSITE CLAY SOLUTION VIA PHASE INVERSION "

In this study, halloysite clay (HC), a reinforcing nanofiller, was mixed with 1 and 1.5% chitin solutions to prepare chitin–halloysite clay composite hydrogel by phase inversion under water vapor atmosphere at 25 °C. Chitin, extracted chemically from crab shell, was dissolved in N,N-dimethyl acetamide in the presence of 5% lithium chloride (DMAc/5% LiCl), and then different amounts of HC solution (0, 3, 5 and 7%) were added to evaluate the effect of HC on the fundamental properties of the obtained composite hydrogels. After the green route of preparation, the diameter and thickness of the hydrogel samples seemed to remain the same, while varying the concentrations of chitin and inorganic filler solution. As a result, in the case of the 1% chitin hydrogel, the tensile strength of the composite films increased from 335.9 to 489.8 kPa, while the elongation was around 61.3 and 86.0% with the addition of 0 and 7% HC solution. With the increase in the chitin content to 1.5%, the mechanical strength of the resultant composite hydrogels was enhanced. Moreover, the reduction in the equilibrium water content confirmed the formation of a composite hydrogel incorporating a well dispersed nanofiller, with good interfacial interaction between the nanofiller and the biopolymer matrix.

Applying Chitin Enhanced Diafiltration Process (CEFP) in Removing Cobalt from Synthetic Wastewater.

This research aims to study the removal of Cobalt (Co) using chitin. The optimum conditions for removing Co were ascertained through batch experiments. This study involves the determination of chitin metal-binding efficiency by using a polymer enhanced diafiltration setup that utilizes a membrane process (ultrafiltration) to keep the Chitin. The effects of several parameters on sorption like pH, the concentrations of chitin, and Co were examined. The best efficiency was reached if the setup was run at pH < 6.3 (i.e., chitin pKa). At acidic conditions and by employing 6 g/L of chitin, Co level (20 mg/L) was decreased at 95%. To further investigate the kinetics of sorption for each gram of chitin, equilibrium experiments were carried out. For 1-100 mM Co, the performed rheological measurements show that chitin was observed to be moderately shear thickening at relatively lower levels (4 and 6 g/L); further, it was moderately shear thinning at slightly more important levels (12 and 20 g/L). Some improvement of the raw polymer will be necessary to enhance sorption to a sustainable limit and make this scheme an economically viable process.

The secreted immune response peptide 1 functions as a phytocytokine in rice immunity.

Small signalling peptides play important roles in various plant processes, but information regarding their involvement in plant immunity is limited. We previously identified a novel small secreted protein in rice, called immune response peptide 1 (IRP1). Here, we studied the function of IRP1 in rice immunity. Rice plants overexpressing IRP1 enhanced resistance to the virulent rice blast fungus. Application of synthetic IRP1 to rice suspension cells triggered the expression of IRP1 itself and the defence gene phenylalanine ammonia-lyase 1 (PAL1). RNA-seq results revealed that 84% of genes up-regulated by IRP1, including 13 OsWRKY transcription factors, were also induced by a microbe-associated molecular pattern (MAMP), chitin, indicating that IRP1 and chitin share a similar signalling pathway. Co-treatment with chitin and IRP1 elevated the expression level of PAL1 and OsWRKYs in an additive manner. The increased chitin concentration arrested the induction of IRP1 and PAL1 expression by IRP1, but did not affect IRP1-triggered mitogen-activated protein kinases (MAPKs) activation. Collectively, our findings indicate that IRP1 functions as a phytocytokine in rice immunity regulating MAPKs and OsWRKYs that can amplify chitin and other signalling pathways, and provide new insights into how MAMPs and phytocytokines cooperatively regulate rice immunity.

