Chitosan Biosynthesis Research Articles

Biosynthesis of Chitosan encapsulated Silver- Nanoparticles using Probiotic-Lactobacillus plantarum strain and It's In vitro Anticancer Assessment on HeLa cells

Biosynthesis of Chitosan encapsulated Silver- Nanoparticles using Probiotic-Lactobacillus plantarum strain and It's In vitro Anticancer Assessment on HeLa cells

Bio synthesis of chitosan from shrimp shell chitin using Streptomyces griseoincarnatus RB7AG:Characterization and application as coating material to enhance shelf life of tomatoes (var. Ruby pusa) in post-harvest storgae condition

In this investigation, chitosan was successfully obtained from shrimp shell chitin through microbial degradation by Streptomyces griseoincarnatus RB7AG. The resulting chitosan showcased remarkable characteristics, including high solubility, a high degree of deacetylation, a well-defined crystalline structure, and a rough surface with minimal pores. The application of chitosan as a coating material for tomatoes demonstrated significant preservation benefits compared to commercially available chitosan. The chitosan coating proved effective in minimizing weight loss, reducing the production of reactive oxygen species (ROS), and preserving essential compounds such as anthocyanin content and antioxidant enzymes in tomatoes. Firstly, the chitosan coating served as a physical barrier, preventing moisture loss and maintaining the tomatoes' structural integrity. Secondly, the antimicrobial properties of chitosan hindered the growth of spoilage-causing microorganisms, extending the shelf life of the tomatoes. Additionally, the chitosan coating exhibited antioxidant properties, scavenging ROS and preserving the natural antioxidants in tomatoes. Furthermore, the chitosan coating facilitated controlled gas exchange, allowing optimal levels of oxygen and carbon dioxide to maintain the freshness of the tomatoes without accelerating the ripening process. With the help of the above-mentioned mechanisms, the potential of the microbially synthesized chitosan to enhance the shelf life of the tomatoes can be explained which was found better than the commercially synthesized chitosan coating. This research underscores the potential of microbially synthesized chitosan as an effective coating material but also provides insights into the intricate mechanisms involved in preserving tomatoes. The findings pave the way for innovative applications of chitosan-based coatings in the food industry, ensuring prolonged freshness and nutritional quality of perishable fruits and vegetables.

Transcriptomic and metabolomic analysis of the mechanisms underlying stress responses of the freshwater snail, Pomacea canaliculata, exposed to different levels of arsenic

Arsenic (As) pollution poses an important problem, but limited information is available about the physiological effects of As on freshwater invertebrates. Here, we investigated the physiological effects of chronic As exposure on Pomacea canaliculata, a freshwater invertebrate. High level of As (Ⅲ, 5 mg/L) inhibited the growth of P. canaliculata, whereas low level of As (Ⅲ, 2 mg/L) promoted growth. Pathological changes in shell and cellular ultrastructure due to As accumulation likely explain the growth inhibition at high As level. Low level of As simulated the expression of genes related to DNA replication and chitosan biosynthesis, potentially accounting for the growth promotion observed. High level of As enrichment pathways primarily involved cytochrome P450, glutathione, and arachidonic acid-mediated metabolism of xenobiotics. ATP-binding cassette (ABC) transporters, specifically the ABCB and ABCC subfamilies, were involved in As transport. Differential metabolites were mainly associated with the metabolism and biosynthesis of amino acids. These findings elucidate the dose-dependent effects of As stress on P. canaliculata growth, with low levels promoting and high levels inhibiting. Additionally, our findings also provide insights into As metabolism and transport in P. canaliculata.

Chitosan-Deficient Cryptococcus as Whole-Cell Vaccines.

Creating a safe and effective vaccine against infection by the fungal pathogen Cryptococcus neoformans is an appealing option that complements the discovery of new small molecule antifungals. Recent animal studies have yielded promising results for a variety of vaccines that include live-attenuated and heat-killed whole-cell vaccines, as well as subunit vaccines formulated around recombinant proteins. Some of the recombinantly engineered cryptococcal mutants in the chitosan biosynthesis pathway are avirulent and very effective at conferring protective immunity. Mice vaccinated with these avirulent chitosan-deficient strains are protected from a lethal pulmonary infection with C. neoformans strain KN99. Heat-killed derivatives of the vaccination strains are likewise effective in a murine model of infection. The efficacy of these whole-cell vaccines, however, is dependent on a number of factors, including the inoculation dose, route of vaccination, frequency of vaccination, and the specific mouse strain used in the study. Here, we present detailed methods for identifying and optimizing various factors influencing vaccine potency and efficacy in various inbred mouse strains using a chitosan-deficient cda1Δcda2Δcda3Δ strain as a whole-cell vaccine candidate. This chapter describes the protocols for immunizing three different laboratory mouse strains with vaccination regimens that use intranasal, orotracheal, and subcutaneous vaccination routes after the animals were sedated using two different types of anesthesia.

PROSPECTS FOR MODIFYING THE STRUCTURE OF CHITIN AND CHITOSAN OF HIGHER FUNGI TO EXPAND THE POTENTIAL OF THEIR APPLIED USE

The review is devoted to summarizing scientific data in the field of the chemical structure and properties of chitin and chitosan obtained from fungal biomass, and to analyzing directions for their modification for use in medicine and the food industry as substances with antibacterial, antiviral, wound-healing and anticoagulant activity. The features of chitin biosynthesis by fungi of the Basidiomycota, Ascomycota, Deuteromycota departments and chitosan biosynthesis by fungi of the Zygomycota department are covered. It has been shown that higher fungi contain chitin in their cell walls in the form of a chitin-glucan complex, while lower fungi (zygomycetes) contain chitin in the form of chitosan-glucan. Effective components of substrates that influence the production of polysaccharides by fungi have been identified - carbohydrates in the form of glucose, sucrose and maltose, organic forms of nitrogen in the form of yeast extract and corn flour, mineral components in the form of dihydrogen phosphate and dipotassium monohydrogen phosphate. Particular attention is paid to methods for isolating chitin and modifying it to chitosan with a detailed description of the physicochemical and biological properties of polymers. The review also presents the main reactions and methods for obtaining carboxymethyl and sulfo derivatives of chitin and chitosan. The biological properties and application of these groups of substances are described. In the carboxymethylation of chitin and chitosan, the choice of appropriate reaction conditions and reagents makes it possible to obtain carboxymethyl chitin, N-, O-, N,O-carboxymethylchitosan, or N,N-dicarboxymethylchitosan. The properties and applications of carboxymethyl derivatives of chitin and chitosan strongly depend on their structure, degree of substitution, and arrangement of amino or hydroxyl groups. The main reagents in the preparation of carboxymethyl derivatives are sodium monochloroacetate, monochloroacetic and glyoxalic acids. Carboxymethyl derivatives of chitin and chitosan are used as drug delivery systems, antimicrobial agents, in tissue engineering, as components of cosmetics and food products. Modification of chitosan with sulfate groups makes it possible to obtain chitosan 2-N-, 6-O-, 2-N-6-O- and 3-O-sulphates. The main sulfonating agents are oleum, pyridine and chlorosulfonic acid. Sulfonic derivatives of chitin and chitosan can be used as a basis for obtaining hemocompatible materials (with antithrombotic and antibacterial activities).

The Tissue Browning and Concomitant Toughening of Yellow Flammulina filiformis Stipes Is Caused by Oxidative Damage-Mediated Metabolic Disorder and Cell Wall Glycan Remodeling.

The browning and associated toughening of fruiting body stipes are the main causes of declines in the commercial production of yellow Flammulina filiformis. The dynamic metabolic changes from the top to bottom stipe sections of yellow F. filiformis fruiting bodies were investigated by integrating physiological, transcriptomic, and metabolomic analyses. The results indicated that oxidative stress levels gradually increased accompanying the degree of tissue browning and toughening from the top to bottom sections of F. filiformis stipes. In-depth analysis showed that there were remarkable changes in the expression of genes, and the content of metabolites correlated with the primary and secondary metabolism of F. filiformis stipes. Interestingly, the expression levels of genes participating in chitosan biosynthesis and the degree of deacetylation of chitosan increased from top to bottom in F. filiformis stipes, implying that cell wall glycan remodeling may contribute to concomitant toughening of the browning of F. filiformis stipes.