Adding Several Types of Insect Flour to Increase the Virulence of Metarhizium anisopliae Local Isolates against Pests Cylas formicarius

The productivity of sweet potato in Timor Tengah Selatan (TTS) Regency is still far below the national productivity. The limiting factor is the attack of the pest Cylas formicarius both in planting and in storage. The aimed of this study was to examine the virulence of local isolates of Metarhizium anisopliae fungus cultured on growth media containing chitin. The study was conducted at the Plant Protection Laboratory from May-December 2021. The experiment was arranged using a Completely Randomized Design with five replications. The treatments were the type and concentration of chitin derived from three types of insects and arranged as follows: T0, control without chitin; T1, chitin from Gryllus assimilis 0.5%; T2, chitin from Gryllus assimilis 1%; T3, chitin from Locusta migratoria 0.5%; T4, chitin from Locusta migratoria 1%; T5, chitin from Oryctes rhinoceros 0.5%; and T6, chitin from Oryctes rhinoceros 1%. The results of this study showed that the addition of a chitin source to PDA growing media was able to increase conidia production, viability, and conidia virulence of M. anisopliae. Chitin also affects the virulence of M. anisopliae in C. formicarius which can increase the mortality percentage from 68% to 187%. Chitin in the form of flour derived from the insect O. rhinoceros with a concentration of 0.5% was an appropriate concentration to increase the viability and virulence of the M. anisopliae, besides that its protein content of 42.43% was high enough to support the production of quality entomopathogenic fungus.

PSXIII-B-1 Black Soldier fly Larvae Were More Digestible Than Adult Flies, and Nutrient Disappearance in Black Soldier fly Larvae can be Predicted Using Acid Detergent Fiber Based on in Vitro Assays for Pigs

Abstract The objectives were to evaluate the nutrient utilization of fish meal, black soldier fly (BSF, Hermetia illucens) larvae, and adult BSF meal and to develop equations for estimating in vitro nutrient disappearance of BSF larvae for pigs. Two-step and 3-step in vitro procedures were used to simulate the digestion and absorption of nutrients in the gastrointestinal tract of pigs. In Exp. 1, defatted BSF larva meal had less (P < 0.05) in vitro ileal disappearance (IVID) of dry matter (DM; 81.2% vs. 91.6% and 61.6%) and crude protein (CP; 81.8% vs. 92.2% and 67.2%) and in vitro total tract disappearance (IVTTD) of DM (82.6% vs. 93.6% and 65.7%) and organic matter (OM; 78.1% vs. 91.5% and 63.2%) than fish meal and greater (P < 0.05) values than adult BSF. In Exp. 2, CP concentrations in BSF larvae were negatively correlated with ether extract (r = −0.91) but positively correlated with acid detergent fiber (ADF; r = 0.98) and chitin (r = 0.95). Acid detergent fiber and chitin concentrations in BSF larvae were negatively correlated with IVID of DM (r = −0.98 and −0.88) and CP (r = −0.87 and −0.84) and IVTTD of DM (r = −1.00 and −0.94) and OM (r = −0.99 and −0.98). Equations for in vitro nutrient disappearance of BSF larvae using ADF (% DM) as an independent variable were developed: IVID of CP, % = −0.95×ADF + 95 (r2 = 0.75 and P = 0.058) and IVTTD of OM, % = −2.60×ADF + 115 (r2 = 0.98 and P < 0.01). In conclusion, in vitro nutrient utilization of defatted BSF larva meal was less compared with fish meal but was greater compared with adult flies and in vitro nutrient utilization of BSF larvae can be predicted using ADF as an independent variable.