Immunization with a heat-killed prm1 deletion strain protects the host from Cryptococcus neoformans infection

ABSTRACT Systemic infection with Cryptococcus neoformans, a dangerous and contagious pathogen found throughout the world, frequently results in lethal cryptococcal pneumonia and meningoencephalitis, and no effective treatments and vaccination of cryptococcosis are available. Here, we describe Prm1, a novel regulator of C. neoformans virulence. C. neoformans prm1Δ cells exhibit extreme sensitivity to various environmental stress conditions. Furthermore, prm1Δ cells show deficiencies in the biosynthesis of chitosan and mannoprotein, which in turn result in impairment of cell wall integrity. Treatment of mice with heat-killed prm1Δ cells was found to facilitate the host immunological defence against infection with wild-type C. neoformans. Further investigation demonstrated that prm1Δ cells strongly promote pulmonary production of interferon-γ, leading to activation of macrophage M1 differentiation and inhibition of M2 polarization. Therefore, our findings suggest that C. neoformans Prm1 may be a viable target for the development of anti-cryptococcosis medications and, cells lacking Prm1 represent a promising candidate for a vaccine.

Engineering chitin deacetylase AsCDA for improving the catalytic efficiency towards crystalline chitin

Chitin deacetylase (CDA) catalyzing the deacetylation of crystal chitin is a crucial step in the biosynthesis of chitosan, and also a scientific problem to be solved, which restricts the high-value utilization of chitin resources. This study aims to improve the catalytic efficiency of AsCDA from Acinetobacter schindleri MCDA01 by a semi-rational design using alanine scanning mutagenesis and saturation mutagenesis. The quadruple mutant M11 displayed a 2.31 and 1.73-fold improvement in kcat/Km and specific activity over AsCDA, which can remove 68 % of the acetyl groups from α-chitin. Furthermore, structural analysis suggested that additional hydrogen bonds, contributing the flexibility of amino acids and increasing the negative charge in M11 increased the catalytic efficiency. The microstructure changes of α-chitin pretreated by the mutant M11 were observed and evaluated using 13C CP/MAS NMR spectroscopy, FT-IR spectroscopy, XRD and SEM, and the results showed that M11 more efficiently catalyzed the release of acetyl groups from α-chitin. This study would provide a theoretical basis for the molecular modification of CDAs and accelerate the process of industrial production of chitosan by CDAs.

The germination of fungal spores in water and enhanced their resistance to chlor(am)ine: Characteristics and mechanisms

Waterborne fungal spores could aggregate and germinate to form biofilms, and current studies mainly focus on the control of dormant and aggregated spores. Herein, the characteristics of fungal spores during germination and the control of germinated spores were investigated. There was no significant difference between the germination percentages at the same carbon concentration (per spore) in the medium and surface water, suggesting the great germination potential of fungal spores in surface water. During germination, the cell size and the complication increased. Besides, the DNA content, esterase activity and intracellular ROS level of fungal spores also increased during germination. Accordingly, the germinated spores had more differential genes and metabolites to regulate germination. The k-Cl2 and the k-NH2Cl of Aspergillus niger spores with 5 %-45 % germination were 0.15–0.23 min−1 and 0.23–0.28 min−1, which were lower than that of dormant spores (0.062 and 0.063 min−1), suggesting that germinated spores were more resistant to chlor(am)ine than dormant spores. The k-Cl2 and k-NH2Cl of germinated spores with different germination percentages were highly negatively related to cell size and the esterase activities. The transcriptome and metabolome analysis showed that germinated spores reprogrammed the related germination genes and promoted chitosan biosynthesis in the cell wall and protein synthesis, thereby enhancing cellular resistance to chlor(am)ine.

Interactions between copper homeostasis and the fungal cell wall affect copper stress resistance.

Copper homeostasis mechanisms are essential for microbial adaption to changing copper levels within the host during infection. In the opportunistic fungal pathogen Cryptococcus neoformans (Cn), the Cn Cbi1/Bim1 protein is a newly identified copper binding and release protein that is highly induced during copper limitation. Recent studies demonstrated that Cbi1 functions in copper uptake through the Ctr1 copper transporter during copper limitation. However, the mechanism of Cbi1 action is unknown. The fungal cell wall is a dynamic structure primarily composed of carbohydrate polymers, such as chitin and chitosan, polymers known to strongly bind copper ions. We demonstrated that Cbi1 depletion affects cell wall integrity and architecture, connecting copper homeostasis with adaptive changes within the fungal cell wall. The cbi1Δ mutant strain possesses an aberrant cell wall gene transcriptional signature as well as defects in chitin / chitosan deposition and exposure. Furthermore, using Cn strains defective in chitosan biosynthesis, we demonstrated that cell wall chitosan modulates the ability of the fungal cell to withstand copper stress. Given the previously described role for Cbi1 in copper uptake, we propose that this copper-binding protein could be involved in shuttling copper from the cell wall to the copper transporter Ctr1 for regulated microbial copper uptake.

Biosynthesis, characterization, biological and photo catalytic investigations of Elsholtzia blanda and chitosan mediated copper oxide nanoparticles

Bio synthesis of nanoparticles using plant parts has gained considerable attention, given the fact that the method is green, environment friendly, cheaper, simple and involves no hazardous substances. The present study involves the green synthesis of copper oxide nanoparticles ( CuO NPs ) using chitosan and the aqueous leaf extract of Elsholtzia blanda , an aromatic medicinal herb. The synthesized E.blanda -chitosan mediated copper oxide nanoparticles ( CPCE ) and E. blanda mediated copper oxide nanoparticles ( PCE ) were subjected to different characterization techniques, Ultraviolet–visible (UV–Vis), Fourier Transform Infrared Spectroscopy (FTIR), X-Ray Diffraction (XRD), Field Emission Scanning Electron Microscopy (FE-SEM), Energy Dispersive X-ray Analysis (EDAX), High Resolution Transmission Electron Microscopy (HRTEM) and Selected Area Electron Diffraction (SAED). The absorbance peaks in UV–Vis spectroscopy at 286 nm and 278 nm for CPCE and PCE respectively indicated the formation of nanoparticles. TEM and SEM employed for studying the surface morphology showed rod-like and spherical morphology bearing average size of 47.71 nm for CPCE and 36.07 nm for PCE. The antibacterial activities of the prepared nanoparticles were tested against Enterococcus faecalis, Staphylococcus aureus , Escherichia coli and Salmonella typhi by agar well diffusion method. The results indicate that CuO NPs possess effective antibacterial potential against all tested bacteria with a maximum zone of inhibition of 18 mm for Enterococcus faecalis . Antioxidant studies revealed the highest DPPH scavenging activity of 89% at 25 μg/mL concentration of the nanoparticles. The percentage of the photo catalytic degradation of Congo red was found to be 95% after 10 h.

Biosynthesis and characterization of Eupatorium adenophorum and chitosan mediated Copper oxide nanoparticles and their antibacterial activity

The biosynthesized metal oxide nanoparticles are of particular interest due to their wide applications in diverse fields. In the present study, copper oxide nanoparticles were successfully synthesized using chitosan and aqueous leaf extract of Eupatorium adenophorum. The green synthesized chitosan-plant extract mediated copper oxide (CPC) nanoparticles were characterized by Ultraviolet–visible (UV–Vis), Fourier Transform Infrared Spectroscopy (FTIR), X-ray Diffraction (XRD), Field Emission Scanning Electron Microscopy (FESEM), Energy Dispersive X-ray Analysis (EDAX), High Resolution Transmission Electron Microscopy (HRTEM) and Selected Area Electron Diffraction (SAED). The FESEM and HR-TEM shows the synthesized nanoparticles structures as Nano flakes and Nano rods with size ranging between 30 nm–80 nm. EDAX shows the purity of the elemental constitutions. Antibacterial efficacy for the synthesized nanoparticles was tested against two-gram positive bacteria viz. Enterococcus faecalis and Staphylococcus aureus and two gram negative bacteria viz. Escherichia coli and Salmonella typhi The maximum zone of inhibition for E. faecalis, S. aureus, E. coli, S. typhi were 18 mm, 12 mm, 11 mm and 12 mm respectively. The synergic effects between chitosan and plant extract in determining the shape and size of the nanoparticles are well established.