Characterization of a novel chitinolytic Serratia marcescens strain TC-1 with broad insecticidal spectrum

The Gram-negative rod-shaped bacterium Serratia marcescens is an opportunistic pathogen of many organisms, including insects. We report the identification and optimal in vitro chitinase production conditions of a novel chitinolytic S. marcescens strain TC-1 isolated from a naturally infected white grub (Anomala corpulenta) collected from a peanut field at Nanyang city, Henan province, China. Strain identification was conducted by morphological, physiological, biochemical and molecular analyses. The amplified 16S rRNA gene of TC-1 showed a similarity greater than 99% with multiple strains of S. marcescens. Based on Neighbor-joining phylogenetic tree analysis of bacterial 16S rRNA gene sequences, TC-1 formed a clade with S. marcescens, clearly separated from other Serratia spp. The strain TC-1 showed larvicidal activities against five insect species (A. corpulenta, Plutella xylostella, Spodoptera exigua, Helicoverpa armigera, Bombyx mori) and the nematode Caenorhabditis elegans, but not against S. litura. The operating parameters of chitinase production by TC-1 were optimized by response surface methodology using a three-factor, three-level Box-Behnken experimental design. The effects of three independent variables i.e. colloidal chitin concentration (7–13 g l−1), incubation time (24–72 h) and incubation temperature (24–32 °C) on chitinase production by TC-1 were investigated. A regression model was proposed to correlate the independent variables for an optimal chitinase activity predicted as 20.946 U ml−1, using a combination of colloidal chitin concentration, incubation time and incubation temperature of 9.06 g l−1, 63.83 h and 28.12 °C, respectively. The latter agreed well with a mean chitinase activity of 20.761 ± 0.102 U ml−1 measured in the culture supernatants of TC-1 grown under similar conditions with a colloidal chitin concentration, incubation time and incubation temperature of 9 g l−1, 64 h and 28 °C, respectively. Our study revealed the S. marcescens strain TC-1 with potential as a biocontrol agent of insect pests and nematodes and demonstrated the proposed regression model's potential to guide chitinase production by this strain.

Morphological and structural characterization of chitin as a substrate for the screening, production, and molecular characterization of chitinase by Bacillus velezensis.

The processing of shellfishery industrial wastes is gaining much interest in recent times due to the presence of valuable components. Chitin is one of the valuable components and is insoluble in most common solvents including water. In this study, a novel gram-positive bacterial strain capable of solubilizing chitin was screened from a prawn shell dumping yard. The chitinolytic activity of the isolated strain was observed through the zone of hydrolysis plate assay. The hyper-producing isolate was identified as Bacillus velezensis through the 16S rRNA sequencing technique. The structural and morphological characterization of raw and colloidal chitin preparation was carried out using FTIR, XRD, and SEM analysis. The residual protein and mineral content, degree of polymerization, and degree of acetylation were reported for both raw and colloidal chitin preparations. There was a linear increase in the chitinase activity with an increase in the colloidal chitin concentration. The maximum activity of chitinase was observed as 38.98 U/mL for the initial colloidal chitin concentration of 1.5%. Supplement of additional carbon sources, viz., glucose and maltose, did not improve the production of chitinase and resulted in a diauxic growth pattern. The maximum chitinase activity was observed to be 33.10 and 30.28 U/mL in the colloidal chitin-containing medium with and without glucose as a secondary carbon source, respectively. Interestingly, the addition of complex nitrogen sources has increased the production of chitinase. A 1.95- and 2.14-fold increase in the enzyme activity was observed with peptone and yeast extract, respectively. The chitinase was confirmed using SDS-PAGE, native PAGE, and zymograms. The optimum pH and temperature for chitinase enzyme activity were found to be 7.0 and 44°C, respectively.

Mechano-Enzymatic Degradation of the Chitin from Crustacea Shells for Efficient Production of N-acetylglucosamine (GlcNAc)