Biosynthesis of chitosan and Eupatorium adenophorum mediated zinc oxide nanoparticles and their biological and photocatalytic activities

Phyto fabricated nanoparticles, bearing the advantage of possessing the plants’ medicinal values and totally following the green approach in nanomaterial synthesis is gaining popularity in the scientific world. The present work investigates the green synthesis of zinc oxide nanoparticles using chitosan and Eupatorium adenophorum. Different spectroscopic and imaging techniques such as UV–Vis, FTIR, XRD, FE-SEM, EDAX, HRTEM and SAED were used to evaluate the structural morphology of the zinc oxide nanoparticles. The UV–Vis spectroscopy displayed absorbance peak at 355 nm. TEM and SEM images showed spherical and hexagonal morphology with average size of 42.60 nm. The synthesized nanoparticles were tested for antibacterial activity and the results showed effective antibacterial potential with a maximum zone of inhibition of 25 mm against Staphylococcus aureus. The maximum DPPH radical scavenging activity was 91.4% at the highest nanoparticle concentration while the α-amylase assay for antidiabetic test showed 68% inhibition at 25 μg/mL concentration of the nanoparticles. After 10 h, the percentage of congo red dye being photocatalytically degraded was found to be 93.4%. To the best of our knowledge this is the first ever study on Chitosan and Eupatorium adenophorum phytochemical mediated green synthesis of ZnO nanoparticles and evaluation of their biological and photocatalytic efficacy.

Improved green biosynthesis of chitosan decorated Ag- and Co3O4-nanoparticles: A relationship between surface morphology, photocatalytic and biomedical applications

AgNPs@Chitosan and Co3O4-NPs@Chitosan were fabricated with Salvia hispanica. Results showed MZI values of 5 and 30 mm for Co3O4-NPs- and AgNPs@Chitosan against S. aureus, and 15 and 21 mm for Co3O4-NPs- and AgNPs@Chitosan against E. coli (24 h, 20 μg/mL), respectively. MTT assays showed up to 80% and 90%, 71% and 75%, and 91% and 94% mammalian cell viability for the green synthesized, chemically synthesized AgNPs and green synthesized AgNPs@Chitosan for HEK-293 and PC12 cells, respectively, and 70% and 71%, 59% and 62%, and 88% and 73% for the related Co3O4-NPs (24 h, 20 μg/mL). The photocatalytic activities showed dye degradation after 135 and 105 min for AgNPs@Chitosan and Co3O4-NPs@Chitosan, respectively. FESEM results showed differences in particle sizes (32 ± 3.0 nm for the AgNPs and 41 ± 3.0 nm for the Co3O4NPs) but AFM results showed lower roughness of the AgNPs@Chitosan (7.639 ± 0.85 nm) compared to Co3O4NPs@Chitosan (9.218 ± 0.93 nm), which resulted in potential biomedical applications.

Biosynthesis and characterization of chitosan based hydrogel: A potential in vitro wound healing agent

Humans are continuously plagued by number of new uncured diseases, resulting in the search of new medicines with zero level side effects. In the present study, an attempt has been made to prepare, characterize and to test the wound healing activity of chitosan based hydrogel. Shrimp shells was collected from Cuddalore (Lat 11° 42́ N Long 79° 46́ E), Annankovil (Lat. 11°30.47 N; Long. 079°47. 02E) and Mudasalodai landing centre (11o29 ′N 79o46 ′E) (Southeast coast of India) for extraction of chitin and chitosan and further characterized by FTIR, 1H – NMR, DSC, SEM and XRD. Poly Ethylene Glycol (PEG) is used as a cross linking agent for the preparation of chitosan hydrogel. The prepared hydrogel was characterized by FTIR, SEM, DSC and its swelling property was tested. Synthesized chitosan hydrogel was shown very good level of growth inhibition activity against human pathogens and wound healing activity within 12 h to 24 h. Thus the present study suggests the possibility of utilizing seafood waste of crustacean shells for the extraction and preparation of useful biologically active compounds like hydrogel for medicinal drug delivery and therapeutic applications.

Bio-Synthesis and Characterization of CS–Ag Nano Hydrogel for Antibacterial Application

An environmental technique of herbal mediated blend of Ag nanoparticles is a substantial stage in the ground of nanotechnology. Chitosan (CS) is a polymer which is biocompatible and antibacterial. In this work, we have synthesized CS–Ag Nano hydrogel which is form with chitosan-based hydrogel merged in the herbal synthesized Ag nanoparticles. These green synthesized Ag-nanoparticles made from Polygonum bistorta plant leaves and described with the assistance of UV-vis spectrophotometer, and Dynamic Light Scattering (DLS). In this work our main focus to synthesized CS–Ag Nano hydrogel. These hydrogel was described by Fourier transform infrared (FTIR) spectroscopic method, X-ray diffraction (XRD) method, and contact angle. Nanoparticle size distribution was within 1 to 100 nm by DLS and the optimum wavelength was noted in 400 to 450 nm by UV-vis spectroscopic readings. A good antibacterial behavior has been displayed by these synthesized CS–Ag Nano hydrogel films against both E. coli (gram –ve) and S. aureus (gram +ve) with the maximum 7 mm inhibition zone.

The Aminoalkylindole BML-190 Negatively Regulates Chitosan Synthesis via the Cyclic AMP/Protein Kinase A1 Pathway in Cryptococcus neoformans.

Cryptococcus neoformans can cause fatal meningoencephalitis in patients with AIDS or other immunocompromising conditions. Current antifungals are suboptimal to treat this disease; therefore, novel targets and new therapies are needed. Previously, we have shown that chitosan is a critical component of the cryptococcal cell wall and is required for survival in the mammalian host and that chitosan deficiency results in rapid clearance from the mammalian host. We had also identified several specific proteins that were required for chitosan biosynthesis, and we hypothesize that screening for compounds that inhibit chitosan biosynthesis would identify additional genes/proteins that influence chitosan biosynthesis. To identify these compounds, we developed a robust and novel cell-based flow cytometry screening method to identify small-molecule inhibitors of chitosan production. We screened the ICCB Known Bioactives library and identified 8 compounds that reduced chitosan in C. neoformans We used flow cytometry-based counterscreens and confirmatory screens, followed by a biochemical secondary screen to refine our primary screening hits to 2 confirmed hits. One of the confirmed hits that reduced chitosan content was the aminoalkylindole BML-190, a known inverse agonist of mammalian cannabinoid receptors. We demonstrated that BML-190 likely targets the C. neoformans G-protein-coupled receptor Gpr4 and, via the cyclic AMP (cAMP)/protein kinase A (PKA) signaling pathway, contributes to an intracellular accumulation of cAMP that results in decreased chitosan. Our discovery suggests that this approach could be used to identify additional compounds and pathways that reduce chitosan biosynthesis and could lead to potential novel therapeutics against C. neoformansIMPORTANCECryptococcus neoformans is a fungal pathogen that kills ∼200,000 people every year. The cell wall is an essential organelle that protects fungi from the environment. Chitosan, the deacetylated form of chitin, has been shown to be an essential component of the cryptococcal cell wall during infection of a mammalian host. In this study, we screened a set of 480 compounds, which are known to have defined biological activities, for activity that reduced chitosan production in C. neoformans Two of these compounds were confirmed using an alternative method of measuring chitosan, and one of these was demonstrated to impact the cAMP signal transduction pathway. This work demonstrates that the cAMP pathway regulates chitosan biosynthesis in C. neoformans and validates that this screening approach could be used to find potential antifungal agents.

Chitosan Biosynthesis and Virulence in the Human Fungal Pathogen Cryptococcus gattii.