Chitin, the second richest polymer in nature, is composed of the monomer N-acetylglucosamine (GlcNAc), which has numerous functions and is widely applied in the medical, food, and chemical industries. However, due to the highly crystalline configuration and low accessibility in water of the chitin resources, such as shrimp and crab shells, the chitin is difficult utilize, and the traditional chemical method causes serious environment pollution and a waste of resources. In the present study, three genes encoding chitinolytic enzymes, including the N-acetylglucosaminidase from Ostrinia furnacalis (OfHex1), endo-chitinase from Trichoderma viride (TvChi1), and multifunctional chitinase from Chitinolyticbacter meiyuanensis (CmChi1), were expressed in the Pichia pastoris system, and the positive transformants with multiple copies were isolated by the PTVA (post-transformational vector amplification) method, respectively. The three recombinants OfHex1, TvChi1, and CmChi1 were induced by methanol and purified by the chitin affinity adsorption method. The purified recombinants OfHex1 and TvChi1 were characterized, and they were further used together for degrading chitin from shrimp and crab shells to produce GlcNAc through liquid-assisted grinding (LAG) under a water-less condition. The substrate chitin concentration reached up to 300 g/L, and the highest yield of the product GlcNAc reached up to 61.3 g/L using the mechano-enzymatic method. A yield rate of up to 102.2 g GlcNAc per 1 g enzyme was obtained.

Trehalose-6-phosphate synthase influences polysaccharide synthesis and cell wall components in Ganoderma lucidum.

Trehalose-6-phosphate synthase (TPS) is a key enzyme that participates in trehalose metabolism, which can synthesize trehalose in a two-step pathway with trehalose phosphatase, but its role in fungi is rarely studied, especially in large basidiomycetes. In this study, the tps gene of Ganoderma lucidum was cloned and named as gltps. And gltps-silenced strains were constructed by RNA interference. In this study, it is found that the extracellular polysaccharidecontent increased 1.6-2-fold, but there was no significant change on intracellular polysaccharidecontent in gltps-silenced strains compared with the wild-type (WT) strain. Furthermore, the cell wall compositions of the gltps-silenced strains were also altered, which showed that the chitin and β-1,3-glucan contents were significantly decreased. Compared with WT, the concentration of chitin decreased by 20%-50% and that of β-1, 3-glucan decreased by 15%-30%. The study found that the cells of gltps-silenced strains were more sensitive to cell wall stress, which might be due to changes in the compounds and structure of the cell wall. These results showed that gltps had an important effect on carbohydrate metabolism of G. lucidum cells.

Effect of elicitors on secondary metabolites biosynthesis in Zataria multiflora Boiss.

Zataria multiflora Boiss. (ZM) is an aromatic herb that is used in various industries, especially in pharmaceuticals and food. However, the inducer effect of elicitors on ZM, with the aim of increasing the bioactive compounds production, is understudied. This study analyzes the influence of a biotic (chitin) and abiotic (copper nanoparticle) elicitor on ZM under in vitro and greenhouse conditions. In the first step, having established in Murashige and Skoog medium (MS) based on the node culture method, the explants transferred to media with different blended to survey the improvements in growth. The modified MS having faster growth and propagation in ZMs was chosen. In the second step, when explants were grown and rooted enough, they were transmitted to the media of elicitors, including four copper oxide nanoparticle concentrations (0, 25, 50, & 100 mg L−1) and four chitin concentrations (0, 50, 100 & 200 mg L−1) at two exposure times (5 and 10 days) to measure biochemical attributes. The analysis showed that the highest contents of the total phenolic (6.44 mg g−1 FW), anthocyanins (39.59 mg g−1 FW), flavonols (0.76 mg g−1 FW), and flavonoids (0.97 mg g−1 FW) compounds were observed in the elicited ZMs. In addition, the components of secondary metabolites were identified by gas chromatography−mass spectrometry (GC–MS) analysis. It was revealed that the highest amount of thymol and carvacrol, as the main compositions of ZM, was in the copper oxide nanoparticle (2.51% and 38.03% respectively), control (2.34% and 28.51% respectively), and chitin (1.92% and 9.84% respectively) ZMs in turn. The present research indicates that choosing an appropriate elicitor, concentration, and exposure time for augmenting the production of bioactive compounds in each plant species is consequential so as to provide the possibility of utilizing in industry.