Cryptococcus gattii R265 is a hypervirulent fungal strain responsible for the recent outbreak of cryptococcosis in Vancouver Island of British Columbia in Canada. It differs significantly from Cryptococcus neoformans in its natural environment, its preferred site in the mammalian host, and its pathogenesis. Our previous studies of C. neoformans have shown that the presence of chitosan, the deacetylated form of chitin, in the cell wall attenuates inflammatory responses in the host, while its absence induces robust immune responses, which in turn facilitate clearance of the fungus and induces a protective response. The results of the present investigation reveal that the cell wall of C. gattii R265 contains a two- to threefold larger amount of chitosan than that of C. neoformans The genes responsible for the biosynthesis of chitosan are highly conserved in the R265 genome; the roles of the three chitin deacetylases (CDAs) have, however, been modified. To deduce their roles, single and double CDA deletion strains and a triple CDA deletion strain were constructed in a R265 background and were subjected to mammalian infection studies. Unlike C. neoformans where Cda1 has a discernible role in fungal pathogenesis, in strain R265, Cda3 is critical for virulence. Deletion of either CDA3 alone or in combination with another CDA (cda1Δ3Δ or cda2Δ3Δ) or both (cda1Δ2Δ3Δ) rendered the fungus avirulent and cleared from the infected host. Moreover, the cda1Δ2Δ3Δ strain of R265 induced a protective response to a subsequent infection with R265. These studies begin to illuminate the regulation of chitosan biosynthesis of C. gattii and its subsequent effect on fungal virulence.IMPORTANCE The fungal cell wall is an essential organelle whose components provide the first line of defense against host-induced antifungal activity. Chitosan is one of the carbohydrate polymers in the cell wall that significantly affects the outcome of host-pathogen interaction. Chitosan-deficient strains are avirulent, implicating chitosan as a critical virulence factor. C. gattii R265 is an important fungal pathogen of concern due to its ability to cause infections in individuals with no apparent immune dysfunction and an increasing geographical distribution. Characterization of the fungal cell wall and understanding the contribution of individual molecules of the cell wall matrix to fungal pathogenesis offer new therapeutic avenues for intervention. In this report, we show that the C. gattii R265 strain has evolved alternate regulation of chitosan biosynthesis under both laboratory growth conditions and during mammalian infection compared to that of C. neoformans.

Synthesis of chitosan incorporated neem seed extract (Azadirachta indica) for medical textiles

In present study, eco-friendly biosynthesis of Chitosan–Neem seed (CS-NS) composite was prepared by co-precipitation method using aqueous neem seed extract. Cotton fabrics were treated with two different crosslinking agents (Glutaraldehyde and Citric acid) then the synthesized composite coated on cotton fabric by chemical linkage between the composite and the cellulose structure. As synthesized composite materials and treated cotton fabrics were characterized by Fourier transform infrared spectroscopy for functional groups confirmation, X-ray diffraction for crystalline behavior determination, UV–vis spectroscopy analysis for optical property and High resolution scanning electron microscopy for Surface morphological properties. The antibacterial activity of CS-NS composite coated cotton fabric and CS-NS composite coated cotton fabric with crosslinking agents were tested against the gram-positive and gram negative bacteria by agar well diffusion method. The results demonstrated that CS-NS composite with crosslinked coated cotton fabric has higher antibacterial activity than without crosslinked cotton fabric. Thus the chitosan-neem seed composite may be applied to the medical textiles.

A Glycosylphosphatidylinositol Anchor Is Required for Membrane Localization but Dispensable for Cell Wall Association of Chitin Deacetylase 2 in Cryptococcus neoformans

ABSTRACTCell wall proteins (CWPs) mediate important cellular processes in fungi, including adhesion, invasion, biofilm formation, and flocculation. The current model of fungal cell wall organization includes a major class of CWPs covalently bound to β-1,6-glucan via a remnant of a glycosylphosphatidylinositol (GPI) anchor. This model was established by studies of ascomycetes more than a decade ago, and relatively little work has been done with other fungi, although the presumption has been that proteins identified in the cell wall which contain a predicted GPI anchor are covalently linked to cell wall glucans. The pathogenic basidiomycete Cryptococcus neoformans encodes >50 putatively GPI-anchored proteins, some of which have been identified in the cell wall. One of these proteins is chitin deacetylase 2 (Cda2), an enzyme responsible for converting chitin to chitosan, a cell wall polymer recently established as a virulence factor for C. neoformans infection of mammalian hosts. Using a combination of biochemistry, molecular biology, and genetics, we show that Cda2 is GPI anchored to membranes but noncovalently associated with the cell wall by means independent of both its GPI anchor and β-1,6-glucan. We also show that Cda2 produces chitosan when localized to the plasma membrane, but association with the cell wall is not essential for this process, thereby providing insight into the mechanism of chitosan biosynthesis. These results increase our understanding of the surface of C. neoformans and provide models of cell walls likely applicable to other undercharacterized basidiomycete pathogenic fungi.